Bone morphogenetic protein and transforming growth factor β inhibitory Smads 6 and 7 are expressed in human adult normal and osteoarthritic cartilage in vivo and are differentially regulated in vitro by interleukin-1β

Objective. Bone morphogenetic protein (BMP) and transforming growth factor beta (TGFbeta) are potent anabolic factors in adult articular chondrocytes. In this study, we investigated whether intracellular inhibitors of BMP and TGFbeta signaling, inhibitory Smad6 (I-Smad6) and I-Smad7, are expressed in articular chondrocytes in normal and osteoarthritic (OA) cartilage, and whether their expression shows a correlation with the anabolic activity of OA chondrocytes in vivo and after interleukin-beta (IL-1beta) stimulation in vitro. Methods. RNA isolated directly from normal and OA human knee cartilage as well as from cultured articular chondrocytes was analyzed by (quantitative) polymerase chain reaction technology. Immunolocalization of the I-Smads was performed on tissue sections and compared with the anabolic cellular activity as documented by in situ hybridization experiments for aggrecan and type II collagen. Results. Both Smad6 and Smad7 were expressed in all samples of normal and OA cartilage. Immunostaining (including confocal microscopy) confirmed the presence of Smad6 and Smad7 in the majority of normal and degenerated articular chondrocytes; localization was mostly cytoplasmic. No correlation between expression of the main anabolic genes and expression of the I-Smads was found. In cultured articular chondrocytes, stimulation with IL-1beta showed up-regulation of Smad7, whereas Smad6 was down-regulated. Conclusion. Both Smad6 and Smad7 are expressed in adult human articular chondrocytes. The primarily cytoplasmic localization suggests permanent activation of the I-Smads in articular cartilage in vivo. No evidence was found that up-regulation or downregulation of I-Smads in OA cartilage correlates directly with the anabolic (or catabolic) activity of articular chondrocytes. The regulation in chondrocytes of Smad6 and Smad7 expression by IL-1beta suggests a potentially important role of IL-1beta signaling in chondrocytes, via indirect influencing of the BMP/TGFbeta signaling cascade.