Simulated Microgravity Disrupts Cytoskeleton Organization and Increases Apoptosis of Rat Neural Crest Stem Cells Via Upregulating CXCR4 Expression and RhoA-ROCK1-p38 MAPK-p53 Signaling.
Neural crest stem cells (NCSCs) are a population of multipotent stem cells that are distributed broadly in many tissues and organs and are capable of differentiating into a variety of cell types that are dispersed throughout three germ layers. We are interested in studying the effects of simulated microgravity on the survival and self-renewal of NCSCs. NCSCs extracted from the hair follicle bulge region of the rat whisker pad were cultured in vitro, respectively, in a 2D adherent environment and a 3D suspension environment using the rotatory cell culture system (RCCS) to simulate microgravity. We found that rat NCSCs (rNCSCs) cultured in the RCCS for 24 h showed disrupted organization of filamentous actin, increased globular actin level, formation of plasma membrane blebbing and neurite-like artifact, as well as decreased levels of cortactin and vimentin. Interestingly, ∼70% of RCCS-cultured rNCSCs co-expressed cleaved (active) caspase-3 and neuronal markers microtubule-associated protein 2 (MAP2) and Tuj1 instead of NCSC markers, suggesting stress-induced formation of neurite-like artifact in rNCSCs. In addition, rNCSCs showed increased C-X-C chemokine receptor 4 (CXCR4) expression, RhoA GTPase activation, Rho-associated kinase 1 (ROCK1) and p38 mitogen-activated protein kinase (MAPK) phosphorylation, and p53 expression in the nucleus. Incubation of rNCSCs with the Gα protein inhibitor pertussis toxin or CXCR4 siRNA during RCCS-culturing prevented cytoskeleton disorganization and plasma membrane blebbing, and it suppressed apoptosis of rNCSCs. Taken together, we demonstrate for the first time that simulated microgravity disrupts cytoskeleton organization and increases apoptosis of rNCSCs via upregulating CXCR4 expression and the RhoA-ROCK1-p38 MAPK-p53 signaling pathway.
1 Graduate Institute of Aerospace and Undersea Medicine, National Defense Medical Center , Neihu District, Taipei City, Taiwan .
Shing-Chen
SC
Lin
2 Graduate Institute of Medical Sciences, National Defense Medical Center , Neihu District, Taipei City, Taiwan .
Guo-Hau
GH
Gou
2 Graduate Institute of Medical Sciences, National Defense Medical Center , Neihu District, Taipei City, Taiwan .
Ching-Wu
CW
Hsia
1 Graduate Institute of Aerospace and Undersea Medicine, National Defense Medical Center , Neihu District, Taipei City, Taiwan .|3 Division of Rehabilitation Medicine, Taoyuan Armed Forces General Hospital , Longtan Township, Taoyuan County, Taiwan .
Cheng-Wen
CW
Ho
1 Graduate Institute of Aerospace and Undersea Medicine, National Defense Medical Center , Neihu District, Taipei City, Taiwan .|4 Department of Undersea and Hyperbaric Medicine, Tri-Service General Hospital , Neihu District, Taipei City, Taiwan .
Kun-Lun
KL
Huang
5 Department of Medical Research, Tri-Service General Hospital , Neihu District, Taipei City, Taiwan .
Yung-Fu
YF
Wu
1 Graduate Institute of Aerospace and Undersea Medicine, National Defense Medical Center , Neihu District, Taipei City, Taiwan .
Shih-Yu
SY
Lee
1 Graduate Institute of Aerospace and Undersea Medicine, National Defense Medical Center , Neihu District, Taipei City, Taiwan .
Yi-Hui
YH
Chen
D000818:Animals; D017209:Apoptosis; D015415:Biomarkers / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism; D002448:Cell Adhesion; D002462:Cell Membrane / Q000378:metabolism; D049109:Cell Proliferation; D048430:Cell Shape; D002470:Cell Survival; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000378:metabolism*; D005260:Female; D009432:Neural Crest / Q000166:cytology*; D058953:Neural Stem Cells / Q000166:cytology* / Q000378:metabolism; D010766:Phosphorylation; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D017207:Rats, Sprague-Dawley; D019718:Receptors, CXCR4 / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D015854:Up-Regulation; D018474:Weightlessness Simulation; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism; D054460:rho-Associated Kinases / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000378:metabolism
D015415:Biomarkers; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D019718:Receptors, CXCR4; D016159:Tumor Suppressor Protein p53; D054460:rho-Associated Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D053148:Caspase 3; D020742:rhoA GTP-Binding Protein