Taladegib controls early chondrocyte hypertrophy via inhibiting smoothened/Gli1 pathway

Chondrocyte hypertrophy is a common phenomenon in osteoarthritis (OA). Indian hedgehog (Ihh) is secreted by pre-hypertrophic chondrocytes, which regulates the hypertrophy and mineralization of chondrocytes during cartilage osteogenesis. Smoothened (Smo) is a connecting protein among the Ihh signaling pathway that triggers glioma-associated oncogene homologue 1 (Gli1) to active the hypertrophic process. In this study, we aimed to examine a new inhibitor of Smo in the prevention of chondrocyte hypertrophy during OA. We collected human joint cartilage from the OA patients undergoing knee arthroplasty. Chondrocytes in different OA degrees were isolated and divided them into mild and severe groups. Alginate beads (ABs) was used to establish a chondrocyte hypertrophy model. The expression of type X collagen, MMP-13, Runx-2, type II collagen, SOX-9, and aggrecan were determined using immunofluorescence, Western blot, real-time polymerase chain reaction (qRT-PCR). Cell counting kit-8 (CCK-8) assay was performed to assess the viability of chondrocytes. ABs treatment accelerated the process of chondrocyte hypertrophy and upregulated the expression of type X collagen, Matrix metalloproteinase-13 (MMP-13), Runx-2 but decreased type II collagen, SOX-9, and aggrecan both in protein and mRNA levels, which abolished by the present of Taladegib with the activation of Smo and Gli1. However, in the severe OA chondrocytes, Taladegib lost the ability to reverse hypertrophic chondrocytes to a healthy state and made no sense to the expression of type X collagen and Gli1. Our results reveal Taladegib as a novel drug in controlling chondrocyte hypertrophy depending on Smo blocking, which plays a vital role in the homeostasis of cartilage and the development of OA. Besides, we found that Taladegib only works in the previous stage of chondrocytes hypertrophy but not in the later of the process.