Annexin A6: a novel therapeutic target for the treatment of osteoarthritis?

Abstract

Purpose: NF-κb and Wnt/β-catenin signaling are two major pathways involved in osteoarthritis (OA). Members of the annexin protein family have been implicated to play essential roles in various disease pathologies based on their function to modulate signaling pathways, including NF-κB. Based on these findings and our findings showing that annexin A6 (AnxA6) is highly expressed in OA cartilage but not in normal healthy articular cartilage, we hypothesized that AnxA6 is a major contributor to cartilage destruction during OA. Methods: The expression of catabolic genes and the activation of NF-κB and Wnt/β-catenin signaling in articular chondrocytes isolated from AnxA6 knockout (-/-) and wild type (WT) mice and in human articular chondrocytes overexpressing AnxA6 and cultured in the absence or presence of interleukin (IL)-1 and Wnt3a were determined by real time PCR and luciferase reporter assays. IL-1 injections into the knee joints of AnxA6-/- and WT mice were used to determine the role of AnxA6 in IL-1-mediated cartilage destruction. The mechanism of how AnxA6 stimulates NF-κB and Wnt/β-catenin signaling was determined by co-immunoprecipitation and immunoblot analysis of plasma membrane, nuclear and cytoplasmic fractions of IL-1 or Wnt3a-treated AnxA6-/- and WT chondrocytes. Cartilage degradation in an OA mouse model using the transection of the medial collateral ligament and partial medial meniscetomy (PMX) was analyzed histologically by safranin O staining. Results: Lack of AnxA6 in articular chondrocytes isolated from AnxA6-/- mice resulted in decreased NF-κB activity after IL- 1 treatment and increased Wnt/β-catenin signaling after Wnt3a treatment. Overexpression of AnxA6 in human articular chondrocytes was sufficient for a marked increase in NF-κB activity and decreased Wnt/β-catenin signaling activity. AnxA6 interaction with p65 resulted in increased nuclear translocation and retention of the active p50/p65 NF-κB complex, whereas plasma membrane-associated AnxA6 interfered with the membrane-association of the Wnt signalosome complex required for the activation of Wnt/β-catenin signaling. IL-1 treatment resulted in a markedly reduced increase of the mRNA levels of catabolic markers, including ADAMTS-5, IL-6, iNOS, and MMP-13, in AnxA6-/- chondrocytes compared with IL-1-treated WT chondrocytes, whereas overexpression of AnxA6 resulted in increased mRNA levels of these catabolic markers in the absence or presence of IL-1. Intraarticular injections of IL-1 into the knee joints of AnxA6-/- mice resulted in decreased proteoglycan loss, cartilage degradation, and MMP-13 expression compared with Il-1 injections into WT knee joints. Contrary, cartilage degradation in the PMX-induced post-traumatic OA model, in which the Wnt/β-catenin signaling plays a major role in mediating cartilage degradation, was markedly accelerated in AnxA6-/- mice compared to WT mice. Finally, we provide evidence that Wnt/β-catenin signaling in human articular chondrocytes inhibited catabolic events, especially NF-κB-mediated catabolic events, whereas in mouse articular chondrocytes Wnt/β-catenin signaling stimulated catabolic events. Conclusions: Our findings show that (a) Wnt/β-catenin signaling acts anti-catabolic in human articular chondrocytes and especially inhibits NF-κB-mediated catabolic events, whereas Wnt/β-catenin signaling in mouse articular chondrocytes stimulates catabolic events, and that (b) AnxA6 via stimulating NF-κB activity and inhibiting Wnt/β-catenin signaling plays a major role in human cartilage degradation during OA pathology. Our findings suggest that targeting AnxA6 function, especially its modulatory function of the NF-κB and Wnt/β-catenin signaling pathways may be a more effective way to control these signaling pathways and their interactions during OA pathology than targeting these individual pathways directly.