Hedgehog signalling is aberrantly activated in osteoarthritis (OA) a chronic degenerative disease that affects articular cartilage [1]. Hedgehog signalling requires the primary cilium [2]. The majority of chondrocytes possess a cilium and cilia incidence and length is increased in osteoarthritic cartilage compared with healthy cartilage [3]. In this study we investigated the hypothesis that Indian Hedgehog (Ihh) gene expression is regulated by mechanical loading in mature articular chondrocytes and that changes in primary cilia length and prevalence influence the capacity of these cells to respond to Hh signals. Articular chondrocytes were harvested from bovine metacarpal-phalangeal joints obtained from an abattoir. Chondrocytes were subjected to cyclic tensile strain using the Flexcell 4000T system and stimulated with 5, 10 or 20% strain at 0.33Hz for 1hr. Primary cilia incidence and length were quantified using confocal images of cells labelled with acetylated a-tubulin (n>300 cells). Quantitative real-time PCR was used to monitor Ihh gene expression and the expression of Patched1 (Ptch1) which provides a measure of Hh pathway activation. For these experiments n=5 replicates with cells pooled from 3 different animals. Statistical analysis was conducted using t-tests. Mechanical loading induced a dose dependant reduction in primary cilia length compared to unloaded controls with statistically significant differences following loading at 10% and 20% strain. Loading had no effect on cilia prevalence. Ihh gene expression was up-regulated only at 10% strain (p<0.05). This was associated with an increase in Ptch1 gene expression (p<0.05) which also occurred at 5% strain (p<0.05). We have shown for the first time that Ihh exhibits magnitude dependent mechanosensitive expression in mature articular chondrocytes and that mechanical loading induces changes in chondrocyte primary cilia length. Hh pathway activation was observed at 5% strain, despite no significant changes in Ihh expression. This suggests that mechanical loading may also be inducing the release of endogenous Ihh. The lack of pathway activation at 20% may be due to reduced cilia length as well as a lack of Ihh expression. Understanding Hh regulation in mature chondrocytes is essential for development of future pharmaceutical treatments for OA based on manipulation of Hh signalling.