Document Detail


Green fluorescent protein incorporation by mouse myoblasts may yield false evidence of myogenic differentiation of human haematopoietic stem cells.
MedLine Citation:
PMID:  18284377     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
AIMS: Haematopoietic CD34+ stem cells are able to differentiate into skeletal muscle, a potentially invaluable tool for treating degenerative diseases such as muscular dystrophy. However, some studies argue that the differentiative potential of these cells might have been overestimated. In vitro studies provide a controlled environment in which to investigate this point. METHODS: CD34+ stem cells from human peripheral blood, labelled with green fluorescent protein (GFP), were co-cultured with mouse myogenic C2C12 cells. The functional properties of mononucleated GFP+ cells were determined using electrophysiological techniques and were related to protein profiling determined by immunofluorescence staining and single-cell RT-PCR. Mouse mesoangioblasts co-cultured with human myotubes provided methodological controls. RESULTS: After 2-4 days, mononucleated adherent GFP+ cells showed acetylcholine-evoked current responses, typical of myogenic cells, as if stem cells had integrated into the host environment. In contrast to this hypothesis, human nuclei could not be detected in adherent GFP+ cells by immunofluorescence. Moreover, single-cell RT-PCR showed that adherent GFP+ cells responsive to acetylcholine expressed mouse markers while loose unresponsive GFP+ cells were of human origin. The transcripts of the human alpha1 subunit of the acetylcholine muscle receptor were not amplified in co-cultures. CONCLUSION: Single-cell analysis of functional properties combined with other markers revealed that, under the co-culture conditions used, GFP was transferred from human CD34+ stem cells to C2C12 myoblasts by mechanisms unrelated to myogenic stem cell differentiation. Our results emphasize the need for careful controls using several markers when investigating the myogenic differentiation of circulating stem cells.
Authors:
A Di Castro; D Bonci; M Musumeci; F Grassi
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2008-02-18
Journal Detail:
Title:  Acta physiologica (Oxford, England)     Volume:  193     ISSN:  1748-1716     ISO Abbreviation:  Acta Physiol (Oxf)     Publication Date:  2008 Jul 
Date Detail:
Created Date:  2008-06-05     Completed Date:  2008-12-04     Revised Date:  2009-02-03    
Medline Journal Info:
Nlm Unique ID:  101262545     Medline TA:  Acta Physiol (Oxf)     Country:  England    
Other Details:
Languages:  eng     Pagination:  249-56     Citation Subset:  IM    
Affiliation:
Department of Human Physiology and Pharmacology, Sapienza University, Rome, Italy.
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MeSH Terms
Descriptor/Qualifier:
Acetylcholine / pharmacology
Adult
Animals
Antigens, CD34 / blood
Artifacts
Cell Adhesion / physiology
Cell Differentiation / physiology
Cells, Cultured
Coculture Techniques
Green Fluorescent Proteins / genetics
Hematopoietic Stem Cells / cytology*,  drug effects,  physiology
Humans
Mice
Muscle Fibers, Skeletal / cytology*,  metabolism
Muscle, Skeletal / cytology
Myoblasts / cytology*,  drug effects,  metabolism,  physiology
Patch-Clamp Techniques
Transfection
Chemical
Reg. No./Substance:
0/Antigens, CD34; 147336-22-9/Green Fluorescent Proteins; 51-84-3/Acetylcholine

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