ILK转染的BM-MSCs通过旁分泌作用抑制CFB增殖和胶原合成

摘要

Objective To investigate the mechanism for attenuating cardiac fibrosis after myocardial infarction with ILK-MSCs transplantation in vitro.Methods BM-MSCs were infected by ad-ILK-GFP in serum-free medium.After cultured for 72 h, the conditioned medium was collected from ILK-MSCs (ILK-MSCs-CM group) and MSCs (MSCs-CM gourp), respectively, which were cocultured with cardiac fibroblast (CFB) for 48 h.The CFB cultured ir the high glucose medium containing 5 ％ fetal bovine serum was taken as the control group.The ability of CFB proliferation was measured with an EdU proliferation Kit.And RT-PCR was performed to assess the gene expressions of types Ⅰ and Ⅲ collagen(Collal andCol3al) ,matrix metalloproteinases(MMPs) ,tissue inhibitor of metalloproteinases(TIMPs) in CFB.Results Compared with control group, CFB proliferation was inhibited and the expressions of Collal, Col3al, TIMP-1 and TIMP-2 were reduced, and the expressions of MMP-2 and MMP-9 were increased in groups of MSCs-CM and ILK-MSCs-CM (P＜0.05).Compared with MSCs-CM group, the inhibition of the expressions of Col3a1 and TIMP-2and the increase of MMP-2 expression were more in ILK-MSCs-CM group (P＜0.05).Conclusion ILK may attenuate fibrosis after myocardial infarction in vivo, possibly through improving the paracrine action of MSCs and reducing CFB and collagen synthesis.%目的 探讨体外移植整合素连接激酶(ILK)转染的骨髓间充质干细胞(BM-MSCs)抑制心肌梗死后心脏纤维化的机制.方法 收集培养72 h的MSCs和ILK-MSCs后的培养基,分别与心脏成纤维细胞(CFB)共培养48 h,用5-乙炔基-2'脱氧尿嘧啶核苷(EdU)增殖试剂盒检测CFB的增殖能力,采用实时荧光定量PCR检测CFB表达金属蛋白酶(MMPs)、金属蛋白酶抑制剂(TIMPs)及细胞外基质的组成成分I型胶原和Ⅲ型胶原的情况.结果 与对照组比较,MSCs培养液组、ILK-MSCs培养液组可抑制CFB的增殖和减少Col1a1、Col3a1、TIMP-1、TIMP-2和a-SMA的基因表达(P<0.05),增加MMP-2、MMP-9的基因表达(P<0.05);与MSCs培养液组比较,ILK-MSCs的培养液组可进一步抑制Col3a1和TIMP-2的基因表达(P<0.05),并增加MMP-2的基因表达(P<0.05).结论 ILK可能通过增加MSCs的旁分泌,抑制CFB的增殖和胶原合成能力,从而在体内发挥降低心肌梗死后纤维化的作用.