Abstract
Transforming growth factor beta (TGFβ) is often used in cartilage tissue engineering to increase matrix formation by cells with various phenotypes. However, adverse effects of TGFβ, such as extensive cross-linking in cultured fibroblasts, have also been reported. Our goal was to study effects of TGFβ on collagen cross-linking and evaluating the role of cellular phenotype and physical environment. We therefore used four different cell populations in two very different physical environments: primary and expanded chondrocytes and fibroblasts embedded in alginate gel and attached to tissue culture plastic. Matrix production, collagen cross-linking, and α-smooth muscle actin (αSMA) were analyzed during 4 weeks with or without 2.5 ng/mL TGFβ2. TGFβ2 did not affect collagen deposition by primary cells. In expanded cells, TGFβ2 increased collagen deposition. Chondrocytes and fibroblasts in monolayer produced more collagen cross-links with TGFβ2. In alginate, primary and expanded cells displayed an unexpected decrease in collagen cross-linking with TGFβ2. αSMA was not present in alginate cultures and barely upregulated by TGFβ2. Organized αSMA fibers were present in all monolayer cultures and became more pronounced with TGFβ2. This study demonstrates that the physical environment determined by the substrate used co-determines the response of cells to TGFβ. The presence of mechanical stress, determined with αSMA-staining, is probably responsible for the increase in collagen cross-linking upon addition of TGFβ.