关键词:

血管紧张素Ⅱ, 血管平滑肌细胞, 增殖, ANO1抑制剂

Abstract:

Objective　To investigate the role of recombinant anoctamin 1 (ANO1) inhibitors in angiotensin Ⅱ (AngⅡ) -induced proliferation of vascular smooth muscle cells (VSMC). Methods　The study period was from July 2019 to June 2022. The rat thoracic aorta smooth muscle cell line A7r5 was cultured in vitro; the AngⅡ stimulated A7r5 cells were used to establish the cell proliferation models; 10, 100, and 500 nmol/L ANO1 inhibitors (A01) were used for the intervention; a control group was set up (intervention with the same amount of normal saline); Hoechst 33342 fluorescence staining was used to observe the morphological changes of VSMC induced by AngⅡ after the intervention of ANO1 inhibitors; the protein western blotting was used to detect the expression levels of apoptosis-related proteins in the VSMC cells induced by AngⅡ after the intervention of ANO1 inhibitors. One-way analysis of variance was used for multi-group comparison, and LSD-t test was used for pairwise comparison between groups check. Results　The Hoechst 33342 fluorescence staining results showed that the ANO1 inhibitor concentration increased, the bright blue cells (chromatin condensation and apoptotic bodies) were getting more, indicating that the number of apoptotic cells increased. The apoptosis rates of the low-, medium-, and high-dose groups were (17.60±1.36)%, (36.30±3.50)%, and (61.25±6.33)%, respectively, which were higher than that of the control group [(4.32±0.51)%], with statistical differences between the groups (all P<0.05), indicating a dose-dependent relationship between AngⅡ-induced post-proliferative apoptosis of VSMC and ANO1 inhibitor concentration. The results of protein western blotting in the detection of AngⅡ-induced apoptosis related proteins in VSMC after ANO1 inhibitor intervention showed that there was a statistical difference in the main effect of EPK, AKT, CDK4, Cyclin D1 protein relative expression levels between the groups (P<0.05); compared with those in the control group, the relative protein expressions of EPK, AKT, CDK4, and Cyclin D1 in the low-, medium-, and high-dose groups decreased in a dose-dependent manner, with statistical differences between the groups (all P<0.05). Conclusions　ANO1 inhibitors can significantly inhibit AngⅡ-induced VSMC proliferation, and the mechanism may be related to the inhibition of the EPK/AKT signaling pathway and cell cycle progression.

Key words:

Angiotensin Ⅱ, Vascular smooth muscle cells, Proliferation, ANO1 inhibitor