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Unique kinetics of Oct3/4 microlocalization following dissociation of human embryonic stem cell colonies.
MedLine Citation:
PMID:  22727933     Owner:  NLM     Status:  Publisher    
Abstract/OtherAbstract:
To investigate the effects of the Rho-dependent protein kinase (ROCK) inhibitor Y-27632 on the kinetics of E-cadherin, F-actin, and Oct3/4 distributions in dissociated human embryonic stem (hES) cells and to analyze their interactions morphologically, Y-27632-treated [R(i) (+)] and untreated [R(i) (-)] cells were immunohistochemically stained for E-cadherin and Oct3/4 within 24h of dissociation and also for F-actin. Furthermore, the gene expression of E-cadherin, Oct3/4, and RhoA was confirmed by quantitative real-time RT-PCR. E-cadherin expression intensified linearly along the membranes of R(i) (+) cells or intercellular junctions in cell clusters. F-actin accumulated along the periphery of cells and expanded in a web-like manner along junctions in cell clusters, and Oct3/4 was restricted to the nucleus within few hours of dissociation. However, R(i) (-) cells exhibited deformation and blebbing and appeared to die over time. E-cadherin exhibited a punctate pattern along the periphery, after which it accumulated on one or both sides of the cytoplasm. Actin filaments were concentrated at the bleb bases. Oct3/4 was detected in the cytoplasm, not in the nucleus the recovery of integrated E-cadherin distribution. Quantitative real-time RT-PCR revealed RhoA upregulation and E-cadherin downregulation at 12h after dissociation. Oct3/4 gene expression was unaffected by ROCK inhibition. These results revealed that the cooperative nature of hES cells is maintained by the E-cadherin-actin cytoskeleton system along with the restricted distribution of Oct3/4 in the nucleus. RhoA activation followed by dissociation disorders this system and accelerates cell death, which is partially suppressed by ROCK inhibition.
Authors:
Hinako Ichikawa; Yoshiya Kanoh; Sakiko Shirasawa; Tadayuki Yokoyama; Fengming Yue; Daihachiro Tomotsune; Katsunori Sasaki
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Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2012-6-5
Journal Detail:
Title:  Annals of anatomy = Anatomischer Anzeiger : official organ of the Anatomische Gesellschaft     Volume:  -     ISSN:  1618-0402     ISO Abbreviation:  -     Publication Date:  2012 Jun 
Date Detail:
Created Date:  2012-6-25     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  100963897     Medline TA:  Ann Anat     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
Copyright Information:
Copyright © 2012 Elsevier GmbH. All rights reserved.
Affiliation:
Department of Histology and Embryology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano 390-8621, Japan.
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