Protection And Part Mechanism Of Tetrahydroxy Stilbene Glucoside On Cerebral Ischemic-reperfusion Injury

Objective:To investigate the effects of tetrahydroxy stilbene glucoside （TSG） on gerbils offocal cerebral ischemia and reperfusion, and to explore protective effects and partmechanism of TSG on the HT22nerve cells. The result showed that TSG could lightencerebral ischemia-reperfusion injury with multiple ways. This study may provide thetheory and the experimental basis for TSG clinical application to resist cerebralischemia effect.Methods:1. Gerbils model of cerebral ischemia-reperfusion injury were established by modifiedLevine method. Rats were randomly divided into six groups: sham group, modelcontrol group, TSG large, medium and small doses （6,3,1.5mg/kg）, the positivecontrol medicine of edaravone injection group （3mg/kg）. The functions of learningand memory were detected by Morris water maze after7days. SOD, GSH-Px activityand MDA contents were detected in brain tissue homogenate. Cerebralpathomorphology was observed by hematoxylin and eosin （HE） staining. The proteinexpression of Nestin was detected by immunohistochemical SABC technology.2. HT22nerve cells of hippocampal ischemic injury model were established withsodium dithionite in DMEM without glucose for24h. The protection of TSG withkinds of doses （10,30,100, and300mg·L^-1） were observed by HT22damaged neurons. MTT test was performed to observe cell proliferation and evaluate theprotection effect of TSG. SOD, GSH-Px activity and MDA contents were detected bythe cell viability. Hoechst33342fluorescence nuclear staining was used to observeHT22cells apoptosis.Results:1. Morris water maze experiment. Compared with sham group, the time of swimmingout and the distance in model group was significantly extended. Compared with modelcontrol group, the time of swimming out and the distance in TSG （6,3mg/kg） treatmentgroup and edaravone control group were obviously reduced.2. Serum and brain tissue homogenate biochemical indicators Compared with shamgroup, SOD, GSH-Px enzyme activity were significantly reduced in gerbils modelbrains, MDA content increased obviously; Compared with model group, TSG （6,3mg/kg） could significantly improve the serum SOD activity and GSH-Px enzymeactivity, TSG （6,3,1.5mg/kg） group could reduce obviously the MDA content.3. HE staining results HE staining results showed that sham group had no infarctionareas in brain tissue and evident pathological change with the normal structure. Inmodel group, the number of neurons at ischemic area decreased, nerve cells shrankand degenerated, apparent interstitial edema between tissues and vacuolization. Butin TSG treatment group, the number of neurons appeared increased, degeneration andnecrosis were significantly decreased when compared with model group.4. Immunohistology results Immunohistology results showed that sham group had noNestin positive cells, model group had few Nestin positive cells. TSG（1.5,3, 6mg·kg^-1） could promote the expression of Nestin positive cells, and Nestin positiveexpressions in TSG （3,6mg·kg-1） were more obviously.5. Influence of HT22hippocampal neuronal cell vitality In normal control group,HT22hippocampal neurons showed cone shape, many bumps, axon staggered intomesh, and branches of synapses extended and thickening obviously. Neurons damageinduced by oxygen-glucose deprivation, lower outstanding refraction, parts of cells felloff, body of the refraction of declined. Compared with model group, TSG(30,100and300mg·L¹)could improve significantly the apoptosis of hippocampal neurons, couldincrease cell survival rate, and increase SOD, GSH-Px enzyme activity, reduce thecontent of MDA, improve HT22hippocampal neuronal cell survival after hypoxiahypoglycemia obviously.6. HT22cell apoptosis rate. Hoechst33342fluorescence staining showed that normalcontrol group of HT22hippocampal neurons presented orbicular-ovate nuclei, diffusedthe uniform blue fluorescence. Model group appeared solid shrink, the nucleardisintegrate, apoptotic body and other typical of the morphological changes, thefracturing of the nuclei showed half or the fragments form, a strong blue fluorescenceintensity. TSG could inhibit the apoptosis of HT22hippocampal neurons, and reducethe number of apoptotic body.Conclusion:1. TSG has positively protective and therapeutic effect on cerebral ischemia-reperfusion injury in gerbils, its mechanism may be relevant with free radicaldamage, inhibit the body of lipid peroxidation and promote the level of nerve cellsrepair. 2. TSG has a significant protective effect on HT22hippocampal nerve cell oxygen-glucose deprivation injury. The mechanism may be related to antioxidation andinhibition of apoptosis.