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PLC?1 inhibition-driven autophagy of IL-1?-treated chondrocyte confers cartilage protection against osteoarthritis, involving AMPK, Erk and Akt.

ABSTRACT: Previous studies identified the involvement of phosphoinositide-specific phospholipase C (PLC) ?1 in some events of chondrocytes. This study aims to investigate whether and how PLC?1 modulates autophagy to execute its role in osteoarthritis (OA) progression. Rat normal or human OA chondrocytes were pretreated with IL-1? for mimicking or sustaining OA pathological condition. Using Western blotting, immunoprecipitation, qPCR, immunofluorescence and Dimethylmethylene blue assays, and ELISA and transmission electron microscope techniques, we found that PLC?1 inhibitor U73122 enhanced Collagen II, Aggrecan and GAG levels, accompanied with increased LC3B-II/I ratio and decreased P62 expression level, whereas autophagy inhibitor Chloroquine partially diminished its effect. Meanwhile, U73122 dissociated Beclin1 from Beclin1-IP3R-Bcl-2 complex and blocked mTOR/ULK1 axis, in which the crosstalk between PLC?1, AMPK, Erk and Akt were involved. Additionally, by haematoxylin and eosin, Safranin O/Fast green, and immunohistochemistry staining, we observed that intra-articular injection of Ad-shPLC?1-1/2 significantly enhanced Collagen and Aggrecan levels, accompanied with increased LC3B and decreased P62 levels in a rat OA model induced by anterior cruciate ligament transection and medial meniscus resection. Consequently, PLC?1 inhibition-driven autophagy conferred cartilage protection against OA through promoting ECM synthesis in OA chondrocytes in vivo and in vitro, involving the crosstalk between PLC?1, AMPK, Erk and Akt.

SUBMITTER: Chen X

PROVIDER: S-EPMC7875910 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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PLCγ1 inhibition-driven autophagy of IL-1β-treated chondrocyte confers cartilage protection against osteoarthritis, involving AMPK, Erk and Akt.

Chen Xiaolei X Wang Yue Y Qu Ning N Zhang Bing B Xia Chun C

Journal of cellular and molecular medicine 20201228 3

Previous studies identified the involvement of phosphoinositide-specific phospholipase C (PLC) γ1 in some events of chondrocytes. This study aims to investigate whether and how PLCγ1 modulates autophagy to execute its role in osteoarthritis (OA) progression. Rat normal or human OA chondrocytes were pretreated with IL-1β for mimicking or sustaining OA pathological condition. Using Western blotting, immunoprecipitation, qPCR, immunofluorescence and Dimethylmethylene blue assays, and ELISA and tran ...[more]

PMID: 33372388

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Project description:BackgroundsAbnormal cartilage calcification is one of the pathological changes of temporomandibular joint (TMJ) osteoarthritis (OA). Recent studies have reported that exosomes can regulate the formation of abnormal calcified nodules in diseases including atherosclerosis and chronic kidney disease. However, the influences of chondrocyte-derived exosomes on abnormal cartilage calcification in TMJ OA are still unclear.MethodsTMJ OA was induced by unilateral anterior crossbite (UAC) for 4, 8, or 12 weeks in rats to observe abnormal calcification in TMJ condylar cartilage and exosome formation. Concomitantly, GW4869, the inhibitor of exosome formation, was locally injected to the TMJ of rats under stimulation of UAC, while the exosomes extracted from primary condylar chondrocytes stimulated with fluid flow shear stress (FFSS) were locally injected to rats TMJ.ResultsAbnormal calcification was enhanced in the degenerative cartilage of TMJ OA in UAC rats, and a large number of exosome-like structures with diameters of 50-150 nm were found in the calcified cartilage together with decreased expression of matrix Gla protein (MGP) and increased expression of CD63, tissue-nonspecific alkaline phosphatase (TNAP) and nucleotide pyrophosphatase/phosphodiesterase-1 (NPP1). After FFSS stimulation, the number of exosomes secreted by chondrocytes and the numbers of calcified nodules were increased in cultured cells, and the protein levels of MGP, TNAP, and NPP1 in exosomes were changed. Inhibition of exosome formation, TNAP, and NPP1 or supplementation with exogenous MGP effectively alleviated FFSS-induced chondrocyte calcification. Local injection of GW4869, the exosome inhibitor, alleviated TMJ OA-related cartilage degeneration and calcification in UAC rats. Local injection of exosomes obtained from chondrocytes stimulated by FFSS to the TMJs of normal rats induced cartilage degeneration and calcification similar to that in TMJ OA.ConclusionsAbnormal biomechanical loading leads to enhanced formation of chondrocyte-derived exosomes, in which promoters of calcification increased and inhibitors decreased, resulting in accelerating abnormal cartilage calcification in TMJ OA. The inhibition of degenerative chondrocyte-derived exosomes is expected to be a new way to prevent and treat TMJ OA.

Dataset Information

PLC?1 inhibition-driven autophagy of IL-1?-treated chondrocyte confers cartilage protection against osteoarthritis, involving AMPK, Erk and Akt.

Publications

PLCγ1 inhibition-driven autophagy of IL-1β-treated chondrocyte confers cartilage protection against osteoarthritis, involving AMPK, Erk and Akt.

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