| Objective To investigate the impact of mesenchymal stem cells conditioned medium on myocardial cell apotosis, the autologous bone marrow mesenchymal stem cells (MSCs) or its conditioned medium were transplanted to swine through Digital Subtraction Angiography after sucessfully creating the model of myocardial infarction in swine by embolization of the left coronary artery with a gelation sponge through cardiac catheter .Methods Thirty swines were randomly divided into six groups. Group A: control 1 group, swines were injected with DMEM after AMI 75 mins; Group B: MSCs 75min group, swines were injected with MSCs after AMI 75 mins; Group C: MSC-CM 75min group, swines were injected with MSC-CM after AMI 75 mins; Group D: control 2 group, swines were injected with DMEM after AMI 4 weeks; Group E: MSCs 4w group , swines were injected with MSCs after AMI 4 weeks; Group F: MSC-CM 4w group , swines were injected with MSC-CM after AMI 4 weeks; Myocardial infarction LAD Model is established by gelatin sponge embolization. Four weeks later, hemodynamics were used to assess the heart function. 30 mL bone marrow was taken from the iliac of each swine, separated and purified by density centrifuge and adherent screening. The autologous bone marrow stem cells (2×106 / mL, 5 mL) or mesenchymal stem cells conditioned medium (5 ml) were injected transcatheterly into the remote embolization of LAD. The change of each index of cadiac function was detected by Powerlab8SP and Millar catheter. In the end , swines'hearts were examined. The area of myocardial infarction was tested. In the 75min group, the product of mycatdial nuclear oxidative stress was determined by immunostaining for 8-hydroxy-2`-deoxyguanosine(8-OHdG). In addition, the protein expression level of phospho-SMAD2 and active caspase 3 were determined by Western blot. In the 4w group, the protein expression of active caspase 3 were also estimated . Vessel number was also counted by immuno-histochemisty staining with anti Factor-Ⅷ antibody. The expression of VEGF and HGF in MSC-CM by ELISA were tested as well.Results 75min groups:(1) Left ventricular function, including LVDP,+dp/dtmax,-dp/dtmax was significantly improved in group C comparied with that in group A and B (P<0.05). (2)MSC-CM treratment significantly reduced the level of 8-OhdG as determined by immunostaining comparied with that of group A and B (P<0.05). (3)The expression of pSMAD2 and active caspase 3 singificantly decreased in group C comparied with that of group A and B. 4w groups: (1)Left ventricular function, including LVDP,+dp/dtmax,-dp/dtmax was significantly improved in group E comparied with that in group D and F (P<0.05).(2)Immunohisto-chemistry showed that the number of vessels in group E significantly increased comparied with that of group D and group F (P<0.05).(3)The expression of active caspase 3 singificantly decreased in group E comparied with that of group D and F (P<0.05).Conclusions Fristly, MSC-CM showed significant anti-apoptotic effects on cardiac myocytes with the possible mechanism of reducing oxidative stress and TGF-βsecretion which influenced the expression of active caspase 3 in the myocytes and enhanced the cardiac function . Secondly, the implantation of MSCs into the infarcted myocardum increased the number of neovascularization, which could improve the supply of blood and oxygen to myocardium. Thirdly, In the acute phase after myocardial infarction , transplanting MSCs can significantly reduce apoptosis of cardiac myocyte through the paracrine effects of MSCs ,the effect should be much better than stem cell transplantation; in the chronic period of myocardial infarction, MSCs showed anti-apoptotic effects on cardiac myocytes by improving the density of angiogenesis in infarction areas, and the effect of transplanting is better than MSCs-CM transplantation;Finally ,there should be a complex mechanism for the cytoprotection of MSC-CM and might be a varity of cytokines.
|
PDF Full Text Request