H₂Se Induces Reductive Stress in HepG2 Cells and Activates Cell Autophagy by Regulating the Redox of HMGB1 Protein under Hypoxia

Abstract

Rationale: Selenium has been shown to have chemotherapeutic effects against cancer. However, the anti-cancer mechanism of selenium is not fully understood, and the role of hydrogen selenide (H 2 Se), which is a common metabolite of dietary selenium compounds, has not been elucidated due to the lack of detection methods. In this study, we revealed a new anti-cancer mechanism of selenite with the help of a H 2 Se fluorescent probe. Methods: HepG2 cells were cultured under a simulated tumor hypoxic microenvironment. The H 2 Se and H 2 O 2 levels were detected by fluorescent probes in living cells and in mice. Autophagic and apoptotic proteins were detected by Western blotting. The redox of HMGB1 protein were analyzed by non-reducing sodium dodecyl sulfate polyacrylamide gel electrophoresis. Results: After pharmacological doses of Na 2 SeO 3 treatment of HepG2 cells under hypoxic conditions, high levels of H 2 Se were produced before cell death. The H 2 Se accumulation resulted in reductive stress instead of oxidative stress, which was induced by Na 2 SeO 3 treatment under normoxic conditions. Furthermore, H 2 Se targeted the HMGB1 protein and induced cell autophagy. H 2 Se could interrupt the disulfide bond in HMGB1 and promote its secretion. The reduced HMGB1 outside the cells stimulated cell autophagy by inhibiting the Akt/mTOR axis. Here, autophagy played a dual role, i.e., mild autophagy inhibited apoptosis, while excessive autophagy led to autophagy-associated cell death. Conclusions: These results show that H 2 Se plays a key role during HepG2 cell death induced by selenite. Our findings reveal a new anti-cancer mechanism of selenite and provide a new research area for selenium studies.

Secretory autophagy machinery and vesicular trafficking are involved in HMGB1 secretion

“…Furthermore, post-translational modifications of HMGB1, such as oxidation, helps its binding to XPO1 [27,48]. Cytoplasmic HMGB1 activates autophagy [48][49][50], and this autophagy activation induces the extracellular secretion of HMGB1. The autophagy machinery ATG5, ATG7, ATG12, and ATG14, and GORASP2 are involved in the packaging of cytosolic HMGB1 into phagophores.…”

Monitoring NAD(P)H by an ultrasensitive fluorescent probe to reveal reductive stress induced by natural antioxidants in HepG2 cells under hypoxia

“…The cell culture was performed according to our previous paper. 39 Briey, HepG2 cells were cultured in DMEM (high glucose) supplemented with 10% FBS and 1% antibiotics (100 U mL À1 penicillin and 100 mg mL À1 streptomycin) at 37 C in a humidied atmosphere of 95% air (20% O 2 ) and 5% CO 2 . For simulating the tumor hypoxic microenvironment, HepG2 cells were cultured in a mixture containing 1% O 2 , 94% N 2 and 5% CO 2 using a low oxygen incubator (Bugbox M, Ruskinn, England).…”

Fluorescent probes based on nucleophilic aromatic substitution reactions for reactive sulfur and selenium species: Recent progress, applications, and design strategies

“…Fluorescent probes can be used to remotely measure analytes and can be used in environments and samples that are difficult to access directly to avoid possible damage when the analyte is taken out of its natural medium. Fluorescent probes have become important tools in chemical and biochemical research due to their non-invasiveness, high sensitivity, high selectivity, operability, low cost, rapid response, high time resolution, and appropriate beneficial characteristics for easy signal detection [60] , [61] , [62] , [63] , [64] , [65] .…”