过表达整合素连接激酶促进心脏c-Kit~+细胞的存活和增殖

目的:探讨整合素连接激酶(ILK)过表达对心脏c-Kit~+细胞存活和增殖的影响,以及移植ILK过表达的心脏c-Kit~+细胞改善心肌梗死(MI)大鼠心功能的作用。方法:从新生SD大鼠分离培养心脏c-Kit~+细胞,构建含人ILK全长的腺病毒载体并感染心脏c-Kit~+细胞。感染48 h后,应用细胞计数和CCK-8法测定细胞活力和增殖,并通过Western blot法检测c-Kit~+细胞中增殖相关蛋白cyclin D1和增殖细胞核抗原(PCNA)的表达。40只8周龄SPF级大鼠经冠状动脉结扎建立MI模型,结扎15 min后于梗死区及梗死边缘区分3点通过心肌注射移植心脏c-Kit~+细胞。采用随机数字表法将大鼠分为假手术组(sham组)、心梗+生理盐水注射组(MI组)、心梗+感染空载体的c-Kit~+细胞注射组(Ad-null-c-Kit~+cell组)和心梗+感染携带ILK腺病毒载体的c-Kit~+细胞注射组(ILKc-Kit~+cell组),每组10只。术后2周,免疫组化法检测梗死区和梗死边缘区c-Kit的表达;术后4周,通过血流动力学检测心脏功能。结果:过表达ILK促进了心脏c-Kit~+细胞的存活和增殖(P0.05)。过表达ILK的心脏cKit+细胞中cyclin D1和PCNA的表达明显增加(P0.05)。细胞移植2周后,ILK-c-Kit~+cell组心肌组织中c-Kit表达明显增加(P0.05);细胞移植4周后,ILK-c-Kit~+cell组大鼠心功能改善比Ad-null-c-Kit~+cell组更明显(P0.05)。结论:过表达ILK通过提高cyclin D1和PCNA的表达,促进了心脏c-Kit~+细胞的存活和增殖。ILK-c-Kit~+细胞移植能更好地改善MI大鼠的心功能。

Overexpression of integrin-linked kinase improves survival and proliferation of cardiac c-Kit+ cells

AIM: To investigate the effects of integrin-linked kinase (ILK) overexpression on survival and proliferation of cardiac c-Kit⁺ cells, and the role of ILK-overexpressing c-Kit⁺ cell transplantation in cardiac function in a rat myocardial infarction (MI) model.METHODS: Cardiac c-Kit⁺ cells were isolated from the hearts of neonatal Sprague-Dawley (SD) rats and cultured to prepare the ILK-c-Kit⁺ cells by infected with recombinant adenoviral vector harboring human wild-type ILK cDNA. The survival and proliferation of cardiac c-Kit⁺ cells were detected by cell counting and CCK-8 assay at 48 h after infection, respectively. The protein levels of cyclin D1 and proliferating cell nuclear antigen (PCNA) in the cardiac c-Kit⁺ cells were examined by Western blot. MI was induced by coronary artery ligation in 40 adult rats. After 15 min, ILK-c-Kit⁺ cells were transplanted into the hearts by myocardial injection at 3 different sites in the infracted zone and border zone. All rats were randomly divided into 4 groups:sham group, MI plus saline injection group (MI group), MI plus null vector-infected cardiac c-Kit⁺ cell injection group (Ad-null-c-Kit⁺ cell group), and MI plus ILK-overexpressing cardiac c-Kit⁺ cells injection group (ILK-c-Kit⁺ cell group), with 10 rats in each group. At 2 weeks after MI, the protein levels of c-Kit in MI hearts were investigated by immunohistochemical assay. At 4 weeks, left ventricular function was examined by hemodynamic measurement.RESULTS: The survival and proliferation of cardiac c-Kit⁺ cells and the protein levels of cyclin D1 and PCNA were enhanced by ILK overexpression compared with Ad-null group. In MI rat model, the number of c-Kit⁺ cells was increased by ILK-c-Kit⁺ cell injection compared with Ad-null-c-Kit⁺ cell group at 2 weeks after MI. Cardiac function was significantly improved in ILK-c-Kit⁺ cell-transplanted rats.CONCLUSION: ILK overexpression improves survival and proliferation of cardiac c-Kit⁺ cells by increasing the protein levels of cyclin D1 and PCNA. ILK-c-Kit⁺ cell transplantation enhances the therapeutic efficiency of cardiac c-Kit⁺ cells in the post-MI hearts of rats.