title: The time-dependent autophagy protects against apoptosis with possible involvement of Sirt1 protein in multiple myeloma under nutrient depletion.
abstract: The absence of researches about autophagy in multiple myeloma promoted us to explore the biological characteristics and role of autophagy induced by nutrient depletion in multiple myeloma (MM) cell line RPMI8226 cells. Both autophagic and apoptotic morphology were observed by TUNEL, transmission electron microscopy (TEM), and monodansylcadervarine (MDC) staining. MDC staining fluorescent intensity and the conversion of LC3-I to LC3-II demonstrated autophagy increased time-dependently at 6, 12, and 18 h then declined at 24 and 48 h. FCM analysis of Annexin V-FITC/PI (AV/PI) dual staining and TUNEL assay demonstrated apoptosis increased time-dependently at 6, 12, 18, 24, and 48 h, and it burst at 24 h when autophagy declined. Bax translocation from cytoplasm to mitochondria and active caspase3 protein level were promoted time dependently along with the increase of apoptosis. We demonstrated the same time-course pattern of Beclin1 protein and mRNA level as autophagy. The phosphorylating rate of S6K1 was inhibited at 6, 12, and 18 h when autophagy increased. Inhibition of autophagy by 3-methyladenine (3-MA) facilitated apoptosis and caspase3 activation at 6, 12, and 18 h but made no effect at 24 or 48 h. Sirt1 protein level in nuclei was gradually enhanced along with the increase of autophagy, however, 3-MA suppressed this enhancement. Taken together, nutrient depletion induced the time-dependent autophagy and apoptosis. Apoptosis was demonstrated as caspase3-mediated and Bax-dependent mitochondrial apoptosis. Autophagy appears to be Beclin1-dependent and related to mTORC1 inhibition. Autophagy protected against the apoptosis in the early period. Sirt1 possibly contributed to autophagy and was involved in pro-survival process mediated by autophagy.
authors.affiliation: Department of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China. zeng_rong_1983@126.com
authors.forename: Rong
authors.initials: R
authors.lastname: Zeng
authors.forename: Jing
authors.initials: J
authors.lastname: He
authors.forename: Jin
authors.initials: J
authors.lastname: Peng
authors.forename: Yan
authors.initials: Y
authors.lastname: Chen
authors.forename: Sha
authors.initials: S
authors.lastname: Yi
authors.forename: Fei
authors.initials: F
authors.lastname: Zhao
authors.forename: Guohui
authors.initials: G
authors.lastname: Cui
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D051017:Apoptosis Regulatory Proteins / Q000235:genetics / Q000378:metabolism; D001343:Autophagy / Q000502:physiology*; D000071186:Beclin-1; D053148:Caspase 3 / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor / Q000378:metabolism / Q000648:ultrastructure; D003470:Culture Media / Q000737:chemistry; D006801:Humans; D000076222:Mechanistic Target of Rapamycin Complex 1; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D009101:Multiple Myeloma / Q000503:physiopathology*; D046912:Multiprotein Complexes; D011506:Proteins / Q000235:genetics / Q000378:metabolism; D056564:Sirtuin 1 / Q000235:genetics / Q000378:metabolism*; D058570:TOR Serine-Threonine Kinases; D013997:Time Factors; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism
chemical_list: D051017:Apoptosis Regulatory Proteins; C491997:BECN1 protein, human; D000071186:Beclin-1; D003470:Culture Media; D008565:Membrane Proteins; D046912:Multiprotein Complexes; D011506:Proteins; D051028:bcl-2-Associated X Protein; D000076222:Mechanistic Target of Rapamycin Complex 1; D058570:TOR Serine-Threonine Kinases; D053148:Caspase 3; C447939:SIRT1 protein, human; D056564:Sirtuin 1