A stimuli-responsive ex vivo model of osteoarthritis demonstrates TLR4-mediated cartilage degradation and a Rapamycin-induced fast matrix recovery

Abstract

BackgroundIn osteoarthritis (OA) TLR4 signaling leads to downstream activation of the phosphoinositide 3-kinases/ protein kinase B/ mammalian target of Rapamycin (PIK3/AKT/mTOR) pathway, a known modulator of autophagic mechanisms in chondrocytes. This paper focuses on creating a realistic ex vivo OA model that mimics elements of the pathophysiology of OA, allowing for further hypotheses-based investigations, and for use as a bench test for new therapeutic targets. ObjectiveTo study the downstream inflammatory and matrix changes in cartilage due to TLR4 signaling and the recovery achieved by a commonly used immunosuppressive drug, Rapamycin. MethodsIn an ex vivo 3D model based on healthy porcine cartilage explants, we mimicked the OA environment by LPS stimulation activating TLR4 signaling. Furthermore, we inhibited mTOR signaling via Rapamycin, which is accepted to attenuate the cartilage response to LPS-TLR4 activation. Histology and immunohistochemistry were used to evaluate the structural and biomolecular modifications driven by LPS and Rapamycin. ResultsThe explant model captured key features of OA, such as extracellular matrix degeneration and altered autophagy. The OA-like changes in the model were driven by TLR4 activation and mTOR signaling, well-known OA-related molecular pathways, and reversed by Rapamycin. ConclusionWe demonstrate that our explant model is responsive to LPS stimulation, leading to activation of OA-related biomolecular pathways, closely mimicking the native physiological processes. This evidence supports the potential of our model to act as a platform for OA studies, in particular related to the gut-joint axis in age-related OA, and for the screening of new disease-modifying molecules.