平滑肌ZFP36通过调控Runx2表达抑制血管钙化

ZFP36 in smooth muscle inhibits vascular calcification by regulating Runx2 expression

摘要:

目的]探究RNA结合蛋白锌指蛋白36(ZFP36)在血管钙化中的作用及机制。 [方法]使用ZFP36过表达病毒和siRNA处理人主动脉平滑肌细胞并给予高磷刺激,观察过表达或敲低ZFP36后对平滑肌细胞钙化的影响；RNA免疫共沉淀和稳定性实验检测Runx2是否为ZFP36的靶基因；制备平滑肌特异性ZFP36敲除小鼠,利用维生素D诱导血管钙化,使用茜素红和Von Kossa染色观察主动脉血管钙化情况,Western blot检测Runx2蛋白的表达。 [结果]ZFP36在钙化刺激条件下表达升高。过表达ZFP36减轻高磷诱导的平滑肌细胞钙化状况,而敲低ZFP36则加重钙化。ZFP36可以结合Runx2 mRNA并促进其降解。在细胞水平,ZFP36通过Runx2抑制平滑肌细胞钙化。在动物水平,平滑肌敲除ZFP36加重了维生素D诱导的血管钙化。 [结论]平滑肌中ZFP36通过靶向Runx2 mRNA抑制其表达从而抑制血管钙化。

Abstract:

Aim To investigate the role and mechanism of RNA binding protein zinc finger protein 36 (ZFP36) in vascular calcification. Methods Human aortic smooth muscle cells were infected with ZFP36-overexpressing virus or transfected with ZFP36 siRNA, followed by high phosphate stimulation to observe the effect of overexpression or knockdown of ZFP36 on smooth muscle cell calcification; RNA immunoprecipitation and stability experiments were used to detect whether Runx2 was the target gene of ZFP36; Smooth muscle specific ZFP36 knockout mice were generated and vitamin D was used to induce vascular calcification. Alizarin Red and Von Kossa staining were used to observe calcification of aortic vessels, and Western blot was used to detect Runx2 protein expression.Results ZFP36 expression was elevated under calcification-stimulating conditions. Overexpression of ZFP36 alleviated high phosphate-induced smooth muscle cell calcification, while knockdown of ZFP36 exacerbated calcification. ZFP36 could bind to Runx2 mRNA and promote its degradation. At the cellular level, ZFP36 could inhibit smooth muscle cell calcification via Runx2. At the animal level, knockout of ZFP36 in smooth muscle exacerbated vascular calcification induced by vitamin D. Conclusion ZFP36 inhibits vascular calcification by targeting Runx2 mRNA and suppressing its expression in smooth muscle cells.