炎症因子刺激IGF-I基因沉默后的hCASMC细胞PAPP-A、IGF-I的表达及细胞学功能改变的意义

Expression of PAPP-A, IGF-I and Their Effects on the Cytological Functions of siRNA Lentiviral Vector of hCASMCs IGF-I after Inflammatory Factor

中文关键词:  炎症因子 人冠状动脉平滑肌细胞 RNA干扰 IGF-I轴 细胞学功能

英文关键词:Inflammation factors Human coronary artery vascular smooth muscle cells RNA interference IGF axis Cytological functions

中文摘要:

      目的 观察炎症因子刺激胰岛素样生长因子-I（IGF-I）基因沉默后的人冠状动脉平滑肌细胞（IGF-I-shRNA-hCASMC）中妊娠相关血浆蛋白-A（PAPP-A）、IGF-I的表达及其细胞学功能改变，为进一步明确IGF轴激素在动脉粥样硬化不稳定斑块破裂中的作用提供实验数据。方法 利用肿瘤坏死因子-α（TNF-α）)、白介素-1β (IL-1β）和/或胰岛素样生长因子结合蛋白-4 (IGFBP4)刺激IGF-I-shRNA-hCASMC，用Western blot及ELISA检测细胞中PAPP-A、IGF-I的表达。用MTT法及流式细胞仪检测TNF-α、IL-1β、IGFBP4对IGF-I-shRNA-hCASMC细胞功能学改变。结果 用TNF-α+IL-1β或TNF-α+IL-1β+IGFBP4刺激空白对照（CON）组细胞、阴性对照（NC）组细胞、RNA干扰（RNAi）组细胞时hCASMCs中PAPP-A均有较强表达， RNAi组的hCASMCs中PAPP-A的表达量少于另外两组。而未行刺激的三组细胞则无PAPP-A的表达。在TNF-α+IL-1β+IGFBP4刺激RNAi组细胞中IGF-I表达量高于仅用TNF-α+IL-1β刺激和未行刺激组，并且差异具有统计学意义。在RNAi组TNF-α+IL-1β+IGFBP4刺激细胞所测的A570值低于TNF-α+IL-1β刺激细胞（P<0.01）。RNAi组细胞经过TNF-α+IL-1β或TNF-α+IL-1β+IGFBP4刺激后表现为显著性凋亡，与CON组及NC组细胞相比具有统计学意义，RNAi组细胞经TNF-α+IL-1β+IGFBP4刺激后与经TNF-α+IL-1β刺激后相比细胞更易凋亡，差异有统计学意义。结论 炎症因子刺激IGF-I-RNAi的hCASMCs可导致细胞增殖力下降，细胞凋亡增加，在局部细胞环境中活性IGF-I增加情况，加剧细胞增殖力下降、细胞凋亡增加。

英文摘要:

      Objective To evaluate expression of PAPP-A, IGF-I and the effect of TNF-α, IL-1β on the cytological functions of hCASMCs with IGF-I gene silencing after inflammatory factor, in order to further study on the action of IGF axis hormone in the rupture of astable atheroxclerosis plaque. Methods A RNA interference (RNAi） aimed at the gene of IGF-I was carried out to have an eukaryon transfection to hCASMCs. When the IGF-I-shRNA-hCASMC were treated by TNF-α, IL-1β, IGFBP4, the expression of PAPP-A and IGF-I were detected with Western blot and ELISA. And then, the effect of TNF-α,IL-1β,IGFBP4 on the proliferation of IGF-I-shRNA-hCASMC were assessed by MTT assay and changing in cell cycle and apoptosis were evaluated by using flow cytometry. Results Significant positive expression of PAPP-A in hCASMCs which were treated by TNF-α+IL-1β or TNF-α+IL-1β+IGFBP4 in Blank control (CON), Negative control (NC) and RNAi group were observed, but lower expression in the RNAi group than that in CON and NC groups. However, there was no positive expression of PAPP-A in hCASMCs of CON, NC, RNAi group treating without TNF-α+IL-1β or TNF-α+IL-1β+IGFBP4. The expression of IGF-I in hCASMCs of CON, NC, RNAi group treated with TNF-α+IL-1β+IGFBP4 were greater than that only with or without TNF-α+IL-1β treatment. In RNAi group, the A₅₇₀ decreased when the hCASMCs treated with TNF-α+IL-1β+IGFBP4 and significant lower than that in hCASMCs treated with TNF-α+IL-1β. When the hCASMCs were treated with TNF-α+IL-1β or TNF-α+IL-1β+IGFBP4, the rate of apoptosis significantly increased in RNAi group, which was significantly higher than that in CON group and NC group. In addition, in RNAi group, the rate of apoptosis in hCASMCs treated with TNF-α+IL-1β+IGFBP4 was significant higher than that in hCASMCs treated only with TNF-α+IL-1β. Conclusion When the IGF-I-shRNA-hCASMCs were stimulated by some inflammation factors, its proliferation decreased but the apoptosis enhanced. So the activated IGF-I level in local microebvironnment increased, which may cause the descend of cell proliferation and the increasing of apoptosis.