CD44 and hyaluronan promote the bmp7 signaling response in chondrocytes

Abstract

Cell-matrix interactions are important in maintaining cartilage homeostasis. Hyaluronan binding to its principal receptor CD44 is essential for such interactions. Hyaluronan-CD44 interactions are required for the retention of proteoglycan in the pericellular matrix, the most active region of matrix assembly and turnover. In previous work, SMAD1, the downstream transcriptional modulator of the canonical bone morphogenetic protein 7 (BMP7) signaling pathway, was found to interact with the cytoplasmic domain of CD44. Therefore, in this study the response to BMP7 stimulation was studied with murine chondrocytes isolated from CD44-/- and BALB/c mice to determine the role of CD44 as well as CD44-hyaluronan interactions in the BMP7 signaling pathway. The two part hypothesis is that chondrocytes from CD44-/- mice will be less responsive to BMP7 compared to wild type chondrocytes and that hyaluronan will have no effect on BMP7 signaling in CD44-/- mouse chondrocytes, but will promote BMP7 signaling in wild type chondrocytes. The results demonstrated that both CD44-/- murine chondrocytes and wild type chondrocytes transfected with CD44 siRNA were less responsive to BMP7 compared to wild type murine chondrocytes. CD44-/- murine chondrocytes transfected with pCD44 showed a BMP7 sensitivity similar to wild type chondrocytes. There were no dose response differences observed between wild type and CD44-/- murine chondrocytes found upon EGF stimulation, thus the hyaluronan-CD44 interaction appears selectively involved in the BMP7 signaling pathway. In both CD44-/- and wild type murine chondrocytes, hyaluronidase treatment decreased cellular responses to BMP7. However, with hyaluronan regrowth, SMAD1 phosphorylation upon BMP7 stimulation was restored. Treatment of chondrocytes with 4-methylumbelliferone in order to reduce synthesis of endogenous hyaluronan, also indicated that hyaluronan promoted the BMP7 signaling pathway. Taken together, these investigations into the mechanisms underlying BMP7 signaling in chondrocytes reveal that while hyaluronan is vital for optimal BMP7 signaling in chondrocytes, the functional significance of CD44 is more subtle and is dose dependent.