Mesenchymal Stem Cells Pretreatment By Ligustrazine Improves The Treatment Of A Rat Model Of Cerebral Ischemia Via Enhanced Migratory Of Implanted Cells

Objective To observe bone marrow mesenchymal stem cells(BMSCs)pretreatment by Ligustrazine improves the treatment of a rat model of cerebral ischemia via enhanced migratory of implanted cells and invstigate its possible mechanism.Methods ①Isolated and cultured BMSCs by the whole bone marrow adherent method,at the third passage,conduct phenotypic identification using Flow cytometry analysis of surface positive antigen of CD29,CD90 and the negative antigen of CD34,CD45.Different of the final concentrations of BrdU(5 μmol/L,10 μmol/L and 20 μmol/L)labeled BMSCs 24 h,48 h and 72 h,measured the BrdU labeling rate.②Rats were subjected to middle cerebral artery occlusion(MCAO)for 90 minutes,sham group,model group,MCAO+BMSCs(BMSCs group),MCAO+BMSCs pretreatment by 50 μM Ligustrazine(50 μM Ligustrazine group),MCAO+BMSCs pretreatment by 100 μM Ligustrazine(100 μM Ligustrazine group),BMSCs pretreat:ment by 50μM Ligustrazine+CXCR4 inhibitor AMD3100(AMD3100 group),n=12,the sham group and model group were injected withlmL PBS,other groups were injected with rats BMSCs solution of ImL(1×106cells)via the tail vein,were administered randomly 24 h after ischemia.③Modified neurological severity score(mNSS),adhensive removal test and the comer test were used to evaluate sensorimotor on 1,7,14 and 28 days after ischemia.④The expression of BrdU was detected by immunofluorescence,and the volume of cerebral infarction were stained with toluidine blue on 28 days after ischemia.⑤Fourteen days after ischemia,the expression of BrdU,SDF-1,vWF,VEGF and BDNF was detected by immunofluorescenceResults ①The whole bone marrow adherent method can obtained the well-growing and get a homogeneous morphology BMSCs.The BMSCs phenotype of the third passage,the positive expression rates of CD29,CD90,CD34 and CD45 were 99.0%,99.3%,0.2%and 4.1%.It’s in line with the characteristics of BMSCs,confirmed the cells we separated were BMSCs.The BrdU labeling rate did not relationship with labeled concentration level and labeled time,a final concentration of BrdU labeled BMSCs at 48 h was 10 μmol/L,labeling highest rate of 74.6%.②Primed with Ligustrazine(50 μM and 100 μM)group significantly ameliorated neurological dysfunction,reduced the time of adhensive removal and the number of right turn significantly reduced at 7,14 and 28 days after ischemia(P<0.01 or P<0.05).③Compared with model group and BMSCs group,Ligustrazine(50 μM and 100 μM)group,the infarct volume significant reduction at 28 days after ischemia(P<0.01 or P<0.05).④Compared with model group,BMSCs group and AMD3100 group,pretreatment by Ligustrazine(50 μM and 100 μM)group the number of BrdU,SDF-1,vWF,VEGF and BDNF was significantly increased(P<0.01)Conclusion Isolated and cultured BMSCs by the whole bone marrow adherent method can obtained the purity of BMSCs.BMSCs group primed with(50 μM and 100 μM)Ligustrazine better than the single BMSCs group in the treatment of Cerebral Ischemia,their could accelerates functional recovery after cerebral ischemic via enhanced migratory of implanted cells,improved angiogenesis,reduce the volume of infarct.BMSCs group primed with Ligustrazine significantly improved neurological function,increased neurogenesis and angiogenesis.The effects may be related with the transplanted cells promoted angiogenesis in the surrounding area of cerebral infarction and increased the expression of vWF,SDF-1,VEGF and BDNF.