Characterization of Transient Receptor Potential Vanilloid-1 (TRPV1) Variant Activation by Coal Fly Ash Particles and Associations with Altered Transient Receptor Potential Ankyrin-1 (TRPA1) Expression and Asthma*

PubMed Central

Stockmann, Chris; Romero, Erin G.; Lu, Zhenyu; Shapiro, Darien; Stone, Bryan L.; Fassl, Bernhard; Nkoy, Flory; Uchida, Derek A.; Ward, Robert M.; Veranth, John M.; Reilly, Christopher A.

2016-01-01

Transient receptor potential (TRP) channels are activated by environmental particulate materials. We hypothesized that polymorphic variants of transient receptor potential vanilloid-1 (TRPV1) would be uniquely responsive to insoluble coal fly ash compared with the prototypical soluble agonist capsaicin. Furthermore, these changes would manifest as differences in lung cell responses to these agonists and perhaps correlate with changes in asthma symptom control. The TRPV1-I315M and -T469I variants were more responsive to capsaicin and coal fly ash. The I585V variant was less responsive to coal fly ash particles due to reduced translation of protein and an apparent role for Ile-585 in activation by particles. In HEK-293 cells, I585V had an inhibitory effect on wild-type TRPV1 expression, activation, and internalization/agonist-induced desensitization. In normal human bronchial epithelial cells, IL-8 secretion in response to coal fly ash treatment was reduced for cells heterozygous for TRPV1-I585V. Finally, both the I315M and I585V variants were associated with worse asthma symptom control with the effects of I315M manifesting in mild asthma and those of the I585V variant manifesting in severe, steroid-insensitive individuals. This effect may be due in part to increased transient receptor potential ankyrin-1 (TRPA1) expression by lung epithelial cells expressing the TRPV1-I585V variant. These findings suggest that specific molecular interactions control TRPV1 activation by particles, differential activation, and desensitization of TRPV1 by particles and/or other agonists, and cellular changes in the expression of TRPA1 as a result of I585V expression could contribute to variations in asthma symptom control. PMID:27758864

Characterization of Transient Receptor Potential Vanilloid-1 (TRPV1) Variant Activation by Coal Fly Ash Particles and Associations with Altered Transient Receptor Potential Ankyrin-1 (TRPA1) Expression and Asthma.

PubMed

Deering-Rice, Cassandra E; Stockmann, Chris; Romero, Erin G; Lu, Zhenyu; Shapiro, Darien; Stone, Bryan L; Fassl, Bernhard; Nkoy, Flory; Uchida, Derek A; Ward, Robert M; Veranth, John M; Reilly, Christopher A

2016-11-25

Transient receptor potential (TRP) channels are activated by environmental particulate materials. We hypothesized that polymorphic variants of transient receptor potential vanilloid-1 (TRPV1) would be uniquely responsive to insoluble coal fly ash compared with the prototypical soluble agonist capsaicin. Furthermore, these changes would manifest as differences in lung cell responses to these agonists and perhaps correlate with changes in asthma symptom control. The TRPV1-I315M and -T469I variants were more responsive to capsaicin and coal fly ash. The I585V variant was less responsive to coal fly ash particles due to reduced translation of protein and an apparent role for Ile-585 in activation by particles. In HEK-293 cells, I585V had an inhibitory effect on wild-type TRPV1 expression, activation, and internalization/agonist-induced desensitization. In normal human bronchial epithelial cells, IL-8 secretion in response to coal fly ash treatment was reduced for cells heterozygous for TRPV1-I585V. Finally, both the I315M and I585V variants were associated with worse asthma symptom control with the effects of I315M manifesting in mild asthma and those of the I585V variant manifesting in severe, steroid-insensitive individuals. This effect may be due in part to increased transient receptor potential ankyrin-1 (TRPA1) expression by lung epithelial cells expressing the TRPV1-I585V variant. These findings suggest that specific molecular interactions control TRPV1 activation by particles, differential activation, and desensitization of TRPV1 by particles and/or other agonists, and cellular changes in the expression of TRPA1 as a result of I585V expression could contribute to variations in asthma symptom control. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Gab-family adapter molecules in signal transduction of cytokine and growth factor receptors, and T and B cell antigen receptors.

PubMed

Hibi, M; Hirano, T

2000-04-01

Gab1 and Gab2 (Grb2 associated binder 1 and 2) are scaffolding adapter molecules that display sequence similarity with Drosophila DOS (daughter of sevenless), which is a potential substrate for the protein tyrosine phosphatase, Corkscrew, Both Gab1 and Gab2, like DOS, have a pleckstrin homology domain and potential binding sites for SH2 and SH3 domains. Gab1 and Gab2 are phosphorylated on tyrosine upon the stimulation of various cytokines, growth factors, and antigen receptors, and interact with signaling molecules, such as Grb2, SHP-2, and PI-3 kinase. Overexpression of Gab1 or Gab2 mimics or enhances growth factor or cytokine-mediated biological processes and activates ERK MAP kinase. These data imply that Gab1 and Gab2 act downstream of a broad range of cytokine and growth factor receptors, as well as T and B antigen receptors, and link these receptors to ERK MAP kinase and biological actions.

Self-renewal of human embryonic stem cells requires insulin-like growth factor-1 receptor and ERBB2 receptor signaling

PubMed Central

Wang, Linlin; Schulz, Thomas C.; Sherrer, Eric S.; Dauphin, Derek S.; Shin, Soojung; Nelson, Angelique M.; Ware, Carol B.; Zhan, Mei; Song, Chao-Zhong; Chen, Xiaoji; Brimble, Sandii N.; McLean, Amanda; Galeano, Maria J.; Uhl, Elizabeth W.; D'Amour, Kevin A.; Chesnut, Jonathan D.; Rao, Mahendra S.

2007-01-01

Despite progress in developing defined conditions for human embryonic stem cell (hESC) cultures, little is known about the cell-surface receptors that are activated under conditions supportive of hESC self-renewal. A simultaneous interrogation of 42 receptor tyrosine kinases (RTKs) in hESCs following stimulation with mouse embryonic fibroblast (MEF) conditioned medium (CM) revealed rapid and prominent tyrosine phosphorylation of insulin receptor (IR) and insulin-like growth factor-1 receptor (IGF1R); less prominent tyrosine phosphorylation of epidermal growth factor receptor (EGFR) family members, including ERBB2 and ERBB3; and trace phosphorylation of fibroblast growth factor receptors. Intense IGF1R and IR phosphorylation occurred in the absence of MEF conditioning (NCM) and was attributable to high concentrations of insulin in the proprietary KnockOut Serum Replacer (KSR). Inhibition of IGF1R using a blocking antibody or lentivirus-delivered shRNA reduced hESC self-renewal and promoted differentiation, while disruption of ERBB2 signaling with the selective inhibitor AG825 severely inhibited hESC proliferation and promoted apoptosis. A simple defined medium containing an IGF1 analog, heregulin-1β (a ligand for ERBB2/ERBB3), fibroblast growth factor-2 (FGF2), and activin A supported long-term growth of multiple hESC lines. These studies identify previously unappreciated RTKs that support hESC proliferation and self-renewal, and provide a rationally designed medium for the growth and maintenance of pluripotent hESCs. PMID:17761519

Transient receptor potential ankyrin receptor 1 is a novel target for pro-tussive agents.

PubMed

Andrè, E; Gatti, R; Trevisani, M; Preti, D; Baraldi, P G; Patacchini, R; Geppetti, P

2009-11-01

The transient receptor potential ankyrin receptor 1 (TRPA1) is a cation channel, co-expressed with the pro-tussive transient receptor potential vanilloid type 1 (TRPV1) channel in primary sensory neurons. TRPA1 is activated by a series of irritant exogenous and endogenous alpha,beta-unsaturated aldehydes which seem to play a role in airway diseases. We investigated whether TRPA1 agonists provoke cough in guinea pigs and whether TRPA1 antagonists inhibit this response. Animals were placed in a Perspex box, and cough sounds were recorded and counted by observers unaware of the treatment used. Inhalation of two selective TRPA1 agonists, allyl isothiocyanate and cinnamaldehyde, dose-dependently caused cough in control guinea pigs, but not in those with airway sensory nerves desensitized by capsaicin. Coughs elicited by TRPA1 agonists were reduced by non-selective (camphor and gentamicin) and selective (HC-030031) TRPA1 antagonists, whereas they were unaffected by the TRPV1 antagonist, capsazepine. Acrolein and crotonaldehyde, two alpha,beta-unsaturated aldehydes recently identified as TRPA1 stimulants and contained in cigarette smoke, air pollution or produced endogenously by oxidative stress, caused a remarkable tussive effect, a response that was selectively inhibited by HC-030031. Part of the cough response induced by cigarette smoke inhalation was inhibited by HC-030031, suggesting the involvement of TRPA1. A novel pro-tussive pathway involves the TRPA1 channel, expressed by capsaicin-sensitive airway sensory nerves and is activated by a series of exogenous (cigarette smoke) and endogenous irritants. These results suggest TRPA1 may be a novel target for anti-tussive medicines.

The transient receptor potential ankyrin-1 mediates mechanical hyperalgesia induced by the activation of B1 receptor in mice.

PubMed

Meotti, Flavia Carla; Figueiredo, Cláudia Pinto; Manjavachi, Marianne; Calixto, João B

2017-02-01

The kinin receptor B 1 and the transient receptor potential ankyrin 1 (TRPA1) work as initiators and gatekeepers of nociception and inflammation. This study reports that the nociceptive transmission induced by activation of B 1 receptor is dependent on TRPA1 ion channel. The mechanical hyperalgesia was induced by intrathecal (i.t.) injection of B 1 agonist des-Arginine 9 -bradykinin (DABK) or TRPA1 agonist cinnamaldehyde and was evaluated by the withdrawal response after von Frey Hair application in the hind paw. After behavioral experiments, lumbar spinal cord and dorsal root ganglia (DRG) were harvested to assess protein expression and mRNA by immunohistochemistry and real time-PCR, respectively. The pharmacological antagonism (HC030031) or the down-regulation of TRPA1 greatly inhibited the mechanical hyperalgesia induced by DABK. Intrathecal injection of DABK up regulated the ionized calcium binding adaptor molecule (Iba-1) in lumbar spinal cord (L5-L6); TRPA1 protein and mRNA in lumbar spinal cord; and B 1 receptor mRNA in both lumbar spinal cord and DRG. The knockdown of TRPA1 prevented microglia activation induced by DABK. Furthermore, the mechanical hyperalgesia induced by either DABK or by cinnamaldehyde was significantly reduced by inhibition of cyclooxygenase (COX), protein kinase C (PKC) or phospholipase C (PLC). In summary, this study revealed that TRPA1 positively modulates the mechanical hyperalgesia induced by B 1 receptor activation in the spinal cord and that the classical GPCR downstream molecules PLC, diacylglycerol (DAG), 3,4,5-inositide phosphate (IP 3 ) and PKC are involved in the nociceptive transmission triggered by these two receptors. Copyright © 2016 Elsevier Inc. All rights reserved.

Vascular Repair After Menstruation Involves Regulation of Vascular Endothelial Growth Factor-Receptor Phosphorylation by sFLT-1

PubMed Central

Graubert, Michael D.; Asuncion Ortega, Maria; Kessel, Bruce; Mortola, Joseph F.; Iruela-Arispe, M. Luisa

2001-01-01

Regeneration of the endometrium after menstruation requires a rapid and highly organized vascular response. Potential regulators of this process include members of the vascular endothelial growth factor (VEGF) family of proteins and their receptors. Although VEGF expression has been detected in the endometrium, the relationship between VEGF production, receptor activation, and endothelial cell proliferation during the endometrial cycle is poorly understood. To better ascertain the relevance of VEGF family members during postmenstrual repair, we have evaluated ligands, receptors, and activity by receptor phosphorylation in human endometrium throughout the menstrual cycle. We found that VEGF is significantly increased at the onset of menstruation, a result of the additive effects of hypoxia, transforming growth factor-α, and interleukin-1β. Both VEGF receptors, FLT-1 and KDR, followed a similar pattern. However, functional activity of KDR, as determined by phosphorylation studies, revealed activation in the late menstrual and early proliferative phases. The degree of KDR phosphorylation was inversely correlated with the presence of sFLT-1. Endothelial cell proliferation analysis in endometrium showed a peak during the late menstrual and early proliferative phases in concert with the presence of VEGF, VEGF receptor phosphorylation, and decrease of sFLT-1. Together, these results suggest that VEGF receptor activation and the subsequent modulation of sFLT-1 in the late menstrual phase likely contributes to the onset of angiogenesis and endothelial repair in the human endometrium. PMID:11290558

TRPV1 recapitulates native capsaicin receptor in sensory neurons in association with Fas-associated factor 1.

PubMed

Kim, Sangsung; Kang, Changjoong; Shin, Chan Young; Hwang, Sun Wook; Yang, Young Duk; Shim, Won Sik; Park, Min-Young; Kim, Eunhee; Kim, Misook; Kim, Byung-Moon; Cho, Hawon; Shin, Youngki; Oh, Uhtaek

2006-03-01

TRPV1, a cloned capsaicin receptor, is a molecular sensor for detecting adverse stimuli and a key element for inflammatory nociception and represents biophysical properties of native channel. However, there seems to be a marked difference between TRPV1 and native capsaicin receptors in the pharmacological response profiles to vanilloids or acid. One plausible explanation for this overt discrepancy is the presence of regulatory proteins associated with TRPV1. Here, we identify Fas-associated factor 1 (FAF1) as a regulatory factor, which is coexpressed with and binds to TRPV1 in sensory neurons. When expressed heterologously, FAF1 reduces the responses of TRPV1 to capsaicin, acid, and heat, to the pharmacological level of native capsaicin receptor in sensory neurons. Furthermore, silencing FAF1 by RNA interference augments capsaicin-sensitive current in native sensory neurons. We therefore conclude that FAF1 forms an integral component of the vanilloid receptor complex and that it constitutively modulates the sensitivity of TRPV1 to various noxious stimuli in sensory neurons.

Isothiocyanates from Wasabia japonica activate transient receptor potential ankyrin 1 channel.

PubMed

Uchida, Kunitoshi; Miura, Yosuke; Nagai, Masashi; Tominaga, Makoto

2012-11-01

6-(Methylsulfinyl)hexyl isothiocyanate (6-MSITC) and 6-(methylthio)hexyl isothiocyanate (6-MTITC) have low pungency and are responsible for the fresh flavor of wasabi (Wasabia japonica [Miq] Matsumura). In this study, we found that these two isothiocyanates activate transient receptor potential ankyrin 1 (TRPA1), and 6-MSITC activates transient receptor potential vanilloid 1 (TRPV1), but not other transient receptor potential channels expressed in sensory neurons. Both 6-MSITC and 6-MTITCinduced intracellular Ca(2+) increases in human embryonic kidney-derived 293 cells expressing mouse TRPA1 (mTRPA1) as measured by Ca(2+) imaging. In whole-cell patch-clamp recordings, 6-MSITC and 6-MTITC dose-dependently activated both mTRPA1 (EC(50) = 147±26 µM for 6-MSITC and 30±3 µM for 6-MTITC) and human TRPA1 (hTRPA1; EC(50) = 39±4 µM for 6-MSITC and 34±3 µM for 6-MTITC). In addition, TRPA1 N-terminal cysteines, which are reported to be important for channel activation by electrophilic ligands, were involved in 6-MSITC- and 6-MTITC-evoked TRPA1 activation. These isothiocyanates also activated endogenous TRPA1 expressed in mouse dorsal root ganglion neurons and intraplantar injection of 10-30 mM 6-MSITC-evoked pain-related behaviors in mice. These results indicate the following: 1) 6-MSITC and 6-MTITC activate both mTRPA1 and hTRPA1; 2) 6-MSITC activates mTRPV1; and 3) the pharmacological functions of these isothiocyanates could be derived from TRPA1 activation.

Targeting the heparin-binding domain of fibroblast growth factor receptor 1 as a potential cancer therapy.

PubMed

Ling, Ling; Tan, Si Kee; Goh, Ting Hwee; Cheung, Edwin; Nurcombe, Victor; van Wijnen, Andre J; Cool, Simon M

2015-07-23

Aberrant activation of fibroblast growth factor receptors (FGFRs) deregulates cell proliferation and promotes cell survival, and may predispose to tumorigenesis. Therefore, selective inactivation of FGFRs is an important strategy for cancer therapy. Here as a proof-of-concept study, we developed a FGFR1 neutralizing antisera, IMB-R1, employing a novel strategy aimed at preventing the access of essential heparan sulfate (HS) co-receptors to the heparin-binding domain on FGFR1. The mRNA and protein expression level of FGFR1 and other FGFRs were examined in several lines of breast cancer and osteosarcoma cells and corresponding normal cells using Taqman real-time quantitative PCR and Western blot analysis. The specificity of IMB-R1 against FGFR1 was assessed with various ELISA-based approaches and Receptor Tyrosine Kinase array. Proliferation assay and apoptosis analysis were performed to assess the effect of IMB-R1 on cancer cell growth and apoptosis, respectively, in comparison with known FGFR1 inhibitors. The IMB-R1 induced alteration of intracellular signaling and gene expression were analysed using Western blot and microarray approaches. Immunohistochemical staining of FGFR1 using IMB-R1 were carried out in different cancer tissues from clinical patients. Throughout the study, statistical differences were determined by Student's t test where appropriate and reported when a p value was less than 0.05. We demonstrate that IMB-R1 is minimally cross-reactive for other FGFRs, and that it potently and specifically inhibits binding of heparin to FGFR1. Furthermore, IMB-R1 blocks the interaction of FGF2 with FGFR1, the kinase activity of FGFR1 and activation of intracellular FGFR signaling. Cancer cells treated with IMB-R1 displayed impaired FGF2 signaling, were unable to grow and instead underwent apoptosis. IMB-R1-induced cell death correlated with a disruption of antioxidative defense networks and increased expression of several tumor suppressors and apoptotic

Synthesis, structure-activity relationships, and in vivo evaluation of N3-phenylpyrazinones as novel corticotropin-releasing factor-1 (CRF1) receptor antagonists.

PubMed

Hartz, Richard A; Ahuja, Vijay T; Arvanitis, Argyrios G; Rafalski, Maria; Yue, Eddy W; Denhart, Derek J; Schmitz, William D; Ditta, Jonathan L; Deskus, Jeffrey A; Brenner, Allison B; Hobbs, Frank W; Payne, Joseph; Lelas, Snjezana; Li, Yu-Wen; Molski, Thaddeus F; Mattson, Gail K; Peng, Yong; Wong, Harvey; Grace, James E; Lentz, Kimberley A; Qian-Cutrone, Jingfang; Zhuo, Xiaoliang; Shu, Yue-Zhong; Lodge, Nicholas J; Zaczek, Robert; Combs, Andrew P; Olson, Richard E; Bronson, Joanne J; Mattson, Ronald J; Macor, John E

2009-07-23

Evidence suggests that corticotropin-releasing factor-1 (CRF(1)) receptor antagonists may offer therapeutic potential for the treatment of diseases associated with elevated levels of CRF such as anxiety and depression. A pyrazinone-based chemotype of CRF(1) receptor antagonists was discovered. Structure-activity relationship studies led to the identification of numerous potent analogues including 12p, a highly potent and selective CRF(1) receptor antagonist with an IC(50) value of 0.26 nM. The pharmacokinetic properties of 12p were assessed in rats and Cynomolgus monkeys. Compound 12p was efficacious in the defensive withdrawal test (an animal model of anxiety) in rats. The synthesis, structure-activity relationships and in vivo properties of compounds within the pyrazinone chemotype are described.

Sigma-1 receptor chaperone and brain-derived neurotrophic factor: emerging links between cardiovascular disease and depression.

PubMed

Hashimoto, Kenji

2013-01-01

Epidemiological studies have demonstrated a close relationship between depression and cardiovascular disease (CVD). Although it is known that the central nervous system (CNS) contributes to this relationship, the detailed mechanisms involved in this process remain unclear. Recent studies suggest that the endoplasmic reticulum (ER) molecular chaperone sigma-1 receptor and brain-derived neurotrophic factor (BDNF) play a role in the pathophysiology of CVD and depression. Several meta-analysis studies have showed that levels of BDNF in the blood of patients with major depressive disorder (MDD) are lower than normal controls, indicating that blood BDNF might be a biomarker for depression. Furthermore, blood levels of BDNF in patients with CVD are also lower than normal controls. A recent study using conditional BDNF knock-out mice in animal models of myocardial infarction highlighted the role of CNS-mediated mechanisms in the cardioprotective effects of BDNF. In addition, a recent study shows that decreased levels of sigma-1 receptor in the mouse brain contribute to the association between heart failure and depression. Moreover, sigma-1 receptor agonists, including the endogenous neurosteroid dehydroepiandosterone (DHEA) and the selective serotonin reuptake inhibitor (SSRI) fluvoxamine, show potent cardioprotective and antidepressive effects in rodents, via sigma-1 receptor stimulation. Interestingly, agonist activation of sigma-1 receptors increased the secretion of mature BDNF from its precursor proBDNF via chaperone activity in the ER. Given the role of ER stress in the pathophysiology of CVD and MDD, the author will discuss the potential link between sigma-1 receptors and BDNF-TrkB pathway in the pathophysiology of these two diseases. Finally, the author will make a case for potent sigma-1 receptor agonists and TrkB agonists as new potential therapeutic drugs for depressive patients with CVD. Copyright © 2012 Elsevier Ltd. All rights reserved.

Decoding Corticotropin-Releasing Factor Receptor Type 1 Crystal 
Structures

PubMed Central

Doré, Andrew S.; Bortolato, Andrea; Hollenstein, Kaspar; Cheng, Robert K.Y.; Read, Randy J.; Marshall, Fiona H.

2017-01-01

The structural analysis of class B G protein-coupled receptors (GPCR), cell surface proteins responding to peptide hormones, has until recently been restricted to the extracellular domain (ECD). Cor-ticotropin-releasing factor receptor type 1 (CRF1R) is a class B receptor mediating stress response and also considered a drug target for depression and anxiety. Here we report the crystal structure of the trans-membrane domain of human CRF1R in complex with the small-molecule antagonist CP-376395 in a hex-agonal setting with translational non-crystallographic symmetry. Molecular dynamics and metadynamics simulations on this novel structure and the existing TMD structure for CRF1R provides insight as to how the small molecule ligand gains access to the induced-fit allosteric binding site with implications for the observed selectivity against CRF2R. Furthermore, molecular dynamics simulations performed using a full-length receptor model point to key interactions between the ECD and extracellular loop 3 of the TMD providing insight into the full inactive state of multidomain class B GPCRs. PMID:28183242

Sensing of blood pressure increase by transient receptor potential vanilloid 1 receptors on baroreceptors.

PubMed

Sun, Hao; Li, De-Pei; Chen, Shao-Rui; Hittelman, Walter N; Pan, Hui-Lin

2009-12-01

The arterial baroreceptor is critically involved in the autonomic regulation of homoeostasis. The transient receptor potential vanilloid 1 (TRPV1) receptor is expressed on both somatic and visceral sensory neurons. Here, we examined the TRPV1 innervation of baroreceptive pathways and its functional significance in the baroreflex. Resiniferatoxin (RTX), an ultrapotent analog of capsaicin, was used to ablate TRPV1-expressing afferent neurons and fibers in adult rats. Immunofluorescence labeling revealed that TRPV1 immunoreactivity was present on nerve fibers and terminals in the adventitia of the ascending aorta and aortic arch, the nodose ganglion neurons, and afferent fibers in the solitary tract of the brainstem. RTX treatment eliminated TRPV1 immunoreactivities in the aorta, nodose ganglion, and solitary tract. Renal sympathetic nerve activity, blood pressure, and heart rate were recorded in anesthetized rats. The baroreflex was triggered by lowering and raising blood pressure through intravenous infusion of sodium nitroprusside and phenylephrine, respectively. Inhibition of sympathetic nerve activity and heart rate by the phenylephrine-induced increase in blood pressure was largely impaired in RTX-treated rats. The maximum gain of the baroreflex function was significantly lower in RTX-treated than vehicle-treated rats. Furthermore, blocking of TRPV1 receptors significantly blunted the baroreflex and decreased the maximum gain of baroreflex function in the high blood pressure range. Our findings provide important new information that TRPV1 is expressed along the entire baroreceptive afferent pathway. TRPV1 receptors expressed on baroreceptive nerve endings can function as mechanoreceptors to detect the increase in blood pressure and maintain the homoeostasis.

Sensing of Blood Pressure Increase by Transient Receptor Potential Vanilloid 1 Receptors on Baroreceptors

PubMed Central

Sun, Hao; Li, De-Pei; Chen, Shao-Rui; Hittelman, Walter N.

2009-01-01

The arterial baroreceptor is critically involved in the autonomic regulation of homoeostasis. The transient receptor potential vanilloid 1 (TRPV1) receptor is expressed on both somatic and visceral sensory neurons. Here, we examined the TRPV1 innervation of baroreceptive pathways and its functional significance in the baroreflex. Resiniferatoxin (RTX), an ultrapotent analog of capsaicin, was used to ablate TRPV1-expressing afferent neurons and fibers in adult rats. Immunofluorescence labeling revealed that TRPV1 immunoreactivity was present on nerve fibers and terminals in the adventitia of the ascending aorta and aortic arch, the nodose ganglion neurons, and afferent fibers in the solitary tract of the brainstem. RTX treatment eliminated TRPV1 immunoreactivities in the aorta, nodose ganglion, and solitary tract. Renal sympathetic nerve activity, blood pressure, and heart rate were recorded in anesthetized rats. The baroreflex was triggered by lowering and raising blood pressure through intravenous infusion of sodium nitroprusside and phenylephrine, respectively. Inhibition of sympathetic nerve activity and heart rate by the phenylephrine-induced increase in blood pressure was largely impaired in RTX-treated rats. The maximum gain of the baroreflex function was significantly lower in RTX-treated than vehicle-treated rats. Furthermore, blocking of TRPV1 receptors significantly blunted the baroreflex and decreased the maximum gain of baroreflex function in the high blood pressure range. Our findings provide important new information that TRPV1 is expressed along the entire baroreceptive afferent pathway. TRPV1 receptors expressed on baroreceptive nerve endings can function as mechanoreceptors to detect the increase in blood pressure and maintain the homoeostasis. PMID:19726694

alpha1B-Adrenergic receptor phosphorylation and desensitization induced by transforming growth factor-beta.

PubMed Central

Romero-Avila, M Teresa; Flores-Jasso, C Fabián; García-Sáinz, J Adolfo

2002-01-01

Transforming growth factor-beta (TGF-beta) induced alpha(1B)-adrenergic receptor phosphorylation in Rat-1 fibroblasts stably expressing these adrenoceptors. This effect of TGF-beta was rapid, reaching a maximum within 30 min and decreasing thereafter, and concentration-dependent (EC(50) 0.3 pM). The phosphoinositide 3-kinase inhibitors wortmannin and LY294002, and the protein kinase C inhibitors staurosporine, Ro 318220 and bisindolylmaleimide, blocked the effect of this growth factor. alpha(1B)-Adrenergic receptor phosphorylation was associated with desensitization, as indicated by a reduction in the adrenergic-mediated production of [(3)H]inositol phosphates. Phosphorylation of alpha(1B)-adrenergic receptors by TGF-beta was also observed in Cos-1 cells transfected with the receptor. Co-transfection of the dominant-negative mutant of the regulatory subunit of phosphoinositide 3-kinase (Deltap85) inhibited the phosphorylation of alpha(1B)-adrenergic receptors induced by TGF-beta. Our results indicate that activation of TGF-beta receptors induces alpha(1B)-adrenergic receptor phosphorylation and desensitization. The data suggest that phosphoinositide 3-kinase and protein kinase C play key roles in this effect of TGF-beta. PMID:12234252

alpha1B-Adrenergic receptor phosphorylation and desensitization induced by transforming growth factor-beta.

PubMed

Romero-Avila, M Teresa; Flores-Jasso, C Fabián; García-Sáinz, J Adolfo

2002-12-01

Transforming growth factor-beta (TGF-beta) induced alpha(1B)-adrenergic receptor phosphorylation in Rat-1 fibroblasts stably expressing these adrenoceptors. This effect of TGF-beta was rapid, reaching a maximum within 30 min and decreasing thereafter, and concentration-dependent (EC(50) 0.3 pM). The phosphoinositide 3-kinase inhibitors wortmannin and LY294002, and the protein kinase C inhibitors staurosporine, Ro 318220 and bisindolylmaleimide, blocked the effect of this growth factor. alpha(1B)-Adrenergic receptor phosphorylation was associated with desensitization, as indicated by a reduction in the adrenergic-mediated production of [(3)H]inositol phosphates. Phosphorylation of alpha(1B)-adrenergic receptors by TGF-beta was also observed in Cos-1 cells transfected with the receptor. Co-transfection of the dominant-negative mutant of the regulatory subunit of phosphoinositide 3-kinase (Deltap85) inhibited the phosphorylation of alpha(1B)-adrenergic receptors induced by TGF-beta. Our results indicate that activation of TGF-beta receptors induces alpha(1B)-adrenergic receptor phosphorylation and desensitization. The data suggest that phosphoinositide 3-kinase and protein kinase C play key roles in this effect of TGF-beta.

Receptor for macrophage colony-stimulating factor transduces a signal decreasing erythroid potential in the multipotent hematopoietic EML cell line.

PubMed

Pawlak, G; Grasset, M F; Arnaud, S; Blanchet, J P; Mouchiroud, G

2000-10-01

To test the hypothesis that hematopoietic growth factors may influence lineage choice in pluripotent progenitor cells, we investigated the effects of macrophage colony-stimulating factor (M-CSF) on erythroid and myeloid potentials of multipotent EML cells ectopically expressing M-CSF receptor (M-CSFR). EML cells are stem cell factor (SCF)-dependent murine cells that give rise spontaneously to pre-B cells, burst-forming unit erythroid (BFU-E), and colony-forming unit granulocyte macrophage (CFU-GM). We determined BFU-E and CFU-GM frequencies among EML cells transduced with murine M-CSFR, human M-CSFR, or chimeric receptors, and cultivated in the presence of SCF, M-CSF, or both growth factors. Effects of specific inhibitors of signaling molecules were investigated. EML cells transduced with murine M-CSFR proliferated in response to M-CSF but also exhibited a sharp and rapid decrease in BFU-E frequency associated with an increase in CFU-GM frequency. In contrast, EML cells expressing human M-CSFR proliferated in response to M-CSF without any changes in erythroid or myeloid potential. Using chimeric receptors between human and murine M-CSFR, we showed that the effects of M-CSF on EML cell differentiation potential are mediated by a large region in the intracellular domain of murine M-CSFR. Furthermore, phospholipase C (PLC) inhibitor U73122 interfered with the negative effects of ligand-activated murine M-CSFR on EML cell erythroid potential. We propose that signaling pathways activated by tyrosine kinase receptors may regulate erythroid potential and commitment decisions in multipotent progenitor cells and that PLC may play a key role in this process.

Epidermal Growth Factor Receptor Transactivation: Mechanisms, Pathophysiology, and Potential Therapies in the Cardiovascular System.

PubMed

Forrester, Steven J; Kawai, Tatsuo; O'Brien, Shannon; Thomas, Walter; Harris, Raymond C; Eguchi, Satoru

2016-01-01

Epidermal growth factor receptor (EGFR) activation impacts the physiology and pathophysiology of the cardiovascular system, and inhibition of EGFR activity is emerging as a potential therapeutic strategy to treat diseases including hypertension, cardiac hypertrophy, renal fibrosis, and abdominal aortic aneurysm. The capacity of G protein-coupled receptor (GPCR) agonists, such as angiotensin II (AngII), to promote EGFR signaling is called transactivation and is well described, yet delineating the molecular processes and functional relevance of this crosstalk has been challenging. Moreover, these critical findings are dispersed among many different fields. The aim of our review is to highlight recent advancements in defining the signaling cascades and downstream consequences of EGFR transactivation in the cardiovascular renal system. We also focus on studies that link EGFR transactivation to animal models of the disease, and we discuss potential therapeutic applications.

Soluble tumor necrosis factor receptor-1 in preterm infants with chronic lung disease.

PubMed

Sato, Miho; Mori, Masaaki; Nishimaki, Shigeru; An, Hiromi; Naruto, Takuya; Sugai, Toshiyuki; Shima, Yoshio; Seki, Kazuo; Yokota, Shumpei

2010-04-01

It is clear that inflammation plays an important role in developing chronic lung disease in preterm infants. The purpose of the present study is to investigate changes of serum soluble tumor necrosis factor receptor-1 levels over time in infants with chronic lung disease. The serum levels of soluble tumor necrosis factor receptor-1 were measured after delivery, and at 7, 14, 21 and 28 days of age in 10 infants with chronic lung disease and in 18 infants without chronic lung disease. The serum level of soluble tumor necrosis factor receptor-1 was significantly higher in infants with chronic lung disease than in infants without chronic lung disease after delivery. The differences between these two groups remained up to 28 days of age. Prenatal inflammation with persistence into postnatal inflammation may be involved in the onset of chronic lung disease.

Cleavage of the urokinase receptor (uPAR) on oral cancer cells: regulation by transforming growth factor - β1 (TGF-β1) and potential effects on migration and invasion.

PubMed

Magnussen, Synnove Norvoll; Hadler-Olsen, Elin; Costea, Daniela Elena; Berg, Eli; Jacobsen, Cristiane Cavalcanti; Mortensen, Bente; Salo, Tuula; Martinez-Zubiaurre, Inigo; Winberg, Jan-Olof; Uhlin-Hansen, Lars; Svineng, Gunbjorg

2017-05-19

Urokinase plasminogen activator (uPA) receptor (uPAR) is up-regulated at the invasive tumour front of human oral squamous cell carcinoma (OSCC), indicating a role for uPAR in tumour progression. We previously observed elevated expression of uPAR at the tumour-stroma interface in a mouse model for OSCC, which was associated with increased proteolytic activity. The tumour microenvironment regulated uPAR expression, as well as its glycosylation and cleavage. Both full-length- and cleaved uPAR (uPAR (II-III)) are involved in highly regulated processes such as cell signalling, proliferation, migration, stem cell mobilization and invasion. The aim of the current study was to analyse tumour associated factors and their effect on uPAR cleavage, and the potential implications for cell proliferation, migration and invasion. Mouse uPAR was stably overexpressed in the mouse OSCC cell line AT84. The ratio of full-length versus cleaved uPAR as analysed by Western blotting and its regulation was assessed by addition of different protease inhibitors and transforming growth factor - β1 (TGF-β1). The role of uPAR cleavage in cell proliferation and migration was analysed using real-time cell analysis and invasion was assessed using the myoma invasion model. We found that when uPAR was overexpressed a proportion of the receptor was cleaved, thus the cells presented both full-length uPAR and uPAR (II-III). Cleavage was mainly performed by serine proteases and urokinase plasminogen activator (uPA) in particular. When the OSCC cells were stimulated with TGF-β1, the production of the uPA inhibitor PAI-1 was increased, resulting in a reduction of uPAR cleavage. By inhibiting cleavage of uPAR, cell migration was reduced, and by inhibiting uPA activity, invasion was reduced. We could also show that medium containing soluble uPAR (suPAR), and cleaved soluble uPAR (suPAR (II-III)), induced migration in OSCC cells with low endogenous levels of uPAR. These results show that soluble factors in

MUC4 potentiates invasion and metastasis of pancreatic cancer cells through stabilization of fibroblast growth factor receptor 1

PubMed Central

Macha, Muzafar A; Ponnusamy, Moorthy P.; Batra, Surinder K

2012-01-01

MUC4 is a type-1 transmembrane mucin differentially expressed in multiple cancers and has previously been shown to potentiate progression and metastasis of pancreatic cancer. In this study, we investigated the molecular mechanisms associated with the MUC4-induced invasion and metastasis in pancreatic cancer. Stable silencing of MUC4 in multiple pancreatic cancer cells resulted in the downregulation of N-cadherin and its interacting partner fibroblast growth factor receptor 1 (FGFR1) through downregulation of partly by pFAK, pMKK7, pJNK and pc-Jun pathway and partly through PI-3K/Akt pathway. The downregulation of FGFR1 in turn led to downregulation of pAkt, pERK1/2, pNF-κB, pIkBα, uPA, MMP-9, vimentin, N-cadherin, Twist, Slug and Zeb1 and upregulation of E-cadherin, Occludin, Cytokeratin-18 and Caspase-9 in MUC4 knockdown BXPC3 and Capan1 cells compared with scramble vector transfected cells. Further, downregulation of FGFR1 was associated with a significant change in morphology and reorganization of the actin-cytoskeleton, leading to a significant decrease in motility (P < 0.00001) and invasion (P < 0.0001) in vitro and decreased tumorigenicity and incidence of metastasis in vivo upon orthotopic implantation in the athymic mice. Taken together, the results of the present study suggest that MUC4 promotes invasion and metastasis by FGFR1 stabilization through the N-cadherin upregulation. PMID:22791819

The heterodimeric sweet taste receptor has multiple potential ligand binding sites.

PubMed

Cui, Meng; Jiang, Peihua; Maillet, Emeline; Max, Marianna; Margolskee, Robert F; Osman, Roman

2006-01-01

The sweet taste receptor is a heterodimer of two G protein coupled receptors, T1R2 and T1R3. This discovery has increased our understanding at the molecular level of the mechanisms underlying sweet taste. Previous experimental studies using sweet receptor chimeras and mutants show that there are at least three potential binding sites in this heterodimeric receptor. Receptor activity toward the artificial sweeteners aspartame and neotame depends on residues in the amino terminal domain of human T1R2. In contrast, receptor activity toward the sweetener cyclamate and the sweet taste inhibitor lactisole depends on residues within the transmembrane domain of human T1R3. Furthermore, receptor activity toward the sweet protein brazzein depends on the cysteine rich domain of human T1R3. Although crystal structures are not available for the sweet taste receptor, useful homology models can be developed based on appropriate templates. The amino terminal domain, cysteine rich domain and transmembrane helix domain of T1R2 and T1R3 have been modeled based on the crystal structures of metabotropic glutamate receptor type 1, tumor necrosis factor receptor, and bovine rhodopsin, respectively. We have used homology models of the sweet taste receptors, molecular docking of sweet ligands to the receptors, and site-directed mutagenesis of the receptors to identify potential ligand binding sites of the sweet taste receptor. These studies have led to a better understanding of the structure and function of this heterodimeric receptor, and can act as a guide for rational structure-based design of novel non-caloric sweeteners, which can be used in the fighting against obesity and diabetes.

Epidermal Growth Factor Receptor Tyrosine Kinase: A Potential Target in Treatment of Non-Small-Cell Lung Carcinoma.

PubMed

Prabhu, Venugopal Vinod; Devaraj, Niranjali

2017-01-01

Lung cancer is responsible for 1.6 million deaths. Approximately 80%-85% of lung cancers are of the non-small-cell variety, which includes squamous cell carcinoma, adenocarcinoma, and large-cell carcinoma. Knowing the stage of cancer progression is a requisite for determining which management approach-surgery, chemotherapy, radiotherapy, and/or immunotherapy-is optimal. Targeted therapeutic approaches with antiangiogenic monoclonal antibodies or tyrosine kinase inhibitors are one option if tumors harbor oncogene mutations. Another, newer approach is directed against cancer-specific molecules and signaling pathways and thus has more limited nonspecific toxicities. This approach targets the epidermal growth factor receptor (EGFR, HER-1/ErbB1), a receptor tyrosine kinase of the ErbB family, which consists of four closely related receptors: HER-1/ErbB1, HER-2/neu/ErbB2, HER-3/ErbB3, and HER-4/ErbB4. Because EGFR is expressed at high levels on the surface of some cancer cells, it has been recognized as an effective anticancer target. EGFR-targeted therapies include monoclonal antibodies (mAbs) and small-molecule tyrosine kinase inhibitors. Tyrosine kinases are an especially important target because they play an important role in the modulation of growth factor signaling. This review highlights various classes of synthetically derived molecules that have been reported in the last few years as potential EGFR-TK inhibitors (TKIs) and their targeted therapies in NSCLC, along with effective strategies for overcoming EGFR-TKI resistance and efforts to develop a novel potent EGFR-TKI as an efficient target of NSCLC treatment in the foreseeable future.

The ubiquitin ligase Nedd4 mediates oxidized low-density lipoprotein-induced downregulation of insulin-like growth factor-1 receptor

PubMed Central

Higashi, Yusuke; Sukhanov, Sergiy; Parthasarathy, Sampath; Delafontaine, Patrice

2008-01-01

Oxidized low-density lipoprotein (LDL) is proatherogenic and induces smooth muscle cell apoptosis, which contributes to atherosclerotic plaque destabilization. We showed previously that oxidized LDL downregulates insulin-like growth factor-1 receptor in human smooth muscle cells and that this is critical for induction of apoptosis. To identify mechanisms, we exposed smooth muscle cells to 60 μg/ml oxidized LDL or native LDL and assessed insulin-like growth factor-1 receptor mRNA levels, protein synthesis rate, and receptor protein stability. Oxidized LDL decreased insulin-like growth factor-1 receptor mRNA levels by 30% at 8 h compared with native LDL, and this decrease was maintained for up to 20 h. However, insulin-like growth factor-1 receptor protein synthesis rate was not altered by oxidized LDL. Pulse-chase labeling experiments revealed that oxidized LDL reduced insulin-like growth factor-1 receptor protein half-life to 12.2 ± 1.7 h from 24.4 ± 4.7 h with native LDL. This destabilization of insulin-like growth factor-1 receptor protein was accompanied by enhanced receptor ubiquitination. Overexpression of dominant-negative Nedd4 prevented oxidized LDL-induced downregulation of insulin-like growth factor-1 receptor, suggesting that Nedd4 was the ubiquitin ligase that mediated receptor downregulation. However, the proteasome inhibitors lactacystin, MG-132, and proteasome inhibitor-1 failed to block oxidized LDL-induced downregulation of insulin-like growth factor-1 receptor. Thus oxidized LDL downregulates insulin-like growth factor-1 receptor by destabilizing the protein via Nedd4-enhanced ubiquitination, leading to degradation via a proteasome-independent pathway. This finding provides novel insights into oxidized LDL-triggered oxidant signaling and mechanisms of smooth muscle cell depletion that contribute to plaque destabilization and coronary events. PMID:18723765

TNF Receptor 2 Makes Tumor Necrosis Factor a Friend of Tumors

PubMed Central

Sheng, Yuqiao; Li, Feng; Qin, Zhihai

2018-01-01

Tumor necrosis factor (TNF) is widely accepted as a tumor-suppressive cytokine via its ubiquitous receptor TNF receptor 1 (TNFR1). The other receptor, TNFR2, is not only expressed on some tumor cells but also on suppressive immune cells, including regulatory T cells and myeloid-derived suppressor cells. In contrast to TNFR1, TNFR2 diverts the tumor-inhibiting TNF into a tumor-advocating factor. TNFR2 directly promotes the proliferation of some kinds of tumor cells. Also activating immunosuppressive cells, it supports immune escape and tumor development. Hence, TNFR2 may represent a potential target of cancer therapy. Here, we focus on expression and role of TNFR2 in the tumor microenvironment. We summarize the recent progress in understanding how TNFR2-dependent mechanisms promote carcinogenesis and tumor growth and discuss the potential value of TNFR2 in cancer treatment. PMID:29892300

Identification of a potential tumor differentiation factor receptor candidate in prostate cancer cells.

PubMed

Sokolowska, Izabela; Woods, Alisa G; Gawinowicz, Mary Ann; Roy, Urmi; Darie, Costel C

2012-07-01

Tumor differentiation factor (TDF) is a pituitary protein that is secreted into the bloodstream and has an endocrine function. TDF and TDF-P1, a 20-residue peptide selected from the ORF of TDF, induce differentiation in human breast and prostate cancer cells, but not in other cells. TDF has no known mechanism of action. In our recent study, we identified heat shock 70 kDa proteins (HSP70s) as TDF receptors (TDF-Rs) in breast cancer cells. Therefore, we sought to investigate whether TDF-R candidates from prostate cancer cells are the same as those identified in breast cancer cells. Here, we used TDF-P1 to purify the potential TDF-R candidates by affinity purification chromatography from DU145 and PC3 steroid-resistant prostate cancer cells, LNCaP steroid-responsive prostate cancer cells, and nonprostate NG108 neuroblastoma and BLK CL.4 fibroblast-like cells. We identified the purified proteins by MS, and validated them by western blotting, immunofluorescence microscopy, immunoaffinity purification chromatography, and structural biology. We identified seven candidate proteins, of which three were from the HSP70 family. These three proteins were validated as potential TDF-R candidates in LNCaP steroid-responsive and in DU145 and PC3 steroid-resistant prostate cancer cells, but not in NG108 neuroblastoma and BLK CL.4 fibroblast-like cells. Our previous study and the current study suggest that GRP78, and perhaps HSP70s, are strong TDF-R candidates, and further suggest that TDF interacts with its receptors exclusively in breast and prostate cells, inducing cell differentiation through a novel, steroid-independent pathway. © 2012 The Authors Journal compilation © 2012 FEBS.

Activation of cannabinoid CB1 receptors modulates evoked action potentials in rat retinal ganglion cells.

PubMed

Jiang, Shu-Xia; Li, Qian; Wang, Xiao-Han; Li, Fang; Wang, Zhong-Feng

2013-08-25

Activation of cannabinoid CB1 receptors (CB1Rs) regulates a variety of physiological functions in the vertebrate retina through modulating various types of ion channels. The aim of the present study was to investigate the effects of this receptor on cell excitability of rat retinal ganglion cells (RGCs) in retinal slices using whole-cell patch-clamp techniques. The results showed that under current-clamped condition perfusing WIN55212-2 (WIN, 5 μmol/L), a CB1R agonist, did not significantly change the spontaneous firing frequency and resting membrane potential of RGCs. In the presence of cocktail synaptic blockers, including excitatory postsynaptic receptor blockers CNQX and D-APV, and inhibitory receptor blockers bicuculline and strychnine, perfusion of WIN (5 μmol/L) hardly changed the frequencies of evoked action potentials by a series of positive current injection (from +10 to +100 pA). Phase-plane plot analysis showed that both average threshold voltage for triggering action potential and delay time to reach threshold voltage were not affected by WIN. However, WIN significantly decreased +dV/dtmax and -dV/dtmax of action potentials, suggestive of reduced rising and descending velocities of action potentials. The effects of WIN were reversed by co-application of SR141716, a CB1R selective antagonist. Moreover, WIN did not influence resting membrane potential of RGCs with synaptic inputs being blocked. These results suggest that activation of CB1Rs may regulate intrinsic excitability of rat RGCs through modulating evoked action potentials.

Resveratrol inhibits proteinase-activated receptor-2-induced release of soluble vascular endothelial growth factor receptor-1 from human endothelial cells

PubMed Central

Al-Ani, Bahjat

2013-01-01

We recently reported that (i) activation of the proinflammatory receptor, proteinase-activated receptor-2 (PAR-2) caused the release of an important biomarker in preeclampsia, soluble vascular endothelial growth factor receptor-1 (sVEGFR-1, also known as sFlt-1) from human umbilical vein endothelial cells (HUVECs), and (ii) that the anti-oxidant and anti-inflammatory agent, resveratrol, is capable of inhibiting the proinflammatory cytokine-induced sVEGFR-1 release from human placenta. Based on these findings and because PAR-2 is upregulated by proinflammatory cytokines, we sought to determine whether resveratrol can inhibit PAR-2-induced sVEGFR-1 release. PAR-2 expressing cells, HUVECs and human embryonic kidney cells (HEK-293) transfected with a human VEGFR-1 promoter-luciferase reporter construct were incubated with PAR-2-activating peptide and/or resveratrol. Cell supernatants were assayed for sVEGFR-1 by enzyme-linked immunosorbent assay (ELISA), and VEGFR-1 promoter-luciferase assay was performed on the harvested cell lysates. Preincubation of HEK-293 cells with resveratrol significantly inhibited PAR-2-induced VEGFR-1 promoter activity without affecting cell viability as assessed by MTT assay. The addition of resveratrol also blocked PAR-2-mediated sVEGFR-1 release from HUVECs. The present study demonstrates that resveratrol suppressed both VEGFR-1 promoter activity and sVEGFR-1 protein release induced by PAR-2 activation, which further endorses our recent findings of a potential therapeutic role for resveratrol in preeclampsia. PMID:26933402

Insulin growth factor-1 (IGF-1) enhances hippocampal excitatory and seizure activity through IGF-1 receptor-mediated mechanisms in the epileptic brain.

PubMed

Jiang, Guohui; Wang, Wei; Cao, Qingqing; Gu, Juan; Mi, Xiujuan; Wang, Kewei; Chen, Guojun; Wang, Xuefeng

2015-12-01

Insulin-like growth factor-1 (IGF-1) is known to promote neurogenesis and survival. However, recent studies have suggested that IGF-1 regulates neuronal firing and excitatory neurotransmission. In the present study, focusing on temporal lobe epilepsy, we found that IGF-1 levels and IGF-1 receptor activation are increased in human epileptogenic tissues, and pilocarpine- and pentylenetetrazole-treated rat models. Using an acute model of seizures, we showed that lateral cerebroventricular infusion of IGF-1 elevates IGF-1 receptor (IGF-1R) signalling before pilocarpine application had proconvulsant effects. In vivo electroencephalogram recordings and power spectrogram analysis of local field potential revealed that IGF-1 promotes epileptiform activities. This effect is diminished by co-application of an IGF-1R inhibitor. In an in vitro electrophysiological study, we demonstrated that IGF-1 enhancement of excitatory neurotransmission and α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid receptor- and N-methyl-D-aspartate receptor-mediated currents is inhibited by IGF-1R inhibitor. Finally, activation of extracellular signal-related kinase (ERK)-1/2 and protein kinase B (Akt) in seizures in rats is increased by exogenous IGF-1 and diminished by picropodophyllin. A behavioural study reveals that the ERK1/2 or Akt inhibitor attenuates seizure activity. These results indicate that increased IGF-1 levels after recurrent hippocampal neuronal firings might, in turn, promote seizure activity via IGF-1R-dependent mechanisms. The present study presents a previously unappreciated role of IGF-1R in the development of seizure activity. © 2015 Authors; published by Portland Press Limited.

Design and Synthesis of Benzimidazoles As Novel Corticotropin-Releasing Factor 1 Receptor Antagonists.

PubMed

Mochizuki, Michiyo; Kori, Masakuni; Kobayashi, Katsumi; Yano, Takahiko; Sako, Yuu; Tanaka, Maiko; Kanzaki, Naoyuki; Gyorkos, Albert C; Corrette, Christopher P; Cho, Suk Young; Pratt, Scott A; Aso, Kazuyoshi

2016-03-24

Benzazole derivatives with a flexible aryl group bonded through a one-atom linker as a new scaffold for a corticotropin-releasing factor 1 (CRF1) receptor antagonist were designed, synthesized, and evaluated. We expected that structural diversity could be expanded beyond that of reported CRF1 receptor antagonists. In a structure-activity relationship study, 4-chloro-N(2)-(4-chloro-2-methoxy-6-methylphenyl)-1-methyl-N(7),N(7)-dipropyl-1H-benzimidazole-2,7-diamine 29g had the most potent binding activity against a human CRF1 receptor and the antagonistic activity (IC50 = 9.5 and 88 nM, respectively) without concerns regarding cytotoxicity at 30 μM. Potent CRF1 receptor-binding activity in brain in an ex vivo test and suppression of stress-induced activation of the hypothalamus-pituitary-adrenocortical (HPA) axis were also observed at 138 μmol/kg of compound 29g after oral administration in mice. Thus, the newly designed benzimidazole 29g showed in vivo CRF1 receptor antagonistic activity and good brain penetration, indicating that it is a promising lead for CRF1 receptor antagonist drug discovery research.

Combined caveolin-1 and epidermal growth factor receptor expression as a prognostic marker for breast cancer.

PubMed

Liang, Ya-Nan; Liu, Yu; Wang, Letian; Yao, Guodong; Li, Xiaobo; Meng, Xiangning; Wang, Fan; Li, Ming; Tong, Dandan; Geng, Jingshu

2018-06-01

Previous studies have indicated that caveolin-1 (Cav-1) is able to bind the signal transduction factor epidermal growth factor receptor (EGFR) to regulate its tyrosine kinase activity. The aim of the present study was to evaluate the clinical significance of Cav-1 gene expression in association with the expression of EGFR in patients with breast cancer. Primary breast cancer samples from 306 patients were analyzed for Cav-1 and EGFR expression using immunohistochemistry, and clinical significance was assessed using multivariate Cox regression analysis, Kaplan-Meier estimator curves and the log-rank test. Stromal Cav-1 was downregulated in 38.56% (118/306) of tumor tissues, whereas cytoplasmic EGFR and Cav-1 were overexpressed in 53.92% (165/306) and 44.12% (135/306) of breast cancer tissues, respectively. EGFR expression was positively associated with cytoplasmic Cav-1 and not associated with stromal Cav-1 expression in breast cancer samples; however, low expression of stromal Cav-1 was negatively associated with cytoplasmic Cav-1 expression in total tumor tissues, and analogous results were identified in the chemotherapy group. Multivariate Cox's proportional hazards model analysis revealed that, for patients in the estrogen receptor (ER)(+) group, the expression of stromal Cav-1 alone was a significant prognostic marker of breast cancer. However, in the chemotherapy, human epidermal growth factor receptor 2 (HER-2)(-), HER-2(+) and ER(-) groups, the use of combined markers was more effective prognostic marker. Stromal Cav-1 has a tumor suppressor function, and the combined marker stromal Cav-1/EGFR expression was identified as an improved prognostic marker in the diagnosis of breast cancer. Parenchymal expression of Cav-1 is able to promote EGFR signaling in breast cancer, potentially being required for EGFR-mediated initiation of mitosis.

Fibroblast growth factor receptor inhibitors.

PubMed

Kumar, Suneel B V S; Narasu, Lakshmi; Gundla, Rambabu; Dayam, Raveendra; J A R P, Sarma

2013-01-01

Fibroblast growth factor receptors (FGFRs) play an important role in embryonic development, angiogenesis, wound healing, cell proliferation and differentiation. The fibroblast growth factor receptor (FGFR) isoforms have been under intense scrutiny for effective anticancer drug candidates. The fibroblast growth factor (FGF) and its receptor (FGFR) provide another pathway that seems critical to monitoring angiogenesis. Recent findings suggest that FGFR mediates signaling, regulates the PKM2 activity, and plays a crucial role in cancer metabolism. The current review also covers the recent findings on the role of FGFR1 in cancer metabolism. This paper reviews the progress, mechanism, and binding modes of recently known kinase inhibitors such as PD173074, SU series and other inhibitors still under clinical development. Some of the structural classes that will be highlighted in this review include Pyrido[2,3-d]pyrimidines, Indolin- 2-one, Pyrrolo[2,1-f][1,2,4]triazine, Pyrido[2,3-d]pyrimidin-7(8H)-one, and 1,6- Naphthyridin-2(1H)-ones.

Endothelin‐1 and its receptors on haemorrhoidal tissue: a potential site for therapeutic intervention

PubMed Central

Lohsiriwat, Varut; Scholefield, John H; Wilson, Vincent G

2017-01-01

Background and Purpose Haemorrhoids is a common anorectal condition affecting millions worldwide. We have studied the effect of endothelin‐1 (ET‐1) and the role of endothelin ETA and ETB receptors in haemorrhoid tissue. Experimental Approach Protein expression of ET‐1, ETA and ETB receptors were compared between haemorrhoids and normal rectal submucosa using Western blot analysis, with the localization of proteins determined by autoradiography and immunohistochemistry. Effects of ET‐1 and sarafotoxin 6a on human colonic and rectal arteries and veins was assessed by wire myography and the involvement of receptor subtypes established by selective antagonists. Key Results Dense binding of [125I]‐ET‐1 to haemorrhoidal sections was reduced by selective receptor antagonists. A higher density of ETB than ETA receptors was found in haemorrhoidal, than in control rectal tissue and confirmed by Western blot analysis. ETA and ETB receptors were localized to smooth muscle of haemorrhoidal arteries and veins, with ETB receptors on the endothelium. Human colonic and rectal arteries and veins were similarly sensitive to ET‐1 and affected by the ETA selective antagonist, but sarafotoxin S6a‐induced contractions were more pronounced in veins and antagonized by a selective ETB receptor antagonist. Conclusions and Implications ETA and ETB receptors are present in human haemorrhoids with ETB receptors predominating. ETA receptors are activated by ET‐1 to mediate a contraction in arteries and veins, but the latter are selectively activated by sarafotoxin S6a – a response that involves ETB receptors at low concentrations. Selective ETB agonists may have therapeutic potential to reduce congestion of the haemorrhoidal venous sinusoids. PMID:28095606

Endothelin-1 and its receptors on haemorrhoidal tissue: a potential site for therapeutic intervention.

PubMed

Lohsiriwat, Varut; Scholefield, John H; Wilson, Vincent G; Dashwood, Michael R

2017-04-01

Haemorrhoids is a common anorectal condition affecting millions worldwide. We have studied the effect of endothelin-1 (ET-1) and the role of endothelin ET A and ET B receptors in haemorrhoid tissue. Protein expression of ET-1, ET A and ET B receptors were compared between haemorrhoids and normal rectal submucosa using Western blot analysis, with the localization of proteins determined by autoradiography and immunohistochemistry. Effects of ET-1 and sarafotoxin 6a on human colonic and rectal arteries and veins was assessed by wire myography and the involvement of receptor subtypes established by selective antagonists. Dense binding of [ 125 I]-ET-1 to haemorrhoidal sections was reduced by selective receptor antagonists. A higher density of ET B than ET A receptors was found in haemorrhoidal, than in control rectal tissue and confirmed by Western blot analysis. ET A and ET B receptors were localized to smooth muscle of haemorrhoidal arteries and veins, with ET B receptors on the endothelium. Human colonic and rectal arteries and veins were similarly sensitive to ET-1 and affected by the ET A selective antagonist, but sarafotoxin S6a-induced contractions were more pronounced in veins and antagonized by a selective ET B receptor antagonist. ET A and ET B receptors are present in human haemorrhoids with ET B receptors predominating. ET A receptors are activated by ET-1 to mediate a contraction in arteries and veins, but the latter are selectively activated by sarafotoxin S6a - a response that involves ET B receptors at low concentrations. Selective ET B agonists may have therapeutic potential to reduce congestion of the haemorrhoidal venous sinusoids. © 2017 The British Pharmacological Society.

Unliganded fibroblast growth factor receptor 1 forms density-independent dimers.

PubMed

Comps-Agrar, Laëtitia; Dunshee, Diana Ronai; Eaton, Dan L; Sonoda, Junichiro

2015-10-02

Fibroblast growth factors receptors (FGFRs) are thought to initiate intracellular signaling cascades upon ligand-induced dimerization of the extracellular domain. Although the existence of unliganded FGFR1 dimers on the surface of living cells has been proposed, this notion remains rather controversial. Here, we employed time-resolved Förster resonance energy transfer combined with SNAP- and ACP-tag labeling in COS7 cells to monitor dimerization of full-length FGFR1 at the cell-surface with or without the coreceptor βKlotho. Using this approach we observed homodimerization of unliganded FGFR1 that is independent of its surface density. The homo-interaction signal observed for FGFR1 was indeed as robust as that obtained for epidermal growth factor receptor (EGFR) and was further increased by the addition of activating ligands or pathogenic mutations. Mutational analysis indicated that the kinase and the transmembrane domains, rather than the extracellular domain, mediate the ligand-independent FGFR1 dimerization. In addition, we observed a formation of a higher order ligand-independent complex by the c-spliced isoform of FGFR1 and βKlotho. Collectively, our approach provides novel insights into the assembly and dynamics of the full-length FGFRs on the cell surface. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

GPER-1 agonist G1 induces vasorelaxation through activation of epidermal growth factor receptor-dependent signalling pathway.

PubMed

Jang, Eun Jin; Seok, Young Mi; Arterburn, Jeffrey B; Olatunji, Lawrence A; Kim, In Kyeom

2013-10-01

The G protein-coupled oestrogen receptor-1 (GPER-1) agonist G1 induces endothelium-dependent relaxation. Activation of the epidermal growth factor (EGF) receptor leads to transduction of signals from the plasma membrane for the release of nitric oxide. We tested the hypothesis that G1 induces endothelium-dependent vasorelaxation through activation of the EGF receptor. Rat aortic rings were mounted in organ baths. After pretreatment with various inhibitors, aortic rings contracted with 11,9-epoxymethano-prostaglandin F2α or KCl were subjected to relaxation by G1. G1 induced endothelium-dependent vasorelaxation, which was attenuated by pretreatment with either L -N(ω) -nitroarginine methyl ester (L -NAME), an inhibitor of nitric oxide synthase, or (3aS,4R,9bR)-4-(6-bromo-1,3-benzodioxol-5-yl)-3a,4,5,9b-tetrahydro-3H-cyclopenta[c]quinoline HB-EGF, heparin-binding EGF-like growth factor, a GPER-1 antagonist. Neither a general oestrogen receptor antagonist, ICI 182 780, nor a selective oestrogen receptor-α antagonist, methyl-piperidino-pyrazole dihydrochloride (MPP), had an effect on G1-induced vasorelaxation. However, pretreatment with EGF receptor blockers, AG1478 or DAPH, resulted in attenuated G1-induced vasorelaxation. In addition, pretreatment with Src inhibitor 4-amino-3-(4-chlorophenyl)-1-(t-butyl)-1H-pyrazolo[3,4-d]pyrimidine, 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine or Akt inhibitor VIII also resulted in attenuated vascular relaxation induced by the cumulative addition of G1. However, neither phosphatidylinositol-3 kinase inhibitors LY294002 and wortmannin nor an extracellular signal-regulated kinase inhibitor 1,4-diamino-2,3-dicyano-1,4-bis(o-aminophenylmercapto) butadiene monoethanolate had effect on vascular relaxation induced by the cumulative addition of G1. G1 induces endothelium-dependent vasorelaxation through Src-mediated activation of the EGF receptor and the Akt pathway in rat aorta. © 2013 Royal Pharmaceutical Society.

All-Atom Structural Models of the Transmembrane Domains of Insulin and Type 1 Insulin-Like Growth Factor Receptors

PubMed Central

Mohammadiarani, Hossein; Vashisth, Harish

2016-01-01

The receptor tyrosine kinase superfamily comprises many cell-surface receptors including the insulin receptor (IR) and type 1 insulin-like growth factor receptor (IGF1R) that are constitutively homodimeric transmembrane glycoproteins. Therefore, these receptors require ligand-triggered domain rearrangements rather than receptor dimerization for activation. Specifically, binding of peptide ligands to receptor ectodomains transduces signals across the transmembrane domains for trans-autophosphorylation in cytoplasmic kinase domains. The molecular details of these processes are poorly understood in part due to the absence of structures of full-length receptors. Using MD simulations and enhanced conformational sampling algorithms, we present all-atom structural models of peptides containing 51 residues from the transmembrane and juxtamembrane regions of IR and IGF1R. In our models, the transmembrane regions of both receptors adopt helical conformations with kinks at Pro961 (IR) and Pro941 (IGF1R), but the C-terminal residues corresponding to the juxtamembrane region of each receptor adopt unfolded and flexible conformations in IR as opposed to a helix in IGF1R. We also observe that the N-terminal residues in IR form a kinked-helix sitting at the membrane–solvent interface, while homologous residues in IGF1R are unfolded and flexible. These conformational differences result in a larger tilt-angle of the membrane-embedded helix in IGF1R in comparison to IR to compensate for interactions with water molecules at the membrane–solvent interfaces. Our metastable/stable states for the transmembrane domain of IR, observed in a lipid bilayer, are consistent with a known NMR structure of this domain determined in detergent micelles, and similar states in IGF1R are consistent with a previously reported model of the dimerized transmembrane domains of IGF1R. Our all-atom structural models suggest potentially unique structural organization of kinase domains in each receptor. PMID

Residues remote from the binding pocket control the antagonist selectivity towards the corticotropin-releasing factor receptor-1

NASA Astrophysics Data System (ADS)

Sun, Xianqiang; Cheng, Jianxin; Wang, Xu; Tang, Yun; Ågren, Hans; Tu, Yaoquan

2015-01-01

The corticotropin releasing factors receptor-1 and receptor-2 (CRF1R and CRF2R) are therapeutic targets for treating neurological diseases. Antagonists targeting CRF1R have been developed for the potential treatment of anxiety disorders and alcohol addiction. It has been found that antagonists targeting CRF1R always show high selectivity, although CRF1R and CRF2R share a very high rate of sequence identity. This has inspired us to study the origin of the selectivity of the antagonists. We have therefore built a homology model for CRF2R and carried out unbiased molecular dynamics and well-tempered metadynamics simulations for systems with the antagonist CP-376395 in CRF1R or CRF2R to address this issue. We found that the side chain of Tyr6.63 forms a hydrogen bond with the residue remote from the binding pocket, which allows Tyr6.63 to adopt different conformations in the two receptors and results in the presence or absence of a bottleneck controlling the antagonist binding to or dissociation from the receptors. The rotameric switch of the side chain of Tyr3566.63 allows the breaking down of the bottleneck and is a perquisite for the dissociation of CP-376395 from CRF1R.

Residues remote from the binding pocket control the antagonist selectivity towards the corticotropin-releasing factor receptor-1

PubMed Central

Sun, Xianqiang; Cheng, Jianxin; Wang, Xu; Tang, Yun; Ågren, Hans; Tu, Yaoquan

2015-01-01

The corticotropin releasing factors receptor-1 and receptor-2 (CRF1R and CRF2R) are therapeutic targets for treating neurological diseases. Antagonists targeting CRF1R have been developed for the potential treatment of anxiety disorders and alcohol addiction. It has been found that antagonists targeting CRF1R always show high selectivity, although CRF1R and CRF2R share a very high rate of sequence identity. This has inspired us to study the origin of the selectivity of the antagonists. We have therefore built a homology model for CRF2R and carried out unbiased molecular dynamics and well-tempered metadynamics simulations for systems with the antagonist CP-376395 in CRF1R or CRF2R to address this issue. We found that the side chain of Tyr6.63 forms a hydrogen bond with the residue remote from the binding pocket, which allows Tyr6.63 to adopt different conformations in the two receptors and results in the presence or absence of a bottleneck controlling the antagonist binding to or dissociation from the receptors. The rotameric switch of the side chain of Tyr3566.63 allows the breaking down of the bottleneck and is a perquisite for the dissociation of CP-376395 from CRF1R. PMID:25628267

Residues remote from the binding pocket control the antagonist selectivity towards the corticotropin-releasing factor receptor-1.

PubMed

Sun, Xianqiang; Cheng, Jianxin; Wang, Xu; Tang, Yun; Ågren, Hans; Tu, Yaoquan

2015-01-28

The corticotropin releasing factors receptor-1 and receptor-2 (CRF1R and CRF2R) are therapeutic targets for treating neurological diseases. Antagonists targeting CRF1R have been developed for the potential treatment of anxiety disorders and alcohol addiction. It has been found that antagonists targeting CRF1R always show high selectivity, although CRF1R and CRF2R share a very high rate of sequence identity. This has inspired us to study the origin of the selectivity of the antagonists. We have therefore built a homology model for CRF2R and carried out unbiased molecular dynamics and well-tempered metadynamics simulations for systems with the antagonist CP-376395 in CRF1R or CRF2R to address this issue. We found that the side chain of Tyr(6.63) forms a hydrogen bond with the residue remote from the binding pocket, which allows Tyr(6.63) to adopt different conformations in the two receptors and results in the presence or absence of a bottleneck controlling the antagonist binding to or dissociation from the receptors. The rotameric switch of the side chain of Tyr356(6.63) allows the breaking down of the bottleneck and is a perquisite for the dissociation of CP-376395 from CRF1R.

PKCepsilon-dependent potentiation of TTX-resistant Nav1.8 current by neurokinin-1 receptor activation in rat dorsal root ganglion neurons.

PubMed

Cang, Chun-Lei; Zhang, Hua; Zhang, Yu-Qiu; Zhao, Zhi-Qi

2009-06-30

Substance P (SP), which mainly exists in a subtype of small-diameter dorsal root ganglion (DRG) neurons, is an important signal molecule in pain processing in the spinal cord. Our previous results have proved the expression of SP receptor neurokinin-1 (NK-1) on DRG neurons and its interaction with transient receptor potential vanilloid 1 (TRPV1) receptor. In this study we investigated the effect of NK-1 receptor agonist on Na(v)1.8, a tetrodotoxin (TTX)-resistant sodium channel, in rat small-diameter DRG neurons employing whole-cell patch clamp recordings. NK-1 agonist [Sar(9), Met(O2)(11)]-substance P (Sar-SP) significantly enhanced the Na(v)1.8 currents in a subgroup of small-diameter DRG neurons under both the normal and inflammatory situation, and the enhancement was blocked by NK-1 antagonist Win51708 and protein kinase C (PKC) inhibitor bisindolylmaleimide (BIM), but not the protein kinase A (PKA) inhibitor H89. In particular, the inhibitor of PKCepsilon, a PKC isoform, completely blocked this effect. Under current clamp model, Sar-SP reduced the amount of current required to evoke action potentials and increased the firing rate in a subgroup of DRG neurons. These data suggest that activation of NK-1 receptor potentiates Na(v)1.8 sodium current via PKCepsilon-dependent signaling pathway, probably participating in the generation of inflammatory hyperalgesia.

Potentiating role of interleukin-1beta (IL-1beta) and IL-1beta type 1 receptors in the medial hypothalamus in defensive rage behavior in the cat.

PubMed

Hassanain, M; Bhatt, S; Zalcman, S; Siegel, A

2005-06-28

Recently, this laboratory provided evidence that interleukin-1beta (IL-1beta), an immune and brain-derived cytokine, microinjected into the medial hypothalamus, potentiates defensive rage behavior in the cat elicited from the midbrain periaqueductal gray (PAG), and that such effects are blocked by a 5-HT2 receptor antagonist. Since this finding represents the first time that a brain cytokine has been shown to affect defensive rage behavior, the present study replicated and extended these findings by documenting the specific potentiating role played by IL-1beta Type 1 receptor (IL-1RI), and the anatomical relationship between IL-1beta and 5-HT2 receptors in the medial hypothalamus. IL-1beta (10 ng) microinjected into the medial hypothalamus induced two separate phases of facilitation, one at 60 min and another at 180 min, post-injection. In turn, these effects were blocked with pretreatment of the selective IL-1 Type I receptor antagonist (IL-1ra) (10 ng), demonstrating the selectivity of the effects of IL-1beta on medial hypothalamic neurons upon PAG-elicited defensive rage behavior. The next stage of the study utilized immunohistochemical methods to demonstrate that IL-1beta and 5-HT2 receptors were present on the same neurons within regions of the medial hypothalamus where IL-1beta and the IL-1beta receptor antagonists were administered. This provided anatomical evidence suggesting a relationship between IL-1RI and 5-HT2 receptors in the medial hypothalamus that is consistent with the previous pharmacological observations in our laboratory. The overall findings show that activation of IL-1RI in the medial hypothalamus potentiates defensive rage behavior in the cat and that these effects may also be linked to the presence of 5-HT2 receptors on the same groups of neurons in this region of hypothalamus.

G-Protein-coupled receptors as potential drug candidates in preeclampsia: targeting the relaxin/insulin-like family peptide receptor 1 for treatment and prevention.

PubMed

Conrad, Kirk P

2016-09-01

expected to (i) enhance endothelial nitric oxide synthesis and bioactivity, as well as directly reduce vascular smooth muscle cytosolic calcium, thus promoting vasodilation; (ii) improve the local angiogenic balance by augmenting arterial vascular endothelial and placental growth factor (VEGF and PLGF) activities; (iii) ameliorate vascular inflammation; (iv) enhance placental peroxisome proliferator-activated receptor gamma, coactivator 1 alpha (PCG1α) expression, and hence, peroxisome proliferator-activated receptor gamma (PPAR-γ) activity and (v) confer cytotrophoblast and endothelial cytoprotection. Insofar as impaired endometrial maturation (decidualization) predisposes to the development of preeclampsia, relaxin administration in the late secretory phase and during early pregnancy would be anticipated to improve decidualization, and hence trophoblast invasion and spiral artery remodeling, thereby reducing the risk of preeclampsia. Relaxin has a favorable safety profile both in the non-pregnant condition and during pregnancy. There is a strong scientific rationale for RXFP1 activation in severe preeclampsia by administration of relaxin, relaxin analogs or small molecule mimetics, in order to mollify the disease pathogenesis for safe prolongation of pregnancy, thus allowing time for more complete fetal maturation, which is a primary therapeutic endpoint in treating the disease. In light of recent data implicating deficient or defective decidualization as a potential etiological factor in preeclampsia and the capacity of relaxin to promote endometrial maturation, the prophylactic application of relaxin to reduce the risk of preeclampsia is a plausible therapeutic approach to consider. Finally, given its pleiotropic and beneficial attributes particularly in the cardiovascular system, relaxin, although traditionally considered as a 'pregnancy' hormone, is likely to prove salutary for several disease indications in the non-pregnant population. © The Author 2016

G-Protein-coupled receptors as potential drug candidates in preeclampsia: targeting the relaxin/insulin-like family peptide receptor 1 for treatment and prevention

PubMed Central

Conrad, Kirk P.

2016-01-01

attributes in preeclampsia, relaxin supplementation is expected to (i) enhance endothelial nitric oxide synthesis and bioactivity, as well as directly reduce vascular smooth muscle cytosolic calcium, thus promoting vasodilation; (ii) improve the local angiogenic balance by augmenting arterial vascular endothelial and placental growth factor (VEGF and PLGF) activities; (iii) ameliorate vascular inflammation; (iv) enhance placental peroxisome proliferator-activated receptor gamma, coactivator 1 alpha (PCG1α) expression, and hence, peroxisome proliferator-activated receptor gamma (PPAR-γ) activity and (v) confer cytotrophoblast and endothelial cytoprotection. Insofar as impaired endometrial maturation (decidualization) predisposes to the development of preeclampsia, relaxin administration in the late secretory phase and during early pregnancy would be anticipated to improve decidualization, and hence trophoblast invasion and spiral artery remodeling, thereby reducing the risk of preeclampsia. Relaxin has a favorable safety profile both in the non-pregnant condition and during pregnancy. WIDER IMPLICATIONS There is a strong scientific rationale for RXFP1 activation in severe preeclampsia by administration of relaxin, relaxin analogs or small molecule mimetics, in order to mollify the disease pathogenesis for safe prolongation of pregnancy, thus allowing time for more complete fetal maturation, which is a primary therapeutic endpoint in treating the disease. In light of recent data implicating deficient or defective decidualization as a potential etiological factor in preeclampsia and the capacity of relaxin to promote endometrial maturation, the prophylactic application of relaxin to reduce the risk of preeclampsia is a plausible therapeutic approach to consider. Finally, given its pleiotropic and beneficial attributes particularly in the cardiovascular system, relaxin, although traditionally considered as a ‘pregnancy’ hormone, is likely to prove salutary for several

Signal transduction by beta1 integrin receptors in human chondrocytes in vitro: collaboration with the insulin-like growth factor-I receptor.

PubMed

Shakibaei, M; John, T; De Souza, P; Rahmanzadeh, R; Merker, H J

1999-09-15

We have examined the mechanism by which collagen-binding integrins co-operate with insulin-like growth factor-I (IGF-I) receptors (IGF-IR) to regulate chondrocyte phenotype and differentiation. Adhesion of chondrocytes to anti-beta1 integrin antibodies or collagen type II leads to phosphorylation of cytoskeletal and signalling proteins localized at focal adhesions, including alpha-actinin, vinculin, paxillin and focal adhesion kinase (FAK). These stimulate docking proteins such as Shc (Src-homology collagen). Moreover, exposure of collagen type II-cultured chondrocytes to IGF-I leads to co-immunoprecipitation of Shc protein with the IGF-IR and with beta1, alpha1 and alpha5 integrins, but not with alpha3 integrin. Shc then associates with growth factor receptor-bound protein 2 (Grb2), an adaptor protein and extracellular signal-regulated kinase. The expression of the docking protein Shc occurs only when chondrocytes are bound to collagen type II or integrin antibodies and increases when IGF-I is added, suggesting a collaboration between integrins and growth factors in a common/shared biochemical signalling pathway. Furthermore, these results indicate that focal adhesion assembly may facilitate signalling via Shc, a potential common target for signal integration between integrin and growth-factor signalling regulatory pathways. Thus, the collagen-binding integrins and IGF-IR co-operate to regulate focal adhesion components and these signalling pathways have common targets (Shc-Grb2 complex) in subcellular compartments, thereby linking to the Ras-mitogen-activated protein kinase signalling pathway. These events may play a role during chondrocyte differentiation.

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

PubMed Central

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

2016-01-01

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. PMID:26749475

Up-regulation of proproliferative genes and the ligand/receptor pair placental growth factor and vascular endothelial growth factor receptor 1 in hepatitis C cirrhosis.

PubMed

Huang, Xiao X; McCaughan, Geoffrey W; Shackel, Nicholas A; Gorrell, Mark D

2007-09-01

Cirrhosis can lead to hepatocellular carcinoma (HCC). Non-diseased liver and hepatitis C virus (HCV)-associated cirrhosis with or without HCC were compared. Proliferation pathway genes, immune response genes and oncogenes were analysed by a quantitative real-time reverse transcriptase-polymerase chain reaction (RT-PCR) and immunostaining. Real-time RT-PCR showed up-regulation of genes in HCV cirrhosis including the proliferation-associated genes bone morphogenetic protein 3 (BMP3), placental growth factor 3 (PGF3), vascular endothelial growth factor receptor 1 (VEGFR1) and soluble VEGFR1, the oncogene FYN, and the immune response-associated genes toll-like receptor 9 (TLR9) and natural killer cell transcript 4 (NK4). Expressions of TLR2 and the oncogenes B-cell CLL/lymphoma 9 (BCL9) and PIM2 were decreased in HCV cirrhosis. In addition, PIM2 and TLR2 were increased in HCV cirrhosis with HCC compared with HCV cirrhosis. The ligand/receptor pair PGF and VEGFR1 was intensely expressed by the portal tract vascular endothelium. VEGFR1 was expressed in reactive biliary epithelial structures in fibrotic septum and in some stellate cells and macrophages. PGF and VEGFR1 may have an important role in the pathogenesis of the neovascular response in cirrhosis.

Components of the CCR4-NOT complex function as nuclear hormone receptor coactivators via association with the NRC-interacting Factor NIF-1.

PubMed

Garapaty, Shivani; Mahajan, Muktar A; Samuels, Herbert H

2008-03-14

CCR4-NOT is an evolutionarily conserved, multicomponent complex known to be involved in transcription as well as mRNA degradation. Various subunits (e.g. CNOT1 and CNOT7/CAF1) have been reported to be involved in influencing nuclear hormone receptor activities. Here, we show that CCR4/CNOT6 and RCD1/CNOT9, members of the CCR4-NOT complex, potentiate nuclear receptor activity. RCD1 interacts in vivo and in vitro with NIF-1 (NRC-interacting factor), a previously characterized nuclear receptor cotransducer that activates nuclear receptors via its interaction with NRC. As with NIF-1, RCD1 and CCR4 do not directly associate with nuclear receptors; however, they enhance ligand-dependent transcriptional activation by nuclear hormone receptors. CCR4 mediates its effect through the ligand binding domain of nuclear receptors and small interference RNA-mediated silencing of endogenous CCR4 results in a marked decrease in nuclear receptor activation. Furthermore, knockdown of CCR4 results in an attenuated stimulation of RARalpha target genes (e.g. Sox9 and HoxA1) as shown by quantitative PCR assays. The silencing of endogenous NIF-1 also resulted in a comparable decrease in the RAR-mediated induction of both Sox9 and HoxA1. Furthermore, CCR4 associates in vivo with NIF-1. In addition, the CCR4-enhanced transcriptional activation by nuclear receptors is dependent on NIF-1. The small interference RNA-mediated knockdown of NIF-1 blocks the ligand-dependent potentiating effect of CCR4. Our results suggest that CCR4 plays a role in the regulation of certain endogenous RARalpha target genes and that RCD1 and CCR4 might mediate their function through their interaction with NIF-1.

Differential roles of vascular endothelial growth factor receptors 1 and 2 in dendritic cell differentiation.

PubMed

Dikov, Mikhail M; Ohm, Joyce E; Ray, Neelanjan; Tchekneva, Elena E; Burlison, Jared; Moghanaki, Drew; Nadaf, Sorena; Carbone, David P

2005-01-01

Impaired Ag-presenting function in dendritic cells (DCs) due to abnormal differentiation is an important mechanism of tumor escape from immune control. A major role for vascular endothelial growth factor (VEGF) and its receptors, VEGFR1/Flt-1 and VEGFR2/KDR/Flk-1, has been documented in hemopoietic development. To study the roles of each of these receptors in DC differentiation, we used an in vitro system of myeloid DC differentiation from murine embryonic stem cells. Exposure of wild-type, VEGFR1(-/-), or VEGFR2(-/-) embryonic stem cells to exogenous VEGF or the VEGFR1-specific ligand, placental growth factor, revealed distinct roles of VEGF receptors. VEGFR1 is the primary mediator of the VEGF inhibition of DC maturation, whereas VEGFR2 tyrosine kinase signaling is essential for early hemopoietic differentiation, but only marginally affects final DC maturation. SU5416, a VEGF receptor tyrosine kinase inhibitor, only partially rescued the mature DC phenotype in the presence of VEGF, suggesting the involvement of both tyrosine kinase-dependent and independent inhibitory mechanisms. VEGFR1 signaling was sufficient for blocking NF-kappaB activation in bone marrow hemopoietic progenitor cells. VEGF and placental growth factor affect the early stages of myeloid/DC differentiation. The data suggest that therapeutic strategies attempting to reverse the immunosuppressive effects of VEGF in cancer patients might be more effective if they specifically targeted VEGFR1.

Fibroblast growth factor receptors, developmental corruption and malignant disease.

PubMed

Kelleher, Fergal C; O'Sullivan, Hazel; Smyth, Elizabeth; McDermott, Ray; Viterbo, Antonella

2013-10-01

Fibroblast growth factors (FGF) are a family of ligands that bind to four different types of cell surface receptor entitled, FGFR1, FGFR2, FGFR3 and FGFR4. These receptors differ in their ligand binding affinity and tissue distribution. The prototypical receptor structure is that of an extracellular region comprising three immunoglobulin (Ig)-like domains, a hydrophobic transmembrane segment and a split intracellular tyrosine kinase domain. Alternative gene splicing affecting the extracellular third Ig loop also creates different receptor isoforms entitled FGFRIIIb and FGFRIIIc. Somatic fibroblast growth factor receptor (FGFR) mutations are implicated in different types of cancer and germline FGFR mutations occur in developmental syndromes particularly those in which craniosynostosis is a feature. The mutations found in both conditions are often identical. Many somatic FGFR mutations in cancer are gain-of-function mutations of established preclinical oncogenic potential. Gene amplification can also occur with 19-22% of squamous cell lung cancers for example having amplification of FGFR1. Ontologic comparators can be informative such as aberrant spermatogenesis being implicated in both spermatocytic seminomas and Apert syndrome. The former arises from somatic FGFR3 mutations and Apert syndrome arises from germline FGFR2 mutations. Finally, therapeutics directed at inhibiting the FGF/FGFR interaction are a promising subject for clinical trials.

Interactions of phosphatidylinositol kinase, GTPase-activating protein (GAP), and GAP-associated proteins with the colony-stimulating factor 1 receptor.

PubMed Central

Reedijk, M; Liu, X Q; Pawson, T

1990-01-01

The interactions of the macrophage colony-stimulating factor 1 (CSF-1) receptor with potential targets were investigated after ligand stimulation either of mouse macrophages or of fibroblasts that ectopically express mouse CSF-1 receptors. In Rat-2 cells expressing the mouse CSF-1 receptor, full activation of the receptor and cellular transformation require exogenous CSF-1, whereas NIH 3T3 cells expressing mouse c-fms are transformed by autocrine stimulation. Activated CSF-1 receptors physically associate with a phosphatidylinositol (PI) 3'-kinase. A mutant CSF-1 receptor with a deletion of the kinase insert region was deficient in its ability to bind functional PI 3'-kinase and to induce PI 3'-kinase activity precipitable with antiphosphotyrosine antibodies. In fibroblasts, CSF-1 stimulation also induced the phosphorylation of the GTPase-activating protein (GAP)-associated protein p62 on tyrosine, although GAP itself was a relatively poor substrate. In contrast to PI 3'-kinase association, phosphorylation of p62 and GAP was not markedly affected by deletion of the kinase insert region. These results indicate that the kinase insert region selectively enhances the CSF-1-dependent association of the CSF-1 receptor with active PI 3'-kinase. The insert deletion mutant retains considerable transforming activity in NIH 3T3 cells (G. Taylor, M. Reedijk, V. Rothwell, L. Rohrschneider, and T. Pawson, EMBO J. 8:2029-2037, 1989). This mutant was more seriously impaired in Rat-2 cell transformation, although mutant-expressing Rat-2 cells still formed small colonies in soft agar in the presence of CSF-1. Therefore, phosphorylation of GAP and p62 through activation of the CSF-1 receptor does not result in full fibroblast transformation. The interaction between the CSF-1 receptor and PI 3'-kinase may contribute to c-fms fibroblast transformation and play a role in CSF-1-stimulated macrophages. Images PMID:2172781

Increased tumor necrosis factor receptor 1 expression in human colorectal adenomas

PubMed Central

Hosono, Kunihiro; Yamada, Eiji; Endo, Hiroki; Takahashi, Hirokazu; Inamori, Masahiko; Hippo, Yoshitaka; Nakagama, Hitoshi; Nakajima, Atsushi

2012-01-01

AIM: To determine the expression statuses of tumor necrosis factor (TNF)-α, its receptors (TNF-R) and downstream effector molecules in human colorectal adenomas. METHODS: We measured the serum concentrations of TNF-α and its receptors in 62 colorectal adenoma patients and 34 healthy controls. The protein expression of TNF-α, TNF-R1, TNF-R2 and downstream signals of the TNF receptors, such as c-Jun N-terminal kinase (JNK), nuclear factor-κ B and caspase-3, were also investigated in human colorectal adenomas and in normal colorectal mucosal tissues by immunohistochemistry. Immunofluorescence confocal microscopy was used to investigate the consistency of expression of TNF-R1 and phospho-JNK (p-JNK). RESULTS: The serum levels of soluble TNF-R1 (sTNF-R1) in adenoma patients were significantly higher than in the control group (3.67 ± 0.86 ng/mL vs 1.57 ± 0.72 ng/mL, P < 0.001). Receiver operating characteristic analysis revealed the high diagnostic sensitivity of TNF-R1 measurements (AUC was 0.928) for the diagnosis of adenoma, and the best cut-off level of TNF-R1 was 2.08 ng/mL, with a sensitivity of 93.4% and a specificity of 82.4%. There were no significant differences in the serum levels of TNF-α or sTNF-R2 between the two groups. Immunohistochemistry showed high levels of TNF-R1 and p-JNK expression in the epithelial cells of adenomas. Furthermore, a high incidence of co-localization of TNF-R1 and p-JNK was identified in adenoma tissue. CONCLUSION: TNF-R1 may be a promising biomarker of colorectal adenoma, and it may also play an important role in the very early stages of colorectal carcinogenesis. PMID:23082052

Phthalocyanine-Peptide Conjugates for Epidermal Growth Factor Receptor Targeting1

PubMed Central

Ongarora, Benson G.; Fontenot, Krystal R.; Hu, Xiaoke; Sehgal, Inder; Satyanarayana-Jois, Seetharama D.; Vicente, M. Graça H.

2012-01-01

Four phthalocyanine (Pc)-peptide conjugates designed to target the epidermal growth factor receptor (EGFR) were synthesized and evaluated in vitro using four cell lines: human carcinoma A431 and HEp2, human colorectal HT-29, and kidney Vero (negative control) cells. Two peptide ligands for EGFR were investigated: EGFR-L1 and -L2, bearing 6 and 13 amino acid residues, respectively. The peptides and Pc-conjugates were shown to bind to EGFR using both theoretical (Autodock) and experimental (SPR) investigations. The Pc-EGFR-L1 conjugates 5a and 5b efficiently targeted EGFR and were internalized, in part due to their cationic charge, whereas the uncharged Pc-EGFR-L2 conjugates 4b and 6a poorly targeted EGFR maybe due to their low aqueous solubility. All conjugates were non-toxic (IC50 > 100 µM) to HT-29 cells, both in the dark and upon light activation (1 J/cm2). Intravenous (iv) administration of conjugate 5b into nude mice bearing A431 and HT-29 human tumor xenografts resulted in a near-IR fluorescence signal at ca. 700 nm, 24 h after administration. Our studies show that Pc-EGFR-L1 conjugates are promising near-IR fluorescent contrast agents for CRC, and potentially other EGFR over-expressing cancers. PMID:22468711

Cyclotides Isolated from an Ipecac Root Extract Antagonize the Corticotropin Releasing Factor Type 1 Receptor

PubMed Central

Fahradpour, Mohsen; Keov, Peter; Tognola, Carlotta; Perez-Santamarina, Estela; McCormick, Peter J.; Ghassempour, Alireza; Gruber, Christian W.

2017-01-01

Cyclotides are plant derived, cystine-knot stabilized peptides characterized by their natural abundance, sequence variability and structural plasticity. They are abundantly expressed in Rubiaceae, Psychotrieae in particular. Previously the cyclotide kalata B7 was identified to modulate the human oxytocin and vasopressin G protein-coupled receptors (GPCRs), providing molecular validation of the plants’ uterotonic properties and further establishing cyclotides as valuable source for GPCR ligand design. In this study we screened a cyclotide extract derived from the root powder of the South American medicinal plant ipecac (Carapichea ipecacuanha) for its GPCR modulating activity of the corticotropin-releasing factor type 1 receptor (CRF1R). We identified and characterized seven novel cyclotides. One cyclotide, caripe 8, isolated from the most active fraction, was further analyzed and found to antagonize the CRF1R. A nanomolar concentration of this cyclotide (260 nM) reduced CRF potency by ∼4.5-fold. In contrast, caripe 8 did not inhibit forskolin-, or vasopressin-stimulated cAMP responses at the vasopressin V2 receptor, suggesting a CRF1R-specific mode-of-action. These results in conjunction with our previous findings establish cyclotides as modulators of both classes A and B GPCRs. Given the diversity of cyclotides, our data point to other cyclotide-GPCR interactions as potentially important sources of drug-like molecules. PMID:29033832

The first three domains of the insulin receptor differ structurally from the insulin-like growth factor 1 receptor in the regions governing ligand specificity

PubMed Central

Lou, Meizhen; Garrett, Thomas P. J.; McKern, Neil M.; Hoyne, Peter A.; Epa, V. Chandana; Bentley, John D.; Lovrecz, George O.; Cosgrove, Leah J.; Frenkel, Maurice J.; Ward, Colin W.

2006-01-01

The insulin receptor (IR) and the type-1 insulin-like growth factor receptor (IGF1R) are homologous multidomain proteins that bind insulin and IGF with differing specificity. Here we report the crystal structure of the first three domains (L1–CR–L2) of human IR at 2.3 Å resolution and compare it with the previously determined structure of the corresponding fragment of IGF1R. The most important differences seen between the two receptors are in the two regions governing ligand specificity. The first is at the corner of the ligand-binding surface of the L1 domain, where the side chain of F39 in IR forms part of the ligand binding surface involving the second (central) β-sheet. This is very different to the location of its counterpart in IGF1R, S35, which is not involved in ligand binding. The second major difference is in the sixth module of the CR domain, where IR contains a larger loop that protrudes further into the ligand-binding pocket. This module, which governs IGF1-binding specificity, shows negligible sequence identity, significantly more α-helix, an additional disulfide bond, and opposite electrostatic potential compared to that of the IGF1R. PMID:16894147

Odontoblasts as sensory receptors: transient receptor potential channels, pannexin-1, and ionotropic ATP receptors mediate intercellular odontoblast-neuron signal transduction.

PubMed

Shibukawa, Yoshiyuki; Sato, Masaki; Kimura, Maki; Sobhan, Ubaidus; Shimada, Miyuki; Nishiyama, Akihiro; Kawaguchi, Aya; Soya, Manabu; Kuroda, Hidetaka; Katakura, Akira; Ichinohe, Tatsuya; Tazaki, Masakazu

2015-04-01

Various stimuli induce pain when applied to the surface of exposed dentin. However, the mechanisms underlying dentinal pain remain unclear. We investigated intercellular signal transduction between odontoblasts and trigeminal ganglion (TG) neurons following direct mechanical stimulation of odontoblasts. Mechanical stimulation of single odontoblasts increased the intracellular free calcium concentration ([Ca(2+)]i) by activating the mechanosensitive-transient receptor potential (TRP) channels TRPV1, TRPV2, TRPV4, and TRPA1, but not TRPM8 channels. In cocultures of odontoblasts and TG neurons, increases in [Ca(2+)]i were observed not only in mechanically stimulated odontoblasts, but also in neighboring odontoblasts and TG neurons. These increases in [Ca(2+)]i were abolished in the absence of extracellular Ca(2+) and in the presence of mechanosensitive TRP channel antagonists. A pannexin-1 (ATP-permeable channel) inhibitor and ATP-degrading enzyme abolished the increases in [Ca(2+)]i in neighboring odontoblasts and TG neurons, but not in the stimulated odontoblasts. G-protein-coupled P2Y nucleotide receptor antagonists also inhibited the increases in [Ca(2+)]i. An ionotropic ATP (P2X3) receptor antagonist inhibited the increase in [Ca(2+)]i in neighboring TG neurons, but not in stimulated or neighboring odontoblasts. During mechanical stimulation of single odontoblasts, a connexin-43 blocker did not have any effects on the [Ca(2+)]i responses observed in any of the cells. These results indicate that ATP, released from mechanically stimulated odontoblasts via pannexin-1 in response to TRP channel activation, transmits a signal to P2X3 receptors on TG neurons. We suggest that odontoblasts are sensory receptor cells and that ATP released from odontoblasts functions as a neurotransmitter in the sensory transduction sequence for dentinal pain.

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

PubMed

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.

Activation of Muscarinic M1 Acetylcholine Receptors Induces Long-Term Potentiation in the Hippocampus

PubMed Central

Dennis, Siobhan H.; Pasqui, Francesca; Colvin, Ellen M.; Sanger, Helen; Mogg, Adrian J.; Felder, Christian C.; Broad, Lisa M.; Fitzjohn, Steve M.; Isaac, John T.R.; Mellor, Jack R.

2016-01-01

Muscarinic M1 acetylcholine receptors (M1Rs) are highly expressed in the hippocampus, and their inhibition or ablation disrupts the encoding of spatial memory. It has been hypothesized that the principal mechanism by which M1Rs influence spatial memory is by the regulation of hippocampal synaptic plasticity. Here, we use a combination of recently developed, well characterized, selective M1R agonists and M1R knock-out mice to define the roles of M1Rs in the regulation of hippocampal neuronal and synaptic function. We confirm that M1R activation increases input resistance and depolarizes hippocampal CA1 pyramidal neurons and show that this profoundly increases excitatory postsynaptic potential-spike coupling. Consistent with a critical role for M1Rs in synaptic plasticity, we now show that M1R activation produces a robust potentiation of glutamatergic synaptic transmission onto CA1 pyramidal neurons that has all the hallmarks of long-term potentiation (LTP): The potentiation requires NMDA receptor activity and bi-directionally occludes with synaptically induced LTP. Thus, we describe synergistic mechanisms by which acetylcholine acting through M1Rs excites CA1 pyramidal neurons and induces LTP, to profoundly increase activation of CA1 pyramidal neurons. These features are predicted to make a major contribution to the pro-cognitive effects of cholinergic transmission in rodents and humans. PMID:26472558

Equine insulin receptor and insulin-like growth factor-1 receptor expression in digital lamellar tissue and insulin target tissues.

PubMed

Kullmann, A; Weber, P S; Bishop, J B; Roux, T M; Norby, B; Burns, T A; McCutcheon, L J; Belknap, J K; Geor, R J

2016-09-01

Hyperinsulinaemia is implicated in the pathogenesis of endocrinopathic laminitis. Insulin can bind to different receptors: two insulin receptor isoforms (InsR-A and InsR-B), insulin-like growth factor-1 receptor (IGF-1R) and InsR/IGF-1R hybrid receptor (Hybrid). Currently, mRNA expression of these receptors in equine tissues and the influence of body type and dietary carbohydrate intake on expression of these receptors is not known. The study objectives were to characterise InsR-A, InsR-B, IGF-1R and Hybrid expression in lamellar tissue (LT) and insulin responsive tissues from horses and examine the effect of dietary nonstructural carbohydrate (NSC) on mRNA expression of these receptors in LT, skeletal muscle, liver and two adipose tissue (AT) depots of lean and obese ponies. In vivo experiment. Lamellar tissue samples were evaluated by quantitative reverse transcription polymerase chain reaction (RT-qPCR) for receptor mRNA expression (n = 8) and immunoblotting for protein expression (n = 3). Archived LT, skeletal muscle, liver and AT from lean and obese mixed-breed ponies fed either a low (~7% NSC as dry matter; 5 lean, 5 obese) or high NSC diet (~42% NSC as dry matter; 6 lean, 6 obese) for 7 days were evaluated by RT-qPCR to determine the effect of body condition and diet on expression of the receptors in different tissues. Significance was set at P≤0.05. Lamellar tissue expresses both InsR isoforms, IGF-1R and Hybrid. LT IGF-1R gene expression was greater than either InsR isoform and InsR-A expression was greater than InsR-B (P≤0.05). Obesity significantly lowered IGF-1R, InsR-A and InsR-B mRNA expression in LT and InsR-A in tailhead AT. High NSC diet lowered expression of all three receptor types in liver; IGF-1R and InsR-A in LT and InsR-A in tailhead AT. Lamellar tissue expresses IGF-1R, InsR isoforms and Hybrids. The functional characteristics of these receptors and their role in endocrinopathic laminitis warrants further investigation. © 2015 EVJ

Insulin-like growth factor-1 inhibits adult supraoptic neurons via complementary modulation of mechanoreceptors and glycine receptors.

PubMed

Ster, Jeanne; Colomer, Claude; Monzo, Cécile; Duvoid-Guillou, Anne; Moos, Françoise; Alonso, Gérard; Hussy, Nicolas

2005-03-02

In the CNS, insulin-like growth factor-1 (IGF-1) is mainly known for its trophic effect both during development and in adulthood. Here, we show than in adult rat supraoptic nucleus (SON), IGF-1 receptor immunoreactivity is present in neurons, whereas IGF-1 immunoreactivity is found principally in astrocytes and more moderately in neurons. In vivo application of IGF-1 within the SON acutely inhibits the activity of both vasopressin and oxytocin neurons, the two populations of SON neuroendocrine cells. Recordings of acutely isolated SON neurons showed that this inhibition occurs through two rapid and reversible mechanisms, both involving the neuronal IGF-1 receptor but different intracellular messengers. IGF-1 inhibits Gd3+-sensitive and osmosensitive mechanoreceptor cation current via phosphatidylinositol-3 (PI3) kinase activation. IGF-1 also potentiates taurine-activated glycine receptor (GlyR) Cl- currents by increasing the agonist sensitivity through a extremely rapid (within a second) PI3 kinase-independent mechanism. Both mechanoreceptor channels and GlyR, which form the excitatory and inhibitory components of SON neuron osmosensitivity, are active at rest, and their respective inhibition and potentiation will both be inhibitory, leading to strong decrease in neuronal activity. It will be of interest to determine whether IGF-1 is released by neurons, thus participating in an inhibitory autocontrol, or astrocytes, then joining the growing family of glia-to-neuron transmitters that modulate neuronal and synaptic activity. Through the opposite and complementary acute regulation of mechanoreceptors and GlyR, IGF-1 appears as a new important neuromodulator in the adult CNS, participating in the complex integration of neural messages that regulates the level of neuronal excitability.

Tumor Necrosis Factor Receptor Levels Are Associated With Carotid Atherosclerosis

PubMed Central

Elkind, Mitchell S.; Cheng, Jianfeng; Boden-Albala, Bernadette; Rundek, Tanja; Thomas, Joyce; Chen, Hong; Rabbani, LeRoy E.; Sacco, Ralph L.

2009-01-01

Background and Purpose Recent evidence suggests that atherosclerosis is an inflammatory condition. Serum levels of inflammatory markers may serve as measures of the severity of atherosclerosis and risk of stroke. We sought to determine whether tumor necrosis factor-α (TNF-α) and TNF receptor levels are associated with carotid plaque thickness. Methods The Northern Manhattan Stroke Study is a community-based study of stroke risk factors. For this cross-sectional analysis, inflammatory marker levels, including TNF-α and TNF receptors 1 and 2, were measured by immunoassay in stroke-free community subjects undergoing carotid duplex Doppler ultrasound. Maximal carotid plaque thickness (MCPT) was measured for each subject. Analyses were stratified by age <70 and ≥70 years. Simple and multiple linear regression analyses were used to calculate the association between marker levels and MCPT. Multiple logistic regression was used to calculate odds ratios and 95% CIs for the association of inflammatory markers with MCPT ≥1.5 mm (>75th percentile), after adjustment for demographic and potential medical confounding factors. Results The mean age of the 279 subjects was 67.6±8.5 years; 49% were men; 63% were Hispanic, 17% black, and 17% white. Mean values for TNF-α and its receptors were as follows: TNF-α, 1.88±3.97 ng/mL; TNF receptor 1, 2.21±0.99 ng/mL; and TNF receptor 2, 4.85±2.23 ng/mL. Mean MCPT was elevated in those in the highest quartiles compared with lowest quartiles of TNF receptor 1 and 2 (1.24 versus 0.79 mm and 1.23 versus 0.80 mm, respectively). Among those aged <70 years, TNF receptor 1 and 2 were associated with an increase in MCPT (mean difference=0.36 mm, P=0.01 for TNF receptor 1 and mean difference=0.10 mm, P=0.04 for TNF receptor 2). After adjustment for sex, race-ethnicity, hypertension, diabetes mellitus, LDL cholesterol, smoking, and body mass index, associations remained (mean difference=0.36 mm, P=0.001 for TNF receptor 1 and mean

Potential therapeutic value of TRPV1 and TRPA1 in diabetes mellitus and obesity.

PubMed

Derbenev, Andrei V; Zsombok, Andrea

2016-05-01

Diabetes mellitus and obesity, which is a major risk factor in the development of type 2 diabetes mellitus, have reached epidemic proportions worldwide including the USA. The current statistics and forecasts, both short- and long-term, are alarming and predict severe problems in the near future. Therefore, there is a race for developing new compounds, discovering new receptors, or finding alternative solutions to prevent and/or treat the symptoms and complications related to obesity and diabetes mellitus. It is well demonstrated that members of the transient receptor potential (TRP) superfamily play a crucial role in a variety of biological functions both in health and disease. In the recent years, transient receptor potential vanilloid type 1 (TRPV1) and transient receptor potential ankyrin 1 (TRPA1) were shown to have beneficial effects on whole body metabolism including glucose homeostasis. TRPV1 and TRPA1 have been associated with control of weight, pancreatic function, hormone secretion, thermogenesis, and neuronal function, which suggest a potential therapeutic value of these channels. This review summarizes recent findings regarding TRPV1 and TRPA1 in association with whole body metabolism with emphasis on obese and diabetic conditions.

L1CAM stimulates glioma cell motility and proliferation through the fibroblast growth factor receptor.

PubMed

Mohanan, Vishnu; Temburni, Murali K; Kappes, John C; Galileo, Deni S

2013-04-01

The L1CAM cell adhesion/recognition molecule (L1, CD171) and fibroblast growth factor receptor (FGFR) both are expressed by human high-grade glioma cells, but their potential actions in controlling cell behavior have not been linked. L1 actions in cancer cells have been attributed mainly to integrin receptors, and we demonstrated previously that L1-stimulated glioma cell migration correlates with integrin expression, increased focal adhesion kinase activation and focal complex turnover. Our analyses of datasets revealed FGFR is overexpressed in glioma regardless of grade, while ADAM10 metalloprotease expression increases with glioma grade. Here, we used dominant-negative and short hairpin RNA approaches to inhibit the activation of FGFR1 and expression of L1, respectively. An L1 peptide that inhibits L1-FGFR interaction and PD173074, a chemical inhibitor of FGFR1 activity, also were used to elucidate the involvement of L1-FGFR interactions on glioma cell behavior. Time-lapse cell motility studies and flow cytometry cell cycle analyses showed that L1 operates to increase glioma cell motility and proliferation through FGFR activation. Shutdown of both L1 expression and FGFR activity in glioma cells resulted in a complete termination of cell migration in vitro. These studies show for the first time that soluble L1 ectodomain (L1LE) acts on glioma cells through FGFRs, and that FGFRs are used by glioma cells for increasing motility as well as proliferation in response to activation by L1LE ligand. Thus, effective treatment of high-grade glioma may require simultaneous targeting of L1, FGFRs, and integrin receptors, which would reduce glioma cell motility as well as proliferation.

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

PubMed

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.

Prostaglandin E₂ regulates cellular migration via induction of vascular endothelial growth factor receptor-1 in HCA-7 human colon cancer cells.

PubMed

Fujino, Hiromichi; Toyomura, Kaori; Chen, Xiao-bo; Regan, John W; Murayama, Toshihiko

2011-02-01

An important event in the development of tumors is angiogenesis, or the formation of new blood vessels. Angiogenesis is also known to be involved in tumor cell metastasis and is dependent upon the activity of the vascular endothelial growth factor (VEGF) signaling pathway. Studies of mice in which the EP3 prostanoid receptors have been genetically deleted have shown a role for these receptors in cancer growth and angiogenesis. In the present study, human colon cancer HCA-7 cells were used as a model system to understand the potential role of EP3 receptors in tumor cell migration. We now show that stimulation of HCA-7 cells with PGE₂ enhanced the up-regulation of VEGF receptor-1 (VEGFR-1) expression by a mechanism involving EP3 receptor-mediated activation of phosphatidylinositol 3-kinase and the extracellular signal-regulated kinases. Moreover, the PGE₂ stimulated increase in VEGFR-1 expression was accompanied by an increase in the cellular migration of HCA-7 cells. Given the known involvement of VEGFR-1 in cellular migration, our results suggest that EP3 receptors may contribute to tumor cell metastasis by increasing cellular migration through the up-regulation of VEGFR-1 signaling. Copyright © 2010 Elsevier Inc. All rights reserved.

Mechanical stretch augments insulin-induced vascular smooth muscle cell proliferation by insulin-like growth factor-1 receptor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Gang; Department of Anesthesiology, First Affiliated Hospital of China Medical University, Shenyang; Hitomi, Hirofumi, E-mail: hitomi@kms.ac.jp

Insulin resistance and hypertension have been implicated in the pathogenesis of cardiovascular disease; however, little is known about the roles of insulin and mechanical force in vascular smooth muscle cell (VSMC) remodeling. We investigated the contribution of mechanical stretch to insulin-induced VSMC proliferation. Thymidine incorporation was stimulated by insulin in stretched VSMCs, but not in un-stretched VSMCs. Insulin increased 2-deoxy-glucose incorporation in both stretched and un-stretched VSMCs. Mechanical stretch augmented insulin-induced extracellular signal-regulated kinase (ERK) and Akt phosphorylation. Inhibitors of epidermal growth factor (EGF) receptor tyrosine kinase and Src attenuated insulin-induced ERK and Akt phosphorylation, as well as thymidine incorporation,more » whereas 2-deoxy-glucose incorporation was not affected by these inhibitors. Moreover, stretch augmented insulin-like growth factor (IGF)-1 receptor expression, although it did not alter the expression of insulin receptor and insulin receptor substrate-1. Insulin-induced ERK and Akt activation, and thymidine incorporation were inhibited by siRNA for the IGF-1 receptor. Mechanical stretch augments insulin-induced VSMC proliferation via upregulation of IGF-1 receptor, and downstream Src/EGF receptor-mediated ERK and Akt activation. Similar to in vitro experiment, IGF-1 receptor expression was also augmented in hypertensive rats. These results provide a basis for clarifying the molecular mechanisms of vascular remodeling in hypertensive patients with hyperinsulinemia. -- Highlights: {yields} Mechanical stretch augments insulin-induced VSMC proliferation via IGF-1 receptor. {yields} Src/EGFR-mediated ERK and Akt phosphorylation are augmented in stretched VSMCs. {yields} Similar to in vitro experiment, IGF-1 receptor is increased in hypertensive rats. {yields} Results provide possible mechanisms of vascular remodeling in hypertension with DM.« less

Hypoxia-inducible factor-1α in vascular smooth muscle regulates blood pressure homeostasis through a peroxisome proliferator-activated receptor-γ-angiotensin II receptor type 1 axis.

PubMed

Huang, Yan; Di Lorenzo, Annarita; Jiang, Weidong; Cantalupo, Anna; Sessa, William C; Giordano, Frank J

2013-09-01

Hypertension is a major worldwide health issue for which only a small proportion of cases have a known mechanistic pathogenesis. Of the defined causes, none have been directly linked to heightened vasoconstrictor responsiveness, despite the fact that vasomotor tone in resistance vessels is a fundamental determinant of blood pressure. Here, we reported a previously undescribed role for smooth muscle hypoxia-inducible factor-1α (HIF-1α) in controlling blood pressure homeostasis. The lack of HIF-1α in smooth muscle caused hypertension in vivo and hyperresponsiveness of resistance vessels to angiotensin II stimulation ex vivo. These data correlated with an increased expression of angiotensin II receptor type I in the vasculature. Specifically, we show that HIF-1α, through peroxisome proliferator-activated receptor-γ, reciprocally defined angiotensin II receptor type I levels in the vessel wall. Indeed, pharmacological blockade of angiotensin II receptor type I by telmisartan abolished the hypertensive phenotype in smooth muscle cell-HIF-1α-KO mice. These data revealed a determinant role of a smooth muscle HIF-1α/peroxisome proliferator-activated receptor-γ/angiotensin II receptor type I axis in controlling vasomotor responsiveness and highlighted an important pathway, the alterations of which may be critical in a variety of hypertensive-based clinical settings.

Increased Eps15 homology domain 1 and RAB11FIP3 expression regulate breast cancer progression via promoting epithelial growth factor receptor recycling.

PubMed

Tong, Dandan; Liang, Ya-Nan; Stepanova, A A; Liu, Yu; Li, Xiaobo; Wang, Letian; Zhang, Fengmin; Vasilyeva, N V

2017-02-01

Recent research indicates that the C-terminal Eps15 homology domain 1 is associated with epithelial growth factor receptor-mediated endocytosis recycling in non-small-cell lung cancer. The aim of this study was to determine the clinical significance of Eps15 homology domain 1 gene expression in relation to phosphorylation of epithelial growth factor receptor expression in patients with breast cancer. Primary breast cancer samples from 306 patients were analyzed for Eps15 homology domain 1, RAB11FIP3, and phosphorylation of epithelial growth factor receptor expression via immunohistochemistry. The clinical significance was assessed via a multivariate Cox regression analysis, Kaplan-Meier curves, and the log-rank test. Eps15 homology domain 1 and phosphorylation of epithelial growth factor receptor were upregulated in 60.46% (185/306) and 53.92% (165/306) of tumor tissues, respectively, as assessed by immunohistochemistry. The statistical correlation analysis indicated that Eps15 homology domain 1 overexpression was positively correlated with the increases in phosphorylation of epithelial growth factor receptor ( r = 0.242, p < 0.001) and RAB11FIP3 ( r = 0.165, p = 0.005) expression. The multivariate Cox proportional hazard model analysis demonstrated that the expression of Eps15 homology domain 1 alone is a significant prognostic marker of breast cancer for the overall survival in the total, chemotherapy, and human epidermal growth factor receptor 2 (-) groups. However, the use of combined expression of Eps15 homology domain 1 and phosphorylation of epithelial growth factor receptor markers is more effective for the disease-free survival in the overall population, chemotherapy, and human epidermal growth factor receptor 2 (-) groups. Moreover, the combined markers are also significant prognostic markers of breast cancer in the human epidermal growth factor receptor 2 (+), estrogen receptor (+), and estrogen receptor (-) groups. Eps15 homology domain

Heritability of Transforming Growth Factor-β1 and Tumor Necrosis Factor-Receptor Type 1 Expression and Vitamin D Levels in Healthy Adolescent Twins.

PubMed

Mills, Natalie T; Wright, Margie J; Henders, Anjali K; Eyles, Darryl W; Baune, Bernhard T; McGrath, John J; Byrne, Enda M; Hansell, Narelle K; Birosova, Eva; Scott, James G; Martin, Nicholas G; Montgomery, Grant W; Wray, Naomi R; Vinkhuyzen, Anna A E

2015-02-01

Cytokines and vitamin D both have a role in modulating the immune system, and are also potentially useful biomarkers in mental illnesses such as major depressive disorder (MDD) and schizophrenia. Studying the variability of cytokines and vitamin D in a healthy population sample may add to understanding the association between these biomarkers and mental illness. To assess genetic and environmental contributions to variation in circulating levels of cytokines and vitamin D (25-hydroxy vitamin D: 25(OH)D3), we analyzed data from a healthy adolescent twin cohort (mean age 16.2 years; standard deviation 0.25). Plasma cytokine measures were available for 400 individuals (85 MZ, 115 DZ pairs), dried blood spot sample vitamin D measures were available for 378 individuals (70 MZ, 118 DZ pairs). Heritability estimates were moderate but significant for the cytokines transforming growth factor-β1 (TGF-β1), 0.57 (95% CI 0.26-0.80) and tumor necrosis factor-receptor type 1 (TNFR1), 0.50 (95% CI 0.11-0.63) respectively. Measures of 25(OH)D3 were within normal range and heritability was estimated to be high (0.86, 95% CI 0.61-0.94). Assays of other cytokines did not generate meaningful results. These potential biomarkers may be useful in mental illness, with further research warranted in larger sample sizes. They may be particularly important in adolescents with mental illness where diagnostic uncertainty poses a significant clinical challenge.

Pharmacological activation of CB1 receptor modulates long term potentiation by interfering with protein synthesis.

PubMed

Navakkode, Sheeja; Korte, Martin

2014-04-01

Cognitive impairment is one of the most important side effects associated with cannabis drug abuse, as well as the serious issue concerning the therapeutic use of cannabinoids. Cognitive impairments and neuropsychiatric symptoms are caused by early synaptic dysfunctions, such as loss of synaptic connections in different brain structures including the hippocampus, a region that is believed to play an important role in certain forms of learning and memory. We report here that metaplastic priming of synapses with a cannabinoid type 1 receptor (CB1 receptor) agonist, WIN55,212-2 (WIN55), significantly impaired long-term potentiation in the apical dendrites of CA1 pyramidal neurons. Interestingly, the CB1 receptor exerts its effect by altering the balance of protein synthesis machinery towards higher protein production. Therefore the activation of CB1 receptor, prior to strong tetanization, increased the propensity to produce new proteins. In addition, WIN55 priming resulted in the expression of late-LTP in a synaptic input that would have normally expressed early-LTP, thus confirming that WIN55 priming of LTP induces new synthesis of plasticity-related proteins. Furthermore, in addition to the effects on protein translation, WIN55 also induced synaptic deficits due to the ability of CB1 receptors to inhibit the release of acetylcholine, mediated by both muscarinic and nicotinic acetylcholine receptors. Taken together this supports the notion that the modulation of cholinergic activity by CB1 receptor activation is one mechanism that regulates the synthesis of plasticity-related proteins. Copyright © 2014 Elsevier Ltd. All rights reserved.

Steroid hormone and epidermal growth factor receptors in meningiomas.

PubMed

Horsfall, D J; Goldsmith, K G; Ricciardelli, C; Skinner, J M; Tilley, W D; Marshall, V R

1989-11-01

A prospective study of steroid hormone and epidermal growth factor receptor expression in 57 meningiomas is presented. Scatchard analysis of radioligand binding identified 20% of meningiomas as expressing classical oestrogen receptors (ER) at levels below that normally accepted for positivity, the remainder being negative. ER could not be visualized in any meningioma using immunocytochemistry. Alternatively, 74% of meningiomas demonstrated the presence of progesterone receptors (PR) by Scatchard analysis, the specificity of which could not be attributed to glucocorticoid or androgen receptors. Confirmation of classical PR presence was determined by immunocytochemical staining. The presence of epidermal growth factor receptor (EGFR) was demonstrated in 100% of meningiomas using immunocytochemical staining. These data are reviewed in the context of previously reported results and are discussed in relation to the potential for medical therapy as an adjunct to surgery.

Expression of neurotensin and NT1 receptor in human breast cancer: a potential role in tumor progression.

PubMed

Souazé, Frédérique; Dupouy, Sandra; Viardot-Foucault, Véronique; Bruyneel, Erik; Attoub, Samir; Gespach, Christian; Gompel, Anne; Forgez, Patricia

2006-06-15

Emerging evidence supports neurotensin as a trophic and antiapoptotic factor, mediating its control via the high-affinity neurotensin receptor (NT1 receptor) in several human solid tumors. In a series of 51 patients with invasive ductal breast cancers, 34% of all tumors were positive for neurotensin and 91% positive for NT1 receptor. We found a coexpression of neurotensin and NT1 receptor in a large proportion (30%) of ductal breast tumors, suggesting a contribution of the neurotensinergic signaling cascade within breast cancer progression. Functionally expressed NT1 receptor, in the highly malignant MDA-MB-231 human breast cancer cell line, coordinated a series of transforming functions, including cellular migration, invasion, induction of the matrix metalloproteinase (MMP)-9 transcripts, and MMP-9 gelatinase activity. Disruption of NT1 receptor signaling by silencing RNA or use of a specific NT1 receptor antagonist, SR48692, caused the reversion of these transforming functions and tumor growth of MDA-MB-231 cells xenografted in nude mice. Our findings support the contribution of neurotensin in human breast cancer progression and point out the utility to develop therapeutic molecules targeting neurotensin or NT1 receptor signaling cascade. These strategies would increase the range of therapeutic approaches and be beneficial for specific patients.

The SRC homology 2 domain of Rin1 mediates its binding to the epidermal growth factor receptor and regulates receptor endocytosis.

PubMed

Barbieri, M Alejandro; Kong, Chen; Chen, Pin-I; Horazdovsky, Bruce F; Stahl, Philip D

2003-08-22

Activated epidermal growth factor receptors (EGFRs) recruit intracellular proteins that mediate receptor signaling and endocytic trafficking. Rin1, a multifunctional protein, has been shown to regulate EGFR internalization (1). Here we show that EGF stimulation induces a specific, rapid, and transient membrane recruitment of Rin1 and that recruitment is dependent on the Src homology 2 (SH2) domain of Rin1. Immunoprecipitation of EGFR is accompanied by co-immunoprecipitation of Rin1 in a time- and ligand-dependent manner. Association of Rin1 and specifically the SH2 domain of Rin1 with the EGFR was dependent on tyrosine phosphorylation of the intracellular domain of the EGFR. The recruitment of Rin1, observed by light microscopy, indicated that although initially cytosolic, Rin1 was recruited to both plasma membrane and endosomes following EGF addition. Moreover, the expression of the SH2 domain of Rin1 substantially impaired the internalization of EGF without affecting internalization of transferrin. Finally, we found that Rin1 co-immunoprecipitated with a number of tyrosine kinase receptors but not with cargo endocytic receptors. These results indicate that Rin1 provides a link via its SH2 domain between activated tyrosine kinase receptors and the endocytic pathway through the recruitment and activation of Rab5a.

Glycine Potentiates AMPA Receptor Function through Metabotropic Activation of GluN2A-Containing NMDA Receptors

PubMed Central

Li, Li-Jun; Hu, Rong; Lujan, Brendan; Chen, Juan; Zhang, Jian-Jian; Nakano, Yasuko; Cui, Tian-Yuan; Liao, Ming-Xia; Chen, Jin-Cao; Man, Heng-Ye; Feng, Hua; Wan, Qi

2016-01-01

NMDA receptors are Ca2+-permeable ion channels. The activation of NMDA receptors requires agonist glutamate and co-agonist glycine. Recent evidence indicates that NMDA receptor also has metabotropic function. Here we report that in cultured mouse hippocampal neurons, glycine increases AMPA receptor-mediated currents independent of the channel activity of NMDA receptors and the activation of glycine receptors. The potentiation of AMPA receptor function by glycine is antagonized by the inhibition of ERK1/2. In the hippocampal neurons and in the HEK293 cells transfected with different combinations of NMDA receptors, glycine preferentially acts on GluN2A-containing NMDA receptors (GluN2ARs), but not GluN2B-containing NMDA receptors (GluN2BRs), to enhance ERK1/2 phosphorylation independent of the channel activity of GluN2ARs. Without requiring the channel activity of GluN2ARs, glycine increases AMPA receptor-mediated currents through GluN2ARs. Thus, these results reveal a metabotropic function of GluN2ARs in mediating glycine-induced potentiation of AMPA receptor function via ERK1/2 activation. PMID:27807405

Acetaminophen Metabolite N-Acylphenolamine Induces Analgesia via Transient Receptor Potential Vanilloid 1 Receptors Expressed on the Primary Afferent Terminals of C-fibers in the Spinal Dorsal Horn.

PubMed

Ohashi, Nobuko; Uta, Daisuke; Sasaki, Mika; Ohashi, Masayuki; Kamiya, Yoshinori; Kohno, Tatsuro

2017-08-01

The widely used analgesic acetaminophen is metabolized to N-acylphenolamine, which induces analgesia by acting directly on transient receptor potential vanilloid 1 or cannabinoid 1 receptors in the brain. Although these receptors are also abundant in the spinal cord, no previous studies have reported analgesic effects of acetaminophen or N-acylphenolamine mediated by the spinal cord dorsal horn. We hypothesized that clinical doses of acetaminophen induce analgesia via these spinal mechanisms. We assessed our hypothesis in a rat model using behavioral measures. We also used in vivo and in vitro whole cell patch-clamp recordings of dorsal horn neurons to assess excitatory synaptic transmission. Intravenous acetaminophen decreased peripheral pinch-induced excitatory responses in the dorsal horn (53.1 ± 20.7% of control; n = 10; P < 0.01), while direct application of acetaminophen to the dorsal horn did not reduce these responses. Direct application of N-acylphenolamine decreased the amplitudes of monosynaptic excitatory postsynaptic currents evoked by C-fiber stimulation (control, 462.5 ± 197.5 pA; N-acylphenolamine, 272.5 ± 134.5 pA; n = 10; P = 0.022) but not those evoked by stimulation of Aδ-fibers. These phenomena were mediated by transient receptor potential vanilloid 1 receptors, but not cannabinoid 1 receptors. The analgesic effects of acetaminophen and N-acylphenolamine were stronger in rats experiencing an inflammatory pain model compared to naïve rats. Our results suggest that the acetaminophen metabolite N-acylphenolamine induces analgesia directly via transient receptor potential vanilloid 1 receptors expressed on central terminals of C-fibers in the spinal dorsal horn and leads to conduction block, shunt currents, and desensitization of these fibers.

Synthesis and binding affinity of new 1,4-disubstituted triazoles as potential dopamine D(3) receptor ligands.

PubMed

Insua, Ignacio; Alvarado, Mario; Masaguer, Christian F; Iglesias, Alba; Brea, José; Loza, María I; Carro, Laura

2013-10-15

A series of new 1,4-disubstituted triazoles was prepared from appropriate arylacetylenes and aminoalkylazides using click chemistry methodology. These compounds were evaluated as potential ligands on several subtypes of dopamine receptors in in vitro competition assays, showing high affinity for dopamine D3 receptors, lower affinity for D2 and D4, and no affinity for the D1 receptors. Compound 18 displayed the highest affinity at the D3 receptor with a Ki value of 2.7 nM, selectivity over D2 (70-fold) and D4 (200-fold), and behaviour as a competitive antagonist in the low nanomolar range. Copyright © 2013 Elsevier Ltd. All rights reserved.

miR-7-1 POTENTIATED ESTROGEN RECEPTOR AGONISTS FOR FUNCTIONAL NEUROPROTECTION IN VSC4.1 MOTONEURONS

PubMed Central

CHAKRABARTI, M.; BANIK, N. L.; RAY, S. K.

2013-01-01

Protection of motoneurons is an important goal in the treatment of spinal cord injury (SCI). We tested whether neuroprotective microRNAs (miRs) like miR-206, miR-17, miR-21, miR-7-1, and miR-106a could enhance efficacy of estrogen receptor (ER) agonists such as 1,3,5-tris (4-hydroxyphenyl)-4-propyl-1H-pyrazole (PPT, ERα agonist), Way200070 (WAY, ERβ agonist), and estrogen (EST, ERα and ERβ agonist) in preventing apoptosis in the calcium ionophore (CI) insulted VSC4.1 motoneurons. We determined that 200 nM CI induced 70% cell death. Treatment with 50 nM PPT, 100 nM WAY, and 150 nM EST induced overexpression of ERα, ERβ, and both receptors, respectively, at mRNA and protein levels. Treatment with ER agonists significantly upregulated miR-206, miR-17, and miR-7-1 in the CI insulted VSC4.1 motoneurons. Transfection with miR-206, miR-17, or miR-7-1 mimic potentiated WAY or EST to inhibit apoptosis in the CI insulted VSC4.1 motoneurons. Overexpression of miR-7-1 maximally increased efficacy of WAY and EST for down regulation of pro-apoptotic Bax and upregulation of anti-apoptotic Bcl-2. A search using miRDB indicated that miR-7-1 could inhibit expression of L-type Ca2+ channel protein alpha 1C (CPα1C). miR-7-1 overexpression and WAY or EST treatment down regulated CPα1C but upregulated p-Akt to trigger cell survival signaling. The same therapeutic strategy increased expression of the Ca2+/calmodulin-dependent protein kinase II beta (CaMKIIβ) and the phosphorylated cAMP response element binding protein (p-CREB) so as to promote Bcl-2 transcription. Whole cell membrane potential and mitochondrial membrane potential studies indicated that miR-7-1 highly potentiated EST to preserve functionality in the CI insulted VSC4.1 motoneurons. In conclusion, our data indicated that miR-7-1 most significantly potentiated efficacy of EST for functional neuroprotection and this therapeutic strategy could be used in the future to attenuate apoptosis of motoneurons in SCI. PMID

Targeting the Insulin-Like Growth Factor 1 Receptor in Ewing's Sarcoma: Reality and Expectations

PubMed Central

Olmos, David; Martins, Ana Sofia; Jones, Robin L.; Alam, Salma; Scurr, Michelle; Judson, Ian R.

2011-01-01

Ewing's sarcoma family of tumours comprises a group of very aggressive diseases that are potentially curable with multimodality treatment. Despite the undoubted success of current treatment, approximately 30% of patients will relapse and ultimately die of disease. The insulin-like growth factor 1 receptor (IGF-1R) has been implicated in the genesis, growth, proliferation, and the development of metastatic disease in Ewing's sarcoma. In addition, IGF1-R has been validated, both in vitro and in vivo, as a potential therapeutic target in Ewing's sarcoma. Phase I studies of IGF-1R monoclonal antibodies reported several radiological and clinical responses in Ewing's sarcoma patients, and initial reports of several Phase II studies suggest that about a fourth of the patients would benefit from IGF-1R monoclonal antibodies as single therapy, with approximately 10% of patients achieving objective responses. Furthermore, these therapies are well tolerated, and thus far severe toxicity has been rare. Other studies assessing IGF-1R monoclonal antibodies in combination with traditional cytotoxics or other targeted therapies are expected. Despite, the initial promising results, not all patients benefit from IGF-1R inhibition, and consequently, there is an urgent need for the identification of predictive markers of response. PMID:21647361

The corticotropin-releasing factor receptor-1 pathway mediates the negative affective states of opiate withdrawal.

PubMed

Contarino, Angelo; Papaleo, Francesco

2005-12-20

The negative affective symptoms of opiate withdrawal powerfully motivate drug-seeking behavior and may trigger relapse to heroin abuse. To date, no medications exist that effectively relieve the negative affective symptoms of opiate withdrawal. The corticotropin-releasing factor (CRF) system has been hypothesized to mediate the motivational effects of drug dependence. The CRF signal is transmitted by two distinct receptors named CRF receptor-1 (CRF1) and CRF2. Here we report that genetic disruption of CRF1 receptor pathways in mice eliminates the negative affective states of opiate withdrawal. In particular, neither CRF1 receptor heterozygous (CRF1+/-) nor homozygous (CRF1-/-) null mutant mice avoided environmental cues repeatedly paired with the early phase of opiate withdrawal. These results were not due to altered associative learning processes because CRF1+/- and CRF1-/- mice displayed reliable, conditioned place aversions to environmental cues paired with the kappa-opioid receptor agonist U-50,488H. We also examined the impact of CRF1 receptor-deficiency upon opiate withdrawal-induced dynorphin activity in the nucleus accumbens, a brain molecular mechanism thought to underlie the negative affective states of drug withdrawal. Consistent with the behavioral indices, we found that, during the early phase of opiate withdrawal, neither CRF1+/- nor CRF1-/- showed increased dynorphin mRNA levels in the nucleus accumbens. This study reveals a cardinal role for CRF/CRF1 receptor pathways in the negative affective states of opiate withdrawal and suggests therapeutic strategies for the treatment of opiate addiction.

Valerian extract Ze 911 inhibits postsynaptic potentials by activation of adenosine A1 receptors in rat cortical neurons.

PubMed

Vissiennon, Z; Sichardt, K; Koetter, U; Brattström, A; Nieber, K

2006-06-01

In this study we evaluated the adenosine A1 receptor-mediated effect of valerian extract (Ze 911) on postsynaptic potentials (PSPs) in pyramidal cells of the rat cingulate cortex in a slice preparation. We first observed that N6-cyclopentyladenosine (CPA, 0.01 - 10 microM), an adenosine A1 receptor agonist, inhibited PSPs in a concentration-dependent manner. The CPA (10 microM)-induced inhibition was antagonized by 1,3-dipropyl-8-cyclopentylxanthine (DPCPX, 0.1 microM), an adenosine A1 receptor antagonist. Ze 911 concentration dependently (0.1 - 15 mg/mL) inhibited PSPs in the presence of the adenosine A2A receptor antagonist 1,3,7-trimethyl-8-(3-chlorostyryl)xanthine (CSC, 0.2 microM) and adenosine deaminase (1 U/mL). The maximal inhibition induced by 10 mg/mL was completely antagonised by DPCPX (0.1 microM), an A1 receptor blocker. The data suggest that activation of adenosine A1 receptors is involved in the pharmacological effects of the valerian extract Ze 911.

Potentiation of Brain Stimulation Reward by Morphine: Effects of Neurokinin-1 Receptor Antagonism

PubMed Central

Robinson, J.E.; Fish, E.W.; Krouse, M.C.; Thorsell, A.; Heilig, M.; Malanga, C.J.

2012-01-01

Rationale The abuse potential of opioids may be due to their reinforcing and rewarding effects, which may be attenuated by neurokinin-1 receptor (NK1R) antagonists. Objective To measure the effects of opioid and neurokinin-1 (NK1R) receptor blockade on the potentiation of brain stimulation reward (BSR) by morphine using the intracranial self-stimulation (ICSS) method. Methods Adult male C57BL/6J mice (n = 15) were implanted with unipolar stimulating electrodes in the lateral hypothalamus and trained to respond for varying frequencies of rewarding electrical stimulation. The BSR threshold (θ0) and maximum response rate (MAX) were determined before and after intraperitoneal administration of saline, morphine (1.0 - 17.0 mg/kg), or the NK1R antagonists L-733,060 (1.0 - 17.0 mg/kg) and L-703,606 (1.0 - 17.0 mg/kg). In morphine antagonism experiments, naltrexone (0.1 – 1.0 mg/kg) or 10.0 mg/kg L-733,060 or L-703,606 was administered 15 minutes before morphine (1.0 - 10.0 mg/kg) or saline. Results Morphine dose-dependently decreased θ0 (maximum effect = 62% of baseline) and altered MAX when compared to saline. L-703,606 and L-733,060 altered θ0 without affecting MAX. 10.0 mg/kg L-733,060 and L-703,606, which did not affect θ0 or MAX, attenuated the effects of 3.0 and 10.0 mg/kg morphine. 1.0 and 0.3 mg/kg naltrexone blocked the effects of 10.0 mg/kg morphine. Naltrexone given before saline did not affect θ0 or MAX. Conclusions The decrease in θ0 by morphine reflects its rewarding effects, which were attenuated by NK1R and opioid receptor blockade. These results demonstrate the importance of substance P signaling during limbic reward system activation by opioids. PMID:21909635

Amadori products promote cellular senescence activating insulin-like growth factor-1 receptor and down-regulating the antioxidant enzyme catalase.

PubMed

Del Nogal-Ávila, María; Troyano-Suárez, Nuria; Román-García, Pablo; Cannata-Andía, Jorge B; Rodriguez-Puyol, Manuel; Rodriguez-Puyol, Diego; Kuro-O, Makoto; Ruiz-Torres, María P

2013-07-01

Activation of the insulin growth factor receptor-1 signaling pathways has been largely related to the aging process. Amadori products are produced in pathological conditions such as diabetes and aging, and are potentially involved in diabetic nephropathy or age-associated decline of renal function. We hypothesize that Amadori products induce senescence in primary human mesangial cells through the activation of IGF-1 receptor and investigate, in the present work, the intracellular mechanism involved after this activation. We treated cultured human mesangial cells with glycated albumin, one of the most abundant Amadori product, and senescence was assessed by determining the senescence associated β-galactosidase activity and the expression of the cell cycle regulators p53 and p21. We demonstrated that prolonged exposition (more than 24h) to glycated albumin induced senescence and, in parallel, incremented the release of IGF-1 and the activation of the IGF-1 receptor. Inhibition of the IGF-1 activation prevented the GA induced senescence. Activation of IGF-1R, after GA addition, promoted a reduction in the catalase content through the constitutive activation of Ras and erk1/2 proteins which were, in turn, responsible of the observed GA-induced senescence. In conclusion, we propose that the Amadori product, glycated albumin, promotes premature cell senescence in mesangial cells through the activation of the IGF-1 receptor and the subsequent reduction in the antioxidant enzyme catalase. Copyright © 2013 Elsevier Ltd. All rights reserved.

Mechanisms of resistance to anti-human epidermal growth factor receptor 2 agents in breast cancer.

PubMed

Mukohara, Toru

2011-01-01

Approximately 20% of breast cancers are characterized by overexpression of human epidermal growth factor receptor 2 (HER2) protein and associated gene amplification, and the receptor tyrosine kinase is believed to play a critical role in the pathogenesis of these tumors. The development and implementation of trastuzumab, a humanized monoclonal antibody against the extracellular domain of HER2 protein, has significantly improved treatment outcomes in patients with HER2-overexpressing breast cancer. However, despite this clinical usefulness, unmet needs for better prediction of trastuzumab's response and overcoming primary and acquired resistance remain. In this review, we discuss several potential mechanisms of resistance to trastuzumab that have been closely studied over the last decade. Briefly, these mechanisms include: impaired access of trastuzumab to HER2 by expression of extracellular domain-truncated HER2 (p95 HER2) or overexpression of MUC4; alternative signaling from insulin-like growth factor-1 receptor, other epidermal growth factor receptor family members, or MET; aberrant downstream signaling caused by loss of phosphatase and tensin homologs deleted from chromosome 10 (PTEN), PIK3CA mutation, or downregulation of p27; or FCGR3A polymorphisms. In addition, we discuss potential strategies for overcoming resistance to trastuzumab. Specifically, the epidermal growth factor receptor/HER2 tyrosine kinase inhibitor lapatinib partially overcame trastuzumab resistance in a clinical setting, so its efficacy results and limited data regarding potential mechanisms of resistance to the drug are also discussed. © 2010 Japanese Cancer Association.

Role of G protein-coupled receptors (GPCR), matrix metalloproteinases 2 and 9 (MMP2 and MMP9), heparin-binding epidermal growth factor-like growth factor (hbEGF), epidermal growth factor receptor (EGFR), erbB2, and insulin-like growth factor 1 receptor (IGF-1R) in trenbolone acetate-stimulated bovine satellite cell proliferation.

PubMed

Thornton, K J; Kamange-Sollo, E; White, M E; Dayton, W R

2015-09-01

Implanting cattle with steroids significantly enhances feed efficiency, rate of gain, and muscle growth. However, the mechanisms responsible for these improvements in muscle growth have not been fully elucidated. Trenbolone acetate (TBA), a testosterone analog, has been shown to increase proliferation rate in bovine satellite cell (BSC) cultures. The classical genomic actions of testosterone have been well characterized; however, our results indicate that TBA may also initiate a quicker, nongenomic response that involves activation of G protein-coupled receptors (GPCR) resulting in activation of matrix metalloproteinases 2 and 9 (MMP2 and MMP9) that release membrane-bound heparin-binding epidermal growth factor-like growth factor (hbEGF), which then binds to and activates the epidermal growth factor receptor (EGFR) and/or erbB2. Furthermore, the EGFR has been shown to regulate expression of the IGF-1 receptor (IGF-1R), which is well known for its role in modulating muscle growth. To determine whether this nongenomic pathway is potentially involved in TBA-stimulated BSC proliferation, we analyzed the effects of treating BSC with guanosine 5'-O-2-thiodiphosphate (GDPβS), an inhibitor of all GPCR; a MMP2 and MMP9 inhibitor (MMPI); CRM19, a specific inhibitor of hbEGF; AG1478, a specific EGFR tyrosine kinase inhibitor; AG879, a specific erbB2 kinase inhibitor; and AG1024, an IGF-1R tyrosine kinase inhibitor on TBA-stimulated proliferation rate (H-thymidine incorporation). Assays were replicated at least 9 times for each inhibitor experiment using BSC cultures obtained from at least 3 different animals. Bovine satellite cell cultures were obtained from yearling steers that had no previous exposure to androgenic or estrogenic compounds. As expected, BSC cultures treated with 10 n TBA showed ( < 0.05) increased proliferation rate when compared with control cultures. Additionally, treatment with 5 ng hbEGF/mL stimulated proliferation in BSC cultures ( < 0.05). Treatment

Cloning and expression of porcine Colony Stimulating Factor-1 (CSF-1) and Colony Stimulating Factor-1 Receptor (CSF-1R) and analysis of the species specificity of stimulation by CSF-1 and Interleukin 34

PubMed Central

Gow, Deborah J.; Garceau, Valerie; Kapetanovic, Ronan; Sester, David P.; Fici, Greg J.; Shelly, John A.; Wilson, Thomas L.; Hume, David A.

2012-01-01

Macrophage Colony Stimulating Factor (CSF-1) controls the survival, differentiation and proliferation of cells of the mononuclear phagocyte system. A second ligand for the CSF-1R, Interleukin 34 (IL-34), has been described, but its physiological role is not yet known. The domestic pig provides an alternative to traditional rodent models for evaluating potential therapeutic applications of CSF-1R agonists and antagonists. To enable such studies, we cloned and expressed active pig CSF-1. To provide a bioassay, pig CSF-1R was expressed in the factor-dependent Ba/F3 cell line. On this transfected cell line, recombinant porcine CSF-1 and human CSF-1 had identical activity. Mouse CSF-1 does not interact with the human CSF-1 receptor but was active on pig. By contrast, porcine CSF-1 was active on mouse, human, cat and dog cells. IL-34 was previously shown to be species-specific, with mouse and human proteins demonstrating limited cross-species activity. The pig CSF-1R was equally responsive to both mouse and human IL-34. Based upon the published crystal structures of CSF-1/CSF-1R and IL34/CSF-1R complexes, we discuss the molecular basis for the species specificity. PMID:22974529

Structure of nerve growth factor complexed with the shared neurotrophin receptor p75.

PubMed

He, Xiao-Lin; Garcia, K Christopher

2004-05-07

Neurotrophins are secreted growth factors critical for the development and maintenance of the vertebrate nervous system. Neurotrophins activate two types of cell surface receptors, the Trk receptor tyrosine kinases and the shared p75 neurotrophin receptor. We have determined the 2.4 A crystal structure of the prototypic neurotrophin, nerve growth factor (NGF), complexed with the extracellular domain of p75. Surprisingly, the complex is composed of an NGF homodimer asymmetrically bound to a single p75. p75 binds along the homodimeric interface of NGF, which disables NGF's symmetry-related second p75 binding site through an allosteric conformational change. Thus, neurotrophin signaling through p75 may occur by disassembly of p75 dimers and assembly of asymmetric 2:1 neurotrophin/p75 complexes, which could potentially engage a Trk receptor to form a trimolecular signaling complex.

Substance P - Neurokinin-1 Receptor Interaction Upregulates Monocyte Tissue Factor

PubMed Central

Khan, Mohammad M; Douglas, Steven D; Benton, Tami D

2011-01-01

Monocytes play an important role in hemostasis. In this study, the prothrombotic effects of the neuropeptide substance P (SP) on human monocytes through neurokinin-1 receptor (NK1-R) were characterized. SP upregulated monocyte tissue factor (TF), the major coagulation cascade stimulator, in a concentration and time dependent manner. Specific inhibition of NK1-R completely blocked TF expression. Monocytes stimulated by SP released cytokines and chemokines. When monocytes were stimulated with cytokines or chemokines, TF was expressed by the cytokines (GM-CSF, IFN-γ and TNF-α). Cytokines may play a major role in the mechanism of SP induced monocyte TF expression. NK1-R antagonists (NK1-RA) may have a role in developing novel therapeutic approaches to patients vulnerable to vaso-occlusive disorders. PMID:22115773

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

PubMed

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.

A Novel Positive Feedback Loop Mediated by the Docking Protein Gab1 and Phosphatidylinositol 3-Kinase in Epidermal Growth Factor Receptor Signaling

PubMed Central

Rodrigues, Gerard A.; Falasca, Marco; Zhang, Zhongtao; Ong, Siew Hwa; Schlessinger, Joseph

2000-01-01

The Gab1 protein is tyrosine phosphorylated in response to various growth factors and serves as a docking protein that recruits a number of downstream signaling proteins, including phosphatidylinositol 3-kinase (PI-3 kinase). To determine the role of Gab1 in signaling via the epidermal growth factor (EGF) receptor (EGFR) we tested the ability of Gab1 to associate with and modulate signaling by this receptor. We show that Gab1 associates with the EGFR in vivo and in vitro via pTyr sites 1068 and 1086 in the carboxy-terminal tail of the receptor and that overexpression of Gab1 potentiates EGF-induced activation of the mitogen-activated protein kinase and Jun kinase signaling pathways. A mutant of Gab1 unable to bind the p85 subunit of PI-3 kinase is defective in potentiating EGFR signaling, confirming a role for PI-3 kinase as a downstream effector of Gab1. Inhibition of PI-3 kinase by a dominant-interfering mutant of p85 or by Wortmannin treatment similarly impairs Gab1-induced enhancement of signaling via the EGFR. The PH domain of Gab1 was shown to bind specifically to phosphatidylinositol 3,4,5-triphosphate [PtdIns(3,4,5)P3], a product of PI-3 kinase, and is required for activation of Gab1-mediated enhancement of EGFR signaling. Moreover, the PH domain mediates Gab1 translocation to the plasma membrane in response to EGF and is required for efficient tyrosine phosphorylation of Gab1 upon EGF stimulation. In addition, overexpression of Gab1 PH domain blocks Gab1 potentiation of EGFR signaling. Finally, expression of the gene for the lipid phosphatase PTEN, which dephosphorylates PtdIns(3,4,5)P3, inhibits EGF signaling and translocation of Gab1 to the plasma membrane. These results reveal a novel positive feedback loop, modulated by PTEN, in which PI-3 kinase functions as both an upstream regulator and a downstream effector of Gab1 in signaling via the EGFR. PMID:10648629

Inhibition of transient receptor potential vanilloid-1 confers neuroprotection, reduces tumor necrosis factor-alpha, and increases IL-10 in a rat stroke model.

PubMed

Hakimizadeh, Elham; Shamsizadeh, Ali; Roohbakhsh, Ali; Arababadi, Mohammad Kazemi; Hajizadeh, Mohammad R; Shariati, Mehdi; Rahmani, Mohammad R; Allahtavakoli, Mohammad

2017-08-01

Stroke is a major cause of mortality and long-term disability in adults. Transient receptor potential vanilloid-1 (TRPV1) plays a crucial role in neuroinflammation. In this study, the effects of TRPV1 agonist (capsaicin) and antagonist (AMG9810) on cerebral ischemia were investigated. Forty male Wistar rats were assigned to the following experimental groups: sham, vehicle) ischemic), AMG9810 (selective TRPV1 antagonist, 0.5 mg/kg; 3 h after stroke), and capsaicin (1 mg/kg; 3 h after stroke). Stroke was induced by permanent middle cerebral artery occlusion and neurological deficits were evaluated 1, 3, and 7 days after stroke. Then, infarct volume, brain edema, body temperature, mRNA expression of TRPV1, and serum concentrations of tumor necrosis factor-alpha (TNF-α) and IL-10 were measured. Compared to the vehicle group, AMG9810 significantly decreased the infarct volume (P < 0.01). Latency for the removal of sticky labels from the forepaw and the hanging time were significantly decreased and increased, respectively, following administration of AMG9810 (P < 0.01 and P < 0.001 vs. vehicle) 3 and 7 days after stroke. Compared to the sham group, the mRNA expression of TRPV1 was significantly increased in vehicle group (P < 0.01). Administration of AMG9810 significantly increased the anti-inflammatory cytokine IL-10 and decreased the inflammatory cytokine TNF-α (P < 0.05). Moreover, our results indicate that AMG9810 might a promising candidate for the hypothermic treatment of stroke. The findings also suggest a key role for AMG9810 in reducing inflammation after stroke and imply that TRPV1 could be a potential target for the treatment of ischemic stroke. © 2017 Société Française de Pharmacologie et de Thérapeutique.

Factor VIII Interacts with the Endocytic Receptor Low-density Lipoprotein Receptor-related Protein 1 via an Extended Surface Comprising "Hot-Spot" Lysine Residues.

PubMed

van den Biggelaar, Maartje; Madsen, Jesper J; Faber, Johan H; Zuurveld, Marleen G; van der Zwaan, Carmen; Olsen, Ole H; Stennicke, Henning R; Mertens, Koen; Meijer, Alexander B

2015-07-03

Lysine residues are implicated in driving the ligand binding to the LDL receptor family. However, it has remained unclear how specificity is regulated. Using coagulation factor VIII as a model ligand, we now study the contribution of individual lysine residues in the interaction with the largest member of the LDL receptor family, low-density lipoprotein receptor-related protein (LRP1). Using hydrogen-deuterium exchange mass spectrometry (HDX-MS) and SPR interaction analysis on a library of lysine replacement variants as two independent approaches, we demonstrate that the interaction between factor VIII (FVIII) and LRP1 occurs over an extended surface containing multiple lysine residues. None of the individual lysine residues account completely for LRP1 binding, suggesting an additive binding model. Together with structural docking studies, our data suggest that FVIII interacts with LRP1 via an extended surface of multiple lysine residues that starts at the bottom of the C1 domain and winds around the FVIII molecule. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

MiR-7-1 potentiated estrogen receptor agonists for functional neuroprotection in VSC4.1 motoneurons.

PubMed

Chakrabarti, M; Banik, N L; Ray, S K

2014-01-03

Protection of motoneurons is an important goal in the treatment of spinal cord injury (SCI). We tested whether neuroprotective microRNAs (miRs) like miR-206, miR-17, miR-21, miR-7-1, and miR-106a could enhance efficacy of estrogen receptor (ER) agonists such as 1,3,5-tris (4-hydroxyphenyl)-4-propyl-1H-pyrazole (PPT, ERα agonist), Way200070 (WAY, ERβ agonist), and estrogen (EST, ERα and ERβ agonist) in preventing apoptosis in the calcium ionophore (CI)-insulted ventral spinal cord 4.1 (VSC4.1) motoneurons. We determined that 200 nM CI induced 70% cell death. Treatment with 50 nM PPT, 100 nM WAY, and 150 nM EST induced overexpression of ERα, ERβ, and both receptors, respectively, at mRNA and protein levels. Treatment with ER agonists significantly upregulated miR-206, miR-17, and miR-7-1 in the CI-insulted VSC4.1 motoneurons. Transfection with miR-206, miR-17, or miR-7-1 mimic potentiated WAY or EST to inhibit apoptosis in the CI-insulted VSC4.1 motoneurons. Overexpression of miR-7-1 maximally increased efficacy of WAY and EST for down regulation of pro-apoptotic Bax and upregulation of anti-apoptotic Bcl-2. A search using microRNA database (miRDB) indicated that miR-7-1 could inhibit the expression of L-type Ca(2+) channel protein alpha 1C (CPα1C). miR-7-1 overexpression and WAY or EST treatment down regulated CPα1C but upregulated p-Akt to trigger cell survival signaling. The same therapeutic strategy increased expression of the Ca(2+)/calmodulin-dependent protein kinase II beta (CaMKIIβ) and the phosphorylated cAMP response element binding protein (p-CREB) so as to promote Bcl-2 transcription. Whole cell membrane potential and mitochondrial membrane potential studies indicated that miR-7-1 highly potentiated EST to preserve functionality in the CI-insulted VSC4.1 motoneurons. In conclusion, our data indicated that miR-7-1 most significantly potentiated efficacy of EST for functional neuroprotection and this therapeutic strategy could be used in the future

Selenoprotein W controls epidermal growth factor receptor surface expression, activation and degradation via receptor ubiquitination

USDA-ARS?s Scientific Manuscript database

Epidermal growth factor (EGF) receptor (EGFR) is the founding member of the ErbB family of growth factor receptors that modulate a complex network of intracellular signaling pathways controlling growth, proliferation and differentiation. Selenoprotein W (SEPW1) is a diet-regulated, highly conserved...

Inflammatory Pathways Regulated by Tumor Necrosis Receptor-Associated Factor 1 Protect From Metabolic Consequences in Diet-Induced Obesity.

PubMed

Anto Michel, Nathaly; Colberg, Christian; Buscher, Konrad; Sommer, Björn; Pramod, Akula Bala; Ehinger, Erik; Dufner, Bianca; Hoppe, Natalie; Pfeiffer, Katharina; Marchini, Timoteo; Willecke, Florian; Stachon, Peter; Hilgendorf, Ingo; Heidt, Timo; von Zur Muhlen, Constantin; von Elverfeldt, Dominik; Pfeifer, Dietmar; Schüle, Roland; Kintscher, Ulrich; Brachs, Sebastian; Ley, Klaus; Bode, Christoph; Zirlik, Andreas; Wolf, Dennis

2018-03-02

The coincidence of inflammation and metabolic derangements in obese adipose tissue has sparked the concept of met-inflammation. Previous observations, however, suggest that inflammatory pathways may not ultimately cause dysmetabolism. We have revisited the relationship between inflammation and metabolism by testing the role of TRAF (tumor necrosis receptor-associated factor)-1, an inhibitory adapter of inflammatory signaling of TNF (tumor necrosis factor)-α, IL (interleukin)-1β, and TLRs (toll-like receptors). Mice deficient for TRAF-1, which is expressed in obese adipocytes and adipose tissue lymphocytes, caused an expected hyperinflammatory phenotype in adipose tissue with enhanced adipokine and chemokine expression, increased leukocyte accumulation, and potentiated proinflammatory signaling in macrophages and adipocytes in a mouse model of diet-induced obesity. Unexpectedly, TRAF-1 -/- mice were protected from metabolic derangements and adipocyte growth, failed to gain weight, and showed improved insulin resistance-an effect caused by increased lipid breakdown in adipocytes and UCP (uncoupling protein)-1-enabled thermogenesis. TRAF-1-dependent catabolic and proinflammatory cues were synergistically driven by β3-adrenergic and inflammatory signaling and required the presence of both TRAF-1-deficient adipocytes and macrophages. In human obesity, TRAF-1-dependent genes were upregulated. Enhancing TRAF-1-dependent inflammatory pathways in a gain-of-function approach protected from metabolic derangements in diet-induced obesity. These findings identify TRAF-1 as a regulator of dysmetabolism in mice and humans and question the pathogenic role of chronic inflammation in metabolism. © 2018 American Heart Association, Inc.

Vestibular receptors contribute to cortical auditory evoked potentials.

PubMed

Todd, Neil P M; Paillard, Aurore C; Kluk, Karolina; Whittle, Elizabeth; Colebatch, James G

2014-03-01

Acoustic sensitivity of the vestibular apparatus is well-established, but the contribution of vestibular receptors to the late auditory evoked potentials of cortical origin is unknown. Evoked potentials from 500 Hz tone pips were recorded using 70 channel EEG at several intensities below and above the vestibular acoustic threshold, as determined by vestibular evoked myogenic potentials (VEMPs). In healthy subjects both auditory mid- and long-latency auditory evoked potentials (AEPs), consisting of Na, Pa, N1 and P2 waves, were observed in the sub-threshold conditions. However, in passing through the vestibular threshold, systematic changes were observed in the morphology of the potentials and in the intensity dependence of their amplitude and latency. These changes were absent in a patient without functioning vestibular receptors. In particular, for the healthy subjects there was a fronto-central negativity, which appeared at about 42 ms, referred to as an N42, prior to the AEP N1. Source analysis of both the N42 and N1 indicated involvement of cingulate cortex, as well as bilateral superior temporal cortex. Our findings are best explained by vestibular receptors contributing to what were hitherto considered as purely auditory evoked potentials and in addition tentatively identify a new component that appears to be primarily of vestibular origin. Copyright © 2013 The Authors. Published by Elsevier B.V. All rights reserved.

Phosphorylation and Intramolecular Stabilization of the Ligand Binding Domain in the Nuclear Receptor Steroidogenic Factor 1

PubMed Central

Desclozeaux, Marion; Krylova, Irina N.; Horn, Florence; Fletterick, Robert J.; Ingraham, Holly A.

2002-01-01

Steroidogenic factor 1 (SF-1) is an orphan nuclear receptor with no known ligand. We showed previously that phosphorylation at serine 203 located N′-terminal to the ligand binding domain (LBD) enhanced cofactor recruitment, analogous to the ligand-mediated recruitment in ligand-dependent receptors. In this study, results of biochemical analyses and an LBD helix assembly assay suggest that the SF-1 LBD adopts an active conformation, with helices 1 and 12 packed against the predicted alpha-helical bundle, in the apparent absence of ligand. Fine mapping of the previously defined proximal activation function in SF-1 showed that the activation function mapped fully to helix 1 of the LBD. Limited proteolyses demonstrate that phosphorylation of S203 in the hinge region mimics the stabilizing effects of ligand on the LBD. Moreover, similar effects were observed in an SF-1/thyroid hormone LBD chimera receptor, illustrating that the S203 phosphorylation effects are transferable to a heterologous ligand-dependent receptor. Our collective data suggest that the hinge together with helix 1 is an individualized specific motif, which is tightly associated with its cognate LBD. For SF-1, we find that this intramolecular association and hence receptor activity are further enhanced by mitogen-activated protein kinase phosphorylation, thus mimicking many of the ligand-induced changes observed for ligand-dependent receptors. PMID:12242296

Coarse Architecture of the Transient Receptor Potential Vanilloid 1 (TRPV1) Ion Channel Determined by Fluorescence Resonance Energy Transfer*

PubMed Central

De-la-Rosa, Víctor; Rangel-Yescas, Gisela E.; Ladrón-de-Guevara, Ernesto; Rosenbaum, Tamara; Islas, León D.

2013-01-01

The transient receptor potential vanilloid 1 ion channel is responsible for the perception of high temperatures and low extracellular pH, and it is also involved in the response to some pungent compounds. Importantly, it is also associated with the perception of pain and noxious stimuli. Here, we attempt to discern the molecular organization and location of the N and C termini of the transient receptor potential vanilloid 1 ion channel by measuring FRET between genetically attached enhanced yellow and cyan fluorescent protein to the N or C terminus of the channel protein, expressed in transfected HEK 293 cells or Xenopus laevis oocytes. The static measurements of the domain organization were mapped into an available cryo-electron microscopy density of the channel with good agreement. These measurements also provide novel insights into the organization of terminal domains and their proximity to the plasma membrane. PMID:23965996

Coarse architecture of the transient receptor potential vanilloid 1 (TRPV1) ion channel determined by fluorescence resonance energy transfer.

PubMed

De-la-Rosa, Víctor; Rangel-Yescas, Gisela E; Ladrón-de-Guevara, Ernesto; Rosenbaum, Tamara; Islas, León D

2013-10-11

The transient receptor potential vanilloid 1 ion channel is responsible for the perception of high temperatures and low extracellular pH, and it is also involved in the response to some pungent compounds. Importantly, it is also associated with the perception of pain and noxious stimuli. Here, we attempt to discern the molecular organization and location of the N and C termini of the transient receptor potential vanilloid 1 ion channel by measuring FRET between genetically attached enhanced yellow and cyan fluorescent protein to the N or C terminus of the channel protein, expressed in transfected HEK 293 cells or Xenopus laevis oocytes. The static measurements of the domain organization were mapped into an available cryo-electron microscopy density of the channel with good agreement. These measurements also provide novel insights into the organization of terminal domains and their proximity to the plasma membrane.

Estrogen receptor β and Liver X receptor β: biology and therapeutic potential in CNS diseases.

PubMed

Warner, M; Gustafsson, J-A

2015-02-01

In the last decade of the twentieth century, two nuclear receptors were discovered in our laboratory and, very surprisingly, were found to have key roles in the central nervous system. These receptors have provided some novel insights into the etiology and progression of neurodegenerative diseases and anxiety disorders. The two receptors are estrogen receptor beta (ERβ) and liver X receptor beta (LXRβ). Both ERβ and LXRβ have potent anti-inflammatory activities and, in addition, LXRβ is involved in the genesis of dopaminergic neurons during development and protection of these neurons against neurodegeneration in adult life. ERβ is involved in migration of cortical neurons and calretinin-positive GABAergic interneurons during development and maintenance of serotonergic neurons in adults. Both receptors are present in magnocellular neurons of the hypothalamic preoptic area including those expressing vasopressin and oxytocin. As both ERβ and LXRβ are ligand-activated transcription factors, their ligands hold great potential in the treatment of diseases of the CNS.

Mechanisms of transient receptor potential vanilloid 1 activation and sensitization by allyl isothiocyanate.

PubMed

Gees, Maarten; Alpizar, Yeranddy A; Boonen, Brett; Sanchez, Alicia; Everaerts, Wouter; Segal, Andrei; Xue, Fenqin; Janssens, Annelies; Owsianik, Grzegorz; Nilius, Bernd; Voets, Thomas; Talavera, Karel

2013-09-01

Allyl isothiocyanate (AITC; aka, mustard oil) is a powerful irritant produced by Brassica plants as a defensive trait against herbivores and confers pungency to mustard and wasabi. AITC is widely used experimentally as an inducer of acute pain and neurogenic inflammation, which are largely mediated by the activation of nociceptive cation channels transient receptor potential ankyrin 1 and transient receptor potential vanilloid 1 (TRPV1). Although it is generally accepted that electrophilic agents activate these channels through covalent modification of cytosolic cysteine residues, the mechanism underlying TRPV1 activation by AITC remains unknown. Here we show that, surprisingly, AITC-induced activation of TRPV1 does not require interaction with cysteine residues, but is largely dependent on S513, a residue that is involved in capsaicin binding. Furthermore, AITC acts in a membrane-delimited manner and induces a shift of the voltage dependence of activation toward negative voltages, which is reminiscent of capsaicin effects. These data indicate that AITC acts through reversible interactions with the capsaicin binding site. In addition, we show that TRPV1 is a locus for cross-sensitization between AITC and acidosis in nociceptive neurons. Furthermore, we show that residue F660, which is known to determine the stimulation by low pH in human TRPV1, is also essential for the cross-sensitization of the effects of AITC and low pH. Taken together, these findings demonstrate that not all reactive electrophiles stimulate TRPV1 via cysteine modification and help understanding the molecular bases underlying the surprisingly large role of this channel as mediator of the algesic properties of AITC.

Inhibiting thyrotropin/insulin-like growth factor 1 receptor crosstalk to treat Graves' ophthalmopathy: studies in orbital fibroblasts in vitro.

PubMed

Place, Robert F; Krieger, Christine C; Neumann, Susanne; Gershengorn, Marvin C

2017-02-01

Crosstalk between thyrotropin (TSH) receptors and insulin-like growth factor 1 (IGF-1) receptors initiated by activation of TSH receptors could be important in the development of Graves' ophthalmopathy (GO). Specifically, TSH receptor activation alone is sufficient to stimulate hyaluronic acid (HA) secretion, a major component of GO, through both IGF-1 receptor-dependent and -independent pathways. Although an anti-IGF-1 receptor antibody is in clinical trials, its effectiveness depends on the relative importance of IGF-1 versus TSH receptor signalling in GO pathogenesis. TSH and IGF-1 receptor antagonists were used to probe TSH/IGF-1 receptor crosstalk in primary cultures of Graves' orbital fibroblasts (GOFs) following activation with monoclonal TSH receptor antibody, M22. Inhibition of HA secretion following TSH receptor stimulation was measured by modified HA elisa. TSH receptor antagonist, ANTAG3 (NCGC00242364), inhibited both IGF-1 receptor -dependent and -independent pathways at all doses of M22; whereas IGF-1 receptor antagonists linsitinib and 1H7 (inhibitory antibody) lost efficacy at high M22 doses. Combining TSH and IGF-1 receptor antagonists exhibited Loewe additivity within the IGF-1 receptor-dependent component of the M22 concentration-response. Similar effects were observed in GOFs activated by autoantibodies from GO patients' sera. Our data support TSH and IGF-1 receptors as therapeutic targets for GO, but reveal putative conditions for anti-IGF-1 receptor resistance. Combination treatments antagonizing both receptors yield additive effects by inhibiting crosstalk triggered by TSH receptor stimulatory antibodies. Combination therapy may be an effective strategy for dose reduction and/or compensate for any loss of anti-IGF-1 receptor efficacy. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.

Plasticity in interactions of fibroblast growth factor 1 (FGF1) N terminus with FGF receptors underlies promiscuity of FGF1.

PubMed

Beenken, Andrew; Eliseenkova, Anna V; Ibrahimi, Omar A; Olsen, Shaun K; Mohammadi, Moosa

2012-01-27

Tissue-specific alternative splicing in the second half of Ig-like domain 3 (D3) of fibroblast growth factor receptors 1-3 (FGFR1 to -3) generates epithelial FGFR1b-FGFR3b and mesenchymal FGFR1c-FGFR3c splice isoforms. This splicing event establishes a selectivity filter to restrict the ligand binding specificity of FGFRb and FGFRc isoforms to mesenchymally and epithelially derived fibroblast growth factors (FGFs), respectively. FGF1 is termed the "universal FGFR ligand" because it overrides this specificity barrier. To elucidate the molecular basis for FGF1 cross-reactivity with the "b" and "c" splice isoforms of FGFRs, we determined the first crystal structure of FGF1 in complex with an FGFRb isoform, FGFR2b, at 2.1 Å resolution. Comparison of the FGF1-FGFR2b structure with the three previously published FGF1-FGFRc structures reveals that plasticity in the interactions of the N-terminal region of FGF1 with FGFR D3 is the main determinant of FGF1 cross-reactivity with both isoforms of FGFRs. In support of our structural data, we demonstrate that substitution of three N-terminal residues (Gly-19, His-25, and Phe-26) of FGF2 (a ligand that does not bind FGFR2b) for the corresponding residues of FGF1 (Phe-16, Asn-22, and Tyr-23) enables the FGF2 triple mutant to bind and activate FGFR2b. These findings taken together with our previous structural data on receptor binding specificity of FGF2, FGF8, and FGF10 conclusively show that sequence divergence at the N termini of FGFs is the primary regulator of the receptor binding specificity and promiscuity of FGFs.

Regulation of insulin-like growth factor I receptors on vascular smooth muscle cells by growth factors and phorbol esters.

PubMed

Ververis, J J; Ku, L; Delafontaine, P

1993-06-01

Insulin-like growth factor I (IGF I) is an important mitogen for vascular smooth muscle cells. To characterize regulation of vascular IGF I receptors, we performed radioligand displacement experiments using rat aortic smooth muscle cells (RASMs). Serum deprivation for 48 hours caused a 40% decrease in IGF I receptor number. Exposure of quiescent RASMs to platelet-derived growth factor (PDGF), fibroblast growth factor (FGF), or angiotensin II (Ang II) caused a 1.5-2.0-fold increase in IGF I receptors per cell. After FGF exposure, there was a marked increase in the mitogenic response to IGF I. IGF I downregulated its receptors in the presence of platelet-poor plasma. Stimulation of protein kinase C (PKC) by exposure of quiescent RASMs to phorbol 12-myristate 13-acetate caused a biphasic response in IGF I binding; there was a 42% decrease in receptor number at 45 minutes and a 238% increase at 24 hours. To determine the role of PKC in growth factor-induced regulation of IGF I receptors, we downregulated PKC by exposing RASMs to phorbol 12,13-dibutyrate (PDBu) for 48 hours. PDGF- and FGF- but not Ang II-mediated upregulation of IGF I receptors was completely inhibited in PDBu-treated cells. Thus, acute PKC activation by phorbol esters inhibits IGF I binding, whereas chronic PKC activation increases IGF I binding. PDGF and FGF but not Ang II regulate vascular IGF I receptors through a PKC-dependent pathway. These data provide new insights into the regulation of vascular smooth muscle cell IGF I receptors in vitro and are of potential importance in characterizing vascular proliferative responses in vivo.

Corticotropin-releasing factor peptide antagonists: design, characterization and potential clinical relevance.

PubMed

Rivier, Jean E; Rivier, Catherine L

2014-04-01

Elusive for more than half a century, corticotropin-releasing factor (CRF) was finally isolated and characterized in 1981 from ovine hypothalami and shortly thereafter, from rat brains. Thirty years later, much has been learned about the function and localization of CRF and related family members (Urocortins 1, 2 and 3) and their 2 receptors, CRF receptor type 1 (CRFR1) and CRF receptor type 2 (CRFR2). Here, we report the stepwise development of peptide CRF agonists and antagonists, which led to the CRFR1 agonist Stressin1; the long-acting antagonists Astressin2-B which is specific for CRFR2; and Astressin B, which binds to both CRFR1 and CRFR2.This analog has potential for the treatment of CRF-dependent diseases in the periphery, such as irritable bowel syndrome. Copyright © 2013 Elsevier Inc. All rights reserved.

Mouse glucocorticoid-induced tumor necrosis factor receptor ligand is costimulatory for T cells

PubMed Central

Tone, Masahide; Tone, Yukiko; Adams, Elizabeth; Yates, Stephen F.; Frewin, Mark R.; Cobbold, Stephen P.; Waldmann, Herman

2003-01-01

Recently, agonist antibodies to glucocorticoid-induced tumor necrosis factor receptor (GITR) (tumor necrosis factor receptor superfamily 18) have been shown to neutralize the suppressive activity of CD4+CD25+ regulatory T cells. It was anticipated that this would be the role of the physiological ligand. We have identified and expressed the gene for mouse GITR ligand and have confirmed that its interaction with GITR reverses suppression by CD4+CD25+ T cells. It also, however, provides a costimulatory signal for the antigen-driven proliferation of naïve T cells and polarized T helper 1 and T helper 2 clones. RT-PCR and mAb staining revealed mouse GITR ligand expression in dendritic cells, macrophages, and B cells. Expression was controlled by the transcription factor NF-1 and potentially by alternative splicing of mRNA destabilization sequences. PMID:14608036

Expression of insulin-like growth factor-1 and insulin-like growth factor-1 receptors in EL4 lymphoma cells overexpressing growth hormone.

PubMed

Weigent, Douglas A; Arnold, Robyn E

2005-03-01

Almost all of the previous studies with growth hormone (GH) have been done with exogenously supplied GH and, therefore, involve actions of the hormone through its receptor. However, the actions of endogenous or lymphocyte GH are still unclear. In a previous study, we showed that overexpression of GH (GHo) in a lymphoid cell line resulted in protection of the cells to apoptosis mediated by nitric oxide (NO). In the present study, we show that the protection from apoptosis could be transferred to control cells with culture fluids obtained from GHo cells and blocked by antibodies to the insulin-like growth factor-1 (IGF-1) or antibodies to the IGF-1-receptor (IGF-1R). Northern and Western blot analysis detected significantly higher levels of IGF-1 in cells overexpressing GH. An increase in the expression of the IGF-1R in GHo cells was also detected by Western blot analysis, (125)I-IGF-1 binding and analysis of IGF-1R promoter luciferase constructs. Transfection of GHo cells with a dominant negative IGF-1R mutant construct blocked the generation of NO and activation of Akt seen in GHo cells compared to vector alone control EL4 cells. The results suggest that one of the consequences of the overexpression of GH, in cells lacking the GH receptor, is an increase in the expression of IGF-1 and the IGF-1R which mediate the protection of EL4 lymphoma cells from apoptosis.

Transient Receptor Potential Canonical (TRPC)/Orai1-dependent Store-operated Ca2+ Channels

PubMed Central

Sabourin, Jessica; Bartoli, Fiona; Antigny, Fabrice; Gomez, Ana Maria; Benitah, Jean-Pierre

2016-01-01

Store-operated Ca2+ entry (SOCE) has emerged as an important mechanism in cardiac pathology. However, the signals that up-regulate SOCE in the heart remain unexplored. Clinical trials have emphasized the beneficial role of mineralocorticoid receptor (MR) signaling blockade in heart failure and associated arrhythmias. Accumulated evidence suggests that the mineralocorticoid hormone aldosterone, through activation of its receptor, MR, might be a key regulator of Ca2+ influx in cardiomyocytes. We thus assessed whether and how SOCE involving transient receptor potential canonical (TRPC) and Orai1 channels are regulated by aldosterone/MR in neonatal rat ventricular cardiomyocytes. Molecular screening using qRT-PCR and Western blotting demonstrated that aldosterone treatment for 24 h specifically increased the mRNA and/or protein levels of Orai1, TRPC1, -C4, -C5, and stromal interaction molecule 1 through MR activation. These effects were correlated with a specific enhancement of SOCE activities sensitive to store-operated channel inhibitors (SKF-96365 and BTP2) and to a potent Orai1 blocker (S66) and were prevented by TRPC1, -C4, and Orai1 dominant negative mutants or TRPC5 siRNA. A mechanistic approach showed that up-regulation of serum- and glucocorticoid-regulated kinase 1 mRNA expression by aldosterone is involved in enhanced SOCE. Functionally, 24-h aldosterone-enhanced SOCE is associated with increased diastolic [Ca2+]i, which is blunted by store-operated channel inhibitors. Our study provides the first evidence that aldosterone promotes TRPC1-, -C4-, -C5-, and Orai1-mediated SOCE in cardiomyocytes through an MR and serum- and glucocorticoid-regulated kinase 1 pathway. PMID:27129253

Transient receptor potential ankyrin 1 channels are involved in spontaneous peptide hormone release from astrocytes.

PubMed

Takizawa, Mai; Harada, Kazuki; Nakamura, Kazuaki; Tsuboi, Takashi

2018-07-02

Astrocytes, a large population of glial cells, detect neurotransmitters and respond by increasing intracellular Ca 2+ concentration ([Ca 2+ ] i ) and releasing chemical molecules called gliotransmitters. Recently discovered Ca 2+ influx through transient receptor potential ankyrin 1 (TRPA1) channels is reported to cause spontaneous [Ca 2+ ] i increase in astrocytes. While several physiological functions of TRPA1-mediated spontaneous Ca 2+ signal have been revealed, relation with gliotransmitter release, especially peptide hormone exocytosis is largely unknown. We therefore explored the [Ca 2+ ] i and exocytosis dynamics in rat astrocyte cell line C6 cells and primary astrocytes. TRPA1-mediated spontaneous [Ca 2+ ] i transients were observed in both C6 cells and primary astrocytes. Total internal reflection fluorescence microscopy revealed that Venus-tagged brain-derived neurotrophic factor and neuropeptide Y were released spontaneously from astrocytes. Activation of TRPA1 channels enhanced the frequency of peptide hormone exocytosis, and inhibition of TRPA1 channels decreased the number of peptide hormone exocytosis. These results suggest that TRPA1-mediated spontaneous [Ca 2+ ] i increase modulates the spontaneous release of peptide hormones from astrocytes. Copyright © 2018 Elsevier Inc. All rights reserved.

Cross-talk between an activator of nuclear receptors-mediated transcription and the D1 dopamine receptor signaling pathway.

PubMed

Schmidt, Azriel; Vogel, Robert; Rutledge, Su Jane; Opas, Evan E; Rodan, Gideon A; Friedman, Eitan

2005-03-01

Nuclear receptors are transcription factors that usually interact, in a ligand-dependent manner, with specific DNA sequences located within promoters of target genes. The nuclear receptors can also be controlled in a ligand-independent manner via the action of membrane receptors and cellular signaling pathways. 5-Tetradecyloxy-2-furancarboxylic acid (TOFA) was shown to stimulate transcription from the MMTV promoter via chimeric receptors that consist of the DNA binding domain of GR and the ligand binding regions of the PPARbeta or LXRbeta nuclear receptors (GR/PPARbeta and GR/LXRbeta). TOFA and hydroxycholesterols also modulate transcription from NF-kappaB- and AP-1-controlled reporter genes and induce neurite differentiation in PC12 cells. In CV-1 cells that express D(1) dopamine receptors, D(1) dopamine receptor stimulation was found to inhibit TOFA-stimulated transcription from the MMTV promoter that is under the control of chimeric GR/PPARbeta and GR/LXRbeta receptors. Treatment with the D(1) dopamine receptor antagonist, SCH23390, prevented dopamine-mediated suppression of transcription, and by itself increased transcription controlled by GR/LXRbeta. Furthermore, combined treatment of CV-1 cells with TOFA and SCH23390 increased transcription controlled by the GR/LXRbeta chimeric receptor synergistically. The significance of this in vitro synergy was demonstrated in vivo, by the observation that SCH23390 (but not haloperidol)-mediated catalepsy in rats was potentiated by TOFA, thus showing that an agent that mimics the in vitro activities of compounds that activate members of the LXR and PPAR receptor families can influence D1 dopamine receptor elicited responses.

Inhibiting thyrotropin/insulin‐like growth factor 1 receptor crosstalk to treat Graves' ophthalmopathy: studies in orbital fibroblasts in vitro

PubMed Central

Place, Robert F; Neumann, Susanne; Gershengorn, Marvin C

2017-01-01

Background and Purpose Crosstalk between thyrotropin (TSH) receptors and insulin‐like growth factor 1 (IGF‐1) receptors initiated by activation of TSH receptors could be important in the development of Graves' ophthalmopathy (GO). Specifically, TSH receptor activation alone is sufficient to stimulate hyaluronic acid (HA) secretion, a major component of GO, through both IGF‐1 receptor‐dependent and ‐independent pathways. Although an anti‐IGF‐1 receptor antibody is in clinical trials, its effectiveness depends on the relative importance of IGF‐1 versus TSH receptor signalling in GO pathogenesis. Experimental Approach TSH and IGF‐1 receptor antagonists were used to probe TSH/IGF‐1 receptor crosstalk in primary cultures of Graves' orbital fibroblasts (GOFs) following activation with monoclonal TSH receptor antibody, M22. Inhibition of HA secretion following TSH receptor stimulation was measured by modified HA elisa. Key Results TSH receptor antagonist, ANTAG3 (NCGC00242364), inhibited both IGF‐1 receptor ‐dependent and ‐independent pathways at all doses of M22; whereas IGF‐1 receptor antagonists linsitinib and 1H7 (inhibitory antibody) lost efficacy at high M22 doses. Combining TSH and IGF‐1 receptor antagonists exhibited Loewe additivity within the IGF‐1 receptor‐dependent component of the M22 concentration‐response. Similar effects were observed in GOFs activated by autoantibodies from GO patients' sera. Conclusions and Implications Our data support TSH and IGF‐1 receptors as therapeutic targets for GO, but reveal putative conditions for anti‐IGF‐1 receptor resistance. Combination treatments antagonizing both receptors yield additive effects by inhibiting crosstalk triggered by TSH receptor stimulatory antibodies. Combination therapy may be an effective strategy for dose reduction and/or compensate for any loss of anti‐IGF‐1 receptor efficacy. PMID:27987211

Prior Activation of Inositol 1,4,5-Trisphosphate Receptors Suppresses the Subsequent Induction of Long-Term Potentiation in Hippocampal CA1 Neurons

ERIC Educational Resources Information Center

Fujii, Satoshi; Yamazaki, Yoshihiko; Goto, Jun-Ichi; Fujiwara, Hiroki; Mikoshiba, Katsuhiko

2016-01-01

We investigated the role of inositol 1,4,5-trisphosphate receptors (IP3Rs) activated by preconditioning low-frequency afferent stimulation (LFS) in the subsequent induction of long-term potentiation (LTP) in CA1 neurons in hippocampal slices from mature guinea pigs. Induction of LTP in the field excitatory postsynaptic potential or the population…

Targeting the fibroblast growth factor receptors for the treatment of cancer.

PubMed

Lemieux, Steven M; Hadden, M Kyle

2013-06-01

Receptor tyrosine kinases (RTKs) are transmembrane proteins that play a critical role in stimulating signal transduction cascades to influence cell proliferation, growth, and differentiation and they have also been shown to promote angiogenesis when they are up-regulated or mutated. For this reason, their dysfunction has been implicated in the development of human cancer. Over the past decade, much attention has been devoted to developing inhibitors and antibodies against several classes of RTKs, including vascular endothelial growth factor receptors (VEGFRs), epidermal growth factor receptors (EGFRs), and platelet-derived growth factor receptors (PDGFRs). More recently, interest in the fibroblast growth factor receptor (FGFR) class of RTKs as a drug target for the treatment of cancer has emerged. Signaling through FGFRs is critical for normal cellular function and their dysregulation has been linked to various malignancies such as breast and prostate cancer. This review will focus on the current state of both small molecules and antibodies as FGFR inhibitors to provide insight into their development and future potential as anti-cancer agents.

Effects of age and insulin-like growth factor-1 on rat neurotrophin receptor expression after nerve injury.

PubMed

Luo, T David; Alton, Timothy B; Apel, Peter J; Cai, Jiaozhong; Barnwell, Jonathan C; Sonntag, William E; Smith, Thomas L; Li, Zhongyu

2016-10-01

Neurotrophin receptors, such as p75(NTR) , direct neuronal response to injury. Insulin-like growth factor-1 receptor (IGF-1R) mediates the increase in p75(NTR) during aging. The aim of this study was to examine the effect of aging and insulin-like growth factor-1 (IGF-1) treatment on recovery after peripheral nerve injury. Young and aged rats underwent tibial nerve transection with either local saline or IGF-1 treatment. Neurotrophin receptor mRNA and protein expression were quantified. Aged rats expressed elevated baseline IGF-1R (34% higher, P = 0.01) and p75(NTR) (68% higher, P < 0.01) compared with young rats. Post-injury, aged animals expressed significantly higher p75(NTR) levels (68.5% above baseline at 4 weeks). IGF-1 treatment suppressed p75(NTR) gene expression at 4 weeks (17.2% above baseline, P = 0.002) post-injury. Local IGF-1 treatment reverses age-related declines in recovery after peripheral nerve injuries by suppressing p75(NTR) upregulation and pro-apoptotic complexes. IGF-1 may be considered a viable adjuvant therapy to current treatment modalities. Muscle Nerve 54: 769-775, 2016. © 2016 Wiley Periodicals, Inc.

Esculetin attenuates receptor activator of nuclear factor kappa-B ligand-mediated osteoclast differentiation through c-Fos/nuclear factor of activated T-cells c1 signaling pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Baek, Jong Min; Park, Sun-Hyang; Cheon, Yoon-Hee

Esculetin exerts various biological effects on anti-oxidation, anti-tumors, and anti-inflammation. However, the involvement of esculetin in the bone metabolism process, particularly osteoclast differentiation has not yet been investigated. In the present study, we first confirmed the inhibitory effect of esculetin on receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast formation. We then revealed the relationship between esculetin and the expression of osteoclast-specific molecules to elucidate its underlying mechanisms. Esculetin interfered with the expression of c-Fos and nuclear factor of activated T cell c1 (NFATc1) both at the mRNA and protein level with no involvement in osteoclast-associated early signaling pathways, suppressingmore » the expression of various transcription factors exclusively expressed in osteoclasts such as tartrate-resistant acid phosphatase (Trap), osteoclast-associated receptor (Oscar), dendritic cell-specific transmembrane protein (Dcstamp), osteoclast stimulatory transmembrane protein (Ocstamp), cathepsin K, αvβ3 integrin, and calcitonin receptor (Ctr). Additionally, esculetin inhibited the formation of filamentous actin (F-actin) ring-positive osteoclasts during osteoclast differentiation. However, the development of F-actin structures and subsequent bone resorbing activity of mature osteoclasts, which are observed in osteoclast/osteoblast co-culture systems were not affected by esculetin. Taken together, our results indicate for the first time that esculetin inhibits RANKL-mediated osteoclastogenesis via direct suppression of c-Fos and NFATc1 expression and exerts an inhibitory effect on actin ring formation during osteoclastogenesis. - Highlights: • We first investigated the effects of esculetin on osteoclast differentiation and function. • Our data demonstrate for the first time that esculetin can suppress osteoclastogenesis in vitro. • Esculetin acts as an inhibitor of c-Fos and NFATc1 activation

Molecular Recognition of Corticotropin releasing Factor by Its G protein-coupled Receptor CRFR1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pioszak, Augen A.; Parker, Naomi R.; Suino-Powell, Kelly

2009-01-15

The bimolecular interaction between corticotropin-releasing factor (CRF), a neuropeptide, and its type 1 receptor (CRFR1), a class B G-protein-coupled receptor (GPCR), is crucial for activation of the hypothalamic-pituitary-adrenal axis in response to stress, and has been a target of intense drug design for the treatment of anxiety, depression, and related disorders. As a class B GPCR, CRFR1 contains an N-terminal extracellular domain (ECD) that provides the primary ligand binding determinants. Here we present three crystal structures of the human CRFR1 ECD, one in a ligand-free form and two in distinct CRF-bound states. The CRFR1 ECD adopts the alpha-beta-betaalpha fold observedmore » for other class B GPCR ECDs, but the N-terminal alpha-helix is significantly shorter and does not contact CRF. CRF adopts a continuous alpha-helix that docks in a hydrophobic surface of the ECD that is distinct from the peptide-binding site of other class B GPCRs, thereby providing a basis for the specificity of ligand recognition between CRFR1 and other class B GPCRs. The binding of CRF is accompanied by clamp-like conformational changes of two loops of the receptor that anchor the CRF C terminus, including the C-terminal amide group. These structural studies provide a molecular framework for understanding peptide binding and specificity by the CRF receptors as well as a template for designing potent and selective CRFR1 antagonists for therapeutic applications.« less

Platelet factor 4 (CXCL4) facilitates human macrophage infection with HIV-1 and potentiates virus replication.

PubMed

Schwartzkopff, Franziska; Grimm, Tobias A; Lankford, Carla S R; Fields, Karen; Wang, Jiun; Brandt, Ernst; Clouse, Kathleen A

2009-12-01

Platelet factor 4 (CXCL4), a member of the CXC chemokine subfamily released in high amounts by activated platelets, has been identified as a monocyte survival factor that induces monocyte differentiation into macrophages. Although CXCL4 has been shown to have biological effects unique to chemokines, nothing is known about the role of CXCL4-derived human macrophages or CXCL4 in human immunodeficiency virus (HIV) disease. In this study, CXCL4-derived macrophages are compared with macrophage-colony stimulating factor (M-CSF)-derived macrophages for their ability to support HIV-1 replication. We show that CXCL4-derived macrophages can be infected with macrophage-tropic HIV-1 that uses either CC-chemokine receptor 5 (CCR5) or CXC-chemokine receptor 4 (CXCR4) as a co-receptor for viral entry. We also find that M-CSF and the chemokines, monocyte chemoattractant protein 1 (MCP-1; CCL2) and macrophage-inflammatory-protein-1-alpha (MIP-1alpha; CCL3) are produced upon R5- and X4-tropic HIV-1 replication in both M-CSF- and CXCL4-derived human macrophages. In addition, CXCL4 added to M-CSF-derived macrophages after virus adsorption and maintained throughout the infection enhances HIV-1 replication. We thus propose a novel role for CXCL4 in HIV disease.

Optimization of 1H-indazol-3-amine derivatives as potent fibroblast growth factor receptor inhibitors.

PubMed

Cui, Jing; Peng, Xia; Gao, Dingding; Dai, Yang; Ai, Jing; Li, Yingxia

2017-08-15

Fibroblast growth factor receptor (FGFR) is a potential target for cancer therapy because of its critical role in promoting cancer formation and progression. In a continuing effort to improve the cellular activity of hit compound 7r bearing an indazole scaffold, which was previously discovered by our group, several compounds harnessing fluorine substituents were designed, synthesized and biological evaluated. Besides, the region extended out to the ATP binding pocket toward solvent was also explored. Among them, compound 2a containing 2,6-difluoro-3-methoxyphenyl residue exhibited the most potent activities (FGFR1: less than 4.1nM, FGFR2: 2.0±0.8nM). More importantly, compound 2a showed an improved antiproliferative effect against KG1 cell lines and SNU16 cell lines with IC 50 values of 25.3±4.6nM and 77.4±6.2nM respectively. Copyright © 2017. Published by Elsevier Ltd.

The cannabinoid receptor CB1 modulates the signaling properties of the lysophosphatidylinositol receptor GPR55.

PubMed

Kargl, Julia; Balenga, Nariman; Parzmair, Gerald P; Brown, Andrew J; Heinemann, Akos; Waldhoer, Maria

2012-12-28

The G protein-coupled receptor (GPCR) 55 (GPR55) and the cannabinoid receptor 1 (CB1R) are co-expressed in many tissues, predominantly in the central nervous system. Seven transmembrane spanning (7TM) receptors/GPCRs can form homo- and heteromers and initiate distinct signaling pathways. Recently, several synthetic CB1 receptor inverse agonists/antagonists, such as SR141716A, AM251, and AM281, were reported to activate GPR55. Of these, SR141716A was marketed as a promising anti-obesity drug, but was withdrawn from the market because of severe side effects. Here, we tested whether GPR55 and CB1 receptors are capable of (i) forming heteromers and (ii) whether such heteromers could exhibit novel signaling patterns. We show that GPR55 and CB1 receptors alter each others signaling properties in human embryonic kidney (HEK293) cells. We demonstrate that the co-expression of FLAG-CB1 receptors in cells stably expressing HA-GPR55 specifically inhibits GPR55-mediated transcription factor activation, such as nuclear factor of activated T-cells and serum response element, as well as extracellular signal-regulated kinases (ERK1/2) activation. GPR55 and CB1 receptors can form heteromers, but the internalization of both receptors is not affected. In addition, we observe that the presence of GPR55 enhances CB1R-mediated ERK1/2 and nuclear factor of activated T-cell activation. Our data provide the first evidence that GPR55 can form heteromers with another 7TM/GPCR and that this interaction with the CB1 receptor has functional consequences in vitro. The GPR55-CB1R heteromer may play an important physiological and/or pathophysiological role in tissues endogenously co-expressing both receptors.

The Cannabinoid Receptor CB1 Modulates the Signaling Properties of the Lysophosphatidylinositol Receptor GPR55*

PubMed Central

Kargl, Julia; Balenga, Nariman; Parzmair, Gerald P.; Brown, Andrew J.; Heinemann, Akos; Waldhoer, Maria

2012-01-01

The G protein-coupled receptor (GPCR) 55 (GPR55) and the cannabinoid receptor 1 (CB1R) are co-expressed in many tissues, predominantly in the central nervous system. Seven transmembrane spanning (7TM) receptors/GPCRs can form homo- and heteromers and initiate distinct signaling pathways. Recently, several synthetic CB1 receptor inverse agonists/antagonists, such as SR141716A, AM251, and AM281, were reported to activate GPR55. Of these, SR141716A was marketed as a promising anti-obesity drug, but was withdrawn from the market because of severe side effects. Here, we tested whether GPR55 and CB1 receptors are capable of (i) forming heteromers and (ii) whether such heteromers could exhibit novel signaling patterns. We show that GPR55 and CB1 receptors alter each others signaling properties in human embryonic kidney (HEK293) cells. We demonstrate that the co-expression of FLAG-CB1 receptors in cells stably expressing HA-GPR55 specifically inhibits GPR55-mediated transcription factor activation, such as nuclear factor of activated T-cells and serum response element, as well as extracellular signal-regulated kinases (ERK1/2) activation. GPR55 and CB1 receptors can form heteromers, but the internalization of both receptors is not affected. In addition, we observe that the presence of GPR55 enhances CB1R-mediated ERK1/2 and nuclear factor of activated T-cell activation. Our data provide the first evidence that GPR55 can form heteromers with another 7TM/GPCR and that this interaction with the CB1 receptor has functional consequences in vitro. The GPR55-CB1R heteromer may play an important physiological and/or pathophysiological role in tissues endogenously co-expressing both receptors. PMID:23161546

Heteromeric Canonical Transient Receptor Potential 1 and 4 Channels Play a Critical Role in Epileptiform Burst Firing and Seizure-Induced Neurodegeneration

PubMed Central

Phelan, Kevin D.; Mock, Matthew M.; Kretz, Oliver; Shwe, U. Thaung; Kozhemyakin, Maxim; Greenfield, L. John; Dietrich, Alexander; Birnbaumer, Lutz; Freichel, Marc; Flockerzi, Veit

2012-01-01

Canonical transient receptor potential channels (TRPCs) are receptor-operated cation channels that are activated in response to phospholipase C signaling. Although TRPC1 is ubiquitously expressed in the brain, TRPC4 expression is the most restrictive, with the highest expression level limited to the lateral septum. The subunit composition of neuronal TRPC channels remains uncertain because of conflicting data from recombinant expression systems. Here we report that the large depolarizing plateau potential that underlies the epileptiform burst firing induced by metabotropic glutamate receptor agonists in lateral septal neurons was completely abolished in TRPC1/4 double-knockout mice, and was abolished in 74% of lateral septal neurons in TRPC1 knockout mice. Furthermore, neuronal cell death in the lateral septum and the cornu ammonis 1 region of hippocampus after pilocarpine-induced severe seizures was significantly ameliorated in TRPC1/4 double-knockout mice. Our data suggest that both TRPC1 and TRPC4 are essential for an intrinsic membrane conductance mediating the plateau potential in lateral septal neurons, possibly as heteromeric channels. Moreover, excitotoxic neuronal cell death, an underlying process for many neurological diseases, is not mediated merely by ionotropic glutamate receptors but also by heteromeric TRPC channels activated by metabotropic glutamate receptors. TRPC channels could be an unsuspected but critical molecular target for clinical intervention for excitotoxicity. PMID:22144671

Heteromeric canonical transient receptor potential 1 and 4 channels play a critical role in epileptiform burst firing and seizure-induced neurodegeneration.

PubMed

Phelan, Kevin D; Mock, Matthew M; Kretz, Oliver; Shwe, U Thaung; Kozhemyakin, Maxim; Greenfield, L John; Dietrich, Alexander; Birnbaumer, Lutz; Freichel, Marc; Flockerzi, Veit; Zheng, Fang

2012-03-01

Canonical transient receptor potential channels (TRPCs) are receptor-operated cation channels that are activated in response to phospholipase C signaling. Although TRPC1 is ubiquitously expressed in the brain, TRPC4 expression is the most restrictive, with the highest expression level limited to the lateral septum. The subunit composition of neuronal TRPC channels remains uncertain because of conflicting data from recombinant expression systems. Here we report that the large depolarizing plateau potential that underlies the epileptiform burst firing induced by metabotropic glutamate receptor agonists in lateral septal neurons was completely abolished in TRPC1/4 double-knockout mice, and was abolished in 74% of lateral septal neurons in TRPC1 knockout mice. Furthermore, neuronal cell death in the lateral septum and the cornu ammonis 1 region of hippocampus after pilocarpine-induced severe seizures was significantly ameliorated in TRPC1/4 double-knockout mice. Our data suggest that both TRPC1 and TRPC4 are essential for an intrinsic membrane conductance mediating the plateau potential in lateral septal neurons, possibly as heteromeric channels. Moreover, excitotoxic neuronal cell death, an underlying process for many neurological diseases, is not mediated merely by ionotropic glutamate receptors but also by heteromeric TRPC channels activated by metabotropic glutamate receptors. TRPC channels could be an unsuspected but critical molecular target for clinical intervention for excitotoxicity.

Laminar shear stress regulates endothelial kinin B1 receptor expression and function: potential implication in atherogenesis

PubMed Central

Duchene, Johan; Cayla, Cécile; Vessillier, Sandrine; Scotland, Ramona; Yamashiro, Kazuo; Lecomte, Florence; Syed, Irfan; Vo, Phuong; Marrelli, Alessandra; Pitzalis, Costantino; Cipollone, Francesco; Schanstra, Joost; Bascands, Jean-Loup; Hobbs, Adrian J; Perretti, Mauro; Ahluwalia, Amrita

2009-01-01

OBJECTIVE The pro-inflammatory phenotype induced by low laminar shear stress (LSS) is implicated in atherogenesis. The kinin B1 receptor (B1R), known to be induced by inflammatory stimuli, exerts many pro-inflammatory effects including vasodilatation and leukocyte recruitment. We investigated whether low LSS is a stimulus for endothelial B1R expression and function. METHODS AND RESULTS Human and mouse atherosclerotic plaques expressed high level of B1R mRNA and protein. In addition, B1R expression was upregulated in the aortic arch (low LSS region) of ApoE-/- mice fed a high fat diet compared to vascular regions of high LSS and animals fed normal chow. Of interest, a greater expression of B1R was noticed in endothelial cells from regions of low LSS in aortic arch of ApoE-/- mice. B1R was also upregulated in human umbilical vein endothelial cells (HUVEC) exposed to low LSS (0-2dyn/cm2) compared to physiological LSS (6-10dyn/cm2): an effect similarly evident in murine vascular tissue perfused ex vivo. Functionally, B1R activation increased prostaglandin and CXCL5 expression in cells exposed to low, but not physiological, LSS. IL-1β and ox-LDL induced B1R expression and function in HUVECs, a response substantially enhanced under low LSS conditions and inhibited by blockade of NFκB activation. CONCLUSION Herein, we show that LSS is a major determinant of functional B1R expression in endothelium. Furthermore, whilst physiological high LSS is a powerful repressor of this inflammatory receptor, low LSS at sites of atheroma are associated with substantial upregulation, identifying this receptor as a potential therapeutic target. CONDENSED ABSTRACT Low laminar shear stress (LSS) underlies the pro-inflammatory processes in atherogenesis. Herein, we demonstrate that whilst physiological LSS represses inflammatory kinin B1 receptor (B1R) expression/function, low atherogenic LSS is associated with profound upregulation of both in atherosclerosis in both humans and animal

Featured Article: Nuclear export of opioid growth factor receptor is CRM1 dependent.

PubMed

Kren, Nancy P; Zagon, Ian S; McLaughlin, Patricia J

2016-02-01

Opioid growth factor receptor (OGFr) facilitates growth inhibition in the presence of its specific ligand opioid growth factor (OGF), chemically termed [Met(5)]-enkephalin. The function of the OGF-OGFr axis requires the receptor to translocate to the nucleus. However, the mechanism of nuclear export of OGFr is unknown. In this study, endogenous OGFr, as well as exogenously expressed OGFr-EGFP, demonstrated significant nuclear accumulation in response to leptomycin B (LMB), an inhibitor of CRM1-dependent nuclear export, suggesting that OGFr is exported in a CRM1-dependent manner. One consensus sequence for a nuclear export signal (NES) was identified. Mutation of the associated leucines, L217 L220 L223 and L225, to alanine resulted in decreased nuclear accumulation. NES-EGFP responded to LMB, indicating that this sequence is capable of functioning as an export signal in isolation. To determine why the sequence functions differently in isolation than as a full length protein, the localization of subNES was evaluated in the presence and absence of MG132, a potent inhibitor of proteosomal degradation. MG132 had no effect of subNES localization. The role of tandem repeats located at the C-terminus of OGFr was examined for their role in nuclear trafficking. Six of seven tandem repeats were removed to form deltaTR. DeltaTR localized exclusively to the nucleus indicating that the tandem repeats may contribute to the localization of the receptor. Similar to the loss of cellular proliferation activity (i.e. inhibition) recorded with subNES, deltaTR also demonstrated a significant loss of inhibitory activity indicating that the repeats may be integral to receptor function. These experiments reveal that OGFr contains one functional NES, L217 L220 L223 and L225 and can be exported from the nucleus in a CRM1-dependent manner. © 2015 by the Society for Experimental Biology and Medicine.

Genetic polymorphisms and protein structures in growth hormone, growth hormone receptor, ghrelin, insulin-like growth factor 1 and leptin in Mehraban sheep.

PubMed

Bahrami, A; Behzadi, Sh; Miraei-Ashtiani, S R; Roh, S-G; Katoh, K

2013-09-15

The somatotropic axis, the control system for growth hormone (GH) secretion and its endogenous factors involved in the regulation of metabolism and energy partitioning, has promising potentials for producing economically valuable traits in farm animals. Here we investigated single nucleotide polymorphisms (SNPs) of the genes of factors involved in the somatotropic axis for growth hormone (GH1), growth hormone receptor (GHR), ghrelin (GHRL), insulin-like growth factor 1 (IGF-I) and leptin (LEP), using polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP) and DNA sequencing methods in 452 individual Mehraban sheep. A nonradioactive method to allow SSCP detection was used for genomic DNA and PCR amplification of six fragments: exons 4 and 5 of GH1; exon 10 of GH receptor (GHR); exon 1 of ghrelin (GHRL); exon 1 of insulin-like growth factor-I (IGF-I), and exon 3 of leptin (LEP). Polymorphisms were detected in five of the six PCR products. Two electrophoretic patterns were detected for GH1 exon 4. Five conformational patterns were detected for GH1 exon 5 and LEP exon 3, and three for IGF-I exon 1. Only GHR and GHRL were monomorphic. Changes in protein structures due to variable SNPs were also analyzed. The results suggest that Mehraban sheep, a major breed that is important for the animal industry in Middle East countries, has high genetic variability, opening interesting prospects for future selection programs and preservation strategies. Copyright © 2013 Elsevier B.V. All rights reserved.

Insulin-like growth factor-1 prevents dorsal root ganglion neuronal tyrosine kinase receptor expression alterations induced by dideoxycytidine in vitro.

PubMed

Liu, Huaxiang; Lu, Jing; He, Yong; Yuan, Bin; Li, Yizhao; Li, Xingfu

2014-03-01

Dideoxycytidine (zalcitabine, ddC) produces neurotoxic effects. It is particularly important to understand the toxic effects of ddC on different subpopulations of dorsal root ganglion (DRG) neurons which express distinct tyrosine kinase receptor (Trk) and to find therapeutic factors for prevention and therapy for ddC-induced peripheral sensory neuropathy. Insulin-like growth factor-1 (IGF-1) has been shown to have neurotrophic effects on DRG sensory neurons. However, little is known about the effects of ddC on distinct Trk (TrkA, TrkB, and TrkC) expression in DRG neurons and the neuroprotective effects of IGF-1 on ddC-induced neurotoxicity. Here, we have tested the extent to which the expression of TrkA, TrkB, and TrkC receptors in primary cultured DRG neurons is affected by ddC in the presence or absence of IGF-1. In this experiment, we found that exposure of 5, 25, and 50 μmol/L ddC caused a dose-dependent decrease of the mRNA, protein, and the proportion of TrkA-, TrkB-, and TrkC-expressing neurons. IGF-1 (20 nmol/L) could partially reverse the decrease of TrkA and TrkB, but not TrkC, expression with ddC exposure. The phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 (10 μmol/L) blocked the effects of IGF-1. These results suggested that the subpopulations of DRG neurons which express distinct TrkA, TrkB, and TrkC receptors were affected by ddC exposure. IGF-1 might relieve the ddC-induced toxicity of TrkA- and TrkB-, but not TrkC-expressing DRG neurons. These data offer new clues for a better understanding of the association of ddC with distinct Trk receptor expression and provide new evidence of the potential therapeutic role of IGF-1 on ddC-induced neurotoxicity.

A desensitization-selective potentiator of AMPA-type glutamate receptors

PubMed Central

Sekiguchi, Masayuki; Nishikawa, Kaori; Aoki, Shunsuke; Wada, Keiji

2002-01-01

We examined the effects of PEPA, an allosteric potentiator of AMPA receptors, on AMPA receptor kinetics. PEPA did not affect the deactivation of glutamate responses but potently attenuated the extent of receptor desensitization without slowing the onset of desensitization in most of the recombinant AMPA receptors (GluR1-flip, GluR1-flop, GluR3-flip, GluR3-flip + GluR2-flip, and GluR3-flop + GluR2-flop) expressed in Xenopus oocytes. For the GluR3-flop subunit, PEPA attenuated the extent of desensitization and only weakly prolonged deactivation (1.3 fold). PEPA did not significantly affect recovery from desensitization in oocytes expressing GluR3-flip, GluR1-flop, and GluR1-flop, but weakly accelerated (2.6 fold) recovery from desensitization in oocytes expressing GluR3-flop. PEPA's effect on desensitization of GluR3-flop-containing receptors is unique in that onset is very slow. Simulation studies using simplified kinetic models for AMPA receptors are utilized to explore the differential effects of PEPA on GluR3-flip and -flop. It is possible to simulate the action on GluR3-flip by modulating two rate constants in a 12-state kinetic model. For simulation of the action on GluR3-flop, the 12-state kinetic model is not enough, and it is necessary to invoke a 13th state, a PEPA-bound receptor to which glutamate cannot bind. These results suggest that attenuation of extent of desensitization represents the principal mechanism underlying the potentiation of AMPA receptors by PEPA, and that PEPA exhibits different mechanisms with respect to GluR3-flip and GluR3-flop. PMID:12145103

Selective over-expression of fibroblast growth factor receptors 1 and 4 in clinical prostate cancer.

PubMed

Sahadevan, K; Darby, S; Leung, H Y; Mathers, M E; Robson, C N; Gnanapragasam, V J

2007-09-01

Fibroblast growth factor receptors (FGFRs) mediate the tumourigenic effects of FGFs in prostate cancer. These receptors are therefore potential therapeutic targets in the development of inhibitors to this pathway. To identify the most relevant targets, we simultaneously investigated FGFR1-4 expression using a prostate cancer tissue microarray (TMA) and in laser capture microdissected (LCM) prostate epithelial cells. In malignant prostates (n = 138) we observed significant FGFR1 and FGFR4 protein over-expression in comparison with benign prostates (n = 58; p < 0.0001). FGFR1 was expressed at high levels in the majority of tumours (69% of grade 3 or less, 74% of grade 4 and 70% of grade 5), while FGFR4 was strongly expressed in 83% of grade 5 cancers but in only 25% of grade 1-3 cancers (p < 0.0001). At the transcript level we observed a similar pattern, with FGFR1 and FGFR4 mRNA over-expressed in malignant epithelial cells compared to benign cells (p < 0.0005 and p < 0.05, respectively). While total FGFR2 was increased in some cancers, there was no association between expression and tumour grade or stage. Transcript analysis, however, revealed a switch in the predominant isoform expressed from FGFR2IIIb to FGFR2IIIc among malignant epithelial cells. In contrast, protein and transcript expression of FGFR3 was very similar between benign and cancer biopsies. The functional effect of targeting FGFR4 in prostate cancer cells has not previously been investigated. In in vitro experiments, suppression of FGFR4 by RNA interference effectively blocked prostate cancer cell proliferation (p < 0.0001) and invasion (p < 0.001) in response to exogenous stimulation. This effect was evident regardless of whether the cells expressed the FGFR4 Arg388 or Gly388 allele. In parallel experiments, FGFR3 suppression had no discernible effect on cancer cell behaviour. These results suggest evidence of selective over-expression of FGFR1 and FGFR4 in clinical prostate cancer and support the

Novel Function for Vascular Endothelial Growth Factor Receptor-1 on Epidermal Keratinocytes

PubMed Central

Wilgus, Traci A.; Matthies, Annette M.; Radek, Katherine A.; Dovi, Julia V.; Burns, Aime L.; Shankar, Ravi; DiPietro, Luisa A.

2005-01-01

Vascular endothelial growth factor (VEGF-A), a potent stimulus for angiogenesis, is up-regulated in the skin after wounding. Although studies have shown that VEGF is important for wound repair, it is unclear whether this is based solely on its ability to promote angiogenesis or if VEGF can also promote healing by acting directly on non-endothelial cell types. By immunohistochemistry and reverse transcriptase-polymerase chain reaction, expression of VEGF receptor-1 (VEGFR-1), but not VEGFR-2, was detected in murine keratinocytes during wound repair and in normal human epidermal keratinocytes (NHEKs). The presence of VEGF receptors on NHEKs was verified by binding studies with 125I-VEGF. In vitro, VEGF stimulated the proliferation of NHEKs, an effect that could be blocked by treatment with neutralizing VEGFR-1 antibodies. A role for VEGFR-1 in keratinocytes was also shown in vivo because treatment of excisional wounds with neutralizing VEGFR-1 antibodies delayed re-epithelialization. Treatment with anti-VEGFR-1 antibodies also reduced the number of proliferating keratinocytes at the leading edge of the wound, suggesting that VEGF sends a proliferative signal to these cells. Together, these data describe a novel role for VEGFR-1 in keratinocytes and suggest that VEGF may play several roles in cutaneous wound repair. PMID:16251410

Breaking the barriers: New role for insulin-like growth factor 1 receptor in vascular permeability.

PubMed

Xavier, Sandhya

2015-05-01

This commentary highlights the article by Liang et al that describes a critical role for insulin-like growth factor 1 receptor in the progression of chronic kidney disease. Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Protein partners in the life history of activated fibroblast growth factor receptors.

PubMed

Vecchione, Anna; Cooper, Helen J; Trim, Kimberley J; Akbarzadeh, Shiva; Heath, John K; Wheldon, Lee M

2007-12-01

Fibroblast growth factor receptors (FGFRs) are a family of four transmembrane (TM) receptor tyrosine kinases (RTKs) which bind to a large family of fibroblast growth factor (FGF) ligands with varying affinity and specificity. FGFR signaling regulates many physiological and pathological processes in development and tissue homeostasis. Understanding FGFR signaling processes requires the identification of partner proteins which regulate receptor function and biological outputs. In this study, we employ an epitope-tagged, covalently dimerized, and constitutively activated form of FGFR1 to identify potential protein partners by MS. By this approach, we sample candidate FGFR effectors throughout the life history of the receptor. Functional classification of the partners identified revealed specific subclasses involved in protein biosynthesis and folding; structural and regulatory components of the cytoskeleton; known signaling effectors and small GTPases implicated in endocytosis and vesicular trafficking. The kinase dependency of the interaction was determined for a subset of previously unrecognized partners by coimmunoprecipitation, Western blotting, and immunocytochemistry. From this group, the small GTPase Rab5 was selected for functional interrogation. We show that short hairpin (sh) RNA-mediated depletion of Rab5 attenuates the activation of the extracellular-regulated kinase (ERK) 1/2 pathway by FGFR signaling. The strategic approach adopted in this study has revealed bona fide novel effectors of the FGFR signaling pathway.

Overexpression of androgen receptor and forkhead-box A1 protein in apocrine breast carcinoma.

PubMed

Sasahara, Manami; Matsui, Akira; Ichimura, Yoshiko; Hirakata, Yuuko; Murata, Yuuya; Marui, Eiji

2014-03-01

Apocrine breast carcinoma often lacks estrogen receptor (ER), progesterone receptor (PgR), and human epidermal growth factor receptor type-2 (HER2) expression. Accordingly, development of a new treatment strategy is important for this type of cancer. The growth stimulus through the androgen receptor (AR) can be a candidate for targeted treatment. Therefore, we examined the factors related to AR transcription. We immunohistochemically evaluated 54 apocrine cancer lesions for ER, PgR, AR, HER2, Ki-67, forkhead-box protein A1 (FOXA1), and prostate-specific antigen (PSA) expression. ER, PgR, and HER2 were expressed at a low level, thus 44 out of 54 (81.4%) cases were of triple-negative breast cancer. AR, PSA and FOXA1 were expressed in 100% (54/54), 48% (26/54) and 93% (50/54) of cases, respectively. Most of apocrine breast carcinomas were immunohistochemically-positive for AR and FOXA1. Anti-androgenic therapies can potentially serve as a cancer-targeting therapy for apocrine breast carcinoma.

The sympathetic nervous system is controlled by transient receptor potential vanilloid 1 in the regulation of body temperature

PubMed Central

Alawi, Khadija M.; Aubdool, Aisah A.; Liang, Lihuan; Wilde, Elena; Vepa, Abhinav; Psefteli, Maria-Paraskevi; Brain, Susan D.; Keeble, Julie E.

2015-01-01

Transient receptor potential vanilloid 1 (TRPV1) is involved in sensory nerve nociceptive signaling. Recently, it has been discovered that TRPV1 receptors also regulate basal body temperature in multiple species from mice to humans. In the present study, we investigated whether TRPV1 modulates basal sympathetic nervous system (SNS) activity. C57BL6/J wild-type (WT) mice and TRPV1 knockout (KO) mice were implanted with radiotelemetry probes for measurement of core body temperature. AMG9810 (50 mg/kg) or vehicle (2% DMSO/5% Tween 80/10 ml/kg saline) was injected intraperitoneally. Adrenoceptor antagonists or vehicle (5 ml/kg saline) was injected subcutaneously. In WT mice, the TRPV1 antagonist, AMG9810, caused significant hyperthermia, associated with increased noradrenaline concentrations in brown adipose tissue. The hyperthermia was significantly attenuated by the β-adrenoceptor antagonist propranolol, the mixed α-/β-adrenoceptor antagonist labetalol, and the α1-adrenoceptor antagonist prazosin. TRPV1 KO mice have a normal basal body temperature, indicative of developmental compensation. d-Amphetamine (potent sympathomimetic) caused hyperthermia in WT mice, which was reduced in TRPV1 KO mice, suggesting a decreased sympathetic drive in KOs. This study provides new evidence that TRPV1 controls thermoregulation upstream of the SNS, providing a potential therapeutic target for sympathetic hyperactivity thermoregulatory disorders.—Alawi, K. M., Aubdool, A. A., Liang, L., Wilde, E., Vepa, A., Psefteli, M.-P., Brain, S. D., Keeble, J. E. The sympathetic nervous system is controlled by transient receptor potential vanilloid 1 in the regulation of body temperature. PMID:26136480

Neuronal expression of fibroblast growth factor receptors in zebrafish.

PubMed

Rohs, Patricia; Ebert, Alicia M; Zuba, Ania; McFarlane, Sarah

2013-12-01

Fibroblast growth factor (FGF) signaling is important for a host of developmental processes such as proliferation, differentiation, tissue patterning, and morphogenesis. In vertebrates, FGFs signal through a family of four fibroblast growth factor receptors (FGFR 1-4), one of which is duplicated in zebrafish (FGFR1). Here we report the mRNA expression of the five known zebrafish fibroblast growth factor receptors at five developmental time points (24, 36, 48, 60, and 72h postfertilization), focusing on expression within the central nervous system. We show that the receptors have distinct and dynamic expression in the developing zebrafish brain, eye, inner ear, lateral line, and pharynx. In many cases, the expression patterns are similar to those of homologous FGFRs in mouse, chicken, amphibians, and other teleosts. Copyright © 2013 Elsevier B.V. All rights reserved.

Structural Determinants of the Transient Receptor Potential 1 (TRPV1) Channel Activation by Phospholipid Analogs*

PubMed Central

Morales-Lázaro, Sara L.; Serrano-Flores, Barbara; Llorente, Itzel; Hernández-García, Enrique; González-Ramírez, Ricardo; Banerjee, Souvik; Miller, Duane; Gududuru, Veeresh; Fells, James; Norman, Derek; Tigyi, Gabor; Escalante-Alcalde, Diana; Rosenbaum, Tamara

2014-01-01

The transient receptor potential vanilloid 1 (TRPV1) ion channel is a polymodal protein that responds to various stimuli, including capsaicin (the pungent compound found in chili peppers), extracellular acid, and basic intracellular pH, temperatures close to 42 °C, and several lipids. Lysophosphatidic acid (LPA), an endogenous lipid widely associated with neuropathic pain, is an agonist of the TRPV1 channel found in primary afferent nociceptors and is activated by other noxious stimuli. Agonists or antagonists of lipid and other chemical natures are known to possess specific structural requirements for producing functional effects on their targets. To better understand how LPA and other lipid analogs might interact and affect the function of TRPV1, we set out to determine the structural features of these lipids that result in the activation of TRPV1. By changing the acyl chain length, saturation, and headgroup of these LPA analogs, we established strict requirements for activation of TRPV1. Among the natural LPA analogs, we found that only LPA 18:1, alkylglycerophosphate 18:1, and cyclic phosphatidic acid 18:1, all with a monounsaturated C18 hydrocarbon chain activate TRPV1, whereas polyunsaturated and saturated analogs do not. Thus, TRPV1 shows a more restricted ligand specificity compared with LPA G-protein-coupled receptors. We synthesized fatty alcohol phosphates and thiophosphates and found that many of them with a single double bond in position Δ9, 10, or 11 and Δ9 cyclopropyl group can activate TRPV1 with efficacy similar to capsaicin. Finally, we developed a pharmacophore and proposed a mechanistic model for how these lipids could induce a conformational change that activates TRPV1. PMID:25035428

Tumour necrosis factor-α regulates human eosinophil apoptosis via ligation of TNF-receptor 1 and balance between NF-κB and AP-1.

PubMed

Kankaanranta, Hannu; Ilmarinen, Pinja; Zhang, Xianzhi; Adcock, Ian M; Lahti, Aleksi; Barnes, Peter J; Giembycz, Mark A; Lindsay, Mark A; Moilanen, Eeva

2014-01-01

Eosinophils play a central role in asthma. The present study was performed to investigate the effect of tumour necrosis factor-α (TNF-α) on longevity of isolated human eosinophils. In contrast to Fas, TNF-α inhibited eosinophil apoptosis as evidenced by a combination of flow cytometry, DNA fragmentation assay and morphological analyses. The effect of TNF-α on eosinophil apoptosis was reversed by a TNF-α neutralising antibody. The anti-apoptotic effect of TNF-α was not due to autocrine release of known survival-prolonging cytokines interleukins 3 and 5 or granulocyte-macrophage-colony-stimulating factor as their neutralisation did not affect the effect of TNF-α. The anti-apoptotic signal was mediated mainly by the TNF-receptor 1. TNF-α induced phosphorylation and degradation of IκB and an increase in NF-κB DNA-binding activity. The survival-prolonging effect of TNF-α was reversed by inhibitors of NF-κB pyrrolidinedithiocarbamate and gliotoxin and by an inhibitor of IκB kinase, BMS-345541. TNF-α induced also an increase in AP-1 DNA-binding activity and the antiapoptotic effect of TNF-α was potentiated by inhibitors of AP-1, SR 11302 and tanshinone IIA and by an inhibitor of c-jun-N-terminal kinase, SP600125, which is an upstream kinase activating AP-1. Our results thus suggest that TNF-α delays human eosinophil apoptosis via TNF-receptor 1 and the resulting changes in longevity depend on yin-yang balance between activation of NF-κB and AP-1.

Something old, something new and something borrowed: emerging paradigm of insulin-like growth factor type 1 receptor (IGF-1R) signaling regulation.

PubMed

Girnita, Leonard; Worrall, Claire; Takahashi, Shin-Ichiro; Seregard, Stefan; Girnita, Ada

2014-07-01

The insulin-like growth factor type 1 receptor (IGF-1R) plays a key role in the development and progression of cancer; however, therapeutics targeting it have had disappointing results in the clinic. As a receptor tyrosine kinase (RTK), IGF-1R is traditionally described as an ON/OFF system, with ligand stabilizing the ON state and exclusive kinase-dependent signaling activation. Newly added to the traditional model, ubiquitin-mediated receptor downregulation and degradation was originally described as a response to ligand/receptor interaction and thus inseparable from kinase signaling activation. Yet, the classical model has proven over-simplified and insufficient to explain experimental evidence accumulated over the last decade, including kinase-independent signaling, unbalanced signaling, or dissociation between signaling and receptor downregulation. Based on the recent findings that IGF-1R "borrows" components of G-protein coupled receptor (GPCR) signaling, including β-arrestins and G-protein-related kinases, we discuss the emerging paradigm for the IGF-1R as a functional RTK/GPCR hybrid, which integrates the kinase signaling with the IGF-1R canonical GPCR characteristics. The contradictions to the classical IGF-1R signaling concept as well as the design of anti-IGF-1R therapeutics treatment are considered in the light of this paradigm shift and we advocate recognition of IGF-1R as a valid target for cancer treatment.

Autoantibodies against β1 receptor and AT1 receptor in type 2 diabetes patients with left ventricular dilatation.

PubMed

Zhao, Linshuang; Xu, Chunyan; Xu, Jinling

2014-01-01

To explore the relationship between the autoantibodies against the β1 and AT1 receptors and left ventricular dilatation in patients with type 2 diabetes (T2DM). The autoantibodies against the β1 and angiotensin II type 1 (AT1) receptors of T2DM patients with and without hypertension were screened by ELISA. Multiple logistic regression was used to analyze the risk factors for left ventricular dilatation. The reversing effect of left ventricular dilatation was evaluated after receptor blocker treatment. The positive rates of autoantibodies against the β1 and AT1 receptors (43.0 and 44.1%, respectively) in T2DM patients with hypertension were significantly higher than those in normotensive patients (16.0 and 10.4%, respectively; all p < 0.01). Furthermore, among T2DM patients with hypertension, the positive rates (61.4 and 64.9%, respectively) in patients with left ventricular dilatation were remarkably higher than those with normal left ventricular dimensions (34.4 and 36.1%, respectively; all p < 0.01). The presence of β1 receptor antibody and AT1 receptor antibody were risk factors for left ventricular dilatation (p < 0.05). The curative effect of metoprolol tartrate and valsartan in reversing left ventricular hypertrophy in the group positive for autoantibodies was much better than in the negative group. The findings show that autoantibodies against the β1 and AT1 receptors may play a role in predicting left ventricular dilatation in T2DM patients in combination with hypertension. Metoprolol tartrate and valsartan are effective and safe in the treatment of these patients. © 2014 S. Karger AG, Basel.

Chewing rescues stress-suppressed hippocampal long-term potentiation via activation of histamine H1 receptor.

PubMed

Ono, Yumie; Kataoka, Tsuyoshi; Miyake, Shinjiro; Sasaguri, Kenichi; Sato, Sadao; Onozuka, Minoru

2009-08-01

We have previously found in rats that chewing, an active behavioral strategy to cope with a stressful situation, rescues long-term potentiation (LTP) in the hippocampus through activating stress-suppressed N-methyl-D-aspartate (NMDA) receptor function. To further examine the mechanisms underlying this ameliorative effect of chewing, we studied the involvement of the histaminergic system, which has been shown to be activated by mastication, in the LTP of hippocampal slices of rats that were allowed to chew a wooden stick during exposure to immobilization stress. Chewing failed to rescue stress-suppressed LTP in the rats treated with histamine H1 receptor (H1R) antagonist pyrilamine (5 mg/kg, i.p.) before exposure to stress, although administration of pyrilamine did not affect LTP in naive rats and in stressed rats that did not chew. However, when pyrilamine was administrated immediately after exposure to stress, chewing rescued LTP whose magnitude was statistically comparable to that in the rats that chewed without drug treatment. These results suggest that chewing-induced histamine release in the hippocampus and the subsequent H1 receptor activation may be essential to rescue stress-suppressed synaptic plasticity.

Epidermal Growth Factor Receptor Transactivation by the Cannabinoid Receptor (CB1) and Transient Receptor Potential Vanilloid 1 (TRPV1) Induces Differential Responses in Corneal Epithelial Cells

DTIC Science & Technology

2010-01-01

induced Ca2þ signaling as well as phospholipase D (PLD)-mediated phosphatidic acid formation (Islam and Akhtar, 2000; Kang et al., 2000, 2001; Mazie et...Epithelial cell motility is triggered by activation of the EGF receptor through phosphatidic acid signaling. J. Cell Sci. 119, 1645e1654. McIntosh, B.T...buffer. Cell lysates were centrifuged and supernatants were collected for measuring proteins with a bichinchoninic acid assay (BCA) protein assay kit

Loss of Dlg-1 in the Mouse Lens Impairs Fibroblast Growth Factor Receptor Signaling

PubMed Central

Lee, SungKyoung; Griep, Anne E.

2014-01-01

Coordination of cell proliferation, differentiation and survival is essential for normal development and maintenance of tissues in the adult organism. Growth factor receptor tyrosine kinase signaling pathways and planar cell polarity pathways are two regulators of many developmental processes. We have previously shown through analysis of mice conditionally null in the lens for the planar cell polarity gene (PCP), Dlg-1, that Dlg-1 is required for fiber differentiation. Herein, we asked if Dlg-1 is a regulator of the Fibroblast growth factor receptor (Fgfr) signaling pathway, which is known to be required for fiber cell differentiation. Western blot analysis of whole fiber cell extracts from control and Dlg-1 deficient lenses showed that levels of the Fgfr signaling intermediates pErk, pAkt, and pFrs2α, the Fgfr target, Erm, and the fiber cell specific protein, Mip26, were reduced in the Dlg-1 deficient fiber cells. The levels of Fgfr2 were decreased in Dlg-1 deficient lenses compared to controls. Conversely, levels of Fgfr1 in Dlg-1 deficient lenses were increased compared to controls. The changes in Fgfr levels were found to be specifically in the triton insoluble, cytoskeletal associated fraction of Dlg-1 deficient lenses. Immunofluorescent staining of lenses from E13.5 embryos showed that expression levels of pErk were reduced in the transition zone, a region of the lens that exhibits PCP, in the Dlg-1 deficient lenses as compared to controls. In control lenses, immunofluorescent staining for Fgfr2 was observed in the epithelium, transition zone and fibers. By E13.5, the intensity of staining for Fgfr2 was reduced in these regions of the Dlg-1 deficient lenses. Thus, loss of Dlg-1 in the lens impairs Fgfr signaling and leads to altered levels of Fgfrs, suggesting that Dlg-1 is a modulator of Fgfr signaling pathway at the level of the receptors and that Dlg-1 regulates fiber cell differentiation through its role in PCP. PMID:24824078

Breast density and parenchymal texture measures as potential risk factors for estrogen-receptor positive breast cancer

NASA Astrophysics Data System (ADS)

Keller, Brad M.; Chen, Jinbo; Conant, Emily F.; Kontos, Despina

2014-03-01

Accurate assessment of a woman's risk to develop specific subtypes of breast cancer is critical for appropriate utilization of chemopreventative measures, such as with tamoxifen in preventing estrogen-receptor positive breast cancer. In this context, we investigate quantitative measures of breast density and parenchymal texture, measures of glandular tissue content and tissue structure, as risk factors for estrogen-receptor positive (ER+) breast cancer. Mediolateral oblique (MLO) view digital mammograms of the contralateral breast from 106 women with unilateral invasive breast cancer were retrospectively analyzed. Breast density and parenchymal texture were analyzed via fully-automated software. Logistic regression with feature selection and was performed to predict ER+ versus ER- cancer status. A combined model considering all imaging measures extracted was compared to baseline models consisting of density-alone and texture-alone features. Area under the curve (AUC) of the receiver operating characteristic (ROC) and Delong's test were used to compare the models' discriminatory capacity for receptor status. The density-alone model had a discriminatory capacity of 0.62 AUC (p=0.05). The texture-alone model had a higher discriminatory capacity of 0.70 AUC (p=0.001), which was not significantly different compared to the density-alone model (p=0.37). In contrast the combined density-texture logistic regression model had a discriminatory capacity of 0.82 AUC (p<0.001), which was statistically significantly higher than both the density-alone (p<0.001) and texture-alone regression models (p=0.04). The combination of breast density and texture measures may have the potential to identify women specifically at risk for estrogen-receptor positive breast cancer and could be useful in triaging women into appropriate risk-reduction strategies.

Computationally Discovered Potentiating Role of Glycans on NMDA Receptors

NASA Astrophysics Data System (ADS)

Sinitskiy, Anton V.; Stanley, Nathaniel H.; Hackos, David H.; Hanson, Jesse E.; Sellers, Benjamin D.; Pande, Vijay S.

2017-04-01

N-methyl-D-aspartate receptors (NMDARs) are glycoproteins in the brain central to learning and memory. The effects of glycosylation on the structure and dynamics of NMDARs are largely unknown. In this work, we use extensive molecular dynamics simulations of GluN1 and GluN2B ligand binding domains (LBDs) of NMDARs to investigate these effects. Our simulations predict that intra-domain interactions involving the glycan attached to residue GluN1-N440 stabilize closed-clamshell conformations of the GluN1 LBD. The glycan on GluN2B-N688 shows a similar, though weaker, effect. Based on these results, and assuming the transferability of the results of LBD simulations to the full receptor, we predict that glycans at GluN1-N440 might play a potentiator role in NMDARs. To validate this prediction, we perform electrophysiological analysis of full-length NMDARs with a glycosylation-preventing GluN1-N440Q mutation, and demonstrate an increase in the glycine EC50 value. Overall, our results suggest an intramolecular potentiating role of glycans on NMDA receptors.

Behavioral, biological, and chemical perspectives on targeting CRF1 receptor antagonists to treat alcoholism

PubMed Central

Zorrilla, Eric P.; Heilig, Markus; de Wit, Harriet; Shaham, Yavin

2013-01-01

Background Alcohol use disorders are chronic disabling conditions for which existing pharmacotherapies have only modest efficacy. In the present review, derived from the 2012 Behavior, Biology and Chemistry “Translational Research in Addiction” symposium, we summarize the anti-relapse potential of corticotropin-releasing factor type 1 (CRF1) receptor antagonists to reduce negative emotional symptoms of acute and protracted alcohol withdrawal and stress-induced relapse to alcohol seeking. Methods We review the biology of CRF1 systems, the activity of CRF1 receptor antagonists in animal models of anxiolytic and antidepressant activity, and experimental findings in alcohol addiction models. We also update the clinical trial status of CRF1 receptor antagonists, including pexacerfont (BMS-562086), emicerfont (GW876008), verucerfont (GSK561679), CP316311, SSR125543A, R121919/NBI30775, R317573/19567470/CRA5626, and ONO-2333Ms. Finally, we discuss the potential heterogeneity and pharmacogenomics of CRF1 receptor pharmacotherapy for alcohol dependence. Results The evidence suggests that brain penetrant-CRF1 receptor antagonists have therapeutic potential for alcohol dependence. Lead compounds with clinically desirable pharmacokinetic properties now exist, and longer receptor residence rates (i.e., slow dissociation) may predict greater CRF1 receptor antagonist efficacy. Functional variants in genes that encode CRF system molecules, including polymorphisms in Crhr1 (rs110402, rs1876831, rs242938) and Crhbp genes (rs10055255, rs3811939) may promote alcohol seeking and consumption by altering basal or stress-induced CRF system activation. Conclusions Ongoing clinical trials with pexacerfont and verucerfont in moderately to highly severe dependent anxious alcoholics may yield insight as to the role of CRF1 receptor antagonists in a personalized medicine approach to treat drug or alcohol dependence. PMID:23294766

Potentiation of NMDA receptor-mediated transmission in striatal cholinergic interneurons

PubMed Central

Oswald, Manfred J.; Schulz, Jan M.; Kelsch, Wolfgang; Oorschot, Dorothy E.; Reynolds, John N. J.

2015-01-01

Pauses in the tonic firing of striatal cholinergic interneurons (CINs) emerge during reward-related learning in response to conditioning of a neutral cue. We have previously reported that augmenting the postsynaptic response to cortical afferents in CINs is coupled to the emergence of a cell-intrinsic afterhyperpolarization (AHP) underlying pauses in tonic activity. Here we investigated in a bihemispheric rat-brain slice preparation the mechanisms of synaptic plasticity of excitatory afferents to CINs and the association with changes in the AHP. We found that high frequency stimulation (HFS) of commissural corticostriatal afferents from the contralateral hemisphere induced a robust long-term depression (LTD) of postsynaptic potentials (PSP) in CINs. Depression of the PSP of smaller magnitude and duration was observed in response to HFS of the ipsilateral white matter or cerebral cortex. In Mg2+-free solution HFS induced NMDA receptor-dependent potentiation of the PSP, evident in both the maximal slope and amplitude of the PSP. The increase in maximal slope corroborates previous findings, and was blocked by antagonism of either D1-like dopamine receptors with SCH23390 or D2-like dopamine receptors with sulpiride during HFS in Mg2+-free solution. Potentiation of the slower PSP amplitude component was due to augmentation of the NMDA receptor-mediated potential as this was completely reversed on subsequent application of the NMDA receptor antagonist AP5. HFS similarly potentiated NMDA receptor currents isolated by blockade of AMPA/kainate receptors with CNQX. The plasticity-induced increase in the slow PSP component was directly associated with an increase in the subsequent AHP. Thus plasticity of cortical afferent synapses is ideally suited to influence the cue-induced firing dynamics of CINs, particularly through potentiation of NMDA receptor-mediated synaptic transmission. PMID:25914618

Potentiation of NMDA receptor-mediated transmission in striatal cholinergic interneurons.

PubMed

Oswald, Manfred J; Schulz, Jan M; Kelsch, Wolfgang; Oorschot, Dorothy E; Reynolds, John N J

2015-01-01

Pauses in the tonic firing of striatal cholinergic interneurons (CINs) emerge during reward-related learning in response to conditioning of a neutral cue. We have previously reported that augmenting the postsynaptic response to cortical afferents in CINs is coupled to the emergence of a cell-intrinsic afterhyperpolarization (AHP) underlying pauses in tonic activity. Here we investigated in a bihemispheric rat-brain slice preparation the mechanisms of synaptic plasticity of excitatory afferents to CINs and the association with changes in the AHP. We found that high frequency stimulation (HFS) of commissural corticostriatal afferents from the contralateral hemisphere induced a robust long-term depression (LTD) of postsynaptic potentials (PSP) in CINs. Depression of the PSP of smaller magnitude and duration was observed in response to HFS of the ipsilateral white matter or cerebral cortex. In Mg(2+)-free solution HFS induced NMDA receptor-dependent potentiation of the PSP, evident in both the maximal slope and amplitude of the PSP. The increase in maximal slope corroborates previous findings, and was blocked by antagonism of either D1-like dopamine receptors with SCH23390 or D2-like dopamine receptors with sulpiride during HFS in Mg(2+)-free solution. Potentiation of the slower PSP amplitude component was due to augmentation of the NMDA receptor-mediated potential as this was completely reversed on subsequent application of the NMDA receptor antagonist AP5. HFS similarly potentiated NMDA receptor currents isolated by blockade of AMPA/kainate receptors with CNQX. The plasticity-induced increase in the slow PSP component was directly associated with an increase in the subsequent AHP. Thus plasticity of cortical afferent synapses is ideally suited to influence the cue-induced firing dynamics of CINs, particularly through potentiation of NMDA receptor-mediated synaptic transmission.

Structural determinants of the transient receptor potential 1 (TRPV1) channel activation by phospholipid analogs.

PubMed

Morales-Lázaro, Sara L; Serrano-Flores, Barbara; Llorente, Itzel; Hernández-García, Enrique; González-Ramírez, Ricardo; Banerjee, Souvik; Miller, Duane; Gududuru, Veeresh; Fells, James; Norman, Derek; Tigyi, Gabor; Escalante-Alcalde, Diana; Rosenbaum, Tamara

2014-08-29

The transient receptor potential vanilloid 1 (TRPV1) ion channel is a polymodal protein that responds to various stimuli, including capsaicin (the pungent compound found in chili peppers), extracellular acid, and basic intracellular pH, temperatures close to 42 °C, and several lipids. Lysophosphatidic acid (LPA), an endogenous lipid widely associated with neuropathic pain, is an agonist of the TRPV1 channel found in primary afferent nociceptors and is activated by other noxious stimuli. Agonists or antagonists of lipid and other chemical natures are known to possess specific structural requirements for producing functional effects on their targets. To better understand how LPA and other lipid analogs might interact and affect the function of TRPV1, we set out to determine the structural features of these lipids that result in the activation of TRPV1. By changing the acyl chain length, saturation, and headgroup of these LPA analogs, we established strict requirements for activation of TRPV1. Among the natural LPA analogs, we found that only LPA 18:1, alkylglycerophosphate 18:1, and cyclic phosphatidic acid 18:1, all with a monounsaturated C18 hydrocarbon chain activate TRPV1, whereas polyunsaturated and saturated analogs do not. Thus, TRPV1 shows a more restricted ligand specificity compared with LPA G-protein-coupled receptors. We synthesized fatty alcohol phosphates and thiophosphates and found that many of them with a single double bond in position Δ9, 10, or 11 and Δ9 cyclopropyl group can activate TRPV1 with efficacy similar to capsaicin. Finally, we developed a pharmacophore and proposed a mechanistic model for how these lipids could induce a conformational change that activates TRPV1. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Activation of Stat1 by mutant fibroblast growth-factor receptor in thanatophoric dysplasia type II dwarfism.

PubMed

Su, W C; Kitagawa, M; Xue, N; Xie, B; Garofalo, S; Cho, J; Deng, C; Horton, W A; Fu, X Y

1997-03-20

The achondroplasia class of chondrodysplasias comprises the most common genetic forms of dwarfism in humans and includes achondroplasia, hypochondroplasia and thanatophoric dysplasia types I and II (TDI and TDII), which are caused by different mutations in a fibroblast growth-factor receptor FGFR3 (ref. 1). The molecular mechanism and the mediators of these FGFR3-related growth abnormalities are not known. Here we show that mutant TDII FGFR3 has a constitutive tyrosine kinase activity which can specifically activate the transcription factor Stat1 (for signal transducer and activator of transcription). Furthermore, expression of TDII FGFR3 induced nuclear translocation of Stat1, expression of the cell-cycle inhibitor p21(WAF1/CIP1), and growth arrest of the cell. Thus, TDII FGFR3 may use Stat1 as a mediator of growth retardation in bone development. Consistent with this, Stat1 activation and increased p21(WAF1/CIP1) expression was found in the cartilage cells from the TDII fetus, but not in those from the normal fetus. Thus, abnormal STAT activation and p21(WAF1/CIP1) expression by the TDII mutant receptor may be responsible for this FGFR3-related bone disease.

Can insulin-like growth factor 1 (IGF-1), IGF-1 receptor connective tissue growth factor and Ki-67 labelling index have a prognostic role in pulmonary carcinoids?

PubMed

Kanakis, Georgios A; Grimelius, Lars; Papaioannou, Dimitrios; Kaltsas, Gregory; Tsolakis, Apostolos V

2018-04-27

Altered expression of Insulin-like Growth Factor-1 (IGF-1), its receptor (IGF-1R), Connective Tissue Growth Factor (CTGF) and Hypoxia Inducible Factor-1 (HIF-1), has been implicated in tumorigenesis. So far, these factors have not been studied systematically in Pulmonary Carcinoids (PCs). To examine IGF-1, IGF-1R, CTGF and HIF-1 expression in PCs, and assess their prognostic value over established factors. Retrospective study of 121 PCs (104 Typical and 17 Atypical). The expression of growth factors was studied immunohistochemically and tumors were considered positive if immunoreactivity appeared in >50% of cells. All studied parameters were expressed in the majority of tumors (IGF-1, IGF-1R, CTGF and HIF-1, in 78.5%, 67%, 72% and 78%, respectively). Their expression tended to be more frequent in TCs and in tumors with Ki-67≤2% (significant only for HIF-1; 82 vs. 53%; p=0.023 and 83 vs. 63%; p=0.025 respectively). CTGF was the only factor correlated with more extensive disease (larger size; presence of lymph node and distant metastases). According to logistic regression analysis, only advanced age, Ki-67≥3.4% and lymph node involvement could predict the development of distant metastases. IGF-1, IGF-1R, CTGF and HIF-1 are avidly expressed in PCs; however, their presence did not appear to be of statistically significant value over established prognostic factors.

Identification of the Niemann-Pick C1-like 1 cholesterol absorption receptor as a new hepatitis C virus entry factor

PubMed Central

Sainz, Bruno; Barretto, Naina; Martin, Danyelle N.; Hiraga, Nobuhiko; Imamura, Michio; Hussain, Snawar; Marsh, Katherine A.; Yu, Xuemei; Chayama, Kazuaki; Alrefai, Waddah A.; Uprichard, Susan L.

2011-01-01

Hepatitis C virus (HCV) is a leading cause of liver disease worldwide. With ~170 million individuals infected and current interferon-based treatment having toxic side-effects and marginal efficacy, more effective antivirals are critically needed1. Although HCV protease inhibitors were just FDA approved, analogous to HIV therapy, optimal HCV therapy likely will require a combination of antivirals targeting multiple aspects of the viral lifecycle. Viral entry represents a promising multi-faceted target for antiviral intervention; however, to date FDA-approved inhibitors of HCV cell entry are unavailable. Here we show that the cellular Niemann-Pick C1-Like 1 (NPC1L1) cholesterol uptake receptor is an HCV entry factor amendable to therapeutic intervention. Specifically, NPC1L1 expression is necessary for HCV infection as silencing or antibody-mediated blocking of NPC1L1 impairs cell-cultured-derived HCV (HCVcc) infection initiation. In addition, the clinically-available FDA-approved NPC1L1 antagonist ezetimibe2,3 potently blocks HCV uptake in vitro via a virion cholesterol-dependent step prior to virion-cell membrane fusion. Importantly, ezetimibe inhibits infection of all major HCV genotypes in vitro, and in vivo delays the establishment of HCV genotype 1b infection in mice with human liver grafts. Thus, we have not only identified NPC1L1 as an HCV cell entry factor, but also discovered a new antiviral target and potential therapeutic agent. PMID:22231557

Cardioprotective role of G-Protein Coupled Estrogen Receptor 1 (GPER1).

PubMed

Koganti, Sivaramakrishna

2015-01-01

G-Protein Coupled Estrogen Receptor 1 (GPER1), also known as G-Protein Coupled Receptor 30 (GPR30) and initially considered an orphan receptor, has become one of the most important pharmacological targets in cardiovascular research. Since the gene encoding this putative receptor was cloned nearly 20 years ago, researchers have addressed its role in various aspects of physiology, including cardioprotection. Although extensive research has been carried out to understand the role of GPER1 as a pharmacological target to treat cardiovascular diseases, there are few current reviews addressing the overall cardioprotective benefits of this receptor and the signaling intermediates involved. This review considers the origins of GPER1, its cell biology, its physiological and pharmacological roles as a therapeutic target in cardiovascular disease, and what future research on GPER1 might entail. More specifically, the review focuses on GPER1 regulation of Angiotensin Type I Receptor (AT1R) and the role of estrogen receptors, epidermal growth factor receptor (EGFR) and matrix metalloproteinases (MMPs) in bringing about the cardioprotective effects of GPER1. Areas where improved knowledge of GPER1 biology is still needed to better understand the receptor's cardioprotective effects are also discussed.

I1 Imidazoline Receptor: Novel Potential Cytoprotective Target of TVP1022, the S-Enantiomer of Rasagiline

PubMed Central

Frolov, Luba; Ovcharenko, Elena; Angel, Itzchak; Youdim, Moussa B. H.; Binah, Ofer

2012-01-01

TVP1022, the S-enantiomer of rasagiline (Azilect®) (N-propargyl-1R-aminoindan), exerts cyto/cardio-protective effects in a variety of experimental cardiac and neuronal models. Previous studies have demonstrated that the protective activity of TVP1022 and other propargyl derivatives involve the activation of p42/44 mitogen-activated protein kinase (MAPK) signaling pathway. In the current study, we further investigated the molecular mechanism of action and signaling pathways of TVP1022 which may account for the cyto/cardio-protective efficacy of the drug. Using specific receptor binding and enzyme assays, we demonstrated that the imidazoline 1 and 2 binding sites (I1 & I2) are potential targets for TVP1022 (IC50 = 9.5E-08 M and IC50 = 1.4E-07 M, respectively). Western blotting analysis showed that TVP1022 (1–20 µM) dose-dependently increased the immunoreactivity of phosphorylated p42 and p44 MAPK in rat pheochromocytoma PC12 cells and in neonatal rat ventricular myocytes (NRVM). This effect of TVP1022 was significantly attenuated by efaroxan, a selective I1 imidazoline receptor antagonist. In addition, the cytoprotective effect of TVP1022 demonstrated in NRVM against serum deprivation-induced toxicity was markedly inhibited by efaroxan, thus suggesting the importance of I1imidazoline receptor in mediating the cardioprotective activity of the drug. Our findings suggest that the I1imidazoline receptor represents a novel site of action for the cyto/cardio-protective efficacy of TVP1022. PMID:23166584

I1 imidazoline receptor: novel potential cytoprotective target of TVP1022, the S-enantiomer of rasagiline.

PubMed

Barac, Yaron D; Bar-Am, Orit; Liani, Esti; Amit, Tamar; Frolov, Luba; Ovcharenko, Elena; Angel, Itzchak; Youdim, Moussa B H; Binah, Ofer

2012-01-01

TVP1022, the S-enantiomer of rasagiline (Azilect®) (N-propargyl-1R-aminoindan), exerts cyto/cardio-protective effects in a variety of experimental cardiac and neuronal models. Previous studies have demonstrated that the protective activity of TVP1022 and other propargyl derivatives involve the activation of p42/44 mitogen-activated protein kinase (MAPK) signaling pathway. In the current study, we further investigated the molecular mechanism of action and signaling pathways of TVP1022 which may account for the cyto/cardio-protective efficacy of the drug. Using specific receptor binding and enzyme assays, we demonstrated that the imidazoline 1 and 2 binding sites (I(1) & I(2)) are potential targets for TVP1022 (IC(50) =9.5E-08 M and IC(50) =1.4E-07 M, respectively). Western blotting analysis showed that TVP1022 (1-20 µM) dose-dependently increased the immunoreactivity of phosphorylated p42 and p44 MAPK in rat pheochromocytoma PC12 cells and in neonatal rat ventricular myocytes (NRVM). This effect of TVP1022 was significantly attenuated by efaroxan, a selective I(1) imidazoline receptor antagonist. In addition, the cytoprotective effect of TVP1022 demonstrated in NRVM against serum deprivation-induced toxicity was markedly inhibited by efaroxan, thus suggesting the importance of I(1)imidazoline receptor in mediating the cardioprotective activity of the drug. Our findings suggest that the I(1)imidazoline receptor represents a novel site of action for the cyto/cardio-protective efficacy of TVP1022.

Insulin-like growth factor 1: common mediator of multiple enterotrophic hormones and growth factors.

PubMed

Bortvedt, Sarah F; Lund, P Kay

2012-03-01

To summarize the recent evidence that insulin-like growth factor 1 (IGF1) mediates growth effects of multiple trophic factors and discuss clinical relevance. Recent reviews and original reports indicate benefits of growth hormone (GH) and long-acting glucagon-like peptide 2 (GLP2) analogs in short bowel syndrome and Crohn's disease. This review highlights the evidence that biomarkers of sustained small intestinal growth or mucosal healing and evaluation of intestinal epithelial stem cell biomarkers may improve clinical measures of intestinal growth or response to trophic hormones. Compelling evidence that IGF1 mediates growth effects of GH and GLP2 on intestine or linear growth in preclinical models of resection or Crohn's disease is presented, along with a concept that these hormones or IGF1 may enhance sustained growth if given early after bowel resection. Evidence that suppressor of cytokine signaling protein induction by GH or GLP2 in normal or inflamed intestine may limit IGF1-induced growth, but protect against risk of dysplasia or fibrosis, is reviewed. Whether IGF1 receptor mediates IGF1 action and potential roles of insulin receptors are addressed. IGF1 has a central role in mediating trophic hormone action in small intestine. Better understanding of benefits and risks of IGF1, receptors that mediate IGF1 action, and factors that limit undesirable growth are needed.

Potential role of follicle-stimulating hormone (FSH) and transforming growth factor (TGFβ1) in the regulation of ovarian angiogenesis.

PubMed

Kuo, Shih-Wei; Ke, Ferng-Chun; Chang, Geen-Dong; Lee, Ming-Ting; Hwang, Jiuan-Jiuan

2011-06-01

Angiogenesis occurs during ovarian follicle development and luteinization. Pituitary secreted FSH was reported to stimulate the expression of endothelial mitogen VEGF in granulosa cells. And, intraovarian cytokine transforming growth factor (TGF)β1 is known to facilitate FSH-induced differentiation of ovarian granulosa cells. This intrigues us to investigate the potential role of FSH and TGFβ1 regulation of granulosa cell function in relation to ovarian angiogenesis. Granulosa cells were isolated from gonadotropin-primed immature rats and treated once with FSH and/or TGFβ1 for 48 h, and the angiogenic potential of conditioned media (granulosa cell culture conditioned media; GCCM) was determined using an in vitro assay with aortic ring embedded in collagen gel and immunoblotting. FSH and TGFβ1 increased the secreted angiogenic activity in granulosa cells (FSH + TGFβ1 > FSH ≈ TGFβ1 >control) that was partly attributed to the increased secretion of pro-angiogenic factors VEGF and PDGF-B. This is further supported by the evidence that pre-treatment with inhibitor of VEGF receptor-2 (Ki8751) or PDGF receptor (AG1296) throughout or only during the first 2-day aortic ring culture period suppressed microvessel growth in GCCM-treated groups, and also inhibited the FSH + TGFβ1-GCCM-stimulated release of matrix remodeling-associated gelatinase activities. Interestingly, pre-treatment of AG1296 at late stage suppressed GCCM-induced microvessel growth and stability with demise of endothelial and mural cells. Together, we provide original findings that both FSH and TGFβ1 increased the secretion of VEGF and PDGF-B, and that in turn up-regulated the angiogenic activity in rat ovarian granulosa cells. This implicates that FSH and TGFβ1 play important roles in regulation of ovarian angiogenesis during follicle development. Copyright © 2010 Wiley-Liss, Inc.

Synergistic apoptosis in head and neck squamous cell carcinoma cells by co-inhibition of insulin-like growth factor-1 receptor signaling and compensatory signaling pathways.

PubMed

Axelrod, Mark J; Mendez, Rolando E; Khalil, Ashraf; Leimgruber, Stephanie S; Sharlow, Elizabeth R; Capaldo, Brian; Conaway, Mark; Gioeli, Daniel G; Weber, Michael J; Jameson, Mark J

2015-12-01

In head and neck squamous cell carcinoma (HNSCC), resistance to single-agent targeted therapy may be overcome by co-targeting of compensatory signaling pathways. A targeted drug screen with 120 combinations was used on 9 HNSCC cell lines. Multiple novel drug combinations demonstrated synergistic growth inhibition. Combining the insulin-like growth factor-1 receptor (IGF-1R) inhibitor, BMS754807, with either the human epidermal growth factor receptor (HER)-family inhibitor, BMS599626, or the Src-family kinase inhibitor, dasatinib, resulted in substantial synergy and growth inhibition. Depending on the cell line, these combinations induced synergistic or additive apoptosis; when synergistic apoptosis was observed, AKT phosphorylation was inhibited to a greater extent than either drug alone. Conversely, when additive apoptosis occurred, AKT phosphorylation was not reduced by the drug combination. Combined IGF-1R/HER family and IGF-1R/Src family inhibition may have therapeutic potential in HNSCC. AKT may be a node of convergence between IGF-1R signaling and pathways that compensate for IGF-1R inhibition. © 2015 Wiley Periodicals, Inc.

Fibroblast growth factor receptor signaling crosstalk in skeletogenesis.

PubMed

Miraoui, Hichem; Marie, Pierre J

2010-11-02

Fibroblast growth factors (FGFs) play important roles in the control of embryonic and postnatal skeletal development by activating signaling through FGF receptors (FGFRs). Germline gain-of-function mutations in FGFR constitutively activate FGFR signaling, causing chondrocyte and osteoblast dysfunctions that result in skeletal dysplasias. Crosstalk between the FGFR pathway and other signaling cascades controls skeletal precursor cell differentiation. Genetic analyses revealed that the interplay of WNT and FGFR1 determines the fate and differentiation of mesenchymal stem cells during mouse craniofacial skeletogenesis. Additionally, interactions between FGFR signaling and other receptor tyrosine kinase networks, such as those mediated by the epidermal growth factor receptor and platelet-derived growth factor receptor α, were associated with excessive osteoblast differentiation and bone formation in the human skeletal dysplasia called craniosynostosis, which is a disorder of skull development. We review the roles of FGFR signaling and its crosstalk with other pathways in controlling skeletal cell fate and discuss how this crosstalk could be pharmacologically targeted to correct the abnormal cell phenotype in skeletal dysplasias caused by aberrant FGFR signaling.

Huntingtin interacting protein 1 is a novel brain tumor marker that associates with epidermal growth factor receptor.

PubMed

Bradley, Sarah V; Holland, Eric C; Liu, Grace Y; Thomas, Dafydd; Hyun, Teresa S; Ross, Theodora S

2007-04-15

Huntingtin interacting protein 1 (HIP1) is a multidomain oncoprotein whose expression correlates with increased epidermal growth factor receptor (EGFR) levels in certain tumors. For example, HIP1-transformed fibroblasts and HIP1-positive breast cancers have elevated EGFR protein levels. The combined association of HIP1 with huntingtin, the protein that is mutated in Huntington's disease, and the known overexpression of EGFR in glial brain tumors prompted us to explore HIP1 expression in a group of patients with different types of brain cancer. We report here that HIP1 is overexpressed with high frequency in brain cancers and that this overexpression correlates with EGFR and platelet-derived growth factor beta receptor expression. Furthermore, serum samples from patients with brain cancer contained anti-HIP1 antibodies more frequently than age-matched brain cancer-free controls. Finally, we report that HIP1 physically associates with EGFR and that this association is independent of the lipid, clathrin, and actin interacting domains of HIP1. These findings suggest that HIP1 may up-regulate or maintain EGFR overexpression in primary brain tumors by directly interacting with the receptor. This novel HIP1-EGFR interaction may work with or independent of HIP1 modulation of EGFR degradation via clathrin-mediated membrane trafficking pathways. Further investigation of HIP1 function in brain cancer biology and validation of its use as a prognostic or predictive brain tumor marker are now warranted.

Serum placental growth factor, vascular endothelial growth factor, soluble vascular endothelial growth factor receptor-1 and -2 levels in periodontal disease, and adverse pregnancy outcomes.

PubMed

Sert, Tuba; Kırzıoğlu, F Yeşim; Fentoğlu, Ozlem; Aylak, Firdevs; Mungan, Tamer

2011-12-01

The aim of this study is the evaluation of levels of serum interleukin (IL)-1β, IL-6, IL-10, tumor necrosis factor (TNF)-α, vascular endothelial growth factor (VEGF), placental growth factor (PIGF), and soluble VEGF receptor (sVEGFR)-1 and -2 in the association between periodontal disease and adverse pregnancy outcomes. One hundred and nine mothers, who recently gave birth, and 51 women who were not recently pregnant, aged 18 to 35 years, were included in this study. The mothers were classified as term birth, preterm birth (PTB), and preterm low birth weight (PLBW) in respect to their gestational age and baby's birth weight. The birth mothers were grouped as having gingivitis or periodontitis. The non-pregnant group also included periodontally healthy patients. Venous blood samples were collected to evaluate serum IL-1β, IL-6, IL-10, TNF-α, VEGF, PIGF, and sVEGFR-1 and -2 levels. Mother's weight, education, and income level were significantly associated with pregnancy outcomes. Serum levels of IL-1β, TNF-α, IL-6, VEGF, and sVEGFR-1 and -2 showed an increase in significance when related to pregnancy. Whereas in the PLBW group IL-1β, VEGF, and sVEGFR-2 levels were increased, in the PTB group sVEGFR-1 levels were increased. Additionally, the patients in the PLBW group with periodontitis had higher serum levels of IL-1β, VEGF, sVEGFR-2, and IL-1β/IL-10. The serum levels of IL-1β, VEGF, and sVEGFR-1 and -2 may have a potential effect on the mechanism of the association between periodontal disease and adverse pregnancy outcomes.

Peroxisome Proliferator Activated Receptor-α/Hypoxia Inducible Factor-1α Interplay Sustains Carbonic Anhydrase IX and Apoliprotein E Expression in Breast Cancer Stem Cells

PubMed Central

Papi, Alessio; Storci, Gianluca; Guarnieri, Tiziana; De Carolis, Sabrina; Bertoni, Sara; Avenia, Nicola; Sanguinetti, Alessandro; Sidoni, Angelo; Santini, Donatella; Ceccarelli, Claudio; Taffurelli, Mario; Orlandi, Marina; Bonafé, Massimiliano

2013-01-01

Aims Cancer stem cell biology is tightly connected to the regulation of the pro-inflammatory cytokine network. The concept of cancer stem cells “inflammatory addiction” leads to envisage the potential role of anti-inflammatory molecules as new anti-cancer targets. Here we report on the relationship between nuclear receptors activity and the modulation of the pro-inflammatory phenotype in breast cancer stem cells. Methods Breast cancer stem cells were expanded as mammospheres from normal and tumor human breast tissues and from tumorigenic (MCF7) and non tumorigenic (MCF10) human breast cell lines. Mammospheres were exposed to the supernatant of breast tumor and normal mammary gland tissue fibroblasts. Results In mammospheres exposed to the breast tumor fibroblasts supernatant, autocrine tumor necrosis factor-α signalling engenders the functional interplay between peroxisome proliferator activated receptor-α and hypoxia inducible factor-1α (PPARα/HIF1α). The two proteins promote mammospheres formation and enhance each other expression via miRNA130b/miRNA17-5p-dependent mechanism which is antagonized by PPARγ. Further, the PPARα/HIF1α interplay regulates the expression of the pro-inflammatory cytokine interleukin-6, the hypoxia survival factor carbonic anhydrase IX and the plasma lipid carrier apolipoprotein E. Conclusion Our data demonstrate the importance of exploring the role of nuclear receptors (PPARα/PPARγ) in the regulation of pro-inflammatory pathways, with the aim to thwart breast cancer stem cells functioning. PMID:23372804

Inhibition of insulin-like growth factor receptor-1 reduces necroptosis-related markers and attenuates LPS-induced lung injury in mice.

PubMed

Lee, Su Hwan; Shin, Ju Hye; Song, Joo Han; Leem, Ah Young; Park, Moo Suk; Kim, Young Sam; Chang, Joon; Chung, Kyung Soo

2018-04-15

Insulin-like growth factor-1 (IGF-1) levels are known to increase in the bronchoalveolar lavage fluid (BALF) of patients with acute respiratory distress syndrome. Herein, we investigated the role of IGF-1 in lipopolysaccharide (LPS)-induced lung injury. In LPS-treated cells, expressions of receptor-interacting protein 3 (RIP3) and phosphorylated mixed lineage kinase domain-like protein (MLKL) were decreased in IGF-1 receptor small interfering RNA (siRNA)-treated cells compared to control cells. The levels of pro-inflammatory cytokines including interleukin (IL)-1β, IL-6, IL-10, tumour necrosis factor-α, and macrophage inflammatory protein 2/C-X-C motif chemokine ligand 2 in the supernatant were significantly reduced in IGF-1 receptor siRNA-treated cells compared to control cells. In LPS-induced murine lung injury model, total cell counts, polymorphonuclear leukocytes counts, and pro-inflammatory cytokine levels in the BALF were significantly lower and histologically detected lung injury was less common in the group treated with IGF-1 receptor monoclonal antibody compared to the non-treated group. On western blotting, RIP3 and phosphorylated MLKL expressions were relatively decreased in the IGF-1 receptor monoclonal antibody group compared to the non-treated group. IGF-1 may be associated with RIP3-mediated necroptosis in vitro, while blocking of the IGF-1 pathway may reduce LPS-induced lung injuries in vivo. Copyright © 2018 Elsevier Inc. All rights reserved.

Epidermal growth factor receptor (EGFR) transactivation by estrogen via the G-protein-coupled receptor, GPR30: a novel signaling pathway with potential significance for breast cancer.

PubMed

Filardo, Edward J

2002-02-01

The biological and biochemical effects of estrogen have been ascribed to its known receptors, which function as ligand-inducible transcription factors. However, estrogen also triggers rapid activation of classical second messengers (cAMP, calcium, and inositol triphosphate) and stimulation of intracellular signaling cascades mitogen-activated protein kinase (MAP K), PI3K and eNOS. These latter events are commonly activated by membrane receptors that either possess intrinsic tyrosine kinase activity or couple to heterotrimeric G-proteins. We have shown that estrogen transactivates the epidermal growth factor receptor (EGFR) to MAP K signaling axis via the G-protein-coupled receptor (GPCR), GPR30, through the release of surface-bound proHB-EGF from estrogen receptor (ER)-negative human breast cancer cells [Molecular Endocrinology 14 (2000) 1649]. This finding is consistent with a growing body of evidence suggesting that transactivation of EGFRs by GPCRs is a recurrent theme in cell signaling. GPCR-mediated transactivation of EGFRs by estrogen provides a previously unappreciated mechanism of cross-talk between estrogen and serum growth factors, and explains prior data reporting the EGF-like effects of estrogen. This novel mechanism by which estrogen activates growth factor-dependent signaling and its implications for breast cancer biology are discussed further in this review.

Aldosterone interaction with epidermal growth factor receptor signaling in MDCK cells.

PubMed

Gekle, Michael; Freudinger, Ruth; Mildenberger, Sigrid; Silbernagl, Stefan

2002-04-01

Epidermal growth factor (EGF) regulates cell proliferation, differentiation, and ion transport by using extracellular signal-regulated kinase (ERK)1/2 as a downstream signal. Furthermore, the EGF-receptor (EGF-R) is involved in signaling by G protein-coupled receptors, growth hormone, and cytokines by means of transactivation. It has been suggested that steroids interact with peptide hormones, in part, by rapid, potentially nongenomic, mechanisms. Previously, we have shown that aldosterone modulates Na(+)/H(+) exchange in Madin-Darby canine kidney (MDCK) cells by means of ERK1/2 in a way similar to growth factors. Here, we tested the hypothesis that aldosterone uses the EGF-R as a heterologous signal transducer in MDCK cells. Nanomolar concentrations of aldosterone induce a rapid increase in ERK1/2 phosphorylation, cellular Ca(2+) concentration, and Na(+)/H(+) exchange activity similar to increases induced by EGF. Furthermore, aldosterone induced a rapid increase in EGF-R-Tyr phosphorylation, and inhibition of EGF-R kinase abolished aldosterone-induced signaling. Inhibition of ERK1/2 phosphorylation reduced the Ca(2+) response, whereas prevention of Ca(2+) influx did not abolish ERK1/2 phosphorylation. Our data show that aldosterone uses the EGF-R-ERK1/2 signaling cascade to elicit its rapid effects in MDCK cells.

Molecular basis for zinc potentiation at strychnine-sensitive glycine receptors.

PubMed

Miller, Paul S; Da Silva, Helena M A; Smart, Trevor G

2005-11-11

The divalent cation Zn(2+) is a potent potentiator at the strychnine-sensitive glycine receptor (GlyR). This occurs at nanomolar concentrations, which are the predicted endogenous levels of extracellular neuronal Zn(2+). Using structural modeling and functional mutagenesis, we have identified the molecular basis for the elusive Zn(2+) potentiation site on GlyRs and account for the differential sensitivity of GlyR alpha(1) and GlyR alpha(2) to Zn(2+) potentiation. In addition, juxtaposed to this Zn(2+) site, which is located externally on the N-terminal domain of the alpha subunit, another residue was identified in the nearby Cys loop, a region that is critical for receptor gating in all Cys loop ligand-gated ion channels. This residue acted as a key control element in the allosteric transduction pathway for Zn(2+) potentiation, enabling either potentiation or overt inhibition of receptor activation depending upon the moiety resident at this location. Overall, we propose that Zn(2+) binds to a site on the extracellular outer face of the GlyR alpha subunit and exerts its positive allosteric effect via an interaction with the Cys loop to increase the efficacy of glycine receptor gating.

An elevation in physical coupling of type 1 inositol 1,4,5-trisphosphate (IP3) receptors to transient receptor potential 3 (TRPC3) channels constricts mesenteric arteries in genetic hypertension.

PubMed

Adebiyi, Adebowale; Thomas-Gatewood, Candice M; Leo, M Dennis; Kidd, Michael W; Neeb, Zachary P; Jaggar, Jonathan H

2012-11-01

Hypertension is associated with an elevation in agonist-induced vasoconstriction, but mechanisms involved require further investigation. Many vasoconstrictors bind to phospholipase C-coupled receptors, leading to an elevation in inositol 1,4,5-trisphosphate (IP(3)) that activates sarcoplasmic reticulum IP(3) receptors. In cerebral artery myocytes, IP(3) receptors release sarcoplasmic reticulum Ca(2+) and can physically couple to canonical transient receptor potential 3 (TRPC3) channels in a caveolin-1-containing macromolecular complex, leading to cation current activation that stimulates vasoconstriction. Here, we investigated mechanisms by which IP(3) receptors control vascular contractility in systemic arteries and IP(3)R involvement in elevated agonist-induced vasoconstriction during hypertension. Total and plasma membrane-localized TRPC3 protein was ≈2.7- and 2-fold higher in mesenteric arteries of spontaneously hypertensive rats (SHRs) than in Wistar-Kyoto (WKY) rat controls, respectively. In contrast, IP(3)R1, TRPC1, TRPC6, and caveolin-1 expression was similar. TRPC3 expression was also similar in arteries of pre-SHRs and WKY rats. Control, IP(3)-induced and endothelin-1 (ET-1)-induced fluorescence resonance energy transfer between IP3R1 and TRPC3 was higher in SHR than WKY myocytes. IP3-induced cation current was ≈3-fold larger in SHR myocytes. Pyr3, a selective TRPC3 channel blocker, and calmodulin and IP(3) receptor binding domain peptide, an IP(3)R-TRP physical coupling inhibitor, reduced IP(3)-induced cation current and ET-1-induced vasoconstriction more in SHR than WKY myocytes and arteries. Thapsigargin, a sarcoplasmic reticulum Ca(2+)-ATPase blocker, did not alter ET-1-stimulated vasoconstriction in SHR or WKY arteries. These data indicate that ET-1 stimulates physical coupling of IP(3)R1 to TRPC3 channels in mesenteric artery myocytes, leading to vasoconstriction. Furthermore, an elevation in IP(3)R1 to TRPC3 channel molecular coupling augments

Ferulic Acid Exerts Anti-Angiogenic and Anti-Tumor Activity by Targeting Fibroblast Growth Factor Receptor 1-Mediated Angiogenesis.

PubMed

Yang, Guang-Wei; Jiang, Jin-Song; Lu, Wei-Qin

2015-10-12

Most anti-angiogenic therapies currently being evaluated target the vascular endothelial growth factor (VEGF) pathway; however, the tumor vasculature can acquire resistance to VEGF-targeted therapy by shifting to other angiogenesis mechanisms. Therefore, other therapeutic agents that block non-VEGF angiogenic pathways need to be evaluated. Here, we identified ferulic acid as a novel fibroblast growth factor receptor 1 (FGFR1) inhibitor and a novel agent with potential anti-angiogenic and anti-cancer activities. Ferulic acid demonstrated inhibition of endothelial cell proliferation, migration and tube formation in response to basic fibroblast growth factor 1 (FGF1). In ex vivo and in vivo angiogenesis assays, ferulic acid suppressed FGF1-induced microvessel sprouting of rat aortic rings and angiogenesis. To understand the underlying molecular basis, we examined the effects of ferulic acid on different molecular components and found that ferulic acid suppressed FGF1-triggered activation of FGFR1 and phosphatidyl inositol 3-kinase (PI3K)-protein kinase B (Akt) signaling. Moreover, ferulic acid directly inhibited proliferation and blocked the PI3K-Akt pathway in melanoma cell. In vivo, using a melanoma xenograft model, ferulic acid showed growth-inhibitory activity associated with inhibition of angiogenesis. Taken together, our results indicate that ferulic acid targets the FGFR1-mediated PI3K-Akt signaling pathway, leading to the suppression of melanoma growth and angiogenesis.

Structural insights into transient receptor potential vanilloid type 1 (TRPV1) from homology modeling, flexible docking, and mutational studies.

PubMed

Lee, Jin Hee; Lee, Yoonji; Ryu, HyungChul; Kang, Dong Wook; Lee, Jeewoo; Lazar, Jozsef; Pearce, Larry V; Pavlyukovets, Vladimir A; Blumberg, Peter M; Choi, Sun

2011-04-01

The transient receptor potential vanilloid subtype 1 (TRPV1) is a non-selective cation channel composed of four monomers with six transmembrane helices (TM1-TM6). TRPV1 is found in the central and peripheral nervous system, and it is an important therapeutic target for pain relief. We describe here the construction of a tetrameric homology model of rat TRPV1 (rTRPV1). We experimentally evaluated by mutational analysis the contribution of residues of rTRPV1 contributing to ligand binding by the prototypical TRPV1 agonists, capsaicin and resiniferatoxin (RTX). We then performed docking analysis using our homology model. The docking results with capsaicin and RTX showed that our homology model was reliable, affording good agreement with our mutation data. Additionally, the binding mode of a simplified RTX (sRTX) ligand as predicted by the modeling agreed well with those of capsaicin and RTX, accounting for the high binding affinity of the sRTX ligand for TRPV1. Through the homology modeling, docking and mutational studies, we obtained important insights into the ligand-receptor interactions at the molecular level which should prove of value in the design of novel TRPV1 ligands.

Structural insights into transient receptor potential vanilloid type 1 (TRPV1) from homology modeling, flexible docking, and mutational studies

NASA Astrophysics Data System (ADS)

Lee, Jin Hee; Lee, Yoonji; Ryu, HyungChul; Kang, Dong Wook; Lee, Jeewoo; Lazar, Jozsef; Pearce, Larry V.; Pavlyukovets, Vladimir A.; Blumberg, Peter M.; Choi, Sun

2011-04-01

The transient receptor potential vanilloid subtype 1 (TRPV1) is a non-selective cation channel composed of four monomers with six transmembrane helices (TM1-TM6). TRPV1 is found in the central and peripheral nervous system, and it is an important therapeutic target for pain relief. We describe here the construction of a tetrameric homology model of rat TRPV1 (rTRPV1). We experimentally evaluated by mutational analysis the contribution of residues of rTRPV1 contributing to ligand binding by the prototypical TRPV1 agonists, capsaicin and resiniferatoxin (RTX). We then performed docking analysis using our homology model. The docking results with capsaicin and RTX showed that our homology model was reliable, affording good agreement with our mutation data. Additionally, the binding mode of a simplified RTX (sRTX) ligand as predicted by the modeling agreed well with those of capsaicin and RTX, accounting for the high binding affinity of the sRTX ligand for TRPV1. Through the homology modeling, docking and mutational studies, we obtained important insights into the ligand-receptor interactions at the molecular level which should prove of value in the design of novel TRPV1 ligands.

Expression and significance of Tie-1 and Tie-2 receptors, and angiopoietins-1, 2 and 4 in colorectal adenocarcinoma: Immunohistochemical analysis and correlation with clinicopathological factors

PubMed Central

Nakayama, Toshiyuki; Hatachi, Go; Wen, Chun-Yang; Yoshizaki, Ayumi; Yamazumi, Kazuyuki; Niino, Daisuke; Sekine, Ichiro

2005-01-01

AIM: There is strong evidence that tyrosine kinases are involved in the regulation of tumor progression, cellular growth and differentiation. Recently, many kinds of tyrosine kinase receptors have been reported, among them Tie-1 and Tie-2 receptors constitute a major class. Angiopoietin (Ang)-1 is known as a ligand of Tie-2 tyrosine kinase receptor. The objective of this study was to establish a comprehensive Tie-1 and Tie-2 and Ang-1, 2 and 4 expression profile in human colorectal adenocarcinomas. METHODS: We examined 96 cases of surgically resected human colorectal adenocarcinoma by immunohistochemistry and investigated the statistical correlation between the expressions of Ties and Angs and clinicopathological factors. RESULTS: Among the 96 cases of adenocarcinoma, 87 (90.6%), 92 (95.8%), 83 (86.5%), 89 (92.7%), and 76 cases (79.2%) showed positive staining in the cytoplasm of carcinoma cells for the Tie-1 and Tie-2 and Ang-1, 2 and 4 proteins, respectively. Histologically, the expressions of Ties and Angs were variable. The expressions of Ties and Angs were correlated with several clinicopathological factors, but did not correlate with the presence of lymph node metastasis. Ties and Angs were highly expressed in human colorectal adenocarcinoma cells. CONCLUSION: These findings suggest that the Tie-Ang receptor-ligand complex is one of the factors involved in the cellular differentiation and progression of human colorectal adenocarcinoma. PMID:15742397

The autoimmunity-associated gene PTPN22 potentiates toll-like receptor-driven, type 1 interferon-dependent immunity.

PubMed

Wang, Yaya; Shaked, Iftach; Stanford, Stephanie M; Zhou, Wenbo; Curtsinger, Julie M; Mikulski, Zbigniew; Shaheen, Zachary R; Cheng, Genhong; Sawatzke, Kristy; Campbell, Amanda M; Auger, Jennifer L; Bilgic, Hatice; Shoyama, Fernanda M; Schmeling, David O; Balfour, Henry H; Hasegawa, Kiminori; Chan, Andrew C; Corbett, John A; Binstadt, Bryce A; Mescher, Matthew F; Ley, Klaus; Bottini, Nunzio; Peterson, Erik J

2013-07-25

Immune cells sense microbial products through Toll-like receptors (TLR), which trigger host defense responses including type 1 interferons (IFNs) secretion. A coding polymorphism in the protein tyrosine phosphatase nonreceptor type 22 (PTPN22) gene is a susceptibility allele for human autoimmune and infectious disease. We report that Ptpn22 selectively regulated type 1 IFN production after TLR engagement in myeloid cells. Ptpn22 promoted host antiviral responses and was critical for TLR agonist-induced, type 1 IFN-dependent suppression of inflammation in colitis and arthritis. PTPN22 directly associated with TNF receptor-associated factor 3 (TRAF3) and promotes TRAF3 lysine 63-linked ubiquitination. The disease-associated PTPN22W variant failed to promote TRAF3 ubiquitination, type 1 IFN upregulation, and type 1 IFN-dependent suppression of arthritis. The findings establish a candidate innate immune mechanism of action for a human autoimmunity "risk" gene in the regulation of host defense and inflammation. Copyright © 2013 Elsevier Inc. All rights reserved.

Allosteric potentiation of quisqualate receptors by a nootropic drug aniracetam.

PubMed

Ito, I; Tanabe, S; Kohda, A; Sugiyama, H

1990-05-01

1. Allosteric potentiation of the ionotropic quisqualate (iQA) receptor by a nootropic drug aniracetam (1-p-anisoyl-2-pyrrolidinone) was investigated using Xenopus oocytes injected with rat brain mRNA and rat hippocampal slices. 2. Aniracetam potentiates the iQA responses induced in Xenopus oocytes by rat brain mRNA in a reversible manner. This effect was observed above the concentrations of 0.1 mM. Kainate. N-methyl-D-aspartate and gamma-aminobutyric acid responses induced in the same oocytes were not affected. 3. The specific potentiation of iQA responses was accompanied by an increase in the conductance change of iQA and alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) responses, but the affinity of receptors for agonist and the ion-selectivity of the channels (reversal potentials) were not changed. 4. Aniracetam reversibly potentiated the iQA responses recorded intracellularly from the pyramidal cells in the CA1 region of rat hippocampal slices. The excitatory postsynaptic potentials (EPSPs) in Schaffer collateral-commissural-CA1 synapses were also potentiated by aniracetam. 5. Population EPSPs recorded in the mossy fibre-CA3 synapses as well as Schaffer-commissural synapses were also potentiated by aniracetam. The amplitudes of the potentiation were not changed by the formation of long-term potentiation.

Transient Receptor Potential Ankyrin 1 Receptor Activation In Vitro and In Vivo by Pro-tussive Agents: GRC 17536 as a Promising Anti-Tussive Therapeutic

PubMed Central

Mukhopadhyay, Indranil; Kulkarni, Abhay; Aranake, Sarika; Karnik, Pallavi; Shetty, Mahesh; Thorat, Sandeep; Ghosh, Indraneel; Wale, Dinesh; Bhosale, Vikram; Khairatkar-Joshi, Neelima

2014-01-01

Cough is a protective reflex action that helps clear the respiratory tract which is continuously exposed to airborne environmental irritants. However, chronic cough presents itself as a disease in its own right and despite its global occurrence; the molecular mechanisms responsible for cough are not completely understood. Transient receptor potential ankyrin1 (TRPA1) is robustly expressed in the neuronal as well as non-neuronal cells of the respiratory tract and is a sensor of a wide range of environmental irritants. It is fast getting acceptance as a key biological sensor of a variety of pro-tussive agents often implicated in miscellaneous chronic cough conditions. In the present study, we demonstrate in vitro direct functional activation of TRPA1 receptor by citric acid which is routinely used to evoke cough in preclinical and clinical studies. We also show for the first time that a potent and selective TRPA1 antagonist GRC 17536 inhibits citric acid induced cellular Ca+2 influx in TRPA1 expressing cells and the citric acid induced cough response in guinea pigs. Hence our data provides a mechanistic link between TRPA1 receptor activation in vitro and cough response induced in vivo by citric acid. Furthermore, we also show evidence for TRPA1 activation in vitro by the TLR4, TLR7 and TLR8 ligands which are implicated in bacterial/respiratory virus pathogenesis often resulting in chronic cough. In conclusion, this study highlights the potential utility of TRPA1 antagonist such as GRC 17536 in the treatment of miscellaneous chronic cough conditions arising due to diverse causes but commonly driven via TRPA1. PMID:24819048

Metabotropic glutamate receptors are required for the induction of long-term potentiation

NASA Technical Reports Server (NTRS)

Zheng, F.; Gallagher, J. P.

1992-01-01

Recent observations have led to the suggestion that the metabotropic glutamate receptor may play a role in the induction or maintenance of long-term potentiation (LTP). However, experimental evidence supporting a role for this receptor in the induction of LTP is still inconclusive and controversial. Here we report that, in rat dorsolateral septal nucleus (DLSN) neurons, which have the highest density of metabotropic receptors and show functional responses, the induction of LTP is not blocked by the NMDA receptor antagonist 2-amino-5-phosphonovalerate, but is blocked by two putative metabotropic glutamate receptor antagonists, L-2-amino-3-phosphonopropionic acid and L-2-amino-4-phosphonobutyrate. Furthermore, superfusion of (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid, a selective metabotropic glutamate agonist, resulted in a long-lasting potentiation of synaptic transmission similar to that induced by tetanic stimuli. Our results demonstrated that activation of postsynaptic metabotropic receptors is both necessary and sufficient for the induction of LTP in the DLSN, and we suggest that such a mechanism may be important at other CNS synapses.

Properties of the intracellular transient receptor potential (TRP) channel in yeast, Yvc1.

PubMed

Chang, Yiming; Schlenstedt, Gabriel; Flockerzi, Veit; Beck, Andreas

2010-05-17

Transient receptor potential (TRP) channels are found among mammals, flies, worms, ciliates, Chlamydomonas, and yeast but are absent in plants. These channels are believed to be tetramers of proteins containing six transmembrane domains (TMs). Their primary structures are diverse with sequence similarities only in some short amino acid sequence motifs mainly within sequences covering TM5, TM6, and adjacent domains. In the yeast genome, there is one gene encoding a TRP-like sequence. This protein forms an ion channel in the vacuolar membrane and is therefore called Yvc1 for yeast vacuolar conductance 1. In the following we summarize its prominent features. Copyright 2009 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

LRP1 protects the vasculature by regulating levels of connective tissue growth factor and HtrA1.

PubMed

Muratoglu, Selen C; Belgrave, Shani; Hampton, Brian; Migliorini, Mary; Coksaygan, Turhan; Chen, Ling; Mikhailenko, Irina; Strickland, Dudley K

2013-09-01

Low-density lipoprotein receptor-related protein 1 (LRP1) is a large endocytic and signaling receptor that is abundant in vascular smooth muscle cells. Mice in which the lrp1 gene is deleted in smooth muscle cells (smLRP1(-/-)) on a low-density lipoprotein receptor-deficient background display excessive platelet derived growth factor-signaling, smooth muscle cell proliferation, aneurysm formation, and increased susceptibility to atherosclerosis. The objectives of the current study were to examine the potential of LRP1 to modulate vascular physiology under nonatherogenic conditions. We found smLRP1(-/-) mice to have extensive in vivo aortic dilatation accompanied by disorganized and degraded elastic lamina along with medial thickening of the arterial vessels resulting from excess matrix deposition. Surprisingly, this was not attributable to excessive platelet derived growth factor-signaling. Rather, quantitative differential proteomic analysis revealed that smLRP1(-/-) vessels contain a 4-fold increase in protein levels of high-temperature requirement factor A1 (HtrA1), which is a secreted serine protease that is known to degrade matrix components and to impair elastogenesis, resulting in fragmentation of elastic fibers. Importantly, our study discovered that HtrA1 is a novel LRP1 ligand. Proteomics analysis also identified excessive accumulation of connective tissue growth factor, an LRP1 ligand and a key mediator of fibrosis. Our findings suggest a critical role for LRP1 in maintaining the integrity of vessels by regulating protease activity as well as matrix deposition by modulating HtrA1 and connective tissue growth factor protein levels. This study highlights 2 new molecules, connective tissue growth factor and HtrA1, which contribute to detrimental changes in the vasculature and, therefore, represent new target molecules for potential therapeutic intervention to maintain vessel wall homeostasis.

Group III mGlu receptor agonists potentiate the anticonvulsant effect of AMPA and NMDA receptor block.

PubMed

De Sarro, Giovambattista; Chimirri, Alba; Meldrum, Brian S

2002-09-06

We report the anticonvulsant action in DBA/2 mice of two mGlu Group III receptor agonists: (R,S)-4-phosphonophenylglycine, (R,S)-PPG, a compound with moderate mGlu8 selectivity, and of (1S,3R,4S)-1-aminocyclopentane-1,2,4-tricarboxylic acid, ACPT-1, a selective agonist for mGlu4alpha receptors. Both compounds, given intracerebroventricularly at doses which did not show marked anticonvulsant activity, produced a consistent shift to the left of the dose-response curves (i.e. enhanced the anticonvulsant properties) of 1-(4'-aminophenyl)-3,5-dihydro-7,8-dimethoxy-4H-2,3-benzodiazepin-4-one hydrochloride, CFM-2, a noncompetitive AMPA receptor antagonist, and 3-((+/-)-2-carboxypiperazin-4-yl)-1-phosphonic acid, CPPene, a competitive NMDA receptor antagonist, in DBA/2 mice. In addition, (R,S)-PPG and ACPT-1 administered intracerebroventricularly prolonged the time course of the anticonvulsant properties of CFM-2 (33 micromol/kg, i.p.) and CPPene (3.3 micromol/kg, i.p.) administered intraperitoneally. We conclude that modest reduction of synaptic glutamate release by activation of Group III metabotropic receptors potentiates the anticonvulsant effect of AMPA and NMDA receptor blockade. Copyright 2002 Elsevier Science B.V.

Evaluation of 11C-Me-NB1 as a Potential PET Radioligand for Measuring GluN2B-Containing NMDA Receptors, Drug Occupancy, and Receptor Cross Talk.

PubMed

Krämer, Stefanie D; Betzel, Thomas; Mu, Linjing; Haider, Ahmed; Herde, Adrienne Müller; Boninsegni, Anna K; Keller, Claudia; Szermerski, Marina; Schibli, Roger; Wünsch, Bernhard; Ametamey, Simon M

2018-04-01

Clinical and preclinical research with modulators at the N -methyl-d-aspartate (NMDA) receptor GluN2B N-terminal domain (NTD) aims for the treatment of various neurologic diseases. The interpretation of the results is hampered by the lack of a suitable NMDA PET tracer for assessing the receptor occupancy of potential drugs. We have developed 11 C-Me-NB1 as a PET tracer for imaging GluN1/GluN2B-containing NMDA receptors and used it to investigate in rats the dose-dependent receptor occupancy of eliprodil, a GluN2B NTD modulator. Methods: 11 C-Me-NB1 was synthesized and characterized by in vitro displacement binding experiments with rat brain membranes, in vitro autoradiography, and blocking and displacement experiments by PET and PET kinetic modeling. Receptor occupancy by eliprodil was studied by PET with 11 C-Me-NB1. Results: 11 C-Me-NB1 was synthesized at 290 ± 90 GBq/μmol molar activity, 7.4 ± 1.9 GBq total activity at the end of synthesis ( n = 17), and more than 99% radiochemical purity. 11 C-Me-NB1 binding in rat brain was blocked in vitro and in vivo by the NTD modulators Ro-25-6981 and eliprodil. Half-maximal receptor occupancy by eliprodil occurred at 1.5 μg/kg. At 1 mg/kg of eliprodil, a dose with reported neuroprotective effects, more than 99.5% of binding sites were occupied. In vitro, 11 C-Me-NB1 binding was independent of the σ-1 receptor (Sigma1R), and the Sigma1R agonist (+)-pentazocine did not compete for high-affinity binding. In vivo, a 2.5 mg/kg dose of (+)-pentazocine abolished 11 C-Me-NB1-specific binding, indicating an indirect effect of Sigma1R on 11 C-Me-NB1 binding. Conclusion: 11 C-Me-NB1 is suitable for the in vivo imaging of NMDA GluN1/GluN2B receptors and the assessment of receptor occupancy by NTD modulators. GluN1/GluN2B NMDA receptors are fully occupied at neuroprotective doses of eliprodil. Furthermore, 11 C-Me-NB1 enables imaging of GluN1/GluN2B NMDA receptor cross talk. © 2018 by the Society of Nuclear Medicine and

Type 1 receptor tyrosine kinases are differentially phosphorylated in mammary carcinoma and differentially associated with steroid receptors.

PubMed Central

Bacus, S. S.; Chin, D.; Yarden, Y.; Zelnick, C. R.; Stern, D. F.

1996-01-01

The neu/erbB-2/HER-2 proto-oncogene is amplified and/or overexpressed in up to 30% of mammary carcinomas and has been variably correlated with poor prognosis. The signaling activity of the encoded receptor tyrosine kinase is regulated by interactions with other type 1 receptors and their ligands. We have used a novel approach, phosphorylation-sensitive anti-Neu antibodies, to quantify signaling by Neu and epidermal growth factor receptor in a panel of frozen sections of mammary carcinoma specimens. We also determined the relationship of Neu, phosphorylated Neu (and epidermal growth factor receptor), and phosphotyrosine to the expression of Neu-related receptors (epidermal growth factor receptor, HER-3, and HER-4) and to prognostic factors (estrogen and progesterone receptor). We found that tyrosine phosphorylation of Neu (and hence signaling activity) is highly variable among mammary carcinomas. Neu and HER-4 were associated with divergent correlates, suggesting that they have profoundly different biological activities. These results have implications for etiology of mammary carcinoma for clinical evaluation of mammary carcinoma patients, and for development of Neu-targeted therapeutic strategies. Images Figure 1 Figure 2 PMID:8579117

Behavioural profiles in the mouse defence test battery suggest anxiolytic potential of 5-HT(1A) receptor antagonists.

PubMed

Griebel, G; Rodgers, R J; Perrault, G; Sanger, D J

1999-05-01

Compounds varying in selectivity as 5-HT1A receptor antagonists have recently been reported to produce anxiolytic-like effects comparable to those of benzodiazepines in the mouse elevated plus-maze procedure. In view of the potential clinical significance of these findings, the present experiments compared the behavioural effects of diazepam (0.5-3.0 mg/kg) with those of several non-selective 5-HT1A receptor antagonists [NAN-190, 0.1-3.0 mg/kg, MM-77, 0.03-1.0 mg/kg, (S)-UH-301, 0.3-3.0 mg/kg and pindobind-5-HT1A, 0.03-1.0 mg/kg], and three selective 5-HT1A receptor antagonists (WAY100635, 0.01-3.0 mg/kg, p-MPPI, 0.1-3.0 mg/kg and SL88.0338, 0.3-3.0 mg/kg) in the mouse defence test battery (MDTB). In this well-validated anxiolytic screening test, Swiss mice are directly confronted with a natural threat (a rat) as well as situations associated with this threat. Primary measures taken during and after rat confrontation were flight, risk assessment (RA), defensive threat/attack and escape attempts. Diazepam significantly decreased flight reactions after the rat was introduced into the runway, reduced RA activities of mice chased by the rat, increased RA responses displayed when subjects were constrained in a straight alley and reduced defensive upright postures and biting upon forced contact. All the selective 5-HT1A receptor antagonists and NAN-190 also reduced flight, RA in the chase test, and defensive threat and attack behaviours. (S)-UH-301 and pindobind-5-HT1A reduced RA in the chase test, but only partially modified defensive threat and attack. Unlike the other drugs tested, MM-77 produced significant effects only at doses which also markedly reduced spontaneous locomotor activity, suggesting a behaviourally non-specific action. In contrast to diazepam, the 5-HT1A receptor ligands failed to affect RA in the straight alley test. Following removal of the rat from the test area, only diazepam and (S)-UH-301 reduced escape behaviour (contextual defence) at doses

Ghrelin-induced stimulation of colonic propulsion is dependent on hypothalamic neuropeptide Y1- and corticotrophin-releasing factor 1 receptor activation.

PubMed

Tebbe, J J; Mronga, S; Tebbe, C G; Ortmann, E; Arnold, R; Schäfer, M K-H

2005-09-01

Peptides participating in the hypothalamic control of feeding behaviour are also involved in the central autonomic control of gastrointestinal functions, such as secretion and motility. An anatomical interaction and functional relationship in the central nervous system between the feeding-related peptides neuropeptide Y and ghrelin is well documented. Furthermore, it has been shown that feeding-related peptides can influence digestive function via central corticotrophin-releasing factor (CRF) pathways. In the present study, we investigated the role of ghrelin in the central autonomic control of colonic motility. Furthermore, we addressed the hypothesis that ghrelin is involved in the hypothalamic control of colonic motor function, utilizing central neuropeptide Y receptors and hypothalamic CRF pathways. Ghrelin (0.03, 0.06 and 0.12 nmol) bilaterally microinjected into the paraventricular nucleus (PVN) induced a significant stimulation of colonic propulsion. In particular, the colonic transit time decreased from 312+/-7 min to 198+/-12 min. Microinjection of the neuropeptide Y1 receptor antagonist, BIBP-3226 (200 pmol), or the nonselective CRF receptor antagonist, astressin (30 pmol), into the PVN abolished the stimulatory effect of ghrelin injected into the PVN on colonic transit time, whereas pretreatment with the selective CRF2 receptor, antisauvagine-30 (28 pmol), failed to affect the effect of PVN-ghrelin injection on colonic propulsion. These results suggest that ghrelin can act as central modulator of gastrointestinal motor functions at the level of the PVN via neuropeptide Y1- and CRF1 receptor-dependent mechanisms.

The Fn14 cytoplasmic tail binds tumour-necrosis-factor-receptor-associated factors 1, 2, 3 and 5 and mediates nuclear factor-kappaB activation.

PubMed Central

Brown, Sharron A N; Richards, Christine M; Hanscom, Heather N; Feng, Sheau-Line Y; Winkles, Jeffrey A

2003-01-01

Fn14 is a growth-factor-inducible immediate-early-response gene encoding a 102-amino-acid type I transmembrane protein. The human Fn14 protein was recently identified as a cell-surface receptor for the tumour necrosis factor (TNF) superfamily member named TWEAK (TNF-like weak inducer of apoptosis). In the present paper, we report that the human TWEAK extracellular domain can also bind the murine Fn14 protein. Furthermore, site-specific mutagenesis and directed yeast two-hybrid interaction assays revealed that the TNFR-associated factor (TRAF) 1, 2, 3 and 5 adaptor molecules bind the murine Fn14 cytoplasmic tail at an overlapping, but non-identical, amino acid sequence motif. We also found that TWEAK treatment of quiescent NIH 3T3 cells stimulates inhibitory kappaBalpha phosphorylation and transcriptional activation of a nuclear factor-kappaB (NF-kappaB) enhancer/luciferase reporter construct. Fn14 overexpression in transiently transfected NIH 3T3 cells also promotes NF-kappaB activation, and this cellular response requires an intact TRAF binding site. These results indicate that Fn14 is a functional TWEAK receptor that can associate with four distinct TRAF family members and stimulate the NF-kappaB transcription factor signalling pathway. PMID:12529173

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.

PubMed

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.

AT1 receptor signaling pathways in the cardiovascular system.

PubMed

Kawai, Tatsuo; Forrester, Steven J; O'Brien, Shannon; Baggett, Ariele; Rizzo, Victor; Eguchi, Satoru

2017-11-01

The importance of the renin angiotensin aldosterone system in cardiovascular physiology and pathophysiology has been well described whereas the detailed molecular mechanisms remain elusive. The angiotensin II type 1 receptor (AT1 receptor) is one of the key players in the renin angiotensin aldosterone system. The AT1 receptor promotes various intracellular signaling pathways resulting in hypertension, endothelial dysfunction, vascular remodeling and end organ damage. Accumulating evidence shows the complex picture of AT1 receptor-mediated signaling; AT1 receptor-mediated heterotrimeric G protein-dependent signaling, transactivation of growth factor receptors, NADPH oxidase and ROS signaling, G protein-independent signaling, including the β-arrestin signals and interaction with several AT1 receptor interacting proteins. In addition, there is functional cross-talk between the AT1 receptor signaling pathway and other signaling pathways. In this review, we will summarize an up to date overview of essential AT1 receptor signaling events and their functional significances in the cardiovascular system. Copyright © 2017 Elsevier Ltd. All rights reserved.

Estrogen receptor 1 (ESR1) regulates VEGFA in adipose tissue.

PubMed

Fatima, L A; Campello, R S; Santos, R de Souza; Freitas, H S; Frank, A P; Machado, U F; Clegg, D J

2017-12-01

Vascular endothelial growth factor A (VEGFA) is a key factor in the regulation of angiogenesis in adipose tissue. Poor vascularization during adipose tissue proliferation causes fibrosis and local inflammation, and is associated with insulin resistance. It is known that 17-beta estradiol (E2) regulates adipose tissue function and VEGFA expression in other tissues; however, the ability of E2 to regulate VEGFA in adipose tissue is currently unknown. In this study, we showed that, in 3T3-L1 cells, E2 and the estrogen receptor 1 (ESR1) agonist PPT induced VEGFA expression, while ESR1 antagonist (MPP), and selective knockdown of ESR1 using siRNA decreased VEGFA and prevented the ability of E2 to modulate its expression. Additionally, we found that E2 and PPT induced the binding of hypoxia inducible factor 1 alpha subunit (HIF1A) in the VEGFA gene promoter. We further found that VEGFA expression was lower in inguinal and gonadal white adipose tissues of ESR1 total body knockout female mice compared to wild type mice. In conclusion, our data provide evidence of an important role for E2/ESR1 in modulating adipose tissue VEGFA, which is potentially important to enhance angiogenesis, reduce inflammation and improve adipose tissue function.

Noradrenaline Modulates the Membrane Potential and Holding Current of Medial Prefrontal Cortex Pyramidal Neurons via β1-Adrenergic Receptors and HCN Channels.

PubMed

Grzelka, Katarzyna; Kurowski, Przemysław; Gawlak, Maciej; Szulczyk, Paweł

2017-01-01

The medial prefrontal cortex (mPFC) receives dense noradrenergic projections from the locus coeruleus. Adrenergic innervation of mPFC pyramidal neurons plays an essential role in both physiology (control of memory formation, attention, working memory, and cognitive behavior) and pathophysiology (attention deficit hyperactivity disorder, posttraumatic stress disorder, cognitive deterioration after traumatic brain injury, behavioral changes related to addiction, Alzheimer's disease and depression). The aim of this study was to elucidate the mechanism responsible for adrenergic receptor-mediated control of the resting membrane potential in layer V mPFC pyramidal neurons. The membrane potential or holding current of synaptically isolated layer V mPFC pyramidal neurons was recorded in perforated-patch and classical whole-cell configurations in slices from young rats. Application of noradrenaline (NA), a neurotransmitter with affinity for all types of adrenergic receptors, evoked depolarization or inward current in the tested neurons irrespective of whether the recordings were performed in the perforated-patch or classical whole-cell configuration. The effect of noradrenaline depended on β 1 - and not α 1 - or α 2 -adrenergic receptor stimulation. Activation of β 1 -adrenergic receptors led to an increase in inward Na + current through hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, which carry a mixed Na + /K + current. The protein kinase A- and C-, glycogen synthase kinase-3β- and tyrosine kinase-linked signaling pathways were not involved in the signal transduction between β 1 -adrenergic receptors and HCN channels. The transduction system operated in a membrane-delimited fashion and involved the βγ subunit of G-protein. Thus, noradrenaline controls the resting membrane potential and holding current in mPFC pyramidal neurons through β 1 -adrenergic receptors, which in turn activate HCN channels via a signaling pathway involving the �

Noradrenaline Modulates the Membrane Potential and Holding Current of Medial Prefrontal Cortex Pyramidal Neurons via β1-Adrenergic Receptors and HCN Channels

PubMed Central

Grzelka, Katarzyna; Kurowski, Przemysław; Gawlak, Maciej; Szulczyk, Paweł

2017-01-01

The medial prefrontal cortex (mPFC) receives dense noradrenergic projections from the locus coeruleus. Adrenergic innervation of mPFC pyramidal neurons plays an essential role in both physiology (control of memory formation, attention, working memory, and cognitive behavior) and pathophysiology (attention deficit hyperactivity disorder, posttraumatic stress disorder, cognitive deterioration after traumatic brain injury, behavioral changes related to addiction, Alzheimer’s disease and depression). The aim of this study was to elucidate the mechanism responsible for adrenergic receptor-mediated control of the resting membrane potential in layer V mPFC pyramidal neurons. The membrane potential or holding current of synaptically isolated layer V mPFC pyramidal neurons was recorded in perforated-patch and classical whole-cell configurations in slices from young rats. Application of noradrenaline (NA), a neurotransmitter with affinity for all types of adrenergic receptors, evoked depolarization or inward current in the tested neurons irrespective of whether the recordings were performed in the perforated-patch or classical whole-cell configuration. The effect of noradrenaline depended on β1- and not α1- or α2-adrenergic receptor stimulation. Activation of β1-adrenergic receptors led to an increase in inward Na+ current through hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, which carry a mixed Na+/K+ current. The protein kinase A- and C-, glycogen synthase kinase-3β- and tyrosine kinase-linked signaling pathways were not involved in the signal transduction between β1-adrenergic receptors and HCN channels. The transduction system operated in a membrane-delimited fashion and involved the βγ subunit of G-protein. Thus, noradrenaline controls the resting membrane potential and holding current in mPFC pyramidal neurons through β1-adrenergic receptors, which in turn activate HCN channels via a signaling pathway involving the βγ subunit. PMID

Insulin-like growth factor type-1 receptor down-regulation associated with dwarfism in Holstein calves.

PubMed

Blum, J W; Elsasser, T H; Greger, D L; Wittenberg, S; de Vries, F; Distl, O

2007-10-01

Perturbations in endocrine functions can impact normal growth. Endocrine traits were studied in three dwarf calves exhibiting retarded but proportionate growth and four phenotypically normal half-siblings, sired by the same bull, and four unrelated control calves. Plasma 3,5,3'-triiodothyronine and thyroxine concentrations in dwarfs and half-siblings were in the physiological range and responded normally to injected thyroid-releasing hormone. Plasma glucagon concentrations were different (dwarfs, controls>half-siblings; P<0.05). Plasma growth hormone (GH), insulin-like growth factor-1 (IGF-1) and insulin concentrations in the three groups during an 8-h period were similar, but integrated GH concentrations (areas under concentration curves) were different (dwarfs>controls, P<0.02; half-siblings>controls, P=0.08). Responses of GH to xylazine and to a GH-releasing-factor analogue were similar in dwarfs and half-siblings. Relative gene expression of IGF-1, IGF-2, GH receptor (GHR), insulin receptor, IGF-1 type-1 and -2 receptors (IGF-1R, IGF-2R), and IGF binding proteins were measured in liver and anconeus muscle. GHR mRNA levels were different in liver (dwarfs1R mRNA abundance in liver in half-siblings and controls was 2.4- and 2.5-fold higher (P=0.003 and P=0.001, respectively) and in muscle tissue was 2.3- and 1.8-fold higher (P=0.01 and P=0.08, respectively) than in dwarfs. Hepatic IGF-1R protein levels (Western blots) in muscle were 2.5-fold higher (P<0.05) and in liver and muscle (quantitative immunohistochemistry) were higher (P<0.02 and P<0.07, respectively) in half-siblings than in dwarfs. The reduced presence of IGF-1R may have been the underlying cause of dwarfism in studied calves.

Synergistic effect of vitamin D and low concentration of transforming growth factor beta 1, a potential role in dermal wound healing.

PubMed

Ding, Jie; Kwan, Peter; Ma, Zengshuan; Iwashina, Takashi; Wang, Jianfei; Shankowsky, Heather A; Tredget, Edward E

2016-09-01

Dermal wound healing, in which transforming growth factor beta 1 (TGFβ1) plays an important role, is a complex process. Previous studies suggest that vitamin D has a potential regulatory role in TGFβ1 induced activation in bone formation, and there is cross-talk between their signaling pathways, but research on their effects in other types of wound healing is limited. The authors therefore wanted to explore the role of vitamin D and its interaction with low concentration of TGFβ1 in dermal fibroblast-mediated wound healing through an in vitro study. Human dermal fibroblasts were treated with vitamin D, TGFβ1, both, or vehicle, and then the wound healing functions of dermal fibroblasts were measured. To further explore possible mechanisms explaining the synergistic effect of vitamin D and TGFβ1, targeted gene silencing of the vitamin D receptor was performed. Compared to either factor alone, treatment of fibroblasts with both vitamin D and low concentration of TGFβ1 increased gene expression of TGFβ1, connective tissue growth factor, and fibronectin 1, and enhanced fibroblast migration, myofibroblast formation, and collagen production. Vitamin D receptor gene silencing blocked this synergistic effect of vitamin D and TGFβ1 on both collagen production and myofibroblast differentiation. Thus a synergistic effect of vitamin D and low TGFβ1 concentration was found in dermal fibroblast-mediated wound healing in vitro. This study suggests that supplementation of vitamin D may be an important step to improve wound healing and regeneration in patients with a vitamin D deficiency. Crown Copyright © 2016. Published by Elsevier Ltd. All rights reserved.

Expression of a transmembrane phosphotyrosine phosphatase inhibits cellular response to platelet-derived growth factor and insulin-like growth factor-1.

PubMed

Mooney, R A; Freund, G G; Way, B A; Bordwell, K L

1992-11-25

Tyrosine phosphorylation is a mechanism of signal transduction shared by many growth factor receptors and oncogene products. Phosphotyrosine phosphatases (PTPases) potentially modulate or counter-regulate these signaling pathways. To test this hypothesis, the transmembrane PTPase CD45 (leukocyte common antigen) was expressed in the murine cell line C127. Hormone-dependent autophosphorylation of the platelet-derived growth factor (PDGF) and insulin-like growth factor-1 (IGF-1) receptors was markedly reduced in cells expressing the transmembrane PTPase. Tyrosine phosphorylation of other PDGF-dependent phosphoproteins (160, 140, and 55 kDa) and IGF-1-dependent phosphoproteins (145 kDa) was similarly decreased. Interestingly, the pattern of growth factor-independent tyrosine phosphorylations was comparable in cells expressing the PTPase and control cells. This suggests a selectivity or accessibility of the PTPase limited to a subset of cellular phosphotyrosyl proteins. The maximum mitogenic response to PDGF and IGF-1 in cells expressing the PTPase was decreased by 67 and 71%, respectively. These results demonstrate that a transmembrane PTPase can both affect the tyrosine phosphorylation state of growth factor receptors and modulate proximal and distal cellular responses to the growth factors.

BDNF released during neuropathic pain potentiates NMDA receptors in primary afferent terminals

PubMed Central

Chen, Wenling; Walwyn, Wendy; Ennes, Helena S.; Kim, Hyeyoung; McRoberts, James A.; Marvizón, Juan Carlos G.

2014-01-01

NMDA receptors in primary afferent terminals can contribute to hyperalgesia by increasing neurotransmitter release. In rats and mice, we found that the ability of intrathecal NMDA to induce neurokinin 1 receptor (NK1R) internalization (a measure of substance P release) required a previous injection of BDNF. Selective knock-down of NMDA receptors in primary afferents decreased NMDA-induced NK1R internalization, confirming the presynaptic location of these receptors. The effect of BDNF was mediated by tropomyosin-related kinase B (trkB) receptors and not p75 neurotrophin receptors (p75NTR), because it was not produced by proBDNF and was inhibited by the trkB antagonist ANA-12 but not by the p75NTR inhibitor TAT-Pep5. These effects are probably mediated through the truncated form of the trkB receptor as there is little expression of full-length trkB in dorsal root ganglion (DRG) neurons. Src family kinase inhibitors blocked the effect of BDNF, suggesting that trkB receptors promote the activation of these NMDA receptors by Src family kinase phosphorylation. Western blots of cultured DRG neurons revealed that BDNF increased Tyr1472 phosphorylation of the NR2B subunit of the NMDA receptor, known to have a potentiating effect. Patch-clamp recordings showed that BDNF, but not proBDNF, increased NMDA receptor currents in cultured DRG neurons. NMDA-induced NK1R internalization was also enabled in a neuropathic pain model or by activating dorsal horn microglia with lipopolysaccharide. These effects were decreased by a BDNF scavenger, a trkB receptor antagonist and an Src family kinase inhibitor, indicating that BDNF released by microglia potentiates NMDA receptors in primary afferents during neuropathic pain. PMID:24611998

Regulatory role of tumor necrosis factor receptor-associated factor 6 in breast cancer by activating the protein kinase B/glycogen synthase kinase 3β signaling pathway.

PubMed

Shen, Hongyu; Li, Liangpeng; Yang, Sujin; Wang, Dandan; Zhou, Siying; Chen, Xiu; Tang, Jinhai

2017-08-01

Tumor necrosis factor receptor-associated factor 6 (TRAF6) is an endogenous adaptor of innate and adaptive immune responses, and serves a crucial role in tumor necrosis factor receptor and toll‑like/interleukin‑1 receptor signaling. Although studies have demonstrated that TRAF6 has oncogenic activity, its potential contributions to breast cancer in human remains largely uninvestigated. The present study examined the expression levels and function of TRAF6 in breast carcinoma (n=32) and adjacent healthy (n=25) tissue samples. Compared with adjacent healthy tissues, TRAF6 protein expression levels were significantly upregulated in breast cancer tissues. Reverse transcription‑quantitative polymerase chain reaction analysis revealed a significant upregulation of the cellular proliferative marker Ki‑67 and proliferation cell nuclear antigen expression levels in breast carcinoma specimens. Furthermore, protein expression levels of the accessory molecule, transforming growth factor β‑activated kinase 1 (TAK1), were significantly increased in breast cancer patients, as detected by western blot analysis. As determined by MTT assay, TRAF6 exerted profoundly proliferative effects in the MCF‑7 breast cancer cell line; however, these detrimental effects were ameliorated by TAK1 inhibition. Notably, protein kinase B (AKT)/glycogen synthase kinase (GSK)3β phosphorylation levels were markedly upregulated in breast cancer samples, compared with adjacent healthy tissues. In conclusion, an altered TRAF6‑TAK1 axis and its corresponding downstream AKT/GSK3β signaling molecules may contribute to breast cancer progression. Therefore, TRAF6 may represent a potential therapeutic target for the treatment of breast cancer.

Allosteric potentiation of quisqualate receptors by a nootropic drug aniracetam.

PubMed Central

Ito, I; Tanabe, S; Kohda, A; Sugiyama, H

1990-01-01

1. Allosteric potentiation of the ionotropic quisqualate (iQA) receptor by a nootropic drug aniracetam (1-p-anisoyl-2-pyrrolidinone) was investigated using Xenopus oocytes injected with rat brain mRNA and rat hippocampal slices. 2. Aniracetam potentiates the iQA responses induced in Xenopus oocytes by rat brain mRNA in a reversible manner. This effect was observed above the concentrations of 0.1 mM. Kainate. N-methyl-D-aspartate and gamma-aminobutyric acid responses induced in the same oocytes were not affected. 3. The specific potentiation of iQA responses was accompanied by an increase in the conductance change of iQA and alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) responses, but the affinity of receptors for agonist and the ion-selectivity of the channels (reversal potentials) were not changed. 4. Aniracetam reversibly potentiated the iQA responses recorded intracellularly from the pyramidal cells in the CA1 region of rat hippocampal slices. The excitatory postsynaptic potentials (EPSPs) in Schaffer collateral-commissural-CA1 synapses were also potentiated by aniracetam. 5. Population EPSPs recorded in the mossy fibre-CA3 synapses as well as Schaffer-commissural synapses were also potentiated by aniracetam. The amplitudes of the potentiation were not changed by the formation of long-term potentiation. PMID:1975272

Epidermal growth factor induces G protein-coupled receptor 30 expression in estrogen receptor-negative breast cancer cells.

PubMed

Albanito, Lidia; Sisci, Diego; Aquila, Saveria; Brunelli, Elvira; Vivacqua, Adele; Madeo, Antonio; Lappano, Rosamaria; Pandey, Deo Prakash; Picard, Didier; Mauro, Loredana; Andò, Sebastiano; Maggiolini, Marcello

2008-08-01

Different cellular receptors mediate the biological effects induced by estrogens. In addition to the classical nuclear estrogen receptors (ERs)-alpha and -beta, estrogen also signals through the seven-transmembrane G-protein-coupled receptor (GPR)-30. Using as a model system SkBr3 and BT20 breast cancer cells lacking the classical ER, the regulation of GPR30 expression by 17beta-estradiol, the selective GPR30 ligand G-1, IGF-I, and epidermal growth factor (EGF) was evaluated. Transient transfections with an expression plasmid encoding a short 5'-flanking sequence of the GPR30 gene revealed that an activator protein-1 site located within this region is required for the activating potential exhibited only by EGF. Accordingly, EGF up-regulated GPR30 protein levels, which accumulated predominantly in the intracellular compartment. The stimulatory role elicited by EGF on GPR30 expression was triggered through rapid ERK phosphorylation and c-fos induction, which was strongly recruited to the activator protein-1 site found in the short 5'-flanking sequence of the GPR30 gene. Of note, EGF activating the EGF receptor-MAPK transduction pathway stimulated a regulatory loop that subsequently engaged estrogen through GPR30 to boost the proliferation of SkBr3 and BT20 breast tumor cells. The up-regulation of GPR30 by ligand-activated EGF receptor-MAPK signaling provides new insight into the well-known estrogen and EGF cross talk, which, as largely reported, contributes to breast cancer progression. On the basis of our results, the action of EGF may include the up-regulation of GPR30 in facilitating a stimulatory role of estrogen, even in ER-negative breast tumor cells.

Design and characteristics of cytotoxic fibroblast growth factor 1 conjugate for fibroblast growth factor receptor-targeted cancer therapy.

PubMed

Szlachcic, Anna; Zakrzewska, Malgorzata; Lobocki, Michal; Jakimowicz, Piotr; Otlewski, Jacek

2016-01-01

Fibroblast growth factor receptors (FGFRs) are attractive candidate cancer therapy targets as they are overexpressed in multiple types of tumors, such as breast, prostate, bladder, and lung cancer. In this study, a natural ligand of FGFR, an engineered variant of fibroblast growth factor 1 (FGF1V), was conjugated to a potent cytotoxic drug, monomethyl auristatin E (MMAE), and used as a targeting agent for cancer cells overexpressing FGFRs, similar to antibodies in antibody-drug conjugates. The FGF1V-valine-citrulline-MMAE conjugate showed a favorable stability profile, bound FGFRs on the cell surface specifically, and efficiently released the drug (MMAE) upon cleavage by the lysosomal protease cathepsin B. Importantly, the conjugate showed a prominent cytotoxic effect toward cell lines expressing FGFR. FGF1V-vcMMAE was highly cytotoxic at concentrations even an order of magnitude lower than those found for free MMAE. This effect was FGFR-specific as cells lacking FGFR did not show any increased mortality.

Design and characteristics of cytotoxic fibroblast growth factor 1 conjugate for fibroblast growth factor receptor-targeted cancer therapy

PubMed Central

Szlachcic, Anna; Zakrzewska, Malgorzata; Lobocki, Michal; Jakimowicz, Piotr; Otlewski, Jacek

2016-01-01

Fibroblast growth factor receptors (FGFRs) are attractive candidate cancer therapy targets as they are overexpressed in multiple types of tumors, such as breast, prostate, bladder, and lung cancer. In this study, a natural ligand of FGFR, an engineered variant of fibroblast growth factor 1 (FGF1V), was conjugated to a potent cytotoxic drug, monomethyl auristatin E (MMAE), and used as a targeting agent for cancer cells overexpressing FGFRs, similar to antibodies in antibody–drug conjugates. The FGF1V–valine–citrulline–MMAE conjugate showed a favorable stability profile, bound FGFRs on the cell surface specifically, and efficiently released the drug (MMAE) upon cleavage by the lysosomal protease cathepsin B. Importantly, the conjugate showed a prominent cytotoxic effect toward cell lines expressing FGFR. FGF1V–vcMMAE was highly cytotoxic at concentrations even an order of magnitude lower than those found for free MMAE. This effect was FGFR-specific as cells lacking FGFR did not show any increased mortality. PMID:27563235

A-kinase anchoring protein 150 mediates transient receptor potential family V type 1 sensitivity to phosphatidylinositol-4,5-bisphosphate.

PubMed

Jeske, Nathaniel A; Por, Elaine D; Belugin, Sergei; Chaudhury, Sraboni; Berg, Kelly A; Akopian, Armen N; Henry, Michael A; Gomez, Ruben

2011-06-08

A-kinase anchoring protein 150 (AKAP150) is a scaffolding protein that controls protein kinase A- and C-mediated phosphorylation of the transient receptor potential family V type 1 (TRPV1), dictating receptor response to nociceptive stimuli. The phospholipid phosphatidylinositol-4,5-bisphosphate (PIP(2)) anchors AKAP150 to the plasma membrane in naive conditions and also affects TRPV1 activity. In the present study, we sought to determine whether the effects of PIP(2) on TRPV1 are mediated through AKAP150. In trigeminal neurons and CHO cells, the manipulation of cellular PIP(2) led to significant changes in the association of AKAP150 and TRPV1. Following PIP(2) degradation, increased TRPV1:AKAP150 coimmunoprecipitation was observed, resulting in increased receptor response to capsaicin treatment. Phospholipase C activation in neurons isolated from AKAP150(-/-) animals indicated that PIP(2)-mediated inhibition of TRPV1 in the whole-cell environment requires expression of the scaffolding protein. Furthermore, the addition of PIP(2) to neurons isolated from AKAP150 wild-type mice reduced PKA sensitization of TRPV1 compared with isolated neurons from AKAP150(-/-) mice. These findings suggest that PIP(2) degradation increases AKAP150 association with TRPV1 in the whole-cell environment, leading to sensitization of the receptor to nociceptive stimuli.

Signaling by Kit protein-tyrosine kinase--the stem cell factor receptor.

PubMed

Roskoski, Robert

2005-11-11

Signaling by stem cell factor and Kit, its receptor, plays important roles in gametogenesis, hematopoiesis, mast cell development and function, and melanogenesis. Moreover, human and mouse embryonic stem cells express Kit transcripts. Stem cell factor exists as both a soluble and a membrane-bound glycoprotein while Kit is a receptor protein-tyrosine kinase. The complete absence of stem cell factor or Kit is lethal. Deficiencies of either produce defects in red and white blood cell production, hypopigmentation, and sterility. Gain-of-function mutations of Kit are associated with several human neoplasms including acute myelogenous leukemia, gastrointestinal stromal tumors, and mastocytomas. Kit consists of an extracellular domain, a transmembrane segment, a juxtamembrane segment, and a protein kinase domain that contains an insert of about 80 amino acid residues. Binding of stem cell factor to Kit results in receptor dimerization and activation of protein kinase activity. The activated receptor becomes autophosphorylated at tyrosine residues that serve as docking sites for signal transduction molecules containing SH2 domains. The adaptor protein APS, Src family kinases, and Shp2 tyrosyl phosphatase bind to phosphotyrosine 568. Shp1 tyrosyl phosphatase and the adaptor protein Shc bind to phosphotyrosine 570. C-terminal Src kinase homologous kinase and the adaptor Shc bind to both phosphotyrosines 568 and 570. These residues occur in the juxtamembrane segment of Kit. Three residues in the kinase insert domain are phosphorylated and attract the adaptor protein Grb2 (Tyr703), phosphatidylinositol 3-kinase (Tyr721), and phospholipase Cgamma (Tyr730). Phosphotyrosine 900 in the distal kinase domain binds phosphatidylinositol 3-kinase which in turn binds the adaptor protein Crk. Phosphotyrosine 936, also in the distal kinase domain, binds the adaptor proteins APS, Grb2, and Grb7. Kit has the potential to participate in multiple signal transduction pathways as a result of

Role of fibroblast growth factor receptors (FGFR) and FGFR like-1 (FGFRL1) in mesenchymal stromal cell differentiation to osteoblasts and adipocytes.

PubMed

Kähkönen, T E; Ivaska, K K; Jiang, M; Büki, K G; Väänänen, H K; Härkönen, P L

2018-02-05

Fibroblast growth factors (FGF) and their receptors (FGFRs) regulate many developmental processes including differentiation of mesenchymal stromal cells (MSC). We developed two MSC lines capable of differentiating to osteoblasts and adipocytes and studied the role of FGFRs in this process. We identified FGFR2 and fibroblast growth factor receptor like-1 (FGFRL1) as possible actors in MSC differentiation with gene microarray and qRT-PCR. FGFR2 and FGFRL1 mRNA expression strongly increased during MSC differentiation to osteoblasts. FGF2 treatment, resulting in downregulation of FGFR2, or silencing FGFR2 expression with siRNAs inhibited osteoblast differentiation. During adipocyte differentiation expression of FGFR1 and FGFRL1 increased and was down-regulated by FGF2. FGFR1 knockdown inhibited adipocyte differentiation. Silencing FGFR2 and FGFR1 in MSCs was associated with decreased FGFRL1 expression in osteoblasts and adipocytes, respectively. Our results suggest that FGFR1 and FGFR2 regulate FGFRL1 expression. FGFRL1 may mediate or modulate FGFR regulation of MSC differentiation together with FGFR2 in osteoblastic and FGFR1 in adipocytic lineage. Copyright © 2017 Elsevier B.V. All rights reserved.

Evaluation of an In Vitro Multi-Receptor and Multi-Species Assay for Potential Endocrine Disruptor Targets (SOT)

EPA Science Inventory

Receptor sequence conservation across species may be a key factor determining susceptibility to potential endocrine disrupting chemicals. Computational approaches that compare receptor sequence similarities (e.g. SeqAPASS; https://seqapass.epa.gov/seqapass/) have been proposed to...

Maitotoxin Is a Potential Selective Activator of the Endogenous Transient Receptor Potential Canonical Type 1 Channel in Xenopus laevis Oocytes

PubMed Central

Flores, Pedro L.; Rodríguez, Emma; Zapata, Estrella; Carbó, Roxana; Farías, José María; Martínez, Martín

2017-01-01

Maitotoxin (MTX) is the most potent marine toxin known to date. It is responsible for a particular human intoxication syndrome called ciguatera fish poisoning (CFP). Several reports indicate that MTX is an activator of non-selective cation channels (NSCC) in different cell types. The molecular identity of these channels is still an unresolved topic, and it has been proposed that the transient receptor potential (TRP) channels are involved in this effect. In Xenopus laevis oocytes, MTX at picomolar (pM) concentrations induces the activation of NSCC with functional and pharmacological properties that resemble the activity of TRP channels. The purpose of this study was to characterize the molecular identity of the TRP channel involved in the MTX response, using the small interference RNA (siRNA) approach and the two-electrode voltage-clamp technique (TEVC). The injection of a specifically designed siRNA to silence the transient receptor potential canonical type 1 (TRPC1) protein expression abolished the MTX response. MTX had no effect on oocytes, even at doses 20-fold higher compared to cells without injection. Total mRNA and protein levels of TRPC1 were notably diminished. The TRPC4 siRNA did not change the MTX effect, even though it was important to note that the protein level was reduced by the silencing of TRPC4. Our results suggest that MTX could be a selective activator of TRPC1 channels in X. laevis oocytes and a useful pharmacological tool for further studies on these TRP channels. PMID:28672825

Maitotoxin Is a Potential Selective Activator of the Endogenous Transient Receptor Potential Canonical Type 1 Channel in Xenopus laevis Oocytes.

PubMed

Flores, Pedro L; Rodríguez, Emma; Zapata, Estrella; Carbó, Roxana; Farías, José María; Martínez, Martín

2017-06-25

Maitotoxin (MTX) is the most potent marine toxin known to date. It is responsible for a particular human intoxication syndrome called ciguatera fish poisoning (CFP). Several reports indicate that MTX is an activator of non-selective cation channels (NSCC) in different cell types. The molecular identity of these channels is still an unresolved topic, and it has been proposed that the transient receptor potential (TRP) channels are involved in this effect. In Xenopus laevis oocytes, MTX at picomolar (pM) concentrations induces the activation of NSCC with functional and pharmacological properties that resemble the activity of TRP channels. The purpose of this study was to characterize the molecular identity of the TRP channel involved in the MTX response, using the small interference RNA (siRNA) approach and the two-electrode voltage-clamp technique (TEVC). The injection of a specifically designed siRNA to silence the transient receptor potential canonical type 1 (TRPC1) protein expression abolished the MTX response. MTX had no effect on oocytes, even at doses 20-fold higher compared to cells without injection. Total mRNA and protein levels of TRPC1 were notably diminished. The TRPC4 siRNA did not change the MTX effect, even though it was important to note that the protein level was reduced by the silencing of TRPC4. Our results suggest that MTX could be a selective activator of TRPC1 channels in X. laevis oocytes and a useful pharmacological tool for further studies on these TRP channels.

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

PubMed

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.

Transient receptor potential channel superfamily: Role in lower urinary tract function.

PubMed

Ogawa, Teruyuki; Imamura, Tetsuya; Nakazawa, Masaki; Hiragata, Shiro; Nagai, Takashi; Minagawa, Tomonori; Yokoyama, Hitoshi; Ishikawa, Masakuni; Domen, Takahisa; Ishizuka, Osamu

2015-11-01

Lower urinary tract symptoms associated with neurogenic bladder and overactive bladder syndrome are mediated in part by members of the transient receptor potential channel superfamily. The best studied member of this superfamily is the vanilloid receptor. Other transient receptor potential channels, such as the melastatin receptor and the ankyrin receptor, are also active in the pathogenesis of lower urinary tract dysfunction. However, the detailed mechanisms by which the transient receptor potential channels contribute to lower urinary tract symptoms are still not clear, and the therapeutic benefits of modulating transient receptor potential channel activity have not been proved in the clinical setting. In the present review, to better understand the pathophysiology and therapeutic potential for lower urinary tract symptoms, we summarize the presence and role of different members of the transient receptor potential channel superfamily in the lower urinary tract. © 2015 The Japanese Urological Association.

Potentiation of adenosine A1 receptor agonist CPA-induced antinociception by paeoniflorin in mice.

PubMed

Liu, Da-Zhi; Zhao, Fei-Li; Liu, Jing; Ji, Xin-Quan; Ye, Yang; Zhu, Xing-Zu

2006-08-01

The effect of paeoniflorin (PF), a major constituent isolated from Paeony radix, on N6-Cyclopentyladenosine (CPA), a selective adenosine A1 receptor (A1 receptor) agonist, induced antinociception was examined in mice. In the tail-pressure test, CPA (0.05, 0.1, 0.2 mg/kg, s.c.) could induce antinociception in a dose-dependent manner. PF (5, 10, 20 mg/kg, s.c.) alone failed to exhibit any antinociceptive effect in mice; however, pretreatment of PF (20 mg/kg, s.c.) could significantly enhance CPA-induced antinociception. Additionally, pretreatment of 8-Cyclopentyl-1,3-dipropylxanthine (DPCPX, 0.25 mg/kg, s.c.), a selective A1 receptor antagonist, could antagonize the antinociceptive effect of combining CPA with PF. Furthermore, in the competitive binding experiments, PF did not displace the binding of [3H]-8-Cyclopentyl-1,3-dipropylxanthine ([3H]-DPCPX) but displaced that of [3H]-2-Chloro-N6-cyclopentyladenosine ([3H]-CCPA, a selective A1 receptor agonist) to the membrane preparation of rat cerebral cortex. These results suggested that PF might selectively increase the binding and antinociceptive effect of CPA by binding with A1 receptor.

Antitumor activity of a novel anti-vascular endothelial growth factor receptor-1 monoclonal antibody that does not interfere with ligand binding

PubMed Central

Tentori, Lucio; Scimeca, Manuel; Dorio, Annalisa S.; Atzori, Maria Grazia; Failla, Cristina M.; Morea, Veronica; Bonanno, Elena; D'Atri, Stefania; Lacal, Pedro M.

2016-01-01

Vascular endothelial growth factor receptor-1 (VEGFR-1) is a tyrosine kinase transmembrane receptor that has also a soluble isoform containing most of the extracellular ligand binding domain (sVEGFR-1). VEGF-A binds to both VEGFR-2 and VEGFR-1, whereas placenta growth factor (PlGF) interacts exclusively with VEGFR-1. In this study we generated an anti-VEGFR-1 mAb (D16F7) by immunizing BALB/C mice with a peptide that we had previously reported to inhibit angiogenesis and endothelial cell migration induced by PlGF. D16F7 did not affect binding of VEGF-A or PlGF to VEGFR-1, thus allowing sVEGFR-1 to act as decoy receptor for these growth factors, but it hampered receptor homodimerization and activation. D16F7 inhibited both the chemotactic response of human endothelial, myelomonocytic and melanoma cells to VEGFR-1 ligands and vasculogenic mimicry by tumor cells. Moreover, D16F7 exerted in vivo antiangiogenic effects in a matrigel plug assay. Importantly, D16F7 inhibited tumor growth and was well tolerated by B6D2F1 mice injected with syngeneic B16F10 melanoma cells. The antitumor effect was associated with melanoma cell apoptosis, vascular abnormalities and decrease of both monocyte/macrophage infiltration and myeloid progenitor mobilization. For all the above, D16F7 may be exploited in the therapy of metastatic melanoma and other tumors or pathological conditions involving VEGFR-1 activation. PMID:27655684

Novel drugs that target the estrogen-related receptor alpha: their therapeutic potential in breast cancer

PubMed Central

May, Felicity EB

2014-01-01

The incidence of breast cancer continues to rise: 1.7 million women were diagnosed with and 521,000 women died from breast cancer in 2012. This review considers first current treatment options: surgery; radiotherapy; and systemic endocrine, anti-biological, and cytotoxic therapies. Clinical management includes prevention, early detection by screening, treatment with curative intent, management of chronic disease, and palliative control of advanced breast cancer. Next, the potential of novel drugs that target DNA repair, growth factor dependence, intracellular and intercellular signal transduction, and cell cycle are considered. Estrogen-related receptor alpha has attracted attention as a therapeutic target in triple-negative breast cancers with de novo resistance to, and in breast cancers with acquired resistance to, endocrine therapies such as antiestrogens and aromatase inhibitors. Estrogen-related receptor alpha is an orphan receptor and transcription factor. Its activity is regulated by coregulator proteins and posttranslational modification. It is an energy sensor that controls adaptation to energy demand and may facilitate glycolytic metabolism and mitochondrial oxidative respiration in breast cancer cells. Estrogen-related receptor alpha increases breast cancer cell migration, proliferation, and tumor development. It is expressed at high levels in estrogen receptor-negative tumors, and is proposed to activate estrogen-responsive genes in endocrine-resistant tumors. The structures and functions of the ligand-binding domains of estrogen receptor alpha and estrogen-related receptor alpha, their ability to bind estrogens, phytoestrogens, and synthetic ligands, and the effects of ligand agonists, antagonists, and inverse agonists on biological activity, are evaluated. Synthetic ligands of estrogen-related receptor alpha have activity in preclinical models of metabolic disorders, diabetes, osteoporosis, and oncology. The clinical settings in which these novel

Neurotensin-induced Erk1/2 phosphorylation and growth of human colonic cancer cells are independent from growth factors receptors activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Massa, Fabienne; Tormo, Aurelie; Beraud-Dufour, Sophie

2011-10-14

Highlights: {yields} We compare intracellular pathways of NT and EGF in HT29 cells. {yields} NT does not transactivate EGFR. {yields} Transactivation of EGFR is not a general rule in cancer cell growth. -- Abstract: Neurotensin (NT) promotes the proliferation of human colonic cancer cells by undefined mechanisms. We already demonstrated that, in the human colon adenocarcinoma cell line HT29, the effects of NT were mediated by a complex formed between the NT receptor-1 (NTSR1) and-3 (NTSR3). Here we examined cellular mechanisms that led to NT-induced MAP kinase phosphorylation and growth factors receptors transactivation in colonic cancer cells and proliferation inmore » HT29 cells. With the aim to identify upstream signaling involved in NT-elicited MAP kinase activation, we found that the stimulatory effects of the peptide were totally independent from the activation of the epidermal growth factor receptor (EGFR) both in the HT29 and the HCT116 cells. NT was unable to promote phosphorylation of EGFR and to compete with EGF for its binding to the receptor. Pharmacological approaches allowed us to differentiate EGF and NT signaling in HT29 cells since only NT activation of Erk1/2 was shown to be sensitive to PKC inhibitors and since only NT increased the intracellular level of calcium. We also observed that NT was not able to transactivate Insulin-like growth factor receptor. Our findings indicate that, in the HT29 and HCT116 cell lines, NT stimulates MAP kinase phosphorylation and cell growth by a pathway which does not involve EGF system but rather NT receptors which transduce their own intracellular effectors. These results indicate that depending on the cell line used, blocking EGFR is not the general rule to inhibit NT-induced cancer cell proliferation.« less

Mortality Benefit of Recombinant Human Interleukin-1 Receptor Antagonist for Sepsis Varies by Initial Interleukin-1 Receptor Antagonist Plasma Concentration.

PubMed

Meyer, Nuala J; Reilly, John P; Anderson, Brian J; Palakshappa, Jessica A; Jones, Tiffanie K; Dunn, Thomas G; Shashaty, Michael G S; Feng, Rui; Christie, Jason D; Opal, Steven M

2018-01-01

interleukin-1 receptor antagonist treatment was not statistically significant. We report a heterogeneous effect of recombinant human interleukin-1 receptor antagonist on 28-day sepsis mortality that is potentially predictable by plasma interleukin-1 receptor antagonist in one trial. A precision clinical trial of recombinant human interleukin-1 receptor antagonist targeted to septic patients with high plasma interleukin-1 receptor antagonist may be worthy of consideration.

MiR-125a TNF receptor-associated factor 6 to inhibit osteoclastogenesis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Guo, Li-Juan; Liao, Lan; Yang, Li

MicroRNAs (miRNAs) play important roles in osteoclastogenesis and bone resorption. In the present study, we found that miR-125a was dramatically down-regulated during macrophage colony stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL) induced osteoclastogenesis of circulating CD14+ peripheral blood mononuclear cells (PBMCs). Overexpression of miR-125a in CD14+ PBMCs inhibited osteoclastogenesis, while inhibition of miR-125a promoted osteoclastogenesis. TNF receptor-associated factor 6 (TRAF6), a transduction factor for RANKL/RANK/NFATc1 signal, was confirmed to be a target of miR-125a. EMSA and ChIP assays confirmed that NFATc1 bound to the promoter of the miR-125a. Overexpression of NFATc1 inhibited miR-125a transcription, and blockmore » of NFATc1 expression attenuated RANKL-regulated miR-125a transcription. Here, we reported that miR-125a played a biological function in osteoclastogenesis through a novel TRAF6/ NFATc1/miR-125a regulatory feedback loop. It suggests that regulation of miR-125a expression may be a potential strategy for ameliorating metabolic disease. - Highlights: • MiR-125a was significantly down-regulated in osteoclastogenesis of CD14+ PBMCs. • MiR-125a inhibited osteoclast differentiation by targeting TRAF6. • NFATc1 inhibited miR-125a transciption by binding to the promoter of miR-125a. • TRAF6/NFATc1 and miR-125a form a regulatory feedback loop in osteoclastogenesis.« less

Transcriptional response to muscarinic acetylcholine receptor stimulation: regulation of Egr-1 biosynthesis by ERK, Elk-1, MKP-1, and calcineurin in carbachol-stimulated human neuroblastoma cells.

PubMed

Rössler, Oliver G; Henss, Isabell; Thiel, Gerald

2008-02-01

Carbachol-mediated activation of type M(3) muscarinic acetylcholine receptors induces the biosynthesis of the transcription factor Egr-1 in human SH-SY5Y neuroblastoma cells involving an activation of extracellular signal-regulated protein kinase. Carbachol triggered the phosphorylation of the ternary complex factor Elk-1, a key transcriptional regulator of serum response element-driven gene transcription, and strikingly enhanced the transcriptional activation potential of Elk-1. Chromatin immunoprecipitation experiments revealed that Elk-1 binds in vivo to the 5'-upstream region of the Egr-1 gene in carbachol-stimulated neuroblastoma cells. Together, these data indicate that Elk-1 connects the intracellular signaling cascade elicited by activation of M(3) muscarinic acetylcholine receptors with the transcription of the Egr-1 gene. Lentiviral-mediated expression of either MAP kinase phosphatase-1 (MKP-1) or a constitutively active mutant of calcineurin A inhibited Egr-1 biosynthesis following carbachol stimulation, indicating that these phosphatases function as shut-off devices of muscarinic acetylcholine receptor signaling. Additionally, carbachol stimulation increased transcription of a chromatin-embedded collagenase promoter/reporter gene, showing that AP-1 activity is enhanced in carbachol-stimulated neuroblastoma. Expression experiments revealed that both MKP-1 and a constitutively active mutant of calcineurin A impaired carbachol-induced upregulation of AP-1 activity. The fact that carbachol stimulation of neuroblastoma cells activates the transcription factors Egr-1 and AP-1 suggests that changes in the gene expression pattern are an integral part of muscarinic acetylcholine receptor signaling.

Arrestin Scaffolds NHERF1 to the P2Y12 Receptor to Regulate Receptor Internalization*

PubMed Central

Nisar, Shaista P.; Cunningham, Margaret; Saxena, Kunal; Pope, Robert J.; Kelly, Eamonn; Mundell, Stuart J.

2012-01-01

We have recently shown in a patient with mild bleeding that the PDZ-binding motif of the platelet G protein-coupled P2Y12 receptor (P2Y12R) is required for effective receptor traffic in human platelets. In this study we show for the first time that the PDZ motif-binding protein NHERF1 exerts a major role in potentiating G protein-coupled receptor (GPCR) internalization. NHERF1 interacts with the C-tail of the P2Y12R and unlike many other GPCRs, NHERF1 interaction is required for effective P2Y12R internalization. In vitro and prior to agonist stimulation P2Y12R/NHERF1 interaction requires the intact PDZ binding motif of this receptor. Interestingly on receptor stimulation NHERF1 no longer interacts directly with the receptor but instead binds to the receptor via the endocytic scaffolding protein arrestin. These findings suggest a novel model by which arrestin can serve as an adaptor to promote NHERF1 interaction with a GPCR to facilitate effective NHERF1-dependent receptor internalization. PMID:22610101

Arrestin scaffolds NHERF1 to the P2Y12 receptor to regulate receptor internalization.

PubMed

Nisar, Shaista P; Cunningham, Margaret; Saxena, Kunal; Pope, Robert J; Kelly, Eamonn; Mundell, Stuart J

2012-07-13

We have recently shown in a patient with mild bleeding that the PDZ-binding motif of the platelet G protein-coupled P2Y(12) receptor (P2Y(12)R) is required for effective receptor traffic in human platelets. In this study we show for the first time that the PDZ motif-binding protein NHERF1 exerts a major role in potentiating G protein-coupled receptor (GPCR) internalization. NHERF1 interacts with the C-tail of the P2Y(12)R and unlike many other GPCRs, NHERF1 interaction is required for effective P2Y(12)R internalization. In vitro and prior to agonist stimulation P2Y(12)R/NHERF1 interaction requires the intact PDZ binding motif of this receptor. Interestingly on receptor stimulation NHERF1 no longer interacts directly with the receptor but instead binds to the receptor via the endocytic scaffolding protein arrestin. These findings suggest a novel model by which arrestin can serve as an adaptor to promote NHERF1 interaction with a GPCR to facilitate effective NHERF1-dependent receptor internalization.

Tumor necrosis factor-α inhibits angiotensin II receptor type 1 expression in dorsal root ganglion neurons via β-catenin signaling.

PubMed

Yang, Y; Wu, H; Yan, J-Q; Song, Z-B; Guo, Q-L

2013-09-17

Both tumor necrosis factor (TNF)-α and the angiotensin (Ang) II/angiotensin II receptor type 1 (AT1) axis play important roles in neuropathic pain and nociception. In the present study, we explored the interaction between the two systems by examining the mutual effects between TNF-α and the Ang II/AT1 receptor axis in dorsal root ganglion (DRG) neurons. Rat DRG neurons were treated with TNF-α in different concentrations for different lengths of time in the presence or absence of transcription inhibitor actinomycin D, TNF receptor 1 (TNFR1) inhibitor SPD304, β-catenin signaling inhibitor CCT031374, or different kinase inhibitors. TNF-α decreased the AT1 receptor mRNA level as well as the AT1a receptor promoter activity in a dose-dependent manner within 30 h, which led to dose-dependent inhibition of Ang II-binding AT1 receptor level on the cell membrane. Actinomycin D (1 mg/ml), SPD304 (50 μM), p38 mitogen-activated protein kinase (MAPK) inhibitor PD169316 (25 μM), and CCT031374 (50 μM) completely abolished the inhibitory effect of TNF-α on AT1 receptor expression. TNF-α dose-dependently increased soluble β-catenin and phosphorylated GSK-3β levels, which was blocked by SPD304 and PD169316. In DRG neurons treated with AT2 receptor agonist CGP421140, or Ang II with or without AT1 receptor antagonist losartan or AT2 receptor antagonist PD123319 for 30 h, we found that Ang II and Ang II+PD123319 significantly decreased TNF-α expression, whereas CPG421140 and Ang II+losartan increased TNF-α expression. In conclusion, we demonstrate that TNF-α inhibits AT1 receptor expression at the transcription level via TNFR1 in rat DRG neurons by increasing the soluble β-catenin level through the p38 MAPK/GSK-3β pathway. In addition, Ang II appears to inhibit and induce TNF-α expression via the AT1 receptor and the AT2 receptor in DRG neurons, respectively. This is the first evidence of crosstalk between TNF-α and the Ang II/AT receptor axis in DRG neurons

Immunization against exon 1 decapeptides from the lutropin/choriogonadotropin receptor or the follitropin receptor as potential male contraceptive.

PubMed

Remy, J J; Couture, L; Rabesona, H; Haertle, T; Salesse, R

1996-11-01

Pituitary gonadotropin hormones lutropin (LH) and follitropin (FSH) control steroidogenesis and gametogenesis in male and female gonads through interaction with G protein-coupled receptors, LHR and FSHR. In the male, LH acts on leydig cells and is mostly responsible for the acquisition of puberty and the production of androgens while FSH, together with androgens, regulates spermatogenesis within Sertoli cells. We have engineered filamentous phages displaying mouse LHR and human FSHR decapeptides chosen in hormone binding regions. Peptides from both receptors displayed on phages belong either to the receptor specific exon 1 (amino acids 18-27) or to the homologous exon 4 (amino acids 98-107). Vaccination of prepubertal BALB/c male mice with hybrid phages using sub-cutaneous or intraperitoneal injections induced immunity against receptors. Anti-receptor immunization produced agonist or antagonist effects depending only on the circulating levels of the antibodies. Both anti-LHR and anti-FSHR vaccines induced efficient as well as reversible male contraception, through different mechanisms: targeting LH receptors inhibited or hyperstimulated Leydig cell testosterone production while targeting FSH receptors did not affect testosterone levels.

Ala67Thr mutation in the poliovirus receptor CD155 is a potential risk factor for vaccine and wild-type paralytic poliomyelitis.

PubMed

Kindberg, Elin; Ax, Cecilia; Fiore, Lucia; Svensson, Lennart

2009-05-01

Poliovirus infections can be asymptomatic or cause severe paralysis. Why some individuals develop paralytic poliomyelitis is unknown, but a role for host genetic factors has been suggested. To investigate if a polymorphism, Ala67Thr, in the poliovirus receptor, which has been found to facilitate increased resistance against poliovirus-induced cell lysis and apoptosis, is associated with increased risk of paralytic poliomyelitis, poliovirus receptor genotyping was undertaken among Italian subjects with vaccine-associated (n = 9), or with wild-type paralytic poliomyelitis (n = 6), and control subjects (n = 71), using RFLP-PCR and pyrosequencing. Heterozygous poliovirus receptor Ala67Thr genotype was found in 13.3% of the patients with paresis and in 8.5% of the controls (Odds Ratio = 1.667). The frequency of Ala67Thr among the controls is in agreement with earlier published data. It is concluded that the Ala67Thr mutation in the poliovirus receptor is a possible risk factor for the development of vaccine-associated or paralytic poliomyelitis associated with wild-type virus. Copyright 2009 Wiley-Liss, Inc.

P2X1 receptor-mediated inhibition of the proliferation of human coronary smooth muscle cells involving the transcription factor NR4A1.

PubMed

Hinze, Annette Viktoria; Mayer, Peter; Harst, Anja; von Kügelgen, Ivar

2013-12-01

Adenine nucleotides acting at P2X1 receptors are potent vasoconstrictors. Recently, we demonstrated that activation of adenosine A2B receptors on human coronary smooth muscle cells inhibits cell proliferation by the induction of the nuclear receptor subfamily 4, group A, member 1 (NR4A1; alternative notation Nur77). In the present study, we searched for long-term effects mediated by P2X1 receptors by analyzing receptor-mediated changes in cell proliferation and in the expression of NR4A1. Cultured human coronary smooth muscle cells were treated with selective receptor ligands. Effects on proliferation were determined by counting cells and measuring changes in impedance. The induction of transcription factors was assessed by qPCR. The P2X receptor agonist α,β-methylene-ATP and its analog β,γ-methylene-ATP inhibited cell proliferation by about 50 % after 5 days in culture with half-maximal concentrations of 0.3 and 0.08 μM, respectively. The effects were abolished or markedly attenuated by the P2X1 receptor antagonist NF449 (carbonylbis-imino-benzene-triylbis-(carbonylimino)tetrakis-benzene-1,3-disulfonic acid; 100 nM and 1 μM). α,β-methylene-ATP and β,γ-methylene-ATP applied for 30 min to 4 h increased the expression of NR4A1; NF449 blocked or attenuated this effect. Small interfering RNA directed against NR4A1 diminished the antiproliferative effects of α,β-methylene-ATP and β,γ-methylene-ATP. α,β-methylene-ATP (0.1 to 30 μM) decreased migration of cultured human coronary smooth muscle cells in a chamber measuring changes in impedance; NF449 blocked the effect. In conclusion, our results demonstrate for the first time that adenine nucleotides acting at P2X1 receptors inhibit the proliferation of human coronary smooth muscle cells via the induction of the early gene NR4A1.

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.

PubMed

Annenkov, Alexander; Rigby, Anne; Amor, Sandra; Zhou, Dun; Yousaf, Nasim; Hemmer, Bernhard; Chernajovsky, Yuti

2011-08-01

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) signalling cascade. Activation of this pro-survival pathway in response to ligand broadly available in the brain might increase neuroregenerative potential of transplanted precursors. The ChR was produced by fusing a MOG-specific single ‎chain antibody with the extracellular boundary of the IGF1R transmembrane segment. The ChR is expressed on the cellular surface, predominantly as a monomer, and is not N-glycosylated. To show MOG-dependent functionality of the ChR, neuroblastoma cells B104 expressing this ChR were stimulated with monolayers of cells expressing recombinant MOG. The ChR undergoes MOG-dependent tyrosine phosphorylation and homodimerisation. It promotes insulin and IGF-independent growth of the oligodendrocyte progenitor cell line CG4. The proposed mode of the ChR activation is by MOG-induced dimerisation which promotes kinase domain transphosphorylation, by-passing the requirement of conformation changes known to be important for IGF1R activation. Another ChR, which contains a segment of the β-chain ectodomain, was produced in an attempt to recapitulate some of these conformational changes, but proved non-functional. 2011 Elsevier B.V. All rights reserved.

Taci Is a Traf-Interacting Receptor for Tall-1, a Tumor Necrosis Factor Family Member Involved in B Cell Regulation

PubMed Central

Xia, Xing-Zhong; Treanor, James; Senaldi, Giorgio; Khare, Sanjay D.; Boone, Tom; Kelley, Michael; Theill, Lars E.; Colombero, Anne; Solovyev, Irina; Lee, Frances; McCabe, Susan; Elliott, Robin; Miner, Kent; Hawkins, Nessa; Guo, Jane; Stolina, Marina; Yu, Gang; Wang, Judy; Delaney, John; Meng, Shi-Yuan; Boyle, William J.; Hsu, Hailing

2000-01-01

We and others recently reported tumor necrosis factor (TNF) and apoptosis ligand–related leukocyte-expressed ligand 1 (TALL-1) as a novel member of the TNF ligand family that is functionally involved in B cell proliferation. Transgenic mice overexpressing TALL-1 have severe B cell hyperplasia and lupus-like autoimmune disease. Here, we describe expression cloning of a cell surface receptor for TALL-1 from a human Burkitt's lymphoma RAJI cell library. The cloned receptor is identical to the previously reported TNF receptor (TNFR) homologue transmembrane activator and calcium modulator and cyclophilin ligand (CAML) interactor (TACI). Murine TACI was subsequently isolated from the mouse B lymphoma A20 cells. Human and murine TACI share 54% identity overall. Human TACI exhibits high binding affinities to both human and murine TALL-1. Soluble TACI extracellular domain protein specifically blocks TALL-1–mediated B cell proliferation without affecting CD40- or lipopolysaccharide-mediated B cell proliferation in vitro. In addition, when injected into mice, soluble TACI inhibits antibody production to both T cell–dependent and –independent antigens. By yeast two-hybrid screening of a B cell library with TACI intracellular domain, we identified that, like many other TNFR family members, TACI intracellular domain interacts with TNFR-associated factor (TRAF)2, 5, and 6. Correspondingly, TACI activation in a B cell line results in nuclear factor κB and c-Jun NH2-terminal kinase activation. The identification and characterization of the receptor for TALL-1 provides useful information for the development of a treatment for B cell–mediated autoimmune diseases such as systemic lupus erythematosus. PMID:10880535

Activation of the chemosensing transient receptor potential channel A1 (TRPA1) by alkylating agents.

PubMed

Stenger, Bernhard; Zehfuss, Franziska; Mückter, Harald; Schmidt, Annette; Balszuweit, Frank; Schäfer, Eva; Büch, Thomas; Gudermann, Thomas; Thiermann, Horst; Steinritz, Dirk

2015-09-01

The transient receptor potential ankyrin 1 (TRPA1) cation channel is expressed in different tissues including skin, lung and neuronal tissue. Recent reports identified TRPA1 as a sensor for noxious substances, implicating a functional role in the molecular toxicology. TRPA1 is activated by various potentially harmful electrophilic substances. The chemical warfare agent sulfur mustard (SM) is a highly reactive alkylating agent that binds to numerous biological targets. Although SM is known for almost 200 years, detailed knowledge about the pathophysiology resulting from exposure is lacking. A specific therapy is not available. In this study, we investigated whether the alkylating agent 2-chloroethyl-ethylsulfide (CEES, a model substance for SM-promoted effects) and SM are able to activate TRPA1 channels. CEES induced a marked increase in the intracellular calcium concentration ([Ca(2+)]i) in TRPA1-expressing but not in TRPA1-negative cells. The TRP-channel blocker AP18 diminished the CEES-induced calcium influx. HEK293 cells permanently expressing TRPA1 were more sensitive toward cytotoxic effects of CEES compared with wild-type cells. At low CEES concentrations, CEES-induced cytotoxicity was prevented by AP18. Proof-of-concept experiments using SM resulted in a pronounced increase in [Ca(2+)]i in HEK293-A1-E cells. Human A549 lung epithelial cells, which express TRPA1 endogenously, reacted with a transient calcium influx in response to CEES exposure. The CEES-dependent calcium response was diminished by AP18. In summary, our results demonstrate that alkylating agents are able to activate TRPA1. Inhibition of TRPA1 counteracted cellular toxicity and could thus represent a feasible approach to mitigate SM-induced cell damage.

Allosteric modulation of sigma-1 receptors elicits anti-seizure activities.

PubMed

Guo, Lin; Chen, Yanke; Zhao, Rui; Wang, Guanghui; Friedman, Eitan; Zhang, Ao; Zhen, Xuechu

2015-08-01

Application of orthosteric sigma-1 receptor agonists as anti-seizure drugs has been hindered by questionable efficacy and potential adverse effects. Here, we have investigated the anti-seizure effects of the novel and potent allosteric modulator of sigma-1 receptors, SKF83959 and its derivative SOMCL-668 (3-methyl-phenyl-2,3,4,5-tetrahydro-1H-benzo[d]azepin-7-ol). The anti-seizure effects of SKF83959 were investigated in three mouse models, maximal electroshock seizures, pentylenetetrazole-induced convulsions and kainic acid-induced 'status epilepticus'. Also, in rats, the cortical epileptiform activity induced by topical application of picrotoxin was recorded in electrocorticograms. In rat hippocampal brain slices, effects of the drugs on the high potassium-evoked epileptiform local field potentials were studied. Anti-seizure activities of SOMCL-668, a newly developed sigma-1 receptor selective allosteric modulator, were also investigated. SKF83959 (20, 40 mg·kg(-1) ) exhibited anti -seizure actitity in the three mouse models and reduced the cortical epileptiform activity without alteration of spontaneous motor activity and motor coordination. These effects were blocked by the sigma-1 receptor antagonist BD1047, but not the dopamine D1 receptor antagonist SCH23390. SKF83959 alone did not directly inhibit the epileptiform firing of CA3 neurons induced by high potassium in hippocampal slices, but did potentiate inhibition by the orthosteric sigma-1 receptor agonist SKF10047. Lastly, a selective sigma-1 receptor allosteric modulator SOMCL-668, which does not bind to dopamine receptors, exerted similar anti-seizure activities. SKF83959 and SOMCL-668 displayed anti-seizure activities, indicating that allosteric modulation of sigma-1 receptors may provide a novel approach for discovering new anti-seizure drugs. © 2015 The British Pharmacological Society.

Synthesis and biological evaluation of aryl isoxazole derivatives as metabotropic glutamate receptor 1 antagonists: a potential treatment for neuropathic pain.

PubMed

Cho, Gyeong Hi; Kim, TaeHun; Son, Woo Seung; Seo, Seon Hee; Min, Sun-Joon; Cho, Yong Seo; Keum, Gyochang; Jeong, Kyu-Sung; Koh, Hun Yeong; Lee, Jiyoun; Pae, Ae Nim

2015-03-15

Glutamate is the major excitatory neurotransmitter and known to activate the metabotropic and ionotropic glutamate receptors in the brain. Among these glutamate receptors, metabotropic glutamate receptor 1 (mGluR1) has been implicated in various brain disorders including anxiety, schizophrenia and chronic pain. Several studies demonstrated that the blockade of mGluR1 signaling reduced pain responses in animal models, suggesting that mGluR1 is a promising target for the treatment of neuropathic pain. In this study, we have developed mGluR1 antagonists with an aryl isoxazole scaffold, and identify several compounds that are orally active in vivo. We believe that these compounds can serve as a useful tool for the investigation of the role of mGluR1 and a promising lead for the potential treatment of neuropathic pain. Copyright © 2015 Elsevier Ltd. All rights reserved.

Agonist- and subunit-dependent potentiation of glutamate receptors by a nootropic drug aniracetam.

PubMed

Tsuzuki, K; Takeuchi, T; Ozawa, S

1992-11-01

GluR1 and GluR2 cDNAs encoding non-NMDA subtypes of glutamate receptor were isolated from a rat brain cDNA library by Boulter et al. (Science, 249 (1990) 1033-1037). Functional receptors activated by kainate, alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) and glutamate were expressed in Xenopus oocytes injected with GluR1, GluR2 or a mixture of GluR1 and GluR2 RNAs. In GluR1-expressed oocytes, 1 mM aniracetam potentiated AMPA-induced currents by 99 +/- 10% (mean +/- S.E.M., n = 5) and glutamate-induced currents by 140 +/- 8% (n = 4), but little affected kainate-induced currents. Aniracetam was effective from a concentration of 0.1 mM, and it exhibited more conspicuous effects with the increase of the dose. In oocytes injected with GluR1 plus GluR2 RNAs, aniracetam more markedly potentiated current responses to AMPA and glutamate than those in oocytes injected with GluR1 RNA alone. For example, 1 mM aniracetam potentiated AMPA-induced currents by 396 +/- 76% (n = 4) and glutamate-induced currents by 970 +/- 65% (n = 5) in oocytes injected with 10% GluR1 and 90% GluR2 RNAs. In these oocytes, however, the potentiation of kainate-induced currents by 1 mM aniracetam was only 8 +/- 5% (n = 4). Thus, we conclude that the potentiation of the AMPA/kainate receptor by aniracetam depends on both species of agonists and subunit composition of the receptor.

Role of IL-1 beta and 5-HT2 receptors in midbrain periaqueductal gray (PAG) in potentiating defensive rage behavior in cat.

PubMed

Bhatt, Suresh; Bhatt, Rekha; Zalcman, Steven S; Siegel, Allan

2008-02-01

Feline defensive rage, a form of aggressive behavior that occurs in response to a threat can be elicited by electrical stimulation of the medial hypothalamus or midbrain periaqueductal gray (PAG). Our laboratory has recently begun a systematic examination of the role of cytokines in the regulation of rage and aggressive behavior. It was shown that the cytokine, interleukin-2 (IL-2), differentially modulates defensive rage when microinjected into the medial hypothalamus and PAG by acting through separate neurotransmitter systems. The present study sought to determine whether a similar relationship exists with respect to interleukin 1-beta (IL-1 beta), whose receptor activation in the medial hypothalamus potentiates defensive rage. Thus, the present study identified the effects of administration of IL-1 beta into the PAG upon defensive rage elicited from the medial hypothalamus. Microinjections of IL-1 beta into the dorsal PAG significantly facilitated defensive rage behavior elicited from the medial hypothalamus in a dose and time dependent manner. In addition, the facilitative effects of IL-1 beta were blocked by pre-treatment with anti-IL-1 beta receptor antibody, while IL-1 beta administration into the PAG had no effect upon predatory attack elicited from the lateral hypothalamus. The findings further demonstrated that IL-1 beta's effects were mediated through 5-HT(2) receptors since pretreatment with a 5-HT(2C) receptors antagonist blocked the facilitating effects of IL-1 beta. An extensive pattern of labeling of IL-1 beta and 5-HT(2C) receptors in the dorsal PAG supported these findings. The present study demonstrates that IL-beta in the dorsal PAG, similar to the medial hypothalamus, potentiates defensive rage behavior and is mediated through a 5-HT(2C) receptor mechanism.

Functional properties of an isolated. cap alpha beta. heterodimeric human placenta insulin-like growth factor 1 receptor complex

DOE Office of Scientific and Technical Information (OSTI.GOV)

Feltz, S.M.; Swanson, M.L.; Wemmie, J.A.

1988-05-03

Treatment of human placenta membranes at pH 8.5 in the presence of 2.0 mM dithiothreitol (DTT) for 5 min, followed by the simultaneous removal of the DTT and pH adjustment of pH 7.6, resulted in the formation of a functional ..cap alpha beta.. heterodimeric insulin-like growth factor 1 (IGF-1) receptor complex from the native ..cap alpha../sub 2/..beta../sub 2/ heterotetrameric disulfide-linked state. The membrane-bound ..cap alpha beta.. heterodimeric complex displayed similar curvilinear /sup 125/I-IGF-1 equilibrium binding compared to the ..cap alpha../sub 2/..beta../sub 2/ heterotetrameric complex. /sup 125/I-IGF-1 binding to both the isolated ..cap alpha../sub 2/..beta../sub 2/ heterotetrameric and ..cap alpha beta..more » heterodimeric complexes demonstrated a marked straightening of the Scatchard plots, compared to the placenta membrane-bound IGF-1 receptors, with a 2-fold increase in the high-affinity binding component. IGF-1 stimulation of IGF-1 receptor autophosphorylation indicated that the ligand-dependent activation of ..cap alpha beta.. heterodimeric protein kinase activity occurred concomitant with the reassociation into a covalent ..cap alpha../sub 2/..beta../sub 2/ heterotetrameric state. These data demonstrate that (i) a combination of alkaline pH and DTT treatment of human placenta membranes results in the formation of an ..cap alpha beta.. heterodimeric IGF-1 receptor complex, (ii) unlike the insulin receptor, high-affinity homogeneous IGF-1 binding occurs in both the ..cap alpha../sub 2/..beta../sub 2/ heterotetrameric and ..cap alpha beta.. heterodimeric complexes, and (iii) IGF-1-dependent autophosphorylation of the ..cap alpha beta.. heterodimeric IGF-1 receptor complex correlates wit an IGF-1 dependent covalent reassociation into an ..cap alpha../sub 2/..beta../sub 2/ heterotetrameric disulfide-linked state.« less

Tumor necrosis factor receptor-1 can function through a G alpha q/11-beta-arrestin-1 signaling complex.

PubMed

Kawamata, Yuji; Imamura, Takeshi; Babendure, Jennie L; Lu, Juu-Chin; Yoshizaki, Takeshi; Olefsky, Jerrold M

2007-09-28

Tumor necrosis factor-alpha (TNFalpha) is a proinflammatory cytokine secreted from macrophages and adipocytes. It is well known that chronic TNFalpha exposure can lead to insulin resistance both in vitro and in vivo and that elevated blood levels of TNFalpha are observed in obese and/or diabetic individuals. TNFalpha has many acute biologic effects, mediated by a complex intracellular signaling pathway. In these studies we have identified new G-protein signaling components to this pathway in 3T3-L1 adipocytes. We found that beta-arrestin-1 is associated with TRAF2 (TNF receptor-associated factor 2), an adaptor protein of TNF receptors, and that TNFalpha acutely stimulates tyrosine phosphorylation of G alpha(q/11) with an increase in G alpha(q/11) activity. Small interfering RNA-mediated knockdown of beta-arrestin-1 inhibits TNFalpha-induced tyrosine phosphorylation of G alpha(q/11) by interruption of Src kinase activation. TNFalpha stimulates lipolysis in 3T3-L1 adipocytes, and beta-arrestin-1 knockdown blocks the effects of TNFalpha to stimulate ERK activation and glycerol release. TNFalpha also led to activation of JNK with increased expression of the proinflammatory gene, monocyte chemoattractant protein-1 and matrix metalloproteinase 3, and beta-arrestin-1 knockdown inhibited both of these effects. Taken together these results reveal novel elements of TNFalpha action; 1) the trimeric G-protein component G alpha(q/11) and the adapter protein beta-arrestin-1 can function as signaling molecules in the TNFalpha action cascade; 2) beta-arrestin-1 can couple TNFalpha stimulation to ERK activation and lipolysis; 3) beta-arrestin-1 and G alpha(q/11) can mediate TNFalpha-induced phosphatidylinositol 3-kinase activation and inflammatory gene expression.

Genetic and pharmacological antagonism of NK1 receptor prevents opiate abuse potential.

PubMed

Sandweiss, A J; McIntosh, M I; Moutal, A; Davidson-Knapp, R; Hu, J; Giri, A K; Yamamoto, T; Hruby, V J; Khanna, R; Largent-Milnes, T M; Vanderah, T W

2017-05-09

Development of an efficacious, non-addicting analgesic has been challenging. Discovery of novel mechanisms underlying addiction may present a solution. Here we target the neurokinin system, which is involved in both pain and addiction. Morphine exerts its rewarding actions, at least in part, by inhibiting GABAergic input onto substance P (SP) neurons in the ventral tegmental area (VTA), subsequently increasing SP release onto dopaminergic neurons. Genome editing of the neurokinin 1 receptor (NK 1 R) in the VTA renders morphine non-rewarding. Complementing our genetic approach, we demonstrate utility of a bivalent pharmacophore with dual activity as a μ/δ opioid agonist and NK 1 R antagonist in inhibiting nociception in an animal model of acute pain while lacking any positive reinforcement. These data indicate that dual targeting of the dopaminergic reward circuitry and pain pathways with a multifunctional opioid agonist-NK 1 R antagonist may be an efficacious strategy in developing future analgesics that lack abuse potential.Molecular Psychiatry advance online publication, 9 May 2017; doi:10.1038/mp.2017.102.

Role of peripheral sigma-1 receptors in ischaemic pain: Potential interactions with ASIC and P2X receptors.

PubMed

Kwon, S G; Roh, D H; Yoon, S Y; Choi, S R; Choi, H S; Moon, J Y; Kang, S Y; Kim, H W; Han, H J; Beitz, A J; Oh, S B; Lee, J H

2016-04-01

The role of peripheral sigma-1 receptors (Sig-1Rs) in normal nociception and in pathologically induced pain conditions has not been thoroughly investigated. Since there is mounting evidence that Sig-1Rs modulate ischaemia-induced pathological conditions, we investigated the role of Sig-1Rs in ischaemia-induced mechanical allodynia (MA) and addressed their possible interaction with acid-sensing ion channels (ASICs) and P2X receptors at the ischaemic site. We used a rodent model of hindlimb thrombus-induced ischaemic pain (TIIP) to investigate their role. Western blot was performed to observe changes in Sig-1R expression in peripheral nervous tissues. MA was measured after intraplantar (i.pl.) injections of antagonists for the Sig-1, ASIC and P2X receptors in TIIP rats or agonists of each receptor in naïve rats. Sig-1R expression significantly increased in skin, sciatic nerve and dorsal root ganglia at 3 days post-TIIP surgery. I.pl. injections of the Sig-1R antagonist, BD-1047 on post-operative days 0-3 significantly attenuated the development of MA during the induction phase, but had no effect on MA when given during the maintenance phase (days 3-6 post-surgery). BD-1047 synergistically increased amiloride (an ASICs blocker)- and TNP-ATP (a P2X antagonist)-induced analgesic effects in TIIP rats. In naïve rats, i.pl. injection of Sig-1R agonist PRE-084 alone did not produce MA; but it did induce MA when co-administered with either an acidic pH solution or a sub-effective dose of αβmeATP. Peripheral Sig-1Rs contribute to the induction of ischaemia-induced MA via facilitation of ASICs and P2X receptors. Thus, peripheral Sig-1Rs represent a novel therapeutic target for the treatment of ischaemic pain. © 2015 European Pain Federation - EFIC®

Transient receptor potential canonical channel-1 (TRPC1) KO mice that exercise are protected from high-fat diet-induced obesity and type 2 diabetes risk

USDA-ARS?s Scientific Manuscript database

Objective: Transient receptor potential canonical channel-1 (TRPC1) is a major class of calcium permeable channels found in key metabolic tissues, including the hypothalamus, adipose tissue, and skeletal muscle, making them likely candidates for the regulation of cellular energy metabolism. The exac...

Functionally important amino acid residues in the transient receptor potential vanilloid 1 (TRPV1) ion channel – an overview of the current mutational data

PubMed Central

2013-01-01

This review aims to create an overview of the currently available results of site-directed mutagenesis studies on transient receptor potential vanilloid type 1 (TRPV1) receptor. Systematization of the vast number of data on the functionally important amino acid mutations of TRPV1 may provide a clearer picture of this field, and may promote a better understanding of the relationship between the structure and function of TRPV1. The review summarizes information on 112 unique mutated sites along the TRPV1, exchanged to multiple different residues in many cases. These mutations influence the effect or binding of different agonists, antagonists, and channel blockers, alter the responsiveness to heat, acid, and voltage dependence, affect the channel pore characteristics, and influence the regulation of the receptor function by phosphorylation, glycosylation, calmodulin, PIP2, ATP, and lipid binding. The main goal of this paper is to publish the above mentioned data in a form that facilitates in silico molecular modelling of the receptor by promoting easier establishment of boundary conditions. The better understanding of the structure-function relationship of TRPV1 may promote discovery of new, promising, more effective and safe drugs for treatment of neurogenic inflammation and pain-related diseases and may offer new opportunities for therapeutic interventions. PMID:23800232

Insulin-like growth factor-1 receptor is regulated by microRNA-133 during skeletal myogenesis.

PubMed

Huang, Mian-Bo; Xu, Hui; Xie, Shu-Juan; Zhou, Hui; Qu, Liang-Hu

2011-01-01

The insulin-like growth factor (IGF) signaling pathway has long been established as playing critical roles in skeletal muscle development. However, the underlying regulatory mechanism is poorly understood. Recently, a large family of small RNAs, named microRNAs (miRNAs), has been identified as key regulators for many developmental processes. Because miRNAs participate in the regulation of various signaling pathways, we hypothesized that miRNAs may be involved in the regulation of IGF signaling in skeletal myogenesis. In the present study, we determined that the cell-surface receptor IGF-1R is directly regulated by a muscle-specific miRNA, microRNA-133 (miR-133). A conserved and functional binding site for miR-133 was identified in the 3'untranslated region (3'UTR) of IGF-1R. During differentiation of C2C12 myoblasts, IGF-1R protein, but not messenger RNA (mRNA) expression, was gradually reduced, concurrent with the upregulation of miR-133. Overexpression of miR-133 in C2C12 cells significantly suppressed IGF-1R expression at the posttranscriptional level. We also demonstrated that both overexpression of miR-133 and knockdown of IGF-1R downregulated the phosphorylation of Akt, the central mediator of the PI3K/Akt signaling pathway. Furthermore, upregulation of miR-133 during C2C12 differentiation was significantly accelerated by the addition of IGF-1. Mechanistically, we found that the expression of myogenin, a myogenic transcription factor reported to transactivate miR-133, was increased by IGF-1 stimulation. Our results elucidate a negative feedback circuit in which IGF-1-stimulated miR-133 in turn represses IGF-1R expression to modulate the IGF-1R signaling pathway during skeletal myogenesis. These findings also suggest that miR-133 may be a potential therapeutic target in muscle diseases.

43.1 GENETIC INSIGHTS LEAD TO DISCOVERY OF SELECTIVE ACTIVATORS OF MGLU1 AND MGLU3 METABOTROPIC GLUTAMATE RECEPTORS AS POTENTIAL TREATMENTS FOR SCHIZOPHRENIA

PubMed Central

Conn, P Jeffrey; Yohn, Samantha; Stansley, Branden; Foster, Dan; Plumley, Hyekyung; Lindsley, Craig

2018-01-01

Abstract Background A large number of clinical and preclinical studies suggest that dysfunction at synapses for the excitatory neurotransmitter glutamate may play a critical role in the pathophysiological changes that underlie each of the major symptom clusters observed in schizophrenia patients. Interestingly, recent genetic studies identified multiple nonsynonymous single nucleotide polymorphisms (SNPs) in the human genes encoding two specific subtypes of metabotropic glutamate (mGlu) receptors that are associated with schizophrenia. These include GRM1 and GRM3, the genes encoding for the mGlu1 and mGlu3 receptor subtypes respectively. Furthermore, postmortem studies suggest that expression of these mGlu receptor subtypes is altered brains of schizophrenia patients compared to controls. Mutations in GRM1 were identified a range of schizophrenia patients, whereas SNPs in the human gene encoding mGlu3 (GRM3) are selectively associated with poor performance on cognitive tests that are dependent on function of the prefrontal cortex (PFC) and hippocampus. These studies raise the possibility that disrupted signaling of mGlu1 and/or mGlu3 could contribute to the symptoms of schizophrenia and that selective modulators of these receptors could provide a novel approach to treatment of this disorder. Methods Wild-type and mutant forms of mGlu receptors were expressed in cell lines and used for discovery and optimization of highly selective positive allosteric modulators (PAMs) of mGlu1 and mGlu3. Optimized mGlu1 and mGlu3 PAMs were then used along with mouse genetic studies to evaluate the roles of these receptors in specific basal ganglia and forebrain circuits that have been implicated in schizophrenia. Finally, these compounds were used in animal models to assess potential efficacy in rodent models that are relevant for reducing positive, negative, and cognitive symptoms that are observed in schizophrenia patients. Results GRM1 mutations associated with schizophrenia

Autoantibody against angiotensin AT1 receptor from preeclamptic patients enhances collagen-induced human platelet aggregation.

PubMed

Bai, Kehua; Wang, Ke; Li, Xiaoyu; Wang, Jie; Zhang, Jie; Song, Li; Wang, Jin; Zhang, Suli; Lau, Wayne Bond; Ma, Xinliang; Liu, Huirong

2013-09-01

Hypercoagulability, platelet activation, and thrombocytopenia are the chief characteristics of preeclampsia, but their responsible underlying molecular mechanisms remain obscure. Recent studies have demonstrated that the autoantibody against angiotensin II type 1 receptor (AT1-AA) constitutes a novel risk factor for preeclampsia. However, the role of AT1-AA in platelet activation and hypercoagulability in preeclampsia has never been investigated. In the present study, we determined whether AT1-AA promotes platelet aggregation in vitro, and dissected the potential underlying mechanisms. AT1-AA was detected by enzyme-linked immunosorbent assay. After immunoglobulin G fractions purified from the preeclamptic patient positive sera were added to platelets isolated from healthy volunteers, platelet aggregation and intracellular Ca(2+) levels were detected. AT1-AA significantly enhanced in vitro collagen-induced platelet aggregation, an effect blocked by the AT1 receptor antagonist losartan. Additionally, AT1-AA increased and maintained collagen-induced cytosolic calcium concentration throughout the experiment. We demonstrated for the first time that AT1-AA significantly promotes collagen-induced platelet aggregation through angiotensin type 1 receptor activation in vitro, potentially via increased intracellular Ca(2+) concentration, supporting AT1-AA as a potential contributor to the hypercoagulable state of preeclampsia.

Modulation of neurosteroid potentiation by protein kinases at synaptic- and extrasynaptic-type GABAA receptors

PubMed Central

Adams, Joanna M.; Thomas, Philip; Smart, Trevor G.

2015-01-01

GABAA receptors are important for inhibition in the CNS where neurosteroids and protein kinases are potent endogenous modulators. Acting individually, these can either enhance or depress receptor function, dependent upon the type of neurosteroid or kinase and the receptor subunit combination. However, in vivo, these modulators probably act in concert to fine-tune GABAA receptor activity and thus inhibition, although how this is achieved remains unclear. Therefore, we investigated the relationship between these modulators at synaptic-type α1β3γ2L and extrasynaptic-type α4β3δ GABAA receptors using electrophysiology. For α1β3γ2L, potentiation of GABA responses by tetrahydro-deoxycorticosterone was reduced after inhibiting protein kinase C, and enhanced following its activation, suggesting this kinase regulates neurosteroid modulation. In comparison, neurosteroid potentiation was reduced at α1β3S408A,S409Aγ2L receptors, and unaltered by PKC inhibitors or activators, indicating that phosphorylation of β3 subunits is important for regulating neurosteroid activity. To determine whether extrasynaptic-type GABAA receptors were similarly modulated, α4β3δ and α4β3S408A,S409Aδ receptors were investigated. Neurosteroid potentiation was reduced at both receptors by the kinase inhibitor staurosporine. By contrast, neurosteroid-mediated potentiation at α4S443Aβ3S408A,S409Aδ receptors was unaffected by protein kinase inhibition, strongly suggesting that phosphorylation of α4 and β3 subunits is required for regulating neurosteroid activity at extrasynaptic receptors. Western blot analyses revealed that neurosteroids increased phosphorylation of β3S408,S409 implying that a reciprocal pathway exists for neurosteroids to modulate phosphorylation of GABAA receptors. Overall, these findings provide important insight into the regulation of GABAA receptors in vivo, and into the mechanisms by which GABAergic inhibitory transmission may be simultaneously tuned by

Expression of the vascular endothelial growth factor receptor neuropilin-1 at the human embryo-maternal interface.

PubMed

Baston-Buest, Dunja M; Porn, Anne C; Schanz, Andrea; Kruessel, Jan-S; Janni, Wolfgang; Hess, Alexandra P

2011-02-01

Angiogenesis is required for successful implantation of the invading blastocyst. Vascular endothelial growth factor (VEGF) is an important key player in angiogenesis and vascular remodeling during the implantation process. Besides its well-characterized receptors VEGFR1 and VEGFR2, neuropilin-1 (NRP-1) has been shown to play an additional role in the signaling process of angiogenesis in human endometrium during the menstrual cycle, as a co-receptor of VEGF. These findings led to the hypothesis that NRP-1 might play a role in the vascular remodeling process during embryo implantation and the establishment of a pregnancy. NRP-1 mRNA transcript and protein expression were investigated in human choriocarcinoma cell lines (JEG-3, Jar and BeWo) aiming to evaluate the expression of NRP-1 in vitro, as well as in human decidua of all three trimesters of pregnancy, by western blot analysis (three samples of each trimester of pregnancy). The localization of NRP-1 in human decidua of all three trimesters of pregnancy was analyzed by immunohistochemistry (five samples of each trimester of pregnancy). NRP-1 transcript and protein were expressed in all cell lines examined. Corresponding to the analysis of human tissue by western blot and the localization by immunohistochemistry, NRP-1 protein higher expressed in samples of early pregnancy in comparison to the end of pregnancy. NRP-1 was expressed in the decidua, villi and invading cytotrophoblast of all samples investigated. This is the first study clearly showing the expression of NRP-1 in human decidua and trophoblast, suggesting an important role for the VEGF co-receptor NRP-1 besides the established receptor VEGFR2 at the embryo-maternal interface during embryonic implantation and placentation. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

Estrogen stabilizes hypoxia-inducible factor 1α through G protein-coupled estrogen receptor 1 in eutopic endometrium of endometriosis.

PubMed

Zhang, Ling; Xiong, Wenqian; Li, Na; Liu, Hengwei; He, Haitang; Du, Yu; Zhang, Zhibing; Liu, Yi

2017-02-01

To investigate whether G protein-coupled estrogen receptor (GPER, also known as GPR30 and GPER1) stabilizes hypoxia-inducible factor 1α (HIF-1α) in eutopic endometrium (EuEM) of endometriosis. Immunohistochemical analysis and experimental in vitro study. University hospital. Patients with or without endometriosis. The EuEM and normal control endometrium (CoEM) were obtained by curettage. Primary cultured endometrial stromal cells (ESCs) were treated with 17β-E 2 , G1, or G15. The EuEM and CoEM were collected for immunohistochemistry. Western blot, polymerase chain reaction, ELISA, and dual luciferase experiments were used to detect expression of GPER, HIF-1α, vascular endothelial growth factor (VEGF), and matrix metalloproteinase 9 (MMP9) in ESCs. Estradiol and G1 were used as agonists of GPER, G15 as an antagonist. Migration of ESCs and endothelial tube formation of human umbilical vein endothelial cells cultured in medium collected from ESCs were measured. Protein levels of GPER and HIF-1α were higher in EuEM than in CoEM. Protein levels of HIF-1α but not HIF-1α mRNA levels increased concurrently with GPER after E 2 and G1 treatment. Furthermore, expression and activity of VEGF and MMP9 increased under E 2 and G1 stimulation. However, these effects disappeared when GPER was blocked. G protein-coupled estrogen receptor stabilizes HIF-1α and thus promotes HIF-1α-induced VEGF and MMP9 in ESCs, which play critical roles in endometriosis. Copyright © 2016 American Society for Reproductive Medicine. All rights reserved.

Quantitative phosphoproteomics analysis reveals a key role of insulin growth factor 1 receptor (IGF1R) tyrosine kinase in human sperm capacitation.

PubMed

Wang, Jing; Qi, Lin; Huang, Shaoping; Zhou, Tao; Guo, Yueshuai; Wang, Gaigai; Guo, Xuejiang; Zhou, Zuomin; Sha, Jiahao

2015-04-01

One of the most important changes during sperm capacitation is the enhancement of tyrosine phosphorylation. However, the mechanisms of protein tyrosine phosphorylation during sperm capacitation are not well studied. We used label-free quantitative phosphoproteomics to investigate the overall phosphorylation events during sperm capacitation in humans and identified 231 sites with increased phosphorylation levels. Motif analysis using the NetworKIN algorithm revealed that the activity of tyrosine phosphorylation kinases insulin growth factor 1 receptor (IGF1R)/insulin receptor is significantly enriched among the up-regulated phosphorylation substrates during capacitation. Western blotting further confirmed inhibition of IGF1R with inhibitors GSK1904529A and NVP-AEW541, which inhibited the increase in tyrosine phosphorylation levels during sperm capacitation. Additionally, sperm hyperactivated motility was also inhibited by GSK1904529A and NVP-AEW541 but could be up-regulated by insulin growth factor 1, the ligand of IGF1R. Thus, the IGF1R-mediated tyrosine phosphorylation pathway may play important roles in the regulation of sperm capacitation in humans and could be a target for improvement in sperm functions in infertile men. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Stress-induced alterations in 5-HT1A receptor transcriptional modulators NUDR and Freud-1.

PubMed

Szewczyk, Bernadeta; Kotarska, Katarzyna; Daigle, Mireille; Misztak, Paulina; Sowa-Kucma, Magdalena; Rafalo, Anna; Curzytek, Katarzyna; Kubera, Marta; Basta-Kaim, Agnieszka; Nowak, Gabriel; Albert, Paul R

2014-11-01

The effect of stress on the mRNA and protein level of the 5-HT1A receptor and two of its key transcriptional modulators, NUDR and Freud-1, was examined in the prefrontal cortex (PFC) and hippocampus (Hp) using rodent models: olfactory bulbectomy (OB) and prenatal stress (PS) in male and female rats; chronic mild stress in male rats (CMS) and pregnancy stress. In PFC, CMS induced the most widespread changes, with significant reduction in both mRNA and protein levels of NUDR, 5-HT1A receptor and in Freud-1 mRNA; while in Hp 5-HT1A receptor and Freud-1 protein levels were also decreased. In male, but not female OB rats PFC Freud-1 and 5-HT1A receptor protein levels were reduced, while in Hp 5-HT1A receptor, Freud-1 and NUDR mRNA's but not protein were reduced. In PS rats PFC 5-HT1A receptor protein was reduced more in females than males; while in Hp Freud-1 protein was increased in females. In pregnancy stress, PFC NUDR, Freud-1 and 5-HT1A protein receptor levels were reduced, and in HP 5-HT1A receptor protein levels were also reduced; in HP only NUDR and Freud-1 mRNA levels were reduced. Overall, CMS and stress during pregnancy produced the most salient changes in 5-HT1A receptor and transcription factor expression, suggesting a primary role for altered transcription factor expression in chronic regulation of 5-HT1A receptor expression. By contrast, OB (in males) and PS (in females) produced gender-specific reductions in PFC 5-HT1A receptor protein levels, suggesting a role for post-transcriptional regulation. These and previous data suggest that chronic stress might be a key regulator of NUDR/Freud-1 gene expression.

Stress-induced alterations in 5-HT1A receptor transcriptional modulators NUDR and Freud-1

PubMed Central

Szewczyk, Bernadeta; Kotarska, Katarzyna; Daigle, Mireille; Misztak, Paulina; Sowa-Kucma, Magdalena; Rafalo, Anna; Curzytek, Katarzyna; Kubera, Marta; Basta-Kaim, Agnieszka; Nowak, Gabriel; Albert, Paul R

2015-01-01

The effect of stress on the mRNA and protein level of the 5-HT1A receptor and two of its key transcriptional modulators, NUDR and Freud-1, was examined in the prefrontal cortex (PFC) and hippocampus (Hp) using rodent models: olfactory bulbectomy (OB) and prenatal stress (PS) in male and female rats; chronic mild stress in male rats (CMS) and pregnancy stress. In PFC, CMS induced the most widespread changes, with significant reduction in both mRNA and protein levels of NUDR, 5-HT1A receptor and in Freud-1 mRNA; while in Hp 5-HT1A receptor and Freud-1 protein levels were also decreased. In male, but not female OB rats PFC Freud-1 and 5-HT1A receptor protein levels were reduced, while in Hp 5-HT1A receptor, Freud-1 and NUDR mRNA’s but not protein were reduced. In PS rats PFC 5-HT1A receptor protein was reduced more in females than males; while in Hp Freud-1 protein was increased in females. In pregnancy stress, PFC NUDR, Freud-1 and 5-HT1A protein receptor levels were reduced, and in HP 5-HT1A receptor protein levels were also reduced; in HP only NUDR and Freud-1 mRNA levels were reduced. Overall, CMS and stress during pregnancy produced the most salient changes in 5-HT1A receptor and transcription factor expression, suggesting a primary role for altered transcription factor expression in chronic regulation of 5-HT1A receptor expression. By contrast, OB (in males) and PS (in females) produced gender-specific reductions in PFC 5-HT1A receptor protein levels, suggesting a role for post-transcriptional regulation. These and previous data suggest that chronic stress might be a key regulator of NUDR/Freud-1 gene expression. PMID:24946016

Neural and receptor cochlear potentials obtained by transtympanic electrocochleography in auditory neuropathy.

PubMed

Santarelli, Rosamaria; Starr, Arnold; Michalewski, Henry J; Arslan, Edoardo

2008-05-01

Transtympanic electrocochleography (ECochG) was recorded bilaterally in children and adults with auditory neuropathy (AN) to evaluate receptor and neural generators. Test stimuli were clicks from 60 to 120dB p.e. SPL. Measures obtained from eight AN subjects were compared to 16 normally hearing children. Receptor cochlear microphonics (CMs) in AN were of normal or enhanced amplitude. Neural compound action potentials (CAPs) and receptor summating potentials (SPs) were identified in five AN ears. ECochG potentials in those ears without CAPs were of negative polarity and of normal or prolonged duration. We used adaptation to rapid stimulus rates to distinguish whether the generators of the negative potentials were of neural or receptor origin. Adaptation in controls resulted in amplitude reduction of CAP twice that of SP without affecting the duration of ECochG potentials. In seven AN ears without CAP and with prolonged negative potential, adaptation was accompanied by reduction of both amplitude and duration of the negative potential to control values consistent with neural generation. In four ears without CAP and with normal duration potentials, adaptation was without effect consistent with receptor generation. In five AN ears with CAP, there was reduction in amplitude of CAP and SP as controls but with a significant decrease in response duration. Three patterns of cochlear potentials were identified in AN: (1) presence of receptor SP without CAP consistent with pre-synaptic disorder of inner hair cells; (2) presence of both SP and CAP consistent with post-synaptic disorder of proximal auditory nerve; (3) presence of prolonged neural potentials without a CAP consistent with post-synaptic disorder of nerve terminals. Cochlear potential measures may identify pre- and post-synaptic disorders of inner hair cells and auditory nerves in AN.

Transcription factor FOXO1 promotes cell migration toward exogenous ATP via controlling P2Y1 receptor expression in lymphatic endothelial cells.

PubMed

Niimi, Kenta; Ueda, Mizuha; Fukumoto, Moe; Kohara, Misaki; Sawano, Toshinori; Tsuchihashi, Ryo; Shibata, Satoshi; Inagaki, Shinobu; Furuyama, Tatsuo

2017-08-05

Sprouting migration of lymphatic endothelial cell (LEC) is a pivotal step in lymphangiogenic process. However, its molecular mechanism remains unclear including effective migratory attractants. Meanwhile, forkhead transcription factor FOXO1 highly expresses in LEC nuclei, but its significance in LEC migratory activity has not been researched. In this study, we investigated function of FOXO1 transcription factor associated with LEC migration toward exogenous ATP which has recently gathered attentions as a cell migratory attractant. The transwell membrane assay indicated that LECs migrated toward exogenous ATP, which was impaired by FOXO1 knockdown. RT-PCR analysis showed that P2Y1, a purinergic receptor, expression was markedly reduced by FOXO1 knockdown in LECs. Moreover, P2Y1 blockage impaired LEC migration toward exogenous ATP. Western blot analysis revealed that Akt phosphorylation contributed to FOXO1-dependent LEC migration toward exogenous ATP and its blockage affected LEC migratory activity. Furthermore, luciferase reporter assay and ChIP assay suggested that FOXO1 directly bound to a conserved binding site in P2RY1 promoter and regulated its activity. These results indicated that FOXO1 serves a pivotal role in LEC migration toward exogenous ATP via direct transcriptional regulation of P2Y1 receptor. Copyright © 2017 Elsevier Inc. All rights reserved.

Antagonism of Lateral Amygdala Alpha1-Adrenergic Receptors Facilitates Fear Conditioning and Long-Term Potentiation

ERIC Educational Resources Information Center

Lazzaro, Stephanie C.; Hou, Mian; Cunha, Catarina; LeDoux, Joseph E.; Cain, Christopher K.

2010-01-01

Norepinephrine receptors have been studied in emotion, memory, and attention. However, the role of alpha1-adrenergic receptors in fear conditioning, a major model of emotional learning, is poorly understood. We examined the effect of terazosin, an alpha1-adrenergic receptor antagonist, on cued fear conditioning. Systemic or intra-lateral amygdala…

CRF1 receptor-deficiency increases cocaine reward.

PubMed

Contarino, Angelo; Kitchener, Pierre; Vallée, Monique; Papaleo, Francesco; Piazza, Pier-Vincenzo

2017-05-01

Stimulant drugs produce reward but also activate stress-responsive systems. The corticotropin-releasing factor (CRF) and the related hypothalamus-pituitary-adrenal (HPA) axis stress-responsive systems are activated by stimulant drugs. However, their role in stimulant drug-induced reward remains poorly understood. Herein, we report that CRF 1 receptor-deficient (CRF 1 -/-), but not wild-type, mice show conditioned place preference (CPP) responses to a relatively low cocaine dose (5 mg/kg, i.p.). Conversely, wild-type, but not CRF 1 -/-, mice display CPP responses to a relatively high cocaine dose (20 mg/kg, i.p.), indicating that CRF 1 receptor-deficiency alters the rewarding effects of cocaine. Acute pharmacological antagonism of the CRF 1 receptor by antalarmin also eliminates cocaine reward. Nevertheless, CRF 1 -/- mice display higher stereotypy responses to cocaine than wild-type mice. Despite the very low plasma corticosterone concentration, CRF 1 -/- mice show higher nuclear glucocorticoid receptor (GR) levels in the brain region of the hippocampus than wild-type mice. Full rescue of wild-type-like corticosterone and GR circadian rhythm and level in CRF 1 -/- mice by exogenous corticosterone does not affect CRF 1 receptor-dependent cocaine reward but induces stereotypy responses to cocaine. These results indicate a critical role for the CRF 1 receptor in cocaine reward, independently of the closely related HPA axis activity. Copyright © 2017 Elsevier Ltd. All rights reserved.

Beta receptor-mediated modulation of the late positive potential in humans.

PubMed

de Rover, Mischa; Brown, Stephen B R E; Boot, Nathalie; Hajcak, Greg; van Noorden, Martijn S; van der Wee, Nic J A; Nieuwenhuis, Sander

2012-02-01

Electrophysiological studies have identified a scalp potential, the late positive potential (LPP), which is modulated by the emotional intensity of observed stimuli. Previous work has shown that the LPP reflects the modulation of activity in extrastriate visual cortical structures, but little is known about the source of that modulation. The present study investigated whether beta-adrenergic receptors are involved in the generation of the LPP. We used a genetic individual differences approach (experiment 1) and a pharmacological manipulation (experiment 2) to test the hypothesis that the LPP is modulated by the activation of β-adrenergic receptors. In experiment 1, we found that LPP amplitude depends on allelic variation in the β1-receptor gene polymorphism. In experiment 2, we found that LPP amplitude was modulated by the β-blocker propranolol in a direction dependent on subjects' level of trait anxiety: In participants with lower trait anxiety, propranolol led to a (nonsignificant) decrease in the LPP modulation; in participants with higher trait anxiety, propranolol increased the emotion-related LPP modulation. These results provide initial support for the hypothesis that the LPP reflects the downstream effects, in visual cortical areas, of β-receptor-mediated activation of the amygdala.

The role of endogenous molecules in modulating pain through transient receptor potential vanilloid 1 (TRPV1)

PubMed Central

Morales-Lázaro, Sara L; Simon, Sidney A; Rosenbaum, Tamara

2013-01-01

Pain is a physiological response to a noxious stimulus that decreases the quality of life of those sufferring from it. Research aimed at finding new therapeutic targets for the treatment of several maladies, including pain, has led to the discovery of numerous molecular regulators of ion channels in primary afferent nociceptive neurons. Among these receptors is TRPV1 (transient receptor potential vanilloid 1), a member of the TRP family of ion channels. TRPV1 is a calcium-permeable channel, which is activated or modulated by diverse exogenous noxious stimuli such as high temperatures, changes in pH, and irritant and pungent compounds, and by selected molecules released during tissue damage and inflammatory processes. During the last decade the number of endogenous regulators of TRPV1's activity has increased to include lipids that can negatively regulate TRPV1, as is the case for cholesterol and PIP2 (phosphatidylinositol 4,5-biphosphate) while, in contrast, other lipids produced in response to tissue injury and ischaemic processes are known to positively regulate TRPV1. Among the latter, lysophosphatidic acid activates TRPV1 while amines such as N-acyl-ethanolamines and N-acyl-dopamines can sensitize or directly activate TRPV1. It has also been found that nucleotides such as ATP act as mediators of chemically induced nociception and pain and gases, such as hydrogen sulphide and nitric oxide, lead to TRPV1 activation. Finally, the products of lipoxygenases and omega-3 fatty acids among other molecules, such as divalent cations, have also been shown to endogenously regulate TRPV1 activity. Here we provide a comprehensive review of endogenous small molecules that regulate the function of TRPV1. Acting through mechanisms that lead to sensitization and desensitization of TRPV1, these molecules regulate pathways involved in pain and nociception. Understanding how these compounds modify TRPV1 activity will allow us to comprehend how some pathologies are associated with

The role of endogenous molecules in modulating pain through transient receptor potential vanilloid 1 (TRPV1).

PubMed

Morales-Lázaro, Sara L; Simon, Sidney A; Rosenbaum, Tamara

2013-07-01

Pain is a physiological response to a noxious stimulus that decreases the quality of life of those sufferring from it. Research aimed at finding new therapeutic targets for the treatment of several maladies, including pain, has led to the discovery of numerous molecular regulators of ion channels in primary afferent nociceptive neurons. Among these receptors is TRPV1 (transient receptor potential vanilloid 1), a member of the TRP family of ion channels. TRPV1 is a calcium-permeable channel, which is activated or modulated by diverse exogenous noxious stimuli such as high temperatures, changes in pH, and irritant and pungent compounds, and by selected molecules released during tissue damage and inflammatory processes. During the last decade the number of endogenous regulators of TRPV1's activity has increased to include lipids that can negatively regulate TRPV1, as is the case for cholesterol and PIP2 (phosphatidylinositol 4,5-biphosphate) while, in contrast, other lipids produced in response to tissue injury and ischaemic processes are known to positively regulate TRPV1. Among the latter, lysophosphatidic acid activates TRPV1 while amines such as N-acyl-ethanolamines and N-acyl-dopamines can sensitize or directly activate TRPV1. It has also been found that nucleotides such as ATP act as mediators of chemically induced nociception and pain and gases, such as hydrogen sulphide and nitric oxide, lead to TRPV1 activation. Finally, the products of lipoxygenases and omega-3 fatty acids among other molecules, such as divalent cations, have also been shown to endogenously regulate TRPV1 activity. Here we provide a comprehensive review of endogenous small molecules that regulate the function of TRPV1. Acting through mechanisms that lead to sensitization and desensitization of TRPV1, these molecules regulate pathways involved in pain and nociception. Understanding how these compounds modify TRPV1 activity will allow us to comprehend how some pathologies are associated with

Haploinsufficiency of the insulin-like growth factor-1 receptor enhances endothelial repair and favorably modifies angiogenic progenitor cell phenotype.

PubMed

Yuldasheva, Nadira Y; Rashid, Sheikh Tawqeer; Haywood, Natalie J; Cordell, Paul; Mughal, Romana; Viswambharan, Hema; Imrie, Helen; Sukumar, Piruthivi; Cubbon, Richard M; Aziz, Amir; Gage, Matthew; Mbonye, Kamatamu Amanda; Smith, Jessica; Galloway, Stacey; Skromna, Anna; Scott, D Julian A; Kearney, Mark T; Wheatcroft, Stephen B

2014-09-01

Defective endothelial regeneration predisposes to adverse arterial remodeling and is thought to contribute to cardiovascular disease in type 2 diabetes mellitus. We recently demonstrated that the type 1 insulin-like growth factor receptor (IGF1R) is a negative regulator of insulin sensitivity and nitric oxide bioavailability. In this report, we examined partial deletion of the IGF1R as a potential strategy to enhance endothelial repair. We assessed endothelial regeneration after wire injury in mice and abundance and function of angiogenic progenitor cells in mice with haploinsufficiency of the IGF1R (IGF1R(+/-)). Endothelial regeneration after arterial injury was accelerated in IGF1R(+/-) mice. Although the yield of angiogenic progenitor cells was lower in IGF1R(+/-) mice, these angiogenic progenitor cells displayed enhanced adhesion, increased secretion of insulin-like growth factor-1, and enhanced angiogenic capacity. To examine the relevance of IGF1R manipulation to cell-based therapy, we transfused IGF1R(+/-) bone marrow-derived CD117(+) cells into wild-type mice. IGF1R(+/-) cells accelerated endothelial regeneration after arterial injury compared with wild-type cells and did not alter atherosclerotic lesion formation. Haploinsufficiency of the IGF1R is associated with accelerated endothelial regeneration in vivo and enhanced tube forming and adhesive potential of angiogenic progenitor cells in vitro. Partial deletion of IGF1R in transfused bone marrow-derived CD117(+) cells enhanced their capacity to promote endothelial regeneration without altering atherosclerosis. Our data suggest that manipulation of the IGF1R could be exploited as novel therapeutic approach to enhance repair of the arterial wall after injury. © 2014 American Heart Association, Inc.

Novel agonistic action of mustard oil on recombinant and endogenous porcine transient receptor potential V1 (pTRPV1) channels.

PubMed

Ohta, Toshio; Imagawa, Toshiaki; Ito, Shigeo

2007-05-15

Neurogenic components play a crucial role in inflammation and nociception. Mustard oil (MO) is a pungent plant extract from mustard seed, horseradish and wasabi, the main constituent of which is allylisothiocyanate. We have characterized the action of MO on transient receptor potential V1 (TRPV1), a key receptor of signal transduction pathways in the nociceptive system, using fura-2-based [Ca(2+)](i) imaging and the patch-clamp technique in a heterologous expression system and sensory neurons. In human embryonic kidney (HEK) 293 cells expressing porcine TRPV1 (pTRPV1), MO evoked increases of [Ca(2+)](i) in a concentration-dependent manner. A high concentration of MO elicited irreversible cell swelling. Capsazepine, ruthenium red and iodoresiniferatoxin dose-dependently suppressed the MO-induced [Ca(2+)](i) increase. MO elicited outward rectified currents in pTRPV1-expressing HEK 293 cells with a reversal potential similar to that of capsaicin. [Ca(2+)](i) responses to MO were completely abolished by the removal of external Ca(2+). MO simultaneously elicited an inward current and increase of [Ca(2+)](i) in the same cells, indicating that MO promoted Ca(2+) influx through TRPV1 channels. In cultured porcine dorsal root ganglion (DRG) neurons, MO elicited a [Ca(2+)](i) increase and inward current. Among DRG neurons responding to MO, 85% were also sensitive to capsaicin. The present data indicate that MO is a novel agonist of TRPV1 channels, and suggest that the action of MO in vivo may be partly mediated via TRPV1. These results provide an insight into the TRPV1-mediated effects of MO on inflammation and hyperalgesia.

The insulin and IGF1 receptor kinase domains are functional dimers in the activated state

NASA Astrophysics Data System (ADS)

Cabail, M. Zulema; Li, Shiqing; Lemmon, Eric; Bowen, Mark E.; Hubbard, Stevan R.; Miller, W. Todd

2015-03-01

The insulin receptor (IR) and insulin-like growth factor-1 receptor (IGF1R) are highly related receptor tyrosine kinases with a disulfide-linked homodimeric architecture. Ligand binding to the receptor ectodomain triggers tyrosine autophosphorylation of the cytoplasmic domains, which stimulates catalytic activity and creates recruitment sites for downstream signalling proteins. Whether the two phosphorylated tyrosine kinase domains within the receptor dimer function independently or cooperatively to phosphorylate protein substrates is not known. Here we provide crystallographic, biophysical and biochemical evidence demonstrating that the phosphorylated kinase domains of IR and IGF1R form a specific dimeric arrangement involving an exchange of the juxtamembrane region proximal to the kinase domain. In this dimer, the active position of α-helix C in the kinase N lobe is stabilized, which promotes downstream substrate phosphorylation. These studies afford a novel strategy for the design of small-molecule IR agonists as potential therapeutic agents for type 2 diabetes.

Transmission to interneurons is via slow excitatory synaptic potentials mediated by P2Y(1) receptors during descending inhibition in guinea-pig ileum.

PubMed

Thornton, Peter D J; Gwynne, Rachel M; McMillan, Darren J; Bornstein, Joel C

2013-01-01

The nature of synaptic transmission at functionally distinct synapses in intestinal reflex pathways has not been fully identified. In this study, we investigated whether transmission between interneurons in the descending inhibitory pathway is mediated by a purine acting at P2Y receptors to produce slow excitatory synaptic potentials (EPSPs). Myenteric neurons from guinea-pig ileum in vitro were impaled with intracellular microelectrodes. Responses to distension 15 mm oral to the recording site, in a separately perfused stimulation chamber and to electrical stimulation of local nerve trunks were recorded. A subset of neurons, previously identified as nitric oxide synthase immunoreactive descending interneurons, responded to both stimuli with slow EPSPs that were reversibly abolished by a high concentration of PPADS (30 μM, P2 receptor antagonist). When added to the central chamber of a three chambered organ bath, PPADS concentration-dependently depressed transmission through that chamber of descending inhibitory reflexes, measured as inhibitory junction potentials in the circular muscle of the anal chamber. Reflexes evoked by distension in the central chamber were unaffected. A similar depression of transmission was seen when the specific P2Y(1) receptor antagonist MRS 2179 (10 μM) was in the central chamber. Blocking either nicotinic receptors (hexamethonium 200 μM) or 5-HT(3) receptors (granisetron 1 μM) together with P2 receptors had no greater effect than blocking P2 receptors alone. Slow EPSPs mediated by P2Y(1) receptors, play a primary role in transmission between descending interneurons of the inhibitory reflexes in the guinea-pig ileum. This is the first demonstration for a primary role of excitatory metabotropic receptors in physiological transmission at a functionally identified synapse.

Sigma receptors: biology and therapeutic potential.

PubMed

Guitart, Xavier; Codony, Xavier; Monroy, Xavier

2004-07-01

More than 20 years after the identification of the sigma receptors as a unique binding site in the brain and in the peripheral organs, several questions regarding this receptor are still open. Only one of the subtypes of the receptor has been cloned to date, but the endogenous ligand still remains unknown, and the possible association of the receptor with a conventional second messenger system is controversial. From the very beginning, the sigma receptors were associated with various central nervous system disorders such as schizophrenia or movement disorders. Today, after hundreds of papers dealing with the importance of sigma receptors in brain function, it is widely accepted that sigma receptors represent a new and different avenue in the possible pharmacological treatment of several brain-related disorders. In this review, what is known about the biology of the sigma receptor regarding its putative structure and its distribution in the central nervous system is summarized first. The role of sigma receptors regulating cellular functions and other neurotransmitter systems is also addressed, as well as a short overview of the possible endogenous ligands. Finally, although no specific sigma ligand has reached the market, different pharmacological approaches to the alleviation and treatment of several central nervous system disorders and deficits, including schizophrenia, pain, memory deficits, etc., are discussed, with an overview of different compounds and their potential therapeutic use.

Low serotonin1B receptor binding potential in the anterior cingulate cortex in drug-free patients with recurrent major depressive disorder.

PubMed

Tiger, Mikael; Farde, Lars; Rück, Christian; Varrone, Andrea; Forsberg, Anton; Lindefors, Nils; Halldin, Christer; Lundberg, Johan

2016-07-30

The pathophysiology of major depressive disorder (MDD) is not fully understood and the diagnosis is largely based on history and clinical examination. So far, several lines of preclinical data and a single imaging study implicate a role for the serotonin1B (5-HT1B) receptor subtype. We sought to study 5-HT1B receptor binding in brain regions of reported relevance in patients with MDD. Subjects were examined at the Karolinska Institutet PET centre using positron emission tomography (PET) and the 5-HT1B receptor selective radioligand [(11)C]AZ10419369. Ten drug-free patients with recurrent MDD and ten control subjects matched for age and sex were examined. The main outcome measure was [(11)C]AZ10419369 binding in brain regions of reported relevance in the pathophysiology of MDD. The [(11)C]AZ10419369 binding potential was significantly lower in the MDD group compared with the healthy control group in the anterior cingulate cortex (20% between-group difference), the subgenual prefrontal cortex (17% between-group difference), and in the hippocampus (32% between-group difference). The low anterior cingulate [(11)C]AZ10419369 binding potential in patients with recurrent MDD positions 5-HT1B receptor binding in this region as a putative biomarker for MDD and corroborate a role of the anterior cingulate cortex and associated areas in the pathophysiology of recurrent MDD. Copyright © 2016 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

Epidermal growth factor- and hepatocyte growth factor-receptor activity in serum-free cultures of human hepatocytes.

PubMed

Runge, D M; Runge, D; Dorko, K; Pisarov, L A; Leckel, K; Kostrubsky, V E; Thomas, D; Strom, S C; Michalopoulos, G K

1999-02-01

Serum-free primary cultures of hepatocytes are a useful tool to study factors triggering hepatocyte proliferation and regeneration. We have developed a chemically defined serum-free system that allows human hepatocyte proliferation in the presence of epidermal growth factor and hepatocyte growth factor. DNA synthesis and accumulation were determined by [3H]thymidine incorporation and fluorometry, respectively. Western blot analyses and co-immunoprecipitations were used to investigate the association of proteins involved in epidermal growth factor and hepatocyte growth factor activation and signaling: epidermal growth factor receptor, hepatocyte growth factor receptor (MET), urokinase-type plasminogen activator and its receptor, and a member of the signal transducer and activator of transcription family, STAT-3. Primary human hepatocytes proliferated under serum-free conditions in a chemically defined medium for up to 12 days. Epidermal growth factor-receptor and MET were present and functional, decreasing over time. MET, urokinase-type plasminogen activator and urokinase-type plasminogen activator receptor co-precipitated to varying degrees during the culture period. STAT-3 co-precipitated with epidermal growth factor-receptor and MET to varying degrees. Proliferation of human hepatocytes can improve by modification of a chemically defined medium originally used for rat hepatocyte cultures. In these long-term cultures of human hepatocytes, hepatocyte growth factor and epidermal growth factor can stimulate growth and differentiation by interacting with their receptors and initiating downstream signaling. This involves complex formation of the receptors with other plasma membrane components for MET (urokinase-type plasminogen activator in context of its receptor) and activation of STAT-3 for both receptors.

Vortioxetine dose-dependently reverses 5-HT depletion-induced deficits in spatial working and object recognition memory: a potential role for 5-HT1A receptor agonism and 5-HT3 receptor antagonism.

PubMed

du Jardin, Kristian Gaarn; Jensen, Jesper Bornø; Sanchez, Connie; Pehrson, Alan L

2014-01-01

We previously reported that the investigational multimodal antidepressant, vortioxetine, reversed 5-HT depletion-induced memory deficits while escitalopram and duloxetine did not. The present report studied the effects of vortioxetine and the potential impact of its 5-HT1A receptor agonist and 5-HT3 receptor antagonist properties on 5-HT depletion-induced memory deficits. Recognition and spatial working memory were assessed in the object recognition (OR) and Y-maze spontaneous alternation (SA) tests, respectively. 5-HT depletion was induced in female Long-Evans rats using 4-cholro-DL-phenylalanine methyl ester HCl (PCPA) and receptor occupancies were determined by ex vivo autoradiography. Rats were acutely dosed with vortioxetine, ondansetron (5-HT3 receptor antagonist) or flesinoxan (5-HT1A receptor agonist). The effects of chronic vortioxetine administration on 5-HT depletion-induced memory deficits were also assessed. 5-HT depletion reliably impaired memory performance in both the tests. Vortioxetine reversed PCPA-induced memory deficits dose-dependently with a minimal effective dose (MED) ≤0.1mg/kg (∼80% 5-HT3 receptor occupancy; OR) and ≤3.0mg/kg (5-HT1A, 5-HT1B, 5-HT3 receptor occupancy: ∼15%, 60%, 95%) in SA. Ondansetron exhibited a MED ≤3.0μg/kg (∼25% 5-HT3 receptor occupancy; OR), but was inactive in the SA test. Flesinoxan had a MED ≤1.0mg/kg (∼25% 5-HT1A receptor occupancy; SA); only 1.0mg/kg ameliorated deficits in the NOR. Chronic p.o. vortioxetine administration significantly improved memory performance in OR and occupied 95%, 66%, and 9.5% of 5-HT3, 5-HT1B, and 5-HT1A receptors, respectively. Vortioxetine's effects on SA performance may involve 5-HT1A receptor agonism, but not 5-HT3 receptor antagonism, whereas the effects on OR performance may involve 5-HT3 receptor antagonism and 5-HT1A receptor agonism. Copyright © 2013 Elsevier B.V. and ECNP. All rights reserved.