MTORC1 coordinates the autophagy and apoptosis signaling in articular chondrocytes in osteoarthritic temporomandibular joint

Abstract

A switch from autophagy to apoptosis is implicated in chondrocytes during the osteoarthritis (OA) progression with currently unknown mechanism(s). In this study we utilized a flow fluid shear stress (FFSS) model in cultured chondrocytes and a unilateral anterior crossbite (UAC) animal model. We found that both FFSS and UAC actively induced endoplasmic reticulum stress (ERS) in the temporomandibular joints (TMJ) chondrocytes, as demonstrated by dramatic increases in expression of HSPA5, p-EIF2AK3, p-ERN1 and ATF6. Interestingly, both FFSS and UAC activated not only pro-death p-EIF2AK3-mediated ERS-apoptosis programs but also pro-survival p-ERN1-mediated autophagic flux in chondrocytes. Data from FFSS demonstrated that MTORC1, a downstream of p-ERN1, suppressed autophagy but promoted p-EIF2AK3 mediated ERS-apoptosis. Data from UAC model demonstrated that at early stage both the p-ERN1 and p-EIF2AK3 were activated and MTORC1 was inhibited in TMJ chondrocytes. At late stage, MTORC1-p-EIF2AK3-mediated ERS apoptosis were predominant, while p-ERN1 and autophagic flux were inhibited. Inhibition of MTORC1 by TMJ local injection of rapamycin in rats or inducible ablation of MTORC1 expression selectively in chondrocytes in mice promoted chondrocyte autophagy and suppressed apoptosis, and reduced TMJ cartilage loss induced by UAC. In contrast, MTORC1 activation by TMJ local administration of MHY1485 or genetic deletion of Tsc1, an upstream MTORC1 suppressor, resulted in opposite effects. Collectively, our results establish that aberrant mechanical loading causes cartilage degeneration by activating, at least in part, the MTORC1 signaling which modulates the autophagy and apoptosis programs in TMJ chondrocytes. Thus, inhibition of MTORC1 provides a novel therapeutic strategy for prevention and treatment of OA.

Moderate Fluid Shear Stress Regulates Heme Oxygenase-1 Expression to Promote Autophagy and ECM Homeostasis in the Nucleus Pulposus Cells

“…*P < 0.05, **P < 0.01, and ***P < 0.001 vs. FSS + IFT88-siRNA group. Zhang et al, 2018;Yang et al, 2019). In line with previous studies, our data demonstrated that moderate FSS activated autophagy in NP cells by increasing the ratio of LC3-II to LC3-I, the protein expression of Beclin-1 and the number of autophagosomes and autolysosomes.…”

“…The cells were cultured in Dulbecco's Modification of Eagle's Medium (10-013-CVR; Corning, United States) containing 10% FBS (10099-141; Gibco, Australia) supplemented with 1% penicillin-streptomycin (SV30010; Hyclone, United States) at 37 • C with 5% CO 2 . FSS experiments were conducted as previously described (Yang et al, 2019). Cells were seeded onto collagen I-coated culture slips (75 mm × 25 mm × 1 mm; FFCS-C; Flexcell, United States) at a density of 3.0 × 10 4 /cm 2 and incubated in a 5% CO 2 incubator at 37 • C. When cells reached up to 85% confluence, the slips were then placed in a parallel plate flow chamber of Streamer R System (STR-4000; Flexcell, United States) ( Figure 1B) and cells are exposed to 12 or 24 dyne/cm 2 FSS for 0, 1, 2, 3, and 4 h. For certain experiments, NP cells were pre-treated with 10 µM CoPP (Sigma, United States, C1900) for 1 h or 500 nM rapamycin (Selleck, United States, S1039) for 12 h before exposure to FSS.…”

“…Fluid shear stress activates autophagy in multiple cell types (Zhang et al, 2018;Yang et al, 2019). Thus, we determined whether FSS activates autophagy in NP cells.…”

Protein delivery in intermittent and continuous enteral nutrition with a protein-rich formula in critically ill patients - A protocol for the prospective randomised controlled proof-of-concept Protein Bolus Nutrition (Pro BoNo) study

“…For the catabolic pathway, forkhead box class O-3, (1,3,52,54) muscle atrophy F-box, (1,3) muscle ring-nger protein-1 (1,3,52) and nuclear factor kappa-beta (1,3) will be measured. For information about autophagy, ULK1, (54,55) light-chain 3 (55,56) and p62 (55,56) will be analysed. For an overview of enzyme regulation, AMP-activated protein kinase (8,25,(53)(54)(55) will be studied.…”

Protein delivery in intermittent and continuous enteral nutrition with a protein-rich formula in critically ill patients - A protocol for the prospective randomised controlled proof-of-concept Protein Bolus Nutrition (Pro BoNo) study

“…For the catabolic pathway, forkhead box class O-3, (1,3,35,37) muscle atrophy F-box, (1,3) muscle ring-finger protein-1 (1,3,35) and nuclear factor kappa-beta (1,3) will be measured. For information about autophagy, ULK1, (37,38) light-chain 3 (38,39) and p62 (38,39) will be analysed. For an overview of enzyme regulation, AMP-activated protein kinase (6,23,(36)(37)(38) will be studied.…”