Linear ubiquitination of LKB1 activates AMPK pathway to inhibit NLRP3 inflammasome response and reduce chondrocyte pyroptosis in osteoarthritis

Osteoarthritis (OA) is the most common chronic disease. It is characterized by high levels of clinical heterogeneity and low inflammation. Therefore, elucidation of the mechanisms that regulate gene expression is critical for developing effective OA therapies. This study aimed to explore the role of LKB1/AMPK in the progression of OA.

LKB1/AMPK pathway significantly ameliorated NLRP3 inflammasome response and chondrocyte injury. Activation of AMPK pathway by linear ubiquitination of LKB1 may be a potential target for OA treatment. The translational potential of this article: This study highlights the importance of the LKB1/AMPK pathway in NLRP3 inflammatory body response and chondrocyte injury. Activation of LKB1 by modulating linear ubiquitination may be a potential target for OA treatment.

Anterior cruciate ligament transection (ACLT) was performed on Sprague Dawley (SD) rats right knee to construct OA model, followed by AICAR [AMP-activated protein kinase (AMPK) activator] treatment. The level changes [AMPK, IL-10, IL-13, IL-1β, TNF-α, IL-6, ASC, Caspase-1, Ki67, and hibit Nod-like receptor protein 3 (NLRP3)] and the degree of tissue injury were assessed by western blot, Immunohistochemical (IHC), Enzyme-linked immunosorbent assay (ELISA), Hematoxylin-eosin staining (HE), Immunofluorescence (IF), Terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay, and Safranin O and Fast Green staining (S-O). Human chondrocytes were induced by LPS to construct a cellular inflammatory model, and then transfected with oe-AMPK or oe-HOIL-1-interacting protein (HOIP). Cell viability/apoptotic and intracellular content of AMPK, HOIP, IL-1β, IL-10, IL-13, TNF-α, IL-6, ASC, NLRP3 and Caspase-1 were measured by western blot, ELISA, CCK-8, IF, flow cytometry and TUNEL assays.

After AICAR treatment with OA rats, the expression of p-AMPK, IL-10, IL-13, Ki67 and Bcl-2 increased, the level of NLRP3 inflammasome, TNF-α, IL-6, Bax and Caspase-3 levels were decreased, and tissue damage and apoptosis were significantly alleviated. After transfected with oe-LKB1, chondrocyte activity and LKB1 linear ubiquitination were enhanced, and the level of HOIP, p-AMPK, IL-10 and IL-13 were increased. In contrast, NLRP3 inflammasome (ASC, NLRP3, Caspase-1, IL-1β, and cleaved Caspase-1), TNF-α, and IL-6 levels decreased, apoptosis rate and TUNEL positive rate were attenuated.