Abstract P224: Connective Tissue Growth Factor Plays a Key Role in Myocardial Fibrosis After Angiotensin II Exposure

Abstract

Background: AngiotensinII (AngII) exposure to rodents is a common model of fibrosis, characterized by hypertension, hypertrophy and eventual deposition of excess extracellular matrix (ECM) proteins resulting in organ dysfunction. We have previously shown that cellular infiltration of bone marrow derived progenitor cells (fibrocytes) occurs prior to ECM deposition and is associated with production of the connective tissue growth factor (CTGF) and transforming growth factor beta (TGF-β). The objective of this project was to characterize the role of CTGF in promoting fibrocyte recruitment and subsequent fibrosis after AngII exposure. Methods: Mice were treated with AngII or saline using an osmotic mini-pump at 2.8mg/kg/day. After 6hr to 7d hearts were excised and embedded in paraffin or prepared for mRNA isolation. Immunohistochemistry was used to determine extent and localization of CTGF protein. Quantitative RT-PCR was used to determine relative CTGF and TGF-β mRNA levels. Primary cardiomyocyte and fibrocyte cultures were isolated from neonatal or 3d AngII exposed animals respectively. Primary cultures were stimulated to determine the source (qRT-PCR) and function (proliferation, migration and differentiation) of CTGF. Results: In animals exposed to AngII, CTGF mRNA peaked the earliest at 6hr (21-fold; p<0.01) when compared to TGFbeta, which peaked at 3d (5-fold; p<0.05). Concurrent CTGF protein expression was evident by 3d of AngII exposure and appeared localized to the cardiomyocytes. Findings were confirmed using isolated cardiomyocytes, which significantly increased expression of CTGF in response to AngII (2-fold; p<0.05). While CTGF did not promote fibrocyte migration in transwell chamber assay it promoted significant fibrocyte proliferation in vitro (2-fold; p<0.05). Conclusion: We provide strong evidence that AngII exposure first results in the production of CTGF by cardiomyocytes. Furthermore, we have shown that CTGF does not promote migration as a chemokine but instead contributes to proliferation of fibrocytes once recruited from the bone marrow into the myocardium.