吡格列酮通过Wnt/β-catenin信号通路减轻大鼠血管平滑肌细胞钙化  

Effect of pioglitazon on calcification of vascular smooth muscle cells through the down-regulation of Wnt / β-catenin signaling pathway

作　　者：高敏[1,2] 陈天雷[3] 吴琳[1] 赵秀芬[1] 邢昌赢[1] 毛慧娟[1] 

机构地区：[1]南京医科大学第一附属医院肾内科,南京210029 [2]常州市武进人民医院肾内科 [3]常州市第一人民医院肾内科

出　　处：《肾脏病与透析肾移植杂志》2016年第4期340-346,共7页Chinese Journal of Nephrology,Dialysis & Transplantation

基　　金：江苏省教育厅六大人才高峰(WSN-056);江苏省临床医学科技专项(BL2014080)

摘　　要：目的:探讨过氧化物酶体增殖物活化受体γ(PPARγ)激动剂吡格列酮(PIO)对高磷诱导大鼠血管平滑肌细胞(VSMC)钙化的作用及其相关机制。方法:利用10 mmol/Lβ甘油磷酸(β-GP)诱导大鼠VSMC钙化,建立钙化模型(即钙化组);同时以完全培养基培养大鼠VSMC为正常对照组,并分别以不同浓度(5μmol/L、10μmol/L、15μmol/L、20μmol/L)PIO干预,观察培养12d,用茜素红染色法检测细胞钙沉积并检测细胞外基质钙离子浓度来观察VSMC钙化程度。Western Blot检测大鼠VSMC的α平滑肌动蛋白(α-SMA)、Runx2、BMP2、Wnt/β-catenin通路相关蛋白(β-catenin、GSK-3β、p-GSK-3β)及核蛋白cyclin-D1的表达情况。选择合适的PIO浓度(20μmol/L)并以PPARγ拮抗剂GW9662(20μmol/L)干预,观察以上指标的变化。结果:(1)钙化组钙离子浓度较正常对照组明显增高(P<0.05),而不同浓度PIO均可减轻VSMC细胞外基质的钙离子浓度(P<0.05),同时钙化组茜素红染色较正常对照组明显,而20μmol/L PIO干预组茜素红染色较钙化组减轻最为明显;(2)钙化组大鼠VSMC表达Runx2、β-catenin、p-GSK-3β、BMP2、cyclin-D1较正常对照组升高;20μmol/L PIO可显著下调钙化大鼠VSMC表达Runx2、β-catenin、p-GSK-3β、BMP2和cyclin-D1,并上调α-SMA的表达;(3)PPARγ拮抗剂GW9662可部分阻断PIO对钙化大鼠VSMC的干预作用。结论:PPARγ激动剂PIO可减轻β-GP诱导的大鼠VSMC的钙化,其作用机制与下调Wnt/β-catenin信号通路活性有关。Objective： To investigate the effect and possible mechanism of pioglitazon on the calcification of rat vascular smooth muscle cells（ VSMCs） in vitro. Methodology： β-glycerophosphate（ 10 mmol / L） was used to induce the calcification of VSMCs,with different concentrations（ 5 μmol / L、10 μmol / L、15 μmol / L、20 μmol / L） of PIO to intervene for 12 d. The calcium deposits were tested by Alizarin red staining. The extracellular calcium content was detected by Calcium Assay Kit. Western Blot was used to measure the expressions of α-smooth muscle actin（ α-SMA）,runt-related transcription factor 2（ Runx2）,bone morphogenetic protein 2（ BMP2）,β-catenin,GSK-3β,p-GSK-3β and cyclin-D1.On the basis of 10 mmol / L β-glycerophosphate and 20 μmol / L PIO,20 μmol / L PPARγ antagonist GW9662 was added to the cell culture media. The changes of the above indexes were observed. Results：（ 1） The calcium content in calcification group was increased significantly compared with that in control＇s（ P〈0. 05）,and all different concentrations of PIO could reduced extracellular calcium content（ P〈0. 05）. Alizarin red staining was strong positive in calcifiedVSMCs,and PIO（ 20 μmol / L） intervention group was almost negative.（ 2） The expressions of Runx2,β-catenin,pGSK-3β,BMP2 and cyclin-D1 were increased significantly in calcification group,and 20 μmol / L PIO group obviously down-regulated the expressions of all the above proteins,while up-regulated the expression of α-SMA.（ 3） PPARγantagonist GW9662 could partly block the effect of PIO on calcified VSMCs. Conclusion： PPARγ agonist PIO can alleviate rat aortic VSMCs calcification induced by β-glycerophosphate via inhibiting the activity of Wnt / β-catenin signaling pathway.

关 键 词：过氧化物酶体增殖物活化受体Γ 血管钙化 血管平滑肌细胞 WNT/Β-CATENIN 信号通路 

分 类 号：R692[医药卫生—泌尿科学]