Sirt1 mitigates hepatic lipotoxic injury induced by high-fat-diet in fish through Ire1α deacetylation

Background Silent information regulator protein 1 (Sirt1) is crucial in regulating lipid metabolism, but its specific role and mechanism in fish hepatic lipotoxic injury remain undefined. Objectives This study aimed to elucidate the regulatory role of Sirt1 and the underlying mechanisms in dietary lipid-induced hepatic lipotoxic injury in a marine teleost black seabream. Methods Black seabream were fed a control diet (12% lipid level), HFD (18% lipid level, oleic acid [OA]-rich), or HFD supplemented with 0.25%, 0.50%, or 1.00% resveratrol (RSV) for 8 weeks. The cultured hepatocytes were stimulated by OA (OA, 200μM), OA supplemented with RSV (20μM), or transfection with sirt1 -small interfering RNA (si sirt1 ). Biochemical indices, gene expression (qPCR), and histology, transmission electron microscope (TEM), immunofluorescence, Western blot (WB), flow cytometry (FCM), and immunoprecipitation assays were conducted to evaluate hepatic lipid deposition, lipid metabolism, endoplasmic reticulum stress (ERS), inflammation and apoptosis, and determine protein interactions between Sirt1 and Ire1α. Results In vivo , RSV supplementation increased mRNA and protein expression levels of sirt1 (236.2%±16.1%, and 53.1%±14.3%) and down-regulated the mRNA and phosphorylated protein expression levels of ire1α / Ire1α (46.0%±7.6% and 38.6%± 7.0%,), jnk /Jnk (57.6±7.3% and 122.1%) and nf-κb /Nf-κb p65 (41.7%±7.1% and 24.6%±0.8%) compared to the HFD group. Similar patterns were found in the in vitro experiments, however, after knockdown of sirt1 , although the cells were incubated with RSV, the expression levels of ire1α / Ire1α , jnk /Jnk and nf-κb /Nf-κb p65 showed no significant differences compared to the OA treatment. Moreover, we found that mutation of K61 to arginine to mimic Ire1α deacetylation confers protection against Ire1α mediated OA-rich HFD-induced inflammation and apoptosis. Conclusion The findings revealed that Sirt1 protects against OA-rich HFD-induced hepatic lipotoxic injury via deacetylation of Ire1α on K61, hence, reducing Ire1α autophosphorylation level, and suppressing Jnk and Nf-κb p65 activation. This mechanism is elucidated for the first time in fish.

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