scholarly journals Proteomic exploration of pancreatic islets in mice null for the α2A adrenergic receptor

2005 ◽  
Vol 35 (1) ◽  
pp. 73-88 ◽  
Author(s):  
Xinran Hu ◽  
David Friedman ◽  
Salisha Hill ◽  
Richard Caprioli ◽  
Wendell Nicholson ◽  
...  
Keyword(s):  
Insulin Release ◽  
Adrenergic Receptor ◽  
Ex Vivo ◽  
Wild Type ◽  
Downstream Signaling ◽  
Glucose Levels ◽  
Endogenous Catecholamines ◽  
Stimulation Of

The present studies extend recent findings that mice null for the α2A adrenergic receptor (α2A AR KO mice) lack suppression of exogenous secretagogue-stimulated insulin secretion in response to α2 AR agonists by evaluating the endogenous secretagogue, glucose, ex vivo, and providing in vivo data that baseline insulin levels are elevated and baseline glucose levels are decreased in α2A AR KO mice. These latter findings reveal that the α2A AR subtype regulates glucose-stimulated insulin release in response to endogenous catecholamines in vivo. The changes in α2A AR responsiveness and resultant changes in insulin/glucose homeostasis encouraged us to utilize proteomics strategies to identify possible α2A AR downstream signaling molecules or other resultant changes due to perturbation of α2A AR expression. Although agonist stimulation of islets from wild type (WT) mice did not significantly alter islet protein profiles, several proteins were enriched in islets from α2A AR KO mice when compared with those from WT mice, including an enzyme participating in insulin protein processing. The present studies document the important role of the α2A AR subtype in tonic suppression of insulin release in response to endogenous catecholamines as well as exogenous α2 agonists and provide insights into pleiotropic changes that result from loss of α2A AR expression and tonic suppression of insulin release.

scholarly journals Endogenous Somatostatin Is Critical in Regulating the Acute Effects of l-Arginine on Growth Hormone and Insulin Release in Mice

Endocrinology ◽  
2013 ◽  
Vol 154 (7) ◽  
pp. 2393-2398 ◽  
Author(s):  
Jose Córdoba-Chacón ◽  
Manuel D. Gahete ◽  
Ana I. Pozo-Salas ◽  
Justo P. Castaño ◽  
Rhonda D. Kineman ◽  
...  
Keyword(s):  
Insulin Release ◽  
Fold Increase ◽  
Second Phase ◽  
Acute Effects ◽  
Wild Type ◽  
Β Cells ◽  
Glucose Levels ◽  
First Phase Insulin Secretion

Abstract l-arginine (l-Arg) rapidly stimulates GH and insulin release in vivo. It has been hypothesized that l-Arg stimulates GH release by lowering hypothalamic somatostatin (SST) tone. l-Arg may also act directly at the pituitary to stimulate GH release. Moreover, l-Arg has a direct stimulatory effect on β-cells, which is thought to be blunted by the release of SST from pancreatic δ-cells. To confirm the role of endogenous SST on l-Arg-induced GH and insulin release, wild-type (WT) and SST-knockout (SST-KO) mice were injected with l-Arg (ip; 0.8 g/kg), and pre-/post-injection GH, insulin, and glucose levels were measured. In WT mice, l-Arg evoked a 6-fold increase in circulating GH. However, there was only a modest increase in GH levels in WT pituitary cell cultures treated with l-Arg. In contrast, l-Arg failed to increase GH in SST-KO beyond their already elevated levels. These results further support the hypothesis that the primary mechanism by which l-Arg acutely increases GH in vivo is by lowering hypothalamic SST input to the pituitary and not via direct pituitary effects. Additionally, l-Arg induced a clear first-phase insulin secretion in WT mice, but not in SST-KO. However, SST-KO, but not WT mice, displayed a robust and sustained second-phase insulin release. These results further support a role for endogenous SST in regulating l-Arg-mediated insulin release.

Abstract P283: Parkin Mediates Mitophagy in Cardioprotection

2011 ◽  
Vol 109 (suppl_1) ◽  
Author(s):  
Allen M Andres ◽  
Chengqun Huang ◽  
Eric P Ratliff ◽  
Genaro Hernandez ◽  
Pamela Lee ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Wild Type ◽  
Simulated Ischemia ◽  
Selective Targeting ◽  
Cardioprotective Effects ◽  
Mitochondrial Turnover ◽  
First Time

Autophagy-dependent mitochondrial turnover in response to cellular stress is necessary for maintaining cellular homeostasis. However, the mechanisms that govern the selective targeting of damaged mitochondria are poorly understood. Parkin, an E3 ubiquitin ligase, has been shown to be essential for the selective clearance of damaged mitochondria. Parkin is expressed in the heart, yet its function has not been investigated in the context of cardioprotection. We previously reported that autophagy is required for cardioprotection by ischemic preconditioning (IPC). In the present study, we used simulated ischemia in vitro and IPC in hearts (in vivo and ex vivo) to investigate the role of Parkin in mediating cardioprotection. In HL-1 cells, simulated ischemia induced Parkin translocation to mitochondria and mitochondrial elimination. Mitochondrial loss was blunted in Atg5-deficient cells, revealing the requirement for autophagy in mitochondrial elimination. Consistent with previous reports implicating p62/SQSTM1 in mitophagy, we found that downregulation of p62 attenuated mitophagy and exacerbated cell death in HL-1 cardiomyocytes subjected to simulated ischemia. While wild type mice showed p62 translocation to mitochondria after IPC, Parkin knockout mice exhibited attenuated translocation of p62 to mitochondria. Importantly, ablation of Parkin in mice abolished the cardioprotective effects of IPC. These results reveal for the first time the crucial role of Parkin and mitophagy in cardioprotection.

scholarly journals Role of the Mannose Receptor in a Murine Model of Cryptococcus neoformans Infection

2008 ◽  
Vol 76 (6) ◽  
pp. 2362-2367 ◽  
Author(s):  
Jennifer M. Dan ◽  
Ryan M. Kelly ◽  
Chrono K. Lee ◽  
Stuart M. Levitz
Keyword(s):  
T Cell ◽  
Cryptococcus Neoformans ◽  
Cell Function ◽  
Ex Vivo ◽  
T Cell Responses ◽  
Mannose Receptor ◽  
Wild Type ◽  
Cell Responses

ABSTRACT Cryptococcus neoformans is an encapsulated fungal pathogen with a predilection to infect persons with suppressed T-cell function. Cryptococcal mannoproteins (MP) are highly mannosylated antigens which elicit T-cell responses in infected mice and in convalescent patients. Key to the immunogenicity of MP is its capacity to bind to the conserved mannose receptor (MR), CD206, on dendritic cells (DCs). To test the role of the MR in the immune response to C. neoformans, wild-type and MR knockout (MR KO) mice were compared by using in vivo and ex vivo models of cryptococcosis. Following a pulmonary challenge with C. neoformans, MR KO mice died significantly faster than wild-type mice and had higher lung fungal burdens after 4 weeks of infection. Uptake of MP was similar when DCs obtained from wild-type and MR KO mice were compared. Additionally, MP did not upregulate the maturation markers major histocompatibility complex class II, CD86, and CD40 in either wild-type or MR KO DCs. However, MP stimulated lymphoproliferation in CD4+ T cells obtained from the peripheral lymph nodes of infected wild-type but not MR KO mice. These studies demonstrate a nonredundant role for the MR in the development of CD4+ T-cell responses to MP and protection from C. neoformans.

Critical Role of CIB1 in Endothelial Cell Function and In Vivo Ischemia-Induced Angiogenesis.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1800-1800
Author(s):  
Mohamed A. Zayed ◽  
Andrew McFadden ◽  
Weiping Yuan ◽  
Mary E. Hartnett ◽  
Dan Chalothorn ◽  
...  
Keyword(s):  
Hind Limb ◽  
Ex Vivo ◽  
Critical Role ◽  
Tube Formation ◽  
Wild Type ◽  
Wild Type Littermate ◽  
Retinal Angiogenesis

Abstract CIB1, a 22kDa EF-hand containing calcium binding protein, was originally identified in a yeast two-hybrid screen as a binding partner for the cytoplasmic tail of the platelet integrin αIIb. CIB1 also associates with a number of kinases and modulates their activity, suggesting that CIB1 is an important regulatory molecule. Recently, we found that CIB1 is expressed in multiple endothelial cell (EC) types. We therefore tested the role of CIB1 in EC function in vitro, and in angiogenesis both ex vivo and in vivo. To test the role of CIB1 in EC function in vitro, we reduced endogenous CIB1 levels in ECs by RNA interference with an shRNA-delivered by lentivirus. CIB1 depletion significantly decreased EC haptotaxis on fibronectin and EC vascular tube formation on growth factor-reduced Matrigel. Treatment with FGF-2, an angiogenic factor, did not counter the observed inhibition of haptotaxis and tube formation by shRNA against CIB1. However, CIB1 overexpression enhanced FGF-2-induced EC haptotaxis relative to control cells. Similarly, ECs derived from CIB1 null mice exhibited a significant decrease in haptotaxis, tube formation, and proliferation compared to ECs isolated from wild-type littermate controls. In ex vivo aortic ring and tibialis anterior muscle culture assays, CIB1 null cultures supplemented with serum or FGF-2 demonstrated reduced blood vessel sprouting compared to wild-type littermate control cultures. Finally, in vivo assays for hyperoxic retinal angiogenesis and hind-limb induced-ischemia revealed a decrease in post-ischemia retinal neovascularization and Doppler hind-limb blood perfusion recovery, although developmental retinal angiogenesis in CIB1 null mice appeared normal. In conclusion, these findings support a critical role for CIB1 in EC function that appears to be important for ischemia-induced angiogenesis.

scholarly journals Reduced allergic lung inflammation and airway responsiveness in mice lacking the cytoskeletal protein gelsolin

2020 ◽  
Vol 319 (5) ◽  
pp. L833-L842
Author(s):  
Maya Mikami ◽  
Gene T. Yocum ◽  
Nicola M. Heller ◽  
Charles W. Emala
Keyword(s):  
Airway Inflammation ◽  
Lung Inflammation ◽  
Ex Vivo ◽  
Cell Types ◽  
Asthma Severity ◽  
Wild Type ◽  
Allergic Lung Inflammation ◽  
Actin Capping

Airway smooth muscle hyperresponsiveness associated with chronic airway inflammation leads to the typical symptoms of asthma including bronchoconstriction and wheezing. Asthma severity is associated with airway inflammation; therefore, reducing airway inflammation is an important therapeutic target. Gelsolin is an actin capping and severing protein that has been reported to be involved in modulation of the inflammatory response. Using mice genetically lacking gelsolin, we evaluated the role of gelsolin in the establishment of house dust mite (HDM) antigen-induced allergic lung inflammation. The genetic absence of gelsolin was found to be protective against HDM sensitization, resulting in reduced lung inflammation, inflammatory cytokines, and Muc5AC protein in bronchoalveolar lavage (BAL) fluid. The number of eosinophils, lymphocytes, and interstitial macrophages in the BAL were increased after HDM sensitization in wild-type mice but were attenuated in gelsolin-null mice. The observed attenuation of inflammation may be partly due to delayed migration of immune cells, because the reduced eosinophils in the BALs from gelsolin-null mice compared with controls occurred despite similar amounts of the chemoattractant eotaxin. Splenic T cells demonstrated similar proliferation rates, but ex vivo alveolar macrophage migration was delayed in gelsolin-null mice. In vivo, the reduced lung inflammation after HDM sensitization in gelsolin-null mice was associated with significantly diminished airway resistance to inhaled methacholine compared with HDM-treated wild-type mice. Our results suggest that modulation of gelsolin expression or function in selective inflammatory cell types that modulate allergic lung inflammation could be a therapeutic approach for asthma.

scholarly journals Stimulation of Platelet Nitric Oxide Production by Nebivolol Prevents Thrombosis

2014 ◽  
Vol 34 (4) ◽  
pp. 820-829 ◽  
Author(s):  
Stefania Momi ◽  
Roberta Caracchini ◽  
Emanuela Falcinelli ◽  
Stefano Evangelista ◽  
Paolo Gresele
Keyword(s):  
Nitric Oxide ◽  
Blood Cells ◽  
Adrenergic Receptor ◽  
Pharmacological Intervention ◽  
Hypotensive Activity ◽  
Wild Type ◽  
Antithrombotic Activity ◽  
Stimulation Of

Objective— dl -Nebivolol, a selective β1-adrenergic receptor antagonist, besides its hypotensive activity exerts vasodilatory and platelet inhibitory effects in vitro by a mechanism involving nitric oxide (NO). Our aim was to evaluate whether nebivolol exerts in vivo antithrombotic effects, to unravel the mechanism of this action and to clarify the relative roles of its 2 enantiomers: d - and l -nebivolol. Methods and Results— In wild-type mice, dl -nebivolol, l -nebivolol, and d -nebivolol, but not bisoprolol, reduced mortality consequent to platelet pulmonary thromboembolism induced by the intravenous injection of collagen plus epinephrine (−44%, −45%, −29%, respectively; P <0.05), whereas in eNOS −/− mice only dl -nebivolol and d -nebivolol were effective. dl -Nebivolol, l - and d -nebivolol reduced photochemical damage-induced femoral artery thrombosis in wild-type mice, whereas in eNOS −/− mice only dl -nebivolol and d -nebivolol were active. Moreover, dl -nebivolol and l -nebivolol increased plasma, urinary-, and platelet-derived nitrites and nitrates (NOx), NO degradation products, in wild-type but not in eNOS −/− mice. In vivo platelet activation, assessed by platelet P-selectin expression, was reduced by dl -nebivolol and l - and d -nebivolol in wild-type mice but only by dl -nebivolol and d -nebivolol in eNOS −/− mice. In bone marrow–transplanted, chimeric mice with only blood cells, and not the endothelium, producing NO dl -nebivolol and l -nebivolol maintained their antithrombotic activity, whereas they lose it in chimeras with only endothelium, and not blood cells, producing NO. In vitro, with isolated platelets, dl -nebivolol and l -nebivolol, but not d -nebivolol and bisoprolol, increased platelet cGMP and NOx formation. Treatment with dl -nebivolol and l -nebivolol increased phophorylated eNOS in platelets. Conclusions— Our data show that dl -nebivolol exerts an antithrombotic activity by stimulating the formation of NO by platelets, and that this effect is generated by its l -enantiomer, whereas the d -enantiomer exerts a weak antiplatelet effect because of β−adrenergic receptor–independent stimulation of adenyly cyclase. These results confirm that platelet-derived NO plays a role in thrombosis prevention and it may represent a target of pharmacological intervention.

scholarly journals Role of Toll-Like Receptor 5 (TLR5) in Experimental Melioidosis

2019 ◽  
Vol 87 (8) ◽  
Author(s):  
Emma Birnie ◽  
Tassili A. F. Weehuizen ◽  
Jacqueline M. Lankelma ◽  
Hanna K. de Jong ◽  
Gavin C. K. W. Koh ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Organ Injury ◽  
Independent Effect ◽  
Wild Type ◽  
Content Type ◽  
Pathogen Invasion ◽  
Toll Like Receptor 5 ◽  
Molecular Patterns

ABSTRACT The Gram-negative intracellular pathogen Burkholderia pseudomallei is the causative agent of melioidosis, an important cause of sepsis in Southeast Asia. Recognition of pathogen-associated molecular patterns by Toll-like receptors (TLRs) is essential for an appropriate immune response during pathogen invasion. In patients with melioidosis, TLR5 is the most abundantly expressed TLR, and a hypofunctional TLR5 variant has been associated with improved survival. Here, we studied the functional role of TLR5 and its ligand flagellin in experimental melioidosis. First, we observed differential TLR5 expression in the pulmonary and hepatic compartments upon infection with B. pseudomallei. Next, we found that B. pseudomallei-challenged TLR5-deficient (Tlr5−/−) mice were more susceptible to infection than wild-type (WT) mice, as demonstrated by higher systemic bacterial loads, increased organ injury, and impaired survival. Lung bacterial loads were not different between the two groups. The phenotype was flagellin independent; no difference in in vivo virulence was observed for the flagellin-lacking mutant MM36 compared to the wild-type B. pseudomallei strain 1026b. Tlr5−/− mice showed a similar impaired antibacterial defense when infected with MM36 or 1026b. Ex vivo experiments showed that TLR5-deficient macrophages display markedly impaired phagocytosis of B. pseudomallei. In conclusion, these data suggest that TLR5 deficiency has a detrimental flagellin-independent effect on the host response against pulmonary B. pseudomallei infection.

Abstract 341: The Role of Nucleotidase in Arterial Thrombosis

2016 ◽  
Vol 36 (suppl_1) ◽  
Author(s):  
Roman Covarrubias ◽  
Elena Chepurko ◽  
Tatiana Novitskaya ◽  
Karen M Dwyer ◽  
Simon C Robson ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Carotid Artery ◽  
Ferric Chloride ◽  
Arterial Thrombosis ◽  
Ex Vivo ◽  
Wild Type ◽  
Circulating Cells ◽  
Clodronate Liposome

Objective: To determine how leukocyte nucleotidase affects arterial thrombosis. Approach and Results: Ectonucleoside triphosphate diphosphohydrolase-1 (CD39) is expressed on circulating cells, endothelium and smooth muscle cells where it hydrolyzes extracellular ATP or ADP to AMP. We have demonstrated that transgenic mice with a global overexpression of human CD39 (hCD39-Tg) are protected against ferric chloride-induced carotid artery thrombosis. Furthermore, transplant of hCD39-Tg bone marrow into WT recipient mice increases the time to thrombosis when compared to recipient mice (wild-type or hCD39-Tg) receiving wild-type bone marrow. Based upon these data and previously published work, we hypothesized that CD39 expression on leukocytes is responsible for the prolongation of the time to thrombosis measured in hCD39-Tg mice. To test this hypothesis, we first performed ex vivo mixing experiments. Addition of hCD39-Tg monocytes to WT blood inhibits the expression of activated glycoprotein IIb/IIIa on platelets in response to ADP as measured by FACS analysis (Baseline: 1224 ± 94.9 MFI vs hCD39-Tg monocytes: 663.5 ± 61.5 activated glycoprotein IIb/IIIa MFI: n=4; p< 0.001). Subsequently, in vivo we demonstrated that monocytes with increased CD39 contribute to extending the time to thrombosis. Clodronate liposome depletion of monocytes (WT: 69% decrease; hCD39-Tg: 63% decrease) resulted in a normalization of the time to thrombosis in hCD39-Tg mice (8.0 ± 1.07 minutes, n = 10) when compared to control loaded liposomes (120.0 ± 0.0, n = 14). No changes in the time to thrombosis were detected in wild-type mice treated with clodronate (8.6 ± 1.35 minutes, n = 8) or control liposomes (7.8 ± 0.80 minutes, n=8). Conclusion: Increased expression of CD39 on monocytes can inhibit platelet activation and extend the time to thrombosis following ferric chloride-induced carotid artery injury.

Catecholamines act via a β-adrenergic receptor to maintain fetal heart rate and survival

2003 ◽  
Vol 284 (6) ◽  
pp. H2069-H2077 ◽  
Author(s):  
Andrea L. Portbury ◽  
Rashmi Chandra ◽  
Marybeth Groelle ◽  
Michael K. McMillian ◽  
Alana Elias ◽  
...  
Keyword(s):  
Heart Rate ◽  
Fetal Heart Rate ◽  
Adrenergic Receptor ◽  
Fetal Heart ◽  
Wild Type ◽  
Reduce Heart Rate ◽  
Specific Manner ◽  
Dose Dependent

Mice lacking catecholamines die before birth, some with cardiovascular abnormalities. To investigate the role of catecholamines in development, embryonic day 12.5 (E12.5) fetuses were cultured and heart rate monitored. Under optimal oxygenation, wild-type and catecholamine-deficient fetuses had the same initial heart rate (200–220 beats/min), which decreased by 15% in wild-type fetuses during 50 min of culture. During the same culture period, catecholamine-deficient fetuses dropped their heart rate by 35%. Hypoxia reduced heart rate of wild-type fetuses by 35–40% in culture and by 20% in utero, assessed by echocardiography. However, catecholamine-deficient fetuses exhibited greater hypoxia-induced bradycardia, reducing their heart rate by 70–75% in culture. Isoproterenol, a β-adrenergic receptor (β-AR) agonist, reversed this extreme bradycardia, restoring the rate of catecholamine-deficient fetuses to that of nonmutant siblings. Moreover, isoproterenol rescued 100% of catecholamine-deficient pups to birth in a dose-dependent, stereo-specific manner when administered in the dam's drinking water. An α-AR agonist was without effect. When wild-type fetuses were cultured with adrenoreceptor antagonists to create pharmacological nulls, blockade of α-ARs with 10 μM phentolamine or β-ARs with 10 μM bupranolol alone or in combination did not reduce heart rate under optimal oxygenation. However, when combined with hypoxia, β-AR blockade reduced heart rate by 35%. In contrast, the muscarinic blocker atropine and the α-AR antagonist phentolamine had no effect. These data suggest that β-ARs mediate survival in vivo and regulate heart rate in culture. We hypothesize that norepinephrine, acting through β-ARs, maintains fetal heart rate during periods of transient hypoxia that occur throughout gestation, and that catecholamine-deficient fetuses die because they cannot withstand hypoxia-induced bradycardia.

The Role of Polyphenols in the Modulation of Plasma Non-Enzymatic Antioxidant Capacity (NEAC)

2012 ◽  
Vol 82 (3) ◽  
pp. 228-232 ◽  
Author(s):  
Mauro Serafini ◽  
Giuseppa Morabito
Keyword(s):  
Antioxidant Capacity ◽  
Dietary Intervention ◽  
Ex Vivo ◽  
The Body ◽  
Dietary Polyphenols ◽  
Enzymatic Antioxidant ◽  
Endogenous Antioxidant

Dietary polyphenols have been shown to scavenge free radicals, modulating cellular redox transcription factors in different in vitro and ex vivo models. Dietary intervention studies have shown that consumption of plant foods modulates plasma Non-Enzymatic Antioxidant Capacity (NEAC), a biomarker of the endogenous antioxidant network, in human subjects. However, the identification of the molecules responsible for this effect are yet to be obtained and evidences of an antioxidant in vivo action of polyphenols are conflicting. There is a clear discrepancy between polyphenols (PP) concentration in body fluids and the extent of increase of plasma NEAC. The low degree of absorption and the extensive metabolism of PP within the body have raised questions about their contribution to the endogenous antioxidant network. This work will discuss the role of polyphenols from galenic preparation, food extracts, and selected dietary sources as modulators of plasma NEAC in humans.

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