Document Detail


Immunomagnetic enrichment of interstitial cells of Cajal.
MedLine Citation:
PMID:  14563669     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Disruptions of networks of interstitial cells of Cajal (ICC), gastrointestinal pacemakers and mediators of neurotransmission, can lead to disordered phasic contractions and peristalsis by reducing and uncoupling electrical slow waves. However, detailed analysis of the ICC network behavior has been hampered by their scarcity, limited accessibility in intact tissues, and contamination with other cell types in culture. Our goal was to develop a simple technique to purify ICC from murine gastrointestinal muscles for functional studies. We identified ICC in live small intestinal muscles or primary cell cultures by Kit immunoreactivity using fluorescent antibodies. Because this technique also labels resident macrophages nonspecifically, parallel studies were performed in which nonfluorescent Kit antibodies and macrophages labeled with fluorescent dextran were used for subtractive analysis of ICC. In both groups, Kit-positive cells were tagged with superparamagnetic antibodies and sorted on magnetic columns. Efficacy was assessed by flow cytometry. ICC enrichment from primary cultures and freshly dissociated tissues was approximately 63-fold and approximately 8-fold, respectively. Unlike the cells derived directly from tissues, cells sorted from cultures frequently yielded extensive, nearly homogenous ICC networks on reseeding. Monitoring oscillations in mitochondrial Ca(2+) or membrane potential by imaging revealed spontaneous rhythmicity in these networks. Cells that did not bind to the columns yielded cultures that were depleted of ICC and dominated by smooth muscle cells. In conclusion, immunomagnetic sorting of primary cultures of ICC results in relatively homogenous, functional ICC networks. This technique is less suitable for obtaining ICC from freshly dispersed cells.
Authors:
Tamás Ordög; Doug Redelman; Nancy N Horowitz; Kenton M Sanders
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, P.H.S.     Date:  2003-10-16
Journal Detail:
Title:  American journal of physiology. Gastrointestinal and liver physiology     Volume:  286     ISSN:  0193-1857     ISO Abbreviation:  Am. J. Physiol. Gastrointest. Liver Physiol.     Publication Date:  2004 Feb 
Date Detail:
Created Date:  2004-01-12     Completed Date:  2004-03-03     Revised Date:  2014-09-14    
Medline Journal Info:
Nlm Unique ID:  100901227     Medline TA:  Am J Physiol Gastrointest Liver Physiol     Country:  United States    
Other Details:
Languages:  eng     Pagination:  G351-60     Citation Subset:  IM    
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MeSH Terms
Descriptor/Qualifier:
Animals
Calcium / metabolism
Cell Separation / methods*
Cells, Cultured
Electrophysiology
Fluorescent Antibody Technique
Gastrointestinal Tract / cytology*,  metabolism
Immunologic Techniques*
Macrophages / cytology
Magnetics*
Membrane Potentials
Mice
Mice, Inbred BALB C
Mitochondria, Muscle / metabolism,  physiology
Muscle, Smooth / cytology*,  metabolism
Oncogene Proteins / metabolism
Periodicity
Proto-Oncogene Proteins c-kit
Grant Support
ID/Acronym/Agency:
DK-58185/DK/NIDDK NIH HHS; P20-RR-16464/RR/NCRR NIH HHS; R01 DK058185/DK/NIDDK NIH HHS; R01 DK058185-02/DK/NIDDK NIH HHS
Chemical
Reg. No./Substance:
0/Oncogene Proteins; EC 2.7.10.1/Proto-Oncogene Proteins c-kit; SY7Q814VUP/Calcium

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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