TGFα effects on cartilage is mechanical stress-dependent

Purpose: Osteoarthritis (OA) has long been viewed as a degenerative disease of the whole joint, including cartilage, subchondral bone and synovium etc. Accumulating evidence indicates thatmechanical stress has a critical role in its pathogenesis. TGFα is a member of the epidermal growth factor (EGF) family and contains the characteristic EGF-like domain which allows it activates a signaling pathway for cell proliferation, differentiation and development through the EGF receptor (EGFR). The aim of present study was to investigate the effects of TGFα on chondrocytes under stress stimulus. Methods: C57/Bl6 Wt and TGFαKO mice were subjected to DMM or SHAM surgery at 12-weeks of age and running wheels were added the day after surgery into each cage. Knee joints were harvested at 2-, 5- and 10-weeks post-surgery. Catwalk gait analyses, Micro-Computed Tomography (μCT), Toluidine Blue (TB), Picrosirius Red (PR) and Tartrate-Resistant Acidic Phosphatase (TRAP) staining of paraffin-embedded sections were used to investigate gait patterns, 3D joint morphology, bone mineral density (BMD) of subchondral bone, histology, collagen organization and activity of osteoclasts. Besides, primary chondrocytes from femoral head of femoral neck fracture patient and interphalangeal jointof hand of baby with polydactyly of hand were obtained. Using the cell stretching system, primary chondrocytes were cultured with/without human recombinant TGFα. Chondrocytes apoptosis were studied through Annexin V FITC Assay and TUNNEL staining. The metabolic and catabolic markers of chondrocyte were detected by Real-time PCR and Western blot. Moreover, the markers of autophagy were determined by Western blot and Cytofluorescence. Results: Our previous data showed protective effects of cartilage degeneration in TGFα KO mice post-DMM surgery. However, when running wheels were added to encourage activity of mice, the protective effects were reversed. Histology showed more severe OA in TGFα KO mice compared to Wt mice at all three time-points (2-, 5- and 10-week post surgery). μCT showed lager osteophyte formation around the joint in DMM mice. Primary chondrocytes in cell stretching system treated with human recombinant TGFα before or after the stress applied showed less cell apoptosis compared to those without treatment of TGFα. Besides, it is more significant in the condition that cultured chondrocytes was treated with human recombinant TGFα before the stress was applied. Results of Annexin V FITC assay using flow cytometry showed that the primary chondrocytesunder the treatment of TGFα and mechanical stress demonstrated less cell appotosis than those under mechanical stress stimulation only. Western blot showed that primary chondrocytes under mechanical stress stimulation showed decreased of MMP13 and increased of ACAN if treated with TGFα . Moreover, TGFα can promoted chondrocyte autophagy induced by mechanical stress, with more significant effects when TGFα were administrated before mechanical stress. Conclusions: TGFα effect on chondrocytes is mechanical stress-dependent. It can protects the chondrocytes under mechanical stress by inhibiting cell apoptosis and decreasing the catabolism of chondrocytes.