IH636 Grape Seed Proanthocyanidin Extract Research Articles

Ingestion of IH636 grape seed proanthocyanidin extract to prevent selenite-induced oxidative stress in experimental cataract

To investigate whether dietary supplementation with IH636 grape seed proanthocyanidin extract (GSPE) prevents selenite-induced cataract. Department of Ophthalmology, Gulhane Military Medical Academy, Ankara, Turkey. Thirty Spraque-Dawley rat litters were put randomly into 3 groups. In group 1 (n = 10), sodium selenite (30 nmol/g body weight) was injected subcutaneously on postpartum day 10. In group 2 (n = 10), sodium selenite (30 nmol/g body weight) was injected on postpartum day 10 and oral GSPE (100 mg/kg body weight) was given for 1 week after sodium selenite injection. Only subcutaneous saline was injected in group 3 (control, n = 10). The development of cataract was assessed for 3 weeks, and its density was graded and photographed with a slitlamp. Removed rat lenses were analyzed for glutathione (GSH) and malondialdehyde (MDA). All of the rats in group 1 had cataract between stage 6 and stage 3. In group 2, only 5 of 10 eyes had cataract between stage 3 and stage 2 and no cataract occurred in the remaining 5 rats. The difference between mean cataract stages in group 1 and group 2 was significant (P<.05). The mean GSH level in group 1 was significantly lower than in group 2 and controls (P<.05). The mean MDA level in group 1 was significantly higher than in group 2 and controls (P<.05). IH636 grape seed proanthocyanidin extract effectively suppressed cataract formation in rats. Routine consumption of grape seed proanthocyanidin extract in the form of food or dietary supplement may offer a prophylactic measure against onset and progression of cataract.

Double-blind, placebo-controlled, randomised phase II trial of IH636 grape seed proanthocyanidin extract (GSPE) in patients with radiation-induced breast induration

Background and purposeTissue hardness (induration), pain and tenderness are common late adverse effects of curative radiotherapy for early breast cancer. The purpose of this study was to test the efficacy of IH636 grape seed proanthocyanidin extract (GSPE) in patients with tissue induration after high-dose radiotherapy for early breast cancer in a double-blind placebo-controlled randomised phase II trial. Patients and methodsSixty-six eligible research volunteers with moderate or marked breast induration at a mean 10.8 years since radiotherapy for early breast cancer were randomised to active drug (n=44) or placebo (n=22). All patients were given grape seed proanthocyanidin extract (GSPE) 100mg three times a day orally, or corresponding placebo capsules, for 6 months. The primary endpoint was percentage change in surface area (cm2) of palpable breast induration measured at the skin surface 12 months after randomisation. Secondary endpoints included change in photographic breast appearance and patient self-assessment of breast hardness, pain and tenderness. ResultsAt 12 months post-randomisation, ≥50% reduction in surface area (cm2) of breast induration was recorded in13/44 (29.5%) GSPE and 6/22 (27%) placebo group patients (NS). At 12 months post-randomisation, there was no significant difference between treatment and control groups in terms of external assessments of tissue hardness, breast appearance or patient self-assessments of breast hardness, pain or tenderness. ConclusionsThe study failed to show efficacy of orally-adminstered GSPE in patients with breast induration following radiotherapy for breast cancer.

Beneficial effects of a novel IH636 grape seed proanthocyanidin extract and a niacin-bound chromium in a hamster atherosclerosis model.

Atherosclerosis is a disease of the arteries in which fatty plaques develop on the inner arterial wall, which eventually obstructs blood flow. Identified risk factors for atherosclerosis include genetics, diet, lifestyle, smoking, circulating lipid and cholesterol levels, and molecular and circulating signals of chronic vascular inflammation. The link between flavonoids and atherosclerosis is based partly on the evidence that some flavonoids possess antioxidant properties and have been shown to be potent inhibitors of LDL oxidation in vitro. Hypercholesterolemia, a significant cardiovascular risk factor is prevalent in the American population. Grape seed proanthocyanidin extracts are known to exhibit a broad spectrum of chemopreventive and cardioprotective properties against oxidative stress. A recent study has shown that a combination of IH636 grape seed proanthocyanidin extract (GSPE) and a niacin-bound chromium (NBC) can decrease total cholesterol, LDL and oxidized LDL levels in hypercholesterolemic human subjects. In this study, we assessed the efficacy of GSPE supplementation in hamsters, singly and in combination with NBC, since these animals have a similar lipid profile to hypercholesterolemic humans when fed a hypercholesterolemic diet of 0.2% cholesterol and 10% coconut oil (HCD). After 10 weeks of feeding HCD, these animals developed foam cells, which is a biomarker of early stages of atherosclerosis. Atherosclerosis (% of aorta covered with foam cells) was reduced by approximately 50% and 63% following supplementation of these animals with 50 mg/kg and 100 mg/kg of GSPE, respectively, in conjunction with a HCD, while approximately 32% reduction was observed following supplementation of GSPE plus NBC. A range of 7-9 animals was used in each study group. GSPE alone and in combination with NBC exerted a pronounced effect on the cholesterol, and triglyceride levels, as well as oxidative lipid damage as demonstrated by the formation of thiobarbituric acid reactive substances (TBARS). This data demonstrates that GSPE and NBC may provide significant health benefits by dramatically ameliorating the incidence of atherosclerosis as demonstrated by reducing the formation of foam cells.

Differential effects of IH636 grape seed proanthocyanidin extract and a DNA repair modulator 4-aminobenzamide on liver microsomal cytochrome 4502E1-dependent aniline hydroxylation.

Previous studies from our laboratories have linked the protective abilities of IH636 grape seed proanthocyanidin extract (GSPE) with inactivation of anti-apoptotic gene bcl-XL, and modification of several other critical molecular targets such as DNA-damage/DNA-repair, lipid peroxidation and intracellular Ca2+ homeostasis. Especially, GSPE provided dramatic protection against acetaminophen (APAP)-induced hepatotoxicity, significantly increased bcl-XL expression in the liver, and antagonized both necrotic and apoptotic deaths of liver cells in vivo. However, it was not clear from this study whether anti-apoptogenic and anti-necrotic effects of GSPE were: (i) due to its interference with endonuclease activity, (ii) due to its antioxidant effect, or, (iii) due to its ability to inhibit microsomal drug metabolizing enzyme(s), such as CYP-4502E1. Since CYP-4502E1 primarily metabolizes acetaminophen in mice and rats, this study specifically focused on CYP-4502E1's catalytic activity in vitro. Overall this investigation compared the in vitro aniline hydroxylation patterns of: (i) in vivo GSPE-exposed and unexposed (control) mouse liver microsomes, (ii) induced (1% acetone in drinking water for 3 days) and uninduced rat liver microsomes in the presence and absence of GSPE in vitro, and (iii) control rat liver microsomes in the presence of an anti-APAP agent 4-aminobenzamide (4-AB) in vitro. For the in vivo assessment, male B6C3F1 mice were fed GSPE diet (ADI 100 mg/kg body wt) for 4 weeks, and liver microsomes were isolated from both control and GSPE-fed mice for aniline hydroxylation, a specific marker of CYP-4502E1 activity. Data show that hydroxylation was 40% less in microsomes from GSPE-exposed livers compared to control microsomes. Similarly, when rat liver microsomes were incubated with various concentrations of GSPE in vitro (100 and 250 microg/ml), aniline hydroxylation was inhibited to various degrees (uninduced: 40 and 60% and induced: 25 and 50%, respectively with 100 and 250 microg/ml). Influence of GSPE on hydroxylation patterns were compared with another hepatoprotective agent 4-aminobenzamide (4-AB), a well-known modulator of nuclear enzyme poly(ADP-ribose) polymerase, and the data shows that 4-AB did not alter aniline hydroxylation at all. Collectively, these results may suggest that GSPE has the ability to inhibit CYP-4502E1, and this is an additional cytoprotective attribute, in conjunction with its novel antioxidant and/or antiendonucleolytic potential.

Protective effects of antioxidants against smokeless tobacco-induced oxidative stress and modulation of Bcl-2 and p53 genes in human oral kerathnocytes

The oral use of chewing tobacco has greatly increased in recent years, and this usage is associated with cancers of the mouth, lip, nasal cavities, esophagus and gut. Oral cancer accounts for 3% of all cancers in U.S.A. and is the seventh most common cancer. Previous studies in our laboratory have demonstrated the protective abilities of a novel IH636 grape seed proanthocyanidin extract (GSPE) against reactive oxygen species both in vitro and in vivo models, and provided significantly better protection as compared to vitamins C, E and β-carotene. In the recent past, we have demonstrated smokeless tobacco (STE)-induced oxidative stress, apoptotic cell death in a primary culture of normal human oral keratinocytes (NHOK), and have compared the protective abilities of vitamins C and E, singly and in combination, and GSPE in this pathobiology [Free Rad. Biol. Med., 26, 992–1000 (1999)]. In the present study, we have assessed the protective role of vitamins C and E, and GSPE against STE-induced modulation of intracellular oxidized states in NHOK cells as demonstrated by laser scanning confocal microscopy. Approximately 11%, 26%, 28% and 50% protection were observed following incubation with vitamin C, vitamin E, a combination of vitamins C plus E, and GSPE, respectively. DNA fragmentation was assessed as an index of oxidative DNA damage and similar results were observed. Furthermore, the cellular viability and functional roles of Bcl-2, p53 and c-myc genes were assessed in STE-induced oxidative stress in NHOK cells. NHOK cells were treated with STE (0–200 μg/ml) for 24h and changes in the expression of Bcl-2, p53 and c-myc genes were measured by reverse transcriptase-polymerase chain reaction (RT-PCR), and the protective effect of GSPE was assessed. Approximately a 2.0-fold increase in p53 gene expression was observed following incubation of the oral keratinocytes with 100 μg/ml of STE, beyond which the expression of p53 decreased, confirming increased apoptotic cell death with a higher concentration of STE as reported earlier. GSPE significantly modulated STE-induced changes in p53. The expression of antiapoptotic Bcl-2 gene decreased with STE treatment and the expression of Bcl-2 gene increased significantly following preincubation with GSPE. No significant change in the expression of transcription factor c-myc gene responsible for cell cycle growth was observed following incubation with STE and/or GSPE. Thus, c-myc may not be involved in STE-induced cytotoxicity towards NHOK cells. These results suggest that antioxidant protection of STE-induced cellular injury is associated with alterations in Bcl-2 and p53 expression.

Free radicals and grape seed proanthocyanidin extract: importance in human health and disease prevention

Free radicals have been implicated in over a hundred disease conditions in humans, including arthritis, hemorrhagic shock, atherosclerosis, advancing age, ischemia and reperfusion injury of many organs, Alzheimer and Parkinson’s disease, gastrointestinal dysfunctions, tumor promotion and carcinogenesis, and AIDS. Antioxidants are potent scavengers of free radicals and serve as inhibitors of neoplastic processes. A large number of synthetic and natural antioxidants have been demonstrated to induce beneficial effects on human health and disease prevention. However, the structure-activity relationship, bioavailability and therapeutic efficacy of the antioxidants differ extensively. Oligomeric proanthocyanidins, naturally occurring antioxidants widely available in fruits, vegetables, nuts, seeds, flowers and bark, have been reported to possess a broad spectrum of biological, pharmacological and therapeutic activities against free radicals and oxidative stress. We have assessed the concentration- or dose-dependent free radical scavenging ability of a novel IH636 grape seed proanthocyanidin extract (GSPE) both in vitro and in vivo models, and compared the free radical scavenging ability of GSPE with vitamins C, E and β-carotene. These experiments demonstrated that GSPE is highly bioavailable and provides significantly greater protection against free radicals and free radical-induced lipid peroxidation and DNA damage than vitamins C, E and β-carotene. GSPE was also shown to demonstrate cytotoxicity towards human breast, lung and gastric adenocarcinoma cells, while enhancing the growth and viability of normal human gastric mucosal cells. The comparative protective effects of GSPE, vitamins C and E were examined on tobacco-induced oxidative stress and apoptotic cell death in human oral keratinocytes. Oxidative tissue damage was determined by lipid peroxidation and DNA fragmentation, while apoptotic cell death was assessed by flow cytometry. GSPE provided significantly better protection as compared to vitamins C and E, singly and in combination. GSPE also demonstrated excellent protection against acetaminophen overdose-induced liver and kidney damage by regulating bcl-X L gene, DNA damage and presumably by reducing oxidative stress. GSPE demonstrated excellent protection against myocardial ischemia-reperfusion injury and myocardial infarction in rats. GSPE was also shown to upregulate bcl 2 gene and downregulate the oncogene c-myc. Topical application of GSPE enhances sun protection factor in human volunteers, as well as supplementation of GSPE ameliorates chronic pancreatitis in humans. These results demonstrate that GSPE provides excellent protection against oxidative stress and free radical-mediated tissue injury.

Amelioration of the cytotoxic effects of chemotherapeutic agents by grape seed proanthocyanidin extract.

Anticancer chemotherapeutic agents are effective in inhibiting growth of cancer cells in vitro and in vivo, however, toxicity to normal cells is a major problem. In this study, we assessed the effect of a novel IH636 grape seed proanthocyanidin extract (GSPE) to ameliorate chemotherapy-induced toxic effects in cultured Chang epithelial cells, established from nonmalignant human tissue. These cells were treated in vitro with idarubicin (Ida) (30 nM) or 4-hydroxyperoxycyclophosphamide (4HC) (1 microg/ml) with or without GSPE (25 microg/ml). The cells were grown in vitro and the growth rate of the cells was determined using the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; thiazolyl blue] assay. Our results showed that GSPE decreased the growth inhibitory and cytotoxic effects of Ida as well as 4HC on Chang epithelial cells in vitro. Because these chemotherapeutic agents are known to induce apoptosis in the target cells, we analyzed the Chang epithelial cells for apoptotic cell population by flow cytometry. There was a significant decrease in the number of cells undergoing apoptosis following treatment with GSPE. We also found increased expression of the anti-apoptotic protein Bcl-2 in GSPE-treated cells using western blot techniques. Thus, these results indicate that GSPE can be a potential candidate to ameliorate the toxic effects associated with chemotherapeutic agents and one of the mechanisms of action of GSPE includes upregulation of Bcl-2 expression.

Acute and chronic stress-induced oxidative gastrointestinal injury in rats, and the protective ability of a novel grape seed proanthocyanidin extract

Reactive oxygen species (ROS) are implicated in the pathogenesis of stress-induced gastrointestinal injury. In the present study, we have investigated the effects of acute stress and chronic stress on the enhanced production of superoxide anion as determined by cytochrome c reduction assay, and correlated the enhanced production of superoxide anion with increased lipid peroxidation, DNA fragmentation and membrane microviscosity, indices of oxidative tissue injury, in gastric mucosa and intestinal mucosa, and determined the protective effects of a novel IH636 grape seed proanthocyanidin extract (GSPE) against mucosal injury. Acute stress was induced by water-immersion restraint stress for 90 min, while chronic stress was induced by water-immersion restraint stress for 15 min/day for 15 consecutive days. Half of the animals exposed to acute stress were pretreated orally with 100 mg GSPE/kg/day for 15 consecutive days. Similarly, half of the animals exposed to chronic stress were pretreated orally with 100 mg GSPE/kg/day for 15 consecutive days. Acute stress produced maximal injury to both gastric mucosa and intestinal mucosa as compared to chronic stress. Acute stress and chronic stress increased cytochrome c reduction by 12.2- and 4.8-fold, respectively, in the gastric mucosa, and 12.1- and 4.6-fold in the intestinal mucosa. Acute stress increased lipid peroxidation, DNA fragmentation and membrane microviscosity by 3.3-, 4.1- and 11.6-fold, respectively, in the gastric mucosa, and 4.4-, 5.2- and 16.6-fold, respectively, in intestinal mucosa. GSPE decreased acute stress-induced lipid peroxidation, DNA fragmentation and membrane microviscosity by 15%, 12% and 13%, respectively, in the gastric mucosa, and by 13%, 14% and 16%, respectively, in the intestinal mucosa. Chronic stress increased lipid peroxidation, DNA fragmentation and membrane microviscosity by 2.9-, 3.3- and 6.3- fold, respectively, in the gastric mucosa, and 3.3-, 4.2- and 9.3-fold, respectively, in the intestinal mucosa. GSPE decreased chronic stress-induced lipid peroxidation, DNA fragmentation and membrane microviscosity by 23%, 21% and 25%, respectively, in the gastric mucosa, and by 26%, 26% and 25%, respectively, in the intestinal mucosa. These results demonstrate that acute stress and chronic stress can induce gastrointestinal oxidative stress and mucosal injury through enhanced production of ROS, and that GSPE provides significant protection against gastrointestinal oxidative stress and mucosal injury by scavenging these ROS.

The cytotoxic effects of a novel IH636 grape seed proanthocyanidin extract on cultured human cancer cells.

Grape seed proanthocyanidins are natural antioxidants which possess a broad spectrum of chemoprotective properties against free radicals and oxidative stress. In this study, we have assessed the cytotoxicity of a novel IH636 grape seed proanthocyanidin extract (GSPE) against MCF-7 human breast cancer cells, A-427 human lung cancer cells, CRL-1739 human gastric adenocarcinoma cells and K562 chronic myelogenous leukemic cells at 25 and 50 mg/lit concentrations for 0-72 h using cytomorphology and MTT cytotoxicity assay. In addition, we compared the effects on normal human gastric mucosal cells and normal J774A.1 murine macrophage cells with the effects on the cancer cell lines. Concentration- and time-dependent cytotoxic effects of GSPE were observed on the MCF-7 breast cancer, A-427 lung cancer and gastric adenocarcinoma cells. Following incubation of the MCF-7 cells with 25 mg/lit of the GSPE approximately 6.5, 30 and 43% inhibitions in cell growth were observed at 24, 48 and 72 h of incubation, respectively, while incubation of the MCF-7 cells with 50 mg/lit of the GSPE resulted in 11, 35 and 47% inhibition in cell growth at these same points, respectively. Similar results were observed in the A-427 and gastric adenocarcinoma cells. GSPE exhibited no cytotoxicity toward the neoplastic K562 myelogenous leukemic cells. However, GSPE enhanced the growth and viability of the normal human gastric mucosal cells and J774A.1 murine macrophage cells. These data demonstrate that GSPE exhibited cytotoxicity towards some cancer cells, while enhancing the growth and viability of the normal cells which were examined.

Hydrogen peroxide-induced modulation of intracellular oxidized states in cultured macrophage J774A.1 and neuroactive PC-12 cells, and protection by a novel grape seed proanthocyanidin extract

We have previously compared selected antioxidants including vitamins C and E, β-carotene and a novel IH636 grape seed proanthocyanidin extract (GSPE) with respect to their scavenging abilities against biochemically generated free radicals in both in vitro and in vivo models. The results demonstrated that GSPE is a significantly more potent oxygen free radical scavenger compared with vitamins C and E and β-carotene. GSPE has been reported to exhibit a wide range of biological and pharmacological activities including free radical scavenging, antibacterial, antiviral, antiinflammatory, antiallergic and vasodilator actions. In this study hydrogen peroxide-induced oxidative damage was assessed in macrophage J774A.1 and neuroactive adrenal pheochromocytoma PC-12 cells, and the concentration-dependent ability of GSPE to protect these cells. Laser scanning confocal microscopy was used to determine intracellular oxidized states. Approximately 5.8- and 4.5-fold increases in fluorescence intensity were observed following incubation of macrophage J774A.1 and PC-12 cells with 0.50 mM H2O2 for 24 h, respectively. Pretreatment of the J774A.1 cells with 50 mg/L and 100 mg/L GSPE decreased H2O2-induced fluorescence intensity by 36% and 70%, respectively, while under these same conditions approximately 50% and 70% decreases in fluorescence intensities were observed in PC-12 cells. The results indicate that hydrogen peroxide significantly increases oxidative stress in these cells as demonstrated by the increase in intracellular oxidized states. Furthermore, GSPE can significantly protect against hydrogen peroxide-induced oxidative stress in cultured J774A.1 macrophage and neuronal PC-12 cells. Copyright © 1998 John Wiley & Sons, Ltd.