scholarly journals PACSIN2-dependent apical endocytosis regulates the morphology of epithelial microvilli

2019 ◽  
Vol 30 (19) ◽  
pp. 2515-2526 ◽  
Author(s):  
Meagan M. Postema ◽  
Nathan E. Grega-Larson ◽  
Leslie M. Meenderink ◽  
Matthew J. Tyska
Keyword(s):  
Intestinal Epithelial Cell ◽  
Intestinal Epithelial Cells ◽  
Intestinal Epithelial ◽  
Endocytic Vesicle ◽  
Bar Domain ◽  
Binding Partner ◽  
Epithelial Cell Lines ◽  
Brush Borders ◽  
Endocytic Machinery ◽  
The Impact

Apical microvilli are critical for the homeostasis of transporting epithelia, yet mechanisms that control the assembly and morphology of these protrusions remain poorly understood. Previous studies in intestinal epithelial cell lines suggested a role for the F-BAR domain protein PACSIN2 in normal microvillar assembly. Here we report the phenotype of PACSIN2 KO mice and provide evidence that through its role in promoting apical endocytosis, this molecule plays a role in controlling microvillar morphology. PACSIN2 KO enterocytes exhibit reduced numbers of microvilli and defects in the microvillar ultrastructure, with membranes lifting away from rootlets of core bundles. Dynamin2, a PACSIN2 binding partner, and other endocytic factors were also lost from their normal localization near microvillar rootlets. To determine whether loss of endocytic machinery could explain defects in microvillar morphology, we examined the impact of PACSIN2 KD and endocytosis inhibition on live intestinal epithelial cells. These assays revealed that when endocytic vesicle scission fails, tubules are pulled into the cytoplasm and this, in turn, leads to a membrane-lifting phenomenon reminiscent of that observed at PACSIN2 KO brush borders. These findings lead to a new model where inward forces generated by endocytic machinery on the plasma membrane control the membrane wrapping of cell surface protrusions.

scholarly journals PACSIN2-dependent apical endocytosis regulates the morphology of epithelial microvilli

2019 ◽  
Author(s):  
Meagan M. Postema ◽  
Nathan E. Grega-Larson ◽  
Leslie M. Meenderink ◽  
Matthew J. Tyska
Keyword(s):  
Critical Role ◽  
Intestinal Epithelial ◽  
Functional Surface ◽  
Bar Domain ◽  
Binding Partner ◽  
Intestinal Brush Border ◽  
Epithelial Cell Lines ◽  
Brush Borders ◽  
Endocytic Machinery ◽  
The Impact

ABSTRACTApical microvilli are critical for the homeostasis of transporting epithelia, yet mechanisms that control the assembly and morphology of these protrusions remain poorly understood. Previous studies in intestinal epithelial cell lines suggested a role for F-BAR domain protein PACSIN2 in normal microvillar assembly. Here we report the phenotype of PACSIN2 KO mice and provide evidence that through its role in promoting apical endocytosis, this molecule functions in controlling microvillar morphology. PACSIN2 KO enterocytes exhibit reduced numbers of microvilli and defects in microvillar ultrastructure, with membranes lifting away from rootlets of core bundles. Dynamin2, a PACSIN2 binding partner, and other endocytic factors were also lost from their normal localization near microvillar rootlets. To determine if loss of endocytic machinery could explain defects in microvillar morphology, we examined the impact of PACSIN2 KD and endocytosis inhibition on live intestinal epithelial cells. These assays revealed that when endocytic vesicle scission fails, tubules are pulled into the cytoplasm and this, in turn, leads to a membrane lifting phenomenon reminiscent of that observed in PACSIN2 KO brush borders. These findings lead to a new model where inward forces generated by endocytic machinery on the plasma membrane control the membrane wrapping of cell surface protrusions.Highlight for TOCApical microvilli increase the functional surface area of transporting epithelia. Here we report that the F-BAR domain-containing protein PACSIN2, through its ability to promote apical endocytosis, plays a critical role in controlling the morphology of intestinal brush border microvilli.

scholarly journals Lactobacillus acidophilus Induces Cytokine and Chemokine Production via NF-κB and p38 Mitogen-Activated Protein Kinase Signaling Pathways in Intestinal Epithelial Cells

2012 ◽  
Vol 19 (4) ◽  
pp. 603-608 ◽  
Author(s):  
Yujun Jiang ◽  
Xuena Lü ◽  
Chaoxin Man ◽  
Linlin Han ◽  
Yi Shan ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
Cell Lines ◽  
Intestinal Epithelial Cell ◽  
Intestinal Epithelial Cells ◽  
Intestinal Epithelial ◽  
Chemokine Production ◽  
Content Type ◽  
Cytokines And Chemokines ◽  
Epithelial Cell Lines

ABSTRACTIntestinal epithelial cells can respond to certain bacteria by producing an array of cytokines and chemokines which are associated with host immune responses.Lactobacillus acidophilusNCFM is a characterized probiotic, originally isolated from human feces. This study aimed to test the ability ofL. acidophilusNCFM to stimulate cytokine and chemokine production in intestinal epithelial cells and to elucidate the mechanisms involved in their upregulation. In experiments using intestinal epithelial cell lines and mouse models, we observed thatL. acidophilusNCFM could rapidly but transiently upregulate a number of effector genes encoding cytokines and chemokines such as interleukin 1α (IL-1α), IL-1β, CCL2, and CCL20 and that cytokines showed lower expression levels withL. acidophilusNCFM treatment than chemokines. Moreover,L. acidophilusNCFM could activate a pathogen-associated molecular pattern receptor, Toll-like receptor 2 (TLR2), in intestinal epithelial cell lines. The phosphorylation of NF-κB p65 and p38 mitogen-activated protein kinase (MAPK) in intestinal epithelial cell lines was also enhanced byL. acidophilusNCFM. Furthermore, inhibitors of NF-κB (pyrrolidine dithiocarbamate [PDTC]) and p38 MAPK (SB203580) significantly reduced cytokine and chemokine production in the intestinal epithelial cell lines stimulated byL. acidophilusNCFM, suggesting that both NF-κB and p38 MAPK signaling pathways were important for the production of cytokines and chemokines induced byL. acidophilusNCFM.

IFN-gamma modulates CD1d surface expression on intestinal epithelia

1996 ◽  
Vol 271 (1) ◽  
pp. C276-C283 ◽  
Author(s):  
S. P. Colgan ◽  
V. M. Morales ◽  
J. L. Madara ◽  
J. E. Polischuk ◽  
S. P. Balk ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Cell Lines ◽  
Intestinal Epithelial Cell ◽  
Surface Expression ◽  
Time Dependent ◽  
Intestinal Epithelial ◽  
Ifn Gamma ◽  
Intestinal Epithelia ◽  
Epithelial Cell Lines ◽  
The Impact

In vivo, epithelial cells that line the intestine are intimately associated with lymphocytes, termed intestinal intraepithelial lymphocytes (iIEL). A putative ligand for iIEL on intestinal epithelial cells is CD1d, and recent studies demonstrate a surface form of this molecule exists on intestinal epithelia. At present, it is not known whether CD1d expression is regulated by cytokines in the intestinal microenvironment. Thus we examined the impact of relevant cytokines on CD1d at the level of mRNA and cell surface expression. Using a sensitive whole cell enzyme-linked immunosorbent assay, we assessed the impact of relevant cytokines on CD1d expression on intestinal epithelial cell lines. We were readily able to detect CD1d on the surface of T84 cells, a cryptlike intestinal epithelial cell line. Epithelial cell exposure to human recombinant interferon-gamma (IFN-gamma) resulted in increased CD1d expression in a dose- and time-dependent manner. Polymerase chain reaction amplification of CD1d cDNA revealed a time-dependent induction after exposure to IFN-gamma. This IFN-gamma effect on CD1d expression was cytokine specific and was evident with epithelial cell lines other than T84, including Caco-2 and HT-29 cells. Finally, we were not able to detect significant surface expression of CD1a, CD1b, or CD1c on intestinal epithelial cell lines in the presence or absence of relevant cytokines. These results indicate that CD1d cell surface protein and cellular mRNA, like other major histocompatibility complex-related molecules, is cytokine regulated in intestinal epithelial cell lines.

scholarly journals Gamma Interferon Augments the Intracellular Pathway for Lipopolysaccharide (LPS) Recognition in Human Intestinal Epithelial Cells through Coordinated Up-Regulation of LPS Uptake and Expression of the Intracellular Toll-Like Receptor 4-MD-2 Complex

2003 ◽  
Vol 71 (6) ◽  
pp. 3503-3511 ◽  
Author(s):  
Manabu Suzuki ◽  
Tadakazu Hisamatsu ◽  
Daniel K. Podolsky
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
Cell Lines ◽  
Intestinal Epithelial Cell ◽  
Intestinal Epithelial Cells ◽  
Intestinal Epithelial ◽  
Toll Like Receptor ◽  
Toll Like Receptor 4 ◽  
Epithelial Cell Lines ◽  
Ifn Γ

ABSTRACT Although some intestinal epithelial cell lines are known to respond to lipopolysaccharide (LPS), understanding of the relationship between LPS responsiveness and the expression of LPS receptors or factors regulating LPS responsiveness of intestinal epithelial cell lines is incomplete. In this study, we demonstrate that commonly studied human intestinal epithelial cell lines can be classified into at least three different types on the basis of LPS responsiveness, Toll-like receptor-4 (TLR4) expression, and the effects of gamma interferon (IFN-γ) on LPS responsiveness. The first phenotype, which includes the HCT-116 and Caco-2 cell lines, is characterized by relative hyporesponsiveness to LPS and diminished expression of TLR4 protein. In these cells, IFN-γ does not induce LPS responsiveness. The second phenotype, which includes cell line SW480, exhibits a highly LPS-responsive phenotype and surface expression of TLR4 protein even in unprimed conditions. These lines are functionally similar to cells of monocytic lineage. In the third phenotype, which includes the HT-29 and Colo205 cell lines, TLR4 protein is largely present in the cytoplasmic fraction and the cells are hyporesponsive to LPS in an unprimed condition. However, priming of these cells with IFN-γ can induce LPS responsiveness through augmentation of LPS uptake and expression of MD-2 mRNA and intracellular TLR4 proteins. Finally, these findings suggest that the Th1 cytokine IFN-γ modulates LPS responsiveness through several mechanisms in intestinal epithelial cells and that these cells may comprise different subpopulations with distinct roles in innate immune responses.

scholarly journals Functional screen of Inflammatory Bowel Disease genes reveals key epithelial functions

2021 ◽  
Author(s):  
Jessy Carol Ntunzwenimana ◽  
Gabrielle Boucher ◽  
Jean Paquette ◽  
Hugues Gosselin ◽  
Azadeh Alikashani ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
Intestinal Epithelial Cell ◽  
Intestinal Epithelial Cells ◽  
Intestinal Epithelial ◽  
Biological Functions ◽  
Functional Screen ◽  
Major Cluster ◽  
Causal Genes ◽  
The Impact

Background: Genetic studies have been tremendously successful in identifying genomic regions associated with a wide variety of phenotypes, although the success of these studies in identifying causal genes, their variants, and their functional impacts have been more limited. Methods: We identified 145 genes from IBD-associated genomic loci having endogenous expression within the intestinal epithelial cell compartment. We evaluated the impact of lentiviral transfer of the open reading frame (ORF) of these IBD genes into the HT-29 intestinal epithelial cell line via transcriptomic analyses. Comparing the genes whose expression was modulated by each ORF, as well as the functions enriched within these gene lists, identified ORFs with shared impacts and their putative disease-relevant biological functions. Results: Analysis of the transcriptomic data for cell lines expressing the ORFs for known causal genes such as HNF4a, IFIH1 and SMAD3 identified functions consistent for what is known for these genes. These analyses also identified two major cluster of genes: Cluster 1 contained the known IBD causal genes IFIH1, SBNO2, NFKB1 and NOD2, as well as genes from other IBD loci (ZFP36L1, IRF1, GIGYF1, OTUD3, AIRE and PITX1), whereas Cluster 2 contained the known causal gene KSR1 and implicated DUSP16 from another IBD locus. Our analyses highlight how multiple IBD gene candidates impact on epithelial structure and function, including the protection of the mucosa from intestinal microbiota, and demonstrate that DUSP16, acts a regulator of MAPK activity and contributes to mucosal defense, in part via its regulation of the polymeric immunoglobulin receptor, involved in the protection of the intestinal mucosa from enteric microbiota. Conclusions: This functional screen, based on expressing IBD genes within an appropriate cellular context, in this instance intestinal epithelial cells, resulted in changes to the cell's transcriptome that are relevant to their endogenous biological function(s). This not only helped in identifying likely causal genes within genetic loci but also provided insight into their biological functions. Furthermore, this work has highlighted the central role of intestinal epithelial cells in IBD pathophysiology, providing a scientific rationale for a drug development strategy that targets epithelial functions in addition to the current therapies targeting immune functions.

scholarly journals Functional screen of inflammatory bowel disease genes reveals key epithelial functions

2021 ◽  
Vol 13 (1) ◽  
Author(s):  
Jessy Carol Ntunzwenimana ◽  
Gabrielle Boucher ◽  
Jean Paquette ◽  
Hugues Gosselin ◽  
Azadeh Alikashani ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
Intestinal Epithelial Cell ◽  
Intestinal Epithelial Cells ◽  
Intestinal Epithelial ◽  
Biological Functions ◽  
Functional Screen ◽  
Endogenous Expression ◽  
Causal Genes ◽  
The Impact

Abstract Background Genetic studies have been tremendously successful in identifying genomic regions associated with a wide variety of phenotypes, although the success of these studies in identifying causal genes, their variants, and their functional impacts has been more limited. Methods We identified 145 genes from IBD-associated genomic loci having endogenous expression within the intestinal epithelial cell compartment. We evaluated the impact of lentiviral transfer of the open reading frame (ORF) of these IBD genes into the HT-29 intestinal epithelial cell line via transcriptomic analyses. By comparing the genes in which expression was modulated by each ORF, as well as the functions enriched within these gene lists, we identified ORFs with shared impacts and their putative disease-relevant biological functions. Results Analysis of the transcriptomic data for cell lines expressing the ORFs for known causal genes such as HNF4a, IFIH1, and SMAD3 identified functions consistent with what is already known for these genes. These analyses also identified two major clusters of genes: Cluster 1 contained the known IBD causal genes IFIH1, SBNO2, NFKB1, and NOD2, as well as genes from other IBD loci (ZFP36L1, IRF1, GIGYF1, OTUD3, AIRE and PITX1), whereas Cluster 2 contained the known causal gene KSR1 and implicated DUSP16 from another IBD locus. Our analyses highlight how multiple IBD gene candidates can impact on epithelial structure and function, including the protection of the mucosa from intestinal microbiota, and demonstrate that DUSP16 acts a regulator of MAPK activity and contributes to mucosal defense, in part via its regulation of the polymeric immunoglobulin receptor, involved in the protection of the intestinal mucosa from enteric microbiota. Conclusions This functional screen, based on expressing IBD genes within an appropriate cellular context, in this instance intestinal epithelial cells, resulted in changes to the cell’s transcriptome that are relevant to their endogenous biological function(s). This not only helped in identifying likely causal genes within genetic loci but also provided insight into their biological functions. Furthermore, this work has highlighted the central role of intestinal epithelial cells in IBD pathophysiology, providing a scientific rationale for a drug development strategy that targets epithelial functions in addition to the current therapies targeting immune functions.

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.

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