scholarly journals The function of macromolecular complex of CFTR-NHERF2-LPA2 in inflammatory responses of intestinal epithelial cells

2017 ◽  
Author(s):  
Shanshan Kong ◽  
Weiqiang Zhang
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
Cell Line ◽  
Human Cell ◽  
Inflammatory Responses ◽  
Intestinal Epithelial Cells ◽  
Human Cell Line ◽  
Macromolecular Complex ◽  
Apical Surface ◽  
Intestinal Epithelial

AbstractCFTR is a cAMP-regulated chloride channel located in the apical surface of intestinal epithelial cells; where it forms a macromolecular complex with NHERF2 and LPA2. CFTR has been shown to play a role in the pathogenies of several types of secretory diarrheas. Inflammatory bowel disease (IBD) is a chronic condition of intestine characterized by severe inflammation and mucosal destruction, genetic analysis has shown that LPA contribute to IBD and patients of cystic fibrosis also display the phenotype of diarrhea. The purpose of this study is to investigate if this complex plays a role in the inflammatory responses of intestinal epithelium.We then explored the role of this complex in maintaining the integrity of tight junction and inflammatory responses in these cells. In vitro assays show that inhibiting CFTR or LPA2 in the intestinal epithelial cell could disrupt the epithelial cell junction, and reduce the TER of intestinal epithelial cells in both mouse and human cell line. EƯSA assay show that intriguing LPA2 through LPS or LPA can increase the secretion of IL-8, while inhibiting or SiRNA knockdown of LPA2 can decrease the secretion of IL-8 in mouse or human intestinal epithelial cells. The CFTR inhibitor can reduce the IL-8 secretion in both mouse and human cell line, the deletion of CFTR in mouse intestine does not affect the IL-8 level, but the knockdown of CFTR in human cell line reduced the IL-8 protein level. The deletion of CFTR in human also reduced the IL-8 mRNA level. This indicates the CFTR-LPA complex is necessary for the expression of IL-8.

Studying Permeability in a Commonly Used Epithelial Cell Line: T84 Intestinal Epithelial Cells

2011 ◽  
pp. 115-137 ◽  
Author(s):  
Rino P. Donato ◽  
Adaweyah El-Merhibi ◽  
Batjargal Gundsambuu ◽  
Kai Yan Mak ◽  
Emma R. Formosa ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
Cell Line ◽  
Intestinal Epithelial Cells ◽  
Epithelial Cell Line ◽  
Intestinal Epithelial

Gene expression of activin, activin receptors, and follistatin in intestinal epithelial cells

2000 ◽  
Vol 278 (1) ◽  
pp. G89-G97 ◽  
Author(s):  
Kei Sonoyama ◽  
Suriya Rutatip ◽  
Takanori Kasai
Keyword(s):  
Gene Expression ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Cell Line ◽  
Intestinal Epithelial Cells ◽  
Activin A ◽  
Intestinal Cell ◽  
Rat Jejunum ◽  
Intestinal Epithelial ◽  
Activin Receptors

Gene expression of activin, activin receptors, and follistatin was investigated in vivo and in vitro using semiquantitative RT-PCR in intestinal epithelial cells. Rat jejunum and the intestinal epithelial cell line IEC-6 expressed mRNA encoding the βA-subunit of activin, α-subunit of inhibin, activin receptors IB and IIA, and follistatin. An epithelial cell isolation study focused along the crypt-villus axis in rat jejunum showed that βA mRNA levels were eight- to tenfold higher in villus cells than in crypt cells. Immunohistochemistry revealed the expression of activin A in upper villus cells. The human intestinal cell line Caco-2 was used as a differentiation model of enterocytes. Four- to fivefold induction of βA mRNA was observed in postconfluent Caco-2 cells grown on filter but not in those cells grown on plastic. In contrast, follistatin mRNA was seen to be reduced after reaching confluence. Exogenous activin A dose-dependently suppressed the proliferation and stimulated the expression of apolipoprotein A-IV gene, a differentiation marker, in IEC-6 cells. These results suggest that the activin system is involved in the regulation of such cellular functions as proliferation and differentiation in intestinal epithelial cells.

scholarly journals Infection of Polarized Cultures of Human Intestinal Epithelial Cells with Hepatitis A Virus: Vectorial Release of Progeny Virions through Apical Cellular Membranes

2000 ◽  
Vol 74 (14) ◽  
pp. 6476-6484 ◽  
Author(s):  
Christian A. Blank ◽  
David A. Anderson ◽  
Michael Beard ◽  
Stanley M. Lemon
Keyword(s):  
Epithelial Cells ◽  
Hepatitis A ◽  
Cytopathic Effect ◽  
Hepatitis A Virus ◽  
Intestinal Epithelial Cells ◽  
Human Cell Line ◽  
Progeny Virus ◽  
Apical Surface ◽  
Intestinal Epithelial ◽  
Virus Release

ABSTRACT Although hepatitis A virus (HAV) is typically transmitted by the fecal-oral route, little is known of its interactions with cells of the gastrointestinal tract. We studied the replication of HAV in polarized cultures of Caco-2 cells, a human cell line which retains many differentiated functions of small intestinal epithelial cells. Virus uptake was 30- to 40-fold more efficient when the inoculum was placed on the apical rather than the basolateral surface of these cells, suggesting a greater abundance of the cellular receptor for HAV on the apical surface. Infection proceeded without cytopathic effect and did not influence transepithelial resistance or the diffusion of inulin across cell monolayers. Nonetheless, there was extensive release of progeny virus, which occurred almost exclusively into apical supernatant fluids (36.4% ± 12.5% of the total virus yield compared with 0.23% ± 0.13% release into basolateral fluids). Brefeldin A caused a profound inhibition of HAV replication, but also selectively reduced apical release of virus. These results indicate that polarized human epithelial cell cultures undergo vectorial infection with HAV and that virus release is largely restricted to the apical membrane. Virus release occurs in the absence of cytopathic effect and may involve cellular vesicular transport mechanisms.

scholarly journals Study on effect of anti-diarrheal medicinal plants on enteropathogenic Escherichia coli induced interleukin-8 secretion by intestinal epithelial cells

2011 ◽  
Vol 1 (1) ◽  
pp. 16 ◽  
Author(s):  
S. Brijesh ◽  
Pundarikakshudu Tetali ◽  
Tannaz J. Birdi
Keyword(s):  
Escherichia Coli ◽  
Epithelial Cells ◽  
Medicinal Plants ◽  
Inflammatory Responses ◽  
Intestinal Epithelial Cells ◽  
Colon Adenocarcinoma ◽  
Cyperus Rotundus ◽  
Health Concern ◽  
Intestinal Epithelial ◽  
Infantile Diarrhea

Diarrhea is a major health concern in developing countries with enteropathogenic <em>Escherichia coli</em> (EPEC) being a leading cause of infantile diarrhea. Much of the pathology of EPEC infection is due to the inflammatory responses of infected intestinal epithelium through secretion of pro-inflammatory cytoki - nes such as interleukin (IL)-8. With medicinal plants gaining popularity as prospective antidiarrheal agents, we aimed to evaluate the effect of anti-diarrheal medicinal plants on secretion of IL-8 by epithelial cells in response to EPEC infection. The effect of the decoctions of four anti-diarrheal medicinal plants viz. <em>Aegle marmelos</em>, <em>Cyperus rotundus</em>, <em>Psidium guajava</em> and <em>Zingiber officinale</em> was studied on secretion of IL-8 by a human colon adenocarcinoma cell line, HT-29 infected with <em>E. coli </em>E2348/69. Two protocols were used viz. pre-incubation and post-incubation. The data obtained demonstrated that out of the four plants used, only <em>P. guajava</em> decreased secretion of IL-8 in the post-incubation protocol although in the pre-incubation protocol an increase was observed. A similar increase was seen with <em>C. rotundus</em> in the preincubation protocol. No effect on IL-8 secretion was observed with <em>A. marmelos</em> and <em>Z. officinale</em> in both protocols and with <em>C. rotundus </em>in the post-incubation protocol. The post-incubation protocol, in terms of clinical relevance, indicates the effect of the plant decoctions when used as treatment. Hence <em>P. guajava</em> may be effective in controlling the acute inflammatory response of the intestinal epithelial cells in response to EPEC infection.<p> </p>

Epithelial sorting of a glycosylphosphatidylinositol-anchored bacterial protein expressed in polarized renal MDCK and intestinal Caco-2 cells

1995 ◽  
Vol 108 (1) ◽  
pp. 369-377 ◽  
Author(s):  
K.L. Soole ◽  
M.A. Jepson ◽  
G.P. Hazlewood ◽  
H.J. Gilbert ◽  
B.H. Hirst
Keyword(s):  
Epithelial Cells ◽  
Cell Surface ◽  
Intestinal Epithelial Cells ◽  
Mdck Cells ◽  
Basolateral Membrane ◽  
Apical Cell ◽  
Apical Surface ◽  
Intestinal Epithelial ◽  
Gpi Anchor ◽  
Sorting Signal

To evaluate whether a glycosylphosphatidylinositol (GPI) anchor can function as a protein sorting signal in polarized intestinal epithelial cells, the GPI-attachment sequence from Thy-1 was fused to bacterial endoglucanase E' (EGE') from Clostridium thermocellum and polarity of secretion of the chimeric EGE'-GPI protein was evaluated. The chimeric EGE'-GPI protein was shown to be associated with a GPI anchor by TX-114 phase-partitioning and susceptibility to phosphoinositol-specific phospholipase C. In polarized MDCK cells, EGE' was localized almost exclusively to the apical cell surface, while in polarized intestinal Caco-2 cells, although 80% of the extracellular form of the enzyme was routed through the apical membrane over a 24 hour period, EGE' was also detected at the basolateral membrane. Rates of delivery of EGE'-GPI to the two membrane domains in Caco-2 cells, as determined with a biotinylation protocol, revealed apical delivery was approximately 2.5 times that of basolateral. EGE' delivered to the basolateral cell surface was transcytosed to the apical surface. These data indicate that a GPI anchor does represent a dominant apical sorting signal in intestinal epithelial cells. However, the mis-sorting of a proportion of EGE'GPI to the basolateral surface of Caco-2 cells provides an explanation for additional sorting signals in the ectodomain of some endogenous GPI-anchored proteins.

MiR-339 attenuates LPS-induced intestinal epithelial cells inflammatory responses and apoptosis by targeting TLR4

2020 ◽  
Vol 42 (9) ◽  
pp. 1097-1105 ◽  
Author(s):  
Meiying Xie ◽  
Lina Zhang ◽  
Luoye Li ◽  
Minhuan Fan ◽  
Lianjie Hou
Keyword(s):  
Epithelial Cells ◽  
Inflammatory Responses ◽  
Intestinal Epithelial Cells ◽  
Intestinal Epithelial

Induction of endothelin-1 synthesis by IL-2 and its modulation of rat intestinal epithelial cell growth

1998 ◽  
Vol 275 (3) ◽  
pp. G556-G563 ◽  
Author(s):  
Takeharu Shigematsu ◽  
Soichiro Miura ◽  
Masahiko Hirokawa ◽  
Ryota Hokari ◽  
Hajime Higuchi ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Cell Growth ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Cell Lines ◽  
Proliferative Response ◽  
Intestinal Epithelial Cell ◽  
Culture Media ◽  
Intestinal Epithelial Cells ◽  
Intestinal Epithelial

Endothelin (ET), a vasoconstrictive peptide, is known to have a variety of biological actions. Although ET is released by vascular endothelial cells, other cell populations also have been reported to synthesize and release ET. In this study, we examined whether ET is synthesized by intestinal epithelial cells and whether it affects induction of epithelial cell proliferation by interleukin-2 (IL-2). Subconfluent monolayers of intestinal epithelial cells (IEC-6 and IEC-18) were maintained in serum-free medium before addition of rat IL-2. Both IEC-6 and IEC-18 cells released ET-1 into the medium under unstimulated conditions, as determined by a sandwich ELISA. IL-2 significantly enhanced ET-1 release in a time-dependent manner. ET-3 was not detectable in the culture media of either cell line. Expression of ET-1 and ET-3 mRNA in epithelial cells was assessed by competitive PCR. Both cell lines were shown to express ET-1 mRNA, but no ET-3 mRNA was detected. IL-2 treatment enhanced ET-1 mRNA expression by both IEC-6 and IEC-18 cells. Both cell lines also expressed mRNA for ETA and ETB receptor subtypes. When cell proliferation was assessed, exogenous ET-1 induced a slight proliferative response in both types of cells that was consistent and significant at low ET-1 concentrations; cell growth was inhibited at a higher concentration (10−7M). IL-2 produced a significant proliferative response in both cell lines. However, the addition of ET-1 (10−7 M) to culture media attenuated the IL-2-induced increase in cell proliferation. ETA-receptor antagonists significantly enhanced cellular proliferation, suggesting involvement of the ETA receptor in modulation of IL-2-induced intestinal epithelial cell growth.

scholarly journals The Hippo Pathway Effector YAP1 Regulates Intestinal Epithelial Cell Differentiation

Cells ◽  
2020 ◽  
Vol 9 (8) ◽  
pp. 1895 ◽  
Author(s):  
Sepideh Fallah ◽  
Jean-François Beaulieu
Keyword(s):  
Stem Cells ◽  
Cell Differentiation ◽  
Epithelial Cells ◽  
Cell Line ◽  
Intestinal Epithelial Cells ◽  
Hippo Pathway ◽  
Intestinal Cell ◽  
Intestinal Epithelial ◽  
The Hippo Pathway

The human intestine is covered by epithelium, which is continuously replaced by new cells provided by stem cells located at the bottom of the glands. The maintenance of intestinal stem cells is supported by a niche which is composed of several signaling proteins including the Hippo pathway effectors YAP1/TAZ. The role of YAP1/TAZ in cell proliferation and regeneration is well documented but their involvement on the differentiation of intestinal epithelial cells is unclear. In the present study, the role of YAP1/TAZ on the differentiation of intestinal epithelial cells was investigated using the HT29 cell line, the only multipotent intestinal cell line available, with a combination of knockdown approaches. The expression of intestinal differentiation cell markers was tested by qPCR, Western blot, indirect immunofluorescence and electron microscopy analyses. The results show that TAZ is not expressed while the abolition of YAP1 expression led to a sharp increase in goblet and absorptive cell differentiation and reduction of some stem cell markers. Further studies using double knockdown experiments revealed that most of these effects resulting from YAP1 abolition are mediated by CDX2, a key intestinal cell transcription factor. In conclusion, our results indicate that YAP1/TAZ negatively regulate the differentiation of intestinal epithelial cells through the inhibition of CDX2 expression.

scholarly journals Mechanism of cholera toxin action on a polarized human intestinal epithelial cell line: role of vesicular traffic

1992 ◽  
Vol 117 (6) ◽  
pp. 1197-1209 ◽  
Author(s):  
WI Lencer ◽  
C Delp ◽  
MR Neutra ◽  
JL Madara
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
Cholera Toxin ◽  
Cell Line ◽  
Intestinal Epithelial Cell ◽  
Time Course ◽  
Epithelial Cell Line ◽  
Intestinal Epithelial ◽  
Vesicular Traffic ◽  
Intestinal Epithelial Cell Line

The massive secretion of salt and water in cholera-induced diarrhea involves binding of cholera toxin (CT) to ganglioside GM1 in the apical membrane of intestinal epithelial cells, translocation of the enzymatically active A1-peptide across the membrane, and subsequent activation of adenylate cyclase located on the cytoplasmic surface of the basolateral membrane. Studies on nonpolarized cells show that CT is internalized by receptor-mediated endocytosis, and that the A1-subunit may remain membrane associated. To test the hypothesis that toxin action in polarized cells may involve intracellular movement of toxin-containing membranes, monolayers of the polarized intestinal epithelial cell line T84 were mounted in modified Ussing chambers and the response to CT was examined. Apical CT at 37 degrees C elicited a short circuit current (Isc: 48 +/- 2.1 microA/cm2; half-maximal effective dose, ED50 integral of 0.5 nM) after a lag of 33 +/- 2 min which bidirectional 22Na+ and 36Cl- flux studies showed to be due to electrogenic Cl- secretion. The time course of the CT-induced Isc response paralleled the time course of cAMP generation. The dose response to basolateral toxin at 37 degrees C was identical to that of apical CT but lag times (24 +/- 2 min) and initial rates were significantly less. At 20 degrees C, the Isc response to apical CT was more strongly inhibited (30-50%) than the response to basolateral CT, even though translocation occurred in both cases as evidenced by the formation of A1-peptide. A functional rhodamine-labeled CT-analogue applied apically or basolaterally at 20 degrees C was visualized only within endocytic vesicles close to apical or basolateral membranes, whereas movement into deeper apical structures was detected at 37 degrees C. At 15 degrees C, in contrast, reduction to the A1-peptide was completely inhibited and both apical and basolateral CT failed to stimulate Isc although Isc responses to 1 nM vasoactive intestinal peptide, 10 microM forskolin, and 3 mM 8Br-cAMP were intact. Re-warming above 32 degrees C restored CT-induced Isc. Preincubating monolayers for 30 min at 37 degrees C before cooling to 15 degrees C overcame the temperature block of basolateral CT but the response to apical toxin remained completely inhibited. These results identify a temperature-sensitive step essential to apical toxin action on polarized epithelial cells. We suggest that this event involves vesicular transport of toxin-containing membranes beyond the apical endosomal compartment.

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