Celastrol promotes apoptosis of human osteosarcoma HOS cells through endoplasmic reticulum stress pathway / 肿瘤

ABSTRACT

Objective: To investigate the effect of celastrol on the apoptosis of human osteosarcoma cell line HOS, and to explore the underlying mechanism. Methods: After treatment with different concentrations (0, 1.5, 2.5, 3.5 and 4.5 μmol/L) of celastrol for 24 h, the proliferation of HOS cells was detected by CCK-8 assay to determine the optimum experimental concentration (3.0 μmol/L). The cell morphology was observed by inverted phase contrast microscopy, and the apoptotic morphology was detected by fluorescence microscopy after Hoechst33258 staining. The apoptotic rate and Ca2+ concentration were analyzed by flow cytometry. The expressions of binding immunoglobulin protein (BIP), calnexin (CNX), endoplasmic reticulum oxidoreductin 1-L alpha (Ero1-Lα), protein disulfide isomerase (PDI), inositol-requiring enzyme 1alpha (IRE1α), protein kinaselike endoplasmic reticulum kinase (PERK), phosphorylated-PERK (p-PERK), CCAAT/enhancerbinding protein homologous protein (CHOP), cleaved-caspase-12 (c-caspase-12) and c-caspase-3 were determined by Western blotting. Results: The different concentrations of celastrol inhibited the proliferation of HOS cells in a dose-dependent manner (all P < 0.05). After treatment with 3 μmol/L celastrol for 24 h, the morphology of HOS cells was obviously atrophic and disordered, the floating cells increased, and the typical morphological changes of apoptotic cells such as nuclear pyknosis and fragmentation appeared. As compared with the untreated control group, the apoptotic rate and Ca2+ concentration in HOS cells treated with 3 μmol/L celastrol for 24 h were significantly increased (both P < 0.05), and the expressions of endoplasmic reticulum stressrelated marker proteins BIP, CNX, Ero1-Lα, PDI, IRE1α, p-PERK and endoplasmic reticulum stress-mediated apoptosis-related proteins c-caspase-12, CHOP and c-caspase-3 were upregulated (all P < 0.05). Conclusion: Celastrol can induce the apoptosis of human osteosarcoma HOS cells, and the mechanism may be related to endoplasmic reticulum stress-mediated apoptosis pathway.