High salt diet alters NF-κB and CREB DNA binding activities in the heart, kidney and hypothalamus of spontaneously hypertensive rats

Objectives: The differentiation of vascular adventitial fibroblasts (AFs) to myofibroblasts (MFs) is a key step in vascular remodeling. Our previous study demonstrated that TGF-β1 and angiotension II (Ang II) were able to induce this differentiation in vitro. The aim of the study was to identify the molecules which might be responsible for the cell phenotypic change and investigated the role of complement C3, one of the related molecules, in differentiation of MFs. Design and methods: Cultured rat AFs were treated with TGF-beta 1 (10 ng/ml) and Ang II (10 − 7 M). TGF-beta 1induced gene expression profiling was studied using Affimetrix oligonucleotide microarrays. Expression profile of complement C3 was verified by real-time RT-PCR. Then, the role of complement C3 in differentiation of AFs to MFs induced by Ang II was investigated using Western-blot, MTT array and Thymidine-Incorporation Assay. Results: Microarray analysis identified 2121 genes with a 2-fold change or above post-TGF-β1 stimulation. Among 1231 genes with known function, the gene expression profile of secreted phosphoprotein 1(APP1) and Rho-associated coiled-coil forming kinase 2 (ROCK2) was same as α-smooth muscleactin, a MF differentiation mark, and genes of potassium voltage gated channel, Shal-related family and member 2(KCND2) were up-regulated. Furthermore, endothelin 1(EDN1), complement C3, NADPH oxidase 4 (NOX4) and NAD(P)H dehydrogenase, quinone 1 (Nqo1) were also found to be involved in MF differentiation. These results were confirmed by realtime quantitative RT-PCR. For the very first time, complement C3 was found to express in vascular adventitial fibroblasts from SHR, WKY and SD rats and increased by angiotension II. Inhibition of complement C3 decreased the expression of α-smooth muscle-actin and reduced the proliferation of AFs from SD rats and WKY rats post treatment of Ang II. Exogenous C3 stimulated the growth of adventitial fibroblasts. Conclusions: Complement C3 contributes to adventitial fibroblast phenotypic differentiation and may be a target of vascular remodeling therapy.