Genetic Deletion of the Collagen Receptor, DDR2, in Mice Leads to Slower Deposition of Fibrillar Collagen by Cardiac Fibroblasts

Abstract

Introduction and Hypothesis: The type 2 discoidin domain receptor (DDR2) is a cell-surface receptor for fibrillar collagens. It is expressed on mesenchymal cells throughout the body, but its expression within the heart is restricted to cardiac fibroblasts. Inhibition of DDR2 function during development leads to dwarfism in mice and humans, an effect believed to be caused by inadequate bone growth. We had generated DDR2 null mice and had observed the expected 20-30% reduction in body weight compared to wildtype littermates. However, we also noticed a significant reduction in whole heart weight and left ventricular mass in these null mice. Although normalization of heart weight to body weight often eliminated statistical significance, we hypothesized that a cardiac phenotype did exist (i.e., a smaller heart) and that it was manifested through the activities of the cardiac fibroblasts during growth. Methods and Results: DDR2 null hearts were compared to wildtype hearts using measures of collagen content such as Masson's Trichrome staining and hydroxyproline assays. Although the absolute content of collagen was lower in the smaller null hearts, relative collagen content (after normalizing to volume or mass) was not statistically different from that of wildtype animals. Hearts were also examined by transmission electron microscopy to observe any abnormalities in collagen fibril structure. Cardiac collagen fibrils from DDR2 null mice appeared normal with an average diameter of 32 ± 3 nm (versus 34 ± 2 nm for wildtypes, n = 3 for both) and a comparable banding periodicity of 50– 60 nm. To ascertain the effect of DDR2 expression on the rate of synthesis of insoluble (i.e., fibrillar) collagen, cardiac fibroblasts were isolated from the ventricles of adult mice and cultured. As determined by hydroxyproline assays, DDR2 null fibroblasts synthesized insoluble collagen at a rate that was 32% lower than that of wildtype cardiac fibroblasts. Studies are ongoing, but the reason for this difference is currently not known. Conclusions: We conclude that the smaller hearts in the DDR2 knockout mice are likely caused by a slower deposition of structurally-normal collagen during growth, which limits tissue expansion. However, adult cardiac fibroblasts also possess this phenotype. Inhibition of DDR2 in adults could slow pathophysiologic cardiac remodeling involving collagen deposition, such as the accumulation of interstitial fibrosis. Therefore, DDR2 may represent a novel therapeutic target. Introduction and Hypothesis: The type 2 discoidin domain receptor (DDR2) is a cell-surface receptor for fibrillar collagens. It is expressed on mesenchymal cells throughout the body, but its expression within the heart is restricted to cardiac fibroblasts. Inhibition of DDR2 function during development leads to dwarfism in mice and humans, an effect believed to be caused by inadequate bone growth. We had generated DDR2 null mice and had observed the expected 20-30% reduction in body weight compared to wildtype littermates. However, we also noticed a significant reduction in whole heart weight and left ventricular mass in these null mice. Although normalization of heart weight to body weight often eliminated statistical significance, we hypothesized that a cardiac phenotype did exist (i.e., a smaller heart) and that it was manifested through the activities of the cardiac fibroblasts during growth. Methods and Results: DDR2 null hearts were compared to wildtype hearts using measures of collagen content such as Masson's Trichrome staining and hydroxyproline assays. Although the absolute content of collagen was lower in the smaller null hearts, relative collagen content (after normalizing to volume or mass) was not statistically different from that of wildtype animals. Hearts were also examined by transmission electron microscopy to observe any abnormalities in collagen fibril structure. Cardiac collagen fibrils from DDR2 null mice appeared normal with an average diameter of 32 ± 3 nm (versus 34 ± 2 nm for wildtypes, n = 3 for both) and a comparable banding periodicity of 50– 60 nm. To ascertain the effect of DDR2 expression on the rate of synthesis of insoluble (i.e., fibrillar) collagen, cardiac fibroblasts were isolated from the ventricles of adult mice and cultured. As determined by hydroxyproline assays, DDR2 null fibroblasts synthesized insoluble collagen at a rate that was 32% lower than that of wildtype cardiac fibroblasts. Studies are ongoing, but the reason for this difference is currently not known. Conclusions: We conclude that the smaller hearts in the DDR2 knockout mice are likely caused by a slower deposition of structurally-normal collagen during growth, which limits tissue expansion. However, adult cardiac fibroblasts also possess this phenotype. Inhibition of DDR2 in adults could slow pathophysiologic cardiac remodeling involving collagen deposition, such as the accumulation of interstitial fibrosis. Therefore, DDR2 may represent a novel therapeutic target.