SLIT3 promotes cardiac fibrosis and differentiation of cardiac fibroblasts by RhoA/ROCK1 signaling pathway

Abstract

Objective(s): Slit guidance ligand 3 (SLIT3) has been identified as a potential therapeutic regulator against fibroblast activity and fibrillary collagen production in an autocrine manner. However, this research aims to investigate the potential role of SLIT3 in cardiac fibrosis and fibroblast differentiation and its underlying mechanism.Materials and Methods: C57BL/6 mice (male, 8-10 weeks, n=47) were subcutaneously infused with Ang II (2.0 mg/kg/day) for 4 weeks. One to two-day-old Sprague-Dawley (SD) rats were anesthetized by intraperitoneal injection of 1% pentobarbital sodium (60 mg/kg) and ketamine (50 mg/kg) and the cardiac fibroblast was isolated aseptically. The mRNA and protein expression were analyzed using RT-qPCR and Western blotting.Results: The SLIT3 expression level was increased in Ang II-induced mice models and cardiac fibroblasts. SLIT3 significantly increased migrated cells and α-smooth muscle actin (α-SMA) expression in cardiac fibroblasts. Ang II-induced increases in mRNA expression of collagen I (COL1A1), and collagen III (COL3A1) was attenuated by SLIT3 inhibition. SLIT3 knockdown attenuated the Ang II-induced increase in mRNA expression of ACTA2 (α-SMA), Fibronectin, and CTGF. SLIT3 suppression potentially reduced DHE expression and decreased malondialdehyde (MDA) content, and the superoxide dismutase (SOD) and catalase (CAT) levels were significantly increased in cardiac fibroblasts. Additionally, SLIT3 inhibition markedly decreased RhoA and ROCK1 protein expression, whereas ROCK inhibitor Y-27632 (10 μM) markedly attenuated the migration of cardiac fibroblasts stimulated by Ang II and SLIT3.Conclusion: The results speculate that SLIT3 could significantly regulate cardiac fibrosis and fibroblast differentiation via the RhoA/ROCK1 signaling pathway.

Keywords

• α-smooth muscle actin
• cardiac fibroblast
• cardiac fibrosis
• collagen i
• collagen iii
• slit guidance ligand 3