The Effect Of Two Kinds Of Tcm Treatments On Nscs Proliferation And Differentiation In Cerebral Ischemia And Reperfusion Rats Based On JAK/STAT Signal Pathway Genes And The Differences Protein Expression

1)Objective This experiment is based on the traditional Chinese medical theory of "essence-brain" and "blood-brain". It is designed to use JAK/STAT to control the proliferation and neuronal differentiation of neural stem cells, and produce a 2-hour focal cerebral ischemia in rats. With the help of gene technology, western blot, AG490 to discuss the effects of Yi Qi Huo Xue recipe and Bu Shen Sheng Sui recipe on the ischemia rats, especially the influence on JAK/STAT signal and their effects on JAK2 and STAT3. This experiment will provide molecular biological evidence for the application of Yi Qi Huo Xue recipe and Bu Shen Sheng Sui recipe.2)Methods Healthy male SD rats 80, weight 280-320. Among them, 8 were randomly made sham operation model, and the other 72 were used to make the model of middle cerebral artery occlusion in rats, The evaluation of the signs of neurological defect, 2~3 were selected. After this, MCAO/R rats were randomly divided into model group, AG490 group and Bushen Shengsui group, Yiqi Huoxue group. 5 groups in all, 6 rats in each group(except for the failure model and the death), 3 rats were used for gene chip, 3 rats were used for Western Blot, and 3 days were administered by gastric lavage. The experiment selected the fresh hippocampus tissue of the rats. The gene descriptiveness vicissitude in JAK/STAT signaling pathway in Hippocampus were detected by PCR gene chips technology in all groups. The protein expression changes of JAK2 and STAT3 in hippocampus were tested using the technique of Western-Blot. The datum from gene chip technique were made an assay by Delta Delta Ct method.and the data from Western-Blot were statistically analyzed by SPSS17.0.3)Results(1)Result of the first experiment89 genes were detected by gene chips technology in JAK/STAT signaling pathway.The expression of 11 kinds of genes(A2m,Cxcl9,Fas,Fcgr1 a,Il2ra,Lrg1,Prlr,Ptprc,Sirpa,Socs4,Socs5) in the sham group was obviously different compared with the model group. from which up-regulated genes have A2 m,Fas,Fcgr1 a,Lrg1,Ptprc,Socs4,Sirpa;down-regulated genes have Cxcl9,Il2 ra,Prlr,Socs5.The expression of 4 kinds of genes( Fcgr1 a,Prlr,Ptprc,Hprt1) in the AG490 blocking agent group was obviously different compared with the model group. from which up-regulated genes have Prlr,Hprt1; down-regulated genes have Ptprc,Fcgr1 a.The expression of 20 kinds of genes(A2m,Crk,Cxcl9,Fas,Fcgr1 a,Il2rg,Il6 st,Irf1,Lrg1,Nfkb1,Nos2,Ptprc,Sirpa,Smad4,Socs4,Sp1,Stam,Stat5 b,Tyk2,Hprt1) in the Yiqi Huoxue group was obviously different compared with the model group. from which up-regulated genes has Hprt1; the rest of genes were down-regulated.The expression of 12 kinds of genes(Prlr,A2 m,Cxcl9,Fas,Fcgr1 a,Jak3,Lrg1,Nos2,Ptprc,Socs1,Socs4,Sp1) in the Bushen Shengsui group was obviously different compared with the model group. from which up-regulated genes has Prlr; the rest of genes were down-regulated.(2)Result of the second experimentDetected the protein of JAK2 and STAT3 in JAK/STAT signal pathway by Western Blot. Compared with the model group, the expression of JAK2 protein and STAT3 protein was significantly decreased(P<0.01) in the sham group. Compared with the model group, the expression of JAK2 protein was significantly decreased(P<0.01) in the AG490 blocking agent group,Yiqi Huoxue group and Busheng Shengsui group,the expression of STAT3 protein was significantly decreased(P<0.01) in the AG490 blocking agent group and Yiqi Huoxue group,and the expression of STAT3 protein was obviously decreased(P<0.05) in the Busheng Shengsui group. Compared with the Yiqi Huoxue group, the expression of JAK2 protein and STAT3 protein have no obvious difference in the Busheng Shengsui group.4)ConclusionThe experiment has confirmed that Yi Qi Huo Xue recipe and Bu Shen Sheng Sui recipe can faciliate the recovery of focal cerebral ischemia and the proliferation and neuronal differentiation of neural stem cells. The above recipes can influence the JAK/STAT signals’ expression through lowering the JAK2 and STAT3 signal and thus influence the proliferation and neuronal differentiation of neural stem cells after stroke.