The Influence Effect Of Di Huang Yin Zi Assosiated With Bone Marrow Stromal Stem Cells On Neuronal Apoptosis And The Expression Of Hsp70 And Vegf After Cerebral Ischemic Injury On Rats

Objective:To explore the protective effects of transplantation of Di Huang Yin Zi incubate bone marrow stromal stem cells on rats ischemic brain injury by means of ischemic brain injury in rats for the research object; by observing the nerve cell apoptosis and the expression changes of HSP70 and VEGF gene after cerebral ischemic injury, to reveal the protective effect of transplantation of Di Huang Yin Zi incubate BMSCs for cerebral ischemic injury in nerve cells and its mechanism.Methods:BMSCs of Wister rat were cultured and purified by attachment method,phenotypers of BMSCs were analysed by flow cytomwetry;to prepare normal rat serum, Di Huang Yin Zi serum.Rat BMSCs passaged to 3 generation were cultured in different drug serum and normal rat serum 72h. Permanent models of middle cerebral artery occlusion by nylon suture embolization were made in healthy Wister rats. The operated rats were divided into the model group, BMSCs cultured by noromal rat serum group(BMSCs group), BMSCs cultured by ATRA group (ATRA group), BMSCs cultured by Di Huang Yin Zi serum group(Di Huang Yin Zi group),and made the sham-operation group. At 24h after MCAO, BMSCs (about 1×106 cells 1ml) were collected and intravenously infushed into the tail vein of rats after digestion and centrifugalization. The sham-operation group and model group were injected the same dose of isotonic Na chloride through vena caudalis.After to breed rats seven days and forteen days, the rats were decapitated then the brains were taken out to make the paraffin sections. To observe the pathological changes of in brain tissue by microscope. Immunohistochemical methed was used to investigate the expressions of Bcl-2,Bax, HSP70 and VEGF in the cerebral mantle and did retrospective analysis using statistic software spss16.0. Results:(1) By observation of the histologic slice, there are no obvious pathological damage in sham-operation group, there are varying degrees of nerve cell damage and structural damage in model group. Compared with the model group, pathological destruction of brain tissue in BMSCs group, ATRA group and Di Huang Yin Zi group are reduced. Compared with BMSCs group, the effect of combined treatment groups are significant. By observation of the microscopy to show the number of nerve cell degeneration and necrosis as well as interstitial edema in BMSCs group,ATRA group and Di Huang Yin Zi group are decreased.(2)Compared with the sham-operation group,the expression of Bcl-2 increased in the model group (P<0.05). The expressions of Bcl-2 in BMSCs group, RA group and Di Huang Yin Zi group were higher than in model group(P<0.05). Compared with BMSCs group, the expression of Bcl-2 in combined treatment group increased significantly(P<0.05).(3) Compared with the sham-operation group,the expression of Bax increased in the model group (P<0.05). The expressions of Bax in BMSCs group, RA group and Di Huang Yin Zi group decreased than in model group(P<0.05); Compared with BMSCs group, the expressions of combined treatment groups decreased significantly(P<0.05).(4) Compared with the sham-operation group,the expression of HSP70 increased in the model group (P<0.05). The expressions of HSP70 in BMSCs group, RA group and Di Huang Yin Zi group were higher than in model group(P<0.05). Compared with BMSCs group, the expressions of combined treatment group increased significantly(P<0.05).(5) Compared with the sham-operation group,the expression of VEGF increased in the model group (P<0.05). The expressions of HSP70 in Di Huang Yin Zi group, RA group and BMSCs group were higher than in model group(P<0.05). and the Di Huang Yin Zi group increased higher than BMSCs group, (P<0.05).Conclusions:Bcl-2 and Bax have the important effect in the procedure of cerebral ischemic injury. Di Huang Yin Zi has protection for the cerebral ischemic injury. To increase the expressions of bcl-2 and decrease the expressions of bax are protection mechanism of Di Huang Yin Zi. To promote the expressions of HSP70 and VEGF protein may be one of protection mechanism with transplantation of Di Huang Yin Zi incubate BMSCs for the treatment of the cerbral infarction.