Objective To investigate the mechanism by which sodium new houttuyfonate (SNH) inhibits hepatocellular carcinoma (HCC) progression through autophagy regulation via ubiquitin-specific peptidase 22 (USP22)-mediated silent mating type information regulation 2 homolog 1 (SIRT1) deubiquitination. Methods Human liver cancer HepG2 cells were treated with SNH or sorafenib for 24 h, cell viability was detected by CCK-8 assay, cell apoptosis was detected by flow cytometry, and cell invasion ability was detected by Transwell assay. After plasmid transfection induced overexpression of USP22, the USP22-SIRT1 interaction was verified by co-immunoprecipitation (Co-IP). USP22 and SIRT1 protein expressions, as well as SIRT1 ubiquitination were detected by Western blotting. After treatment with SIRT1 activator, autophagy activity was evaluated through mRFP-GFP-LC3 dual fluorescence labeling, adenosine triphosphate (ATP) level measurement, lactate production assay, and analysis of USP22, SIRT1, microtubule-associated protein light chain 3 (LC3)-II/I and p62 protein expressions. For in vivo experiments, 40 BALB/c-nu nude mice were eutectopically transplanted with HepG2 cells and intervened with SNH or sorafenib for 12 d. Tumor volume and weight were monitored every 3 d. Hematoxylin-eosin (HE) staining and TUNEL staining were performed on tumor tissues, and immunohistochemistry was used to detect the protein expressions of nuclear proliferation antigens Ki67, USP22 and SIRT1. Results SNH dose-dependently suppressed HepG2 cell viability and invasion (P < 0.01, 0.001), induced cell apoptosis (P < 0.001). The effect of high-dose SNH was comparable to sorafenib. Co-IP confirmed USP22-SIRT1 protein interaction, which was significantly reduced by high-dose SNH (P < 0.01). High-dose SNH significantly down-regulated USP22 and SIRT1 protein expressions (P < 0.01, 0.001), increased SIRT1 ubiquitination (P < 0.001). These effects were reversed by USP22 overexpression (P < 0.05, 0.001). SNH inhibited autophagy in HepG2 cells, manifested by enhanced GFP/mRFP fluorescence signal (P < 0.01), decreased ATP level (P < 0.001), increased lactate level (P < 0.001), increased p62 expression and decreased expression levels of USP22, SIRT1 and LC3-II/I proteins (P < 0.05, 0.001). SIRT1 activation partially counteracted the autophagic inhibition of SNH (P < 0.05, 0.01, 0.001). In vivo, high-dose SNH significantly reduced tumor volume, weight and malignancy degree (P < 0.001), induced tumor cell apoptosis and decreased Ki67, USP22, and SIRT1 expressions (P < 0.001). Conclusion SNH inhibits HCC progression by suppressing autophagy through USP22-mediated regulation of SIRT1 deubiquitination, providing a potential therapeutic strategy for HCC.

sodium new houttuyfonate  /  hepatocellular carcinoma  /  autophagy  /  USP22  /  SIRT1