Three-dimensional organization and functional relationships of endocytotic compartments in pig intestinal epithelial cells

1992 ◽  
Vol 50 (1) ◽  
pp. 576-577
Author(s):  
Julian P. Heath ◽  
Buford L. Nichols ◽  
László G. Kömüves
Keyword(s):  
Electron Microscopy ◽  
Epithelial Cells ◽  
Intestinal Epithelial Cells ◽  
Three Dimensional ◽  
Intestinal Epithelial ◽  
Cell Surfaces ◽  
Basal Surface ◽  
Functional Relationships

The newborn pig intestine is adapted for the rapid and efficient absorption of nutrients from colostrum. In enterocytes, colostral proteins are taken up into an apical endocytotic complex of channels that transports them to target organelles or to the basal surface for release into the circulation. The apical endocytotic complex of tubules and vesicles clearly is a major intersection in the routes taken by vesicles trafficking to and from the Golgi, lysosomes, and the apical and basolateral cell surfaces.Jejunal tissues were taken from piglets suckled for up to 6 hours and prepared for electron microscopy and immunocytochemistry as previously described.

Comparison of gene expression and activation of transcription factors in organoid-derived monolayer intestinal epithelial cells and organoids

2021 ◽  
Author(s):  
Yu Takahashi ◽  
Yu Inoue ◽  
Keitaro Kuze ◽  
Shintaro Sato ◽  
Makoto Shimizu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Gene Expression Regulation ◽  
Intestinal Epithelial Cells ◽  
Three Dimensional ◽  
Culture Conditions ◽  
Dependent Manner ◽  
Intestinal Epithelial

Abstract Intestinal organoids better represent in vivo intestinal properties than conventionally used established cell lines in vitro. However, they are maintained in three-dimensional culture conditions that may be accompanied by handling complexities. We characterized the properties of human organoid-derived two-dimensionally cultured intestinal epithelial cells (IECs) compared with those of their parental organoids. We found that the expression of several intestinal markers and functional genes were indistinguishable between monolayer IECs and organoids. We further confirmed that their specific ligands equally activate intestinal ligand-activated transcriptional regulators in a dose-dependent manner. The results suggest that culture conditions do not significantly influence the fundamental properties of monolayer IECs originating from organoids, at least from the perspective of gene expression regulation. This will enable their use as novel biological tools to investigate the physiological functions of the human intestine.

scholarly journals Failure of propagation of human norovirus in intestinal epithelial cells with microvilli grown in three-dimensional cultures

2013 ◽  
Vol 159 (2) ◽  
pp. 257-266 ◽  
Author(s):  
Sayaka Takanashi ◽  
Linda J. Saif ◽  
John H. Hughes ◽  
Tea Meulia ◽  
Kwonil Jung ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Intestinal Epithelial Cells ◽  
Three Dimensional ◽  
Intestinal Epithelial ◽  
Human Norovirus

scholarly journals Immunocytochemical localization of ornithine transcarbamylase in rat intestinal mucosa. Light and electron microscopic study.

1988 ◽  
Vol 36 (1) ◽  
pp. 29-35 ◽  
Author(s):  
Y Hamano ◽  
H Kodama ◽  
M Yanagisawa ◽  
Y Haraguchi ◽  
M Mori ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Small Intestine ◽  
Epithelial Cells ◽  
Light Microscopy ◽  
Intestinal Mucosa ◽  
Intestinal Epithelial Cells ◽  
Protein A ◽  
Intestinal Epithelial ◽  
Electron Microscopic ◽  
Small Intestinal

We investigated light and electron microscopic localization of ornithine transcarbamylase (OTC) in rat intestinal mucosa. In the immunoblotting assay of OTC-related protein, a single protein band with a molecular weight of about 36,500 is observed in extracts of liver and small intestinal mucosa but is not observed in those of stomach and large intestine. For light microscopy, tissue slices of the digestive system were embedded in Epon and stained by using anti-bovine OTC rabbit IgG and the immunoenzyme technique. For electron microscopy, slices of these and the liver tissues were embedded in Lowicryl K4M and stained by the protein A-gold technique. By light microscopy, the absorptive epithelial cells of duodenum, jejunum, and ileum stained positively for OTC, but stomach, large intestine, rectum, and propria mucosa of small intestine were not stained. Electron microscopy showed that gold particles representing the antigenic sites for OTC were confined to the mitochondrial matrix of hepatocytes and small intestinal epithelial cells. However, the enzyme was detected in mitochondria of neither liver endothelial cells, submucosal cells of small intestine, nor large intestinal epithelial cells. Labeling density of mitochondria in the absorptive epithelial cells of duodenum, jejunum, and ileum was about half of that in liver cells.

scholarly journals A Three-Dimensional Cell Culture Model To Study Enterovirus Infection of Polarized Intestinal Epithelial Cells

mSphere ◽  
2015 ◽  
Vol 1 (1) ◽  
Author(s):  
Coyne G. Drummond ◽  
Cheryl A. Nickerson ◽  
Carolyn B. Coyne
Keyword(s):  
Cell Culture ◽  
Epithelial Cells ◽  
Intestinal Epithelium ◽  
Intestinal Epithelial Cells ◽  
Three Dimensional ◽  
Intestinal Epithelial ◽  
Cell Culture Model ◽  
Culture Model ◽  
Gastrointestinal Epithelium

ABSTRACT Coxsackievirus B (CVB), a member of the enterovirus family of RNA viruses, is associated with meningitis, pericarditis, diabetes, dilated cardiomyopathy, and myocarditis, among other pathologies. CVB is transmitted via the fecal-oral route and encounters the epithelium lining the gastrointestinal tract early in infection. The lack of suitable in vivo and in vitro models to study CVB infection of the gastrointestinal epithelium has limited our understanding of the events that surround infection of these specialized cells. Here, we report on the development of a three-dimensional (3-D) organotypic cell culture model of human intestinal epithelial cells that better models the gastrointestinal epithelium in vivo. By applying this 3-D model, which recapitulates many aspects of the gastrointestinal epithelium in vivo, to the study of CVB infection, our work provides a new cell system to model the mechanisms by which CVB infects the intestinal epithelium, which may have a profound impact on CVB pathogenesis. Despite serving as the primary entry portal for coxsackievirus B (CVB), little is known about CVB infection of the intestinal epithelium, owing at least in part to the lack of suitable in vivo models and the inability of cultured cells to recapitulate the complexity and structure associated with the gastrointestinal (GI) tract. Here, we report on the development of a three-dimensional (3-D) organotypic cell culture model of Caco-2 cells to model CVB infection of the gastrointestinal epithelium. We show that Caco-2 cells grown in 3-D using the rotating wall vessel (RWV) bioreactor recapitulate many of the properties of the intestinal epithelium, including the formation of well-developed tight junctions, apical-basolateral polarity, brush borders, and multicellular complexity. In addition, transcriptome analyses using transcriptome sequencing (RNA-Seq) revealed the induction of a number of genes associated with intestinal epithelial differentiation and/or intestinal processes in vivo when Caco-2 cells were cultured in 3-D. Applying this model to CVB infection, we found that although the levels of intracellular virus production were similar in two-dimensional (2-D) and 3-D Caco-2 cell cultures, the release of infectious CVB was enhanced in 3-D cultures at early stages of infection. Unlike CVB, the replication of poliovirus (PV) was significantly reduced in 3-D Caco-2 cell cultures. Collectively, our studies show that Caco-2 cells grown in 3-D using the RWV bioreactor provide a cell culture model that structurally and transcriptionally represents key aspects of cells in the human GI tract and can thus be used to expand our understanding of enterovirus-host interactions in intestinal epithelial cells. IMPORTANCE Coxsackievirus B (CVB), a member of the enterovirus family of RNA viruses, is associated with meningitis, pericarditis, diabetes, dilated cardiomyopathy, and myocarditis, among other pathologies. CVB is transmitted via the fecal-oral route and encounters the epithelium lining the gastrointestinal tract early in infection. The lack of suitable in vivo and in vitro models to study CVB infection of the gastrointestinal epithelium has limited our understanding of the events that surround infection of these specialized cells. Here, we report on the development of a three-dimensional (3-D) organotypic cell culture model of human intestinal epithelial cells that better models the gastrointestinal epithelium in vivo. By applying this 3-D model, which recapitulates many aspects of the gastrointestinal epithelium in vivo, to the study of CVB infection, our work provides a new cell system to model the mechanisms by which CVB infects the intestinal epithelium, which may have a profound impact on CVB pathogenesis. Podcast: A podcast concerning this article is available.

scholarly journals Massive programmed cell death in intestinal epithelial cells induced by three-dimensional growth conditions: suppression by mutant c-H-ras oncogene expression.

1995 ◽  
Vol 131 (6) ◽  
pp. 1587-1598 ◽  
Author(s):  
J Rak ◽  
Y Mitsuhashi ◽  
V Erdos ◽  
S N Huang ◽  
J Filmus ◽  
...  
Keyword(s):  
Cell Culture ◽  
Cell Death ◽  
Epithelial Cells ◽  
Programmed Cell Death ◽  
Intestinal Epithelial Cells ◽  
Three Dimensional ◽  
Intestinal Epithelial ◽  
Ras Oncogene ◽  
Multicellular Spheroids ◽  
Monolayer Cell Culture

Deregulation of molecular pathways controlling cell survival and death, including programmed cell death, are thought to be important factors in tumor formation, disease progression, and response to therapy. Studies devoted to analyzing the role of programmed cell death in cancer have been carried out primarily using conventional monolayer cell culture systems. However the majority of cancers grow as three-dimensional solid tumors. Because gene expression, and possibly function, can be significantly altered under such conditions, we decided to analyze the control and characteristics of cell death using a compatible three-dimensional tissue culture system (multicellular spheroids) and compare the results obtained to those using two-dimensional monolayer cell culture. To do so we selected for study an immortalized, but nontumorigenic line of rat intestinal epithelial cells, called IEC-18, and several tumorigenic variants of IEC-18 obtained by transfection with a mutant (activated) c-H-ras oncogene. The rationale for choosing these cell lines was based in part on the fact that intestinal epithelial cells grow in vivo in a monolayer-like manner and form solid tumors only after sustaining certain genetic mutations, including those involving the ras gene family. We found that the IEC-18 cells, which grow readily and survive in monolayer cell culture, undergo massive cell death within 48-72 h when cultured as multicellular spheroids on a nonadhesive surface. This process was accompanied by a number of features associated with programmed cell death including chromatin condensation (Hoechst 33258 staining) apoptotic morphology, DNA degradation, and a virtual complete loss of colony forming (clonogenic) ability in the absence of apparent membrane damage as well as accumulation of lipid containing vacuoles in the cytoplasm. Moreover, enforced over-expression of a transfected bcl-2 gene could prevent this cell death process from taking place. In marked contrast, three different stably transfected ras clones of IEC-18 survived when grown as multicellular spheroids. In addition, an IEC cell line (called clone 25) carrying its mutant transfected ras under a glucocorticoid inducible promoter survived in three-dimensional culture only when the cells were exposed to dexamethasone. If exposure to dexamethasone was delayed for as long as 48 h the cells nevertheless survived, whereas the cells became irreversibly committed to programmed cell death (PCD) if exposed to dexamethasone after 72 h. These results suggest that intestinal epithelial cells may be programmed to activate a PCD pathway upon detachment from a physiologic two-dimensional monolayer configuration, and that this process of adhesion regulated programmed cell death (ARPCD) can be substantially suppressed by expression of a mutant ras oncogene.(ABSTRACT TRUNCATED AT 400 WORDS)

Engineering a Biomimetic Villus Array for In Vitro Three-Dimensional Culture of Intestinal Epithelial Cells

2012 ◽  
Vol 21 (6) ◽  
pp. 1418-1425
Author(s):  
Wen-Cheng Yang ◽  
Yuan-Ching Chen ◽  
Yu-Sheng Huang ◽  
Ya-Yuan Fu ◽  
Shiue-Cheng Tang ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Intestinal Epithelial Cells ◽  
Three Dimensional ◽  
Intestinal Epithelial ◽  
Three Dimensional Culture

Epithelial cell basal processes in the duodenum of NZB mice

1978 ◽  
Vol 36 (2) ◽  
pp. 534-535
Author(s):  
B. T. Partridge ◽  
L. O. Simpson
Keyword(s):  
Epithelial Cells ◽  
Basement Membrane ◽  
Intestinal Epithelial Cells ◽  
Mouse Strains ◽  
Peptic Ulcers ◽  
Intestinal Epithelial ◽  
Focal Lesions ◽  
Basal Surface ◽  
Scanning Electron Microscope Study ◽  
Human Small Intestine

Although spontaneous peptic ulcers in NZB mice were described first in 1965 and the incidence of the lesion reported two years later no studies of the fine structure of the lesions have been reported. During the course of a scanning electron microscope study of the proximal duodenum of NZB and related mouse strains focal lesions previously unreported were observed. Early lesions were characterised by loss of epithelial cells from the tips of villi. Recently denuded basement membrane had a very rough surface and it was noted that pores 0. 5 to 1. 0 microns in diameter, (Fig. 1) were arranged in rows with regular interspaces between the pores. Epithelial cells which had fallen from the villi in sheets revealing their basement membrane aspect had elongated basal processes (Fig. 2, 3). In a study on the morphology of the human small intestine (Trier, 1968) it was noted that, “Occasionally small pseudopod-like extensions of cytoplasm may penetrate the basement membrane and project into the lamina propria from the basal surface of intestinal epithelial cells”.

scholarly journals Intestinal cyto differentiation in vitro of chick stomach endoderm induced by the duodenal mesenchyme

Development ◽  
1984 ◽  
Vol 82 (1) ◽  
pp. 163-176
Author(s):  
Atsuko Ishizuya-Oka ◽  
Takeo Mizuno
Keyword(s):  
Electron Microscopy ◽  
Epithelial Cells ◽  
Intestinal Epithelium ◽  
Intestinal Epithelial Cells ◽  
Goblet Cells ◽  
Secretory Cells ◽  
Chick Embryos ◽  
Intestinal Epithelial ◽  
Self Differentiation

The inductive action of duodenal mesenchyme on the cytodifferentiation of stomach endoderm in chick embryos was investigated in vitro with electron microscopy and immunofluorescence. Morphologically undifferentiated endoderm of the stomach of a 4-day embryo could differentiate only into a mucous secretory epithelium when cultured in the absence of mesenchyme. However, when cultivated in recombination with 6-day duodenal mesenchyme, most cells of 4-day stomach endoderm differentiated into intestinal absorptive cells possessing striated border and sucrase, and goblet cells, but not into stomach-type mucous secretory cells. In contrast, when 4-day stomach endoderm was cultured recombined with mesenchyme of embryonic digestive organs other than intestine, none of the stomach endoderm cells differentiated into intestinal epithelial cells. The competence of stomach endoderm for intestinal cytodifferentiation decreased rapidly with development, but remained until relatively later stages in the gizzard region. The present investigation demonstrates that duodenal mesenchyme can induce stomach endoderm, which has acquired the potency for self-differentiation into stomach-type epithelium, to cytodifferentiate into intestinal epithelium.

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