Increased Complement 3 With Suppression of miR-145 Induces the Synthetic Phenotype in Vascular Smooth Muscle Cells From Spontaneously Hypertensive Rats

Abstract

Background: We previously reported that vascular smooth muscle cells (VSMCs) from spontaneously hypertensive rats (SHRs) show the increased expression of complement 3 (C3) and the synthetic phenotype. We targeted the SHR C3 gene (C3 knockout [C3KO] SHRs) by the zinc finger gene editing method. In the current study, we investigated the mechanisms underlying the increased expression of C3 and the role of endogenous C3 in the synthetic phenotype of SHR VSMCs in comparison to cells from Wistar-Kyoto (WKY) rats and C3KO SHRs. Methods and Results: Nonmuscle myosin heavy chain staining of aortas from SHRs at 1 day after birth was stronger in comparison to WKY rats and C3KO SHRs. DNA synthesis in VSMCs from SHRs was significantly higher in comparison to WKY rats and C3KO SHRs. Immunohistochemical staining of renin and liver X receptor α in VSMCs from SHRs was stronger in comparison to WKY rats and C3KO SHRs. The expression of renin, Krüppel-like factor 5, and liver X receptor α proteins in VSMCs from SHRs was significantly higher in comparison to WKY rats and C3KO SHRs. The expression of synthetic phenotype markers osteopontin, matrix gla, and l-caldesmon, growth factors transforming growth factor-β1 and platelet-derived growth factor-A, transcription factors Krüppel-like factor 5 and liver X receptor α, and angiotensinogen mRNAs in VSMCs from SHRs was significantly higher in comparison to WKY rats and C3KO SHRs. The expression of miR-145 mRNA in VSMCs from SHRs was suppressed in comparison to cells from WKY rats. miR-145 inhibitor significantly increased the expression of C3 in VSMCs from WKY rats, but not in cells from SHRs. Conclusions: These findings indicate that the increased C3 with the suppression of miR-145 induces the synthetic phenotype through Krüppel-like factor 5 and the activation of the renin-angiotensin system through liver X receptor α in VSMCs from SHRs.