[Interferon-α induces disulfidptosis occurrence in human liver cancer cells]

Objective: To investigate the potential molecular regulatory mechanism of interferon-α (IFN-α)-induced disulfidptosis occurrence in human liver cancer cells. Methods: Glucose starvation and IFN-α treatment models were constructed using human hepatocellular carcinoma cell lines HepG2 and Huh7, respectively. Western blotting (WB), NADPH content measurement, and immunofluorescence staining were employed to evaluate disulfidptosis-related phenotypes, such as intracellular disulfide bond accumulation, NADPH depletion, and filamentous actin (F-actin) skeleton collapse. The reversibility of the phenotypes was validated by combining the disulfidptosis inhibitor D-penicillamine (D-Pen). The expression of solute carrier family 7 member 11 (SLC7A11) was downregulated using siRNA interference to evaluate the dependence of IFN-α-induced disulfidptosis on the SLC7A11 pathway. Furthermore, the GEO transcriptome dataset was employed to conduct differential gene analysis for screening candidate molecules involved in the regulation of IFN-α-induced disulfidptosis. Quantitative data are presented as mean ± standard deviation (SD). Comparisons between two groups were performed using the independent-samples t test, while comparisons among multiple groups were analyzed by one-way analysis of variance (ANOVA), followed by Dunnett's t test for pairwise comparisons. Results: Compared with the control group, glucose starvation and IFN-α treatment both induced disulfidptosis-related phenotypes in HepG2 and Huh7 cells, characterized by increased intracellular disulfide bond levels, decreased NADPH content, and F-actin cytoskeleton collapse. Following IFN-α treatment, the accumulation of high-molecular-weight protein aggregates increased in a time- and dose-dependent manner. Compared with IFN-α treatment alone, combined D-Pen intervention partially alleviated intracellular disulfide accumulation, NADPH depletion, and F-actin cytoskeletal collapse. Under conditions of SLC7A11 knockdown, IFN-α-induced high-molecular-weight protein aggregation was further aggravated, suggesting that regulatory mechanisms independent of SLC7A11 may also be involved in this process. Transcriptomic differential expression analysis revealed that β2-microglobulin (B2M), ubiquitin-specific peptidase 18 (USP18) and proteasome activator subunit 1 (PSME1) were upregulated following IFN-α stimulation, among which PSME1 also showed increased protein expression after IFN-α treatment. Conclusion: IFN-α can induce disulfidptosis-related phenotypic changes in HepG2 and Huh7 human hepatocellular carcinoma cells. The upregulation of B2M, USP18, and PSME1 suggests that these molecules may be involved in this process; however, the specific underlying mechanisms require further investigation.