Life Extension Magazine®

Issue: Aug 2014


By Life Extension.

(-)-Epigallocatechin-3-gallate (EGCG) sensitizes melanoma cells to interferon induced growth inhibition in a mouse model of human melanoma.

Melanoma incidence has increased over the last few decades and metastatic melanoma is one of the hardest malignancies to treat. Thus, novel approaches are needed for an effective management of melanoma. Interferon-alpha2b (IFN), an immunomodulatory cytokine commonly used in melanoma treatment, has shown marginal efficacy and often results in discontinuation of therapy due to toxicity. We earlier demonstrated that epigallocatechin-3-gallate (EGCG), the major polyphenolic constituent of green tea, caused cell cycle arrest and apoptosis of human melanoma cells via modulation in cki-cyclin-cdk machinery and Bcl-2 family proteins. This study was undertaken to determine if EGCG could enhance the anti-proliferative effects of IFN. In this study, we demonstrated that EGCG and/or IFN treatments to melanoma cells resulted in a marked (1) decrease in cell proliferation and colony formation ability, and (2) induction of apoptosis. Interestingly, the combination was found to be more effective than either of the agents alone. Further, the anti-proliferative effects of EGCG and/or IFN were accompanied with an increase in Fas protein levels and a decrease in nuclear factor NFkappaB/p65 in the nucleus as well as NFkappaB promoter activity. EGCG and/or IFN also resulted in an increase in Fas-L mediated apoptosis. Further, EGCG and/or IFN treatments resulted in a decrease in melanoma tumor growth and protein levels of proliferation marker PCNA, in athymic nude mice implanted with melanoma tumors. The combination of the two modalities demonstrated a better response than either of them alone. Our data suggest that EGCG could impart therapeutic advantage if used in conjunction with IFN.

Cell Cycle. 2009 Jul 1;8(13):2057-63

Multifunctional effect of epigallocatechin-3-gallate (EGCG) in downregulation of gelatinase- A (MMP-2) in human breast cancer cell line MCF-7.

AIMS: The tumor inhibiting property of green tea polyphenol epigallocatechin-3-gallate (EGCG) is well documented. Studies reveal that matrix-metalloproteinases (MMPs) play pivotal roles in tumor invasion through degradation of basement membranes and extracellular matrix (ECM). We studied the effect of EGCG on matrixmetalloproteinases-2 (MMP-2), the factors involved in activation, secretion and signaling molecules that might be involved in the regulation of MMP-2 in human breast cancer cell line, MCF-7. MAIN METHODS: MCF-7 was treated with EGCG (20 muM, 24 h), the effect of EGCG on MMP-2 expression, activity and its regulatory molecules were studied by gelatin zymography, Western blot, quantitative and semi-quantitative real time RT-PCR, immunoflourescence and cell adhesion assay. KEY FINDINGS: EGCG treatment reduced the activity, protein expression and mRNA expression level of MMP-2. EGCG treatment reduced the expression of focal adhesion kinase (FAK), membrane type-1-matrix metalloproteinase (MT1-MMP), nuclear factor-kappa B (NF-kB), vascular endothelial growth factor (VEGF) and reduced the adhesion of MCF-7 cells to ECM, fibronectin and vitronectin. Real time RT-PCR revealed a reduced expression of integrin receptors alpha5, beta1, alphav and beta3 due to EGCG treatment. SIGNIFICANCE: Down regulation of expression of MT1-MMP, NF-kB, VEGF and disruption of functional status of integrin receptors may indicate decreased MMP-2 activation; low levels of FAK expression might indicate disruption in FAK-induced MMP-2 secretion and decrease in activation of phosphatidyl-inositol-3-kinase (PI-3K), extracellular regulated kinase (ERK) indicates probable hindrance in MMP-2 regulation and induction. We propose EGCG as potential inhibitor of expression and activity of pro-MMP-2 by a process involving multiple regulatory molecules in MCF-7.

Life Sci. 2009 Feb 13;84(7-8):194-204

Tea polyphenols inhibit cyclooxygenase-2 expression and block activation of nuclear factor-kappa B and Akt in diethylnitrosoamine induced lung tumors in Swiss mice.

BACKGROUND: Due to lack of validated screening methods and hence poor prognosis, treatment of lung cancer has not still improved up to the expectations. Therefore, risk of lung cancer needs to be minimized by efficient preventive measures. Tea (Camellia sinensis) and its bioactive polyphenols have been associated with prevention of human cancer for several organs. Thus, intake of tea polyphenols seems to be a viable mean to control lung cancer burden. In the present study, we studied the chemopreventive effects of green tea polyphenols (GTP) and black tea polyphenols (BTP) against diethylnitrosoamine (DEN) induced lung tumors in Swiss albino mice. RESULTS: Chemopreventive potential of tea polyphenols, was recorded as evident by, low incidence of alveologenic tumors in lungs of animals at tested doses (0.1% and 0.2% of both GTP and BTP) when compared with DEN (20 mg/kg b wt) treated animals. As a mechanism of cancer chemoprevention cellular signaling pathways were also targeted. GTP and BTP treatment inhibited the expression of Akt, cyclooxygenase-2 and inactivated nuclear factor-kappa B via blocking phosphorylation and subsequent degradation of IkappaB alpha. CONCLUSION: Thus, the study suggests that polyphenolic constituents of both cultivars of tea, i.e. green and black, have chemopreventive effects in DEN induced lung tumorigenesis in Swiss albino mice.

Invest New Drugs. 2010 Aug;28(4):466-71

Insights into antiamyloidogenic properties of the green tea extract (-)-epigallocatechin-3-gallate toward metal-associated amyloid-b species.

Despite the significance of Alzheimer’s disease, the link between metal-associated amyloid-b (metal-Ab) and disease etiology remains unclear. To elucidate this relationship, chemical tools capable of specifically targeting and modulating metal-Ab species are necessary, along with a fundamental understanding of their mechanism at the molecular level. Herein, we investigated and compared the interactions and reactivities of the green tea extract, (-)-epigallocatechin-3-gallate [(2R,3R)-5,7-dihydroxy-2-(3,4,5-trihydroxyphenyl)-3,4-dihydro-2H-1-benzopyran-3-yl 3,4,5-trihydroxybenzoate; EGCG], with metal [Cu(II) and Zn(II)]-Ab and metal-free Ab species. We found that EGCG interacted with metal-Ab species and formed small, unstructured Ab aggregates more noticeably than in metal-free conditions in vitro. In addition, upon incubation with EGCG, the toxicity presented by metal-free Ab and metal-Ab was mitigated in living cells. To understand this reactivity at the molecular level, structural insights were obtained by ion mobility-mass spectrometry (IM-MS), 2D NMR spectroscopy, and computational methods. These studies indicated that (i) EGCG was bound to Ab monomers and dimers, generating more compact peptide conformations than those from EGCG-untreated Ab species; and (ii) ternary EGCG-metal-Ab complexes were produced. Thus, we demonstrate the distinct antiamyloidogenic reactivity of EGCG toward metal-Ab species with a structure-based mechanism.

Proc Natl Acad Sci U S A. 2013 Mar 5;110 (10):3743-8

Neural effects of green tea extract on dorsolateral prefrontal cortex.

BACKGROUND/OBJECTIVES: Green tea is being recognized as a beverage with potential benefits for human health and cognitive functions. In vivo studies provide preliminary evidence that green tea intake may have a positive role in improving effects on cognitive functions. We aimed to examine the neural effects of green tea extract on brain activation in humans. SUBJECTS/METHODS: Functional magnetic resonance imaging was recorded while 12 healthy volunteers performed a working memory task following administration of 250 or 500 ml of a milk whey based green tea containing soft drink or milk whey based soft drink without green tea as control in a double-blind, controlled repeated measures within-subject design with counterbalanced order of substance administration. A whole-brain analysis with a cluster-level threshold of P<0.001 (unadjusted) was followed by an a priori-defined region of interest (ROI) analysis of the dorsolateral prefrontal cortex (DLPFC) including a cluster-level threshold of P<0.05 and family-wise error (FWE) adjustment for multiple comparisons. RESULTS: Whole-brain analyses revealed no significant effects after correction for multiple comparisons (FWE P<0.05). Using a ROI approach, green tea extract increased activation in the DLPFC relative to a control condition (FWE P<0.001). This neural effect was related to green tea dosage. Green tea extract was not associated with any significant attenuation in regional activation relative to control condition. CONCLUSIONS: These data suggest that green tea extract may modulate brain activity in the DLPFC, a key area that mediates working memory processing in the human brain. Moreover, this is the first neuroimaging study implicating that functional neuroimaging methods provide a means of examining how green tea extract acts on the brain.

Eur J Clin Nutr. 2012 Nov;66(11):1187-92

Green tea consumption and cognitive function: a cross-sectional study from the Tsurugaya Project 1.

BACKGROUND: Although considerable experimental and animal evidence shows that green tea may possess potent activities of neuroprotection, neurorescue, and amyloid precursor protein processing that may lead to cognitive enhancement, no human data are available. OBJECTIVE: The objective was to examine the association between green tea consumption and cognitive function in humans. DESIGN: We analyzed cross-sectional data from a community-based Comprehensive Geriatric Assessment (CGA) conducted in 2002. The subjects were 1003 Japanese subjects aged > or =70 y. They completed a self-administered questionnaire that included questions about the frequency of green tea consumption. We evaluated cognitive function by using the Mini-Mental State Examination with cutoffs of <28, <26, and <24 and calculated multivariate-adjusted odds ratios (ORs) of cognitive impairment. RESULTS: Higher consumption of green tea was associated with a lower prevalence of cognitive impairment. At the <26 cutoff, after adjustment for potential confounders, the ORs for the cognitive impairment associated with different frequencies of green tea consumption were 1.00 (reference) for < or =3 cups/wk, 0.62 (95% CI: 0.33, 1.19) for 4-6 cups/wk or 1 cup/d, and 0.46 (95% CI: 0.30, 0.72) for > or =2 cups/d (P for trend = 0.0006). Corresponding ORs were 1.00 (reference), 0.60 (95% CI: 0.35, 1.02), and 0.87 (95% CI: 0.55, 1.38) (P for trend = 0.33) for black or oolong tea and 1.00 (reference), 1.16 (95% CI: 0.78, 1.73), and 1.03 (95% CI: 0.59, 1.80) (P for trend = 0.70) for coffee. The results were essentially the same at cutoffs of <28 and <24. CONCLUSION: A higher consumption of green tea is associated with a lower prevalence of cognitive impairment in humans.

Am J Clin Nutr. 2006 Feb;83(2):355-61

Green tea epigallocatechin-3-gallate (EGCG) promotes neural progenitor cell proliferation and sonic hedgehog pathway activation during adult hippocampal neurogenesis.

SCOPE: Adult hippocampal neurogenesis is a lifelong feature of brain plasticity that appears to be critically involved in adult brain function and neurological disease. Recent studies suggest that (-)-epigallocatechin-3-gallate (EGCG), which is the main polyphenolic constituent of green tea, may be used for the prevention and treatment of various neurodegenerative diseases. We hypothesized that EGCG promotes adult neurogenesis, which may be beneficial to hippocampus-dependent learning and memory. METHODS AND RESULTS: We show that EGCG treatment significantly increased the number of 5-bromo-2’-deoxyuridine (BrdU)-labeled cells in adult hippocampal neural progenitor cell (NPC) cultures and in the dentate gyrus of adult mice. Meanwhile, EGCG markedly improved spatial cognition in mice. These events are associated with the sonic hedgehog (Shh) signaling pathway. We observed that EGCG triggered a robust upregulation of Shh receptor (Patched) mRNA and protein expression in cultured NPCs as well as an upregulation of the downstream Shh transcriptional target Gli1. These changes were further confirmed in the hippocampus of mice administered EGCG. The blockage of the Shh signal with the pharmacological inhibitor cyclopamine attenuated EGCG-induced hippocampal neurogenesis. CONCLUSION: Our results provide strong evidence that EGCG enhances adult hippocampal neurogenesis.

Mol Nutr Food Res. 2012 Aug;56(8):1292-303

Green tea consumption and the risk of incident functional disability in elderly Japanese: the Ohsaki Cohort 2006 Study.

BACKGROUND: Previous studies have reported that green tea consumption is associated with a lower risk of diseases that cause functional disability, such as stroke, cognitive impairment, and osteoporosis. Although it is expected that green tea consumption would lower the risk of incident functional disability, this has never been investigated directly. OBJECTIVE: The objective was to determine the association between green tea consumption and incident functional disability in elderly individuals. DESIGN: We conducted a prospective cohort study in 13,988 Japanese individuals aged ≥65 y. Information on daily green tea consumption and other lifestyle factors was collected via questionnaire in 2006. Data on functional disability were retrieved from the public Long-term Care Insurance database, in which subjects were followed up for 3 y. We used Cox proportional hazards regression analysis to investigate the association between green tea consumption and functional disability. RESULTS: The 3-y incidence of functional disability was 9.4% (1316 cases). The multiple-adjusted HR (95% CI) of incident functional disability was 0.90 (0.77, 1.06) among respondents who consumed 1-2 cups green tea/d, 0.75 (0.64, 0.88) for those who consumed 3-4 cups/d, and 0.67 (0.57, 0.79) for those who consumed ≥5 cups/d in comparison with those who consumed <1 cup/d (P-trend < 0.001). CONCLUSION: Green tea consumption is significantly associated with a lower risk of incident functional disability, even after adjustment for possible confounding factors.

Am J Clin Nutr. 2012 Mar;95(3):732-9

Recent findings of green tea extract AR25 (Exolise) and its activity for the treatment of obesity.

The green tea extract AR25 is an 80% ethanolic dry extract standardized at 25% catechins expressed as epigallocatechin gallate (EGCG). In vitro, green tea extract AR25 exerts a direct inhibition of gastric and pancreatic lipases and a stimulation of thermogenesis. In an open study, the effects of extract AR25 were evaluated in moderately obese patients. After 3 months, body weight was decreased by 4.6% and waist circumference by 4.48%. These results suggest the green tea extract AR25 to be a natural product for the treatment of obesity, which exerts its activity by several ways: inhibition of lipases and stimulation of thermogenesis.

Phytomedicine. 2002 Jan;9(1):3-8

Mechanisms of hypolipidemic and anti-obesity effects of tea and tea polyphenols.

Among the health-promoting effects of tea and tea polyphenols, the cancer-chemopreventive effects in various animal model systems have been intensively investigated; meanwhile, the hypolipidemic and antiobesity effects in animals and humans have also become a hot issue for molecular nutrition and food research. It has been demonstrated that the body weights of rats and their plasma triglyceride, cholesterol, and LDL-cholesterol have been significantly reduced by feedings of oolong, black, pu-erh, and green tea leaves to the animals. It has been suggested that the inhibition of growth and suppression of lipogenesis in MCF-7 breast cancer cells may be through down-regulation of fatty acid synthase gene expression in the nucleus and stimulation of cell energy expenditure in the mitochondria. The experimental data indicated that the molecular mechanisms of fatty acid synthase gene suppression by tea polyphenols (EGCG, theaflavins) may invite down-regulation of EGFR/PI3K/Akt/Sp-1 signal transduction pathways.

Mol Nutr Food Res. 2006 Feb;50(2):211-7

Effects of licorice on relief and recurrence of menopausal hot flashes.

Vasomotor hot flash is the most common and distressful complication of menopausal women. Its treatment is the most frequent clinical challenge. As a result, an effective and harmless therapy is needed. This double-blind controlled clinical trial was conducted to determine the effects of licorice roots on the relief and recurrence of hot flash in menopausal women referring to the healthcare centers affiliated to Shahid Beheshti Medical University in 2010. Ninety menopausal women complaining of hot flash were selected by reviewing their records in healthcare centers and randomly divided into 2 licorices (3 capsules daily containing 330 mg licorice abstract) and placebo (3 capsules daily containing 330 mg starch) groups over the 8 weeks of intervention and 4 weeks of follow-up. Two weeks prior to the intervention, the severity as well as frequency of hot flashes and the foods taken were asked and documented with questionnaires and data sheets. Data within and between the groups were analyzed by ANOVA with repeated measurements and t-test respectively. Means of age and body mass index (BMI) of the subjects in licorice and placebo groups were 53 ± 3.2, 52.69 ± 2.8, 24.71 ± 3.2 and 23.61 ± 3.3, respectively. The groups were similar in terms of intervening variables. The frequency of hot flash decreased significantly in the experimental (than the placebo group) and this lasted for 2 weeks after the administration of the capsules. The severity of hot flash decreased in the licorice group as well. This decrease was also seen in the placebo group in the first week of the intervention. Decreased hot flash in the placebo group was only significant after the 1(st) week of intervention compared to the previous period. Recurrence of frequency and severity of hot flashes occurred 2 weeks after the termination of therapy. The significant decrease in the placebo group after the 1(st) week of the intervention may be attributed to the psychological effects of placebo. Licorice roots decreased the frequency and severity of hot flashes. The administration of this harmless, inexpensive herb well accepted by the menopausal women together with the appropriate and continuous physical activities and consumption of dairy products are recommended for relieving this complication.

Iran J Pharm Res. 2012 Spring;11(2):541-8

The effects of compounded bioidentical transdermal hormone therapy on hemostatic, inflammatory, immune factors; cardiovascular biomarkers; quality-of-life measures; and health outcomes in perimenopausal and postmenopausal women.

Menopause impacts 25 million women world wide each year, and the World Health Organization estimates 1.2 billion women will be postmenopausal by 2030. Menopause has been associated with symptoms of hot flashes, night sweats, dysphoric mood, sleep disturbance, and conditions of cardiovascular disease, depression, osteoporosis, osteoarthritis, depression, dementia, and frailty. Conventional hormone replacement therapy results in increased thrombotic events, and an increased risk of breast cancer and dementia as evidenced in large prospective clinical trials including Heart and Estrogen/Progestin Replacement Study I and the Women’s Health Initiative. A possible mechanism for these adverse events is the unfavorable net effects of conjugated equine estrogens and medroxyprogesterone acetate on the hemostatic balance and inflammatory and immune factors. Physiologic sex steroid therapy with transdermal delivery for peri/postmenopausal women may offer a different risk/benefit profile, yet long-term studies of this treatment model are lacking. The objective of this study was to examine the long-term effects of compounded bioidentical transdermal sex steroid therapy including estriol, estradiol, progesterone, DHEA, and testosterone on cardiovascular biomarkers, hemostatic, inflammatory, immune signaling factors; quality-of-life measures; and health outcomes in peri/postmenopausal women within the context of a hormone restoration model of care. A prospective, cohort, closed-label study received approval from the Human Subjects Committee. Recruitment from outpatient clinics at an academic medical center and the community at large resulted in three hundred women giving signed consent. Seventy-five women who met strict inclusion/exclusion criteria were enrolled. Baseline hormone evaluation was performed along with baseline experimental measures. Following this, women received compounded transdermal bioidentical hormone therapy of BiEst (80%Estriol/20%Estradiol), and/or Progesterone for eight weeks to meet established physiologic reference ranges for the luteal phase in premenopausal women. The luteal phase hormone ratios were selected based on animal and epidemiologic studies demonstrating favorable outcomes related to traumatic, ischemic, or neuronal injury. Follow-up testing was performed at eight weeks and adjustment to hormone regimens were made including addition of androgens of DHEA and Testosterone if indicated. Experimental subjects were monitored for 36 months. Baseline, 2-month, and annual values were obtained for: blood pressure, body mass index, fasting glucose, Homeostasis Metabolic Assessment of Insulin Resistance (HOMA-IR), fasting triglycerides, total Factor VII, Factor VIII, fibrinogen, Antithrombin III, Plasminogen Activator Inhibitor1(PAL-1), C-reactive protein (CRP), Interleukin-6 (IL-6), Matrix Metalloproteinase-9 (MMP-9), Tumor Necrosis Factor-alpha (TNF), Insulin-like Growth Factor (IGF-1), and sex steroid levels. Psychosocial measures included: Greene Climacteric Scale, Visual Analog Pain Scale, Hamilton Anxiety Scale, Hamilton Depression Scale, Holmes Rahe Stress Scale, Job Strain, and Home Strain. Health outcome measures included the number of prescribed medications used, number of co-morbidities, and endometrial thickness in postmenopausal women with intact uteri. Subjects receiving compounded transdermal bioidentical hormone therapy showed significant favorable changes in: Greene Climacteric Scale scores, Hamilton Anxiety Scale, Hamilton Depression Scale, Visual Analog Pain Scale, fasting glucose, fasting triglycerides, MMP-9, C-reactive Protein, fibrinogen, Factor VII, Factor VIII, Insulin-Like Growth Factor 1, and health outcomes of co-morbidities and a number of prescribed medications. Antithrombin III levels were significantly decreased at 36 months. All other measures did not exhibit significant effects. Administration of compounded transdermal bioidentical hormone therapy in doses targeted to physiologic reference ranges administered in a daily dose significantly relieved menopausal symptoms in peri/postmenopausal women. Cardiovascular biomarkers, inflammatory factors, immune signaling factors, and health outcomes were favorably impacted, despite very high life stress, and home and work strain in study subjects. The therapy did not adversely alter the net prothrombotic potential, and there were no associated adverse events. This model of care warrants consideration as an effective and safe clinical therapy for peri/postmenopausal women especially in populations with high perceived stress and a history of stressful life events prior to, or during the menopausal transition.

Int J Pharm Compd. 2013 Jan-Feb;17(1):74-85

Risk of long-term hot flashes after natural menopause: evidence from the Penn Ovarian Aging Study cohort.

OBJECTIVE: This study aims to estimate the risk of hot flashes relative to natural menopause and to evaluate the associations of hormone levels, behavioral variables, and demographic variables with the risk of hot flashes after menopause. METHODS: We performed annual assessment of 255 women who were premenopausal at baseline and reached natural menopause within 16 years of follow-up. RESULTS: The prevalence of moderate/severe hot flashes increased in each premenopausal year, reaching a peak of 46% in the first 2 years after the final menstrual period (FMP). Hot flashes decreased slowly after menopause and did not return to premenopausal levels until 9 years after the FMP. The mean (SD) duration of moderate/severe hot flashes after the FMP was 4.6 (2.9) years (for any hot flashes, 4.9 [3.1] y). One third of women at 10 years or more after menopause continued to experience moderate/severe hot flashes. African-American women (obese and nonobese) and obese white women had significantly greater risks of hot flashes compared with nonobese white women (interaction, P = 0.01). In multivariable analysis, increasing follicle-stimulating hormone levels before the FMP (P < 0.001), decreasing estradiol (odds ratio, 0.87; 95% CI, 0.78-0.96; P = 0.008), and increasing anxiety (odds ratio, 1.05; 95% CI, 1.03-1.06; P < 0.001) were significant risk factors for hot flashes, whereas higher education levels were protective (odds ratio, 0.66; 95% CI, 0.47-0.91; P = 0.011). CONCLUSIONS: Moderate/severe hot flashes continue, on average, for nearly 5 years after menopause; more than one third of women observed for 10 years or more after menopause have moderate/severe hot flashes. Continuation of hot flashes for more than 5 years after menopause underscores the importance of determining individual risks/benefits when selecting hormone or nonhormone therapy for menopausal symptoms.

Menopause. 2014 Jan 27

Evaluation of contextual and demographic factors on licorice effects on reducing hot flashes in postmenopause women.

Menopause is an important stage in the life of every woman. Hot flashes are the most common climacteric symptom and a major cause of suffering in postmenopausal women. Licorice is one of the plants that is used to relieve menopausal symptoms. The present study was undertaken to evaluate the effects of licorice on hot flash symptoms in menopausal women. The participants of this randomized, double blind, clinical trial study were 60 menopausal women randomly allocated to licorice or hormone replacement therapy (HRT) groups. The participants in this trial received licorice (1,140 mg/day) or HRT (a conjugated estrogen 0.312 mg/day and Medroxyprogesterone 2.5 mg/day) for 90 days. In this study we observed that licorice is not very different from hormones in terms of reducing the number and duration of hot flashes, but that HRT can reduce the severity of hot flashes significantly better than licorice can. In addition, there was no significant difference between age, education level, marital status, occupation, income, number of pregnancies, time from cessation of menstruation, and severity of hot flashes in the two groups. We observed that licorice seems more effective than HRT in improving hot flash duration, but that HRT can reduce the duration and severity of hot flashes more than licorice.

Health Care Women Int. 2014 Jan;35(1):87-99

Menstruation and the menopausal transition.

This paper characterizes changes in menstrual bleeding during perimenopause,including bleeding changes that represent markers of the menopausal transition. Recent results from the Study of Women’s Health Across the Nation and other cohort studies are reviewed. Emerging data describing subpopulation differences in the transition experience are highlighted. When treating women in the midlife, clinicians should pay careful attention to medical factors, including both conditions and treatment, that may increase menstrual blood loss or alter menstrual cycle characteristics sufficiently to obscure the onset of the menopausal transition or the final menstrual period.

Obstet Gynecol Clin North Am. 2011 Sep;38 (3):595-607

Menopausal hormone therapy for the primary prevention of chronic conditions. U.S. Preventive Services Task Force recommendation statement.

Since the early 20th century, scientists have been tantalized with the hypothesis that premenopausal health benefits in women can be preserved in postmenopausal women with the supplementation of exogenous hormone replacement therapy (HRT) of estrogen (alone/with progesterone). This hypothesis was shattered when the results of 2 large randomized controlled trials (RCTs), the Heart Estrogen/Progesterone Replacement Study (HERS) and Women’s Health Initiative (WHI), reported an increased risk of adverse clinical outcomes including coronary heart disease, thromboembolic events, stroke, dementia, urinary incontinence, gallbladder disease, and breast cancer. However, since the WHI was published, firestorms of critique, controversy, and multiple subgroup analyses have populated the medical literature, predominantly focused around the analysis of the age of women at entry into the trials (hypothesized as an effect modifier) and suggesting lower-dose preparations including using bioidentical hormones. Recently, the U.S. Preventive Services Task Force (USPSTF) along with other professional groups have issued recommendations against the use of HRT to prevent chronic conditions. In this review, we review the most recent evidence, including the long-term follow-up data from RCTs along a multitude of health outcomes.

Pol Arch Med Wewn. 2013;123(3):112-7

Hop extracts and hop substances in treatment of menopausal complaints.

Hop extract is a long used medicinal product and, regarding hormonal activities, in 1999 a number of prenylflavanones have been identified as its major constituents with 8-prenylnaringenin (8-PN) being the main active estrogenic compound. There have been several in vivo studies performed that demonstrate the potential of hop extract and the single compound 8-PN to alleviate climacteric symptoms like osteoporosis, vasomotoric complaints, and sexual motivation. On the other hand, only a few clinical studies have been performed so far, and these mainly focused on menopausal discomforts, especially hot flushes, yielding rather inconclusive results. Despite preferentially activating estrogen receptor a, 8-PN is only slightly uterotrophic, but it also elucidates estrogenic effects on the mammary gland. In conclusion, although hop extract and especially 8-PN are promising candidates as a relief for climacteric symptoms, data on the safety and efficacy is still scarce.

Planta Med. 2013 May;79(7):576-9

Identification of a potent phytoestrogen in hops (Humulus lupulus L.) and beer.

The female flowers of the hop plant are used as a preservative and as a flavoring agent in beer. However, a recurring suggestion has been that hops have a powerful estrogenic activity and that beer may also be estrogenic. In this study, sensitive and specific in vitro bioassays for estrogens were used for an activity-guided fractionation of hops via selective solvent extraction and appropriate HPLC separation. We have identified a potent phytoestrogen in hops, 8-prenylnaringenin, which has an activity greater than other established plant estrogens. The estrogenic activity of this compound was reflected in its relative binding affinity to estrogen receptors from rat uteri. The presence of 8-prenylnaringenin in hops may provide an explanation for the accounts of menstrual disturbances in female hop workers. This phytoestrogen can also be detected in beer, but the levels are low and should not pose any cause for concern.

J Clin Endocrinol Metab. 1999 Jun;84 (6):2249-52

EST analysis of hop glandular trichomes identifies an O-methyltransferase that catalyzes the biosynthesis of xanthohumol.

The glandular trichomes (lupulin glands) of hop (Humulus lupulus) synthesize essential oils and terpenophenolic resins, including the bioactive prenylflavonoid xanthohumol. To dissect the biosynthetic processes occurring in lupulin glands, we sequenced 10,581 ESTs from four trichome-derived cDNA libraries. ESTs representing enzymes of terpenoid biosynthesis, including all of the steps of the methyl 4-erythritol phosphate pathway, were abundant in the EST data set, as were ESTs for the known type III polyketide synthases of bitter acid and xanthohumol biosynthesis. The xanthohumol biosynthetic pathway involves a key O-methylation step. Four S-adenosyl-l-methionine-dependent O-methyltransferases (OMTs) with similarity to known flavonoid-methylating enzymes were present in the EST data set. OMT1, which was the most highly expressed OMT based on EST abundance and RT-PCR analysis, performs the final reaction in xanthohumol biosynthesis by methylating desmethylxanthohumol to form xanthohumol. OMT2 accepted a broad range of substrates, including desmethylxanthohumol, but did not form xanthohumol. Mass spectrometry and proton nuclear magnetic resonance analysis showed it methylated xanthohumol to 4-O-methylxanthohumol, which is not known from hop. OMT3 was inactive with all substrates tested. The lupulin gland-specific EST data set expands the genomic resources for H. lupulus and provides further insight into the metabolic specialization of glandular trichomes.

Plant Cell. 2008 Jan;20(1):186-200

Morphologic changes induced by oral long-term treatment with 8-prenylnaringenin in the uterus, vagina, and mammary gland of castrated rats.

OBJECTIVE: The flavonoid 8-prenylnaringenin (8-PN) is found in hops, and hence in beer, and is also increasingly consumed as a food supplement. It is the strongest known phytoestrogen, which makes it a good candidate as an alternative to hormone therapy. Its putatively undesired estrogenic effects in the uterus and mammary gland have not yet been thoroughly investigated. Therefore, we performed a long-term oral administration experiment. DESIGN: Rats were ovariectomized and fed for 3 months with soy-free chow containing estradiol (E(2)) or 8-PN, both in two doses (8-PN: 6.77 mg or 68.42 mg/kg body weight; E(2): 0.17 mg or 0.7 mg/kg body weight) or no additives. Analysis was mainly focused on morphologic and immunocytochemical parameters. Expression of proliferating cell nuclear antigen as a proliferation marker and of progesterone receptor was quantified in the mammary gland. RESULTS: Uteri of animals treated with both E(2) doses and the high 8-PN dose had increased weight and showed histologic estrogen-induced features. 8-PN at the high dose induced epithelial polypoid formation unique to this group. Compared to the atrophic controls, both E(2) doses and the high 8-PN dose induced hyperplastic epithelia in the vagina. The high doses of E(2) and 8-PN caused secretion in the mammary gland, whereas proliferation and progesterone receptor expression were stimulated by both E(2) doses and the high 8-PN dose. CONCLUSIONS: E(2) and 8-PN share many effects in the three studied organs, but some differences in the mechanism of action appear to exist

Menopause. 2006 Jul-Aug;13(4):669-77

Vitamin E d-tocotrienol prolongs survival in the LSL-KrasG12D/+;LSL-Trp53R172H/+;Pdx-1-Cre (KPC) transgenic mouse model of pancreatic cancer.

Previous work has shown that vitamin E d-tocotrienol (VEDT) prolongs survival and delays progression of pancreatic cancer in the LSL-Kras(G12D)(/+);Pdx-1-Cre mouse model of pancreatic cancer. However, the effect of VEDT alone or in combination with gemcitabine in the more aggressive LSL-Kras(G12D)(/+);LSL-Trp53(R172H)(/+);Pdx-1-Cre (KPC) mouse model is unknown. Here, we studied the effects of VEDT and the combination of VEDT and gemcitabine in the KPC mice. KPC mice were randomized into four groups: (i) vehicle [olive oil, 1.0 mL/kg per os twice a day and PBS 1.0 mL/kg intrapertoneally (i.p.) twice a week], (ii) gemcitabine (100 mg/kg i.p. twice a week), (iii) VEDT (200 mg/kg per os twice a day), and (iv) gemcitabine + VEDT. Mice received treatment until they displayed symptoms of impending death from pancreatic cancer, at which point animals were euthanized. At 16 weeks, survival was 10% in the vehicle group, 30% in the gemcitabine group, 70% in the VEDT group (P < 0.01), and 90% in the VEDT combined with gemcitabine group (P < 0.05). VEDT alone and combined with gemcitabine resulted in reversal of epithelial-to-mesenchymal transition in tumors. Biomarkers of apoptosis (plasma CK18), PARP1 cleavage, and Bax expression were more greatly induced in tumors subjected to combined treatment versus individual treatment. Combined treatment induced cell-cycle inhibitors (p27(Kip1) and p21(Cip1)) and inhibited VEGF, vascularity (CD31), and oncogenic signaling (pAKT, pMEK, and pERK) greater than individual drugs. No significant differences in body weight gain between drug treatment and control mice were observed. These results strongly support further investigation of VEDT alone and in combination with gemcitabine for pancreatic cancer prevention and treatment.

Cancer Prev Res (Phila). 2013 Oct;6(10):1074-83

Palm tocotrienol-rich fraction reduced plasma homocysteine and heart oxidative stress in rats fed with a high-methionine diet.

Oxidative stress contributes to cardiovascular diseases. We aimed to study the effects of palm tocotrienol-rich fraction (TRF) on plasma homocysteine and cardiac oxidative stress in rats fed with a high-methionine diet. Forty-two male Wistar rats were divided into six groups. The first group was the control. Groups 2-6 were fed 1% methionine diet for 10 weeks. From week 6 onward, folate (8 mg/kg diet) or palm TRF (30, 60 and 150 mg/kg diet) was added into the diet of groups 3, 4, 5 and 6. The rats were then killed. Palm TRF at 150 mg/kg and folate supplementation prevented the increase in plasma total homocysteine (4.14 ± 0.33 and 4.30 ± 0.26 vs 5.49 ± 0.25 mmol/L, p < 0.05) induced by a high-methionine diet. The increased heart thiobarbituric acid reactive substance in rats fed with high-methionine diet was also prevented by the supplementations of palm TRF (60 and 150 mg/kg) and folate. The high-methionine group had a lower glutathione peroxidase activity (49 ± 3 vs 69 ± 4 pmol/mg protein/min) than the control group. This reduction was reversed by palm TRF at 60 and 150 mg/kg diet (p < 0.05), but not by folate. Catalase and superoxide dismutase activities were unaffected by both methionine and vitamin supplementations. In conclusion, palm TRF was comparable to folate in reducing high-methionine diet-induced hyperhomocysteinemia and oxidative stress in the rats’ hearts. However, palm TRF was more effective than folate in preserving the heart glutathione peroxidase enzyme activity.

J Physiol Biochem. 2013 Sep;69(3):441-9

Natural vitamin E a-tocotrienol protects against ischemic stroke by induction of multidrug resistance-associated protein 1.

BACKGROUND AND PURPOSE: a-Tocotrienol (TCT) represents the most potent neuroprotective form of natural vitamin E that is Generally Recognized As Safe certified by the U.S. Food and Drug Administration. This work addresses a novel molecular mechanism by which a-TCT may be protective against stroke in vivo. Elevation of intracellular oxidized glutathione (GSSG) triggers neural cell death. Multidrug resistance-associated protein 1 (MRP1), a key mediator of intracellular oxidized glutathione efflux from neural cells, may therefore possess neuroprotective functions. METHODS: Stroke-dependent brain tissue damage was studied in MRP1-deficient mice and a-TCT-supplemented mice. RESULTS: Elevated MRP1 expression was observed in glutamate-challenged primary cortical neuronal cells and in stroke-affected brain tissue. MRP1-deficient mice displayed larger stroke-induced lesions, recognizing a protective role of MRP1. In vitro, protection against glutamate-induced neurotoxicity by a-TCT was attenuated under conditions of MRP1 knockdown; this suggests the role of MRP1 in a-TCT-dependent neuroprotection. In vivo studies demonstrated that oral supplementation of a-TCT protected against murine stroke. MRP1 expression was elevated in the stroke-affected cortical tissue of a-TCT-supplemented mice. Efforts to elucidate the underlying mechanism identified MRP1 as a target of microRNA (miR)-199a-5p. In a-TCT-supplemented mice, miR-199a-5p was downregulated in stroke-affected brain tissue. CONCLUSIONS: This work recognizes MRP1 as a protective factor against stroke. Furthermore, findings of this study add a new dimension to the current understanding of the molecular bases of a-TCT neuroprotection in 2 ways: by identifying MRP1 as a a-TCT-sensitive target and by unveiling the general prospect that oral a-TCT may regulate miR expression in stroke-affected brain tissue.

Stroke. 2011 Aug;42(8):2308-14

Tocotrienol vitamin E protects against preclinical canine ischemic stroke by inducing arteriogenesis.

Vitamin E consists of tocopherols and tocotrienols, in which a-tocotrienol is the most potent neuroprotective form that is also effective in protecting against stroke in rodents. As neuroprotective agents alone are insufficient to protect against stroke, we sought to test the effects of tocotrienol on the cerebrovascular circulation during ischemic stroke using a preclinical model that enables fluoroscopy-guided angiography. Mongrel canines (mean weight=26.3±3.2 kg) were supplemented with tocotrienol-enriched (TE) supplement (200 mg b.i.d, n=11) or vehicle placebo (n=9) for 10 weeks before inducing transient middle cerebral artery (MCA) occlusion. Magnetic resonance imaging was performed 1 hour and 24 hours post reperfusion to assess stroke-induced lesion volume. Tocotrienol-enriched supplementation significantly attenuated ischemic stroke-induced lesion volume (P<0.005). Furthermore, TE prevented loss of white matter fiber tract connectivity after stroke as evident by probabilistic tractography. Post hoc analysis of cerebral angiograms during MCA occlusion revealed that TE-supplemented canines had improved cerebrovascular collateral circulation to the ischemic MCA territory (P<0.05). Tocotrienol-enriched supplementation induced arteriogenic tissue inhibitor of metalloprotease 1 and subsequently attenuated the activity of matrix metalloproteinase-2. Outcomes of the current preclinical trial set the stage for a clinical trial testing the effects of TE in patients who have suffered from transient ischemic attack and are therefore at a high risk for stroke.

J Cereb Blood Flow Metab. 2011 Nov;31 (11):2218-30

Tocotrienol supplementation in postmenopausal osteoporosis: evidence from a laboratory study.

OBJECTIVE: Accelerated bone loss that occurs in postmenopausal women has been linked to oxidative stress and increased free radicals. We propose the use of antioxidants to prevent and reverse postmenopausal osteoporosis. This study aimed to examine the effects of tocotrienol, a vitamin E analog, on bone loss due to estrogen deficiency. Our previous study showed that tocotrienol increased the trabecular bone volume and trabecular number in ovariectomized rats. In the current study, we investigated the effects of tocotrienol supplementation on various biochemical parameters in a postmenopausal osteoporosis rat model. MATERIALS AND METHODS: A total of 32 female Wistar rats were randomly divided into four groups. The baseline group was sacrificed at the start of the study, and another group was sham operated. The remaining rats were ovariectomized and either given olive oil as a vehicle or treated with tocotrienol at a dose of 60 mg/kg body weight. After four weeks of treatment, blood was withdrawn for the measurement of interleukin-1 (IL1) and interleukin-6 (IL6) (bone resorbing cytokines), serum osteocalcin (a bone formation marker) and pyridinoline (a bone resorption marker). RESULTS: Tocotrienol supplementation in ovariectomized rats significantly reduced the levels of osteocalcin, IL1 and IL6. However, it did not alter the serum pyridinoline level. CONCLUSION: Tocotrienol prevented osteoporotic bone loss by reducing the high bone turnover rate associated with estrogen deficiency. Therefore, tocotrienol has the potential to be used as an anti-osteoporotic agent in postmenopausal women.

Clinics (Sao Paulo). 2013 Oct;68(10):1338-43

Tocotrienols in health and disease: the other half of the natural vitamin E family.

Tocochromanols encompass a group of compounds with vitamin E activity essential for human nutrition. Structurally, natural vitamin E includes eight chemically distinct molecules: alpha-, beta-, gamma- and delta-tocopherol; and alpha-, beta-, gamma- and delta-tocotrienol. Symptoms caused by alpha-tocopherol deficiency can be alleviated by tocotrienols. Thus, tocotrienols may be viewed as being members of the natural vitamin E family not only structurally but also functionally. Palm oil and rice bran oil represent two major nutritional sources of natural tocotrienol. Taken orally, tocotrienols are bioavailable to all vital organs. The tocotrienol forms of natural vitamin E possesses powerful hypocholesterolemic, anti-cancer and neuroprotective properties that are often not exhibited by tocopherols. Oral tocotrienol protects against stroke-associated brain damage in vivo. Disappointments with outcomes-based clinical studies testing the efficacy of alpha-tocopherol need to be handled with caution and prudence recognizing the untapped opportunities offered by the other forms of natural vitamin E. Although tocotrienols represent half of the natural vitamin E family, work on tocotrienols account for roughly 1% of the total literature on vitamin E. The current state of knowledge warrants strategic investment into investigating the lesser known forms of vitamin E.

Mol Aspects Med. 2007 Oct-Dec;28(5-6):692-728

Tocotrienols: Vitamin E beyond tocopherols.

In nature, eight substances have been found to have vitamin E activity: alpha-, beta-, gamma- and delta-tocopherol; and alpha-, beta-, gamma- and delta-tocotrienol. Yet, of all papers on vitamin E listed in PubMed less than 1% relate to tocotrienols. The abundance of alpha-tocopherol in the human body and the comparable efficiency of all vitamin E molecules as antioxidants, led biologists to neglect the non-tocopherol vitamin E molecules as topics for basic and clinical research. Recent developments warrant a serious reconsideration of this conventional wisdom. Tocotrienols possess powerful neuroprotective, anti-cancer and cholesterol lowering properties that are often not exhibited by tocopherols. Current developments in vitamin E research clearly indicate that members of the vitamin E family are not redundant with respect to their biological functions. alpha-Tocotrienol, gamma-tocopherol, and delta-tocotrienol have emerged as vitamin E molecules with functions in health and disease that are clearly distinct from that of alpha-tocopherol. At nanomolar concentration, alpha-tocotrienol, not alpha-tocopherol, prevents neurodegeneration. On a concentration basis, this finding represents the most potent of all biological functions exhibited by any natural vitamin E molecule. An expanding body of evidence support that members of the vitamin E family are functionally unique. In recognition of this fact, title claims in manuscripts should be limited to the specific form of vitamin E studied. For example, evidence for toxicity of a specific form of tocopherol in excess may not be used to conclude that high-dosage “vitamin E” supplementation may increase all-cause mortality. Such conclusion incorrectly implies that tocotrienols are toxic as well under conditions where tocotrienols were not even considered. The current state of knowledge warrants strategic investment into the lesser known forms of vitamin E. This will enable prudent selection of the appropriate vitamin E molecule for studies addressing a specific need.

Life Sci. 2006 Mar 27;78(18):2088-98

Cancer preventive effects of vitamin E.

Vitamin E is well known as an antioxidant, with 8 natural isoforms, such as a-, bgr;-, g- and d-tocopherols and a-, b-, gamma;- and d-tocotrienols. It has been suggested that both tocopherols and tocotrienols have anti-tumor effects due to the antioxidant effect. The results of several studies have indicated that the tocotrienols may have a stronger bioactivity than the tocopherols. Both types have shown antiproliferative, proapoptotic and cyclooxygenase-2- inhibiting effects in in vitro studies. Several animal studies have demonstrated that vitamin E has cancer-preventing effects. However, clinical trials have not shown similar results for the cancer prevention effect of tocopherol. Although the Linxian Trials demonstrated that the supplementation of b-carotene, a-tocopherol and selenium reduced cancer risk, the beneficial effects of a- tocopherol on prostate cancer disappeared after several years in the Alpha-Tocopherol, Beta-Carotene Cancer Prevention Study. Vitamin E, especially tocotrienols, seems to be a potent agent for cancer prevention, however no large-scale clinical trial on the cancer prevention effect of tocotrienols has been conducted yet. Therefore it is expected that clinical trials overcoming the lower bioavailability of tocotrienols will be conducted, and it is urgently needed to assess the safety and the efficacy of the administration of the tocotrienols as a part of a cancer prevention regimen.

Curr Pharm Biotechnol. 2012 Jan;13(1):156-64

Tocotrienol as a potential anticancer agent.

Vitamin E is composed of two structurally similar compounds: tocopherols (TPs) and tocotrienols (T3). Despite being overshadowed by TP over the past few decades, T3 is now considered to be a promising anticancer agent due to its potent effects against a wide range of cancers. A growing body of evidence suggests that in addition to its antioxidative and pro-apoptotic functions, T3 possesses a number of anticancer properties that make it superior to TP. These include the inhibition of epithelial-to-mesenchymal transitions, the suppression of vascular endothelial growth factor tumor angiogenic pathway and the induction of antitumor immunity. More recently, T3, but not TP, has been shown to have chemosensitization and anti-cancer stem cell effects, further demonstrating the potential of T3 as an effective anticancer therapeutic agent. With most of the previous clinical studies on TP producing disappointing results, research has now focused on testing T3 as the next generation vitamin E for chemoprevention and cancer treatment. This review will summarize recent developments in the understanding of the anticancer effects of T3. We will also discuss current progress in clinical trials involving T3 as an adjuvant to conventional cancer therapy.

Carcinogenesis. 2012 Feb;33(2):233-9

Potential role of tocotrienols in the treatment and prevention of breast cancer.

Vitamin E is a generic term that refers to a family of compounds that is further divided into two subgroups called tocopherols and tocotrienols. Although all natural forms of vitamin E display potent antioxidant activity, tocotrienols are significantly more potent than tocopherols in inhibiting tumor cell growth and viability, and anticancer activity of tocotrienols is mediated independently of their antioxidant activity. In addition, the anticancer effects of tocotrienols are observed using treatment doses that have little or no effect on normal cell function or viability. This review will summarize experimental studies that have identified the intracellular mechanism mediating the anticancer effects of tocotrienols. Evidence is also provided showing that combined treatment of tocotrienol with other cancer chemotherapies can result in a synergistic inhibition in cancer cell growth and viability. Taken together, these findings strongly indicate that tocotrienols may provide significant health benefits in the prevention and/or treatment of cancer when used either alone as monotherapy or in combination with other anticancer agents.

Biofactors. 2014 Jan-Feb;40(1):49-58

Immunosenescence, inflammation and Alzheimer’s disease.

Ageing impacts negatively on the development of the immune system and its ability to fight pathogens. Progressive changes in the T-cell and B-cell systems over the lifespan of individuals have a major impact on the capacity to respond to immune challenges. The cumulative age-associated changes in immune competence are termed immunosenescence that is characterized by changes where adaptive immunity deteriorates, while innate immunity is largely conserved or even upregulated with age. On the other hand, ageing is also characterized by “inflamm-ageing”, a term coined to explain the inflammation commonly present in many age-associated diseases. It is believed that immune inflammatory processes are relevant in Alzheimer’s disease, the most common cause of dementia in older people. In the present paper we review data focusing on changes of some immunoinflammatory parameters observed in patients affected by Alzheimer’s disease.

Longev Healthspan. 2012 Nov 1;1:8

Inflammation and cardiovascular disease mechanisms.

The traditional view of atherosclerosis as a lipid storage disease crumbles in the face of extensive and growing evidence that inflammation participates centrally in all stages of this disease, from the initial lesion to the end-stage thrombotic complications. Investigators now appreciate that narrowing arteries do not necessarily presage myocardial infarction and that simply treating narrowed blood vessels does not prolong life. Although invasive approaches such as angioplasty and coronary artery bypass will remain necessary in some cases, we now understand that at least some of the cardiovascular benefits attributable to medical treatment and lifestyle modification (diet and physical activity) may result from reductions in inflammatory processes.

Am J Clin Nutr. 2006 Feb;83(2):456S-460S

Immunosenescence: emerging challenges for an ageing population.

It is now becoming apparent that the immune system undergoes age-associated alterations, which accumulate to produce a progressive deterioration in the ability to respond to infections and to develop immunity after vaccination, both of which are associated with a higher mortality rate in the elderly. Immunosenescence, defined as the changes in the immune system associated with age, has been gathering interest in the scientific and health-care sectors alike. The rise in its recognition is both pertinent and timely given the increasing average age and the corresponding failure to increase healthy life expectancy. This review attempts to highlight the age-dependent defects in the innate and adaptive immune systems. While discussing the mechanisms that contribute to immunosenescence, with emphasis on the extrinsic factors, particular attention will be focused on thymic involution. Finally, we illuminate potential therapies that could be employed to help us live a longer, fuller and healthier life.

Immunology. 2007 Apr;120(4):435-46

Immunosenescence is associated with altered gene expression and epigenetic regulation in primary and secondary immune organs.

Deterioration of the immune system (immunosenescence) with age is associated with an increased susceptibility to infection, autoimmune disease and cancer, and reduced responsiveness to vaccination. Immunosenescence entails a reduced supply of naïve T cells from the thymus and increased specialization of peripheral T cell clones. Both thymic involution and peripheral T cell homeostasis are thought to involve cellular senescence. In order to analyze this at the molecular level, we studied gene expression profiles, epigenetic status, and genome stability in the thymus and spleen of 1-, 4-, and 18-month-old Long Evans rats. In the thymus, altered gene expression, DNA and histone H3K9 hypomethylation, increased genome instability, and apoptosis were observed in 18-month-old animals compared to 1- and 4-month-old animals. In the spleen, alterations in gene expression and epigenetic regulation occurred already by the age of 4 months compared to 1 month and persisted in 18-month-old compared to 1-month-old rats. In both organs, these changes were accompanied by the altered composition of resident T cell populations. Our study suggests that both senescence and apoptosis may be involved in altered organ function.

Front Genet. 2013 Oct 18;4:211

Ganoderma lucidum polysaccharides prevent platelet-derived growth factor-stimulated smooth muscle cell proliferation in vitro and neointimal hyperplasia in the endothelial-denuded artery in vivo.

Ganoderma lucidum is used in traditional Chinese medicine to prevent or treat a variety of diseases, including cardiovascular disorders. We previously demonstrated that a glucan-containing extract of Reishi polysaccharides (EORP) has the potent anti-inflammatory action of reducing ICAM-1 expression in lipopolysaccharide (LPS)-treated human aortic smooth muscle cells (HASMCs) and LPS-treated mice. In the present study, we examined whether EORP inhibited platelet-derived growth factor-BB (PDGF)-stimulated HASMC proliferation and the mechanism involved. EORP dose-dependently reduced cell numbers and DNA synthesis of PDGF-treated HASMCs in vitro. EORP also arrested cell cycle progression in the G0/G1 phase, and this was associated with decreased expression of cyclin D1, cyclin E, CDK2, CDK4, and p21(Cip1) and upregulation of the cyclin-dependent kinase inhibitor p27(Kip1). The anti-proliferative effect of EORP was partly mediated by downregulation of PDGF-induced JNK phosphorylation. In in vivo studies, the femoral artery of C57BL/6 mice was endothelial-denuded and the mice were fed a diet containing 100 mg/kg/day of EORP. On day 14, both cell proliferation (proliferating cell nuclear antigen-positive cells) in the neointima and the neointima/media area ratio (0.67 ± 0.03 vs. 1.46 ± 0.30) were significantly reduced. Our data show that EORP interferes with the mitogenic activation of JNK, preventing entry of HASMCs into the cell cycle in vitro and reducing cell proliferation in the neointima and decreasing the neointimal area in vivo. Thus, EORP may represent a safe and effective novel approach to the prevention and treatment of vascular proliferative diseases.

J Cell Physiol. 2012 Aug;227(8):3063-71

Ganoderma lucidum polysaccharides attenuate endotoxin-induced intercellular cell adhesion molecule-1 expression in cultured smooth muscle cells and in the neointima in mice.

The expression of adhesion molecules on vessels and subsequent leukocyte recruitment are critical events in vascular diseases and inflammation. The aim of the present study was to examine the effects of an extract of Ganoderma lucidum (Reishi) polysaccharides (EORP), which is effective against cancer and immunological disorders, on adhesion molecule expression by human aortic smooth muscle cells (HASMCs) and the underlying mechanism. EORP significantly suppressed lipopolysaccharide (LPS)-induced intercellular cell adhesion molecule-1 (ICAM-1) mRNA and protein expression and reduced the binding of human monocytes to LPS-stimulated HASMCs. Immunoprecipitation and real-time polymerase chain reaction demonstrated that EORP markedly reduced the interaction of human antigen R protein (HuR) with the 3’-UTR of ICAM-1 mRNA in LPS-stimulated HASMCs. EORP treatment also suppressed extracellular signal-regulated kinase (ERK) phosphorylation and reduced the density of the shifted bands of nuclear factor (NF)-kappaB after LPS-induced activation. In an endothelial-denuded artery model in LPS-treated mice, daily oral administration of EORP for 2 weeks decreased neointimal hyperplasia and ICAM-1 expression in the plasma and neointima. These results provide evidence that EORP attenuates LPS-induced adhesion molecule expression and monocyte adherence and that this protective effect is mediated by decreased ERK phosphorylation and NF-kappaB activation. These findings suggest that EORP has anti-inflammatory properties and could prove useful in the prevention of vascular diseases and inflammatory responses.

J Agric Food Chem. 2010 Sep 8;58(17):9563-71

Effect of 26-oxygenosterols from Ganoderma lucidum and their activity as cholesterol synthesis inhibitors.

Ganoderma lucidum is a medicinal fungus belonging to the Polyporaceae family which has long been known in Japan as Reishi and has been used extensively in traditional Chinese medicine. We report the isolation and identification of the 26-oxygenosterols ganoderol A, ganoderol B, ganoderal A, and ganoderic acid Y and their biological effects on cholesterol synthesis in a human hepatic cell line in vitro. We also investigated the site of inhibition in the cholesterol synthesis pathway. We found that these oxygenated sterols from G. lucidum inhibited cholesterol biosynthesis via conversion of acetate or mevalonate as a precursor of cholesterol. By incorporation of 24,25-dihydro-[24,25-3H2]lanosterol and [3-3H]lathosterol in the presence of ganoderol A, we determined that the point of inhibition of cholesterol synthesis is between lanosterol and lathosterol. These results demonstrate that the lanosterol 14alpha-demethylase, which converts 24,25-dihydrolanosterol to cholesterol, can be inhibited by the 26-oxygenosterols from G. lucidum. These 26-oxygenosterols could lead to novel therapeutic agents that lower blood cholesterol.

Appl Environ Microbiol. 2005 Jul;71(7):3653-8

Isolation and structure determination of a cholesterol esterase inhibitor from Ganoderma lucidum.

Bioassay-guided fractionation of methanol extract of Ganoderma lucidum gave a pure cholesterol esterase inhibitor. On the basis of spectroscopic analysis and comparison with data from the literature, its structure was identified as 5alpha,8alpha-epidioxyergosta-6,22-dien-3beta-ol (compound I). Compound (I) inhibited cholesterol esterase with an IC50 value of 42 micronM. A Lineweaver-Burk plot revealed that the compound (I) is a noncompetitive inhibitor. Findings from this study suggest that compound (I) may be the active principle of the hypocholesterolemic effect of Ganoderma lucidum.

J Microbiol Biotechnol. 2010 Nov;20 (11):1521-3

Ganoderma lucidum polysaccharides exert anti-hyperglycemic effect on streptozotocin-induced diabetic rats through affecting b-cells.

Previous studies have demonstrated that Ganoderma lucidum polysaccharides (Gl-PS) exhibited potential antihyperglycemic effect in rats. The aim of the present study was to investigate the mechanism of the hypoglycemic effect of a low- molecular-weight Gl-PS in streptozotocin (STZ)-induced diabetic Sprague-Dawley (SD) rats. Gl-PS was extracted and purified from Ganodema lucidum fruiting body. 50 male SD rats were included in the study; 10 were taken as healthy controls; 40 were induced to diabetes by a single injection of 65 mg/kg STZ, of which 30 were selected as successful diabetic rat models. The 30 diabetic rats were divided into three groups: Gl-PS (200 mg/kg Gl-PS), metformin (100 mg/kg metformin) and diabetic control (n = 10 per group). After eight weeks’ oral administration, plasma concentrations of fasting glucose, triacylglyceride, total cholesterol and nitric oxide were significantly decreased in Gl-PS and metformin groups. Pancreatic superoxide dismutase, catalase and glutathione peroxidase were significantly increased in Gl-PS and metformin groups. Histopathological results showed that Gl-PS and metformin had protective effect on b-cells. The mRNA expressions of Bcl-2 and PDX-1 in pancreas were up-regulated, but Bax, iNOS and Casp-3 down-regulated in Gl- PS and metformin groups compared to diabetic control group. The present results suggested that Gl-PS had a hypoglycemic effect in STZ-induced diabetic rats through preventing apoptosis of pancreatic b-cells and enhancing b-cells regeneration.

Comb Chem High Throughput Screen. 2012 Aug;15(7):542-50

Ganoderic acid Mf and S induce mitochondria mediated apoptosis in human cervical carcinoma HeLa cells.

In this work, the effects of a pair of positional isomer of ganoderic acids (GAs), namely ganoderic acid Mf (GA-Mf) and ganoderic acid S (GA-S) purified from the fermented mycelia of Ganoderma lucidum, on induction of cell apoptosis and the apoptotic pathway in HeLa cells were investigated. The results demonstrate that both isomers decreased cell population growth on various human carcinoma cell lines by MTT assay, while GA-Mf had better selectivity between normal and cancer cells. The flow cytometry analysis indicated that treatment of HeLa cells with GA-S caused cell cycle arrest in the S phase, while GA-Mf caused cell cycle arrest in the G1 phase. Compared with GA-S, GA-Mf had more potent increase in the number of early and late apoptotic cells. Treatment of HeLa cells with each isomer decreased the mitochondria membrane potential and caused the release of cytochrome c from mitochondria into the cytosol. In addition, stimulation of caspase-3 and caspase-9 activity was observed. The Bax/Bcl-2 ratio was also increased in GA-treated HeLa cells. The results demonstrated that both isomers GA-Mf and GA-S induced apoptosis of human HeLa cells through a mitochondria mediated pathway, but they had the different cell cycle arrest specificity. The findings will be helpful to the development of useful cancer chemopreventive compounds from G. lucidum.

Phytomedicine. 2011 Mar 15;18(5):349-55

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