The Study Of Astragaloside Ⅳ Of Protective Effects Of Bone Marrow Stromal Stem Cells Apoptosis Which Is Induced Of Hypoxia Ischemia Through MAPK Pathways

Objective Even though existing studies show that bone marrow stromal stem cells play a role in repairing cardiomyocytes by transplanting to cardiomyocytes,bone marrow stromal stem cells(bone marrow stem cells,BMSCs)have a poor viability after transplant.The aim of this study is to explore whether As-Ⅳ(Astragaloside),a traditional Chinese medicine astragalus extract with main pharmacological effects of enhancing immunity,resisting virus,inflammation and oxidation,can protect BMSCs via MAPK pathway.Methods Cultured the BMSCs taken from SD rats’bone marrow in tibia and femur.Used those BMSCs cultured for three generations at least on this experiment.Then took each dish of cells as a single group.Model group is group which makes BMSCs apoptosis by serum deprivation and hypoxia.Blocking agent group is group which achieves the effect of blocking pathway through testing blockers on each level of Unicom Roads under MAPK:ERK,JNK and P38.Once finished reorgainsing two groups,we used flow cytometry and CCK8 method to detect each group’s cell apoptosis rate and cell viability respectively.Under flow cytometry,cells were divided into normal group,model group,model+As-Ⅳ(10ug/ml)group,model+As-Ⅳ(20ug/ml)group,model+As-Ⅳ(40ug/ml)group and model+As-Ⅳ(80ug/ml)group.Afterwards,detected each group of apoptosis rate.Under CCK8 method,cells were divided into normal group,DMSO group,model+As-Ⅳ(lOug/ml)group,model+As-Ⅳ(20ug/ml)group,model+As-Ⅳ(40ug/ml)group and model+As-Ⅳ(80ug/ml)group.Then detected cell viability.Under Western blot analysis,cells were detected their P38,JNK and ERK1/2,p-p38,p-JNK,p-ERK1/2,mitochondrial CytC,Bcl-2 and Bax protein expression level after researches done by normal group,model group,model group+As-Ⅳ,model+blocking agent group,model+blocking agent+As-Ⅳ group.Results Under flow cytometry,cell apoptosis rates increased(P<0.01)in model group,model+As-Ⅳ(10ug/ml)group,model+As-Ⅳ(20ug/ml)group,model+As-Ⅳ(40ug/ml)group while no significant change in model+As-Ⅳ(80ug/ml)group when comparing with normal group.The number of cell apoptosis achieved its top level at the 6 hour after model built,which can be found via hoechst33342 decoration method.Under CCK8 method,we observed cell viability in its 24h,48h,72h.Model+As-Ⅳ(80ug/ml)group had the greatest viability(P<0.01)when comparing with other groups in the time of 24h and 48h respectively.There were no statistical significance among model+As-Ⅳ(80ug/ml)group,model+As-Ⅳ(20ug/ml)group and model+As-Ⅳ(40ug/ml)group in the time of 72h,while comparing with normal group,model+As-IV(80ug/ml)group cell viability increased and had a statistical difference of P<0.05.Within these three time periods,cell vaibilties have no statical significane in model+As-Ⅳ(80ug/ml)group.As to Western blot method,we gained that under Unicom Roads of ERK and JNK,model group p-ERK,p-JNK got higher,mitochondrial cyt c,Bcl-2 and Bax lower than normal group on protein expression level;in model+As-Ⅳ(80ug/ml)group,p-ERK,mitochondrial Cyt C,Bcl-2 and Bax higher than model group,p-JNK lower.In model+blocking agent group,p-ERK lower than model group while mitochondrial cyt c,Bcl-2 and Bax higher,no siginificant difference in p-JNK.In model+blocking agent+As-Ⅳ group,p-ERK,p-JNK lower than those in model+blocking agent group,however,mitochondrial Cyt C,Bcl-2 and Bax higher.All of five groups in Unicome Road P38 level had no statistical difference.Conclusion The protective effect of As-Ⅳ on BMSC reaches its strongest when concentration is 80ug/ml.MAPK pathway was involved in the progress of preventing BMSCs apoptosis as well.It is through the activation of ERK Unicom Road that As-Ⅳ inhibited JNK Unicom Road to resist BMSCs apoptosis,not through the level of P38.