Agonist-induced polarized trafficking and surface expression of the adenosine 2b receptor in intestinal epithelial cells: role of SNARE proteins

2004 ◽  
Vol 287 (5) ◽  
pp. G1100-G1107 ◽  
Author(s):  
Lixin Wang ◽  
Vasantha Kolachala ◽  
Baljit Walia ◽  
Srividya Balasubramanian ◽  
Randy A. Hall ◽  
...  
Keyword(s):  
Adenosine Receptor ◽  
Intestinal Inflammation ◽  
Apical Membrane ◽  
Snare Proteins ◽  
Cell Fractionation ◽  
Camp Synthesis ◽  
Agonist Stimulation ◽  
A2b Receptor ◽  
In Cells

Adenosine, acting through the A2b receptor, induces vectorial chloride and IL-6 secretion in intestinal epithelia and may play an important role in intestinal inflammation. We have previously shown that apical or basolateral adenosine receptor stimulation results in the recruitment of the A2b receptor to the plasma membrane. In this study, we examined domain specificity of recruitment and the role of soluble N-ethylmaleimide (NEM) attachment receptor (SNARE) proteins in the agonist-mediated recruitment of the A2b receptor to the membrane. The colonic epithelial cell line T84 was used because it only expresses the A2b-subtype adenosine receptor. Cell fractionation, biotinylation, and electron microscopic studies showed that the A2b receptor is intracellular at rest and that apical or basolateral adenosine stimulation resulted in the recruitment of the receptor to the apical membrane. Upon agonist stimulation, the A2b receptor is enriched in the vesicle fraction containing vesicle-associated membrane protein (VAMP)-2. Furthermore, in cells stimulated with apical or basolateral adenosine, we demonstrate a complex consisting of VAMP-2, soluble NEM-sensitive factor attachment protein (SNAP)-23, and A2b receptor that is coimmunoprecipitated in cells stimulated with adenosine within 5 min and is no longer detected within 15 min. Inhibition of trafficking with NEM or nocodazole inhibits cAMP synthesis induced by apical or basolateral adenosine by 98 and 90%, respectively. cAMP synthesis induced by foskolin was not affected, suggesting that generalized signaling is not affected under these conditions. Collectively, our data suggest that 1) the A2b receptor is intracellular at rest; 2) apical or basolateral agonist stimulation induces recruitment of the A2b receptor to the apical membrane; 3) the SNARE proteins, VAMP-2 and SNAP-23, participate in the recruitment of the A2b receptor; and 4) the SNARE-mediated recruitment of the A2b receptor may be required for its signaling.

scholarly journals Role of adenosine A1 and A3 receptors in regulation of cardiomyocyte homeostasis after mitochondrial respiratory chain injury

2005 ◽  
Vol 288 (6) ◽  
pp. H2792-H2801 ◽  
Author(s):  
Vladimir Shneyvays ◽  
Dorit Leshem ◽  
Tova Zinman ◽  
Liaman K. Mamedova ◽  
Kenneth A. Jacobson ◽  
...  
Keyword(s):  
Respiratory Chain ◽  
Sodium Azide ◽  
Adenosine Receptor ◽  
Mitochondrial Respiratory Chain ◽  
Receptor Activation ◽  
Atp Level ◽  
Intracellular Ca ◽  
Mitochondrial Respiratory ◽  
In Cells

Activation of either the A1 or the A3 adenosine receptor (A1R or A3R, respectively) elicits delayed cardioprotection against infarction, ischemia, and hypoxia. Mitochondrial contribution to the progression of cardiomyocyte injury is well known; however, the protective effects of adenosine receptor activation in cardiac cells with a respiratory chain deficiency are poorly elucidated. The aim of our study was to further define the role of A1R and A3R activation on functional tolerance after inhibition of the terminal link of the mitochondrial respiratory chain with sodium azide, in a state of normoxia or hypoxia, compared with the effects of the mitochondrial ATP-sensitive K+ channel opener diazoxide. Treatment with 10 mM sodium azide for 2 h in normoxia caused a considerable decrease in the total ATP level; however, activation of adenosine receptors significantly attenuated this decrease. Diazoxide (100 μM) was less effective in protection. During treatment of cultured cardiomyocytes with hypoxia in the presence of 1 mM sodium azide, the A1R agonist 2-chloro- N6-cyclopentyladenosine was ineffective, whereas the A3R agonist 2-chloro- N6-iodobenzyl-5′- N-methylcarboxamidoadenosine (Cl-IB-MECA) attenuated the decrease in ATP level and prevented cell injury. Cl-IB-MECA delayed the dissipation in the mitochondrial membrane potential during hypoxia in cells impaired in the mitochondrial respiratory chain. In cells with elevated intracellular Ca2+ concentration after hypoxia and treatment with NaN3 or after application of high doses of NaN3, Cl-IB-MECA immediately decreased the elevated intracellular Ca2+ concentration toward the diastolic control level. The A1R agonist was ineffective. This may be especially important for the development of effective pharmacological agents, because mitochondrial dysfunction is a leading factor in the pathophysiological cascade of heart disease.

scholarly journals P060 A2B adenosine receptor deficiency ameliorates chronic murine Crohn’s-like-ileitis

2021 ◽  
Vol 15 (Supplement_1) ◽  
pp. S166-S167
Author(s):  
L O Connell ◽  
K Olli ◽  
C Wilkinson ◽  
C Collins ◽  
P Jedlicka ◽  
...  
Keyword(s):  
Chronic Disease ◽  
Chronic Inflammation ◽  
Adenosine Receptor ◽  
Intestinal Inflammation ◽  
Receptor Expression ◽  
A2b Adenosine Receptor ◽  
Murine Colitis ◽  
Cell Counts ◽  
A2b Receptor ◽  
Small Intestinal

Abstract Background The A2B adenosine receptor (Adora2b) has been demonstrated to play an immunomodulatory and epithelial barrier protective role in acute murine colitis. A2B agonist treatment during acute murine colitis has been demonstrated to decrease colonic inflammation and damage suggesting targeting the A2B adenosine receptor may protect the intestinal barrier in acute intestinal inflammation. Crohn’s disease (CD) is associated with chronic inflammation and damage in the small intestine, most often the ileum. To date the role, if any, of the A2B adenosine receptor in the small intestine and in CD is unknown. Methods The TNFΔARE model of Crohn’s-like ileitis was used to investigate a possible role for A2B in small intestinal inflammation. TNFΔARE mice develop discontinuous, transmural inflammation in the distal ileum that progresses with age and is representative of the chronic inflammation observed in CD. Whole body A2B receptor deficient (Adora2b -/-) mice were crossed on to the TNFΔARE background to generate TNFΔARE mice deficient in the A2B adenosine receptor (Adora2b -/-/TNFΔARE+/-). Whole ileum was harvested from Adora2b -/-/TNFΔARE+/- and wildtype TNFΔARE mice for histological analysis during early (6–8 weeks), active (10–12 weeks) and chronic disease (>20 weeks). Histological scoring was performed by a board-certified pathologist blinded to the study. Whole tissue was also harvested for digestion to perform live cell counts and cytokine analysis. Results Histological analysis revealed no significant difference between wildtype and Adora2b deficient TNFΔARE mice during early or active disease. However, during chronic disease Adora2b -/-/TNFΔARE+/- histological scores for active inflammation, chronic inflammation and degree of tissue damage were significantly reduced (approximately 25%) compared to wildtype TNFΔARE mice. This dramatic improvement in histological tissue health in mice with chronic disease was accompanied by an approximately 50% reduction in live cell counts in the ileum of Adora2b -/-/TNFΔARE+/- mice compared to wildtype TNFΔARE mice with a corresponding reduction in pro-inflammatory mediators CXCL1 and IL-1β Conclusion Our findings demonstrate that sustained loss of A2B receptor expression improves the outcome of chronic small intestinal inflammation. Previous studies demonstrate that A2B receptor expression promotes pro-fibrotic responses in chronic lung fibrosis. Our future studies will address the possible functional role of A2B in pro-fibrotic responses associated with chronic intestinal inflammation.

scholarly journals Role of purinergic signaling pathways in V-ATPase recruitment to apical membrane of acidifying epididymal clear cells

2010 ◽  
Vol 298 (4) ◽  
pp. C817-C830 ◽  
Author(s):  
Clémence Belleannée ◽  
Nicolas Da Silva ◽  
Winnie W. C. Shum ◽  
Dennis Brown ◽  
Sylvie Breton
Keyword(s):  
Epithelial Cells ◽  
Kinase Inhibitor ◽  
Apical Membrane ◽  
Purinergic Signaling ◽  
Hydrolysis Product ◽  
Clear Cells ◽  
A2b Receptor ◽  
P1 Receptor ◽  
And Storage

Extracellular purinergic agonists regulate a broad range of physiological functions via P1 and P2 receptors. Using the epididymis as a model system in which luminal acidification is essential for sperm maturation and storage, we show here that extracellular ATP and its hydrolysis product adenosine trigger the apical accumulation of vacuolar H+-ATPase (V-ATPase) in acidifying clear cells. We demonstrate that the epididymis can hydrolyze luminal ATP into other purinergic agonists such as ADP via the activity of nucleotidases located in the epididymal fluid and in the apical membrane of epithelial cells. Alkaline phosphatase activity and abundant ecto-5′-nucleotidase protein were detected in the apical pole of principal cells. In addition, we show that nine nucleotidase genes ( Nt5e, Alpl, Alpp, Enpp1, 2, and 3, and Entpd 2, 4, and 5), seven ATP P2 receptor genes (P2X1, P2X2, P2X3, P2X4, P2X6, P2Y2, P2Y5), and three adenosine P1 receptor genes (A1, A2B, and A3) are expressed in epithelial cells isolated by laser cut microdissection (LCM). The calcium chelator BAPTA-AM abolished the apical V-ATPase accumulation induced by ATP, supporting the contribution of P2X or P2Y in this response. The PKA inhibitor myristoylated protein kinase inhibitor (mPKI) inhibited adenosine-dependent V-ATPase apical accumulation, indicating the participation of the P1 A2B receptor. Altogether, these results suggest that the activation of P1 and P2 purinergic receptors by ATP and adenosine might play a significant role in luminal acidification in the epididymis, a process that is crucial for the establishment of male fertility.

Role of PKC isozymes in water transport in toad urinary bladder

1991 ◽  
Vol 49 ◽  
pp. 302-303
Author(s):  
A.J. Mia ◽  
L.X. Oakford ◽  
T. Yorio
Keyword(s):  
Urinary Bladder ◽  
Apical Membrane ◽  
Membrane Surface ◽  
Protein A ◽  
Cell Types ◽  
Leukemic Cells ◽  
Toad Urinary Bladder ◽  
Pkc Isozymes ◽  
Antibody Labeling

Protein kinase C (PKC) isozymes, when activated, are translocated to particulate membrane fractions for transport to the apical membrane surface in a variety of cell types. Evidence of PKC translocation was demonstrated in human megakaryoblastic leukemic cells, and in cardiac myocytes and fibroblasts, using FTTC immunofluorescent antibody labeling techniques. Recently, we reported immunogold localizations of PKC subtypes I and II in toad urinary bladder epithelia, following 60 min stimulation with Mezerein (MZ), a PKC activator, or antidiuretic hormone (ADH). Localization of isozyme subtypes I and n was carried out in separate grids using specific monoclonal antibodies with subsequent labeling with 20nm protein A-gold probes. Each PKC subtype was found to be distributed singularly and in discrete isolated patches in the cytosol as well as in the apical membrane domains. To determine if the PKC isozymes co-localized within the cell, a double immunogold labeling technique using single grids was utilized.

Role of CD5 expression on TCRγδ T cell-differentiation and development of chronic intestinal inflammation

2001 ◽  
Vol 120 (5) ◽  
pp. A517-A517
Author(s):  
A MIZOGUCHI ◽  
E MIZOGUCHI ◽  
Y DEJONG ◽  
H TAKEDATSU ◽  
F PREFFER ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
T Cell ◽  
Intestinal Inflammation ◽  
T Cell Differentiation ◽  
Chronic Intestinal Inflammation

scholarly journals Role of Dietary Nutritional Treatment on Hepatic and Intestinal Damage in Transplantation with Steatotic and Non-Steatotic Liver Grafts from Brain Dead Donors

Nutrients ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 2554
Author(s):  
Marc Micó-Carnero ◽  
Araní Casillas-Ramírez ◽  
Albert Caballeria-Casals ◽  
Carlos Rojano-Alfonso ◽  
Alfredo Sánchez-González ◽  
...  
Keyword(s):  
Growth Factor ◽  
Gut Microbiota ◽  
Intestinal Inflammation ◽  
Mucosal Damage ◽  
Hepatic Damage ◽  
Intestinal Damage ◽  
Edema Formation ◽  
Steatotic Liver ◽  
Damage And Failure

Herein, we investigate whether: (1) the administration of glucose or a lipid emulsion is useful in liver transplantation (LT) using steatotic (induced genetically or nutritionally) or non-steatotic livers from donors after brain death (DBDs); and (2) any such benefits are due to reductions in intestinal damage and consequently to gut microbiota preservation. In recipients from DBDs, we show increased hepatic damage and failure in the maintenance of ATP, glycogen, phospholipid and growth factor (HGF, IGF1 and VEGFA) levels, compared to recipients from non-DBDs. In recipients of non-steatotic grafts from DBDs, the administration of glucose or lipids did not protect against hepatic damage. This was associated with unchanged ATP, glycogen, phospholipid and growth factor levels. However, the administration of lipids in steatotic grafts from DBDs protected against damage and ATP and glycogen drop and increased phospholipid levels. This was associated with increases in growth factors. In all recipients from DBDs, intestinal inflammation and damage (evaluated by LPS, vascular permeability, mucosal damage, TLR4, TNF, IL1, IL-10, MPO, MDA and edema formation) was not shown. In such cases, potential changes in gut microbiota would not be relevant since neither inflammation nor damage was evidenced in the intestine following LT in any of the groups evaluated. In conclusion, lipid treatment is the preferable nutritional support to protect against hepatic damage in steatotic LT from DBDs; the benefits were independent of alterations in the recipient intestine.

scholarly journals Helminth Interactions with Bacteria in the Host Gut Are Essential for Its Immunomodulatory Effect

2021 ◽  
Vol 9 (2) ◽  
pp. 226
Author(s):  
Milan Jirků ◽  
Zuzana Lhotská ◽  
Lucia Frgelecová ◽  
Oldřiška Kadlecová ◽  
Klára Judita Petrželková ◽  
...  
Keyword(s):  
Protective Effect ◽  
Intestinal Inflammation ◽  
Hymenolepis Diminuta ◽  
Clinical Parameters ◽  
Th2 Response ◽  
Histological Changes ◽  
Immunological Markers ◽  
Bacterial Microbiota ◽  
Inflammatory Effect

Colonization by the benign tapeworm, Hymenolepis diminuta, has been associated with a reduction in intestinal inflammation and changes in bacterial microbiota. However, the role of microbiota in the tapeworm anti-inflammatory effect is not yet clear, and the aim of this study was to determine whether disruption of the microflora during worm colonization can affect the course of intestinal inflammation. We added a phase for disrupting the intestinal microbiota using antibiotics to the experimental design for which we previously demonstrated the protective effect of H. diminuta. We monitored the immunological markers, clinical parameters, bacterial microbiota, and histological changes in the colon of rats. After a combination of colonization, antibiotics, and colitis induction, we had four differently affected experimental groups. We observed a different course of the immune response in each group, but no protective effect was found. Rats treated with colonization and antibiotics showed a strong induction of the Th2 response as well as a significant change in microbial diversity. The microbial results also revealed differences in the richness and abundance of some bacterial taxa, influenced by various factors. Our data suggest that interactions between the tapeworm and bacteria may have a major impact on its protective effect.

scholarly journals 5-LO-derived LTB4 plays a key role in MCP-1 expression in HMGB1-exposed VSMCs via a BLTR1 signaling axis

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Jong Min Choi ◽  
Seung Eun Baek ◽  
Ji On Kim ◽  
Eun Yeong Jeon ◽  
Eun Jeong Jang ◽  
...  
Keyword(s):  
Tissue Injury ◽  
Vascular Inflammation ◽  
High Mobility ◽  
Leukotriene B4 ◽  
Monocyte Chemoattractant Protein 1 ◽  
Signaling Axis ◽  
Cellular Source ◽  
Leukotriene Receptor ◽  
In Cells

AbstractMonocyte chemoattractant protein-1 (MCP-1) plays an important role in initiating vascular inflammation; however, its cellular source in the injured vasculatures is unclear. Given the importance of high mobility group box 1 (HMGB1) in tissue injury, we investigated the role of vascular smooth muscle cells (VSMCs) in MCP-1 production in response to HMGB1. In primary cultured rat aortic VSMCs stimulated with HMGB1, the expression of MCP-1 and 5-lipoxygenase (LO) was increased. The increased MCP-1 expression in HMGB1 (30 ng/ml)-stimulated cells was significantly attenuated in 5-LO-deficient cells as well as in cells treated with zileuton, a 5-LO inhibitor. Likewise, MCP-1 expression and production were also increased in cells stimulated with exogenous leukotriene B4 (LTB4), but not exogenous LTC4. LTB4-induced MCP-1 expression was attenuated in cells treated with U75302, a LTB4 receptor 1 (BLTR1) inhibitor as well as in BLTR1-deficient cells, but not in 5-LO-deficient cells. Moreover, HMGB1-induced MCP-1 expression was attenuated in BLTR1-deficient cells or by treatment with a BLTR1 inhibitor, but not other leukotriene receptor inhibitors. In contrast to MCP-1 expression in response to LTB4, the increased MCP-1 production in HMGB1-stimulated VSMC was markedly attenuated in 5-LO-deficient cells, indicating a pivotal role of LTB4-BLTR1 signaling in MCP-1 expression in VSMCs. Taken together, 5-LO-derived LTB4 plays a key role in MCP-1 expression in HMGB1-exposed VSMCs via BLTR1 signaling, suggesting the LTB4-BLTR1 signaling axis as a potential therapeutic target for vascular inflammation in the injured vasculatures.

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