M1760 Sphingosine-1-Phosphate Promotes Paracellular Junctional Protein Expression and Intestinal Epithelial Cell Barrier Function

2009 ◽  
Vol 136 (5) ◽  
pp. A-906
Author(s):  
Ruiyun Li ◽  
Jose Greenspon ◽  
Rao N. Jaladanki ◽  
Jian-Ying Wang ◽  
Douglas J. Turner
Keyword(s):  
Epithelial Cell ◽  
Protein Expression ◽  
Barrier Function ◽  
Intestinal Epithelial Cell ◽  
Intestinal Epithelial ◽  
Sphingosine 1 Phosphate ◽  
Junctional Protein

scholarly journals Translocation of verotoxin-1 across T84 monolayers: mechanism of bacterial toxin penetration of epithelium

1997 ◽  
Vol 273 (6) ◽  
pp. G1349-G1358 ◽  
Author(s):  
Dana J. Philpott ◽  
Cameron A. Ackerley ◽  
Amanda J. Kiliaan ◽  
Mohamed A. Karmali ◽  
Mary H. Perdue ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Barrier Function ◽  
Pathogenic Bacteria ◽  
Intestinal Epithelial Cell ◽  
Cell Function ◽  
Bacterial Toxin ◽  
General Mechanism ◽  
Epithelial Cell Line ◽  
Intestinal Epithelial ◽  
T84 Cells

Verotoxin-producing Escherichia coli (VTEC) are pathogenic bacteria associated with diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome. Verotoxins (VTs) elaborated by these organisms produce cytopathic effects on a restricted number of cell types, including endothelial cells lining the microvasculature of the bowel and the kidney. Because human intestinal epithelial cells lack the globotriaosylceramide receptor for VT binding, it is unclear how the toxin moves across the intestinal mucosa to the systemic circulation. The aims of this study were to determine the effects of VT-1 on intestinal epithelial cell function and to characterize VT-1 translocation across monolayers of T84 cells, an intestinal epithelial cell line. VT-1 at concentrations up to 1 μg/ml had no effect on the barrier function of T84 monolayers as assessed by measuring transmonolayer electrical resistance (102 ± 8% of control monolayers). In contrast, both VT-positive and VT-negative VTEC bacterial strains lowered T84 transmonolayer resistance (45 ± 7 and 38 ± 6% of controls, respectively). Comparable amounts of toxin moved across monolayers of T84 cells, exhibiting high-resistance values, as monolayers with VTEC-induced decreases in barrier function, suggesting a transcellular mode of transport. Translocation of VT-1 across T84 monolayers paralleled the movement of a comparably sized protein, horseradish peroxidase. Immunoelectron microscopy confirmed transcellular transport of VT-1, since the toxin was observed within endosomes and associated with specific intracellular targets, including the Golgi network and endoplasmic reticulum. These data present a mode of VT-1 uptake by toxin-insensitive cells and suggest a general mechanism by which bacterial toxins lacking specific intestinal receptors can penetrate the intestinal epithelial barrier.

scholarly journals Different Sources of Copper Effect on Intestinal Epithelial Cell: Toxicity, Oxidative Stress, and Metabolism

Metabolites ◽  
2019 ◽  
Vol 10 (1) ◽  
pp. 11
Author(s):  
Runxian Li ◽  
Yang Wen ◽  
Gang Lin ◽  
Chengzhen Meng ◽  
Pingli He ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Gene Expression ◽  
Epithelial Cell ◽  
Protein Expression ◽  
Intestinal Epithelial Cell ◽  
Cell Damage ◽  
Intestinal Epithelial ◽  
Protein Expression Level ◽  
Treatment Groups ◽  
All Treatment

Copper (Cu) is widely used in the swine industry to improve the growth performance of pigs. However, high doses of copper will induce cell damage and toxicity. The aim of this study was to evaluate toxicity, bioavailability, and effects on metabolic processes of varying copper sources using porcine intestinal epithelial cells (IPEC-J2) as a model. The IPEC-J2 were treated with two doses (30 and 120 μM) of CuSO4, Cu Glycine (Cu-Gly), and Cu proteinate (Cu-Pro) for 10 h, respectively. Cell damage and cellular copper metabolism were measured by the changes in cell viability, copper uptake, oxidative stress biomarkers, and gene/protein expression levels. The results showed that cell viability and ratio of reduced and oxidized glutathione (GSH/GSSG) decreased significantly in all treatment groups; intracellular copper content increased significantly in all treatment groups; total superoxide dismutase (SOD) activity increased significantly in the 120 μM exposed groups; SOD1 protein expression levels were significantly upregulated in 30 μM Cu-Pro, 120 μM Cu-Gly, and 120 μM Cu-Pro treatment groups; intracellular reactive oxygen species (ROS) generation and malondialdehyde (MDA) content increased significantly in 30 μM treatment groups and 120 μM CuSO4 treatment group. CTR1 and ATP7A gene expression were significantly downregulated in the 120 μM exposed groups. While upregulation of ATOX1 expression was observed in the presence of 120 μM Cu-Gly and Cu-Pro. ASCT2 gene expression was significantly upregulated after 120 μM Cu-Glycine and CuSO4 exposure, and PepT1 gene expression was significantly upregulated after Cu-Pro exposure. In addition, CTR1 protein expression level decreased after 120 μM CuSO4 and Cu-Gly exposure. PepT1 protein expression level was only upregulated after 120 μM Cu-Pro exposure. These findings indicated that extra copper supplementation can induce intestinal epithelial cell injury, and different forms of copper may have differing effects on cell metabolism.

scholarly journals The PTEN Phosphatase Controls Intestinal Epithelial Cell Polarity and Barrier Function: Role in Colorectal Cancer Progression

PLoS ONE ◽  
2010 ◽  
Vol 5 (12) ◽  
pp. e15742 ◽  
Author(s):  
Marie-Josée Langlois ◽  
Sébastien Bergeron ◽  
Gérald Bernatchez ◽  
François Boudreau ◽  
Caroline Saucier ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Epithelial Cell ◽  
Cell Polarity ◽  
Cancer Progression ◽  
Barrier Function ◽  
Intestinal Epithelial Cell ◽  
Intestinal Epithelial ◽  
Epithelial Cell Polarity ◽  
Colorectal Cancer Progression

scholarly journals Gallic acid affects intestinal-epithelial-cell integrity and selected amino-acid uptake in porcine in vitro and ex vivo permeability models

2020 ◽  
pp. 1-9
Author(s):  
Marco Tretola ◽  
Giuseppe Bee ◽  
Paolo Silacci
Keyword(s):  
Amino Acid ◽  
Epithelial Cell ◽  
Protein Expression ◽  
Gallic Acid ◽  
Intestinal Epithelial Cell ◽  
Ex Vivo ◽  
Amino Acid Transporter ◽  
Intestinal Epithelial ◽  
Cell Integrity

Abstract Gallic acid (GA) is widely used as a dietary supplement due to several health-promoting effects, although its effects on intestinal-epithelial-cell integrity and transport remain mostly unknown. The present study aims to clarify the effects of GA on tight junctions and intestinal nutrient uptake through in vitro and ex vivo models. Both intestinal porcine enterocyte cell line-J2 cells and porcine middle-jejunum segments were treated with 5 (T5), 25 (T25) and 50 (T50) µm GA and mounted in Ussing chambers to determine transepithelial resistance (TEER), claudin-1 (CLDN1), occludin (OCLN), zonula occludens-1 (ZO-1) protein (in tissues and cells) and mRNA (in cells) expression. In addition, uptake of l-glutamate (l-Glut), l-arginine (l-Arg), l-lysine (l-Lys) and l-methionine (l-Meth) together with cationic-amino-acid transporter-1 (CAT-1) and excitatory-amino-acid transporter-3 (EAAT3) expression was evaluated. No apoptosis was observed in GA-treated cells, but TEER and CLDN1 protein abundance was lower with T50 compared with untreated cells. l-Arg and l-Lys uptake was greater with T5 than with T25 and T50. Ex vivo, T50 decreased the TEER values and the protein levels of CLDN1, OCLN and ZO-1, whereas T5 and T25 only decreased CLDN1 protein expression compared with untreated tissues. Moreover, T25 increased l-Glut and l-Arg uptake, the latter confirmed by an increased protein expression of CAT-1. GA influences intestinal uptake of the tested cationic amino acids at low concentrations and decreases the intestinal-cell barrier function at high concentrations. Similarities were observed between in vitro and ex vivo, but different treatment times and structures must be considered.

14 COLONIC EPITHELIAL CELL-SPECIFIC AFTIPHILIN KNOCKDOWN REGULATES EPITHELIAL BARRIER FUNCTION THROUGH MYOSIN LIGHT CHAIN-ASSOCIATED ACTIN ORGANIZATION IN VITRO AND INTESTINAL LENGTH IN VIVO

2020 ◽  
Vol 26 (Supplement_1) ◽  
pp. S28-S28
Author(s):  
Ivy Ka Man Law ◽  
Carl Rankin ◽  
Charalabos Pothoulakis
Keyword(s):  
Epithelial Cell ◽  
Barrier Function ◽  
Intestinal Epithelial Cell ◽  
Epithelial Barrier ◽  
Intestinal Epithelial ◽  
Knock Out ◽  
Epithelial Barrier Function ◽  
Actin Organization

Abstract Background and Aims Colonic epithelial integrity is often compromised during colonic inflammation and Inflammatory Bowel Disease. Aftiphilin (AFTPH) is a downstream target of microRNA-133a and its expression is reduced in colonic tissues of wild type mice from experimental colitis models and colonic biopsies from patients with ulcerative colitis. We have previously shown that AFTPH is involved in regulating intestinal epithelial barrier function and actin organization in human colonic epithelial cells in vitro (DDW 2016). On the other hand, our results suggested that global aftiphilin knock-out is embryonic lethal in mouse models (DDW 2019). Here, we further examined the role of AFTPH in regulating actin organization in vitro and characterize the colonic epithelial cell-specific aftiphilin knock-out mice. Methods Human colonic epithelial NCM460 cells were transfected with si-RNA against AFTPH to achieve transient AFTPH gene-silencing. Stable AFTPH knock-down clones were generated by transducing Caco2-BBE cells with recombinant lentivirus carrying sh-AFTPH or control sh-RNA. To create intestinal epithelial cell-specific aftiphilin knock-out mice, Aftph flox/flox mice were cross-bred with B6.Cg-Tg(Vil1-cre)997Gum/J mice, which express Villin-driven Cre recombinase (Vil-Cre), to generate intestinal epithelial cell-specific aftiphilin knock-out mice (Aftph Vil-/Vil-). Protein expression of F- and G-actin and p70S6K were detected using Western blot. Tissues from various organs were collected with Aftph Vil-/Vil- and its wildtype counterparts at 12 weeks. Results Results from western blot analysis showed that F-/G-actin ratio in AFTPH gene-silenced NCM460 cells were 0.6±0.17 fold, when compared to the treatment control. In addition, AFTPH gene-silencing in human colonic epithelial cells activated p70S6K, a kinase that is involved in actin organization, when compared to treatment control (1.2±0.15 vs. 2.0±0.15, p=0.0354). Furthermore, transepithelial electric resistance (TER) of Caco2-BBE cells deficient in AFTPH is significantly lower than that of control cells (0.5±0.07 fold). Lastly, in vivo intestinal epithelial cell-specific Aftph knock-out increased the length of small intestine, when compared to that of wild type mice (30.7±0.33 vs. 34.8±0.97, p=0.02), while the tissue weight of spleen to body weight was reduced (0.30±0.011 vs. 0.26±0.006, p=0.0169). Summary and Conclusions Our results indicate that AFTPH directly regulates epithelial barrier function and actin organization through mediating F-/G-actin ratio in human colonic epithelial cells, possibly through p70S6K. Importantly, intestinal epithelial cell-specific knock-out in vivo increased intestinal length and reduced size of the spleen. Our results suggested that AFTPH is crucial in regulating colonic epithelial barrier function in vitro and intestinal homeostasis.

Intestinal fibroblasts regulate intestinal epithelial cell proliferation via hepatocyte growth factor

1998 ◽  
Vol 274 (5) ◽  
pp. G809-G818 ◽  
Author(s):  
Michael Göke ◽  
Michiyuki Kanai ◽  
Daniel K. Podolsky
Keyword(s):  
Cell Proliferation ◽  
Growth Factor ◽  
Epithelial Cell ◽  
Hepatocyte Growth Factor ◽  
Protein Expression ◽  
Conditioned Medium ◽  
Intestinal Epithelial Cell ◽  
Intestinal Epithelial ◽  
Cell Counts ◽  
Hepatocyte Growth

Although the presence of subepithelial intestinal fibroblasts has been well recognized, the effects of fibroblasts on intestinal epithelial cell (IEC) growth are incompletely understood. In vitro studies were undertaken to evaluate the effects of fibroblasts on the proliferation of model IEC lines. IECs (Caco-2, T84, and IEC-6) were grown alone or in the presence of human intestinal (CCD-18), lung (CCD-37), or skin explant-derived fibroblasts. Cocultures were carried out directly on irradiated fibroblasts or by Transwell coculture technique with fibroblasts and epithelial cells separated by a porous filter. Cell proliferation was assessed by [3H]thymidine incorporation and cell counts. Hepatocyte growth factor (HGF) and c- met transcript expression in IECs and fibroblasts was examined by RT-PCR and Northern blotting; protein expression was evaluated by immunoblotting. Intestinal as well as lung and skin fibroblasts substantially stimulated proliferation of Caco-2, T84, and IEC-6 cells in both direct and Transwell cocultures. In addition, fibroblast-conditioned medium stimulated IEC proliferation, suggesting a paracrine mechanism. Anti-human HGF-neutralizing antibodies blocked the growth-promoting effects in both fibroblasts and fibroblast-conditioned medium. Recombinant human HGF dose dependently promoted IEC proliferation. HGF mRNA and protein expression was restricted to fibroblasts. High levels of c- met expression were found in Caco-2 and T84 cells; in contrast, expression in fibroblasts was weak. In summary, fibroblasts stimulate IEC proliferation through a paracrine mechanism mediated predominantly by HGF.

Sign in / Sign up

Export Citation Format

Share Document