Jichu yixue yu linchuang (Apr 2023)

Circadian clock protein CRY1 inhibits Ang Ⅱ-induced phenotypic transformation of mouse aorta smooth muscular cells

  • YUE Xiuqing, SONG Qitai, LIU Chaoli, LI Sangrou, YANG Fan, CHEN Muhu, ZHONG Wu

DOI
https://doi.org/10.16352/j.issn.1001-6325.2023.04.0588
Journal volume & issue
Vol. 43, no. 4
pp. 588 – 595

Abstract

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Objective To investigate the effect of circadian clock protein cryptochrome 1 (CRY1) on angiotensin Ⅱ(Ang Ⅱ)-induced phenotypic transformation of mouse aorta smooth muscular cells. Methods VSMCs were divided into control group and experimental group. VSMCs were treated with different concentrations of Ang Ⅱ for various time, α-smooth muscle actin (α-SMA), osteopontin (OPN) and CRY1 expressions in VSMCs were detected via RT-qPCR and Western blot. VSMCs were transfected with knockdown vector siCRY1 and siYAP1 or overexpression vector, and the corresponding gene expression was detected with RT-PCR and Western blot. VSMCs were divided into 4 groups: Control group, Ang Ⅱ group, Ang Ⅱ+siCRY1 group and Ang Ⅱ+pcDNA-CRY1 group. siCRY1, siYAP1, pcDNA-CRY1 were transfected into VSMCs induced by Ang Ⅱ for observation on α-SMA, CRY1, OPN and YAP1 at mRNA and protein levels detected by RT-PCR and Western blot. CCK-8 method were used to measure proliferation capacity. Results VSMCs were treated with different concentrations of Ang Ⅱ for various time, the expression of CRY1 was decreased in a model of Ang Ⅱ-induced VSMC phenotypic transformation, which contractile markers down-regulated and synthetic markers up-regulated at mRNA and protein levels. After VSMCs were transfected with siCRY1, siYAP1 and pcDNA-CRY1, the expression of CRY1 decreased in siCRY1 group and increased in pcDNA-CRY1 group, while the expression of YAP1 decreased in siYAP1 group and increased in siCRY1 group.Knockdown of CRY1 could stimulate the transformation of VSMCs from contractile phenotype into synthetic phenotypes by further activating expression of synthetic markers and CRY1(P＜0.05), inhibiting expression of contractile markers and YAP1(P＜0.05), the proliferation of VSMCs increased significantly(P＜0.05). While overexpressed CRY1 could block the phenotypic transformation, the expression of synthetic markers and CRY1 was reduced and contractile markers and YAP1 was increased both at mRNA and protein levels(P＜0.05), the proliferation of VSMCs inhibited(P＜0.05). Conclusions CRY1 may inhibit switch of mouse VSMCs phenotypes induced by Ang Ⅱ and inhibit VSMCs proliferation, which is potentially related to the Hippo-YAP signaling pathway.

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