Isolation, identification and quantitative evaluation of specific cell types from the mammalian gastric mucosa

Abstract

Functional in vitro studies with isolated gastric mucosal cells require cytological identification of different cell types in suspension or primary culture. Since suitable techniques have not been well established, different staining methods for the discrimination of dispersed pig and guinea pig gastric cells have been developed on the basis of modified previous protocols for enzymatic cell dispersion. Chief and parietal cells were visualized by combined periodic acid-Schiff stains. Surface mucous and mucous neck cells were identified by affinity-labelling, using lectins with selective staining properties in situ. Two of the lectins were found to be specific markers for gastric polymorphonuclear cells. The following vital tests were found to be useful: succinic dehydrogenase for parietal cells, Nile blue/brilliant cresyl blue stains for chief cells, and different phagocytosis assays for endothelial cells and gastric phagocytes. Endocrine cells were characterized by immunocytochemistry using specific antibodies against gastrin, somatostatin, histamine and serotonin. The same technique using a vimentin antibody was performed for the identification of fibroblasts. Proliferation of mucosal cells in primary culture was monitored by the incorporation of bromo-deoxyuridine, which was subsequently detected by a monoclonal antibody.

A molecular profile of the mouse gastric parietal cell with and without exposure to Helicobacter pylori

“…Cells from PC ϩ and PC Ϫ preparations were smeared onto glass slides, fixed in 70% ethanol, rehydrated in PBS, and stained using a modified Maxwell technique (10) to score the fractional representation of parietal, zymogenic, and mucus-producing cells.…”

Phospholipid Synthesis in Isolated Porcine Gastric Mucous Cells

“…Different cell types were identified by PAS/Alcian blue staining [9], The mucous cell fraction had a 90-95% purity, with parietal cells as the main contami nants. The ratio between neutral mucous cells and mucous neck cells was 60:40.…”

“…Isolated gastric mucous cells from porcine stom achs were prepared by counterflow elutriation accord ing to an updated version of the method of Beinbom et al [9], In brief, fresh pig stomachs were provided by the local slaughterhouse. Fundic mucosa was carefully separated from muscle and connective tissue, rinsed and transported to the laboratory in buffer I .…”

Recovery of Cholecystokinin Response of Porcine Gastric Chief Cells during Monolayer Culture

“…Fraction F3, which consisted mainly of chief cells (74.4 ± 2.9%. n = 9) and w'as practically devoid of parietal cells as described previously [22], was used for the further experiments. Chief cells were detected by staining with brillant cresyl blue [22], Dead cells were detected by the trypan blue exclusion test [22] and amounted to 4.0 ± 0.7% (n = 9).…”

“…Cells were prepared by a modified method of Beinborn et al [22], The following buffers were used: buffer A consisted of (mmol/1): NaCl 104: NaHCO} 4.5: KCI 5; NaH: P 0 4().5: N a;H P 0 4 I; N-(2-hydroxycthyl)piperazine-N'-2-ethanesu!fonic acid (HEPES)25; glu cose 11.1: indometacin 0.01: penicillin G 100 U/ml, and streptomy cin 100 pg/ml. Buffer B had the same composition but contained additionally (mmol/1): CaCI; 1; MgCI; 1.5, and bovine serum albu min 2 mg/ml.…”