Midkine promotes articular chondrocyte proliferation through the MK-LRP1-nucleolin signaling pathway

Osteoarthritis (OA) is the most common disease of joint tissues; unfortunately, there are currently no curative therapies available for OA. Chondrocytes, the only cell type residing in cartilage, secrete many types of collagen (the mainly one is type II collagen) and aggrecan, which are the main components of the cartilage matrix. Chondrocyte apoptosis can lead to OA degenerative progression. We previously indicated that recombinant human midkine (rhMK), as a chondrocyte growth factor has a significant reparative effect on cartilage injury animal models. However, the molecular mechanism of this restorative function remains under investigation. Herein, we focused on the molecular mechanism underlying the role of MK in promoting the proliferation of chondrocytes cultured in vitro. Chondrocytes from rats and OA patients were successfully isolated by the digestion of articular cartilage using type II collagenase, and their proliferation was evaluated by a CCK8 assay and flow cytometry. rhMK stimulated the proliferation of chondrocytes from both OA patients and rats. Furthermore, qRT-PCR, shRNA-mediated knockdown, Western blot and immunoprecipitation (IP) assays were performed to identify the receptor and key elements responsible for the role of MK in promoting chondrocyte proliferation. Low-density lipoprotein receptor-related protein 1 (LRP1) was identified as the dominant MK receptor in chondrocytes that, as a translocator, mediates the endocytosis of MK. After being transferred into chondrocytes, MK was shown to form a complex with nucleolin that interacts with the active form of K-Ras. Upon the activation of ERK1/2, cyclin D1 expression was upregulated, promoting the chondrocyte cell cycle. Our data reveal for the first time the role of the MK-LRP1-nucleolin signaling pathway in facilitating MK-induced chondrocyte proliferation, thus providing a strong theoretical foundation for the further use of MK in OA clinical therapy.