Inhibition of CTGF ameliorates peritoneal fibrosis through suppression of fibroblast and myofibroblast accumulation and angiogenesis

Norihiko Sakai,Yasunori Iwata,Taito Miyake,Akinori Hara,Shinji Kitajima,Yasutaka Kamikawa,Yasuyuki Shinozaki,Kengo Furuichi,Andrew M Tager,Tadashi Toyama,Akihiro Sagara,Kenneth E Lipson,Miki Nakamura,Shuichi Kaneko,Miho Shimizu,Takashi Wada

doi:10.1038/s41598-017-05624-2

Abstract

Peritoneal fibrosis (PF) is a serious complication in various clinical settings, but the mechanisms driving it remain to be fully determined. Connective tissue growth factor (CTGF) is known to regulate fibroblast activities. We therefore examined if CTGF inhibition has anti-fibrotic effects in PF. PF was induced by repetitive intraperitoneal injections of chlorhexidine gluconate (CG) in mice with type I pro-collagen promoter-driven green fluorescent protein (GFP) expression to identify fibroblasts. FG-3019, an anti-CTGF monoclonal antibody, was used to inhibit CTGF. CG-induced PF was significantly attenuated in FG-3019-treated mice. CG challenges induced marked accumulations of proliferating fibroblasts and of myofibroblasts, which were both reduced by FG-3019. Levels of peritoneal CTGF expression were increased by CG challenges, and suppressed in FG-3019-treated mice. FG-3019 treatment also reduced the number of CD31+ vessels and VEGF-A-positive cells in fibrotic peritoneum. In vitro studies using NIH 3T3 fibroblasts and peritoneal mesothelial cells (PMCs) showed that CTGF blockade suppressed TGF-β1-induced fibroblast proliferation and myofibroblast differentiation, PMC mesothelial-to-mesenchymal transition, and VEGF-A production. These findings suggest that the inhibition of CTGF by FG-3019 might be a novel treatment for PF through the regulation of fibroblast and myofibroblast accumulation and angiogenesis.

Highlights

Peritoneal fibrosis (PF) is a serious complication in various clinical settings, but the mechanisms driving it remain to be fully determined
Since peritoneal fibrosis is already established by day 7 of the 21-day chlorhexidine gluconate (CG) model[27], we examined the therapeutic potential of Connective tissue growth factor (CTGF) inhibition for peritoneal fibrosis by administering FG-3019 beginning 7 days after CG challenge onset in a therapeutic regimen
The percentage of myofibroblasts among total fibroblasts, calculated as the percentage of α-smooth muscle actin (αSMA)+green fluorescent protein (GFP)+ cells among total GFP+ cells, was significantly reduced by the blockade of CTGF (Fig. 3d). These results suggest that myofibroblast accumulation is dependent on CTGF, and that FG-3019 can inhibit the accumulation of these cells during the development of peritoneal fibrosis as well

Summary

Introduction

Peritoneal fibrosis (PF) is a serious complication in various clinical settings, but the mechanisms driving it remain to be fully determined. In vitro studies using NIH 3T3 fibroblasts and peritoneal mesothelial cells (PMCs) showed that CTGF blockade suppressed TGF-β1induced fibroblast proliferation and myofibroblast differentiation, PMC mesothelial-to-mesenchymal transition, and VEGF-A production. These findings suggest that the inhibition of CTGF by FG-3019 might be a novel treatment for PF through the regulation of fibroblast and myofibroblast accumulation and angiogenesis. We used an inhibitory monoclonal antibody, FG-3019, targeting CTGF22 to evaluate the contribution of CTGF to peritoneal fibrosis induced in mice by intraperitoneal injection of chlorhexidine gluconate (CG), a well-described model of peritoneal fibrosis[20, 23]. This study shows that FG-3019 could provide a beneficial therapeutic strategy to combat peritoneal fibrosis through the blockade of CTGF

Methods

Results

Conclusion