Regulation of human jejunal transmucosal resistance and MLC phosphorylation by Na+-glucose cotransport

2001 ◽  
Vol 281 (6) ◽  
pp. G1487-G1493 ◽  
Author(s):  
Jessica J. Berglund ◽  
Martin Riegler ◽  
Yevgeny Zolotarevsky ◽  
Etienne Wenzl ◽  
Jerrold R. Turner
Keyword(s):  
Cell Lines ◽  
Critical Role ◽  
Paracellular Permeability ◽  
Intestinal Epithelial ◽  
Physiological Regulation ◽  
Ussing Chambers ◽  
Epithelial Cell Lines ◽  
Double Label ◽  
Mlc Phosphorylation

Na+- nutrient cotransport-dependent regulation of paracellular permeability has been demonstrated in rodent intestine and human intestinal epithelial cell lines. In cell lines this regulation is associated with phosphorylation of myosin II regulatory light chain (MLC). However, the subcellular localization of phosphorylated MLC during this regulation has not been studied and regulation of paracellular permeability and MLC phosphorylation has not been studied in isolated human intestine. To evaluate these events in human jejunum, isolated mucosa was mounted in Ussing chambers, characterized electrophysiologically, and then immunostained using anti-phosphorylated MLC and anti-total MLC antisera. MLC phosphorylation was assessed by calculating the ratio of anti-phosphorylated MLC signal to anti-total MLC signal within defined regions. Transmucosal resistance of mucosae without active Na+-glucose cotransport was 37 ± 3% greater than that of mucosae with active Na+-glucose cotransport within 15 min. Quantitative double-label immunofluorescence showed that the phosphorylated MLC-to-total MLC ratio increased by 45 ± 4% within the perijunctional actomyosin ring when Na+-glucose cotransport was active. Thus regulation of transmucosal resistance by Na+-glucose cotransport is accompanied by increased MLC phosphorylation within the perijunctional actomyosin ring. These data support the proposed critical role of the perijunctional cytoskeleton in physiological regulation of human small intestinal paracellular permeability.

Asymmetrical targeting of type II Na-Picotransporters in renal and intestinal epithelial cell lines

2000 ◽  
Vol 278 (3) ◽  
pp. F361-F368 ◽  
Author(s):  
N. Hernando ◽  
S. Sheikh ◽  
Z. Karim-Jimenez ◽  
H. Galliker ◽  
J. Forgo ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Cell Lines ◽  
Fluorescent Protein ◽  
Intestinal Epithelial ◽  
Type Ii ◽  
Opossum Kidney ◽  
Epithelial Cell Lines ◽  
Molecular Determinants ◽  
Cellular Context ◽  
Green Fluorescent

Targeting of newly synthesized transporters to either the apical or basolateral domains of polarized cells is crucial for the function of epithelia, such as in the renal proximal tubule or in the small intestine. Recently, different sodium-phosphate cotransporters have been identified. Type II cotransporters can be subdivided into two groups: type IIa and type IIb. Type IIa is predominantly expressed in renal proximal tubules, whereas type IIb is located on the intestinal and lung epithelia. To gain some insights into the polarized targeting of the type II cotransporters, we have transiently expressed type IIa and type IIb cotransporters in several epithelial cell lines: two lines derived from renal proximal cells (opossum kidney and LLC-PK1), one from renal distal cells (Madin-Darby canine kidney), and one from colonic epithelium (CaCo-2). We studied the expression of the transporters fused to the enhanced green fluorescent protein. Our data indicate that the polarized targeting is dependent on molecular determinants most probably located at the COOH terminus of the cotransporters as well as on the cellular context.

scholarly journals Drosophila Gut—A Nexus Between Dietary Restriction and Lifespan

2018 ◽  
Vol 19 (12) ◽  
pp. 3810 ◽  
Author(s):  
Ting Lian ◽  
Qi Wu ◽  
Brian Hodge ◽  
Kenneth Wilson ◽  
Guixiang Yu ◽  
...  
Keyword(s):  
Digestive Tract ◽  
Dietary Restriction ◽  
Healthy Aging ◽  
Fat Body ◽  
Critical Role ◽  
Vital Role ◽  
Intestinal Epithelial ◽  
Beneficial Effects ◽  
Accumulation Of Damage

Aging is often defined as the accumulation of damage at the molecular and cellular levels which, over time, results in marked physiological impairments throughout the organism. Dietary restriction (DR) has been recognized as one of the strongest lifespan extending therapies observed in a wide array of organisms. Recent studies aimed at elucidating how DR promotes healthy aging have demonstrated a vital role of the digestive tract in mediating the beneficial effects of DR. Here, we review how dietary restriction influences gut metabolic homeostasis and immune function. Our discussion is focused on studies of the Drosophila digestive tract, where we describe in detail the potential mechanisms in which DR enhances maintenance of the intestinal epithelial barrier, up-regulates lipid metabolic processes, and improves the ability of the gut to deal with damage or stress. We also examine evidence of a tissue-tissue crosstalk between gut and neighboring organs including brain and fat body. Taken together, we argue that the Drosophila gut plays a critical role in DR-mediated lifespan extension.

scholarly journals Thymus leukemia antigen controls intraepithelial lymphocyte function and inflammatory bowel disease

2008 ◽  
Vol 105 (46) ◽  
pp. 17931-17936 ◽  
Author(s):  
Danyvid Olivares-Villagómez ◽  
Yanice V. Mendez-Fernandez ◽  
Vrajesh V. Parekh ◽  
Saif Lalani ◽  
Tiffaney L. Vincent ◽  
...  
Keyword(s):  
Inflammatory Bowel Disease ◽  
Bowel Disease ◽  
Intestinal Inflammation ◽  
Genetic Model ◽  
Critical Role ◽  
Intraepithelial Lymphocytes ◽  
Lymphocyte Function ◽  
Intestinal Epithelial ◽  
Inflammatory Bowel

Intestinal intraepithelial lymphocytes (IEL) bear a partially activated phenotype that permits them to rapidly respond to antigenic insults. However, this phenotype also implies that IEL must be highly controlled to prevent misdirected immune reactions. It has been suggested that IEL are regulated through the interaction of the CD8αα homodimer with the thymus leukemia (TL) antigen expressed by intestinal epithelial cells. We have generated and characterized mice genetically-deficient in TL expression. Our findings show that TL expression has a critical role in maintaining IEL effector functions. Also, TL deficiency accelerated colitis in a genetic model of inflammatory bowel disease. These findings reveal an important regulatory role of TL in controlling IEL function and intestinal inflammation.

C/EBPb Is a Critical Mediator of Resistance to IMiD® Immunomodulatory Compounds and Affected by IMiD Compounds Via Control of Protein Translation

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 135-135
Author(s):  
Shirong Li ◽  
Rekha Pal ◽  
Sara Monaghan ◽  
Peter Schafer ◽  
Hongjiao Ouyang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Proliferation ◽  
Cell Lines ◽  
Translational Control ◽  
Critical Role ◽  
Protein Translation ◽  
Thymidine Uptake ◽  
Double Labeling ◽  
Equity Ownership

Abstract Abstract 135 Background: Lenalidomide and pomalidomide are IMiD® immunomodulatory compounds that have been shown to be highly active in the treatment of multiple myeloma (MM). IMiD compounds exert their anti-tumor effects via acting on costimulatory proteins of T cells and NK cells, augmenting both the adaptive and innate immune system. But the mechanisms by which IMiD compounds directly inhibit MM cell proliferation are still unclear. Here we focused on the direct effects of IMiD compounds alone on MM cells. Results and Methods: We found that IMiDs, at concentrations as low as 0.01 μ M, induce significant inhibition of DNA synthesis in MM cells as shown by thymidine uptake. Since our previous work demonstrated that C/EBPβ is an important transcription factor which controls the growth and proliferation of myeloma cells, we analyzed the effects of IMiD compounds on C/EBPβ. We found that both pomalidomide and lenalidomide significantly decreased the protein level of C/EBPβ LAP-isoform in MM cell lines and primary MM cells. IMiD compound-induced suppression of C/EBPβ protein expression led to impaired transcription of the downstream IRF4, and subsequently to downregulation of BLIMP1 and XBP1, which are all critical for MM survival. To confirm our findings in vivo, we analyzed IRF4 expression by double labeling (IRF4+/CD138+) immunohistochemical staining of bone marrow biopsy samples of 23 myeloma patients prior to therapy and during therapy with lenalidomide. During lenalidomide therapy, the bone marrow MM cells showed a significantly weaker staining intensity for IRF4 in comparison to prior therapy. This was quantified by a significant (p<0.001) decrease of the staining score from 176 to 152, respectively. To confirm the critical role of C/EBPβ in MM we stably overexpressed C/EBPβ in MM cells. Overexpression of C/EBPβ prevented IMiD compound-induced inhibition of MM cell proliferation, indicating that C/EBPβ is critical in mediating resistance to IMiD compounds. This was supported by the fact that C/EBPβ was not down regulated in IMiD-resistant cell lines by IMiD treatment. Dissection of the C/EBPβ protein regulation revealed that IMiD compounds shut down C/EBPβ protein translation by decreasing eIF-4E. Knockdown experiments of eIF-4e resulted in downregulation of C/EBPβ, suggesting that C/EBPβ is under translational control in MM. Conclusions: Our studies, for the first time, provide evidence that IMiD compounds inhibit MM cell proliferation and survival by affecting the translation of C/EBPβ and subsequently multiple downstream transcription factors including IRF4, BLIMP1 and XBP1. Due to the critical role of C/EBPβ in mediating effects of IMiD compounds in MM, it might be a target to overcome drug resistance to IMiD compounds. The fact that pomalidomide can overcome resistance to lenalidomide in MM requires still further evaluation. Disclosures: Schafer: Celgene Corporation: Employment, Equity Ownership. Mapara:Gentium: Equity Ownership. Lentzsch:Celgene Corp: Research Funding.

Potentiation of bortezomib's Anti-Myeloma Effects in Combination with Anti-DR5 Agonistic Antibody: The Role of ER Stress in DR5 Editing and DR-Mediated Apoptotic Signaling

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 5104-5104
Author(s):  
Hirokazu Miki ◽  
Masahiro Abe ◽  
Kumiko Kagawa ◽  
Asuka Oda ◽  
Hiroe Amou ◽  
...  
Keyword(s):  
Er Stress ◽  
Cell Lines ◽  
Critical Role ◽  
Protein Translation ◽  
Chemotherapeutic Agents ◽  
Mrna Levels ◽  
Trail Receptor ◽  
Normal Peripheral Blood ◽  
Agonistic Antibody

Abstract Abstract 5104 Bortezomib (BTZ) is widely used in the treatment of myeloma (MM) with marked response rates in both relapsed/refractory as well as newly diagnosed MM. However, significant numbers of patients still remain outside benefit of the BTZ treatment; and various combinatory treatments with BTZ have been implemented to improve BTZ's anti-MM effects. On the other hand, immunotherapies seem attractive for yet incurable malignancies by chemotherapeutic agents such as MM and their clinical application has been studied. One such approach is a TNF-related apoptosis-inducing ligand (TRAIL)-mediated immunotherapy. In the present study, we therefore explored the role of BTZ on TRAIL receptor editing and its downstream signaling with special reference to endoplasmic reticulum (ER) stress and the cytotoxic effects of BTZ and anti-TRAIL receptor agonistic antibody in combination on MM cells. Most MM cells expressed DR4 but weakly DR5, while normal peripheral blood mononuclear cells expressed neither DR4 nor DR5. BTZ at 10 nM markedly up-regulated the surface levels of DR5 and its mRNA expression but not those of DR4 in MM cell lines and primary MM cells. Furthermore, BTZ decreased the levels of c-FLIP, an inhibitor of DISC, along with activation of caspase-8 and caspase-3, suggesting potentiation of the DR-mediated extrinsic apoptotic pathway. Consistently, BTZ and anti-DR5 agonistic antibody cooperatively enhanced the cytotoxicity against MM cells. BTZ induced phosphorylation of eIF2alpha, ATF4 and CHOP, along with disappearance of anti-apoptotic proteins including Mcl-1 in MM cells, suggesting the enhancement of ER stress and subsequent suppression of protein translation by BTZ. However, such induction of ER stress by BTZ was not observed in BTZ-resistant MM cell lines, KMS-11/BTZ and OPM-2/BTZ, with a point mutation in BTZ-binding proteasome beta5 subunit (Ri et al. Leukemia 2010). In KMS-11/BTZ and OPM-2/BTZ, surface protein as well as mRNA levels of DR5 were not up-regulated by BTZ, suggesting a critical role of ER stress in up-regulation of DR5 expression by BTZ. Because DR5 expression has been shown to be transcriptionally up-regulated by CHOP, the up-regulation of DR5 mRNA and protein in MM cells by BTZ is suggested to be at least in part due to CHOP induced by BTZ-mediated ER stress. Although BTZ exerts its anti-MM effects through induction of ER stress, the present study demonstrates that induction of ER stress by BTZ is also able to sensitize MM cells to TRAIL-mediated immunotherapy. Therefore, the combination of BTZ and TRAIL-mediated immunotherapy is warranted for further study. Disclosures: No relevant conflicts of interest to declare.

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