Combination Of Ultrasound Microbubble Mediated Granulocyte Colony-stimulating Factor And Mesenchymal Stem Cells Transplantation To Treat Acute Myocardial Infarction In Rats

Part ? : Isolation,cultivation and identification of bone marrow source MSCs in ratsObjective:To explore the approaches of isolation,cultivation and identification of the bone marrow mesenchymal stem cells(MSCs)in rats.Methods: The isolation,purification and cultivation of bone marrow MSCs in rat bone marrow were carried out by density gradient centrifuge joint wall culture method,and its biological characteristics were detected.Cell surface markers were detected by cellular immunochemical and flow cytometry.Results: The culture of MSCs adhered to the wall growth,mainly in the spindle shape;The CD44 is highly expressed on Cell surface.The CD45 is lower expressed and the CD34 is not expressed.Conclusion: Using the method of applying the density of 1.073 g/ml Percoll separation fluid gradient centrifuge,the bone marrow MSCs with higher purity were obtained by the method of lining the wall.It can be expanded to 107 cell orders of magnitude in 2w,which is in line with the requirements of cell transplantation of myocardial infarction cells.PART ? : The study of ultrasound microbubble mediated G-CSF to promote MSCs homing rat ischemic myocardiumObjective: To observe the role of ultrasound microbubble mediated granulocyte colony-stimulating factor(G-CSF)to the myocardial ischemia in rats with bone marrow mesenchymal stem cells in rats.Methods: Wistar rats successfully prepared with 30 MI models were randomly divided into 5 groups: A: blank control group(C),B: simple MSCs group(MSCs),C:ultrasound microbubble+MSCs group(US/MB+MSCs),D: ultrasound microbubble +G-CSF group(US/MB+G-CSF),E: ultrasound microbubble+G-CSF+MSCs group(US/MB+G-CSF+MSCs);After 48 h transplantation,the animals were killed and the marginal tissue of myocardial infarction was cut.The distribution of MSCs in this site(myocardial ischemia)was observed by confocal laser scanning microscope..Results: The MSCs of ultrasound microbubble+G-CSF+MSCs group were significantly higher than those of other groups by laser confocal microscope.Conclusion: Ultrasound microbubble mediated G-CSF can significantly promote MSCs homing to myocardial ischemia in rats.PART III: Combination of ultrasound microbubble mediated G-CSF and MSCs transplantation to treat acute myocardial infarction in ratsObjective: To study the feasibility of the combination of ultrasound microbubble mediated G-CSF and MSCs to treat acute myocardial infarction in rats.Methods: Wistar rats successfully prepared with 30 MI models were randomly divided into 5 groups: A: blank control group(C),B: simple MSCs group(MSCs),C:ultrasound microbubble+MSCs group(US/MB+MSCs),D: ultrasound microbubble+G-CSF group(US/MB+G-CSF),E: ultrasound microbubble + G-CSF + MSCs group(US/MB+G-CSF+MSCs);The left heart function and heart size of all the rats were detected by echocardiography 4w after implantation.Then the rats were killed and the myocardium were harvested.The CD34 expression was detected by immunohistochemistry,and microvessel density(MVD)was counted in high-powerfield on microscopy.The expression level of VEGF protein was detected by Westernblot and ELISA.Results: Echocardiography showed that cardiac function improved significantly in ultrasound microbubble+G-CSF+MSCs group,ultrasound microbubble+MSCs group and ultrasound+G-CSF group.The expression of CD34 and VEGF protein improved significantly and microvessel density is also higher than the other groups,microbubbles+G-CSF+MSCs group was the highest,the difference was statistically meaning(P<0.05).There was no significant difference between ultrasound microbubble + MSCs group and ultrasound microbubble+G-CSF group.Conclusion: Ultrasound microbubble mediated G-CSF and MSCs transplantation can be used to treat myocardial infarction and improve cardiac function by promoting cardiac angiogenesis.