Elevated platelets may signal increased cancer risk

Platelets as cancer predictorPlatelets (thrombocytes) are active for more than just adhesion and cohesion in the formation of a ‘hemostatic plug’ (blood clot), along with activation of coagulation mechanisms. Platelets also have important secretory functions that release growth factors and communicate with white blood cells and cells that line blood vessels (endothelial cells). Through this they promote inflammation and tissue proliferation (as in wound healing). Now an important study published in the British Journal of General Practice that an increase in platelet count is clinical risk marker for cancer. The authors note:

“The commonest route to cancer diagnosis follows the development of symptoms, and definitive diagnosis by biopsy and access to specialist care often rely on a primary care physician to recognise the possibility of cancer. It is generally accepted that delay in symptomatic diagnosis is harmful. One feature of possible cancer has only recently been recognised to have diagnostic potential: a raised platelet count, or thrombocytosis.”

Platelets as predictors

Earlier studies have shown the predictive value of thrombocytosis for certain cancers, but none have looked at cancer in general.

“Revised UK national guidance for suspected cancer incorporates thrombocytosis in some of its recommendations for lung, oesophagogastric, and uterine cancers. However, no study has examined thrombocytosis in primary care for all cancers. This study aimed to address that gap.”

The authors examined 1-year data for two groups of subjects: 40,000 patients aged ≥40 years with a platelet count of more than 400 × 109/L (109/L = 10³/uL) and 10,000 matched patients with a normal platelet count. Clinicians, note the reference range: >400 x 10³/uL = thrombocytosisTheir data did show that elevated platelets should be regarded as a cancer risk factor, especially for lung and colorectal cancer.

“A total of 1098 out of 9435 males with thrombocytosis were diagnosed with cancer (11.6%), compared with 106 of 2599 males without thrombocytosis (4.1%). A total of 1355 out of 21 826 females with thrombocytosis developed cancer (6.2%). The risk of cancer increased to 18.1% for males and 10.1% for females, when a second raised platelet count was recorded within 6 months. Lung and colorectal cancer were more commonly diagnosed with thrombocytosis.”

Very importantly:

One-third of patients with thrombocytosis and lung or colorectal cancer had no other symptoms indicative of malignancy.”

The authors summarize their findings:

“This large-scale cohort study is the first from primary care to report the overall risk of cancer in patients with thrombocytosis, compared with those with normal platelet counts. Males with thrombocytosis had an 11.6% incidence of cancer in the following year, and females had an incidence of 6.2%: this compares with 4.1% of males with normal platelet counts. The incidence of cancer rose with age and with a higher platelet count, and at least one-third of patients with lung and colorectal cancer with pre-diagnosis thrombocytosis had no other symptoms indicative of malignancy.”

Commenting in Medscape Family Medicine

“Lead author, Sarah Bailey, MPH, PhD, research fellow at the University of Exeter Medical School, United Kingdom, said in a statement:  “We know that early diagnosis is absolutely key in whether people survive cancer. Our research suggests that substantial numbers of people could have their cancer diagnosed up to three months earlier if thrombocytosis prompted investigation for cancer. This time could make a vital difference in achieving earlier diagnosis.”

Prediabetes, chronic inflammation and hemoglobin A1c

PrediabetesPrediabetes, blood glucose is slightly higher than normal but not enough to qualify for diabetes, is associated with an increased systemic burden of inflammation and elevated risk for cardiovascular, cancer, dementia and other diseases. The first study described in this post, published in the European Journal of Nutrition, highlights the link between prediabetes, chronic inflammation and mortality from a range of diseases tied to HgbA1c (hemoglobin A1c, glycosylated hemoglobin), the key biomarker for glucose regulation. The authors state:

Chronic inflammation is associated with increased risk of cancer, cardiovascular disease (CVD), and diabetes. The role of pro-inflammatory diet in the risk of cancer mortality and CVD mortality in prediabetics is unclear. We examined the relationship between diet-associated inflammation, as measured by dietary inflammatory index (DII) score, and mortality, with special focus on prediabetics.”

Pro-inflammatory diet plus prediabetes (increased HgbA1c)

Of great significance is the effect they reveal when a pro-inflammatory diet, measured by the dietary inflammatory index (DII) score, is consumed when there is elevated HgbA1c. They categorized 13,280 subjects between the ages 20 of and 90 years according to whether or not they were prediabetic, which they defined as a HgbA1c percentage of 5.7–6.4. Their data highlighted this connection between all-cause mortality, a pro-inflammatory diet and prediabetes:

“The prevalence of prediabetes was 20.19 %. After controlling for age, sex, race, HgbA1c, current smoking, physical activity, BMI, and systolic blood pressure, DII scores in tertile III (vs tertile I) was significantly associated with mortality from all causes (HR 1.39, 95 % CI 1.13, 1.72), CVD (HR 1.44, 95 % CI 1.02, 2.04), all cancers (HR 2.02, 95 % CI 1.27, 3.21), and digestive-tract cancer (HR 2.89, 95 % CI 1.08, 7.71). Findings for lung cancer (HR 2.01, 95 % CI 0.93, 4.34) suggested a likely effect.”

The authors conclude:

“A pro-inflammatory diet, as indicated by higher DII scores, is associated with an increased risk of all-cause, CVD, all-cancer, and digestive-tract cancer mortality among prediabetic subjects.”

 Prediabetes and cardiovascular risk

Research published in The BMJ (British Medical Journal) focusses on the substantial impact of prediabetes on the risk of heart attack and ischemic stroke. The authors set out to…

“…evaluate associations between different definitions of prediabetes and the risk of cardiovascular disease and all cause mortality…”

…by analyzing 53 prospective cohort studies with 1,611,339 individuals that passed the screening tests for validity. In this study they applied several definitions of prediabetes:

“Prediabetes was defined as impaired fasting glucose according to the criteria of the American Diabetes Association (IFG-ADA; fasting glucose 5.6-6.9 mmol/L = 101-124 mg/dL), the WHO expert group (IFG-WHO; fasting glucose 6.1-6.9 mmol/L = 110-124 mg/dL), impaired glucose tolerance (2 hour plasma glucose concentration 7.8-11.0 mmol/L = 141-198 mg/dL during an oral glucose tolerance test), or raised haemoglobin A1c (HbA1c) of 39-47 mmol/mol [5.7-6.4%] according to ADA criteria or 42-47 mmol/mol [6.0-6.4%] according to the National Institute for Health and Care Excellence (NICE) guideline.”

Their data show that prediabetes with a ‘mildly’ elevated HgbA1c was clearly associated with increased cardiovascular risk:

“Compared with normoglycaemia, prediabetes (impaired glucose tolerance or impaired fasting glucose according to IFG-ADA or IFG-WHO criteria) was associated with an increased risk of composite cardiovascular disease (relative risk 1.13, 1.26, and 1.30 for IFG-ADA, IFG-WHO, and impaired glucose tolerance, respectively), coronary heart disease (1.10, 1.18, and 1.20, respectively), stroke (1.06, 1.17, and 1.20, respectively), and all cause mortality (1.13, 1.13 and 1.32, respectively). Increases in HBA1c to 39-47 mmol/mol [5.7-6.4%] or 42-47 mmol/mol [6.0-6.4%] were both associated with an increased risk of composite cardiovascular disease (1.21 and 1.25, respectively) and coronary heart disease (1.15 and 1.28, respectively), but not with an increased risk of stroke and all cause mortality.”

Interestingly, risk of stroke does not emerge from these data, suggesting other factors promoting vascular inflammation. The authors conclude:

“…we found that prediabetes defined as impaired fasting glucose or impaired glucose tolerance is associated with an increased risk of composite cardiovascular events, coronary heart disease, stroke, and all cause mortality. There was an increased risk in people with fasting plasma glucose as low as 5.6 mmol/L [100 mg/dL]. Additionally, the risk of composite cardiovascular events and coronary heart disease increased in people with raised HbA1c. These results support the lower cut-off point for impaired fasting glucose according to ADA criteria as well as the incorporation of HbA1c in defining prediabetes.”

HgbA1c and risk of all-cause and cause-specific mortality without diabetes

Similar results were obtained in a study published in Scientific Reports. Here the authors concluded:

“We found evidence of a non-linear association between HbA1c and mortality from all causes, CVD and cancer in this meta-analysis. The dose-response curves were relatively flat for HbA1c less than around 5.7%, and rose steeply thereafter. This fact reveals a clear threshold effect for the association of HbA1clevels with mortality. In addition, from the perspective of mortality benefit and health care burden, it suggests that the most appropriate HbA1c level of initiating intervention is approximately 5.7%…higher HbA1c level is associated with increased mortality from all causes, CVD, and cancer among subjects without known diabetes. However, this association is influenced by those with undiagnosed diabetes or prediabetes .Because of limited studies, the results in relation to cancer mortality should be treated with caution, and more studies are therefore warranted to investigate whether higher HbA1c level is associated with increased cancer mortality.”

 

Ketone supplementation and the ketogenic diet for cancer

Clinical Cancer ResearchKetone enhancement by diet and supplementation can dramatically improve cancer survival. An easily implemented low carb high fat approach can produce results comparable to a more strict diet and be further enhanced by ketone supplementation.

Glucose consumption at a ravenous rate is characteristic of the peculiar metabolism of cancer cells. Fresh evidence for the ‘Warburg Effect’ the metabolic theory of cancer is eclipsing the earlier ascendant somatic mutation theory. Accordingly, ketogenic LCHF (low carb high fat) diets are gathering momentum in the treatment and prevention of malignancies. A study recently published in Clinical Cancer Research offers evidence that an easily implemented LCHF ketogenic diet supplemented with MCTs (medium chain triglycerides; sHFLC) may render significant benefit in the treatment of glioblastoma, a very malignant and metabolically active brain cancer, while being easier to maintain than a more strict ketogenic diet. The authors state:

“Dysregulated energetics coupled with uncontrolled proliferation has become a hallmark of cancer, leading to increased interest in metabolic therapies. Glioblastoma (GB) is highly malignant, very metabolically active, and typically resistant to current therapies. Dietary treatment options based on glucose deprivation have been explored using a restrictive ketogenic diet (KD), with positive anticancer reports. However, negative side effects and a lack of palatability make the KD difficult to implement in an adult population. Hence, we developed a less stringent, supplemented high-fat low-carbohydrate (sHFLC) diet that mimics the metabolic and antitumor effects of the KD, maintains a stable nutritional profile, and presents an alternative clinical option for diverse patient populations…We report a dietary intervention that produces low circulating glucose while elevating ketones and results in a substantial reduction in GB cellular proliferation.”

Glucose metabolism and the Warburg Effect

The bizarre metabolism of cancer cells has been recognized has an especially promising vulnerable target for treatment.

“Glucose metabolism and the Warburg Effect have gained traction as a potential tumor weakness and exploitable treatment area. Where normal cells utilize glucose for high-yield energy production in the mitochondria (1:36ATP), tumor cells demand higher levels of glucose for diminished energy production, via lactate in the cytosol (1:4ATP) and nucleotide synthesis in the pentose phosphate pathway. This metabolic characteristic, termed the Warburg Effect, is an essential byproduct of rapid cellular proliferation and promoted during tumorigenesis by oncogenic metabolic reprogramming. Hence tumor cells acquire the ability to sustain proliferative signaling mechanisms, which subsequently promotes malignant glycolysis. “

Ketogenic diet

The ketogenic diet has been around has been demonstrated to be safe and effective but in its strict form can be difficult to maintain.

“Research into dysregulated cellular metabolism has given rise to the notion that dietary therapies for cancer patients may have significant clinical utility. GB has been proposed to be a promising candidate for dietary intervention due to its substantial reliance and utilization of glucose. At the forefront of dietary anticancer therapy is the ketogenic diet (KD), which is a high-fat, low-carbohydrate, low-protein diet, used for decades to treat refractory epileptic seizures. Extreme carbohydrate restriction mimics a fasting state, resulting in reduction of blood glucose and induction of ketone bodies (e.g., b-hydroxybutyrate/BHB). Ketone bodies are suitable energy replacements for normal cells with functional mitochondria, but have been shown to be unsuitable for tumor cells, as tumor cell mitochondrial functions are dysregulated. Existing preclinical data support the KD and a calorie-restricted KD (RKD) in the treatment of brain cancer by diminishing tumor growth and increasing animal survival. Clinical report, case reports, and pilot trials have demonstrated that the KD is safe, has low toxicity, and is applicable to cancer patients.”

But because of challenges to implementation of the strict ketogenic diet the authors sought to investigate the effect of a more easily maintained less restrictive LCHF diet with supplemented with medium chain triglycerides (MCT).

“…a less restrictive KD-like diet that would exhibit the same physiologic phenotype and antitumor efficacy. By supplementing a high-fat, low-carbohydrate (sHFLC), moderate protein diet with specialized medium-chain triglycerides [MCT; 60%(30%):30%:10%::Fat(MCT): Protein:Carb], we hypothesize that a more balanced diet can be implemented, resulting in diminished tumor progression. MCTs were specifically chosen based on carbon chain lengths (C8: C10::97%:3%), which allow them to rapidly diffuse from the gastrointestinal tract into the hepatic portal system and travel directly to the liver where they are converted into ketone bodies). We believe it is possible to provide a more nutritionally complete, flexible, and palatable anticancer diet with the sHFLC, which could target a diverse patient population and increase patient compliance.”

Tumor cell growth significantly reduced

tumor-volume-with-high-fat-low-carb-low-glucose-ketogenicThey tested glioblastoma cells both in vivo and in vitro resulting in a significant decrease in the proliferation of both the tumor cells, and very importantly the tumor stem cells.

Lowering glucose concentrations resulted in a significant reduction in Ki-67 and MCM2 expression, in both PG and LG as well as a significant increase in active caspase-3 between NG and LG. Therefore, alterations in glucose availability, to levels equivalent to a low glucose state, are sufficient to slow the proliferation of gliomaspheres while concomitantly increasing apoptosis…Quantification of the self-renewing stem cell symmetrical division rate demonstrated a significant decrease in cancer stem cell expansion under reduced glucose conditions. The combination of diminished stem cell division rates, cellular fold expansion, and proliferation markers indicates that lowering glucose affects not only the putative stem cell population, but also the non-stem cell population.”

Low carb high fat benefits comparable to strict ketone diet

This is a very important point for practical implementation of an effective therapeutic diet.

“Animals placed on the sHFLC and KD had a significant reduction in blood glucose, above hypoglycemic levels, compared to controls, with no difference between the sHFLC and KD. Blood ketones were significantly increased in mice maintained on the sHFLC and KD to a safe level, with a statistical difference between the groups.”

The employed a glucose ketone index (GKI) to compare dietary interventions:

“To compare blood glucose and ketone levels among different anticancer dietary therapies, a simple glucose ketone index (GKI) is used. The GKI is a single number, and can be used both clinically and preclinically, to identify a therapeutic zone. On the basis of this formulation, our average calculated GKIs are 24.4 ± 7.14, 3.1 ± 1.07, and 1.94 ± 0.67 for the control, sHFLC and KD, respectively.”

Adding metformin yielded no additional benefit

Those following the topic of metformin as an anticancer therapy should note that it appears to be rendered unnecessary by this dietary approach:

“It has been proposed that a potential combinatorial treatment of metformin with carbohydrate restriction could result in enhanced antitumor efficacy. Mice fed the KD and sHFLC alone demonstrated a significant increase in survival compared with the control-fed mice, statistically equal to metformin alone. In both xenograft models, metformin alone was able to reduce blood glucose, reduce tumor progression, and increase survival, yet the combination sHFLC diet and metformin showed no additive or synergistic effects…These data indicate that the sHFLC diet is capable of increasing animal survival while minimizing tumor burden, is as effective as metformin, and may mechanistically overlap in AMPK-mediated inactivation of the mTOR pathway.”

Most importantly…

“The sHFLC diet slows tumor progression, increases survival, and reduces tumor burden in subcutaneous and orthotopic xenograft models.”

Low carb high fat with MCT oil much easier to maintain than strict ketogenic

Low carb high fat supplemented with MCT oil is as effective as the stricter ketogenic diet and has nutritional advantages.

“Here, we demonstrate that a high-fat, low-carbohydrate diet supplemented with MCT oil (sHFLC) is able to slow tumor progression and increase survival. In vivo, the sHFLC diet was similar to the ketogenic diet (KD) in antitumor efficacy, but showed nutritional advantages in body weight, organ enzyme levels, and lipid profile. Finally, we demonstrate that the sHFLC diet affects the mTOR signaling pathway by reducing expression of upstream regulators and translational downstream effectors…We designed the sHFLC diet for long-term sustainable maintenance of GB and for increased flexibility and palatability.”

Clinical note: Clearly, glucose control is crucial in case management for treatment and prevention of malignancies.

Excess glucose, as seen in GB patients with persistent hyperglycemia, leads to poor patient survival. It has also been suggested that diets with a high glycemic index may increase the risk of tumorigenesis, and low-carbohydrate, high-protein diets that limit circulating glucose can delay cancer development and progression.”

Moreover, regarding the hugely important issue of restricting cancer stem cell expansion:

“Treatment of GB stem cell lines with a constant physiologic concentration of BHB (4 mmol/L), as seen in KD patients, resulted in reduction of clonogenic frequency and symmetrical stem cell divisions, suggesting that elevated ketones affect the putative cancer stem cell population.”

Effective for diverse malignancies

Because a dependence on glycolysis is characteristic of all cancers these principles can be broadly applied.

“The combination of reduced glucose and increased ketone bodies has shown an enhanced anticancer effect…The KD mimics these biologic effects and has been proposed as a treatment for GB and other cancers…numerous groups have investigated the antitumor efficacy of a KD and an RKD in several types of cancer. RKD in experimental mouse models of glioma has been shown to be antitumorigenic, antiangiogenic, and pro survival, while also being anti-invasive, anti-inflammatory, and proapoptic by targeting signaling pathways related to glucose and glutamine metabolism. In animal models, feeding with the KD ad libitum has been reported to increase survival and reduce tumor growth. Other preclinical animal models such as gastric cancer, colon cancer, and metastatic cancer have used the KD, reporting similar antitumorigenic effects.”

Effects on the mTOR pathway is especially important:

“The mTOR pathway is one of the largest and most utilized pathways in cellular signaling, with two complexes (mTORC1/mTORC2) that have demonstrated a role in tumorigenesis. Recently, it’s been shown that inhibition of both mTORC1/mTORC2 signaling results in dramatically reduced cell viability in glioma cell lines, as well as inhibition of tumor growth in vivo. In our assessment of the sHFLC diet’s effects on the mTOR pathway, we found significant reduction in both mTORC1/mTORC2 signaling…Taken together, these findings indicate that inhibition of the mTORC1/2 pathway can be achieved through dietary intervention, resulting in a potent anti-cancer treatment.”

The authors conclusions highlight fundamental concerns in case management:

“Our work demonstrates that there is a distinct relationship between metabolism and proliferation that can be exploited by changing the energy sources in the body. Further research into the biochemical reactions of metabolic intermediates may shed more light on how ketone bodies are differentially utilized by tumor cells, as the role of mitochondria in tumor propagation and carcinogenesis is multifaceted and incompletely understood. Nevertheless, we effectively show that a combination of low glucose and high ketones results in negative proliferative effects on gliomaspheres, which can be translated in vivo with the sHFLC diet. This diet reduces overall tumor burden and increases survival, equivalent to a strict 1:6 KD, and has a complete nutritional profile. Hence we propose that dietary therapy, such as the sHFLC diet, could be utilized in the management of GB.”

The Warburg effect, foundation of the benefits of a low glycemic ketogenic diet

international-journal-of-cancerAn excellent study published in the International Journal of Cancer documenting decreased tumor cell viability and prolonged survival with supplemental ketones includes a fine review of the Warburg effect.

 “A century ago, Otto Warburg discovered that cancer cells display a unique metabolic phenotype of lactate fermentation in the presence of oxygen. This phenotype, known as the Warburg effect, enables tumor visualization using fluorodeoxyglucose positron emission tomography (FDG-PET) scans owing to the elevated rate of glucose consumption in most cancers. Metabolic therapies can exploit this phenotype, offering novel therapeutic directions aside from the classically targeted cytotoxic and gene-based therapies. The Warburg effect exposes a fundamental weakness of cancer cells, reliance on excess glucose for survival and maximal proliferation. Fasting, calorie restriction (CR) and the carbohydrate-restricted ketogenic diet have been successfully used to limit glucose availability and slow cancer progression in a variety of animal models and human studies.”

Importantly, the ketogenic effect is additive to the benefits of low glucose…

“Previously, the anticancer effects of these dietary manipulations have largely been attributed to decreased circulating blood glucose, which limits energy substrates for cancer cells. New evidence suggests, however, that the physiological state of ketosis and elevated circulating ketones also have anticancer effects.”

Ketogenic effect may be primary

For anti-cancer effects elevated ketones may be even more important than low glucose.

“Recently, Fine et al. demonstrated that a carbohydrate-restricted ketogenic diet inhibited disease progression and promoted partial remission in patients with advanced metastatic cancers from various tissue origins. [10] On average, the patients did not exhibit a drop in glucose from baseline, suggesting that decreased glucose availability was not the sole or primary cause of efficacy. Interestingly, the study found that the most important factor dictating the patients’ response to therapy was the degree of elevated ketosis from baseline. Indeed, a prominent metabolic shift to higher levels of ketosis correlated with reduced disease progression, stable disease or partial regression.”

Cancer as a mitochondrial disorder

The metabolic theory of cancer (as opposed to the somatic mutation theory) posits mitochondrial dysfunction as an instigator in the shift of normal to cancer cells.

“Although ketone bodies are efficient energy substrates for healthy extrahepatic tissues, cancer cells cannot effectively use them for energy. Widespread mitochondrial pathology has been observed in most if not all tumors examined, including decreased mitochondrial number, abnormal ultrastructural morphology, mitochondrial swelling, abnormal fusion–fission, partial or total cristolysis, mtDNA mutations, altered mitochondrial membrane potential and abnormal mitochondrial enzyme presence or function, among others.These defects in mitochondrial structure and function impair respiratory capacity and force a reliance on substrate-level phosphorylation for survival. As ketone bodies are metabolized exclusively within the mitochondria, cancer cells with impaired mitochondrial function are unable to efficiently metabolize ketone bodies for energy. Indeed, unlike healthy cells, ketone bodies fail to rescue glioma cells from glucose withdrawal-induced death.”

Ketone bodies oppose cancer in multiple ways

A ketogenic diet does more than just starve cancer cells…

  1. “Ketone bodies inhibit glycolysis, thus decreasing the main pathway of energy production for cancer cells.
  2. Cancer cells thrive in an environment of elevated reactive oxygen species (ROS) production but are very sensitive to even small changes in redox status. Ketones decrease mitochondrial ROS production and enhance endogenous antioxidant defenses in normal cells, but not in cancer cells. Ketone metabolism in healthy cells near the tumor may inhibit cancer cell growth by creating a less favorable redox environment for their survival.
  3. Ketone bodies are transported into the cell through the monocarboxylate transporters (MCTs), which are also responsible for lactate export. It has been shown that inhibiting MCT1 activity or inhibiting lactate export from the cell dramatically decreases cancer cell growth and survival. Ketones may impair cancer cells indirectly by competitive inhibition of the MCTs, decreasing critical lactate export from the cell.
  4. Recently, Verdin and coworkers demonstrated that βHB acts as an endogenous HDAC inhibitor at millimolar concentrations easily achieved through fasting, CR or ketone supplementation such as with a ketone ester (KE). Thus, ketone bodies may elicit their anticancer effects by altering the expression of oncogenes and tumor suppressor genes under control of the cancer epigenome.”

“Clearly, ketone bodies exhibit several unique characteristics that support their use as a metabolic therapy for cancer.”

Ketones oppose metastasis

Effectiveness against metastasis is critical for successful cancer therapy.

The Warburg effect is especially prevalent in aggressive cancers and metastatic cells. Metastasis, the spreading of a primary tumor to distal locations, is the primary cause of cancer morbidity and mortality and is responsible for more than 90% of cancer-related deaths.”

Ketones can be easily increased by diet and supplementation, so authors set out to investigate in vivo effectiveness:

It is possible to raise blood ketone levels without the need for carbohydrate restriction by administering a source of supplemental ketones or ketone precursors. 1,3-Butanediol (BD) is a commercially available food additive and hypoglycemic agent that is converted to βHB by the liver. The KE [ketone ester] elevates both AcAc and βHB in a dose-dependent manner to levels beyond what can be achieved with the KD or therapeutic fasting. Oral administrations of BD and KE have been shown to elevate blood ketones for at least 240 min in rats. As ketone bodies appear to elicit anticancer effects, and metastasis is the most significant obstacle in the successful treatment of neoplasms, we tested the efficacy of ketone supplementation in the VM-M3 cell line and mouse model of metastatic cancer.”

They measured proliferation and viability in highly metastatic cells cultured in the presence and absence of β-hydroxybutyrate (βHB). Also adult male inbred VM mice were implanted subcutaneously with firefly luciferase-tagged syngeneic VM-M3 cells and fed a standard diet supplemented with either 1,3-butanediol (BD) or a ketone ester (KE) which are metabolized into βHB and acetoacetate. They then monitored tumor growth in vivo bioluminescent imaging, and documented Survival time, tumor growth rate, blood glucose, blood βHB and body weight.

Ketone supplementation prolonged survival and reduced tumor burden

Tumor burden with ketone supplementation

Effect of supplemental ketones on tumor bioluminescence. (CR calorie restriction, BD 1,3-butanediol, and KE ketone ester).

The results were amazing, even without reducing glucose and calorie restriction:

Ketone supplementation decreased proliferation and viability of the VM-M3 cells grown in vitro, even in the presence of high glucose. Dietary ketone supplementation with BD and KE prolonged survival in VM-M3 mice with systemic metastatic cancer by 51 and 69%, respectively (p < 0.05). Ketone administration elicited anticancer effects in vitro and in vivo independent of glucose levels or calorie restriction.”

The authors discuss the profound clinical implications:

“The Warburg effect is the most ubiquitous cancer phenotype, exhibited by most if not all cancer types. Exploiting the metabolic deficiencies of cancer cells should be prioritized, because this therapeutic strategy would likely prove effective against most cancers. Mitochondrial dysfunction underlies many aspects of cancer metabolic deficiency and prevents cancer cells from effectively using ketone bodies for energy. In our study, ketone supplementation decreased VM-M3 cell proliferation and viability, confirming similar results demonstrated in other cancer types in vitro. Therefore, we hypothesized that dietary administration of ketone body precursors would inhibit disease progression in vivo. Indeed, dietary administration of ketone precursors, BD and KE, increased mean survival time by 51 and 69%, respectively, in VM-M3 mice with metastatic cancer. These data support the use of supplemental ketone administration as a feasible and efficacious cancer therapy, which should be further investigated…”

Although carbohydrate restriction has other important metabolic benefits, ketone supplementation was effective in prolonging survival even without it.

“Ketone supplementation decreased blood glucose after acute administration, decreased body weight with chronic administration and sustained ketosis in vivo, even when administered with a high-carbohydrate rodent chow in both healthy (VM/Dk) and cancer (VM-M3) mice. Our study demonstrates the ability of dietary administration of BD and KE to significantly elevate ketone bodiesin vivo for at least 12 hr in healthy VM/Dk mice and 7 days in VM-M3 cancer mice.”

Moreover, ketone supplementation on its own reduces weight, diminishes appetite and improves insulin sensitivity.

“It is important to note that the metabolic changes associated with acute and chronic ketosis are vast and can dramatically affect blood metabolite concentrations. In previous studies, chronic BD and βHB administration has been shown to decrease food intake in the rat and pigmy goat. Similarly, Veech and coworkers demonstrated that feeding a KE-supplemented diet increased malonyl-CoA, an anorexigenic metabolite known to decrease food intake. Ketone-induced appetite suppression may account for the decreased blood glucose and body weight seen in treated VM-M3 cancer mice. Additionally, prior studies suggest that ketones increase insulin sensitivity, which may be contributing to the decreased circulating blood glucose in KE-fed mice…Furthermore, chronic ketosis enhances ketone utilization by tissues, known as keto-adaptation, resulting in lower blood ketone concentrations.”

A ketogenic high fat diet with ketone supplementation may be more effective than calorie restriction

In the past, dietary treatment in cancer has emphasized carbohydrate or calorie restriction to exploit the Warburg effect, but this may not be the best approach.

“Interestingly, although CR [calorie restriction] decreased blood glucose and elevated blood ketones, CR mice exhibited a trend of increased latency to disease progression and increased survival that was not statistically significant from controls in our study. As described, some data suggest that elevated ketones are responsible for much of the anticancer efficacy of the ketogenic diet.Perhaps elevating ketones with exogenous sources such as ketone supplementation or a ketogenic diet, rather than elevating ketones endogenously through lipolysis such as occurs with CR, provides a more effective anticancer strategy. Additionally, ketone supplementation may preserve lean muscle mass to a greater degree than CR, and may therefore support overall health of the organism in this way…These data support the in vitro and in vivo conclusions of Fine et al. suggesting that ketone bodies can inhibit cancer progression independently of other factors such as carbohydrate restriction or CR.

Enhancement of radiation and chemotherapy

Ketogenic diet and ketone supplementation can enhance the cytotoxic effects of the increase in ROS (reactive oxygen species) by radiation and chemotherapy.

“The ketogenic diet has been shown to enhance the efficacy of both radiation and chemotherapy in vivo. As supplemental ketones mimic the physiological ketosis induced by the ketogenic diet, combining supplemental ketone therapy with standard of care could produce similar effects, even if administered with a SD [standard diet]. Furthermore, the neuroprotective effects of ketone metabolism have been widely documented. Ketone metabolism protects normal cells from oxidative damage by decreasing mitochondrial ROS production and enhancing endogenous antioxidant defenses. Radiation and chemotherapy work in large part by inducing ROS production in the tumor, but simultaneously incur damage to normal tissue. Ketone metabolism by healthy tissue would likely mitigate some of the adverse side effects of standard of care as ketones have been shown to protect against oxidative stress.”

The authors’ conclusion needs to be appreciated by any practitioner involvement in cancer case management:

Our data strongly suggest that supplemental ketone administration could provide a safe, feasible and cost-effective adjuvant to standard care that should be further investigated in preclinical and clinical settings.”

Exploiting Cancer Metabolism with Ketosis—Dr. Angela Poff

Chemotherapy: how much does it actually improve survival?

BMJ chemotherapy and survivalChemotherapy drugs have had little effect on cancer survival in adults according to an analysis just published in BMJ (British Journal of Medicine).

The author notes that although there have been advances in chemotherapy in recent years, a thorough examination of the data reveals that…

“Despite considerable investment and innovation, chemotherapy drugs have had little effect on survival in adults with metastatic cancer.”

In fact, the data on the survival benefits of chemotherapy are so disturbing that they raise questions about ethics, drug trials and approval, and patient consent.

The approval of drugs with such small survival benefits raises ethical questions, including whether recipients are aware of the drugs’ limited benefits, whether the high cost:benefit ratios are justified, and whether trials are providing the right information.”

Key messages include:

  • “Advances in chemotherapy have contributed little to population cancer survival

  • Responses in clinical trials may not apply to patients treated in the community

  • Evaluation outside trial centres is essential to ensure that scarce resources are not squandered

  • Stricter approval criteria are needed to achieve ethical treatment and reduce cancer costs

  • Ethical informed consent and empowerment of patients must be promoted”

The author strongly advises a more clinically sound and ethical way forward:
Many irregularities and competing interests—in pharma, in trials, in government approval, and in the clinical use of cancer drugs—impact ethically on the care and costs of patients with cancer. Non-representative clinical trials with imprecise endpoints and misinformed patients with unrealistic expectations compel interventions that are mostly not in their best interests. Spending a six figure sum to prolong life by a few weeks or months is already unaffordable, and inappropriate for many of the 20% of the (Western) population who will almost inevitably die from solid tumour metastases.”

 

Patients deserve better information and more supportive treatment….

Ethical cancer care demands empowerment of patients with accurate, impartial information followed by genuinely informed consent in both the clinical trial and therapeutic settings. Intensified prevention, earlier detection, more prompt and radical treatment of localised and regional disease, together with highly skilled, earlier, supportive care are the important yet underfinanced priorities in cancer control. Ethical impediments to sound practice need to be addressed and corrected. Above all, the efficacy bar for approval needs to be raised for both new and existing cancer drugs—by using more meaningful statistical and disease specific criteria of risk-benefit and cost-benefit. Finally, aggressively targeting the less than ethical actions of stakeholders in the heavily veiled medical-industrial complex may be the only way forward: current market driven rather than health driven priorities and practices do not benefit cancer patients.”

The entire paper can be downloaded here, and an illuminating, brief interview with the author can heard by clicking on the arrow in the upper left corner of the image below:

 

the-bmjThe editor-in-chief of BMJ in an accompanying editorial states:

“People with cancer are living longer now than 40 years ago. This is clearly good news. But how much of this improvement can we attribute to drug treatment? Not much, concludes Peter Wise this week in an article I humbly suggest all oncologists should read. The nearly 20% improvement in five year survival over the past four decades is probably mainly due to improved early diagnosis and treatment rather than developments in cytotoxic chemotherapy, he says. And patients are being badly misled by over-enthusiastic accounts of what chemotherapy can achieve. Many expect a cure. In reality they will gain on average only a few months of extra life.”

Moreover…

Unjustified enthusiasm for cancer drug treatments comes at huge cost, financial and personal (including treatment related deaths and reduced quality of life), and increased risk of dying in hospital rather than at home. Many patients don’t realise that opting for supportive rather than active treatment—often called “refusal”—is an option and may give them longer as well as better quality life than chemotherapy. Conflicts of interest among clinicians compound their reluctance to have tricky conversations.”

The editor summarizes the author’s recommendations in light of this data:

“Wise concludes with a call for higher bars for drug approval for new and existing drugs. Ethical cancer care demands empowerment of patients, he says, with accurate, impartial information followed by genuinely informed consent. And funds and attention should shift to prevention, early detection, prompt and radical treatment of localised and regional disease, and early provision of supportive care. Only then will cancer care serve patients rather than governments and industry.”

ONCOblot cancer test, sensitive and specific, explained in brief video

This brief video of less than 3 minutes offers a quick explanation of the valuable ONCOblot test for cancer detection.

The ONCOblot test has demonstrated extraordinary accuracy down to as little as 2 million cells or less, with identification of tissue of origin.

BioMed Central paper on ONCOblot test for early cancer detectionA recent study published in Clinical Proteomics illustrates how this test can detect cancer years in advance of a clinical diagnosis.

“In a population of asbestos-exposed subjects who eventually developed malignant mesothelioma, ENOX2 protein transcript variants characteristic of malignant mesothelioma were present in serum 4–10 years in advance of clinical symptoms.”

For further information see ONCOblot® Labs. Practitioner colleagues who would like to discuss our use of this test are welcome to contact.

Nuts reduce inflammation and all-cause mortality

Asia Pacific Journal of Clinical NutritionNuts have been shown to confer multiple health benefits, so it’s disconcerting to see  some apparently popular paleo diet plans that forbid them. In the absence of a nut allergy it’s a shame to forgo the benefit of such a healthful and convenient food. The intent of the paleo diet is to reduce inflammation, so it’s worth considering a paper published in the Asia Pacific Journal of Clinical Nutrition offering evidence that nuts reduce inflammation. The authors note:

“Several large epidemiological studies have associated the frequency of nut consumption with reduced risk of coronary heart disease (CHD), CVD, myocardial infarction, sudden death, and all causes of mortality, Type 2 diabetes (T2D) and other chronic disease.

Nuts are anti-inflammatory

Key inflammatory markers including CRP and IL-6 are reduced by nut consumption:

“Epidemiological and clinical studies suggest that some dietary factors, such as n–3 polyunsaturated fatty acids, antioxidant vitamins, dietary fiber, L-arginine and magnesium may play an important role in modulating inflammation. The relationship observed between frequent nut consumption and the reduced risk of cardiovascular mortality and type 2 diabetes in some prospective studies could be explained by the fact that nuts are rich in all of these modulator nutrients. In fact, frequent nut consumption has been associated with lower concentrations of some peripheral inflammation markers in cross-sectional studies. Nut consumption has also been shown to decrease the plasma concentration of CRP, IL-6 and some endothelial markers in recent clinical trials.”

Nuts also benefit cholesterol and lipids

“In the last two decades, a considerable number of clinical trials have consistently demonstrated beneficial effects on blood lipids and lipoproteins, primarily a decrease in Low-density lipoprotein (LDL) cholesterol, a classical CHD risk factor. This effect has been demonstrated consistently in different population groups, using different types of nuts (walnuts, hazelnuts, almonds, pecan, pistachio and macadamia nuts) and study designs. The favourable effects of tree nuts or tree nut oils on plasma lipid and lipoprotein profiles is a mechanism that appears to account for some of the cardio protective effects observed in the epidemiological studies.”

Nuts and olive oil are a great combination for cardiovascular risk:

“…in a cross-sectional study we evaluated the association between components of the Mediterranean diet and circulating markers of inflammation in a large cohort of asymptomatic subjects with high risk of cardiovascular disease. Subjects with the highest consumption of nuts and virgin olive oil showed the lowest concentrations of VCAM-1, ICAM-1, IL-6 and CRP; although this difference was statistically significant for ICAM-1 only in the case of nuts and for VCAM-1 in the case of olive oil.”

After reviewing several other studies documenting improvements in inflammation and endothelial function the authors conclude:

“In conclusion, nuts are complex food matrices containing diverse nutrients and other chemical constituents that may favourably influence human physiology. These sub- stances may inhibit the activation of the innate immune system, probably by decreasing the production of proinflammatory cytokines such as CRP, IL-6, TNF-α or IL-18, and increase the production of antiinflammatory cytokines such as adiponectin. This may improve the proinflammatory milieu, which in turn ameliorates endothelial dysfunction at the vascular level, and ultimately decreases the risk of insulin resistance, type 2 diabetes and coronary heart disease. The capacity of nuts to modulate inflammation may explain at least in part why frequent nut consumption is associated with reduced risk of diabetes and cardiovascular disease in epidemiological studies.”

Nut consumption reduces total and cause-specific mortality

New England Journal of MedicineA paper published earlier this year in The New England Journal of Medicine add more extensive data presenting evidence that eating nuts reduces death from cancer, heart disease, respiratory disease and ‘all causes’.

“Observational and intervention studies of nut consumption have also shown reductions in various mediators of chronic diseases, including oxidative stress, inflammation, visceral adiposity, hyperglycemia, insulin resistance, and endothelial dysfunction. In prospective cohort studies, increased nut intake has been associated with reduced risks of type 2 diabetes mellitus, the metabolic syndrome, colon cancer, hypertension, gallstone disease, diverticulitis, and death from inflammatory diseases.”

To extend the data to encompass the effects of eating nuts and all causes of death the authors:

“…examined the association of nut consumption with total and cause-specific mortality in two large, independent cohort studies of nurses and other health professionals. These studies provide repeated measures of diet (including separate data on peanuts and tree nuts), extensive data on known or suspected confounding variables, 30 years of follow-up, and data on more than 27,000 deaths for analysis.”

Their data suggest that nuts are among the healthiest foods to eat:

“In two large prospective U.S. cohorts, we found a significant, dose-dependent inverse association between nut consumption and total mortality, after adjusting for potential confounders. As compared with participants who did not eat nuts, those who consumed nuts seven or more times per week had a 20% lower death rate. Inverse associations were observed for most major causes of death, including heart disease, cancer, and respiratory diseases. Results were similar for peanuts and tree nuts, and the inverse association persisted across all subgroups.”

Some nuts every day was the best:

“Our results are consistent with the findings in previous, smaller studies. The Adventist Health Study showed that, as compared with nut consumption less than once per week, consumption five or more times per week was associated with reduced total mortality among whites, blacks, and elderly persons, with hazard ratios ranging from 0.56 to 0.82. Similarly, a study of a U.K. cohort, the Iowa Women’s Health Study, the Netherlands Cohort Study, and an earlier analysis of the NHS all showed significant inverse associations between nut intake and total mortality. Finally, in a recent secondary analysis within the PREDIMED (Prevención con Dieta Mediterránea) trial, a hazard ratio for death of 0.61 (95% CI, 0.45 to 0.83) was found for consumption of more than three servings of nuts per week, as compared with no nut consumption.”

Bottom line: ‘paleo’ and ‘autoimmune’ paleo diets can be fine healing diets for many, but like everything else should not be applied dogmatically or in a ‘rubber stamp’, ‘one-size-fits-all’ manner. In the absence of allergy, the evidence supports the consumption of nuts as wholesome foods with anti-inflammatory and metabolic benefits, exactly what paleo diets intend to accomplish.

Prediabetes increases cancer risk

DiabetologiaPrediabetes, elevated levels of blood sugar that are still ‘within’ the normal range, increases cancer risk among its mob of other afflictions as further validated by a meta-analysis just published in Diabetologia. The authors state:

Prediabetes is a general term that refers to an intermediate stage between normoglycaemia and overt diabetes mellitus. It includes individuals with impaired glucose tolerance (IGT), impaired fasting glucose (IFG) or a combination of the two. In 2003, the ADA redefined the range of fasting plasma glucose (FPG) concentration for diagnosing IFG from 6.1– 6.9 mmol/l to 5.6–6.9 mmol/l [101-124 mg/dL] in order to better identify individuals at risk of developing diabetes.”

Because this lower range has been disputed with inconsistencies in previous studies, the authors set out to…

“…to evaluate the putative association between different definitions of prediabetes and risk of cancer.”

Their data adds yet more weight to the vital clinical importance of regulating blood sugar and insulin:

“In this meta-analysis of 16 prospective cohort studies comprising more than 890,000 individuals, we found that the presence of prediabetes at baseline was significantly associated with increased risks of cancer in the general population, particularly for liver cancer and stomach or colorectal cancer. The risks were increased when a lower FPG value of 5.6– 6.9 mmol/l [101-124 mg/dL] was used, according to the current ADA definition of IFG, as well as in participants with IGT. The results were consistent across cancer endpoints, age, study characteristics, follow-up duration and ethnicity.”

Much has been written here about the importance of glucose and insulin regulation for a wide range of conditions. The authors echo these themes in comments about likely mechanisms:

Hyperglycemia, advanced glycation end-products and oxidative damage

“First, chronic hyperglycaemia and its related conditions, such as chronic oxidative stress and the accumulation of advanced glycation end-products, may act as carcinogenic factors. It has been reported that diabetes is associated with an increased production of reactive oxygen species and greater oxidative damage to DNA. Recently, it has also been reported that the overall frequency of DNA damage and cytotoxicity correlates with the level of HbA1c in people with prediabetes.”

Insulin resistance

“Second, insulin resistance is a core defect responsible for the development of diabetes, and is established in individuals with prediabetes. The compensatory hyperinsulinaemia and increased level of bioavailable IGF 1 related to insulin resistance may promote the proliferation of cancer cells and may also relate to worsened cancer outcomes.”

Genetics

Third, genetic ‘interferences’ may also play an important role in the development of cancer in prediabetic individuals. A recent study has suggested that nuclear receptor coactivator 5 is a haploinsufficient tumour suppressor, and that a deficiency of nuclear receptor coactivator 5 increases susceptibility to both glucose intolerance and hepatocellular carcinoma, partially by increasing IL-6 expression.”

The public health implications of their results are enormous:

“These findings have important clinical and public health implications. In the US population aged ≥18 years, the age- adjusted prevalence of prediabetes increased from 29.2% in 1999–2002 to 36.2% in 2007–2010. Considering the high prevalence of prediabetes, as well as the robust and significant association between prediabetes and cancer dem- onstrated in our study, successful intervention in this large population could have a major public health impact. The ADA suggest that lifestyle intervention is the mainstay of treatment for prediabetes in the general population, and metformin is recommended for delaying progression to overt diabetes if individuals present with other related risk factors, such as a BMI ≥35 kg/m2, dyslipidaemia, hypertension, a family history of diabetes or an HbA1c >6% (42 mmol/mol)]. It should be noted that metformin is now considered as having some ‘protective’ anticancer properties. Notably, metformin mediates an approximately 30% reduction in the lifetime risk of cancer in diabetic patients. However, whether this is true in prediabetic individuals is not yet known. Long-term, large- scale studies of high-risk individuals, especially those with IGT or a combination of IGT and IFG, are urgently needed…”

Of course, functional practitioners have a number of resources besides metformin to help recover insulin sensitivity and restore healthier blood glucose regulation. The authors conclude:

“Overall, prediabetes was associated with an increased risk of cancer, especially liver, endometrial and stomach/colorectal cancer.’

Inflammation and diabetes

Diabetes Research and Clinical PracticeConsidering that chronic inflammation is a key common denominator in diabetes, prediabetes (metabolic syndrome) and cancer, it’s edifying to reflect on a paper published recently in Diabetes Research and Clinical Practice:

“It is recognized that a chronic low-grade inflammation and an activation of the immune system are involved in the pathogenesis of obesity-related insulin resistance and type 2 diabetes. Systemic inflammatory markers are risk factors for the development of type 2 diabetes and its macrovascular complications. Adipose tissue, liver, muscle and pancreas are themselves sites of inflammation in presence of obesity. An infiltration of macrophages and other immune cells is observed in these tissues associated with a cell population shift from an anti-inflammatory to a pro-inflammatory profile. These cells are crucial for the production of pro-inflammatory cytokines, which act in an autocrine and paracrine manner to interfere with insulin signaling in peripheral tissues or induce β-cell dysfunction and subsequent insulin deficiency. Particularly, the pro-inflammatory interleukin-1β is implicated in the pathogenesis of type 2 diabetes through the activation of the NLRP3 inflammasome. The objectives of this review are to expose recent data supporting the role of the immune system in the pathogenesis of insulin resistance and type 2 diabetes and to examine various mechanisms underlying this relationship. If type 2 diabetes is an inflammatory disease, anti-inflammatory therapies could have a place in prevention and treatment of type 2 diabetes.”

Nigella sativa, a true ‘wonder medicine’?

Nigella sativa flower and seedsNigella sativa, also known as black cumin, produces seeds with a mind-boggling wealth of medicinal virtues. For colleagues and others who may not be familiar with the abundance of scientific evidence for the use of Nigella sativa seed extract in clinical practice, this selection of citations serves as an introduction to its wide range of indications.

An illustrious history

Asian Pacific Journal of Tropical MedicineTraditional uses of Nigella sativa are surveyed in a paper published in the Asian Pacific Journal of Tropical Medicine:

Nigella sativa (N. sativa) (Family Ranunculaceae) is a widely used medicinal plant throughout the world. It is very popular in various traditional systems of medicine like Unani and Tibb, Ayurveda and Siddha. Seeds and oil have a long history of folklore usage in various systems of medicines and food. The seeds of N. sativa have been widely used in the treatment of different diseases and ailments. In Islamic literature, it is considered as one of the greatest forms of healing medicine. It has been recommended for using on regular basis in Tibb-e-Nabwi (Prophetic Medicine). It has been widely used as antihypertensive, liver tonics, diuretics, digestive, anti-diarrheal, appetite stimulant, analgesics, anti-bacterial and in skin disorders. Extensive studies on N. sativa have been carried out by various researchers and a wide spectrum of its pharmacological actions have been explored which may include antidiabetic, anticancer, immunomodulator, analgesic, antimicrobial, anti-inflammatory, spasmolytic, bronchodilator, hepato-protective, renal protective, gastro-protective, antioxidant properties, etc. Due to its miraculous power of healing, N. sativa has got the place among the top ranked evidence based herbal medicines. This is also revealed that most of the therapeutic properties of this plant are due to the presence of thymoquinone which is major bioactive component of the essential oil. The present review is an effort to provide a detailed survey of the literature on scientific researches of pharmacognostical characteristics, chemical composition and pharmacological activities of the seeds of this plant.”

Critical Reviews in Food Science and NutritionA paper published in the journal Critical Reviews in Food Science and Nutrition also suggests Nigella sativa’s wide scope of use:

“…It possesses a nutritional dense profile as its fixed oil (lipid fraction), is rich in unsaturated fatty acids while essential oil contains thymoquinone and carvacrol as antioxidants. N. sativa seeds also contain proteins, alkaloids (nigellicines and nigelledine), and saponins (α-hederin) in substantial amounts. Recent pharmacological investigations suggested its potential role, especially for the amelioration of oxidative stress through free radical scavenging activity, the induction of apoptosis to cure various cancer lines, the reduction of blood glucose, and the prevention of complications from diabetes. It regulates hematological and serological aspects and can be effective in dyslipidemia and respiratory disorders. Moreover, its immunopotentiating and immunomodulating role brings balance in the immune system. Evidence is available supporting the utilization of Nigella sativa and its bioactive components in a daily diet for health improvement. This review is intended to focus on the composition of Nigella sativa and to elaborate its possible therapeutic roles as a functional food to prevent an array of maladies.”

Anti-inflammatory activity

Molecular Biology ReportsChronic inflammation is a hallmark of most chronic degenerative diseases. A study published in Molecular Biology Reports demonstrates that Nigella sativa reduces inflammation triggered by LPS (lipopolysaccharide), of particular relevance for autoimmunity.

“Inflammation has an important role in many diseases such as cystic fibrosis, allergies and cancer. The free radicals produced during inflammation, can induce gene mutations and posttranslational modifications of cancer related proteins. Nigella sativa L. (N. sativa) is herbaceous plant and commonly used as a natural food. It has many pharmacological effects including antibacterial, antifungal, antitumor, analgesic, antipyretic activity. The aim of this study was to investigate the anti-inflammatuar and anti-oxidant activity of N. sativa in acute inflammation. Thus we used the experimental lipopolysaccharides (LPS)-induced model. Intraperitoneal LPS 1 mg/kg was administered to groups. N. sativa (500 mg/kg) and essential oil (5 ml/kg) were given orally to treatment groups, after 24-h of intraperitoneal LPS-injection. To determine the lung inflammation, 18F-fluoro-deoxy-d-glucose (0.8 ml/kg) was administrated under the anesthesia before the 1 h of PET-scanning. After the FDG-PET, samples were collected. Lung and liver18F-FDG-uptake was calculated. Serum AST, ALT, LDH and hcCRP levels were determined and liver, lung and erythrocyte SOD, MDA and CAT levels were measured. Liver and lung NO and DNA fragmentation levels were determined. MDA levels were decreased in treated inflammation groups whereas increased in untreated inflammation group. SOD and CAT activities in untreated inflammation group were significantly lower. According to the control group, increased AST and ALT levels were found in untreated inflammation group. 18F-FDG uptake of inflammation groups were increased when compare the control group… We conclude that, in LPS-induced inflammation, N. sativa have therapeutic and anti-oxidant effects.”

Immunomodulatory effects of Nigella sativa

Chinese Journal of Integrative MedicineA fascinating study in the Chinese Journal of Integrative Medicine offers evidence that Nigella sativa, beyond having simply an anti-inflammatory effect, is an immunomodulator that may help to restore healthier immune regulation:

“Cells isolated from human PBMCs which were treated with methanolic extract of NS for 48 h into two separate environments (PHA and non-PHA stimulated). Flow cytometry (for T helper/inducer cells and natural killer cells) and real time-polymerase chain reaction (PCR) assays for a few selected proinflammatory gene expressions were performed. Extracts from NS had an immunostimulating effect on non-PHA-stimulated proliferation of human PBMCs. In contrast, immunosuppressive activity was observed on PHA-stimulated proliferation of human PBMCs.”

Antimicrobial activity

BioMed Research InternationalNigella sativa has also shown good effect in the treatment of infections. A study recently published in Biomed Research International validates its antibacterial and antifungal properties:

“…major components in black cumin essential oils which were thymoquinone (37.6%) followed by p-cymene (31.2%), α-thujene (5.6%), thymohydroquinone (3.4%), and longifolene (2.0%), whereas the oleoresins extracted in different solvents contain linoleic acid as a major component….The essential oil showed up to 90% zone inhibition against Fusarium moniliforme in inverted petri plate method. Using agar well diffusion method for evaluating antibacterial activity, the essential oil was found to be highly effective against Gram-positive bacteria.”

The authors summarize their findings by concluding:

“The results obtained in antimicrobial investigations of black cumin oil and oleoresins were in good agreement with the previous reported work…Seeds of black cumin seem to possess magical properties and have been worked out extensively. This study revealed that black cumin essential oil and its oleoresins constitute a good alternative source of essential fatty acids compared with common vegetable oil. The present results showed that essential oil and oleoresins of black cumin exhibited higher antioxidant activity than synthetic antioxidants. These findings could be used to prepare multipurpose products for pharmaceutical applications and its usage as dietary source of antioxidant should be considered largely for alleviating and ameliorating diseases.”

World Journal of GastroenterologyPotent antiviral effects of Nigella sativa are in evidence in a study published in the World Journal of Gastroenterology on hepatitis C:

“Thirty patients with hepatitis C virus (HCV) infection, who were not eligible for IFN/ribavirin therapy, were included in the present study…Various parameters, including clinical parameters, complete blood count, liver function, renal function, plasma glucose, total antioxidant capacity (TAC), and polymerase chain reaction, were all assessed at baseline and at the end of the study. Clinical assessment included: hepato and/or splenomegaly, jaundice, palmar erythema, flapping tremors, spider naevi, lower-limb edema, and ascites. N. sativa was administered for three successive months at a dose of (450 mg three times daily). Clinical response and incidence of adverse drug reactions were assessed initially, periodically, and at the end of the study.”

The improvements noted were outstanding:

“N. sativa administration significantly improved HCV viral load. After N. sativa administration, the following laboratory parameters improved: total protein, albumin, red blood cell count, and platelet count. Fasting blood glucose and postprandial blood glucose were significantly decreased in both diabetic and non-diabetic HCV patients. Patients with lower-limb edema decreased significantly from baseline compared with after treatment. Adverse drug reactions were unremarkable except for a few cases of epigastric pain and hypoglycemia that did not affect patient compliance.”

Clinicians involved in case management of HCV should note their conclusion:

N. sativa administration in patients with HCV was tolerable, safe, decreased viral load, and improved oxidative stress, clinical condition and glycemic control in diabetic patients.”

 Amelioration of metabolic disorders

Plant Foods for Human NutritionNigella sativa possesses remarkable properties that improve metabolic disorders ranging including insulin resistance and diabetes, obesity, and liver fibrosis. From a paper in Plant Foods for Human Nutrition:

“Obesity is closely associated with increased incidence of cardiovascular diseases, cancer, insulin resistance, and immune dysfunction, and thus obesity-mitigation strategies should take into account these secondary pathologies in addition to promoting weight loss. Recent studies indicate that black cumin (Nigella sativa) has cardio-protective, anti-cancer, anti-diabetic, antioxidant, and immune-modulatory properties.”

 Diabetes

Evidence-Based Complementary and Alternative MedicineEvidence for its benefit in diabetes is offered in a study published in Evidence-Based Complementary and Alternative Medicine:

“The main objective of this instant study was to explore the antidiabetic potential of Nigella sativa fixed oil (NSFO) and essential oil (NSEO). Three experimental groups of rats received diets during the entire study duration, that is, D1 (control), D2 (NSFO: 4.0%), and D3 (NSEO: 0.30%). Experimental diets (NSFO & NSEO) modulated the lipid profile, while decreasing the antioxidant damage. However, production of free radicals, that is, MDA, and conjugated dienes increased by 59.00 and 33.63%, respectively, in control. On the contrary, NSFO and NSEO reduced the MDA levels by 11.54 and 26.86% and the conjugated dienes levels by 32.53 and 38.39%, respectively. N. sativa oils improved the health and showed some promising anti-diabetic results.”

BMC Complementary & Alternative MedicineAnother study on Nigella sativa and diabetes was recently published in BMC Complementary and Alternative Medicine.

Nigella sativa fixed (NSFO) and essential (NSEO) oils have been used to treat diabetes mellitus and its complications. Present study was undertaken to explore and validate these folkloric uses…Sprague dawley rats having streptozotocin (STZ) induced diabetes mellitus were used to assess the role of NSFO and NSEO in the management of diabetes complications.”

Of note is its ability to increase levels of glutathione:

“The results indicated that STZ decreased the glutathione contents (25.72%), while NSFO and NSEO increased the trait significantly. Experimental diets increased the tocopherol contents and enhanced the expression of hepatic enzymes. Correlation matrix further indicated that antioxidant potential is positively associated responsible for the modulation of hepatic enzymes and the decrease of the nitric oxide production thus controlling the diabetes complications.”

Nigella sativa lowers cholesterol

Advanced Pharmaceutical BulletinCholesterol along with blood glucose was lowered in a study on Nigella sative for metabolic syndrome in menopausal women published in the Advanced Pharmaceutical Bulletin:

“Thirty subjects who were menopausal women within the age limit of 45-60 were participated in this study and randomly allotted into two experimental groups. The treatment group was orally administered with N. sativa seeds powder in the form of capsules at a dose of 1g per day after breakfast for period of two months and compared to control group given placebo…significant improvement was observed in total cholesterol (TC), triglycerides (TG), low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C), and blood glucose…These results suggested that treatment with N. sativa exert a protective effect by improving lipid profile and blood glucose which are in higher risk to be elevated during menopausal period.”

Journal of Translational MedicineImprovements in hypercholesterolemia in menopause were also documented in a study recently published in the Journal of Translational Medicine:

“In this study, Nigella sativa was evaluated for its hypolipidemic effects among menopausal women. In a randomised trial, hyperlipidemic menopausal women were assigned to treatment (n = 19) or placebo groups (n = 18), and given N. sativa or placebo for two months after their informed consents were sought. At baseline, blood samples were taken and at one month intervals thereafter until one month after the end of the study…The results showed that N. sativa significantly improved lipid profiles of menopausal women (decreased total cholesterol, low density lipoprotein cholesterol and triglyceride, and increased high density lipoprotein cholesterol) more than the placebo treatment over 2 months of intervention.”

These benefits persisted for a month after treatment with Nigella sativa was discontinued:

One month after cessation of treatment, the lipid profiles in the N. sativa-treated group tended to change towards the pretreatment levels.”

The authors conclude:

“N. sativa is thought to have multiple mechanisms of action and is cost-effective. Therefore, it could be used by menopausal women to remedy hypercholesterolemia, with likely more benefits than with single pharmacological agents that may cause side effects. The use of N. sativa as an alternative therapy for hypercholesterolemia could have profound impact on the management of CVD among menopausal women especially in countries where it is readily available.”

International Journal of Preventive MedicineAnd a study in the International Journal of Preventive Medicine documented improvements in lipid metabolism and oxygen utilization:

“In this randomized, double-blind, controlled trial…20 sedentary overweight females were divided into two groups and assigned to N. sativa supplementation (N. sativa capsules) or a placebo for the 8 weeks, both groups participated in an aerobic training program (3 times/week)…. Blood lipids and VO2 max were determined at baseline and at the end of 8 weeks…N. sativa supplementation lowered total cholesterol (TC), triglyceride, low-density lipoprotein (LDL) and body mass index and increased high density lipoprotein (HDL) and VO2 max.”

It’s worth noting that the diet of the study subjects remained the same:

Since we asked all subjects not to change their usual daily diet, it seems that this changes may be due to the result of consuming black seeds and regular aerobic training.”

Interestingly in regard to lowering cholesterol:

“The hypotriglyceridemic effect of N. sativa is possibly due to its choleretic activity. The choleretic function of N. sativa is either by reducing the synthesis of cholesterol by hepatocytes or by decreasing its fractional reabsorption from the small intestine.”

Nigella sativa’s thymoquinone ameliorates liver fibrosis

International ImmunopharmacologyWith the proliferation of NAFLD and NASH medicines that sustainably alleviate hepatic fibrosis are in urgent need. A study published in International Immunopharmacology offers evidence that thymoquinone, a principal compound in Nigella sativa, has potent hepatic anti-fibrotic effects:

Thymoquinone (TQ) is the major active compound derived from the medicinal Nigella sativa. In the present study, we investigated the anti-fibrotic mechanism of TQ in lipopolysaccharide (LPS)-activated rat hepatic stellate cells line, T-HSC/Cl-6. T-HSC/Cl-6 cells were treated with TQ (3.125, 6.25 and 12.5 μM) prior to LPS (1 μg/ml). Our data demonstrated that TQ effectively decreased activated T-HSC/Cl-6 cell viability. TQ significantly attenuated the expression of CD14 and Toll-like receptor 4 (TLR4). TQ also significantly inhibited phosphatidylinositol 3-kinase (PI3K) and serine/threonine kinase-protein kinase B (Akt) phosphorylation. The expression of α-SMA and collagen-I were significantly decreased by TQ. Furthermore, TQ decreased X linked inhibitor of apoptosis (XIAP) and cellular FLIP (c-FLIPL) expression, which are related with the regulation of apoptosis. Furthermore, TQ significantly increased the survival against LPS challenge in d-galactosamine (d-GlaN)-sensitized mice, and decreased the levels of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST), which were in line with in vitro results. Our data demonstrated that TQ attenuates liver fibrosis partially via blocking TLR4 expression and PI3K phosphorylation on the activated HSCs. Therefore, TQ may be a potential candidate for the therapy of hepatic fibrosis.

A follow-up study published recently in the same journal added more evidence to Nigella sativa’s benefits for hepatic fibrosis:Hepatic fibrosis attenuated by thymoquinone

“The current study was conducted to investigate the anti-fibrotic effect and its possible underlying mechanisms of thymoquinone (TQ) against hepatic fibrosis in vivo. TQ is the major active compound derived from the medicinal Nigella sativa. Liver fibrosis was induced in male Kunming mice by intraperitoneal injections of thioacetamide (TAA, 200 mg/kg). Mice were treated concurrently with TAA alone or TAA plus TQ (20 mg/kg or 40 mg/kg) given daily by oral gavage. Our data demonstrated that TQ treatment obviously reversed liver tissue damage compared with TAA alone group, characterized by less inflammatory infiltration and accumulation of extracellular matrix (ECM) proteins. TQ significantly attenuated TAA-induced liver fibrosis, accompanied by reduced protein and mRNA expression of α-smooth muscle actin (α-SMA), collagen-І and tissue inhibitor of metalloproteinase-1 (TIMP-1). TQ downregulated the expression of toll-like receptor 4 (TLR4) and remarkably decreased proinflammatory cytokine levels as well. TQ also significantly inhibited phosphatidylinositol 3-kinase (PI3K) phosphorylation. Furthermore, TQ enhanced the phosphorylation adenosine monophosphate-activated protein kinase (AMPK) and liver kinase B (LKB)-1. In conclusion, TQ may reduce ECM accumulation, and it may be at least regulated by phosphorylation of AMPK signaling pathways, suggesting that TQ may be a potential candidate for the therapy of hepatic fibrosis.

 Protection against diabetic kidney damage

Ultrastructural PathologyThymoquinone in Nigella sativa also reduced experimentally induced kidney damage in models of diabetes as reported in a study published in Ultrastructural Pathology:

“Diabetic rats exhibited morphological changes in both renal glomeruli and tubules with immunohistochemical expression of the mesenchymal markers Fsp1, desmin, and MMP-17 and disappearance of the epithelial marker ZO-1 largely in the glomeruli of diabetic kidneys. Treatment with TQ significantly attenuated renal morphological and immunohistochemical changes in STZ-induced diabetic ratsThymoquinone has protective effects on experimental diabetic nephropathy. Both mesenchymal and epithelial markers serve as excellent predictors of early kidney damage and indicators of TQ responsiveness in STZ-induced diabetic nephropathy.”

Hypertension and Oxidative Stress

Regarding the anti-hypertensive effects of Nigella sativa, from a paperEvidence-Based Complementary and Alternative Medicine in Evidence-Based Complementary & Alternative Medicine:

Excessive production of reactive oxygen species reduces nitric oxide bioavailability leading to an endothelial dysfunction and a subsequent increase in total peripheral resistance…Nigella sativa (NS) and its active constituents have been documented to exhibit antioxidant, hypotensive, calcium channel blockade and diuretic properties which may contribute to reduce blood pressure. This suggests a potential role of NS in the management of hypertension…”

Protection Against Heart Damage

Pakistan Journal of Pharmaceutical SciencesNot surprisingly, thymoquinone in Nigella sativa appears to exert protective effects against heart damage associated with coronary insufficiency and stress as documented by a study in the Pakistan Journal of Pharmaceutical Sciences. Here again the beneficial effects include support for glutathione:

“Myocardial injury constitutes a major cause of morbidity and mortality in humans. Present study aimed to investigate protective role of thymoquinone, which is an active principle of Nigella sativa (N. sativa) seed (Commonly called as black seed), in isoproterenol induced myocardial injury, a classical example of excess catecholamines related coronary insufficiency and stress cardiomyopathy. Thymoquinone, in olive oil, was administered orally (12.5, 25 and 50mg/kg) to three groups of Wistar albino rats for 7 days, while two control groups were given plain olive oil. Thereafter, thymoquinone receiving groups and one control group were injected, subcutaneously, with isoproterenol (125mg/kg) for 2 days. Myocardial injury was assessed by biochemical markers (plasma LDH, TBARS, GR & SOD and myocardial GSH/GSSG ratio) and cardiac histopathology. Plasma LDH, TBARS and GR increased in control groups receiving isoproterenol, while there was a dose related decrease in these markers in thymoquinone treated groups, down to levels in controls given olive oil only. Decrease in plasma SOD and myocardial GSH/GSSG ratio and histological changes produced with isoproternol were also reversed in thymoquinone treated rats. Results of our study revealed that thymoquinone protects the heart from injury induced by isoproterenol.”

Anti-cancer effects of Nigella sativa

Drug Discovery TodayThere is a wealth of evidence supporting the use Nigella sativa and its active compound thymoquinone as an adjunctive treatment in numerous malignancies as noted in a paper published earlier this year in Drug Discovery Today:

“Thymoquinone (TQ), the main active constituent of black seed essential oil, exhibits promising effects against inflammatory diseases and cancer. TQ, modulates signaling pathways that are key to cancer progression, and enhances the anticancer potential of clinical drugs while reducing their toxic side effects. Considering that TQ was isolated 50 years ago, this review focuses on TQ’s chemical and pharmacological properties and the latest advances in TQ analog design and nanoformulation. We discuss our current state of knowledge of TQ’s adjuvant potential and in vivo antitumor activity and highlight its ability to modulate the hallmarks of cancer.

  • This year marks 50 years since thymoquinone was isolated from black seed.
  • Thymoquinone has had a long history of battling cancer in vitro and in vivo.
  • Thymoquinone modulates nine of the ten hallmarks of cancer.”

American Journal of Chinese MedicineA paper in the American Journal of Chinese Medicine reviews Nigella sativa’s anticancer activities:

“…quite a few pharmacological effects of N. sativa seed, its oil, various extracts and active components have been identified to include immune stimulation, anti-inflammation, hypoglycemic, antihypertensive, antiasthmatic, antimicrobial, antiparasitic, antioxidant and anticancer effects…A literature search has revealed that a lot more studies have been recently carried out related to the anticancer activities of N. sativa and some of its active compounds, such as thymoquinone and alpha-hederin. Acute and chronic toxicity studies have recently confirmed the safety of N. sativa oil and its most abundant active component, thymoquinone, particularly when given orally. The present work is aimed at summarizing the extremely valuable work done by various investigators on the effects of N. sativa seed, its extracts and active principles against cancer. Those related to the underlying mechanism of action, derivatives of thymoquinone, nano thymoquinone and combinations of thymoquinone with the currently used cytotoxic drugs are of particular interest.”

Thymoquinone mechanisms of actionA paper in the African Journal of Traditional, Complementary and Alternative Medicines describes its activity against a number of malignancies and the molecular mechanisms involved:

“Nigella sativa has been used as traditional medicine for centuries. The crude oil and thymoquinone (TQ) extracted from its seeds and oil are effective against many diseases like cancer, cardiovascular complications, diabetes, asthma, kidney disease etc. It is effective against cancer in blood system, lung, kidney, liver, prostate, breast, cervix, skin with much safety. The molecular mechanisms behind its anticancer role is still not clearly understood, however, some studies showed that TQ has antioxidant role and improves body’s defense system, induces apoptosis and controls Akt pathway. Although the anti-cancer activity of N. sativa components was recognized thousands of years ago but proper scientific research with this important traditional medicine is a history of last 2∼3 decades…In this article, we have summarized the actions of TQ and crude oil of N. sativa against different cancers with their molecular mechanisms.”

Pharmacognosy ReviewsA review article in Pharmacognosy Review notes the anti-cancer potential implied by numerous investigations:

“Thymoquinone (TQ) is the bioactive phytochemical constituent of the seeds oil of Nigella sativa. In vitro and in vivo research has thoroughly investigated the anticancer effects of TQ against several cancer cell lines and animal models. As a result, a considerable amount of information has been generated from research thus providing a better understanding of the anti-proliferating activity of this compound. Therefore, it is appropriate that TQ should move from testing on the bench to clinical experiments. The purpose of this review is to highlight the potential of TQ as an anticancer agent and the chances of this compound in the clinical treatment of cancer, with special attention on breast cancer treatment.”

Evidence-Based Complementary and Alternative MedicineA paper in Evidence-Based Complementary and Alternative Medicine outlines mechanisms by which thymoquinone in Nigella sativa can act to prevent cancer:

Earlier studies have shown that N. sativa and its constituent thymoquinone (TQ) have important roles in the prevention and treatment of cancer by modulating cell signaling pathways. In this review, we summarize the role of N. sativa and its constituents TQ in the prevention of cancer through the activation or inactivation of molecular cell signaling pathways.”

Upregulation of tumor suppressor gene and inhibition of VEGF, Akt/PI3K pathways:

Upregulation of tumor suppresor geneThymoquinone role in prevention of cancer via modulation of phase I and phase II enzymes:

Thymoquinone's role in cancer prevention

Osteosarcoma, angiogenesis and NF-κB

Oncology ReportsEvidence for thymoquinone’s benefit in osteosarcoma through inhibition of tumor angiogenesis and tumor growth by suppressing NF-κB is offered by a study published in Oncology Reports:

“Recent studies reported that thymoquinone exhibited inhibitory effects on the cell proliferation of several cancer cell lines. This study was performed to investigate the antitumor and anti-angiogenic effects of thymoquinone on osteosarcoma in vitro and in vivo. Our results showed that thymoquinone induced a higher percentage of growth inhibition and apoptosis in the human osteosarcoma cell line SaOS-2 compared to that of control, and thymoquinone significantly blocked human umbilical vein endothelial cell (HUVEC) tube formation in a dose-dependent manner. To investigate the possible mechanisms involved in these events, we performed electrophoretic mobility shift assay (EMSA) and western blot analysis, and found that thymoquinone significantly downregulated NF-κB DNA-binding activity, XIAP, survivin and VEGF in SaOS-2 cells. Moreover, the expression of cleaved caspase-3 and Smac were upregulated in SaOS-2 cells after treatment with thymoquinone. In addition to these in vitro results, we also found that thymoquinone inhibits tumor angiogenesis and tumor growth through suppressing NF-κB and its regulated molecules. Collectively, our results demonstrate that thymoquinone effectively inhibits tumor growth and angiogenesis both in vitro and in vivo. Moreover, inhibition of NF-κB and downstream effector molecules is a possible underlying mechanism of the antitumor and anti-angiogenic activity of thymoquinone in osteosarcoma.”

Cytotoxic prooxidant effects of thymoquinone in copper rich malignant tissues

Cell Death & DiseaseUsing prostate cancer cells, a fascinating study published in Cell Death & Disease demonstrates that thymoquinone has a beneficial prooxidant cytoxic effect in copper-rich malignant tissue:

“Thymoquinone (TQ) is the major bioactive constituent of volatile oil of Nigella sativa and has been shown to exert various pharmacological properties, such as anti-inflammatory, cardiovascular, analgesic, anti-neoplastic, anticancer and chemopreventive…TQ is a known antioxidant at lower concentrations and most of the studies elucidating the mechanism have centered on the antioxidant property. However, recent publications have shown that TQ may act as a prooxidant at higher Nigella sativa flower 2concentrations. It is well known that plant-derived antioxidants can switch to prooxidants even at low concentrations in the presence of transition metal ions such as copper. It is well established that tissue, cellular and serum copper levels are considerably elevated in various malignancies. Copper is an important metal ion present in the chromatin and is closely associated with DNA bases, particularly guanine. Using human peripheral lymphocytes and comet assay, we first show that TQ is able to cause oxidative cellular DNA breakage. Such a DNA breakage can be inhibited by copper-chelating agents, neocuproine and bathocuproine, and scavengers of reactive oxygen species. Further, it is seen that TQ targets cellular copper in prostate cancer cell lines leading to a prooxidant cell death.”

Interestingly…

“We believe that such a prooxidant cytotoxic mechanism better explains the anticancer activity of plant-derived antioxidants.”

Inhibition of cell proliferation in liver cancer

Toxicology LettersMarked inhibition of tumor multiplicity in hepatocellular carcinoma was shown in a study published in Toxicology Letters:

“…agents that inhibit cell proliferation and restrain hepatic tumorigenesis through cell cycle regulation have a beneficial effect in the treatment of hepatocellular carcinogenesis. The present study was aimed to investigate the efficacy of thymoquinone (TQ), an active compound derived from the medicinal plant Nigella sativa, on N-nitrosodiethylamine (NDEA) [0.01% in drinking water for 16 weeks]-induced hepatocarcinogenesis in experimental rats. After experimental period, the hepatic nodules, liver injury markers and tumor markers levels were substantially increased in NDEA induced liver tumors in rats. However, TQ (20 mg/kg body weight) treatment greatly reduced liver injury markers and decreased tumor markers and prevented hepatic nodule formation and reduced tumor multiplicity in NDEA induced hepatic cancer bearing rats and this was evident from argyrophilic nucleolar organizer region (AgNORs) staining. Moreover…TQ significantly reduced the detrimental alterations by abrogating cell proliferation, which strongly induced G1/S arrest in cell cycle transition. In conclusion, our results suggest that TQ has a potent anti proliferative activity by regulating the G1/S phase cell cycle transition and exhibits a beneficial role in the treatment of HCC.”

Thymoquinone induces glioblastoma cell death

PLOS ONEA fascinating study in PLoS One demonstrates that thymoquinone is a rare agent that can inhibit autophagy (the cellular ‘housecleaning’ process by which degraded cellular components are removed) to promote malignant cell death in the brain cancer gliosblastoma:

“Glioblastoma is the most aggressive and common type of malignant brain tumor in humans, with a median survival of 15 months. There is a great need for more therapies for the treatment of glioblastoma…TQ has anti-oxidant, anti-inflammatory and anti-neoplastic actions with selective cytotoxicity for human cancer cells compared to normal cells. Here, we show that TQ selectively inhibits the clonogenicity of glioblastoma cells as compared to normal human astrocytes. Also, glioblastoma cell proliferation could be impaired by chloroquine, an autophagy inhibitor, suggesting that glioblastoma cells may be dependent on the autophagic pathway for survival…TQ also caused an accumulation of the LC3-associated protein p62, confirming the inhibition of autophagy. Furthermore, the levels of Beclin-1 protein expression were unchanged, indicating that TQ interferes with a later stage of autophagy. Finally, treatment with TQ induces lysosome membrane permeabilization…which mediates caspase-independent cell death… TQ induced apoptosis…”

Inhibition of autophagy by thymoquinoneThe authors note an important difference between the action of thymoquinone and other cytotoxic therapies:

Ionizing radiation and temozolomide have both been shown to increase a cytoprotective autophagy response in glioblastoma cells, leading to resistant tumors. In addition, many other chemotherapeutics, such as rapamycin, tamoxifen, and etoposide, induce a protective autophagic response in cancer cells. Therefore, inhibitors of autophagy, both alone and in combination with standard therapies, may provide a viable and promising new strategy in cancer treatment…To the best of our knowledge, this report represents the first finding of TQ as an autophagy inhibitor, and provides a platform for which to extend studies in the treatment of glioblastoma with TQ.”

The authors conclude:

“Inhibition of autophagy is an exciting and emerging strategy in cancer therapy. In this vein, our results describe a novel mechanism of action for TQ as an autophagy inhibitor selectively targeting glioblastoma cells.

Nigella sativa induces apoptosis in cervical cancer

Natural Product CommunicationsAccording to a study published in Natural Product Communications, Nigella sativa inhibits proliferation of cervical cancer cells by inducing apoptosis:

“Nigella sativa (NS) showed an 88.3% inhibition of proliferation of SiHa human cervical cancer cells at a concentration of 125 microL/mL methanolic extract at 24 h, and an IC50 value 93.2 microL/mL. NS exposure increased the expression of caspase-3, -8 and -9 several-fold. The analysis of apoptosis by Dead End terminal transferase-mediated dUTP-digoxigenin end labeling (TUNEL) assay was used to further confirm that NS induced apoptosis. Thus, NS was concluded to induce apoptosis in SiHa cell through both p53 and caspases activation. NS could potentially be an alternative source of medicine for cervical cancer therapy.”

Suppression of melanoma metastasis by inhibition of the NLRP3 inflammasome

Toxicology and Applied PharmacologyIn an exciting study published in Toxicology and Applied Pharmacology that has implications for a wide range of conditions, investigators report suppression of metastasis in melanoma inhibiting the proinflammatory activity of the NLRP3 inflammasome:

“The inflammasome is a multi-protein complex which when activated regulates caspase-1 activation and IL-1β and IL-18 secretion. The NLRP3 (NACHT, LRR, and pyrin domain-containing protein 3) inflammasome is constitutively assembled and activated in human melanoma cells. We have examined the inhibitory effect of thymoquinone (2-isopropyl-5-methylbenzo-1,4-quinone), a major ingredient of black seed obtained from the plant Nigella sativa on metastatic human (A375) and mouse (B16F10) melanoma cell lines. We have assessed whether thymoquinone inhibits metastasis of melanoma cells by targeting NLRP3 subunit of inflammasomes. Using an in vitro cell migration assay, we found that thymoquinone inhibited the migration of both human and mouse melanoma cells…The inhibition of migration of melanoma cells by thymoquinone was accompanied by a decrease in expression of NLRP3 inflammasome resulting in decrease in proteolytic cleavage of caspase-1. Inactivation of caspase-1 by thymoquinone resulted in inhibition of IL-1β and IL-18. Treatment of mouse melanoma cells with thymoquinone also inhibited NF-κB activity. Furthermore, inhibition of reactive oxygen species (ROS) by thymoquinone resulted in partial inactivation of NLRP3 inflammasome. Thus, thymoquinone exerts its inhibitory effect on migration of human and mouse melanoma cells by inhibition of NLRP3 inflammasome. Thus, our results indicate that thymoquinone can be a potential immunotherapeutic agent not only as an adjuvant therapy for melanoma, but also, in the control and prevention of metastatic melanoma.”

Readers will recall that activation of the inflammasome is a mechanism shared by many autoimmune and malignant disorders.

Nigella sativa attenuates iNOS pathway inflammation in liver cancer

Environmental Health and Preventive MedicineBecause iNOS activation of inflammation is a key process in a multitude of inflammatory disorders including a host of autoimmune diseases, a study published in Environmental Health and Preventative Medicine showing value in hepatocellular carcinoma is of is of particular importance:

“Nitric oxide (NO) and inducible nitric oxide synthase enzyme (iNOS) have been implicated in various tumors….Nigella sativa (NS) has been shown to have specific health benefits. The aim of this study was to investigate the in vivo modulation of the iNOS pathway by NS ethanolic extract (NSEE) and the implications of this effect as an antitumor therapeutic approach against diethylnitrosamine (DENA)-induced hepatocarcinogenesis…Serum AFP, NO, TNF-α, and IL-6 levels and iNOS enzyme activity were significantly increased in rats treated with DENA. Significant up-regulation of liver iNOS mRNA and protein expression was also observed. Subsequent treatment with NSEE significantly reversed these effects and improved the histopathological changes in malignant liver tissue which appeared after treatment with DENA, without any toxic effect when given alone.”

This data inspired the authors to conclude:

“These results provide evidence that attenuation of the iNOS pathway and suppression of the inflammatory response mediated by TNF-α, and IL-6 could be implicated in the antitumor effect of NSEE. As such, our findings hold great promise for the utilization of NS as an effective natural therapeutic agent in the treatment of hepatocarcinogenesis.”

Cytotoxic effect against lung cancer

Asian Pacific Journal of Cancer PreventionAuthors of a study just published in the Asian Pacific Journal of Cancer Prevention report that Nigella sativa seed extract significantly reduces the viability of lung cancer cells:

Nigella sativa (N sativa), commonly known as black seed, has been used in traditional medicine to treat many diseases. The antioxidant, anti-inflammatory, and antibacterial activities of N sativa extracts are well known. Therefore, the present study was designed to investigate the anticancer activity of seed extract (NSE) and seed oil (NSO) of N sativa against a human lung cancer cell line…The results showed NSE and NSO significantly reduce the cell viability and alter the cellular morphology of A-549 cells in a concentration dependent manner. The percent cell viability was recorded as 75%, 50%, and 26% at 0.25, 0.5, and 1 mg/ml of NSE by MTT assay and 73%, 48%, and 23% at 0.25, 0.5, and 1 mg/ml of NSE by NRU assay. Exposure to NSO concentrations of 0.1 mg/ml and above for 24 h was also found to be cytotoxic. The decrease in cell viability at 0.1, 0.25, 0.5, and 1 mg/ml of NSO was recorded to be 89%, 52%, 41%, and 13% by MTT assay and 85%, 52%, 38%, and 11% by NRU assay, respectively. A-549 cells exposed to 0.25, 0.5 and 1 mg/ml of NSE and NSO lost their typical morphology and appeared smaller in size. The data revealed that the treatment of seed extract (NSE) and seed oil (NSO) of Nigella sativa significantly reduce viability of human lung cancer cells.

Nigella sativa inhibits breast cancer

PLOS ONEEvidence is mounting for the use of Nigella sativa against breast cancer. Similar to the prooxidant effect described above, a study published in PLoS One describes how thymoquinone inhibits tumor growth and induces apoptosis in breast cancer cells through p38 phosphorylation and ROS production:

“Due to narrow therapeutic window of cancer therapeutic agents and the development of resistance against these agents, there is a need to discover novel agents to treat breast cancer. The antitumor activities of thymoquinone (TQ), a compound isolated from Nigella sativa oil, were investigated in breast carcinoma in vitro and in vivo. Cell responses after TQ treatment were assessed by using different assays including MTT assay, annexin V-propidium iodide staining, Mitosox staining and Western blot. The antitumor effect was studied by breast tumor xenograft mouse model, and the tumor tissues were examined by histology and immunohistochemistry. The level of anti-oxidant enzymes/molecules in mouse liver tissues was measured by commercial kits. Here, we show that TQ induced p38 phosphorylation and ROS production in breast cancer cells. These inductions were found to be responsible for TQ’s anti-proliferative and pro-apoptotic effects. Moreover, TQ-induced ROS production regulated p38 phosphorylation but not vice versa. TQ treatment was found to suppress the tumor growth and this effect was further enhanced by combination with doxorubicin. TQ also inhibited the protein expression of anti-apoptotic genes, such as XIAP, survivin, Bcl-xL and Bcl-2, in breast cancer cells and breast tumor xenograft. Reduced Ki67 and increased TUNEL staining were observed in TQ-treated tumors. TQ was also found to increase the level of catalase, superoxide dismutase and glutathione in mouse liver tissues.”

Again we see increases in the profoundly important glutatione under the influence of thymoquinone. Note also that the antitumor effect of the conventional chemotherapeutic agent was enhanced.

“In conclusion, our study provides evidence for the mechanism of action of TQ in suppressing human breast carcinoma in both in vitro and in vivo models. We demonstrated that the anti-proliferative and pro-apoptotic effects of TQ are mediated through its induction effect on p38 and ROS signaling. Our results also indicate the anti-tumor effects of TQ in breast tumor xenograft mice and its ability to potentiate the antitumor effect of doxorubicin. TQ serves as a promising anticancer agent and further studies may provide important leads for its clinical application.”

Journal of Medicinal FoodA study published in the Journal of Medicinal Food also reports proapoptotic and antimetastatic effects of Nigella sativa for breast cancer:

“This study investigated the apoptotic, antimetastatic, and anticancer activities of supercritical carbon dioxide (SC-CO2) extracts of the seeds of N. sativa Linn. against estrogen-dependent human breast cancer cells (MCF-7)….Of the 12 extracts, 1 extract (A3) that was prepared at 60°C and 2500 psi (~17.24 MPa) showed selective antiproliferative activity against MCF-7 cells with an IC50 of 53.34±2.15 μg/mL. Induction of apoptosis was confirmed by evaluating caspases activities and observing the cells under a scanning electron microscope. In vitro antimetastatic properties of A3 were investigated by colony formation, cell migration, and cell invasion assays. The elevated levels of caspases in A3 treated MCF-7 cells suggest that A3 is proapoptotic. Further nuclear condensation and fragmentation studies confirmed that A3 induces cytotoxicity through the apoptosis pathway. A3 also demonstrated remarkable inhibition in migration and invasion assays of MCF-7 cells at subcytotoxic concentrations. Thus, this study highlights the therapeutic potentials of SC-CO2 extract of N. sativa in targeting breast cancer.”

Pharmacognosy ResearchAnd authors of a study published in Pharmacognosy Research also report activity of thymoquinone against breast cancer:

“The study addressed the anti-cancer efficiency of long-term in vitro treatment with thymoquinone towards human breast cancer cell lines MCF-7...The 50% inhibitory concentration (IC50) value determined using the proliferation assay was 25 μM thymoquinone. Late apoptotic cell percentage increased rapidly when treatment duration was increased to 24 h with 25 and 100 μM thymoquinone. Further analysis using cell cycle assay showed thymoquinone inhibition of breast cancer cell proliferation at minimal dose 25 μM and led to S phase arrest significantly at 72 h treatment. It was also noted elevation sub-G1 peak following treatment with 25 μM thymoquinone for 12 h. Increase in thymoquinone to 50 μM caused G2 phase arrest at each time-point studied…In general thymoquinone showed sustained inhibition of breast cancer cell proliferation with long-term treatment. Specificity of phase arrest was determined by thymoquinone dose.”

Asian Pacific Journal of Cancer PreventionAntiproliferative effects against breast cancer cells were also shown in a study published in the Asian Pacific Journal of Cancer Prevention:

“Our data showed that Nigella sativa extracts significantly inhibited human breast cancer MDA-MB-231 cell proliferation at doses of 2.5-5 μg/mL. Apoptotic induction in MDA-MB-231 cells was observed in a dose-dependent manner after exposure to Nigella sativa extracts for 48 h. Real time PCR and flow cytometry analyses suggested that Nigella sativa extracts possess the ability to suppress the proliferation of human breast cancer cells through induction of apoptosis.”

Nigella sativa protects against liver damage caused by tamoxifen

Canadian Journal of Physiology and PharmacologyProtection against the harmful toxic effects of chemotherapy is a critical component of cancer case management. A welcome study published in the Canadian Journal of Physiology and Pharmacology shows that thymoquinone from Nigella sativa protects against the hepatotoxicity of tamoxifen:

“One of the major reasons for terminating a clinical trial is the liver toxicity induced by chemotherapy. Tamoxifen (TAM) is an anti-estrogen used in the treatment and prevention of hormone-dependent breast cancer. Tamoxifen therapy may cause hepatic injury. The seeds of Nigella sativa, which contain the active ingredient thymoquinone (TQ), have been used in folk medicine for diverse ailments. TQ is reported to possess anticancer and hepatoprotective effects. In this study, the protective effects of TQ against TAM-induced hepatotoxicity in female rats were evaluated. Four groups of rats were used: control; TAM; TQ; TAM+TQ. TAM (45 mg·(kg body mass)(-1)·day(-1), by intraperitoneal injection (i.p.), for 10 consecutive days) resulted in elevated serum levels of alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, lactate dehydrogenase, total bilirubin, and gamma glutamyl transferase, as well as depletion of reduced glutathione in the liver and accumulation of lipid peroxides. Also, TAM treatment inhibited the hepatic activity of superoxide dismutase. Further, it raised the levels of tumor necrosis factor alpha in the liver and induced histopathological changes. Pretreatment with TQ (50 mg·(kg body mass)(-1)·day(-1); orally, for 20 consecutive days, starting 10 days before TAM injection) significantly prevented the elevation in serum activity of the assessed enzymes. TQ significantly inhibited TAM-induced hepatic GSH depletion and LPO accumulation. Consistently, TQ normalized the activity of SOD, inhibited the rise in TNF-α and ameliorated the histopathological changes. In conclusion, TQ protects against TAM-induced hepatotoxicity.”

Again we see beneficial effects on glutatione metabolism.

Protection against kidney toxicity of cisplatin

Iranian Journal of Kidney DiseasesWhile on the topic of protection unwanted against damage done by cytotoxic chemotherapy, we can appreciate a study published in the Iranian Journal of Kidney Diseases reporting evidence that Nigella sativa offers some protection against the nephrotoxic effects of cisplatin:

“Thirty rats were divided into 3 groups to receive distilled water (control group), cisplatin (3 mg/kg per body weight for 3 days), and cisplatin and alcoholic extract of NS (100 mg/kg per body weight). Biochemical and histopathologic parameters were compared between the three groups on days 14 and 42 of the study…Cisplatin-induced nephrotoxicity was confirmed in our study…Histology of the kidneys exposed to cisplatin showed significant kidney injury, but the rats treated with NS showed a relatively well-preserved architectureNigella sativa seeds had nonsignificant effects on biochemical parameters, although the histopathologic properties of the kidneys relatively recovered after NS use.”

Nigella sativa benefits for the brain, mood and cognition

Journal of EthnopharmacologyConsidering the immune-regulating and anti-inflammatory virtues of Nigella sativa it stands to reason that there would be benefits for the brain. A study published in the Journal of Ethnopharmacology reports that it helps stabilize mood, reduce anxiety and cognition in adolescent males.

“Previous studies conducted on animals linked consumption of Nigella sativa L. seeds (NS) to decreased anxiety and improved memory. The present study, which was carried out at a boarding school in Bangladesh, was designed to examine probable effect of NS on mood, anxiety and cognition in adolescent human males…Forty-eight healthy adolescent human males aged between 14 to 17 years were randomly recruited as volunteers and were randomly split into two groups: A (n=24) and B (n=24). The treatment procedure for group A and B were one capsule of 500 mg placebo and 500 mg NS respectively once daily for four weeks. All the volunteers were assessed for cognition with modified California verbal learning test-II (CVLT-II), mood with Bond–Lader scale and anxiety with State–Trait Anxiety Inventory (STAI) at the beginning and after four weeks of either NS or placebo ingestion…Over the 4 weeks study period, the use of NS as a nutritional supplement been observed to- stabilize mood, decrease anxiety and modulate cognition positively.”

Relieving neuroinflammation of depression

Journal of Pharmacy & BioAllied SciencesIt’s well known than neuroimmune inflammation plays a fundamental role in depression. Authors of a study published in the Journal of Pharmacy & BioAllied Sciences present welcome evidence that Nigella sativa and thymoquinone may relieve depression by reducing neuroinflammation:

Neuroimmune factors have been proposed as contributors to the pathogenesis of depression. Beside other therapeutic effects including neuroprotective, antioxidant, anticonvulsant and analgesic effects, Nigella sativa and its main ingredient, thymoquinone (TQ), have been shown to have anti-inflammatory effects. In the present study, the effects of Nigella sativa hydro-alcoholic extract and thymoquinone was investigated on lipopolysaccharide- induced depression like behavior in rats…The results of the present study showed that hydro-alcoholic extract of Nigella sativa can prevent LPS-induced depression like behavior in rats. These results support the traditional belief on the beneficial effects of Nigella sativa in the nervous system.”

Thymoquinone ameliorates lead-induced brain damage

Experimental and Toxicologic PathologyEnvironmental toxicity is a concern for brain health; an exciting study published Experimental and Toxicologic Pathology indicates that thymoquinone from Nigella sativa protects against brain damage from lead:

“The present study aims to investigate the protective effects of thymoquinone, the major active ingredient of Nigella sativa seeds, against lead-induced brain damage in Sprague-Dawley rats. In which, 40 rats were divided into four groups (10 rats each). The first group served as control. The second, third and fourth groups received lead acetate, lead acetate and thymoquinone, and thymoquinone only, respectively, for one month. Lead acetate was given in drinking water at a concentration of 0.5 g/l (500 ppm). Thymoquinone was given daily at a dose of 20 mg/kg b.w. in corn oil by gastric tube. Control and thymoquinone-treated rats showed normal brain histology. Treatment of rats with lead acetate was shown to produce degeneration of endothelial lining of brain blood vessels with peri-vascular cuffing of mononuclear cells consistent to lymphocytes, congestion of choroid plexus blood vessels, ischemic brain infarction, chromatolysis and neuronal degeneration, microglial reaction and neuronophagia, degeneration of hippocampal and cerebellar neurons, and axonal demyelination. On the other hand, co-administration of thymoquinone with lead acetate markedly decreased the incidence of lead acetate-induced pathological lesions.”

Protection against Parkinson’s disease α-synuclein-induced synapse damage

Neuroscience LettersAgents that offer protection against α-synuclein toxicity are welcome in the treatment of Parkinson’s disease and dementia. A study recently published in Neuroscience Letters presents evidence that thymoquinone from Nigella sativa has this property:

“The present study aimed to determine whether TQ protects against α-synuclein (αSN)-induced synaptic toxicity in rat hippocampal and human induced pluripotent stem cell (hiPSC)-derived neurons. Here, we report that αSN decreased the level of synaptophysin, a protein used as an indicator of synaptic density, in cultured hippocampal and hiPSC-derived neurons. However, simultaneous treatment with αSN and TQ protected neurons against αSN-induced synapse damage, as revealed by immunostaining. Moreover, administration of TQ efficiently induced protection in these cells against αSN-induced inhibition of synaptic vesicle recycling in hippocampal and hiPSC-derived neurons as well as against mutated P123H β-synuclein (βSN) in hippocampal neurons, as revealed by experiments using the fluorescent dye FM1-43. Using a multielectrode array, we further demonstrated that the treatment of hiPSC-derived neurons with αSN induced a reduction in spontaneous firing activity, and cotreatment with αSN and TQ partially reversed this loss. These results suggest that TQ protects cultured rat primary hippocampal and hiPSC-derived neurons against αSN-induced synaptic toxicity and could be a promising therapeutic agent for patients with Parkinson’s disease and dementia with Lewy bodies.

Thymoquinone prevents β-amyloid neurotoxicity of Alzheimer’s disease

Cellular and Molecular NeurobiologyOf great interest in the prevention of Alzheimer’s disease are agents that may protect agains β-amyloid neurotoxicity. Here too thymoquinone has effect as reported in a study published in Cellular and Molecular Neurobiology:

Thymoquinone (TQ), a bioactive constituent of Nigella sativa Linn (N. sativa) has demonstrated several neuropharmacological attributes. In the present study, the neuroprotective properties of TQ were investigated by studying its anti-apoptotic potential to diminish β-amyloid peptide 1-40 sequence (Aβ1-40)-induced neuronal cell death in primary cultured cerebellar granule neurons (CGNs)…Pretreatment of CGNs with TQ (0.1 and 1 μM) and subsequent exposure to 10 μM Aβ1-40 protected the CGNs against the neurotoxic effects of the latter. In addition, the CGNs were better preserved with intact cell bodies, extensive neurite networks, a loss of condensed chromatin and less free radical generation than those exposed to Aβ1-40 alone. TQ pretreatment inhibited Aβ1-40-induced apoptosis of CGNs via both extrinsic and intrinsic caspase pathways. Thus, the findings of this study suggest that TQ may prevent neurotoxicity and Aβ1-40-induced apoptosis. TQ is, therefore, worth studying further for its potential to reduce the risks of developing Alzheimer’s disease.”

 Nigella sativa protects and promotes healing from nerve trauma

Pathologie BiologieA study published Pathologie Biologie reports that Nigella sativa improves the neurodegeneration typical after nerve trauma:

“The aim of this study was designed to evaluate the possible protective effects of Nigella sativa (NS) on the neuronal injury in the sciatic nerve of rats. The rats were randomly allotted into one of the three experimental groups: A (control), B (only trauma) and C (trauma and treated with NS); each group contain 10 animals… To date, no histopathological changes of neurodegeneration in the sciatic nerve after trauma in rats by NS treatment have been reported. Results showed in the group B (only trauma), the neurons of sciatic nerve tissue became extensively dark and degenerated with picnotic nuclei. Treatment of NS markedly reduced degenerating neurons after trauma and the distorted nerve cells were mainly absent in the NS-treated rats. The morphology of neurons in groups treated with NS was well protected, but not as neurons of the control group. The number of neurons in sciatic nerve tissue of group B (only trauma) was significantly less than both control and treated with NS groups. The morphology of neurons revealed that the number of neurons were significantly less in group B compared to control and group C rats’ motor neurons anterior horn spinal cord tissue. We conclude that NS therapy causes morphologic improvement on neurodegeneration in sciatic nerve after trauma in rats.”

Nigella sativa for osteoporosis

Evidence-Based Complementary and Alternative MedicineConsidering that inflammation plays a key role in osteoporosis, it’s reasonable to investigate the use Nigella sativa as described in a paper in Evidence-Based Complementary and Alternative Medicine:

“Animal studies have shown that NS and TQ may be used for the treatment of diabetes-induced osteoporosis and for the promotion of fracture healing. The mechanism involved is unclear, but it was postulated that the antioxidative, and anti-inflammatory activities may play some roles in the treatment of osteoporosis as this bone disease has been linked to oxidative stress and inflammation. This paper highlights studies on the antiosteoporotic effects of NS and TQ, the mechanisms behind these effects and their safety profiles. NS and TQ were shown to inhibit inflammatory cytokines such as interleukin-1 and 6 and the transcription factor, nuclear factor κB. NS and TQ were found to be safe at the current dosage for supplementation in human with precautions in children and pregnant women. Both NS and TQ have shown potential as antiosteoporotic agent but more animal and clinical studies are required to further assess their antiosteoporotic efficacies.”

Inhibition of osteoporosis by Nigella sativa

BMC Complementary & Alternative MedicineIn an exciting study published in the BMC Complementary and Alternative Medicine, investigators report the reversal of osteoporosis in subjects whose ovaries had been removed:

“There is a direct relationship between the lack of estrogen after menopause and the development of osteoporosis…Nigella Sativa (NS) has been shown to have beneficial effects on bone and joint diseases. The present study was conducted to elucidate the protective effect of Nigella Sativa on osteoporosis produced by ovariectomy in rats…Female Wistar rats aged 12-14 months were divided into three groups: sham-operated control (SHAM), ovariectomized (OVX), and ovariectomized supplemented with nigella sativa (OVX-NS) orally for 12 weeks; 4 weeks before ovariectomy and 8 weeks after…OVX rats showed significant decrease in plasma Ca(+2), accompanied by a significant increase in plasma ALP, amino terminal collagen type 1 telopeptide, MDA, nitrates, TNF-α and IL-6. These changes were reversed by NS supplementation in OVX-NS group to be near SHAM levels. Histological examination of the tibias revealed discontinuous eroded bone trabeculae with widened bone marrow spaces in OVX rats accompanied by a significant decrease in both cortical and trabecular bone thickness compared to Sham rats. These parameters were markedly reversed in OVX-NS rats. Histological examination of the liver showed mononuclear cellular infiltration and congestion of blood vessels at the portal area in OVX rats which were not found in OVX-NS rats.”

Their data supported this exciting conclusion:

“It can be concluded that NS has shown potential as a safe and effective antiosteoporotic agent, which can be attributed to its high content of unsaturated fatty acids as well as its antioxidant and anti-inflammatory properties.”

Nigella sativa helps with psoriasis

Pharmacognosy MagazineConsidering its antiinflammatory and immunomodulating characteristics it seems a good bet that Nigella sativa would help with psoriasis as described in a study published in Pharmacognosy Magazine:

“The screening of antipsoriatic activity of 95% of ethanolic extract of Nigella sativa seeds by using mouse tail model for psoriasis and in vitro antipsoriatic activity was carried out by SRB Assay using HaCaT human keratinocyte cell lines….The ethanolic extract of Nigella sativa seeds extract produced a significant epidermal differentiation, from its degree of orthokeratosis (71.36±2.64) when compared to the negative control (17.30±4.09%)…The 95% ethanolic extract of Nigella sativa shown IC50 239 μg/ml, with good antiproliferant activity compared to Asiaticoside as positive control which showed potent activity with IC50 value of 20.13 μg/ml. From the present study it can be said that topical application of 95% ethanolic extract of Nigella sativa seeds has antipsoriatic activity and the external application is be beneficial in the management of psoriasis.”

Assists in treatment of vitiligo

Iranian Red Crescent Medical JournalNIgella sativa is an agent to consider in case management of any autoimmune disorder including vitiligo, for which it showed benefit in a study published in the Iranian Red Crescent Medical Journal:

Vitiligo is one of the autoimmune skin diseases that destroy the melanocytes of the skin…The aim of this study was to compare the effect of Nigella sativa and fish oil on vitiligo lesions of the patients referred to a dermatology clinic…After six months, a mean score of VASI decreased from 4.98 to 3.75 in patients applying topical Nigella sativa and from 4.98 to 4.62 in those using topical fish oil…In the current study, administration of Nigella sativa and fish oil significantly decreased skin lesions size, indicating an improvement in clinical condition…the depigmented areas were reduced over time and the skin color showed improvement. One reason for this positive response to treatment is the thymoquinone component of Nigella sativa…Thymoquinone can simulate the activity of acetylcholine, which causes the release of melanin and darkening of the skin through stimulation of cholinergic receptors. In addition, Nigella sativa oil administration was tolerable as well as safe and improved oxidative stress and clinical condition of patients…It was also shown that this type of treatment has no significant side effects and resulted in high patient satisfaction and acceptance.”

The authors state in conclusion:

“Nigella sativa oil and fish oil were effective in reduction the size of patient’s lesions; however, Nigella sativa was more effective in comparison to the fish oil. Therefore, using Nigella sativa with the major drugs in the treatment of vitiligo is recommended.”

Topical treatment of allergic rhinitis

Anti-Inflammatory & Anti-Allergy Agents in Medicinal ChemistryAllergic rhinitis as a chronic inflammatory disorder also responds to Nigella sativa applied topically as reported in Anti-Inflammatory & Anti-Allergy Agents in Medicinal Chemistry:

Allergic rhinitis (AR) is the most common manifestation of atopic reaction to inhaled allergens. It is a chronic inflammatory disease which may first appear at any age, but the onset is usually during childhood or adolescence…The individuals in the active group received N. sativa oil and the control group individuals received ordinary food oil in the form of nasal drops for 6 weeks…After the 6 weeks treatment course, 100% of the patients in the mild active group became symptoms free; while in moderate active group 68.7% became symptoms free and 25% were improved; while in severe active group 58.3% became symptoms free and 25% were improved. In addition, 92.1% of total patients in the active group demonstrated improvement in their symptoms or were symptoms free, while the corresponding value was 30.1% in the control group. At the end of 6 weeks of treatment with topical use, the improvement in tolerability of allergen exposure in active group became 55.2% which was significant as compared with control group which was accounted for 20% at the same time…Topical application of black seed oil was effective in the treatment of allergic rhinitis, with minimal side effects.”

Nigella sativa protects against radiation damage

Journal of Investigative SurgeryRadiation therapy can produce substantial ‘collateral damage’. Authors of a study just published in the Journal of Investigative Surgery demonstrate that Nigella sativa reduces oxidative stress in animals subjected to total head irradiation:

“Many cancer patients treated with radiotherapy suffer severe side effects during and after their treatment. The aim of this study was to investigate the effects of irradiation and the addition of Nigella sativa oil (NSO) on the oxidant/antioxidant system in the liver tissue of irradiated rats…The control group received neither NSO nor irradiation but received 1-ml saline orally. The irradiation group (IR) received total head 5 gray (Gy) of gamma irradiation as a single dose, plus 1-ml saline orally. The IR plus NSO group received both total head 5 Gy of gamma irradiation as a single dose and 1 g/kg/day NSO orally through an orogastric tube starting one hour before irradiation and continuing for 10 days…Conclusions: NSO reduces oxidative stress markers and has antioxidant effects, which also augments the antioxidant capacity in the liver tissue of rats.”

Cutaneous and Ocular ToxicologyNigella sativa was shown to reduce radiation-induced cataracts in a study published in Cutaneous and Ocular Toxicology:

“The aim of this study was to investigate the antioxidant and radioprotective effects of Nigella sativa oil (NSO) and thymoquinone (TQ) against ionizing radiation-induced cataracts in lens after total cranium irradiation (IR) of rats with a single dose of 5 gray (Gy)…At the end of the 10th d, cataract developed in 80% of the rats in IR group only. After IR, cataract rate dropped to 20% and 50% in groups which were treated with NSO and TQ, respectively, and was limited at grades 1 and 2. Nitric oxide synthase activity, nitric oxide and peroxynitrite levels in the radiotherapy group were higher than those of all other groups. Conclusions: The results implicate a major role for NSO and TQ in preventing cataractogenesis in ionizing radiation-induced cataracts in the lenses of rats, wherein NSO were found to be more potent.”

PhytomedicineAnd protection from radiation-induced damage to brain tissue was demonstrated in a study recently published in the journal Phytomedicine designed…

“To investigate Nigella sativa oil (NSO) and Thymoquinone (TQ) for their antioxidant effects on the brain tissue of rats exposed to ionizing radiation….Levels of NO· and ONOO(-), and enzyme activity of NOS in brain tissue of the rats treated with NSO or TQ were found to be lower than in received IR alone (p<0.002) Nigella sativa oil (NSO) and its active component, TQ, clearly protect brain tissue from radiation-induced nitrosative stress.

 Activity against Staphylococcal and fungal skin infections

Pakistan Journal of Biological SciencesNigella sativa is a benevolent agent in the treatment of skin infection and inflammation as documented by a study published in the Pakistan Journal of Biological Sciences:

“Nigella sativa has been used for a long time in Jordanian folk medicine to treat skin diseases like microbial infections and inflammation. Therefore, the present study was conducted to assess the healing efficacy of petroleum ether extract of Nigella sativa seeds (fixed oil) on staphylococcal-infected skin. Male BALB/c mice were infected with 100 microL of Staphylococcus aureus (ATCC 6538)… Application of treatments for each group (100 microL sterile saline, 100 microL chloramphenicol (10 microg/mouse) and Nigella sativa fixed oil at a dose of 50, 100 or 150 microL/mouse) was performed at the site of infection… At day 3 and 5 after infection, total White Blood Cells (WBCs) count; differential and absolute differential WBC counts and the number of viable bacteria present in the skin area were measured…Results indicated that fixed oil of Nigella sativa seeds enhance healing of staphylococcal-infected skin by reducing total and absolute differential WBC counts, local infection and inflammation, bacterial expansion and tissue impairment. These effects provide scientific basis for the use of Nigella sativa in traditional medicine to treat skin infections and inflammations.

Journal of EthnopharmacologyThe authors of a study published in the Journal of Ethnopharmacology report effectiveness against fungal skin infections (dermatophytes):

“The antifungal activity of ether extract of Nigella sativa seed and its active principle thymoquinone was tested against eight species of dermatophytes: four species of Trichophyton rubrum and one each of Trichophyton interdigitale, Trichophyton mentagrophytes, Epidermophyton floccosum and Microsporum canis. Agar diffusion method with serial dilutions of ether extract of Nigella sativa, thymoquinone and griseofulvin was employed…The minimum inhibitory concentration (MIC) was considered as the minimum concentration of the drug, which inhibited 80–100% of the fungal growth. The MICs of the ether extract of Nigella sativa and thymoquinone were between 10 and 40 and 0.125 and 0.25 mg/ml…These results denote the potentiality of Nigella sativa as a source for antidermatophyte drugs and support its use in folk medicine for the treatment of fungal skin infections.”

Case report of seroreversion in HIV

Afr J Tradit Complement Altern Med.A case report published in the African Journal of Traditional, Complementary, and Alternative Medicines presents unexpected results in the treatment of HIV:

“Nigella sativa had been documented to possess many therapeutic functions in medicine but the least expected is sero-reversion in HIV infection which is very rare despite extensive therapy with highly active anti-retroviral therapy (HAART). This case presentation is to highlight the complete recovery and sero-reversion of adult HIV patient after treatment with Nigella sativa concoction for the period of six months. The patient presented to the herbal therapist with history of chronic fever, diarrhoea, weight loss and multiple papular pruritic lesions of 3 months duration. Examination revealed moderate weight loss, and the laboratory tests of ELISA (Genscreen) and western blot (new blot 1 & 2) confirmed sero-positivity to HIV infection with pre-treatment viral (HIV-RNA) load and CD4 count of 27,000 copies/ml and CD4 count of 250 cells/ mm(3) respectively. The patient was commenced on Nigella sativa concoction 10 mls twice daily for 6 months. He was contacted daily to monitor side-effects and drug efficacy. Fever, diarrhoea and multiple pruritic lesions disappeared on 5th, 7th and 20th day respectively on Nigella sativa therapy. The CD4 count decreased to 160 cells/ mm3 despite significant reduction in viral load (≤1000 copies/ml) on 30th day on N. sativa. Repeated EIA and Western blot tests on 187th day on Nigella sativa therapy was sero-negative. The post therapy CD4 count was 650 cells/ mm(3) with undetectable viral (HIV-RNA) load. Several repeats of the HIV tests remained sero-negative, aviraemia and normal CD4 count since 24 months without herbal therapy. This case report reflects the fact that there are possible therapeutic agents in Nigella sativa that may effectively control HIV infection.

Improvement in semen quality

PhytomedicineAnother study published in Phytomedicine presents evidence from a double-blind, placebo-controlled that Nigella sativa improves abnormal semen quality in infertility:

“Since Nigella sativa L. seed (N. sativa) has many uses including infertility in traditional medicine, the effects of Nigella sativa L. seed oil on abnormal semen quality in infertile men with abnormal semen quality are of interest. This study was conducted on Iranian infertile men with inclusion criteria of abnormal sperm morphology less than 30% or sperm counts below 20×10(6)/ml or type A and B motility less than 25% and 50% respectively. The patients in N. sativa oil group (n=34) received 2.5mlN. sativa oil and placebo group (n=34) received 2.5ml liquid paraffin two times a day orally for 2 months. At baseline and after 2 months, the sperm count, motility and morphology and semen volume, pH and round cells as primary outcomes were determined in both groups. Results showed that sperm count, motility and morphology and semen volume, pH and round cells were improved significantly in N. sativa oil treated group compared with placebo group after 2 months. It is concluded that daily intake of 5ml N. sativa oil for two months improves abnormal semen quality in infertile men without any adverse effects.”

Is Nigella sativa safe?

Advanced Pharmaceutical BulletinA study investigating the potential for liver toxicity was reported last year in the journal Advanced Pharmaceutical Bulletin:

“The aim of this study was to determine the toxic effect of Nigella sativa powder on the liver function which was evaluated by measuring liver enzymes and through histopathological examination of liver tissue…Twenty four male Sprague Dawley rats were allotted randomly to four groups including: control (taking normal diet); low dose (supplemented with 0.01 g/kg/day Nigella sativa); normal dose (supplemented with 0.1 g/kg/day Nigella sativa) and high dose (supplemented with 1 g/kg/day Nigella sativa)…To assess liver toxicity, liver enzymes measurement and histological study were done at the end of supplementation…The study showed that supplementation of Nigella sativa up to the dose of 1 g/kg supplemented for a period of 28 days resulted no changes in liver enzymes level and did not cause any toxicity effect on the liver function

The authors stated this conclusion regarding human consumption of Nigella sativa:

“With the evidence of normal ALT and AST level in blood and normal liver tissue in histology examination for all treatment groups, it is suggested that there are no toxic effect on liver function of Nigella sativa at different doses for 4 weeks period. As a conclusion, popular consumption of Nigella sativa powder by human did not cause any toxicity effect on the liver function and safe to be consumed for many purposes.”

 Protection against alcohol-induced liver injury

Chinese Journal of Natural MedicinesNot only is Nigella sativa safe for the liver, but a study published in the Chinese Journal of Natural Medicines provides data showing that it protects the liver against oxidative damage caused by alcohol:

Nigella sativa L. (Ranunculaceae) is considered as a therapeutic plant-based medicine for liver damage. In this study, the aim was to study the effect of Nigella sativa oil (NSO) pretreatment on ethanol-induced hepatotoxicity in rats…Rats were given Nigella sativa oil at doses of 2.5 and 5.0 mL·kg(-1), orally for 3 weeks, followed by oral ethanol (EtOH) administration (5 g·kg(-1)) every 12 h three times (binge model).”

Amazingly…

Binge ethanol application caused significant increases in plasma transaminase activities and hepatic triglyceride and malondialdehyde (MDA) levels. It decreased hepatic glutathione (GSH) levels, but did not change vitamins E and vitamin C levels and antioxidant enzyme activities. NSO (5.0 mL·kg(-1)) pretreatment significantly decreased plasma transaminase activities, hepatic MDA, and triglyceride levels together with amelioration in hepatic histopathological findings.”

Based on these findings the authors conclude:

“NSO pretreatment may be effective in protecting oxidative stress-induced hepatotoxicity after ethanol administration.”

Practical use of Nigella sativa

Nigella sativa seeds 3The foregoing sampling of studies from the scientific literature on Nigella sativa should not be construed as an endorsement for its use in any specific case or condition. It is a presentation of the extraordinary scope of action and clinical potential of an agent that I am finding valuable in practice. Colleagues who are interested in knowing the particular Nigella sativa whole seed extract that I am using are welcome to contact me. For the general reader, I caution against taking anything (especially something found on the internet) without having first discussed it with your knowledgeable health care practitioner who has the background and depth to advise on how this may fit into your treatment or health maintenance plan.

Antioxidants in excess can increase inflammation and blunt benefits of exercise

PLOS ONEAntioxidants, even glutathione, taken in excess can increase rather than ameliorate harmful inflammation as attested by two revealing papers recently published in PLOS (Public Library of Science). In a fascinating study on experimental colitis, the authors demonstrate that certain levels of ROS (reactive oxygen species, well known to be damaging at higher levels) are critically necessary to regulate the Treg (regulatory T cell) function that controls autoimmune inflammation. Moreover, of premiere clinical importance is their observation that higher levels of glutathione that excessively dampened ROS blocked Treg function and thus worsened the inflammation of inflammatory bowel disease (IBD). Antioxidants, of course, are employed to keep the lid on ROS. And with good reason:

Reactive oxygen species (ROS) are highly reactive and interact with many bio-molecules. At high concentrations, they are likely to destroy biological structures, promoting cellular damage and tissue destruction. Traditionally, ROS have been implicated in ageing and the progression of inflammatory and autoimmune diseases, including inflammatory bowel diseases (IBD).

However…

Meanwhile, many recent observations are opposing the traditional concept on ROS, suggesting the protective role of ROS in immune-mediated inflammatory diseases.”

The authors relate the fascinating background to this study on ROS, antioxidants and autoimmune inflammation:

“Mice with lower level of ROS than WT mice due to defects in ROS-producing enzyme system, such as Ncf1−/− or Nox2−/−, are more susceptible to autoimmune diseases, such as arthritis and encephalomyelitis. Humans with lower levels ROS than normal persons, such as chronic granulomatous disease (CGD) patients and carriers, are also more susceptible to autoimmune diseases. By contrast, mice with higher level ROS than WT mice due to the defect in a ROS metabolizing enzyme, glutathione peroxidase-1 (GPx-1), are resistant to immune-mediated inflammatory diseases, such as allergen-induced airway inflammation and high fat diet-induced atherosclerosis. In particular, mice with higher level of ROS due to defect of a non-enzymatic cellular anti-oxidant, peroxiredoxin (Prx) II, are resistant to dextran sodium sulfate (DSS)-induced colitis.”

In other words, more enzyme activity producing antioxidants such as glutathione and peroxiredoxin increased susceptibility to autoimmune inflammatory disorders. Moreover…

“These clinical or experimental observations implicated the immunoregulatory role of ROS, and adoptive-transfer of CD4+ cells from rats with lower ROS level induced arthritis in rats with normal ROS level, demonstrating the key role of CD4+ cells in the hyperinflammatory response in lowered levels of ROS. On the other hand, oxidative stress induces T cell hyporesponsiveness in several human pathologies (e.g. cancer, rheumatoid arthritis, AIDS and leprosy). Accordingly, ROS level is supposed to be closely associated with T cell responsiveness. In particular, regulatory T cell (Treg) function seems to be closely linked to ROS level. Tregs isolated from mice with lower level of ROS, such as Ncf1−/− mice, were hypofunctional than WT Tregs. Tregs were also hypofunctional in vitro at lowered levels of ROS by adding antioxidants or NADPH oxidase inhibitors. Differentiation of inducible Treg (iTreg) seems also linked to the level of ROS. Induction of FoxP3+ iTreg was attenuated, whereas that of Th17 cells was enhanced in lowered levels of ROS due to Nox2 deficiency or addition of apocynin. By contrast, induction of FoxP3+ Treg was enhanced in elevated levels of ROS due to PrxII deficiency.”

Thus, whereas excessive oxidative stress (ROS) with insufficient antioxidants can contribute to T cell hypoactivity in some pathologies including cancer and AIDS, levels of regulatory T cell (FoxP3+ iTreg) that rein in autoimmune Th17 inflammation are suppressed in the presence to ROS levels that are too low.

Colitis inflammation attenuated in KO mice, aggravated by NACSo the authors set out to investigate the suppressive function of Tregs isolated from mice with elevated levels of ROS due to defects in the enzymes that make the antioxidatns glutathione (GPx1) and catalase. Their results illuminated the aspects of ROS and antioxidants that are critical to case management of inflammatory disorders:

“In the present study, we demonstrated for the first time that Tregs were hyperfunctional in elevated level of ROS by using GPx1−/− × Cat−/− Tregs. As it has been already reported that Tregs were hypofunctional in lowered levels of ROS, it could be argued that Treg function is closely linked to ROS level. Actually in the present study, IP injection of NAC [N-acetylcysteine] into GPx1−/− × Cat−/− mice reduced the suppressive function of Tregs to the level comparable to WT Tregs. Administration of NAC also has made GPx1−/− × Cat−/− mice, which are naturally resistant, susceptible to DSS-induced colitis, suggesting the critical role of ROS in the prevention of DSS-induced colitis. The importance of Tregs in the maintenance of intestinal immune balance has been already shown in many other studies. Consequently, ROS level might be critical in the maintenance of intestinal immune homeostasis, providing an insight for the immunomodulation by ROS.”

In other words, elevated ROS increase the antiinflammatory activity of Treg cells, while administration of NAC (which increases glutathione production) suppressed Tregs by reducing ROS, resulting in increased intestinal inflammation.

Clinical note: Excessive suppression of ROS by overenthusiastic application of antioxidants can disturb immune homeostasis resulting in increased autoimmune inflammation.

The authors also shed light on the association of ROS and inflammation with tryptophan metabolism:

At molecular level, the expression of an immunoregulatory enzyme, IDO, is also associated with ROS level. IDO catabolizes the essential amino acid tryptophan into the stable metabolite, kynurenine. Consequently, IDO depletes tryptophan from the environment, thus starving effector cells. It was also found that tryptophan depletion resulted in inhibition of Th17 cell differentiation and expansion of Foxp3+ Tregs….Thus, IDO expression might be induced as a consequence of the inflammatory reaction, contributing to the feedback regulation…Elevated levels of ROS not only contribute to the induction but also enhance the enzyme activity of IDO, as superoxide radical acts as a cofactor of IDO. Therefore, high expression and strong activity of IDO from the beginning in GPx1−/− × Cat−/− mice might contribute to the preparation of immunosuppressive environment preventing inflammatory tissue damage during treatment with DSS.”

Clinical note: antioxidant and ROS status can be investigated with a urinary organic acid analysis that includes p-Hydroxyphenyllactate, 8-Hydroxy-2′-deoxyguanosine, kynurenate and quinolinate; and a blood assay for oxidative stress that includes cysteine, cystine, glutathione, glutathione peroxidase, lipid peroxides, sulfate, superoxide dismutase (SOD), and total antioxidant capacity (TAC).

The authors summarize implications for modulation of ROS by antioxidants:

“Actually in the present study, the frequency of FoxP3+ cells was significantly increased in parallel with significantly attenuated inflammatory reaction in the lesions of DSS-induced colitis in mice with elevated level of ROS due to defects in GPx1 and Cat. By contrast, IP injection of NAC significantly reduced the frequency of FoxP3+ cells and aggravated inflammatory reaction in the lesions of DSS-induced colitis… we demonstrated an experimental colitis was attenuated in elevated level of ROS. Enhancement of Treg function and IDO expression, investigated in the present study, might be involved in the underlying mechanism…Taken together, the results of the present study suggest the potential therapeutic strategy for IBD through immunomodulation by ROS.”

ROS and Psoriasis

The same team of scientists a study also in PLOS One similarly investigating the role of ROS and by implication the use of antioxidants in regard to psoriatic dermatitis:

Psoriasis is a chronic inflammatory skin disease resulting from immune dysregulation. Regulatory T cells (Tregs) are important in the prevention of psoriasis. Traditionally, reactive oxygen species (ROS) are known to be implicated in the progression of inflammatory diseases, including psoriasis, but many recent studies suggested the protective role of ROS in immune-mediated diseases. In particular, severe cases of psoriasis vulgaris have been reported to be successfully treated by hyperbaric oxygen therapy (HBOT), which raises tissue level of ROS. Also it was reported that Treg function was closely associated with ROS level. However, it has been only investigated in lowered levels of ROS so far.”

The authors state:

Psoriasis is known to develop as a result of immune dysregulation, in particular hyperfunction of T helper 17 (Th17) cells. In steady state, immune homeostasis is maintained by regulatory T cells (Tregs) that suppress immune effectors including Th17 cells. It was also reported that psoriasis is associated with impaired suppressive function of Tregs. Therefore, in order to restore the dysregulated immune status in psoriasis, it is necessary to suppress immune effectors including Th17 cells and/or to enhance Tregs.”

Consonant with their study on colitis:

“Traditionally, reactive oxygen species (ROS) are known to be implicated in the progression of many inflammatory diseases. As ROS are highly reactive and interact with many bio-molecules, they are likely to destroy biological structures, promoting cellular damage and tissue destruction. In contrast, many recent evidences are accumulating on the protective role of ROS in immune-mediated diseases. Autoimmune arthritis was aggravated in rodents with lower levels of ROS than wildtype (WT) mice due to defects in ROS-producing enzyme system, such as mutation in the neutrophil cytosolic factor (Ncf)-1 or NADPH oxidase (NOX)2. In human, too, many autoimmune diseases develop more frequently in chronic granulomatous disease (CGD) patients with lower level of ROS than normal persons due to defect in ROS-producing NOX. To the contrary, experimentally induced asthmatic inflammation was attenuated in mice with higher level of ROS than WT mice due to the defect of a ROS metabolizing enzyme, glutathione peroxidase-1 (GPx-1). Atherosclerotic lesions induced by high-fat diet were also decreased in GPx-1−/− mice. In addition, experimental colitis was attenuated in mice with a higher level of ROS due to defect in a non-enzymatic anti-oxidant, peroxiredoxin II.”

So they set out to…

“…clarify the relationship between ROS level and Treg function, as well as their role in the pathogenesis of psoriasis, we investigated imiquimod-induced psoriatic dermatitis (PD) in association with Treg function both in elevated and lowered levels of ROS by using knockout mice, such as glutathione peroxidase-1−/− and neutrophil cytosolic factor-1−/− mice, as well as by using HBOT or chemicals, such as 2,3-dimethoxy-1,4-naphthoquinone and N-acetylcysteine.

Reactive Oxygen Species Prevent Imiquimod-Induced Psoriatic Dermatitis through Enhancing Regulatory T Cell FunctionHere too they found that moderately high levels of ROS were protective against autoimmune inflammation PD (psoriatic dermatitis):

The results consistently showed Tregs were hyperfunctional in elevated levels of ROS, whereas hypofunctional in lowered levels of ROS. In addition, imiquimod-induced PD was attenuated in elevated levels of ROS, whereas aggravated in lowered levels of ROS. For the molecular mechanism that may link ROS level and Treg function, we investigated the expression of an immunoregulatory enzyme, indoleamine 2,3-dioxygenase (IDO) which is induced by ROS, in PD lesions. Taken together, it was implied that appropriately elevated levels of ROS might prevent psoriasis through enhancing IDO expression and Treg function.”

Reflecting on the clinical significance of these results and the crucial difference between moderately high levels of ROS which are protective versus higher levels that are damaging:

“In the present study, we demonstrated that imiquimod-induced PD was attenuated in elevated levels of ROS, whereas aggravated in lowered levels of ROS. This observation provides experimental evidence supporting the immunoregulatory role of ROS, that is contradictory to the traditional concept. Traditionally, ROS is implicated in the progression of inflammatory diseases by promoting cellular damage and tissue destruction as well as ageing. At the moment, it is necessary to establish a new conceptual framework where the recent observations and the traditional concept can be compromised…Taken together, we can imagine a threshold level of ROS that divides the moderately high tolerable range and the intolerably higher levels. In the higher levels such as GPx-1−/−×GPx-2−/− mice, inflammatory reactions are augmented due to the direct tissue damage by ROS. Many evident previous observations that contribute to establish the traditional concept of ROS, such as vascular reperfusion injury and other in vitro observations, might fall in this range of intolerably higher levels of ROS. In contrast, as demonstrated previously by others and by us in the present study, inflammatory diseases are attenuated in the moderately high tolerably ranges such as in GPx-1−/− or PrxII−/− mice. Thus, we suppose some kinds of anti-inflammatory mechanisms are operating in the moderately high tolerable range of ROS. As ROS can induce direct tissue damages at high levels, it would be natural to develop defensive or compensatory mechanisms counteracting the destructive effects of ROS in the body.”

Clinical note: Here is where the rubber meets the road in clinical case management of autoimmune diseases in respect to ROS versus excessive use of antioxidants:

“In this aspect, enhancement of Treg function depending on ROS level is quite pertinent to counteract the destructive damages induced by ROS, as Tregs suppress every arm of immune response, including Th1, Th2, Th17, B, NK cells and DCsTregs play a critical role in the prevention of autoimmune diseases, and functional impairment of Tregs is important in the pathogenesis of psoriasis. Therefore, restoration or strengthening of impaired Treg function would be a desirable therapeutic strategy for psoriasis. The results of the present study suggested appropriately elevated levels of ROS could enhance Treg function, and thus might attenuate psoriasis.”

In particular for clinicians involved in case management of autoimmune disorders, from a personal communication with one of the lead authors Ju-young Seoh, MD, Ph,D.:

“Your understanding, “too much as well as too little antioxidant activity can be harmful, and too aggressive use of antioxidant agent, such as vitamin C, could impair Treg activity and accelerate autoimmune inflammatory activity.” is exact our message.”

 

Clinical Chimica ActaThe authors of a paper just published in Clinica Chimica Acta also note the distinction between low and high levels of ROS and the role of antioxidants:

“Oxidative stress plays a pivotal role in the development of human diseases. Reactive oxygen species (ROS) that includes hydrogen peroxide, hyphochlorus acid, superoxide anion, singlet oxygen, lipid peroxides, hypochlorite and hydroxyl radical are involved in growth, differentiation, progression and death of the cell. They can react with membrane lipids, nucleic acids, proteins, enzymes and other small molecules. Low concentrations of ROS has an indispensable role in intracellular signalling and defence against pathogens, while, higher amounts of ROS play a role in number of human diseases, including arthritis, cancer, diabetes, atherosclerosis, ischemia, failures in immunity and endocrine functions. Antioxidants presumably act as safeguard against the accumulation of ROS and their elimination from the system.”

Antioxidants and Exercise

Journal of the International Society of Sports NutritionIt’s edifying in this context to consider the effects of antioxidants on desirable physiological adaptations to the stress of exercise. In a study recently published in the Journal of the International Society of Sports Nutrition the investigators documented that while antioxidants increased the strength of muscle contraction acutely, there was also a suppression of the desirable growth hormone (GH) response to exercise (which increases muscle mass).

Antioxidant supplementation is known to increase human endogenous antioxidant (AOX) capacity providing a means of blunting exercise induced reactive oxygen species (ROS). The purpose of this study was to compare the effects of a single acute dose of an AOX (vs blinded placebo) on muscle contractile performance and hormonal responses to a single bout of lower limb ‘hypertrophic’ resistance training (RT).”

Their data should be considered in the context of antioxidant supplementation and exercise:

“It was found that in comparison to a placebo mixture, subjects were able to perform 3.75% more work (W), and generate greater mean concentric power and velocity throughout the HTS after consuming the AOX mixture…however circulating GH levels was significantly reduced in the AOX trial compared to the placebo trial…This would suggest that the GH results from this study indicate they may be undesirable in regards to promoting muscular hypertrophy. It is therefore of interest for future studies to examine whether this decreased circulating GH would affect muscular hypertrophy after a prolonged period of use or whether it acutely affects IGF-1 levels. Moreover, recent research suggests excessive AOX supplementation may hinder important physiological training adaptations. This has prompted the suggestion that optimal oxidant content for maximal force production exists within the muscle…GH secretion is involved in MH and strength development and its attenuation may negatively impact training adaptations…These recent findings and the GH results in this study, highlight the need to further our understanding of the effect of AOX supplementation on training adaptations.”

 

Sports MedicineThese results further validate the findings documented in paper published a couple of years earlier in the journal Sports Medicine:

“High levels of reactive oxygen species (ROS) produced in skeletal muscle during exercise have been associated with muscle damage and impaired muscle function. Supporting endogenous defence systems with additional oral doses of antioxidants has received much attention as a noninvasive strategy to prevent or reduce oxidative stress, decrease muscle damage and improve exercise performance…The consistent finding is that antioxidant supplementation attenuates exercise-induced oxidative stress. However, any physiological implications of this have yet to be consistently demonstrated, with most studies reporting no effects on exercise- induced muscle damage and performance. Moreover, a growing body of evidence indicates detrimental effects of antioxidant supplementation on the health and performance benefits of exercise training. Indeed, although ROS are associated with harmful biological events, they are also essential to the development and optimal function of every cell. The aim of this review is to present and discuss 23 studies that have shown that antioxidant supplementation interferes with exercise training-induced adaptations.”

Here again we see that antioxidants have both positive and negative effects:

“The main findings of these studies are that, in certain situations, loading the cell with high doses of antioxidants leads to a blunting of the positive effects of exercise training and interferes with important ROS-mediated physiological processes, such as vasodilation and insulin signalling. More research is needed to produce evidence-based guidelines regarding the use of antioxidant supplementation during exercise training.”

 

Bottom line: The use of antioxidants must be calibrated with careful consideration of the balance between protective and suppressive effects according to the needs of the individual patient by observing appropriate lab values for ROS and oxidative damage, outcomes for regulation of inflammation, and the patient’s subjective impression of energy versus fatigue.

Vagal nerve activity moderates brain-immune relationships and is measured by heart rate variability

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 Brain-immune interactions control inflammation and the response to stress. An exciting study with tremendous practical significance was just published in the Journal of Neuroimmunology that shows how vagal nerve activity, which can be measured in the clinic by heart rate variability analysis (HRV), is a key moderator of the brain-immune web and determines the immune and physiological responses to acute stress. Highlights include:

  • Vagal tone indexed by heart rate variability reflects biological regulatory capacity.
  • Vagal tone is linked with flexible immune and physiological stress responses.
  • Frontal-striatal network mediates effects of vagal tone on stress responses.

Journal of NeuroimmunologyThe authors note:

“The bidirectional communication between the immune and nervous systems is considered to involve neural pathways that link these systems and expression of receptors for ligands such as cytokines and neurotransmitters. The brain produces immune-regulatory effects, and immunity has sensory functions (Haddad, 2008). Specifically, descending neural influences on immunity include neural innervation of lymphatic organs (Madden et al., 1995), expression of receptors for neurotransmitters on immune cells (Levite, 2008; Tracey, 2009) and differential left versus right hemisphere influences on immunity (Davidson et al., 1999; Sumner et al., 2011). Ascending immune-to-brain pathways include immune signals entering brain regions that lack a blood-brain barrier (BBB), prostaglandins on both sides of the BBB that mediate inflammatory signals, and an immune-to-brain conversion of inflammatory information by the vagus nerve (Ek et al., 1998; Dantzer et al., 2000; Davidson et al., 2001; Tracey, 2009).”

Fortunately there is increasing interest in understanding the brain-immune web and how the brain modulates the immune system during acute stress. Earlier studies have shown that the brain regions involved in executive functions and stress coping also modulate adaptive immune activities, and are responsible for making the physiological response to stress flexible and appropriate. The authors observe:

“Such associations between the neural and immune systems may depend on and be affected by a third variable, relevant to both systems, specifically tonic activity of the vagus nerve (Thayer and Sternberg, 2010). The vagus nerve expresses receptors for interleukin-1, enabling it to convert immune to nerve information via ascending acetylcholine signals to the brain stem (Ek et al., 1998). In return, the descending vagus modulates the activity of peripheral leukocytes and inflammation via the HPA axis and neural routes that inhibit cytokine production by macrophages (Tracey, 2009). Importantly, brain regions regulating activity of the vagus nerve partly overlap with brain regions involved in immune regulation, including the medial prefrontal cortex (MPFC) and DLPFC (Lane et al., 2009; Ohira et al., 2009). Given the strategic location of the vagus nerve mediating between the periphery and the brain and given its neuroimmunomodulatory roles, we hypothesized that tonic activity of the vagus nerve, indexed by heart rate variability (HRV) in a resting state, moderates transient brain–immune relationships accompanying acute stress.”

Moreover…

“…it was previously reported that individuals with a higher resting HRV showed faster recovery in their acute stress responses of immune, neuroendocrine, and cardiovascular parameters (Weber et al., 2010). These data suggest that higher resting HRV is associated with context-appropriate responses including adaptive recovery after termination of stress, and that the autonomic and endocrine systems mediate the associations between brain and immunity.”

Correlations between regional cerebral blood flow with proportion of natural killer cells and concentration of adrenocorticotropic hormone in high heart rate variability group.

Correlations between regional cerebral blood flow with proportion of natural killer cells and concentration of adrenocorticotropic hormone in high heart rate variability group.

So they set out to investigate whether vagus nerve activity as measured by HRV modulates brain-immune associations including the autonomic nervous system and endocrine (HPA) responses to acute stress. They subjected their study subjects to a learning task that has been proven to psychological and physiological stress and serve as a valid acute stressor for study purposes. The participants underwent PET scans of the brain, had blood sampled after each stress for the ratio of NK cells (natural killer cells) and helper T cells, and amounts of norepinephrine as an index of sympathetic activity and ACTH as an index of endocrine (HPA) activity. Their findings are fascinating:

“There were two main findings of the present study. First, low tonic vagal activity (low resting HRV) was associated with blunted responses in NK cells, norepinephrine, and ACTH to an acute stressor, whereas high tonic vagal activity (high resting HRV) was associated with more sensitive responses in those physiological parameters. Second, low and high tonic vagal activity was related to a qualitatively different neural matrix associated with immune, sympathetic, and endocrine changes. While low HRV participants showed only a correlation between ACTH and activity in the VLPFC, high HRV participants showed stronger associations between their brain activities and NK cells and ACTH. Specifically, in the high HRV participants, NK cell proportions were correlated with activity in the rostral ACC which is a portion of the MPFC and the dorsal striatum (nucleus caudate). The ACTH levels of the high HRV participants correlated with activation in the insula, OFC, cerebellum, and dorsal ACC. To the best of our knowledge, this is the first study to demonstrate that tonic vagal activity moderates brain–immune and brain–neuroendocrine associations accompanying acute stress.”

They discuss some of the important implications of their findings:

“Importantly, we observed that high HRV participants showed…associations between NK cell proportions and activity in [several brain regions]…By contrast, in low HRV participants, NK cell proportions showed no correlation with brain activity. These findings suggest that in individuals with high tonic vagal activity, immune responses to stress are associated with a higher and more complex regulatory neural network that…may enable regulation of NK-cell responses.”

Furthermore…

“The observation that individuals with high HRV initially showed a reduced NK cell response to an ongoing stressor during the initial learning task suggests that high HRV reflects the ability to habituate to stress. This is consistent with a previous finding by Weber et al. (2010) indicating that individuals with high HRV recovered cardiovascular, endocrine, and immune responses more rapidly after termination of an acute stressor than individuals with low HRV…By contrast, low HRV individuals demonstrated blunted immune, sympathetic, and endocrine reactivity to the stressor. These data suggest a greater physiological adaptability of in high HRV individuals and a potential moderating role of the vagus nerve in neuroimmuno-endocrine responses to stress.”

It was a similar story for the excitatory neurotransmitter norepinephrine:

Changes of norepinephrine due to stress showed a similar pattern to that in NK cells, with an initial decrease followed by increase after reversal of contingency between options and outcomes in the high HRV group, compared to a more blunted reactivity in the low HRV group.”

And for ACTH (adrenocorticotrophic hormone produced in the pituitary that stimulates adrenal production of cortisol):

Values of ACTH showed a continuous decrease in the high HRV group, reflecting a habituation process, but not in the low HRV group.”

For clinicians reading this who wisely do HRV assessments in their practice:

“…rating of subjective stress at baseline (before measurement of baseline HRV) did not differ between the low and high HRV groups. This suggest that baseline HRV, which was measured before the experimental procedure, might reflect the basic characteristics of an individuals’ vagal tone, rather than individual differences in phasic reactivity of HRV affected by anticipatory anxiety.”

In other words, this implies that heart rate variability assessments really does give us objective data about the patient’s vagal and parasympathetic resources. Other insights that emerge include:

low HRV participants had some impairment in the connections between the brain and peripheral physiology, with consequent differential patterns in physiological responses to the stressor…The high HRV group manifested greater sensitivity in their immune and physiological responses and greater association and possibly regullation by the brain over these responses. Although whether this is an adaptive response is an open question, it is possible that a high tonic vagal activity is a prerequisite for top-down rapid regulation of immune, autonomic, and endocrine responses to acute stress. By contrast, lower vagal activity may result in slower recovery (Weber et al., 2010) or lack of changes of immune responses to environmental challenges, possibly because of impairment in neuro-immune circuits.”

And regarding the premiere factor of inflammation:

An impaired regulation of immune responses can result in inflammation, which is etiologic to various chronic diseases, such as coronary-artery disease, cancer, and dementia, in which the vagus nerve was recently postulated to play a protective role via regulation of multiple basic processes (De Couck et al., 2012).”

Summing up their findings regarding vagal actvity as measured by HRV and the brain-immune response to stress:

“…our study revealed that tonic vagal nerve activity may be an important determinant of neuro- immune and neurophysiological associations and the regulation of the multisystem responses under acute stress.”

Since we can easily measure vagal (parasympathetic) tone in the clinic with HRV and we have sustainable interventions to increase vagal activity (BioCranial Therapy and many others), it’s hard to overemphasize the practical significance of this research.

Readers may also enjoy earlier posts on HRV including Nervous system regulation of inflammation, cytokines, and heart rate variability showing how vagal tone correlates with inflammatory cytokines in the bloodstream.