Triptolide impedes high glucose-induced cell function in HK2 cells through PRKN-mediated ubiquitination of ACSL1

Triptolide (TP), a bioactive compound, demonstrates efficacy in ameliorating diabetic nephropathy (DN). This study aimed to investigate the role of TP in renal tubular injury during DN and elucidate the underlying mechanism involving acyl-CoA synthetase long-chain family member 1 (ACSL1) and parkin (PRKN). DN model was induced in HK2 cells by high glucose (HG, 30 mmol/L). Cell counting kit-8, EdU assay, flow cytometry were used to assess cell viability, proliferation, and apoptosis. Inflammatory cytokines were measured via enzyme-linked immunosorbent assay. Ferroptosis was assessed by detecting reactive oxygen species (ROS), lipid peroxidation (MDA), Fe2+, and glutathione (GSH) using kits. The mRNA and protein examination was performed by real-time quantitative PCR and western blotting. Co-immunoprecipitation assay was conducted for protein interaction and ubiquitination detection. DN in mice was established by high-fat diet and streptozocin injection. The effects of TP on mice were analyzed by histopathology analysis, biochemical analysis, and protein detection. TP mitigated HG-induced apoptosis, inflammation, and ferroptosis in HK2 cells. The protective effects of TP against HG-induced injury in HK2 cells were mediated by the inhibition of ACSL1. PRKN promoted ubiquitination of ACSL1 to reduce the protein level of ACSL1. PRKN/ACSL1 inhibited HG-induced HK2 cell dysfunction. The protective effect of TP in HG-stimulated HK2 cells was mediated by the upregulation of PRKN. TP activated anti-ferroptosis NRF2/SLC7A11/GPX4 pathway by targeting ACSL1. TP could ameliorate kidney injury in DN mice through modulating PRKN, ACSL1, and NRF2/SLC7A11/GPX4 pathway. All these discoveries suggested that TP protected HK2 cells from HG-triggered kidney injury through upregulating PRKN that further promoted ubiquitination of ACSL1.