Studies On Neuroprotective Mechanisms Of Resveratrol Against Cerebral Ischemia Reperfusion Injury In Different Phases In Mice

Ischemic brain stroke is one of the frequently occurred diseases of central nervous system, which can cause mortality and chronic disability. In the western world, 250~400 of every 0.1 million people die of ischemic brain stroke which is the third cause of death, and death rate of ischemic brain stroke is approximately 30%. Despite numerous clinical trials, the treatment is still elusive and largely empirical because the pathophysiology involved therein is complex and not yet well understood. The focus of current research is, therefore, to find novel drug to be used in ischemic stroke. Resveratrol (3, 5, 4 -trihydroxy-trans-stilbene, Res), a natural polyphenolic compound, is found in a few edible materials and is well known for its phytoestrogenic and antioxidant properties. Since Res was discovered in 1940, dozens of reports have shown that Res exerts such a range of beneficial effect across species and disease models, including inhibiting carcinogenesis at multiple stages in a wide variety of cancer models, decreasing platelet aggregation enhancing stress resistance and extending the lifespans of various organisms from yeast to vertebrates. In recently years, a lot of researches indicate that Res exhibits neuroprotective effect in a variety of in vitro and in vivo model for cerebral ischemic stroke. In this study, we further researched the protective effects of Res against ischemia reperfusion injury developed in the mouse brains in different phases in vivo. Moreover, we further investigated the relationship of Res and PPAR, MMP-2 and VEGF. The aim of this study is to discuss mechanisms of the protective effect of Res on cerebral ischemia reperfusion in different phases, which will provide an academic basis for Res to be used in treatment of cerebral ischemic disease.ⅠResveratrol's effect on acute phases of focal cerebral ischemic injury in miceObjective To discuss the influence of Res on acute phases of focal cerebral ischemic injury in mice. Methods Ninety Balb/c healthy male mice, weighting 18-22 g, were randomly divided into three groups: Res group, fenofibrate group and ischemia group. There were thirty mice in each group. The ischemic model of mice was induced with an 8/0 nylon monofilament coated with silicone hardener mixture via the internal carotid artery. 24 hours after reperfusion, the infarct area and edema extent were shown by 2, 3, 5-triphenyl tetrazolium chlorid (TTC) staining; Evans blue staining was used for showing the damage of blood brain barrier and the express of peroxisome proliferator activated receptor (PPAR) alpha was shown by RT-PCR. Results The infarct volume in resveratrol group and fenofibrate group were lower than ischemia group(43.80±5.31 mm3, 45.57±5.63 mm3 VS 63.87±9.33 mm3, P<0.01). In resveratrol group and fenofibrate group, the extent and quality of Evans blue extravasation were reduced compared with ischemia group(58.66±4.65 mm3, 78.72±5.78 mm3 VS 102.06±6.07 mm3, P<0.01 and 6.40±0.92 ug/g, 12.47±1.24 ug/g VS 15.25±1.51 ug/g, P<0.01), PPARαmRNAs were prominently expressed compared with ischemia group (0.893±0.028, 1.117±0.042 VS 0.467±0.013, P<0.01). Conclusion The study suggests that Res has protective effects against acute ischemic stroke, which could be attributed to its property increasing PPARα. Thus, Res may be a potential agent for the treatment of neuronal injury associated with stroke.ⅡResveratrol attenuates ischemic brain damage in the delayed phase after stroke and induces mRNA and protein express for angiogenic factorsObjective To investigate the effects and possible protective mechanism of Res on the delayed phase after focal cerebral ischemia injury in mice. Methods All mice were randomly assigned to five groups according to the time of administration of Res. Control group: Mice receiving corresponding volume of saline solution (0.9% NaCl) containing 20% hydroxypropyl h-cyclodextrin by gavage were used as control, and exposed to middle cerebral artery occlusion middle cerebral artery (MCA) occlusion and reperfusion injury; Ischemia group: Mice were treated with Res till the 7th day (50 mg/kg/d, gavage, the first time was 5 min prior to MCA reperfusion); Reperfusion group: Mice were treated with Res till the 7th day (50 mg/kg/d, gavage, the first time was 5 min prior to MCA reperfusion); 1st day group: Mice were treated with Res till the 7th day (50 mg/kg/d, gavage, the first time was 24 hours after MCA reperfusion); 3rd day group: Mice were treated with Res till the 7th day (50 mg/kg/d, gavage, the first time was 72 hours after MCA reperfusion). At the 7th day after reperfusion, brain injury was evaluated by triphenyltetrazolium chloride staining (TTC) and neurological examination. Microvascular number was examined with immunohistochemistry staining. Effect of resveratrol on MMP-2 and VEGF gene expression were investigated with RT-PCR and western blot. Results The mean neurological scores and infarct volume of ischemia group and reperfusion group were lower than that of control group at the 7th day after MCA reperfusion (P < 0.05). Immunohistochemistry staining showed significantly less reduction of the number of the microvessels in the cortical area of mice on ischemia group, reperfusion group and 1st day group compared with the control group. In the ischemic hemispheres of ischemia group and reperfusion group, the level of mRNA and protein of MMP-2 and VEGF were significantly elevated (P < 0.05). Conclusions This study demonstrated that Res administration by gavage provided an important neuroprotective effect on focal cerebral ischemia injury in the delayed phase. Furthermore, the elevated MMP-2 and VEGF might play important role in the neuroprotective effect of Res administration by inducing angiogenesis.