File Download

There are no files associated with this item.

  Links for fulltext
     (May Require Subscription)
Supplementary

Article: Long-term culture and partial characterization of dog gallbladder epithelial cells

TitleLong-term culture and partial characterization of dog gallbladder epithelial cells
Authors
Issue Date1991
PublisherNature Publishing Group. The Journal's web site is located at http://www.nature.com/labinvest/
Citation
Laboratory Investigation, 1991, v. 64 n. 5, p. 682-692 How to Cite?
AbstractWe describe the successful isolation and maintenance of primary cultures of dog gallbladder epithelial cells. The surgically removed gallbladder was treated with trypsin/EDTA for 45 minutes and epithelial cells were collected and resuspended in Eagle's minimum essential medium with 10% fetal calf serum, and plated on Vitrogen-coated culture dishes. Each gallbladder yielded approximately 12 to 15 X 106 columnar epithelial cells, >95% of which were viable by trypan blue exclusion. In culture, cells maintained their polarity. They were arranged and grew in small and tight clusters that coalesced at confluency. When examined using transmission electron microscopy, prominent and numerous microville were identified on the apical portion of the plasma membrane. Cells were connected by well-formed desmosomes. Scanning electron microscopy revealed clusters of polyhedral cells with numerous papillary projections. Immunohistochemical studies demonstrated uniform staining of cells to keratin 35BH11 and AE1. Histochemical studies were positive for γ-glutamyl transpeptidase and negative for glucose-6-phosphatase and albumin. Cells incorporated [3H]uridine into intracellular proteins and [14C]glucosamine into tissue and secreted mucous glycoproteins linearly over 2 to 24 hours. Flow cytometry studies demonstrated a consistent and reproducible number of cells (10 to 12%) at S-phase. However, the number of cells at S-phase was dramatically reduced to almost negligible as cells reached confluency. This method of culturing primary dog gallbladder epithelial cells is highly reproducible and reliable. These cells preserve their state of differentiation, polarity, histochemical and immunohistochemical profile, morphologic, and metabolic integrity with repeated passaging or after being frozen. [3H]Thymidine uptake is well maintained, although doubling time shows a trend of decreased cell duplication with time. This technique offers the opportunity to study the electrophysiologic, metabolic, and immunologic properties of epithelial cells.
Persistent Identifierhttp://hdl.handle.net/10722/175657
ISSN
2015 Impact Factor: 4.202
2015 SCImago Journal Rankings: 2.133
ISI Accession Number ID

 

DC FieldValueLanguage
dc.contributor.authorOda, Den_US
dc.contributor.authorLee, SPen_US
dc.contributor.authorHayashi, Aen_US
dc.date.accessioned2012-11-26T09:00:18Z-
dc.date.available2012-11-26T09:00:18Z-
dc.date.issued1991en_US
dc.identifier.citationLaboratory Investigation, 1991, v. 64 n. 5, p. 682-692en_US
dc.identifier.issn0023-6837en_US
dc.identifier.urihttp://hdl.handle.net/10722/175657-
dc.description.abstractWe describe the successful isolation and maintenance of primary cultures of dog gallbladder epithelial cells. The surgically removed gallbladder was treated with trypsin/EDTA for 45 minutes and epithelial cells were collected and resuspended in Eagle's minimum essential medium with 10% fetal calf serum, and plated on Vitrogen-coated culture dishes. Each gallbladder yielded approximately 12 to 15 X 106 columnar epithelial cells, >95% of which were viable by trypan blue exclusion. In culture, cells maintained their polarity. They were arranged and grew in small and tight clusters that coalesced at confluency. When examined using transmission electron microscopy, prominent and numerous microville were identified on the apical portion of the plasma membrane. Cells were connected by well-formed desmosomes. Scanning electron microscopy revealed clusters of polyhedral cells with numerous papillary projections. Immunohistochemical studies demonstrated uniform staining of cells to keratin 35BH11 and AE1. Histochemical studies were positive for γ-glutamyl transpeptidase and negative for glucose-6-phosphatase and albumin. Cells incorporated [3H]uridine into intracellular proteins and [14C]glucosamine into tissue and secreted mucous glycoproteins linearly over 2 to 24 hours. Flow cytometry studies demonstrated a consistent and reproducible number of cells (10 to 12%) at S-phase. However, the number of cells at S-phase was dramatically reduced to almost negligible as cells reached confluency. This method of culturing primary dog gallbladder epithelial cells is highly reproducible and reliable. These cells preserve their state of differentiation, polarity, histochemical and immunohistochemical profile, morphologic, and metabolic integrity with repeated passaging or after being frozen. [3H]Thymidine uptake is well maintained, although doubling time shows a trend of decreased cell duplication with time. This technique offers the opportunity to study the electrophysiologic, metabolic, and immunologic properties of epithelial cells.en_US
dc.languageengen_US
dc.publisherNature Publishing Group. The Journal's web site is located at http://www.nature.com/labinvest/en_US
dc.relation.ispartofLaboratory Investigationen_US
dc.subject.meshAnimalsen_US
dc.subject.meshCell Cycleen_US
dc.subject.meshCell Divisionen_US
dc.subject.meshCell Separation - Methodsen_US
dc.subject.meshCells, Cultureden_US
dc.subject.meshDna - Metabolismen_US
dc.subject.meshDogsen_US
dc.subject.meshEdetic Acid - Diagnostic Useen_US
dc.subject.meshEpithelial Cellsen_US
dc.subject.meshEpithelium - Ultrastructureen_US
dc.subject.meshFemaleen_US
dc.subject.meshFlow Cytometryen_US
dc.subject.meshGallbladder - Cytology - Metabolism - Ultrastructureen_US
dc.subject.meshGlycoproteins - Metabolismen_US
dc.subject.meshImmunohistochemistry - Methodsen_US
dc.subject.meshKeratins - Metabolismen_US
dc.subject.meshMaleen_US
dc.subject.meshMicroscopy, Electronen_US
dc.subject.meshMicroscopy, Electron, Scanningen_US
dc.subject.meshTritium - Diagnostic Useen_US
dc.subject.meshTrypsin - Diagnostic Useen_US
dc.subject.meshUridine - Metabolismen_US
dc.subject.meshGamma-Glutamyltransferase - Metabolismen_US
dc.titleLong-term culture and partial characterization of dog gallbladder epithelial cellsen_US
dc.typeArticleen_US
dc.identifier.emailLee, SP: sumlee@hku.hken_US
dc.identifier.authorityLee, SP=rp01351en_US
dc.description.naturelink_to_subscribed_fulltexten_US
dc.identifier.pmid1709426-
dc.identifier.scopuseid_2-s2.0-0025781354en_US
dc.identifier.volume64en_US
dc.identifier.issue5en_US
dc.identifier.spage682en_US
dc.identifier.epage692en_US
dc.identifier.isiWOS:A1991FM28000011-
dc.publisher.placeUnited Kingdomen_US
dc.identifier.scopusauthoridOda, D=7006186359en_US
dc.identifier.scopusauthoridLee, SP=7601417497en_US
dc.identifier.scopusauthoridHayashi, A=14022812400en_US

Export via OAI-PMH Interface in XML Formats


OR


Export to Other Non-XML Formats