论文部分内容阅读
目的利用β-甘油磷酸盐处理人血管平滑肌细胞(human vascular smooth muscle cells,HVSMCs)制备体外血管钙化模型。方法用组织贴壁法从人胚胎脐带动脉中分离原代人主动脉平滑肌细胞,原代细胞通过抗体α-sm-actin染色鉴定,将传4～6代的细胞分为钙化组和对照组,对照组用正常DMEM培养基培养,钙化组加入10mmol/Lβ-甘油磷酸盐诱导细胞钙化,连续培养10d。茜素红S染色及碱性磷酸酶法鉴定。结果:分离的原代细胞经S-P染色鉴定为阳性,呈淡黄色。钙化组细胞诱导钙化后,细胞增殖缓慢,并形成囊泡结构,茜素红S染色形成红色的钙化结节,钙化组碱性磷酸酶活性较对照组在不同时间点(4,6,8,10d)都有所增加(P<0.01)。结论:β-甘油磷酸盐体外能够诱导HVSMC钙化,此方法诱导的钙化模型是一种良好的研究血管疾病方面的体外模型。
OBJECTIVE To prepare in vitro vascular calcification models by using β-glycerophosphate to treat human vascular smooth muscle cells (HVSMCs). Methods Primary human aortic smooth muscle cells were isolated from human umbilical artery by tissue adhesion method. Primary cells were identified by α-sm-actin staining. Cells from passage 4 to passage 6 were divided into the calcification group and the control group. The control group was cultured in normal DMEM medium, the calcification group was added with 10mmol / L β-glycerophosphate to induce cell calcification and cultured continuously for 10 days. Alizarin red S staining and alkaline phosphatase identification. Results: Isolated primary cells were identified as positive by S-P staining and light yellow. After calcified, the cell proliferation was slow and the vesicle structure was formed. Alizarin red S stained to form red calcified nodules. Calcification group showed higher alkaline phosphatase activity than control group at different time points (4, 6, 8, 10d) increased (P <0.01). CONCLUSION: β-glycerophosphate can induce HVSMC calcification in vitro. The calcified model induced by this method is a good in vitro model for the study of vascular diseases.