scholarly journals DES2 protein is responsible for phytoceramide biosynthesis in the mouse small intestine

2004 ◽  
Vol 379 (3) ◽  
pp. 687-695 ◽  
Author(s):  
Fumio OMAE ◽  
Masao MIYAZAKI ◽  
Ayako ENOMOTO ◽  
Minoru SUZUKI ◽  
Yusuke SUZUKI ◽  
...  
Keyword(s):  
Small Intestine ◽  
Epithelial Cells ◽  
Intestinal Epithelial ◽  
Vitro Assay ◽  
Peptide Antibody ◽  
Mouse Small Intestine ◽  
Mrna Content ◽  
Crypt Cells

The C-4 hydroxylation of sphinganine and dihydroceramide is a rate-limiting reaction in the biosynthesis of phytosphingolipids. Mouse DES1 (MDES1) cDNA homologous to the Drosophila melanogaster degenerative spermatocyte gene-1 (des-1) cDNA leads to sphingosine Δ4-desaturase activity, and another mouse homologue, MDES2, has bifunctional activity, producing C-4 hydroxysphinganine and Δ4-sphingenine in yeast [Ternes, Franke, Zahringer, Sperling and Heinz (2002) J. Biol. Chem. 277, 25512–25518]. Here, we report the characterization of mouse DES2 (MDES2) using an in vitro assay with a homogenate of COS-7 cells transfected with MDES2 cDNA and N-octanoyl-sphinganine and sphinganine as substrates. MDES2 protein prefers dihydroceramide as a substrate to sphinganine, and exhibits dihydroceramide Δ4-desaturase and C-4 hydroxylase activities. MDES2 mRNA content was high in the small intestine and abundant in the kidney. In situ hybridization detected signals of MDES2 mRNA in the crypt cells. Immunohistochemistry using an anti-MDES2 peptide antibody stained the crypt cells and the adjacent epithelial cells. These results suggest that MDES2 is the dihydroceramide C-4 hydroxylase responsible for the biosynthesis of enriched phytosphingoglycolipids in the microvillous membranes of intestinal epithelial cells.

Effect of leptin on intestinal apolipoprotein AIV in response to lipid feeding

2001 ◽  
Vol 281 (3) ◽  
pp. R753-R759 ◽  
Author(s):  
Takashi Doi ◽  
Min Liu ◽  
Randy J. Seeley ◽  
Stephen C. Woods ◽  
Patrick Tso
Keyword(s):  
Small Intestine ◽  
Epithelial Cells ◽  
Intestinal Epithelial Cells ◽  
Regional Distribution ◽  
Intestinal Epithelial ◽  
Lipid Infusion ◽  
Intraduodenal Infusion ◽  
Crypt Cells

We determined apolipoprotein AIV (apo AIV) content in intestinal epithelial cells using immunohistochemistry when leptin was administered intravenously. Most of the apo AIV immunoreactivity in the untreated intestine was located in the villous cells as opposed to the crypt cells. Regional distribution of apo AIV immunostaining revealed low apo AIV content in the duodenum and high content in the jejunum that gradually decreases caudally toward the ileum. Intraduodenal infusion of lipid (4 h) significantly increased apo AIV immunoreactivity in the jejunum and ileum. Simultaneous intravenous leptin infusion plus duodenal lipid infusion markedly suppressed apo AIV immunoreactivity. Duodenal lipid infusion increased plasma apo AIV significantly (measured by ELISA), whereas simultaneous leptin infusion attenuated the increase. These findings suggest that leptin may regulate circulating apo AIV by suppressing apo AIV synthesis in the small intestine.

An adapted in vitro assay to assess Campylobacter jejuni interaction with intestinal epithelial cells: Taking into stimulation with TNFα

2018 ◽  
Vol 149 ◽  
pp. 67-72 ◽  
Author(s):  
Ramila Cristiane Rodrigues ◽  
Anne-Lise Pocheron ◽  
Jean-Michel Cappelier ◽  
Odile Tresse ◽  
Nabila Haddad
Keyword(s):  
Epithelial Cells ◽  
Campylobacter Jejuni ◽  
Intestinal Epithelial Cells ◽  
In Vitro Assay ◽  
Intestinal Epithelial ◽  
Vitro Assay

scholarly journals 62 Evaluating detoxification efficacy of deoxynivalenol by encapsulated sodium metabisulfite (SMBS) using an in vitro intestinal epithelial cell IPEC-J2 model

2020 ◽  
Vol 98 (Supplement_4) ◽  
pp. 38-39
Author(s):  
Peng Lu ◽  
Changning Yu ◽  
Shangxi Liu ◽  
Joshua Gong ◽  
Song Liu ◽  
...  
Keyword(s):  
Small Intestine ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Intestinal Epithelial Cell ◽  
Intestinal Epithelial Cells ◽  
In Vitro Release ◽  
Simulated Gastric Fluid ◽  
Sodium Metabisulfite ◽  
Intestinal Epithelial

Abstract Deoxynivalenol (DON) contamination occurs on feed ingredients and causes a reduction in growth performance, damage to the intestinal epithelial cells, and increased susceptibility to enteric pathogen challenge. Sodium metabisulfite (SMBS) has been successfully used to destroy DON in processed grains or feeds. However, SMBS degrades quickly under aqueous acid conditions, such as pig stomachs, and when SMBS is added to diet, little will remain intact in the small intestine where an optimal pH environment exists for detoxification by SMBS. Thus, this study was to encapsulate SMBS into microparticles to deliver intact SMBS to the small intestine and evaluate its efficacy of DON detoxification in the simulated intestine fluid (SIF) using an in vitro intestinal epithelial cell (IPEC-J2) model. The results showed that around 40% of the SMBS loading capacity was achieved in the microparticles. In vitro release studies showed that 1.61% of encapsulated SMBS was released in the simulated gastric fluid (SGF), and the majority of encapsulated SMBS (75.52%) was progressively released in the SIF within 6 h at 37 °C. In vitro cell experiments showed that DON treated with the SIF containing 0.5% SMBS for 2 h completely attenuated the DON-induced cytotoxicity. When DON was treated with the SGF containing 0.5% encapsulated SMBS for 2 h and then the mixture was mixed with the SIF (1:1) and incubated for 2 h, it also completely attenuated the DON-induced cytotoxicity. Moreover, DON treated with the simulated fluid containing 0.5% encapsulated SMBS completely attenuated the gene expression inflammatory cytokines upregulated by DON and restored trans-epithelial electrical resistance (TEER) and tight junction and cytoskeleton. In summary, the encapsulation of SMBS was stable in SGF and allowed a progressive release of SMBS in the SIF. Moreover, the released SMBS in the SIF effectively attenuated the adverse effects induced by DON in the intestinal epithelial cells.

Isolation of intestinal epithelial cells for the study of differential gene expression along the crypt-villus axis

1991 ◽  
Vol 260 (6) ◽  
pp. G895-G903 ◽  
Author(s):  
P. G. Traber ◽  
D. L. Gumucio ◽  
W. Wang
Keyword(s):  
Epithelial Cells ◽  
Intestinal Epithelial Cells ◽  
Light And Electron Microscopy ◽  
Intestinal Epithelial ◽  
Isolated Cells ◽  
Intestinal Functions ◽  
Differential Gene ◽  
Cell Fractions ◽  
Crypt Cells

Methods for the differential isolation of intestinal epithelial cells from crypt and villus compartments of small intestine have been used in the study of intestinal functions. However, the use of different methods has resulted in discrepant conclusions as to the localization of expressed genes. Therefore, we undertook a careful comparison of two methods of intestinal epithelial cell isolation, the distended intestinal sac method and the everted intestinal sac method. The isolated cell fractions (distended sac fractions 1-10, everted sac fractions 1-5) were evaluated for the expression of two mRNAs whose localization along the crypt-villus axis had been previously elucidated by in situ hybridization: cytochrome P-450IIB1 (expressed in villus cells) and cryptdin (expressed in crypt cells). Northern blots of total or poly(A)+ RNA from each cell population showed that the first fractions from both methods contained P-450IIB1 mRNA, suggesting that both methods allowed the isolation of cells originating from the villus. Cryptdin mRNA was detected only in cell fractions 5-10 using the distended sac method and was not detected in any fractions from the everted sac method. [3H]thymidine incorporation demonstrated that dividing (crypt) cells were successfully removed by the distended sac method, but remained with the everted sac intestinal remnant. Finally, light and electron microscopy of the isolated cells as well as the intestinal remnants confirmed that while undifferentiated crypt cells were present in distended sac cell fractions 9 and 10, they remained with the everted sac remnant. Thus the distended sac protocol was useful for the isolation of cells from tip and crypt compartments.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Fibroblast growth factor 10 alters the balance between goblet and Paneth cells in the adult mouse small intestine

2015 ◽  
Vol 308 (8) ◽  
pp. G678-G690 ◽  
Author(s):  
Denise Al Alam ◽  
Soula Danopoulos ◽  
Kathy Schall ◽  
Frederic G. Sala ◽  
Dana Almohazey ◽  
...  
Keyword(s):  
Small Intestine ◽  
Cell Differentiation ◽  
Epithelial Cell ◽  
Paneth Cells ◽  
Intestinal Epithelial ◽  
Mouse Small Intestine ◽  
Fibroblast Growth Factor 10 ◽  
Goblet Cell Differentiation

Intestinal epithelial cell renewal relies on the right balance of epithelial cell migration, proliferation, differentiation, and apoptosis. Intestinal epithelial cells consist of absorptive and secretory lineage. The latter is comprised of goblet, Paneth, and enteroendocrine cells. Fibroblast growth factor 10 (FGF10) plays a central role in epithelial cell proliferation, survival, and differentiation in several organs. The expression pattern of FGF10 and its receptors in both human and mouse intestine and their role in small intestine have yet to be investigated. First, we analyzed the expression of FGF10, FGFR1, and FGFR2, in the human ileum and throughout the adult mouse small intestine. We found that FGF10, FGFR1b, and FGFR2b are expressed in the human ileum as well as in the mouse small intestine. We then used transgenic mouse models to overexpress Fgf10 and a soluble form of Fgfr2b, to study the impact of gain or loss of Fgf signaling in the adult small intestine. We demonstrated that overexpression of Fgf10 in vivo and in vitro induces goblet cell differentiation while decreasing Paneth cells. Moreover, FGF10 decreases stem cell markers such as Lgr5, Lrig1, Hopx, Ascl2, and Sox9. FGF10 inhibited Hes1 expression in vitro, suggesting that FGF10 induces goblet cell differentiation likely through the inhibition of Notch signaling. Interestingly, Fgf10 overexpression for 3 days in vivo and in vitro increased the number of Mmp7/Muc2 double-positive cells, suggesting that goblet cells replace Paneth cells. Further studies are needed to determine the mechanism by which Fgf10 alters cell differentiation in the small intestine.

Sucrase-isomaltase gene expression along crypt-villus axis of human small intestine is regulated at level of mRNA abundance

1992 ◽  
Vol 262 (1) ◽  
pp. G123-G130 ◽  
Author(s):  
P. G. Traber ◽  
L. Yu ◽  
G. D. Wu ◽  
T. A. Judge
Keyword(s):  
Gene Expression ◽  
Small Intestine ◽  
In Situ Hybridization ◽  
Epithelial Cells ◽  
Stem Cell Population ◽  
Specific Gene ◽  
Mrna Levels ◽  
Human Small Intestine ◽  
Crypt Cells

The mucosal lining of the small intestine is a complex epithelium that is continually renewed by division of a stem cell population located in intestinal crypts, migration of daughter cells along the villus, and, finally, extrusion of senescent cells into the lumen. The majority of cells in both crypt and villus cell compartments are enterocytes that acquire differentiated functions as they migrate out of the crypt. Sucrase-isomaltase (SI) is an enterocyte-specific, brush-border enzyme that has little activity in crypt cells and maximal activity in low and mid villus cells. The mechanism by which enterocytes acquire SI enzymatic activity as they move from crypt to villus is controversial. In this study we examined the distribution of SI mRNA along the crypt-villus axis of human small intestine using isolated epithelial cells and in situ hybridization. A complementary DNA to the 5' portion of the human SI mRNA was amplified and cloned using the polymerase chain reaction. Hybridization analysis of RNA extracted from human intestinal epithelial cells showed that the cloned cDNA recognized a single 6.5-kb mRNA. In situ hybridization of duodenal biopsy specimens was performed using a single-stranded RNA probe derived from this cDNA. This analysis showed that there was little SI mRNA in crypt cells and appearance of mRNA in enterocytes located at the crypt-villus junction. The mRNA levels were maximal in lower and mid villus cells with decreased levels noted in villus tip cells. These results are identical to those previously described in rat intestine and suggest that expression of the SI gene as enterocytes emerge from intestinal crypts is regulated primarily at the level of mRNA accumulation. Study of SI gene regulation may provide a useful model to investigate the mechanisms that regulate enterocyte-specific gene expression and intestinal differentiation.

Electron probe x-ray microanalysis and electron microscopy of amino acid absorption and transport by the small intestine: inhibition by chemical poisons and correlation to the elemental composition of the epithelial cells

1986 ◽  
Vol 44 ◽  
pp. 134-135
Author(s):  
A. J. Tousimis
Keyword(s):  
Small Intestine ◽  
Amino Acid ◽  
Epithelial Cells ◽  
Elemental Composition ◽  
Isotope Labeling ◽  
Structural Components ◽  
X Ray ◽  
Human Small Intestine ◽  
Transport Studies

The elemental composition of amino acids is similar to that of the major structural components of the epithelial cells of the small intestine and other tissues. Therefore, their subcellular localization and concentration measurements are not possible by x-ray microanalysis. Radioactive isotope labeling: I131-tyrosine, Se75-methionine and S35-methionine have been successfully employed in numerous absorption and transport studies. The latter two have been utilized both in vitro and vivo, with similar results in the hamster and human small intestine. Non-radioactive Selenomethionine, since its absorption/transport behavior is assumed to be the same as that of Se75- methionine and S75-methionine could serve as a compound tracer for this amino acid.

The protease phenotype and crystalline nature of the granules of intraepithelial mast cells in Trichinella spiralis-infected mice

1995 ◽  
Vol 53 ◽  
pp. 818-819
Author(s):  
D.S. Friend ◽  
N. Ghildyal ◽  
M.F. Gurish ◽  
K.F. Austen ◽  
R.L. Stevens
Keyword(s):  
Small Intestine ◽  
Mast Cells ◽  
Epithelial Cells ◽  
Lamina Propria ◽  
Trichinella Spiralis ◽  
Intestinal Epithelial ◽  
Carboxypeptidase A ◽  
C3h Mice ◽  
Granule Morphology ◽  
Crystalline Nature

Trichinella spiralis induces a profound mastocytosis and eosinophilia in the small intestine of the infected mouse. Mouse mast cells (MC) store in their granules various combinations of at least five chymotryptic chymases [designated mouse MC protease (mMCP) 1 to 5], two tryptic proteases designated mMCP-6 and mMCP-7 and an exopeptidase, carboxypeptidase A (mMC-CPA). Using antipeptide, protease -specific antibodies to these MC granule proteases, immunohistochemistry was done to determine the distribution, number and protease phenotype of the MCs in the small intestine and spleen 10 to >60 days after Trichinella infection of BALB/c and C3H mice. TEM was performed to evaluate the granule morphology of the MCs between intestinal epithelial cells and in the lamina propria (mucosal MCs) and in the submucosa, muscle and serosa of the intestine (submucosal MCs).As noted in the table below, the number of submucosal MCs remained constant throughout the study. In contrast, on day 14, the number of MCs in the mucosa increased ~25 fold. Increased numbers of MCs were observed between epithelial cells in the mucosal crypts, in the lamina propria and to a lesser extent, between epithelial cells of the intestinal villi.

scholarly journals Protective Effects of Lactoferrin against SARS-CoV-2 Infection In Vitro

Nutrients ◽  
2021 ◽  
Vol 13 (2) ◽  
pp. 328 ◽  
Author(s):  
Claudio Salaris ◽  
Melania Scarpa ◽  
Marina Elli ◽  
Alice Bertolini ◽  
Simone Guglielmetti ◽  
...  
Keyword(s):  
Immune Response ◽  
Epithelial Cells ◽  
Intestinal Epithelial Cells ◽  
Plaque Assay ◽  
Immune Response Gene ◽  
Intestinal Epithelial ◽  
Antiviral Immune Response ◽  
Qrt Pcr ◽  
Anti Inflammatory

SARS-CoV-2 is a newly emerging virus that currently lacks curative treatments. Lactoferrin (LF) is a naturally occurring non-toxic glycoprotein with broad-spectrum antiviral, immunomodulatory and anti-inflammatory effects. In this study, we assessed the potential of LF in the prevention of SARS-CoV-2 infection in vitro. Antiviral immune response gene expression was analyzed by qRT-PCR in uninfected Caco-2 intestinal epithelial cells treated with LF. An infection assay for SARS-CoV-2 was performed in Caco-2 cells treated or not with LF. SARS-CoV-2 titer was determined by qRT-PCR, plaque assay and immunostaining. Inflammatory and anti-inflammatory cytokine production was determined by qRT-PCR. LF significantly induced the expression of IFNA1, IFNB1, TLR3, TLR7, IRF3, IRF7 and MAVS genes. Furthermore, LF partially inhibited SARS-CoV-2 infection and replication in Caco-2 intestinal epithelial cells. Our in vitro data support LF as an immune modulator of the antiviral immune response with moderate effects against SARS-CoV-2 infection.

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