scholarly journals Organization of actin, myosin, and intermediate filaments in the brush border of intestinal epithelial cells.

1982 ◽  
Vol 94 (2) ◽  
pp. 425-443 ◽  
Author(s):  
N Hirokawa ◽  
L G Tilney ◽  
K Fujiwara ◽  
J E Heuser
Keyword(s):  
Epithelial Cells ◽  
Intermediate Filaments ◽  
Actin Filaments ◽  
Intestinal Epithelial Cells ◽  
Intestinal Cell ◽  
Intestinal Epithelial ◽  
Replication Method ◽  
Terminal Web ◽  
Filament Bundles ◽  
Lateral Margins

Terminal webs prepared from mouse intestinal epithelial cells were examined by the quick-freeze, deep-etch, and rotary-replication method. The microvilli of these cells contain actin filaments that extend into the terminal web in compact bundles. Within the terminal web these bundles remain compact; few filaments are separated from the bundles and fewer still bend towards the lateral margins of the cell. Decoration with subfragment 1 (S1) of myosin confirmed that relatively few actin filaments travel horizontally in the web. Instead, between actin bundles there are complicated networks of the fibrils. Here we present two lines of evidence which suggest that myosin is one of the major cross-linkers in the terminal web. First, when brush borders are exposed to 1 mM ATP in 0.3 M KCl, they lose their normal ability to bind antimyosin antibodies as judged by immunofluorescence, and they lose the thin fibrils normally found in deep-etch replicas. Correspondingly, myosin is released into the supernatant as judged by SDS gel electrophoresis. Second, electron microscope immunocytochemistry with antimyosin antibodies followed by ferritin-conjugated second antibodies leads to ferritin deposition mainly on the fibrils at the basal part of rootlets. Deep-etching also reveals that the actin filament bundles are connected to intermediate filaments by another population of cross-linkers that are not extracted by ATP in 0.3 M KCl. From these results we conclude that myosin in the intestinal cell may not only be involved in a short range sliding-filament type of motility, but may also play a purely structural role as a long range cross-linker between microvillar rootlets.

scholarly journals The spectrin-related molecule, TW-260/240, cross-links the actin bundles of the microvillus rootlets in the brush borders of intestinal epithelial cells.

1983 ◽  
Vol 96 (5) ◽  
pp. 1491-1496 ◽  
Author(s):  
J R Glenney ◽  
P Glenney ◽  
K Weber
Keyword(s):  
Epithelial Cells ◽  
Actin Filaments ◽  
Intestinal Epithelial Cells ◽  
Intestinal Epithelial ◽  
Cross Link ◽  
Actin Bundles ◽  
Terminal Web ◽  
Brush Borders ◽  
Cross Links

Previous studies have shown that molecules related to erythrocyte spectrin are present in the cortical cytoplasm of nonerythroid cells. We report here the localization by immunoelectron microscopy of one such molecule, TW-260/240, in the brush border of intestinal epithelial cells. Using highly specific antibodies against TW-260 and TW-240 as well as antibodies against fodrin, another spectrinlike molecule, we have found that the TW-260/240 molecules are displayed between rootlets at all levels of the terminal web. Occasionally, extended structures appear labeled suggestive of the fine filaments known to cross-link actin bundles. These results are in line with previous in vitro studies showing that TW-260/240 binds to, and cross-links, actin filaments. The results are discussed in terms of a model in which rootlets are immobilized in the terminal web in a matrix of TW-260/240.

scholarly journals Organization of an actin filament-membrane complex. Filament polarity and membrane attachment in the microvilli of intestinal epithelial cells.

1975 ◽  
Vol 67 (3) ◽  
pp. 725-743 ◽  
Author(s):  
M S Mooseker ◽  
L G Tilney
Keyword(s):  
Epithelial Cells ◽  
Actin Filament ◽  
Brush Border ◽  
Actin Filaments ◽  
Functional Organization ◽  
Intestinal Epithelial Cells ◽  
Intestinal Epithelial ◽  
Membrane Complex ◽  
Brush Borders ◽  
Filament Bundles

The association of actin filaments with membranes is now recognized as an important parameter in the motility of nonmuscle cells. We have investigated the organization of one of the most extensive and highly ordered actin filament-membrane complexes in nature, the brush border of intestinal epithelial cells. Through the analysis of isolated, demembranated brush borders decorated with the myosin subfragment, S1, we have determined that all the microvillar actin filaments have the same polarity. The S1 arrowhead complexes point away from the site of attachment of actin filaments at the apical tip of the microvillar membrane. In addition to the end-on attachment of actin filaments at the tip of the microvillus, these filaments are also connected to the plasma membrane all along their lengths by periodic (33 nm) cross bridges. These bridges were best observed in isolated brush borders incubated in high concentrations of Mg++. Their visibility is attributed to the induction of actin paracrystals in the filament bundles of the microvilli. Finally, we present evidence for the presence of myosinlike filaments in the terminal web region of the brush border. A model for the functional organization of actin and myosin in the brush border is presented.

scholarly journals Quick-freeze, deep-etch visualization of the cytoskeleton beneath surface differentiations of intestinal epithelial cells.

1981 ◽  
Vol 91 (2) ◽  
pp. 399-409 ◽  
Author(s):  
N Hirokawa ◽  
J E Heuser
Keyword(s):  
Epithelial Cells ◽  
Intestinal Epithelial Cells ◽  
Intestinal Epithelial ◽  
Terminal Web ◽  
Cell Biol ◽  
Myosin Subfragment ◽  
Quick Freezing ◽  
New Finding ◽  
Filament Bundles ◽  
Replication Technique

The cytoskeleton that supports microvilli in intestinal epithelial cells was visualized by the quick-freeze, deep-etch, rotary-replication technique (Heuser and Salpeter. 1979. J. Cell Biol. 82: 150). Before quick freezing, cells were exposed to detergents or broken open physically to clear away the granular material in their cytoplasm that would otherwise obscure the view. After such extraction, cells still displayed a characteristic organization of cytoskeletal filaments in their interiors. Platinum replicas of these cytoskeletons had sufficient resolution to allow us to identify the filament types present, and to determine their characteristic patterns of interaction. The most important new finding was that the apical "terminal web" in these cells, which supports the microvilli via their core bundles of actin filaments, does not itself contain very much actin but instead is comprised largely of narrow strands that interconnect adjacent actin bundles with one another and with the underlying base of intermediate filaments. These strands are slightly thinner than actin, do not display actin's 53A periodicity, and do not decorate with myosin subfragment S1. On the contrary, two lines of evidence suggested that these strands, could include myosin molecules. First, other investigators have shown that myosin is present in the terminal web (Mooseker et al. 1978. J. Cell Biol. 79: 444-453), yet we could find no thick filaments in this area. Second, we found that the strands were removed completely in the process of decorating the core filament bundles with the myosin subfragment S1, suggesting that they had been competitively displaced by exogenous myosin. We conclude that myosin may play a structural role in these cells, via its cross-linking distribution, in addition to whatever role it plays in microvillar motility.

Etomidate Alleviates Ischemia-Anoxia Reperfusion Injury in Intestinal Epithelial Cells by Inhibiting the Activation of traf6-Regulated NF-KB Signaling

2022 ◽  
Vol 12 (5) ◽  
pp. 1015-1021
Author(s):  
Gen Lin ◽  
Ruichun Long ◽  
Xiaoqing Yang ◽  
Songsong Mao ◽  
Hongying Li
Keyword(s):  
Oxidative Stress ◽  
Epithelial Cells ◽  
Reperfusion Injury ◽  
Ischemia Reperfusion Injury ◽  
Intestinal Epithelial Cells ◽  
Ischemia Reperfusion ◽  
Stress Factors ◽  
Inflammatory Factors ◽  
Intestinal Cell ◽  
Intestinal Epithelial

Objective: The present study aimed to investigate the role of etomidate in intestinal cell ischemia and hypoxia-reperfusion injury and potential mechanisms. Method: In this study, we establish the intestinal epithelial cells ischemia-reperfusion model in vitro. CCK8 was used to detect cell viability and flow cytometry assay was used to detect apoptosis levels of treated OGD/R model cells. ELISA measured the expression level of oxidative stress factors and inflammatory factors. Furthermore, western blot assay was used to detect the expression the apoptosis-related factors and TNFR-associated factors in treated OGD/R model cells. Result: Etomidate does not affect the activity of intestinal epithelial cells, and can protect intestinal epithelial cells to reduce ischemiareperfusion injury, and the expression of inflammatory factors and oxidative stress in cells with mild intestinal epithelial ischemia-reperfusion injury. Etomidate alleviates apoptosis of intestinal epithelial ischemia-reperfusion injury cells. Etomidate inhibits the activation of traf6-mediated NF-κB signal during ischemia-anoxia reperfusion of intestinal epithelial cells. Conclusion: Taken together, our study demonstrated that etomidate attenuates inflammatory response and apoptosis in intestinal epithelial cells during ischemic hypoxia-reperfusion injury and inhibits activation of NF-κB signaling regulated by TRAF6.

A thermal injury-induced circulating factor(s) compromises intestinal cell morphology, proliferation, and migration

1999 ◽  
Vol 277 (1) ◽  
pp. G175-G182 ◽  
Author(s):  
Maryam Varedi ◽  
George H. Greeley ◽  
David N. Herndon ◽  
Ella W. Englander
Keyword(s):  
Epithelial Cells ◽  
Thermal Injury ◽  
Intestinal Epithelial Cells ◽  
Intestinal Cell ◽  
Intestinal Epithelial ◽  
Rat Serum ◽  
In Vitro Effects ◽  
Mucosal Morphology ◽  
And Migration

The effects of a 60% body surface area thermal injury in rats on the morphology and proliferation of the epithelium of the small intestine and the in vitro effects of serum collected from scalded rats on intestinal epithelial cells were investigated. Scald injury caused significant reductions in duodenal villus width and crypt dimensions, villus enterocytes changed in shape from columnar to cuboidal, and the number of goblet cells decreased. The proportion of bromodeoxyuridine-labeled S phase cells in crypts was also diminished. In vitro, incubation of intestinal epithelial cells (IEC-6) with scalded rat serum (SRS) collected at either 12 or 24 h after injury caused a disruption in the integrity of the confluent culture and induced the appearance of large denuded areas. SRS also decreased DNA synthesis and delayed wound closure in an in vitro wound-healing model. The thermal injury-induced changes in intestinal mucosal morphology and epithelial cell growth characteristics described in this study may underlie, in part, the mechanism(s) involved in the diminished absorption of nutrients, increased intestinal permeability, and sepsis in patients with thermal injury.

The Fine-Structural Organization of the Brush Border of Intestinal Epithelial Cells

1971 ◽  
Vol 8 (3) ◽  
pp. 573-599
Author(s):  
T. M. MUKHERJEE ◽  
L. A. STAEHELIN
Keyword(s):  
Plasma Membrane ◽  
Epithelial Cells ◽  
Brush Border ◽  
Intestinal Epithelial Cells ◽  
Membrane Surface ◽  
Intestinal Epithelial ◽  
The Core ◽  
Unit Structure ◽  
Terminal Web ◽  
Freeze Etching

The fine structure of the brush border of intestinal epithelial cells of the mouse has been studied with both normal sectioning and freeze-etching techniques. Freeze-etching reveals the plasma membrane of the microvilli as consisting of a continuous layer, that is split during the cleaving process, in which numerous particles, 5-9 nm in diameter, are embedded, while other particle-like structures, with diameters of 7-10 nm, appear attached to the true outer membrane surface. The mucopolysaccharide surface coats of the microvilli show up more clearly in sectioned material than in freeze-etched specimens. Inside each microvillus 2 different filament systems can be demonstrated: (1) bundles of fairly closely packed and straight core microfilaments, which lead into the tip of the microvillus, and (2) short cross-filaments. Under suitable conditions the core microfilaments display a sub-unit structure with a repeating distance of approximately 6 nm. The diameter of a microfilament can vary along its length from 6 to 11 nm. Two strands of globular particles wound helically around each other seem to make up each microfilament. These and other data support the idea that the core microfilaments are actin-like. No substructure has been found on the cross-filaments, which have an orientation approximately radial to the axis of the microvilli and seem to be attached at one end to the core microfilaments and at the other to the inner surface of the microvillous membrane. The interwoven terminal web filaments also show no substructure. They form a continuous flexible platform-like structure into which the bundles of core microfilaments extend. Some terminal web filaments appear attached to the plasma membrane between the microvilli. It is suggested that the core microfilaments represent mechanical supporting elements and that the terminal web and cross-filaments are tensile elements of the brush border. In addition all 3 filament systems may also be involved in possible contractile movements of the microvilli.

Na(+)-K(+)-ATPase gene expression in rat intestine and Caco-2 cells: response to thyroid hormone

1993 ◽  
Vol 265 (4) ◽  
pp. G775-G782 ◽  
Author(s):  
R. A. Giannella ◽  
J. Orlowski ◽  
M. L. Jump ◽  
J. B. Lingrel
Keyword(s):  
Thyroid Hormone ◽  
Epithelial Cells ◽  
Enzymatic Activity ◽  
Intestinal Epithelial Cells ◽  
Blot Hybridization ◽  
Intestinal Cell ◽  
Transcriptional Level ◽  
Intestinal Epithelial ◽  
Subunit Mrna ◽  
Atpase Gene

Expression of the Na(+)-K(+)-adenosinetriphosphatase (ATPase) gene family in rat intestinal epithelial cells was examined using RNA blot hybridization analyses. Rat intestinal epithelial cells express only the alpha 1- and beta 1-subunit mRNAs. A gradient in expression of alpha 1- and beta 1-subunit mRNA was seen along the villus-crypt unit in both jejunum and ileum, i.e., villus tip >> crypt cells. Regional differences in expression were observed along the intestine. alpha 1- and beta 1-subunit mRNA abundance was similar in jejunum, ileum, and colon while enzymatic activity was highest in the jejunum and lowest in the ileum. Administration of thyroid hormone to thyroidectomized rats increased the expression of alpha 1- and beta 1-subunit mRNAs in jejunum but not in colon. Hypothyroidism had no effect on subunit mRNA expression. The human intestinal cell line Caco-2 was also studied. These cells also expressed only the alpha 1- and beta 1-isoform mRNAs and demonstrated a developmental profile in both mRNA and enzymatic activity. Furthermore, in Caco-2 cells both alpha 1- and beta 1-mRNAs and Na(+)-K(+)-ATPase enzymatic activity were stimulated by thyroid hormone. Caco-2 cells transfected with 5' flanking regions of the human Na(+)-K(+)-ATPase beta 1-gene linked to the chloramphenicol acetyltransferase (CAT) reporter gene responded to 3,5,3'-triiodothyronine (T3) treatment with increased expression of CAT activity. This suggests that the 5' flanking region of the beta 1-gene contains a thyroid hormone response element and that T3 upregulation occurs at the transcriptional level.(ABSTRACT TRUNCATED AT 250 WORDS)

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 Intermediate Filaments Interact with Dormant Ezrin in Intestinal Epithelial Cells

2005 ◽  
Vol 16 (9) ◽  
pp. 4096-4107 ◽  
Author(s):  
Flavia A. Wald ◽  
Andrea S. Oriolo ◽  
M. Llanos Casanova ◽  
Pedro J.I. Salas
Keyword(s):  
Epithelial Cells ◽  
Intermediate Filaments ◽  
Brush Border ◽  
Antisense Rna ◽  
Apical Membrane ◽  
Intestinal Epithelial Cells ◽  
Intestinal Epithelial ◽  
Undifferentiated Cells ◽  
Responsive Element

Ezrin connects the apical F-actin scaffold to membrane proteins in the apical brush border of intestinal epithelial cells. Yet, the mechanisms that recruit ezrin to the apical domain remain obscure. Using stable CACO-2 transfectants expressing keratin 8 (K8) antisense RNA under a tetracycline-responsive element, we showed that the actin-ezrin scaffold cannot assemble in the absence of intermediate filaments (IFs). Overexpression of ezrin partially rescued this phenotype. Overexpression of K8 in mice also disrupted the assembly of the brush border, but ezrin distributed away from the apical membrane in spots along supernumerary IFs. In cytochalasin D-treated cells ezrin localized to a subapical compartment and coimmunoprecipitated with IFs. Overexpression of ezrin in undifferentiated cells showed a Triton-insoluble ezrin compartment negative for phospho-T567 (dormant) ezrin visualized as spots along IFs. Pulse-chase analysis showed that Triton-insoluble, newly synthesized ezrin transiently coimmunoprecipitates with IFs during the first 30 min of the chase. Dormant, but not active (p-T567), ezrin bound in vitro to isolated denatured keratins in Far-Western analysis and to native IFs in pull-down assays. We conclude that a transient association to IFs is an early step in the polarized assembly of apical ezrin in intestinal epithelial cells.

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