title: Assembly and functional analysis of tight junction in a colon adenocarcinoma cell line: effect of glucose depletion.
abstract: We describe morphologic and biochemical changes in the colonic epithelial HCT-116 cell line following depletion of glucose from the culture medium. Cultured cells under permissive differentiation conditions (inosine-supplemented glucose-free medium) exhibited, after confluence, an enterocytic differentiation, in contrast to cells grown under standard culture conditions, where they remain in an undifferentiated state. The differentiated phenotype was characterized by the presence of a monolayer of polarized cells displaying an apical tight junction, and by the presence of alkaline phosphatase, a well known brush border marker. We demonstrated that the formed tight junctions were functional using the following criteria: a) labeling of the junctions with antibodies recognizing the tight juntion proteins occludin and ZO-1, as observed by immunofluorescence and immunoblotting analysis; b) characteristic organization of the tight junction strands, as observed in freeze-fracture replicas; c) increase ofthe transepithelial resistance across the monolayer; d) not permeation of the ruthenium red stain across the tight junction, and e) presence of the hyperphosphorylated form of occludin.
authors.affiliation: Divisão de Biologia Celular, Coordenação de Pesquisa, Instiuto Nacional de Câncer, Praça da Cruz Vermelha, 23, 6o Andar, 20230 130, Rio de Janeiro, Brazil.
authors.forename: R
authors.initials: R
authors.lastname: Ancillotti
authors.forename: L
authors.initials: L
authors.lastname: Goto-Silva
authors.forename: G
authors.initials: G
authors.lastname: Montesano
authors.forename: S
authors.initials: S
authors.lastname: Oliveira
authors.forename: P A
authors.initials: PA
authors.lastname: Redondo
authors.forename: L A
authors.initials: LA
authors.lastname: Barbosa
authors.forename: W
authors.initials: W
authors.lastname: De Souza
authors.forename: J A
authors.initials: JA
authors.lastname: Morgado-Díaz
mesh_terms: D000230:Adenocarcinoma / Q000378:metabolism* / Q000648:ultrastructure; D003110:Colonic Neoplasms / Q000378:metabolism* / Q000648:ultrastructure; D005947:Glucose / Q000172:deficiency*; D006801:Humans; D008854:Microscopy, Electron; D019108:Tight Junctions / Q000378:metabolism* / Q000648:ultrastructure
chemical_list: D005947:Glucose