Effect Of MiR-107 Transfected Bone Marrow Mesenchymal Stem Cells On Brain Microvascular Endothelial Cells Under Simulated Intracerebral Hemorrhage

Objective: Intracerebral hemorrhage is a type of cerebrovascular disease with high morbidity,morbidity and mortality.Foreign and domestic studies have confirmed that bone marrow mesenchymal stem cells can promote neurological recovery in rats under intracerebral hemorrhage through a variety of mechanisms.With further research,we have found that micro RNA has a certain relationship with the recovery of cerebral hemorrhage.Previous studies have shown that mi R-107 is decreased in the peripheral blood with cerebral hemorrhage in human beings.So far,mi R-107 has been found in Alzheimer’s disease,ischemic cardiovascular disease,glioma and other tumors,which plays in different ways.However,the role is not yet clear in cerebral hemorrhage.Therefore,we transfected the rno-mi R-107 to bone marrow mesenchymal stem cells in vitro,and observed the changes of vascular endothelial growth factor(VEGF)expression and the proliferation ability of co-cultured rats brain microvascular endothelial cells.It may provide a new theoretical basis for the future treatment of cerebral hemorrhage by bone marrow mesenchymal stem cells.Methods: The experiment is studied in vitro of intracerebral hemorrhage.Firstly,rno-mi R-107 mimics,inhibitor,Nc are transfected into BMSCs under normoxia and the transfections of gene are detected by real-time quantitative PCR(q RT-PCR).Further,establishing the moderl of cerebral hemorrhage in vitro,the expression of vascular endothelial growth factor(VEGF)secreted by BMSCs is detected by q RT-PCR and Western blot.Finally,the transfected BMSCs are co-cultured with RBMECs.The changes of VEGF expression and proliferation of RBMECs are detected by Western-blot and CCK-8.Results: 1.In this study,rno-mi R-107 negative control(Nc group),rno-mi R-107 mimics(mimics group)and rno-mi R-107 inhibitor(inhibitor group)were transfected in BMSCs.We detected transfection efficiencies of them by q RT-PCR.The results showed that mi R-107 expression was significantly increased in mimics group compared with Nc group and mi R-107 expression was decreased in inhibitor group.The difference between the two groups was statistically significant(P < 0.05).Compared with the control group,the expression of mi R-107 in Nc group had no significant difference(P> 0.05).We established mi R-107 overexpression and silencing BMSCs model.2.Establishing the model of intracerebral hemorrhage in vitro,BMSCs were exposed to the condition of oxygen-glucose deprivation(OGD)4 hours and hemin(10u M).Expression of VEGF were detected by using q RT-PCR and Western blot.The results showed that compared with Nc group,the m RNA and protein expression of VEGF in mimics group were increased and VEGF m RNA and protein expression in inhibitor group were decreased(P <0.05).Compared with Control group,VEGF expression in Nc group had no significant difference(P> 0.05).It was suggested that overexpression of mi R-107 can increase the secretion of VEGF in BMSCs,while silence of mi R-107 can decrease the secretion of VEGF in BMSCs.3.RBMECs were co-cultured with BMSCs in system of transwell.The experiment was divided into five groups: RBMECs group(no BMSCs co-cultured,only RBMECs),RBMECs+ Control-BMSCs group(untransfected BMSCs co-cultured with RBMECs),RBMECs+ mi R-107 Nc-BMSCs group(RBMECs co-cultured with BMSCs transfected with rno-mi R-107 negative control),RBMECs+ mi R-107 mimics-BMSCs group(RBMECs co-cultured with BMSCs transfected with rno-mi R-107 mimics)and RBMECs+ mi R-107 inhibitor-BMSCs group(RBMECs co-cultured with BMSCs transfected with rno-mi R-107 inhibitor).We detected expression of VEGF in RBMECs by Western blot.The results showed that compared with the RBMECs group,the expression of VEGF in other four groups co-cultured with BMSCs were significantly increased(P <0.05).Compared with RBMECs+ mi R-107 Nc-BMSCs group,the expression of VEGF was increased in mimics group and decreased in inhibitor group,and the difference between the two groups was statistically significant(P <0.05).Compared with RBMECs+ Control-BMSCs group,VEGF expression in Nc group had no significant difference(P> 0.05).4.The co-culture systems were same(such as 3).The proliferations of RBMECs were detected by CCK-8.The results showed that compared with the RBMECs group,proliferation abilities of other four groups co-cultured with BMSCs were increased(P<0.05).Compared with RBMECs+ mi R-107 Nc-BMSCs group,the cell proliferation of mimics group was enhanced and the inhibitor group was weaker than that of Nc group(P<0.05).There was no significant difference in cell proliferation between Nc group and control group(P> 0.05).Conclusions: 1.Under the simulated model of intracerebral hemorrhage in vitro,overexpression of mi R-107 can increase the expression of VEGF.2.BMSCs and RBMECs were co-cultured in vitro with intracerebral hemorrhage.RBMECs co-cultured with BMSCs showed more VEGF expression than that of in RBMECs only.RBMECs co-cultured with overexpression mi R-107 showed stronger ability of cell proliferation.