title: Bufalin induces the interplay between apoptosis and autophagy in glioma cells through endoplasmic reticulum stress.
abstract: Malignant gliomas are common primary tumors of the central nervous system. The prognosis of patients with malignant glioma is poor in spite of current intensive therapy and thus novel therapeutic modalities are necessary. Bufalin is the major component of Chan-Su (a traditional Chinese medicine) extracts from the venom of Bufo gargarizan. In this study, we evaluated the growth inhibitory effect of bufalin on glioma cells and explored the underlying molecular mechanisms. Our results showed that bufalin inhibited the growth of glioma cells significantly. Mechanistic studies demonstrated that bufalin induced apoptosis through mitochondrial apoptotic pathway. In addition, bufalin was also found to induce ER stress-mediated apoptosis, which was supported by the up- regulation of ER stress markers, CHOP and GRP78, and augmented phosphorylation of PERK and eIF2α as well as cleavage of caspase-4. Downregulation of CHOP using siCHOP RNA attenuated bufalin-induced apoptosis, further confirming the role of ER stress response in mediating bufalin-induced apoptosis. Evidence of bufalin-induced autophagy included formation of the acidic vesicular organelles, increase of autophagolysosomes and LC3-II accumulation. Further experiments showed that the mechanism of bufalin-induced autophagy associated with ATP deleption involved an increase in the active form of AMPK, decreased phosphorylation levels of mTOR and its downstream targets 4EBP1 and p70S6K1. Furthermore, TUDC and silencing of eIF2α or CHOP partially blocked bufalin-induced accumulation of LC3-II, which indicated that ER stress preceded bufalin-induced autophagy and PERK/eIF2α/CHOP signaling pathway played a major part in the process. Blockage of autophagy increased expression of ER stress associated proteins and the ratio of apoptosis, indicating that autophagy played a cytoprotective role in bufalin induced ER stress and cell death. In conclusion, bufalin inhibits glioma cell growth and induces interplay between apoptosis and autophagy through endoplasmic reticulum stress. It will provide molecular bases for developing bufalin into a drug candidate for the treatment of maglinant glioma.
authors.affiliation: 1. Institute of Biochemistry, Zhejiang University, Hangzhou, 310058, PR China;
authors.forename: Shuying
authors.initials: S
authors.lastname: Shen
authors.affiliation: 1. Institute of Biochemistry, Zhejiang University, Hangzhou, 310058, PR China;
authors.forename: Yi
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: 1. Institute of Biochemistry, Zhejiang University, Hangzhou, 310058, PR China;
authors.forename: Zhen
authors.initials: Z
authors.lastname: Wang
authors.affiliation: 2. Department of Neurosurgery, The First Affiliated Hospital of Nanjing Medical University, No. 300 Guangzhou Road, Nanjing 210029, China.
authors.forename: Rui
authors.initials: R
authors.lastname: Liu
authors.affiliation: 1. Institute of Biochemistry, Zhejiang University, Hangzhou, 310058, PR China;
authors.forename: Xingguo
authors.initials: X
authors.lastname: Gong
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology* / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects*; D001343:Autophagy / Q000187:drug effects*; D002018:Bufanolides / Q000494:pharmacology* / Q000627:therapeutic use; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D000091342:Endoplasmic Reticulum Chaperone BiP; D059865:Endoplasmic Reticulum Stress / Q000187:drug effects*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D005910:Glioma / Q000188:drug therapy / Q000473:pathology*; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008928:Mitochondria / Q000187:drug effects; D034622:RNA Interference; D015398:Signal Transduction / Q000187:drug effects; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism; D051742:Transcription Factor CHOP / Q000235:genetics / Q000378:metabolism
chemical_list: D000970:Antineoplastic Agents; D002018:Bufanolides; C497249:DDIT3 protein, human; D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; D006360:Heat-Shock Proteins; D051742:Transcription Factor CHOP; C546842:MTOR protein, human; D058570:TOR Serine-Threonine Kinases; C022777:bufalin
keywords: ER stress; apoptosis; autophagy; bufalin; glioma cancer.