Interactions between primary cilia length and hedgehog signalling in response to mechanical and thermal stress.

Abstract

The primary cilium is a microtubule-based organelle present on the majority of interphase cells where it functions as a hub for numerous signalling pathways including hedgehog signalling. Chondrocytes, the unique cellular component of articular cartilage, possess primary cilia which are required for mechanotransduction and maintenance of a healthy extracellular matrix. However in osteoarthritis there is an increase in primary cilia length and prevalence associated with aberrant activation of hedgehog signalling which promotes cartilage degradation. The aim of this thesis was therefore to examine the influence of biophysical stimuli on chondrocyte primary cilia structure and function, relating changes in ciliary length to perturbations in hedgehog signalling. An in vitro mechanical loading model was established to study the influence of cyclic tensile strain on chondrocyte primary cilia. Loading at 10% strain activated hedgehog signalling measured by expression of Gli1 and Ptch1. Cilia progressively disassembled in response to increasing levels of mechanical strain in a manner dependent upon tubulin deacetylation. Cilia disassembly at 20% strain was associated with the loss of mechanosensitive hedgehog signalling despite continued expression of hedgehog ligand (Ihh). Therefore this behaviour may function as a protective mechanism limiting hedgehog-mediated cartilage degradation in response to high levels of mechanical strain. To further understand the influence the extracellular environment exerts over ciliary function, a second in vitro model was developed investigating the effects of thermal stress. In chondrocytes and fibroblasts, primary cilia underwent rapid resorption in response to elevated temperature and ligand mediated hedgehog signalling was inhibited. These studies demonstrate that regulated disassembly of the cilium in response to physical stress modulates both cilia size and function. In particular, the findings suggest that changes in the chondrocyte physical environment affect cilia structure and function and may therefore be an important factor in the aetiology of cartilage disease.