Prenatal nicotinic exposure upregulates pulmonary C-fiber NK1R expression to prolong pulmonary C-fiber-mediated apneic response

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Zhao, Lei; Zhuang, Jianguo; Zang, Na; Lin, Yong; Lee, Lu-Yuan; Xu, Fadi

2016-01-01

Prenatal nicotinic exposure (PNE) prolongs bronchopulmonary C-fiber (PCF)-mediated apneic response to intra-atrial bolus injection of capsaicin in rat pups. The relevant mechanisms remain unclear. Pulmonary substance P and adenosine and their receptors (neurokinin-A receptor, NK1R and ADA 1 receptor, ADA 1 R) and transient receptor potential cation channel subfamily V member 1 (TRPV1) expressed on PCFs are critical for PCF sensitization and/or activation. Here, we compared substance P and adenosine in BALF and NK1R, ADA 1 R, and TRPV1 expression in the nodose/jugular (N/J) ganglia (vagal pulmonary C-neurons retrogradely labeled) between Ctrl and PNE pups. We found that PNE failed to change BALF substance P and adenosine content, but significantly upregulated both mRNA and protein TRPV1 and NK1R in the N/J ganglia and only NK1R mRNA in pulmonary C-neurons. To define the role of NK1R in the PNE-induced PCF sensitization, the apneic response to capsaicin (i.v.) without or with pretreatment of SR140333 (a peripheral and selective NK1R antagonist) was compared and the prolonged apnea by PNE significantly shortened by SR140333. To clarify if the PNE-evoked responses depended on action of nicotinic acetylcholine receptors (nAChRs), particularly α7nAChR, mecamylamine or methyllycaconitine (a general nAChR or a selective α7nAChR antagonist) was administrated via another mini-pump over the PNE period. Mecamylamine or methyllycaconitine eliminated the PNE-evoked mRNA and protein responses. Our data suggest that PNE is able to elevate PCF NK1R expression via activation of nAChRs, especially α7nAChR, which likely contributes to sensitize PCFs and prolong the PCF-mediated apneic response to capsaicin. - Highlights: • PNE upregulated NK1R and TRPV1 gene and protein expression in the N/J ganglia. • PNE only elevated NK1R mRNA in vagal pulmonary C-neurons. • Blockage of peripheral NK1R reduced the PNE-induced PCF sensitization. • PNE induced gene and protein changes in

Neurokinin-1 (NK-1 receptor and brain-derived neurotrophic factor (BDNF gene expression is differentially modulated in the rat spinal dorsal horn and hippocampus during inflammatory pain

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McCarson Kenneth E

2007-10-01

Full Text Available Abstract Persistent pain produces complex alterations in sensory pathways of the central nervous system (CNS through activation of various nociceptive mechanisms. However, the effects of pain on higher brain centers, particularly the influence of the stressful component of pain on the limbic system, are poorly understood. Neurokinin-1 (NK-1 receptors and brain-derived neurotrophic factor (BDNF, known neuromediators of hyperalgesia and spinal central sensitization, have also been implicated in the plasticity and neurodegeneration occurring in the hippocampal formation during exposures to various stressors. Results of this study showed that injections of complete Freund's adjuvant (CFA into the hind paw increased NK-1 receptor and BDNF mRNA levels in the ipsilateral dorsal horn, supporting an important role for these nociceptive mediators in the amplification of ascending pain signaling. An opposite effect was observed in the hippocampus, where CFA down-regulated NK-1 receptor and BDNF gene expression, phenomena previously observed in immobilization models of stress and depression. Western blot analyses demonstrated that in the spinal cord, CFA also increased levels of phosphorylated cAMP response element-binding protein (CREB, while in the hippocampus the activation of this transcription factor was significantly reduced, further suggesting that tissue specific transcription of either NK-1 or BDNF genes may be partially regulated by common intracellular transduction mechanisms mediated through activation of CREB. These findings suggest that persistent nociception induces differential regional regulation of NK-1 receptor and BDNF gene expression and CREB activation in the CNS, potentially reflecting varied roles of these neuromodulators in the spinal cord during persistent sensory activation vs. modulation of the higher brain structures such as the hippocampus.

Bladder inflammatory transcriptome in response to tachykinins: Neurokinin 1 receptor-dependent genes and transcription regulatory elements

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Dozmorov Igor

2007-05-01

Full Text Available Abstract Background Tachykinins (TK, such as substance P, and their neurokinin receptors which are ubiquitously expressed in the human urinary tract, represent an endogenous system regulating bladder inflammatory, immune responses, and visceral hypersensitivity. Increasing evidence correlates alterations in the TK system with urinary tract diseases such as neurogenic bladders, outflow obstruction, idiopathic detrusor instability, and interstitial cystitis. However, despite promising effects in animal models, there seems to be no published clinical study showing that NK-receptor antagonists are an effective treatment of pain in general or urinary tract disorders, such as detrusor overactivity. In order to search for therapeutic targets that could block the tachykinin system, we set forth to determine the regulatory network downstream of NK1 receptor activation. First, NK1R-dependent transcripts were determined and used to query known databases for their respective transcription regulatory elements (TREs. Methods An expression analysis was performed using urinary bladders isolated from sensitized wild type (WT and NK1R-/- mice that were stimulated with saline, LPS, or antigen to provoke inflammation. Based on cDNA array results, NK1R-dependent genes were selected. PAINT software was used to query TRANSFAC database and to retrieve upstream TREs that were confirmed by electrophoretic mobility shift assays. Results The regulatory network of TREs driving NK1R-dependent genes presented cRel in a central position driving 22% of all genes, followed by AP-1, NF-kappaB, v-Myb, CRE-BP1/c-Jun, USF, Pax-6, Efr-1, Egr-3, and AREB6. A comparison between NK1R-dependent and NK1R-independent genes revealed Nkx-2.5 as a unique discriminator. In the presence of NK1R, Nkx2-5 _01 was significantly correlated with 36 transcripts which included several candidates for mediating bladder development (FGF and inflammation (PAR-3, IL-1R, IL-6, α-NGF, TSP2. In the absence of

β-Arrestin 1 has an essential role in neurokinin-1 receptor-mediated glioblastoma cell proliferation and G2/M phase transition.

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Zhang, Yi-Xin; Li, Xiao-Fang; Yuan, Guo-Qiang; Hu, Hui; Song, Xiao-Yun; Li, Jing-Yi; Miao, Xiao-Kang; Zhou, Tian-Xiong; Yang, Wen-Le; Zhang, Xiao-Wei; Mou, Ling-Yun; Wang, Rui

2017-05-26

Glioblastoma is the most common malignant brain tumor and has a poor prognosis. Tachykinin receptor neurokinin-1 (NK1R) is a promising target in glioblastoma therapy because of its overexpression in human glioblastoma. NK1R agonists promote glioblastoma cell growth, whereas NK1R antagonists efficiently inhibit cell growth both in vitro and in vivo However, the molecular mechanisms involved in these effects are incompletely understood. β-Arrestins (ARRBs) serve as scaffold proteins and adapters to mediate intracellular signal transduction. Here we show that the ARRB1-mediated signaling pathway is essential for NK1-mediated glioblastoma cell proliferation. ARRB1 knockdown significantly inhibited NK1-mediated glioblastoma cell proliferation and induced G 2 /M phase cell cycle arrest. ARRB1 knockdown cells showed remarkable down-regulation of CDC25C/CDK1/cyclin B1 activity. We also demonstrated that ARRB1 mediated prolonged phosphorylation of ERK1/2 and Akt in glioblastoma cells induced by NK1R activation. ERK1/2 and Akt phosphorylation are involved in regulating CDC25C/CDK1/cyclin B1 activity. The lack of long-term ERK1/2 and Akt activation in ARRB1 knockdown cells was at least partly responsible for the delayed cell cycle progression and proliferation. Moreover, we found that ARRB1-mediated ERK1/2 and Akt phosphorylation regulated the transcriptional activity of both NF-κB and AP-1, which were involved in cyclin B1 expression. ARRB1 deficiency increased the sensitivity of glioblastoma cells to the treatment of NK1R antagonists. Taken together, our results suggest that ARRB1 plays an essential role in NK1R-mediated cell proliferation and G 2 /M transition in glioblastoma cells. Interference with ARRB1-mediated signaling via NK1R may have potential significance for therapeutic strategies targeting glioblastoma. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Two active molecular phenotypes of the tachykinin NK1 receptor revealed by G-protein fusions and mutagenesis.

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Holst, B; Hastrup, H; Raffetseder, U; Martini, L; Schwartz, T W

2001-06-08

The NK1 neurokinin receptor presents two non-ideal binding phenomena, two-component binding curves for all agonists and significant differences between agonist affinity determined by homologous versus heterologous competition binding. Whole cell binding with fusion proteins constructed between either Galpha(s) or Galpha(q) and the NK1 receptor with a truncated tail, which secured non-promiscuous G-protein interaction, demonstrated monocomponent agonist binding closely corresponding to either of the two affinity states found in the wild-type receptor. High affinity binding of both substance P and neurokinin A was observed in the tail-truncated Galpha(s) fusion construct, whereas the lower affinity component was displayed by the tail-truncated Galpha(q) fusion. The elusive difference between the affinity determined in heterologous versus homologous binding assays for substance P and especially for neurokinin A was eliminated in the G-protein fusions. An NK1 receptor mutant with a single substitution at the extracellular end of TM-III-(F111S), which totally uncoupled the receptor from Galpha(s) signaling, showed binding properties that were monocomponent and otherwise very similar to those observed in the tail-truncated Galpha(q) fusion construct. Thus, the heterogenous pharmacological phenotype displayed by the NK1 receptor is a reflection of the occurrence of two active conformations or molecular phenotypes representing complexes with the Galpha(s) and Galpha(q) species, respectively. We propose that these molecular forms do not interchange readily, conceivably because of the occurrence of microdomains or "signal-transductosomes" within the cell membrane.

Prenatal nicotinic exposure upregulates pulmonary C-fiber NK1R expression to prolong pulmonary C-fiber-mediated apneic response

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Zhao, Lei; Zhuang, Jianguo; Zang, Na; Lin, Yong [Pathophysiology Program, Lovelace Respiratory Research Institute, Albuquerque, NM (United States); Lee, Lu-Yuan [Department of Physiology, University of Kentucky, Lexington, KY (United States); Xu, Fadi, E-mail: fxu@lrri.org [Pathophysiology Program, Lovelace Respiratory Research Institute, Albuquerque, NM (United States); Department of Physiology, University of Kentucky, Lexington, KY (United States)

2016-01-01

Prenatal nicotinic exposure (PNE) prolongs bronchopulmonary C-fiber (PCF)-mediated apneic response to intra-atrial bolus injection of capsaicin in rat pups. The relevant mechanisms remain unclear. Pulmonary substance P and adenosine and their receptors (neurokinin-A receptor, NK1R and ADA{sub 1} receptor, ADA{sub 1}R) and transient receptor potential cation channel subfamily V member 1 (TRPV1) expressed on PCFs are critical for PCF sensitization and/or activation. Here, we compared substance P and adenosine in BALF and NK1R, ADA{sub 1}R, and TRPV1 expression in the nodose/jugular (N/J) ganglia (vagal pulmonary C-neurons retrogradely labeled) between Ctrl and PNE pups. We found that PNE failed to change BALF substance P and adenosine content, but significantly upregulated both mRNA and protein TRPV1 and NK1R in the N/J ganglia and only NK1R mRNA in pulmonary C-neurons. To define the role of NK1R in the PNE-induced PCF sensitization, the apneic response to capsaicin (i.v.) without or with pretreatment of SR140333 (a peripheral and selective NK1R antagonist) was compared and the prolonged apnea by PNE significantly shortened by SR140333. To clarify if the PNE-evoked responses depended on action of nicotinic acetylcholine receptors (nAChRs), particularly α7nAChR, mecamylamine or methyllycaconitine (a general nAChR or a selective α7nAChR antagonist) was administrated via another mini-pump over the PNE period. Mecamylamine or methyllycaconitine eliminated the PNE-evoked mRNA and protein responses. Our data suggest that PNE is able to elevate PCF NK1R expression via activation of nAChRs, especially α7nAChR, which likely contributes to sensitize PCFs and prolong the PCF-mediated apneic response to capsaicin. - Highlights: • PNE upregulated NK1R and TRPV1 gene and protein expression in the N/J ganglia. • PNE only elevated NK1R mRNA in vagal pulmonary C-neurons. • Blockage of peripheral NK1R reduced the PNE-induced PCF sensitization. • PNE induced gene and protein

Cellular and behavioural profile of the novel, selective neurokinin1 receptor antagonist, vestipitant: a comparison to other agents.

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Brocco, Mauricette; Dekeyne, Anne; Mannoury la Cour, Clotilde; Touzard, Manuelle; Girardon, Sylvie; Veiga, Sylvie; de Nanteuil, Guillaume; deJong, Trynke R; Olivier, Berend; Millan, Mark J

2008-10-01

This study characterized the novel neurokinin (NK)(1) antagonist, vestipitant, under clinical evaluation for treatment of anxiety and depression. Vestipitant possessed high affinity for human NK(1) receptors (pK(i), 9.4), and potently blocked Substance P-mediated phosphorylation of Extracellular-Regulated-Kinase. In vivo, it occupied central NK(1) receptors in gerbils (Inhibitory Dose(50), 0.11 mg/kg). At similar doses, it abrogated nociception elicited by formalin in gerbils, and blocked foot-tapping and locomotion elicited by the NK(1) agonist, GR73632, in gerbils and guinea pigs, respectively. Further, vestipitant attenuated fear-induced foot-tapping in gerbils, separation-induced distress-vocalizations in guinea pigs, marble-burying behaviour in mice, and displayed anxiolytic actions in Vogel conflict and fear-induced ultrasonic vocalization procedures in rats. These actions were mimicked by CP99,994, L733,060 and GR205,171 which acted stereoselectively vs its less active isomer, GR226,206. In conclusion, vestipitant is a potent NK(1) receptor antagonist: its actions support the utility of NK(1) receptor blockade in the alleviation of anxiety and, possibly, depression.

Targeting NK-1 Receptors to Prevent and Treat Pancreatic Cancer: A New Therapeutic Approach

International Nuclear Information System (INIS)

Muñoz, Miguel; Coveñas, Rafael

2015-01-01

Pancreatic cancer (PC) is the fourth leading cause of cancer related-deaths in both men and women, and the 1- and 5-year relative survival rates are 25% and 6%, respectively. It is known that smoking, alcoholism and psychological stress are risk factors that can promote PC and increase PC progression. To date, the prevention of PC is crucial because there is no curative treatment. After binding to the neurokinin-1 (NK-1) receptor (a receptor coupled to the stimulatory G-protein Gαs that activates adenylate cyclase), the peptide substance P (SP)—at high concentrations—is involved in many pathophysiological functions, such as depression, smoking, alcoholism, chronic inflammation and cancer. It is known that PC cells and samples express NK-1 receptors; that the NK-1 receptor is overexpressed in PC cells in comparison with non-tumor cells, and that nanomolar concentrations of SP induce PC cell proliferation. By contrast, NK-1 receptor antagonists exert antidepressive, anxiolytic and anti-inflammatory effects and anti-alcohol addiction. These antagonists also exert an antitumor action since in vitro they inhibit PC cell proliferation (PC cells death by apoptosis), and in a xenograft PC mouse model they exert both antitumor and anti-angiogenic actions. NK-1 receptor antagonists could be used for the treatment of PC and hence the NK-1 receptor could be a new promising therapeutic target in PC

Targeting NK-1 Receptors to Prevent and Treat Pancreatic Cancer: A New Therapeutic Approach

Energy Technology Data Exchange (ETDEWEB)

Muñoz, Miguel, E-mail: mmunoz@cica.es [Research Laboratory on Neuropeptides (IBIS), Virgen del Rocío University Hospital, 41013 Sevilla (Spain); Coveñas, Rafael [Laboratory of Neuroanatomy of the Peptidergic System (Lab. 14), Institute of Neurosciences of Castilla y León (INCYL), University of Salamanca, 37008 Salamanca (Spain)

2015-07-06

Pancreatic cancer (PC) is the fourth leading cause of cancer related-deaths in both men and women, and the 1- and 5-year relative survival rates are 25% and 6%, respectively. It is known that smoking, alcoholism and psychological stress are risk factors that can promote PC and increase PC progression. To date, the prevention of PC is crucial because there is no curative treatment. After binding to the neurokinin-1 (NK-1) receptor (a receptor coupled to the stimulatory G-protein Gαs that activates adenylate cyclase), the peptide substance P (SP)—at high concentrations—is involved in many pathophysiological functions, such as depression, smoking, alcoholism, chronic inflammation and cancer. It is known that PC cells and samples express NK-1 receptors; that the NK-1 receptor is overexpressed in PC cells in comparison with non-tumor cells, and that nanomolar concentrations of SP induce PC cell proliferation. By contrast, NK-1 receptor antagonists exert antidepressive, anxiolytic and anti-inflammatory effects and anti-alcohol addiction. These antagonists also exert an antitumor action since in vitro they inhibit PC cell proliferation (PC cells death by apoptosis), and in a xenograft PC mouse model they exert both antitumor and anti-angiogenic actions. NK-1 receptor antagonists could be used for the treatment of PC and hence the NK-1 receptor could be a new promising therapeutic target in PC.

Atomoxetine reduces hyperactive/impulsive behaviours in neurokinin-1 receptor ‘knockout’ mice

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Pillidge, Katharine; Porter, Ashley J.; Vasili, Temis; Heal, David J.; Stanford, S. Clare

2014-01-01

Background Mice with functional ablation of the neurokinin-1 receptor gene (NK1R−/−) display behavioural abnormalities which resemble the hyperactivity, inattention and impulsivity seen in Attention Deficit Hyperactivity Disorder (ADHD). Here, we investigated whether the established ADHD treatment, atomoxetine, alleviates these abnormalities when tested in the light/dark exploration box (LDEB) and 5-Choice Serial Reaction-Time Task (5-CSRTT). Methods Separate cohorts of mice were tested in the 5-CSRTT and LDEB after treatment with no injection, vehicle or atomoxetine (5-CSRTT: 0.3, 3 or 10 mg/kg; LDEB: 1, 3 or 10 mg/kg). Results Atomoxetine reduced the hyperactivity displayed by NK1R−/− mice in the LDEB at a dose (3 mg/kg) which did not affect the locomotor activity of wildtypes. Atomoxetine (10 mg/kg) also reduced impulsivity in NK1R−/− mice, but not wildtypes, in the 5-CSRTT. No dose of drug affected attention in either genotype. Conclusions This evidence that atomoxetine reduces hyperactive/impulsive behaviours in NK1R−/− mice consolidates the validity of using NK1R−/− mice in research of the aetiology and treatment of ADHD. PMID:25450119

Activation of neurokinin-1 receptors during ozone inhalation contributes to epithelial injury and repair.

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Oslund, Karen L; Hyde, Dallas M; Putney, Leialoha F; Alfaro, Mario F; Walby, William F; Tyler, Nancy K; Schelegle, Edward S

2008-09-01

We investigated the importance of neurokinin (NK)-1 receptors in epithelial injury and repair and neutrophil function. Conscious Wistar rats were exposed to 1 ppm ozone or filtered air for 8 hours, followed by an 8-hour postexposure period. Before exposure, we administered either the NK-1 receptor antagonist, SR140333, or saline as a control. Ethidium homodimer was instilled into lungs as a marker of necrotic airway epithelial cells. After fixation, whole mounts of airway dissected lung lobes were immunostained for 5-bromo-2'-deoxyuridine, a marker of epithelial proliferation. Both ethidium homodimer and 5-bromo-2'-deoxyuridine-positive epithelial cells were quantified in specific airway generations. Rats treated with the NK-1 receptor antagonist had significantly reduced epithelial injury and epithelial proliferation compared with control rats. Sections of terminal bronchioles showed no significant difference in the number of neutrophils in airways between groups. In addition, staining ozone-exposed lung sections for active caspase 3 showed no apoptotic cells, but ethidium-positive cells colocalized with the orphan nuclear receptor, Nur77, a marker of nonapoptotic, programmed cell death mediated by the NK-1 receptor. An immortalized human airway epithelial cell line, human bronchial epithelial-1, showed no significant difference in the number of oxidant stress-positive cells during exposure to hydrogen peroxide and a range of SR140333 doses, demonstrating no antioxidant effect of the receptor antagonist. We conclude that activation of the NK-1 receptor during acute ozone inhalation contributes to epithelial injury and subsequent epithelial proliferation, a critical component of repair, but does not influence neutrophil emigration into airways.

PET-scan shows peripherally increased neurokinin 1 receptor availability in chronic tennis elbow: visualizing neurogenic inflammation?

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Magnus Peterson

Full Text Available In response to pain, neurokinin 1 (NK1 receptor availability is altered in the central nervous system. The NK1 receptor and its primary agonist, substance P, also play a crucial role in peripheral tissue in response to pain, as part of neurogenic inflammation. However, little is known about alterations in NK1 receptor availability in peripheral tissue in chronic pain conditions and very few studies have been performed on human beings. Ten subjects with chronic tennis elbow were therefore examined by positron emission tomography (PET with the NK1 specific radioligand [(11C]GR205171 before and after treatment with graded exercise. The radioligand signal intensity was higher in the affected arm as compared with the unaffected arm, measured as differences between the arms in volume of voxels and signal intensity of this volume above a reference threshold set as 2.5 SD above mean signal intensity of the unaffected arm before treatment. In the eight subjects examined after treatment, pain ratings decreased in all subjects but signal intensity decreased in five and increased in three. In conclusion, NK1 receptors may be activated, or up-regulated in the peripheral, painful tissue of a chronic pain condition. This up-regulation does, however, have moderate correlation to pain ratings. The increased NK1 receptor availability is interpreted as part of ongoing neurogenic inflammation and may have correlation to the pathogenesis of chronic tennis elbow.ClinicalTrials.gov NCT00888225 http://clinicaltrials.gov/

Adrenocortical and behavioural response to chronic restraint stress in neurokinin-1 receptor knockout mice.

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Delgado-Morales, Raúl; del Río, Eva; Gómez-Román, Almudena; Bisagno, Verónica; Nadal, Roser; de Felipe, Carmen; Armario, Antonio

2012-02-01

Brain substance P and its receptor (neurokinin-1, NK1) have a widespread brain distribution and are involved in an important number of behavioural and physiological responses to emotional stimuli. However, the role of NK1 receptors in the consequences of exposure to chronic stress has not been explored. The present study focused on the role of these receptors in the hypothalamic-pituitary-adrenal (HPA) response to daily repeated restraint stress (evaluated by plasma corticosterone levels), as well as on the effect of this procedure on anxiety-like behaviour, spatial learning and memory in the Morris water maze (MWM), a hippocampus-dependent task. Adult null mutant NK1-/- mice, with a C57BL/6J background, and the corresponding wild-type mice showed similar resting corticosterone levels and, also, did not differ in corticosterone response to a first restraint. Nevertheless, adaptation to the repeated stressor was faster in NK1-/- mice. Chronic restraint modestly increased anxiety-like behaviour in the light-dark test, irrespective of genotype. Throughout the days of the MWM trials, NK1-/- mice showed a similar learning rate to that of wild-type mice, but had lower levels of thigmotaxis and showed a better retention in the probe trial. Chronic restraint stress did not affect these variables in either genotype. These results indicate that deletion of the NK1 receptor does not alter behavioural susceptibility to chronic repeated stress in mice, but accelerates adaptation of the HPA axis. In addition, deletion may result in lower levels of thigmotaxis and improved short-term spatial memory, perhaps reflecting a better learning strategy in the MWM. Copyright © 2011 Elsevier Inc. All rights reserved.

From the Cover: Prenatal Nicotinic Exposure Attenuates Respiratory Chemoreflexes Associated With Downregulation of Tyrosine Hydroxylase and Neurokinin 1 Receptor in Rat Pup Carotid Body.

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Zhao, Lei; Zhuang, Jianguo; Gao, Xiuping; Ye, Chunyan; Lee, Lu-Yuan; Xu, Fadi

2016-09-01

Maternal cigarette smoke is the major risk of sudden infant death syndrome (SIDS). A depressed ventilatory response to hypoxia (HVR) and hypercapnia (HCVR) is thought to be responsible for the pathogenesis of SIDS and the carotid body is critically involved in these responses. We have recently reported that prenatal nicotinic exposure (PNE) over the full gestation induces depressed HVR in rat pups. Here, we asked whether PNE (1) depressed not only HVR but also HCVR that were dependent on the carotid body, (2) affected some important receptors and neurochemicals expressed in the carotid body, such as tyrosine hydroxylase (TH), neurokinin-1 receptor (NK1R), and α7 nicotinic acetylcholine receptor (α7nAChR), and (3) blunted the ventilatory responses to activation of these receptors. To this end, HVR and HCVR in Ctrl and PNE pups were measured with plethysmography before and after carotid body ablation (Series I), mRNA expression and/or immunoreactivity (IR) of TH, NK1R, and α7nAChR in the carotid body were examined by RT-PCR and immunohistochemistry (Series II), and the ventilatory responses were tested before and after intracarotid injection of substance P (NK1R agonist) and AR-R17779 (α7nAChR agonist) (Series III). Our results showed that PNE (1) significantly depressed both HVR and HCVR and these depressions were abolished by carotid body ablation, (2) reduced the relative population of glomus cells, mRNA NK1R, and α7nAChR and IR of NK1R and TH in the carotid body, and (3) decreased ventilatory responses to intracarotid injection of substance P or AR-R17779. These results suggest that PNE acting via the carotid body could strikingly blunt HVR and HCVR, likely through downregulating TH and NK1R. © The Author 2016. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Two active molecular phenotypes of the tachykinin NK1 receptor revealed by G-protein fusions and mutagenesis

DEFF Research Database (Denmark)

Holst, B; Hastrup, H; Raffetseder, U

2001-01-01

The NK1 neurokinin receptor presents two non-ideal binding phenomena, two-component binding curves for all agonists and significant differences between agonist affinity determined by homologous versus heterologous competition binding. Whole cell binding with fusion proteins constructed between ei...

Upregulation of neurokinin-1 receptor expression in the lungs of patients with sarcoidosis.

LENUS (Irish Health Repository)

O'Connor, Terence M

2012-02-03

Substance P (SP) is a proinflammatory neuropeptide that is secreted by sensory nerves and inflammatory cells. Increased levels of SP are found in sarcoid bronchoalveolar lavage fluid. SP acts by binding to the neurokinin-1 receptor and increases secretion of tumor necrosis factor-alpha in many cell types. We sought to determine neurokinin-1 receptor expression in patients with sarcoidosis compared with normal controls. Neurokinin-1 receptor messenger RNA and protein expression were below the limits of detection by reverse transcriptase-polymerase chain reaction and immunohistochemistry in peripheral blood mononuclear cells of healthy volunteers (n = 9) or patients with stage 1 or 2 pulmonary sarcoidosis (n = 10), but were detected in 1\\/9 bronchoalveolar lavage cells of controls compared with 8\\/10 patients with sarcoidosis (p = 0.012) and 2\\/9 biopsies of controls compared with 9\\/10 patients with sarcoidosis (p = 0.013). Immunohistochemistry localized upregulated neurokinin-1 receptor expression to bronchial and alveolar epithelial cells, macrophages, lymphocytes, and sarcoid granulomas. The patient in whom neurokinin-1 receptor was not detected was taking corticosteroids. Incubation of the type II alveolar and bronchial epithelial cell lines A549 and SK-LU 1 with dexamethasone downregulated neurokinin-1 receptor expression. Upregulated neurokinin-1 receptor expression in patients with sarcoidosis may potentiate substance P-induced proinflammatory cytokine production in patients with sarcoidosis.

Phosphorylation of ERK in neurokinin 1 receptor-expressing neurons in laminae III and IV of the rat spinal dorsal horn following noxious stimulation

Directory of Open Access Journals (Sweden)

Watanabe Masahiko

2007-02-01

Full Text Available Abstract Background There is a population of large neurons with cell bodies in laminae III and IV of the spinal dorsal horn which express the neurokinin 1 receptor (NK1r and have dendrites that enter the superficial laminae. Although it has been shown that these are all projection neurons and that they are innervated by substance P-containing (nociceptive primary afferents, we know little about their responses to noxious stimuli. In this study we have looked for phosphorylation of extracellular signal-regulated kinases (ERKs in these neurons in response to different types of noxious stimulus applied to one hindlimb of anaesthetised rats. The stimuli were mechanical (repeated pinching, thermal (immersion in water at 52°C or chemical (injection of 2% formaldehyde. Results Five minutes after each type of stimulus we observed numerous cells with phosphorylated ERK (pERK in laminae I and IIo, together with scattered positive cells in deeper laminae. We found that virtually all of the lamina III/IV NK1r-immunoreactive neurons contained pERK after each of these stimuli and that in the great majority of cases there was internalisation of the NK1r on the dorsal dendrites of these cells. In addition, we also saw neurons in lamina III that were pERK-positive but lacked the NK1r, and these were particularly evident in animals that had had the pinch stimulus. Conclusion Our results demonstrate that lamina III/IV NK1r-immunoreactive neurons show receptor internalisation and ERK phosphorylation after mechanical, thermal or chemical noxious stimuli.

The NK-1 Receptor Antagonist L-732,138 Induces Apoptosis and Counteracts Substance P-Related Mitogenesis in Human Melanoma Cell Lines

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Miguel Muñoz

2010-04-01

Full Text Available It has been recently demonstrated that substance P (SP and neurokinin-1 (NK-1 receptor antagonists induce cell proliferation and cell inhibition in human melanoma cells, respectively. However, the antitumor action of the NK-1 receptor antagonist L-732,138 on such cells is unknown. The aim of this study was to demonstrate an antitumor action of L-732,138 against three human melanoma cell lines (COLO 858, MEL HO, COLO 679. We found that L-732,138 elicits cell growth inhibition in a concentration dependent manner in the melanoma cells studied. Moreover, L-732,138 blocks SP mitogen stimulation. The specific antitumor action of L-732,138 occurred through the NK-1 receptor and melanoma cell death was by apoptosis. These findings indicate that the NK-1 receptor antagonist L-732,138 could be a new antitumor agent in the treatment of human melanoma.

The NK-1 Receptor Antagonist L-732,138 Induces Apoptosis and Counteracts Substance P-Related Mitogenesis in Human Melanoma Cell Lines

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Muñoz, Miguel, E-mail: mmunoz@cica.es; Rosso, Marisa; González-Ortega, Ana [Research Laboratory on Neuropeptides, Virgen del Rocío University Hospital, Sevilla (Spain); Coveñas, Rafael [Institute of Neurosciences of Castilla y León (INCYL), Laboratory of Neuroanatomy of the Peptidergic Systems (Laboratory 14), Salamanca (Spain)

2010-04-20

It has been recently demonstrated that substance P (SP) and neurokinin-1 (NK-1) receptor antagonists induce cell proliferation and cell inhibition in human melanoma cells, respectively. However, the antitumor action of the NK-1 receptor antagonist L-732,138 on such cells is unknown. The aim of this study was to demonstrate an antitumor action of L-732,138 against three human melanoma cell lines (COLO 858, MEL HO, COLO 679). We found that L-732,138 elicits cell growth inhibition in a concentration dependent manner in the melanoma cells studied. Moreover, L-732,138 blocks SP mitogen stimulation. The specific antitumor action of L-732,138 occurred through the NK-1 receptor and melanoma cell death was by apoptosis. These findings indicate that the NK-1 receptor antagonist L-732,138 could be a new antitumor agent in the treatment of human melanoma.

Casopitant: a novel NK1-receptor antagonist in the prevention of chemotherapy-induced nausea and vomiting

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Christina Ruhlmann

2009-05-01

Full Text Available Christina Ruhlmann, Jørn HerrstedtOdense University Hospital, Department of Oncology, Odense, DenmarkAbstract: Chemotherapy-induced nausea and vomiting (CINV are among the most feared and distressing symptoms experienced by patients with cancer. The knowledge of the pathogenesis and neuropharmacology of CINV has expanded enormously over the last decades, the most significant discoveries being the role of 5-hydroxytryptamine (5-HT3- and neurokinin (NK1 receptors in the emetic reflex arch. This has led to the development of two new classes of antiemetics acting as highly selective antagonists at one of these receptors. These drugs have had a huge impact in the protection from chemotherapy-induced vomiting, whereas the effect on nausea seems to be limited. The first NK1 receptor antagonist, aprepitant, became clinically available in 2003, and casopitant, the second in this class of antiemetics, has now completed phase III trials. This review delineates the properties and clinical use of casopitant in the prevention of CINV.Keywords: casopitant, GW679769, NK1 receptor antagonist, chemotherapy, emesis

The angiotensin converting enzyme inhibitor, captopril, prevents the hyperactivity and impulsivity of neurokinin-1 receptor gene 'knockout' mice: sex differences and implications for the treatment of attention deficit hyperactivity disorder.

Science.gov (United States)

Porter, Ashley J; Pillidge, Katharine; Grabowska, Ewelina M; Stanford, S Clare

2015-04-01

Mice lacking functional neurokinin-1 receptors (NK1R-/-) display behavioural abnormalities resembling attention deficit hyperactivity disorder (ADHD): locomotor hyperactivity, impulsivity and inattentiveness. The preferred ligand for NK1R, substance P, is metabolised by angiotensin converting enzyme (ACE), which forms part of the brain renin angiotensin system (BRAS). In view of evidence that the BRAS modulates locomotor activity and cognitive performance, we tested the effects of drugs that target the BRAS on these behaviours in NK1R-/- and wildtype mice. We first tested the effects of the ACE inhibitor, captopril, on locomotor activity. Because there are well-established sex differences in both ADHD and ACE activity, we compared the effects of captopril in both male and female mice. Locomotor hyperactivity was evident in male NK1R-/- mice, only, and this was abolished by treatment with captopril. By contrast, male wildtypes and females of both genotypes were unaffected by ACE inhibition. We then investigated the effects of angiotensin AT1 (losartan) and AT2 (PD 123319) receptor antagonists on the locomotor activity of male NK1R-/- and wildtype mice. Both antagonists increased the locomotor activity of NK1R-/- mice, but neither affected the wildtypes. Finally, we tested the effects of captopril on the performance of male NK1R-/- and wildtype mice in the 5-choice serial reaction-time task (5-CSRTT) and found that ACE inhibition prevented the impulsivity of NK1R-/- mice. These results indicate that certain behaviours, disrupted in ADHD, are influenced by an interaction between the BRAS and NK1R, and suggest that ACE inhibitors could provide a novel treatment for this disorder. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

Prevention of chemotherapy-induced nausea: the role of neurokinin-1 (NK1) receptor antagonists.

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Bošnjak, Snežana M; Gralla, Richard J; Schwartzberg, Lee

2017-05-01

Chemotherapy-induced nausea (CIN) has a significant negative impact on the quality of life of cancer patients. The use of 5-hydroxytryptamine-3 (5-HT 3 ) receptor antagonists (RAs) has reduced the risk of vomiting, but (except for palonosetron) their effect on nausea, especially delayed nausea, is limited. This article reviews the role of NK 1 RAs when combined with 5-HT 3 RA-dexamethasone in CIN prophylaxis. Aprepitant has not shown consistent superiority over a two-drug (ondansetron-dexamethasone) combination in nausea control after cisplatin- or anthracycline-cyclophosphamide (AC)-based highly emetogenic chemotherapy (HEC). Recently, dexamethasone and dexamethasone-metoclopramide were demonstrated to be non-inferior to aprepitant and aprepitant-dexamethasone, respectively, for the control of delayed nausea after HEC (AC/cisplatin), and are now recognized in the guidelines. The potential impact of the new NK 1 RAs rolapitant and netupitant (oral fixed combination with palonosetron, as NEPA) in CIN prophylaxis is discussed. While the clinical significance of the effect on nausea of the rolapitant-granisetron-dexamethasone combination after cisplatin is not conclusive, rolapitant addition showed no improvement in nausea prophylaxis after AC or moderately emetogenic chemotherapy (MEC). NEPA was superior to palonosetron in the control of nausea after HEC (AC/cisplatin). Moreover, the efficacy of NEPA in nausea control was maintained over multiple cycles of HEC/MEC. Recently, NK 1 RAs have been challenged by olanzapine, with olanzapine showing superior efficacy in nausea prevention after HEC. Fixed antiemetic combinations (such as NEPA) or new antiemetics with a long half-life that may be given once per chemotherapy cycle (rolapitant or NEPA) may improve patient compliance with antiemetic treatment.

Role of Neurokinin 3 Receptor Signaling in Oral Squamous Cell Carcinoma.

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Obata, Kyoichi; Shimo, Tsuyoshi; Okui, Tatsuo; Matsumoto, Kenichi; Takada, Hiroyuki; Takabatake, Kiyofumi; Kunisada, Yuki; Ibaragi, Soichiro; Yoshioka, Norie; Kishimoto, Koji; Nagatsuka, Hitoshi; Sasaki, Akira

2017-11-01

The neurokinin 3 receptor (NK-3R) is differentially expressed in the central nervous system including cases of human oral squamous cell carcinoma. However, the role of NK-3R signaling in oral squamous cell carcinoma is not well known. NK-3R expression in surgically resected oral squamous cell carcinoma was examined immunohistochemically and the strength of the expression was quantified. We evaluated the function of NK-3R signaling using NK-3R antagonist in human oral squamous cell carcinoma bone invasion mouse model. NK-3R was significantly expressed in tumor cells that had invaded the bone matrix compared to the oral side tumor cells. SB222200, a selective antagonist of NK-3R, significantly suppressed the radiographic osteolytic lesion and tumorigenesis. NK-3R signaling is a potential target for the treatment of oral squamous cell carcinoma in cases of bone destruction. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Targeted deletion of neurokinin-1 receptor expressing nucleus tractus solitarii neurons precludes somatosensory depression of arterial baroreceptor-heart rate reflex.

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Potts, J T; Fong, A Y; Anguelov, P I; Lee, S; McGovern, D; Grias, I

2007-03-30

Neurokinin-1 receptor (NK1-R) expressing neurons are densely distributed throughout the nucleus tractus solitarii (NTS). However, their fundamental role in arterial baroreflex function remains debated. Previously, our group has shown that activation of contraction-sensitive somatic afferents evoke substance P (SP) release in the NTS and resets the arterial baroreflex via activation of a GABAergic NTS circuit. Based on these findings, we hypothesized that modulation of arterial baroreflex function by somatic afferents is mediated by NK1-R dependent inhibition of barosensitive NTS circuits. In the present study, SP-conjugated saporin toxin (SP-SAP) was used to ablate NK1-R expressing NTS neurons. Contraction-sensitive somatic afferents were activated by electrically-evoked muscle contraction and the arterial baroreceptor-heart rate reflex was assessed by constructing reflex curves using a decerebrate, arterially-perfused preparation. Baseline baroreflex sensitivity was significantly attenuated in SP-SAP-treated rats compared with control rats receiving either unconjugated SAP or vehicle. Muscle contraction significantly attenuated baroslope in SAP and vehicle-treated animals and shifted the baroreflex curves to higher systemic pressure. In contrast, somatic afferent stimulation failed to alter baroslope or shift the baroreflex curves in SP-SAP-treated animals. Moreover, when reflex sensitivity was partially restored in SP-SAP animals, somatic stimulation failed to attenuate baroreflex bradycardia. In contrast, SP-SAP and somatic stimulation failed to blunt the reflex bradycardia evoked by the peripheral chemoreflex. Immunohistochemistry revealed that pretreatment with SP-SAP significantly reduced the number of NK1-R expressing neurons in the caudal NTS, while sparing NK1-R expressing neurons rostral to the injection site. This was accompanied by a significant reduction in the number of glutamic acid decarboxylase (GAD67) expressing neurons at equivalent levels of the

Discovery of Indazoles as Potent, Orally Active Dual Neurokinin 1 Receptor Antagonists and Serotonin Transporter Inhibitors for the Treatment of Depression.

Science.gov (United States)

Degnan, Andrew P; Tora, George O; Huang, Hong; Conlon, David A; Davis, Carl D; Hanumegowda, Umesh M; Hou, Xiaoping; Hsiao, Yi; Hu, Joanna; Krause, Rudolph; Li, Yu-Wen; Newton, Amy E; Pieschl, Rick L; Raybon, Joseph; Rosner, Thorsten; Sun, Jung-Hui; Taber, Matthew T; Taylor, Sarah J; Wong, Michael K; Zhang, Huiping; Lodge, Nicholas J; Bronson, Joanne J; Macor, John E; Gillman, Kevin W

2016-12-21

Combination studies of neurokinin 1 (NK1) receptor antagonists and serotonin-selective reuptake inhibitors (SSRIs) have shown promise in preclinical models of depression. Such a combination may offer important advantages over the current standard of care. Herein we describe the discovery and optimization of an indazole-based chemotype to provide a series of potent dual NK1 receptor antagonists/serotonin transporter (SERT) inhibitors to overcome issues of ion channel blockade. This effort culminated in the identification of compound 9, an analogue that demonstrated favorable oral bioavailability, excellent brain uptake, and robust in vivo efficacy in a validated depression model. Over the course of this work, a novel heterocycle-directed asymmetric hydrogenation was developed to facilitate installation of the key stereogenic center.

Neurokinin-1 enables measles virus trans-synaptic spread in neurons

International Nuclear Information System (INIS)

Makhortova, Nina R.; Askovich, Peter; Patterson, Catherine E.; Gechman, Lisa A.; Gerard, Norma P.; Rall, Glenn F.

2007-01-01

Measles virus (MV), a morbillivirus that remains a significant human pathogen, can infect the central nervous system, resulting in rare but often fatal diseases, such as subacute sclerosing panencephalitis. Previous work demonstrated that MV was transmitted trans-synaptically and that, while a cellular receptor for the hemagglutinin (H) protein was required for MV entry, it was dispensable for subsequent cell-to-cell spread. Here, we explored what role the other envelope protein, fusion (F), played in trans-synaptic transport. We made the following observations: (1) MV-F expression in infected neurons was similar to that seen in infected fibroblasts; (2) fusion inhibitory peptide (FIP), an inhibitor of MV fusion, prevented both infection and spread in primary neurons; (3) Substance P, a neurotransmitter with the same active site as FIP, also blocked neuronal MV spread; and (4) both genetic deletion and pharmacological inhibition of the Substance P receptor, neurokinin-1 (NK-1), reduced infection of susceptible mice. Together, these data implicate a role for NK-1 in MV CNS infection and spread, perhaps serving as an MV-F receptor or co-receptor on neurons

Casopitant: a novel NK(1)-receptor antagonist in the prevention of chemotherapy-induced nausea and vomiting

DEFF Research Database (Denmark)

Ruhlmann, Christina; Herrstedt, Jørn

2009-01-01

Chemotherapy-induced nausea and vomiting (CINV) are among the most feared and distressing symptoms experienced by patients with cancer. The knowledge of the pathogenesis and neuropharmacology of CINV has expanded enormously over the last decades, the most significant discoveries being the role of 5......-hydroxytryptamine (5-HT)(3)- and neurokinin (NK)(1) receptors in the emetic reflex arch. This has led to the development of two new classes of antiemetics acting as highly selective antagonists at one of these receptors. These drugs have had a huge impact in the protection from chemotherapy-induced vomiting...

Neurokinin-1 Receptor Immunoreactive Neuronal Elements in the Superficial Dorsal Horn of the Chicken Spinal Cord: With Special Reference to Their Relationship with the Tachykinin-containing Central Axon Terminals in Synaptic Glomeruli

International Nuclear Information System (INIS)

Sakamoto, Hiroshi; Kawate, Toyoko; Li, Yongnan; Atsumi, Saoko

2009-01-01

Synaptic glomeruli that involve tachykinin-containing primary afferent central terminals are numerous in lamina II of the chicken spinal cord. Therefore, a certain amount of noxious information is likely to be modulated in these structures in chickens. In this study, we used immunohistochemistry with confocal and electron microscopy to investigate whether neurokinin-1 receptor (NK-1R)-expressing neuronal elements are in contact with the central primary afferent terminals in synaptic glomeruli of the chicken spinal cord. We also investigated which neuronal elements (axon terminals, dendrites, cell bodies) and which neurons in the spinal cord possess NK-1R, and are possibly influenced by tachykinin in the glomeruli. By confocal microscopy, NK-1R immunoreactivities were seen in a variety of neuronal cell bodies, their dendrites and smaller fibers of unknown origin. Some of the NK-1R immunoreactive profiles also expressed GABA immunoreactivities. A close association was observed between the NK-1R-immunoreactive neurons and tachykinin-immunoreactive axonal varicosities. By electron microscopy, NK-1R immunoreactivity was seen in cell bodies, conventional dendrites and vesicle-containing dendrites in laminae I and II. Among these elements, dendrites and vesicle-containing dendrites made contact with tachykinin-containing central terminals in the synaptic glomeruli. These results indicate that tachykinin-containing central terminals in the chicken spinal cord can modulate second-order neuronal elements in the synaptic glomeruli

Do Substance P and Neurokinin A Play Important Roles in the Control of LH Secretion in Ewes?

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Fergani, Chrysanthi; Mazzella, Leanne; Coolen, Lique M.; McCosh, Richard B.; Hardy, Steven L.; Newcomb, Nora; Grachev, Pasha; Lehman, Michael N.

2016-01-01

There is now general agreement that neurokinin B (NKB) acts via neurokinin-3-receptor (NK3R) to stimulate secretion of GnRH and LH in several species, including rats, mice, sheep, and humans. However, the roles of two other tachykinins, substance P (SP) and neurokinin A, which act primarily via NK1R and NK2R, respectively, are less clear. In rodents, these signaling pathways can stimulate LH release and substitute for NKB signaling; in humans, SP is colocalized with kisspeptin and NKB in the mediobasal hypothalamus. In this study, we examined the possible role of these tachykinins in control of the reproductive axis in sheep. Immunohistochemistry was used to describe the expression of SP and NK1R in the ovine diencephalon and determine whether these proteins are colocalized in kisspeptin or GnRH neurons. SP-containing cell bodies were largely confined to the arcuate nucleus, but NK1R-immunoreactivity was more widespread. However, there was very low coexpression of SP or NK1R in kisspeptin cells and none in GnRH neurons. We next determined the minimal effective dose of these three tachykinins that would stimulate LH secretion when administered into the third ventricle of ovary-intact anestrous sheep. A much lower dose of NKB (0.2 nmol) than of neurokinin A (2 nmol) or SP (10 nmol) consistently stimulated LH secretion. Moreover, the relative potency of these three neuropeptides parallels the relative selectivity of NK3R. Based on these anatomical and pharmacological data, we conclude that NKB-NK3R signaling is the primary pathway for the control of GnRH secretion by tachykinins in ewes. PMID:27704950

The substance P/NK-1 receptor system

Indian Academy of Sciences (India)

2015-04-27

Apr 27, 2015 ... NK-1 receptor may be a promising target in the treatment of cancer; NK-1 ... the contribution of chemotherapy for adult malignancies .... nisms that regulate cellular excitability and function. ..... positive expression of Ki-67 in dysplastic epithelium ..... emotional behaviour (behaviour traits, such as depression),.

Identification of a Novel Substance P–Neurokinin-1 Receptor MicroRNA-221-5p Inflammatory Network in Human Colonic Epithelial CellsSummary

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Kai Fang

2015-09-01

Full Text Available Background & Aims: Substance P (SP, a neuropeptide member of the tachykinin family, plays a critical role in colitis. MicroRNAs (miRNAs are small noncoding RNAs that negatively regulate gene expression. We examined whether SP modulates expression of microRNAs in human colonic epithelial cells. Methods: We performed microRNA profiling analysis of SP-stimulated human colonic epithelial NCM460 cells overexpressing neurokinin-1 receptor (NCM460-NK-1R. Targets of SP-regulated microRNAs were validated by real-time polymerase chain reaction (RT-PCR. Functions of miRNAs were tested in NCM460-NK-1R cells and the trinitrobenzene sulfonic acid (TNBS and dextran sulfate sodium (DSS models of colitis. Results: SP stimulated differential expression of 29 microRNAs, including miR-221-5p, the highest up-regulated miR (by 12.6-fold upon SP stimulation. Bioinformatic and luciferase reporter analyses identified interleukin-6 receptor (IL-6R mRNA as a direct target of miR-221-5p in NCM460 cells. Accordingly, SP exposure of NCM460-NK-1R cells increased IL-6R mRNA expression, and overexpression of miR-221-5p reduced IL-6R expression. Nuclear factor κB and c-Jun N-terminal kinase inhibition decreased SP-induced miR-221-5p expression. MiR-221-5p expression was increased in both TNBS- and DSS-induced colitis and in colonic biopsy samples from ulcerative colitis but not Crohn’s disease patients compared with controls. In mice, intracolonic administration of a miR-221-5p chemical inhibitor exacerbated TNBS- and DSS-induced colitis and increased colonic tumor necrosis factor-α, C-X-C motif chemokine 10 (Cxcl10, and collagen, type II, α 1 (Col2α1 mRNA expression. In situ hybridization in TNBS- and DSS-exposed colons revealed increased miR-221-5p expression primarily in colonocytes. Conclusions: Our results reveal a novel NK-1R-miR-221-5p-IL-6R network that protects from colitis. The use of miR-221-5p mimics may be a promising approach for colitis

The Neurokinin-1 Receptor Contributes to the Early Phase of Lipopolysaccharide-Induced Fever via Stimulation of Peripheral Cyclooxygenase-2 Protein Expression in Mice

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Eszter Pakai

2018-02-01

Full Text Available Neurokinin (NK signaling is involved in various inflammatory processes. A common manifestation of systemic inflammation is fever, which is usually induced in animal models with the administration of bacterial lipopolysaccharide (LPS. A role for the NK1 receptor was shown in LPS-induced fever, but the underlying mechanisms of how the NK1 receptor contributes to febrile response, especially in the early phase, have remained unknown. We administered LPS (120 µg/kg, intraperitoneally to mice with the Tacr1 gene, i.e., the gene encoding the NK1 receptor, either present (Tacr1+/+ or absent (Tacr1−/− and measured their thermoregulatory responses, serum cytokine levels, tissue cyclooxygenase-2 (COX-2 expression, and prostaglandin (PG E2 concentration. We found that the LPS-induced febrile response was attenuated in Tacr1−/− compared to their Tacr1+/+ littermates starting from 40 min postinfusion. The febrigenic effect of intracerebroventricularly administered PGE2 was not suppressed in the Tacr1−/− mice. Serum concentration of pyrogenic cytokines did not differ between Tacr1−/− and Tacr1+/+ at 40 min post-LPS infusion. Administration of LPS resulted in amplification of COX-2 mRNA expression in the lungs, liver, and brain of the mice, which was statistically indistinguishable between the genotypes. In contrast, the LPS-induced augmentation of COX-2 protein expression was attenuated in the lungs and tended to be suppressed in the liver of Tacr1−/− mice compared with Tacr1+/+ mice. The Tacr1+/+ mice responded to LPS with a significant surge of PGE2 production in the lungs, whereas Tacr1−/− mice did not. In conclusion, the NK1 receptor is necessary for normal fever genesis. Our results suggest that the NK1 receptor contributes to the early phase of LPS-induced fever by enhancing COX-2 protein expression in the periphery. These findings advance the understanding of the crosstalk between NK signaling and the “cytokine-COX-2

Characterization of species-related differences in the pharmacology of tachykinin NK receptors 1, 2 and 3

OpenAIRE

2009-01-01

Abstract Tachykinin NK receptors (NKR) differ to a large degree among species with respect to their affinities for small molecule antagonists. The aims of the present study were to clone NKRs from gerbil (NK2R and NK3R) and dog (NK1R, NK2R, NK3R) in which the sequence was previously unknown and to investigate the potency of several NKR antagonists at all known human, dog, gerbil and rat NKRs. The NKR protein coding sequences were cloned and expressed in CHO cells. The in...

Neurokinin B receptor antagonism decreases luteinising hormone pulse frequency and amplitude and delays puberty onset in the female rat.

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Li, S Y; Li, X F; Hu, M H; Shao, B; Poston, L; Lightman, S L; O'Byrne, K T

2014-08-01

The neural mechanisms controlling puberty onset remain enigmatic. Humans with loss of function mutations in TAC3 or TACR3, the genes encoding neurokinin B (NKB) or its receptor, neurokinin-3 receptor (NK3R), respectively, present with severe congenital gonadotrophin deficiency and pubertal failure. Animal studies have shown ambiguous actions of NKB-NK3R signalling with respect to controlling puberty onset. The present study aimed to determine the role of endogenous NKB-NK3R signalling in the control of pulsatile luteinising hormone (LH) secretion and the timing of puberty onset, and also whether precocious pubertal onset as a result of an obesogenic diet is similarly regulated by this neuropeptide system. Prepubertal female rats, chronically implanted with i.c.v. cannulae, were administered SB222200, a NK3R antagonist, or artificial cerebrospinal fluid via an osmotic mini-pump for 14 days. SB222200 significantly delayed the onset of vaginal opening and first oestrus (as markers of puberty) compared to controls in both normal and high-fat diet fed animals. Additionally, serial blood sampling, via chronic indwelling cardiac catheters, revealed that the increase in LH pulse frequency was delayed and that the LH pulse amplitude was reduced in response to NK3R antagonism, regardless of dietary status. These data suggest that endogenous NKB-NK3R signalling plays a role in controlling the timing of puberty and the associated acceleration of gonadotrophin-releasing hormone pulse generator frequency in the female rat. © 2014 British Society for Neuroendocrinology.

Neurokinin-3 Receptor Activation in the Retrochiasmatic Area is Essential for the Full Preovulatory LH Surge in Ewes

Science.gov (United States)

Porter, Katrina L.; Hileman, Stanley M.; Hardy, Steven L.; Nestor, Casey C; Lehman, Michael N.; Goodman, Robert L.

2014-01-01

Neurokinin B (NKB) is essential for human reproduction and has been shown to stimulate LH secretion in several species, including sheep. Ewes express the neurokinin-3 receptor (NK3R) in the retrochiasmatic area (RCh) and there is one report that placement of senktide, an NK3R agonist, therein stimulates LH secretion that resembles an LH surge in ewes. In this study, we first confirmed that local administration of senktide to the RCh produced a surge-like increase in LH secretion, and then tested the effects of this agonist in two other areas implicated in the control of LH secretion and where NK3R is found in high abundance: the preoptic area (POA) and arcuate nucleus (ARC). Bilateral microimplants containing senktide induced a dramatic surge-like increase in LH when given in the POA similar to that seen with RCh treatment. In contrast, senktide treatment in the ARC resulted in a much smaller, but significant, increase in LH concentrations suggestive of an effect on tonic secretion. The possible role of POA and RCh NK3R activation in the LH surge was next tested by treating ewes with SB222200, an NK3R antagonist, in each area during an E2-induced LH surge. SB222200 in the RCh, but not in the POA, reduced LH surge amplitude by about 40% compared to controls, indicating that NK3R activation in the former region is essential for full expression of the preovulatory LH surge. Based on these data, we propose that NKB actions in the RCh are an important component of the preovulatory LH surge in ewes. PMID:25040132

TGF-β1 Downregulates the Expression of CX3CR1 by Inducing miR-27a-5p in Primary Human NK Cells.

Science.gov (United States)

Regis, Stefano; Caliendo, Fabio; Dondero, Alessandra; Casu, Beatrice; Romano, Filomena; Loiacono, Fabrizio; Moretta, Alessandro; Bottino, Cristina; Castriconi, Roberta

2017-01-01

Activity of human natural killer (NK) cells against cancer cells is deeply suppressed by TGF-β1, an immunomodulatory cytokine that is released and activated in the tumor microenvironment. Moreover, our previous data showed that TGF-β1 modifies the chemokine receptor repertoire of NK cells. In particular, it decreases the expression of CX 3 CR1 that drives these effectors toward peripheral tissues, including tumor sites. To identify possible mechanisms mediating chemokine receptors modulation, we analyzed the microRNA profile of TGF-β1-treated primary NK cells. The analysis pointed out miR-27a-5p as a possible modulator of CX 3 CR1. We demonstrated the functional interaction of miR-27a-5p with the 3' untranslated region (3'UTR) of CX 3 CR1 mRNA by two different experimental approaches: by the use of a luciferase assay based on a reporter construct containing the CX 3 CR1 3'UTR and by transfection of primary NK cells with a miR-27a-5p inhibitor. We also showed that the TGF-β1-mediated increase of miR-27a-5p expression is a consequence of miR-23a-27a-24-2 cluster induction. Moreover, we demonstrated that miR-27a-5p downregulates the surface expression of CX 3 CR1. Finally, we showed that neuroblastoma cells induced in resting NK cells a downregulation of the CX 3 CR1 expression that was paralleled by a significant increase of miR-27a-5p expression. Therefore, the present study highlights miR-27a-5p as a pivotal TGF-β1-induced regulator of CX 3 CR1 expression.

Role of Kisspeptin and Neurokinin B in Puberty in Female Non-Human Primates

OpenAIRE

Ei Terasawa; Ei Terasawa; James P. Garcia; Stephanie B. Seminara; Kim L. Keen

2018-01-01

In human patients, loss-of-function mutations in the genes encoding kisspeptin (KISS1) and neurokinin B (NKB) and their receptors (KISS1R and NK3R, respectively) result in an abnormal timing of puberty or the absence of puberty. To understand the neuroendocrine mechanism of puberty, we investigated the contribution of kisspeptin and NKB signaling to the pubertal increase in GnRH release using rhesus monkeys as a model. Direct measurements of GnRH and kisspeptin in the median eminence of the h...

Radiodinated L-703,606: a potent selective antagonist to the human NK[sub 1] receptor

Energy Technology Data Exchange (ETDEWEB)

Francis, B E; Burns, H D [Merck Research Labs., West Point, PA (United States). Dept. of Radiopharmacology; Swain, C; Sabin, V [Merck Sharp and Dohme Research Labs., Harlow (United Kingdom). The Neuroscience Centre

1994-01-01

A new, radioiodinated, NK[sub 1] selective radiotracer ([[sup 125]I]L-703,606) was prepared. L-703,606 is an iodinated analog of the NK[sub 1] antagonist CP-96,345 in which the methoxy group has been replaced by an iodine substituent. [[sup 125]I]L-703,606 was made from the corresponding trimethylsilyl compound by treatment with no carrier added Na[sub 125]I and an Iodobead in TFA. The tracer was prepared at a specific activity of approx. 1100 Ci/mmol and preliminary binding studies demonstrated that [[sup 125]I]L=703,606 binds selectively to NK[sub 1] receptors. These results suggest that this radioligand will be useful for the biochemical and pharmacological characterization of the human NK[sub 1] receptor and, if labeled with I-123, may be useful for non-invasive NK[sub 1] receptor imaging via SPECT. (author).

Sex differences in the neurokinin B system in the human infundibular nucleus

NARCIS (Netherlands)

Taziaux, Melanie; Swaab, Dick F.; Bakker, Julie

2012-01-01

The recent report that loss-of-function mutations in either the gene encoding neurokinin B (NKB) or its receptor (NK3R) produce gonadotropin deficiencies in humans strongly points to NKB as a key regulator of GnRH release. We used NKB immunohistochemistry on postmortem human brain tissue to

KHYG-1 and NK-92 represent different subtypes of LFA-1-mediated NK cell adhesiveness.

Science.gov (United States)

Suck, Garnet; Tan, Suet-Mien; Chu, Sixian; Niam, Madelaine; Vararattanavech, Ardcharaporn; Lim, Tsyr Jong; Koh, Mickey B C

2011-01-01

Novel cancer cellular therapy approaches involving long-term ex vivo IL-2 stimulated highly cytotoxic natural killer (NK) cells are emerging. However, adhesion properties of such NK cells are not very well understood. Herein, we describe the novel observation of permanently activated alphaLbeta2 integrin leukocyte function-associated antigen (LFA)-1 adhesion receptor in long-term IL-2 activated NK cells and the permanent NK cell lines KHYG-1 and NK-92. We show that such cytokine activated NK effectors constitutively adhered to the LFA-1-ligand ICAM-1, whereas binding to the lower affinity ligand ICAM-3 required additional exogenous activating conditions. The results demonstrate an extended conformation and an intermediate affinity state for the LFA-1 population expressed by the NK cells. Interestingly, adhesion to ICAM-1 or K562 induced pronounced cell spreading in KHYG-1, but not in NK-92, and partially in long-term IL-2 stimulated primary NK cells. It is conceivable that such differential adhesion characteristics may impact motility potential of such NK effectors with relevance to clinical tumor targeting. KHYG-1 could be a useful model in planning future targeted therapeutic approaches involving NK effectors with augmented functions.

The Adhesion G Protein-Coupled Receptor GPR56/ADGRG1 Is an Inhibitory Receptor on Human NK Cells

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Gin-Wen Chang

2016-05-01

Full Text Available Natural killer (NK cells possess potent cytotoxic mechanisms that need to be tightly controlled. Here, we explored the regulation and function of GPR56/ADGRG1, an adhesion G protein-coupled receptor implicated in developmental processes and expressed distinctively in mature NK cells. Expression of GPR56 was triggered by Hobit (a homolog of Blimp-1 in T cells and declined upon cell activation. Through studying NK cells from polymicrogyria patients with disease-causing mutations in ADGRG1, encoding GPR56, and NK-92 cells ectopically expressing the receptor, we found that GPR56 negatively regulates immediate effector functions, including production of inflammatory cytokines and cytolytic proteins, degranulation, and target cell killing. GPR56 pursues this activity by associating with the tetraspanin CD81. We conclude that GPR56 inhibits natural cytotoxicity of human NK cells.

Tachykinins activate guinea-pig alveolar macrophages: involvement of NK2 and NK1 receptors.

OpenAIRE

Brunelleschi, S.; Vanni, L.; Ledda, F.; Giotti, A.; Maggi, C. A.; Fantozzi, R.

1990-01-01

1. The effects of substance P (SP), neurokinin A (NKA) and neurokinin B (NKB) were evaluated on superoxide anion (O2-.) production by guinea-pig alveolar macrophages (AM). 2. SP dose-dependently (ED50 = 0.7 nM) evoked O2-. production from guinea-pig AM; the N-terminal heptapeptide, SP(1-7), was ineffective. In the presence of thiorphan (10(-5) M), an enkephalinase inhibitor, the stimulating effects of SP were not significantly modified. NKA and NKB were both able to induce O2-. production fro...

Synthesis of two potential NK1-receptor ligands using [1-11C]ethyl iodide and [1-11C]propyl iodide and initial PET-imaging

Directory of Open Access Journals (Sweden)

Genchel Tove

2007-07-01

Full Text Available Abstract Background The previously validated NK1-receptor ligand [O-methyl-11C]GR205171 binds with a high affinity to the NK1-receptor and displays a slow dissociation from the receptor. Hence, it cannot be used in vivo for detecting concentration changes in substance P, the endogenous ligand for the NK1-receptor. A radioligand used for monitoring these changes has to enable displacement by the endogenous ligand and thus bind reversibly to the receptor. Small changes in the structure of a receptor ligand can lead to changes in binding characteristics and also in the ability to penetrate the blood-brain barrier. The aim of this study was to use carbon-11 labelled ethyl and propyl iodide with high specific radioactivity in the synthesis of two new and potentially reversible NK1-receptor ligands with chemical structures based on [O-methyl-11C]GR205171. Methods [1-11C]Ethyl and [1-11C]propyl iodide with specific radioactivities of 90 GBq/μmol and 270 GBq/μmol, respectively, were used in the synthesis of [O-methyl-11C]GR205171 analogues by alkylation of O-desmethyl GR205171. The brain uptake of the obtained (2S,3S-N-(1-(2- [1-11C]ethoxy-5-(3-(trifluoromethyl-4H-1,2,4-triazol-4-ylphenylethyl-2-phenylpiperidin-3-amine (I and (2S,3S-2-phenyl-N-(1-(2- [1-11C]propoxy-5-(3-(trifluoromethyl-4H-1,2,4-triazol-4-ylphenylethylpiperidin-3-amine (II was studied with PET in guinea pigs and rhesus monkeys and compared to the uptake of [O-methyl-11C]GR205171. Results All ligands had similar uptake distribution in the guinea pig brain. The PET-studies in rhesus monkeys showed that (II had no specific binding in striatum. Ligand (I had moderate specific binding compared to the [O-methyl-11C]GR205171. The ethyl analogue (I displayed reversible binding characteristics contrary to the slow dissociation rate shown by [O-methyl-11C]GR205171. Conclusion The propyl-analogue (II cannot be used for detecting changes in NK1-ligand levels, while further studies should be

The Murine Natural Cytotoxic Receptor NKp46/NCR1 Controls TRAIL Protein Expression in NK Cells and ILC1s

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Sam Sheppard

2018-03-01

Full Text Available Summary: TRAIL is an apoptosis-inducing ligand constitutively expressed on liver-resident type 1 innate lymphoid cells (ILC1s and a subset of natural killer (NK cells, where it contributes to NK cell anti-tumor, anti-viral, and immunoregulatory functions. However, the intrinsic pathways involved in TRAIL expression in ILCs remain unclear. Here, we demonstrate that the murine natural cytotoxic receptor mNKp46/NCR1, expressed on ILC1s and NK cells, controls TRAIL protein expression. Using NKp46-deficient mice, we show that ILC1s lack constitutive expression of TRAIL protein and that NK cells activated in vitro and in vivo fail to upregulate cell surface TRAIL in the absence of NKp46. We show that NKp46 regulates TRAIL expression in a dose-dependent manner and that the reintroduction of NKp46 in mature NK cells deficient for NKp46 is sufficient to restore TRAIL surface expression. These studies uncover a link between NKp46 and TRAIL expression in ILCs with potential implications in pathologies involving NKp46-expressing cells. : Sheppard et al. find that mice deficient in the activating receptor NCR1/NKp46 (Ncr1−/− fail to express the apoptosis-inducing ligand TRAIL at the surface of group 1 innate lymphoid cells (ILC1s. Keywords: NK cell, natural killer cell, NKp46, ILC1, TRAIL, IL-15, IL-2

Mapping Substance P Binding Sites on the Neurokinin-1 Receptor Using Genetic Incorporation of a Photoreactive Amino Acid

DEFF Research Database (Denmark)

Valentin-Hansen, Louise; Park, Minyoung; Huber, Thomas

2014-01-01

that the binding site for SP includes multiple domains in the N-terminal (Nt) segment and the second extracellular loop (ECLII) of NK1. To map precisely the NK1 residues that interact with SP, we applied a novel receptor-based targeted photocross-linking approach. We used amber codon suppression to introduce...... the photoreactive unnatural amino acid p-benzoyl-l-phenylalanine (BzF) at 11 selected individual positions in the Nt tail (residues 11-21) and 23 positions in the ECLII (residues 170(C-10)-193(C+13)) of NK1. The 34 NK1 variants were expressed in mammalian HEK293 cells and retained the ability to interact...

Peripheral substance P and neurokinin-1 receptors have a role in inflammatory and neuropathic orofacial pain models.

Science.gov (United States)

Teodoro, Fernanda C; Tronco Júnior, Marcos F; Zampronio, Aleksander R; Martini, Alessandra C; Rae, Giles A; Chichorro, Juliana G

2013-06-01

There is accumulating evidence that substance P released from peripheral sensory neurons participates in inflammatory and neuropathic pain. In this study it was investigated the ability of substance P to induce orofacial nociception and thermal and mechanical hyperalgesia, as well as the role of NK1 receptors on models of orofacial inflammatory and neuropathic pain. Substance P injected into the upper lip at 1, 10 and 100 μg/50 μL failed to induce nociceptive behavior. Also, substance P (0.1-10 μg/50 μL) injected into the upper lip did not evoke orofacial cold hyperalgesia and when injected at 1 μg/50 μL did not induce mechanical hyperalgesia. However, substance P at this latter dose induced orofacial heat hyperalgesia, which was reduced by the pre-treatment of rats with a non-peptide NK1 receptor antagonist (SR140333B, 3mg/kg). Systemic treatment with SR140333B (3 mg/kg) also reduced carrageenan-induced heat hyperalgesia, but did not exert any influence on carrageenan-induced cold hyperalgesia. Blockade of NK1 receptors with SR140333B also reduced by about 50% both phases of the formalin response evaluated in the orofacial region. Moreover, heat, but not cold or mechanical, hyperalgesia induced by constriction of the infraorbital nerve, a model of trigeminal neuropathic pain, was abolished by pretreatment with SR140333B. Considering that substance P was peripherally injected (i.e. upper lip) and the NK1 antagonist used lacks the ability to cross the blood-brain-barrier, our results demonstrate that the peripheral SP/NK1 system participates in the heat hyperalgesia associated with inflammation or nerve injury and in the persistent pain evoked by formalin in the orofacial region. Copyright © 2012 Elsevier Ltd. All rights reserved.

AHR prevents human IL-1R1hi ILC3 differentiation to natural killer cells

Science.gov (United States)

Hughes, Tiffany; Briercheck, Edward L.; Freud, Aharon G.; Trotta, Rossana; McClory, Susan; Scoville, Steven D.; Keller, Karen; Deng, Youcai; Cole, Jordan; Harrison, Nicholas; Mao, Charlene; Zhang, Jianying; Benson, Don M.; Yu, Jianhua; Caligiuri, Michael A.

2014-01-01

SUMMARY Accumulating evidence indicates that human natural killer (NK) cells develop in secondary lymphoid tissue (SLT) through a so-called “stage 3” developmental intermediate minimally characterized by a CD34-CD117+CD94- immunophenotype that lacks mature NK cell function. This stage 3 population is heterogeneous, potentially composed of functionally distinct innate lymphoid cell (ILC) types that includes interleukin-1 receptor (IL-1R1) positive, IL-22-producing ILC3s. Whether human ILC3s are developmentally related to NK cells is a subject of ongoing investigation. Here we show that antagonism of the aryl hydrocarbon receptor (AHR) or silencing of AHR gene expression promotes differentiation of tonsillar IL-22-producing IL-1R1hi human ILC3s to CD56brightCD94+ IFN-gamma-producing cytolytic mature NK cells expressing eomesodermin (EOMES) and T-Box Protein 21 (TBX21 or TBET). Hence, AHR is a transcription factor that prevents human IL-1R1hi ILC3s from differentiating into NK cells. PMID:24953655

Expression Profiles of Neuropeptides, Neurotransmitters, and Their Receptors in Human Keratocytes In Vitro and In Situ.

Science.gov (United States)

Słoniecka, Marta; Le Roux, Sandrine; Boman, Peter; Byström, Berit; Zhou, Qingjun; Danielson, Patrik

2015-01-01

Keratocytes, the quiescent cells of the corneal stroma, play a crucial role in corneal wound healing. Neuropeptides and neurotransmitters are usually associated with neuronal signaling, but have recently been shown to be produced also by non-neuronal cells and to be involved in many cellular processes. The aim of this study was to assess the endogenous intracellular and secreted levels of the neuropeptides substance P (SP) and neurokinin A (NKA), and of the neurotransmitters acetylcholine (ACh), catecholamines (adrenaline, noradrenaline and dopamine), and glutamate, as well as the expression profiles of their receptors, in human primary keratocytes in vitro and in keratocytes of human corneal tissue sections in situ. Cultured keratocytes expressed genes encoding for SP and NKA, and for catecholamine and glutamate synthesizing enzymes, as well as genes for neuropeptide, adrenergic and ACh (muscarinic) receptors. Keratocytes in culture produced SP, NKA, catecholamines, ACh, and glutamate, and expressed neurokinin-1 and -2 receptors (NK-1R and NK-2R), dopamine receptor D2, muscarinic ACh receptors, and NDMAR1 glutamate receptor. Human corneal sections expressed SP, NKA, NK-1R, NK-2R, receptor D2, choline acetyl transferase (ChAT), M3, M4 and M5 muscarinic ACh receptors, glutamate, and NMDAR1, but not catecholamine synthesizing enzyme or the α1 and β2 adrenoreceptors, nor M1 receptor. In addition, expression profiles assumed significant differences between keratocytes from the peripheral cornea as compared to those from the central cornea, as well as differences between keratocytes cultured under various serum concentrations. In conclusion, human keratocytes express an array of neuropeptides and neurotransmitters. The cells furthermore express receptors for neuropeptides/neurotransmitters, which suggests that they are susceptible to stimulation by these substances in the cornea, whether of neuronal or non-neuronal origin. As it has been shown that neuropeptides

DNAM-1 Expression Marks an Alternative Program of NK Cell Maturation

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Ludovic Martinet

2015-04-01

Full Text Available Natural killer (NK cells comprise a heterogeneous population of cells important for pathogen defense and cancer surveillance. However, the functional significance of this diversity is not fully understood. Here, we demonstrate through transcriptional profiling and functional studies that the activating receptor DNAM-1 (CD226 identifies two distinct NK cell functional subsets: DNAM-1+ and DNAM-1− NK cells. DNAM-1+ NK cells produce high levels of inflammatory cytokines, have enhanced interleukin 15 signaling, and proliferate vigorously. By contrast, DNAM-1− NK cells that differentiate from DNAM-1+ NK cells have greater expression of NK-cell-receptor-related genes and are higher producers of MIP1 chemokines. Collectively, our data reveal the existence of a functional program of NK cell maturation marked by DNAM-1 expression.

Neurokinin-1 receptor blocker CP-99 994 improved emesis induced by cisplatin via regulating the activity of gastric distention responsive neurons in the dorsal motor nucleus of vagus and enhancing gastric motility in rats.

Science.gov (United States)

Sun, X; Xu, L; Guo, F; Luo, W; Gao, S; Luan, X

2017-10-01

Nowadays, chemotherapy induced nausea and vomiting (CINV) is still common in patients with cancer. It was reported that substance P mediated CINV via neurokinin-1 (NK 1 ) receptor and antagonists of NK 1 receptor has been proved useful for treating CINV but the mechanism are not fully understood. This study aimed to examine the role of NK 1 receptor blocker, CP-99 994, when administrated into dorsal motor nucleus of vagus (DMNV), on the cisplatin-induced emesis in rats and the possible mechanism. Rats' kaolin intake, food intake, and bodyweight were recorded every day; gastric contraction activity was recorded in conscious rats through a force transducer implanted into the stomach; gastric emptying was monitored using the phenol red method; single unit extracellular firing in the DMNV were recorded. DMNV microinjection of CP-99 994 reduced the changes of increased kaolin consumption and suppressed food intake in cisplatin-treated rats; enhanced the gastric contraction activity dose-dependently in control and cisplatin-treated rats but enhanced gastric emptying only in cisplatin-treated rats; reduced the firing rate of gastric distention inhibited (GD-I) neurons but increased the firing rate of GD excited (GD-E) neurons in the DMNV. The effects of CP-99 994 on gastric motility and neuronal activity were stronger in cisplatin-treated rats than those of control rats. Our results suggested that CP-99 994 could improve emesis induced by cisplatin by regulating gastric motility and gastric related neuronal activity in the DMNV. © 2017 John Wiley & Sons Ltd.

A neurokinin 3 receptor-selective agonist accelerates pulsatile luteinizing hormone secretion in lactating cattle.

Science.gov (United States)

Nakamura, Sho; Wakabayashi, Yoshihiro; Yamamura, Takashi; Ohkura, Satoshi; Matsuyama, Shuichi

2017-07-01

Pulsatile gonadotropin-releasing hormone (GnRH) secretion, which is indispensable for follicular development, is suppressed in lactating dairy and beef cattle. Neurokinin B (NKB) neurons in the arcuate nucleus of the hypothalamus are considered to play an essential role in generating the pulsatile mode of GnRH/luteinizing hormone (LH) secretion. The present study aimed to clarify the role of NKB-neurokinin 3 receptor (NK3R) signaling in the pulsatile pattern of GnRH/gonadotropin secretion in postpartum lactating cattle. We examined the effects of the administration of an NK3R-selective agonist, senktide, on gonadotropin secretion in lactating cattle. The lactating cattle, at approximately 7 days postpartum, were intravenously infused with senktide (30 or 300 nmol/min) or vehicle for 24 h. The administration of 30 or 300 nmol/min senktide significantly increased LH pulse frequency compared to in the control group during 0-4 or 20-24 h after infusion, respectively. Moreover, LH and follicle-stimulating hormone levels were gradually increased by 300 nmol/min administration of senktide during the 0-4-h sampling period. Ultrasonography of the ovaries was performed to identify the first postpartum ovulation in senktide-administered lactating cattle. The interval from calving to first postpartum ovulation was significantly shorter in the 300 nmol/min senktide-administered group than in the control group. Taken together, these findings suggest that senktide infusion elicits an increase in LH pulse frequency that may stimulate follicular development and, in turn, induce the first postpartum ovulation in lactating cattle. © The Authors 2017. Published by Oxford University Press on behalf of Society for the Study of Reproduction. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Development and labeling of EP-00652218 analogues, NK1 receptors antagonist, for PET and SPECT imaging

International Nuclear Information System (INIS)

Bagot-Gueret, C.

2001-12-01

The aim of this work was the synthesis and radiosynthesis of compounds labelled either with a positron emitter (fluorine-18, t 1/2 = 109 minutes) or with a gamma emitter (iodine-123, t 1/2 = 16.2 hours), for Positron Emission Tomography (PET) and Single Photon Emission Computed Tomography (SPECT) studies. EP-00652218 is a novel potent antagonist, with a sub-nano-molar affinity towards the NK 1 receptors. In order to develop ligands that could be used either in PET or SPECT, we undertook the synthesis of poly-halogenated analogues of EP-00652218. Compound 17 was synthesized through two different synthetic pathways. A series of original compounds has been obtained from compound 17 by halogen exchanges on the naphthyridone or the benzene ring. These molecules were tested to determine their in vitro affinity towards NK 1 receptors. Compound 21 was labelled with fluorine-18 in 135 minutes and with a 20% radiochemical yield. Compound 26 was radioiodinated following reaction with Na 125 I (t 1/2 = 60.14 days) in a 18% radiochemical yield. Despite expectation, these analogues of EP-00652218 exhibited an insufficient affinity for NK 1 receptors (IC 50 = 10 -7 M) and thus unlikely usable for in vivo studies with PET and SPECT. (author)

Tachykinin NK2 receptor and functional mechanisms in human colon: changes with indomethacin and in diverticular disease and ulcerative colitis.

Science.gov (United States)

Burcher, Elizabeth; Shang, Fei; Warner, Fiona J; Du, Qin; Lubowski, David Z; King, Denis W; Liu, Lu

2008-01-01

Neurokinin A (NKA) is an important spasmogen in human colon. We examined inflammatory disease-related changes in the tachykinin NK(2) receptor system in human sigmoid colon circular muscle, using functional, radioligand binding, and quantitative reverse transcription-polymerase chain reaction methods. In circular muscle strips, indomethacin enhanced contractile responses to NKA (p diverticular disease (DD) specimens, indicating NK(2) receptor-mediated release of relaxant prostanoids. Contractile responses to both tachykinins were reduced in strips from DD (p disease patients, demonstrating that the change in responsiveness to tachykinins in disease is specifically mediated by the NK(2) receptor. In membranes from UC specimens, receptor affinity for (125)I-NKA (median K(D) 0.91 nM, n = 16) was lower (p disease-related changes in receptor number (B(max)) were found (mean, 2.0-2.5 fmol/mg of wet weight tissue), suggesting that the reduced contractile responses in disease are not due to a loss of receptor number. Different mechanisms may account for the reduced contractility in DD compared with UC. A gender-related difference in receptor density was seen in controls, with B(max) lower in females (1.77 fmol/mg, n = 15) than in males (2.60 fmol/mg, n = 25, p = 0.01). In contrast, no gender-related differences were seen in NK(2) receptor mRNA in control colonic muscle, indicating that the gender difference is a post-translational event.

Novel method for the study of receptor Ca2+ signalling exemplified by the NK1 receptor

DEFF Research Database (Denmark)

Heding, A; Elling, C E; Schwartz, T W

2002-01-01

We have used a novel technology (NovoStar from BMG Labtechnologies) for the study of the Ca2+ signalling of the human tackykinin NK1 (hNK-I receptor). The NovoStar is a microplate reader based on fluorescence and luminescence. The instrument implements a robotic pipettor arm and two microplate...... carriers, typically one for samples and one for cells. The robotic pipettor arm can transfer sample (agonist or antagonist) from the sample plate or other liquid containers to the cell plate, facilitating the study of Ca2+ signalling to such a degree that the instrument can be used for Medium Throughput...

Involvement of midbrain tectum neurokinin-mediated mechanisms in fear and anxiety

International Nuclear Information System (INIS)

Brenes, J.C.; Broiz, A.C.; Bassi, G.S.; Schwarting, R.K.W.; Brandão, M.L.

2012-01-01

Electrical stimulation of midbrain tectum structures, particularly the dorsal periaqueductal gray (dPAG) and inferior colliculus (IC), produces defensive responses, such as freezing and escape behavior. Freezing also ensues after termination of dPAG stimulation (post-stimulation freezing). These defensive reaction responses are critically mediated by Y -aminobutyric acid and 5-hydroxytryptamine mechanisms in the midbrain tectum. Neurokinins (NKs) also play a role in the mediation of dPAG stimulation-evoked fear, but how NK receptors are involved in the global processing and expression of fear at the level of the midbrain tectum is yet unclear. The present study investigated the role of NK-1 receptors in unconditioned defensive behavior induced by electrical stimulation of the dPAG and IC of male Wistar rats. Spantide (100 pmol/0.2 µL), a selective NK-1 antagonist, injected into these midbrain structures had anti-aversive effects on defensive responses and distress ultrasonic vocalizations induced by stimulation of the dPAG but not of the IC. Moreover, intra-dPAG injections of spantide did not influence post-stimulation freezing or alter exploratory behavior in rats subjected to the elevated plus maze. These results suggest that NK-1 receptors are mainly involved in the mediation of defensive behavior organized in the dPAG. Dorsal periaqueductal gray-evoked post-stimulation freezing was not affected by intra-dPAG injections of spantide, suggesting that NK-1-mediated mechanisms are only involved in the output mechanisms of defensive behavior and not involved in the processing of ascending aversive information from the dPAG

Involvement of midbrain tectum neurokinin-mediated mechanisms in fear and anxiety

Energy Technology Data Exchange (ETDEWEB)

Brenes, J.C. [Experimental and Physiological Psychology, Philipps-University of Marburg, Marburg (Germany); Broiz, A.C.; Bassi, G.S. [Instituto de Neurociências e Comportamento, Campus USP, Ribeirão Preto, SP (Brazil); Laboratório de Psicobiologia, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP (Brazil); Schwarting, R.K.W. [Experimental and Physiological Psychology, Philipps-University of Marburg, Marburg (Germany); Brandão, M.L. [Instituto de Neurociências e Comportamento, Campus USP, Ribeirão Preto, SP (Brazil); Laboratório de Psicobiologia, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP (Brazil)

2012-03-09

Electrical stimulation of midbrain tectum structures, particularly the dorsal periaqueductal gray (dPAG) and inferior colliculus (IC), produces defensive responses, such as freezing and escape behavior. Freezing also ensues after termination of dPAG stimulation (post-stimulation freezing). These defensive reaction responses are critically mediated by {sub Y}-aminobutyric acid and 5-hydroxytryptamine mechanisms in the midbrain tectum. Neurokinins (NKs) also play a role in the mediation of dPAG stimulation-evoked fear, but how NK receptors are involved in the global processing and expression of fear at the level of the midbrain tectum is yet unclear. The present study investigated the role of NK-1 receptors in unconditioned defensive behavior induced by electrical stimulation of the dPAG and IC of male Wistar rats. Spantide (100 pmol/0.2 µL), a selective NK-1 antagonist, injected into these midbrain structures had anti-aversive effects on defensive responses and distress ultrasonic vocalizations induced by stimulation of the dPAG but not of the IC. Moreover, intra-dPAG injections of spantide did not influence post-stimulation freezing or alter exploratory behavior in rats subjected to the elevated plus maze. These results suggest that NK-1 receptors are mainly involved in the mediation of defensive behavior organized in the dPAG. Dorsal periaqueductal gray-evoked post-stimulation freezing was not affected by intra-dPAG injections of spantide, suggesting that NK-1-mediated mechanisms are only involved in the output mechanisms of defensive behavior and not involved in the processing of ascending aversive information from the dPAG.

A microscale synthesis of a promising radiolabelled antitumor drug: cis-1,1-cyclobutanedicarboxylato (2R)-2-methyl-1,4-butanediamine platinum(II), NK121

International Nuclear Information System (INIS)

Suwa, Masato; Kogawa, Osamu; Hashimoto, Yutaka

1992-01-01

A promising antitumor drug, cis-1,1-cyclobutane-dicarboxylato (2R)-2-methyl-1,4-butanediamine platinum (II), NK121, was synthesized from radionuclides of platinum such as 193m Pt, 195m Pt and 191 Pt which were produced by neutron irradiation of enriched 192 Pt. The overall yield was 38.6% in a synthesis time of 10 hours. The radioactivities present in 8.39 mg of NK121 were 115.3 μCi as 193m Pt, 29.9 μCi as 197 Pt, 22.0 μCi as 195m Pt, and 4.8 μCi as 191 Pt at the end of synthesis. The specific activity of the NK121 was 13.7 μCi ( 193m Pt)/mg NK121 at the end of synthesis. The radiochemical purity of NK121 was typically 99%. HPLC analyses confirmed that NK121 was in an adequate chemical purity and suitable for animal experimentation. (author)

A microscale synthesis of a promising radiolabelled antitumor drug: cis-1,1-cyclobutanedicarboxylato (2R)-2-methyl-1,4-butanediamine platinum(II), NK121

Energy Technology Data Exchange (ETDEWEB)

Suwa, Masato; Kogawa, Osamu; Hashimoto, Yutaka (Nippon Kayaku Co. Ltd., Tokyo (Japan). Research Labs. of Pharmaceuticals Group); Nowatari, Hiroyoshi (Nippon Kayaku Co. Ltd., Takasaki, Gumma (Japan). Takasaki Research Labs.); Murase, Yuko; Homma, Yoshio (Kyoritsu Coll. of Pharmacy, Tokyo (Japan))

1992-05-01

A promising antitumor drug, cis-1,1-cyclobutane-dicarboxylato (2R)-2-methyl-1,4-butanediamine platinum (II), NK121, was synthesized from radionuclides of platinum such as {sup 193m}Pt, {sup 195m}Pt and {sup 191}Pt which were produced by neutron irradiation of enriched {sup 192}Pt. The overall yield was 38.6% in a synthesis time of 10 hours. The radioactivities present in 8.39 mg of NK121 were 115.3 {mu}Ci as {sup 193m}Pt, 29.9 {mu}Ci as {sup 197}Pt, 22.0 {mu}Ci as {sup 195m}Pt, and 4.8 {mu}Ci as {sup 191}Pt at the end of synthesis. The specific activity of the NK121 was 13.7 {mu}Ci ({sup 193m}Pt)/mg NK121 at the end of synthesis. The radiochemical purity of NK121 was typically 99%. HPLC analyses confirmed that NK121 was in an adequate chemical purity and suitable for animal experimentation. (author).

The neurokinin-3 receptor agonist senktide facilitates the integration of memories for object, place and temporal order into episodic memory.

Science.gov (United States)

Chao, Owen Y; Nikolaus, Susanne; Huston, Joseph P; de Souza Silva, Maria A

2014-10-01

Senktide, a potent neurokinin-3 receptor (NK3-R) agonist, has been shown to have promnestic effects in adult and aged rodents and to facilitate episodic-like memory (ELM) in mice when administrated before the learning trial. In the present study we assessed the effects of senktide on memory consolidation by administering it post-trial (after the learning trial) in adult rats. We applied an ELM test, based on the integrated memory for object, place and temporal order, which we developed (Kart-Teke, de Souza Silva, Huston, & Dere, 2006). This test involves two learning trials and one test trial. We examined intervals of 1h and 23 h between the learning and test trials (experiment 1) in untreated animals and found that they exhibited intact ELM after a delay of 1 h, but not 23 h. In another test for ELM performed 7 days later, vehicle or senktide (0.2 mg/kg, s.c.) was applied immediately after the second learning trial and the test was conducted 23 h later (experiment 2). Senktide treatment recovered components of ELM (memory for place and object) compared with vehicle-treated animals. After one more week, vehicle or senktide (0.2 mg/kg, s.c.) was applied post-trial and the test conducted 6h later (experiment 3). The senktide-treated group exhibited intact ELM, unlike the vehicle-treated group. Finally, animals received post-trial treatment with either vehicle or SR142801, a selective NK3-R antagonist (6 mg/kg, i.p.), 1 min before senktide injection (0.2 mg/kg, s.c.) in the ELM paradigm and were tested 6h later (experiment 4). The vehicle+senktide group showed intact ELM, while the SR142801+senktide group did not. The results indicate that senktide facilitated the consolidation or the expression of ELM and that the senktide effect was NK3-R dependent. Copyright © 2014 Elsevier Inc. All rights reserved.

Addition of the Neurokinin-1-Receptor Antagonist (RA) Aprepitant to a 5-Hydroxytryptamine-RA and Dexamethasone in the Prophylaxis of Nausea and Vomiting Due to Radiation Therapy With Concomitant Cisplatin

Energy Technology Data Exchange (ETDEWEB)

Jahn, Franziska, E-mail: franziska.jahn@uk-halle.de [Department of Hematology/Oncology, Martin-Luther-University Halle-Wittenberg, Halle (Saale) (Germany); Riesner, Anica [Department of Gastroenterology, Martin-Luther-University Halle-Wittenberg, Halle (Saale) (Germany); Jahn, Patrick [Nursing Research Unit, Martin-Luther-University Halle-Wittenberg, Halle (Saale) (Germany); Sieker, Frank; Vordermark, Dirk [Department of Radiation Oncology, Martin-Luther-University Halle-Wittenberg, Halle (Saale) (Germany); Jordan, Karin [Department of Hematology/Oncology, Martin-Luther-University Halle-Wittenberg, Halle (Saale) (Germany)

2015-08-01

Purpose: To assess, in a prospective, observational study, the safety and efficacy of the addition of the neurokinin-1-receptor antagonist (NK1-RA) aprepitant to concomitant radiochemotherapy, for the prophylaxis of radiation therapy–induced nausea and vomiting. Patients and Methods: This prospective observational study compared the antiemetic efficacy of an NK1-RA (aprepitant), a 5-hydroxytryptamine-RA, and dexamethasone (aprepitant regimen) versus a 5-hydroxytryptamine-RA and dexamethasone (control regimen) in patients receiving concomitant radiochemotherapy with cisplatin at the Department of Radiation Oncology, University Hospital Halle (Saale), Germany. The primary endpoint was complete response in the overall phase, defined as no vomiting and no use of rescue therapy in this period. Results: Fifty-nine patients treated with concomitant radiochemotherapy with cisplatin were included in this study. Thirty-one patients received the aprepitant regimen and 29 the control regimen. The overall complete response rates for cycles 1 and 2 were 75.9% and 64.5% for the aprepitant group and 60.7% and 54.2% for the control group, respectively. Although a 15.2% absolute difference was reached in cycle 1, a statistical significance was not detected (P=.22). Furthermore maximum nausea was 1.58 ± 1.91 in the control group and 0.73 ± 1.79 in the aprepitant group (P=.084); for the head-and-neck subset, 2.23 ± 2.13 in the control group and 0.64 ± 1.77 in the aprepitant group, respectively (P=.03). Conclusion: This is the first study of an NK1-RA–containing antiemetic prophylaxis regimen in patients receiving concomitant radiochemotherapy. Although the primary endpoint was not obtained, the absolute difference of 10% in efficacy was reached, which is defined as clinically meaningful for patients by international guidelines groups. Randomized phase 3 studies are necessary to further define the potential role of an NK1-RA in this setting.

γ-Preprotachykinin-(72-92)-peptide amide: An endogenous preprotachykinin I gene-derived peptide that preferentially binds to neurokinin-2 receptors

International Nuclear Information System (INIS)

Dam, T.V.; Takeda, Y.; Krause, J.E.; Escher, E.; Quirion, R.

1990-01-01

The presence of N-terminally extended forms of neurokinin A has recently been reported in the mammalian brain. Among them, gamma-preprotachykinin-(72-92)-peptide amide [gamma-PPT-(72-92)-NH2], a peptide derived by posttranslational processing of gamma-preprotachykinin, is most prominent. We report here that this peptide most likely acts on neurokinin-2 receptor sites since neurokinin A (a putative neurokinin-2 agonist) and gamma-PPT-(72-92)-NH2 are potent competitors of 125I-labeled gamma-PPT-(72-92)-NH2 binding whereas selective neurokinin-1 and -3 agonists are not. Moreover, the distribution of 125I-labeled gamma-PPT-(72-92)-NH2 and 125I-labeled neurokinin A binding sites are very similar in rat brain. On the other hand, 125I-labeled Bolton-Hunter-substance P (a neurokinin-1 ligand) and 125I-labeled Bolton-Hunter-eledoisin (a neurokinin-3 ligand) binding sites are differentially located in this tissue. Thus, it appears that gamma-PPT-(72-92)-NH2 binds to neurokinin-2 receptors and should be considered as a putative endogenous ligand for this receptor class

Targeting the neurokinin receptor 1 with aprepitant: a novel antipruritic strategy.

Science.gov (United States)

Ständer, Sonja; Siepmann, Dorothee; Herrgott, Ilka; Sunderkötter, Cord; Luger, Thomas A

2010-06-04

Chronic pruritus is a global clinical problem with a high impact on the quality of life and lack of specific therapies. It is an excruciating and frequent symptom of e.g. uncurable renal, liver and skin diseases which often does not respond to conventional treatment with e.g. antihistamines. Therefore antipruritic therapies which target physiological mechanisms of pruritus need to be developed. Substance P (SP) is a major mediator of pruritus. As it binds to the neurokinin receptor 1 (NKR1), we evaluated if the application of a NKR1 antagonist would significantly decrease chronic pruritus. Twenty hitherto untreatable patients with chronic pruritus (12 female, 8 male; mean age, 66.7 years) were treated with the NKR1 antagonist aprepitant 80 mg for one week. 16 of 20 patients (80%) experienced a considerable reduction of itch intensity, as assessed by the visual analog scale (VAS, range 0 to 10). Considering all patients, the mean value of pruritus intensity was significantly reduced from 8.4 VAS points (SD +/-1.7) before treatment to 4.9 VAS points (SD +/-3.2) (pprofit from the treatment. Side-effects were mild (nausea, vertigo, and drowsiness) and only occurred in three patients. The high response rate in patients with therapy refractory pruritus suggests that the NKR1 antagonist aprepitant may indeed exhibit antipruritic effects and may present a novel, effective treatment strategy based on pathophysiology of chronic pruritus. The results are promising enough to warrant confirming the efficacy of NKR1 antagonists in a randomized, controlled clinical trial.

Inhaled corticosteroids inhibit substance P receptor expression in asthmatic rat airway smooth muscle cells

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Li Miao

2012-12-01

Full Text Available Abstract Background Neurokinins (NKs participate in asthmatic airway inflammation, but the effects of NKs on airway smooth muscle cells (ASMCs and those of corticosteroids on NKs are unknown. Methods To investigate the effect of budesonide on substance P (NK-1 receptor (NK-1R expression in the lung and ASMCs, 45 Wistar rats were randomly divided into three groups: control, asthmatic, and budesonide treatment. Aerosolized ovalbumin was used to generate the asthmatic rat model, and budesonide was administered after ovalbumin inhalation. On day 21, bronchial responsiveness tests, bronchoalveolar lavage, and cell counting were conducted. NK-1R protein expression in the lung was investigated by immunohistochemistry and image analysis. Primary rat ASMC cultures were established, and purified ASMCs of the fourth passage were collected for mRNA and protein studies via real-time RT-PCR, immunocytochemistry, and image analysis. Results NK-1R mRNA and protein expression in the budesonide treatment group rat’s lung and ASMCs were less than that in the asthmatic group but greater than that in the control group. Conclusions NK-1R is involved in the pathogenesis of asthma and that budesonide may downregulate the expression of NK-1R in the ASMCs and airways of asthmatic rats, which may alleviate neurogenic airway inflammation.

Mas-related G protein coupled receptor-X2: A potential new target for modulating mast cell-mediated allergic and inflammatory diseases.

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Ali, Hydar

2016-12-01

Mast cells (MCs) are tissue resident immune cells that are best known for their roles in allergic and inflammatory diseases. In addition to the high affinity IgE receptor (FcεRI), MCs express numerous G protein coupled receptors (GPCRs), which are the most common targets of drug therapy. Neurokinin 1 receptor (NK-1R) is expressed on MCs and contributes to IgE and non-IgE-mediated responses in mice. Although NK-1R antagonists are highly effective in modulating experimental allergic and inflammatory responses in mice they lack efficacy in humans. This article reviews recent findings that demonstrate that while neuropeptides (NPs) activate murine MCs via NK-1R and Mas related G protein coupled receptor B2 (MrgprB2), they activate human MCs via Mas-related G protein coupled receptor X2 (MRGPRX2). Interestingly, conventional NK-1R antagonists have off-target activity against mouse MrgprB2 but not human MRGPRX2. These findings suggest that the failure to translate studies with NK-1R antagonists from in vivo mouse studies to the clinic likely reflects their lack of effect on human MRGPRX2. A unique feature of MRGPRX2 that distinguishes it from other GPCRs is that it is activated by a diverse group of ligands that include; neuropeptides, cysteine proteases, antimicrobial peptides and cationic proteins released from activated eosinophils. Thus, the development of small molecule MRGPRX2-specific antagonists or neutralizing antibodies may provide new targets for the treatment of MC-mediated allergic and inflammatory diseases.

The role of NK cells in HIV-1 protection: autologous, allogeneic or both?

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Hens, Jef; Jennes, Wim; Kestens, Luc

2016-01-01

Natural killer (NK) cells specialize in killing virally infected- or tumor cells and are part of the innate immune system. The activational state of NK cells is determined by the balance of incoming activating and inhibitory signals mediated by receptor-ligand binding with the target cell. These receptor-ligand bonds mainly consist of the killer immunoglobulin-like receptors (KIR), which are expressed at the cell surface of NK cells, and their ligands: the highly variable human leukocyte antigen -class I molecules (HLA). Absence of an inhibitory receptor-ligand bond lowers the NK cell activation threshold, whereas an activating receptor-ligand bond stimulates the cell, potentially overcoming this threshold and triggering NK cell activation. NK cells influence the course of infection as well as the acquisition of HIV-1. Several lines of evidence relate the activating NK cell receptor KIR3DS1, in the presence or absence of its putative ligand HLA-Bw4, with slower disease progression as well as resistance to HIV-1 infection. Overall, resistance to HIV-1 infection predominantly correlates with activating KIR/HLA profiles, consisting of e.g. activating KIRs, group B haplotypes, or inhibitory KIRs in absence of their ligands. Such a conclusion is less evident for studies of HIV-1 disease progression, with studies reporting beneficial as well as detrimental effects of activating KIR/HLA genotypes. It is likely that KIR/HLA association studies are complicated by the complexity of the KIR and HLA loci and their mutual interactions, as well as by additional factors like route of HIV exposure, immune activation, presence of co-infections, and the effect of anti-HIV-1 antibodies. One newly discovered NK cell activation pathway associated with resistance to HIV-1 infection involves the presence of an iKIR/HLA mismatch between partners. The absence of such an iKIR/HLA bond renders donor-derived allogeneic HIV-1 infected cells vulnerable to NK cell responses during HIV-1

Targeting the neurokinin receptor 1 with aprepitant: a novel antipruritic strategy.

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Sonja Ständer

2010-06-01

Full Text Available Chronic pruritus is a global clinical problem with a high impact on the quality of life and lack of specific therapies. It is an excruciating and frequent symptom of e.g. uncurable renal, liver and skin diseases which often does not respond to conventional treatment with e.g. antihistamines. Therefore antipruritic therapies which target physiological mechanisms of pruritus need to be developed. Substance P (SP is a major mediator of pruritus. As it binds to the neurokinin receptor 1 (NKR1, we evaluated if the application of a NKR1 antagonist would significantly decrease chronic pruritus.Twenty hitherto untreatable patients with chronic pruritus (12 female, 8 male; mean age, 66.7 years were treated with the NKR1 antagonist aprepitant 80 mg for one week. 16 of 20 patients (80% experienced a considerable reduction of itch intensity, as assessed by the visual analog scale (VAS, range 0 to 10. Considering all patients, the mean value of pruritus intensity was significantly reduced from 8.4 VAS points (SD +/-1.7 before treatment to 4.9 VAS points (SD +/-3.2 (p<0.001, CI 1.913-5.187. Patients with dermatological diseases (e.g. atopic diathesis, prurigo nodularis had the best profit from the treatment. Side-effects were mild (nausea, vertigo, and drowsiness and only occurred in three patients.The high response rate in patients with therapy refractory pruritus suggests that the NKR1 antagonist aprepitant may indeed exhibit antipruritic effects and may present a novel, effective treatment strategy based on pathophysiology of chronic pruritus. The results are promising enough to warrant confirming the efficacy of NKR1 antagonists in a randomized, controlled clinical trial.

Phase III trial of casopitant, a novel neurokinin-1 receptor antagonist, for the prevention of nausea and vomiting in patients receiving moderately emetogenic chemotherapy

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Herrstedt, Jørn; Apornwirat, Wichit; Shaharyar, Ahmed

2009-01-01

PURPOSE: The purpose of this phase III trial was to evaluate the efficacy and safety of regimens containing casopitant, a novel neurokinin-1 receptor antagonist, for the prevention of chemotherapy-induced nausea and vomiting during the first cycle in patients receiving moderately emetogenic chemo...

Antagonist profile of ibodutant at the tachykinin NK2 receptor in guinea pig isolated bronchi.

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Santicioli, Paolo; Meini, Stefania; Giuliani, Sandro; Lecci, Alessandro; Maggi, Carlo Alberto

2013-10-24

In this study we have characterized the pharmacological profile of the non-peptide tachykinin NK 2 receptor antagonist ibodutant (MEN15596) in guinea pig isolated main bronchi contractility. The antagonist potency of ibodutant was evaluated using the selective NK 2 receptor agonist [βAla 8 ]NKA(4-10)-mediated contractions of guinea pig isolated main bronchi. In this assay ibodutant (30, 100 and 300nM) induced a concentration-dependent rightward shift of the [βAla 8 ]NKA(4-10) concentration-response curves without affecting the maximal contractile effect. The analysis of the results yielded a Schild-plot linear regression with a slope not different from unity (0.95, 95% c.l. 0.65-1.25), thus indicating a surmountable behaviour. The calculated apparent antagonist potency as pK B value was 8.31±0.05. Ibodutant (0.3-100nM), produced a concentration-dependent inhibition of the nonadrenergic-noncholinergic (NANC) contractile response induced by electrical field stimulation (EFS) of intrinsic airway nerves in guinea pig isolated main bronchi. At the highest concentration tested (100nM) ibodutant almost abolished the EFS-induced bronchoconstriction (95±4% inhibition), the calculated IC 50 value was 2.98nM (95% c.l. 1.73-5.16nM). In bronchi from ovalbumin (OVA) sensitized guinea pigs ibodutant (100nM) did not affect the maximal contractile response to OVA, but completely prevented the slowing in the fading of the motor response induced by phosphoramidon pretreatment linked to the endogenous neurokinin A release. Altogether, the present study demonstrate that ibodutant is a potent NK 2 receptor antagonist in guinea pig airways. © 2013 Published by Elsevier B.V.

HIV-1 adaptation to NK cell-mediated immune pressure

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Elemans, Marjet; Boelen, Lies; Rasmussen, Michael

2017-01-01

The observation, by Alter et al., of the enrichment of NK cell “escape” variants in individuals carrying certain Killer-cell Immunoglobulin-like Receptor (KIR) genes is compelling evidence that natural killer (NK) cells exert selection pressure on HIV-1. Alter et al hypothesise that variant pepti...

Constitutively polarized granules prime KHYG-1 NK cells.

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Suck, Garnet; Branch, Donald R; Aravena, Paola; Mathieson, Mark; Helke, Simone; Keating, Armand

2006-09-01

The major mechanism for NK cell lysis of tumor cells is granule-mediated cytotoxicity. Polarization of granules is a prelude to the release of their cytotoxic contents in response to target-cell binding. We describe the novel observation of constitutive granule polarization in the cytotoxic NK cell line, KHYG-1. Continuous degranulation of KHYG-1 cells, however, does not occur and still requires target-cell contact. Disruption of microtubules with colcemid is sufficient to disperse the granules in KHYG-1 and significantly decreases cytotoxicity. A similar effect is not obtained by inhibiting extracellular signal-related kinase 2 (ERK2), the most distal kinase investigated in the cytolytic pathway. Disruption of microtubules significantly down-regulates activation receptors, NKp44 and NKG2D, implicating them as potential microtubule-trafficking receptors. Such changes in upstream receptor expression may have caused deactivation of ERK2, since NKG2D cross-linking also leads to receptor down-regulation and diminished ERK phosphorylation. Thus, a functional role for NKG2D in KHYG-1 cytotoxicity is demonstrated. Moreover, the novel primed state may contribute to the high cytotoxicity exhibited by KHYG-1.

Escape of HIV-1-infected dendritic cells from TRAIL-mediated NK cell cytotoxicity during NK-DC cross-talk--a pivotal role of HMGB1.

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Marie-Thérèse Melki

2010-04-01

Full Text Available Early stages of Human Immunodeficiency Virus-1 (HIV-1 infection are associated with local recruitment and activation of important effectors of innate immunity, i.e. natural killer (NK cells and dendritic cells (DCs. Immature DCs (iDCs capture HIV-1 through specific receptors and can disseminate the infection to lymphoid tissues following their migration, which is associated to a maturation process. This process is dependent on NK cells, whose role is to keep in check the quality and the quantity of DCs undergoing maturation. If DC maturation is inappropriate, NK cells will kill them ("editing process" at sites of tissue inflammation, thus optimizing the adaptive immunity. In the context of a viral infection, NK-dependent killing of infected-DCs is a crucial event required for early elimination of infected target cells. Here, we report that NK-mediated editing of iDCs is impaired if DCs are infected with HIV-1. We first addressed the question of the mechanisms involved in iDC editing, and we show that cognate NK-iDC interaction triggers apoptosis via the TNF-related apoptosis-inducing ligand (TRAIL-Death Receptor 4 (DR4 pathway and not via the perforin pathway. Nevertheless, once infected with HIV-1, DC(HIV become resistant to NK-induced TRAIL-mediated apoptosis. This resistance occurs despite normal amounts of TRAIL released by NK cells and comparable DR4 expression on DC(HIV. The escape of DC(HIV from NK killing is due to the upregulation of two anti-apoptotic molecules, the cellular-Flice like inhibitory protein (c-FLIP and the cellular inhibitor of apoptosis 2 (c-IAP2, induced by NK-DC(HIV cognate interaction. High-mobility group box 1 (HMGB1, an alarmin and a key mediator of NK-DC cross-talk, was found to play a pivotal role in NK-dependent upregulation of c-FLIP and c-IAP2 in DC(HIV. Finally, we demonstrate that restoration of DC(HIV susceptibility to NK-induced TRAIL killing can be obtained either by silencing c-FLIP and c-IAP2 by specific

mGluR1 receptors contribute to non-purinergic slow excitatory transmission to submucosal VIP neurons of guinea-pig ileum

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Jaime Pei Pei Foong

2009-05-01

Full Text Available Vasoactive intestinal peptide (VIP immunoreactive secretomotor neurons in the submucous plexus are involved in mediating bacterial toxin-induced hypersecretion leading to diarrhoea. VIP neurons become hyperexcitable after the mucosa is exposed to cholera toxin, which suggests that the manipulation of the excitability of these neurons may be therapeutic. This study used standard intracellular recording methods to systematically characterize slow excitatory postsynaptic potentials (EPSPs evoked in submucosal VIP neurons by different stimulus regimes (1, 3 and 15 pulse 30 Hz stimulation, together with their associated input resistances and pharmacology. All slow EPSPs were associated with a significant increase in input resistance compared to baseline values. Slow EPSPs evoked by a single stimulus were confirmed to be purinergic, however, slow EPSPs evoked by 15 pulse trains were non-purinergic and those evoked by 3 pulse trains were mixed. NK1 or NK3 receptor antagonists did not affect slow EPSPs. The group I mGluR receptor antagonist, PHCCC reduced the amplitude of purinergic and non-purinergic slow EPSPs. Blocking mGluR1 receptors depressed the overall response to 3 and 15 pulse trains, but this effect was inconsistent, while blockade of mGluR5 receptors had no effect on the non-purinergic slow EPSPs. Thus, although other receptors are almost certainly involved, our data indicate that there are at least two pharmacologically distinct types of slow EPSPs in the VIP secretomotor neurons: one mediated by P2Y receptors and the other in part by mGluR1 receptors.

Intrathymic selection of NK1.1+α/β T cell antigen receptor (TCR)+ cells in transgenic mice bearing TCR specific for chicken ovalbumin and restricted to I-Ad

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Iwabuchi, Chikako; Iwabuchi, Kazuya; Nakagawa, Ken-ichi; Takayanagi, Toshiaki; Nishihori, Hiroki; Tone, Saori; Ogasawara, Kazumasa; Good, Robert A.; Onoé, Kazunori

1998-01-01

Generation and negative selection of NK1.1+α/β T cell receptor (TCR)+ thymocytes were analyzed using TCR-transgenic (B10.D2 × DO10)F1 and (C57BL/6 × DO10)F1 mice and Rag-1−/−/DO10 mice, which had been established by breeding and backcrossing between Rag-1−/− and DO10 mice. Almost all T cells from these mice were shown to bear Vα13/Vβ8.2 that is specific for chicken ovalbumin (cOVA) and restricted to I-Ad. A normal proportion of the NK1.1+ Vα13/Vβ8.2+ thymocytes was generated in these mice. However, the actual cell number of both NK1.1+ and NK1.1− thymocytes in I-Ad/d mice (positive selecting background) was larger than that in I-Ab/d mice (negative selecting background). Markedly low but significant proportions of NK1.1+ Vα13/Vβ8.2+ cells were detected in the spleens from I-Ad/d and I-Ab/d mice. It was shown that the splenic NK1.1+ T cells of the I-Ab/d mice were anergized against stimulation through TCR. When (B10.D2 × DO10)F1 and (C57BL/6 × DO10)F1 mice were given cOVA, extensive or intermediate elimination of NK1.1+α/βTCR+ thymocytes was induced in I-Ad/d or I-Ab/d mice, respectively. However, the clonal elimination was not as complete as that seen in the major NK1.1− thymocyte population. The present findings indicate that normal generation of NK1.1+α/βTCR+ thymocytes occurs in the absence of Vα14-Jα281 and that substantial negative selection operates on the NK1.1+α/βTCR+ cells. PMID:9653164

Recruitment of activation receptors at inhibitory NK cell immune synapses.

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Nicolas Schleinitz

2008-09-01

Full Text Available Natural killer (NK cell activation receptors accumulate by an actin-dependent process at cytotoxic immune synapses where they provide synergistic signals that trigger NK cell effector functions. In contrast, NK cell inhibitory receptors, including members of the MHC class I-specific killer cell Ig-like receptor (KIR family, accumulate at inhibitory immune synapses, block actin dynamics, and prevent actin-dependent phosphorylation of activation receptors. Therefore, one would predict inhibition of actin-dependent accumulation of activation receptors when inhibitory receptors are engaged. By confocal imaging of primary human NK cells in contact with target cells expressing physiological ligands of NK cell receptors, we show here that this prediction is incorrect. Target cells included a human cell line and transfected Drosophila insect cells that expressed ligands of NK cell activation receptors in combination with an MHC class I ligand of inhibitory KIR. The two NK cell activation receptors CD2 and 2B4 accumulated and co-localized with KIR at inhibitory immune synapses. In fact, KIR promoted CD2 and 2B4 clustering, as CD2 and 2B4 accumulated more efficiently at inhibitory synapses. In contrast, accumulation of KIR and of activation receptors at inhibitory synapses correlated with reduced density of the integrin LFA-1. These results imply that inhibitory KIR does not prevent CD2 and 2B4 signaling by blocking their accumulation at NK cell immune synapses, but by blocking their ability to signal within inhibitory synapses.

Modulation of respiratory frequency by peptidergic input to rhythmogenic neurons in the preBötzinger complex

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Gray, P A; Rekling, J C; Bocchiaro, C M

1999-01-01

Neurokinin-1 receptor (NK1R) and mu-opioid receptor (muOR) agonists affected respiratory rhythm when injected directly into the preBötzinger Complex (preBötC), the hypothesized site for respiratory rhythmogenesis in mammals. These effects were mediated by actions on preBötC rhythmogenic neurons....... The distribution of NK1R+ neurons anatomically defined the preBötC. Type 1 neurons in the preBötC, which have rhythmogenic properties, expressed both NK1Rs and muORs, whereas type 2 neurons expressed only NK1Rs. These findings suggest that the preBötC is a definable anatomic structure with unique physiological...... function and that a subpopulation of neurons expressing both NK1Rs and muORs generate respiratory rhythm and modulate respiratory frequency....

Neurokinin-3 Receptor Binding in Guinea Pig, Monkey, and Human Brain: In Vitro and in Vivo Imaging Using the Novel Radioligand, [18F]Lu AF10628.

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Varnäs, Katarina; Finnema, Sjoerd J; Stepanov, Vladimir; Takano, Akihiro; Tóth, Miklós; Svedberg, Marie; Møller Nielsen, Søren; Khanzhin, Nikolay A; Juhl, Karsten; Bang-Andersen, Benny; Halldin, Christer; Farde, Lars

2016-08-01

Previous autoradiography studies have suggested a marked interspecies variation in the neuroanatomical localization and expression levels of the neurokinin 3 receptor, with high density in the brain of rat, gerbil, and guinea pig, but at the time offered no conclusive evidence for its presence in the human brain. Hitherto available radioligands have displayed low affinity for the human neurokinin 3 receptor relative to the rodent homologue and may thus not be optimal for cross-species analyses of the expression of this protein. A novel neurokinin 3 receptor radioligand, [(18)F]Lu AF10628 ((S)-N-(cyclobutyl(3-fluorophenyl)methyl)-8-fluoro-2-((3-[(18)F]-fluoropropyl)amino)-3-methyl-1-oxo-1,2-dihydroisoquinoline-4-carboxamide), was synthesized and used for autoradiography studies in cryosections from guinea pig, monkey, and human brain as well as for positron emission tomography studies in guinea pig and monkey. The results confirmed previous observations of interspecies variation in the neurokinin 3 receptor brain localization with more extensive distribution in guinea pig than in primate brain. In the human brain, specific binding to the neurokinin 3 receptor was highest in the amygdala and in the hypothalamus and very low in other regions examined. Positron emission tomography imaging showed a pattern consistent with that observed using autoradiography. The radioactivity was, however, found to accumulate in skull bone, which limits the use of this radioligand for in vivo quantification of neurokinin 3 receptor binding. Species differences in the brain distribution of neurokinin 3 receptors should be considered when using animal models for predicting human neurokinin 3 receptor pharmacology. For positron emission tomography imaging of brain neurokinin 3 receptors, additional work is required to develop a radioligand with more favorable in vivo properties. © The Author 2016. Published by Oxford University Press on behalf of CINP.

DX5+NKT cells display phenotypical and functional differences between spleen and liver as well as NK1.1-Balb/c and NK1.1+ C57Bl/6 mice.

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Werner, Jens M; Busl, Elisabeth; Farkas, Stefan A; Schlitt, Hans J; Geissler, Edward K; Hornung, Matthias

2011-04-29

Natural killer T cells represent a linkage between innate and adaptive immunity. They are a heterogeneous population of specialized T lymphocytes composed of different subsets. DX5+NKT cells are characterized by expression of the NK cell marker DX5 in the context of CD3. However, little is known about the phenotype and functional capacity of this unique cell population. Therefore, we investigated the expression of several T cell and NK cell markers, as well as functional parameters in spleen and liver subsets of DX5+NKT cells in NK1.1- Balb/c mice and compared our findings to NK1.1+ C57Bl/6 mice. In the spleen 34% of DX5+NKT cells expressed CD62L and they up-regulated the functional receptors CD154 as well as CD178 upon activation. In contrast, only a few liver DX5+NKT cells expressed CD62L, and they did not up-regulate CD154 upon activation. A further difference between spleen and liver subsets was observed in cytokine production. Spleen DX5+NKT cells produced more Th1 cytokines including IL-2, IFN-γ and TNF-α, while liver DX5+NKT cells secreted more Th2 cytokines (e.g. IL-4) and even the Th17 cytokine, IL-17a. Furthermore, we found inter-strain differences. In NK1.1+ C57Bl/6 mice DX5+NKT cells represented a distinct T cell population expressing less CD4 and more CD8. Accordingly, these cells showed a CD178 and Th2-type functional capacity upon activation. These results show that DX5+NKT cells are a heterogeneous population, depending on the dedicated organ and mouse strain, that has diverse functional capacity.

Requirement for Innate Immunity and CD90+ NK1.1− Lymphocytes to Treat Established Melanoma with Chemo-Immunotherapy

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Moskalenko, Marina; Pan, Michael; Fu, Yichun; de Moll, Ellen H.; Hashimoto, Daigo; Mortha, Arthur; Leboeuf, Marylene; Jayaraman, Padmini; Bernardo, Sebastian; Sikora, Andrew G.; Wolchok, Jedd; Bhardwaj, Nina; Merad, Miriam; Saenger, Yvonne

2015-01-01

We sought to define cellular immune mechanisms of synergy between tumor-antigen–targeted monoclonal antibodies and chemotherapy. Established B16 melanoma in mice was treated with cytotoxic doses of cyclophosphamide in combination with an antibody targeting tyrosinase-related protein 1 (αTRP1), a native melanoma differentiation antigen. We find that Fcγ receptors are required for efficacy, showing that antitumor activity of combination therapy is immune mediated. Rag1−/− mice deficient in adaptive immunity are able to clear tumors, and thus innate immunity is sufficient for efficacy. Furthermore, previously treated wild-type mice are not significantly protected against tumor reinduction, as compared with mice inoculated with irradiated B16 alone, consistent with a primarily innate immune mechanism of action of chemo-immunotherapy. In contrast, mice deficient in both classical natural killer (NK) lymphocytes and nonclassical innate lymphocytes (ILC) due to deletion of the IL2 receptor common gamma chain IL2γc−/−) are refractory to chemo-immunotherapy. Classical NK lymphocytes are not critical for treatment, as depletion of NK1.1+ cells does not impair antitumor effect. Depletion of CD90+NK1.1− lymphocytes, however, both diminishes therapeutic benefit and decreases accumulation of macrophages within the tumor. Tumor clearance during combination chemo-immunotherapy with monoclonal antibodies against native antigen is mediated by the innate immune system. We highlight a novel potential role for CD90+NK1.1− ILCs in chemo-immunotherapy. PMID:25600438

Role of Kisspeptin and Neurokinin B in Puberty in Female Non-Human Primates

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Ei Terasawa

2018-04-01

Full Text Available In human patients, loss-of-function mutations in the genes encoding kisspeptin (KISS1 and neurokinin B (NKB and their receptors (KISS1R and NK3R, respectively result in an abnormal timing of puberty or the absence of puberty. To understand the neuroendocrine mechanism of puberty, we investigated the contribution of kisspeptin and NKB signaling to the pubertal increase in GnRH release using rhesus monkeys as a model. Direct measurements of GnRH and kisspeptin in the median eminence of the hypothalamus with infusion of agonists and antagonists for kisspeptin and NKB reveal that kisspeptin and NKB signaling stimulate GnRH release independently or collaboratively by forming kisspeptin and NKB neuronal networks depending on the developmental age. For example, while in prepubertal females, kisspeptin and NKB signaling independently stimulate GnRH release, in pubertal females, the formation of a collaborative kisspeptin and NKB network further accelerates the pubertal increase in GnRH release. It is speculated that the collaborative mechanism between kisspeptin and NKB signaling to GnRH neurons is necessary for the complex reproductive function in females.

Lymphocytes Negatively Regulate NK Cell Activity via Qa-1b following Viral Infection

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Haifeng C. Xu

2017-11-01

Full Text Available NK cells can reduce anti-viral T cell immunity during chronic viral infections, including infection with the lymphocytic choriomeningitis virus (LCMV. However, regulating factors that maintain the equilibrium between productive T cell and NK cell immunity are poorly understood. Here, we show that a large viral load resulted in inhibition of NK cell activation, which correlated with increased expression of Qa-1b, a ligand for inhibitory NK cell receptors. Qa-1b was predominantly upregulated on B cells following LCMV infection, and this upregulation was dependent on type I interferons. Absence of Qa-1b resulted in increased NK cell-mediated regulation of anti-viral T cells following viral infection. Consequently, anti-viral T cell immunity was reduced in Qa-1b- and NKG2A-deficient mice, resulting in increased viral replication and immunopathology. NK cell depletion restored anti-viral immunity and virus control in the absence of Qa-1b. Taken together, our findings indicate that lymphocytes limit NK cell activity during viral infection in order to promote anti-viral T cell immunity.

Fine tuning a well-oiled machine: Influence of NK1.1 and NKG2D on NKT cell development and function.

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Joshi, Sunil K; Lang, Mark L

2013-10-01

Natural killer T cells (NKT) represent a group of CD1d-restricted T-lineage cells that provide a functional interface between innate and adaptive immune responses in infectious disease, cancer, allergy and autoimmunity. There have been remarkable advances in understanding the molecular events that underpin NKT development in the thymus and in the complex array of functions in the periphery. Most functional studies have focused on activation of T cell antigen receptors expressed by NKT cells and their responses to CD1d presentation of glycolipid and related antigens. Receiving less attention has been several molecules that are hallmarks of Natural Killer (NK) cells, but nonetheless expressed by NKT cells. These include several activating and inhibitory receptors that may fine-tune NKT development and survival, as well as activation via antigen receptors. Herein, we review the possible roles of the NK1.1 and NKG2D receptors in regulating development and function of NKT cells in health and disease. We suggest that pharmacological alteration of NKT activity should consider the potential complexities commensurate with NK1.1 and NKG2D expression. Published by Elsevier B.V.

Endometrial natural killer (NK) cells reveal a tissue-specific receptor repertoire.

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Feyaerts, D; Kuret, T; van Cranenbroek, B; van der Zeeuw-Hingrez, S; van der Heijden, O W H; van der Meer, A; Joosten, I; van der Molen, R G

2018-02-13

Is the natural killer (NK) cell receptor repertoire of endometrial NK (eNK) cells tissue-specific? The NK cell receptor (NKR) expression profile in pre-pregnancy endometrium appears to have a unique tissue-specific phenotype, different from that found in NK cells in peripheral blood, suggesting that these cells are finely tuned towards the reception of an allogeneic fetus. NK cells are important for successful pregnancy. After implantation, NK cells encounter extravillous trophoblast cells and regulate trophoblast invasion. NK cell activity is amongst others regulated by C-type lectin heterodimer (CD94/NKG2) and killer cell immunoglobulin-like (KIR) receptors. KIR expression on decidual NK cells is affected by the presence of maternal HLA-C and biased towards KIR2D expression. However, little is known about NKR expression on eNK cells prior to pregnancy. In this study, matched peripheral and menstrual blood (a source of endometrial cells) was obtained from 25 healthy females with regular menstrual cycles. Menstrual blood was collected during the first 36 h of menstruation using a menstrual cup, a non-invasive technique to obtain endometrial cells. KIR and NKG2 receptor expression on eNK cells was characterized by 10-color flow cytometry, and compared to matched pbNK cells of the same female. KIR and HLA-C genotypes were determined by PCR-SSOP techniques. Anti-CMV IgG antibodies in plasma were measured by chemiluminescence immunoassay. KIR expression patterns of eNK cells collected from the same female do not differ over consecutive menstrual cycles. The percentage of NK cells expressing KIR2DL2/L3/S2, KIR2DL3, KIR2DL1, LILRB1 and/or NKG2A was significantly higher in eNK cells compared to pbNK cells, while no significant difference was observed for NKG2C, KIR2DL1/S1, and KIR3DL1. The NKR repertoire of eNK cells was clearly different from pbNK cells, with eNK cells co-expressing more than three NKR simultaneously. In addition, outlier analysis revealed 8 and 15 NKR

Intrathymic selection of NK1.1+α/β T cell antigen receptor (TCR)+ cells in transgenic mice bearing TCR specific for chicken ovalbumin and restricted to I-Ad

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Iwabuchi, Chikako; Iwabuchi, Kazuya; Nakagawa, Ken-ichi; Takayanagi, Toshiaki; Nishihori, Hiroki; Tone, Saori; Ogasawara, Kazumasa; Good, Robert A.; Onoé, Kazunori

1998-01-01

Generation and negative selection of NK1.1+α/β T cell receptor (TCR)+ thymocytes were analyzed using TCR-transgenic (B10.D2 × DO10)F1 and (C57BL/6 × DO10)F1 mice and Rag-1−/−/DO10 mice, which had been established by breeding and backcrossing between Rag-1−/− and DO10 mice. Almost all T cells from these mice were shown to bear Vα13/Vβ8.2 that is specific for chicken ovalbumin (cOVA) and restricted to I-Ad. A normal proportion of the NK1.1+ Vα13/Vβ8.2+ thymocytes was generated in these mice. Ho...

Monosodium Urate Crystals Induce Upregulation of NK1.1-Dependent Killing by Macrophages and Support Tumor-Resident NK1.1+ Monocyte/Macrophage Populations in Antitumor Therapy.

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Steiger, Stefanie; Kuhn, Sabine; Ronchese, Franca; Harper, Jacquie L

2015-12-01

Macrophages display phenotypic and functional heterogeneity dependent on the changing inflammatory microenvironment. Under some conditions, macrophages can acquire effector functions commonly associated with NK cells. In the current study, we investigated how the endogenous danger signal monosodium urate (MSU) crystals can alter macrophage functions. We report that naive, primary peritoneal macrophages rapidly upregulate the expression of the NK cell-surface marker NK1.1 in response to MSU crystals but not in response to LPS or other urate crystals. NK1.1 upregulation by macrophages was associated with mechanisms including phagocytosis of crystals, NLRP3 inflammasome activation, and autocrine proinflammatory cytokine signaling. Further analysis demonstrated that MSU crystal-activated macrophages exhibited NK cell-like cytotoxic activity against target cells in a perforin/granzyme B-dependent manner. Furthermore, analysis of tumor hemopoietic cell populations showed that effective, MSU-mediated antitumor activity required coadministration with Mycobacterium smegmatis to induce IL-1β production and significant accumulation of monocytes and macrophages (but not granulocytes or dendritic cells) expressing elevated levels of NK1.1. Our findings provide evidence that MSU crystal-activated macrophages have the potential to develop tumoricidal NK cell-like functions that may be exploited to boost antitumor activity in vivo. Copyright © 2015 by The American Association of Immunologists, Inc.

Evolution of the C-Type Lectin-Like Receptor Genes of the DECTIN-1 Cluster in the NK Gene Complex

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Susanne Sattler

2012-01-01

Full Text Available Pattern recognition receptors are crucial in initiating and shaping innate and adaptive immune responses and often belong to families of structurally and evolutionarily related proteins. The human C-type lectin-like receptors encoded in the DECTIN-1 cluster within the NK gene complex contain prominent receptors with pattern recognition function, such as DECTIN-1 and LOX-1. All members of this cluster share significant homology and are considered to have arisen from subsequent gene duplications. Recent developments in sequencing and the availability of comprehensive sequence data comprising many species showed that the receptors of the DECTIN-1 cluster are not only homologous to each other but also highly conserved between species. Even in Caenorhabditis elegans, genes displaying homology to the mammalian C-type lectin-like receptors have been detected. In this paper, we conduct a comprehensive phylogenetic survey and give an up-to-date overview of the currently available data on the evolutionary emergence of the DECTIN-1 cluster genes.

Células NK: generalidades y papel durante la infección por el virus de la inmunodeficiencia humana tipo 1 (VIH-1 NK cells: characteristics and role during the infection by type-1 human immunodeficiency virus (HIV-1

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María Teresa Rugeles López

2007-03-01

Full Text Available Las células NK exhiben actividad espontánea contra células tumorales o células infectadas, particularmente por virus. Ellas se caracterizan por la expresión de las moléculas CD16 y CD56, y se subdividen en dos poblaciones, CD16Low/CD56Hi y CD16Hi/CD56Low, que difieren en las citoquinas que producen y en la capacidad citotóxica. La activación de las células NK está regulada por la expresión de receptores inhibidores y activadores que interactúan con diferentes ligandos de las células blanco. La actividad efectora de estas células incluye la lisis de las células blanco por diferentes mecanismos y la producción de citoquinas; las células NK participan por medio de estos factores solubles en diversos procesos fisiológicos, como la hematopoyesis y la regulación de otras células del sistema inmune. Durante la infección por el VIH-1, las células NK ayudan al control de la replicación viral tanto por mecanismos citotóxicos como por la producción de citoquinas, particularmente -quimoquinas. Sin embargo, el VIH- 1 ha desarrollado mecanismos para evadir la respuesta antiviral mediada por las células NK. Adicionalmente, esta infección induce anormalidades cuantitativas y funcionales en estas células que pueden presentarse muy temprano en la evolución de la enfermedad y que hacen parte de la inmunosupresión severa característica del SIDA. NK cells exhibit spontaneous activity against tumor and infected cells, particularly with virus. They are characterized by the expression of the CD16 and CD56 molecules. Two NK cell subpopulations have been described: CD16Low/CD56Hi and CD16Hi/ CD56Low that differ in the cytokines produced and their cytotoxic ability. NK cell activation is regulated by the expression of inhibition and activation receptors, which interact with different ligands on the target cells. The effector activity of these cells includes lysis of target cells by different mechanisms and the production of cytokines

Mouse Clr-g, a ligand for NK cell activation receptor NKR-P1F: crystal structure and biophysical properties

Czech Academy of Sciences Publication Activity Database

Skálová, Tereza; Kotýnková, Kristýna; Dušková, Jarmila; Hašek, Jindřich; Koval, Tomáš; Kolenko, Petr; Novák, P.; Man, Petr; Hanč, Pavel; Vaněk, Ondřej; Bezouška, K.; Dohnálek, Jan

2012-01-01

Roč. 189, č. 10 (2012), s. 4881-4889 ISSN 0022-1767 R&D Projects: GA AV ČR KJB101120805; GA ČR GAP302/11/0855; GA ČR GD305/09/H008; GA ČR GA305/07/1073; GA ČR(CZ) GAP207/10/1040; GA MŠk EE2.3.30.0029; GA ČR GA303/09/0477 EU Projects: European Commission(XE) 31220 - SPINE2-COMPLEXES Institutional research plan: CEZ:AV0Z40500505; CEZ:AV0Z50200510 Institutional support: RVO:61389013 ; RVO:61388971 Keywords : Clr-g * C-type lectin like fold * NK receptor Subject RIV: CE - Biochemistry; EC - Immunology (MBU-M) Impact factor: 5.520, year: 2012

Distinct human and mouse membrane trafficking systems for sweet taste receptors T1r2 and T1r3.

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Shimizu, Madoka; Goto, Masao; Kawai, Takayuki; Yamashita, Atsuko; Kusakabe, Yuko

2014-01-01

The sweet taste receptors T1r2 and T1r3 are included in the T1r taste receptor family that belongs to class C of the G protein-coupled receptors. Heterodimerization of T1r2 and T1r3 is required for the perception of sweet substances, but little is known about the mechanisms underlying this heterodimerization, including membrane trafficking. We developed tagged mouse T1r2 and T1r3, and human T1R2 and T1R3 and evaluated membrane trafficking in human embryonic kidney 293 (HEK293) cells. We found that human T1R3 surface expression was only observed when human T1R3 was coexpressed with human T1R2, whereas mouse T1r3 was expressed without mouse T1r2 expression. A domain-swapped chimera and truncated human T1R3 mutant showed that the Venus flytrap module and cysteine-rich domain (CRD) of human T1R3 contain a region related to the inhibition of human T1R3 membrane trafficking and coordinated regulation of human T1R3 membrane trafficking. We also found that the Venus flytrap module of both human T1R2 and T1R3 are needed for membrane trafficking, suggesting that the coexpression of human T1R2 and T1R3 is required for this event. These results suggest that the Venus flytrap module and CRD receive taste substances and play roles in membrane trafficking of human T1R2 and T1R3. These features are different from those of mouse receptors, indicating that human T1R2 and T1R3 are likely to have a novel membrane trafficking system.

Downregulation of miR-15a due to LMP1 promotes cell proliferation and predicts poor prognosis in nasal NK/T-cell lymphoma.

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Komabayashi, Yuki; Kishibe, Kan; Nagato, Toshihiro; Ueda, Seigo; Takahara, Miki; Harabuchi, Yasuaki

2014-01-01

Nasal NK/T-cell lymphoma (NNKTL) is an Epstein-Barr virus (EBV)-associated malignancy and has distinct clinical and histological features. However, its genetic features are hitherto unclear. MicroRNAs (miRNAs) play a crucial role in the pathogenesis of several malignancies via regulating gene expression. In this study, we investigated whether the specific microRNAs were related to the tumor behaviors in NNKTL. MiRNA array and Quantitative RT-PCR analyses revealed that miR-15a was expressed at a much lower level in NNKTL cells (SNK-1, SNK-6, and SNT-8) than in normal peripheral NK cells and EBV-negative NK cell line KHYG-1. Quantitative PCR and western blot analyses showed that the expression of MYB and cyclin D1, which are validated targets of miR-15a, was higher in NNKTL cells. Transfection of NNKTL cells (SNK-6 and SNT-8) with a miR-15a precursor decreased MYB and cyclin D1 levels, thereby blocking G1/S transition and cell proliferation. Knockdown of EBV-encoded latent membrane protein 1 (LMP1) significantly increased miR-15a expression in SNK-6 cells. In NNKTL tissues, we found that reduced miR-15a expression, which correlated with MYB and cyclin D1 expression, was associated with poor prognosis of NNKTL patients. These data suggest that downregulation of miR-15a, possibly due to LMP1, implicates in the pathogenesis of NNKTL by inducing cell proliferation via MYB and cyclin D1. Thus, miR-15a could be a potential target for antitumor therapy and a prognostic predictor for NNKTL. Copyright © 2013 Wiley Periodicals, Inc.

Homoeologous Recombination of the V1r1-V1r2 Gene Cluster of Pheromone Receptors in an Allotetraploid Lineage of Teleosts

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Lei Zhong

2017-11-01

Full Text Available In contrast to other olfactory receptor families that exhibit frequent lineage-specific expansions, the vomeronasal type 1 receptor (V1R family exhibits a canonical six-member repertoire in teleosts. V1r1 and V1r2 are present in no more than one copy in all examined teleosts, including salmons, which are ancient polyploids, implying strict evolutionary constraints. However, recent polyploids have not been examined. Here, we identified a young allotetraploid lineage of weatherfishes and investigated their V1r1-V1r2 cluster. We found a novel pattern that the parental V1r1-V1r2 clusters had recombined in the tetraploid genome and that the recombinant was nearly fixed in the tetraploid population. Subsequent analyses suggested strong selective pressure, for both a new combination of paralogs and homogeneity among gene duplicates, acting on the V1r1-V1r2 pair.

Cell-Autonomous Regulation of Mu-Opioid Receptor Recycling by Substance P

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Bowman, Shanna L.; Soohoo, Amanda L.; Shiwarski, Daniel J.; Schulz, Stefan; Pradhan, Amynah A.; Puthenveedu, Manojkumar A.

2015-01-01

SUMMARY How neurons coordinate and reprogram multiple neurotransmitter signals is an area of broad interest. Here, we show that substance P (SP), a neuropep-tide associated with inflammatory pain, reprograms opioid receptor recycling and signaling. SP, through activation of the neurokinin 1 (NK1R) receptor, increases the post-endocytic recycling of the muopioid receptor (MOR) in trigeminal ganglion (TG) neurons in an agonist-selective manner. SP-mediated protein kinase C (PKC) activation is both required and sufficient for increasing recycling of exogenous and endogenous MOR in TG neurons. The target of this cross-regulation is MOR itself, given that mutation of either of two PKC phosphorylation sites on MOR abolishes the SP-induced increase in recycling and resensitization. Furthermore, SP enhances the resensitization of fentanyl-induced, but not morphine-induced, antinociception in mice. Our results define a physiological pathway that cross-regulates opioid receptor recycling via direct modification of MOR and suggest a mode of homeo-static interaction between the pain and analgesic systems. PMID:25801029

Kinetic properties of 'dual' orexin receptor antagonists at OX1R and OX2R orexin receptors.

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Gabrielle Elizabeth Callander

2013-12-01

Full Text Available Orexin receptor antagonists represent attractive targets for the development of drugs for the treatment of insomnia. Both efficacy and safety are crucial in clinical settings and thorough investigations of pharmacokinetics and pharmacodynamics can predict contributing factors such as duration of action and undesirable effects. To this end, we studied the interactions between various ‘dual’ orexin receptor antagonists and the orexin receptors, OX1R and OX2R, over time using saturation and competition radioligand binding with [3H]-BBAC ((S-N-([1,1'-biphenyl]-2-yl-1-(2-((1-methyl-1H-benzo[d]imidazol-2-ylthioacetylpyrrolidine-2-carboxamide. In addition, the kinetics of these compounds were investigated in cells expressing human, mouse and rat OX1R and OX2R using FLIPR® assays for calcium accumulation. We demonstrate that almorexant reaches equilibrium very slowly at OX2R, whereas SB-649868, suvorexant and filorexant may take hours to reach steady state at both orexin receptors. By contrast, compounds such as BBAC or the selective OX2R antagonist IPSU ((2-((1H-Indol-3-ylmethyl-9-(4-methoxypyrimidin-2-yl-2,9-diazaspiro[5.5]undecan-1-one bind rapidly and reach equilibrium very quickly in both binding and / or functional assays. Overall, the dual antagonists tested here tend to be rather unselective under non-equilibrium conditions and reach equilibrium very slowly. Once equilibrium is reached, each ligand demonstrates a selectivity profile that is however, distinct from the non-equilibrium condition. The slow kinetics of the dual antagonists tested suggest that in vitro receptor occupancy may be longer lasting than would be predicted. This raises questions as to whether pharmacokinetic studies measuring plasma or brain levels of these antagonists are accurate reflections of receptor occupancy in vivo.

Compensatory insulin receptor (IR) activation on inhibition of insulin-like growth factor-1 receptor (IGF-1R): rationale for cotargeting IGF-1R and IR in cancer.

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Buck, Elizabeth; Gokhale, Prafulla C; Koujak, Susan; Brown, Eric; Eyzaguirre, Alexandra; Tao, Nianjun; Rosenfeld-Franklin, Maryland; Lerner, Lorena; Chiu, M Isabel; Wild, Robert; Epstein, David; Pachter, Jonathan A; Miglarese, Mark R

2010-10-01

Insulin-like growth factor-1 receptor (IGF-1R) is a receptor tyrosine kinase (RTK) and critical activator of the phosphatidylinositol 3-kinase-AKT pathway. IGF-1R is required for oncogenic transformation and tumorigenesis. These observations have spurred anticancer drug discovery and development efforts for both biological and small-molecule IGF-1R inhibitors. The ability for one RTK to compensate for another to maintain tumor cell viability is emerging as a common resistance mechanism to antitumor agents targeting individual RTKs. As IGF-1R is structurally and functionally related to the insulin receptor (IR), we asked whether IR is tumorigenic and whether IR-AKT signaling contributes to resistance to IGF-1R inhibition. Both IGF-1R and IR(A) are tumorigenic in a mouse mammary tumor model. In human tumor cells coexpressing IGF-1R and IR, bidirectional cross talk was observed following either knockdown of IR expression or treatment with a selective anti-IGF-1R antibody, MAB391. MAB391 treatment resulted in a compensatory increase in phospho-IR, which was associated with resistance to inhibition of IRS1 and AKT. In contrast, treatment with OSI-906, a small-molecule dual inhibitor of IGF-1R/IR, resulted in enhanced reduction in phospho-IRS1/phospho-AKT relative to MAB391. Insulin or IGF-2 activated the IR-AKT pathway and decreased sensitivity to MAB391 but not to OSI-906. In tumor cells with an autocrine IGF-2 loop, both OSI-906 and an anti-IGF-2 antibody reduced phospho-IR/phospho-AKT, whereas MAB391 was ineffective. Finally, OSI-906 showed superior efficacy compared with MAB391 in human tumor xenograft models in which both IGF-1R and IR were phosphorylated. Collectively, these data indicate that cotargeting IGF-1R and IR may provide superior antitumor efficacy compared with targeting IGF-1R alone.

The cognition-enhancing activity of E1R, a novel positive allosteric modulator of sigma-1 receptors

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Zvejniece, L; Vavers, E; Svalbe, B; Vilskersts, R; Domracheva, I; Vorona, M; Veinberg, G; Misane, I; Stonans, I; Kalvinsh, I; Dambrova, M

2014-01-01

Background and Purpose Here, we describe the in vitro and in vivo effects of (4R,5S)-2-(5-methyl-2-oxo-4-phenyl-pyrrolidin-1-yl)-acetamide (E1R), a novel positive allosteric modulator of sigma-1 receptors. Experimental Approach E1R was tested for sigma receptor binding activity in a [3H](+)-pentazocine assay, in bradykinin (BK)-induced intracellular Ca2+ concentration ([Ca2+]i) assays and in an electrically stimulated rat vas deferens model. E1R's effects on cognitive function were tested using passive avoidance (PA) and Y-maze tests in mice. A selective sigma-1 receptor antagonist (NE-100), was used to study the involvement of the sigma-1 receptor in the effects of E1R. The open-field test was used to detect the effects of E1R on locomotion. Key Results Pretreatment with E1R enhanced the selective sigma-1 receptor agonist PRE-084's stimulating effect during a model study employing electrically stimulated rat vasa deferentia and an assay measuring the BK-induced [Ca2+]i increase. Pretreatment with E1R facilitated PA retention in a dose-related manner. Furthermore, E1R alleviated the scopolamine-induced cognitive impairment during the PA and Y-maze tests in mice. The in vivo and in vitro effects of E1R were blocked by treatment with the selective sigma-1 receptor antagonist NE-100. E1R did not affect locomotor activity. Conclusion and Implications E1R is a novel 4,5-disubstituted derivative of piracetam that enhances cognition and demonstrates efficacy against scopolamine-induced cholinergic dysfunction in mice. These effects are attributed to its positive modulatory action on the sigma-1 receptor and this activity may be relevant when developing new drugs for treating cognitive symptoms related to neurodegenerative diseases. PMID:24490863

IL-15 inhibits pre-B cell proliferation by selectively expanding Mac-1+B220+ NK cells

International Nuclear Information System (INIS)

Nakajima, Shinsuke; Hida, Shigeaki; Taki, Shinsuke

2008-01-01

Natural killer (NK) cells are the cells critical for inhibition of repopulation of allogenic bone marrow cells. However, it is not well known if NK cells affect autologous lymphopoiesis. Here, we observed that NK cells could inhibit pre-B cell proliferation in vitro driven by interleukin (IL)-7 in a manner dependent on IL-15. Interestingly, the great majority of expanding NK cells were Mac-1 + B220 + , a recently identified potent interferon (IFN)-γ producer. Indeed, IFN-γ was produced in those cultures, and pre-B cells lacking IFN-γ receptors, but not those lacking type I IFN receptors, were resistant to such an inhibition. Furthermore, even NK cells from mice lacking β2-microglobulin, which were known to be functionally dampened, inhibited pre-B cell proliferation as well. Thus, activated NK cells, which were expanded selectively by IL-15, could potentially regulate B lymphopoiesis through IFN-γ beyond the selection imposed upon self-recognition

Antiallodynic Effects of Bee Venom in an Animal Model of Complex Regional Pain Syndrome Type 1 (CRPS-I).

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Lee, Sung Hyun; Lee, Jae Min; Kim, Yun Hong; Choi, Jung Hyun; Jeon, Seung Hwan; Kim, Dong Kyu; Jeong, Hyeon Do; Lee, You Jung; Park, Hue Jung

2017-09-15

Neuropathic pain in a chronic post-ischaemic pain (CPIP) model mimics the symptoms of complex regional pain syndrome type I (CRPS I). The administration of bee venom (BV) has been utilized in Eastern medicine to treat chronic inflammatory diseases accompanying pain. However, the analgesic effect of BV in a CPIP model remains unknown. The application of a tight-fitting O-ring around the left ankle for a period of 3 h generated CPIP in C57/Bl6 male adult mice. BV (1 mg/kg ; 1, 2, and 3 times) was administered into the SC layer of the hind paw, and the antiallodynic effects were investigated using the von Frey test and by measuring the expression of neurokinin type 1 (NK-1) receptors in dorsal root ganglia (DRG). The administration of BV dose-dependently reduced the pain withdrawal threshold to mechanical stimuli compared with the pre-administration value and with that of the control group. After the development of the CPIP model, the expression of NK-1 receptors in DRG increased and then decreased following the administration of BV. SC administration of BV results in the attenuation of allodynia in a mouse model of CPIP. The antiallodynic effect was objectively proven through a reduction in the increased expression of NK-1 receptors in DRG.

Acute inflammation induces segmental, bilateral, supraspinally mediated opioid release in the rat spinal cord, as measured by μ-opioid receptor internalization

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Chen, Wenling; Marvizón, Juan Carlos G.

2009-01-01

The objective of this study was to measure opioid release in the spinal cord during acute and long-term inflammation using μ-opioid receptor (MOR) internalization. In particular, we determined whether opioid release occurs in the segments receiving the noxious signals or in the entire spinal cord, and whether it involves supraspinal signals. Internalization of neurokinin 1 receptors (NK1Rs) was measured to track the intensity of the noxious stimulus. Rats received peptidase inhibitors intrathecally to protect opioids from degradation. Acute inflammation of the hindpaw with formalin induced moderate MOR internalization in the L5 segment bilaterally, whereas NK1R internalization occurred only ipsilaterally. MOR internalization was restricted to the lumbar spinal cord, regardless of whether the peptidase inhibitors were injected in a lumbar or thoracic site. Formalin-induced MOR internalization was substantially reduced by isoflurane anesthesia. It was also markedly reduced by a lidocaine block of the cervical-thoracic spinal cord (which did not affect the evoked NK1R internalization) indicating that spinal opioid release is mediated supraspinally. In the absence of peptidase inhibitors, formalin and hindpaw clamp induced a small amount of MOR internalization, which was significantly higher than in controls. To study spinal opioid release during chronic inflammation, we injected Complete Freund's Adjuvant (CFA) in the hindpaw and peptidase inhibitors intrathecally. Two days later, no MOR or NK1R internalization was detected. Furthermore, CFA inflammation decreased MOR internalization induced by clamping the inflamed hindpaw. These results show that acute inflammation, but not chronic inflammation, induce segmental opioid release in the spinal cord that involves supraspinal signals. PMID:19298846

Acute inflammation induces segmental, bilateral, supraspinally mediated opioid release in the rat spinal cord, as measured by mu-opioid receptor internalization.

Science.gov (United States)

Chen, W; Marvizón, J C G

2009-06-16

The objective of this study was to measure opioid release in the spinal cord during acute and long-term inflammation using mu-opioid receptor (MOR) internalization. In particular, we determined whether opioid release occurs in the segments receiving the noxious signals or in the entire spinal cord, and whether it involves supraspinal signals. Internalization of neurokinin 1 receptors (NK1Rs) was measured to track the intensity of the noxious stimulus. Rats received peptidase inhibitors intrathecally to protect opioids from degradation. Acute inflammation of the hind paw with formalin induced moderate MOR internalization in the L5 segment bilaterally, whereas NK1R internalization occurred only ipsilaterally. MOR internalization was restricted to the lumbar spinal cord, regardless of whether the peptidase inhibitors were injected in a lumbar or thoracic site. Formalin-induced MOR internalization was substantially reduced by isoflurane anesthesia. It was also markedly reduced by a lidocaine block of the cervical-thoracic spinal cord (which did not affect the evoked NK1R internalization) indicating that spinal opioid release is mediated supraspinally. In the absence of peptidase inhibitors, formalin and hind paw clamp induced a small amount of MOR internalization, which was significantly higher than in controls. To study spinal opioid release during chronic inflammation, we injected complete Freund's adjuvant (CFA) in the hind paw and peptidase inhibitors intrathecally. Two days later, no MOR or NK1R internalization was detected. Furthermore, CFA inflammation decreased MOR internalization induced by clamping the inflamed hind paw. These results show that acute inflammation, but not chronic inflammation, induces segmental opioid release in the spinal cord that involves supraspinal signals.

The NK1R-/- mouse phenotype suggests that small body size, with a sex- and diet-dependent excess in body mass and fat, are physical biomarkers for a human endophenotype with vulnerability to attention deficit hyperactivity disorder.

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Pillidge, Katharine; Heal, David J; Stanford, S Clare

2016-09-01

The abnormal behaviour of NK1R-/- mice (locomotor hyperactivity, inattentiveness and impulsivity in the 5-Choice Serial Reaction-Time Test) is arguably analogous to that of patients with attention deficit hyperactivity disorder (ADHD). Evidence suggests that small body size and increased body weight are risk factors for ADHD. Here, we compared the body size, body mass and body composition of male and female NK1R-/- mice and their wildtypes that had been fed either standard laboratory chow or a high-fat (45%: 'Western') diet. Male NK1R-/- mice from both cohorts were approximately 7% shorter than wildtypes. A similar trend was evident in females. Male NK1R-/- mice fed the normal diet weighed less than wildtypes but the 'body mass index' ('mBMI': weight (mg)/length (cm)(2)) of female NK1R-/- mice was higher than wildtypes. When given the high-fat diet, the mBMI of both male and female NK1R-/- mice was higher than wildtypes. There were no consistent genotype or sex differences in protein, ash or water content of mice from the two cohorts. However, the fat content of male NK1R-/- mice on the Western diet was considerably (35%) higher than wildtypes and resembled that of females from both genotypes. We conclude that a lack of functional NK1R is associated with small body size but increases vulnerability to an increase in mBMI and fat content, especially in males. This phenotype could also be evident in ADHD patients with polymorphism(s) of the TACR1 gene (the human equivalent of Nk1r). © The Author(s) 2016.

Dopamine receptors modulate cytotoxicity of natural killer cells via cAMP-PKA-CREB signaling pathway.

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Wei Zhao

Full Text Available Dopamine (DA, a neurotransmitter in the nervous system, has been shown to modulate immune function. We have previously reported that five subtypes of DA receptors, including D1R, D2R, D3R, D4R and D5R, are expressed in T lymphocytes and they are involved in regulation of T cells. However, roles of these DA receptor subtypes and their coupled signal-transduction pathway in modulation of natural killer (NK cells still remain to be clarified. The spleen of mice was harvested and NK cells were isolated and purified by negative selection using magnetic activated cell sorting. After NK cells were incubated with various drugs for 4 h, flow cytometry measured cytotoxicity of NK cells against YAC-1 lymphoma cells. NK cells expressed the five subtypes of DA receptors at mRNA and protein levels. Activation of D1-like receptors (including D1R and D5R with agonist SKF38393 enhanced NK cell cytotoxicity, but activation of D2-like receptors (including D2R, D3R and D4R with agonist quinpirole attenuated NK cells. Simultaneously, SKF38393 elevated D1R and D5R expression, cAMP content, and phosphorylated cAMP-response element-binding (CREB level in NK cells, while quinpirole reduced D3R and D4R expression, cAMP content, and phosphorylated CREB level in NK cells. These effects of SKF38393 were blocked by SCH23390, an antagonist of D1-like receptors, and quinpirole effects were abolished by haloperidol, an antagonist of D2-like receptors. In support these results, H89, an inhibitor of phosphokinase A (PKA, prevented the SKF38393-dependent enhancement of NK cells and forskolin, an activator of adenylyl cyclase (AC, counteracted the quinpirole-dependent suppression of NK cells. These findings show that DA receptor subtypes are involved in modulation of NK cells and suggest that D1-like receptors facilitate NK cells by stimulating D1R/D5R-cAMP-PKA-CREB signaling pathway and D2-like receptors suppress NK cells by inhibiting D3R/D4R-cAMP-PKA-CREB signaling pathway. The

The role of NK1.1+ cells in the protection against MHC class I+ HPV16-associated tumours

Czech Academy of Sciences Publication Activity Database

Šímová, Jana; Bubeník, Jan; Bieblová, Jana; Jandlová, Táňa

2004-01-01

Roč. 50, č. 6 (2004), s. 200-202 ISSN 0015-5500 R&D Projects: GA AV ČR IAA5052203; GA AV ČR KSK5011112; GA MZd NC7148; GA MZd NR7807 Institutional research plan: CEZ:AV0Z5052915 Keywords : HPV16 * MHC class I expression * NK1.1+ cells Subject RIV: EC - Immunology Impact factor: 0.507, year: 2004

Recent Advances of Colony-Stimulating Factor-1 Receptor (CSF-1R) Kinase and Its Inhibitors.

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El-Gamal, Mohammed I; Al-Ameen, Shahad K; Al-Koumi, Dania M; Hamad, Mawadda G; Jalal, Nouran A; Oh, Chang-Hyun

2018-01-17

Colony stimulation factor-1 receptor (CSF-1R), which is also known as FMS kinase, plays an important role in initiating inflammatory, cancer, and bone disorders when it is overstimulated by its ligand, CSF-1. Innate immunity, as well as macrophage differentiation and survival, are regulated by the stimulation of the CSF-1R. Another ligand, interlukin-34 (IL-34), was recently reported to activate the CSF-1R receptor in a different manner. The relationship between CSF-1R and microglia has been reviewed. Both CSF-1 antibodies and small molecule CSF-1R kinase inhibitors have now been tested in animal models and in humans. In this Perspective, we discuss the role of CSF-1 and IL-34 in producing cancer, bone disorders, and inflammation. We also review the newly discovered and improved small molecule kinase inhibitors and monoclonal antibodies that have shown potent activity toward CSF-1R, reported from 2012 until 2017.

ADCC employing an NK cell line (haNK) expressing the high affinity CD16 allele with avelumab, an anti-PD-L1 antibody.

Science.gov (United States)

Jochems, Caroline; Hodge, James W; Fantini, Massimo; Tsang, Kwong Y; Vandeveer, Amanda J; Gulley, James L; Schlom, Jeffrey

2017-08-01

NK-92 cells, and their derivative, designated aNK, were obtained from a patient with non-Hodgkin lymphoma. Prior clinical studies employing adoptively transferred irradiated aNK cells have provided evidence of clinical benefit and an acceptable safety profile. aNK cells have now been engineered to express IL-2 and the high affinity (ha) CD16 allele (designated haNK). Avelumab is a human IgG1 anti-PD-L1 monoclonal antibody, which has shown evidence of clinical activity in a range of human tumors. Prior in vitro studies have shown that avelumab has the ability to mediate antibody-dependent cell-mediated cytotoxicity (ADCC) of human tumor cells when combined with NK cells. In the studies reported here, the ability of avelumab to enhance the lysis of a range of human carcinoma cells by irradiated haNK cells via the ADCC mechanism is demonstrated; this ADCC is shown to be inhibited by anti-CD16 blocking antibody and by concanamycin A, indicating the use of the granzyme/perforin pathway in tumor cell lysis. Studies also show that while NK cells have the ability to lyse aNK or haNK cells, the addition of NK cells to irradiated haNK cells does not inhibit haNK-mediated lysis of human tumor cells, with or without the addition of avelumab. Avelumab-mediated lysis of tumor cells by irradiated haNK cells is also shown to be similar to that of NK cells bearing the V/V Fc receptor high affinity allele. These studies thus provide the rationale for the clinical evaluation of the combined use of avelumab with that of irradiated adoptively transferred haNK cells. © 2017 UICC.

NK1.1+ cells promote sustained tissue injury and inflammation after trauma with hemorrhagic shock.

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Chen, Shuhua; Hoffman, Rosemary A; Scott, Melanie; Manson, Joanna; Loughran, Patricia; Ramadan, Mostafa; Demetris, Anthony J; Billiar, Timothy R

2017-07-01

Various cell populations expressing NK1.1 contribute to innate host defense and systemic inflammatory responses, but their role in hemorrhagic shock and trauma remains uncertain. NK1.1 + cells were depleted by i.p. administration of anti-NK1.1 (or isotype control) on two consecutive days, followed by hemorrhagic shock with resuscitation and peripheral tissue trauma (HS/T). The plasma levels of IL-6, MCP-1, alanine transaminase (ALT), and aspartate aminotransferase (AST) were measured at 6 and 24 h. Histology in liver and gut were examined at 6 and 24 h. The number of NK cells, NKT cells, neutrophils, and macrophages in liver, as well as intracellular staining for TNF-α, IFN-γ, and MCP-1 in liver cell populations were determined by flow cytometry. Control mice subjected to HS/T exhibited end organ damage manifested by marked increases in circulating ALT, AST, and MCP-1 levels, as well as histologic evidence of hepatic necrosis and gut injury. Although NK1.1 + cell-depleted mice exhibited a similar degree of organ damage as nondepleted animals at 6 h, NK1.1 + cell depletion resulted in marked suppression of both liver and gut injury by 24 h after HS/T. These findings indicate that NK1.1 + cells contribute to the persistence of inflammation leading to end organ damage in the liver and gut. © Society for Leukocyte Biology.

Tachykinins stimulate a subset of mouse taste cells.

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Jeff Grant

Full Text Available The tachykinins substance P (SP and neurokinin A (NKA are present in nociceptive sensory fibers expressing transient receptor potential cation channel, subfamily V, member 1 (TRPV1. These fibers are found extensively in and around the taste buds of several species. Tachykinins are released from nociceptive fibers by irritants such as capsaicin, the active compound found in chili peppers commonly associated with the sensation of spiciness. Using real-time Ca(2+-imaging on isolated taste cells, it was observed that SP induces Ca(2+ -responses in a subset of taste cells at concentrations in the low nanomolar range. These responses were reversibly inhibited by blocking the SP receptor NK-1R. NKA also induced Ca(2+-responses in a subset of taste cells, but only at concentrations in the high nanomolar range. These responses were only partially inhibited by blocking the NKA receptor NK-2R, and were also inhibited by blocking NK-1R indicating that NKA is only active in taste cells at concentrations that activate both receptors. In addition, it was determined that tachykinin signaling in taste cells requires Ca(2+-release from endoplasmic reticulum stores. RT-PCR analysis further confirmed that mouse taste buds express NK-1R and NK-2R. Using Ca(2+-imaging and single cell RT-PCR, it was determined that the majority of tachykinin-responsive taste cells were Type I (Glial-like and umami-responsive Type II (Receptor cells. Importantly, stimulating NK-1R had an additive effect on Ca(2+ responses evoked by umami stimuli in Type II (Receptor cells. This data indicates that tachykinin release from nociceptive sensory fibers in and around taste buds may enhance umami and other taste modalities, providing a possible mechanism for the increased palatability of spicy foods.

TRPV1 channels and the progesterone receptor Sig-1R interact to regulate pain.

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Ortíz-Rentería, Miguel; Juárez-Contreras, Rebeca; González-Ramírez, Ricardo; Islas, León D; Sierra-Ramírez, Félix; Llorente, Itzel; Simon, Sidney A; Hiriart, Marcia; Rosenbaum, Tamara; Morales-Lázaro, Sara L

2018-02-13

The Transient Receptor Potential Vanilloid 1 (TRPV1) ion channel is expressed in nociceptors where, when activated by chemical or thermal stimuli, it functions as an important transducer of painful and itch-related stimuli. Although the interaction of TRPV1 with proteins that regulate its function has been previously explored, their modulation by chaperones has not been elucidated, as is the case for other mammalian TRP channels. Here we show that TRPV1 physically interacts with the Sigma 1 Receptor (Sig-1R), a chaperone that binds progesterone, an antagonist of Sig-1R and an important neurosteroid associated to the modulation of pain. Antagonism of Sig-1R by progesterone results in the down-regulation of TRPV1 expression in the plasma membrane of sensory neurons and, consequently, a decrease in capsaicin-induced nociceptive responses. This is observed both in males treated with a synthetic antagonist of Sig-1R and in pregnant females where progesterone levels are elevated. This constitutes a previously undescribed mechanism by which TRPV1-dependent nociception and pain can be regulated.

HMGB1-dependent triggering of HIV-1 replication and persistence in dendritic cells as a consequence of NK-DC cross-talk.

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Héla Saïdi

Full Text Available HIV-1 has evolved ways to exploit DCs, thereby facilitating viral dissemination and allowing evasion of antiviral immunity. Recently, the fate of DCs has been found to be extremely dependent on the interaction with autologous NK cells, but the mechanisms by which NK-DC interaction controls viral infections remain unclear. Here, we investigate the impact of NK-DC cross-talk on maturation and functions of HIV-infected immature DCs.Immature DCs were derived from primary monocytes, cultured in the presence of IL-4 and GM-CSF. In some experiments, DCs were infected with R5-HIV-1(BaL or X4-HIV-1(NDK, and viral replication, proviral HIV-DNA and the frequency of infected DCs were measured. Autologous NK cells were sorted and either kept unstimulated in the presence of suboptimal concentration of IL-2, or activated by a combination of PHA and IL-2. The impact of 24 h NK-DC cross-talk on the fate of HIV-1-infected DCs was analyzed. We report that activated NK cells were required for the induction of maturation of DCs, whether uninfected or HIV-1-infected, and this process involved HMGB1. However, the cross-talk between HIV-1-infected DCs and activated NK cells was functionally defective, as demonstrated by the strong impairment of DCs to induce Th1 polarization of naïve CD4 T cells. This was associated with the defective production of IL-12 and IL-18 by infected DCs. Moreover, the crosstalk between activated NK cells and HIV-infected DCs resulted in a dramatic increase in viral replication and proviral DNA expression in DCs. HMGB1, produced both by NK cells and DCs, was found to play a pivotal role in this process, and inhibition of HMGB1 activity by glycyrrhizin, known to bind specifically to HMGB1, or blocking anti-HMGB1 antibodies, abrogated NK-dependent HIV-1 replication in DCs.These observations provide evidence for the crucial role of NK-DC cross-talk in promoting viral dissemination, and challenge the question of the in vivo involvement of HMGB1

Tachykinins in the porcine pancreas

DEFF Research Database (Denmark)

Schmidt, P T; Tornøe, K; Poulsen, Steen Seier

2000-01-01

The localization, release, and effects of substance P and neurokinin A were studied in the porcine pancreas and the localization of substance P immunoreactive nerve fibers was examined by immunohistochemistry. The effects of electrical vagus stimulation and capsaicin infusion on tachykinin release...... and the effects of substance P and neurokinin A infusion on insulin, glucagon, somatostatin, and exocrine secretion were studied using the isolated perfused porcine pancreas with intact vagal innervation. NK-1 and NK-2 receptor antagonists were used to investigate receptor involvement. Substance P immunoreactive...

Angiotensin II reduces cardiac AdipoR1 expression through AT1 receptor/ROS/ERK1/2/c-Myc pathway.

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Li Li

Full Text Available Adiponectin, an abundant adipose tissue-derived protein, exerts protective effect against cardiovascular disease. Adiponectin receptors (AdipoR1 and AdipoR2 mediate the beneficial effects of adiponectin on the cardiovascular system. However, the alteration of AdipoRs in cardiac remodeling is not fully elucidated. Here, we investigated the effect of angiotensin II (AngII on cardiac AdipoRs expression and explored the possible molecular mechanism. AngII infusion into rats induced cardiac hypertrophy, reduced AdipoR1 but not AdipoR2 expression, and attenuated the phosphorylations of adenosine monophosphate-activated protein kinase and acetyl coenzyme A carboxylase, and those effects were all reversed by losartan, an AngII type 1 (AT1 receptor blocker. AngII reduced expression of AdipoR1 mRNA and protein in cultured neonatal rat cardiomyocytes, which was abolished by losartan, but not by PD123319, an AT2 receptor antagonist. The antioxidants including reactive oxygen species (ROS scavenger NAC, NADPH oxidase inhibitor apocynin, Nox2 inhibitor peptide gp91 ds-tat, and mitochondrial electron transport chain complex I inhibitor rotenone attenuated AngII-induced production of ROS and phosphorylation of extracellular signal-regulated kinase (ERK 1/2. AngII-reduced AdipoR1 expression was reversed by pretreatment with NAC, apocynin, gp91 ds-tat, rotenone, and an ERK1/2 inhibitor PD98059. Chromatin immunoprecipitation assay demonstrated that AngII provoked the recruitment of c-Myc onto the promoter region of AdipoR1, which was attenuated by PD98059. Moreover, AngII-induced DNA binding activity of c-Myc was inhibited by losartan, NAC, apocynin, gp91 ds-tat, rotenone, and PD98059. c-Myc small interfering RNA abolished the inhibitory effect of AngII on AdipoR1 expression. Our results suggest that AngII inhibits cardiac AdipoR1 expression in vivo and in vitro and AT1 receptor/ROS/ERK1/2/c-Myc pathway is required for the downregulation of AdipoR1 induced by AngII.

Substance p regulates puberty onset and fertility in the female mouse.

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Simavli, Serap; Thompson, Iain R; Maguire, Caroline A; Gill, John C; Carroll, Rona S; Wolfe, Andrew; Kaiser, Ursula B; Navarro, Víctor M

2015-06-01

Puberty is a tightly regulated process that leads to reproductive capacity. Kiss1 neurons are crucial in this process by stimulating GnRH, yet how Kiss1 neurons are regulated remains unknown. Substance P (SP), an important neuropeptide in pain perception, induces gonadotropin release in adult mice in a kisspeptin-dependent manner. Here, we assessed whether SP, through binding to its receptor NK1R (neurokinin 1 receptor), participates in the timing of puberty onset and fertility in the mouse. We observed that 1) selective NK1R agonists induce gonadotropin release in prepubertal females; 2) the expression of Tac1 (encoding SP) and Tacr1 (NK1R) in the arcuate nucleus is maximal before puberty, suggesting increased SP tone; 3) repeated exposure to NK1R agonists prepubertally advances puberty onset; and 4) female Tac1(-/-) mice display delayed puberty; moreover, 5) SP deficiency leads to subfertility in females, showing fewer corpora lutea and antral follicles and leading to decreased litter size. Thus, our findings support a role for SP in the stimulation of gonadotropins before puberty, acting via Kiss1 neurons to stimulate GnRH release, and its involvement in the attainment of full reproductive capabilities in female mice.

Cell-Autonomous Regulation of Mu-Opioid Receptor Recycling by Substance P

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Shanna L. Bowman

2015-03-01

Full Text Available How neurons coordinate and reprogram multiple neurotransmitter signals is an area of broad interest. Here, we show that substance P (SP, a neuropeptide associated with inflammatory pain, reprograms opioid receptor recycling and signaling. SP, through activation of the neurokinin 1 (NK1R receptor, increases the post-endocytic recycling of the mu-opioid receptor (MOR in trigeminal ganglion (TG neurons in an agonist-selective manner. SP-mediated protein kinase C (PKC activation is both required and sufficient for increasing recycling of exogenous and endogenous MOR in TG neurons. The target of this cross-regulation is MOR itself, given that mutation of either of two PKC phosphorylation sites on MOR abolishes the SP-induced increase in recycling and resensitization. Furthermore, SP enhances the resensitization of fentanyl-induced, but not morphine-induced, antinociception in mice. Our results define a physiological pathway that cross-regulates opioid receptor recycling via direct modification of MOR and suggest a mode of homeostatic interaction between the pain and analgesic systems.

NETUPITANT, A POTENT AND HIGHLY SELECTIVE NK1 RECEPTOR ANTAGONIST, ALLEVIATES ACETIC ACID-INDUCED BLADDER OVERACTIVITY IN ANESTHETIZED GUINEA-PIGS

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Stefano Palea

2016-08-01

Full Text Available Introduction. Tachykinins potently contract the isolated urinary bladder from a number of animal species and play an important role in the regulation of the micturition reflex. On the guinea-pig isolated urinary bladder we examined the effects of a new potent and selective NK1 receptor antagonist (netupitant on the contractions induced by a selective NK1 receptor agonist, SP-methylester (SP-OMe. Moreover, the effects of netupitant and another selective NK1 antagonist (L-733,060 were studied in anesthetized guinea-pigs using two experimental models, the isovolumetric bladder contractions and a model of bladder overactivity induced by intravesical administration of acetic acid (AA. Methods and Results. Detrusor muscle strips were mounted in 5 mL organ baths and isometric contractions to cumulative concentrations of SP-OME were recorded before and after incubation with increasing concentrations of netupitant. In anesthetized female guinea-pigs, reflex bladder activity was examined under isovolumetric conditions with the bladder distended with saline or during cystometry using intravesical infusion of acetic acid (AA. After a 30 min stabilization period, netupitant (0.1-3 mg/kg, i.v. or L-733,060 (3-10 mg/kg, i.v. were administered. In the detrusor muscle, netupitant produced a concentration-dependent inhibition (mean pKB = 9.24 of the responses to SP-OMe. Under isovolumetric conditions, netupitant or L-733,060 reduced bladder contraction frequency in a dose-dependent manner, but neither drug changed bladder contraction amplitude. In the AA model, netupitant dose-dependently increased intercontraction interval (ICI but had no effect on the amplitude of micturition (AM. L-733,060 dose-dependently increased ICI also but this effect was paralleled by a significant reduction of AM. Conclusion. Netupitant decreases the frequency of reflex bladder contractions without altering their amplitude, suggesting that this drug targets the afferent limb of the

Regulation of NKG2D-Dependent NK Cell Functions: The Yin and the Yang of Receptor Endocytosis

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Rosa Molfetta

2017-08-01

Full Text Available Natural-killer receptor group 2, member D (NKG2D is a well characterized natural killer (NK cell activating receptor that recognizes several ligands poorly expressed on healthy cells but up-regulated upon stressing stimuli in the context of cancer or viral infection. Although NKG2D ligands represent danger signals that render target cells more susceptible to NK cell lysis, accumulating evidence demonstrates that persistent exposure to ligand-expressing cells causes the decrease of NKG2D surface expression leading to a functional impairment of NKG2D-dependent NK cell functions. Upon ligand binding, NKG2D is internalized from the plasma membrane and sorted to lysosomes for degradation. However, receptor endocytosis is not only a mechanism of receptor clearance from the cell surface, but is also required for the proper activation of signalling events leading to the functional program of NK cells. This review is aimed at providing a summary of current literature relevant to the molecular mechanisms leading to NKG2D down-modulation with particular emphasis given to the role of NKG2D endocytosis in both receptor degradation and signal propagation. Examples of chronic ligand-induced down-regulation of NK cell activating receptors other than NKG2D, including natural cytotoxicity receptors (NCRs, DNAX accessory molecule-1 (DNAM1 and CD16, will be also discussed.

HMB-45, S-100, NK1/C3, and MART-1 in metastatic melanoma.

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Zubovits, Judit; Buzney, Elizabeth; Yu, Lawrence; Duncan, Lyn M

2004-02-01

The diagnosis of melanoma metastatic to lymph node remains a difficult problem given its histological diversity. We examined the staining patterns of S-100, NK1/C3, HMB-45, and MART-1 (DC10) in melanoma metastases to lymph nodes. Immunohistochemical stains were performed on tissue sections of 126 formalin-fixed lymph nodes from 126 patients with an established diagnosis of metastatic melanoma. A total of 98% of cases (123 of 126) stained positive for S-100, 93% (117 of 125) stained positive for NK1/C3, 82% (103 of 126) stained positive for MART-1, and 76% (95 of 125) stained positive for HMB-45. The distribution and intensity of staining varied among these markers. A diffuse staining pattern, defined as >50% of tumor cells stained, was observed in 83% of MART-1-positive cases but in only 56% of S-100-positive cases, 48% of NK1/C3-positive cases, and 34% of HMB-45-positive cases. A maximally intense signal was almost always observed for MART-1 (83% of positive cases) but was rarely observed for NK1/C3 (20%). S-100 and HMB-45 showed maximally intense staining in 50% and 54% of cases, respectively. S-100 and NK1/C3 stained both histiocytes and melanocytes, whereas MART-1 and HMB-45 stained only melanocytes. Seventy-eight cases (63%) stained positive for all 4 markers, 17 cases (14%) stained for all markers except HMB-45, 13 cases (10%) stained for all markers except MART-1, 6 cases (5%) stained only with S-100 and NK1/C3, 4 cases (3%) stained only with S-100 and HMB-45, and 2 cases stained for all markers except S-100. One case each stained for the following: only S-100, only S-100 and HMB-45, and all markers except NK1/C3. One case exhibited absence of staining for any of these markers. We demonstrate that lymph node metastases of melanoma are heterogeneous with regard to tumor marker expression. S-100 and NK1/C3 were the most sensitive stains for detecting metastatic melanoma; however, they both also stain other nontumor cells in lymph nodes. MART-1 did not stain

Changes in glutamate receptor subunits within the medulla in goats after section of the carotid sinus nerves

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Miller, Justin Robert; Neumueller, Suzanne; Muere, Clarissa; Olesiak, Samantha; Pan, Lawrence; Bukowy, John D.; Daghistany, Asem O.; Hodges, Matthew R.

2014-01-01

The mechanisms which contribute to the time-dependent recovery of resting ventilation and the ventilatory CO2 chemoreflex after carotid body denervation (CBD) are poorly understood. Herein we tested the hypothesis that there are time-dependent changes in the expression of specific AMPA, NMDA, and/or neurokinin-1 (NK1R) receptors within respiratory-related brain stem nuclei acutely or chronically after CBD in adult goats. Brain stem tissues were collected acutely (5 days) or chronically (30 days) after sham or bilateral CBD, immunostained with antibodies targeting AMPA (GluA1 or GluA2), NMDA (GluN1), or NK-1 receptors, and optical density (OD) compared. Physiological measurement confirmed categorization of each group and showed ventilatory effects consistent with bilateral CBD (Miller et al. J Appl Physiol 115: 1088–1098, 2013). Acutely after CBD, GluA1 OD was unchanged or slightly increased, but GluA2 and GluN1 OD were reduced 15–30% within the nucleus tractus solitarius (NTS) and in other medullary respiratory nuclei. Chronically after CBD, GluA1 was reduced (P < 0.05) within the caudal NTS and in other nuclei, but there was significant recovery of GluA2 and GluN1 OD. NK1 OD was not significantly different from control after CBD. We conclude that the initial decrease in GluA2 and GluN1 after CBD likely contributes to hypoventilation and the reduced CO2 chemoreflex. The partial recovery of ventilation and the CO2 chemoreflex after CBD parallel a time-dependent return of these receptors to near control levels but likely depend upon additional initiating and maintenance factors for neuroplasticity. PMID:24790015

Metalloelastase in lungs and alveolar macrophages is modulated by extracellular substance P in mice.

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Xu, J; Xu, F; Barrett, E

2008-07-01

Metalloelastase (MMP-12), mainly produced by macrophages, has been shown to play a key role in the pathogenesis of emphysema in animal models. Chronic cigarette smoke increases pulmonary MMP-12, which is closely correlated with an elevation of pulmonary substance P (SP). Because alveolar macrophages (AMs) contain the neurokinin-1 receptor (NK1R), we tested whether SP was able to trigger the upregulation of MMP-12 synthesis in AMs by acting on the NK1R. AMs isolated from bronchoalveolar lavage cells in C3H/HeN mice were cultured with control medium or SP that was coupled without or with NK1R antagonists (CP-99,994 or aprepitant) for 24 h. We found that SP significantly increased the mRNA of MMP-12 and NK1R by 11-fold and 82%, respectively, in AMs (PNCAP) to degenerate PCFs, respectively. Our results show that NCAP treatment significantly decreased mRNA and protein levels of SP associated with a reduction NK1R and MMP-12 in the lungs and AMs. These findings suggest that SP has a modulatory effect on pulmonary MMP-12 by acting on NK1R to trigger MMP-12 syntheses in the AMs.

NK1.1+ cells are important for the development of protective immunity against MHC I-deficient, HPV16-associated tumours

Czech Academy of Sciences Publication Activity Database

Indrová, Marie; Šímová, Jana; Bieblová, Jana; Bubeník, Jan; Reiniš, Milan

2011-01-01

Roč. 25, č. 1 (2011), s. 281-288 ISSN 1021-335X R&D Projects: GA AV ČR IAA500520807 Grant - others:EU-FP6-NOE(XE) Project 018933 Institutional research plan: CEZ:AV0Z50520514 Keywords : MHC class I-deficient tumours * CD8+, CD4+, NK1.1+cell subpopulations * interferon gamma Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.835, year: 2011

Simultaneous detection of decidual Th1/Th2 and NK1/NK2 immunophenotyping in unknown recurrent miscarriage using 8-color flow cytometry with FSC/Vt extended strategy.

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Dong, Peng; Wen, Xi; Liu, Jia; Yan, Cui-Yan; Yuan, Jing; Luo, Lan-Rong; Hu, Qiao-Fei; Li, Jian

2017-06-30

Th1/Th2 imbalance is considered as a mechanism for recurrent miscarriage. The NK1/NK2 paradigm is hypothesised to play an important role in pregnancy. However, few results showed simultaneous changes of these subsets in vivo in decidual tissues. The present study aimed to detect the decidual mononuclear cells (dMo), and the Th1/Th2, and NK1/NK2 paradigm simultaneously using multiparametric flow cytometry (MFC) in unexplained recurrent miscarriages (URM). Mononuclear cells were isolated from the decidual tissues of URM cases and early pregnant women. The mononuclear cell percent was demonstrated by detecting the expression of CD3, CD4, CD8, CD56, and CD16 extracellular markers, interferon (IFN)-γ, and interleukin (IL)-4 intracellular markers in live cells using 8-color flow cytometry with forward scatter (FSC)/side scatter (SSC) and FSC/viability (Vt) initial gating strategies, and the ratios of Th1/Th2 and decidual NK1 (dNK1)/decidual NK2 (dNK2) cells were compared between the subject groups. Two initial gating strategies of the FSC/SSC or FSC/Vt, with central or extended gating scales, were adapted, and there was no main effect or interaction for the cell proportions, except for the type 1 and type 2 subsets in the FSC/Vt extended gating strategy. There was no significant difference of the proportions of the decidual T, dNK, NKT-like, Th, and Tc cells between the two groups. However, the Th1/Th2 and dNK1/dNK2 ratios in the URM patients were higher compared with the normal group when using the FSC/Vt extended gating strategy. The present study provides means to detect Th1/Th2 and dNK1/dNK2 simultaneously in URM patients for large sample investigations in the future. © 2017 The Author(s).

Partial agonism through a zinc-Ion switch constructed between transmembrane domains III and VII in the tachykinin NK(1) receptor

DEFF Research Database (Denmark)

Holst, B; Elling, C E; Schwartz, T W

2000-01-01

switch located exactly one helical turn below the two previously identified interaction points for Substance P in, respectively, TM-III and -VII. The metal-ion chelator, phenantroline, which in the beta(2)-adrenergic receptor increased both the potency and the agonistic efficacy of Zn(2+) or Cu(2......Partly due to lack of detailed knowledge of the molecular recognition of ligands the structural basis for partial versus full agonism is not known. In the beta(2)-adrenergic receptor the agonist binding site has previously been structurally and functionally exchanged with an activating metal....... In contrast to the similarly mutated beta(2)-adrenergic receptor, signal transduction-i.e., inositol phosphate turnover-could be stimulated by both Zn(2+) and by the natural agonist, Substance P in the mutated NK(1) receptor. The metal-ion acted as a 25% partial agonist through binding to the bidentate zinc...

Defensive and pathological functions of the gastrointestinal NK3 receptor.

Science.gov (United States)

Sanger, Gareth J; Tuladhar, Bishwa R; Bueno, Lionel; Furness, John B

2006-10-01

In general, normal gut functions are unaffected by selective NK(3) receptor antagonists such as talnetant (SB-223412), osanetant (SR 142901) or SB-235375. However, NK(3) receptors may mediate certain defensive or pathological intestinal processes. The precise mechanisms, by which this role is achieved, are not fully understood. In summary, intense stimulation of the intrinsic primary afferent neurones (IPANs) of the enteric nervous system is thought to release tachykinins from these neurones, to induce slow excitation (slow EPSPs) of connecting IPANs. This is hypothesised to cause hypersensitivity and disrupt intestinal motility, at least partly explaining why NK(3) receptor antagonism can reduce the level of disruption caused by supramaximal distension pressures in vitro. Tachykinin release from IPANs may also increase C-fibre sensitivity, directly or indirectly. Thus, NK(3) receptor antagonists can inhibit nociception associated with intestinal distension, in normal animals or after pre-sensitisation by restraint stress. Importantly, such inhibition has been found with SB-235375, a peripherally restricted antagonist. SB-235375 can also reduce a visceromotor response to brief colorectal distension without affecting similar responses to skin pinch, providing additional evidence for intestinal-specific activity. NK(3) receptor biology is, therefore, revealing a novel pathway by which disruptions in intestinal motility and nociception can be induced.

Crystal Structure of Human Interferon-[lamda]1 in Complex with Its High-Affinity Receptor Interferon-[lamda]R1

Energy Technology Data Exchange (ETDEWEB)

Miknis, Zachary; Magracheva, Eugenia; Li, Wei; Zdanov, Alexander; Kotenko, Sergei V.; Wlodawer, Alexander (NJMS); (NCI)

2010-12-01

Interferon (IFN)-{lambda}1 [also known as interleukin (IL)-29] belongs to the recently discovered group of type III IFNs. All type III IFNs initiate signaling processes through formation of specific heterodimeric receptor complexes consisting of IFN-{lambda}R1 and IL-10R2. We have determined the structure of human IFN-{lambda}1 complexed with human IFN-{lambda}R1, a receptor unique to type III IFNs. The overall structure of IFN-{lambda}1 is topologically similar to the structure of IL-10 and other members of the IL-10 family of cytokines. IFN-{lambda}R1 consists of two distinct domains having fibronectin type III topology. The ligand-receptor interface includes helix A, loop AB, and helix F on the IFN site, as well as loops primarily from the N-terminal domain and inter-domain hinge region of IFN-{lambda}R1. Composition and architecture of the interface that includes only a few direct hydrogen bonds support an idea that long-range ionic interactions between ligand and receptor govern the process of initial recognition of the molecules while hydrophobic interactions finalize it.

A Novel System of Polymorphic and Diverse NK Cell Receptors in Primates

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Rosner, Cornelia; Neff, Jennifer; Roos, Christian; Eberle, Manfred; Aujard, Fabienne; Münch, Claudia; Schempp, Werner; Carrington, Mary; Shiina, Takashi; Inoko, Hidetoshi; Knaust, Florian; Coggill, Penny; Sehra, Harminder; Beck, Stephan; Abi-Rached, Laurent; Reinhardt, Richard; Walter, Lutz

2009-01-01

There are two main classes of natural killer (NK) cell receptors in mammals, the killer cell immunoglobulin-like receptors (KIR) and the structurally unrelated killer cell lectin-like receptors (KLR). While KIR represent the most diverse group of NK receptors in all primates studied to date, including humans, apes, and Old and New World monkeys, KLR represent the functional equivalent in rodents. Here, we report a first digression from this rule in lemurs, where the KLR (CD94/NKG2) rather than KIR constitute the most diverse group of NK cell receptors. We demonstrate that natural selection contributed to such diversification in lemurs and particularly targeted KLR residues interacting with the peptide presented by MHC class I ligands. We further show that lemurs lack a strict ortholog or functional equivalent of MHC-E, the ligands of non-polymorphic KLR in “higher” primates. Our data support the existence of a hitherto unknown system of polymorphic and diverse NK cell receptors in primates and of combinatorial diversity as a novel mechanism to increase NK cell receptor repertoire. PMID:19834558

HMGB1 Is Involved in IFN-α Production and TRAIL Expression by HIV-1-Exposed Plasmacytoid Dendritic Cells: Impact of the Crosstalk with NK Cells.

Directory of Open Access Journals (Sweden)

Héla Saïdi

2016-02-01

Full Text Available Plasmacytoid dendritic cells (pDCs are innate sensors of viral infections and important mediators of antiviral innate immunity through their ability to produce large amounts of IFN-α. Moreover, Toll-like receptor 7 (TLR7 and 9 (TLR9 ligands, such as HIV and CpG respectively, turn pDCs into TRAIL-expressing killer pDCs able to lyse HIV-infected CD4+ T cells. NK cells can regulate antiviral immunity by modulating pDC functions, and pDC production of IFN-α as well as cell-cell contact is required to promote NK cell functions. Impaired pDC-NK cell crosstalk was reported in the setting of HIV-1 infection, but the impact of HIV-1 on TRAIL expression and innate antiviral immunity during this crosstalk is unknown. Here, we report that low concentrations of CCR5-tropic HIV-1Ba-L promote the release of pro-inflammatory cytokines such as IFN-α, TNF-α, IFN-γ and IL-12, and CCR5-interacting chemokines (MIP-1α and MIP-1β in NK-pDCs co-cultures. At high HIV-1BaL concentrations, the addition of NK cells did not promote the release of these mediators, suggesting that once efficiently triggered by the virus, pDCs could not integrate new activating signals delivered by NK cells. However, high HIV-1BaL concentrations were required to trigger IFN-α-mediated TRAIL expression at the surface of both pDCs and NK cells during their crosstalk. Interestingly, we identified the alarmin HMGB1, released at pDC-NK cell synapse, as an essential trigger for the secretion of IFN-α and IFN-related soluble mediators during the interplay of HIV-1 exposed pDCs with NK cells. Moreover, HMGB1 was found crucial for mTRAIL translocation to the plasma membrane of both pDCs and NK cells during their crosstalk following pDC exposure to HIV-1. Data from serum analyses of circulating HMGB1, HMGB1-specific antibodies, sTRAIL and IP-10 in a cohort of 67 HIV-1+ patients argue for the in vivo relevance of these observations. Altogether, these findings identify HMGB1 as a trigger for IFN

HMGB1 Is Involved in IFN-α Production and TRAIL Expression by HIV-1-Exposed Plasmacytoid Dendritic Cells: Impact of the Crosstalk with NK Cells.

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Saïdi, Héla; Bras, Marlène; Formaglio, Pauline; Melki, Marie-Thérèse; Charbit, Bruno; Herbeuval, Jean-Philippe; Gougeon, Marie-Lise

2016-02-01

Plasmacytoid dendritic cells (pDCs) are innate sensors of viral infections and important mediators of antiviral innate immunity through their ability to produce large amounts of IFN-α. Moreover, Toll-like receptor 7 (TLR7) and 9 (TLR9) ligands, such as HIV and CpG respectively, turn pDCs into TRAIL-expressing killer pDCs able to lyse HIV-infected CD4+ T cells. NK cells can regulate antiviral immunity by modulating pDC functions, and pDC production of IFN-α as well as cell-cell contact is required to promote NK cell functions. Impaired pDC-NK cell crosstalk was reported in the setting of HIV-1 infection, but the impact of HIV-1 on TRAIL expression and innate antiviral immunity during this crosstalk is unknown. Here, we report that low concentrations of CCR5-tropic HIV-1Ba-L promote the release of pro-inflammatory cytokines such as IFN-α, TNF-α, IFN-γ and IL-12, and CCR5-interacting chemokines (MIP-1α and MIP-1β) in NK-pDCs co-cultures. At high HIV-1BaL concentrations, the addition of NK cells did not promote the release of these mediators, suggesting that once efficiently triggered by the virus, pDCs could not integrate new activating signals delivered by NK cells. However, high HIV-1BaL concentrations were required to trigger IFN-α-mediated TRAIL expression at the surface of both pDCs and NK cells during their crosstalk. Interestingly, we identified the alarmin HMGB1, released at pDC-NK cell synapse, as an essential trigger for the secretion of IFN-α and IFN-related soluble mediators during the interplay of HIV-1 exposed pDCs with NK cells. Moreover, HMGB1 was found crucial for mTRAIL translocation to the plasma membrane of both pDCs and NK cells during their crosstalk following pDC exposure to HIV-1. Data from serum analyses of circulating HMGB1, HMGB1-specific antibodies, sTRAIL and IP-10 in a cohort of 67 HIV-1+ patients argue for the in vivo relevance of these observations. Altogether, these findings identify HMGB1 as a trigger for IFN

SUMO-modified insulin-like growth factor 1 receptor (IGF-1R) increases cell cycle progression and cell proliferation.

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Lin, Yingbo; Liu, Hongyu; Waraky, Ahmed; Haglund, Felix; Agarwal, Prasoon; Jernberg-Wiklund, Helena; Warsito, Dudi; Larsson, Olle

2017-10-01

Increasing number of studies have shown nuclear localization of the insulin-like growth factor 1 receptor (nIGF-1R) in tumor cells and its links to adverse clinical outcome in various cancers. Any obvious cell physiological roles of nIGF-1R have, however, still not been disclosed. Previously, we reported that IGF-1R translocates to cell nucleus and modulates gene expression by binding to enhancers, provided that the receptor is SUMOylated. In this study, we constructed stable transfectants of wild type IGF1R (WT) and triple-SUMO-site-mutated IGF1R (TSM) using igf1r knockout mouse fibroblasts (R-). Cell clones (R-WT and R-TSM) expressing equal amounts of IGF-1R were selected for experiments. Phosphorylation of IGF-1R, Akt, and Erk upon IGF-1 stimulation was equal in R-WT and R-TSM. WT was confirmed to enter nuclei. TSM did also undergo nuclear translocation, although to a lesser extent. This may be explained by that TSM heterodimerizes with insulin receptor, which is known to translocate to cell nuclei. R-WT proliferated substantially faster than R-TSM, which did not differ significantly from the empty vector control. Upon IGF-1 stimulation G1-S-phase progression of R-WT increased from 12 to 38%, compared to 13 to 20% of R-TSM. The G1-S progression of R-WT correlated with increased expression of cyclin D1, A, and CDK2, as well as downregulation of p27. This suggests that SUMO-IGF-1R affects upstream mechanisms that control and coordinate expression of cell cycle regulators. Further studies to identify such SUMO-IGF-1R dependent mechanisms seem important. © 2017 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals Inc.

Effects of substance P and Sar-Met-SP, a NK1 agonist, in distinct amygdaloid nuclei on anxiety-like behavior in rats.

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Bassi, Gabriel Shimizu; de Carvalho, Milene Cristina; Brandão, Marcus Lira

2014-05-21

The amygdala, together with the dorsal periaqueductal gray (dPAG), medial hypothalamus, and deep layers of the superior and inferior colliculi, constitutes the encephalic aversion system, which has been considered the main neural substrate for the organization of fear and anxiety. The basolateral nucleus of the amygdala (BLA) acts as a filter for aversive stimuli to higher structures while the central (CeA) and the medial (MeA) nuclei constitute the output for the autonomic and somatic components of the emotional reaction through major projections to the limbic and brainstem regions. Although some findings point to the distinct participation of the substance P (SP) and the NK1 receptors system in the different nuclei of the amygdala on the expression of emotional behaviors, it is not clear if this system modulates anxiety-like responses in the distinct nuclei of the amygdala as well as the dPAG. Thus, it was investigated if the injection of SP into the BLA, CeA, or MeA affects the expression of anxiety-like responses of rats submitted to the elevated plus-maze (EPM) test and, if the effects are mediated by NK1 receptors. The results showed that SP and Sar-Met-SP (NK1 receptor selective agonist) injected into the CeA and MeA, but not into the BLA, caused anxiogenic-like effects in the EPM. Altogether, the data indicates that the SP may mimic the effects of anxiogenic stimuli via NK1 receptor activation only in the CeA and MeA (amygdala's nuclei output) and may activate the neural mechanisms involved in the defensive reaction genesis. The SP/NK1 receptors system activation may be phasically involved in very specific aspects of anxiety behaviors. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Improved Preparation of Beta-D-ManNAc-(1-4)-D-Glc and Beta-d-TalNAc-(1-4)-D-Glc Disaccharides and Evaluation of Their Activating Properties on the Natural Killer Cells NKR-P1 and CD69 Receptors

Czech Academy of Sciences Publication Activity Database

Attolino, E.; Bonaccorsi, F.; Catelani, G.; D'Andrea, F.; Křenek, Karel; Bezouška, Karel; Křen, Vladimír

2008-01-01

Roč. 27, č. 3 (2008), s. 156-171 ISSN 0732-8303 R&D Projects: GA AV ČR IAA400200503; GA MŠk(CZ) LC06010; GA MŠk OC 136 Grant - others:GA MŠk(CZ) COST D34 Institutional research plan: CEZ:AV0Z50200510 Keywords : reductions * nkr-p1 receptor * nk cells Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 1.114, year: 2008

Involvement of Activating NK Cell Receptors and Their Modulation in Pathogen Immunity

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Francesco Marras

2011-01-01

Full Text Available Natural Killer (NK cells are endowed with cell-structure-sensing receptors providing inhibitory protection from self-destruction (inhibitory NK receptors, iNKRs, including killer inhibitory receptors and other molecules and rapid triggering potential leading to functional cell activation by Toll-like receptors (TLRs, cytokine receptors, and activating NK cell receptors including natural cytotoxicity receptors (NCRs, i.e., NKp46, NKp46, and NKp44. NCR and NKG2D recognize ligands on infected cells which may be endogenous or may directly bind to some structures derived from invading pathogens. In this paper, we address the known direct or indirect interactions between activating receptors and pathogens and their expression during chronic HIV and HCV infections.

Neuropeptide substance P stimulates the formation of osteoclasts via synovial fibroblastic cells

International Nuclear Information System (INIS)

Matayoshi, Takaaki; Goto, Tetsuya; Fukuhara, Eiji; Takano, Hiroshi; Kobayashi, Shigeru; Takahashi, Tetsu

2005-01-01

The present study was designed to evaluate the effects of neuropeptide substance P (Sp) on the formation of osteoclasts via synovial fibroblastic cells. Synovial fibroblastic cells derived from rat knee joint expressed the Sp receptor, neurokinin-1 receptor (NK 1 -R). The addition of Sp stimulated the proliferation of synovial fibroblastic cells and this effect was inhibited by Sp or NK 1 -R antagonists. Increased expression of the receptor activator of nuclear factor κB ligand (Rankle) in synovial fibroblastic cells after the addition of Sp was demonstrated by reverse transcriptase-polymerase chain reaction and immunofluorescence staining. Osteoprotegerin expression in synovial fibroblastic cells was decreased after incubation with SP. In co-cultures of synovial fibroblastic cells and rat peripheral blood monocytes, SP stimulated osteoclastogenesis. These results suggest that SP in the joint cavity may cause both hypertrophy of the synovium and induction of increased osteoclast formation through the increased expression of RANKL in the synovium

The pharmacology of neurokinin receptors in addiction: prospects for therapy

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Sandweiss AJ

2015-09-01

Full Text Available Alexander J Sandweiss, Todd W VanderahDepartment of Pharmacology, College of Medicine, University of Arizona, Tucson, AZ, USAAbstract: Addiction is a chronic disorder in which consumption of a substance or a habitual behavior becomes compulsive and often recurrent, despite adverse consequences. Substance p (SP is an undecapeptide and was the first neuropeptide of the neurokinin family to be discovered. The subsequent decades of research after its discovery implicated SP and its neurokinin relatives as neurotransmitters involved in the modulation of the reward pathway. Here, we review the neurokinin literature, giving a brief historical perspective of neurokinin pharmacology, localization in various brain regions involved in addictive behaviors, and the functional aspects of neurokinin pharmacology in relation to reward in preclinical models of addiction that have shaped the rational drug design of neurokinin antagonists that could translate into human research. Finally, we will cover the clinical investigations using neurokinin antagonists and discuss their potential as a therapy for drug abuse.Keywords: reward, substance p, alcohol, morphine, cocaine, dopamine

High-level expression of soluble form of mouse natural killer cell receptor NKR-P1C(B6) in Escherichia coli

Czech Academy of Sciences Publication Activity Database

Rozbeský, Daniel; Kavan, Daniel; Chmelík, Josef; Novák, Petr; Vaněk, Ondřej; Bezouška, Karel

2011-01-01

Roč. 77, č. 2 (2011), s. 178-184 ISSN 1046-5928 R&D Projects: GA ČR GA303/09/0477; GA ČR GD305/09/H008; GA ČR GAP207/10/1040; GA AV ČR IAA500200620; GA MŠk LC07017; GA MŠk LC545; GA MŠk 1M0505 Institutional research plan: CEZ:AV0Z50200510 Keywords : Natural killer cell * NKR-P1C receptor * NK1.1 antigen Subject RIV: EC - Immunology Impact factor: 1.587, year: 2011

Recent developments in the clinical pharmacology of rolapitant: subanalyses in specific populations

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Rapoport BL

2017-09-01

Full Text Available Bernardo Leon Rapoport,1 Matti Aapro,2 Martin R Chasen,3 Karin Jordan,4 Rudolph M Navari,5 Ian Schnadig,6 Lee Schwartzberg7 1The Medical Oncology Centre of Rosebank, Johannesburg, South Africa; 2Breast Center, Genolier Cancer Center, Genolier, Switzerland; 3Palliative Care, William Osler Health Services, Brampton, ON, Canada; 4Department of Medicine V, University of Heidelberg, Heidelberg, Germany; 5Division of Hematology Oncology, University of Alabama School of Medicine, Birmingham, AL, USA; 6Compass Oncology, US Oncology Research, Tualatin, OR, USA; 7West Clinic, Memphis, TN, USA Abstract: Knowledge of the involvement of the neurokinin substance P in emesis has led to the development of the neurokinin-1 receptor antagonists (NK-1 RAs for control of chemotherapy-induced nausea and vomiting (CINV, in combination with serotonin type 3 receptor antagonists and corticosteroids. The NK-1 RA rolapitant, recently approved in oral formulation, has nanomolar affinity for the NK-1 receptor, as do the other commercially available NK-1 RAs, aprepitant and netupitant. Rolapitant is rapidly absorbed and has a long half-life in comparison to aprepitant and netupitant. All three NK-1 RAs undergo metabolism by cytochrome P450 (CYP 3A4, necessitating caution with the concomitant use of CYP3A4 inhibitors, but in contrast to aprepitant and netupitant, rolapitant does not inhibit or induce CYP3A4. However, rolapitant is a moderate inhibitor of CYP2D6, and concomitant use with CYP2D6 substrates with narrow therapeutic indices should be avoided. Aprepitant, netupitant, and rolapitant have all demonstrated efficacy in the control of delayed CINV in patients receiving moderately and highly emetogenic chemotherapy in randomized controlled trials, including over multiple cycles of chemotherapy. We reviewed recent post hoc analyses of clinical trial data demonstrating that rolapitant is efficacious in the control of CINV in patient populations with specific tumor types

Differential pharmacology and clinical utility of rolapitant in chemotherapy-induced nausea and vomiting

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Rapoport BL

2017-02-01

Full Text Available Bernardo Leon Rapoport The Medical Oncology Centre of Rosebank, Johannesburg, South Africa Abstract: Chemotherapy-induced nausea and vomiting (CINV is a debilitating side effect of many cytotoxic chemotherapy regimens. CINV typically manifests during two well-defined time periods (acute and delayed phases. The acute phase is the first 24 hours after chemotherapy and is largely managed with 5-hydroxytryptamine 3 receptor antagonists. The delayed phase, a 5-day at-risk period during which patients are not often in direct contact with their health care provider, remains a significant unmet medical need. Neurokinin-1 (NK-1 receptor antagonists have demonstrated protection against acute and delayed CINV in patients treated with highly emetogenic chemotherapy and moderately emetogenic chemotherapy when used in combination with a 5-hydroxytryptamine 3 receptor antagonist and dexamethasone. Furthermore, recent data indicate that this protection is maintained over multiple treatment cycles. Rolapitant, a selective and long-acting NK-1 receptor antagonist, is approved as oral formulation for the prevention of delayed CINV in adults. This review discusses the differential pharmacology and clinical utility of rolapitant in preventing CINV compared with other NK-1 receptor antagonists. Keywords: antiemetics, highly emetogenic chemotherapy, moderately emetogenic chemotherapy, delayed chemotherapy-induced nausea and vomiting, emesis, neurokinin-1 receptor antagonists

Deoxynojirimycin and its hexosaminyl derivatives bind to natural killer cell receptors rNKR-P1A and hCD69

Czech Academy of Sciences Publication Activity Database

Catelani, G.; D’Andrea, F.; Griselli, A.; Guazelli, L.; Němcová, P.; Bezouška, K.; Křenek, Karel; Křen, Vladimír

2010-01-01

Roč. 20, č. 15 (2010), s. 4645-4648 ISSN 0960-894X R&D Projects: GA MŠk OC 136; GA MŠk(CZ) LC06010 Institutional research plan: CEZ:AV0Z50200510 Keywords : Deoxynojirimycin * NK cells * hCD69 receptors Subject RIV: CC - Organic Chemistry Impact factor: 2.661, year: 2010

(S-1-[3,5-Bis(trifluoromethylphenyl]-N-methylethylamine–(R-2-hydroxybutanedioic acid (1/1

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Hai-Bin Zhu

2009-01-01

Full Text Available In the title compound, C11H11F6N·C4H6O5, a key intermediate in the synthesis of the NK1 receptor antagonist of casopitant, the F atoms of the trifluoromethyl groups are disordered over two sites with equal occupancies. In the crystal, the components are linked by bifurcated N—H...(O,O hydrogen bonds.

Increased Numbers of NK Cells, NKT-Like Cells, and NK Inhibitory Receptors in Peripheral Blood of Patients with Chronic Obstructive Pulmonary Disease

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Ying Tang

2013-01-01

Full Text Available T cells and B cells participate in the pathogenesis of COPD. Currently, NK cells and NKT cells have gained increasing attention. In the present study, 19 COPD patients and 12 healthy nonsmokers (HNS were recruited, and their pulmonary function was assessed. The frequencies of CD3+ T, CD4+ T, CD8+ T, B, NK, and NKT-like cells were determined using flow cytometry. The frequencies of spontaneous and inducible IFN-γ+ or CD107a+ NK and NKT-like cells as well as activating or inhibitory receptors were also detected. The potential association of lymphocyte subsets with disease severity was further analyzed. Significantly decreased numbers of CD3+ and CD4+ T cells, and the CD4+/CD8+ ratio, but increased numbers of CD3−CD56+ NK and CD3+CD56+ NKT-like cells were observed in COPD patients compared to HNS. The frequencies of inducible IFN-γ-secreting NK and NKT-like cells were less in COPD patients. The frequencies of CD158a and CD158b on NK cells and CD158b on NKT-like cells were greater. The frequency of CD158b+ NK cells was negatively correlated with FEV1% prediction and FEV1/FVC. Our data indicate that COPD patients have immune dysfunction, and higher frequencies of inhibitory NK cells and NKT-like cells may participate in the pathogenesis of COPD.

Characterization of V1R receptor (ora) genes in Lake Victoria cichlids.

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Ota, Tomoki; Nikaido, Masato; Suzuki, Hikoyu; Hagino-Yamagishi, Kimiko; Okada, Norihiro

2012-05-15

Although olfaction could play a crucial role in underwater habitats by allowing fish to sense a variety of nonvolatile chemical signals, the importance of olfaction in species-rich cichlids is still controversial. In particular, examining whether cichlids rely on olfaction for reproduction is of primary interest to understand the mechanisms of speciation. In the present study, we explored the V1R (also known as ora) genes, which are believed to encode reproductive pheromone receptors in fish, in the genomes of Lake Victoria cichlids. By screening a bacterial artificial chromosome library, we identified all six intact V1R genes (V1R1 to V1R6) that have been reported in other teleost fish. Furthermore, RT-PCR and in situ hybridization analyses showed that all of the V1R genes were expressed in the olfactory epithelium, indicating that these receptors are functional in cichlids. These observations indicate that cichlids use V1R-mediated olfaction in some ways for their social behaviors. Copyright © 2012 Elsevier B.V. All rights reserved.

Activation of TRPV1 by capsaicin induces functional Kinin B1 receptor in rat spinal cord microglia

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Talbot Sébastien

2012-01-01

.o., glutamate NMDA receptor (DL-AP5, 10 μg/site, i.t., substance P NK-1 receptor (RP-67580, 10 μg/site, i.t. and nitric oxide synthase (L-NNA, 10 μg/site, i.t.. The B1R fluorescent agonist was co-localized with an immunomarker of microglia (Iba-1 in spinal cord dorsal horn of capsaicin-treated rats. Conclusion This study highlights a new mechanism for B1R induction via TRPV1 activation and establishes a link between these two pro-nociceptive receptors in inflammatory pain.

NK cell activation: distinct stimulatory pathways counterbalancing inhibitory signals.

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Bakker, A B; Wu, J; Phillips, J H; Lanier, L L

2000-01-01

A delicate balance between positive and negative signals regulates NK cell effector function. Activation of NK cells may be initiated by the triggering of multiple adhesion or costimulatory molecules, and can be counterbalanced by inhibitory signals induced by receptors for MHC class I. A common pathway of inhibitory signaling is provided by immunoreceptor tyrosine-based inhibitory motifs (ITIMs) in the cytoplasmic domains of these receptors which mediate the recruitment of SH2 domain-bearing tyrosine phosphate-1 (SHP-1). In contrast to the extensive progress that has been made regarding the negative regulation of NK cell function, our knowledge of the signals that activate NK cells is still poor. Recent studies of the activating receptor complexes have shed new light on the induction of NK cell effector function. Several NK receptors using novel adaptors with immunoreceptor tyrosine-based activation motifs (ITAMs) and with PI 3-kinase recruiting motifs have been implicated in NK cell stimulation.

A Novel Toll-Like Receptor 9 Agonist, MGN1703, Enhances HIV-1 Transcription and NK Cell-Mediated Inhibition of HIV-1-Infected Autologous CD4+ T Cells.

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Offersen, Rasmus; Nissen, Sara Konstantin; Rasmussen, Thomas A; Østergaard, Lars; Denton, Paul W; Søgaard, Ole Schmeltz; Tolstrup, Martin

2016-05-01

Toll-like receptor (TLR) agonists are potent enhancers of innate antiviral immunity and may also reverse HIV-1 latency. Therefore, TLR agonists have a potential role in the context of a "shock-and-kill" approach to eradicate HIV-1. Our extensive preclinical evaluation suggests that a novel TLR9 agonist, MGN1703, may indeed perform both functions in an HIV-1 eradication trial. Peripheral blood mononuclear cells (PBMCs) from aviremic HIV-1-infected donors on antiretroviral therapy (ART) that were incubated with MGN1703 ex vivo exhibited increased secretion of interferon alpha (IFN-α) (P= 0.005) and CXCL10 (P= 0.0005) in culture supernatants. Within the incubated PBMC pool, there were higher proportions of CD69-positive CD56(dim)CD16(+)NK cells (P= 0.001) as well as higher proportions of CD107a-positive (P= 0.002) and IFN-γ-producing (P= 0.038) NK cells. Incubation with MGN1703 also increased the proportions of CD69-expressing CD4(+)and CD8(+)T cells. Furthermore, CD4(+)T cells within the pool of MGN1703-incubated PBMCs showed enhanced levels of unspliced HIV-1 RNA (P= 0.036). Importantly, MGN1703 increased the capacity of NK cells to inhibit virus spread within a culture of autologous CD4(+)T cells assessed by using an HIV-1 p24 enzyme-linked immunosorbent assay (ELISA) (P= 0.03). In conclusion, we show that MGN1703 induced strong antiviral innate immune responses, enhanced HIV-1 transcription, and boosted NK cell-mediated suppression of HIV-1 infection in autologous CD4(+)T cells. These findings support clinical testing of MGN1703 in HIV-1 eradication trials. We demonstrate that MGN1703 (a TLR9 agonist currently undergoing phase 3 clinical testing for the treatment of metastatic colorectal cancer) induces potent antiviral responses in immune effector cells from HIV-1-infected individuals on suppressive antiretroviral therapy. The significantly improved safety and tolerability profiles of MGN1703 versus TLR9 agonists of the CpG-oligodeoxynucleotide (CpG-ODN) family

Transcription Factor Foxo1 Is a Negative Regulator of NK Cell Maturation and Function

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Deng, Youcai; Kerdiles, Yann; Chu, Jianhong; Yuan, Shunzong; Wang, Youwei; Chen, Xilin; Mao, Hsiaoyin; Zhang, Lingling; Zhang, Jianying; Hughes, Tiffany; Deng, Yafei; Zhang, Qi; Wang, Fangjie; Zou, Xianghong; Liu, Chang-Gong; Freud, Aharon G.; Li, Xiaohui; Caligiuri, Michael A; Vivier, Eric; Yu, Jianhua

2015-01-01

SUMMARY Little is known about the role of negative regulators in controlling natural killer (NK) cell development and effector functions. Foxo1 is a multifunctional transcription factor of the forkhead family. Using a mouse model of conditional deletion in NK cells, we found that Foxo1 negatively controlled NK cell differentiation and function. Immature NK cells expressed abundant Foxo1 and little Tbx21 relative to mature NK cells, but these two transcription factors reversed their expression as NK cells proceeded through development. Foxo1 promoted NK cell homing to lymph nodes through upregulating CD62L expression, and impaired late-stage maturation and effector functions by repressing Tbx21 expression. Loss of Foxo1 rescued the defect in late-stage NK cell maturation in heterozygous Tbx21+/− mice. Collectively, our data reveal a regulatory pathway by which the negative regulator Foxo1 and the positive regulator Tbx21 play opposing roles in controlling NK cell development and effector functions. PMID:25769609

Differential inhibitory and activating NK cell receptor levels and NK/NKT-like cell functionality in chronic and recovered stages of chikungunya.

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Thanapati, Subrat; Ganu, Mohini A; Tripathy, Anuradha S

2017-01-01

The role of natural killer (NK; CD3-CD56+)/NKT (CD3+CD56+)-like cells in chikungunya virus (CHIKV) disease progression/recovery remains unclear. Here, we investigated the expression profiles and function of NK and NKT-like cells from 35 chronic chikungunya patients, 30 recovered individuals, and 69 controls. Percentage of NKT-like cells was low in chronic chikungunya patients. NKp30+, CD244+, DNAM-1+, and NKG2D+ NK cell percentages were also lower (MFI and/or percentage), while those of CD94+ and NKG2A+ NKT-like cells were higher (MFI and/or percentage) in chronic patients than in recovered subjects. IFN-γ and TNF-α expression on NKT-like cells was high in the chronic patients, while only IFN-γ expression on NK cells was high in the recovered individuals. Furthermore, percentage of perforin+NK cells was low in the chronic patients. Lower cytotoxic activity was observed in the chronic patients than in the controls. CD107a expression on NK and NKT-like cells post anti-CD94/anti-NKG2A blocking was comparable among the patients and controls. Upregulated inhibitory and downregulated activating NK receptor expressions on NK/NKT-like cells, lower perforin+ and CD107a+NK cells are likely responsible for inhibiting the NK and NKT-like cell function in the chronic stage of chikungunya. Therefore, deregulation of NKR expression might underlie CHIKV-induced chronicity.

Functional Characterization of MC1R-TUBB3 Intergenic Splice Variants of the Human Melanocortin 1 Receptor.

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Cecilia Herraiz

Full Text Available The melanocortin 1 receptor gene (MC1R expressed in melanocytes is a major determinant of skin pigmentation. It encodes a Gs protein-coupled receptor activated by α-melanocyte stimulating hormone (αMSH. Human MC1R has an inefficient poly(A site allowing intergenic splicing with its downstream neighbour Tubulin-β-III (TUBB3. Intergenic splicing produces two MC1R isoforms, designated Iso1 and Iso2, bearing the complete seven transmembrane helices from MC1R fused to TUBB3-derived C-terminal extensions, in-frame for Iso1 and out-of-frame for Iso2. It has been reported that exposure to ultraviolet radiation (UVR might promote an isoform switch from canonical MC1R (MC1R-001 to the MC1R-TUBB3 chimeras, which might lead to novel phenotypes required for tanning. We expressed the Flag epitope-tagged intergenic isoforms in heterologous HEK293T cells and human melanoma cells, for functional characterization. Iso1 was expressed with the expected size. Iso2 yielded a doublet of Mr significantly lower than predicted, and impaired intracellular stability. Although Iso1- and Iso2 bound radiolabelled agonist with the same affinity as MC1R-001, their plasma membrane expression was strongly reduced. Decreased surface expression mostly resulted from aberrant forward trafficking, rather than high rates of endocytosis. Functional coupling of both isoforms to cAMP was lower than wild-type, but ERK activation upon binding of αMSH was unimpaired, suggesting imbalanced signaling from the splice variants. Heterodimerization of differentially labelled MC1R-001 with the splicing isoforms analyzed by co-immunoprecipitation was efficient and caused decreased surface expression of binding sites. Thus, UVR-induced MC1R isoforms might contribute to fine-tune the tanning response by modulating MC1R-001 availability and functional parameters.

Pharmacological properties of homomeric and heteromeric GluR1o and GluR3o receptors

DEFF Research Database (Denmark)

Nielsen, B S; Banke, T G; Schousboe, A

1998-01-01

Homomeric and heteromeric alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptor subunits GluR1o and GluR3o were expressed in Spodoptera frugiperda (Sf9) insect cells. Membranes containing the recombinant receptors showed a doublet of bands of the expected size (99-109 kDa) after...

Substance P induces rapid and transient membrane blebbing in U373MG cells in a p21-activated kinase-dependent manner.

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John Meshki

Full Text Available U373MG astrocytoma cells endogenously express the full-length neurokinin 1 receptor (NK1R. Substance P (SP, the natural ligand for NK1R, triggers rapid and transient membrane blebbing and we report that these morphological changes have different dynamics and intracellular signaling as compared to the changes that we have previously described in HEK293-NK1R cells. In both cell lines, the SP-induced morphological changes are Gq-independent, and they require the Rho, Rho-associated coiled-coil kinase (ROCK signaling pathway. Using confocal microscopy we have demonstrated that tubulin is phosphorylated subsequent to cell stimulation with SP and that tubulin accumulates inside the blebs. Colchicine, a tubulin polymerization inhibitor, blocked SP-induced blebbing in U373MG but not in HEK293-NK1R cells. Although p21-activated kinase (PAK is expressed in both cell lines, SP induced rapid phosphorylation of PAK in U373MG, but failed to phosphorylate PAK in HEK293-NK1R cells. The cell-permeable Rho inhibitor C3 transferase inhibited SP-induced PAK phosphorylation, but the ROCK inhibitor Y27632 had no effect on PAK phosphorylation, suggesting that Rho activates PAK in a ROCK-independent manner. Our study demonstrates that SP triggers rapid changes in cell morphology mediated by distinct intracellular signaling mechanisms in U373MG versus HEK293-NK1R cells.

Chronic In Vivo Interaction of Dendritic Cells Expressing the Ligand Rae-1ε with NK Cells Impacts NKG2D Expression and Function.

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Morvan, Maelig G; Champsaur, Marine; Reizis, Boris; Lanier, Lewis L

2017-05-01

To investigate how dendritic cells (DCs) interact with NK cells in vivo, we developed a novel mouse model in which Rae-1ε, a ligand of the NKG2D receptor, is expressed in cells with high levels of CD11c. In these CD11c-Rae1 mice, expression of Rae-1 was confirmed on all subsets of DCs and a small subset of B and T cells, but not on NK cells. DC numbers and activation status were unchanged, and NK cells in these CD11c-Rae1 mice presented the same Ly49 repertoire and maturation levels as their littermate wildtype controls. Early NK cell activation after mouse CMV infection was slightly lower than in wildtype mice, but NK cell expansion and viral control were comparable. Notably, we demonstrate that chronic interaction of NK cells with NKG2D ligand-expressing DCs leads to a reversible NKG2D down-modulation, as well as impaired NKG2D-dependent NK cell functions, including tumor rejection. In addition to generating a useful mouse model, our studies reveal in vivo the functional importance of the NK cell and DC cross-talk.

Quantitative Study of NPY-Expressing GABAergic Neurons and Axons in Rat Spinal Dorsal Horn*

OpenAIRE

Polg?r, Erika; Sardella, Thomas CP; Watanabe, Masahiko; Todd, Andrew J

2010-01-01

Between 25?40% of neurons in laminae I?III are GABAergic, and some of these express neuropeptide Y (NPY). We previously reported that NPY-immunoreactive axons form numerous synapses on lamina III projection neurons that possess the neurokinin 1 receptor (NK1r). The aims of this study were to determine the proportion of neurons and GABAergic boutons in this region that contain NPY, and to look for evidence that they selectively innervate different neuronal populations. We found that 4?6% of ne...

Growth hormone-releasing hormone as an agonist of the ghrelin receptor GHS-R1a.

Science.gov (United States)

Casanueva, Felipe F; Camiña, Jesus P; Carreira, Marcos C; Pazos, Yolanda; Varga, Jozsef L; Schally, Andrew V

2008-12-23

Ghrelin synergizes with growth hormone-releasing hormone (GHRH) to potentiate growth hormone (GH) response through a mechanism not yet fully characterized. This study was conducted to analyze the role of GHRH as a potential ligand of the ghrelin receptor, GHS-R1a. The results show that hGHRH(1-29)NH(2) (GHRH) induces a dose-dependent calcium mobilization in HEK 293 cells stably transfected with GHS-R1a an effect not observed in wild-type HEK 293 cells. This calcium rise is also observed using the GHRH receptor agonists JI-34 and JI-36. Radioligand binding and cross-linking studies revealed that calcium response to GHRH is mediated by the ghrelin receptor GHS-R1a. GHRH activates the signaling route of inositol phosphate and potentiates the maximal response to ghrelin measured in inositol phosphate turnover. The presence of GHRH increases the binding capacity of (125)I-ghrelin in a dose dependent-fashion showing a positive binding cooperativity. In addition, confocal microscopy in CHO cells transfected with GHS-R1a tagged with enhanced green fluorescent protein shows that GHRH activates the GHS-R1a endocytosis. Furthermore, the selective GHRH-R antagonists, JV-1-42 and JMR-132, act also as antagonists of the ghrelin receptor GHS-R1a. Our findings suggest that GHRH interacts with ghrelin receptor GHS-R1a, and, in consequence, modifies the ghrelin-associated intracellular signaling pathway. This interaction may represent a form of regulation, which could play a putative role in the physiology of GH regulation and appetite control.

On the Emerging Role of the Taste Receptor Type 1 (T1R Family of Nutrient-Sensors in the Musculoskeletal System

Directory of Open Access Journals (Sweden)

Shoichiro Kokabu

2017-03-01

Full Text Available The special sense of taste guides and guards food intake and is essential for body maintenance. Salty and sour tastes are sensed via ion channels or gated ion channels while G protein-coupled receptors (GPCRs of the taste receptor type 1 (T1R family sense sweet and umami tastes and GPCRs of the taste receptor type 2 (T2R family sense bitter tastes. T1R and T2R receptors share similar downstream signaling pathways that result in the stimulation of phospholipase-C-β2. The T1R family includes three members that form heterodimeric complexes to recognize either amino acids or sweet molecules such as glucose. Although these functions were originally described in gustatory tissue, T1R family members are expressed in numerous non-gustatory tissues and are now viewed as nutrient sensors that play important roles in monitoring global glucose and amino acid status. Here, we highlight emerging evidence detailing the function of T1R family members in the musculoskeletal system and review these findings in the context of the musculoskeletal diseases sarcopenia and osteoporosis, which are major public health problems among the elderly that affect locomotion, activities of daily living, and quality of life. These studies raise the possibility that T1R family member function may be modulated for therapeutic benefit.

Fc gamma receptor activation induces the tyrosine phosphorylation of both phospholipase C (PLC)-gamma 1 and PLC-gamma 2 in natural killer cells

OpenAIRE

1992-01-01

Crosslinking of the low affinity immunoglobulin G (IgG) Fc receptor (Fc gamma R type III) on natural killer (NK) cells initiates antibody- dependent cellular cytotoxicity. During this process, Fc gamma R stimulation results in the rapid activation of phospholipase C (PLC), which hydrolyzes membrane phosphoinositides, generating inositol-1,4,5- trisphosphate and sn-1,2-diacylglycerol as second messengers. We have recently reported that PLC activation after Fc gamma R stimulation can be inhibit...

Correlation between the Appearance of Neuropeptides in the Rat Trigeminal Ganglion and Reinnervation of the Healing Root Socket after Tooth Extraction

International Nuclear Information System (INIS)

Gunjigake, Kaori K.; Goto, Tetsuya; Nakao, Kayoko; Konoo, Tetsuro; Kobayashi, Shigeru; Yamaguchi, Kazunori

2006-01-01

The neuropeptide substance P (SP) modulates bone metabolism. This study examined the temporal appearance of the neuropeptides SP and brain-derived nerve growth factor (BDNF) and their receptors (neurokinin-1 receptor (NK 1 -R) and Trk B, respectively) in the rat trigeminal ganglion to investigate the role of neuropeptides in healing after tooth extraction. Rats were anesthetized and their upper right first molars were extracted; the rats were sacrificed 3 hours and 1–21 days after extraction. Their trigeminal ganglion and maxilla were removed, and cryosections were prepared and immunostained using specific antibodies against SP, BDNF, NK 1 -R, and Trk B. In the tooth sockets after extraction, new bone and a few SP-immunoreactive nerve fibers were first seen at day 7, and bone completely filled the sockets at day 21. In the trigeminal ganglion, the proportions of NK 1 -R-, BDNF-, and Trk B-immunoreactive neurons changed similarly, i.e., they initially decreased, increased rapidly to maximum levels by day 3, and then decreased gradually to control levels until 21 days. These findings suggest that the appearance of neuropeptides in the trigeminal ganglion, the reinnervation of SP-immunoreactive nerve fibers, and bone repair in the tooth socket during healing after extraction were correlated

The strength of small: Improved targeting of Insulin-like Growth Factor-1 Receptor (IGF-1R) with F(ab')2-R1507 fragments in Ewing sarcomas

NARCIS (Netherlands)

Fleuren, Emmy D. G.; Versleijen-Jonkers, Yvonne M. H.; Heskamp, Sandra; Roeffen, Melissa H. S.; Bouwman, Wilbert H.; Molkenboer-Kuenen, Janneke D. M.; van Laarhoven, Hanneke W. M.; Oyen, Wim J. G.; Boerman, Otto C.; van der Graaf, Winette T. A.

2013-01-01

To investigate whether F(ab')2-fragments of the monoclonal Insulin-like Growth Factor-1 Receptor (IGF-1R) antibody R1507 (F(ab')2-R1507) can successfully target IGF-1R in Ewing sarcomas (ES). BALB/c nude mice were subcutaneously implanted with IGF-1R-expressing human ES xenografts (EW-5 and EW-8)

The Plant Leucine-Rich Repeat Receptor-Like Kinase PSY1R from Head to Toe

DEFF Research Database (Denmark)

Oehlenschlæger, Christian Berg

PSY1R belongs to the family of plant leucine-rich repeat receptor-like kinases that play important roles in processes such as growth regulation and plant immunity response. PSY1R was proposed to be the receptor of the plant peptide hormone PSY1 which promotes cell expansion. PSY1R was furthermore...... is activated. This work provides the first study of the direct interaction between PSY1R and the peptide ligand PSY1. The binding was evaluated both for full length PSY1R expressed in plants and for the isolated extracellular domain expressed in insect cells. PSY1 binds to the extracellular domain of PSY1R...... shown to phosphorylate and regulate the activity of the plasma membrane localized H+-ATPase, AHA2. While the mechanism of PSY1R-mediated AHA2 phosphorylation has previously been studied in detail, little is known about how PSY1R binds PSY1 peptide ligand and how the intracellular PSY1R kinase domain...

TRAIL-receptor preferences in pancreatic cancer cells revisited: Both TRAIL-R1 and TRAIL-R2 have a licence to kill

International Nuclear Information System (INIS)

Mohr, Andrea; Yu, Rui; Zwacka, Ralf M.

2015-01-01

TRAIL is a potent and specific inducer of apoptosis in tumour cells and therefore is a possible new cancer treatment. It triggers apoptosis by binding to its cognate, death-inducing receptors, TRAIL-R1 and TRAIL-R2. In order to increase its activity, receptor-specific ligands and agonistic antibodies have been developed and some cancer types, including pancreatic cancer, have been reported to respond preferentially to TRAIL-R1 triggering. The aim of the present study was to examine an array of TRAIL-receptor specific variants on a number of pancreatic cancer cells and test the generality of the concept of TRAIL-R1 preference in these cells. TRAIL-R1 and TRAIL-R2 specific sTRAIL variants were designed and tested on a number of pancreatic cancer cells for their TRAIL-receptor preference. These sTRAIL variants were produced in HEK293 cells and were secreted into the medium. After having measured and normalised the different sTRAIL variant concentrations, they were applied to pancreatic and control cancer cells. Twenty-four hours later apoptosis was measured by DNA hypodiploidy assays. Furthermore, the specificities of the sTRAIL variants were validated in HCT116 cells that were silenced either for TRAIL-R1 or TRAIL-R2. Our results show that some pancreatic cancer cells use TRAIL-R1 to induce cell death, whereas other pancreatic carcinoma cells such as AsPC-1 and BxPC-3 cells trigger apoptosis via TRAIL-R2. This observation extended to cells that were naturally TRAIL-resistant and had to be sensitised by silencing of XIAP (Panc1 cells). The measurement of TRAIL-receptor expression by FACS revealed no correlation between receptor preferences and the relative levels of TRAIL-R1 and TRAIL-R2 on the cellular surface. These results demonstrate that TRAIL-receptor preferences in pancreatic cancer cells are variable and that predictions according to cancer type are difficult and that determining factors to inform the optimal TRAIL-based treatments still have to be identified

Impaired Glucose Metabolism in Mice Lacking the Tas1r3 Taste Receptor Gene.

Science.gov (United States)

Murovets, Vladimir O; Bachmanov, Alexander A; Zolotarev, Vasiliy A

2015-01-01

The G-protein-coupled sweet taste receptor dimer T1R2/T1R3 is expressed in taste bud cells in the oral cavity. In recent years, its involvement in membrane glucose sensing was discovered in endocrine cells regulating glucose homeostasis. We investigated importance of extraorally expressed T1R3 taste receptor protein in age-dependent control of blood glucose homeostasis in vivo, using nonfasted mice with a targeted mutation of the Tas1r3 gene that encodes the T1R3 protein. Glucose and insulin tolerance tests, as well as behavioral tests measuring taste responses to sucrose solutions, were performed with C57BL/6ByJ (Tas1r3+/+) inbred mice bearing the wild-type allele and C57BL/6J-Tas1r3tm1Rfm mice lacking the entire Tas1r3 coding region and devoid of the T1R3 protein (Tas1r3-/-). Compared with Tas1r3+/+ mice, Tas1r3-/- mice lacked attraction to sucrose in brief-access licking tests, had diminished taste preferences for sucrose solutions in the two-bottle tests, and had reduced insulin sensitivity and tolerance to glucose administered intraperitoneally or intragastrically, which suggests that these effects are due to absence of T1R3. Impairment of glucose clearance in Tas1r3-/- mice was exacerbated with age after intraperitoneal but not intragastric administration of glucose, pointing to a compensatory role of extraoral T1R3-dependent mechanisms in offsetting age-dependent decline in regulation of glucose homeostasis. Incretin effects were similar in Tas1r3+/+ and Tas1r3-/- mice, which suggests that control of blood glucose clearance is associated with effects of extraoral T1R3 in tissues other than the gastrointestinal tract. Collectively, the obtained data demonstrate that the T1R3 receptor protein plays an important role in control of glucose homeostasis not only by regulating sugar intake but also via its extraoral function, probably in the pancreas and brain.

Impaired Glucose Metabolism in Mice Lacking the Tas1r3 Taste Receptor Gene.

Directory of Open Access Journals (Sweden)

Vladimir O Murovets

Full Text Available The G-protein-coupled sweet taste receptor dimer T1R2/T1R3 is expressed in taste bud cells in the oral cavity. In recent years, its involvement in membrane glucose sensing was discovered in endocrine cells regulating glucose homeostasis. We investigated importance of extraorally expressed T1R3 taste receptor protein in age-dependent control of blood glucose homeostasis in vivo, using nonfasted mice with a targeted mutation of the Tas1r3 gene that encodes the T1R3 protein. Glucose and insulin tolerance tests, as well as behavioral tests measuring taste responses to sucrose solutions, were performed with C57BL/6ByJ (Tas1r3+/+ inbred mice bearing the wild-type allele and C57BL/6J-Tas1r3tm1Rfm mice lacking the entire Tas1r3 coding region and devoid of the T1R3 protein (Tas1r3-/-. Compared with Tas1r3+/+ mice, Tas1r3-/- mice lacked attraction to sucrose in brief-access licking tests, had diminished taste preferences for sucrose solutions in the two-bottle tests, and had reduced insulin sensitivity and tolerance to glucose administered intraperitoneally or intragastrically, which suggests that these effects are due to absence of T1R3. Impairment of glucose clearance in Tas1r3-/- mice was exacerbated with age after intraperitoneal but not intragastric administration of glucose, pointing to a compensatory role of extraoral T1R3-dependent mechanisms in offsetting age-dependent decline in regulation of glucose homeostasis. Incretin effects were similar in Tas1r3+/+ and Tas1r3-/- mice, which suggests that control of blood glucose clearance is associated with effects of extraoral T1R3 in tissues other than the gastrointestinal tract. Collectively, the obtained data demonstrate that the T1R3 receptor protein plays an important role in control of glucose homeostasis not only by regulating sugar intake but also via its extraoral function, probably in the pancreas and brain.

Effect of casopitant, a novel NK-1 antagonist, on the pharmacokinetics of dolasetron and granisetron.

Science.gov (United States)

Adams, Laurel M; Johnson, Brendan; Zhang, Ke; Yue, Lin; Kirby, Lyndon C; Lebowitz, Peter; Stoltz, Randall

2009-09-01

The objective of this study was to characterize the impact of casopitant, a novel neurokinin-1 receptor antagonist under investigation for the prevention of postoperative and chemotherapy-induced nausea and vomiting, on the pharmacokinetics of the commonly prescribed 5-hydroxytryptamine receptor 3 receptor antagonists, dolasetron or granisetron. In a phase I, open-label, two-part, two-period, single-sequence study, two cohorts of healthy subjects received either oral dolasetron (100 mg once daily for 3 days) or oral granisetron (2 mg once daily for 3 days) alone (period 1) and combined with oral casopitant, 150 mg day 1, 50 mg days 2 and 3 (period 2). Pharmacokinetics of hydrodolasetron and granisetron were assessed on days 1 and 3 of each period. Log-transformed area under the curve (AUC) and Cmax were statistically analyzed by performing an analysis of variance. Eighteen subjects were enrolled in the dolasetron cohort; nine subjects were CYP2D6 extensive metabolizers (EMs) and nine subjects were CYP2D6 poor metabolizers. Nineteen subjects were enrolled in the granisetron cohort. The largest changes in hydrodolasetron exposure after coadministration with casopitant were seen in CYP2D6 EMs, with a 24% increase in hydrodolasetron AUC on day 1 and 30% increase in Cmax on days 1 and 3. All other changes in hydrodolasetron exposure were granisetron exposure was not altered to any relevant extent (granisetron was well tolerated.

CD5−NK1.1+ γδ T Cells that Develop in a Bcl11b-Independent Manner Participate in Early Protection against Infection

Directory of Open Access Journals (Sweden)

Shinya Hatano

2017-10-01

Full Text Available Summary: We recently found that a unique subset of innate-like γδ T cells develops from the DN2a stage of the fetal thymus independently of the zinc-finger transcription factor B cell leukemia/lymphoma 11b (Bcl11b. Herein, we characterize these Bcl11b-independent γδ T cells in the periphery as CD5−NK1.1+ and Granzyme B+, and we show that they are capable of producing interferon (IFN-γ upon T cell receptor stimulation without Ca2+ influx. In wild-type mice, these cells were sparse in lymphoid tissues but abundant in non-lymphoid tissues, such as the liver. Bcl11b-independent CD5−NK1.1+ γδ T cells appeared and contributed to early protection before Bcl11b-dependent CD5+NK1.1− γδ T cells following Listeria monocytogenes infection, resembling their sequential appearance during development in the thymus. : Bcl11b is essential for transition from the DN2a to the DN2b stage in the thymus. Hatano et al. find that CD5−NK1.1+ γδ T cells develop from the DN2a stage in a Bcl11b-independent manner and participate in host defense at an early stage after bacterial infection in periphery. Keywords: innate immunity, γδ T cell, Bcl11b, DN2a, IFN-γ, Granzyme, IL-17A, host defense, bacteria, Listeria monocytogenes

CCR6 and NK1.1 distinguish between IL-17A and IFN-gamma-producing gammadelta effector T cells.

Science.gov (United States)

Haas, Jan D; González, Frano H Malinarich; Schmitz, Susanne; Chennupati, Vijaykumar; Föhse, Lisa; Kremmer, Elisabeth; Förster, Reinhold; Prinz, Immo

2009-12-01

Gammadelta T cells are a potent source of innate IL-17A and IFN-gamma, and they acquire the capacity to produce these cytokines within the thymus. However, the precise stages and required signals that guide this differentiation are unclear. Here we show that the CD24(low) CD44(high) effector gammadelta T cells of the adult thymus are segregated into two lineages by the mutually exclusive expression of CCR6 and NK1.1. Only CCR6+ gammadelta T cells produced IL-17A, while NK1.1+ gammadelta T cells were efficient producers of IFN-gamma but not of IL-17A. Their effector phenotype correlated with loss of CCR9 expression, particularly among the NK1.1+ gammadelta T cells. Accordingly, both gammadelta T-cell subsets were rare in gut-associated lymphoid tissues, but abundant in peripheral lymphoid tissues. There, they provided IL-17A and IFN-gamma in response to TCR-specific and TCR-independent stimuli. IL-12 and IL-18 induced IFN-gamma and IL-23 induced IL-17A production by NK1.1+ or CCR6+ gammadelta T cells, respectively. Importantly, we show that CCR6+ gammadelta T cells are more responsive to TCR stimulation than their NK1.1+ counterparts. In conclusion, our findings support the hypothesis that CCR6+ IL-17A-producing gammadelta T cells derive from less TCR-dependent selection events than IFN-gamma-producing NK1.1+ gammadelta T cells.

A chimeric antigen receptor for TRAIL-receptor 1 induces apoptosis in various types of tumor cells.

Science.gov (United States)

Kobayashi, Eiji; Kishi, Hiroyuki; Ozawa, Tatsuhiko; Hamana, Hiroshi; Nakagawa, Hidetoshi; Jin, Aishun; Lin, Zhezhu; Muraguchi, Atsushi

2014-10-31

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) and its associated receptors (TRAIL-R/TR) are attractive targets for cancer therapy because TRAIL induces apoptosis in tumor cells through TR while having little cytotoxicity on normal cells. Therefore, many agonistic monoclonal antibodies (mAbs) specific for TR have been produced, and these induce apoptosis in multiple tumor cell types. However, some TR-expressing tumor cells are resistant to TR-specific mAb-induced apoptosis. In this study, we constructed a chimeric antigen receptor (CAR) of a TRAIL-receptor 1 (TR1)-specific single chain variable fragment (scFv) antibody (TR1-scFv-CAR) and expressed it on a Jurkat T cell line, the KHYG-1 NK cell line, and human peripheral blood lymphocytes (PBLs). We found that the TR1-scFv-CAR-expressing Jurkat cells killed target cells via TR1-mediated apoptosis, whereas TR1-scFv-CAR-expressing KHYG-1 cells and PBLs killed target cells not only via TR1-mediated apoptosis but also via CAR signal-induced cytolysis, resulting in cytotoxicity on a broader range if target cells than with TR1-scFv-CAR-expressing Jurkat cells. The results suggest that TR1-scFv-CAR could be a new candidate for cancer gene therapy. Copyright © 2014 Elsevier Inc. All rights reserved.

An Examination of the Role of L-Glutamate and Inosine 5'-Monophosphate in Hedonic Taste-Guided Behavior by Mice Lacking the T1R1 + T1R3 Receptor.

Science.gov (United States)

Blonde, Ginger D; Spector, Alan C

2017-06-01

The heterodimeric T1R1 + T1R3 receptor is considered critical for normal signaling of L-glutamate and 5'-ribonucleotides in the oral cavity. However, some taste-guided responsiveness remains in mice lacking one subunit of the receptor, suggesting that other receptors are sufficient to support some behaviors. Here, mice lacking both receptor subunits (KO) and wild-type (WT, both n = 13) mice were tested in a battery of behavioral tests. Mice were trained and tested in gustometers with a concentration series of Maltrin-580, a maltodextrin, in a brief-access test (10-s trials) as a positive control. Similar tests followed with monosodium glutamate (MSG) with and without the ribonucleotide inosine 5'-monophosphate (IMP), but always in the presence of the epithelial sodium channel blocker amiloride (A). Brief-access tests were repeated following short-term (30-min) and long-term (48-h) exposures to MSG + A + IMP and were also conducted with sodium gluconate replacing MSG. Finally, progressive ratio tests were conducted with Maltrin-580 or MSG + A + IMP, to assess appetitive behavior while minimizing satiation. Overall, MSG generated little concentration-dependent responding in either food-restricted WT or KO mice, even in combination with IMP. However, KO mice licked less to the amino acid stimuli, a measure of consummatory behavior in the brief-access tests. In contrast, both groups initiated a similar number of trials and had a similar breakpoint in the progressive ratio task, both measures of appetitive (approach) behavior. Collectively, these results suggest that while the T1R1 + T1R3 receptor is necessary for consummatory responding to MSG (+IMP), other receptors are sufficient to maintain appetitive responding to this "umami" stimulus complex in food-restricted mice. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Recombinant Expression, In Vitro Refolding and Characterizing Disulfide Bonds of a Mouse Inhibitory C-Type Lectin-Like Receptor Nkrp1b

Czech Academy of Sciences Publication Activity Database

Hernychová, Lucie; Mrázek, Hynek; Ivanova, Ljubina; Kukačka, Zdeněk; Chmelík, Josef; Novák, Petr

2015-01-01

Roč. 64, č. 2015 (2015), s. 85-93 ISSN 0862-8408 R&D Projects: GA MŠk(CZ) EE2.3.30.0003; GA MŠk(CZ) ED1.1.00/02.0109; GA MŠk LO1509 Grant - others:OPPC(XE) CZ.2.16/3.1.00/24023 Institutional support: RVO:61388971 Keywords : NK cell * C-type lectin-like receptor (CTLR) * Nkrp1 Subject RIV: CE - Biochemistry Impact factor: 1.643, year: 2015

NKL homeobox gene MSX1 acts like a tumor suppressor in NK-cell leukemia.

Science.gov (United States)

Nagel, Stefan; Pommerenke, Claudia; Meyer, Corinna; Kaufmann, Maren; MacLeod, Roderick A F; Drexler, Hans G

2017-09-15

NKL homeobox gene MSX1 is physiologically expressed in lymphoid progenitors and subsequently downregulated in developing T- and B-cells. In contrast, elevated expression levels of MSX1 persist in mature natural killer (NK)-cells, indicating a functional role in this compartment. While T-cell acute lymphoblastic leukemia (T-ALL) subsets exhibit aberrant overexpression of MSX1, we show here that in malignant NK-cells the level of MSX1 transcripts is aberrantly downregulated. Chromosomal deletions at 4p16 hosting the MSX1 locus have been described in NK-cell leukemia patients. However, NK-cell lines analyzed here showed normal MSX1 gene configurations, indicating that this aberration might be uncommon. To identify alternative MSX1 regulatory mechanisms we compared expression profiling data of primary normal NK-cells and malignant NK-cell lines. This procedure revealed several deregulated genes including overexpressed IRF4, MIR155HG and MIR17HG and downregulated AUTS2, EP300, GATA3 and HHEX. As shown recently, chromatin-modulator AUTS2 is overexpressed in T-ALL subsets where it mediates aberrant transcriptional activation of MSX1. Here, our data demonstrate that in malignant NK-cell lines AUTS2 performed MSX1 activation as well, but in accordance with downregulated MSX1 transcription therein we detected reduced AUTS2 expression, a small genomic deletion at 7q11 removing exons 3 and 4, and truncating mutations in exon 1. Moreover, genomic profiling and chromosomal analyses of NK-cell lines demonstrated amplification of IRF4 at 6p25 and deletion of PRDM1 at 6q21, highlighting their potential oncogenic impact. Functional analyses performed via knockdown or forced expression of these genes revealed regulatory network disturbances effecting downregulation of MSX1 which may underlie malignant development in NK-cells.

Enhanced lysis of herpes simplex virus type 1-infected mouse cell lines by NC and NK effectors

Energy Technology Data Exchange (ETDEWEB)

Colmenares, C.; Lopez, C.

1986-05-01

Spontaneously cytotoxic murine lymphocytes lysed certain cell types infected by herpes simplex virus type 1 (HSV-1) better than uninfected cells. Although HSV-1 adsorbed to the surface of all the target cells, those in which the virus replicated more efficiently were lysed to a greater extent. As targets, the authors used cell lines that, when uninfected, were spontaneously lysed by NK cells (YAC-1) or by NC cells (WEHI-164). They also used a fibroblastoid cell line (M50) and a monocytic tumor line (PU51R), which were not spontaneously killed. NK cells lysed HSV-1-infected YAC cells better than uninfected cells, and an NC-like activity selectively lysed HSV-1-infected WEHI cells. These findings were consistent with the results of experiments performed to define the role of interferon in induction of virus-augmented cytolysis. Increased lysis of YAC-HSV and PU51R-HSV was entirely due to interferon activation and was completely abolished by performing the /sup 51/Cr-release assay in the presence of anti-interferon serum. The data show that HSV-1 infection of NK/NC targets induces increased cytotoxity, but the effector cell responsible for lysis is determined by the uninfected target, or by an interaction between the virus and target cell, rather than by a viral determinant alone.

Synthesis, radiosynthesis and biological evaluation of quinoline carboxamide analogues of Talnetant (SB 223412) to study by PET or SPECT imaging the NK-3 receptor

International Nuclear Information System (INIS)

Bennacef, Idriss

2003-01-01

With the aim of visualizing NK-3 receptors in vivo by medical imaging techniques (positron emission tomography and single photon emission computed tomography), the synthesis of novel 2-phenylquinoline-4-carboxamides, derived from SB 223412 or SB 222 200, two non peptide potent (hNK-3-CHO: K i = 1.0 nM; K i = 4.4 nM respectively) and selective (NK-3/NK-1 ≥10 5 ) human NK-3 receptor antagonists, has been studied. The compounds obtained are characterized by the presence, on the quinoline moiety, of an atom of carbon, fluorine or iodine that can be substituted by their β + emitter (carbon-11, t(1/2) = 20 min; fluorine-18, t(1/2) = 110 min) or γ emitter (iodine-123, t(1/2) = 13 h) radio-isotopes. Monohalogenated (fluorinated or iodinated) quinolines were prepared using an efficient synthesis based on the Pfitzinger reaction. Di-halogenated analogues, bearing both an iodine and a fluorine atom, were obtained via a regioselective ortho-metalation reaction. Biological evaluation of these compounds led to the determination of potent and selective structures towards the NK-3 receptor, for which labeling will be envisaged. Functionalization in position 3 of quinoline-4-carboxamides, using a directed metalation reaction, allowed the introduction of various (hetero)elements (chlorine, bromine, iodine, trialkystannyle or silyle moiety). This research allowed to perform the radiosynthesis of [ 11 C]SB 222200 using a Stille coupling with [ 11 C]iodomethane. (author) [fr

Absence of torsion for NK_1(R) over associative rings

OpenAIRE

Basu, Rabeya

2010-01-01

When R is a commutative ring with identity, and if k is a natural number with kR = R, then C. Weibel proved that SK_1(R[X]) has no k-torsion. We reprove his result for any associative ring R with identity in which kR = R.

Treatment with the NK1 antagonist emend reduces blood brain barrier dysfunction and edema formation in an experimental model of brain tumors.

Directory of Open Access Journals (Sweden)

Elizabeth Harford-Wright

Full Text Available The neuropeptide substance P (SP has been implicated in the disruption of the blood-brain barrier (BBB and development of cerebral edema in acute brain injury. Cerebral edema accumulates rapidly around brain tumors and has been linked to several tumor-associated deficits. Currently, the standard treatment for peritumoral edema is the corticosteroid dexamethasone, prolonged use of which is associated with a number of deleterious side effects. As SP is reported to increase in many cancer types, this study examined whether SP plays a role in the genesis of brain peritumoral edema. A-375 human melanoma cells were injected into the right striatum of male Balb/c nude mice to induce brain tumor growth, with culture medium injected in animals serving as controls. At 2, 3 or 4 weeks following tumor cell inoculation, non-treated animals were perfusion fixed for immunohistochemical detection of Albumin, SP and NK1 receptor. A further subgroup of animals was treated with a daily injection of the NK1 antagonist Emend (3 mg/kg, dexamethasone (8 mg/kg or saline vehicle at 3 weeks post-inoculation. Animals were sacrificed a week later to determine BBB permeability using Evan's Blue and brain water content. Non-treated animals demonstrated a significant increase in albumin, SP and NK1 receptor immunoreactivity in the peritumoral area as well as increased perivascular staining in the surrounding brain tissue. Brain water content and BBB permeability was significantly increased in tumor-inoculated animals when compared to controls (p<0.05. Treatment with Emend and dexamethasone reduced BBB permeability and brain water content when compared to vehicle-treated tumor-inoculated mice. The increase in peritumoral staining for both SP and the NK1 receptor, coupled with the reduction in brain water content and BBB permeability seen following treatment with the NK1 antagonist Emend, suggests that SP plays a role in the genesis of peritumoral edema, and thus warrants

Expression and Purification of Functional Ligand-binding Domains of T1R3 Taste Receptors

Energy Technology Data Exchange (ETDEWEB)

Nie,Y.; Hobbs, J.; Vigues, S.; Olson, W.; Conn, G.; Munger, S.

2006-01-01

Chemosensory receptors, including odor, taste, and vomeronasal receptors, comprise the largest group of G protein-coupled receptors (GPCRs) in the mammalian genome. However, little is known about the molecular determinants that are critical for the detection and discrimination of ligands by most of these receptors. This dearth of understanding is due in part to difficulties in preparing functional receptors suitable for biochemical and biophysical analyses. Here we describe in detail two strategies for the expression and purification of the ligand-binding domain of T1R taste receptors, which are constituents of the sweet and umami taste receptors. These class C GPCRs contain a large extracellular N-terminal domain (NTD) that is the site of interaction with most ligands and that is amenable to expression as a separate polypeptide in heterologous cells. The NTD of mouse T1R3 was expressed as two distinct fusion proteins in Escherichia coli and purified by column chromatography. Spectroscopic analysis of the purified NTD proteins shows them to be properly folded and capable of binding ligands. This methodology should not only facilitate the characterization of T1R ligand interactions but may also be useful for dissecting the function of other class C GPCRs such as the large family of orphan V2R vomeronasal receptors.

R1507, an Anti-Insulin-Like Growth Factor-1 Receptor (IGF-1R) Antibody, and EWS/FLI-1 siRNA in Ewing's Sarcoma: Convergence at the IGF/IGFR/Akt Axis

Science.gov (United States)

Rodon, Jordi; Sun, Michael; Kuenkele, Klaus-Peter; Parsons, Henrique A.; Trent, Jonathan C.; Kurzrock, Razelle

2011-01-01

A subset of patients with Ewing's sarcoma responds to anti-insulin-like growth factor-1 receptor (IGF-1R) antibodies. Mechanisms of sensitivity and resistance are unknown. We investigated whether an anti-IGF-1R antibody acts via a pathway that could also be suppressed by small interfering (si) RNA against the EWS/FLI-1 fusion protein, the hallmark of Ewing's sarcoma. The growth of two Ewing's sarcoma cell lines (TC-32 and TC-71) was inhibited by the fully human anti-IGF-1R antibody, R1507 (clonogenic and MTT assays). TC-32 and TC-71 cells express high levels of IGF-2, while RD-ES and A4573 Ewing's cell lines, which were less responsive to R1507 in our assays, express low or undetectable IGF-2, respectively. TC-71 cells also expressed high levels of IGF-1R, and R1507 decreased steady-state levels of this receptor by internalization/degradation, an effect which was associated with a decrease in p-IGF-1R, p-IRS-1, and p-Akt. EWS/FLI-1 siRNA also decreased p-Akt, due to its ability to increase IGF-BP3 levels and subsequently decrease IGF-1 and IGF-2 levels, thus inhibiting signaling through p-IGF-1R. This inhibition correlated with growth suppression and apoptosis. The attenuation of Akt activation was confirmed in TC-71 and HEK-293 (human embryonic kidney) cells by transfecting them with IGF-1R siRNA. We conclude that antibodies and siRNA to IGF-1R, as well as siRNA to EWS/FLI-1, act via intersecting IGF/IGF-1R signals that suppress a common point in this pathway, namely the phosphorylation of Akt. PMID:22022506

R1507, an anti-insulin-like growth factor-1 receptor (IGF-1R antibody, and EWS/FLI-1 siRNA in Ewing's sarcoma: convergence at the IGF/IGFR/Akt axis.

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Helen J Huang

Full Text Available A subset of patients with Ewing's sarcoma responds to anti-insulin-like growth factor-1 receptor (IGF-1R antibodies. Mechanisms of sensitivity and resistance are unknown. We investigated whether an anti-IGF-1R antibody acts via a pathway that could also be suppressed by small interfering (si RNA against the EWS/FLI-1 fusion protein, the hallmark of Ewing's sarcoma. The growth of two Ewing's sarcoma cell lines (TC-32 and TC-71 was inhibited by the fully human anti-IGF-1R antibody, R1507 (clonogenic and MTT assays. TC-32 and TC-71 cells express high levels of IGF-2, while RD-ES and A4573 Ewing's cell lines, which were less responsive to R1507 in our assays, express low or undetectable IGF-2, respectively. TC-71 cells also expressed high levels of IGF-1R, and R1507 decreased steady-state levels of this receptor by internalization/degradation, an effect which was associated with a decrease in p-IGF-1R, p-IRS-1, and p-Akt. EWS/FLI-1 siRNA also decreased p-Akt, due to its ability to increase IGF-BP3 levels and subsequently decrease IGF-1 and IGF-2 levels, thus inhibiting signaling through p-IGF-1R. This inhibition correlated with growth suppression and apoptosis. The attenuation of Akt activation was confirmed in TC-71 and HEK-293 (human embryonic kidney cells by transfecting them with IGF-1R siRNA. We conclude that antibodies and siRNA to IGF-1R, as well as siRNA to EWS/FLI-1, act via intersecting IGF/IGF-1R signals that suppress a common point in this pathway, namely the phosphorylation of Akt.

Effect Of IGF-1 On Expression Of Gh Receptor, IGF-1, IGF-1 ...

African Journals Online (AJOL)

... and the skin expression of growth hormone receptor (GHR), insulin-like growth factor1 (IGF-1), insulin-like growth factor receptor (IGF- R), KAP3.2 and KAP6-1 mRNA were measured by RT-PCR. The results indicated that IGF-1 could degrade GHR gene expression, have no effect of IGF-1 and IGF-1R gene expression, ...

A Multinational Study to Measure the Value that Patients with Cancer Place on Improved Emesis Control Following Cisplatin Chemotherapy

OpenAIRE

George Dranitsaris; Pauline Leung; Renoto Ciotti; Ana Ortega; Maria Spinthouri; Lycurgus Liaropoulos; Roberto Labianca; Antonello Quadri

2001-01-01

Background: The neurokinin-1 (NK1) receptor antagonists are a new class of agents designed to reduce the risk of emesis following chemotherapy, particularly with cisplatin. Early data from double-blind randomised trials suggest that an orally administered NK1 antagonist can reduce the absolute risk of acute and delayed emesis following cisplatin by 20 and 30%, respectively. Objective: To measure the value that patients with cancer place on improved emesis control and quality of life. Design: ...

Cannabinoid-1 receptor (CB1R) blockers as medicines: beyond obesity and cardiometabolic disorders to substance abuse/drug addiction with CB1R neutral antagonists.

Science.gov (United States)

Janero, David R

2012-03-01

Addiction to chemical substances with abuse potential presents medical needs largely unsolved by extant therapeutic strategies. Signal transmission through the cannabinoid-1 receptor (CB1R) in the central nervous system (CNS) modulates neurotransmitters/neuronal pathways contributing to the rewarding properties and hedonic effects of certain nondrug stimuli (e.g., food) and many prototypical addictive drugs, promoting excessive intake and its pathological consequences. Typical CB1R antagonists/inverse agonists reduce the rewarding effects and normalize behavioral phenotypes associated with food and abused drugs, but carry an unacceptable adverse-event profile that may reflect, at least partly, their intrinsic ability to alter basal homeostatic CB1R signaling in the CNS and elicit a negative efficacy response. Alternatively, peripherally biased CB1R inverse agonists with limited CNS permeability and putative CB1R neutral antagonists expressing modest (if any) inverse-agonist efficacy are garnering attention for treating obesity and related cardiometabolic complications with a potentially enhanced benefit-to-risk profile. This mini-review calls attention to the proposition that CB1R neutral antagonists offer attractive opportunities for pharmacotherapeutic exploitation in the substance abuse/drug addiction space, whereas the restricted CNS accessibility of peripherally biased CB1R inverse agonists circumscribes their therapeutic utility for this indication. The unique preclinical pharmacology, efficacy profiles, and reduced adverse-event risk of CB1R neutral antagonists make them worthy of translational study for their potential therapeutic application beyond obesity/cardiometabolic disease to include substance-abuse/drug-addiction disorders.

Differential expression of ligands for NKG2D and DNAM-1 receptors by epithelial ovarian cancer-derived exosomes and its influence on NK cell cytotoxicity.

Science.gov (United States)

Labani-Motlagh, Alireza; Israelsson, Pernilla; Ottander, Ulrika; Lundin, Eva; Nagaev, Ivan; Nagaeva, Olga; Dehlin, Eva; Baranov, Vladimir; Mincheva-Nilsson, Lucia

2016-04-01

Cancers constitutively produce and secrete into the blood and other biofluids 30-150 nm-sized endosomal vehicles called exosomes. Cancer-derived exosomes exhibit powerful influence on a variety of biological mechanisms to the benefit of the tumors that produce them. We studied the immunosuppressive ability of epithelial ovarian cancer (EOC) exosomes on two cytotoxic pathways of importance for anticancer immunity-the NKG2D receptor-ligand pathway and the DNAM-1-PVR/nectin-2 pathway. Using exosomes, isolated from EOC tumor explant and EOC cell-line culture supernatants, and ascitic fluid from EOC patients, we studied the expression of NKG2D and DNAM-1 ligands on EOC exosomes and their ability to downregulate the cognate receptors. Our results show that EOC exosomes differentially and constitutively express NKG2D ligands from both MICA/B and ULBP families on their surface, while DNAM-1 ligands are more seldom expressed and not associated with the exosomal membrane surface. Consequently, the NKG2D ligand-bearing EOC exosomes significantly downregulated the NKG2D receptor expression on peripheral blood mononuclear cells (PBMC) while the DNAM-1 receptor was unaffected. The downregulation of NKG2D receptor expression was coupled to inhibition of NKG2D receptor-ligand-mediated degranulation and cytotoxicity measured in vitro with OVCAR-3 and K562 cells as targets. The EOC exosomes acted as a decoy impairing the NKG2D mediated cytotoxicity while the DNAM-1 receptor-ligand system remained unchanged. Taken together, our results support and explain the mechanism behind the recently reported finding that in EOC, NK-cell recognition and killing of tumor cells was mainly dependent on DNAM-1 signaling while the contribution of the NKG2D receptor-ligand pathway was complementary and uncertain.

A Significant Role of the Truncated Ghrelin Receptor GHS-R1b in Ghrelin-induced Signaling in Neurons*

Science.gov (United States)

Navarro, Gemma; Aguinaga, David; Angelats, Edgar; Medrano, Mireia; Moreno, Estefanía; Mallol, Josefa; Cortés, Antonio; Canela, Enric I.; Casadó, Vicent; McCormick, Peter J.; Lluís, Carme; Ferré, Sergi

2016-01-01

The truncated non-signaling ghrelin receptor growth hormone secretagogue R1b (GHS-R1b) has been suggested to simply exert a dominant negative role in the trafficking and signaling of the full and functional ghrelin receptor GHS-R1a. Here we reveal a more complex modulatory role of GHS-R1b. Differential co-expression of GHS-R1a and GHS-R1b, both in HEK-293T cells and in striatal and hippocampal neurons in culture, demonstrates that GHS-R1b acts as a dual modulator of GHS-R1a function: low relative GHS-R1b expression potentiates and high relative GHS-R1b expression inhibits GHS-R1a function by facilitating GHS-R1a trafficking to the plasma membrane and by exerting a negative allosteric effect on GHS-R1a signaling, respectively. We found a preferential Gi/o coupling of the GHS-R1a-GHS-R1b complex in HEK-293T cells and, unexpectedly, a preferential Gs/olf coupling in both striatal and hippocampal neurons in culture. A dopamine D1 receptor (D1R) antagonist blocked ghrelin-induced cAMP accumulation in striatal but not hippocampal neurons, indicating the involvement of D1R in the striatal GHS-R1a-Gs/olf coupling. Experiments in HEK-293T cells demonstrated that D1R co-expression promotes a switch in GHS-R1a-G protein coupling from Gi/o to Gs/olf, but only upon co-expression of GHS-R1b. Furthermore, resonance energy transfer experiments showed that D1R interacts with GHS-R1a, but only in the presence of GHS-R1b. Therefore, GHS-R1b not only determines the efficacy of ghrelin-induced GHS-R1a-mediated signaling but also determines the ability of GHS-R1a to form oligomeric complexes with other receptors, promoting profound qualitative changes in ghrelin-induced signaling. PMID:27129257

Functional enhancement of AT1R potency in the presence of the TPαR is revealed by a comprehensive 7TM receptor co-expression screen.

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Jonas Tind Hansen

Full Text Available BACKGROUND: Functional cross-talk between seven transmembrane (7TM receptors can dramatically alter their pharmacological properties, both in vitro and in vivo. This represents an opportunity for the development of novel therapeutics that potentially target more specific biological effects while causing fewer adverse events. Although several studies convincingly have established the existence of 7TM receptor cross-talk, little is known about the frequencey and biological significance of this phenomenon. METHODOLOGY/PRINCIPAL FINDINGS: To evaluate the extent of synergism in 7TM receptor signaling, we took a comprehensive approach and co-expressed 123 different 7TM receptors together with the angiotensin II type 1 receptor (AT1R and analyzed how each receptor affected the angiotensin II (AngII response. To monitor the effect we used integrative receptor activation/signaling assay called Receptor Selection and Amplification Technology (R-SAT. In this screen the thromboxane A2α receptor (TPαR was the only receptor which significantly enhanced the AngII-mediated response. The TPαR-mediated enhancement of AngII signaling was significantly reduced when a signaling deficient receptor mutant (TPαR R130V was co-expressed instead of the wild-type TPαR, and was completely blocked both by TPαR antagonists and COX inhibitors inhibiting formation of thromboxane A2 (TXA2. CONCLUSIONS/SIGNIFICANCE: We found a functional enhancement of AT1R only when co-expressed with TPαR, but not with 122 other 7TM receptors. In addition, the TPαR must be functionally active, indicating the AT1R enhancement is mediated by a paracrine mechanism. Since we only found one receptor enhancing AT1R potency, our results suggest that functional augmentation through 7TM receptor cross-talk is a rare event that may require specific conditions to occur.

Differentiation of Forebrain and Hippocampal Dopamine 1-Class Receptors, D1R and D5R, in Spatial Learning and Memory

Science.gov (United States)

Sariñana, Joshua; Tonegawa, Susumu

2017-01-01

Activation of prefrontal cortical (PFC), striatal, and hippocampal dopamine 1-class receptors (D1R and D5R) is necessary for normal spatial information processing. Yet the precise role of the D1R versus the D5R in the aforementioned structures, and their specific contribution to the water-maze spatial learning task remains unknown. D1R- and D5R- specific in situ hybridization probes showed that forebrain restricted D1R and D5R KO mice (F-D1R/D5R KO) displayed D1R mRNA deletion in the medial (m)PFC, dorsal and ventral striatum, and the dentate gyrus (DG) of the hippocampus. D5R mRNA deletion was limited to the mPFC, the CA1 and DG hippocampal subregions. F-D1R/D5R KO mice were given water-maze training and displayed subtle spatial latency differences between genotypes and spatial memory deficits during both regular and reversal training. To differentiate forebrain D1R from D5R activation, forebrain restricted D1R KO (F-D1R KO) and D5R KO (F-D5R KO) mice were trained on the water-maze task. F-D1R KO animals exhibited escape latency deficits throughout regular and reversal training as well as spatial memory deficits during reversal training. F-D1R KO mice also showed perseverative behavior during the reversal spatial memory probe test. In contrast, F-D5R KO animals did not present observable deficits on the water-maze task. Because F-D1R KO mice showed water-maze deficits we tested the necessity of hippocampal D1R activation for spatial learning and memory. We trained DG restricted D1R KO (DG-D1R KO) mice on the water-maze task. DG-D1R KO mice did not present detectable spatial memory deficit, but did show subtle deficits during specific days of training. Our data provides evidence that forebrain D5R activation plays a unique role in spatial learning and memory in conjunction with D1R activation. Moreover, these data suggest that mPFC and striatal, but not DG D1R activation are essential for spatial learning and memory. PMID:26174222

Role of type I interferon receptor signaling on NK cell development and functions.

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Jean Guan

Full Text Available Type I interferons (IFN are unique cytokines transcribed from intronless genes. They have been extensively studied because of their anti-viral functions. The anti-viral effects of type I IFN are mediated in part by natural killer (NK cells. However, the exact contribution of type I IFN on NK cell development, maturation and activation has been somewhat difficult to assess. In this study, we used a variety of approaches to define the consequences of the lack of type I interferon receptor (IFNAR signaling on NK cells. Using IFNAR deficient mice, we found that type I IFN affect NK cell development at the pre-pro NK stage. We also found that systemic absence of IFNAR signaling impacts NK cell maturation with a significant increase in the CD27+CD11b+ double positive (DP compartment in all organs. However, there is tissue specificity, and only in liver and bone marrow is the maturation defect strictly dependent on cell intrinsic IFNAR signaling. Finally, using adoptive transfer and mixed bone marrow approaches, we also show that cell intrinsic IFNAR signaling is not required for NK cell IFN-γ production in the context of MCMV infection. Taken together, our studies provide novel insights on how type I IFN receptor signaling regulates NK cell development and functions.

BmpR1A is a major type 1 BMP receptor for BMP-Smad signaling during skull development.

Science.gov (United States)

Pan, Haichun; Zhang, Honghao; Abraham, Ponnu; Komatsu, Yoshihiro; Lyons, Karen; Kaartinen, Vesa; Mishina, Yuji

2017-09-01

Craniosynostosis is caused by premature fusion of one or more sutures in an infant skull, resulting in abnormal facial features. The molecular and cellular mechanisms by which genetic mutations cause craniosynostosis are incompletely characterized, and many of the causative genes for diverse types of syndromic craniosynostosis have not yet been identified. We previously demonstrated that augmentation of BMP signaling mediated by a constitutively active BMP type IA receptor (ca-BmpR1A) in neural crest cells (ca1A hereafter) causes craniosynostosis and superimposition of heterozygous null mutation of Bmpr1a rescues premature suture fusion (ca1A;1aH hereafter). In this study, we superimposed heterozygous null mutations of the other two BMP type I receptors, Bmpr1b and Acvr1 (ca1A;1bH and ca1A;AcH respectively hereafter) to further dissect involvement of BMP-Smad signaling. Unlike caA1;1aH, ca1A;1bH and ca1A;AcH did not restore the craniosynostosis phenotypes. In our in vivo study, Smad-dependent BMP signaling was decreased to normal levels in mut;1aH mice. However, BMP receptor-regulated Smads (R-Smads; pSmad1/5/9 hereafter) levels were comparable between ca1A, ca1A;1bH and ca1A;AcH mice, and elevated compared to control mice. Bmpr1a, Bmpr1b and Acvr1 null cells were used to examine potential mechanisms underlying the differences in ability of heterozygosity for Bmpr1a vs. Bmpr1b or Acvr1 to rescue the mut phenotype. pSmad1/5/9 level was undetectable in Bmpr1a homozygous null cells while pSmad1/5/9 levels did not decrease in Bmpr1b or Acvr1 homozygous null cells. Taken together, our study indicates that different levels of expression and subsequent activation of Smad signaling differentially contribute each BMP type I receptor to BMP-Smad signaling and craniofacial development. These results also suggest differential involvement of each type 1 receptor in pathogenesis of syndromic craniosynostoses. Copyright © 2017 Elsevier Inc. All rights reserved.

A chimeric receptor of the insulin-like growth factor receptor type 1 (IGFR1) and a single chain antibody specific to myelin oligodendrocyte glycoprotein activates the IGF1R signalling cascade in CG4 oligodendrocyte progenitors

NARCIS (Netherlands)

Annenkov, A.; Rigby, A.; Amor, S.; Zhou, D.M.; Yousaf, N.; Hemmer, B.; Chernajovsky, Y.

2011-01-01

In order to generate neural stem cells with increased ability to survive after transplantation in brain parenchyma we developed a chimeric receptor (ChR) that binds to myelin oligodendrocyte glycoprotein (MOG) via its ectodomain and activates the insulin-like growth factor receptor type 1 (IGF1R)

The D1 family dopamine receptor, DopR, potentiates hind leg grooming behavior in Drosophila.

Science.gov (United States)

Pitmon, E; Stephens, G; Parkhurst, S J; Wolf, F W; Kehne, G; Taylor, M; Lebestky, T

2016-03-01

Drosophila groom away debris and pathogens from the body using their legs in a stereotyped sequence of innate motor behaviors. Here, we investigated one aspect of the grooming repertoire by characterizing the D1 family dopamine receptor, DopR. Removal of DopR results in decreased hind leg grooming, as substantiated by quantitation of dye remaining on mutant and RNAi animals vs. controls and direct scoring of behavioral events. These data are also supported by pharmacological results that D1 receptor agonists fail to potentiate grooming behaviors in headless DopR flies. DopR protein is broadly expressed in the neuropil of the thoracic ganglion and overlaps with TH-positive dopaminergic neurons. Broad neuronal expression of dopamine receptor in mutant animals restored normal grooming behaviors. These data provide evidence for the role of DopR in potentiating hind leg grooming behaviors in the thoracic ganglion of adult Drosophila. This is a remarkable juxtaposition to the considerable role of D1 family dopamine receptors in rodent grooming, and future investigations of evolutionary relationships of circuitry may be warranted. © 2016 The Authors. Genes, Brain and Behavior published by International Behavioural and Neural Genetics Society and John Wiley & Sons Ltd.

Impaired associative fear learning in mice with complete loss or haploinsufficiency of AMPA GluR1 receptors

Directory of Open Access Journals (Sweden)

Michael Feyder

2007-12-01

Full Text Available There is compelling evidence that L-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA glutamate receptors containing the GluR1 subunit contribute to the molecular mechanisms associated with learning. AMPA GluR1 glutamate receptor knockout mice (KO exhibit abnormal hippocampal and amygdala plasticity, and deficits on various assays for cognition including Pavlovian fear conditioning. Here we examined associative fear learning in mice with complete absence (KO or partial loss (heterozygous mutant, HET of GluR1 on multiple fear conditioning paradigms. After multi-trial delay or trace conditioning, KO displayed impaired tone and context fear recall relative to WT, whereas HET were normal. After one-trial delay conditioning, both KO and HET showed impaired tone and context recall. HET and KO showed normal nociceptive sensitivity in the hot plate and tail flick tests. These data demonstrate that the complete absence of GluR1 subunit-containing receptors prevents the formation of associative fear memories, while GluR1 haploinsufficiency is sufficient to impair one-trial fear learning. These findings support growing evidence of a major role for GluR1-containing AMPA receptors in amygdalamediated forms of learning and memory.

Activation of type 2 cannabinoid receptors (CB2R) promotes fatty acid oxidation through the SIRT1/PGC-1α pathway

Energy Technology Data Exchange (ETDEWEB)

Zheng, Xuqin [Department of Endocrinology, First Affiliated Hospital, Nanjing Medical University, Nanjing, Jiangsu Province 210029 (China); Sun, Tao [Department of Neurology, Jinling Hospital, Nanjing University School of Medicine, Nanjing, Jiangsu Province 210002 (China); Wang, Xiaodong, E-mail: xdwang666@hotmail.com [Department of Endocrinology, First Affiliated Hospital, Nanjing Medical University, Nanjing, Jiangsu Province 210029 (China)

2013-07-05

Highlights: •TC, a CB2R specific agonist, stimulates SIRT1 activity by PKA/CREB pathway. •TC promotes PGC-1α transcriptional activity by increasing its deacetylation. •TC increases the expression of genes linked to FAO and promotes the rate of FAO. •The effects of TC in FAO are dependent on CB2R. •Suggesting CB2R as a target to treat diseases with lipid dysregulation. -- Abstract: Abnormal fatty acid oxidation has been associated with obesity and type 2 diabetes. At the transcriptional level, peroxisome proliferator-activated receptor-gamma coactivator 1α (PGC-1α) has been reported to strongly increase the ability of hormone nuclear receptors PPARα and ERRα to drive transcription of fatty acid oxidation enzymes. In this study, we report that a specific agonist of the type 2 cannabinoid receptor (CB2R) can lead to fatty acid oxidation through the PGC-1α pathway. We have found that CB2R is expressed in differentiated C2C12 myotubes, and that use of the specific agonist trans-caryophyllene (TC) stimulates sirtuin 1 (SIRT1) deacetylase activity by increasing the phosphorylation of cAMP response element-binding protein (CREB), thus leading to increased levels of PGC-1α deacetylation. This use of TC treatment increases the expression of genes linked to the fatty acid oxidation pathway in a SIRT1/PGC-1α-dependent mechanism and also drastically accelerates the rate of complete fatty acid oxidation in C2C12 myotubes, neither of which occur when CB2R mRNA is knocked down using siRNA. These results reveal that activation of CB2R by a selective agonist promotes lipid oxidation through a signaling/transcriptional pathway. Our findings imply that pharmacological manipulation of CB2R may provide therapeutic possibilities to treat metabolic diseases associated with lipid dysregulation.

NK-cell-dependent killing of colon carcinoma cells is mediated by natural cytotoxicity receptors (NCRs) and stimulated by parvovirus infection of target cells

International Nuclear Information System (INIS)

Bhat, Rauf; Rommelaere, Jean

2013-01-01

Investigating how the immune system functions during malignancies is crucial to developing novel therapeutic strategies. Natural killer (NK) cells, an important component of the innate immune system, play a vital role in immune defense against tumors and virus-infected cells. The poor survival rate in colon cancer makes it particularly important to develop novel therapeutic strategies. Oncolytic viruses, in addition to lysing tumor cells, may have the potential to augment antitumor immune responses. In the present study, we investigate the role of NK cells and how parvovirus H-1PV can modulate NK-cell mediated immune responses against colon carcinoma. Human NK cells were isolated from the blood of healthy donors. The cytotoxicity and antibody-mediated inhibition of NK cells were measured in chromium release assays. Phenotypic assessment of colon cancer and dendritic cells was done by FACS. The statistical significance of the results was calculated with Student’s t test (*p <0.05; **, p < 0.01; ***, p < 0.001). We show that IL-2-activated human NK cells can effectively kill colon carcinoma cells. Killing of colon carcinoma cells by NK cells was further enhanced upon infection of the former cells with parvovirus H-1PV. H-1PV has potent oncolytic activity against various tumors, yet its direct killing effect on colon carcinoma cells is limited. The cytotoxicity of NK cells towards colon carcinoma cells, both mock- and H-1PV-infected, was found to be mostly mediated by a combination of natural cytotoxicity receptors (NCRs), namely NKp30, 44, and 46. Colon carcinoma cells displayed low to moderate expression of NK cell ligands, and this expression was modulated upon H-1PV infection. Lysates of H-1PV-infected colon carcinoma cells were found to increase MHC class II expression on dendritic cells. Altogether, these data suggest that IL-2-activated NK cells actively kill colon carcinoma cells and that this killing is mediated by several natural cytotoxicity receptors

A Significant Role of the Truncated Ghrelin Receptor GHS-R1b in Ghrelin-induced Signaling in Neurons.

Science.gov (United States)

Navarro, Gemma; Aguinaga, David; Angelats, Edgar; Medrano, Mireia; Moreno, Estefanía; Mallol, Josefa; Cortés, Antonio; Canela, Enric I; Casadó, Vicent; McCormick, Peter J; Lluís, Carme; Ferré, Sergi

2016-06-17

The truncated non-signaling ghrelin receptor growth hormone secretagogue R1b (GHS-R1b) has been suggested to simply exert a dominant negative role in the trafficking and signaling of the full and functional ghrelin receptor GHS-R1a. Here we reveal a more complex modulatory role of GHS-R1b. Differential co-expression of GHS-R1a and GHS-R1b, both in HEK-293T cells and in striatal and hippocampal neurons in culture, demonstrates that GHS-R1b acts as a dual modulator of GHS-R1a function: low relative GHS-R1b expression potentiates and high relative GHS-R1b expression inhibits GHS-R1a function by facilitating GHS-R1a trafficking to the plasma membrane and by exerting a negative allosteric effect on GHS-R1a signaling, respectively. We found a preferential Gi/o coupling of the GHS-R1a-GHS-R1b complex in HEK-293T cells and, unexpectedly, a preferential Gs/olf coupling in both striatal and hippocampal neurons in culture. A dopamine D1 receptor (D1R) antagonist blocked ghrelin-induced cAMP accumulation in striatal but not hippocampal neurons, indicating the involvement of D1R in the striatal GHS-R1a-Gs/olf coupling. Experiments in HEK-293T cells demonstrated that D1R co-expression promotes a switch in GHS-R1a-G protein coupling from Gi/o to Gs/olf, but only upon co-expression of GHS-R1b. Furthermore, resonance energy transfer experiments showed that D1R interacts with GHS-R1a, but only in the presence of GHS-R1b. Therefore, GHS-R1b not only determines the efficacy of ghrelin-induced GHS-R1a-mediated signaling but also determines the ability of GHS-R1a to form oligomeric complexes with other receptors, promoting profound qualitative changes in ghrelin-induced signaling. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Comparative functional analysis of two fibroblast growth factor receptor 1 (FGFR1) mutations affecting the same residue (R254W and R254Q) in isolated hypogonadotropic hypogonadism (IHH).

Science.gov (United States)

Koika, Vasiliki; Varnavas, Petros; Valavani, Helen; Sidis, Yisrael; Plummer, Lacey; Dwyer, Andrew; Quinton, Richard; Kanaka-Gantenbein, Christine; Pitteloud, Nelly; Sertedaki, Amalia; Dacou-Voutetakis, Catherine; Georgopoulos, Neoklis A

2013-03-01

FGFR1 mutations have been identified in both Kallmann syndrome and normosmic HH (nIHH). To date, few mutations in the FGFR1 gene have been structurally or functionally characterized in vitro to identify molecular mechanisms that contribute to the disease pathogenesis. We attempted to define the in vitro functionality of two FGFR1 mutants (R254W and R254Q), resulting from two different amino acid substitutions of the same residue, and to correlate the in vitro findings to the patient phenotypes. Two unrelated GnRH deficient probands were found to harbor mutations in FGFR1 (R254W and R254Q). Mutant signaling activity and expression levels were evaluated in vitro and compared to a wild type (WT) receptor. Signaling activity was determined by a FGF2/FGFR1 dependent transcription reporter assay. Receptor total expression levels were assessed by Western blot and cell surface expression was measured by a radiolabeled antibody binding assay. The R254W maximal receptor signaling capacity was reduced by 45% (p<0.01) while R254Q activity was not different from WT. However, both mutants displayed diminished total protein expression levels (40 and 30% reduction relative to WT, respectively), while protein maturation was unaffected. Accordingly, cell surface expression levels of the mutant receptors were also significantly reduced (35% p<0.01 and 15% p<0.05, respectively). The p.R254W and p.R254Q are both loss-of-function mutations as demonstrated by their reduced overall and cell surface expression levels suggesting a deleterious effect on receptor folding and stability. It appears that a tryptophan substitution at R254 is more disruptive to receptor structure than the more conserved glutamine substitution. No clear correlation between the severity of in vitro loss-of-function and phenotypic presentation could be assigned. Copyright © 2012 Elsevier B.V. All rights reserved.

Re-evaluation of the involvement of NK cells and C-type lectin-like NK receptors in modulation of immune responses by multivalent GlcNAc-terminated oligosaccharides

Czech Academy of Sciences Publication Activity Database

Grobárová, Valeria; Benson, Veronika; Rozbeský, Daniel; Novák, Petr; Černý, O.

2013-01-01

Roč. 156, 1-2 (2013), s. 110-117 ISSN 0165-2478 R&D Projects: GA MŠk ED1.1.00/02.0109 Institutional support: RVO:61388971 Keywords : Anti-tumor immunity * Carbohydrate dendrimer * NK cells Subject RIV: EC - Immunology Impact factor: 2.367, year: 2013

Multi-cellular natural killer (NK) cell clusters enhance NK cell activation through localizing IL-2 within the cluster

Science.gov (United States)

Kim, Miju; Kim, Tae-Jin; Kim, Hye Mi; Doh, Junsang; Lee, Kyung-Mi

2017-01-01

Multi-cellular cluster formation of natural killer (NK) cells occurs during in vivo priming and potentiates their activation to IL-2. However, the precise mechanism underlying this synergy within NK cell clusters remains unclear. We employed lymphocyte-laden microwell technologies to modulate contact-mediated multi-cellular interactions among activating NK cells and to quantitatively assess the molecular events occurring in multi-cellular clusters of NK cells. NK cells in social microwells, which allow cell-to-cell contact, exhibited significantly higher levels of IL-2 receptor (IL-2R) signaling compared with those in lonesome microwells, which prevent intercellular contact. Further, CD25, an IL-2R α chain, and lytic granules of NK cells in social microwells were polarized toward MTOC. Live cell imaging of lytic granules revealed their dynamic and prolonged polarization toward neighboring NK cells without degranulation. These results suggest that IL-2 bound on CD25 of one NK cells triggered IL-2 signaling of neighboring NK cells. These results were further corroborated by findings that CD25-KO NK cells exhibited lower proliferation than WT NK cells, and when mixed with WT NK cells, underwent significantly higher level of proliferation. These data highlights the existence of IL-2 trans-presentation between NK cells in the local microenvironment where the availability of IL-2 is limited.

Activation of the LRR Receptor-Like Kinase PSY1R Requires Transphosphorylation of Residues in the Activation Loop

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Christian B. Oehlenschlæger

2017-11-01

Full Text Available PSY1R is a leucine-rich repeat (LRR receptor-like kinase (RLK previously shown to act as receptor for the plant peptide hormone PSY1 (peptide containing sulfated tyrosine 1 and to regulate cell expansion. PSY1R phosphorylates and thereby regulates the activity of plasma membrane-localized H+-ATPases. While this mechanism has been studied in detail, little is known about how PSY1R itself is activated. Here we studied the activation mechanism of PSY1R. We show that full-length PSY1R interacts with members of the SERK co-receptor family in planta. We identified seven in vitro autophosphorylation sites on serine and threonine residues within the kinase domain of PSY1R using mass spectrometry. We furthermore show that PSY1R autophosphorylation occurs in trans and that the initial transphosphorylation takes place within the activation loop at residues Ser951, Thr959, and Thr963. While Thr959 and Thr963 are conserved among other related plant LRR RLKs, Ser951 is unique to PSY1R. Based on homology modeling we propose that phosphorylation of Ser951 stabilize the inactive conformation of PSY1R.

Transmembrane and ubiquitin-like domain-containing protein 1 (Tmub1/HOPS facilitates surface expression of GluR2-containing AMPA receptors.

Directory of Open Access Journals (Sweden)

Hyunjeong Yang

Full Text Available Some ubiquitin-like (UBL domain-containing proteins are known to play roles in receptor trafficking. Alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs undergo constitutive cycling between the intracellular compartment and the cell surface in the central nervous system. However, the function of UBL domain-containing proteins in the recycling of the AMPARs to the synaptic surface has not yet been reported.Here, we report that the Transmembrane and ubiquitin-like domain-containing 1 (Tmub1 protein, formerly known as the Hepatocyte Odd Protein Shuttling (HOPS protein, which is abundantly expressed in the brain and which exists in a synaptosomal membrane fraction, facilitates the recycling of the AMPAR subunit GluR2 to the cell surface. Neurons transfected with Tmub1/HOPS-RNAi plasmids showed a significant reduction in the AMPAR current as compared to their control neurons. Consistently, the synaptic surface expression of GluR2, but not of GluR1, was significantly decreased in the neurons transfected with the Tmub1/HOPS-RNAi and increased in the neurons overexpressing EGFP-Tmub1/HOPS. The altered surface expression of GluR2 was speculated to be due to the altered surface-recycling of the internalized GluR2 in our recycling assay. Eventually, we found that GluR2 and glutamate receptor interacting protein (GRIP were coimmunoprecipitated by the anti-Tmub1/HOPS antibody from the mouse brain. Taken together, these observations show that the Tmub1/HOPS plays a role in regulating basal synaptic transmission; it contributes to maintain the synaptic surface number of the GluR2-containing AMPARs by facilitating the recycling of GluR2 to the plasma membrane.

HIV-1 Control by NK Cells via Reduced Interaction between KIR2DL2 and HLA-C∗12:02/C∗14:03.

Science.gov (United States)

Lin, Zhansong; Kuroki, Kimiko; Kuse, Nozomi; Sun, Xiaoming; Akahoshi, Tomohiro; Qi, Ying; Chikata, Takayuki; Naruto, Takuya; Koyanagi, Madoka; Murakoshi, Hayato; Gatanaga, Hiroyuki; Oka, Shinichi; Carrington, Mary; Maenaka, Katsumi; Takiguchi, Masafumi

2016-11-22

Natural killer (NK) cells control viral infection in part through the interaction between killer cell immunoglobulin-like receptors (KIRs) and their human leukocyte antigen (HLA) ligands. We investigated 504 anti-retroviral (ART)-free Japanese patients chronically infected with HIV-1 and identified two KIR/HLA combinations, KIR2DL2/HLA-C ∗ 12:02 and KIR2DL2/HLA-C ∗ 14:03, that impact suppression of HIV-1 replication. KIR2DL2 + NK cells suppressed viral replication in HLA-C ∗ 14:03 + or HLA-C ∗ 12:02 + cells to a significantly greater extent than did KIR2DL2 - NK cells in vitro. Functional analysis showed that the binding between HIV-1-derived peptide and HLA-C ∗ 14:03 or HLA-C ∗ 12:02 influenced KIR2DL2 + NK cell activity through reduced expression of the peptide-HLA (pHLA) complex on the cell surface (i.e., reduced KIR2DL2 ligand expression), rather than through reduced binding affinity of KIR2DL2 to the respective pHLA complexes. Thus, KIR2DL2/HLA-C ∗ 12:02 and KIR2DL2/HLA-C ∗ 14:03 compound genotypes have protective effects on control of HIV-1 through a mechanism involving KIR2DL2-mediated NK cell recognition of virus-infected cells, providing additional understanding of NK cells in HIV-1 infection. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

Immuno-localization of galanin receptor-1 (GALR1) in rat brain

International Nuclear Information System (INIS)

Larm, J.M.; Gundlach, A.L.

2002-01-01

Full text: Galanin is expressed in discrete areas throughout the central nervous system and has several putative physiological actions including effects on hormone secretion, reproduction and cognition, via actions at multiple G-protein-coupled receptors. Currently, three galanin receptors - GalR1, -R2, -R3 - have been identified that differ in pharmacology, signalling and distribution. The distribution of [ 125 I]-galanin binding sites presumably represents multiple receptors and so the precise regional and cellular localization of each receptor subtype is unknown. This study examined the distribution in rat brain of GalR1 receptors by immunohistochemistry, using polyclonal antibodies raised against short peptide sequences from the third intracellular loop and the proximal C-terminal. Adult rats were deeply anaesthetized (pentobarbitone 60 mg/kg, ip.) and perfusion-fixed with 4% paraformaldehyde. Specific GalR1 immunoreactivity (IR) was detected in neurons in various brain regions including cells within the olfactory bulb, piriform cortex, dorsomedial thalamus, hypothalamus (PVN, SON, ARC), midbrain/pons (intense staining in ventrolateral/medial PAG) and medulla. The localization pattern was qualitatively similar with both antisera and was consistent with that observed for GalR1 mRNA in normal rat brain. Recent evidence also reveals that GalR1- mRNA and -IR levels are coordinately altered after neuronal stimulation. These studies demonstrate a method for the identification of GalR1-containing cells that should assist in better differentiating the phenotype of galanin-receptive neurons. Copyright (2002) Australian Neuroscience Society

Determination of taste receptor type 1 member 1 (TAS1R1) gene ...

African Journals Online (AJOL)

Jane

2011-10-10

Oct 10, 2011 ... 1Institute of Cellular and Molecular Biology, College of Life Science, Xuzhou Normal University, ... TAS1R1 and TAS1R3 form an L-amino acid sensor, ... 0.5 × TBE buffer (89 mM tris–borate, 2 mM EDTA, pH 8.3) for 2 h at.

PKCɛ mediates substance P inhibition of GABAA receptors-mediated current in rat dorsal root ganglion.

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Li, Li; Zhao, Lei; Wang, Yang; Ma, Ke-tao; Shi, Wen-yan; Wang, Ying-zi; Si, Jun-qiang

2015-02-01

The mechanism underlying the modulatory effect of substance P (SP) on GABA-activated response in rat dorsal root ganglion (DRG) neurons was investigated. In freshly dissociated rat DRG neurons, whole-cell patch-clamp technique was used to record GABA-activated current and sharp electrode intracellular recording technique was used to record GABA-induced membrane depolarization. Application of GABA (1-1000 μmol/L) induced an inward current in a concentration-dependent manner in 114 out of 127 DRG neurons (89.8 %) examined with whole-cell patch-clamp recordings. Bath application of GABA (1-1000 μmol/L) evoked a depolarizing response in 236 out of 257 (91.8%) DRG neurons examined with intracellular recordings. Application of SP (0.001-1 μmol/L) suppressed the GABA-activated inward current and membrane depolarization. The inhibitory effects were concentration-dependent and could be blocked by the selective neurokinin 1 (NK1) receptors antagonist spantide but not by L659187 and SR142801 (1 μmol/L, n=7), selective antagonists of NK2 and NK3. The inhibitory effect of SP was significantly reduced by the calcium chelator BAPTA-AM, phospholipase C (PLC) inhibitor U73122, and PKC inhibitor chelerythrine, respectively. The PKA inhibitor H-89 did not affect the SP effect. Remarkably, the inhibitory effect of SP on GABA-activated current was nearly completely removed by a selective PKCε inhibitor epilon-V1-2 but not by safingol and LY333531, selective inhibitors of PKCα and PKCβ. Our results suggest that NK1 receptor mediates SP-induced inhibition of GABA-activated current and membrane depolarization by activating intracellular PLC-Ca²⁺-PKCε cascade. SP might regulate the excitability of peripheral nociceptors through inhibition of the "pre-synaptic inhibition" evoked by GABA, which may explain its role in pain and neurogenic inflammation.

Functional characterisation of homomeric ionotropic glutamate receptors GluR1-GluR6 in a fluorescence-based high throughput screening assay

DEFF Research Database (Denmark)

Strange, Mette; Bräuner-Osborne, Hans; Jensen, Anders A.

2006-01-01

We have constructed stable HEK293 cell lines expressing the rat ionotropic glutamate receptor subtypes GluR1(i), GluR2Q(i), GluR3(i), GluR4(i), GluR5Q and GluR6Q and characterised the pharmacological profiles of the six homomeric receptors in a fluorescence-based high throughput screening assay...... assay reported to date. We propose that high throughput screening of compound libraries at the six GluR-HEK293 cell lines could be helpful in the search for structurally and pharmacologically novel ligands acting at the receptors....

Sphingosine-1-phosphate (S1P) displays sustained S1P1 receptor agonism and signaling through S1P lyase-dependent receptor recycling.

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Gatfield, John; Monnier, Lucile; Studer, Rolf; Bolli, Martin H; Steiner, Beat; Nayler, Oliver

2014-07-01

The sphingosine-1-phosphate (S1P) type 1 receptor (S1P1R) is a novel therapeutic target in lymphocyte-mediated autoimmune diseases. S1P1 receptor desensitization caused by synthetic S1P1 receptor agonists prevents T-lymphocyte egress from secondary lymphoid organs into the circulation. The selective S1P1 receptor agonist ponesimod, which is in development for the treatment of autoimmune diseases, efficiently reduces peripheral lymphocyte counts and displays efficacy in animal models of autoimmune disease. Using ponesimod and the natural ligand S1P, we investigated the molecular mechanisms leading to different signaling, desensitization and trafficking behavior of S1P1 receptors. In recombinant S1P1 receptor-expressing cells, ponesimod and S1P triggered Gαi protein-mediated signaling and β-arrestin recruitment with comparable potency and efficiency, but only ponesimod efficiently induced intracellular receptor accumulation. In human umbilical vein endothelial cells (HUVEC), ponesimod and S1P triggered translocation of the endogenous S1P1 receptor to the Golgi compartment. However, only ponesimod treatment caused efficient surface receptor depletion, receptor accumulation in the Golgi and degradation. Impedance measurements in HUVEC showed that ponesimod induced only short-lived Gαi protein-mediated signaling followed by resistance to further stimulation, whereas S1P induced sustained Gαi protein-mediated signaling without desensitization. Inhibition of S1P lyase activity in HUVEC rendered S1P an efficient S1P1 receptor internalizing compound and abrogated S1P-mediated sustained signaling. This suggests that S1P lyase - by facilitating S1P1 receptor recycling - is essential for S1P-mediated sustained signaling, and that synthetic agonists are functional antagonists because they are not S1P lyase substrates. Copyright © 2014 Elsevier Inc. All rights reserved.

Synthesis and Preliminary Evaluation of a New 99mTc Labeled Substance P Analogue as a Potential Tumor Imaging Agent

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Mozaffari, Saeed; Erfani, Mostafa; Beiki, Davood; Johari Daha, Fariba; Kobarfard, Farzad; Balalaie, Saeed; Fallahi, Babak

2015-01-01

Neurokinin 1 receptors (NK1R) are overexpressed on several types of important human cancer cells. Substance P (SP) is the most specific endogenous ligand known for NK1Rs. Accordingly,a new SP analogue was synthesized and evaluated for detection of NK1R positive tumors.[6-hydrazinopyridine-3-carboxylic acid (HYNIC)-Tyr8-Met(O)11-SP] was synthesized and radiolabeled with 99mTc using ethylenediamine-N,N'-diacetic acid (EDDA)and Tricine as coligands. Common physicochemical properties of radioconjugate were studied and in-vitro cell line biological tests were accomplished to determine the receptor mediated characteristics. In-vivo biodistribution in normal and tumor bearingnude mice was also assessed. The cold peptide was prepared in high purity (>99%) and radiolabeled with 99mTc at high specific activities (84-112GBq/µmol) with an acceptable labeling yield (>95%). The radioconjugate was stable in-vitro in the presence of human serum and showed 44% protein binding to human serumalbumin. In-vitro cell line studies on U373MG cells showed an acceptable uptake up to 4.91 ± 0.22% with the ratio of 60.21 ± 1.19% for its specific fraction and increasing specific internalization during 4 h. Receptor binding assays on U373MG cells indicated a mean Kd of 2.46 ± 0.43 nM and Bmax of 128925 ± 8145 sites/cell. In-vivo investigations determined the specific tumor uptake in 3.36 percent of injected dose per gram (%ID/g) for U373MG cells and noticeable accumulations of activity in the intestines and lung. Predominant renal excretion pathway was demonstrated. Therefore, this new radiolabeled peptide could be a promising radiotracer for detection of NK1R positive primary or secondary tumors. PMID:25561916

M1 muscarinic receptor facilitates cognitive function by interplay with AMPA receptor GluA1 subunit.

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Zhao, Lan-Xue; Ge, Yan-Hui; Xiong, Cai-Hong; Tang, Ling; Yan, Ying-Hui; Law, Ping-Yee; Qiu, Yu; Chen, Hong-Zhuan

2018-03-06

M1 muscarinic acetylcholine receptors (M1 mAChRs) are the most abundant muscarinic receptors in the hippocampus and have been shown to have procognitive effects. AMPA receptors (AMPARs), an important subtype of ionotropic glutamate receptors, are key components in neurocognitive networks. However, the role of AMPARs in procognitive effects of M1 mAChRs and how M1 mAChRs affect the function of AMPARs remain poorly understood. Here, we found that basal expression of GluA1, a subunit of AMPARs, and its phosphorylation at Ser845 were maintained by M1 mAChR activity. Activation of M1 mAChRs promoted membrane insertion of GluA1, especially to postsynaptic densities. Impairment of hippocampus-dependent learning and memory by antagonism of M1 mAChRs paralleled the reduction of GluA1 expression, and improvement of learning and memory by activation of M1 mAChRs was accompanied by the synaptic insertion of GluA1 and its increased phosphorylation at Ser845. Furthermore, abrogation of phosphorylation of Ser845 residue of GluA1 ablated M1 mAChR-mediated improvement of learning and memory. Taken together, these results show a functional correlation of M1 mAChRs and GluA1 and the essential role of GluA1 in M1 mAChR-mediated cognitive improvement.-Zhao, L.-X., Ge, Y.-H., Xiong, C.-H., Tang, L., Yan, Y.-H., Law, P.-Y., Qiu, Y., Chen, H.-Z. M1 muscarinic receptor facilitates cognitive function by interplay with AMPA receptor GluA1 subunit.

Substance P increases liver fibrosis by differential changes in senescence of cholangiocytes and hepatic stellate cells.

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Wan, Ying; Meng, Fanyin; Wu, Nan; Zhou, Tianhao; Venter, Julie; Francis, Heather; Kennedy, Lindsey; Glaser, Trenton; Bernuzzi, Francesca; Invernizzi, Pietro; Glaser, Shannon; Huang, Qiaobing; Alpini, Gianfranco

2017-08-01

Substance P (SP) is involved in the proliferation of cholangiocytes in bile duct-ligated (BDL) mice and human cholangiocarcinoma growth by interacting with the neurokinin-1 receptor (NK-1R). To identify whether SP regulates liver fibrosis during cholestasis, wild-type or NK-1R knockout (NK-1R -/- ) mice that received BDL or sham surgery and multidrug resistance protein 2 knockout (Mdr2 -/- ) mice treated with either an NK-1R antagonist (L-733,060) or saline were used. Additionally, wild-type mice were treated with SP or saline intraperitoneally. In vivo, there was increased expression of tachykinin precursor 1 (coding SP) and NK-1R in both BDL and Mdr2 -/- mice compared to wild-type mice. Expression of tachykinin precursor 1 and NK-1R was significantly higher in liver samples from primary sclerosing cholangitis patients compared to healthy controls. Knockout of NK-1R decreased BDL-induced liver fibrosis, and treatment with L-733,060 resulted in decreased liver fibrosis in Mdr2 -/- mice, which was shown by decreased sirius red staining, fibrosis gene and protein expression, and reduced transforming growth factor-β1 levels in serum and cholangiocyte supernatants. Furthermore, we observed that reduced liver fibrosis in NK-1R -/- mice with BDL surgery or Mdr2 -/- mice treated with L-733,060 was associated with enhanced cellular senescence of hepatic stellate cells and decreased senescence of cholangiocytes. In vitro, L-733,060 inhibited SP-induced expression of fibrotic genes in hepatic stellate cells and cholangiocytes; treatment with L-733,060 partially reversed the SP-induced decrease of senescence gene expression in cultured hepatic stellate cells and the SP-induced increase of senescence-related gene expression in cultured cholangiocytes. Collectively, our results demonstrate the regulatory effects of the SP/NK-1R axis on liver fibrosis through changes in cellular senescence during cholestatic liver injury. (Hepatology 2017;66:528-541). © 2017 by the American

Analyses of functions of an anti-PD-L1/TGFβR2 bispecific fusion protein (M7824).

Science.gov (United States)

Jochems, Caroline; Tritsch, Sarah R; Pellom, Samuel Troy; Su, Zhen; Soon-Shiong, Patrick; Wong, Hing C; Gulley, James L; Schlom, Jeffrey

2017-09-26

M7824 (MSB0011359C) is a novel first-in-class bifunctional fusion protein consisting of a fully human IgG1 anti-PD-L1 monoclonal antibody (with structural similarities to avelumab) linked to the extracellular domain of two TGFβ receptor 2 (TGFβR2) molecules serving as a TGFβ Trap. Avelumab has demonstrated clinical activity in a range of human cancers and has been approved by the Food and Drug Administration for the therapy of Merkel cell and bladder carcinomas. Preclinical studies have shown this anti-PD-L1 is capable of mediating antibody-dependent cell-mediated cytotoxicity (ADCC). In the studies reported here, it is shown that M7824 is also capable of mediating ADCC of a wide range of human carcinoma cells in vitro , employing natural killer (NK) cells as effectors, albeit not as potent as anti-PD-L1 employing some tumor cells as targets. The addition of the IL-15 superagonist fusion protein complex ALT-803 enhanced the ADCC capacity of both anti-PD-L1 and M7824, and to levels that both agents now demonstrated similar levels of ADCC of tumor cells. TGFβ is a known immunosuppressive entity. Studies reported here show TGFβ1 induced reduction of several NK activation markers as well as reduction of endogenous NK lytic activity and NK-mediated ADCC of tumor cells. These phenomena could be reduced or mitigated, however, by M7824, but not by anti-PD-L1. M7824, but not anti-PD-L1, was also shown to reduce the immunosuppressive activity of regulatory T cells on human CD4 + T-cell proliferation. These studies thus demonstrate the dual functionalities of M7824 and provide the rationale for its further clinical development.

Preclinical evaluation of melanocortin-1 receptor (MC1-R) specific 68Ga- and 44Sc-labeled DOTA-NAPamide in melanoma imaging.

Science.gov (United States)

Nagy, Gábor; Dénes, Noémi; Kis, Adrienn; Szabó, Judit P; Berényi, Ervin; Garai, Ildikó; Bai, Péter; Hajdu, István; Szikra, Dezső; Trencsényi, György

2017-08-30

Alpha melanocyte stimulating hormone (α-MSH) enhances melanogenesis in melanoma malignum by binding to melanocortin-1 receptors (MC1-R). Earlier studies demonstrated that alpha-MSH analog NAPamide molecule specifically binds to MC1-R receptor. Radiolabeled NAPamide is a promising radiotracer for the non-invasive detection of melanin producing melanoma tumors by Positron Emission Tomography (PET). In this present study the MC1-R selectivity of the newly developed Sc-44-labeled DOTA-NAPamide was investigated in vitro and in vivo using melanoma tumors. DOTA-NAPamide was labeled with Ga-68 and Sc-44 radionuclides. The MC1-R specificity of Ga-68- and Sc-44-labeled DOTA-NAPamide was investigated in vitro and in vivo using MC1-R positive (B16-F10) and negative (A375) melanoma cell lines. For in vivo imaging studies B16-F10 and A375 tumor-bearing mice were injected with 44 Sc/ 68 Ga-DOTA-NAPamide (in blocking studies with α-MSH) and whole body PET/MRI scans were acquired. Radiotracer uptake was expressed in terms of standardized uptake values (SUVs). 44 Sc/ 68 Ga-labeled DOTA-NAPamide were produced with high specific activity (approx. 19 GBq/μmol) and with excellent radiochemical purity (99%DOTA-NAPamide (SUVmean: 0.38±0.02), and Sc-44-DOTA-NAPamide (SUVmean: 0.52±0.13) uptake was observed in subcutaneously growing B16-F10 tumors, than in receptor negative A375 tumors, where the SUVmean values of Ga-68-DOTA-NAPamide and Sc-44-DOTA-NAPamide were 0.04±0.01 and 0.07±0.01, respectively. Tumor-to-muscle (T/M SUVmean) ratios were approximately 15-fold higher in B16-F10 tumor-bearing mice, than that of A375 tumors, and this difference was also significant (p≤0.01) using both radiotracers after 60 min incubation time. Our newly synthesized 44 Sc-labeled DOTA-NAPamide probe showed excellent binding properties to melanocortin-1 receptor (MC1-R) positive melanoma cell and tumors. Due to its high specificity and sensitivity 44 Sc-DOTA-NAPamide is a promising radiotracer in

Neuroblastoma Cell Lines Are Refractory to Genotoxic Drug-Mediated Induction of Ligands for NK Cell-Activating Receptors

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Irene Veneziani

2018-01-01

Full Text Available Neuroblastoma (NB, the most common extracranial solid tumor of childhood, causes death in almost 15% of children affected by cancer. Treatment of neuroblastoma is based on the combination of chemotherapy with other therapeutic interventions such as surgery, radiotherapy, use of differentiating agents, and immunotherapy. In particular, adoptive NK cell transfer is a new immune-therapeutic approach whose efficacy may be boosted by several anticancer agents able to induce the expression of ligands for NK cell-activating receptors, thus rendering cancer cells more susceptible to NK cell-mediated lysis. Here, we show that chemotherapeutic drugs commonly used for the treatment of NB such as cisplatin, topotecan, irinotecan, and etoposide are unable to induce the expression of activating ligands in a panel of NB cell lines. Consistently, cisplatin-treated NB cell lines were not more susceptible to NK cells than untreated cells. The refractoriness of NB cell lines to these drugs has been partially associated with the abnormal status of genes for ATM, ATR, Chk1, and Chk2, the major transducers of the DNA damage response (DDR, triggered by several anticancer agents and promoting different antitumor mechanisms including the expression of ligands for NK cell-activating receptors. Moreover, both the impaired production of reactive oxygen species (ROS in some NB cell lines and the transient p53 stabilization in response to our genotoxic drugs under our experimental conditions could contribute to inefficient induction of activating ligands. These data suggest that further investigations, exploiting molecular strategies aimed to potentiate the NK cell-mediated immunotherapy of NB, are warranted.

Neuroblastoma Cell Lines Are Refractory to Genotoxic Drug-Mediated Induction of Ligands for NK Cell-Activating Receptors

Science.gov (United States)

Veneziani, Irene; Brandetti, Elisa; Ognibene, Marzia; Pezzolo, Annalisa; Pistoia, Vito

2018-01-01

Neuroblastoma (NB), the most common extracranial solid tumor of childhood, causes death in almost 15% of children affected by cancer. Treatment of neuroblastoma is based on the combination of chemotherapy with other therapeutic interventions such as surgery, radiotherapy, use of differentiating agents, and immunotherapy. In particular, adoptive NK cell transfer is a new immune-therapeutic approach whose efficacy may be boosted by several anticancer agents able to induce the expression of ligands for NK cell-activating receptors, thus rendering cancer cells more susceptible to NK cell-mediated lysis. Here, we show that chemotherapeutic drugs commonly used for the treatment of NB such as cisplatin, topotecan, irinotecan, and etoposide are unable to induce the expression of activating ligands in a panel of NB cell lines. Consistently, cisplatin-treated NB cell lines were not more susceptible to NK cells than untreated cells. The refractoriness of NB cell lines to these drugs has been partially associated with the abnormal status of genes for ATM, ATR, Chk1, and Chk2, the major transducers of the DNA damage response (DDR), triggered by several anticancer agents and promoting different antitumor mechanisms including the expression of ligands for NK cell-activating receptors. Moreover, both the impaired production of reactive oxygen species (ROS) in some NB cell lines and the transient p53 stabilization in response to our genotoxic drugs under our experimental conditions could contribute to inefficient induction of activating ligands. These data suggest that further investigations, exploiting molecular strategies aimed to potentiate the NK cell-mediated immunotherapy of NB, are warranted. PMID:29805983

Ghrelin receptor (GHS-R1A) agonists show potential as interventive agents during aging.

Science.gov (United States)

Smith, Roy G; Sun, Yuxiang; Jiang, Hong; Albarran-Zeckler, Rosie; Timchenko, Nikolai

2007-11-01

Administration of an orally active agonist (MK-0677) of the growth hormone secretagogue receptor (GHS-R1a) to elderly subjects restored the amplitude of endogenous episodic growth hormone (GH) release to that of young adults. Functional benefits include increased lean mass and bone density and modest improvements in strength. In old mice, a similar agonist partially restored function to the thymus and reduced tumor cell growth and metastasis. Treatment of old mice with the endogenous GHS-R1a agonist ghrelin restored a young liver phenotype. The mechanism involves inhibition of cyclin D3:cdk4/cdk6 activity and increased protein phosphatase-2A (PP2A) activity in liver nuclei, which stabilizes the dephosphorylated form of the transcription factor C/EBPalpha preventing the age-dependent formation of the C/EBPalpha-Rb-E2F4-Brm nuclear complex. By inhibiting formation of this complex, repression of E2F target genes is de-repressed and C/EBPalpha regulated expression of Pepck, a regulator of gluconeogenesis, is normalized, thereby restoring a young liver phenotype. In the brain, aging is associated with decline in dopamine function. We investigated the potential neuromodulatory role of GHS-R1a on dopamine action. Neurons were identified in the hippocampus, cortex, substantia nigra, and ventral tegmental areas that coexpressed GHS-R1a and dopamine receptor subtype-1 (D1R). Cell culture studies showed that, in the presence of ghrelin and dopamine, GHS-R and D1R form heterodimers, which modified G-protein signal transduction resulting in amplification of dopamine signaling. We speculate that aging is associated with deficient endogenous ghrelin signaling that can be rescued by intervention with GHS-R1a agonists to improve quality of life and maintain independence.

Sigma-1 receptor and inflammatory pain.

Science.gov (United States)

Gris, Georgia; Cobos, Enrique José; Zamanillo, Daniel; Portillo-Salido, Enrique

2015-06-01

The sigma-1 receptor (Sig-1R) is a unique ligand-regulated molecular chaperone that interacts with several protein targets such as G protein-coupled receptors and ion channels to modulate their activity. Sig-1R is located in areas of the central and peripheral nervous system that are key to pain control. Previous preclinical studies have suggested a potential therapeutic use of Sig-1R antagonists for the management of neuropathic pain. Recent studies using pharmacological and genetic tools have explored the role of Sig-1R in inflammatory pain conditions. Mice lacking the Sig-1R have shown different patterns of phenotypic responses to inflammatory injury. Systemic or peripheral administration of several Sig-1R antagonists, including the selective Sig-1R antagonist S1RA, inhibited both mechanical and thermal hypersensitivity in several preclinical models of inflammatory pain. These recent studies are summarized in the present commentary. Central and peripheral pharmacological blockade of Sig-1R could be an effective option to treat inflammatory pain.

HUMAN NK CELLS: FROM SURFACE RECEPTORS TO THE THERAPY OF LEUKEMIAS AND SOLID TUMORS

Directory of Open Access Journals (Sweden)

LORENZO eMORETTA

2014-03-01

Full Text Available Natural Killer (NK cells are major effector cells of the innate immunity. The discovery, over two decades ago, of MHC-class I specific NK receptors and subsequently of activating receptors, recognizing ligands expressed by tumor or virus-infected cells, paved the way to our understanding of the mechanisms of selective recognition and killing of tumor cells. Although NK cells can efficiently kill tumor cells of different histotypes in vitro, their activity may be limited in vivo by their inefficient trafficking to tumor lesions and by the inhibition of their function induced by tumor cells themselves and by the tumor microenvironment. On the other hand, the important role of NK cells has been clearly demonstrated in the therapy of high risk leukemias in the haploidentical hematopoietic cell (HSC transplantation setting. NK cells derived from donor HSC kill leukemic cells residual after the conditioning regimen, thus preventing leukemia relapses. In addition, they also kill residual dendritic cells and T lymphocytes, thus preventing both GvHD and graft rejection.

Enrichment of variations in KIR3DL1/S1 and KIR2DL2/L3 among H1N1/09 ICU patients: an exploratory study.

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David La

Full Text Available BACKGROUND: Infection by the pandemic influenza A (H1N1/09 virus resulted in significant pathology among specific ethnic groups worldwide. Natural Killer (NK cells are important in early innate immune responses to viral infections. Activation of NK cells, in part, depend on killer-cell immunoglobulin-like receptors (KIR and HLA class I ligand interactions. To study factors involved in NK cell dysfunction in overactive immune responses to H1N1 infection, KIR3DL1/S1 and KIR2DL2/L3 allotypes and cognate HLA ligands of H1N1/09 intensive-care unit (ICU patients were determined. METHODOLOGY AND FINDINGS: KIR3DL1/S1, KIR2DL2/L3, and HLA -B and -C of 51 H1N1/09 ICU patients and 105 H1N1-negative subjects (St. Theresa Point, Manitoba were characterized. We detected an increase of 3DL1 ligand-negative pairs (3DL1/S1(+ Bw6(+ Bw4(-, and a lack of 2DL1 HLA-C2 ligands, among ICU patients. They were also significantly enriched for 2DL2/L3 ligand-positive pairs (PVA, P=0.024, Pc=0.047; Odds Ratio:2.563, CI95%:1.109-5.923, 3DL1*00101 (Ab>VA, PSTh, P=0.034, Pc=0.268, and 3DL1*029 (Ab>STh, P=0.039, Pc=0.301. Aboriginal patients ligand-positive for 3DL1/S1 and 2DL1 had the lowest probabilities of death (R(d (R(d=28%, compared to patients that were 3DL1/S1 ligand-negative (R(d=52% or carried 3DL1*029 (R(d=52%. Relative to Caucasoids (CA, two allotypes were enriched among non-aboriginal ICU patients (NAb: 3DL1*00401 (NAb>CA, P<0.001, Pc<0.001 and 3DL1*01502 (CA1/S1, 2DL2/L3, and 2DL1 had the lowest probabilities of death (R(d=36%, compared to subjects with 3DL1*01502 (R(d=48% and/or 3DL1*00401 (R(d=58%. CONCLUSIONS: Specific KIR3DL1/S1 allotypes, 3DL1/S1 and 2DL1 ligand-negative pairs, and 2DL2/L3 ligand-positive pairs were enriched among ICU patients. This suggests a possible association with NK cell dysfunction in patients with overactive immune responses to H1N1/09, leading to

Humans Can Taste Glucose Oligomers Independent of the hT1R2/hT1R3 Sweet Taste Receptor.

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Lapis, Trina J; Penner, Michael H; Lim, Juyun

2016-08-23

It is widely accepted that humans can taste mono- and disaccharides as sweet substances, but they cannot taste longer chain oligo- and polysaccharides. From the evolutionary standpoint, the ability to taste starch or its oligomeric hydrolysis products would be highly adaptive, given their nutritional value. Here, we report that humans can taste glucose oligomer preparations (average degree of polymerization 7 and 14) without any other sensorial cues. The same human subjects could not taste the corresponding glucose polymer preparation (average degree of polymerization 44). When the sweet taste receptor was blocked by lactisole, a known sweet inhibitor, subjects could not detect sweet substances (glucose, maltose, and sucralose), but they could still detect the glucose oligomers. This suggests that glucose oligomer detection is independent of the hT1R2/hT1R3 sweet taste receptor. Human subjects described the taste of glucose oligomers as "starchy," while they describe sugars as "sweet." The dose-response function of glucose oligomer was also found to be indistinguishable from that of glucose on a molar basis. © The Author 2016. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Lower numbers of circulating natural killer T (NK T) cells in individuals with human T lymphotropic virus type 1 (HTLV-1) associated neurological disease

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Ndhlovu, L C; Snyder-Cappione, J E; Carvalho, K I; Leal, F E; Loo, C P; bruno, F R; Jha, A R; Devita, D; Hasenkrug, A M; Barbosa, H M R; Segurado, A C; Nixon, D F; Murphy, E L; Kallas, E G

2009-01-01

Human T lymphotropic virus type 1 (HTLV-1) infects 10–20 million people worldwide. The majority of infected individuals are asymptomatic; however, approximately 3% develop the debilitating neurological disease HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). There is also currently no cure, vaccine or effective therapy for HTLV-1 infection, and the mechanisms for progression to HAM/TSP remain unclear. NK T cells are an immunoregulatory T cell subset whose frequencies and effector functions are associated critically with immunity against infectious diseases. We hypothesized that NK T cells are associated with HAM/TSP progression. We measured NK T cell frequencies and absolute numbers in individuals with HAM/TSP infection from two cohorts on two continents: São Paulo, Brazil and San Francisco, CA, USA, and found significantly lower levels when compared with healthy subjects and/or asymptomatic carriers. Also, the circulating NK T cell compartment in HAM/TSP subjects is comprised of significantly more CD4+ and fewer CD8+ cells than healthy controls. These findings suggest that lower numbers of circulating NK T cells and enrichment of the CD4+ NK T subset are associated with HTLV-1 disease progression. PMID:19778295

T1r3 taste receptor involvement in gustatory neural responses to ethanol and oral ethanol preference.

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Brasser, Susan M; Norman, Meghan B; Lemon, Christian H

2010-05-01

Elevated alcohol consumption is associated with enhanced preference for sweet substances across species and may be mediated by oral alcohol-induced activation of neurobiological substrates for sweet taste. Here, we directly examined the contribution of the T1r3 receptor protein, important for sweet taste detection in mammals, to ethanol intake and preference and the neural processing of ethanol taste by measuring behavioral and central neurophysiological responses to oral alcohol in T1r3 receptor-deficient mice and their C57BL/6J background strain. T1r3 knockout and wild-type mice were tested in behavioral preference assays for long-term voluntary intake of a broad concentration range of ethanol, sucrose, and quinine. For neurophysiological experiments, separate groups of mice of each genotype were anesthetized, and taste responses to ethanol and stimuli of different taste qualities were electrophysiologically recorded from gustatory neurons in the nucleus of the solitary tract. Mice lacking the T1r3 receptor were behaviorally indifferent to alcohol (i.e., ∼50% preference values) at concentrations typically preferred by wild-type mice (5-15%). Central neural taste responses to ethanol in T1r3-deficient mice were significantly lower compared with C57BL/6J controls, a strain for which oral ethanol stimulation produced a concentration-dependent activation of sweet-responsive NTS gustatory neurons. An attenuated difference in ethanol preference between knockouts and controls at concentrations >15% indicated that other sensory and/or postingestive effects of ethanol compete with sweet taste input at high concentrations. As expected, T1r3 knockouts exhibited strongly suppressed behavioral and neural taste responses to sweeteners but did not differ from wild-type mice in responses to prototypic salt, acid, or bitter stimuli. These data implicate the T1r3 receptor in the sensory detection and transduction of ethanol taste.

Characterization of the Sweet Taste Receptor Tas1r2 from an Old World Monkey Species Rhesus Monkey and Species-Dependent Activation of the Monomeric Receptor by an Intense Sweetener Perillartine.

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Chenggu Cai

Full Text Available Sweet state is a basic physiological sensation of humans and other mammals which is mediated by the broadly acting sweet taste receptor-the heterodimer of Tas1r2 (taste receptor type 1 member 2 and Tas1r3 (taste receptor type 1 member 3. Various sweeteners interact with either Tas1r2 or Tas1r3 and then activate the receptor. In this study, we cloned, expressed and functionally characterized the taste receptor Tas1r2 from a species of Old World monkeys, the rhesus monkey. Paired with the human TAS1R3, it was shown that the rhesus monkey Tas1r2 could respond to natural sugars, amino acids and their derivates. Furthermore, similar to human TAS1R2, rhesus monkey Tas1r2 could respond to artificial sweeteners and sweet-tasting proteins. However, the responses induced by rhesus monkey Tas1r2 could not be inhibited by the sweet inhibitor amiloride. Moreover, we found a species-dependent activation of the Tas1r2 monomeric receptors of human, rhesus monkey and squirrel monkey but not mouse by an intense sweetener perillartine. Molecular modeling and sequence analysis indicate that the receptor has the conserved domains and ligand-specific interactive residues, which have been identified in the characterized sweet taste receptors up to now. This is the first report of the functional characterization of sweet taste receptors from an Old World monkey species.

Characterization of the Sweet Taste Receptor Tas1r2 from an Old World Monkey Species Rhesus Monkey and Species-Dependent Activation of the Monomeric Receptor by an Intense Sweetener Perillartine.

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Cai, Chenggu; Jiang, Hua; Li, Lei; Liu, Tianming; Song, Xuejie; Liu, Bo

2016-01-01

Sweet state is a basic physiological sensation of humans and other mammals which is mediated by the broadly acting sweet taste receptor-the heterodimer of Tas1r2 (taste receptor type 1 member 2) and Tas1r3 (taste receptor type 1 member 3). Various sweeteners interact with either Tas1r2 or Tas1r3 and then activate the receptor. In this study, we cloned, expressed and functionally characterized the taste receptor Tas1r2 from a species of Old World monkeys, the rhesus monkey. Paired with the human TAS1R3, it was shown that the rhesus monkey Tas1r2 could respond to natural sugars, amino acids and their derivates. Furthermore, similar to human TAS1R2, rhesus monkey Tas1r2 could respond to artificial sweeteners and sweet-tasting proteins. However, the responses induced by rhesus monkey Tas1r2 could not be inhibited by the sweet inhibitor amiloride. Moreover, we found a species-dependent activation of the Tas1r2 monomeric receptors of human, rhesus monkey and squirrel monkey but not mouse by an intense sweetener perillartine. Molecular modeling and sequence analysis indicate that the receptor has the conserved domains and ligand-specific interactive residues, which have been identified in the characterized sweet taste receptors up to now. This is the first report of the functional characterization of sweet taste receptors from an Old World monkey species.

α1b-Adrenergic Receptor Localization and Relationship to the D1-Dopamine Receptor in the Rat Nucleus Accumbens.

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Mitrano, Darlene A; Jackson, Kelsey; Finley, Samantha; Seeley, Allison

2018-02-10

The α1-adrenergic receptors (α1ARs) have been implicated in numerous actions of the brain, including attention and wakefulness. Additionally, they have been identified as contributing to disorders of the brain, such as drug addiction, and recent work has shown a role of these receptors in relapse to psychostimulants. While some functionality is known, the actual subcellular localization of the subtypes of the α1ARs remains to be elucidated. Further, their anatomical relationship to receptors for other neurotransmitters, such as dopamine (DA), remains unclear. Therefore, using immunohistochemistry and electron microscopy techniques, this study describes the subcellular localization of the α1b-adrenergic receptor (α1bAR), the subtype most tied to relapse behaviors, as well as its relationship to the D1-dopamine receptor (D1R) in both the shell and core of the rat nucleus accumbens (NAc). Overall, α1bARs were found in unmyelinated axons and axon terminals with some labeling in dendrites. In accordance with other studies of the striatum, the D1R was found mainly in dendrites and spines; therefore, colocalization of the D1R with the α1bAR was rare postsynaptically. However, in the NAc shell, when the receptors were co-expressed in the same neuronal elements there was a trend for both receptors to be found on the plasma membrane, as opposed to the intracellular compartment. This study provides valuable anatomical information about the α1bAR and its relationship to the D1R and the regulation of DA and norepinephrine (NE) neurotransmission in the brain which have been examined previously. Published by Elsevier Ltd.

Dissection of the role of PfEMP1 and ICAM-1 in the sensing of Plasmodium-falciparum-infected erythrocytes by natural killer cells.

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Myriam Baratin

Full Text Available BACKGROUND: Host innate immunity contributes to malaria clinical outcome by providing protective inflammatory cytokines such as interferon-gamma, and by shaping the adaptive immune response. Plasmodium falciparum (Pf is the etiologic agent of the most severe forms of human malaria. Natural Killer (NK cells are lymphocytes of the innate immune system that are the first effectors to produce interferon-gamma in response to Pf. However, the molecular bases of Pf-NK cell recognition events are unknown. Our study focuses on the role of Pf erythrocyte membrane protein 1 (PfEMP1, a major Pf virulence factor. PfEMP1 is expressed on parasitized-erythrocytes and participates to vascular obstruction through the binding to several host receptors. PfEMP1 is also a pivotal target for host antibody response to Pf infection. METHODOLOGY/PRINCIPAL FINDINGS: Using genetically-engineered parasite mutant strains, a human genetic deficiency, and blocking antibodies, we identified two receptor-ligand pairs involved in two uncoupled events occurring during the sensing of Pf infection by NK cells. First, PfEMP1 interaction with one of its host receptor, chondroitin sulfate A, mediates the cytoadhesion of Pf-infected erythrocytes to human NK cell lines, but is not required for primary NK cell activation. Second, intercellular adhesion molecule-1 (ICAM-1, another host receptor for PfEMP1, is mandatory for NK cell interferon-gamma response. In this case, ICAM-1 acts via its engagement with its host ligand, LFA-1, and not with PfEMP1, consistent with the obligatory cross-talk of NK cells with macrophages for their production of interferon-gamma. CONCLUSION/SIGNIFICANCE: PfEMP1-independent but ICAM-1/LFA-1-dependent events occurring during NK cell activation by Pf highlight the fundamental role of cellular cooperation during innate immune response to malaria.

Substance P Promotes the Proliferation, but Inhibits Differentiation and Mineralization of Osteoblasts from Rats with Spinal Cord Injury via RANKL/OPG System.

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Hai-Juan Liu

Full Text Available Spinal cord injury (SCI causes a significant amount of bone loss, which results in osteoporosis (OP. The neuropeptide substance P (SP and SP receptors may play important roles in the pathogenesis of OP after SCI. To identify the roles of SP in the bone marrow mesenchymal stem cell derived osteoblasts (BMSC-OB in SCI rats, we investigated the expression of neurokinin-1 receptors (NK1R in BMSC-OB and the effects of SP on bone formation by development of BMSC-OB cultures. Sixty young male Sprague-Dawley rats were randomized into two groups: SHAM and SCI. The expression of NK1R protein in BMSC-OB was observed using immunohistochemistry and Western blot analysis. The dose- and time-dependent effects of SP on the proliferation, differentiation and mineralization of BMSC-OB and the expression of osteoblastic markers by in vitro experiments. The expression of NK1R in BMSC-OB was observed on plasma membranes and in cytoplasm. One week after osteogenic differentiation, the expression of NK1R was significantly increased after SCI at mRNA and protein levels. However, this difference was gradually attenuated at 2 or 3 weeks later. SP have the function to enhance cell proliferation, inhibite cell differentiation and mineralization at a proper concentration and incubation time, and this effect would be inhibited by adding SP or NK1R antagonist. The expression of RANKL/OPG was significantly increased in tibiae after SCI. Similarly, the RANKL/OPG expression in SCI rats was significantly increased when treating with 10-8 M SP. SP plays a very important role in the pathogenesis of OP after SCI. The direct effect of SP may lead to increased bone resorption through the RANKL/OPG axis after SCI. In addition, high expression of SP also results in the suppression of osteogenesis in SCI rats. Then, the balance between bone resorption and bone formation was broken and finally osteoporosis occurred.

KIR2DL4 differentially signals downstream functions in human NK cells through distinct structural modules.

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Miah, S M Shahjahan; Hughes, Tracey L; Campbell, Kerry S

2008-03-01

KIR2DL4 (2DL4) is a member of the killer cell Ig-like receptor (KIR) family in human NK cells. It can stimulate potent cytokine production and weak cytolytic activity in resting NK cells, but the mechanism for 2DL4-mediated signaling remains unclear. In this study we characterized the signaling pathways stimulated by 2DL4 engagement. In a human NK-like cell line, KHYG-1, cross-linking of 2DL4 activated MAPKs including JNK, ERK, and p38. Furthermore, 2DL4 cross-linking resulted in phosphorylation of IkappaB kinase beta (IKKbeta) and the phosphorylation and degradation of IkappaBalpha, which indicate activation of the classical NF-kappaB pathway. Engagement of 2DL4 was also shown to activate the transcription and translation of a variety of cytokine genes, including TNF-alpha, IFN-gamma, MIP1alpha, MIP1beta, and IL-8. Pharmacological inhibitors of JNK, MEK1/2 and p38, blocked IFN-gamma, IL-8, and MIP1alpha production, suggesting that MAPKs are regulating 2DL4-mediated cytokine production in a nonredundant manner. Activation of both p38 and ERK appear to be upstream of the stimulation of NF-kappaB. Mutation of a transmembrane arginine in 2DL4 to glycine (R/G mutant) abrogated FcepsilonRI-gamma association, as well as receptor-mediated cytolytic activity and calcium responses. Surprisingly, the R/G mutant still activated MAPKs and the NF-kappaB pathway and selectively stimulated the production of MIP1alpha, but not that of IFN-gamma or IL-8. In conclusion, we provide evidence that the activating functions of 2DL4 can be compartmentalized into two distinct structural modules: 1) through transmembrane association with FcepsilonRI-gamma; and 2) through another receptor domain independent of the transmembrane arginine.

Safety evaluation of aprepitant for the prevention of chemotherapy-induced nausea and vomiting

DEFF Research Database (Denmark)

Ruhlmann, Christina H; Herrstedt, Jørn

2011-01-01

INTRODUCTION: Aprepitant is the only neurokinin (NK(1)) receptor antagonist (RA) approved for prevention of chemotherapy-induced nausea and vomiting (CINV). Aprepitant is co-administered with a 5-HT(3) RA and a corticosteroid. Although aprepitant is safe, in most clinical settings potential drug......A4, MK-0869, neurokinin(1) receptor antagonist, safety and tolerability. EXPERT OPINION: The recommended antiemetic regimen of aprepitant, a 5-HT(3) RA and a corticosteroid is safe. The combination of aprepitant, a 5-HT(3) RA and dexamethasone is now the gold standard of antiemetic treatment...... in prevention of CINV induced by HEC, or by the combination of an anthracycline and cyclophosphamide. The intravenous formulation of aprepitant used as a single dose is expected to be of benefit to cancer patients....

Cocaine Disrupts Histamine H3 Receptor Modulation of Dopamine D1 Receptor Signaling: σ1-D1-H3 Receptor Complexes as Key Targets for Reducing Cocaine's Effects

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Moreno, Estefanía; Moreno-Delgado, David; Navarro, Gemma; Hoffmann, Hanne M.; Fuentes, Silvia; Rosell-Vilar, Santi; Gasperini, Paola; Rodríguez-Ruiz, Mar; Medrano, Mireia; Mallol, Josefa; Cortés, Antoni; Casadó, Vicent; Lluís, Carme; Ferré, Sergi; Ortiz, Jordi; Canela, Enric

2014-01-01

The general effects of cocaine are not well understood at the molecular level. What is known is that the dopamine D1 receptor plays an important role. Here we show that a key mechanism may be cocaine's blockade of the histamine H3 receptor-mediated inhibition of D1 receptor function. This blockade requires the σ1 receptor and occurs upon cocaine binding to σ1-D1-H3 receptor complexes. The cocaine-mediated disruption leaves an uninhibited D1 receptor that activates Gs, freely recruits β-arrestin, increases p-ERK 1/2 levels, and induces cell death when over activated. Using in vitro assays with transfected cells and in ex vivo experiments using both rats acutely treated or self-administered with cocaine along with mice depleted of σ1 receptor, we show that blockade of σ1 receptor by an antagonist restores the protective H3 receptor-mediated brake on D1 receptor signaling and prevents the cell death from elevated D1 receptor signaling. These findings suggest that a combination therapy of σ1R antagonists with H3 receptor agonists could serve to reduce some effects of cocaine. PMID:24599455

Ly49Q, an ITIM-bearing NK receptor, positively regulates osteoclast differentiation

International Nuclear Information System (INIS)

Hayashi, Mikihito; Nakashima, Tomoki; Kodama, Tatsuhiko; Makrigiannis, Andrew P.; Toyama-Sorimachi, Noriko; Takayanagi, Hiroshi

2010-01-01

Osteoclasts, multinucleated cells that resorb bone, play a key role in bone remodeling. Although immunoreceptor tyrosine-based activation motif (ITAM)-mediated signaling is critical for osteoclast differentiation, the significance of immunoreceptor tyrosine-based inhibitory motif (ITIM) has not been well understood. Here we report the function of Ly49Q, an Ly49 family member possessing an ITIM motif, in osteoclastogenesis. Ly49Q is selectively induced by receptor activator of nuclear factor-κB (NF-κB) ligand (RANKL) stimulation in bone marrow-derived monocyte/macrophage precursor cells (BMMs) among the Ly49 family of NK receptors. The knockdown of Ly49Q resulted in a significant reduction in the RANKL-induced formation of tartrate-resistance acid phosphatase (TRAP)-positive multinucleated cells, accompanied by a decreased expression of osteoclast-specific genes such as Nfatc1, Tm7sf4, Oscar, Ctsk, and Acp5. Osteoclastogenesis was also significantly impaired in Ly49Q-deficient cells in vitro. The inhibitory effect of Ly49Q-deficiency may be explained by the finding that Ly49Q competed for the association of Src-homology domain-2 phosphatase-1 (SHP-1) with paired immunoglobulin-like receptor-B (PIR-B), an ITIM-bearing receptor which negatively regulates osteoclast differentiation. Unexpectedly, Ly49Q deficiency did not lead to impaired osteoclast formation in vivo, suggesting the existence of a compensatory mechanism. This study provides an example in which an ITIM-bearing receptor functions as a positive regulator of osteoclast differentiation.

The receptor-like kinase SOBIR1 interacts with Brassica napus LepR3 and is required for Leptosphaeria maculans AvrLm1-triggered immunity

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Lisong eMa

2015-10-01

Full Text Available AbstractThe fungus Leptosphaeria maculans (L. maculans is the causal agent of blackleg disease of canola/oilseed rape (Brassica napus worldwide. We previously reported cloning of the B. napus blackleg resistance gene, LepR3, which encodes a receptor-like protein. LepR3 triggers localised cell death upon recognition of its cognate Avr protein, AvrLm1. Here, we exploited the Nicotiana benthamiana model plant to investigate the recognition mechanism of AvrLm1 by LepR3. Co-expression of the LepR3/AvrLm1 gene pair in N. benthamiana resulted in development of a hypersensitive response (HR. However, a truncated AvrLm1 lacking its indigenous signal peptide was compromised in its ability to induce LepR3-mediated HR, indicating that AvrLm1 is perceived by LepR3 extracellularly. Structure-function analysis of the AvrLm1 protein revealed that the C-terminal region of AvrLm1 was required for LepR3-mediated HR in N. benthamiana and for resistance to L. maculans in B. napus. LepR3 was shown to be physically interacting with the B. napus receptor like kinase, SOBIR1 (BnSOBIR1. Silencing of NbSOBIR1 or NbSERK3 (BAK1 compromised LepR3-AvrLm1-dependent HR in N. benthamiana, suggesting that LepR3-mediated resistance to L. maculans in B. napus requires SOBIR1 and BAK1/SERK3. Using this model system, we determined that BnSOBIR1 and SERK3/BAK1 are essential partners in the LepR3 signalling complex and were able to define the AvrLm1 effector domain.

The role of substance P in the maintenance of colonic hypermotility induced by repeated stress in rats.

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Lu, Ping; Luo, Hesheng; Quan, Xiaojing; Fan, Han; Tang, Qincai; Yu, Guang; Chen, Wei; Xia, Hong

2016-04-01

The mechanism underlying chronic stress-induced gastrointestinal (GI) dysmotility has not been fully elucidated and GI hormones have been indicated playing a role in mediating stress-induced changes in GI motor function. Our objective was to study the possible role of substance P (SP) in the colonic hypermotility induced by repeated water avoidance stress (WAS) which mimics irritable bowel syndrome. Male Wistar rats were submitted to WAS or sham WAS (SWAS) (1h/day) for up to 10 consecutive days. Enzyme Immunoassay Kit was used to detect the serum level of SP. The expression of neurokinin-1 receptor (NK1R) was investigated by Immunohistochemistry and Western blotting. The spontaneous contraction of muscle strip was studied in an organ bath system. L-type calcium channel currents (ICa,L) of smooth muscle cells (SMCs) were recorded by whole-cell patch-clamp technique. Fecal pellet expulsion and spontaneous contraction of proximal colon in rats were increased after repeated WAS. The serum level of SP was elevated following WAS. Immunohistochemistry proved the expression of NK1R in mucosa, muscularis and myenteric plexus. Western blotting demonstrated stress-induced up-regulation of NK1R in colon devoid of mucosa and submucosa. Repeated WAS increased the contractile activities of longitudinal muscle and circular muscle strips induced by SP and this effect was reversed by a selective NK1R antagonist. The ICa,L of SMCs in the WAS rats were drastically increased compared to controls after addition of SP. Increased serum SP level and up-regulated NK1R in colon may contribute to stress-induced colonic hypermotility. And L-type calcium channels play a potentially important role in the process of WAS-induced dysmotility. Copyright © 2016 Elsevier Ltd. All rights reserved.

Comprehensive behavioral analysis of Ox1r-/- mice showed implication of orexin receptor-1 in mood, anxiety and social behavior

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Md Golam Abbas

2015-12-01

Full Text Available Neuropeptides orexin A and orexin B, which are exclusively produced by neurons in the lateral hypothalamic area, play an important role in the regulation of a wide range of behaviors and homeostatic processes, including regulation of sleep/wakefulness states and energy homeostasis. The orexin system has close anatomical and functional relationships with systems that regulate the autonomic nervous system, emotion, mood, the reward system and sleep/wakefulness states. Recent pharmacological studies using selective antagonists have suggested that orexin receptor-1 (OX1R is involved in physiological processes that regulate emotion, the reward system and autonomic nervous system. Here, we examined Ox1r-/- mice with a comprehensive behavioral test battery to screen additional OX1R functions. Ox1r-/- mice showed increased anxiety-like behavior, altered depression-like behavior, slightly decreased spontaneous locomotor activity, reduced social interaction, increased startle response and decreased prepulse inhibition. These results suggest that OX1R plays roles in social behaviour and sensory motor gating in addition to roles in mood and anxiety.

Changes in the Ratio of Tc1/Tc2 and Th1/Th2 Cells but Not in Subtypes of NK-Cells in Preeclampsia

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Yayi Hou

2007-06-01

Full Text Available It has been suggested that natural killer (NK cell activity and Th1 immunitymay be involved in the pathogenesis of preeclampsia. This study aimed to investigate theimmunophenotypes of NK cells and type 1/type 2 immunity in both decidua and maternalperipheral blood between normal (n=11 and preeclamptic pregnant women (n=20 by flowcytometry. The results showed that no significant difference was observed between patientsand controls by detecting CD56+ CD69+ and CD56+ CD94+ NK cells in both peripheralblood and decidua. Moreover, in preeclamptic patients, decreased percentages of Tc2 andTh2 cells and the increased ratios of Tc1/Tc2 were determined in both decidua andmaternal peripheral blood. In addition, the ratio of Th1/Th2 in peripheral blood alsoincreased. There was no significant difference of immunophenotypes of uNK cells betweenpreeclampsia and normal pregnancy. Local decidua and systematic immunity did notcorrelate with each other. These results suggest that the type 1/type 2 immunity shifted totype 1 immunity including Th1 and Tc1 cells may contribute to the patho-genesis ofpreeclampsia.

Region-specific proteolysis differentially regulates type 1 inositol 1,4,5-trisphosphate receptor activity.

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Wang, Liwei; Wagner, Larry E; Alzayady, Kamil J; Yule, David I

2017-07-14

The inositol 1,4,5 trisphosphate receptor (IP 3 R) is an intracellular Ca 2+ release channel expressed predominately on the membranes of the endoplasmic reticulum. IP 3 R1 can be cleaved by caspase or calpain into at least two receptor fragments. However, the functional consequences of receptor fragmentation are poorly understood. Our previous work has demonstrated that IP 3 R1 channels, formed following either enzymatic fragmentation or expression of the corresponding complementary polypeptide chains, retain tetrameric architecture and are still activated by IP 3 binding despite the loss of peptide continuity. In this study, we demonstrate that region-specific receptor fragmentation modifies channel regulation. Specifically, the agonist-evoked temporal Ca 2+ release profile and protein kinase A modulation of Ca 2+ release are markedly altered. Moreover, we also demonstrate that activation of fragmented IP 3 R1 can result in a distinct functional outcome. Our work suggests that proteolysis of IP 3 R1 may represent a novel form of modulation of IP 3 R1 channel function and increases the repertoire of Ca 2+ signals achievable through this channel. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

The mechanisms behind decreased internalization of angiotensin II type 1 receptor.

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Bian, Jingwei; Zhang, Suli; Yi, Ming; Yue, Mingming; Liu, Huirong

2018-04-01

The internalization of angiotensin II type 1 receptor (AT 1 R) plays an important role in maintaining cardiovascular homeostasis. Decreased receptor internalization is closely related to cardiovascular diseases induced by the abnormal activation of AT 1 R, such as hypertension. However, the mechanism behind reduced AT 1 R internalization is not fully understood. This review focuses on four parts of the receptor internalization process (the combination of agonists and receptors, receptor phosphorylation, endocytosis, and recycling) and summarizes the possible mechanisms by which AT 1 R internalization is reduced based on these four parts of the process. (1) The agonist has a large molecular weight or a stronger ability to hydrolyze phosphatidylinositol 4,5-bisphosphate (PtdIns (4,5) P 2 ), which can increase the consumption of PtdIns (4,5) P 2 . (2) AT 1 R phosphorylation is weakened because of an abnormal function of phosphorylated kinase or changes in phospho-barcoding and GPCR-β-arrestin complex conformation. (3) The abnormal formation of vesicles or AT 1 R heterodimers with fewer endocytic receptors results in less AT 1 R endocytosis. (4) The enhanced activity and upregulated expression of small GTP-binding protein 4 (Rab4) and 11 (Rab11), which regulate receptor recycling, and phosphatidylinositol 3-kinase increase AT 1 R recycling. In addition, lower expression of AT 1 R-associated protein (ATRAP) or higher expression of AT 1 R-associated protein 1 (ARAP1) can reduce receptor internalization. Copyright © 2018 Elsevier Inc. All rights reserved.

Metabotropic glutamate receptor 1 splice variants mGluR1a and mGluR1b combine in mGluR1a/b dimers in vivo

Czech Academy of Sciences Publication Activity Database

Techlovská, Šárka; Chambers, Jayne Nicole; Dvořáková, Michaela; Petralia, R.S.; Wang, Y.X.; Hájková, Alena; Franková, Daniela; Prezeau, L.; Blahoš, Jaroslav

2014-01-01

Roč. 86, November (2014), s. 329-326 ISSN 0028-3908 R&D Projects: GA ČR GAP303/12/2408 Institutional support: RVO:68378050 Keywords : Glutamate receptors * GPCR * alternative splicing Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.106, year: 2014

The biology of NK cells and their receptors affects clinical outcomes after hematopoietic cell transplantation (HCT).

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Foley, Bree; Felices, Martin; Cichocki, Frank; Cooley, Sarah; Verneris, Michael R; Miller, Jeffrey S

2014-03-01

Natural killer (NK) cells were first identified for their capacity to reject bone marrow allografts in lethally irradiated mice without prior sensitization. Subsequently, human NK cells were detected and defined by their non-major histocompatibility complex (MHC)-restricted cytotoxicity toward transformed or virally infected target cells. Karre et al. later proposed 'the missing self hypothesis' to explain the mechanism by which self-tolerant cells could kill targets that had lost self MHC class I. Subsequently, the receptors that recognize MHC class I to mediate tolerance in the host were identified on NK cells. These class I-recognizing receptors contribute to the acquisition of function by a dynamic process known as NK cell education or licensing. In the past, NK cells were assumed to be short lived, but more recently NK cells have been shown to mediate immunologic memory to secondary exposures to cytomegalovirus infection. Because of their ability to lyse tumors with aberrant MHC class I expression and to produce cytokines and chemokines upon activation, NK cells may be primed by many stimuli, including viruses and inflammation, to contribute to a graft-versus-tumor effect. In addition, interactions with other immune cells support the therapeutic potential of NK cells to eradicate tumor and to enhance outcomes after hematopoietic cell transplantation. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

AMP is an adenosine A1 receptor agonist.

Science.gov (United States)

Rittiner, Joseph E; Korboukh, Ilia; Hull-Ryde, Emily A; Jin, Jian; Janzen, William P; Frye, Stephen V; Zylka, Mark J

2012-02-17

Numerous receptors for ATP, ADP, and adenosine exist; however, it is currently unknown whether a receptor for the related nucleotide adenosine 5'-monophosphate (AMP) exists. Using a novel cell-based assay to visualize adenosine receptor activation in real time, we found that AMP and a non-hydrolyzable AMP analog (deoxyadenosine 5'-monophosphonate, ACP) directly activated the adenosine A(1) receptor (A(1)R). In contrast, AMP only activated the adenosine A(2B) receptor (A(2B)R) after hydrolysis to adenosine by ecto-5'-nucleotidase (NT5E, CD73) or prostatic acid phosphatase (PAP, ACPP). Adenosine and AMP were equipotent human A(1)R agonists in our real-time assay and in a cAMP accumulation assay. ACP also depressed cAMP levels in mouse cortical neurons through activation of endogenous A(1)R. Non-selective purinergic receptor antagonists (pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid and suramin) did not block adenosine- or AMP-evoked activation. Moreover, mutation of His-251 in the human A(1)R ligand binding pocket reduced AMP potency without affecting adenosine potency. In contrast, mutation of a different binding pocket residue (His-278) eliminated responses to AMP and to adenosine. Taken together, our study indicates that the physiologically relevant nucleotide AMP is a full agonist of A(1)R. In addition, our study suggests that some of the physiological effects of AMP may be direct, and not indirect through ectonucleotidases that hydrolyze this nucleotide to adenosine.

Soluble intercellular adhesion molecule-1 (sICAM-1) and soluble interleukin-2 receptors (sIL-2R) in scleroderma skin

DEFF Research Database (Denmark)

Søndergaard, Klaus; Deleuran, Mette; Heickendorff, Lene

1998-01-01

In order to investigate whether soluble intercellular adhesion molecule-1 (sICAM-1) and soluble interleukin-2 receptors (sIL-2R) were present in scleroderma skin, and to compare their levels to concentrations measured in plasma and clinical parameters, we examined suction blister fluid and plasma...... from 13 patients with systemic sclerosis and 11 healthy volunteers. Suction blisters and biopsies were from the transition zone between normal skin and scleroderma, and uninvolved abdominal skin. The levels of sICAM-1 and sIL-2R were significantly increased in both plasma and suction blister fluid from...

Calcium-Dependent Energetics of Calmodulin Domain Interactions with Regulatory Regions of the Ryanodine Receptor Type 1 (RyR1)

Science.gov (United States)

Newman, Rhonda A.; Sorensen, Brenda R.; Kilpatrick, Adina M.; Shea, Madeline A.

2014-01-01

Calmodulin (CaM) plays a vital role in calcium homeostasis by allosterically modulating intracellular calcium channels including the homo-tetrameric human Ryanodine Receptor Type 1 (hRyR1). Apo (calcium-free) CaM activates hRyR1 while calcium-saturated CaM inhibits it. Two CaM-binding regions (residues 1975–1999 and 3614–3643) identified in each RyR1 monomer were proposed to allow CaM to bridge adjacent RyR1 subunits. We explored the distinct roles of CaM domains by using fluorescence anisotropy to determine the affinity of CaM1–148 (full-length), CaM1–80 (N-domain) and CaM76–148 (C-domain) for peptides encompassing hRyR1 residues 1975–1999 or 3614–3643. Both CaM1–148 and CaM76–148 associated in a calcium-independent manner with similar affinities for hRyR1(3614–3643)p while CaM1–80 required calcium and bound ~250-fold more weakly. Association of CaM1–148, CaM1–80 and CaM76–148 with hRyR1(1975–1999)p was much less favorable than with hRyR1(3614–3643)p; differences between the two CaM domains were smaller. Equilibrium calcium titrations monitored by steady-state fluorescence demonstrated that both hRyR1 peptides increased the calcium-binding affinity of both CaM domains. These thermodynamic properties support a prior model in which the CaM C-domain associates with RyR1(3614–3643) at low levels of calcium, positioning CaM to rapidly respond to calcium efflux. However, the affinity of the N-domain of CaM for hRyR1(1975–1999)p is insufficient to explain a model in which CaM bridges adjacent RyR1 subunits within the tetramer. This indicates that other protein factors or properties of the tertiary or quaternary structure of hRyR1 contribute to the energetics of CaM-mediated regulation. PMID:25145833

Glutamate Receptors GluR1 and GluR4 in the Hamster Superior Colliculus: Distribution and Co-localization with Calcium-Binding Proteins and GABA

International Nuclear Information System (INIS)

Choi, Jae-Sik; Lee, Jea-Young; Jeon, Chang-Jin

2009-01-01

We investigated the distributions of AMPA glutamate receptor subtypes GluR1 and GluR4 in the hamster superior colliculus (SC) with antibody immunocytochemistry and the effect of enucleation on these distributions. We compared these labelings to those of GluR2/3 in our previous report (Park et al., 2004, Neurosci Res., 49:139–155) and calcium-binding proteins calbindin D28K, calretinin, parvalbumin, and GABA. Anti-GluR1-immunoreactive (IR) cells were scattered throughout the SC. By contrast, anti-GluR4-IR cells formed distinct clusters within the lower lateral stratum griseum intermediale (SGI) and lateral stratum album intermediale (SAI). The GluR1- and GluR4-IR neurons varied in size and morphology. The average diameter of the GluR1-IR cells was 13.00 µm, while the GluR4-IR cells was 20.00 µm. The large majority of IR neurons were round or oval cells, but they also included stellate, vertical fusiform and horizontal cells. Monocular enucleation appeared to have no effect on the GluR1 and GluR4 immunoreactivity. Some GluR1-IR cells expressed calbindin D28K (9.50%), calretinin (6.59%), parvalbumin (2.53%), and GABA (20.54%). By contrast, no GluR4-IR cells expressed calcium-binding proteins or GABA. Although the function of the AMPA receptor subunits in SC is not yet clear, the distinct segregation of the GluR subunits, its differential colocalization with calcium-binding proteins and GABA, and differential responses to enucleation suggest the functional diversity of the receptor subunits in visuo-motor integration in the SC

Human GH Receptor-IGF-1 Receptor Interaction: Implications for GH Signaling

Science.gov (United States)

Gan, Yujun; Buckels, Ashiya; Liu, Ying; Zhang, Yue; Paterson, Andrew J.; Jiang, Jing; Zinn, Kurt R.

2014-01-01

GH signaling yields multiple anabolic and metabolic effects. GH binds the transmembrane GH receptor (GHR) to activate the intracellular GHR-associated tyrosine kinase, Janus kinase 2 (JAK2), and downstream signals, including signal transducer and activator of transcription 5 (STAT5) activation and IGF-1 gene expression. Some GH effects are partly mediated by GH-induced IGF-1 via IGF-1 receptor (IGF-1R), a tyrosine kinase receptor. We previously demonstrated in non-human cells that GH causes formation of a GHR-JAK2-IGF-1R complex and that presence of IGF-1R (even without IGF-1 binding) augments proximal GH signaling. In this study, we use human LNCaP prostate cancer cells as a model system to further study the IGF-1R's role in GH signaling. GH promoted JAK2 and GHR tyrosine phosphorylation and STAT5 activation in LNCaP cells. By coimmunoprecipitation and a new split luciferase complementation assay, we find that GH augments GHR/IGF-1R complex formation, which is inhibited by a Fab of an antagonistic anti-GHR monoclonal antibody. Short hairpin RNA-mediated IGF-1R silencing in LNCaP cells reduced GH-induced GHR, JAK2, and STAT5 phosphorylation. Similarly, a soluble IGF-1R extracellular domain fragment (sol IGF-1R) interacts with GHR in response to GH and blunts GH signaling. Sol IGF-1R also markedly inhibits GH-induced IGF-1 gene expression in both LNCaP cells and mouse primary osteoblast cells. On the basis of these and other findings, we propose a model in which IGF-1R augments GH signaling by allowing a putative IGF-1R-associated molecule that regulates GH signaling to access the activated GHR/JAK2 complex and envision sol IGF-1R as a dominant-negative inhibitor of this IGF-1R-mediated augmentation. Physiological implications of this new model are discussed. PMID:25211187

HIV exposed seronegative (HESN compared to HIV infected individuals have higher frequencies of telomeric Killer Immunoglobulin-like Receptor (KIR B motifs; Contribution of KIR B motif encoded genes to NK cell responsiveness.

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Elise Jackson