Mechanical forces exerted by the extracellular matrix (ECM) on cells play an essential role in development, wound healing, and tissue engineering. The ECM in mammalian connective tissue is primarily composed of fibers of collagen type 1. Several collagen binding proteins are known to influence collagen fiber structure and content. How these changes in collagen fiber affect forces exerted by the ECM is not well understood. We recently established that the collagen binding proteins, Discoidin domain receptors (DDR1 and DDR2) alter the native structure and mechanical properties of collagen fibers. The objective of this study was to evaluate how alteration of the ECM environment by DDRs affects mechanical forces exerted on cells. Cell lines stably expressing the extracellular domain (ECD) of DDR1 or DDR2 and DDR2/-KD (DDR2 lacking kinase domain) were used in collagen gel contraction assays. While both DDR2-ECD and DDR2/-KD expressing cells inhibited collagen gel contraction, DDR1-ECD enhanced contraction as compared to controls. To further our understanding of DDR2-ECD affect on cell-matrix interaction we employed DDR2 antibody with the collagen gel contraction assays. DDR2 antibody affects contraction of both nontransfected and transfected cell lines. To confirm that modulation of collagen gel contraction by DDR-ECD expressing cells was due to changes in collagen morphology and not due to changes in the cell cytoskeleton, we performed actin staining assays with each cell line with collagen stimulation. Transfected cell lines demonstrated changes in actin organization compared to the nontransfected cell lines. To evaluate the viscoelastic properties of the ECM altered by DDRs, a micro-rheology technique employing optical tweezers was utilized. We demonstrated that the alteration of the ECM by DDRs influences the mechanical forces experienced in cell-matrix interactions.No embarg
