| Abstrakt: |
Nonalcoholic fatty liver disease (NAFLD) is a global health concern with the acid sphingomyelinase (ASM)/ceramide (CE) pathway and the NOD-like receptor family, pyrin domain-containing protein 3 (NLRP3) inflammasome identified as pivotal players in lipid disorders and inflammation. This study explores the interaction mechanism between the ASM/CE pathway and NLRP3 in NAFLD cell models, aiming to understand the impact of amitriptyline (Ami), an ASM inhibitor, on lipid deposition and hepatocyte injury by regulating the ASM/CE-NLRP3 pathway. Methods: HepG2 and HL-7702 cells were exposed to free fatty acids (FFAs) to establish the NAFLD model. The cells were divided into 5 groups: control group, model group, Ami group, tumor necrosis factoralpha (TNF-α) group, and Ami + TNF-α group. Intracellular lipid droplets were visualized using Oil Red O staining, and Western blot analysis quantified ASM, NLRP3, and caspase 1 protein expression. Enzyme linked immunosorbent assay (ELISA) was measured CE and ASM levels, while qRT-PCR assessed mRNA expression. The apoptotic rate was evaluated by flow cytometry (FCM). Results: Following FFAs incubation, significant increases in ASM and CE levels were observed in HepG2 and HL-7702 cells, accompanied by elevated expression of NLRP3, and caspase 1, and IL-1ß. TNF-α treatment further amplified these indicators. Ami demonstrated a reduction in lipid deposition, suppressed ASM/CE pathway activation, downregulated NLRP3 and caspase 1 expression, and improved apoptosis. Additionally, MCC950, a selective inhibitor of the NLRP3, mitigated NLRP3, caspase 1, and IL-1β expression, alleviating lipid deposition and apoptosis in the NAFLD cell model. Conclusion: The ASM/CE-NLRP3 pathway in NAFLD cells promotes hepatocyte steatosis, inflammation, and cell damage. Ami emerges as a promising therapeutic agent by inhibiting the ASM/CE-NLRP3 pathway, underscoring its potential as a key target for NAFLD treatment. [ABSTRACT FROM AUTHOR] |
