title: Antagonism between apoptotic (Bax/Bcl-2) and anti-apoptotic (IAP) signals in human osteoblastic cells under vector-averaged gravity condition.
abstract: A functional disorder associated with weightlessness is well documented in osteoblasts. The apototic features of this disorder are poorly understood. Harmful stress induces apoptosis in cells via mitochondria and/or Fas. The Bax triggers cytochrome c release from mitochondria, which can be blocked by the Bcl-2. Released cytochrome c then activates the initiator caspase, caspase-9, which can be blocked by the anti-apototic (IAP) family of molecules. The effector caspase, caspase-3, finally exerts DNA fragmentation. We conducted this study to examine the apoptotic effects of vector-averaged gravity on normal human osteoblastic cells. Cell culture flasks were incubated on the clinostat, which generated vector-averaged gravity condition (simulated microgravity) for 12, 24, 48, and 96 hours. Upon termination of clinostat cultures, the cell number and cell viability were assessed. DNA fragmentation was analyzed on the agarose-gel electrophoresis. The mRNA levels for Bax, Bcl-2, XIAP, and caspase-3 genes were analyzed by semi-quantitative RT-PCR. Twenty-four hours after starting clinostat rotation, the ratios of Bax/Bcl-2 mRNA levels (indicator of apoptosis) were significantly increased to 136% of the 1G static controls. However, the XIAP mRNA levels (anti-apoptotic molecule) were increased concomitantly to 138% of the 1G static controls. Thus, cell proliferation or cell viability was not affected by vector-averaged gravity. DNA fragmentation was not observed in clinostat group as well as in control group. Finally, the caspase-3 mRNA levels were not affected by vector-averaged gravity. Simulated microgravity might modulate some apoptotic signals upstream the mitochondrial pathway.
authors.affiliation: Section of Orthopedic Spinal Surgery, Department of Frontier Surgical Therapeutics, Division of Advanced Therapeutical Sciences, Graduate School of Tokyo Medical and Dental University, Tokyo 113-8519, Japan. nakamura.orth@tmd.ac.jp
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Nakamura
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Kumei
authors.forename: Sadao
authors.initials: S
authors.lastname: Morita
authors.forename: Hitoyata
authors.initials: H
authors.lastname: Shimokawa
authors.forename: Keiichi
authors.initials: K
authors.lastname: Ohya
authors.forename: Kenichi
authors.initials: K
authors.lastname: Shinomiya
mesh_terms: D000328:Adult; D017209:Apoptosis / Q000502:physiology*; D002455:Cell Division / Q000502:physiology; D002470:Cell Survival / Q000502:physiology; D002478:Cells, Cultured; D005786:Gene Expression Regulation / Q000502:physiology*; D006112:Gravitation; D006801:Humans; D007700:Kinetics; D008297:Male; D010006:Osteoblasts / Q000166:cytology* / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051028:bcl-2-Associated X Protein
chemical_list: C494826:BAX protein, human; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D051028:bcl-2-Associated X Protein