|
Purpose: To study on that intravenous injection of radix salviae miltiorrhizae, bone marrow mesenchymal stem cells(BMMSCs) by vein transplantation,and this traditional chinese madichine combined with BMMSCs are how to effect on myocardial collagen accumulation after acute myocardial infarction in rabbits,in order to explore a novel therapeutic avenue against myocardial fibrosis after myocardial infarction.Material and method:1.Materials:A total of 72 healthy Japanese White Rabbits were purchased from Experimental Animal Center,Liaoning University of Traditional Chinese Medicine.2.Methods:Firstly,bone marrow mesenchymal stem cells(BMMSCs) were obtained from the bone marrow of two rabbits by isolation,culture,proliferation,identification,purification,and labelled with BrdU before vein transplantation.Secondly,ten rabbits randomly serves as a normal group,and sixty rabbits were used to established the models of acute myocardial infarction by ligation of the left ventricular branch of coronary artery.Forty rabbits which were sucessfully replicated animal model of acute myocardial infarction were randomly divided into four groups of ten rabbits each:model control group,intravenous injection of radix salviae miltiorrhizae group,BMMSCs by vein transplantation group, radix salviae miltiorrhizae combined with BMMSCs group.After the surgery,experimental groups,immediately through their ear vein,were intravenously injected danshen injections or(and)1ml BMMSCs(2×106 cells). 24 hours after myocardial infarction,the expression of peripheral blood serum creatine kinase-MB(CK-MB),matrix metalloproteinase-1(MMP-1), tissue inhibitor of metalloproteinase-1(TIMP-1),C-terminal propeptide of procollagen typeⅠ(PⅠCP) and N-terminal peptide of procollagen typeⅢ(PⅢNP) were detected by ELISA analysis.Five weeks later, the levels of peripheral blood serum MMP-1, TIMP-1, PⅠCP and PⅢNP were analyzed again. Meantime,the left ventricular tissues were sampled to experiment on that:the myocardium of infarct area was tested by BrdU immunohistochemical staining,and positive area would be brown yellow;Myocardial cell degeneration and necrosis were observed by HE staining;the fibrous structure of myocardial stroma was observed,and collagen volume fraction(CVF) was measured by masson’s trichrome staining.Results:1.The content of peripheral blood serum CK-MB was significantly increased at 24 hours after establishment of AMI that suggested massive necrosis of myocardial cells.2.BrdU-positive BMMSCs were appeared in and around the infarcted zone of cell transplantation groups.3.HE staining and masson’s trichrome staining showed that,in experimental groups, there were different degrees of collagenous fibers deposition in the infarcted zone, on the contrary, many myocardial fibers in order rules were lost.4.CVF:5 weeks after AMI, the differences among groups were not statistically significance(P>0.05);but compared with the normal group,collagen volume fraction tended to decrease in other groups.5.Blood serum MMP-1:24 hours after AMI,compared with the model control group or the BMMSCs transplantation group, the expression of blood serum MMP-1 were dramatically decreased in the danshen group and combined treatment group(P<0.01).At 5 weeks following AMI, the differences among groups were not statistically significance(P>0.05).6.Blood serum TIMP-1:24 hours or 5 weeks after AMI, the differences among groups were not statistically significance(P>0.05).7.Blood serum PⅠCP:24 hours after AMI,compared with the model control group or the BMMSCs transplantation group,the expression of blood serum PⅠCP were dramatically decreased in the danshen group and combined treatment group(P<0.01). At 5 weeks following AMI, compared with the model control group, the levels of blood serum PⅠCP were lower in the combined treatment group(P<0.01)and higher in the danshen group(P<0.05);and compared with the combined treatment group,the levels were dramatically higher in the combined treatment group than in the model or BMMSCs transplantation group(P<0.01).8.Blood serum PⅢNP:24 hours after AMI, compared with the model control group,the level of blood serum PⅢNP was dramatically higher in the BMMSCs transplantation group(P<0.01);compared with the BMMSCs transplantation group, the levels were dramatically decreased in the model control group, danshen group and combined treatment group(P<0.01). At 5 weeks following AMI, the differences among groups were not statistically significance(P>0.05).Conclusion:1.The models of acute myocardial infarction could be induced successfully by ligation of the left ventricular branch of coronary artery.2.Transplanted BMMSCs intravenously could migrate into the infarcted myocardium.3.Radix salviae miltiorrhizae attenuated myocardial fibrosis after myocardial infarction through regulation of MMPs/TIMPs system and inhibition of collagen synthesis.4.In this study,it was not found that BMMSCs transplantation though vien immediately after AMI would improve myocardial fibrosis,which may be associated with the timing of transplantation BMMSCs.5.Radix salviae miltiorrhizae combined with BMMSCs transplantation could also attenuat myocardial fibrosis after myocardial infarction through regulation of MMPs/TIMPs system and inhibition of collagen synthesis.But, because of limited role of BMMSCs in this experiment, do not rule out the major role to play by danshen.
|