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

Journal Article · Wed May 01 00:00:00 EDT 1991 · Laboratory Investigation (A Journal of Experimental Methods and Pathology); (USA)

OSTI ID:5639266

Oda, D; Lee, S P; Hayashi, A ^[1]

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1. Univ. of Washington, Seattle (USA)

We 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 10{sup 6} columnar epithelial cells, greater than 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 gamma-glutamyl transpeptidase and negative for glucose-6-phosphatase and albumin. Cells incorporated ({sup 3}H)uridine into intracellular proteins and ({sup 14}C)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.

• OSTI ID: 5639266
• Journal Information: Laboratory Investigation (A Journal of Experimental Methods and Pathology); (USA), Vol. 64:5; ISSN 0023-6837
• Country of Publication: United States
• Language: English

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59 BASIC BIOLOGICAL SCIENCES
ANIMAL CELLS
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CARBON 14 COMPOUNDS
CELL CULTURES
CELL CYCLE
CELL FLOW SYSTEMS
DNA
DOGS
EDTA
EPITHELIUM
GLUCOSAMINE
GLYCOPROTEINS
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TRYPSIN
URIDINE
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ANIMAL TISSUES
ANIMALS
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BODY
CARBOHYDRATES
CARBON COMPOUNDS
CARBOXYLIC ACIDS
CHELATING AGENTS
DIGESTIVE SYSTEM
ELECTRON MICROSCOPY
ENZYMES
HETEROCYCLIC COMPOUNDS
HEXOSAMINES
HEXOSES
HYDROGEN COMPOUNDS
HYDROLASES
HYDROXY COMPOUNDS
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
MAMMALS
MICROSCOPY
MONOSACCHARIDES
NUCLEIC ACIDS
NUCLEOSIDES
NUCLEOTIDES
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PEPTIDE HYDROLASES
PROTEINS
PYRIMIDINES
RIBOSIDES
SACCHARIDES
SCLEROPROTEINS
SERINE PROTEINASES
TISSUES
URACILS
VERTEBRATES
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