YAP activation inhibits inflammatory signalling and cartilage breakdown associated with reduced primary cilia expression

Abstract

Objective: To clarify the role of YAP in modulating cartilage inflammation and degradation and the involvement of primary cilia and associated intraflagellar transport (IFT).

Methods: Isolated primary chondrocytes were cultured on substrates of different stiffness (6-1000 kPa) or treated with YAP agonist lysophosphatidic acid (LPA) or YAP antagonist verteporfin (VP), or genetically modified by YAP siRNA, all ± IL1β. Nitric oxide (NO) and prostaglandin E₂ (PGE₂) release were measured to monitor IL1β response. YAP activity was quantified by YAP nuclear/cytoplasmic ratio and percentage of YAP-positive cells. Mechanical properties of cartilage explants were tested to confirm cartilage degradation. The involvement of primary cilia and IFT was analysed using IFT88 siRNA and ORPK cells with hypomorphic mutation of IFT88.

Results: Treatment with LPA, or increasing polydimethylsiloxane (PDMS) substrate stiffness, activated YAP nuclear expression and inhibited IL1β-induced release of NO and PGE₂, in isolated chondrocytes. Treatment with LPA also inhibited IL1β-mediated inflammatory signalling in cartilage explants and prevented matrix degradation and the loss of cartilage biomechanics. YAP activation reduced expression of primary cilia, knockdown of YAP in the absence of functional cilia/IFT failed to induce an inflammatory response.

Conclusions: We demonstrate that both pharmaceutical and mechanical activation of YAP blocks pro-inflammatory signalling induced by IL1β and prevents cartilage breakdown and the loss of biomechanical functionality. This is associated with reduced expression of primary cilia revealing a potential anti-inflammatory mechanism with novel therapeutic targets for treatment of osteoarthritis (OA).

Keywords: Extracellular matrix stiffness; Inflammatory response; Matrix degradation; Primary cilia; Yes-associated protein (YAP).