Kazuhiro Yamamoto, Salvatore Santamaria, Kenneth A. Botkjaer, Jayesh Dudhia, Linda Troeberg, Yoshifumi Itoh, Gillian Murphy, Hideaki Nagase

The aggrecanase ADAMTS-5 and the collagenase MMP-13 are constitutively secreted by chondrocytes in normal cartilage, but rapidly endocytosed via the cell surface endocytic receptor low-density lipoprotein receptor-related protein 1 (LRP1) and subsequently degraded. This endocytic system is impaired in osteoarthritic (OA) cartilage due to increased ectodomain 'shedding' of LRP1. The aim of this study is to identify the LRP1 sheddase(s) in human cartilage, and to test whether inhibition of LRP1 shedding prevents the cartilage degradation in OA. Cell-associated LRP1 and soluble LRP1 (sLRP1) released from human cartilage explants and chondrocytes were measured by Western blot analysis. LRP1 sheddases were identified by protease inhibitor profiling and gene silencing with siRNAs. Specific monoclonal antibodies were used to selectively inhibit the sheddases. Degradation of aggrecan and collagen in human OA cartilage was measured by Western blot analysis using an aggrecan neo-epitope antibody and hydroxyproline assay, respectively. The shedding of LRP1 was increased in OA cartilage compared to normal tissue. Shed sLRP1 bound to ADAMTS-5 and MMP-13 and prevented their endocytosis without interfering with their proteolytic activities. Two membrane-bound metalloproteinases, ADAM17 and MMP-14, have been identified as the LRP1 sheddases in cartilage. Inhibition of their activities restored the endocytic capacity of chondrocytes and reduced degradation of aggrecan and collagen in OA cartilage. The shedding of LRP1 is a key link to OA progression. Local inhibition of LRP1 sheddase activities of ADAM17 and MMP-14 is a unique way to reverse matrix degradation in OA cartilage and could be effective as a therapeutic approach. This article is protected by copyright. All rights reserved.