A monolayer culture of human gastric epithelial cells

Abstract

Our aim was to develop a fibroblast-free monolayer culture of human gastric mucosal cells, using the specimens obtained by routine endoscopic biopsy. Human gastric mucosa obtained from normal volunteers by endoscopic biopsy was dissociated from collagenase and hyaluronidase. Dissociated cells were cultured in supplemented Coon's modified Ham's F-12 medium. Within 24 hr of inoculation, the cells were attached to the culture dishes. This was followed by cellular outgrowth. On phase-contrast microscopy, all cells had epithelial characteristics and fibroblasts were not observed. Ninety percent of cells contained periodic acid Schiff reaction-positive mucous granules after diastase digestion consistent with mucous epithelial cells. Two percent of the cells gave a strong reaction for succinic dehydrogenase activity (parietal cells). Immunohistochemical staining for pepsinogen in cultured cells was negative. On EM, microvilli-like projections, junctional complexes, Golgi apparatus, and mucous granules were apparent in the majority of cells. Mitotic figures were observed by day 3 with Giemsa staining. Autoradiographically, these cells were able to incorporate [3H]TdR into the nuclei. Cells were capable of synthesizing DNA, and this function was inhibited by cycloheximide. Cells could be cultured for up to two weeks without fibroblast contamination. A method of primary monolayer culture of human gastric mucosa obtained by a routine endoscopic biopsy has been successfully developed.

Transformed immortalized gastric epithelial cells by virulence factor CagA of Helicobacter pylori through Erk mitogen-activated protein kinase pathway

“…Using carcinogenic agents to induce the transformation of normal cells has been a routine method for the study of tumors in vitro (Male et al, 1987;Narayan et al, 2004). The isolation of normal, primary gastric epithelial cells, however, is not suitable for the study of carcinogenesis mechanisms as these cells rapidly enter into a senescence phase (Terano et al, 1983;Rutten et al, 1996;Smoot et al, 2000). In our previous study (Zhu et al, 2004a), we utilized telomerase reverse transcriptase (hTERT) and simian virus 40 (SV40) large T antigen to establish successfully a conditionally immortalized, normal human gastric epithelial cell line (NEC), which also expressed c-src/ Lyn kinase.…”

Protective role of intracellular superoxide dismutase against extracellular oxidants in cultured rat gastric cells.

“…Histochemical study with succinate dehydrogenase activity identified 5% of cells as parietal cells, whereas immunohistochemical study with pepsinogen failed to reveal any chief cells (25). These cells possessed the capability to synthesize DNA as well as cyclic nucleotides (25,36), to produce and secrete mucous glycoprotein (25), and to generate prostaglandins (36).…”

“…The cells reached confluency on day 3. Over 90% ofthe cells were identified as mucous-producing epithelial cells (surface epithelial cells and mucous neck cells) by PAS staining and ultrastructural studies (25,35). Histochemical study with succinate dehydrogenase activity identified 5% of cells as parietal cells, whereas immunohistochemical study with pepsinogen failed to reveal any chief cells (25).…”

“…Primary culture of the gastric fundic mucosa from 7-10-d old rats was prepared by the method of Terano et al (25). In brief, the intact corpus (oxyntic glandular mucosa) area was excised from the antrum and forestomach, and the full wall thickness was minced into 2-to 3-mm3 pieces.…”

Acetaminophen Directly Protects Human Gastric Epithelial Cell Monolayers against Damage Induced by Sodium Taurocholate

“…have been successfully used to study cyto protection in vitro [2,3], Monolayer cultures of gastric mucous cells have also been pre pared from humans, either from biopsy or surgical specimens [4][5][6] but, to our knowl edge, they have not been used to study drug damage and protection of gastric epithelial cells in vitro.…”