Study on the Mechanism of PRDX5 Regulating ACSL4 Ubiquitination via UFD1 in Ferroptosis Associated with Contrast-Induced Nephropathy

Background: Contrast-induced nephropathy (CIN) is a common complication caused by iodinated contrast media, with no specific treatment available. Ferroptosis may contribute to its pathogenesis, and peroxiredoxin 5 (PRDX5) is downregulated in the urine of CIN patients. This study investigates whether PRDX5 affects ferroptosis in CIN by regulating the ubiquitination of acyl-CoA synthetase long-chain family member 4 (ACSL4). Methods: HK-2 cells and a mouse CIN model (induced by iohexol) were established. Groups included: control, CIN, CIN + ferroptosis inhibitor (Fer-1), PRDX5 overexpression, and PRDX5 overexpression + CIN. PRDX5, ACSL4, GPX4, and SLC7A11 expression were analyzed (qPCR/Western blot). Cell viability was measured via MTT assay. Fe2+and reactive oxygen species (ROS) levels were quantified. Co-immunoprecipitation (Co-IP) combined with LC-MS/MS identified UFD1 as a PRDX5-interacting protein. Immunofluorescence confirmed co-localization. Ubiquitination assays evaluated UFD1-mediated regulation of ACSL4. Renal function and histopathology were assessed in mice. Results: 1.Ferroptosis in CIN: The CIN group exhibited reduced cell proliferation, increased ACSL4, decreased GPX4/SLC7A11, and elevated Fe2+/ROS levels, which were reversed by Fer-1 (P<0.05). 2.Protective role of PRDX5: PRDX5 expression was downregulated in CIN. Its overexpression alleviated cellular damage, suppressed ACSL4, restored GPX4/SLC7A11 levels, and reduced Fe2+/ROS accumulation. In mice, PRDX5 overexpression improved renal injury and lowered serum creatinine. 3.PRDX5-UFD1 interaction: UFD1 was identified as a PRDX5-binding partner via mass spectrometry. Co-IP and immunofluorescence confirmed their co-localization, which weakened in CIN. 4.UFD1 regulates ACSL4 ubiquitination: UFD1 knockdown reduced ACSL4 ubiquitination and degradation, leading to its accumulation. UFD1 overexpression or proteasome inhibitor MG132 reversed this effect. Immunofluorescence demonstrated co-localization of UFD1 with ACSL4 and ubiquitin, suggesting UFD1 binds ACSL4’s intracellular domain. Conclusion: PRDX5 inhibits ferroptosis and mitigates CIN by binding UFD1 to promote ACSL4 ubiquitination and degradation. These findings propose novel therapeutic strategies targeting the PRDX5-UFD1-ACSL4 axis in CIN. Funding: Government Support – Non-U.S.