A human colonic tumor cell line that maintains vectorial electrolyte transport

Dharmsathaphorn, Kiertisin; McRoberts, James A.; Mandel, Kenneth G.; Tisdale, L. D.; Masui, Hideo

doi:10.1152/ajpgi.1984.246.2.g204

Cited by 282 publications

(238 citation statements)

References 16 publications

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“…The human intestinal epithelial T84 colon carcinoma-derived cell line was used in these experiments (6). Cells of the Caco-2 colon carcinoma cell line were grown in high glucose Dulbecco-Vogt modified Eagle's medium (DMEM) supplemented with 10% fetal calf serum.…”

Section: Methodsmentioning

confidence: 99%

Comparative Analysis of EspF from Enteropathogenic and Enterohemorrhagic Escherichia coli in Alteration of Epithelial Barrier Function

et al. 2004

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Enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli (EHEC) are related intestinal pathogens that harbor highly similar pathogenicity islands known as the locus of enterocyte effacement (LEE). Despite their genetic similarity, these two pathogens disrupt epithelial tight junction barrier function with distinct kinetics. EHEC-induced reduction in transepithelial electrical resistance (TER), a measure of barrier function disruption, is significantly slower and more modest in comparison to that induced by EPEC. The variation in bacterial adherence only partially accounted for these differences. The LEE-encoded effector protein EspF has been shown to be critical for EPEC-induced alterations in TER. EspF from both EPEC and EHEC is expressed and secreted upon growth in tissue culture medium. The mutation of EHEC cesF suggested that the optimal expression and secretion of EHEC EspF required its chaperone CesF, as has been shown for EPEC. In contrast to EPEC espF and cesF, mutation of the corresponding EHEC homologs did not dramatically alter the decrease in TER. These differences could possibly be explained by the presence of additional espF-like sequences (designated U-and M-espF, where the letter designations refer to the specific cryptic prophage sequences on the EHEC chromosome closest to the respective genes) in EHEC. Reverse transcription-PCR analyses revealed coordinate regulation of EHEC U-espF and the LEE-encoded espF, with enhanced expression in bacteria grown in Dulbecco-Vogt modified Eagle's medium compared to bacteria grown in Luria broth. Both EHEC espF and U-espF complemented an EPEC espF deletion strain for barrier function alteration. The overexpression of U-espF, but not espF, in wild-type EHEC potentiated the TER response. These studies reveal further similarities and differences in the pathogenesis of EPEC and EHEC.

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Section: Methodsmentioning

confidence: 99%

Comparative Analysis of EspF from Enteropathogenic and Enterohemorrhagic Escherichia coli in Alteration of Epithelial Barrier Function

et al. 2004

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“…Calcium antagonists (e.g., verapamil), calmodulin inhibitors (e.g., trifluoperazine) and other agents (e.g., reserpine) inhibit the active drug efflux and restore drug sensitivity in multidrug resistant cells (Tsuruo, 1983;Kessel & Wilberding, 1984;Kessel & Wilberding, 1985;Bellamy et al, 1988;Twentyman et al, 1986). (Madara et al, 1987;Chan et al, 1989 Dharmsathaphorn et al (1984) and Simmons (1990). Cells were seeded at high density (1 x 106 cells ml-', 3 ml well) into rat-tail collagen coated filtercups, Transwell 24.5 mm diam.…”

mentioning

confidence: 99%

Epithelial secretion of vinblastine by human intestinal adenocarcinoma cell (HCT-8 and T84) layers expressing P-glycoprotein

Hunter

Hirst²,

Simmons³

1991

Br J Cancer

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Summary P-glycoprotein expression was demonstrated in two human intestinal adenocarcinoma cell-lines ileocaecal and T84, colonic) by immunoprecipitation of a 170-180 kDa protein with monoclonal antibody JSB-1. Both HCT-8 and T84 formed functional epithelial cell layers of high transepithelial electrical resistance (>700 Q cm2) when grown on permeable matrices. These epithelial layers demonstrated vectorial secretion (net vinblastine fluxes in the basal-to-apical direction of 0.135 and 0.452 pmol h-' cm-2 in HCT-8 and T84 cell layers, respectively, from bathing solutions containing 10 nM vinblastine). These vectorial vinblastine secretions were sensitive to inhibition by verapamil. Passive transepithelial vinblastine permeation was limited by the presence of intercellular (tight) junctions, as demonstrated by the high transepithelial electrical resistance, and verapamil increased this passive vinblastine permeation concomitant with a reduction in the electrical resistance. Cellular vinblastine loading was significantly greater from the basal side, and this was also susceptible to inhibition by basal verapamil. The demonstration of vectorial transport of vinblastine in human intestinal colonic adenocarcinoma cell layers is direct evidence in favour of the hypothesis that the function of mdrl in epithelia from the gastrointestinal tract is to promote detoxification by a process of epithelial secretion. This study also highlights that cellular vinblastine accumulation depends not only upon P-glycoprotein function, but also upon differential apparent membrane permeabilities and the presence of intercellular (tight) junctions that may restrict drug permeation and cellular accumulation to apical or basal membrane domains.

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“…T84 cells reproduce the cAMP-and Ca 2+ -regulated Cl -secretion observed in native intestinal tissue [13] and increase Cl -secretion in response to cholera toxin, a cAMP agonist. We tested whether secramine B (Fig.…”

Section: Secramine B Inhibits Camp-induced CL -Secretion From Intestimentioning

confidence: 78%

The Cdc42 inhibitor secramine B prevents cAMP-induced K+ conductance in intestinal epithelial cells

Pelish

Ciesla

Tanaka

et al. 2006

Biochemical Pharmacology

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Cyclic AMP-(cAMP) and calcium-dependent agonists stimulate chloride secretion through the coordinated activation of distinct apical and basolateral membrane channels and ion transporters in mucosal epithelial cells. Defects in the regulation of Cl -transport across mucosal surfaces occur with cystic fibrosis and V. cholerae infection and can be life threatening. Here we report that secramine B, a small molecule that inhibits activation of the Rho GTPase Cdc42, reduced cAMPstimulated chloride secretion in the human intestinal cell line T84. Secramine B interfered with a cAMP-gated and Ba 2+ -sensitive K + channel, presumably KCNQ1/KCNE3. This channel is required to maintain the membrane potential that sustains chloride secretion. In contrast, secramine B did not affect the Ca 2+ -mediated chloride secretion pathway, which requires a separate K + channel activity from that of cAMP. Pirl1, another small molecule structurally unrelated to secramine B that also inhibits Cdc42 activation in vitro, similarly inhibited cAMPdependent but not Ca 2+ -dependent chloride secretion. These results suggest that Rho GTPases may be involved in the regulation of the chloride secretory response and identify secramine B an inhibitor of cAMP-dependent K + conductance in intestinal epithelial cells.

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A human colonic tumor cell line that maintains vectorial electrolyte transport

Cited by 282 publications

References 16 publications

Comparative Analysis of EspF from Enteropathogenic and Enterohemorrhagic Escherichia coli in Alteration of Epithelial Barrier Function

Comparative Analysis of EspF from Enteropathogenic and Enterohemorrhagic Escherichia coli in Alteration of Epithelial Barrier Function

Epithelial secretion of vinblastine by human intestinal adenocarcinoma cell (HCT-8 and T84) layers expressing P-glycoprotein

The Cdc42 inhibitor secramine B prevents cAMP-induced K+ conductance in intestinal epithelial cells

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