Characterization of new human gastric epithelial cell lines

• Author(s): Saraiva-Pava, Kathy ; Navabi, N. ; Lindén, Sarah ; Oleastro, Mónica ; Roxo-Rosa, Mónica
• Date: 2011
• Persistent ID: http://hdl.handle.net/10400.18/365
• Origin: Repositório Científico do Instituto Nacional de Saúde
• Subject(s): Helicobacter pylori; Gastric epithelial cell line; Infecções Gastrointestinais
• Document Type: Conference Object
• Language: English

Description

The lack of a cellular model which correctly mimics the natural niche of the pathogen Helicobacter pylori is still limitative for the study of this infection. Aiming to overcome this limitation, we have previously isolated clones of a subpopulation of the widely used heterogenic NCI-N87 (ATCC CRL-5822) gastric cell line1, those presenting typical epithelial markers and a progenitor-like phenotype (simultaneous synthesis of mucus and zymogens). For that, we stably-transduced the NCIN87 cells with human telomerase reverse-transcriptase (hTERT) catalytic subunit (pGRN145 plasmid, ATCC MBA-141), using the FuGENE-HD reagent (Roche). The two most promising NCI-N87-derived clones (C5 and C6) were shown to be composed of cells with homogenous phenotype with ability to grow in adherent monolayers, to produce gastric zymogens (hematoxylin staining) and to produce and secrete neutral mucins (Periodic-Acid-Schiff staining). Preliminary results have also shown that they are able to generate transepithelial electrical resistance and the ability of C5 to produce and secrete acidic mucins (Alcian-Blue staining). We are now clarifying the identity of the mucin species C5 and C6 produce by immunohistochemical analysis and zymogens (Pepsinogen) by western-blot. Moreover, the subcellular localization (immunocytochemistry) of adherens and tight-junctions’ proteins (E-cadherin and ZO-1) and the polarization status of both clones is now under evaluation. Due to their improved properties, compared to the heterogeneous parental line, these NCI-N87-derived clones are promising models of the human gastric epithelium.