Role And Mechanism Of Conditioned Medium Of Human Umbilical Cord-Derived Mesenchymal Stromal Cells On Irradiation-Induced Myocardial Fibrosis

PART I Effects of Conditioned Medium of Human Umbilical Cord–Derived Mesenchymal Stromal Cells on Cardiac Function,Inflammatory Response and Myocardial Fibrosis in Rats with Irradiation-Induced Cardiac InjuryObjective: To investigate the effects of myocardial injection of conditioned medium of human umbilical cord–derived mesenchymal stromal cells on cardiac function,inflammatory response and myocardial fibrosis in rats with irradiation-induced cardiac injury.Methods: Human umbilical cord-derived mesenchymal stem cells were isolated and cultured,and conditioned medium was thencollected.SD rats were randomly divided into sham operation group(Sham group),X-ray irradiation group(Ir group),X-ray irradiation with control medium injection group(Ir+DMEM group),irradiation with conditioned medium of human umbilical cord–derived mesenchymal stromal cells injection group(Ir+MSCCM group).From the early stage of feeding to 10 weeks after irradiation,the cardiac function of each group was measured by echocardiography,and the expression levels of inflammatory factors and myocardial injury markers in serum of each group were detected by serological examination.HE and PSR staining of myocardial tissue were observed for the changes of myocardial structure,and the real-time quantitative PCR was used to detect the m RNA content of fibrosis markers in myocardial tissue of rats.Results: Compared with the control medium,echocardiographic results showed that conditioned medium of human umbilical cord–derived mesenchymal stromal cells injection significantly improved left ventricular ejection fraction and inhibited the level of cardiac hypertrophy in rats with irradiation-induced heart injury.Pathological staining and PCR results also found that the conditioned medium injection significantly inhibited the degree of myocardial fibrosis in rats with irradiation-induced heart injury.In addition,serological examination revealed that conditioned medium also significantly reduced the inflammatory state and myocardial damage in rats with irradiation-induced heart injury.Conclusion: Conditioned medium of human umbilical cord–derived mesenchymal stromal cells could reduce the degree of myocardial damage,inflammatory state and myocardial fibrosis,which thus promoting the recovery of cardiac function in rats with irradiation-induced heart injury.PART Ⅱ Effects of Conditioned Medium of Human Umbilical Cord–Derived Mesenchymal Stromal Cells on the Activity,Antioxidant Capacity,Inflammatory Response and Collagen Synthesis of Cardiac Fibroblasts after IrradiationObjective: To investigate the effects of conditioned medium of human umbilical cord–derived mesenchymal stromal cells on the activity,antioxidant capacity,inflammatory response and collagen synthesis of cardiac fibroblasts after X-ray irradiation.Methods: Human umbilical cord–derived mesenchymal stromal cellswere isolated,cultured and identified.Conditioned medium of human umbilical cord–derived mesenchymal stem cells was collected.Heart fibroblasts were randomly divided into control group(Control group),X-ray irradiation group(Ir group),andirradiation with conditioned medium of human umbilical cord-derived mesenchymal stem cells.After intervention,CCK8 was used to measure the cell viability of cardiac fibroblasts.Cell supernatant was used to detect the content of MDA,total SOD,CAT and GPx in cardiac fibroblasts after irradiation,and real-time quantitative PCR was used to detect the fibrosis markers in cardiac fibroblasts after irradiation.The m RNA contents of oxidases SOD1,SOD2,CAT and GPx were used to evaluate the antioxidant capacity of cardiac fibroblasts;ELISA assay and PCR were combined to detect the inflammatory factors IL-6,IL-8 in the supernatant of cardiac fibroblasts.Finally,the ELISA kit combined with real-time PCR was to detect the collagen level including Col1A1 and Col1A2 in supernatant ofcardiac fibroblast.Results: Compared with the irradiated group,conditioned medium of human umbilical cord-derived mesenchymal stem cells significantly inhibited the death number of irradiation-induced cardiac fibroblast,improved the anti-oxidative capacity,and inhibited the synthesis of profibrotic proteins such as TGF-β1,Col1A1 and Col1A2 in cardiac fibroblastation after irradiation.Conclusion: Conditioned medium ofhuman umbilical cord-derived mesenchymal stem cells can significantly improve the vitality,antioxidant capacity,and inhibited inflammation degree and collagen accumulation of cardiac fibroblasts induced by irradiation.PART Ⅲ Conditioned Medium of Human Umbilical Cord-Derived Mesenchymal Stem Cells may improve radioactive myocardial fibrosis by inhibiting NF-κB signaling pathwayObjective: To investigate the possible mechanism of conditioned medium of human umbilical cord-derived mesenchymal stem cells in improving irradiation-induced myocardial fibrosis in vitro.Methods: Human umbilical cord-derived mesenchymal stem cells were isolated,cultured and identified.Conditioned medium of human umbilical cord mesenchymal stem cell was collected.Cardiac fibroblasts were randomly divided into control group(Control group),X-ray irradiation group(Ir group)and irradiation with conditioned medium of human umbilical cord-derived mesenchymal stem cells group(Ir+MSCCM group).The expression of NF-κB p65 was detected in each group of cardiac fibroblasts.Then,NF-κB inhibitor was administered tofurther confirme whether the NF-κB signaling pathway actually affect the synthesis of profibrotic factors in cardiac fibroblasts after Thus,it was also to detecte the content of TGF-β1 in the control group,X-ray group and irradiation with NF-κB inhibitor in the cardiac fibroblasts.Results: Compared with the control group,the NF-κB signaling pathway was activated in the cardiac fibroblasts after irradiation.Treated with NF-κB inhibitor,the synthesis of TGF-β1 in cardiac fibroblasts was significantly reduced after irradiation;while conditioned medium ofhuman umbilical cord-derived mesenchyme stem cells significantly inhibited activation of the NF-κB signaling pathway following irradiation induction.Conclusion: Conditioned medium of human umbilical cord mesenchymal stem cells might enhance the activity of cardiac fibroblasts,increase the antioxidant capacity of cells,reduce the accumulation of collagen,and inhibit the inflammatory reaction by inhibiting NF-κB signaling pathway,which then possibly iinhibit myocardial fibrosis induced by irradiation.