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  1. Novel toll-like receptor 9 agonist induces epidermal growth factor receptor (EGFR) inhibition and synergistic antitumor activity with EGFR inhibitors.

    PubMed

    Damiano, Vincenzo; Caputo, Rosa; Bianco, Roberto; D'Armiento, Francesco P; Leonardi, Antonio; De Placido, Sabino; Bianco, A Raffaele; Agrawal, Sudhir; Ciardiello, Fortunato; Tortora, Giampaolo

    2006-01-15

    Immunostimulating Toll-like receptor 9 (TLR9) agonists cause antitumor activity interfering also with cancer proliferation and angiogenesis by mechanisms still incompletely understood. We hypothesized that modified TLR9 agonists could impair epidermal growth factor receptor (EGFR) signaling and, by this means, greatly enhance EGFR inhibitors effect, acting on both the receptor targeting and the immunologic arm. We used a novel second-generation, modified, immunomodulatory TLR9 agonist (IMO), alone and in combination with the anti-EGFR monoclonal antibody cetuximab or tyrosine kinase inhibitor gefitinib, on the growth of GEO and cetuximab-resistant derivatives GEO-CR colon cancer xenografts. We have also evaluated the expression of several proteins critical for cell proliferation, apoptosis, and angiogenesis, including EGFR, mitogen-activated protein kinase, Akt, bcl-2, cyclooxygenase-2, vascular endothelial growth factor, and nuclear factor-kappaB. IMO inhibited GEO growth and signaling by EGFR and the other proteins critical for cell proliferation and angiogenesis. IMO plus the anti-EGFR antibody cetuximab synergistically inhibited tumor growth, signaling proteins, and microvessel formation. EGFR signaling inhibition by IMO is relevant because IMO cooperated also with EGFR tyrosine kinase inhibitor gefitinib in GEO tumors, while it was inactive against GEO-CR xenografts. On the other hand, IMO boosted the non-EGFR-dependent cetuximab activity, causing a cooperative antitumor effect in GEO-CR cells. Finally, combination of IMO, cetuximab and chemotherapeutic irinotecan eradicated the tumors in 90% of mice. IMO interferes with EGFR-related signaling and angiogenesis and has a synergistic antitumor effect with EGFR inhibitors, especially with cetuximab, boosting both the EGFR dependent and independent activity of this agent. Moreover, this therapeutic strategy could be translated in patients affected by colorectal cancer.

  2. Toll-Like Receptor 9 Agonists for Cancer Therapy

    PubMed Central

    Melisi, Davide; Frizziero, Melissa; Tamburrino, Anna; Zanotto, Marco; Carbone, Carmine; Piro, Geny; Tortora, Giampaolo

    2014-01-01

    The immune system has acquired increasing importance as a key player in cancer maintenance and growth. Thus, modulating anti-tumor immune mediators has become an attractive strategy for cancer treatment. Toll-like receptors (TLRs) have gradually emerged as potential targets of newer immunotherapies. TLR-9 is preferentially expressed on endosome membranes of B-cells and plasmacytoid dendritic cells (pDC) and is known for its ability to stimulate specific immune reactions through the activation of inflammation-like innate responses. Several synthetic CpG oligonucleotides (ODNs) have been developed as TLR-9 agonists with the aim of enhancing cancer immune surveillance. In many preclinical models, CpG ODNs were found to suppress tumor growth and proliferation both in monotherapy and in addition to chemotherapies or target therapies. TLR-9 agonists have been also tested in several clinical trials in patients with solid tumors. These agents showed good tolerability and usually met activity endpoints in early phase trials. However, they have not yet been demonstrated to significantly impact survival, neither as single agent treatments, nor in combination with chemotherapies or cancer vaccines. Further investigations in larger prospective studies are required. PMID:28548068

  3. Toll-Like Receptor 9 Agonists for Cancer Therapy.

    PubMed

    Melisi, Davide; Frizziero, Melissa; Tamburrino, Anna; Zanotto, Marco; Carbone, Carmine; Piro, Geny; Tortora, Giampaolo

    2014-08-04

    The immune system has acquired increasing importance as a key player in cancer maintenance and growth. Thus, modulating anti-tumor immune mediators has become an attractive strategy for cancer treatment. Toll-like receptors (TLRs) have gradually emerged as potential targets of newer immunotherapies. TLR-9 is preferentially expressed on endosome membranes of B-cells and plasmacytoid dendritic cells (pDC) and is known for its ability to stimulate specific immune reactions through the activation of inflammation-like innate responses. Several synthetic CpG oligonucleotides (ODNs) have been developed as TLR-9 agonists with the aim of enhancing cancer immune surveillance. In many preclinical models, CpG ODNs were found to suppress tumor growth and proliferation both in monotherapy and in addition to chemotherapies or target therapies. TLR-9 agonists have been also tested in several clinical trials in patients with solid tumors. These agents showed good tolerability and usually met activity endpoints in early phase trials. However, they have not yet been demonstrated to significantly impact survival, neither as single agent treatments, nor in combination with chemotherapies or cancer vaccines. Further investigations in larger prospective studies are required.

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

    PubMed

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

    2016-05-01

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

  5. Methamphetamine Inhibits Toll-Like Receptor 9-Mediated Anti-HIV Activity in Macrophages

    PubMed Central

    Cen, Ping; Ye, Li; Su, Qi-Jian; Wang, Xu; Li, Jie-Liang; Lin, Xin-Qin

    2013-01-01

    Abstract Toll-like receptor 9 (TLR9) is one of the key sensors that recognize viral infection/replication in the host cells. Studies have demonstrated that methamphetamine (METH) dysregulated host cell innate immunity and facilitated HIV infection of macrophages. In this study, we present new evidence that METH suppressed TLR9-mediated anti-HIV activity in macrophages. Activation of TLR9 by its agonist CpG-ODN 2216 inhibits HIV replication, which was demonstrated by increased expression of TLR9, interferon (IFN)-α, IFN regulatory factor-7 (IRF-7), myeloid differentiation factor 88 (MyD88), and myxovirus resistance gene A (MxA) in macrophages. However, METH treatment of macrophages greatly compromised the TLR9 signaling-mediated anti-HIV effect and inhibited the expression of TLR9 downstream signaling factors. Dopamine D1 receptor (D1R) antagonists (SCH23390) could block METH-mediated inhibition of anti-HIV activity of TLR9 signaling. Investigation of the underlying mechanisms of the METH action showed that METH treatment selectively down-regulated the expression of TLR9 on macrophages, whereas it had little effect on the expression of other TLRs. Collectively, our results provide further evidence that METH suppresses host cell innate immunity against HIV infection by down-regulating TLR9 expression and its signaling-mediated antiviral effect in macrophages. PMID:23751096

  6. Toll-like receptor 9 agonist IMO cooperates with cetuximab in K-ras mutant colorectal and pancreatic cancers.

    PubMed

    Rosa, Roberta; Melisi, Davide; Damiano, Vincenzo; Bianco, Roberto; Garofalo, Sonia; Gelardi, Teresa; Agrawal, Sudhir; Di Nicolantonio, Federica; Scarpa, Aldo; Bardelli, Alberto; Tortora, Giampaolo

    2011-10-15

    K-Ras somatic mutations are a strong predictive biomarker for resistance to epidermal growth factor receptor (EGFR) inhibitors in patients with colorectal and pancreatic cancer. We previously showed that the novel Toll-like receptor 9 (TLR9) agonist immunomodulatory oligonucleotide (IMO) has a strong in vivo activity in colorectal cancer models by interfering with EGFR-related signaling and synergizing with the anti-EGFR monoclonal antibody cetuximab. In the present study, we investigated, both in vitro and in vivo, the antitumor effect of IMO alone or in combination with cetuximab in subcutaneous colon and orthotopic pancreatic cancer models harboring K-Ras mutations and resistance to EGFR inhibitors. We showed that IMO was able to significantly restore the sensitivity of K-Ras mutant cancer cells to cetuximab, producing a marked inhibition of cell survival and a complete suppression of mitogen-activated protein kinase phosphorylation, when used in combination with cetuximab. IMO interfered with EGFR-dependent signaling, modulating the functional interaction between TLR9 and EGFR. In vivo, IMO plus cetuximab combination caused a potent and long-lasting cooperative antitumor activity in LS174T colorectal cancer and in orthotopic AsPC1 pancreatic cancer. The capability of IMO to restore cetuximab sensitivity was further confirmed by using K-Ras mutant colorectal cancer cell models obtained through homologous recombination technology. We showed that IMO markedly inhibits growth of K-Ras mutant colon and pancreatic cancers in vitro and in nude mice and cooperates with cetuximab via multiple mechanisms of action. Therefore, we propose IMO plus cetuximab as a therapeutic strategy for K-Ras wild-type as well for K-Ras mutant, cetuximab-resistant colorectal and pancreatic cancers. ©2011 AACR.

  7. A novel toll-like receptor 9 agonist cooperates with trastuzumab in trastuzumab-resistant breast tumors through multiple mechanisms of action.

    PubMed

    Damiano, Vincenzo; Garofalo, Sonia; Rosa, Roberta; Bianco, Roberto; Caputo, Rosa; Gelardi, Teresa; Merola, Gerardina; Racioppi, Luigi; Garbi, Corrado; Kandimalla, Ekambar R; Agrawal, Sudhir; Tortora, Giampaolo

    2009-11-15

    Resistance to anti-HER2 monoclonal antibody trastuzumab is a relevant issue in breast cancer patients. Among the mechanisms implicated in trastuzumab resistance, increasing evidence supports a role of tumor microenvironment. We previously found that a novel toll-like receptor 9 agonist, referred to as immune modulatory oligonucleotide (IMO) and currently under clinical investigation, acts through epidermal growth factor receptor (EGFR) and shows direct antiangiogenic effects by cooperating with anti-EGFR or anti-VEGF drugs, thus interfering with cancer cells and microenvironment. In this study, we used KPL-4 and JIMT-1 trastuzumab-resistant breast cancer cells to evaluate the combination IMO plus trastuzumab as a therapeutic option for trastuzumab-resistant breast cancers. IMO inhibits KPL-4 and JIMT-1 xenografts growth and potentiates trastuzumab antitumor effect, with complete suppression of tumor growth, potent enhancement of trastuzumab-mediated antibody-dependent cell-mediated cytotoxicity, and strong inhibition of EGFR/HER2-related signaling. In KPL-4 xenografts, IMO alone interferes with HER signal transduction, whereas trastuzumab is ineffective. IMO induces an HER-dependent signal inhibition also in vitro by modulating a functional interaction between toll-like receptor 9 and HER receptors occurring at membrane level. Finally, IMO plus trastuzumab produces a cooperative antiangiogenic effect related to suppression of endothelial HER-related signaling. We showed a cooperative effect of IMO plus trastuzumab in trastuzumab-resistant breast cancers due to IMO direct antitumor and antiangiogenic activity and antibody-dependent cell-mediated cytotoxicity enhancement. Moreover, we provided first evidence of a toll-like receptor 9/HER interaction at membrane level as novel mechanism of action. Altogether, we propose IMO plus trastuzumab as an effective strategy in trastuzumab-resistant breast cancers.

  8. CPG-7909 (PF-3512676, ProMune): toll-like receptor-9 agonist in cancer therapy.

    PubMed

    Murad, Yanal M; Clay, Timothy M; Lyerly, H Kim; Morse, Michael A

    2007-08-01

    Stimulation of toll-like receptor (TLR)9 activates human plasmacytoid dendritic cells and B cells, and induces potent innate immune responses in preclinical tumor models and in patients. CpG oligodeoxynucleotides (ODNs) are TLR9 agonists that show promising results as vaccine adjuvants and in the treatment of cancers, infections, asthma and allergy. PF-3512676 (ProMune) was developed as a TLR9 agonist for the treatment of cancer as monotherapy and as an adjuvant in combination with chemo- and immunotherapy. Phase I and II trials have tested this drug in several hematopoietic and solid tumors. Pfizer has initiated Phase III trials to test PF-3512676 in combination with standard chemotherapy for non-small-cell lung cancer.

  9. Toll-like receptor 9 agonist IMO cooperates with everolimus in renal cell carcinoma by interfering with tumour growth and angiogenesis.

    PubMed

    Damiano, V; Rosa, R; Formisano, L; Nappi, L; Gelardi, T; Marciano, R; Cozzolino, I; Troncone, G; Agrawal, S; Veneziani, B M; De Placido, S; Bianco, R; Tortora, G

    2013-04-30

    Targeting the mammalian target of rapamycin by everolimus is a successful approach for renal cell carcinoma (RCC) therapy. The Toll-like receptor 9 agonist immune modulatory oligonucleotide (IMO) exhibits direct antitumour and antiangiogenic activity and cooperates with both epidermal growth factor receptor (EGFR) and vascular endothelial growth factor (VEGF) inhibitors. We tested the combination of IMO and everolimus on models of human RCC with different Von-Hippel Lindau (VHL) gene status, both in vitro and in nude mice. We studied their direct antiangiogenic effects on human umbilical vein endothelial cells. Both IMO and everolimus inhibited in vitro growth and survival of RCC cell lines, and their combination produced a synergistic inhibitory effect. Moreover, everolimus plus IMO interfered with EGFR-dependent signaling and reduced VEGF secretion in both VHL wild-type and mutant cells. In RCC tumour xenografts, IMO plus everolimus caused a potent and long-lasting cooperative antitumour activity, with reduction of tumour growth, prolongation of mice survival and inhibition of signal transduction. Furthermore, IMO and everolimus impaired the main endothelial cell functions. A combined treatment with everolimus and IMO is effective in VHL wild-type and mutant models of RCC by interfering with tumour growth and angiogenesis, thus representing a potentially effective, rationale-based combination to be translated in the clinical setting.

  10. Administration of a Toll-like receptor 9 agonist decreases the proviral reservoir in virologically suppressed HIV-infected patients.

    PubMed

    Winckelmann, Anni A; Munk-Petersen, Lærke V; Rasmussen, Thomas A; Melchjorsen, Jesper; Hjelholt, Thomas J; Montefiori, David; Østergaard, Lars; Søgaard, Ole S; Tolstrup, Martin

    2013-01-01

    Toll-like receptor (TLR) agonists can reactivate HIV from latently infected cells in vitro. We aimed to investigate the TLR-9 agonist, CPG 7909's in vivo effect on the proviral HIV reservoir and HIV-specific immunity. This was a post-hoc analysis of a double-blind randomized controlled vaccine trial. HIV-infected adults were randomized 1:1 to receive pneumococcal vaccines with or without 1 mg CPG 7909 as adjuvant at 0, 3 and 9 months. In patients on suppressive antiretroviral therapy we quantified proviral DNA at 0, 3, 4, 9, and 10 months (31 subjects in the CPG group and 37 in the placebo-adjuvant group). Furthermore, we measured HIV-specific antibodies, characterized T cell phenotypes and HIV-specific T cell immunity. We observed a mean reduction in proviral DNA in the CPG group of 12.6% (95% CI: -23.6-0.0) following each immunization whereas proviral DNA in the placebo-adjuvant group remained largely unchanged (6.7% increase; 95% CI: -4.2-19.0 after each immunization, p = 0.02). Among participants with additional cryo-preserved PBMCs, HIV-specific CD8+ T cell immunity as indicated by increased expression of degranulation marker CD107a and macrophage inflammatory protein 1β (MIP1β) tended to be up-regulated following immunization with CPG 7909 compared with placebo as adjuvant. Further, increasing proportion of HIV-specific CD107a and MIP1β-expressing CD8+ T cells were strongly correlated with decreasing proviral load. No changes were observed in T cell phenotype distribution, HIV-specific CD4+ T cell immunity, or HIV-specific antibodies. TLR9-adjuvanted pneumococcal vaccination decreased proviral load. Reductions in proviral load correlated with increasing levels of HIV specific CD8+ T cells. Further investigation into the potential effect of TLR9 agonists on HIV latency is warranted.

  11. Administration of a Toll-Like Receptor 9 Agonist Decreases the Proviral Reservoir in Virologically Suppressed HIV-Infected Patients

    PubMed Central

    Winckelmann, Anni A.; Munk-Petersen, Lærke V.; Rasmussen, Thomas A.; Melchjorsen, Jesper; Hjelholt, Thomas J.; Montefiori, David; Østergaard, Lars; Søgaard, Ole S.; Tolstrup, Martin

    2013-01-01

    Toll-like receptor (TLR) agonists can reactivate HIV from latently infected cells in vitro. We aimed to investigate the TLR-9 agonist, CPG 7909's in vivo effect on the proviral HIV reservoir and HIV-specific immunity. This was a post-hoc analysis of a double-blind randomized controlled vaccine trial. HIV-infected adults were randomized 1∶1 to receive pneumococcal vaccines with or without 1 mg CPG 7909 as adjuvant at 0, 3 and 9 months. In patients on suppressive antiretroviral therapy we quantified proviral DNA at 0, 3, 4, 9, and 10 months (31 subjects in the CPG group and 37 in the placebo-adjuvant group). Furthermore, we measured HIV-specific antibodies, characterized T cell phenotypes and HIV-specific T cell immunity. We observed a mean reduction in proviral DNA in the CPG group of 12.6% (95% CI: −23.6–0.0) following each immunization whereas proviral DNA in the placebo-adjuvant group remained largely unchanged (6.7% increase; 95% CI: −4.2–19.0 after each immunization, p = 0.02). Among participants with additional cryo-preserved PBMCs, HIV-specific CD8+ T cell immunity as indicated by increased expression of degranulation marker CD107a and macrophage inflammatory protein 1β (MIP1β) tended to be up-regulated following immunization with CPG 7909 compared with placebo as adjuvant. Further, increasing proportion of HIV-specific CD107a and MIP1β-expressing CD8+ T cells were strongly correlated with decreasing proviral load. No changes were observed in T cell phenotype distribution, HIV-specific CD4+ T cell immunity, or HIV-specific antibodies. TLR9-adjuvanted pneumococcal vaccination decreased proviral load. Reductions in proviral load correlated with increasing levels of HIV specific CD8+ T cells. Further investigation into the potential effect of TLR9 agonists on HIV latency is warranted. PMID:23637967

  12. Clinical Effects of a Topically Applied Toll-like Receptor 9 Agonist in Active Moderate-to-Severe Ulcerative Colitis

    PubMed Central

    Atreya, Raja; Bloom, Stuart; Scaldaferri, Franco; Gerardi, Viviana; Admyre, Charlotte; Karlsson, Åsa; Knittel, Thomas; Kowalski, Jan; Lukas, Milan; Löfberg, Robert; Nancey, Stephane; Petryka, Robert; Rydzewska, Grazyna; Schnabel, Robert; Seidler, Ursula; Neurath, Markus F.

    2016-01-01

    Background and Aims: Toll-like receptors [TLRs] are potential drug targets for immunomodulation. We determined the safety and efficacy of the TLR-9 agonist DNA-based immunomodulatory sequence 0150 [DIMS0150] in ulcerative colitis [UC] patients refractory to standard therapy. Methods: In this randomized, double-blind, placebo-controlled trial, 131 patients with moderate-to-severe active UC were randomized to receive two single doses of the oligonucleotide DIMS0150 [30 mg] or placebo administered topically during lower GI endoscopy at baseline and Week 4. The primary endpoint was clinical remission, defined as Clinical Activity Index [CAI] ≤4, at Week 12. Secondary endpoints included mucosal healing and symptomatic remission of key patient-reported outcomes [absence of blood in stool and weekly stool frequency <35]. Results: There was no statistical significant difference between the groups in the induction of clinical remission at Week 12, with 44.4% in the DIMS0150 group vs. 46.5% in the placebo group. However, the proportion of patients who achieved symptomatic remission was 32.1% in the DIMS0150 group vs. 14.0% in the placebo group at Week 4 [p = 0.020], and 44.4% vs. 27.9% at Week 8 [p = 0.061]. More patients on DIMS0150 compared with those on placebo had mucosal healing [34.6% vs. 18.6%; p = 0.09] and histological improvement regarding the Geboes score [30.9% vs. 9.3%; p = 0.0073] at Week 4. Significantly more patients on DIMS0150 were in clinical remission with mucosal healing at Week 4: 21% vs. 4.7% in the placebo group [p = 0.02]. DIMS0150 was well tolerated, and no safety signals compared with placebo were evident. Conclusions: Therapy with the topically applied TLR-9 agonist DIMS0150 is a promising and well-tolerated novel therapeutic option for treatment-refractory, chronic active UC patients, warranting further clinical trials. PMID:27208386

  13. The activation of liver X receptors inhibits toll-like receptor-9-induced foam cell formation.

    PubMed

    Sorrentino, Rosalinda; Morello, Silvana; Chen, Shuang; Bonavita, Eduardo; Pinto, Aldo

    2010-04-01

    Toll-like receptors (TLRs) are related to foam cell formation (FCF), key event in the establishment/progression of atherosclerosis. The activation of TLR2 and TLR4 can increase FCF. The aim of this study was to evaluate the role of TLR9 in FCF. Murine macrophages were treated with CpG-ODN, TLR9 agonist, and oxidized particles of LDL (Paz-PC) and FCF was analyzed by means of Oil Red O staining. The administration of CpG-ODN plus Paz-PC onto macrophages increased the amount of lipid droplets, correlated to increased levels of tumor necrosis factor (TNF)-alpha, IFNbeta, and IP-10. The underlying mechanism by which TLR9 ligation influenced Paz-PC in the FCF was NF-kappaB- and IRF7-dependent, as observed by higher levels of phosphorylated IkappaBalpha, increased nuclear translocation of the p65 subunit, lower levels of the total IKKalpha protein and higher release of interferon-dependent cytokines, such as IP-10. Liver X receptors (LXRs) regulate lipid cellular transport and negatively modulate TLR-dependent signaling pathways. Indeed, the addition of GW3965, synthetic LXRs agonist, significantly reduced FCF after CpG-ODN plus Paz-PC stimulation. In this condition, we observed decreased levels of the nuclear translocation of the p65 subunit, related to the higher presence of LXRalpha into the nucleus. TNF-alpha, IP-10, and IFNbeta levels were reduced by the administration of GW3965 following CpG-ODN and Paz-PC treatment. In conclusion, the activation of TLR9 facilitates the formation of foam cells in an NF-kappaB- and IRF7-dependent manner, countered by the activation of LXRs. This study further support LXRs as potential anti-atherosclerotic target.

  14. High-dose irradiation in combination with toll-like receptor 9 agonist CpG oligodeoxynucleotide 7909 downregulates PD-L1 expression via the NF-κB signaling pathway in non-small cell lung cancer cells

    PubMed Central

    Chen, Xue; Zhang, Qi; Luo, Youjun; Gao, Caixia; Zhuang, Xibing; Xu, Guoxiong; Qiao, Tiankui

    2016-01-01

    Objectives Irradiation resistance appears as local recurrence and distant metastasis in advanced stages of non-small cell lung cancer (NSCLC). High-dose irradiation combined with immunotherapy improved overall survival and local control of NSCLC. This study explored the underlying molecular mechanism by which the effect of high-dose irradiation plus toll-like receptor 9 (TLR9) agonist CpG oligodeoxynucleotide (CpG ODN) 7909 on NSCLC. Materials and methods NSCLC H460 cells were exposed to constant high-dose irradiation (6.37 Gy) in irradiation (IR) group and the irradiation plus CpG group. Gene expression was assessed using quantitative reverse transcriptase-polymerase chain reaction and Western blot. Knockdown of nuclear factor kappa B (NF-κB) p65 expression was conducted using p65 siRNA. Results Expression of programmed death-ligand 1 (PD-L1) mRNA was significantly decreased in IR combined with CpG ODN 7909 group compared with the control or IR-alone groups (P<0.05). TLR9 expression was also obviously increased in the combination group compared with the control (P<0.05). Moreover, expression of NF-κB p65 was apparently reduced in the combination group compared with the control (P<0.05). However, expression of PD-L1 was significantly decreased after knockdown of p65 in IR group (P<0.05), but increased in the combination group (P<0.05) and slightly increased in CpG ODN-alone group (P<0.05), which was opposite to that without p65 knockdown group. Conclusion This study demonstrated that radiotherapy combined with CpG ODN 7909 was able to downregulate PD-L1 expression through inhibition via the NF-κB signaling pathway. PMID:27799798

  15. Polygonum cuspidatum and Its Active Components Inhibit Replication of the Influenza Virus through Toll-Like Receptor 9-Induced Interferon Beta Expression

    PubMed Central

    Lin, Chao-jen; Lin, Hui-Ju; Chen, Ter-Hsin; Hsu, Yu-An; Liu, Chin-San; Hwang, Guang-Yuh; Wan, Lei

    2015-01-01

    Influenza virus infection is a global public health issue. The effectiveness of antiviral therapies for influenza has been limited by the emergence of drug-resistant viral strains. Therefore, there is an urgent need to identify novel antiviral therapies. Here we tested the effects of 300 traditional Chinese medicines on the replication of various influenza virus strains in a lung cell line, A549, using an influenza-specific luciferase reporter assay. Of the traditional medicines tested, Polygonum cuspidatum (PC) and its active components, resveratrol and emodin, were found to attenuate influenza viral replication in A549 cells. Furthermore, they preferentially inhibited the replication of influenza A virus, including clinical strains isolated in 2009 and 2011 in Taiwan and the laboratory strain A/WSN/33 (H1N1). In addition to inhibiting the expression of hemagglutinin and neuraminidase, PC, emodin, and resveratrol also increased the expression of interferon beta (IFN-β) through Toll-like receptor 9 (TLR9). Moreover, the anti-viral activity of IFN-β or resveratrol was reduced when the A549 cells were treated with neutralizing anti-IFN-β antibodies or a TLR9 inhibitor, suggesting that IFN-β likely acts synergistically with resveratrol to inhibit H1N1 replication. This potential antiviral mechanism, involving direct inhibition of virus replication and simultaneous activation of the host immune response, has not been previously described for a single antiviral molecule. In conclusion, our data support the use of PC, resveratrol or emodin for inhibiting influenza virus replication directly and via TLR-9–induced IFN-β production. PMID:25658356

  16. Angiotensin-converting Enzyme Inhibition Down-regulates the Pro-atherogenic Chemokine Receptor 9 (CCR9)-Chemokine Ligand 25 (CCL25) Axis*

    PubMed Central

    Abd Alla, Joshua; Langer, Andreas; Elzahwy, Sherif S.; Arman-Kalcek, Gökhan; Streichert, Thomas; Quitterer, Ursula

    2010-01-01

    Many experimental and clinical studies suggest a relationship between enhanced angiotensin II release by the angiotensin-converting enzyme (ACE) and the pathophysiology of atherosclerosis. The atherosclerosis-enhancing effects of angiotensin II are complex and incompletely understood. To identify anti-atherogenic target genes, we performed microarray gene expression profiling of the aorta during atherosclerosis prevention with the ACE inhibitor, captopril. Atherosclerosis-prone apolipoprotein E (apoE)-deficient mice were used as a model to decipher susceptible genes regulated during atherosclerosis prevention with captopril. Microarray gene expression profiling and immunohistology revealed that captopril treatment for 7 months strongly decreased the recruitment of pro-atherogenic immune cells into the aorta. Captopril-mediated inhibition of plaque-infiltrating immune cells involved down-regulation of the C-C chemokine receptor 9 (CCR9). Reduced cell migration correlated with decreased numbers of aorta-resident cells expressing the CCR9-specific chemoattractant factor, chemokine ligand 25 (CCL25). The CCL25-CCR9 axis was pro-atherogenic, because inhibition of CCR9 by RNA interference in hematopoietic progenitors of apoE-deficient mice significantly retarded the development of atherosclerosis. Analysis of coronary artery biopsy specimens of patients with coronary artery atherosclerosis undergoing bypass surgery also showed strong infiltrates of CCR9-positive cells in atherosclerotic lesions. Thus, the C-C chemokine receptor, CCR9, exerts a significant role in atherosclerosis. PMID:20504763

  17. UV Radiation Activates Toll-Like Receptor 9 Expression in Primary Human Keratinocytes, an Event Inhibited by Human Papillomavirus 38 E6 and E7 Oncoproteins.

    PubMed

    Pacini, Laura; Ceraolo, Maria Grazia; Venuti, Assunta; Melita, Giusi; Hasan, Uzma A; Accardi, Rosita; Tommasino, Massimo

    2017-10-01

    Several lines of evidence indicate that cutaneous human papillomavirus (HPV) types belonging to the beta genus of the HPV phylogenetic tree synergize with UV radiation in the development of skin cancer. Accordingly, the E6 and E7 oncoproteins from some beta HPV types are able to deregulate pathways related to immune response and cellular transformation. Toll-like receptor 9 (TLR9), in addition to playing a role in innate immunity, has been shown to be involved in the cellular stress response. Using primary human keratinocytes as experimental models, we have shown that UV irradiation (and other cellular stresses) activates TLR9 expression. This event is closely linked to p53 activation. Silencing the expression of p53 or deleting its encoding gene affected the activation of TLR9 expression after UV irradiation. Using various strategies, we have also shown that the transcription factors p53 and c-Jun are recruited onto a specific region of the TLR9 promoter after UV irradiation. Importantly, the E6 and E7 oncoproteins from beta HPV38, by inducing the accumulation of the p53 antagonist ΔNp73α, prevent the UV-mediated recruitment of these transcription factors onto the TLR9 promoter, with subsequent impairment of TLR9 gene expression. This study provides new insight into the mechanism that mediates TLR9 upregulation in response to cellular stresses. In addition, we show that HPV38 E6 and E7 are able to interfere with this mechanism, providing another explanation for the possible cooperation of beta HPV types with UV radiation in skin carcinogenesis.IMPORTANCE Beta HPV types have been suggested to act as cofactors in UV-induced skin carcinogenesis by altering several cellular mechanisms activated by UV radiation. We show that the expression of TLR9, a sensor of damage-associated molecular patterns produced during cellular stress, is activated by UV radiation in primary human keratinocytes (PHKs). Two transcription factors known to be activated by UV radiation, p53 and

  18. Cardamonin inhibits agonist-induced vascular contractility via Rho-kinase and MEK inhibition.

    PubMed

    Je, Hyun Dong; Jeong, Ji Hoon

    2016-01-01

    The present study was undertaken to investigate the influence of cardamonin on vascular smooth muscle contractility and to determine the mechanism(s) involved. Denuded aortic rings from male rats were used and isometric contractions were recorded and combined with molecular experiments. Cardamonin significantly relaxed fluoride-, phenylephrine-, and phorbol ester-induced vascular contractions, suggesting that it has an anti-hypertensive effect on agonist-induced vascular contraction regardless of endothelial nitric oxide synthesis. Furthermore, cardamonin significantly inhibited the fluoride-induced increase in pMYPT1 level and phenylephrine-induced increase in pERK1/2 level, suggesting inhibition of Rho-kinase and MEK activity and subsequent phosphorylation of MYPT1 and ERK1/2. This study provides evidence that the relaxing effect of cardamonin on agonist-induced vascular contraction regardless of endothelial function involves inhibition of Rho-kinase and MEK activity.

  19. Use-dependent inhibition of P2X3 receptors by nanomolar agonist.

    PubMed

    Pratt, Emily B; Brink, Thaddeus S; Bergson, Pamela; Voigt, Mark M; Cook, Sean P

    2005-08-10

    P2X3 receptors desensitize within 100 ms of channel activation, yet recovery from desensitization requires several minutes. The molecular basis for this slow rate of recovery is unknown. We designed experiments to test the hypothesis that this slow recovery is attributable to the high affinity (< 1 nM) of desensitized P2X3 receptors for agonist. We found that agonist binding to the desensitized state provided a mechanism for potent inhibition of P2X3 current. Sustained applications of 0.5 nM ATP inhibited > 50% of current to repetitive applications of P2X3 agonist. Inhibition occurred at 1000-fold lower agonist concentrations than required for channel activation and showed strong use dependence. No inhibition occurred without previous activation and desensitization. Our data are consistent with a model whereby inhibition of P2X3 by nanomolar [agonist] occurs by the rebinding of agonist to desensitized channels before recovery from desensitization. For several ATP analogs, the concentration required to inhibit P2X3 current inversely correlated with the rate of recovery from desensitization. This indicates that the affinity of the desensitized state and recovery rate primarily depend on the rate of agonist unbinding. Consistent with this hypothesis, unbinding of [32P]ATP from desensitized P2X3 receptors mirrored the rate of recovery from desensitization. As expected, disruption of agonist binding by site-directed mutagenesis increased the IC50 for inhibition and increased the rate of recovery.

  20. Inhibition by TRPA1 agonists of compound action potentials in the frog sciatic nerve

    SciTech Connect

    Matsushita, Akitomo; Ohtsubo, Sena; Fujita, Tsugumi; Kumamoto, Eiichi

    2013-04-26

    Highlights: •TRPA1 agonists inhibited compound action potentials in frog sciatic nerves. •This inhibition was not mediated by TRPA1 channels. •This efficacy was comparable to those of lidocaine and cocaine. •We found for the first time an ability of TRPA1 agonists to inhibit nerve conduction. -- Abstract: Although TRPV1 and TRPM8 agonists (vanilloid capsaicin and menthol, respectively) at high concentrations inhibit action potential conduction, it remains to be unknown whether TRPA1 agonists have a similar action. The present study examined the actions of TRPA1 agonists, cinnamaldehyde (CA) and allyl isothiocyanate (AITC), which differ in chemical structure from each other, on compound action potentials (CAPs) recorded from the frog sciatic nerve by using the air-gap method. CA and AITC concentration-dependently reduced the peak amplitude of the CAP with the IC{sub 50} values of 1.2 and 1.5 mM, respectively; these activities were resistant to a non-selective TRP antagonist ruthenium red or a selective TRPA1 antagonist HC-030031. The CA and AITC actions were distinct in property; the latter but not former action was delayed in onset and partially reversible, and CA but not AITC increased thresholds to elicit CAPs. A CAP inhibition was seen by hydroxy-α-sanshool (by 60% at 0.05 mM), which activates both TRPA1 and TRPV1 channels, a non-vanilloid TRPV1 agonist piperine (by 20% at 0.07 mM) and tetrahydrolavandulol (where the six-membered ring of menthol is opened; IC{sub 50} = 0.38 mM). It is suggested that TRPA1 agonists as well as TRPV1 and TRPM8 agonists have an ability to inhibit nerve conduction without TRP activation, although their agonists are quite different in chemical structure from each other.

  1. Negative regulation of inflammatory responses by immunoglobulin A receptor (FcαRI) inhibits the development of Toll-like receptor-9 signalling-accelerated glomerulonephritis.

    PubMed

    Watanabe, T; Kanamaru, Y; Liu, C; Suzuki, Y; Tada, N; Okumura, K; Horikoshi, S; Tomino, Y

    2011-11-01

    Myeloid FcαRI, a receptor for immunoglobulin (Ig)A, mediates cell activation or inhibition depending on the type of ligand interaction, which can be either multivalent or monovalent. Anti-inflammatory signalling is triggered by monomeric targeting using anti-FcαRI Fab or IgA ligand binding, which inhibits immune and non-immune-mediated renal inflammation. The participation of Toll-like receptors (TLRs) in kidney pathology in experimental models and various forms of human glomerular nephritis has been discussed. However, little is known about negative regulation of innate-immune activation. In the present study, we generated new transgenic mice that express FcαRI(R209L) /FcRγ chimeric protein and showed that the monovalent targeting of FcαRI exhibited inhibitory effects in an in vivo model of TLR-9 signalling-accelerated nephritis. Mouse monoclonal anti-FcαRI MIP8a Fab improved urinary protein levels and reduced the number of macrophages and immunoglobulin deposition in the glomeruli. Monovalent targeting using MIP8a Fab attenuates the TLR-9 signalling pathway and is associated with phosphorylation of extracellular signal-related protein kinases [extracellular signal-regulated kinase (ERK), P38, c-Jun N-terminal kinase (JNK)] and the activation of nuclear factor (NF)-κB. The inhibitory mechanism involves recruitment of tyrosine phosphatase Src homology 2 domain-containing phosphatase-1 (SHP-1) to FcαRI. Furthermore, cell transfer studies with macrophages pretreated with MIP8a Fab showed that blockade of FcαRI signalling in macrophages prevents the development of TLR-9 signalling-accelerated nephritis. These results suggest a role of anti-FcαRI Fab as a negative regulator in controlling the magnitude of the innate immune response and a new type of anti-inflammatory drug for treatment of kidney disease. © 2011 The Authors. Clinical and Experimental Immunology © 2011 British Society for Immunology.

  2. Activation of Toll-like receptor 9 inhibits LPS-induced receptor activator of NF-κB ligand expression in rat B lymphocytes

    PubMed Central

    Yu, Xiaoqian; Lin, Jiang; Yu, Qing; Kawai, Toshihisa; Taubman, Martin A.; Han, Xiaozhe

    2014-01-01

    B lymphocytes express multiple Toll-like receptors (TLRs) that regulate cytokine production by these B cells. We investigated the effect of TLR4 and TLR9 activation on receptor activator of NF-κB ligand (RANKL) expression by rat spleen B cells. Splenocytes or purified spleen B cells from Rowett rats were cultured with TLR4 ligand E. coli LPS and/or TLR9 ligand CpG-oligodeoxynucleotide (CpG-ODN) for 2 days. RANKL mRNA expressions and the percentage of RANKL-positive B cells were increased in rat splenocytes challenged by E. coli LPS alone. Such increase was diminished when cells were treated with both CpG-ODN and E. coli LPS. Microarray results revealed that expressions of multiple cyclin-dependent kinase (CDK) pathway-related genes were up-regulated only in cells treated with both E. coli LPS and CpG-ODN. This study suggests that CpG-ODN inhibit LPS-induced RANKL expression in rat B cells via regulation of CDK pathway. PMID:24661200

  3. Rapid kinetics of 2-adrenergic agonist binding and inhibition of adenylate cyclase

    SciTech Connect

    Thomsen, W.; Neubig, R.R.

    1987-05-01

    Activation of 2-adrenergic receptors in human platelets results in inhibition of adenylate cyclase (AC). To elucidate the relation between agonist binding and response, the authors have used a novel rapid-mix quench method to compare the kinetics of binding and response. At functionally effective concentrations, the time course of binding of the full 2-agonist, (TH)UK14,304 (UK), to purified platelet membranes was faster than could be measured manually. Using the rapid-mix quench method, agonist binding was quantitated for times for 0.3 to 60 seconds. UK binding exhibited biexponential kinetics. The rate constant of the fast binding component increases linearly with agonist concentration from 1 to 100 nM with a second order rate constant and 7 x 10WM s (at 25C). The slow rate constant was nearly independent of agonist concentration. The half times of the fast and slow components of binding for 100 nM UK are 1.5 seconds and approximately 2 minutes respectively. The rate and magnitude of the fast binding was unaffected by 10 M GTP whereas the magnitude of the slow phase was markedly reduced. Inhibition of forskolin stimulated AC by 100 M epinephrine occurs with a lag of 5-10 seconds in the presence of 10 M GTP. At lower GTP concentrations, this lag is prolonged. The observation that the fast component of agonist binding precedes inhibition even at agonist concentrations 20-fold lower than the EC40 for responses indicates that the rate limiting step in inhibition of AC is distal to the binding of agonist.

  4. Prophylactic Herpes Simplex Virus 2 (HSV-2) Vaccines Adjuvanted with Stable Emulsion and Toll-Like Receptor 9 Agonist Induce a Robust HSV-2-Specific Cell-Mediated Immune Response, Protect against Symptomatic Disease, and Reduce the Latent Viral Reservoir.

    PubMed

    Hensel, Michael T; Marshall, Jason D; Dorwart, Michael R; Heeke, Darren S; Rao, Eileen; Tummala, Padmaja; Yu, Li; Cohen, Gary H; Eisenberg, Roselyn J; Sloan, Derek D

    2017-02-22

    Several prophylactic vaccines targeting HSV-2 have failed in the clinic to demonstrate a sustained depression in viral shedding or protection from recurrences. Although these vaccines have generated high titers of neutralizing antibodies, their induction of robust CD8 T cells has largely been unreported, even though evidence for the importance of HSV-2 antigen-specific CD8 T cells is mounting in animal models and in translational studies involving subjects with active HSV-2-specific immune responses. We developed a subunit vaccine composed of the neutralizing antibody (nAb) targets gD and gB, the novel T cell antigen and tegument protein UL40, and we compared this to a whole-inactivated virus vaccine (FI-HSV-2). We evaluated different formulations in combination with several Th1-inducing TLR agonists in vivo. In mice, the TLR9 agonist cytosine-phosphate-guanine (CpG) oligodeoxynucleotide formulated in a squalene-based oil-in-water emulsion promoted the most robust, functional HSV-2 antigen-specific CD8 T cell responses and high neutralizing antibodies, demonstrating superiority to vaccines adjuvanted by monophosphoryl lipid A (MPL)/alum. We further established that FI-HSV-2 alone or in combination with adjuvants as well as adjuvanted subunit vaccines were successful in the induction of nAbs and T cell responses in guinea pigs. These immunological responses were coincident with a suppression of vaginal HSV-2 shedding, low lesion scores, and a reduction in latent HSV-2 DNA in dorsal root ganglia to undetectable levels. These data support the further preclinical and clinical development of prophylactic HSV-2 vaccines that contain appropriate antigen and adjuvant components responsible for programming elevated CD8 T cell responses.IMPORTANCE Millions of people worldwide are infected with herpes simplex virus type 2 (HSV-2), and to date, an efficacious prophylactic vaccine has not met the rigors of clinical trials. Attempts to develop a vaccine have focused primarily on

  5. Beta-adrenergic agonists inhibit corticosteroid-induced apoptosis of airway epithelial cells.

    PubMed

    Tse, Roberta; Marroquin, Bertha A; Dorscheid, Delbert R; White, Steven R

    2003-08-01

    Airway epithelial damage is a feature of persistent asthma. Treatment with inhaled and oral corticosteroids may suppress inflammation and gain clinical control despite continued epithelial damage. We have previously demonstrated that corticosteroids elicit apoptosis of airway epithelial cells in culture. beta-Adrenergic receptor agonists are commonly used in asthma therapy and can inhibit corticosteroid-induced apoptosis of eosinophils. We tested the hypothesis that beta-adrenergic agonists would inhibit corticosteroid-induced airway epithelial cell apoptosis in cultured primary airway epithelial cells and in the cell line 1HAEo-. Albuterol treatment inhibited dexamethasone-induced apoptosis completely but did not inhibit apoptosis induced by Fas receptor activation. The protective effect of albuterol was duplicated by two different analogs of protein kinase A. The protective effect was not associated with increased translocation of the glucocorticoid receptor to the nucleus nor with changes in glucocorticoid receptor-mediated transcriptional activation or repression. We demonstrate that beta-adrenergic agonists can inhibit corticosteroid-induced apoptosis but not apoptosis induced by Fas activation. These data suggest that one potential deleterious effect of corticosteroid therapy in asthma can be prevented by concomitant beta-adrenergic agonist treatment.

  6. Inhibition by TRPA1 agonists of compound action potentials in the frog sciatic nerve.

    PubMed

    Matsushita, Akitomo; Ohtsubo, Sena; Fujita, Tsugumi; Kumamoto, Eiichi

    2013-04-26

    Although TRPV1 and TRPM8 agonists (vanilloid capsaicin and menthol, respectively) at high concentrations inhibit action potential conduction, it remains to be unknown whether TRPA1 agonists have a similar action. The present study examined the actions of TRPA1 agonists, cinnamaldehyde (CA) and allyl isothiocyanate (AITC), which differ in chemical structure from each other, on compound action potentials (CAPs) recorded from the frog sciatic nerve by using the air-gap method. CA and AITC concentration-dependently reduced the peak amplitude of the CAP with the IC50 values of 1.2 and 1.5mM, respectively; these activities were resistant to a non-selective TRP antagonist ruthenium red or a selective TRPA1 antagonist HC-030031. The CA and AITC actions were distinct in property; the latter but not former action was delayed in onset and partially reversible, and CA but not AITC increased thresholds to elicit CAPs. A CAP inhibition was seen by hydroxy-α-sanshool (by 60% at 0.05 mM), which activates both TRPA1 and TRPV1 channels, a non-vanilloid TRPV1 agonist piperine (by 20% at 0.07 mM) and tetrahydrolavandulol (where the six-membered ring of menthol is opened; IC50=0.38 mM). It is suggested that TRPA1 agonists as well as TRPV1 and TRPM8 agonists have an ability to inhibit nerve conduction without TRP activation, although their agonists are quite different in chemical structure from each other.

  7. Gender differences in the effects of presynaptic and postsynaptic dopamine agonists on latent inhibition in rats.

    PubMed

    Wang, Ying-Chou; He, Bo-Han; Chen, Chih-Chung; Huang, Andrew Chih Wei; Yeh, Yu-Chi

    2012-04-04

    The present study investigated gender differences in the effects of presynaptic and postsynaptic DA agonists on latent inhibition in the passive avoidance paradigm. During the preexposure phase, 32 male and 32 female Wistar rats were exposed to a passive avoidance box (or a different context) and received drug injections in three trials: the control group received an injection of 10% ascorbic acid in a different context. The experimental groups received injections of 10% ascorbic acid (latent inhibition [LI] group), 1mg/kg of the postsynaptic DA D(1)/D(2) agonist apomorphine (APO group), and 1.5mg/kg of the presynaptic DA agonist methamphetamine (METH group) in a passive avoidance box. All experimental groups were placed in the light compartment of the passive avoidance box and were allowed to enter into the dark compartment to receive a footshock (1mA, 2s) in five trials over 5 days. The latency to enter into the dark compartment was recorded in these five trials. The latent inhibition occurred in the female LI group but not in the male LI group. Regardless of gender, the APO group exhibited an increase in latent inhibition. Male rats in the METH group exhibited a decrease in latent inhibition, but female rats in the METH group exhibited an increase in latent inhibition, indicating that the METH group exhibited sexual dimorphism. The gender factor interacted only with the METH group and not the LI or APO group. The present paper discusses whether gender, the postsynaptic DA D(1)/D(2) agonist APO, and presynaptic DA agonist METH may be related to schizophrenia.

  8. Toll-Like Receptor 9 Mediated Responses in Cardiac Fibroblasts

    PubMed Central

    Ohm, Ingrid Kristine; Alfsnes, Katrine; Belland Olsen, Maria; Ranheim, Trine; Sandanger, Øystein; Dahl, Tuva Børresdatter; Aukrust, Pål; Finsen, Alexandra Vanessa; Yndestad, Arne; Vinge, Leif Erik

    2014-01-01

    Altered cardiac Toll-like receptor 9 (TLR9) signaling is important in several experimental cardiovascular disorders. These studies have predominantly focused on cardiac myocytes or the heart as a whole. Cardiac fibroblasts have recently been attributed increasing significance in mediating inflammatory signaling. However, putative TLR9-signaling through cardiac fibroblasts remains non-investigated. Thus, our aim was to explore TLR9-signaling in cardiac fibroblasts and investigate the consequence of such receptor activity on classical cardiac fibroblast cellular functions. Cultivated murine cardiac fibroblasts were stimulated with different TLR9 agonists (CpG A, B and C) and assayed for the secretion of inflammatory cytokines (tumor necrosis factor α [TNFα], CXCL2 and interferon α/β). Expression of functional cardiac fibroblast TLR9 was proven as stimulation with CpG B and –C caused significant CXCL2 and TNFα-release. These responses were TLR9-specific as complete inhibition of receptor-stimulated responses was achieved by co-treatment with a TLR9-antagonist (ODN 2088) or chloroquine diphosphate. TLR9-stimulated responses were also found more potent in cardiac fibroblasts when compared with classical innate immune cells. Stimulation of cardiac fibroblasts TLR9 was also found to attenuate migration and proliferation, but did not influence myofibroblast differentiation in vitro. Finally, results from in vivo TLR9-stimulation with subsequent fractionation of specific cardiac cell-types (cardiac myocytes, CD45+ cells, CD31+ cells and cardiac fibroblast-enriched cell-fractions) corroborated our in vitro data and provided evidence of differentiated cell-specific cardiac responses. Thus, we conclude that cardiac fibroblast may constitute a significant TLR9 responder cell within the myocardium and, further, that such receptor activity may impact important cardiac fibroblast cellular functions. PMID:25126740

  9. Toll-like receptor 9 mediated responses in cardiac fibroblasts.

    PubMed

    Ohm, Ingrid Kristine; Alfsnes, Katrine; Belland Olsen, Maria; Ranheim, Trine; Sandanger, Øystein; Dahl, Tuva Børresdatter; Aukrust, Pål; Finsen, Alexandra Vanessa; Yndestad, Arne; Vinge, Leif Erik

    2014-01-01

    Altered cardiac Toll-like receptor 9 (TLR9) signaling is important in several experimental cardiovascular disorders. These studies have predominantly focused on cardiac myocytes or the heart as a whole. Cardiac fibroblasts have recently been attributed increasing significance in mediating inflammatory signaling. However, putative TLR9-signaling through cardiac fibroblasts remains non-investigated. Thus, our aim was to explore TLR9-signaling in cardiac fibroblasts and investigate the consequence of such receptor activity on classical cardiac fibroblast cellular functions. Cultivated murine cardiac fibroblasts were stimulated with different TLR9 agonists (CpG A, B and C) and assayed for the secretion of inflammatory cytokines (tumor necrosis factor α [TNFα], CXCL2 and interferon α/β). Expression of functional cardiac fibroblast TLR9 was proven as stimulation with CpG B and -C caused significant CXCL2 and TNFα-release. These responses were TLR9-specific as complete inhibition of receptor-stimulated responses was achieved by co-treatment with a TLR9-antagonist (ODN 2088) or chloroquine diphosphate. TLR9-stimulated responses were also found more potent in cardiac fibroblasts when compared with classical innate immune cells. Stimulation of cardiac fibroblasts TLR9 was also found to attenuate migration and proliferation, but did not influence myofibroblast differentiation in vitro. Finally, results from in vivo TLR9-stimulation with subsequent fractionation of specific cardiac cell-types (cardiac myocytes, CD45+ cells, CD31+ cells and cardiac fibroblast-enriched cell-fractions) corroborated our in vitro data and provided evidence of differentiated cell-specific cardiac responses. Thus, we conclude that cardiac fibroblast may constitute a significant TLR9 responder cell within the myocardium and, further, that such receptor activity may impact important cardiac fibroblast cellular functions.

  10. Inhibition of tubuloglomerular feedback by the D1 agonist fenoldopam in chronically salt-loaded rats.

    PubMed Central

    Häberle, D A; Königbauer, B

    1991-01-01

    1. Chronic dietary NaCl loading in rats is paralleled by an increase of the dopamine concentration in the tubular fluid and humorally mediated inhibition of the tubuloglomerular feedback mechanism at the macula densa. Since these two phenomena are causally linked, the alterations in the tubuloglomerular feedback response by the luminal application of dopamine, the D1 agonist fenoldopam, the D2 agonist bromocriptine and the D1 and D2 antagonists SCH 23390 and metoclopramide were further investigated using the micropuncture technique. 2. Very similar, concentration-dependent inhibition of the tubuloglomerular feedback response was observed for dopamine and fenoldopam. Half-maximal inhibition was achieved at 10(-11) M and the slope factors of the sigmoid concentration-response curves were comparable. Bromocriptine was ineffective. 3. The inhibition of TGF by both agonists could be antagonized very similarly and concentration dependently by the D1 antagonist SCH 23390. At equimolar concentrations of 10(-9) M the inhibition was reduced by approximately 50%. Raising the SCH 23390 concentration to 10(-6) M completely abolished the TGF inhibition. In contrast, TGF inhibition by 10(-9) M-fenoldopam or dopamine was not significantly affected by an equimolar concentration of the D2 antagonist metoclopramide. Increasing metoclopramide concentration to 10(-6) M attenuated tubuloglomerular feedback inhibition by approximately 55%. 4. It is concluded that the inhibition of tubuloglomerular feedback seen during chronic dietary salt loading can be ascribed to the binding of endogenous dopamine to luminal D1 receptors on the macula densa cells. PMID:1687747

  11. Cyclic AMP agonist inhibition increases at low levels of histamine release from human basophils

    SciTech Connect

    Tung, R.S.; Lichtenstein, L.M.

    1981-09-01

    The relationship between the intensity of the signal for antigen-induced immunoglobulin E-mediated histamine release from human basophils and the concentration of agonist needed to inhibit release has been determined. The agonists, prostaglandin E1, dimaprit, fenoterol, isobutylmethylxanthine and dibutyryl cyclic AMP, all act by increasing the cyclic AMP level. Each agonist was 10- to 1000-fold more potent (relative ID50) at low levels of histamine release (5-10% of total histamine) than at high levels (50-80%). Thus, the inhibitory potential of a drug is a function of the concentration of antigen used to initiate the response. Our results are now more in accord with the inhibitory profile of these drugs in human lung tissue. It is suggested that in vivo release is likely to be low and that this is the level at which to evaluate drugs in vitro.

  12. The effects of dopamine agonists on prepulse inhibition in healthy men depend on baseline PPI values.

    PubMed

    Bitsios, Panos; Giakoumaki, Stella G; Frangou, Sophia

    2005-10-01

    Dopamine (DA) agonists reliably disrupt prepulse inhibition (PPI) of the startle reflex in animals but less so in humans despite cross-species similarities in the neural regulation of PPI. This study examines whether individual variation in baseline PPI may account for the inconsistencies in DA agonist-induced PPI disruption in humans. Baseline PPI measures were obtained from 32 healthy adult men. Subjects were subsequently tested in three sessions after ingestion of placebo or active drug in a balanced double-blind design. Seventeen subjects were given 0.05 and 0.1 mg of pergolide (a direct DA agonist) and 15 subjects were given 100 and 200 mg of amantadine (an indirect DA agonist). In each treatment group, subjects were assigned to "high" and "low" PPI subgroups based on the median split of their baseline PPI. Amantadine and pergolide disrupted PPI in high- but not in low-PPI subjects. In contrast, low-PPI subjects showed a trend towards PPI facilitation especially with pergolide. Our results suggest that baseline PPI is an important determinant of the effect of DA agonists on PPI.

  13. Free fatty acid receptor (FFAR) agonists inhibit proliferation of human ovarian cancer cells.

    PubMed

    Hopkins, Mandi M; Meier, Kathryn E

    2017-07-01

    Many cellular actions of omega-3 fatty acids are mediated by two G protein-coupled receptors, FFA1 and FFA4, free fatty acid receptor (FFAR) family members that are activated by these dietary constituents. FFAR agonists inhibit proliferation of human prostate and breast cancer cells. Since omega-3 fatty acids can inhibit ovarian cancer cell growth, the current study tested the potential role of FFARs in the response. OVCAR3 and SKOV3 human ovarian cancer cell lines express mRNA for FFA1; FFA4 mRNA was detected at low levels in SKOV3 but not OVCAR3. Lysophosphatidic acid (LPA) and epidermal growth factor (EGF) stimulated proliferation of both cell lines; these responses were inhibited by eicosopentaneoic acid (EPA) and by GW9508, a synthetic FFAR agonist. The LPA antagonist Ki16425 also inhibited LPA- and EGF-induced proliferation; FFAR agonists had no further effect when added with Ki16425. The results suggest that FFARs are potential targets for ovarian cancer therapy. Copyright © 2017. Published by Elsevier Ltd.

  14. Treating enhanced GABAergic inhibition in Down syndrome: use of GABA α5-selective inverse agonists.

    PubMed

    Martínez-Cué, Carmen; Delatour, Benoît; Potier, Marie-Claude

    2014-10-01

    Excess inhibition in the brain of individuals carrying an extra copy of chromosome 21 could be responsible for cognitive deficits observed throughout their lives. A change in the excitatory/inhibitory balance in adulthood would alter synaptic plasticity, potentially triggering learning and memory deficits. γ-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the mature central nervous system and binds to GABAA receptors, opens a chloride channel, and reduces neuronal excitability. In this review we discuss methods to alleviate neuronal inhibition in a mouse model of Down syndrome, the Ts65Dn mouse, using either an antagonist (pentylenetetrazol) or two different inverse agonists selective for the α5-subunit containing receptor. Both inverse agonists, which reduce inhibitory GABAergic transmission, could rescue learning and memory deficits in Ts65Dn mice. We also discuss safety issues since modulation of the excitatory-inhibitory balance to improve cognition without inducing seizures remains particularly difficult when using GABA antagonists.

  15. Inhibition of forskolin-stimulated adenylate cyclase activity by 5-HT receptor agonists.

    PubMed

    Devivo, M; Maayani, S

    1985-12-17

    We measured the inhibition of forskolin-stimulated adenylate cyclase activity in guinea pig hippocampal membranes by 5-HT, 5-carboxamidotryptamine (CAT) and 8-hydroxy-2-(di-n-propylamino) tetralin (PAT). Low concentrations of these agonists inhibited forskolin-stimulated adenylate cyclase activity in a concentration-dependent and saturable manner. The antagonist spiperone shifted the concentration-response curve to CAT to the right in a parallel manner. The EC50 values of CAT, PAT and 5-HT and the KB of spiperone suggest that this receptor may correspond to the 5-HT1A binding site.

  16. Neurotransmitter agonists inhibit inositol phosphate formation in the brain of bupropione-treated rats

    SciTech Connect

    Butler, P.D.; Hungund, B.; Suckow, R.; Barkai, A.I.

    1986-03-05

    Bupropione is a chemically unique antidepressant whose mechanism of action is not known. In this study they have evaluated the effect of chronic treatment with bupropione on the receptor-mediated release of inositol phosphates (IP) from brain slices in rats. Animals were implanted with Alzet osmotic pumps that delivered bupropione at a constant rate (40mg/kg/day) for 2 weeks. Cross-chopped slices of cerebral cortex from control and drug-treated rats were prelabelled with myo-/sup 3/H-inositol in HEPES buffer containing 11 mM LiCl. Accumulation of IP was measured in the presence and absence of the following agonists: Carbamylcholine (100..mu..m); norepinephrine (5..mu..M) and serotonin (10..mu..M). All agonists stimulated release of IP from slices of control animals but appeared to inhibit IP release in bupropione-treated rats. These results indicate that a phospholipase C inhibitor may appear following the activation of this enzyme by the agonist, and that the agonist-induced formation of the apparent inhibitor may be markedly enhanced after treatment with bupropione.

  17. Buprenorphine is a weak partial agonist that inhibits opioid receptor desensitization

    PubMed Central

    Virk, Michael S.; Arttamangkul, Seksiri; Birdsong, William T.; Williams, John T.

    2009-01-01

    Buprenorphine is a weak partial agonist at mu-opioid receptors that is used for treatment of pain and addiction. Intracellular and whole cell recordings were made from locus coeruleus (LC) neurons in rat brain slices to characterize the actions of buprenorphine. Acute application of buprenorphine caused a hyperpolarization that was prevented by previous treatment of slices with the irreversible opioid antagonist, β-chlornaltrexamine (β-CNA), but was not reversed by a saturating concentration of naloxone. As expected for a partial agonist, sub-saturating concentrations of buprenorphine decreased the [Met]5 enkephalin (ME) induced hyperpolarization or outward current. When the ME induced current was decreased below a critical value, desensitization and internalization of μ-opioid receptors (MOR) was eliminated. The inhibition of desensitization by buprenorphine was not the result of prior desensitization, slow dissociation from the receptor, or elimination of receptor reserve. Treatment of slices with sub-saturating concentrations of etorphine, methadone, oxymorphone or β-CNA also reduced the current induced by ME but did not block ME-induced desensitization. Treatment of animals with buprenorphine for a week resulted in the inhibition of the current induced by ME and a block of desensitization that was not different from the acute application of buprenorphine to brain slices. These observations show the unique characteristics of buprenorphine and further demonstrate the range of agonist selective actions that are possible through G-protein coupled receptors. PMID:19494155

  18. 3,5-Dihydroxybenzoic acid, a specific agonist for hydroxycarboxylic acid 1, inhibits lipolysis in adipocytes.

    PubMed

    Liu, Changlu; Kuei, Chester; Zhu, Jessica; Yu, Jingxue; Zhang, Li; Shih, Amy; Mirzadegan, Taraneh; Shelton, Jonathan; Sutton, Steven; Connelly, Margery A; Lee, Grace; Carruthers, Nicholas; Wu, Jiejun; Lovenberg, Timothy W

    2012-06-01

    Niacin raises high-density lipoprotein and lowers low-density lipoprotein through the activation of the β-hydroxybutyrate receptor hydroxycarboxylic acid 2 (HCA2) (aka GPR109a) but with an unwanted side effect of cutaneous flushing caused by vascular dilation because of the stimulation of HCA2 receptors in Langerhans cells in skin. HCA1 (aka GPR81), predominantly expressed in adipocytes, was recently identified as a receptor for lactate. Activation of HCA1 in adipocytes by lactate results in the inhibition of lipolysis, suggesting that agonists for HCA1 may be useful for the treatment of dyslipidemia. Lactate is a metabolite of glucose, suggesting that HCA1 may also be involved in the regulation of glucose metabolism. The low potency of lactate to activate HCA1, coupled with its fast turnover rate in vivo, render it an inadequate tool for studying the biological role of lactate/HCA1 in vivo. In this article, we demonstrate the identification of 3-hydroxybenzoic acid (3-HBA) as an agonist for both HCA2 and HCA1, whereas 3,5-dihydroxybenzoic acid (3,5-DHBA) is a specific agonist for only HCA1 (EC(50) ∼150 μM). 3,5-DHBA inhibits lipolysis in wild-type mouse adipocytes but not in HCA1-deficient adipocytes. Therefore, 3,5-DHBA is a useful tool for the in vivo study of HCA1 function and offers a base for further HCA1 agonist design. Because 3-HBA and 3,5-DHBA are polyphenolic acids found in many natural products, such as fruits, berries, and coffee, it is intriguing to speculate that other heretofore undiscovered natural substances may have therapeutic benefits.

  19. A3 adenosine receptor agonist reduces brain ischemic injury and inhibits inflammatory cell migration in rats.

    PubMed

    Choi, In-Young; Lee, Jae-Chul; Ju, Chung; Hwang, Sunyoung; Cho, Geum-Sil; Lee, Hyuk Woo; Choi, Won Jun; Jeong, Lak Shin; Kim, Won-Ki

    2011-10-01

    A3 adenosine receptor (A3AR) is recognized as a novel therapeutic target for ischemic injury; however, the mechanism underlying anti-ischemic protection by the A3AR agonist remains unclear. Here, we report that 2-chloro-N(6)-(3-iodobenzyl)-5'-N-methylcarbamoyl-4'-thioadenosine (LJ529), a selective A3AR agonist, reduces inflammatory responses that may contribute to ischemic cerebral injury. Postischemic treatment with LJ529 markedly reduced cerebral ischemic injury caused by 1.5-hour middle cerebral artery occlusion, followed by 24-hour reperfusion in rats. This effect was abolished by the simultaneous administration of the A3AR antagonist MRS1523, but not the A2AAR antagonist SCH58261. LJ529 prevented the infiltration/migration of microglia and monocytes occurring after middle cerebral artery occlusion and reperfusion, and also after injection of lipopolysaccharides into the corpus callosum. The reduced migration of microglia by LJ529 could be related with direct inhibition of chemotaxis and down-regulation of spatiotemporal expression of Rho GTPases (including Rac, Cdc42, and Rho), rather than by biologically relevant inhibition of inflammatory cytokine/chemokine release (eg, IL-1β, TNF-α, and MCP-1) or by direct inhibition of excitotoxicity/oxidative stress (not affected by LJ529). The present findings indicate that postischemic activation of A3AR and the resultant reduction of inflammatory response should provide a promising therapeutic strategy for the treatment of ischemic stroke.

  20. PPARgamma agonists inhibit TGF-beta induced pulmonary myofibroblast differentiation and collagen production: implications for therapy of lung fibrosis.

    PubMed

    Burgess, Heather A; Daugherty, Louis Eugene; Thatcher, Thomas H; Lakatos, Heather F; Ray, Denise M; Redonnet, Michelle; Phipps, Richard P; Sime, Patricia J

    2005-06-01

    Pulmonary fibrosis is a progressive life-threatening disease for which no effective therapy exists. Myofibroblasts are one of the key effector cells in pulmonary fibrosis and are the primary source of extracellular matrix production. Drugs that inhibit the differentiation of fibroblasts to myofibroblasts have potential as antifibrotic therapies. Peroxisome proliferator-activated receptor (PPAR)-gamma is a transcription factor that upon ligation with PPARgamma agonists activates target genes containing PPAR response elements. PPARgamma agonists have anti-inflammatory activities and may have potential as antifibrotic agents. In this study, we examined the abilities of PPARgamma agonists to block two of the most important profibrotic activities of TGF-beta on pulmonary fibroblasts: myofibroblast differentiation and production of excess collagen. Both natural (15d-PGJ2) and synthetic (ciglitazone and rosiglitazone) PPARgamma agonists inhibited TGF-beta-driven myofibroblast differentiation, as determined by alpha-smooth muscle actin-specific immunocytochemistry and Western blot analysis. PPARgamma agonists also potently attenuated TGF-beta-driven type I collagen protein production. A dominant-negative PPARgamma partially reversed the inhibition of myofibroblast differentiation by 15d-PGJ2 and rosiglitazone, but the irreversible PPARgamma antagonist GW-9662 did not, suggesting that the antifibrotic effects of the PPARgamma agonists are mediated through both PPARgamma-dependent and independent mechanisms. Thus PPARgamma agonists have novel and potent antifibrotic effects in human lung fibroblasts and may have potential for therapy of fibrotic diseases in the lung and other tissues.

  1. Transforming growth factor-β1 and cigarette smoke inhibit the ability of β2-agonists to enhance epithelial permeability.

    PubMed

    Unwalla, Hoshang J; Ivonnet, Pedro; Dennis, John S; Conner, Gregory E; Salathe, Matthias

    2015-01-01

    Chronic bronchitis, caused by cigarette smoke exposure, is characterized by mucus hypersecretion and reduced mucociliary clearance (MCC). Effective MCC depends, in part, on adequate airway surface liquid. Cystic fibrosis transmembrane conductance regulator (CFTR) provides the necessary osmotic gradient for serosal to mucosal fluid transport through its ability to both secrete Cl(-) and regulate paracellular permeability, but CFTR activity is attenuated in chronic bronchitis and in smokers. β2-adrenergic receptor (β2-AR) agonists are widely used for managing chronic obstructive pulmonary disease, and can activate CFTR, stimulate ciliary beat frequency, and increase epithelial permeability, thereby stimulating MCC. Patients with chronic airway diseases and cigarette smokers demonstrate increased transforming growth factor (TGF)-β1 signaling, which suppresses β2-agonist-mediated CFTR activation and epithelial permeability increases. Restoring CFTR function in these diseases can restore the ability of β2-agonists to enhance epithelial permeability. Human bronchial epithelial cells, fully redifferentiated at the air-liquid interface, were used for (14)C mannitol flux measurements, Ussing chamber experiments, and quantitative RT-PCR. β2-agonists enhance epithelial permeability by activating CFTR via the β2-AR/adenylyl cyclase/cAMP/protein kinase A pathway. TGF-β1 inhibits β2-agonist-mediated CFTR activation and epithelial permeability enhancement. Although TGF-β1 down-regulates both β2-AR and CFTR mRNA, functionally it only decreases CFTR activity. Cigarette smoke exposure inhibits β2-agonist-mediated epithelial permeability increases, an effect reversed by blocking TGF-β signaling. β2-agonists enhance epithelial permeability via CFTR activation. TGF-β1 signaling inhibits β2-agonist-mediated CFTR activation and subsequent increased epithelial permeability, potentially limiting the ability of β2-agonists to facilitate paracellular transport in disease

  2. Nanoparticles Containing a Liver X Receptor Agonist Inhibit Inflammation and Atherosclerosis

    PubMed Central

    Zhang, Xue-Qing; Even-Or, Orli; Xu, Xiaoyang; van Rosmalen, Mariska; Lim, Lucas; Gadde, Suresh

    2015-01-01

    Liver X receptor (LXR) signaling pathways regulate lipid metabolism and inflammation, which has generated widespread interest in developing synthetic LXR agonists as potential therapeutics for the management of atherosclerosis. In this study, we demonstrate that nanoparticles (NPs) containing the synthetic LXR agonist GW3965 (NP-LXR) exert anti-inflammatory effects and inhibit the development of atherosclerosis without causing hepatic steatosis. These NPs were engineered through self-assembly of a biodegradable diblock poly(lactide-co-glycolide)-b-poly(ethylene glycol) (PLGA-b-PEG) copolymer. NP-LXR was significantly more effective than free GW3965 at inducing LXR target gene expression and suppressing inflammatory factors in macrophages in vitro and in vivo. Addtionally, the NPs elicited negligible lipogenic gene stimulation in the liver. Using the Ldlr−/− mouse model of atherosclerosis, we saw abundant co-localization of fluorescently labeled NPs within plaque macrophages following systemic administration. Notably, six intravenous injections of NP-LXR over two weeks markedly reduced the CD68-positive cell (macrophage) content of plaques (by 50%) without increasing total cholesterol or triglycerides in the liver and plasma. Together, these findings identify GW3965-encapsulated PLGA-b-PEG NPs as a promising nanotherapeutic approach to combat atherosclerosis, providing the benefits of LXR agonists without their adverse effects on hepatic and plasma lipid metabolism. PMID:25156796

  3. Soluble Guanylate Cyclase Agonists Inhibit Expression and Procoagulant Activity of Tissue Factor

    PubMed Central

    Sovershaev, Mikhail A.; Egorina, Elena M.; Hansen, John-Bjarne; Østerud, Bjarne; Pacher, Pál; Stasch, Johannes-Peter; Evgenov, Oleg V.

    2010-01-01

    Objective Tissue factor (TF), a major initiator of blood coagulation, contributes to inflammation, atherosclerosis, angiogenesis, and vascular remodeling. Pharmacological agonists of soluble guanylate cyclase (sGC) attenuate systemic and pulmonary hypertension, vascular remodeling, and platelet aggregation. However, the influence of these novel pharmacophores on TF is unknown. Methods and Results We evaluated effects of BAY 41-2272 and BAY 58-2667 on expression and activity of TF in human monocytes and umbilical vein endothelial cells (HUVECs). Both compounds reduced expression of active TF protein in monocytes stimulated with lipopolysaccharide, as demonstrated by immunoblotting and a TF procoagulant activity assay. In-cell Western assay revealed that this effect was associated with a marked reduction of total and surface TF presentation. Furthermore, BAY 41-2272 and BAY 58-2667 decreased TF protein expression and the TF-dependent procoagulant activity in HUVECs stimulated with TNF-α. The sGC agonists also suppressed transcriptional activity of NF-κB. A siRNA-mediated knockdown of the α1-subunit of sGC in monocytes and HUVECs confirmed that the inhibitory effect of BAY 41-2272 and BAY 58-2667 on TF expression is mediated through the sGC-dependent mechanisms. Conclusions Inhibition of TF expression and activity by sGC agonists might provide therapeutic benefits in cardiovascular diseases associated with enhanced procoagulant and inflammatory response. PMID:19592462

  4. Transient receptor potential a1 (TRPA1) agonists inhibit contractions of the isolated human ureter.

    PubMed

    Weinhold, Philipp; Hennenberg, Martin; Strittmatter, Frank; Stief, Christian G; Gratzke, Christian; Hedlund, Petter

    2017-07-03

    Mechanoafferent and peristaltic mechanisms of the human ureter involve transient receptor potential V1 (TRPV1)- and purinoceptor-mediated functions. Hydrogen sulphide, an endogenous TRPA1 ligand, is linked to inhibitory neurotransmission of the pig ureter. No information is available on TRPA1 activity in the human ureter. We therefore examined the distribution and function of TRPA1 in the human ureter. Expression of TRPA1 in human ureter tissue was studied by Western blot and immunofluorescence. The TRPA1 distribution was compared to TRPV1, calcitonin gene related peptide (CGRP), tyrosine hydroxylase (TH), and vimentin. Effects of the TRPA1 agonists allyl isothiocyanate (AI), cinnamaldehyde (CA), sodium hydrogen sulfide (NaHS), and capsaicin (TRPV1 agonist) on human ureter preparations were studied in organ baths. By Western blot, bands were detected at the expected molecular weight for TRPA1. TRPA1- and TRPV1-immunoreactivities were located on CGRP-positive nerves, but not on TH-positive nerves. TRPA1 was also located in vimentin-positive interstitial cells. In functional experiments, neither of the TRPA1-agonists (1-100 μM) had any direct effects on ureter tension (baseline/potassium-induced contractions). However, CA, AI, NaHS, and capsaicin (10 μM) decreased (P < 0.01-0.05) tetrodotoxin-sensitive electrically induced (2,4,8,16,32 Hz) contractions. Inhibitory activities were 50-61% (CA), 30-56% (AI), 30-40% (NaHS), and 37-67% (Capsaicin). In the human ureter, TRPA1 is located to sensory nerves and interstitial cells. TRPA1 agonists inhibited electrically induced contractions but had no direct effect on smooth muscle tension of the human ureter. A role for TRPA1 in modulating neurotransmission and possibly peristalsis of the human ureter is proposed. © 2017 Wiley Periodicals, Inc.

  5. GLP-1 agonists inhibit ox-LDL uptake in macrophages by activating protein kinase A.

    PubMed

    Dai, Yao; Dai, Dongsheng; Wang, Xianwei; Ding, Zufeng; Li, Chunlin; Mehta, Jawahar L

    2014-07-01

    Oxidized low-density lipoprotein (ox-LDL) uptake by monocytes/macrophages plays a pivotal role in atherogenesis. This study was designed to examine the effect of glucagon-like peptide-1 (GLP-1) agonists on ox-LDL uptake in macrophages. Human primary monocytes/macrophages were incubated with native GLP-1 (nGLP-1) or GLP-1 agonist liraglutide to evaluate their effect on ox-LDL uptake and the expression of scavenger receptors (SRs), such as SR-A, CD36, and lectin-like ox-LDL SR-1, in this process. Our study showed a decrease in ox-LDL uptake and CD36 expression in macrophages treated with nGLP-1 or liraglutide. However, nGLP-1 and liraglutide did not affect the expression of other SRs SR-A and lectin-like ox-LDL SR-1. Simultaneously, there was an increase in the expression of activated protein kinase A (PKA). To examine the role of PKA in the effects of nGLP-1 or liraglutide, we treated macrophages with PK inhibitor (6-22) amide, a PKA inhibitor, followed by treatment with nGLP-1 or liraglutide. Inhibition of PKA activation markedly reversed the effect of nGLP-1 or liraglutide on ox-LDL uptake and enhanced the expression of CD36. Our results suggest that GLP-1 agonism inhibits ox-LDL uptake through PKA/CD36 pathway in macrophages. This study provides a novel insight in the mechanism of foam cell formation and the role by GLP-1 agonists therein.

  6. Inhibition of the production of endothelium-derived hyperpolarizing factor by cannabinoid receptor agonists

    PubMed Central

    Fleming, I; Schermer, B; Popp, R; Busse, R

    1999-01-01

    The endogenous cannabinoid, anandamide, has been reported to induce an 'endothelium-derived hyperpolarizing factor (EDHF)-like' relaxation in vitro. We therefore investigated the effects of cannabinoid CB1 receptor agonists; HU 210, Δ9-tetrahydrocannabinol (Δ9-THC) and anandamide, and a CB1 antagonist/inverse agonist, SR 141716A, on nitric oxide (NO) and EDHF-mediated relaxation in precontracted rings of porcine coronary, rabbit carotid and mesenteric arteries. In rings of mesenteric artery HU 210 and Δ9-THC induced endothelium- and cyclo-oxygenase-independent relaxations which were sensitive to SR 141716A. Anandamide (0.03–30 μM) induced a slowly developing, endothelium-independent relaxation which was abolished by diclofenac and was therefore mediated by cyclo-oxygenase product(s). None of the CB1 agonists tested affected the tone of precontracted rings of rabbit carotid or porcine coronary artery. In endothelium-intact segments, HU 210, Δ9-THC and anandamide did not affect NO-mediated responses but under conditions of continuous NO synthase/cyclo-oxygenase blockade, significantly inhibited acetylcholine and bradykinin-induced relaxations which are attributed to the production of EDHF. The effects of HU 210 and Δ9-THC were not observed when experiments were performed in the presence of SR 141716A suggesting the involvement of the CB1 receptor. In a patch clamp bioassay of EDHF production, HU 210 decreased the EDHF-mediated hyperpolarization of detector smooth muscle cells when applied to the donor segment but was without effect on the membrane potential of detector cells. The inhibition of EDHF production was unrelated to alterations in Ca2+-signalling or cytochrome P450 activity. These results suggest that the activation of endothelial CB1 receptors appears to be negatively coupled to the production of EDHF. PMID:10193775

  7. Cannabinoid agonists showing BuChE inhibition as potential therapeutic agents for Alzheimer's disease.

    PubMed

    González-Naranjo, Pedro; Pérez-Macias, Natalia; Campillo, Nuria E; Pérez, Concepción; Arán, Vicente J; Girón, Rocio; Sánchez-Robles, Eva; Martín, María Isabel; Gómez-Cañas, María; García-Arencibia, Moisés; Fernández-Ruiz, Javier; Páez, Juan A

    2014-02-12

    Designing drugs with a specific multi-target profile is a promising approach against multifactorial illnesses as Alzheimer's disease. In this work, new indazole ethers that possess dual activity as both cannabinoid agonists CB2 and inhibitors of BuChE have been designed by computational methods. On the basis of this knowledge, the synthesis, pharmacological evaluation and docking studies of a new class of indazoles has been performed. Pharmacological evaluation includes radioligand binding assays with [(3)H]-CP55940 for CB1R and CB2R and functional activity for cannabinoid receptors on isolated tissue. Additionally, in vitro inhibitory assays of AChE/BuChE and the corresponding competition studies have been carried out. The results of pharmacological tests have revealed that three of these derivatives behave as CB2 cannabinoid agonists and simultaneously show BuChE inhibition. In particular, compounds 3 and 24 have emerged as promising candidates as novel cannabinoids that inhibit BuChE by a non-competitive or mixed mechanism, respectively. On the other hand, both molecules show antioxidant properties.

  8. Sigma 1 receptor agonists act as neuroprotective drugs through inhibition of inducible nitric oxide synthase.

    PubMed

    Vagnerova, Kamila; Hurn, Patricia D; Bhardwaj, Anish; Kirsch, Jeffrey R

    2006-08-01

    Postischemic administration of the sigma-1 agonists reduces ischemic brain injury; however, the mechanism is unclear. We hypothesized that the sigma-1 agonist (+)isoform of pentazocine (P(+)) reduces damage in part by ameliorating cell death mediated via inducible nitric oxide synthase (iNOS) and that the (-)isoform (P(-)) lacks this effect. We compared treatment with P(+) with or without the iNOS inhibitor aminoguanidine (AG) and also the effects of P(+) in iNOS deficient (iNOSKO) mice. A possible mechanism of neuroprotection is inhibition of iNOS expression. Male C57/Bl6 mice were subjected to transient middle cerebral artery occlusion (90 min) and drugs were administered with reperfusion: 1) P(+) with AG (P+/AG), 2) P(+), 3) P(-), 4) AG, or 5) placebo. iNOSKOs were treated with either P(+) or placebo. Infarction (triphenyltetrazolium chloride histology, 72 h) was reduced by P(+) treatment in striatum by 44% and in neocortex by 23% versus placebo (P < 0.05), a reduction comparable to AG effect. P(-) did not attenuate brain injury. There was no difference in P(+)/AG treatment compared with showed the same level of neuroprotection as P(+) alone. P(+) also did not provide further neuroprotection for iNOSKOs. We conclude that postischemic administration of P(+) reduces infarct volume in mice. Because AG provides no additional benefit to P(+) treatment and iNOSKOs do not benefit from P(+), we speculate that P(+) acts by suppressing cell death resulting from iNOS toxicity.

  9. A novel TRPV4-specific agonist inhibits monocyte adhesion and atherosclerosis

    PubMed Central

    Xu, Suowen; Liu, Bin; Yin, Meimei; Koroleva, Marina; Mastrangelo, Michael; Ture, Sara; Morrell, Craig N.; Zhang, David X.; Fisher, Edward A.; Jin, Zheng Gen

    2016-01-01

    TRPV4 ion channel mediates vascular mechanosensitivity and vasodilation. Here, we sought to explore whether non-mechanical activation of TRPV4 could limit vascular inflammation and atherosclerosis. We found that GSK1016790A, a potent and specific small-molecule agonist of TRPV4, induces the phosphorylation and activation of eNOS partially through the AMPK pathway. Moreover, GSK1016790A inhibited TNF-α-induced monocyte adhesion to human endothelial cells. Mice given GSK1016790A showed increased phosphorylation of eNOS and AMPK in the aorta and decreased leukocyte adhesion to TNF-α-inflamed endothelium. Importantly, oral administration of GSK1016790A reduced atherosclerotic plaque formation in ApoE deficient mice fed a Western-type diet. Together, the present study suggests that pharmacological activation of TRPV4 may serve as a potential therapeutic approach to treat atherosclerosis. PMID:27191895

  10. Aripiprazole, an Antipsychotic and Partial Dopamine Agonist, Inhibits Cancer Stem Cells and Reverses Chemoresistance.

    PubMed

    Suzuki, Shuhei; Okada, Masashi; Kuramoto, Kenta; Takeda, Hiroyuki; Sakaki, Hirotsugu; Watarai, Hikaru; Sanomachi, Tomomi; Seino, Shizuka; Yoshioka, Takashi; Kitanaka, Chifumi

    2016-10-01

    There is a growing interest in repurposing antipsychotic dopamine antagonists for cancer treatment; however, antipsychotics are often associated with an increased risk of fatal events. The anticancer activities of aripiprazole, an antipsychotic drug with partial dopamine agonist activity and an excellent safety profile, remain unknown. The effects of aripiprazole alone or in combination with chemotherapeutic agents on the growth, sphere-forming ability and stem cell/differentiation/chemoresistance marker expression of cancer stem cells, serum-cultured cancer cells from which they were derived, and normal cells were examined. At concentrations non-toxic to normal cells, aripiprazole inhibited the growth of serum-cultured cancer cells and cancer stem cells. Furthermore, aripiprazole induced differentiation and inhibited sphere formation, as well as stem cell marker expression of cancer stem cells while inhibiting their survivin expression and sensitizing them to chemotherapeutic agents. Repurposing aripiprazole as an anticancer stem cell drug may merit further consideration. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  11. Inhibition of the voltage-dependent calcium currents in isolated frog sensory neurons by GABA-related agonistic compounds.

    PubMed

    Maruyama, T; Behrends, J C; Akaike, N

    1988-12-01

    Effects of GABAA-, barbiturate- and benzodiazepine receptor agonists and GABAB agonist, baclofen, on voltage-dependent Ca2+ current (ICa) were studied in isolated frog sensory neurons after suppression of Na+ and K+ currents using single-electrode voltage-clamp. GABA, muscimol, taurine and pentobarbital (PB) dose-dependently induced a transient Cl- current (ICl), while baclofen and diazepam (DZP) did not elicit any currents. With GABAA agonists such as GABA, muscimol and taurine, ICa was suppressed transiently, and the maximum inhibition of ICa occurred within 1 min. The suppression of ICa by all GABAA agonists was neither voltage dependent nor attenuated in the presence of either bicuculline or picrotoxin. In addition, there was no correlation between GABA- and baclofen-induced suppressions of ICa. The results suggest that the inhibition of ICa by GABAA receptor agonists is not due to either GABAA or GABAB receptor activation at least. The inhibition of ICa by baclofen, PB and DZP was persistent. PB suppressed the amplitude of ICa and also facilitated the inactivation process, suggesting that PB behaves as a Ca channel blocker. However, the mechanisms of ICa suppression by baclofen and DZP are the subject for a future study. The potency order of the drugs in reducing ICa was muscimol greater than GABA = DZP greater than baclofen greater than PB greater than taurine.

  12. Agonistic Anti-TIGIT Treatment Inhibits T Cell Responses in LDLr Deficient Mice without Affecting Atherosclerotic Lesion Development

    PubMed Central

    Foks, Amanda C.; Ran, Ingrid A.; Frodermann, Vanessa; Bot, Ilze; van Santbrink, Peter J.; Kuiper, Johan; van Puijvelde, Gijs H. M.

    2013-01-01

    Objective Co-stimulatory and co-inhibitory molecules are mainly expressed on T cells and antigen presenting cells and strongly orchestrate adaptive immune responses. Whereas co-stimulatory molecules enhance immune responses, signaling via co-inhibitory molecules dampens the immune system, thereby showing great therapeutic potential to prevent cardiovascular diseases. Signaling via co-inhibitory T cell immunoglobulin and ITIM domain (TIGIT) directly inhibits T cell activation and proliferation, and therefore represents a novel therapeutic candidate to specifically dampen pro-atherogenic T cell reactivity. In the present study, we used an agonistic anti-TIGIT antibody to determine the effect of excessive TIGIT-signaling on atherosclerosis. Methods and Results TIGIT was upregulated on CD4+ T cells isolated from mice fed a Western-type diet in comparison with mice fed a chow diet. Agonistic anti-TIGIT suppressed T cell activation and proliferation both in vitro and in vivo. However, agonistic anti-TIGIT treatment of LDLr−/− mice fed a Western-type diet for 4 or 8 weeks did not affect atherosclerotic lesion development in comparison with PBS and Armenian Hamster IgG treatment. Furthermore, elevated percentages of dendritic cells were observed in the blood and spleen of agonistic anti-TIGIT-treated mice. Additionally, these cells showed an increased activation status but decreased IL-10 production. Conclusions Despite the inhibition of splenic T cell responses, agonistic anti-TIGIT treatment does not affect initial atherosclerosis development, possibly due to increased activity of dendritic cells. PMID:24376654

  13. Nitric oxide inhibits isoproterenol-stimulated adipocyte lipolysis through oxidative inactivation of the beta-agonist.

    PubMed Central

    Klatt, P; Cacho, J; Crespo, M D; Herrera, E; Ramos, P

    2000-01-01

    Nitric oxide has been implicated in the inhibition of catecholamine-stimulated lipolysis in adipose tissue by as yet unknown mechanisms. In the present study, it is shown that the nitric oxide donor, 2,2-diethyl-1-nitroso-oxyhydrazine, antagonized isoproterenol (isoprenaline)-induced lipolysis in rat adipocytes, freshly isolated from white adipose tissue, by decreasing the potency of the beta-agonist without affecting its efficacy. These data suggest that nitric oxide did not act downstream of the beta-adrenoceptor but reduced the effective concentration of isoproterenol. In support of the latter hypothesis, we found that pre-treatment of isoproterenol with nitric oxide abolished the lipolytic activity of the catecholamine. Spectroscopic data and HPLC analysis confirmed that the nitric oxide-mediated inactivation of isoproterenol was in fact because of the modification of the catecholamine through a sequence of oxidation reactions, which apparently involved the generation of an aminochrome. Similarly, aminochrome was found to be the primary product of isoproterenol oxidation by 3-morpholinosydnonimine and peroxynitrite. Finally, it was shown that nitric oxide released from cytokine-stimulated adipocytes attenuated the lipolytic effect of isoproterenol by inactivating the catecholamine. In contrast with very recent findings, which suggest that nitric oxide impairs the beta-adrenergic action of isoproterenol through intracellular mechanisms and not through a chemical reaction between NO and the catecholamine, we showed that nitric oxide was able to attenuate the pharmacological activity of isoproterenol in vitro as well as in a nitric oxide-generating cellular system through oxidation of the beta-agonist. These findings should be taken into account in both the design and interpretation of studies used to investigate the role of nitric oxide as a modulator of isoproterenol-stimulated signal transduction pathways. PMID:11023835

  14. Peripheral Administration of a Long-Acting Peptide Oxytocin Receptor Agonist Inhibits Fear-Induced Freezing

    PubMed Central

    Modi, Meera E.; Majchrzak, Mark J.; Fonseca, Kari R.; Doran, Angela; Osgood, Sarah; Vanase-Frawley, Michelle; Feyfant, Eric; McInnes, Heather; Darvari, Ramin; Buhl, Derek L.

    2016-01-01

    Oxytocin (OT) modulates the expression of social and emotional behaviors and consequently has been proposed as a pharmacologic treatment of psychiatric diseases, including autism spectrum disorders and schizophrenia; however, endogenous OT has a short half-life in plasma and poor permeability across the blood-brain barrier. Recent efforts have focused on the development of novel drug delivery methods to enhance brain penetration, but few efforts have aimed at improving its half-life. To explore the behavioral efficacy of an OT analog with enhanced plasma stability, we developed PF-06655075 (PF1), a novel non–brain-penetrant OT receptor agonist with increased selectivity for the OT receptor and significantly increased pharmacokinetic stability. PF-06478939 was generated with only increased stability to disambiguate changes to selectivity versus stability. The efficacy of these compounds in evoking behavioral effects was tested in a conditioned fear paradigm. Both central and peripheral administration of PF1 inhibited freezing in response to a conditioned fear stimulus. Peripheral administration of PF1 resulted in a sustained level of plasma concentrations for greater than 20 hours but no detectable accumulation in brain tissue, suggesting that plasma or cerebrospinal fluid exposure was sufficient to evoke behavioral effects. Behavioral efficacy of peripherally administered OT receptor agonists on conditioned fear response opens the door to potential peripheral mechanisms in other behavioral paradigms, whether they are mediated by direct peripheral activation or feed-forward responses. Compound PF1 is freely available as a tool compound to further explore the role of peripheral OT in behavioral response. PMID:27217590

  15. Nitric oxide inhibits isoproterenol-stimulated adipocyte lipolysis through oxidative inactivation of the beta-agonist.

    PubMed

    Klatt, P; Cacho, J; Crespo, M D; Herrera, E; Ramos, P

    2000-10-15

    Nitric oxide has been implicated in the inhibition of catecholamine-stimulated lipolysis in adipose tissue by as yet unknown mechanisms. In the present study, it is shown that the nitric oxide donor, 2,2-diethyl-1-nitroso-oxyhydrazine, antagonized isoproterenol (isoprenaline)-induced lipolysis in rat adipocytes, freshly isolated from white adipose tissue, by decreasing the potency of the beta-agonist without affecting its efficacy. These data suggest that nitric oxide did not act downstream of the beta-adrenoceptor but reduced the effective concentration of isoproterenol. In support of the latter hypothesis, we found that pre-treatment of isoproterenol with nitric oxide abolished the lipolytic activity of the catecholamine. Spectroscopic data and HPLC analysis confirmed that the nitric oxide-mediated inactivation of isoproterenol was in fact because of the modification of the catecholamine through a sequence of oxidation reactions, which apparently involved the generation of an aminochrome. Similarly, aminochrome was found to be the primary product of isoproterenol oxidation by 3-morpholinosydnonimine and peroxynitrite. Finally, it was shown that nitric oxide released from cytokine-stimulated adipocytes attenuated the lipolytic effect of isoproterenol by inactivating the catecholamine. In contrast with very recent findings, which suggest that nitric oxide impairs the beta-adrenergic action of isoproterenol through intracellular mechanisms and not through a chemical reaction between NO and the catecholamine, we showed that nitric oxide was able to attenuate the pharmacological activity of isoproterenol in vitro as well as in a nitric oxide-generating cellular system through oxidation of the beta-agonist. These findings should be taken into account in both the design and interpretation of studies used to investigate the role of nitric oxide as a modulator of isoproterenol-stimulated signal transduction pathways.

  16. Exposure to D2-like dopamine receptor agonists inhibits swimming in Daphnia magna.

    PubMed

    Barrozo, Enrico R; Fowler, David A; Beckman, Matthew L

    2015-10-01

    Daphnia are freshwater crustaceans that have been used for decades in ecotoxicology research. Despite the important role that Daphnia have played in environmental toxicology studies, very little is known about the neurobiology of Daphnia. Although many studies have investigated the swimming movements of these "water fleas", few studies have examined the underlying neurochemical basis for these movements. To characterize the locomotor effect of drugs in Daphnia, a two-dimensional video imaging tool was developed and animal tracking was performed with freely available software, CTRAX. Due to the central role that dopamine plays in the movement of animals, we sought to determine the role of dopamine receptor signaling in Daphnia movement by characterizing the effect of ten drugs that are agonists or antagonists of dopamine receptors. At 1, 2, and 6h of treatment with a 10μM drug, several dopamine receptor agonists with documented effects on the D2-like class of receptors decreased the movement. Further, we determined behavioral inhibition values (IC50) at 1h of treatment for (1R,3S)-1-(aminomethyl)-3-phenyl-3,4-dihydro-1H-isochromene-5,6-diol (A68930) to be 1.4μM and for bromocriptine to be 6.6μM. This study describes a new method to study Daphnia swimming and establishes this organism as a useful model for studies of dopaminergic signaling. Specifically, this study shows that a dopamine receptor signaling pathway, mediated by putative D2-like receptors, is involved in the control of Daphnia swimming behavior. Due to its ease of use and its rich motor program we propose that Daphnia should be considered for future studies of dopamine neuron toxicity and protection. Copyright © 2015 Elsevier Inc. All rights reserved.

  17. Organ fibrosis inhibited by blocking transforming growth factor-β signaling via peroxisome proliferator-activated receptor γ agonists.

    PubMed

    Deng, Yi-Lei; Xiong, Xian-Ze; Cheng, Nan-Sheng

    2012-10-01

    Organ fibrosis has been viewed as one of the major medical problems, which can lead to progressive dysfunction of the liver, lung, kidney, skin, heart, and eventually death of patients. Fibrosis is initiated by a variety of pathological, physiological, biochemical, and physical factors. Regardless of their different etiologies, they all share a common pathogenetic process: excessive activation of the key profibrotic cytokine, transforming growth factor-beta (TGF-beta). Peroxisome proliferator-activated receptor gamma (PPARgamma), a ligand-activated transcription factor of the nuclear receptor superfamily, has received particular attention in recent years, because the activation of PPARgamma by both natural and synthetic agonists could effectively inhibit TGF-beta-induced profibrotic effects in many organs. The English-language medical databases, PubMed, Elsevier and SpringerLink were searched for articles on PPARgamma, TGF-beta, and fibrosis, and related topics. TGF-beta is recognized as a key profibrotic cytokine. Excessive activation of TGF-beta increases synthesis of extracellular matrix proteins and decreases their degradation, associated with a gradual destruction of normal tissue architecture and function, whereas PPARgamma agonists inhibit TGF-beta signal transduction and are effective antifibrogenic agents in many organs including the liver, lung, kidney, skin and heart. The main antifibrotic activity of PPARgamma agonists is to suppress the TGF-beta signaling pathway by so-called PPARgamma-dependent effect. In addition, PPARgamma agonists, especially 15d-PGJ2, also exert potentially antifibrotic activity independent of PPARgamma activation. TGF-beta1/Smads signaling not only plays many essential roles in multiple developmental processes, but also forms cross-talk networks with other signal pathways, and their inhibition by PPARgamma agonists certainly affects the cytokine networks and causes non-suspected side-effects. Anti-TGF-beta therapies with

  18. Inhibition of experimental auto-immune uveitis by the A3 adenosine receptor agonist CF101.

    PubMed

    Bar-Yehuda, Sara; Luger, Dror; Ochaion, Avivit; Cohen, Shira; Patokaa, Renana; Zozulya, Galina; Silver, Phyllis B; de Morales, Jose Maria Garcia Ruiz; Caspi, Rachel R; Fishman, Pnina

    2011-11-01

    Uveitis is an inflammation of the middle layer of the eye with a high risk of blindness. The Gi protein associated A3 adenosine receptor (A3AR) is highly expressed in inflammatory cells whereas low expression is found in normal cells. CF101 is a highly specific agonist at the A3AR known to induce a robust anti-inflammatory effect in different experimental animal models. The CF101 mechanism of action entails down-regulation of the NF-κB-TNF-α signaling pathway, resulting in inhibition of pro-inflammatory cytokine production and apoptosis of inflammatory cells. In this study the effect of CF101 on the development of retinal antigen interphotoreceptor retinoid-binding protein (IRBP)-induced experimental autoimmune uveitis (EAU) was assessed. Oral treatment with CF101 (10 µg/kg, twice daily), initiated upon disease onset, improved uveitis clinical score measured by fundoscopy and ameliorated the pathological manifestations of the disease. Shortly after treatment with CF101 A3AR expression levels were down-regulated in the lymph node and spleen cells pointing towards receptor activation. Downstream events included a decrease in PI3K and STAT-1 and proliferation inhibition of IRPB auto-reactive T cells ex vivo. Inhibition of interleukin-2, tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ) production and up-regulation of interleukin-10 was found in cultured splenocytes derived from CF101-treated animals. Overall, the present study data point towards a marked anti-inflammatory effect of CF101 in EAU and support further exploration of this small molecule drug for the treatment of uveitis.

  19. The melanocortin MC1 receptor agonist BMS-470539 inhibits leucocyte trafficking in the inflamed vasculature

    PubMed Central

    Leoni, G; Voisin, M-B; Carlson, K; Getting, SJ; Nourshargh, S; Perretti, M

    2010-01-01

    Background and purpose: Over three decades of research evaluating the biology of melanocortin (MC) hormones and synthetic peptides, activation of the MC type 1 (MC1) receptor has been identified as a viable target for the development of novel anti-inflammatory therapeutic agents. Here, we have tested a recently described selective agonist of MC1 receptors, BMS-470539, on leucocyte/post-capillary venule interactions in murine microvascular beds. Experimental approach: Intravital microscopy of two murine microcirculations were utilized, applying two distinct modes of promoting inflammation. The specificity of the effects of BMS-470539 was assessed using mice bearing mutant inactive MC1 receptors (the recessive yellow e/e colony). Key results: BMS-470539, given before an ischaemia–reperfusion protocol, inhibited cell adhesion and emigration with no effect on cell rolling, as assessed 90 min into the reperfusion phase. These properties were paralleled by inhibition of tissue expression of both CXCL1 and CCL2. Confocal investigations of inflamed post-capillary venules revealed immunostaining for MC1 receptors on adherent and emigrated leucocytes. Congruently, the anti-inflammatory properties of BMS-470539 were lost in mesenteries of mice bearing the inactive mutant MC1 receptors. Therapeutic administration of BMS-470539 stopped cell emigration, but did not affect cell adhesion in the cremasteric microcirculation inflamed by superfusion with platelet-activating factor. Conclusions and implications: Activation of MC1 receptors inhibited leucocyte adhesion and emigration. Development of new chemical entities directed at MC1 receptors could be a viable approach in the development of novel anti-inflammatory therapeutic agents with potential application to post-ischaemic conditions. PMID:20331604

  20. Transforming Growth Factor-β1 and Cigarette Smoke Inhibit the Ability of β2-Agonists to Enhance Epithelial Permeability

    PubMed Central

    Ivonnet, Pedro; Dennis, John S.; Conner, Gregory E.; Salathe, Matthias

    2015-01-01

    Chronic bronchitis, caused by cigarette smoke exposure, is characterized by mucus hypersecretion and reduced mucociliary clearance (MCC). Effective MCC depends, in part, on adequate airway surface liquid. Cystic fibrosis transmembrane conductance regulator (CFTR) provides the necessary osmotic gradient for serosal to mucosal fluid transport through its ability to both secrete Cl− and regulate paracellular permeability, but CFTR activity is attenuated in chronic bronchitis and in smokers. β2-adrenergic receptor (β2-AR) agonists are widely used for managing chronic obstructive pulmonary disease, and can activate CFTR, stimulate ciliary beat frequency, and increase epithelial permeability, thereby stimulating MCC. Patients with chronic airway diseases and cigarette smokers demonstrate increased transforming growth factor (TGF)-β1 signaling, which suppresses β2-agonist–mediated CFTR activation and epithelial permeability increases. Restoring CFTR function in these diseases can restore the ability of β2-agonists to enhance epithelial permeability. Human bronchial epithelial cells, fully redifferentiated at the air–liquid interface, were used for 14C mannitol flux measurements, Ussing chamber experiments, and quantitative RT-PCR. β2-agonists enhance epithelial permeability by activating CFTR via the β2-AR/adenylyl cyclase/cAMP/protein kinase A pathway. TGF-β1 inhibits β2-agonist–mediated CFTR activation and epithelial permeability enhancement. Although TGF-β1 down-regulates both β2-AR and CFTR mRNA, functionally it only decreases CFTR activity. Cigarette smoke exposure inhibits β2-agonist–mediated epithelial permeability increases, an effect reversed by blocking TGF-β signaling. β2-agonists enhance epithelial permeability via CFTR activation. TGF-β1 signaling inhibits β2-agonist–mediated CFTR activation and subsequent increased epithelial permeability, potentially limiting the ability of β2-agonists to facilitate paracellular transport in

  1. Appeasing pheromone inhibits cortisol augmentation and agonistic behaviors during social stress in adult miniature pigs.

    PubMed

    Yonezawa, Tomohiro; Koori, Miyuki; Kikusui, Takefumi; Mori, Yuji

    2009-11-01

    Pairing and physical confrontation In adult sows causes social stress reactions and aggressive behaviors. Recently, maternal pig skin secretions were Isolated and a mixture containing several fatty acids, now called pig appeasing pheromone (PAP), was synthesized. In this study, we Investigated the effects of PAP on social and Immune stress response In adult female miniature pigs. PAP or vehicle solvents were sprayed Into the pens of Individually housed adult sows. A two-week exposure to the pheromone did not alter basal salivary Cortisol levels or clrcadlan rhythms. Following this treatment, the animals were paired and placed In a new pen that was divided with a wire-mesh fence. Although salivary cortisol Increased markedly In the vehicle-treated group, the PAP-treated group exhibited a drastic Inhibition of cortisol secretion. This effect was sustained even after they were allowed to physically Interact following fence removal. Moreover, the latency time of agonistic behaviors, such as escaping or biting, was significantly extended after PAP exposure. When lipopolysaccharide was Injected Intramuscularly, Cortisol levels, rectal temperatures, and lying time lengths Increased substantially. No differences were observed between the pheromone-treated and untreated groups. These results suggest that this synthetic pheromone alleviates social stress In adult pigs, although It does not affect Immune stress responses. Our findings demonstrate the potential benefit of this pheromone In field applications and clinical disciplines relating to adult female pigs.

  2. Inverse Agonist of Nuclear Receptor ERRγ Mediates Antidiabetic Effect Through Inhibition of Hepatic Gluconeogenesis

    PubMed Central

    Kim, Don-Kyu; Gang, Gil-Tae; Ryu, Dongryeol; Koh, Minseob; Kim, Yo-Na; Kim, Su Sung; Park, Jinyoung; Kim, Yong-Hoon; Sim, Taebo; Lee, In-Kyu; Choi, Cheol Soo; Park, Seung Bum; Lee, Chul-Ho; Koo, Seung-Hoi; Choi, Hueng-Sik

    2013-01-01

    Type 2 diabetes mellitus (T2DM) is a progressive metabolic disorder with diverse pathological manifestations and is often associated with abnormal regulation of hepatic glucose production. Many nuclear receptors known to control the hepatic gluconeogenic program are potential targets for the treatment of T2DM and its complications. Nevertheless, the therapeutic potential of the estrogen-related receptor γ (ERRγ) in T2DM remains unknown. In this study, we show that the nuclear receptor ERRγ is a major contributor to hyperglycemia under diabetic conditions by controlling hepatic glucose production. Hepatic ERRγ expression induced by fasting and diabetic conditions resulted in elevated levels of gluconeogenic gene expression and blood glucose in wild-type mice. Conversely, ablation of hepatic ERRγ gene expression reduced the expression of gluconeogenic genes and normalized blood glucose levels in mouse models of T2DM: db/db and diet-induced obesity (DIO) mice. In addition, a hyperinsulinemic-euglycemic clamp study and long-term studies of the antidiabetic effects of GSK5182, the ERRγ-specific inverse agonist, in db/db and DIO mice demonstrated that GSK5182 normalizes hyperglycemia mainly through inhibition of hepatic glucose production. Our findings suggest that the ability of GSK5182 to control hepatic glucose production can be used as a novel therapeutic approach for the treatment of T2DM. PMID:23775767

  3. PPAR-gamma agonists inhibit profibrotic phenotypes in human lung fibroblasts and bleomycin-induced pulmonary fibrosis.

    PubMed

    Milam, Jami E; Keshamouni, Venkateshwar G; Phan, Sem H; Hu, Biao; Gangireddy, Srinivasa R; Hogaboam, Cory M; Standiford, Theodore J; Thannickal, Victor J; Reddy, Raju C

    2008-05-01

    Pulmonary fibrosis is characterized by alterations in fibroblast phenotypes resulting in excessive extracellular matrix accumulation and anatomic remodeling. Current therapies for this condition are largely ineffective. Peroxisome proliferator-activated receptor-gamma (PPAR-gamma) is a member of the nuclear hormone receptor superfamily, the activation of which produces a number of biological effects, including alterations in metabolic and inflammatory responses. The role of PPAR-gamma as a potential therapeutic target for fibrotic lung diseases remains undefined. In the present study, we show expression of PPAR-gamma in fibroblasts obtained from normal human lungs and lungs of patients with idiopathic interstitial pneumonias. Treatment of lung fibroblasts and myofibroblasts with PPAR-gamma agonists results in inhibition of proliferative responses and induces cell cycle arrest. In addition, PPAR-gamma agonists, including a constitutively active PPAR-gamma construct (VP16-PPAR-gamma), inhibit the ability of transforming growth factor-beta1 to induce myofibroblast differentiation and collagen secretion. PPAR-gamma agonists also inhibit fibrosis in a murine model, even when administration is delayed until after the initial inflammation has largely resolved. These observations indicate that PPAR-gamma is an important regulator of fibroblast/myofibroblast activation and suggest a role for PPAR-gamma ligands as novel therapeutic agents for fibrotic lung diseases.

  4. The cannabinoid TRPA1 agonist cannabichromene inhibits nitric oxide production in macrophages and ameliorates murine colitis

    PubMed Central

    Romano, B; Borrelli, F; Fasolino, I; Capasso, R; Piscitelli, F; Cascio, MG; Pertwee, RG; Coppola, D; Vassallo, L; Orlando, P; Di Marzo, V; Izzo, AA

    2013-01-01

    Background and Purpose The non-psychotropic cannabinoid cannabichromene is known to activate the transient receptor potential ankyrin-type1 (TRPA1) and to inhibit endocannabinoid inactivation, both of which are involved in inflammatory processes. We examined here the effects of this phytocannabinoid on peritoneal macrophages and its efficacy in an experimental model of colitis. Experimental Approach Murine peritoneal macrophages were activated in vitro by LPS. Nitrite levels were measured using a fluorescent assay; inducible nitric oxide (iNOS), cyclooxygenase-2 (COX-2) and cannabinoid (CB1 and CB2) receptors were analysed by RT-PCR (and/or Western blot analysis); colitis was induced by dinitrobenzene sulphonic acid (DNBS). Endocannabinoid (anandamide and 2-arachidonoylglycerol), palmitoylethanolamide and oleoylethanolamide levels were measured by liquid chromatography-mass spectrometry. Colonic inflammation was assessed by evaluating the myeloperoxidase activity as well as by histology and immunohistochemistry. Key Results LPS caused a significant production of nitrites, associated to up-regulation of anandamide, iNOS, COX-2, CB1 receptors and down-regulation of CB2 receptors mRNA expression. Cannabichromene significantly reduced LPS-stimulated nitrite levels, and its effect was mimicked by cannabinoid receptor and TRPA1 agonists (carvacrol and cinnamaldehyde) and enhanced by CB1 receptor antagonists. LPS-induced anandamide, iNOS, COX-2 and cannabinoid receptor changes were not significantly modified by cannabichromene, which, however, increased oleoylethanolamide levels. In vivo, cannabichromene ameliorated DNBS-induced colonic inflammation, as revealed by histology, immunohistochemistry and myeloperoxidase activity. Conclusion and Implications Cannabichromene exerts anti-inflammatory actions in activated macrophages – with tonic CB1 cannabinoid signalling being negatively coupled to this effect – and ameliorates experimental murine colitis. PMID:23373571

  5. Novel selective glucocorticoid receptor agonists (SEGRAs) with a covalent warhead for long-lasting inhibition.

    PubMed

    Ryabtsova, Oksana; Joossens, Jurgen; Van Der Veken, Pieter; Vanden Berghe, Wim; Augustyns, Koen; De Winter, Hans

    2016-10-15

    The synthesis and in vitro properties of six analogues of the selective glucocorticoid receptor (GR) agonist GSK866, bearing a warhead for covalent linkage to the glucocorticoid receptor, is described.

  6. PPAR agonists stimulate adipogenesis at the expense of osteoblast differentiation while inhibiting osteoclast formation and activity.

    PubMed

    Patel, Jessal J; Butters, Oliver R; Arnett, Timothy R

    2014-06-01

    Drugs used in the treatment of type 2 diabetes and cardiovascular disease, specifically peroxisome proliferator-activated receptor (PPAR) agonists, have been reported to affect bone cell function and fracture risk. In this study, we assessed the direct effects of PPAR-γ agonists (rosiglitazone and troglitazone), used in the treatment of diabetes, and a PPAR-α agonist (fenofibrate), used to treat hyperlipidaemia, on the function of primary osteoblasts and osteoclasts. Formation of 'trabecular' bone structures by rat calvarial osteoblasts was reduced by up to 85% in cultures treated with rosiglitazone and by 45% in troglitazone-treated or fenofibrate-treated cultures; at the same time, lipid droplet formation was increased by 40-70%. The expression of key osteogenic markers was similarly downregulated in cultures treated with PPAR agonists, whereas adipogenesis markers were upregulated. Formation of osteoclasts in cultures derived from mouse marrow diminished with fenofibrate treatment, whereas both glitazones reduced resorptive activity without affecting osteoclast number. Metformin, although not a PPAR agonist, is also commonly used in the treatment of type 2 diabetes. Here, metformin was found to have no effect on bone cell function. Taken together, these data suggest that PPAR-γ agonists may enhance bone loss via increased adipogenesis at the expense of osteoblast formation. In contrast, PPAR-α agonists may prevent bone loss. Given that the prevalence of diabetes and cardiovascular disease is expected to rise significantly, greater attention may need to be paid to the effects of PPAR agonists on bone homeostasis. Copyright © 2014 John Wiley & Sons, Ltd.

  7. Omega-3 fatty acids and other FFA4 agonists inhibit growth factor signaling in human prostate cancer cells.

    PubMed

    Liu, Ze; Hopkins, Mandi M; Zhang, Zhihong; Quisenberry, Chrystal B; Fix, Louise C; Galvan, Brianna M; Meier, Kathryn E

    2015-02-01

    Omega-3 fatty acids (n-3 FAs) are proposed to have many beneficial effects on human health. However, the mechanisms underlying their potential cancer preventative effects are unclear. G protein-coupled receptors (GPCRs) of the free fatty acid receptor (FFAR) family, FFA1/GPR40 and FFA4/GPR120, specifically bind n-3 FAs as agonist ligands. In this study, we examined the effects of n-3 FAs in human prostate cancer cell lines. Initial studies established that the long-chain n-3 FAs, eicosapentaenoic acid (EPA) and docosahexaenoic acid, inhibit proliferation of DU145 cells in response to lysophosphatidic acid (LPA), a mitogenic lipid mediator. When added alone to serum-starved DU145 cells, EPA transiently activates signaling events, including p70S6K phosphorylation. However, when added 15 minutes prior to LPA, EPA suppresses LPA-induced activating phosphorylations of ERK, FAK, and p70S6K, and expression of the matricellular protein CCN1. The rapid onset of the inhibitory action of EPA suggested involvement of a GPCR. Further studies showed that DU145 and PC-3 cells express mRNA and protein for both FFA4 and FFA1. TUG-891 (4-[(4-fluoro-4'-methyl[1,1'-biphenyl]-2-yl)methoxy]-benzenepropanoic acid), a selective agonist for FFA4, exerts inhibitory effects on LPA- and epidermal growth factor-induced proliferation and migration, similar to EPA, in DU145 and PC-3 cells. The effects of TUG-891 and EPA are readily reversible. The FFA1/FFA4 agonist GW9508 (4-[[(3-phenoxyphenyl)methyl]amino]-benzenepropranoic acid) likewise inhibits proliferation at doses that block FFA4. Knockdown of FFA4 expression prevents EPA- and TUG-891-induced inhibition of growth and migration. Together, these results indicate that activation of FFA4 initiates signaling events that can inhibit growth factor-induced signaling, providing a novel mechanism for suppression of cancer cell proliferation.

  8. Marked increases in mucociliary clearance produced by synergistic secretory agonists or inhibition of the epithelial sodium channel

    PubMed Central

    Joo, Nam Soo; Jeong, Jin Hyeok; Cho, Hyung-Ju; Wine, Jeffrey J.

    2016-01-01

    Mucociliary clearance (MCC) is a critical host innate defense mechanism in airways, and it is impaired in cystic fibrosis (CF) and other obstructive lung diseases. Epithelial fluid secretion and absorption modify MCC velocity (MCCV). We tested the hypotheses that inhibiting fluid absorption accelerates MCCV, whereas inhibiting fluid secretion decelerates it. In airways, ENaC is mainly responsible for fluid absorption, while anion channels, including CFTR and Ca2+-activated chloride channels mediate anion/fluid secretion. MCCV was increased by the cAMP-elevating agonists, forskolin or isoproterenol (10 μM) and by the Ca2+-elevating agonist, carbachol (0.3 μM). The CFTR-selective inhibitor, CFTRinh-172, modestly reduced MCCV-increases induced by forskolin or isoproterenol but not increases induced by carbachol. The ENaC inhibitor benzamil increased basal MCCV as well as MCCV increases produced by forskolin or carbachol. MCC velocity was most dramatically accelerated by the synergistic combination of forskolin and carbachol, which produced near-maximal clearance rates regardless of prior treatment with CFTR or ENaC inhibitors. In CF airways, where CFTR-mediated secretion (and possibly synergistic MCC) is lost, ENaC inhibition via exogenous agents may provide therapeutic benefit, as has long been proposed. PMID:27830759

  9. Effects of an intrathecally administered benzodiazepine receptor agonist, antagonist and inverse agonist on morphine-induced inhibition of a spinal nociceptive reflex.

    PubMed Central

    Moreau, J. L.; Pieri, L.

    1988-01-01

    1. The effects of an intrathecally administered benzodiazepine receptor (BZR) agonist (midazolam, up to 50 micrograms), antagonist (flumazenil, Ro 15-1788, 5 micrograms) and inverse agonist (Ro 19-4603, 15 micrograms) on nociception and on morphine-induced antinociception were studied in rats. 2. By themselves, none of these compounds significantly altered pain threshold. 3. The BZR agonist midazolam enhanced the morphine-induced antinociceptive effect whereas the antagonist flumazenil did not alter it. In contrast, the BZR inverse agonist Ro 19-4603 decreased the morphine-induced antinociceptive effect. 4. Naloxone (1 mg kg-1 i.p.) completely reversed all these effects. 5. These results demonstrate that BZR agonists and inverse agonists are able to affect, by allosteric up- or down-modulation of gamma-aminobutyric acidA (GABAA)-receptors, the transmission of nociceptive information at the spinal cord level, when this transmission is depressed by mu-opioid receptor activation. PMID:2898960

  10. Inverse PPARβ/δ agonists suppress oncogenic signaling to the ANGPTL4 gene and inhibit cancer cell invasion

    PubMed Central

    Adhikary, T; Brandt, D T; Kaddatz, K; Stockert, J; Naruhn, S; Meissner, W; Finkernagel, F; Obert, J; Lieber, S; Scharfe, M; Jarek, M; Toth, P M; Scheer, F; Diederich, W E; Reinartz, S; Grosse, R; Müller-Brüsselbach, S; Müller, R

    2013-01-01

    Besides its established functions in intermediary metabolism and developmental processes, the nuclear receptor peroxisome proliferator-activated receptor β/δ (PPARβ/δ) has a less defined role in tumorigenesis. In the present study, we have identified a function for PPARβ/δ in cancer cell invasion. We show that two structurally divergent inhibitory ligands for PPARβ/δ, the inverse agonists ST247 and DG172, strongly inhibit the serum- and transforming growth factor β (TGFβ)-induced invasion of MDA-MB-231 human breast cancer cells into a three-dimensional matrigel matrix. To elucidate the molecular basis of this finding, we performed chromatin immunoprecipitation sequencing (ChIP-Seq) and microarray analyses, which identified the gene encoding angiopoietin-like 4 (ANGPTL4) as the major transcriptional PPARβ/δ target in MDA-MB-231 cells, previously implicated in TGFβ-mediated tumor progression and metastatic dissemination. We show that the induction of ANGPTL4 by TGFβ and other oncogenic signals is strongly repressed by ST247 and DG172 in a PPARβ/δ-dependent fashion, resulting in the inhibition of ANGPTL4 secretion. This effect is attributable to these ligands' ability to induce a dominant transcriptional repressor complex at the site of transcription initiation that blocks preinitiation complex formation through an histone deacetylase-independent, non-canonical mechanism. Repression of ANGPTL4 transcription by inverse PPARβ/δ agonists is functionally linked to the inhibition of cancer cell invasion into a three-dimensional matrix, as (i) invasion of MDA-MB-231 cells is critically dependent on ANGPTL4 expression, (ii) recombinant ANGPTL4 stimulates invasion, and (iii) reverses the inhibitory effect of ST247 and DG172. These findings indicate that a PPARβ/δ–ANGPTL4 pathway is involved in the regulation of tumor cell invasion and that its pharmacological manipulation by inverse PPARβ/δ agonists is feasible. PMID:23208498

  11. Disruption of prepulse inhibition of the startle reflex by the preferential D(3) agonist ropinirole in healthy males.

    PubMed

    Giakoumaki, Stella G; Roussos, Panos; Frangou, Sophia; Bitsios, Panos

    2007-10-01

    Emerging evidence from agonist-antagonist studies suggests a role for the dopamine D(3) receptor subtype in the regulation of PPI in animals, but such evidence is lacking for human subjects. This study examines the effect of the preferential D(3) agonist ropinirole on PPI in humans. PPI was tested in 12 healthy men in three sessions associated with ropinirole 0.25 mg, ropinirole 0.5 mg, or placebo according to a balanced, crossover, double-blind design. Two prepulses (75- and 85-dB white noise bursts) and two lead intervals (50 and 80 ms) were employed. Ropinirole 0.5 mg significantly reduced prepulse inhibition (PPI) with both prepulses at the 80-ms lead intervals. There was no effect of treatment on startle amplitude and habituation. These results suggest a role for the dopamine D(3) receptor in the mediation of human PPI, although a contribution from ropinirole's agonistic activity at the D(2) receptor cannot be entirely excluded. Firm conclusions on the role of the D(3) receptor in the modulation of human PPI can only be drawn with the use of genetic approaches or more selective ligands for this receptor.

  12. Correction for Inhibition Leads to an Allosteric Co-Agonist Model for Pentobarbital Modulation and Activation of α1β3γ2L GABAA Receptors

    PubMed Central

    Ziemba, Alexis M.; Forman, Stuart A.

    2016-01-01

    Background Pentobarbital, like propofol and etomidate, produces important general anesthetic effects through GABAA receptors. Photolabeling also indicates that pentobarbital binds to some of the same sites where propofol and etomidate act. Quantitative allosteric co-agonist models for propofol and etomidate account for modulatory and agonist effects in GABAA receptors and have proven valuable in establishing drug site characteristics and for functional analysis of mutants. We therefore sought to establish an allosteric co-agonist model for pentobarbital activation and modulation of α1β3γ2L receptors, using a novel approach to first correct pentobarbital activation data for inhibitory effects in the same concentration range. Methods Using oocyte-expressed α1β3γ2L GABAA receptors and two-microelectrode voltage-clamp, we quantified modulation of GABA responses by a low pentobarbital concentration and direct effects of high pentobarbital concentrations, the latter displaying mixed agonist and inhibitory effects. We then isolated and quantified pentobarbital inhibition in activated receptors using a novel single-sweep “notch” approach, and used these results to correct steady-state direct activation for inhibition. Results Combining results for GABA modulation and corrected direct activation, we estimated receptor open probability and optimized parameters for a Monod-Wyman-Changeux allosteric co-agonist model. Inhibition by pentobarbital was consistent with two sites with IC50s near 1 mM, while co-agonist model parameters suggest two allosteric pentobarbital agonist sites characterized by KPB ≈ 5 mM and high efficacy. The results also indicate that pentobarbital may be a more efficacious agonist than GABA. Conclusions Our novel approach to quantifying both inhibitory and co-agonist effects of pentobarbital provides a basis for future structure-function analyses of GABAA receptor mutations in putative pentobarbital binding sites. PMID:27110714

  13. The selective PAC1 receptor agonist maxadilan inhibits neurogenic vasodilation and edema formation in the mouse skin.

    PubMed

    Banki, E; Hajna, Zs; Kemeny, A; Botz, B; Nagy, P; Bolcskei, K; Toth, G; Reglodi, D; Helyes, Zs

    2014-10-01

    We have earlier shown that PACAP-38 decreases neurogenic inflammation. However, there were no data on its receptorial mechanism and the involvement of its PAC1 and VPAC1/2 receptors (PAC1R, VPAC1/2R) in this inhibitory effect. Neurogenic inflammation in the mouse ear was induced by topical application of the Transient Receptor Potential Ankyrin 1 (TRPA1) receptor activator mustard oil (MO). Consequent neurogenic edema, vasodilation and plasma leakage were assessed by measuring ear thickness with engineer's micrometer, detecting tissue perfusion by laser Doppler scanning and Evans blue or indocyanine green extravasation by intravital videomicroscopy or fluorescence imaging, respectively. Myeloperoxidase activity, an indicator of neutrophil infiltration, was measured from the ear homogenates with spectrophotometry. The selective PAC1R agonist maxadilan, the VPAC1/2R agonist vasoactive intestinal polypeptide (VIP) or the vehicle were administered i.p. 15 min before MO. Substance P (SP) concentration of the ear was assessed by radioimmunoassay. Maxadilan significantly diminished MO-induced neurogenic edema, increase of vascular permeability and vasodilation. These inhibitory effects of maxadilan may be partially due to the decreased substance P (SP) levels. In contrast, inhibitory effect of VIP on ear swelling was moderate, without any effect on MO-induced plasma leakage or SP release, however, activation of VPAC1/2R inhibited the increased microcirculation caused by the early arteriolar vasodilation. Neither the PAC1R, nor the VPAC1/2R agonist influenced the MO-evoked increase in tissue myeloperoxidase activity. These results clearly show that PAC1R activation inhibits acute neurogenic arterial vasodilation and plasma protein leakage from the venules, while VPAC1/2R stimulation is only involved in the attenuation of vasodilation. Copyright © 2014 Elsevier Ltd. All rights reserved.

  14. 5HT4 agonists inhibit interferon-gamma-induced MHC class II and B7 costimulatory molecules expression on cultured astrocytes.

    PubMed

    Zeinstra, Esther M; Wilczak, Nadine; Wilschut, Jan C; Glazenburg, Lisa; Chesik, Daniel; Kroese, Frans G M; De Keyser, Jacques

    2006-10-01

    A failure of tight control of MHC class II expression on astrocytes may play a role in the development of autoimmune responses in multiple sclerosis. The 5-HT(4) serotonin receptor agonists cisapride and prucalopride, at concentrations between 10(-10) M and 10(-8) M, reduced interferon-gamma-induced MHC class II immunostaining in cultured astrocytes derived from newborn Wistar rats by approximately 50-60%. The magnitude of MHC class II inhibition by 5-HT(4) agonists was comparable to that of interferon-beta. The alpha(1)-adrenergic receptor agonist phenylephrine was without effect. Cisapride (10(-9) M) also prevented interferon-gamma-induced B7-1 and B7-2 immunostaining. Our results suggest that 5-HT(4) agonists may have therapeutic potential in multiple sclerosis by inhibiting the up-regulation of immune responsiveness of astrocytes in the central nervous system.

  15. UV-filter benzophenone-3 inhibits agonistic behavior in male Siamese fighting fish (Betta splendens).

    PubMed

    Chen, Te-Hao; Wu, Yea-Ting; Ding, Wang-Hsien

    2016-03-01

    Benzophenone-3 (BP-3) is a widely used organic UV-filter compound. Despite the frequent occurrence of BP-3 in aquatic environments, little is known about its effect on fish behavior. The aim of this study was to investigate the endocrine disrupting effects of BP-3 in male fighting fish (Betta splendens) with a focus on agonistic behavior. Male fighting fish were exposed to 10, 100, and 1000 μg/L BP-3, as well as a solvent control (0.1% ethanol) and a positive control (100 ng/L 17α-ethynylestradiol, EE2), for 28 days. At the beginning and the end of exposure, standard length and body mass of the fish were measured for calculating the condition factor (CF). In addition, spontaneous swimming activity (total distance moved) and agonistic behavior (maximum velocity and duration of opercular display in front of a mirror) were also quantified. At the end of exposure, the fish gonads were sampled for gonadosomatic index (GSI) measurement and histology. After the exposure, CF was significantly decreased in the 1000 μg/L BP-3 groups. Spontaneous swimming activity was not affected. However, maximum velocity was significantly reduced in the EE2 and 1000 μg/L BP-3 treatments; duration of opercular display was significantly decreased in the EE2 and 10 and 1000 μg/L BP-3 treatments. GSI was not significantly different between groups. There was a slight but statistically significant decrease of relative proportion of mature spermatozoa in testicular tissue in the 100 μg/L BP-3 treatment. Collectively, our results demonstrate that BP-3 can disrupt agonistic behavior of male fighting fish, indicating the endocrine disrupting activity of this compound.

  16. Selective postsynaptic inhibition of tonic-firing neurons in substantia gelatinosa by mu-opioid agonist.

    PubMed

    Santos, Sónia F A; Melnick, Igor V; Safronov, Boris V

    2004-11-01

    Spinal substantia gelatinosa (SG) is a site of action of administered and endogenous opioid agonists and is an important element in the system of antinociception. However, little is known about the types of neurons serving as specific postsynaptic targets for opioid action within the SG. To study the spinal mechanisms of opioidergic analgesia, the authors compared the action of mu-opioid agonist [D-Ala, N-Me-Phe, Gly-ol]-enkephalin (DAMGO) on SG neurons with different intrinsic firing properties. Whole cell patch clamp recordings from spinal cord slices of Wistar rats were used to study the sensitivity of SG neurons to DAMGO. Three groups of neurons with distinct distributions in SG were classified: tonic-, adapting-, and delayed-firing neurons. DAMGO at 1 microm concentration selectively hyperpolarized all tonic-firing neurons tested, whereas none of the adapting- or delayed-firing neurons were affected. The effect of DAMGO on tonic-firing neurons was due to activation of G protein-coupled inward-rectifier K conductance, which could be blocked by 500 microm Ba and 500 microm Cs but increased by 50 microm baclofen. As a functional consequence of DAMGO action, a majority of tonic-firing neurons changed their pattern of intrinsic firing from tonic to adapting. It is suggested that tonic-firing neurons, presumably functioning as excitatory interneurons, are primary postsynaptic targets for administered and endogenous opioid agonists in spinal SG. Functional transition of cells in this group from tonic to adapting firing mode may represent an important mechanism facilitating opioidergic analgesia.

  17. Peroxisome proliferator-activated receptor-γ (PPAR-γ) agonist inhibits collagen synthesis in human hypertrophic scar fibroblasts by targeting Smad3 via miR-145

    SciTech Connect

    Zhu, Hua-Yu; Li, Chao; Zheng, Zhao; Zhou, Qin; Guan, Hao; Su, Lin-Lin; Han, Jun-Tao; Zhu, Xiong-Xiang; Wang, Shu-yue; Li, Jun Hu, Da-Hai

    2015-03-27

    The transcription factor peroxisome proliferator-activated receptor-γ (PPAR-γ) functions to regulate cell differentiation and lipid metabolism. Recently, its agonist has been documented to regulate extracellular matrix production in human dermal fibroblasts. This study explored the underlying molecular mechanisms and gene interactions in hypertrophic scar fibroblasts (HSFBs) in vitro. HSFBs were cultured and treated with or without PPAR-γ agonist or antagonist for gene expression. Bioinformatical analysis predicted that miR-145 could target Smad3 expression. Luciferase assay was used to confirm such an interaction. The data showed that PPAR-γ agonist troglitazone suppressed expression of Smad3 and Col1 in HSFBs. PPAR-γ agonist induced miR-145 at the gene transcriptional level, which in turn inhibited Smad3 expression and Col1 level in HSFBs. Furthermore, ELISA data showed that Col1 level in HSFBs was controlled by a feedback regulation mechanism involved in PPAR-γ agonist and antagonist-regulated expression of miR-145 and Smad3 in HSFBs. These findings indicate that PPAR-γ-miR-145-Smad3 axis plays a role in regulation of collagen synthesis in HSFBs. - Highlights: • PPAR-γ agonist inhibits collagen synthesis in HSFBs. • Smad3 and type I collagen expression are decreased by PPAR-γ agonist. • miR-145 expression is increased by PPAR-γ agonist in HSFBs. • Increased miR-145 inhibits collagen synthesis by targeting Smad3. • miR-145 regulates collagen synthesis.

  18. Rosiglitazone inhibits hypercholesterolaemia-induced myeloperoxidase upregulation—a novel mechanism for the cardioprotective effects of PPAR agonists

    PubMed Central

    Liu, Hui-Rong; Tao, Ling; Gao, Erhe; Qu, Yan; Lau, Wayne Bond; Lopez, Bernard L.; Christopher, Theodore A.; Koch, Walter; Yue, Tian-Li; Ma, Xin-Liang

    2009-01-01

    Aims Hypercholesterolaemia and myeloperoxidase (MPO) overexpression are two well-recognized risk factors for ischaemic heart disease. Peroxisome proliferator-activated receptor-γ (PPARγ) agonists have recently been shown to reduce ischaemic heart injury in hypercholesterolaemic animals. However, whether PPARγ agonists may exert their cardioprotective effects by eliminating those risk factors that increase ischaemic injury remains unknown. Methods and results Male New Zealand rabbits were fed with a normal or a high-cholesterol diet for 8 weeks, treated with vehicle or rosiglitazone (RSG, 3 mg/kg/day for the last 5 weeks) and subjected to myocardial ischaemia/reperfusion (1 h/4 h). MPO expression, activity, and distribution, cardiac caspase-3 activity, and myocardial infarct size were determined. Diet-induced hypercholesterolaemia caused a significant increase in neutrophil MPO expression/activity (7.2-/5.4-fold). Hypercholesterolaemia also tripled MPO activity in ischaemic/reperfused hearts when compared with rabbits fed with a normal diet. Surprisingly, MPO immunostaining was not only observed in perivascular and extracellular spaces in ischaemic/reperfused hearts, but also in cardiomyocytes. This intracardiomyocyte MPO staining was further intensified by hypercholesterolaemia. There is a strong positive correlation between cardiac MPO activity and caspase-3 activity, and treatment with an MPO inhibitor significantly reduced post-ischaemic caspase-3 activation. Treatment with RSG markedly inhibited hypercholesterolaemia-induced leucocyte MPO overexpression and activation, reduced MPO activity in ischaemic/reperfused hearts, decreased caspase-3 activity, and reduced myocardial infarct size (P < 0.01). Conclusion Our results demonstrated that hypercholesterolaemia and MPO overexpression are causally related and that PPARγ agonists may have great therapeutic value in ischaemic heart disease patients with multiple complications such as hypercholesterolaemia and

  19. Rosiglitazone inhibits hypercholesterolaemia-induced myeloperoxidase upregulation--a novel mechanism for the cardioprotective effects of PPAR agonists.

    PubMed

    Liu, Hui-Rong; Tao, Ling; Gao, Erhe; Qu, Yan; Lau, Wayne Bond; Lopez, Bernard L; Christopher, Theodore A; Koch, Walter; Yue, Tian-Li; Ma, Xin-Liang

    2009-02-01

    Hypercholesterolaemia and myeloperoxidase (MPO) overexpression are two well-recognized risk factors for ischaemic heart disease. Peroxisome proliferator-activated receptor-gamma (PPARgamma) agonists have recently been shown to reduce ischaemic heart injury in hypercholesterolaemic animals. However, whether PPARgamma agonists may exert their cardioprotective effects by eliminating those risk factors that increase ischaemic injury remains unknown. Male New Zealand rabbits were fed with a normal or a high-cholesterol diet for 8 weeks, treated with vehicle or rosiglitazone (RSG, 3 mg/kg/day for the last 5 weeks) and subjected to myocardial ischaemia/reperfusion (1 h/4 h). MPO expression, activity, and distribution, cardiac caspase-3 activity, and myocardial infarct size were determined. Diet-induced hypercholesterolaemia caused a significant increase in neutrophil MPO expression/activity (7.2-/5.4-fold). Hypercholesterolaemia also tripled MPO activity in ischaemic/reperfused hearts when compared with rabbits fed with a normal diet. Surprisingly, MPO immunostaining was not only observed in perivascular and extracellular spaces in ischaemic/reperfused hearts, but also in cardiomyocytes. This intracardiomyocyte MPO staining was further intensified by hypercholesterolaemia. There is a strong positive correlation between cardiac MPO activity and caspase-3 activity, and treatment with an MPO inhibitor significantly reduced post-ischaemic caspase-3 activation. Treatment with RSG markedly inhibited hypercholesterolaemia-induced leucocyte MPO overexpression and activation, reduced MPO activity in ischaemic/reperfused hearts, decreased caspase-3 activity, and reduced myocardial infarct size (P < 0.01). Our results demonstrated that hypercholesterolaemia and MPO overexpression are causally related and that PPARgamma agonists may have great therapeutic value in ischaemic heart disease patients with multiple complications such as hypercholesterolaemia and diabetes.

  20. Inhibition of indoleamine 2,3-dioxygenase activity enhances the anti-tumour effects of a Toll-like receptor 7 agonist in an established cancer model.

    PubMed

    Ito, Hiroyasu; Ando, Tatsuya; Arioka, Yuko; Saito, Kuniaki; Seishima, Mitsuru

    2015-04-01

    Toll-like receptor (TLR) agonists have been shown to have anti-tumour activity in basic research and clinical studies. However, TLR agonist monotherapy does not sufficiently eliminate tumours. Activation of the innate immune response by TLR agonists is effective at driving adaptive immunity via interleukin-12 (IL-12) or IL-1, but is counteracted by the simultaneous induction of immunosuppressive cytokines and other molecules, including IL-10, transforming growth factor-β, and indoleamine 2,3-dioxygenase (IDO). In the present study, we evaluated the anti-cancer effect of the TLR7 agonist, imiquimod (IMQ), in the absence of IDO activity. The administration of IMQ in IDO knockout (KO) mice inoculated with tumour cells significantly suppressed tumour progression compared with that in wild-type (WT) mice, and improved the survival rate. Moreover, injection with IMQ enhanced the tumour antigen-specific T helper type 1 response in IDO-KO mice with tumours. Combination therapy with IMQ and an IDO inhibitor also significantly inhibited tumour growth. Our results indicated that the enhancement of IDO expression with TLR agonists in cancer treatment might impair host anti-tumour immunity while the inhibition of IDO could enhance the therapeutic efficacy of TLR agonists via the increase of T helper type 1 immune response. © 2014 John Wiley & Sons Ltd.

  1. Physiological evidence of a postsynaptic inhibition of the tail flick reflex by a cannabinoid receptor agonist

    PubMed Central

    Dableh, Liliane J.; Yashpal, Kiran; Henry, James L.

    2016-01-01

    Current evidence indicates that cannabinoids are antinociceptive and this effect is in part mediated by spinal mechanisms. Anatomical studies have localized cannabinoid CB1 receptors to pre- and postsynaptic sites within the spinal cord. However, behavioural tests have not clearly indicated the relative importance of each of these sites. In this study, the tail flick test was used as a model of acute pain in the rat to determine the site of action of WIN 55,212-2 ((R)-(+)-[2,3-dihydro-5-methyl-3[(4-morpholinyl)methyl]pyrolo[1,2,3-de]-1,4-benzoxazinyl]-(1-naphthalenyl)methanone mesylate), a synthetic cannabinoid receptor agonist. WIN 55,212-2 (3 mg/kg, i.p.) increased the latency of tail withdrawal from a noxious radiant heat source, indicating it is antinociceptive in this model. Using the same paradigm, WIN 55,212-2 was then tested against the synaptically-induced nociceptive hypersensitivity in response to noxious thermal stimulation of the tail (hot water tail immersion). WIN 55,212-2 blocked this hypersensitivity, confirming a spinal site of action of the cannabinoid receptor agonist. Further, WIN 55,212-2 blocked the nociceptive hypersensitivity induced by intrathecal administration of substance P. As substance P acts on postsynaptic tachykinin NK1 receptors in the dorsal horn of the spinal cord, the data are interpreted to suggest that WIN 55,212-2 expressed its anti-hypersensitivity effects at least partially via a postsynaptic site of action; the data do not rule out a presynaptic site of action. This study suggests that cannabinoids may induce analgesia via a postsynaptic site of action in the spinal cord, as well as the possibility that they may interact with substance P signaling. PMID:19027734

  2. Systems Analysis of a RIG-I Agonist Inducing Broad Spectrum Inhibition of Virus Infectivity

    PubMed Central

    Goulet, Marie-Line; Olagnier, David; Xu, Zhengyun; Paz, Suzanne; Belgnaoui, S. Mehdi; Lafferty, Erin I.; Janelle, Valérie; Arguello, Meztli; Paquet, Marilene; Ghneim, Khader; Richards, Stephanie; Smith, Andrew; Wilkinson, Peter; Cameron, Mark; Kalinke, Ulrich; Qureshi, Salman; Lamarre, Alain; Haddad, Elias K.; Sekaly, Rafick Pierre; Peri, Suraj; Balachandran, Siddharth; Lin, Rongtuan; Hiscott, John

    2013-01-01

    The RIG-I like receptor pathway is stimulated during RNA virus infection by interaction between cytosolic RIG-I and viral RNA structures that contain short hairpin dsRNA and 5′ triphosphate (5′ppp) terminal structure. In the present study, an RNA agonist of RIG-I was synthesized in vitro and shown to stimulate RIG-I-dependent antiviral responses at concentrations in the picomolar range. In human lung epithelial A549 cells, 5′pppRNA specifically stimulated multiple parameters of the innate antiviral response, including IRF3, IRF7 and STAT1 activation, and induction of inflammatory and interferon stimulated genes - hallmarks of a fully functional antiviral response. Evaluation of the magnitude and duration of gene expression by transcriptional profiling identified a robust, sustained and diversified antiviral and inflammatory response characterized by enhanced pathogen recognition and interferon (IFN) signaling. Bioinformatics analysis further identified a transcriptional signature uniquely induced by 5′pppRNA, and not by IFNα-2b, that included a constellation of IRF7 and NF-kB target genes capable of mobilizing multiple arms of the innate and adaptive immune response. Treatment of primary PBMCs or lung epithelial A549 cells with 5′pppRNA provided significant protection against a spectrum of RNA and DNA viruses. In C57Bl/6 mice, intravenous administration of 5′pppRNA protected animals from a lethal challenge with H1N1 Influenza, reduced virus titers in mouse lungs and protected animals from virus-induced pneumonia. Strikingly, the RIG-I-specific transcriptional response afforded partial protection from influenza challenge, even in the absence of type I interferon signaling. This systems approach provides transcriptional, biochemical, and in vivo analysis of the antiviral efficacy of 5′pppRNA and highlights the therapeutic potential associated with the use of RIG-I agonists as broad spectrum antiviral agents. PMID:23633948

  3. Anandamide (arachidonylethanolamide), a brain cannabinoid receptor agonist, reduces sperm fertilizing capacity in sea urchins by inhibiting the acrosome reaction.

    PubMed Central

    Schuel, H; Goldstein, E; Mechoulam, R; Zimmerman, A M; Zimmerman, S

    1994-01-01

    Anandamide (arachidonylethanolamide) is an endogenous cannabinoid receptor agonist in mammalian brain. Sea urchin sperm contain a high-affinity cannabinoid receptor similar to the cannabinoid receptor in mammalian brain. (-)-delta 9-Tetrahydrocannabinol (THC), the primary psychoactive cannabinoid in marihuana, reduces the fertilizing capacity of sea urchin sperm by blocking the acrosome reaction that normally is stimulated by a specific ligand in the egg's jelly coat. We now report that anandamide produces effects similar to those previously obtained with THC in Strongylocentrotus purpuratus in reducing sperm fertilizing capacity and inhibiting the egg jelly-stimulated acrosome reaction. Arachidonic acid does not inhibit the acrosome reaction under similar conditions. The adverse effects of anandamide on sperm fertilizing capacity and the acrosome reaction are reversible. The receptivity of unfertilized eggs to sperm and sperm motility are not impaired by anandamide. Under conditions where anandamide completely blocks the egg jelly-stimulated acrosome reaction, it does not inhibit the acrosome reaction artificially initiated by ionomycin, which promotes Ca2+ influx, and nigericin, which activates K+ channels in sperm. These findings provide additional evidence that the cannabinoid receptor in sperm plays a role in blocking the acrosome reaction, indicate that anandamide or a related molecule may be the natural ligand for the cannabinoid receptor in sea urchin sperm, and suggest that binding of anandamide to the cannabinoid receptor modulates stimulus-secretion-coupling in sperm by affecting an event prior to ion channel opening. PMID:8052642

  4. A novel PPARα agonist propane-2-sulfonic acid octadec-9-enyl-amide inhibits inflammation in THP-1 cells.

    PubMed

    Zhao, Yun; Yan, Lu; Luo, Xiu-Mei; Peng, Lu; Guo, Han; Jing, Zuo; Yang, Li-Chao; Hu, Rong; Wang, Xuan; Huang, Xue-Feng; Wang, Yi-Qing; Jin, Xin

    2016-10-05

    Our group synthesized propane-2-sulfonic acid octadec-9-enyl-amide (N15), a novel peroxisome proliferator activated receptor alpha (PPARα) agonist. Because PPARα activation is associated with inflammation control, we hypothesize that N15 may have anti-inflammatory effects. We investigated the effect of N15 on the regulation of inflammation in THP-1 cells stimulated with lipopolysaccharide (LPS). In particular, we assessed the production of chemokines, adhesion molecules and proinflammatory cytokines, three important types of cytokines that are released from monocytes and are involved in the development of atherosclerosis. The results showed that N15 remarkably reduced the mRNA expression of chemokines, such as monocyte chemotactic protein 1 (MCP-1 or CCL2), interleukin-8 (IL-8) and interferon-inducible protein-10 (IP-10 or CXCL10), and proinflammatory cytokines, such as tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6). N15 also decreased the protein expression of vascular cell adhesion molecule (VCAM) and matrix metalloproteinase (MMP) 2 and 9. The reduction in the expression of cytokine mRNAs observed following N15 treatment was abrogated in THP-1 cells treated with PPARα siRNA, indicating that the anti-inflammatory effects of N15 are dependent on PPARα activation. Toll-like receptor 4 (TLR4)/nuclear factor-κB (NF-κB) and signal transducer and activator of transcription 3 (STAT3) inhibition, which are dependent on PPARα activation, were also involved in the mechanism underlying the anti-inflammatory effects of N15. In conclusion, the novel PPARα agonist, N15, exerts notable anti-inflammatory effects, which are mediated via PPARα activation and TLR4/NF-κB and STAT3 inhibition, in LPS-stimulated THP-1 cells. In our study, N15 exhibits promise for the treatment of atherosclerosis.

  5. Peroxisome proliferator-activated receptor-{gamma} agonists inhibit the replication of respiratory syncytial virus (RSV) in human lung epithelial cells

    SciTech Connect

    Arnold, Ralf . E-mail: ralf.arnold@medizin.uni-magdeburg.de; Koenig, Wolfgang

    2006-07-05

    We have previously shown that peroxisome proliferator-activated receptor-{gamma} (PPAR{gamma}) agonists inhibited the inflammatory response of RSV-infected human lung epithelial cells. In this study, we supply evidence that specific PPAR{gamma} agonists (15d-PGJ{sub 2}, ciglitazone, troglitazone, Fmoc-Leu) efficiently blocked the RSV-induced cytotoxicity and development of syncytia in tissue culture (A549, HEp-2). All PPAR{gamma} agonists under study markedly inhibited the cell surface expression of the viral G and F protein on RSV-infected A549 cells. This was paralleled by a reduced cellular amount of N protein-encoding mRNA determined by real-time RT-PCR. Concomitantly, a reduced release of infectious progeny virus into the cell supernatants of human lung epithelial cells (A549, normal human bronchial epithelial cells (NHBE)) was observed. Similar results were obtained regardless whether PPAR{gamma} agonists were added prior to RSV infection or thereafter, suggesting that the agonists inhibited viral gene expression and not the primary adhesion or fusion process.

  6. Denatonium and 6-n-Propyl-2-thiouracil, Agonists of Bitter Taste Receptor, Inhibit Contraction of Various Types of Smooth Muscles in the Rat and Mouse.

    PubMed

    Sakai, Hiroyasu; Sato, Ken; Kai, Yuki; Chiba, Yoshihiko; Narita, Minoru

    2016-01-01

    Recently the global expression of taste 2 receptors (TAS2Rs) on smooth muscle cells in human airways was demonstrated. Here, the effects of agonists of taste receptor, type 2, denatonium and 6-n-propyl-2-thiouracil, on smooth-muscle contraction were examined in the rat and mouse. Contractions induced by carbachol (CCh), high K(+), and sodium fluoride, but not calyculin-A, were inhibited significantly in the presence of a TAS2R agonist in the bronchial smooth muscle of mice. The contraction induced by CCh was inhibited by TAS2R agonists in ileal smooth muscle. Phenylephrine-induced contraction was also inhibited by TAS2R agonists in aortic smooth muscle. Gastrointestinal motility and blood pressure were attenuated by administration of TAS2R agonists in vivo. These findings suggest that TAS2R may be receptor for endogenous biologically active substances as well as for bitter tastes on the tongue. TAS2R signaling could be employed in the development of anti-asthmatic, anti-spasmodic, and anti-hypertensive drugs.

  7. Denatonium and 6-n-Propyl-2-thiouracil, Agonists of Bitter Taste Receptor, Inhibit Contraction of Various Types of Smooth Muscles in the Rat and Mouse.

    PubMed

    Sakai, Hiroyasu; Sato, Ken; Kai, Yuki; Chiba, Yoshihiko; Narita, Minoru

    2016-01-01

    Recently the global expression of taste 2 receptors (TAS2Rs) on smooth muscle cells in human airways was demonstrated. Here, the effects of agonists of taste receptor, type 2, denatonium and 6-n-propyl-2-thiouracil, on smooth-muscle contraction were examined in the rat and mouse. Contractions induced by carbachol (CCh), high K, and sodium fluoride, but not calyculin-A, were inhibited significantly in the presence of a TAS2R agonist in the bronchial smooth muscle of mice. The contraction induced by CCh was inhibited by TAS2R agonists in ileal smooth muscle. Phenylephrine-induced contraction was also inhibited by TAS2R agonists in aortic smooth muscle. Gastrointestinal motility and blood pressure were attenuated by administration of TAS2R agonists in vivo. These findings suggest that TAS2R may be receptor for endogenous biologically active substances as well as for bitter tastes on the tongue. TAS2R signaling could be employed in the development of anti-asthmatic, anti-spasmodic, and anti-hypertensive drugs.

  8. Beryllium competitively inhibits brain myo-inositol monophosphatase, but unlike lithium does not enhance agonist-induced inositol phosphate accumulation.

    PubMed Central

    Faraci, W S; Zorn, S H; Bakker, A V; Jackson, E; Pratt, K

    1993-01-01

    Despite limiting side-effects, lithium is the drug of choice for the treatment of bipolar depression. Its action may be due, in part, to its ability to dampen phosphatidylinositol turnover by inhibiting myo-inositol monophosphatase. Beryllium has been identified as a potent inhibitor of partially purified myo-inositol monophosphatase isolated from rat brain (Ki = 150 nM), bovine brain (Ki = 35 nM), and from the human neuroblastoma cell line SK-N-SH (Ki = 85 nM). It is over three orders of magnitude more potent than LiCl (Ki = 0.5-1.2 mM). Kinetic analysis reveals that beryllium is a competitive inhibitor of myo-inositol monophosphatase, in contrast with lithium which is an uncompetitive inhibitor. Inhibition of exogenous [3H]inositol phosphate hydrolysis by beryllium (IC50 = 250-300 nM) was observed to the same maximal extent as that seen with lithium in permeabilized SK-N-SH cells, reflecting inhibition of cellular myo-inositol monophosphatase. However, in contrast with that observed with lithium, agonist-induced accumulation of inositol phosphate was not observed with beryllium in permeabilized and non-permeabilized SK-N-SH cells and in rat brain slices. Similar results were obtained in permeabilized SK-N-SH cells when GTP-gamma-S was used as an alternative stimulator of inositol phosphate accumulation. The disparity in the actions of beryllium and lithium suggest that either (1) selective inhibition of myo-inositol monophosphatase does not completely explain the action of lithium on the phosphatidylinositol cycle, or (2) that uncompetitive inhibition of myo-inositol monophosphatase is a necessary requirement to observe functional lithium mimetic activity. PMID:8387266

  9. Combination of roflumilast with a beta-2 adrenergic receptor agonist inhibits proinflammatory and profibrotic mediator release from human lung fibroblasts

    PubMed Central

    2012-01-01

    Background Small airway narrowing is an important pathology which impacts lung function in chronic obstructive pulmonary disease (COPD). The accumulation of fibroblasts and myofibroblasts contribute to inflammation, remodeling and fibrosis by production and release of mediators such as cytokines, profibrotic factors and extracellular matrix proteins. This study investigated the effects of the phosphodiesterase 4 inhibitor roflumilast, combined with the long acting β2 adrenergic agonist indacaterol, both approved therapeutics for COPD, on fibroblast functions that contribute to inflammation and airway fibrosis. Methods The effects of roflumilast and indacaterol treatment were characterized on transforming growth factor β1 (TGFβ1)-treated normal human lung fibroblasts (NHLF). NHLF were evaluated for expression of the profibrotic mediators endothelin-1 (ET-1) and connective tissue growth factor (CTGF), expression of the myofibroblast marker alpha smooth muscle actin, and fibronectin (FN) secretion. Tumor necrosis factor-α (TNF-α) was used to induce secretion of chemokine C-X-C motif ligand 10 (CXCL10), chemokine C-C motif ligand 5 (CCL5) and granulocyte macrophage colony-stimulating factor (GM-CSF) from NHLF and drug inhibition was assessed. Results Evaluation of roflumilast (1-10 μM) showed no significant inhibition alone on TGFβ1-induced ET-1 and CTGF mRNA transcripts, ET-1 and FN protein production, alpha smooth muscle expression, or TNF-α-induced secretion of CXCL10, CCL5 and GM-CSF. A concentration-dependent inhibition of ET-1 and CTGF was shown with indacaterol treatment, and a submaximal concentration was chosen for combination studies. When indacaterol (0.1 nM) was added to roflumilast, significant inhibition was seen on all inflammatory and fibrotic mediators evaluated, which was superior to the inhibition seen with either drug alone. Roflumilast plus indacaterol combination treatment resulted in significantly elevated phosphorylation of the

  10. PPARγ Agonists as an Anti-Inflammatory Treatment Inhibiting Rotavirus Infection of Small Intestinal Villi

    PubMed Central

    Gómez, Dory; Muñoz, Natalia; Guerrero, Rafael; Acosta, Orlando; Guerrero, Carlos A.

    2016-01-01

    Rotavirus infection has been reported to induce an inflammatory response in the host cell accompanied by the increased expression or activation of some cellular molecules including ROS, NF-κB, and COX-2. PPARγ stimulation and N-acetylcysteine (NAC) treatment have been found to interfere with viral infections including rotavirus infection. Small intestinal villi isolated from in vivo infected mice with rotavirus ECwt were analyzed for the percentage of ECwt-infected cells, the presence of rotavirus antigens, and infectious virion yield following treatment with pioglitazone. Isolated villi were also infected in vitro and treated with PPARγ agonists (PGZ, TZD, RGZ, DHA, and ALA), all-trans retinoic acid (ATRA), and NAC. After treatments, the expression of cellular proteins including PPARγ, NF-κB, PDI, Hsc70, and COX-2 was analyzed using immunochemistry, ELISA, immunofluorescence, and Western blotting. The results showed that rotavirus infection led to an increased accumulation of the cellular proteins studied and ROS. The virus infection-induced accumulation of the cellular proteins studied and ROS was reduced upon pioglitazone treatment, causing also a concomitant reduction of the infectious virion yield. We hypothesized that rotavirus infection is benefiting from the induction of a host cell proinflammatory response and that the interference of the inflammatory pathways involved leads to decreased infection. PMID:27382365

  11. A 5-HT(1B) receptor agonist inhibits light-induced suppression of pineal melatonin production.

    PubMed

    Rea, M A; Pickard, G E

    2000-03-10

    Serotonin (5-HT) modulates the phase adjusting effects of light on the mammalian circadian clock through the activation of presynaptic 5-HT(1B) receptors located on retinal terminals in the suprachiasmatic nucleus (SCN). The current study was conducted to determine whether activation of 5-HT(1B) receptors also alters photic regulation of nocturnal pineal melatonin production. Systemic administration of the 5-HT(1B) receptor agonist TFMPP attenuated the inhibitory effect of light on pineal melatonin synthesis in a dose-related manner with an apparent ED(50) value of 0.9 mg/kg. The effect of TFMPP on light-induced melatonin suppression was blocked by the 5-HT(1) receptor antagonist, methiothepin, but not by the 5-HT(1A) antagonist, WAY 100,635, consistent with the involvement of 5-HT(1B) receptors. The results are consistent with the interpretation that activation of presynaptic 5-HT(1B) receptors on retinal terminals in the SCN attenuates the effect of light on pineal melatonin production, as well as on circadian phase.

  12. Zebrafish Cardiotoxicity: The Effects of CYP1A Inhibition and AHR2 Knockdown Following Exposure to Weak Aryl Hydrocarbon Receptor Agonists

    PubMed Central

    Clark, Bryan William; Van Tiem Garner, Lindsey; Di Giulio, Richard Thomas

    2014-01-01

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that mediates many of the toxic effects of dioxin-like compounds (DLCs) and some polycyclic aromatic hydrocarbons (PAHs). Strong AHR agonists, such as certain polychlorinated biphenyls and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), cause severe cardiac teratogenesis in fish embryos. Moderately strong AHR agonists, such as benzo[a]pyrene and β-naphthoflavone, have been shown to cause similar cardiotoxic effects when coupled with a cytochrome P450 1A (CYP1A) inhibitor, such as fluoranthene (FL). We sought to determine if weak AHR agonists, when combined with a CYP1A inhibitor (FL) or CYP1A morpholino gene knockdown, are capable of causing cardiac deformities similar to moderately strong AHR agonists (Wassenberg and Di Giulio 2004; Wassenberg and Di Giulio 2004; Billiard, Timme-Laragy et al. 2006; Van Tiem and Di Giulio 2011). The weak AHR agonists included the following: carbaryl, phenanthrene, 2-methylindole, 3-methylindole, indigo, and indirubin. The results showed a complex pattern of cardiotoxic response to weak agonist inhibitor exposure and morpholino-knockdown. Danio rerio (zebrafish) embryos were first exposed to weak AHR agonists at equimolar concentrations. The agonists were assessed for their relative potency as inducers of CYP1 enzyme activity, measured by the ethoxyresorufin-o-deethylase (EROD) assay, and cardiac deformities. Carbaryl, 2-methylindole, and 3-methylindole induced the highest CYP1A activity in zebrafish. Experiments were then conducted to determine the individual cardiotoxicity of each compound. Next, zebrafish were co-exposed to each agonist (at concentrations below those determined to be cardiotoxic) and FL in combination to assess if CYP1A inhibition could induce cardiac deformities. Carbaryl, 2-methylindole, 3-methylindole, and phenanthrene significantly increased pericardial edema relative to controls when combined with FL. To further evaluate the

  13. Zebrafish cardiotoxicity: the effects of CYP1A inhibition and AHR2 knockdown following exposure to weak aryl hydrocarbon receptor agonists.

    PubMed

    Brown, Daniel R; Clark, Bryan W; Garner, Lindsey V T; Di Giulio, Richard T

    2015-06-01

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that mediates many of the toxic effects of dioxin-like compounds (DLCs) and some polycyclic aromatic hydrocarbons (PAHs). Strong AHR agonists, such as certain polychlorinated biphenyls and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), cause severe cardiac teratogenesis in fish embryos. Moderately strong AHR agonists, such as benzo[a]pyrene and β-naphthoflavone, have been shown to cause similar cardiotoxic effects when coupled with a cytochrome P450 1A (CYP1A) inhibitor, such as fluoranthene (FL). We sought to determine if weak AHR agonists, when combined with a CYP1A inhibitor (FL) or CYP1A morpholino gene knockdown, are capable of causing cardiac deformities similar to moderately strong AHR agonists (Wassenberg and Di Giulio Environ Health Perspect 112(17):1658-1664, 2004a; Wassenberg and Di Giulio Res 58(2-5):163-168, 2004b; Billiard et al. Toxicol Sci 92(2):526-536, 2006; Van Tiem and Di Giulio Toxicol Appl Pharmacol 254(3):280-287, 2011). The weak AHR agonists included the following: carbaryl, phenanthrene, 2-methylindole, 3-methylindole, indigo, and indirubin. Danio rerio (zebrafish) embryos were first exposed to weak AHR agonists at equimolar concentrations. The agonists were assessed for their relative potency as inducers of CYP1 enzyme activity, measured by the ethoxyresorufin-O-deethylase (EROD) assay, and cardiac deformities. Carbaryl, 2-methylindole, and 3-methylindole induced the highest CYP1A activity in zebrafish. Experiments were then conducted to determine the individual cardiotoxicity of each compound. Next, zebrafish were coexposed to each agonist (at concentrations below those determined to be cardiotoxic) and FL in combination to assess if CYP1A inhibition could induce cardiac deformities. Carbaryl, 2-methylindole, 3-methylindole, and phenanthrene significantly increased pericardial edema relative to controls when combined with FL. To further evaluate the interaction

  14. The PPARalpha Agonist Fenofibrate Preserves Hippocampal Neurogenesis and Inhibits Microglial Activation After Whole-Brain Irradiation

    SciTech Connect

    Ramanan, Sriram; Kooshki, Mitra; Zhao Weiling; Hsu, F.-C.; Riddle, David R.; Robbins, Mike E.

    2009-11-01

    Purpose: Whole-brain irradiation (WBI) leads to cognitive impairment months to years after radiation. Numerous studies suggest that decreased hippocampal neurogenesis and microglial activation are involved in the pathogenesis of WBI-induced brain injury. The goal of this study was to investigate whether administration of the peroxisomal proliferator-activated receptor (PPAR) alpha agonist fenofibrate would prevent the detrimental effect of WBI on hippocampal neurogenesis. Methods and Materials: For this study, 129S1/SvImJ wild-type and PPARalpha knockout mice that were fed either regular or 0.2% wt/wt fenofibrate-containing chow received either sham irradiation or WBI (10-Gy single dose of {sup 137}Cs gamma-rays). Mice were injected intraperitoneally with bromodeoxyuridine to label the surviving cells at 1 month after WBI, and the newborn neurons were counted at 2 months after WBI by use of bromodeoxyuridine/neuronal nuclei double immunofluorescence. Proliferation in the subgranular zone and microglial activation were measured at 1 week and 2 months after WBI by use of Ki-67 and CD68 immunohistochemistry, respectively. Results: Whole-brain irradiation led to a significant decrease in the number of newborn hippocampal neurons 2 months after it was performed. Fenofibrate prevented this decrease by promoting the survival of newborn cells in the dentate gyrus. In addition, fenofibrate treatment was associated with decreased microglial activation in the dentate gyrus after WBI. The neuroprotective effects of fenofibrate were abolished in the knockout mice, indicating a PPARalpha-dependent mechanism or mechanisms. Conclusions: These data highlight a novel role for PPARalpha ligands in improving neurogenesis after WBI and offer the promise of improving the quality of life for brain cancer patients receiving radiotherapy.

  15. L-stepholidine, a natural dopamine receptor D1 agonist and D2 antagonist, inhibits heroin-induced reinstatement.

    PubMed

    Ma, Baomiao; Yue, Kai; Chen, Lin; Tian, Xiang; Ru, Qin; Gan, Yongping; Wang, Daisong; Jin, Guozhang; Li, Chaoying

    2014-01-24

    L-Stepholidine (l-SPD), an alkaloid extract of the Chinese herb Stephania intermedia, is the first compound known to exhibit mixed dopamine D1 receptor agonist/D2 antagonist properties and is a potential medication for the treatment of opiate addiction. The aim of the present study was to investigate the effects of pretreatment with L-SPD on heroin-seeking behavior induced by heroin priming. Male Sprague-Dawley rats were trained to self-administer heroin (0.05mg/kg per infusion) under a fixed ratio 1 schedule for 12 consecutive days and nose-poke responding was extinguished for 12 days, after which reinstatement of drug seeking was induced by heroin priming. Pretreatment with L-SPD (2.5, 5.0 and 10.0mg/kg, i.p.) inhibited the heroin-induced reinstatement of heroin-seeking behavior. Importantly, L-SPD did not affect locomotion, indicating that the observed effects of L-SPD on reinstatement are not the result of motor impairments. The present data suggested that l-SPD inhibits heroin-induced reinstatement and its potential for the treatment of heroin relapse.

  16. Peroxisome proliferator-activated receptor δ agonists inhibit T helper type 1 (Th1) and Th17 responses in experimental allergic encephalomyelitis

    PubMed Central

    Kanakasabai, Saravanan; Chearwae, Wanida; Walline, Crystal C; Iams, Wade; Adams, Suzanne M; Bright, John J

    2010-01-01

    Multiple sclerosis (MS) is a neurological disorder that affects more than a million people world-wide. The aetiology of MS is not known and there is no medical treatment available that can cure MS. Experimental autoimmune encephalomyelitis (EAE) is a T-cell-mediated autoimmune disease model of MS. The pathogenesis of EAE/MS is a complex process involving activation of immune cells, secretion of inflammatory cytokines and destruction of myelin sheath in the central nervous system (CNS). Peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptor transcription factors that regulate cell growth, differentiation and homeostasis. PPAR agonists have been used in the treatment of obesity, diabetes, cancer and inflammation. We and others have shown that PPARγ, α and δ agonists inhibit CNS inflammation and demyelination in the EAE model of MS. In this study we show that the PPARδ agonists GW501516 and L165041 ameliorate MOGp35-55-induced EAE in C57BL/6 mice by blocking interferon (IFN)-γ and interleukin (IL)-17 production by T helper type 1 (Th1) and Th17 cells. The inhibition of EAE by PPARδ agonists was also associated with a decrease in IL-12 and IL-23 and an increase in IL-4 and IL-10 expression in the CNS and lymphoid organs. These findings indicate that PPARδ agonists modulate Th1 and Th17 responses in EAE and suggest their use in the treatment of MS and other autoimmune diseases. PMID:20406305

  17. Diabetes decreases sensitivity of adipocyte lipolysis to inhibition by Gi-linked receptor agonists.

    PubMed

    Saggerson, D; Orford, M; Chatzipanteli, K; Shepherd, J

    1991-01-01

    (1) Streptozotocin-diabetes decreased the responsiveness of noradrenaline- or forskolin-stimulated lipolysis to inhibition by phenylisopropyladenosine (PIA), prostaglandin E1 (PGE1) and nicotinate in rat adipocytes. (2) Diabetes had no effect on high affinity binding of [3H]PIA to adipocyte plasma membranes. (3) Plasma membranes from diabetic animals had increased abundance of alpha-subunits of Gi1 and Gi2. The effect of pertussis toxin in overcoming inhibition of lipolysis by PIA was delayed in adipocytes from diabetic rats. (4) Diabetes decreased the GTP-dependent right-wards shift in the dose-curve for displacement of the antagonist [3H]DPCPX by PIA in adipocyte plasma membranes. (5) It is concluded that, despite increased abundance of Gi in diabetic adipocytes, less of this is functional. This may contribute to reduced sensitivity to PIA, PGE1 and nicotinate and explains some of the loss of control of lipolysis in insulin-dependent diabetes.

  18. Inhibition of adult liver progenitor (oval) cell growth and viability by an agonist of the peroxisome proliferator activated receptor (PPAR) family member gamma, but not alpha or delta.

    PubMed

    Knight, Belinda; Yeap, Bu B; Yeoh, George C; Olynyk, John K

    2005-10-01

    Multifaceted evidence links the development of liver tumours to the activation and proliferation of adult liver progenitor (oval) cells during the early stages of chronic liver injury. The aim of this study was to examine the role of the peroxisome proliferator activated receptors (PPARs): PPARalpha, delta and gamma, in mediating the behaviour of liver progenitor cells during pre-neoplastic disease and to investigate their potential as therapeutic targets for the treatment of chronic liver injury. We observed increased liver expression of PPARalpha and gamma in concert with expanding oval cell numbers during the first 21 days following commencement of the choline deficient, ethionine supplemented (CDE) dietary model of carcinogenic liver injury in mice. Both primary and immortalized liver progenitor cells were found to express PPARalpha, delta and gamma, but not gamma2, the alternate splice form of PPARgamma. WY14643 (PPARalpha agonist), GW501516 (PPARdelta agonist) and ciglitazone (PPARgamma agonist) were tested for their ability to modulate the behaviour of p53-immortalized liver (PIL) progenitor cell lines in vitro. Both PPARdelta and gamma agonists induced dose-dependent growth inhibition and apoptosis of PIL cells. In contrast, the PPARalpha agonist had no effect on PIL cell growth. None of the drugs affected the maturation of PIL cells along either the hepatocytic or biliary lineages, as judged by their patterns of hepatic gene expression prior to and following treatment. Administration of the PPARgamma agonist ciglitazone to mice fed with the CDE diet for 14 days resulted in a significantly diminished oval cell response and decreased fibrosis compared with those receiving placebo. In contrast, GW501516 did not affect oval cell numbers or liver fibrosis, but inhibited CDE-induced hepatic steatosis. In summary, PPARgamma agonists reduce oval cell proliferation and fibrosis during chronic liver injury and may be useful in the prevention of hepatocellular

  19. Adaptations in corticospinal excitability and inhibition are not spatially confined to the agonist muscle following strength training.

    PubMed

    Mason, Joel; Frazer, Ashlyn; Horvath, Deanna M; Pearce, Alan J; Avela, Janne; Howatson, Glyn; Kidgell, Dawson

    2017-07-01

    We used transcranial magnetic stimulation (TMS) to determine the corticospinal responses from an agonist and synergist muscle following strength training of the right elbow flexors. Motor-evoked potentials were recorded from the biceps brachii and flexor carpi radialis during a submaximal contraction from 20 individuals (10 women, 10 men, aged 18-35 years; training group; n = 10 and control group; n = 10) before and after 3 weeks of strength training at 80% of 1-repetition maximum (1-RM). To characterise the input-output properties of the corticospinal tract, stimulus-response curves for corticospinal excitability and inhibition of the right biceps brachii and flexor carpi radialis were constructed and assessed by examining the area under the recruitment curve (AURC). Strength training resulted in a 29% (P < 0.001) increase in 1-RM biceps brachii strength and this was accompanied by a 19% increase in isometric strength of the wrist flexors (P = 0.001). TMS revealed an increase in corticospinal excitability AURC and a decrease in silent period duration AURC for the biceps brachii and flexor carpi radialis following strength training (all P < 0.05). However, the changes in corticospinal function were not associated with increased muscle strength. These findings show that the corticospinal responses to strength training of a proximal upper limb muscle are not spatially restricted, but rather, results in a change in connectivity, among an agonist and a synergistic muscle relevant to force production.

  20. Treatment with PPARα Agonist Clofibrate Inhibits the Transcription and Activation of SREBPs and Reduces Triglyceride and Cholesterol Levels in Liver of Broiler Chickens

    PubMed Central

    Zhang, Lijun; Li, Chunyan; Wang, Fang; Zhou, Shenghua; Shangguan, Mingjun; Xue, Lina; Zhang, Bianying; Ding, Fuxiang; Hui, Dequan; Liang, Aihua; He, Dongchang

    2015-01-01

    PPARα agonist clofibrate reduces cholesterol and fatty acid concentrations in rodent liver by an inhibition of SREBP-dependent gene expression. In present study we investigated the regulation mechanisms of the triglyceride- and cholesterol-lowering effect of the PPARα agonist clofibrate in broiler chickens. We observed that PPARα agonist clofibrate decreases the mRNA and protein levels of LXRα and the mRNA and both precursor and nuclear protein levels of SREBP1 and SREBP2 as well as the mRNA levels of the SREBP1 (FASN and GPAM) and SREBP2 (HMGCR and LDLR) target genes in the liver of treated broiler chickens compared to control group, whereas the mRNA level of INSIG2, which inhibits SREBP activation, was increased in the liver of treated broiler chickens compared to control group. Taken together, the effects of PPARα agonist clofibrate on lipid metabolism in liver of broiler chickens involve inhibiting transcription and activation of SREBPs and SREBP-dependent lipogenic and cholesterologenic gene expression, thereby resulting in a reduction of the triglyceride and cholesterol levels in liver of broiler chickens. PMID:26693219

  1. Opioid Regulation of Spinal Cord Plasticity: Evidence the Kappa-2 Opioid Receptor Agonist GR89696 Inhibits Learning within the Rat Spinal Cord

    PubMed Central

    Washburn, Stephanie N.; Maultsby, Marissa L.; Puga, Denise A.; Grau, James W.

    2007-01-01

    Spinal cord neurons can support a simple form of instrumental learning. In this paradigm, rats completely transected at the second thoracic vertebra learn to minimize shock exposure by maintaining a hindlimb in a flexed position. Prior exposure to uncontrollable shock (shock independent of leg position) disrupts this learning. This learning deficit lasts for at least 24 hours and depends on the NMDA receptor. Intrathecal application of an opioid antagonist blocks the expression, but not the induction, of the learning deficit. A comparison of selective opioid antagonists implicated the kappa opioid receptor. The present experiments further explore how opioids affect spinal instrumental learning using selective opioid agonists. Male Sprague Dawley rats were given an intrathecal injection (30 nmol) of a kappa-1 (U69593), a kappa-2 (GR89696), a mu (DAMGO), or a delta opioid receptor agonist (DPDPE) 10 minutes prior to instrumental testing. Only the kappa-2 opioid receptor agonist GR89696 inhibited acquisition (Experiment 1). GR89696 inhibited learning in a dose dependent fashion (Experiment 2), but had no effect on instrumental performance in previously trained subjects (Experiment 3). Pretreatment with an opioid antagonist (naltrexone) blocked the GR89696-induced learning deficit (Experiment 4). Administration of GR89696 did not produce a lasting impairment (Experiment 5) and a moderate dose of GR89696 (6 nmol) reduced the adverse consequences of uncontrollable nociceptive stimulation (Experiment 6). The results suggest that a kappa-2 opioid agonist inhibits neural modifications within the spinal cord. PMID:17983769

  2. Inhibition of rotavirus infection in cultured cells by N-acetyl-cysteine, PPARγ agonists and NSAIDs.

    PubMed

    Guerrero, Carlos A; Guererero, Carlos A; Murillo, Andrea; Acosta, Orlando

    2012-10-01

    Although the current rotavirus vaccines have shown good tolerance and significant efficacy, it would be useful to develop alternative or complementary strategies aimed at preventing or treating acute diarrhoeal disease caused by this viral agent. A variety of antiviral strategies other than vaccines have been assayed for rotavirus infection management. The recently demonstrated sensitivity of rotavirus infectivity to thiol/disulfide reagents prompted assays for screening drugs that potentially affect cellular redox reactions. MA104 or Caco-2 cells were inoculated with the rotavirus strains RRV, Wa, Wi or M69 and then incubated with different concentrations of drugs belonging to a selected group of 60 drugs that are currently used in humans for purposes other than rotavirus infection treatment. Eighteen of these drugs were able to inhibit rotavirus infectivity to different extents. A more systematic evaluation was performed with drugs that could be used in children such as N-acetylcysteine and ascorbic acid, in addition to ibuprofen, pioglitazone and rosiglitazone, all of which affecting cellular pathways potentially needed by the rotavirus infection process. Evidence is provided here that rotavirus infectivity is significantly inhibited by NAC in different cell-culture systems. These findings suggest that NAC has the potential to be used as a therapeutic tool for treatment and prevention of rotavirus disease in children. Copyright © 2012 Elsevier B.V. All rights reserved.

  3. Inhibition of ERK activity enhances the cytotoxic effect of peroxisome proliferator-activated receptor γ (PPARγ) agonists in HeLa cells.

    PubMed

    Chang, Ha Kyun; Kim, Dae Seong; Chae, Jung Jun; Kim, Minsun; Myong, Jun-Pyo; Lee, Keun Ho; Lee, Myoung Woo; Park, Tae Chul

    2017-01-22

    In this study, we examined whether the peroxisome proliferator-activated receptor γ (PPARγ) agonists, ciglitazone (CGZ) and troglitazone (TGZ), induce cell death in human cervical cancer HeLa cells. The cells were treated with a range of CGZ or TGZ doses for 24 or 48 h. Low concentrations of CGZ (≤10 μM) or TGZ (≤20 μM) had no effect on cell viability whereas higher doses induced cell death in a time- and dose-dependent manner as evidenced by the detection of activated caspase-3 and PARP cleavage. Treatment with the PPARγ antagonist GW9662 followed by PPARγ agonists did not increase CGZ- or TGZ-induced cell death, indicating that PPARγ agonists induced HeLa cell death independently of PPARγ. Moreover, ERK1/2 activation was observed at a CGZ concentration of 25 μM and a TGZ concentration of 35 μM, both of which induced cell death. To elucidate the role of ERK1/2 activated by the two PPARγ agonists, the effect of U0126, an inhibitor of ERK1/2, on PPARγ-agonist-induced cell death was examined. Treatment with 10 or 20 μM U0126 followed by CGZ or TGZ induced the down-regulation of ERK1/2 activity and a decrease in Bcl-2 expression accompanied by the collapse of mitochondrial membrane potential, which in turn significantly enhanced CGZ- or TGZ-induced apoptotic cell death. Our results suggest that PPARγ agonists are capable of inducing apoptotic cell death in HeLa cells independently of PPARγ and that inhibition of ERK1/2 activity offers a strategy to enhance the cytotoxicity of PPARγ agonists in the treatment of cervical cancer. Copyright © 2016 Elsevier Inc. All rights reserved.

  4. Peroxisome-proliferator-activated receptor-{gamma} agonists inhibit the release of proinflammatory cytokines from RSV-infected epithelial cells

    SciTech Connect

    Arnold, Ralf . E-mail: ralf.arnold@medizin.uni-magdeburg.de; Koenig, Wolfgang

    2006-03-15

    The epithelial cells of the airways are the target cells for respiratory syncytial virus (RSV) infection and the site of the majority of the inflammation associated with the disease. Recently, peroxisome-proliferator-activated receptor {gamma} (PPAR{gamma}), a member of the nuclear hormone receptor superfamily, has been shown to possess anti-inflammatory properties. Therefore, we investigated the role of PPAR{gamma} agonists (15d-PGJ{sub 2}, ciglitazone and troglitazone) on the synthesis of RSV-induced cytokine release from RSV-infected human lung epithelial cells (A549). We observed that all PPAR{gamma} ligands inhibited dose-dependently the release of TNF-{alpha}, GM-CSF, IL-1{alpha}, IL-6 and the chemokines CXCL8 (IL-8) and CCL5 (RANTES) from RSV-infected A549 cells. Concomitantly, the PPAR{gamma} ligands diminished the cellular amount of mRNA encoding for IL-6, CXCL8 and CCL5 and the RSV-induced binding activity of the transcription factors NF-{kappa}B (p65/p50) and AP-1 (c-fos), respectively. Our data presented herein suggest a potential application of PPAR{gamma} ligands in the anti-inflammatory treatment of RSV infection.

  5. Pioglitazone, a PPARγ agonist, inhibits growth and invasion of human hepatocellular carcinoma via blockade of the rage signaling.

    PubMed

    Yang, Yuan; Zhao, Ling-Hao; Huang, Bo; Wang, Ruo-Yu; Yuan, Sheng-Xian; Tao, Qi-Fei; Xu, Yong; Sun, Han-Yong; Lin, Chuan; Zhou, Wei-Ping

    2015-12-01

    Pioglitazone (PGZ), a synthetic PPARγ ligand, is known to have anti-tumor activity. However, it is unclear how it acts against hepatocellular carcinoma (HCC). We hypothesized that the pathological receptor for advanced glycation end products (RAGE) is involved in the PGZ anti-tumor process. To test this notion, human primary HCC tissues and corresponding adjacent non-cancerous tissues (ANCT) from 75 consecutive cases were analyzed. The expression and clinical significance of RAGE was assessed by immunohistochemical assay through tissue microarray. After HCC cells were pretreated with different concentrations of PGZ, cell proliferation, apoptosis, cell invasion, and cell cycle distribution were evaluated by multiple assays. The results showed that, the positive expression of RAGE was significantly higher in HCC tissues than in ANCT (66.7% vs. 36.0%, P < 0.001), and was closely associated with pathological staging (P = 0.014) and lymph-vascular space invasion (P = 0.003). Moreover, PGZ inhibited proliferative activity and invasive potential, and induced apoptosis and cell cycle arrest in HCC cells resulting in increased expression of PPARγ and decreased expression of RAGE, NF-κB, HMGB1, p38MAPK, Ki-67, MMP-2, and CyclinD1. Furthermore, knockdown of RAGE or NF-κB by siRNA effectively suppressed cell proliferation and invasion, and mediated the inhibitory effects of PGZ in HCC cells. Taken together, our findings suggest that, RAGE is overexpressed in human HCC tissues, and is closely associated with the pathological staging and tumor invasion of HCC. In addition, PGZ as a PPARγ agonist may inhibit growth and invasion of HCC cells via blockade of the RAGE signaling. © 2014 Wiley Periodicals, Inc.

  6. Phosphocholine-Modified Macromolecules and Canonical Nicotinic Agonists Inhibit ATP-Induced IL-1β Release.

    PubMed

    Hecker, Andreas; Küllmar, Mira; Wilker, Sigrid; Richter, Katrin; Zakrzewicz, Anna; Atanasova, Srebrena; Mathes, Verena; Timm, Thomas; Lerner, Sabrina; Klein, Jochen; Kaufmann, Andreas; Bauer, Stefan; Padberg, Winfried; Kummer, Wolfgang; Janciauskiene, Sabina; Fronius, Martin; Schweda, Elke K H; Lochnit, Günter; Grau, Veronika

    2015-09-01

    IL-1β is a potent proinflammatory cytokine of the innate immune system that is involved in host defense against infection. However, increased production of IL-1β plays a pathogenic role in various inflammatory diseases, such as rheumatoid arthritis, gout, sepsis, stroke, and transplant rejection. To prevent detrimental collateral damage, IL-1β release is tightly controlled and typically requires two consecutive danger signals. LPS from Gram-negative bacteria is a prototypical first signal inducing pro-IL-1β synthesis, whereas extracellular ATP is a typical second signal sensed by the ATP receptor P2X7 that triggers activation of the NLRP3-containing inflammasome, proteolytic cleavage of pro-IL-1β by caspase-1, and release of mature IL-1β. Mechanisms controlling IL-1β release, even in the presence of both danger signals, are needed to protect from collateral damage and are of therapeutic interest. In this article, we show that acetylcholine, choline, phosphocholine, phosphocholine-modified LPS from Haemophilus influenzae, and phosphocholine-modified protein efficiently inhibit ATP-mediated IL-1β release in human and rat monocytes via nicotinic acetylcholine receptors containing subunits α7, α9, and/or α10. Of note, we identify receptors for phosphocholine-modified macromolecules that are synthesized by microbes and eukaryotic parasites and are well-known modulators of the immune system. Our data suggest that an endogenous anti-inflammatory cholinergic control mechanism effectively controls ATP-mediated release of IL-1β and that the same mechanism is used by symbionts and misused by parasites to evade innate immune responses of the host. Copyright © 2015 by The American Association of Immunologists, Inc.

  7. Hydrogen sulfide decreases β-adrenergic agonist-stimulated lung liquid clearance by inhibiting ENaC-mediated transepithelial sodium absorption.

    PubMed

    Agné, Alisa M; Baldin, Jan-Peter; Benjamin, Audra R; Orogo-Wenn, Maria C; Wichmann, Lukas; Olson, Kenneth R; Walters, Dafydd V; Althaus, Mike

    2015-04-01

    In pulmonary epithelia, β-adrenergic agonists regulate the membrane abundance of the epithelial sodium channel (ENaC) and, thereby, control the rate of transepithelial electrolyte absorption. This is a crucial regulatory mechanism for lung liquid clearance at birth and thereafter. This study investigated the influence of the gaseous signaling molecule hydrogen sulfide (H2S) on β-adrenergic agonist-regulated pulmonary sodium and liquid absorption. Application of the H2S-liberating molecule Na2S (50 μM) to the alveolar compartment of rat lungs in situ decreased baseline liquid absorption and abrogated the stimulation of liquid absorption by the β-adrenergic agonist terbutaline. There was no additional effect of Na2S over that of the ENaC inhibitor amiloride. In electrophysiological Ussing chamber experiments with native lung epithelia (Xenopus laevis), Na2S inhibited the stimulation of amiloride-sensitive current by terbutaline. β-adrenergic agonists generally increase ENaC abundance by cAMP formation and activation of PKA. Activation of this pathway by forskolin and 3-isobutyl-1-methylxanthine increased amiloride-sensitive currents in H441 pulmonary epithelial cells. This effect was inhibited by Na2S in a dose-dependent manner (5-50 μM). Na2S had no effect on cellular ATP concentration, cAMP formation, and activation of PKA. By contrast, Na2S prevented the cAMP-induced increase in ENaC activity in the apical membrane of H441 cells. H441 cells expressed the H2S-generating enzymes cystathionine-β-synthase, cystathionine-γ-lyase, and 3-mercaptopyruvate sulfurtransferase, and they produced H2S amounts within the employed concentration range. These data demonstrate that H2S prevents the stimulation of ENaC by cAMP/PKA and, thereby, inhibits the proabsorptive effect of β-adrenergic agonists on lung liquid clearance.

  8. The serotonin receptor agonist 5-methoxy-N,N-dimethyltryptamine facilitates noradrenaline release from rat spinal cord slices and inhibits monoamine oxidase activity.

    PubMed

    Reimann, W; Schneider, F

    1993-03-01

    1. The influences of the purported serotonergic agonist 5-methoxy-N,N-dimethyltryptamine (MeODMT) on noradrenaline release and metabolism were investigated in a rat spinal cord release model and a monoamine oxidase (MAO) assay. 2. MeODMT inhibited the basal outflow of tritium from rat spinal cord slices preincubated with [3H]noradrenaline and enhanced the electrically-evoked overflow. 3. Effects on basal outflow were not observed, when monoamine oxidase (MAO) was inhibited by pargyline. Effects on the evoked overflow were not observed in the presence of metitepine or phentolamine. 4. Preferential inhibition by MeODMT of MAO A-type enzyme activity was found in a direct assay. 5. The results provide evidence for two different effects by which MeODMT reinforces noradrenergic neurotransmission in the rat spinal cord: facilitation of stimulation-evoked noradrenaline release and inhibition of noradrenaline metabolism by MAO inhibition.

  9. Mycobacterium tuberculosis and TLR2 Agonists Inhibit Induction of Type I IFN and Class I MHC Antigen Cross Processing by TLR9

    PubMed Central

    Simmons, Daimon P.; Canaday, David H.; Liu, Yi; Li, Qing; Huang, Alex; Boom, W. Henry; Harding, Clifford V.

    2010-01-01

    Dendritic cells (DCs) cross process exogenous Ags and present them by class I MHC (MHC-I) molecules to CD8+ T cells specific for Ags from viruses and bacteria such as Mycobacterium tuberculosis. Unmethylated CpG DNA signals through TLR9 to induce type I IFN (IFN-α/β), which enhances MHC-I Ag cross processing, but lipoproteins that signal through TLR2 do not induce IFN-α/β. In these studies we observed that M. tuberculosis, which expresses agonists of both TLR9 and TLR2, did not induce production of IFN-α/β or cross processing by murine DCs. Furthermore, M. tuberculosis and TLR2 agonists inhibited induction of IFN-α/β and DC cross processing by CpG DNA. Exogenous IFN-α/β effectively enhanced cross processing of M. bovis bacillus Calmette-Guérin expressing OVA, bypassing the inhibition of induction of endogenous IFN-α/β. In addition, inhibition of TLR9-induced cross processing of M. bovis bacillus Calmette-Guérin expressing OVA could be circumvented by pretreating cells with CpG DNA to induce IFN-α/β and MHC-I cross processing before inhibitory mycobacterial TLR2 agonists were present. Inhibition of the response to one TLR by another may affect the ultimate response to pathogens like M. tuberculosis that express agonists of multiple TLRs, including TLR2 and TLR9. This mechanism may contribute to immune evasion and explain why IFN-α/β provides little contribution to host immunity to M. tuberculosis. However, downregulation of certain TLR responses may benefit the host by preventing detrimental excessive inflammation that may occur in the presence of persistent infection. PMID:20660347

  10. Further pharmacological characterization of 5-HT2C receptor agonist-induced inhibition of 5-HT neuronal activity in the dorsal raphe nucleus in vivo

    PubMed Central

    Quérée, P; Peters, S; Sharp, T

    2009-01-01

    Background and purpose: Recent experiments using non-selective 5-hydroxytryptamine (5-HT)2C receptor agonists including WAY 161503 suggested that midbrain 5-HT neurones are under the inhibitory control of 5-HT2C receptors, acting via neighbouring gamma-aminobutyric acid (GABA) neurones. The present study extended this pharmacological characterization by comparing the actions of WAY 161503 with the 5-HT2C receptor agonists, Ro 60-0275 and 1-(3-chlorophenyl) piperazine (mCPP), as well as the non-selective 5-HT agonist lysergic acid diethylamide (LSD) and the 5-HT releasing agent 3,4-methylenedioxymethamphetamine (MDMA). Experimental approach: 5-HT neuronal activity was measured in the dorsal raphe nucleus (DRN) using extracellular recordings in anaesthetized rats. The activity of DRN GABA neurones was assessed using double-label immunohistochemical measurements of Fos and glutamate decarboxylase (GAD). Key results: Ro 60-0175, like WAY 161503, inhibited 5-HT neurone firing, and the 5-HT2C antagonist SB 242084 reversed this effect. mCPP also inhibited 5-HT neurone firing (∼60% neurones) in a SB 242084-reversible manner. LSD inhibited 5-HT neurone firing; however, this effect was not altered by either SB 242084 or the 5-HT2A/C receptor antagonist ritanserin but was reversed by the 5-HT1A receptor antagonist WAY 100635. Similarly, MDMA inhibited 5-HT neurone firing in a manner reversible by WAY 100635, but not SB 242084 or ritanserin. Finally, both Ro 60-0275 and mCPP, like WAY 161503, increased Fos expression in GAD-positive DRN neurones. Conclusions and implications: These data strengthen the hypothesis that midbrain 5-HT neurones are under the inhibitory control of 5-HT2C receptors, and suggest that the 5-HT2C agonists Ro 60-0175, mCPP and WAY 161503, but not LSD or MDMA, are useful probes of the mechanism(s) involved. PMID:19845681

  11. The G Protein-Coupled Estrogen Receptor Agonist G-1 Inhibits Nuclear Estrogen Receptor Activity and Stimulates Novel Phosphoproteomic Signatures

    PubMed Central

    Smith, L. Cody; Ralston-Hooper, Kimberly J.; Ferguson, P. Lee; Sabo-Attwood, Tara

    2016-01-01

    Estrogen exerts cellular effects through both nuclear (ESR1 and ESR2) and membrane-bound estrogen receptors (G-protein coupled estrogen receptor, GPER); however, it is unclear if they act independently or engage in crosstalk to influence hormonal responses. To investigate each receptor’s role in proliferation, transcriptional activation, and protein phosphorylation in breast cancer cells (MCF-7), we employed selective agonists for ESR1 propyl-pyrazole-triol (PPT), ESR2 diarylpropionitrile (DPN), and GPER (G-1) and also determined the impact of xenoestrogens bisphenol-A (BPA) and genistein on these effects. As anticipated, 17β-estradiol (E2), PPT, DPN, BPA, and genistein each enhanced proliferation and activation of an ERE-driven reporter gene whereas G-1 had no significant impact. However, G-1 significantly reduced E2-, PPT-, DPN-, BPA-, and genistein-induced proliferation and ERE activation at doses greater than 500 nM indicating that G-1 mediated inhibition is not ESR isotype specific. As membrane receptors initiate cascades of phosphorylation events, we performed a global phosphoproteomic analysis on cells exposed to E2 or G-1 to identify potential targets of receptor crosstalk via downstream protein phosphorylation targets. Of the 211 phosphorylated proteins identified, 40 and 13 phosphoproteins were specifically modified by E2 and G-1, respectively. Subnetwork enrichment analysis revealed several processes related to cell cycle were specifically enriched by G-1 compared with E2. Further there existed a number of newly identified proteins that were specifically phosphorylated by G-1. These phosphorylation networks highlight specific proteins that may modulate the inhibitory effects of G-1 and suggest a novel role for interference with nuclear receptor activity driven by E2 and xenoestrogens. PMID:27026707

  12. Ciproxifan, a histamine H3 receptor antagonist and inverse agonist, presynaptically inhibits glutamate release in rat hippocampus.

    PubMed

    Lu, Cheng-Wei; Lin, Tzu-Yu; Chang, Chia-Ying; Huang, Shu-Kuei; Wang, Su-Jane

    2017-03-15

    Ciproxifan is an H3 receptor antagonist and inverse agonist with antipsychotic effects in several preclinical models; its effect on glutamate release has been investigated in the rat hippocampus. In a synaptosomal preparation, ciproxifan reduced 4-aminopyridine (4-AP)-evoked Ca(2+)-dependent glutamate release and cytosolic Ca(2+) concentration elevation but did not affect the membrane potential. The inhibitory effect of ciproxifan on 4-AP-evoked glutamate release was prevented by the Gi/Go-protein inhibitor pertussis toxin and Cav2.2 (N-type) and Cav2.1 (P/Q-type) channel blocker ω-conotoxin MVIIC, but was not affected by the intracellular Ca(2+)-release inhibitors dantrolene and CGP37157. Furthermore, the phospholipase A2 (PLA2) inhibitor OBAA, prostaglandin E2 (PGE2), PGE2 subtype 2 (EP2) receptor antagonist PF04418948, and extracellular signal-regulated kinase (ERK) inhibitor FR180204 eliminated the inhibitory effect of ciproxifan on glutamate release. Ciproxifan reduced the 4-AP-evoked phosphorylation of ERK and synapsin I, a presynaptic target of ERK. The ciproxifan-mediated inhibition of glutamate release was prevented in synaptosomes from synapsin I-deficient mice. Moreover, ciproxifan reduced the frequency of miniature excitatory postsynaptic currents without affecting their amplitude in hippocampal slices. Our data suggest that ciproxifan, acting through the blockade of Gi/Go protein-coupled H3 receptors present on hippocampal nerve terminals, reduces voltage-dependent Ca(2+) entry by diminishing PLA2/PGE2/EP2 receptor pathway, which subsequently suppresses the ERK/synapsin I cascade to decrease the evoked glutamate release.

  13. Btk inhibition suppresses agonist-induced human macrophage activation and inflammatory gene expression in RA synovial tissue explants.

    PubMed

    Hartkamp, Linda M; Fine, Jay S; van Es, Inge E; Tang, Man Wai; Smith, Michael; Woods, John; Narula, Satwant; DeMartino, Julie; Tak, Paul P; Reedquist, Kris A

    2015-08-01

    Bruton's tyrosine kinase (Btk) is required for B lymphocyte and myeloid cell contributions to pathology in murine models of arthritis. Here, we examined the potential contributions of synovial Btk expression and activation to inflammation in rheumatoid arthritis (RA). Btk was detected by immunohistochemistry and digital image analysis in synovial tissue from biologically naive RA (n=16) and psoriatic arthritis (PsA) (n=12) patients. Cell populations expressing Btk were identified by immunofluorescent double labelling confocal microscopy, quantitative (q-) PCR and immunoblotting. The effects of a Btk-specific inhibitor, RN486, on gene expression in human macrophages and RA synovial tissue explants (n=8) were assessed by qPCR, ELISA and single-plex assays. Btk was expressed at equivalent levels in RA and PsA synovial tissue, restricted to B lymphocytes, monocytes, macrophages and mast cells. RN486 significantly inhibited macrophage IL-6 production induced by Fc receptor and CD40 ligation. RN486 also reduced mRNA expression of overlapping gene sets induced by IgG, CD40 ligand (CD40L) and RA synovial fluid, and significantly suppressed macrophage production of CD40L-induced IL-8, TNF, MMP-1 and MMP-10, LPS-induced MMP-1, MMP-7 and MMP-10 production, and spontaneous production of IL-6, PDGF, CXCL-9 and MMP-1 by RA synovial explants. Btk is expressed equivalently in RA and PsA synovial tissue, primarily in macrophages. Btk activity is needed to drive macrophage activation in response to multiple agonists relevant to inflammatory arthritis, and promotes RA synovial tissue cytokine and MMP production. Pharmacological targeting of Btk may be of therapeutic benefit in the treatment of RA and other inflammatory diseases. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

  14. Inhibition of some aspects of acute inflammation of guinea-pig lung by intraperitoneal dexamethasone and mifepristone: demonstration of agonist activity of mifepristone in the guinea-pig.

    PubMed

    Whelan, C J; Hughes, S C; Wren, G P

    1995-03-01

    We have determined the inhibitory activity of dexamethasone as an inhibitor of histamine-induced plasma protein extravasation (PPE) in guinea-pig lung and skin, and of lipopolysaccharide (LPS)-induced neutrophilia and platelet activating factor (PAF)-induced eosinophilia in guinea-pig lungs. Dexamethasone inhibited PAF-induced eosinophilia in guinea-pig lung (ED50 1.4 mg/kg i.p.). Higher doses of dexamethasone were required to inhibit LPS-induced neutrophilia (ED50 10.8 mg/kg i.p.). However, at doses up to 150 mg/kg i.p. dexamethasone did not inhibit histamine-induced plasma protein extravasation (PPE) in guinea-pig lung, but did inhibit PPE in guinea-pig skin. These preparations have previously been shown to be equally sensitive to inhibition by the beta 2-adrenoceptor agonist salmeterol. Dexamethasone inhibited PAF-induced eosinophilia (5 mg/kg) or LPS-induced neutrophilia (50 mg/kg) when given 3 h or 1 h prior to challenge. Inhibitory activity was lost when dexamethasone was administered 23 h prior to LPS or 1 h after PAF. The glucocorticoid antagonist mifepristone (1-100 mg/kg i.p.) caused dose-related inhibition of PAF-induced eosinophilia but not of LPS-induced neutrophilia. The highest dose of mifepristone used (100 mg/kg) did not reverse the inhibitory actions of dexamethasone (50 mg/kg) on LPS-induced neutrophilia. We suggest that the different inhibitory activity of dexamethasone in the preparations studied indicates differences in the sensitivity of the target cells involved to inhibition by dexamethasone. We also suggest that inhibition of PAF-induced eosinophilia by mifepristone reflects the partial agonist activity of this agent, demonstrated by others in different experimental systems.

  15. Candida glabrata binds to glycosylated and lectinic receptors on the coronary endothelial luminal membrane and inhibits flow sense and cardiac responses to agonists.

    PubMed

    Torres-Tirado, David; Knabb, Maureen; Castaño, Irene; Patrón-Soberano, Araceli; De Las Peñas, Alejandro; Rubio, Rafael

    2016-01-01

    Candida glabrata (CG) is an opportunistic fungal pathogen that initiates infection by binding to host cells via specific lectin-like adhesin proteins. We have previously shown the importance of lectin-oligosaccharide binding in cardiac responses to flow and agonists. Because of the lectinic-oligosaccharide nature of CG binding, we tested the ability of CG to alter the agonist- and flow-induced changes in cardiac function in isolated perfused guinea pig hearts. Both transmission and scanning electron microscopy showed strong attachment of CG to the coronary endothelium, even after extensive washing. CG shifted the coronary flow vs. auricular-ventricular (AV) delay relationship upward, indicating that greater flow was required to achieve the same AV delay. This effect was completely reversed with mannose, partially reversed with galactose and N-acetylgalactosamine, but hyaluronan had no effect. Western blot analysis was used to determine binding of CG to isolated coronary endothelial luminal membrane (CELM) receptors, and the results indicate that flow-sensitive CELM receptors, ANG II type I, α-adrenergic 1A receptor, endothelin-2, and VCAM-1 bind to CG. In addition, CG inhibited agonist-induced effects of bradykinin, angiotensin, and phenylephrine on AV delay, coronary perfusion pressure, and left ventricular pressure. Mannose reversed the inhibitory effects of CG on the agonist responses. These results suggest that CG directly binds to flow-sensitive CELM receptors via lectinic-oligosaccharide interactions with mannose and disrupts the lectin-oligosaccharide binding necessary for flow-induced cardiac responses.

  16. TRA-418, a thromboxane A2 receptor antagonist and prostacyclin receptor agonist, inhibits platelet-leukocyte interaction in human whole blood.

    PubMed

    Miyamoto, Mitsuko; Ohno, Michihiro; Yamada, Naohiro; Ohtake, Atsushi; Matsushita, Teruo

    2010-10-01

    TRA-418, a compound with both thromboxane A2 receptor (TP receptor) antagonistic and prostacyclin receptor (IP receptor) agonistic activities, was synthesised in our laboratory as a new antithrombotic agent. In this study, we examined the effects of TRA-418 on platelet-leukocyte interactions in human whole blood. Platelet-leukocyte interactions were induced by U-46619 in the presence of epinephrine (U-46619 + epinephrine) or with thrombin receptor agonist peptide 1-6 (TRAP). Platelet-leukocyte interactions were assessed by flow cytometry, with examination of both platelet-neutrophil and platelet-monocyte complexes. In a control experiment, the TP receptor antagonist SQ-29548 significantly inhibited the induction of platelet-leukocyte complexes by the combination of U-46619 and epinephrine, but not TRAP-induced formation of platelet-leukocyte complexes. Conversely, the IP receptor agonist beraprost sodium inhibited platelet-leukocyte complex formation induced by both methods, although the IC50 values of beraprost sodium for U-46619 + epinephrine were at least 10-fold greater than for TRAP. Under such conditions, TRA-418 inhibited both U-46619 + epinephrine-induced and TRAP-induced platelet-leukocyte complex formation in a concentration-dependent manner, in a similar range. These results suggest that TRA-418 exerts its inhibitory effects on platelet-leukocyte interactions by acting as a TP receptor antagonist as well as an IP receptor agonist in an additive or synergistic manner. These inhibitory effects of TRA-418 on formation of platelet-leukocyte complexes suggest the compound is beneficial effects as an antithrombotic agent.

  17. Central administration of pan-somatostatin agonist ODT8-SST prevents abdominal surgery-induced inhibition of circulating ghrelin, food intake and gastric emptying in rats.

    PubMed

    Stengel, A; Goebel-Stengel, M; Wang, L; Luckey, A; Hu, E; Rivier, J; Taché, Y

    2011-07-01

      Activation of brain somatostatin receptors (sst(1-5) ) with the stable pan-sst(1-5) somatostatin agonist, ODT8-SST blocks acute stress and central corticotropin-releasing factor (CRF)-mediated activation of endocrine and adrenal sympathetic responses. Brain CRF signaling is involved in delaying gastric emptying (GE) immediately post surgery. We investigated whether activation of brain sst signaling pathways modulates surgical stress-induced inhibition of gastric emptying and food intake. Fasted rats were injected intracisternally (i.c.) with somatostatin agonists and underwent laparotomy and 1-min cecal palpation. Gastric emptying of a non-nutrient solution and circulating acyl and desacyl ghrelin levels were assessed 50min post surgery. Food intake was monitored for 24 h. The abdominal surgery-induced inhibition of GE (65%), food intake (73% at 2h) and plasma acyl ghrelin levels (67%) was completely prevented by ODT8-SST (1μg per rat, i.c.). The selective sst(5) agonist, BIM-23052 prevented surgery-induced delayed GE, whereas selective sst(1) , sst(2) , or sst(4) agonists had no effect. However, the selective sst(2) agonist, S-346-011 (1μg per rat, i.c.) counteracted the abdominal surgery-induced inhibition of acyl ghrelin and food intake but not the delayed GE. The ghrelin receptor antagonist, [D-Lys(3) ]-GHRP-6 (0.93mg kg(-1) , intraperitoneal, i.p.) blocked i.p. ghrelin-induced increased GE, while not influencing i.c. ODT8-SST-induced prevention of delayed GE and reduced food intake after surgery. ODT8-SST acts in the brain to prevent surgery-induced delayed GE likely via activating sst(5) . ODT8-SST and the sst(2) agonist prevent the abdominal surgery-induced decrease in food intake and plasma acyl ghrelin indicating dissociation between brain somatostatin signaling involved in preventing surgery-induced suppression of GE and feeding response. © 2011 Blackwell Publishing Ltd.

  18. Central administration of pansomatostatin agonist ODT8-SST prevents abdominal surgery-induced inhibition of circulating ghrelin, food intake and gastric emptying in rats

    PubMed Central

    STENGEL, A.; GOEBEL-STENGEL, M.; WANG, L.; LUCKEY, A.; HU, E.; RIVIER, J.; TACHÉ, Y.

    2011-01-01

    Background Activation of brain somatostatin receptors (sst1-5) with the stable pan-sst1-5 somatostatin agonist, ODT8-SST blocks acute stress and central corticotropin-releasing factor (CRF)-mediated activation of endocrine adrenal sympathetic responses. Brain CRF signaling is involved in delaying gastric emptying (GE) immediately post surgery. We investigated whether activation of brain sst signaling pathways modulates surgical stress-induced inhibition of gastric emptying and food intake. Methods Fasted rats were injected intracisternally (i.c.) with somatostatin agonists and underwent laparotomy and 1-min cecal palpation. GE of a non-nutrient solution and circulating acyl and desacyl ghrelin levels were assessed 50 min post surgery. Food intake was monitored for 24h. Key results The abdominal surgery-induced inhibition of GE (65%), food intake (73% at 2h) and plasma acyl ghrelin levels (67%) was completely prevented by ODT8-SST (1μg/rat, i.c.). The selective sst5 agonist, BIM-23052 prevented surgery-induced delayed GE, whereas selective sst1, sst2 or sst4 agonists had no effect. However, the selective sst2 agonist, S-346-011 (1μg/rat, i.c.) counteracted the abdominal surgery-induced inhibition of acyl ghrelin and food intake but not the delayed GE. The ghrelin receptor antagonist, [D-Lys3]-GHRP-6 (0.93 mg/kg, intraperitoneal, i.p.) blocked i.p. ghrelin-induced increased GE, while not influencing i.c. ODT8-SST-induced prevention of delayed GE and reduced food intake after surgery. Conclusions & Inferences ODT8-SST acts in the brain to prevent surgery-induced delayed GE likely via activating sst5. ODT8-SST and the sst2 agonist prevent the abdominal surgery-induced decrease in food intake and plasma acyl ghrelin indicating dissociation between brain somatostatin signaling involved in preventing surgery-induced suppression of GE and feeding response. PMID:21569179

  19. Cisapride, a selective serotonin 5-HT4-receptor agonist, inhibits voltage-dependent K(+) channels in rabbit coronary arterial smooth muscle cells.

    PubMed

    Kim, Hye Won; Li, Hongliang; Kim, Han Sol; Shin, Sung Eun; Jung, Won-Kyo; Ha, Kwon-Soo; Han, Eun-Taek; Hong, Seok-Ho; Choi, Il-Whan; Park, Won Sun

    2016-09-23

    We investigated the effect of cisapride, a selective serotonin 5-HT4-receptor agonist, on voltage-dependent K(+) (Kv) channels using freshly isolated smooth muscle cells from the coronary arteries of rabbits. The amplitude of Kv currents was reduced by cisapride in a concentration-dependent manner, with an IC50 value of 6.77 ± 6.01 μM and a Hill coefficient of 0.51 ± 0.18. The application of cisapride shifted the steady-state inactivation curve toward a more negative potential, but had no significant effect on the steady-state activation curve. This suggested that cisapride inhibited the Kv channel in a closed state by changing the voltage sensitivity of Kv channels. The application of another selective serotonin 5-HT4-receptor agonist, prucalopride, did not affect the basal Kv current and did not alter the inhibitory effect of cisapride on Kv channels. From these results, we concluded that cisapride inhibited vascular Kv current in a concentration-dependent manner by shifting the steady-state inactivation curve, independent of its own function as a selective serotonin 5-HT4-receptor agonist. Copyright © 2016 Elsevier Inc. All rights reserved.

  20. FXR agonists enhance the sensitivity of biliary tract cancer cells to cisplatin via SHP dependent inhibition of Bcl-xL expression

    PubMed Central

    Wang, Wei; Zhan, Ming; Li, Qi; Chen, Wei; Chu, Huiling; Huang, Qihong; Hou, Zhaoyuan; Man, Mohan; Wang, Jian

    2016-01-01

    Chemoresistance is common in patients with biliary tract cancer (BTC) including gallbladder cancer (GBC) and cholangiocarcinoma (CC). Therefore, it is necessary to identify effective chemotherapeutic agents for BTC. In the present study, we for the first time tested the effect of farnesoid X receptor (FXR) agonists GW4064 and CDCA (chenodeoxycholic acid) in combination with cisplatin (CDDP) on increasing the chemosensitivity in BTC. Our results show that co-treatment of CDDP with FXR agonists remarkably enhance chemosensitivity of BTC cells. Mechanistically, we found that activation of FXR induced expression of small heterodimer partner (SHP), which in turn inhibited signal transducer and activator of transcription 3 (STAT3) phosphorylation and resulted in down-regulation of Bcl-xL expression in BTC cells, leading to increased susceptibility to CDDP. Moreover, the experiments on tumor-bearing mice showed that GW4064/CDDP co-treatment inhibited the tumor growth in vivo by up-regulating SHP expression and down-regulating STAT3 phosphorylation. These results suggest CDDP in combination with FXR agonists could be a potential new therapeutic strategy for BTC. PMID:27127878

  1. Inhibition of human lung cancer cell growth by the peroxisome proliferator-activated receptor-gamma agonists through induction of apoptosis.

    PubMed

    Tsubouchi, Y; Sano, H; Kawahito, Y; Mukai, S; Yamada, R; Kohno, M; Inoue, K; Hla, T; Kondo, M

    2000-04-13

    Peroxisome proliferator-activated receptors (PPARs), members of the nuclear hormone receptors superfamily, have an important regulatory role in adipogenesis and inflammation. PPAR-gamma ligands induce terminal differentiation and growth inhibition of human breast cancer cells and prostatic cancer cells. In this study, we demonstrated that PPAR-gamma, but not PPAR-alpha, was expressed in human lung cancer cell lines by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis. We also found that the synthetic PPAR-gamma agonist thiazolidinedione compounds (troglitazone) and the endogenous PPAR-gamma ligand, 15-deoxy-Delta(12,14)-prostaglandin J(2) (15d-PGJ(2)), inhibited the growth of human lung cancer cells through the induction of apoptosis. However, PPAR-alpha agonist (bezafibrate) and other prostanoids (PGE(2), PGF(2alpha)) did not induce apoptosis. These findings suggest that PPAR-gamma may play an important role in the pathogenesis of lung cancer and that PPAR-gamma agonist may be useful therapeutic agents in the treatment of human lung cancer. Copyright 2000 Academic Press.

  2. OL3, a novel low-absorbed TGR5 agonist with reduced side effects, lowered blood glucose via dual actions on TGR5 activation and DPP-4 inhibition

    PubMed Central

    Ma, Shan-yao; Ning, Meng-meng; Zou, Qing-an; Feng, Ying; Ye, Yang-liang; Shen, Jian-hua; Leng, Ying

    2016-01-01

    Aim: TGR5 agonists stimulate intestinal glucagon-like peptide-1 (GLP-1) release, but systemic exposure causes unwanted side effects, such as gallbladder filling. In the present study, linagliptin, a DPP-4 inhibitor with a large molecular weight and polarity, and MN6, a previously described TGR5 agonist, were linked to produce OL3, a novel low-absorbed TGR5 agonist with reduced side-effects and dual function in lowering blood glucose by activation of TGR5 and inhibition of DPP-4. Methods: TGR5 activation was assayed in HEK293 cells stably expressing human or mouse TGR5 and a CRE-driven luciferase gene. DPP-4 inhibition was assessed based on the rate of hydrolysis of a surrogate substrate. GLP-1 secretion was measured in human enteroendocrine NCI-H716 cells. OL3 permeability was tested in Caco-2 cells. Acute glucose-lowering effects of OL3 were evaluated in ICR and diabetic ob/ob mice. Results: OL3 activated human and mouse TGR5 with an EC50 of 86.24 and 17.36 nmol/L, respectively, and stimulated GLP-1 secretion in human enteroendocrine NCI-H716 cells (3–30 μmol/L). OL3 inhibited human and mouse DPP-4 with IC50 values of 18.44 and 69.98 μmol/L, respectively. Low permeability of OL3 was observed in Caco-2 cells. In ICR mice treated orally with OL3 (150 mg/kg), the serum OL3 concentration was 101.10 ng/mL at 1 h, and decreased to 13.38 ng/mL at 5.5 h post dose, confirming the low absorption of OL3 in vivo. In ICR mice and ob/ob mice, oral administration of OL3 significantly lowered the blood glucose levels, which was a synergic effect of activating TGR5 that stimulated GLP-1 secretion in the intestine and inhibiting DPP-4 that cleaved GLP-1 in the plasma. In ICR mice, oral administration of OL3 did not cause gallbladder filling. Conclusion: OL3 is a low-absorbed TGR5 agonist that lowers blood glucose without inducing gallbladder filling. This study presents a new strategy in the development of potent TGR5 agonists in treating type 2 diabetes, which target to the

  3. A retinoic acid receptor β agonist (CD2019) overcomes inhibition of axonal outgrowth via phosphoinositide 3-kinase signalling in the injured adult spinal cord

    PubMed Central

    Agudo, Marta; Yip, Ping; Davies, Meirion; Bradbury, Elizabeth; Doherty, Patrick; McMahon, Stephen; Maden, Malcolm; Corcoran, Jonathan P.T.

    2010-01-01

    After spinal cord injury in the adult mammal, axons do not normally regrow and this commonly leads to paralysis. Retinoic acid (RA) can stimulate neurite outgrowth in vitro of both the embryonic central and peripheral nervous system, via activation of the retinoic acid receptor (RAR) β2. We show here that regions of the adult CNS, including the cerebellum and cerebral cortex, express RARβ2. We show that when cerebellar neurons are grown in the presence of myelin-associated glycoprotein (MAG) which inhibits neurite outgrowth, RARβ can be activated in a dose dependent manner by a RARβ agonist (CD2019) and neurite outgrowth can occur via phosphoinositide 3-kinase (PI3K) signalling. In a model of spinal cord injury CD2019 also acts through PI3K signalling to induce axonal outgrowth of descending corticospinal fibres and promote functional recovery. Our data suggest that RARβ agonists may be of therapeutic potential for human spinal cord injuries. PMID:19800972

  4. Inhibition of beta-oxidative respiration is a therapeutic window associated with the cancer chemo-preventive activity of PPARgamma agonists.

    PubMed

    Andela, Valentine B; Altuwaijri, Saleh; Wood, James; Rosier, Randy N

    2005-03-14

    We demonstrate expression and coordinate induction of PPARgamma and lipogenic enzymes (HMG-CoA synthase, HMG-CoA reductase and fatty acid synthase) in a murine lung alveolar carcinoma cell line (Line 1) treated with the PPARgamma agonist troglitazone (TRO) [0-100 microM]. We postulate that TRO induces a shift in cellular energy metabolism towards fatty acid oxidation (beta-oxidative respiration). Accordingly, co-treatment with TRO [30 microM] and increasing concentrations of trimetazidine (TMZ) [0.1-3 mM], an inhibitor of beta-oxidation, results in a dose dependent decrease cellular ATP levels and a dose dependent induction of apoptosis. These findings, suggest that inhibition of beta-oxidative respiration is a therapeutic window associated with the cancer chemo-preventive activity of PPARgamma agonists.

  5. A retinoic acid receptor beta agonist (CD2019) overcomes inhibition of axonal outgrowth via phosphoinositide 3-kinase signalling in the injured adult spinal cord.

    PubMed

    Agudo, Marta; Yip, Ping; Davies, Meirion; Bradbury, Elizabeth; Doherty, Patrick; McMahon, Stephen; Maden, Malcolm; Corcoran, Jonathan P T

    2010-01-01

    After spinal cord injury in the adult mammal, axons do not normally regrow and this commonly leads to paralysis. Retinoic acid (RA) can stimulate neurite outgrowth in vitro of both the embryonic central and peripheral nervous system, via activation of the retinoic acid receptor (RAR) beta2. We show here that regions of the adult CNS, including the cerebellum and cerebral cortex, express RARbeta2. We show that when cerebellar neurons are grown in the presence of myelin-associated glycoprotein (MAG) which inhibits neurite outgrowth, RARbeta can be activated in a dose dependent manner by a RARbeta agonist (CD2019) and neurite outgrowth can occur via phosphoinositide 3-kinase (PI3K) signalling. In a model of spinal cord injury CD2019 also acts through PI3K signalling to induce axonal outgrowth of descending corticospinal fibres and promote functional recovery. Our data suggest that RARbeta agonists may be of therapeutic potential for human spinal cord injuries.

  6. Discovery and Characterization of a G Protein–Biased Agonist That Inhibits β-Arrestin Recruitment to the D2 Dopamine Receptor

    PubMed Central

    Chun, Lani S.; Moritz, Amy E.; Miller, Brittney N.; Doyle, Trevor B.; Conroy, Jennie L.; Padron, Adrian; Meade, Julie A.; Xiao, Jingbo; Hu, Xin; Dulcey, Andrés E.; Han, Yang; Duan, Lihua; Titus, Steve; Bryant-Genevier, Melanie; Barnaeva, Elena; Ferrer, Marc; Javitch, Jonathan A.; Beuming, Thijs; Shi, Lei; Southall, Noel T.; Marugan, Juan J.; Sibley, David R.

    2014-01-01

    A high-throughput screening campaign was conducted to interrogate a 380,000+ small-molecule library for novel D2 dopamine receptor modulators using a calcium mobilization assay. Active agonist compounds from the primary screen were examined for orthogonal D2 dopamine receptor signaling activities including cAMP modulation and β-arrestin recruitment. Although the majority of the subsequently confirmed hits activated all signaling pathways tested, several compounds showed a diminished ability to stimulate β-arrestin recruitment. One such compound (MLS1547; 5-chloro-7-[(4-pyridin-2-ylpiperazin-1-yl)methyl]quinolin-8-ol) is a highly efficacious agonist at D2 receptor–mediated G protein–linked signaling, but does not recruit β-arrestin as demonstrated using two different assays. This compound does, however, antagonize dopamine-stimulated β-arrestin recruitment to the D2 receptor. In an effort to investigate the chemical scaffold of MLS1547 further, we characterized a set of 24 analogs of MLS1547 with respect to their ability to inhibit cAMP accumulation or stimulate β-arrestin recruitment. A number of the analogs were similar to MLS1547 in that they displayed agonist activity for inhibiting cAMP accumulation, but did not stimulate β-arrestin recruitment (i.e., they were highly biased). In contrast, other analogs displayed various degrees of G protein signaling bias. These results provided the basis to use pharmacophore modeling and molecular docking analyses to build a preliminary structure-activity relationship of the functionally selective properties of this series of compounds. In summary, we have identified and characterized a novel G protein–biased agonist of the D2 dopamine receptor and identified structural features that may contribute to its biased signaling properties. PMID:24755247

  7. Guanine nucleotide regulation of dopamine receptor agonist affinity states in rat estradiol-induced pituitary tumors

    SciTech Connect

    Di Paolo, T.; Falardeau, P.

    1987-08-31

    The authors have investigated dopamine (DA) receptor agonist high- and low-affinity states in female rate estradiol-induced prolactin (PRL)-secreting pituitary tumors and intact pituitary tissue. Estradiol treatment increased the anterior pituitary weight 9-fold and plasma prolactin levels 74-fold and these measures are correlated (R = 0.745, n = 73, p < 0.001). Competition for (/sup 3/H)-spiperone binding to the DA receptor by apomorphine was compared in normal and adenomatous pituitary tissue. The inhibition constants (Ki) and the proportions of the two apomorphine sites are unchanged in tumors compared to intact pituitary tissue. Guanosine 5'-(..beta..-..gamma..-imino)triphosphate (Gpp(NH)p) causes complete conversion of the high into low affinity dopaminergic agonist site in normal pituitary and in tumors. These results suggest that rats with primary estradiol-induced pituitary tumors have normal and functional DA receptors. 9 references, 2 tables.

  8. Differential Desensitization Observed at Multiple Effectors of Somatic μ-Opioid Receptors Underlies Sustained Agonist-Mediated Inhibition of Proopiomelanocortin Neuron Activity.

    PubMed

    Fox, Philip D; Hentges, Shane T

    2017-09-06

    Activation of somatic μ-opioid receptors (MORs) in hypothalamic proopiomelanocortin (POMC) neurons leads to the activation of G-protein-coupled inward rectifier potassium (GIRK) channels and hyperpolarization, but in response to continued signaling MORs undergo acute desensitization resulting in robust reduction in the peak GIRK current after minutes of agonist exposure. We hypothesized that the attenuation of the GIRK current would lead to a recovery of neuronal excitability whereby desensitization of the receptor would lead to a new steady state of POMC neuron activity reflecting the sustained GIRK current observed after the initial decline from peak with continued agonist exposure. However, electrophysiologic recordings and GCaMP6f Ca(2+) imaging in POMC neurons in mouse brain slices indicate that maximal inhibition of cellular activity by these measures can be maintained after the GIRK current declines. Blockade of the GIRK current by Ba(2+) or Tertiapin-Q did not disrupt the sustained inhibition of Ca(2+) transients in the continued presence of agonist, indicating the activation of an effector other than GIRK channels. Use of an irreversible MOR antagonist and Furchgott analysis revealed a low receptor reserve for the activation of GIRK channels but a >90% receptor reserve for the inhibition of Ca(2+) events. Altogether, the data show that somatodendritic MORs in POMC neurons inhibit neuronal activity through at least two effectors with distinct levels of receptor reserve and that differentially reflect receptor desensitization. Thus, in POMC cells, the decline in the GIRK current during prolonged MOR agonist exposure does not reflect an increase in cellular activity as expected.SIGNIFICANCE STATEMENT Desensitization of the μ-opioid receptor (MOR) is thought to underlie the development of cellular tolerance to opiate therapy. The present studies focused on MOR desensitization in hypothalamic proopiomelanocortin (POMC) neurons as these neurons produce the

  9. Inhibition of K+-stimulated [3H]dopamine and [14C]acetylcholine release by the putative dopamine autoreceptor agonist, B-HT 920.

    PubMed

    Schmidt, C J; Lobur, A; Lovenberg, W

    1986-12-01

    The inhibition of K+-stimulated [3H]dopamine and [14C]acetylcholine release from preloaded rat striatal slices was used to examine the presynaptic selectivity of the putative dopamine autoreceptor agonist, B-HT 920. In the micromolar range, B-HT 920 caused a concentration-dependent inhibition of the release of both labeled neurotransmitters as evoked by 20 mM K+. The effect of B-HT 920 on both [3H]dopamine and [14C]acetylcholine release was completely blocked by (+) butaclamol but not by (-) butaclamol. Sulpiride, a selective D2 antagonist, similarly blocked the inhibitory effect of B-HT 920 on the release of both labeled neurotransmitters indicating both responses were mediated by D2 receptors. (+) Butaclamol alone elevated stimulated [3H]dopamine release suggesting a significant amount of autoreceptor occupancy by endogenously released dopamine. Experiments with tolazoline and the alpha 2 agonist, B-HT 933, did not suggest any involvement of alpha-adrenoceptor activity in the inhibitory effects of B-HT 920 on the release of either transmitter. Inhibition of release was a selective effect of B-HT 920 as the drug was without effect on the K+-stimulated release of [3H]serotonin. The results indicate that in vitro B-HT 920 is active of both pre- and postsynaptic dopamine receptors in contrast to the pattern of effects observed after its in vivo administration.

  10. The antilipolytic agent 3,5-dimethylpyrazole inhibits insulin release in response to both nutrient secretagogues and cyclic adenosine monophosphate agonists in isolated rat islets.

    PubMed

    Masiello, P; Novelli, M; Bombara, M; Fierabracci, V; Vittorini, S; Prentki, M; Bergamini, E

    2002-01-01

    This study intended to test the hypothesis that intracellular lipolysis in the pancreatic beta cells is implicated in the regulation of insulin secretion stimulated by nutrient secretagogues or cyclic adenosine monophosphate (cAMP) agonists. Indeed, although lipid signaling molecules were repeatedly reported to influence beta-cell function, the contribution of intracellular triglycerides to the generation of these molecules has remained elusive. Thus, we have studied insulin secretion of isolated rat pancreatic islets in response to various secretagogues in the presence or absence of 3,5-dimethylpyrazole (DMP), a water-soluble and highly effective antilipolytic agent, as previously shown in vivo. In vitro exposure of islets to DMP resulted in an inhibition (by approximately 50%) of the insulin release stimulated not only by high glucose, but also by another nutrient secretagogue, 2-ketoisocaproate, as well as the cAMP agonists 3-isobutyl-1-methylxanthine and glucagon. The inhibitory effect of DMP, which was not due to alteration of islet glucose oxidation, could be reversed upon addition of sn-1,2-dioctanoylglycerol, a synthetic diglyceride, which activates protein kinase C. The results provide direct pharmacologic evidence supporting the concept that endogenous beta-cell lipolysis plays an important role in the generation of lipid signaling molecules involved in the control of insulin secretion in response to both fuel stimuli and cAMP agonists.

  11. Receptor reserve for 5-hydroxytryptamine1A-mediated inhibition of serotonin synthesis: possible relationship to anxiolytic properties of 5-hydroxytryptamine1A agonists.

    PubMed

    Meller, E; Goldstein, M; Bohmaker, K

    1990-02-01

    The irreversible receptor antagonist N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) was used to determine the relationship between receptor occupancy and response at central 5-hydroxytryptamine1A (5-HT1A) serotonin receptors mediating the inhibition of serotonin synthesis in rat cortex and hippocampus. Rats were treated with vehicle or EEDQ (2 or 6 mg/kg) and 24 hr later dose-response curves were constructed for inhibition of 5-hydroxytrytophan (5-HTP) accumulation (after decarboxylase inhibition with NSD-1015) by the selective 5-HT1A agonists 8-hydroxy-2(di-n-propylamino)tetralin (8-OH-DPAT) (0.01-3 mg/kg), buspirone (0.1-7.5 mg/kg), and ipsapirone (0.1-6.25 mg/kg) and the 5-HT1A agonist/antagonist BMY 7378 (0.015-5 mg/kg). In vehicle-pretreated rats, a similar maximal inhibition of 5-HT synthesis (range, 52-59%) was observed in both brain areas with 8-OH-DPAT, buspirone, and ipsapirone. These three agonists were also more potent in reducing 5-HTP accumulation in the cortex than in the hippocampus (ED50, 8-OH-DPAT, 14 and 30 microgram/kg; buspirone, 0.42 and 0.63 mg/kg; ipsapirone, 0.44 and 1.26 mg/kg, respectively). In the cortex, EEDQ treatment shifted the dose-response curves for 8-OH-DPAT, buspirone, and ipsapirone 8.6-, 2.0-, and 2.8-fold to the right, respectively. Corresponding rightward shifts in the hippocampus were smaller, 6.0-, 1.6-, and 2.1-fold, respectively. The EEDQ-induced shifts in the dose-response curves were accompanied by reductions in maximal response. In contrast, whereas the maximal inhibition of cortical 5-HTP accumulation by BMY 7378 (55%) was similar to that obtained with the agonists, maximal response in the hippocampus was much smaller (32%). Furthermore, in both brain regions EEDQ reduced the maximal response to BMY 7378 without shifting the dose-response curves. Analysis of the data by the double-reciprocal method of Furchgott, followed by calculation of fractional receptor occupancy for each dose of agonist, revealed a

  12. 5HT1A receptor agonist differentially increases cyclic AMP concentration in intact and lesioned goldfish retina. In vitro inhibition of outgrowth by forskolin.

    PubMed

    Urbina, M; Schmeer, C; Lima, L

    1996-11-01

    5HT1A receptors occur in the retina of various species and the administration of 5HT1A agonists results in the inhibition of outgrowth from postcrush goldfish retinal explants. The levels of cyclic AMP (cAMP) play a role in the modulation of the outgrowth of the nevous system. Moreover, the stimulation of central 5HT1A receptors with the agonist 8-hydroxy-2-(di-n-propylamino)tetralin has been reported to produce an increase or decrease in the activity of adenylate cyclase. In the present investigation we studied the effect of adenylate cyclase stimulation by forskolin, as well as the modulatory effects of 5HT1A receptor agonists and antagonists on the production of cAMP in the goldfish retina, and on the outgrowth of this tissue in vitro. 8-Hydroxy-2-(di-n-propylamino)tetralin produced a significant and dose-dependent increase in cAMP concentration. This effect was not additive to the stimulation produced by forskolin. By contrast, as previously described, the 5HT1A agonist decreased cAMP concentration in the hippocampus of the rat. Both effects were significantly impaired by the 5HT1A antagonist WAY-100,135. A significant effect of the antagonist alone was observed only in the goldfish retina. The increase in cAMP levels was greater in the intact than in the postcrush retina. In addition, forskolin decreased the outgrowth of postcrush retinal explants in a dose-dependent manner, suggesting the importance of critical levels of cAMP in this process. Taken together, 5HT1A receptors seem to be positively coupled to adenylate cyclase in the goldfish retina, where cAMP plays a role as a modulator of outgrowth and regeneration. The inhibitory effect of 5HT1A receptor agonists on retinal outgrowth might be mediated through the production of cAMP. The activation of other subtypes of 5HT receptors positively coupled to adenylate cyclase by the 5HT1A agonist, such as 5HT7, cannot be discarded.

  13. Inhibition of cyst growth in PCK and Wpk rat models of polycystic kidney disease with low doses of peroxisome proliferator-activated receptor γ agonists

    PubMed Central

    Flaig, Stephanie M.; Gattone, Vincent H.

    2016-01-01

    Abstract Background and Objectives The studies were designed to test the efficacy of two peroxisome proliferator-activated receptor γ (PPARγ) agonists in two rodent models of polycystic kidney disease (PKD). Materials and Methods The PCK rat is a slowly progressing cystic model while the Wpk-/- rat is a rapidly progressing model. PCK rats were fed with a pharmacological (0.4 mg/kg body weight [BW]) and a sub-pharmacological (0.04 mg/kg BW) dose of rosiglitazone (week 4–28). Wpk-/- rats were fed with pharmacological (2.0 mg/kg BW) and sub-pharmacologic (0.2 mg/kg BW) doses of pioglitazone from day 5 to 18. At termination, kidney weights of treated versus untreated cystic animals were used to determine efficacy. The current studies were also compared with previous studies containing higher doses of PPARγ agonists. The concentrations used in the animals were calculated with reference to equivalent human doses for both drugs. Results The current studies demonstrate: 1) that low, pharmacologically relevant, doses of the PPARγ agonists effectively inhibit cyst growth; 2) there is a class action of the drugs with both commercially available PPARγ agonists, rosiglitazone, and pioglitazone, inhibiting cyst growth; 3) the drugs showed efficacy in two different preclinical cystic models. In the PCK rat, animals fed with a sub-pharmacological dose of rosiglitazone for 24 weeks had significantly lower kidney weights than untreated animals (3.68 ± 0.13 g vs. 4.17 ± 0. 11 g, respectively, P < 0.01) while treatment with a pharmacologic dose had no significant effect on kidney weight. The rapidly progressing Wpk-/- rats were fed with pharmacological and sub-pharmacologic doses of pioglitazone from day 5 to 18 and the kidneys were compared with non-treated, cystic animals. Kidney weights on the pharmacologic dose were not statistically lower than the untreated animals while rats fed a sub-pharmacologic dose showed a significant decrease compared with untreated animals (3

  14. PPAR-α and -γ but not -δ agonists inhibit airway inflammation in a murine model of asthma: in vitro evidence for an NF-κB-independent effect

    PubMed Central

    Trifilieff, Alexandre; Bench, Anne; Hanley, Marcus; Bayley, Debbie; Campbell, Emma; Whittaker, Paul

    2003-01-01

    Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors that have been proposed to regulate inflammation by antagonising the nuclear factor-κB (NF-κB) signalling pathway. We investigated the role of PPARs using synthetic agonists in murine models of airway inflammation, and addressed the possible effect on NF-κB signalling in vitro using a human epithelial cell line, A549. Sensitised BALB/c mice exposed to an aerosol solution of ovalbumin had an increased number of airway eosinophils, neutrophils and lymphocytes. When given intranasally an hour before the aerosol challenge, a PPAR-α (GW 9578) and PPAR-γ (GI 262570) selective agonist as well as a dual PPAR-α/γ (GW 2331) agonist selectively inhibited allergen-induced bronchoalveolar lavage eosinophil and lymphocyte but not neutrophil influx. In contrast, a PPAR-δ agonist (GW 501516) was inactive. When given intranasally an hour before challenge, PPAR-α and PPAR-γ selective agonists as well as a dual PPAR-α/γ agonist did not inhibit lipopolysaccharide-induced bronchoalveolar lavage neutrophil influx or tumour necrosis factor-α (TNF-α) and KC production. In A549 cells, selective agonists for PPAR-α, -γ and -δ did not inhibit intracellular adhesion molecule-1 expression following stimulation with proinflammatory cytokines. In addition, IL-8 release and the activation of an NF-κB-responsive reporter gene construct were inhibited only at micromolar concentrations, suggesting that these effects were not PPAR-mediated. Our in vivo data show that agonists of PPAR-α and -γ, but not -δ, inhibit allergen-induced bronchoalveolar lavage eosinophil and lymphocyte influx. In vitro data suggest that this effect might not be mediated by antagonism of the NF-κB pathway. PMID:12746235

  15. Toll-Like Receptor 9-Mediated Inflammation Triggers Alveolar Bone Loss in Experimental Murine Periodontitis

    PubMed Central

    Kim, Paul D.; Xia-Juan, Xia; Crump, Katie E.; Abe, Toshiharu; Hajishengallis, George

    2015-01-01

    Chronic periodontitis is a local inflammatory disease induced by a dysbiotic microbiota and leading to destruction of the tooth-supporting structures. Microbial nucleic acids are abundantly present in the periodontium, derived through release after phagocytic uptake of microbes and/or from biofilm-associated extracellular DNA. Binding of microbial DNA to its cognate receptors, such as Toll-like receptor 9 (TLR9), can trigger inflammation. In this study, we utilized TLR9 knockout (TLR9−/−) mice and wild-type (WT) controls in a murine model of Porphyromonas gingivalis-induced periodontitis and report the first in vivo evidence that TLR9 signaling mediates the induction of periodontal bone loss. P. gingivalis-infected WT mice exhibited significantly increased bone loss compared to that in sham-infected WT mice or P. gingivalis-infected TLR9−/− mice, which were resistant to bone loss. Consistent with this, the expression levels of interleukin 6 (IL-6), tumor necrosis factor (TNF), and receptor-activator of nuclear factor kappa B ligand (RANKL) were significantly elevated in the gingival tissues of the infected WT mice but not in infected TLR9−/− mice compared to their levels in controls. Ex vivo studies using splenocytes and bone marrow-derived macrophages revealed significantly diminished cytokine production in TLR9−/− cells relative to the cytokine production in WT cells in response to P. gingivalis, thereby implicating TLR9 in inflammatory responses to this organism. Intriguingly, compared to the cytokine production in WT cells, TLR9−/− cells exhibited significantly decreased proinflammatory cytokine production upon challenge with lipopolysaccharide (LPS) (TLR4 agonist) or Pam3Cys (TLR2 agonist), suggesting possible cross talk between TLR9, TLR4, and TLR2. Collectively, our results provide the first proof-of-concept evidence implicating TLR9-triggered inflammation in periodontal disease pathogenesis, thereby identifying a new potential

  16. The effects of central aromatic amino acid DOPA decarboxylase inhibition on the motor actions of L-DOPA and dopamine agonists in MPTP-treated primates

    PubMed Central

    Treseder, Sarah A; Jackson, Michael; Jenner, Peter

    2000-01-01

    Endogenous L-DOPA may act as a neuromodulator contributing to the production of motor activity. We now investigate the effects of the centrally acting aromatic amino acid dopa decarboxylase (AADC) inhibitor NSD-1015 (3-hydroxybenzyl hydrazine) on the motor actions of L-DOPA and dopamine agonist drugs in MPTP treated common marmosets.Pretreatment with NSD-1015 (10–50 mg kg−1; i.p.) worsened baseline motor deficits in MPTP-treated common marmosets. Similarly, it abolished L-DOPA (5–18 mg kg−1 s.c.) induced locomotor activity and reversal of disability. NSD-1015 pretreatment inhibited dopamine formation and elevated L-DOPA levels in plasma.The increase in locomotor activity and improvement in disability produced by the administration of the D-1 agonist A-86929 (0.03–0.04 mg kg−1 s.c.) or the D-2 agonist quinpirole (0.05–0.3 mg kg−1 i.p.) was abolished by NSD-1015 (25 mg kg−1 i.p.) pretreatment. While the effects of a low dose combination of A-86929 (0.04 mg kg−1 s.c.) and quinpirole (0.05 mg kg−1 i.p.) were inhibited by NSD-1015 (25 mg kg−1 i.p.), there was little effect on the action of a high dose combination of these drugs (0.08 mg kg−1 A-86929 and 0.1 mg kg−1 quinpirole).Following central AADC inhibition with NSD-1015 (25 mg kg−1 i.p.), locomotor behaviour induced by administration of high dose combinations of A-86929 (0.08 mg kg−1 s.c.) and quinpirole (0.1 mg kg−1 i.p.) was unaffected by L-DOPA (5 mg kg−1 s.c.) pretreatment.These results do not support a role for endogenous L-DOPA in spontaneous or drug induced locomotor activity. Rather, they strengthen the argument for the importance of endogenous dopaminergic tone in the motor actions of dopamine agonists. PMID:10742291

  17. δ-Opioid receptor agonists inhibit migraine-related hyperalgesia, aversive state and cortical spreading depression in mice

    PubMed Central

    Pradhan, Amynah A; Smith, Monique L; Zyuzin, Jekaterina; Charles, Andrew

    2014-01-01

    Background and Purpose Migraine is an extraordinarily common brain disorder for which treatment options continue to be limited. Agonists that activate the δ-opioid receptor may be promising for the treatment of migraine as they are highly effective for the treatment of chronic rather than acute pain, do not induce hyperalgesia, have low abuse potential and have anxiolytic and antidepressant properties. The aim of this study was to investigate the therapeutic potential of δ-opioid receptor agonists for migraine by characterizing their effects in mouse migraine models. Experimental Approach Mechanical hypersensitivity was assessed in mice treated with acute and chronic doses of nitroglycerin (NTG), a known human migraine trigger. Conditioned place aversion to NTG was also measured as a model of migraine-associated negative affect. In addition, we assessed evoked cortical spreading depression (CSD), an established model of migraine aura, in a thinned skull preparation. Key Results NTG evoked acute and chronic mechanical and thermal hyperalgesia in mice, as well as conditioned place aversion. Three different δ-opioid receptor agonists, SNC80, ARM390 and JNJ20788560, significantly reduced NTG-evoked hyperalgesia. SNC80 also abolished NTG-induced conditioned place aversion, suggesting that δ-opioid receptor activation may also alleviate the negative emotional state associated with migraine. We also found that SNC80 significantly attenuated CSD, a model that is considered predictive of migraine preventive therapies. Conclusions and Implications These data show that δ-opioid receptor agonists modulate multiple basic mechanisms associated with migraine, indicating that δ-opioid receptors are a promising therapeutic target for this disorder. PMID:24467301

  18. A comparative review of the tolerability profiles of dopamine agonists in the treatment of hyperprolactinaemia and inhibition of lactation.

    PubMed

    Webster, J

    1996-04-01

    Dopamine agonists are the treatment of choice for the majority of patients with hyperprolactinaemic disorders. Although characterised by a relatively high incidence of adverse effects, most commonly gastrointestinal, cardiovascular and neurological, these are usually mild and transient, and can be minimised by starting with a low dose and gradually increasing it, or taking the drug with food or while recumbent. Bromocriptine, introduced in 1971, is the reference preparation against which newer dopamine agonists are compared. It is effective in suppressing prolactin secretion, reducing prolactinoma size and restoring gonadal function. However, up to 12% of patients cannot tolerate the drug at therapeutic dosages. Cabergoline, a long-acting dopamine agonist administered once or twice weekly, has been shown to be significantly more effective than bromocriptine in suppressing prolactin secretion in hyperprolactinaemic patients, and is better tolerated, particularly in terms of nausea and vomiting. In suppressing physiological lactation, cabergoline is at least as effective as bromocriptine, and is associated with significantly fewer rebound symptoms and adverse effects. Quinagolide is a non-ergot dopamine agonist that is administered once daily. It has similar efficacy to bromocriptine, but is probably less effective than cabergoline in hyperprolactinaemic patients; it is not licensed for suppression of lactation. It is better tolerated than twice-daily bromocriptine, but is probably inferior to cabergoline in this regard. Neither bromocriptine, cabergoline nor quinagolide has been associated with any detrimental effect on pregnancy or fetal development. However, experience with bromocriptine is far more extensive; thus, for women requiring treatment for subfertility, this drug remains the treatment of choice in most centres, with cabergoline and quinagolide as acceptable second-line drugs in bromocriptine-intolerant patients. In hyperprolactinaemic men

  19. Sustained Toll-Like Receptor 9 Activation Promotes Systemic and Cardiac Inflammation, and Aggravates Diastolic Heart Failure in SERCA2a KO Mice

    PubMed Central

    Dhondup, Yangchen; Sjaastad, Ivar; Scott, Helge; Sandanger, Øystein; Zhang, Lili; Haugstad, Solveig Bjærum; Aronsen, Jan Magnus; Ranheim, Trine; Holmen, Sigve Dhondup; Alfsnes, Katrine; Ahmed, Muhammad Shakil; Attramadal, Håvard; Gullestad, Lars; Aukrust, Pål; Christensen, Geir; Yndestad, Arne; Vinge, Leif Erik

    2015-01-01

    Aim Cardiac inflammation is important in the pathogenesis of heart failure. However, the consequence of systemic inflammation on concomitant established heart failure, and in particular diastolic heart failure, is less explored. Here we investigated the impact of systemic inflammation, caused by sustained Toll-like receptor 9 activation, on established diastolic heart failure. Methods and Results Diastolic heart failure was established in 8–10 week old cardiomyocyte specific, inducible SERCA2a knock out (i.e., SERCA2a KO) C57Bl/6J mice. Four weeks after conditional KO, mice were randomized to receive Toll-like receptor 9 agonist (CpG B; 2μg/g body weight) or PBS every third day. After additional four weeks, echocardiography, phase contrast magnetic resonance imaging, histology, flow cytometry, and cardiac RNA analyses were performed. A subgroup was followed, registering morbidity and death. Non-heart failure control groups treated with CpG B or PBS served as controls. Our main findings were: (i) Toll-like receptor 9 activation (CpG B) reduced life expectancy in SERCA2a KO mice compared to PBS treated SERCA2a KO mice. (ii) Diastolic function was lower in SERCA2a KO mice with Toll-like receptor 9 activation. (iii) Toll-like receptor 9 stimulated SERCA2a KO mice also had increased cardiac and systemic inflammation. Conclusion Sustained activation of Toll-like receptor 9 causes cardiac and systemic inflammation, and deterioration of SERCA2a depletion-mediated diastolic heart failure. PMID:26461521

  20. Quercetin acutely relaxes airway smooth muscle and potentiates β-agonist-induced relaxation via dual phosphodiesterase inhibition of PLCβ and PDE4

    PubMed Central

    Emala, Charles W.

    2013-01-01

    Asthma is a disease of the airways with symptoms including exaggerated airway narrowing and airway inflammation. Early asthma therapies used methylxanthines to relieve symptoms, in part, by inhibiting cyclic nucleotide phosphodiesterases (PDEs), the enzyme responsible for degrading cAMP. The classification of tissue-specific PDE subtypes and the clinical introduction of PDE-selective inhibitors for chronic obstructive pulmonary disease (i.e., roflumilast) have reopened the possibility of using PDE inhibition in the treatment of asthma. Quercetin is a naturally derived PDE4-selective inhibitor found in fruits, vegetables, and tea. We hypothesized that quercetin relaxes airway smooth muscle via cAMP-mediated pathways and augments β-agonist relaxation. Tracheal rings from male A/J mice were mounted in myographs and contracted with acetylcholine (ACh). Addition of quercetin (100 nM-1 mM) acutely and concentration-dependently relaxed airway rings precontracted with ACh. In separate studies, pretreatment with quercetin (100 μM) prevented force generation upon exposure to ACh. In additional studies, quercetin (50 μM) significantly potentiated isoproterenol-induced relaxations. In in vitro assays, quercetin directly attenuated phospholipase C activity, decreased inositol phosphate synthesis, and decreased intracellular calcium responses to Gq-coupled agonists (histamine or bradykinin). Finally, nebulization of quercetin (100 μM) in an in vivo model of airway responsiveness significantly attenuated methacholine-induced increases in airway resistance. These novel data show that the natural PDE4-selective inhibitor quercetin may provide therapeutic relief of asthma symptoms and decrease reliance on short-acting β-agonists. PMID:23873842

  1. Further Advances in Optimizing (2-Phenylcyclopropyl)methylamines as Novel Serotonin 2C Agonists: Effects on Hyperlocomotion, Prepulse Inhibition, and Cognition Models.

    PubMed

    Cheng, Jianjun; Giguere, Patrick M; Schmerberg, Claire M; Pogorelov, Vladimir M; Rodriguiz, Ramona M; Huang, Xi-Ping; Zhu, Hu; McCorvy, John D; Wetsel, William C; Roth, Bryan L; Kozikowski, Alan P

    2016-01-28

    A series of novel compounds with two halogen substituents have been designed and synthesized to further optimize the 2-phenylcyclopropylmethylamine scaffold in the quest for drug-like 5-HT2C agonists. Compound (+)-22a was identified as a potent 5-HT2C receptor agonist, with good selectivity against the 5-HT2B and the 5-HT2A receptors. ADMET assays showed that compound (+)-22a possessed desirable properties in terms of its microsomal stability, and CYP and hERG inhibition, along with an excellent brain penetration profile. Evaluation of (+)-22a in animal models of schizophrenia-related behaviors revealed that it had a desirable activity profile, as it reduced d-amphetamine-stimulated hyperlocomotion in the open field test, it restored d-amphetamine-disrupted prepulse inhibition, it induced cognitive improvements in the novel object recognition memory test in NR1-KD animals, and it produced very little catalepsy relative to haloperidol. These data support the further development of (+)-22a as a drug candidate for the treatment of schizophrenia.

  2. The PPARβ/δ agonist GW501516 attenuates peritonitis in peritoneal fibrosis via inhibition of TAK1-NFκB pathway in rats.

    PubMed

    Su, Xuesong; Zhou, Guangyu; Wang, Yanqiu; Yang, Xu; Li, Li; Yu, Rui; Li, Detian

    2014-06-01

    Peritoneal fibrosis is a common consequence of long-term peritoneal dialysis (PD), and peritonitis is a factor in its onset. Agonist-bound peroxisome proliferator-activated receptors (PPARs) function as key regulators of energy metabolism and inflammation. Here, we examined the effects of PPARβ/δ agonist GW501516 on peritonitis in a rat peritoneal fibrosis model. Peritoneal fibrosis secondary to inflammation was induced into uremic rats by daily injection of Dianeal 4.25% PD solutions along with six doses of lipopolysaccharide before commencement of GW501516 treatment. Normal non-uremic rats served as control, and all rats were fed with a control diet or a GW501516-containing diet. Compared to control group, exposure to PD fluids caused peritoneal fibrosis that was accompanied by increased mRNA levels of monocyte chemoattractant protein-1, tumor necrotic factor-α, and interleukin-6 in the uremic rats, and these effects were prevented by GW501516 treatment. Moreover, GW501516 was found to attenuate glucose-stimulated inflammation in cultured rat peritoneal mesothelial cells via inhibition of transforming growth factor-β-activated kinase 1 (TAK1), and nuclear factor kappa B (NFκB) signaling pathway (TAK1-NFκB pathway), a main inflammation regulatory pathway. In conclusion, inhibition of TAK1-NFκB pathway with GW501516 may represent a novel therapeutic approach to ameliorate peritonitis-induced peritoneal fibrosis for patients on PD.

  3. The C-terminus of murine S100A9 protein inhibits hyperalgesia induced by the agonist peptide of protease-activated receptor 2 (PAR2)

    PubMed Central

    Dale, C S; Cenac, N; Britto, L R G; Juliano, M A; Juliano, L; Vergnolle, N; Giorgi, R

    2006-01-01

    Background and purpose: S100A9 protein induces anti-nociception in rodents, in different experimental models of inflammatory pain. Herein, we investigated the effects of a fragment of the C-terminus of S100A9 (mS100A9p), on the hyperalgesia induced by serine proteases, through the activation of protease-activated receptor-2 (PAR2). Experimental approach: Mechanical and thermal hyperalgesia induced by PAR2 agonists (SLIGRL-NH2 and trypsin) was measured in rats submitted to the paw pressure or plantar tests, and Egr-1 expression was determined by immunohistochemistry in rat spinal cord dorsal horn. Calcium flux in human embryonic kidney cells (HEK), which naturally express PAR2, in Kirsten virus-transformed kidney cells, transfected (KNRK-PAR2) or not (KNRK) with PAR2, and in mouse dorsal root ganglia neurons (DRG) was measured by fluorimetric methods. Key results: mS100A9p inhibited mechanical hyperalgesia induced by trypsin, without modifying its enzymatic activity. Mechanical and thermal hyperalgesia induced by SLIGRL-NH2 were inhibited by mS100A9p. SLIGRL-NH2 enhanced Egr-1 expression, a marker of nociceptor activation, and this effect was inhibited by concomitant treatment with mS100A9p. mS100A9p inhibited calcium mobilization in DRG neurons in response to the PAR2 agonists trypsin and SLIGRL-NH2, but also in response to capsaicin and bradykinin, suggesting a direct effect of mS100A9 on sensory neurons. No effect on the calcium flux induced by trypsin or SLIGRL in HEK cells or KNRK-PAR2 cells was observed. Conclusions and implications: These data demonstrate that mS100A9p interferes with mechanisms involved in nociception and hyperalgesia and modulates, possibly directly on sensory neurons, the PAR2-induced nociceptive signal. PMID:16967049

  4. Chrysin, a PPAR-γ agonist improves myocardial injury in diabetic rats through inhibiting AGE-RAGE mediated oxidative stress and inflammation.

    PubMed

    Rani, Neha; Bharti, Saurabh; Bhatia, Jagriti; Nag, T C; Ray, Ruma; Arya, Dharamvir Singh

    2016-04-25

    AGE-RAGE interaction mediated oxidative stress and inflammation is the key mechanism involved in the pathogenesis of cardiovascular disease in diabetes. Inhibition of AGE-RAGE axis by several PPAR-γ agonists has shown positive results in ameliorating cardio-metabolic disease conditions. Chrysin, a natural flavonoid has shown to possess PPAR-γ agonist activity along with antioxidant and anti-inflammatory effect. Therefore, the present study was designed to evaluate the effect of chrysin in isoproterenol-induced myocardial injury in diabetic rats. In male albino Wistar rats, diabetes was induced by single injection of streptozotocin (70 mg/kg, i.p.). After confirmation of the diabetes, rats were treated with vehicle (1.5 mL/kg, p.o.), chrysin (60 mg/kg, p.o.) or PPAR-γ antagonist GW9662 (1 mg/kg, i.p.) for 28 days. Simultaneously, on 27th and 28th day myocardial injury was induced by isoproterenol (85 mg/kg, s.c.). Chrysin significantly ameliorated cardiac dysfunction as reflected by improved MAP, ±LVdP/dtmax and LVEDP in diabetic rats. This improvement was associated with increased PPAR-γ expression and reduced RAGE expression in diabetic rats. Chrysin significantly decreased inflammation through inhibiting NF-κBp65/IKK-β expression and TNF-α level. Additionally, chrysin significantly reduced apoptosis as indicated by augmented Bcl-2 expression and decreased Bax and caspase-3 expressions. Furthermore, chrysin inhibited nitro-oxidative stress by normalizing the alteration in 8-OHdG, GSH, TBARS, NO and CAT levels and Nox4, MnSOD, eNOS and NT expressions. Co-administration of GW9662 significantly blunted the chrysin mediated cardioprotective effect as there was increase in oxidative stress, inflammation and apoptosis markers. Chrysin significantly ameliorated isoproterenol-induced myocardial injury in diabetic rats via PPAR-γ activation and inhibition of AGE-RAGE mediated oxidative stress and inflammation.

  5. β2-Adrenoceptor agonists inhibit release of eosinophil-activating cytokines from human airway smooth muscle cells

    PubMed Central

    Hallsworth, Matthew P; Twort, Charles H C; Lee, Tak H; Hirst, Stuart J

    2001-01-01

    Airway smooth muscle (ASM) is a potential source of multiple pro-inflammatory cytokines during airway inflammation. β-Adrenoceptor agonist hyporesponsiveness is a characteristic feature of asthma, and interleukin (IL)-1β and tumour necrosis factor (TNF)-α are implicated in its cause. Here, the capacity of β-adrenoceptor agonists to prevent release of GM-CSF, RANTES, eotaxin and IL-8, elicited by IL-1β or TNFα, was examined in human ASM cells. Isoprenaline (∼EC50 150 nM), a non-selective β-adrenoceptor agonist, and salbutamol (∼EC50 25 nM), a selective β2-adrenoceptor agonist, attenuated release of GM-CSF, RANTES and eotaxin, but not IL-8 (EC50 >1 μM). The maximum extent of attenuation was RANTES ⩾ eotaxin > GM-CSF >> IL-8, and was prevented by either propranolol (1 μM), a non-selective β-adrenoceptor antagonist, or ICI 118511 (IC50 15 nM), a selective β2-adrenoceptor antagonist. The cyclic AMP-elevating agents, dibutyryl cyclic AMP (∼EC50 135 μM), forskolin (∼EC50 530 nM) and cholera toxin (∼EC50 575 pg ml−1) abolished IL-1β-induced release of GM-CSF, RANTES and eotaxin, but not IL-8. IL-1β (1 ng ml−1) attenuated early increases (up to 1 h) in cyclic AMP formation induced by salbutamol (1 μM), but not by forskolin (10 μM). The cyclo-oxygenase inhibitor, indomethacin (1 μM) prevented later increases (3 – 12 h) in IL-1β-stimulated cyclic AMP content, but did not prevent the attenuation by salbutamol of IL-1β-induced cytokine release. We conclude in human ASM cells that activation of β2-adrenoceptors and generation of cyclic AMP is negatively-linked to the release, elicited by IL-1β or TNFα, of eosinophil-activating cytokines such as GM-CSF, RANTES and eotaxin, but not IL-8. PMID:11159726

  6. Role of protein kinase C (PKC) in short- and long-term cellular responses: inhibition of agonist-mediated calcium transients and down-regulation of PKC

    SciTech Connect

    Fabbro, D.; Mazurek, N.; Borner, C.; Conscience, J.F.; Erne, P.

    1988-01-01

    Active tumor promoters such as 12-O-tetradecanoyl-phorbol-13-acetate (TPA) or membrane-diffusible synthetic diacylglycerols such as 1,2-dioctanoyl-sn-glycerol (DiC8), which specifically activate protein kinase C (PKC), inhibited the agonist-mediated rise in cytosolic calcium ((Ca2+)i) in a mast cell line (PB-3c) and human platelets. TPA inhibition of agonist-mediated calcium transient in platelets was readily reversed by the PKC inhibitor staurosporine. In contrast to DiCs, only active tumor promoters induced a time- and dose-dependent translocation of cytosolic PKC to membranes as determined both enzymatically or by immunoblotting. However, the concentration of TPA required to induce a half-maximal subcellular redistribution of immunodetectable PKC activity was an order of magnitude greater than the half-maximal dose required to inhibit the intracellular rise in (Ca2+)i. Thus, activation of PKC seems not to be exclusively coupled to its translocation to membranes, suggesting that translocation of PKC is mainly involved in the down-regulation of PKC. Down-regulation of immunoprecipitable PKC was studied in various human breast cancer cell lines that display differential growth inhibitory responses toward the tumor promoter. TPA induced translocation of (35S)methionine-prelabeled cytosolic 80 kDa PKC to membranes followed by complete degradation of the enzyme (t1/2 = 2 h) without affecting PKC synthesis. During prolonged TPA exposure, 20-80% of total 80 kDa PKC of control cells was still synthetized as a membrane-bound 74/80 kDa PKC doublet. Although both proteins lacked PKC activity and phorbol ester binding, they revealed structural similarity with the active 80 kDa PKC form of untreated cells.

  7. Eicosopentaneoic Acid and Other Free Fatty Acid Receptor Agonists Inhibit Lysophosphatidic Acid- and Epidermal Growth Factor-Induced Proliferation of Human Breast Cancer Cells.

    PubMed

    Hopkins, Mandi M; Zhang, Zhihong; Liu, Ze; Meier, Kathryn E

    2016-01-26

    Many key actions of ω-3 (n-3) fatty acids have recently been shown to be mediated by two G protein-coupled receptors (GPCRs) in the free fatty acid receptor (FFAR) family, FFA1 (GPR40) and FFA4 (GPR120). n-3 Fatty acids inhibit proliferation of human breast cancer cells in culture and in animals. In the current study, the roles of FFA1 and FFA4 were investigated. In addition, the role of cross-talk between GPCRs activated by lysophosphatidic acid (LPA), and the tyrosine kinase receptor activated by epidermal growth factor (EGF), was examined. In MCF-7 and MDA-MB-231 human breast cancer cell lines, both LPA and EGF stimulated proliferation, Erk activation, Akt activation, and CCN1 induction. LPA antagonists blocked effects of LPA and EGF on proliferation in MCF-7 and MDA-MB-231, and on cell migration in MCF-7. The n-3 fatty acid eicosopentaneoic acid inhibited LPA- and EGF-induced proliferation in both cell lines. Two synthetic FFAR agonists, GW9508 and TUG-891, likewise inhibited LPA- and EGF-induced proliferation. The data suggest a major role for FFA1, which was expressed by both cell lines. The results indicate that n-3 fatty acids inhibit breast cancer cell proliferation via FFARs, and suggest a mechanism involving negative cross-talk between FFARS, LPA receptors, and EGF receptor.

  8. Eicosopentaneoic Acid and Other Free Fatty Acid Receptor Agonists Inhibit Lysophosphatidic Acid- and Epidermal Growth Factor-Induced Proliferation of Human Breast Cancer Cells

    PubMed Central

    Hopkins, Mandi M.; Zhang, Zhihong; Liu, Ze; Meier, Kathryn E.

    2016-01-01

    Many key actions of ω-3 (n-3) fatty acids have recently been shown to be mediated by two G protein-coupled receptors (GPCRs) in the free fatty acid receptor (FFAR) family, FFA1 (GPR40) and FFA4 (GPR120). n-3 Fatty acids inhibit proliferation of human breast cancer cells in culture and in animals. In the current study, the roles of FFA1 and FFA4 were investigated. In addition, the role of cross-talk between GPCRs activated by lysophosphatidic acid (LPA), and the tyrosine kinase receptor activated by epidermal growth factor (EGF), was examined. In MCF-7 and MDA-MB-231 human breast cancer cell lines, both LPA and EGF stimulated proliferation, Erk activation, Akt activation, and CCN1 induction. LPA antagonists blocked effects of LPA and EGF on proliferation in MCF-7 and MDA-MB-231, and on cell migration in MCF-7. The n-3 fatty acid eicosopentaneoic acid inhibited LPA- and EGF-induced proliferation in both cell lines. Two synthetic FFAR agonists, GW9508 and TUG-891, likewise inhibited LPA- and EGF-induced proliferation. The data suggest a major role for FFA1, which was expressed by both cell lines. The results indicate that n-3 fatty acids inhibit breast cancer cell proliferation via FFARs, and suggest a mechanism involving negative cross-talk between FFARS, LPA receptors, and EGF receptor. PMID:26821052

  9. Compound A, a selective glucocorticoid receptor agonist, inhibits immunoinflammatory diabetes, induced by multiple low doses of streptozotocin in mice

    PubMed Central

    Saksida, T; Vujicic, M; Nikolic, I; Stojanovic, I; Haegeman, G; Stosic-Grujicic, S

    2014-01-01

    Background and Purpose Type 1 diabetes is a multifactorial inflammatory disease that develops as a result of deregulated immune responses, causing progressive autoimmune destruction of insulin-producing beta cells of pancreas. 2-((4-acetoxyphenyl)-2-chloro-N-methyl) ethylammonium chloride, compound A (CpdA), is a selective glucocorticoid receptor (GR) agonist that displays strong anti-inflammatory and immunomodulatory activities. We investigated the therapeutic effectiveness of CpdA in a pharmacological model of type 1 diabetes in mice. Experimental Approach The utility of CpdA in diabetes prevention was evaluated in vivo through its prophylactic administration to male C57BL/6 mice that received multiple low doses of streptozotocin for immunoinflammatory diabetes induction. The effect of CpdA on disease development was studied by measuring blood glucose and insulin level, histopathological examination, determination of the nature of infiltrating cells, pro- and anti-inflammatory cytokine production, and signalling pathways. Key Results Prophylactic in vivo therapy with CpdA conferred protection against development of immunoinflammatory diabetes in mice by dampening the M1/Th1/Th17 immune response and switching it towards an anti-inflammatory M2/Th2/Treg profile, thus preserving beta cell function. Conclusions and Implications Anti-diabetic properties of CpdA are mediated through modulation of immune cell-mediated pathways, but without triggering adverse events. These findings provide basic information for the therapeutic use of selective GR agonists in the amelioration of islet-directed autoimmunity. PMID:25158597

  10. Soluble epoxide hydrolase inhibition and peroxisome proliferator activated receptor γ agonist improve vascular function and decrease renal injury in hypertensive obese rats.

    PubMed

    Imig, John D; Walsh, Katie A; Hye Khan, Md Abdul; Nagasawa, Tasuku; Cherian-Shaw, Mary; Shaw, Sean M; Hammock, Bruce D

    2012-12-01

    Cardiometabolic syndrome occurs with obesity and consists of pathophysiological factors that increase the risk for cardiovascular events. Soluble epoxide hydrolase inhibition (sEHi) is a novel therapeutic approach that exerts renal and cardiovascular protection. Although sEHi as a therapeutic approach is promising, it could be more effective for the treatment of cardiometabolic syndrome when combined with peroxisome proliferator activated receptor γ (PPARγ) agonists. We hypothesized that the PPARγ agonist, rosiglitazone in combination with a sEHi (tAUCB) will provide synergistic actions to decrease blood pressure, improve vascular function, decrease inflammation, and prevent renal damage in spontaneously hypertensive obese rats (SHROB). SHROB were treated with rosiglitazone, tAUCB or the combination of tAUCB and rosiglitazone for four-weeks and compared with spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) rats. Blood pressure increased in SHROB (164 ± 7 mmHg) and decreased 10 mmHg when treated with rosiglitazone, tAUCB, or tAUCB and rosiglitazone. Mesenteric artery dilation to the K(ATP) channel opener pinacidil was attenuated in SHROB (E(Max) = 77 ± 7%), compared with WKY (E(Max) = 115 ± 19) and SHR (E(Max) = 93 ± 12%). Vasodilation to pinacidil was improved by rosiglitazone (E(Max) = 92 ± 14%) but not tAUCB. Renal macrophage infiltration increased in SHROB and significantly decreased with rosiglitazone or tAUCB and rosiglitazone treatment. Albuminuria was increased in SHROB (90 ± 20 mg/d) and was significantly decreased by the combination of tAUCB and rosiglitazone (37 ± 9 mg/d). Glomerular injury in SHROB was also significantly decreased by tAUCB and rosiglitazone. These results indicate that even though sEHi or PPARγ agonist have benefits when used individually, the combination is more beneficial for the multidisease features in cardiometabolic syndrome.

  11. Pro-cognitive and antipsychotic efficacy of the alpha7 nicotinic partial agonist SSR180711 in pharmacological and neurodevelopmental latent inhibition models of schizophrenia.

    PubMed

    Barak, Segev; Arad, Michal; De Levie, Amaya; Black, Mark D; Griebel, Guy; Weiner, Ina

    2009-06-01

    Schizophrenia symptoms can be segregated into positive, negative and cognitive, which exhibit differential sensitivity to drug treatments. Accumulating evidence points to efficacy of alpha7 nicotinic receptor (nAChR) agonists for cognitive deficits in schizophrenia but their activity against positive symptoms is thought to be minimal. The present study examined potential pro-cognitive and antipsychotic activity of the novel selective alpha7 nAChR partial agonist SSR180711 using the latent inhibition (LI) model. LI is the reduced efficacy of a previously non-reinforced stimulus to gain behavioral control when paired with reinforcement, compared with a novel stimulus. Here, no-drug controls displayed LI if non-reinforced pre-exposure to a tone was followed by weak but not strong conditioning (2 vs 5 tone-shock pairings). MK801 (0.05 mg/kg, i.p.) -treated rats as well as rats neonatally treated with nitric oxide synthase inhibitor L-NoArg (10 mg/kg, s.c.) on postnatal days 4-5, persisted in displaying LI with strong conditioning, whereas amphetamine (1 mg/kg) -treated rats failed to show LI with weak conditioning. SSR180711 (0.3, 1, 3 mg/kg, i.p.) was able to alleviate abnormally persistent LI produced by acute MK801 and neonatal L-NoArg; these models are believed to model cognitive aspects of schizophrenia and activity here was consistent with previous findings with alpha7-nAChR agonists. In addition, unexpectedly, SSR180711 (1, 3 mg/kg, i.p.) potentiated LI with strong conditioning in no-drug controls and reversed amphetamine-induced LI disruption, two effects considered predictive of activity against positive symptoms of schizophrenia. These findings suggest that SSR180711 may be beneficial not only for the treatment of cognitive symptoms in schizophrenia, as reported multiple times previously, but also positive symptoms.

  12. Corticosteroid dependent and independent effects of a cannabinoid agonist on core temperature, motor activity, and prepulse inhibition of the acoustic startle reflex in Wistar rats.

    PubMed

    Avdesh, Avdesh; Cornelisse, Vincent; Martin-Iverson, Mathew Thomas

    2012-03-01

    There are inconsistent reports on the effects of cannabinoid agonists on prepulse inhibition of the startle reflex (PPI) with increases, decreases, and no effects. It has been hypothesized that the conflicting observations may be as a result of modulation of the effects of cannabinoid agonists by the regulation of corticosteroid release. The purpose of the present study was to determine the effects of CP55940, a cannabinoid agonist, and metyrapone, a corticosteroid synthesis inhibitor on core temperature, motor activity, the startle reflex, and PPI. Startle responses were measured in 64 male Wistar rats while varying startling stimulus intensities, analogous to dose-response curves. A stimulus potency measure (ES(50)) and a response measure, the maximal achievable response (R (MAX)) were derived from the stimulus-response curves. CP55940 reduced core temperature and motor activity; these effects were potentiated by metyrapone. CP55940 increased R (MAX) of startle in the absence of a prepulse by a corticosteroid-dependent mechanism but decreased it when metyrapone was administered before CP55940, a corticosteroid-independent mechanism. The inverse of stimulus potency (ES(50)) was not affected by either drug alone but was increased by the combined drugs. CP55940 increased the prepulse motor gating effects and decreased the prepulse sensory gating effects of the same prepulses but only when given after metyrapone. The most parsimonious interpretation of these effects is that CP55940 has some effects through corticosteroid-dependent actions and opposite effects by corticosteroid-independent actions. These two putative sites of actions affect stimulus gating opposite to their effects on response gating.

  13. Soluble epoxide hydrolase inhibition and peroxisome proliferator activated receptor γ agonist improve vascular function and decrease renal injury in hypertensive obese rats

    PubMed Central

    Imig, John D; Walsh, Katie A; Khan, Md Abdul Hye; Nagasawa, Tasuku; Cherian-Shaw, Mary; Shaw, Sean M; Hammock, Bruce D

    2013-01-01

    Cardiometabolic syndrome occurs with obesity and consists of pathophysiological factors that increase the risk for cardiovascular events. Soluble epoxide hydrolase inhibition (sEHi) is a novel therapeutic approach that exerts renal and cardiovascular protection. Although sEHi as a therapeutic approach is promising, it could be more effective for the treatment of cardiometabolic syndrome when combined with peroxisome proliferator activated receptor γ (PPARγ) agonists. We hypothesized that the PPARγ agonist, rosiglitazone in combination with a sEHi (tAUCB) will provide synergistic actions to decrease blood pressure, improve vascular function, decrease inflammation, and prevent renal damage in spontaneously hypertensive obese rats (SHROB). SHROB were treated with rosiglitazone, tAUCB or the combination of tAUCB and rosiglitazone for four-weeks and compared with spontaneously hypertensive (SHR) and Wistar–Kyoto (WKY) rats. Blood pressure increased in SHROB (164 ±7 mmHg) and decreased 10 mmHg when treated with rosiglitazone, tAUCB, or tAUCB and rosiglitazone. Mesenteric artery dilation to the KATP channel opener pinacidil was attenuated in SHROB (EMax = 77 ±7%), compared with WKY (EMax = 115 ±19) and SHR (EMax = 93 ±12%). Vasodilation to pinacidil was improved by rosiglitazone (EMax = 92 ±14%) but not tAUCB. Renal macrophage infiltration increased in SHROB and significantly decreased with rosiglitazone or tAUCB and rosiglitazone treatment. Albuminuria was increased in SHROB (90 ±20 mg/d) and was significantly decreased by the combination of tAUCB and rosiglitazone (37 ±9 mg/d). Glomerular injury in SHROB was also significantly decreased by tAUCB and rosiglitazone. These results indicate that even though sEHi or PPARγ agonist have benefits when used individually, the combination is more beneficial for the multidisease features in cardiometabolic syndrome. PMID:23354399

  14. NO synthase inhibition attenuates EDHF-mediated relaxation induced by TRPV4 channel agonist GSK1016790A in the rat pulmonary artery: Role of TxA2.

    PubMed

    Addison, M Pule; Singh, Thakur Uttam; Parida, Subhashree; Choudhury, Soumen; Kasa, Jaya Kiran; Sukumaran, Susanth V; Darzi, Sajad Ahmad; Kandasamy, Kannan; Singh, Vishakha; Kumar, Dinesh; Mishra, Santosh Kumar

    2016-06-01

    The aim of the present study was to observe the concomitant activation of nitric oxide (NO) and endothelium-derived hyperpolarizing factor (EDHF) pathways by TRPV4 channel agonist GSK1016790A in the rat pulmonary artery and explore the mechanism by which NO synthase inhibition attenuates EDHF-mediated relaxation in endothelium-intact rat pulmonary artery. Tension experiments were conducted on the pulmonary artery from male Wistar rats. TRPV4 channel agonist GSK1016790A (GSK) caused concentration-dependent relaxation (Emax 86.9±4.6%; pD2 8.7±0.24) of the endothelium-intact rat pulmonary artery. Combined presence of apamin and TRAM-34 significantly attenuated the relaxation (Emax 61.1±6.0%) to GSK. l-NAME (100μM) significantly attenuated (8.2±2.9%) the relaxation response to GSK that was resistant to apamin plus TRAM-34. However, presence of ICI192605 or furegrelate alongwith l-NAME revealed the GSK-mediated EDHF-response (Emax of 28.5±5.2%; Emax 24.5±4.3%) in this vessel, respectively. Further, these two TxA2 modulators (ICI/furegrelate) alongwith l-NAME had no effect on SNP-induced endothelium-independent relaxation in comparison to l-NAME alone. This EDHF-mediated relaxation was sensitive to inhibition by K(+) channel blockers apamin and TRAM-34 or 60mMK(+) depolarizing solution. Further, combined presence of apamin and TRAM-34 in U46619 pre-contracted pulmonary arterial rings significantly reduced the maximal relaxation (Emax 71.6±6.9%) elicited by GSK, but had no effect on the pD2 (8.1±0.03) of the TRPV4 channel agonist in comparison to controls (Emax, 92.4±4.3% and pD2, 8.3±0.06). The present study suggests that NO and EDHF are released concomitantly and NO synthase inhibition attenuates GSK-induced EDHF response through thromboxane pathway in the rat pulmonary artery. Copyright © 2016 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

  15. The cAMP-producing agonist beraprost inhibits human vascular smooth muscle cell migration via exchange protein directly activated by cAMP

    PubMed Central

    McKean, Jenny S.; Murray, Fiona; Gibson, George; Shewan, Derryck A.; Tucker, Steven J.; Nixon, Graeme F.

    2015-01-01

    Aims During restenosis, vascular smooth muscle cells (VSMCs) migrate from the vascular media to the developing neointima. Preventing VSMC migration is therefore a therapeutic target for restenosis. Drugs, such as prostacyclin analogues, that increase the intracellular concentration of cyclic adenosine monophosphate (cAMP) can inhibit VSMC migration, but the mechanisms via which this occurs are unknown. Two main downstream mediators of cAMP are protein kinase A (PKA) and exchange protein directly activated by cAMP (Epac). This study has examined the effects of the prostacyclin analogue beraprost on VSMC migration and investigated the intracellular pathways involved. Methods and results In a chemotaxis chamber, human saphenous vein VSMC migrated towards a platelet-derived growth-factor-BB (PDGF) chemogradient. Incubation with therapeutically relevant concentrations of cAMP-producing agonist beraprost significantly decreased PDGF-induced migration. Direct activation of either PKA or Epac inhibited migration whereas inhibition of PKA did not prevent the anti-migratory effect of beraprost. Direct activation of Epac also prevented hyperplasia in ex vivo serum-treated human veins. Using fluorescence resonance energy transfer, we demonstrated that beraprost activated Epac but not PKA. The mechanisms of this Epac-mediated effect involved activation of Rap1 with subsequent inhibition of RhoA. Cytoskeletal rearrangement at the leading edge of the cell was consequently inhibited. Interestingly, Epac1 was localized to the leading edge of migrating VSMC. Conclusions These results indicate that therapeutically relevant concentrations of beraprost can inhibit VSMC migration via a previously unknown mechanism involving the cAMP mediator Epac. This may provide a novel target that could blunt neointimal formation. PMID:26092100

  16. Obesity-induced p53 activation in insulin-dependent and independent tissues is inhibited by beta-adrenergic agonist in diet-induced obese rats.

    PubMed

    Zand, Hamid; Homayounfar, Reza; Cheraghpour, Makan; Jeddi-Tehrani, Mahmood; Ghorbani, Arman; Pourvali, Katayoun; Soltani, Sama Reza

    2016-02-15

    The purpose of this study was to assay the role of beta-adrenergic receptor signaling in the regulation of obesity-induced p53 in high fat feeding obese rats. The role of beta-adrenergic receptor/cyclic AMP in the regulation of p53 and its downstream mediators was evaluated by western blot and real-time quantitative RT-PCR among diet induced rats. Beta-adrenergic receptor agonist, isoproterenol, and an adenylate cyclase activator, forskolin, at a single dose significantly reduced insulin resistance consistent with a decrease in total and phospho-p53 levels in insulin and non-insulin metabolic target tissues. The decrease of p53 signaling was consistent with the elevation of AKT and subsequent activation. Obese rats exposed to fasting also exhibited improvement in insulin action despite a slight effect on p53 level. Results of the present study obviously showed that beta-adrenergic receptor agonist/cAMP prevented obesity-induced p53 activation. Although this effect in metabolic insulin target tissues tempted us to consider them as insulin sensitizers in obesity-related diabetes, p53 inhibition in non-insulin target tissues warned about the impairment of anti-cancer mechanisms in obese subjects. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. A Cannabinoid Receptor 2 Agonist Prevents Thrombin-Induced Blood-Brain Barrier Damage via the Inhibition of Microglial Activation and Matrix Metalloproteinase Expression in Rats.

    PubMed

    Li, Lin; Tao, Yihao; Tang, Jun; Chen, Qianwei; Yang, Yang; Feng, Zhou; Chen, Yujie; Yang, Liming; Yang, Yunfeng; Zhu, Gang; Feng, Hua; Chen, Zhi

    2015-12-01

    Thrombin mediates the life-threatening cerebral edema and blood-brain barrier (BBB) damage that occurs after intracerebral hemorrhage (ICH). We previously found that the selective cannabinoid receptor 2 (CB2R) agonist JWH-133 reduced brain edema and neurological deficits following germinal matrix hemorrhage (GMH). We explored whether CB2R stimulation ameliorated thrombin-induced brain edema and BBB permeability as well as the possible molecular mechanism involved. A total of 144 Sprague-Dawley (S-D) rats received a thrombin (20 U) injection in the right basal ganglia. JWH-133 (1.5 mg/kg) or SR-144528 (3.0 mg/kg) and vehicle were intraperitoneally (i.p.) injected 1 h after surgery. Brain water content measurement, Evans blue (EB) extravasation, Western blot, and immunofluorescence were used to study the effects of a CB2R agonist 24 h after surgery. The results demonstrated that JWH-133 administration significantly decreased thrombin-induced brain edema and reduced the number of Iba-1-positive microglia. JWH-133 also decreased the number of P44/P42(+)/Iba-1(+) microglia, lowered Evans blue extravasation, and inhibited the elevated matrix metallopeptidase (MMP)-9 and matrix metallopeptidase (MMP)-12 activities. However, a selective CB2R antagonist (SR-144528) reversed these effects. We demonstrated that CB2R stimulation reduced thrombin-induced brain edema and alleviated BBB damage. We also found that matrix metalloproteinase suppression may be partially involved in these processes.

  18. Inhibition of autophagy and enhancement of endoplasmic reticulum stress increase sensitivity of osteosarcoma Saos-2 cells to cannabinoid receptor agonist WIN55,212-2.

    PubMed

    Zhang, Guodong; Bi, Haiyong; Gao, Ji; Lu, Xing; Zheng, Yanping

    2016-07-01

    WIN55,212-2, a cannabinoid receptor agonist, can activate cannabinoid receptors, which has proven anti-tumour effects in several tumour types. Studies showed that WIN can inhibit tumour cell proliferation and induce apoptosis in diverse cancers. However, the role and mechanism of WIN in osteosarcoma are still unclear. In this study, we examined the effect of WIN55,212-2 on osteosarcoma cell line Saos-2 in terms of cell viability and apoptosis. Meanwhile, we further explored the role of endoplasmic reticulum stress and autophagy in apoptosis induced by WIN55,212-2. Our results showed that the cell proliferation of Saos-2 was inhibited by WIN55,212-2 in a dose-dependent and time-dependent manner. WIN55,212-2-induced Saos-2 apoptosis through mitochondrial apoptosis pathway. Meanwhile, WIN55,212-2 can induce endoplasmic reticulum stress and autophagy in Saos-2 cells. Inhibition of autophagy and enhancement of endoplasmic reticulum stress increased apoptosis induced by WIN55,212-2 in Saos-2 cells. These findings indicated that WIN55,212-2 in combination with autophagic inhibitor or endoplasmic reticulum stress activator may shed new light on osteosarcoma treatment. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  19. Peroxisome proliferator-activated receptor-γ agonist inhibits the mammalian target of rapamycin signaling pathway and has a protective effect in a rat model of status epilepticus

    PubMed Central

    SAN, YONG-ZHI; LIU, YU; ZHANG, YU; SHI, PING-PING; ZHU, YU-LAN

    2015-01-01

    Peroxisome proliferator-activated receptor γ (PPAR-γ) has a protective role in several neurological diseases. The present study investigated the effect of the PPAR-γ agonist, pioglitazone, on the mammalian target of rapamycin (mTOR) signaling pathway in a rat model of pentylenetetrazol (PTZ)-induced status epilepticus (SE). The investigation proceeded in two stages. First, the course of activation of the mTOR signaling pathway in PTZ-induced SE was examined to determine the time-point of peak activity, as reflected by phopshorylated (p)-mTOR/mTOR and p-S6/S6 ratios. Subsequently, pioglitazone was administrated intragastrically to investigate its effect on the mTOR signaling pathway, through western blot and immunochemical analyses. The levels of the interleukin (IL)-1β and IL-6 inflammatory cytokines were detected using ELISA, and neuronal loss was observed via Nissl staining. In the first stage of experimentation, the mTOR signaling pathway was activated, and the p-mTOR/mTOR and p-S6/S6 ratios peaked on the third day. Compared with the vehicle treated-SE group, pretreatment with pioglitazone was associated with the loss of fewer neurons, lower levels of IL-1β and IL-6, and inhibition of the activation of the mTOR signaling pathway. Therefore, the mTOR signaling pathway was activated in the PTZ-induced SE rat model, and the PPAR-γ agonist, pioglitazone, had a neuroprotective effect, by inhibiting activation of the mTOR pathway and preventing the increase in the levels of IL-1β and IL-6. PMID:25891824

  20. In vitro inhibition of methadone and oxycodone cytochrome P450-dependent metabolism: reversible inhibition by H2-receptor agonists and proton-pump inhibitors.

    PubMed

    Moody, David E; Liu, Fenyun; Fang, Wenfang B

    2013-10-01

    In vitro inhibition of oxycodone metabolism to noroxycodone and oxymorphone and R- and S-methadone metabolism to R- and S-2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) was measured for four H2-receptor antagonists and five proton-pump inhibitors (PPIs) using human liver microsomes (HLM) and cDNA-expressed human cytochrome P450s (rCYPs). Inhibitors were first incubated with HLM at three concentrations with and without preincubation of inhibitor, enzyme source and reducing equivalents to also screen for time-dependent inhibition (TDI). Cimetidine and famotidine (10-1,000 µM) inhibited all the four pathways >50%. Nizatidine and ranitidine did not. All the five PPIs (1-200 µM) inhibited one or more pathways >50%. Half maximal inhibitory concentrations (IC50s) were then determined using rCYPs. Cimetidine and famotidine both inhibited CYP3A4-mediated formation of noroxycodone and CYP2D6-mediated formation of oxymorphone, and famotidine inhibited CYP3A4-mediated formation of R- and S-EDDP, but IC50s were so high that only >10× therapeutic concentrations may have potential for reversible in vivo inhibition. The PPIs were more potent inhibitors; many have the potential for reversible in vivo inhibition at therapeutic concentrations. Omeprazole, esomeprazole and pantoprazole had greater effects on CYP3A4-mediated reactions, whereas lansoprazole was selective for CYP2D6-mediated formation of oxymorphone. Preincubation enhanced cimetidine inhibition of noroxycodone formation and rabeprazole inhibition of all pathways. Future studies will explore irreversible TDI.

  1. Further evidence for prejunctional GABA-B inhibition of cholinergic and peptidergic bronchoconstriction in guinea pigs: studies with new agonists and antagonists.

    PubMed

    Chapman, R W; Danko, G; del Prado, M; Egan, R W; Kreutner, W; Rizzo, C A; Hey, J A

    1993-06-01

    We examined the effect of the potent and selective GABA-B agonists, baclofen, 3-aminopropylphosphinic acid (3-APPi) and 3-aminopropyl (methyl) phosphinic acid (SKF 97541), and the GABA-B antagonists, 3-aminopropyl (diethoxymethyl) phosphinic acid (CGP 35348), 2-hydroxysaclofen and 3-aminopropylphosphonic acid (3-APPA) on cholinergic and peptidergic contractile responses in the airways of guinea pigs. Electrical field stimulation of the isolated guinea pig trachea produced cholinergic contractions that were inhibited by baclofen (EC50 = 5 mumol/l), 3-APPi (EC50 = 0.3 mumol/l) and SKF 97541 (EC50 = 0.4 mumol/l). The inhibition by baclofen (30 mumol/l) was reduced by CGP 35348 (IC50 = 65 mumol/l), 2-hydroxysaclofen (IC50 = 273 mumol/l) and 3-APPA (IC50 = 355 mumol/l). The in vivo cholinergic bronchoconstrictor response to vagal nerve stimulation (5 V, 20 Hz, 0.5 ms for 5 s) was attenuated by intravenous baclofen (ED50 = 1.7 mg/kg), 3-APPi (ED50 = 0.9 mg/kg) and SKF 97541 (ED50 = 0.2 mg/kg). The inhibition of vagally induced bronchoconstriction by baclofen was blocked by CGP 35348 (1-10 mg/kg, i.v.) and 2-hydroxysaclofen (10 mg/kg, i.v.). A cholinergic bronchoconstriction induced by CNS stimulation (400 microA, 2 ms, 32 Hz for 5 s) was inhibited by baclofen (ED50 = 5.1 mg/kg, i.v.) and 3-APPi (ED50 = 0.6 mg/kg, i.v.). The effect of baclofen was attenuated by 3-APPA (5 mg/kg, i.v.). A peptidergic bronchoconstriction was evoked by intravenous nicotine (1 mg/kg) in animals treated with ipratropium and phosphoramidon.(ABSTRACT TRUNCATED AT 250 WORDS)

  2. The Dectin 1 Agonist Curdlan Regulates Osteoclastogenesis by Inhibiting Nuclear Factor of Activated T cells Cytoplasmic 1 (NFATc1) through Syk Kinase

    PubMed Central

    Yamasaki, Toru; Ariyoshi, Wataru; Okinaga, Toshinori; Adachi, Yoshiyuki; Hosokawa, Ryuji; Mochizuki, Shinichi; Sakurai, Kazuo; Nishihara, Tatsuji

    2014-01-01

    Several immune system cell surface receptors are reported to be associated with osteoclastogenesis. Dectin 1, a lectin receptor for β-glucan, is found predominantly on cells of the myeloid lineage. In this study, we examined the effect of the dectin 1 agonist curdlan on osteoclastogenesis. In mouse bone marrow cells and dectin 1-overexpressing RAW 264.7 cells (d-RAWs), curdlan suppressed receptor activator of NF-κB ligand (RANKL)-induced osteoclast differentiation, bone resorption, and actin ring formation in a dose-dependent manner. This was achieved within non-growth inhibitory concentrations at the early stage. Conversely, curdlan had no effect on macrophage colony-stimulating factor-induced differentiation. Furthermore, curdlan inhibited RANKL-induced nuclear factor of activated T cell cytoplasmic 1 (NFATc1) expression, thereby decreasing osteoclastogenesis-related marker gene expression, including tartrate-resistant acid phosphatase, osteoclast stimulatory transmembrane protein, cathepsin K, and matrix metallopeptidase 9. Curdlan inhibited RANKL-induced c-fos expression, followed by suppression of NFATc1 autoamplification, without significantly affecting the NF-κB signaling pathway. We also observed that curdlan treatment decreased Syk protein in d-RAWs. Inhibition of the dectin 1-Syk kinase pathway by Syk-specific siRNA or chemical inhibitors suppressed osteoclast formation and NFATc1 expression stimulated by RANKL. In conclusion, our results demonstrate that curdlan potentially inhibits osteoclast differentiation, especially NFATc1 expression, and that Syk kinase plays a crucial role in the transcriptional pathways. This suggests that the activation of dectin 1-Syk kinase interaction critically regulates the genes required for osteoclastogenesis. PMID:24821724

  3. The dectin 1 agonist curdlan regulates osteoclastogenesis by inhibiting nuclear factor of activated T cells cytoplasmic 1 (NFATc1) through Syk kinase.

    PubMed

    Yamasaki, Toru; Ariyoshi, Wataru; Okinaga, Toshinori; Adachi, Yoshiyuki; Hosokawa, Ryuji; Mochizuki, Shinichi; Sakurai, Kazuo; Nishihara, Tatsuji

    2014-07-04

    Several immune system cell surface receptors are reported to be associated with osteoclastogenesis. Dectin 1, a lectin receptor for β-glucan, is found predominantly on cells of the myeloid lineage. In this study, we examined the effect of the dectin 1 agonist curdlan on osteoclastogenesis. In mouse bone marrow cells and dectin 1-overexpressing RAW 264.7 cells (d-RAWs), curdlan suppressed receptor activator of NF-κB ligand (RANKL)-induced osteoclast differentiation, bone resorption, and actin ring formation in a dose-dependent manner. This was achieved within non-growth inhibitory concentrations at the early stage. Conversely, curdlan had no effect on macrophage colony-stimulating factor-induced differentiation. Furthermore, curdlan inhibited RANKL-induced nuclear factor of activated T cell cytoplasmic 1 (NFATc1) expression, thereby decreasing osteoclastogenesis-related marker gene expression, including tartrate-resistant acid phosphatase, osteoclast stimulatory transmembrane protein, cathepsin K, and matrix metallopeptidase 9. Curdlan inhibited RANKL-induced c-fos expression, followed by suppression of NFATc1 autoamplification, without significantly affecting the NF-κB signaling pathway. We also observed that curdlan treatment decreased Syk protein in d-RAWs. Inhibition of the dectin 1-Syk kinase pathway by Syk-specific siRNA or chemical inhibitors suppressed osteoclast formation and NFATc1 expression stimulated by RANKL. In conclusion, our results demonstrate that curdlan potentially inhibits osteoclast differentiation, especially NFATc1 expression, and that Syk kinase plays a crucial role in the transcriptional pathways. This suggests that the activation of dectin 1-Syk kinase interaction critically regulates the genes required for osteoclastogenesis. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  4. The GLP-1 receptor agonist liraglutide inhibits progression of vascular disease via effects on atherogenesis, plaque stability and endothelial function in an ApoE(-/-) mouse model.

    PubMed

    Gaspari, Tracey; Welungoda, Iresha; Widdop, Robert E; Simpson, Richard W; Dear, Anthony E

    2013-07-01

    Liraglutide, a once-daily glucagon-like peptide-1 receptor (GLP-1R) agonist, has been approved as a new treatment for type 2 diabetes and is the subject of a clinical trial programme to evaluate the effects on cardiovascular disease and safety. The current study aimed to determine the in vivo effect of liraglutide on progression of atherosclerotic vascular disease in the apolipoprotein E-deficient (ApoE(-/-)) mouse model and identify underlying mechanisms responsible. Liraglutide treatment inhibited progression of early onset, low-burden atherosclerotic disease in a partially GLP-1R-dependent manner in the ApoE(-/-) mouse model. In addition, liraglutide treatment inhibited progression of atherosclerotic plaque formation and enhanced plaque stability, again in a partially GLP-1R-dependent manner. No significant effect of liraglutide on progression of late onset, high-burden atherosclerotic disease was observed. In addition, no significant endothelial cell dysfunction was identified in ApoE(-/-) mice with early onset, low-burden atherosclerotic disease, although significant prevention of weight gain was observed in liraglutide-treated mice using this dietary protocol. Taken together, these results suggest a potential role for liraglutide in the prevention and stabilisation of atherosclerotic vascular disease together with possible protection against major cardiovascular events.

  5. A novel PPAR{gamma} agonist, KR62776, suppresses RANKL-induced osteoclast differentiation and activity by inhibiting MAP kinase pathways

    SciTech Connect

    Park, Ju-Young; Bae, Myung-Ae; Cheon, Hyae Gyeong; Kim, Sung Soo; Hong, Jung-Min; Kim, Tae-Ho; Choi, Je-Yong; Kim, Sang-Hyun; Lim, Jiwon; Choi, Chang-Hyuk; Shin, Hong-In; Kim, Shin-Yoon Park, Eui Kyun

    2009-01-16

    We investigated the effects of a novel peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}) agonist, KR62776, on osteoclast differentiation and function, and on the underlying signaling pathways. KR62776 markedly suppressed differentiation into osteoclasts in various osteoclast model systems, including bone marrow mononuclear (BMM) cells and a co-culture of calvarial osteoblasts and BMM cells. KR62776 suppressed the activation of tartrate-resistant acid phosphatase (TRAP) and the expression of genes associated with osteoclast differentiation, such as TRAP, dendritic cell-specific transmembrane protein (DC-STAMP), and osteoclast-associated receptor (OSCAR). Furthermore, KR62776 reduced resorption pit formation in osteoclasts, and down-regulated genes essential for osteoclast activity, such as Src and {alpha}v{beta}3 integrin. An analysis of a signaling pathway showed that KR62776 inhibited the receptor activator of nuclear factor-{kappa}B ligand (RANKL)-induced activation of p38 mitogen-activated protein kinase (p38MAPK), extracellular regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and nuclear factor-{kappa}B (NF-{kappa}B). Together, these results demonstrate that KR62776 negatively affects osteoclast differentiation and activity by inhibiting the RANKL-induced activation of MAP kinases and NF-{kappa}B.

  6. The effects of the preferential 5-HT2A agonist psilocybin on prepulse inhibition of startle in healthy human volunteers depend on interstimulus interval.

    PubMed

    Vollenweider, Franz X; Csomor, Philipp A; Knappe, Bernhard; Geyer, Mark A; Quednow, Boris B

    2007-09-01

    Schizophrenia patients exhibit impairments in prepulse inhibition (PPI) of the startle response. Hallucinogenic 5-HT(2A) receptor agonists are used for animal models of schizophrenia because they mimic some symptoms of schizophrenia in humans and induce PPI deficits in animals. Nevertheless, one report indicates that the 5-HT(2A) receptor agonist psilocybin increases PPI in healthy humans. Hence, we investigated these inconsistent results by assessing the dose-dependent effects of psilocybin on PPI in healthy humans. Sixteen subjects each received placebo or 115, 215, and 315 microg/kg of psilocybin at 4-week intervals in a randomized and counterbalanced order. PPI at 30-, 60-, 120-, 240-, and 2000-ms interstimulus intervals (ISIs) was measured 90 and 165 min after drug intake, coinciding with the peak and post-peak effects of psilocybin. The effects of psilocybin on psychopathological core dimensions and sustained attention were assessed by the Altered States of Consciousness Rating Scale (5D-ASC) and the Frankfurt Attention Inventory (FAIR). Psilocybin dose-dependently reduced PPI at short (30 ms), had no effect at medium (60 ms), and increased PPI at long (120-2000 ms) ISIs, without affecting startle reactivity or habituation. Psilocybin dose-dependently impaired sustained attention and increased all 5D-ASC scores with exception of Auditory Alterations. Moreover, psilocybin-induced impairments in sustained attention performance were positively correlated with reduced PPI at the 30 ms ISI and not with the concomitant increases in PPI observed at long ISIs. These results confirm the psilocybin-induced increase in PPI at long ISIs and reveal that psilocybin also produces a decrease in PPI at short ISIs that is correlated with impaired attention and consistent with deficient PPI in schizophrenia.

  7. Administration of caffeine inhibited adenosine receptor agonist-induced decreases in motor performance, thermoregulation, and brain neurotransmitter release in exercising rats.

    PubMed

    Zheng, Xinyan; Hasegawa, Hiroshi

    2016-01-01

    We examined the effects of an adenosine receptor agonist on caffeine-induced changes in thermoregulation, neurotransmitter release in the preoptic area and anterior hypothalamus, and endurance exercise performance in rats. One hour before the start of exercise, rats were intraperitoneally injected with either saline alone (SAL), 10 mg kg(-1) caffeine and saline (CAF), a non-selective adenosine receptor agonist (5'-N-ethylcarboxamidoadenosine [NECA]: 0.5 mg kg(-1)) and saline (NECA), or the combination of caffeine and NECA (CAF+NECA). Rats ran until fatigue on the treadmill with a 5% grade at a speed of 18 m min(-1) at 23 °C. Compared to the SAL group, the run time to fatigue (RTTF) was significantly increased by 52% following caffeine administration and significantly decreased by 65% following NECA injection (SAL: 91 ± 14.1 min; CAF: 137 ± 25.8 min; NECA: 31 ± 13.7 min; CAF+NECA: 85 ± 11.8 min; p<0.05). NECA decreased the core body temperature (Tcore), oxygen consumption, which is an index of heat production, tail skin temperature, which is an index of heat loss, and extracellular dopamine (DA) release at rest and during exercise. Furthermore, caffeine injection inhibited the NECA-induced decreases in the RTTF, Tcore, heat production, heat loss, and extracellular DA release. Neither caffeine nor NECA affected extracellular noradrenaline or serotonin release. These results support the findings of previous studies showing improved endurance performance and overrides in body limitations after caffeine administration, and imply that the ergogenic effects of caffeine may be associated with the adenosine receptor blockade-induced increases in brain DA release.

  8. Complete reversal of muscle wasting in experimental cancer cachexia: Additive effects of activin type II receptor inhibition and β-2 agonist.

    PubMed

    Toledo, Míriam; Busquets, Sílvia; Penna, Fabio; Zhou, Xiaolan; Marmonti, Enrica; Betancourt, Angelica; Massa, David; López-Soriano, Francisco J; Han, H Q; Argilés, Josep M

    2016-04-15

    Formoterol is a highly potent β2-adrenoceptor-selective agonist, which is a muscle growth promoter in many animal species. Myostatin/activin inhibition reverses skeletal muscle loss and prolongs survival of tumor-bearing animals. The aim of this investigation was to evaluate the effects of a combination of the soluble myostatin receptor ActRIIB (sActRIIB) and the β2-agonist formoterol in the cachectic Lewis lung carcinoma model. The combination of formoterol and sActRIIB was extremely effective in reversing muscle wasting associated with experimental cancer cachexia in mice. Muscle weights from tumor-bearing animals were completely recovered following treatment and this was also reflected in the measured grip strength. This combination increased food intake in both control and tumor-bearing animals. The double treatment also prolonged survival significantly without affecting the weight and growth of the primary tumor. In addition, it significantly reduced the number of metastasis. Concerning the mechanisms for the preservation of muscle mass during cachexia, the effects of formoterol and sActRIIB seemed to be additive, since formoterol reduced the rate of protein degradation (as measured in vitro as tyrosine release, using incubated isolated individual muscles) while sActRIIB only affected protein synthesis (as measured in vivo using tritiated phenylalanine). Formoterol also increased the rate of protein synthesis and this seemed to be favored by the presence of sActRIIB. Combining formoterol and sActRIIB seemed to be a very promising treatment for experimental cancer cachexia. Further studies in human patients are necessary and may lead to a highly effective treatment option for muscle wasting associated with cancer.

  9. Inhibition of NAD(P)H oxidase potentiates AT2 receptor agonist-induced natriuresis in Sprague-Dawley rats.

    PubMed

    Sabuhi, Rifat; Asghar, Mohammad; Hussain, Tahir

    2010-10-01

    A positive association between renin-angiotensin system, especially AT1 receptor, and oxidative stress in the pathogenesis of hypertension and cardiovascular/renal diseases has been suggested. However, the role of oxidative stress, especially superoxide radicals in renal sodium handling in response to AT1 and AT2 receptors, is not known. Therefore, the present study was designed to investigate the role of NAD(P)H oxidase (NOX), a major superoxide radical producing enzyme, in AT1 and AT2 receptor function on natriuresis/diuresis in Sprague-Dawley rats. The rats under anesthesia were intravenously infused with NOX inhibitor apocynin (3.5 μg·kg(-1)·min(-1)), the AT1 receptor antagonist candesartan (100 μg/kg; bolus), and the AT2 receptor agonist CGP-42112A (1 μg·kg(-1)·min(-1)) alone and in combinations. Candesartan alone significantly increased urinary flow (UF; μl/30 min) by 53 and urinary Na excretion (U(Na)V; μmol/min) by 0.4 over basal. Preinfusion of apocynin had no effect on the net increase in UF or U(Na)V in response to candesartan. On the other hand, apocynin preinfusion caused profound increases in CGP-42112A-induced UF by 72, U(Na)V by 1.14, and fractional excretion of Na by 7.8. Apocynin and CGP-42112A alone did not cause significant increase in UF or U(Na)V over the basal. CGP-42112A infusion in the presence of apocynin increased urinary nitrite/nitrates and cGMP over basal. The infusion of candesartan, apocynin, and CGP-42112A alone or in combinations had no effect on the blood pressure or the glomerular filtration rate, suggesting tubular effects on natriuresis/diuresis. The data suggest that NOX may have an antagonistic role in AT2 receptor-mediated natriuresis/diuresis possibly via neutralizing nitric oxide and thereby influence fluid-Na homeostasis.

  10. The GABA(B) receptor agonist, baclofen, and the positive allosteric modulator, CGP7930, inhibit visceral pain-related responses to colorectal distension in rats.

    PubMed

    Brusberg, Mikael; Ravnefjord, Anna; Martinsson, Rakel; Larsson, Håkan; Martinez, Vicente; Lindström, Erik

    2009-02-01

    Activation of GABA(B) receptors by the selective agonist baclofen produces anti-nociceptive effects in animal models of somatic pain. The aim of the present study was to evaluate the effect of baclofen and the GABA(B) receptor positive allosteric modulator CGP7930 on pseudo-affective responses to colorectal distension in rats. Female Sprague-Dawley rats were subjected to repeated, noxious colorectal distension (CRD) (12 distensions at 80 mmHg, for 30 s with 5 min intervals). The visceromotor response (VMR) and cardiovascular responses (mean arterial blood pressure (ABP) and heart rate (HR)) to CRD were monitored in conscious, telemetrized animals. Baclofen (0.3-3 micromol/kg, i.v.) reduced the VMR to CRD dose-dependently, reaching a 61% maximal inhibition (p < 0.001). The highest doses of baclofen attenuated CRD-evoked increases in ABP by 17% (p > 0.05) and reduced the change in HR by 48% (p < 0.01). CGP7930 (3-30 micromol/kg, i.v.) reduced the VMR to CRD in a dose-dependent fashion with a maximal inhibition of 31% (p < 0.05). The highest dose of CGP7930 also attenuated the increase in ABP by 18% (p > 0.05) and inhibited the increase in HR by 24% (p < 0.05) associated with CRD. Neither baclofen nor CGP7930 affected colorectal compliance. The results suggest that activation of GABA(B) receptors produces anti-nociceptive effects in a rat model of mechanically induced visceral pain. While CGP7930 was less efficacious than baclofen overall, positive allosteric modulation of GABA(B) receptors may represent a valid approach in the treatment of visceral pain conditions, with the possibility of an improved safety profile compared to full agonism.

  11. The Mechanism of Chemokine Receptor 9 Internalization Triggered by Interleukin 2 and Interleukin 4

    PubMed Central

    Tong, Xiaoling; Zhang, Lijun; Zhang, Li; Hu, Meng; Leng, Jun; Yu, Beibei; Zhou, Beibei; Hu, Yi; Zhang, Qiuping

    2009-01-01

    In previous study, we found that the chemokine receptor 9 (CCR9) was highly expressed on CD4+ T cells from patients with T-cell lineage acute lymphocytic leukemia (T-ALL) and mediated leukemia cell infiltration and metastasis. Combined use of interleukin 2 (IL-2) and IL-4 promoted the internalization of CCR9 and therefore attenuated leukemia cell infiltration and metastasis. In this study, we preliminarily investigated the mechanism of internalization of CCR9 on MOLT4 cell model (a human leukemia T-cell line, naturally expresses CCR9) and found that IL-2 upregulated the cell surface expression of IL-4Rα (CD124) greatly, whereas IL-4 had no significant influence on α (CD25) and β subunits (CD122) of IL-2R. Moreover, specific inhibitors, such as staurosporine, H89 and heparin, inhibited internalization of CCR9, which indicated a role of protein kinase C (PKC) and G protein-coupled kinase 2 (GRK2), respectively. Furthermore, GRK2 was upregulated and translocated to cell membrane in IL-2 and IL-4 treated cells which indicated that PKC could be a prerequisite for GRK2 activity. PMID:19567201

  12. Suppression of inflammation response by a novel A₃ adenosine receptor agonist thio-Cl-IB-MECA through inhibition of Akt and NF-κB signaling.

    PubMed

    Lee, Hak-Sun; Chung, Hwa-Jin; Lee, Hyuk Woo; Jeong, Lak Shin; Lee, Sang Kook

    2011-09-01

    Adenosine, a purine nucleoside, is released from metabolically active cells into extracellular space and plays an important role in various pathophysiological processes. Adenosine regulates many biological responses including inflammation by the interaction with their receptors such as A₁, A(2A), A(2B), and A₃. Especially, A₃ adenosine receptor (A₃AR) is considered to be expressed in macrophage cells. To the end, A₃AR agonists have been reported to have an anti-inflammatory activity. In our continuous efforts to develop new anti-inflammatory agents, we found a novel adenosine analog, 2-chloro-N⁶-(3-iodobenzyl)-4'-thioadenosine-5'-N-methyluronamide (thio-Cl-IB-MECA), was a potent human A₃AR agonist. The study was designed to investigate whether thio-Cl-IB-MECA has an anti-inflammatory potential in mouse macrophage RAW 264.7 cells and mouse sepsis model in vivo. Thio-Cl-IB-MECA exhibited an effective anti-inflammatory activity. The expression of pro-inflammatory biomarkers including inducible nitric oxide synthase (iNOS), interleukin-1β (IL-1β), and tumor necrosis factor (TNF-α) was suppressed by the treatment of thio-Cl-IB-MECA in the protein and mRNA levels in lipopolysaccharide (LPS)-stimulated mouse macrophage RAW 264.7 cells. Further examination revealed that thio-Cl-IB-MECA inhibited LPS-induced phosphatidylinositol 3-kinase (PI3 kinase)/Akt activation, NF-kB binding activity, and β-catenin expression. In addition, in in vivo LPS-induced mouse endotoxemia model, thio-Cl-IB-MECA exerted the increase of survival rate compared to vehicle-treated mouse. The analysis of the protein levels of iNOS, IL-1β, and TNF-α was also suppressed by the compound-treated groups in lung tissues. These results suggest that thio-Cl-IB-MECA might have an anti-inflammatory activity through the inhibition of pro-inflammatory cytokine expression by modulating PI3K/Akt and NF-κB signaling pathways.

  13. H3 histamine receptor agonist inhibits biliary growth of BDL rats by downregulation of the cAMP-dependent PKA/ERK1/2/ELK-1 pathway.

    PubMed

    Francis, Heather; Franchitto, Antonio; Ueno, Yoshiyuki; Glaser, Shannon; DeMorrow, Sharon; Venter, Julie; Gaudio, Eugenio; Alvaro, Domenico; Fava, Giammarco; Marzioni, Marco; Vaculin, Bradley; Alpini, Gianfranco

    2007-05-01

    Histamine regulates many functions by binding to four histamine G-coupled receptor proteins (H1R, H2R, H3R and H4R). As H3R exerts their effects by coupling to Galpha(i/o) proteins reducing adenosine 3', 5'-monophosphate (cAMP) levels (a key player in the modulation of cholangiocyte hyperplasia/damage), we evaluated the role of H3R in the regulation of biliary growth. We posed the following questions: (1) Do cholangiocytes express H3R? (2) Does in vivo administration of (R)-(alpha)-(-)-methylhistamine dihydrobromide (RAMH) (H3R agonist), thioperamide maleate (H3R antagonist) or histamine, in the absence/presence of thioperamide maleate, to bile duct ligated (BDL) rats regulate cholangiocyte proliferation? and (3) Does RAMH inhibit cholangiocyte proliferation by downregulation of cAMP-dependent phosphorylation of protein kinase A (PKA)/extracellular signal-regulated kinase 1/2 (ERK1/2)/ets-like gene-1 (Elk-1)? The expression of H3R was evaluated in liver sections by immunohistochemistry and immunofluorescence, and by real-time PCR in cholangiocyte RNA from normal and BDL rats. BDL rats (immediately after BDL) were treated daily with RAMH, thioperamide maleate or histamine in the absence/presence of thioperamide maleate for 1 week. Following in vivo treatment of BDL rats with RAMH for 1 week, and in vitro stimulation of BDL cholangiocytes with RAMH, we evaluated cholangiocyte proliferation, cAMP levels and PKA, ERK1/2 and Elk-1 phosphorylation. Cholangiocytes from normal and BDL rats express H3R. The expression of H3R mRNA increased in BDL compared to normal cholangiocytes. Histamine decreased cholangiocyte growth of BDL rats to a lower extent than that observed in BDL RAMH-treated rats; histamine-induced inhibition of cholangiocyte growth was partly blocked by thioperamide maleate. In BDL rats treated with thioperamide maleate, cholangiocyte hyperplasia was slightly higher than that of BDL rats. In vitro, RAMH inhibited the proliferation of BDL cholangiocytes. RAMH

  14. The gamma-aminobutyric acid type B (GABAB) receptor agonist baclofen inhibits morphine sensitization by decreasing the dopamine level in rat nucleus accumbens

    PubMed Central

    2012-01-01

    Background Repeated morphine exposure can induce behavioral sensitization. There are evidences have shown that central gamma-aminobutyric acid (GABA) system is involved in morphine dependence. However, the effect of a GABAB receptor agonist baclofen on morphine-induced behavioral sensitization in rats is unclear. Methods We used morphine-induced behavioral sensitization model in rat to investigate the effects of baclofen on behavioral sensitization. Moreover, dopamine release in the shell of the nucleus accumbens was evaluated using microdialysis assay in vivo. Results The present study demonstrated that morphine challenge (3 mg/kg, s.c.) obviously enhanced the locomotor activity following 4-day consecutive morphine administration and 3-day withdrawal period, which indicated the expression of morphine sensitization. In addition, chronic treatment with baclofen (2.5, 5 mg/kg) significantly inhibited the development of morphine sensitization. It was also found that morphine challenge 3 days after repeated morphine administration produced a significant increase of extracellular dopamine release in nucleus accumbens. Furthermore, chronic treatment with baclofen decreased the dopamine release induced by morphine challenge. Conclusions Our results indicated that gamma-aminobutyric acid system plays an important role in the morphine sensitization in rat and suggested that behavioral sensitization is a promising model to study the mechanism underlying drug abuse. PMID:22559224

  15. Dopamine receptor 2 activation inhibits ovarian vascular endothelial growth factor secretion in vitro: implications for treatment of ovarian hyperstimulation syndrome with dopamine receptor 2 agonists.

    PubMed

    Ferrero, Hortensia; García-Pascual, Carmen M; Gómez, Raúl; Delgado-Rosas, Francisco; Cauli, Omar; Simón, Carlos; Gaytán, Francisco; Pellicer, Antonio

    2014-05-01

    To ascertain whether vascular endothelial growth factor (VEGF) secretion by luteinized granulosa cells (GCs) is modulated by the dopaminergic system in a dose-dependent fashion and how this is related to the differential efficacy of dopamine receptor 2 (D2)-agonists (D2-ag) in preventing ovarian hyperstimulation syndrome (OHSS). The relationship between the dopaminergic system and VEGF secretion in luteinized GCs was evaluated. Archived human ovaries were immunostained to characterize D2 expression. University affiliated infertility center. Premenopausal women and egg donors. Luteinized GCs were cultured with the D2-ag cabergoline. Human ovarian sections were immunostained for D2. The VEGF was measured by ELISA and D2 expression was evaluated by In-Cell ELISA. The D2 expression throughout the luteal phase was characterized by immunohistochemistry. The VEGF secretion was decreased by the D2-ag in a dose-dependent fashion. The efficiency of this process was correlated with the amount of D2 expressed by luteinized GCs. A decrease in D2 expression in ovarian sections was observed during the late luteal phase. The efficacy of D2-ags in preventing OHSS might rely on their capacity to inhibit VEGF secretion by luteinized GCs. Because this capacity is dose-dependent, increasing the intraovarian concentration of D2-ags should be explored as a means of increasing the efficacy of these drugs in preventing OHSS. Copyright © 2014 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  16. NEU-P11, a novel melatonin agonist, inhibits weight gain and improves insulin sensitivity in high-fat/high-sucrose-fed rats.

    PubMed

    She, Meihua; Deng, Xiaojian; Guo, Zhenyu; Laudon, Moshe; Hu, Zhuowei; Liao, Duanfang; Hu, Xiaobo; Luo, Yi; Shen, Qingyun; Su, Zehong; Yin, Weidong

    2009-04-01

    Evidences indicate that a complex relationship exists among sleep disorders, obesity and insulin resistance. NEU-P11 is a novel melatonin agonist used in treatment of psychophysiological insomnia, and in animal studies NEU-P11 showed sleep-promoting effect. In this study, we applied NEU-P11 on obese rats to assess its potential melatoninergic effects in vivo. Obese models were established using high-fat/high-sucrose-fed for 5 months. NEU-P11 (10mg/kg)/melatonin (4mg/kg)/vehicle were administered by a daily intraperitoneal injection respectively for 8 weeks. Our results showed that NEU-P11 or melatonin inhibited both body weight gain and deposit of abdominal fat with no influence on food intake. The impaired insulin sensitivity and antioxidative potency were improved and the levels of plasma glucose, total cholesterol (TC), triglycerides (TG) decreased with an increased in HDL-cholesterol (HDL-c) after NEU-P11 or melatonin administration. These data suggest that NEU-P11, like melatonin, decreased body weight gain and improved insulin sensitivity and metabolic profiles in obese rats. We conclude that NEU-P11 has a melatoninergic effect on regulating body weight in obese rats and also improving metabolic profiles and efficiently enhancing insulin sensitivity.

  17. Influence of central inhibition of sympathetic nervous activity on myocardial metabolism in chronic heart failure: acute effects of the imidazoline I1-receptor agonist moxonidine.

    PubMed

    Mobini, Reza; Fu, Michael; Jansson, Per-Anders; Bergh, Claes-Håkan; Scharin Täng, Margareta; Waagstein, Finn; Andersson, Bert

    2006-03-01

    Although beta-adrenergic blockade is beneficial in heart failure, inhibition of central sympathetic outflow using moxonidine has been associated with increased mortality. In the present study, we studied the acute effects of the imidazoline-receptor agonist moxonidine on haemodynamics, NA (noradrenaline) kinetics and myocardial metabolism. Fifteen patients with CHF (chronic heart failure) were randomized to a single dose of 0.6 mg of sustained-release moxonidine or matching placebo. Haemodynamics, NA kinetics and myocardial metabolism were studied over a 2.5 h time period. There was a significant reduction in pulmonary and systemic arterial pressures, together with a decrease in cardiac index in the moxonidine group. Furthermore, there was a simultaneous reduction in systemic and cardiac net spillover of NA in the moxonidine group. Analysis of myocardial consumption of substrates in the moxonidine group showed a significant increase in non-esterified fatty acid consumption and a possible trend towards an increase in myocardial oxygen consumption compared with the placebo group (P=0.16). We conclude that a single dose of moxonidine (0.6 mg) in patients already treated with a beta-blocker reduced cardiac and overall sympathetic activity. The finding of increased lipid consumption without decreased myocardial oxygen consumption indicates a lack of positive effects on myocardial metabolism under these conditions. We suggest this might be a reason for the failure of moxonidine to prevent deaths in long-term studies in CHF.

  18. Novel mixed NOP/MOP agonist BU08070 alleviates pain and inhibits gastrointestinal motility in mouse models mimicking diarrhea-predominant irritable bowel syndrome symptoms.

    PubMed

    Sobczak, Marta; Cami-Kobeci, Gerta; Sałaga, Maciej; Husbands, Stephen M; Fichna, Jakub

    2014-08-05

    The opioid and nociceptin systems play a crucial role in the maintenance of homeostasis in the gastrointestinal (GI) tract. The aim of this study was to characterize the effect of BU08070, a novel mixed MOP/NOP agonist, on mouse intestinal contractility in vitro and GI motility in vivo in physiological conditions and in animal models mimicking symptoms of irritable bowel syndrome (IBS), including diarrhea and abdominal pain. The effect of BU08070 on muscle contractility in vitro was characterized in the ileum and colon. To assess the effect of BU08070 in vivo, the following parameters were assessed: whole GI transit, gastric emptying, geometric center, colonic bead expulsion, fecal pellet output and time to castor oil-induced diarrhea. The antinociceptive activity of BU08070 was characterized in the mustard oil (MO)-induced abdominal pain model and the writhing test, alone and in the presence of MOP and NOP antagonists. in vitro, BU08070 (10(-10)-10(-6) M) inhibited colonic and ileal smooth muscle contractions in a concentration-dependent manner. in vivo, BU08070 prolonged the whole GI transit and inhibited colonic bead expulsion. The antitransit and antidiarrheal effects of BU08070 were observed already at the dose of 0.1 mg/kg (i.p.). BU08070 reversed hypermotility and reduced pain in mouse models mimicking IBS-D symptoms. Our results suggest that BU08070 has a potential of becoming an efficient drug in IBS-D therapy. Here we also validate mixed NOP/MOP receptor targeting as possible future treatment of functional GI diseases. Copyright © 2014 Elsevier B.V. All rights reserved.

  19. Novel mixed NOP/MOP agonist BU08070 alleviates pain and inhibits gastrointestinal motility in mouse models mimicking diarrhea-predominant irritable bowel syndrome symptoms

    PubMed Central

    Sobczak, Marta; Cami-Kobeci, Gerta; Sałaga, Maciej; Husbands, Stephen M.; Fichna, Jakub

    2015-01-01

    Background The opioid and nociceptin systems play a crucial role in the maintenance of homeostasis in the gastrointestinal (GI) tract. The aim of this study was to characterize the effect of BU08070, a novel mixed MOP/NOP agonist, on mouse intestinal contractility in vitro and GI motility in vivo in physiological conditions and in animal models mimicking symptoms of irritable bowel syndrome (IBS), including diarrhea and abdominal pain. Methods The effect of BU08070 on muscle contractility in vitro was characterized in the ileum and colon. To assess the effect of BU08070 in vivo, the following parameters were assessed: whole GI transit, gastric emptying, geometric center, colonic bead expulsion, fecal pellet output and time to castor oil-induced diarrhea. The antinociceptive activity of BU08070 was characterized in the mustard oil (MO)-induced abdominal pain model and the writhing test, alone and in the presence of MOP and NOP antagonists. Results In vitro, BU08070 (10−10–10−6 M) inhibited colonic and ileal smooth muscle contractions in a concentration-dependent manner. In vivo, BU08070 prolonged the whole GI transit and inhibited colonic bead expulsion. The antitransit and antidiarrheal effect of BU08070 was observed already at the dose of 0.1 mg/kg (i.p.). BU08070 reversed hypermotility and reduced pain in mouse models mimicking IBS-D symptoms. Conclusion Our results suggest that BU08070 has a potential of becoming an efficient drug in IBS-D therapy. Here we also validate mixed NOP/MOP receptor targeting as possible future treatment of functional GI diseases. PMID:24815321

  20. Anti-inflammatory effect of cannabinoid agonist WIN55, 212 on mouse experimental colitis is related to inhibition of p38MAPK.

    PubMed

    Feng, Ya-Jing; Li, Yong-Yu; Lin, Xu-Hong; Li, Kun; Cao, Ming-Hua

    2016-11-21

    To investigate the anti-inflammatory effect and the possible mechanisms of an agonist of cannabinoid (CB) receptors, WIN55-212-2 (WIN55), in mice with experimental colitis, so as to supply experimental evidence for its clinical use in future. We established the colitis model in C57BL/6 mice by replacing the animals' water supply with 4% dextran sulfate sodium (DSS) for 7 consecutive days. A colitis scoring system was used to evaluate the severity of colon local lesion. The plasma levels of proinflammatory cytokines, such as tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6), and the myeloperoxidase (MPO) activity in colon tissue were measured. The expressions of cannabinoid receptors, claudin-1 protein, p38 mitogen-activated protein kinase (p38MAPK) and its phosphorylated form (p-p38) in colon tissue were determined by immunohistochemistry and Western blot. In addition, the effect of SB203580 (SB), an inhibitor of p38, was investigated in parallel experiments, and the data were compared with those from intervention groups of WIN55 and SB alone or used together. The results demonstrated that WIN55 or SB treatment alone or together improved the pathological changes in mice with DSS colitis, decreased the plasma levels of TNF-α, and IL-6, and MPO activity in colon. The enhanced expression of claudin-1 and the inhibited expression of p-p38 in colon tissues were found in the WIN55-treated group. Besides, the expression of CB1 and CB2 receptors was enhanced in the colon after the induction of DSS colitis, but reduced when p38MAPK was inhibited. These results confirmed the anti-inflammatory effect and protective role of WIN55 on the mice with experimental colitis, and revealed that this agent exercises its action at least partially by inhibiting p38MAPK. Furthermore, the results showed that SB203580, affected the expression of CB1 and CB2 receptors in the mouse colon, suggesting a close linkage and cross-talk between the p38MAPK signaling pathway and the

  1. Anti-inflammatory effect of cannabinoid agonist WIN55, 212 on mouse experimental colitis is related to inhibition of p38MAPK

    PubMed Central

    Feng, Ya-Jing; Li, Yong-Yu; Lin, Xu-Hong; Li, Kun; Cao, Ming-Hua

    2016-01-01

    AIM To investigate the anti-inflammatory effect and the possible mechanisms of an agonist of cannabinoid (CB) receptors, WIN55-212-2 (WIN55), in mice with experimental colitis, so as to supply experimental evidence for its clinical use in future. METHODS We established the colitis model in C57BL/6 mice by replacing the animals’ water supply with 4% dextran sulfate sodium (DSS) for 7 consecutive days. A colitis scoring system was used to evaluate the severity of colon local lesion. The plasma levels of proinflammatory cytokines, such as tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6), and the myeloperoxidase (MPO) activity in colon tissue were measured. The expressions of cannabinoid receptors, claudin-1 protein, p38 mitogen-activated protein kinase (p38MAPK) and its phosphorylated form (p-p38) in colon tissue were determined by immunohistochemistry and Western blot. In addition, the effect of SB203580 (SB), an inhibitor of p38, was investigated in parallel experiments, and the data were compared with those from intervention groups of WIN55 and SB alone or used together. RESULTS The results demonstrated that WIN55 or SB treatment alone or together improved the pathological changes in mice with DSS colitis, decreased the plasma levels of TNF-α, and IL-6, and MPO activity in colon. The enhanced expression of claudin-1 and the inhibited expression of p-p38 in colon tissues were found in the WIN55-treated group. Besides, the expression of CB1 and CB2 receptors was enhanced in the colon after the induction of DSS colitis, but reduced when p38MAPK was inhibited. CONCLUSION These results confirmed the anti-inflammatory effect and protective role of WIN55 on the mice with experimental colitis, and revealed that this agent exercises its action at least partially by inhibiting p38MAPK. Furthermore, the results showed that SB203580, affected the expression of CB1 and CB2 receptors in the mouse colon, suggesting a close linkage and cross-talk between the p38

  2. Involvement of pallidotegmental neurons in methamphetamine- and MK-801-induced impairment of prepulse inhibition of the acoustic startle reflex in mice: reversal by GABAB receptor agonist baclofen.

    PubMed

    Arai, Sawako; Takuma, Kazuhiro; Mizoguchi, Hiroyuki; Ibi, Daisuke; Nagai, Taku; Takahashi, Kenji; Kamei, Hiroyuki; Nabeshima, Toshitaka; Yamada, Kiyofumi

    2008-12-01

    We have previously demonstrated that pallidotegmental GABAergic neurons play a crucial role in prepulse inhibition (PPI) of the startle reflex in mice through the activation of GABA(B) receptors in pedunculopontine tegmental neurons. In this study, we investigated whether PPI disruption induced by methamphetamine (METH) or MK-801 is associated with the dysfunction of pallidotegmental neurons. Furthermore, we examined the effects of baclofen, a GABA(B) receptor agonist, on METH- and MK-801-induced PPI impairment. Acute treatment with METH (3 mg/kg, subcutaneouly (s.c.)) and MK-801 (>0.3 mg/kg, s.c.) significantly disrupted PPI, accompanied by the suppression of c-Fos expression in lateral globus pallidus induced by PPI. Furthermore, acute treatment with METH and MK-801 stimulated c-Fos expression in the caudal pontine reticular nucleus (PnC) in mice subjected to the PPT test, although PPI alone had no effect on c-Fos expression. Repeated treatment with 1 mg/kg METH for 7 days, which did not affect PPI acutely, showed similar effects on PPI and c-Fos expression to acute treatment with METH (3 mg/kg). Baclofen dose-dependently ameliorated PPI impairment induced by acute treatment with METH (3 mg/kg) and MK-801 (1 mg/kg), and decreased METH- and MK-801-stimulated c-Fos expression in PnC to the basal level. These results suggest that dysfunction of pallidotegmental neurons is involved in PPI disruption caused by METH and MK-801 in mice. GABA(B) receptor may constitute a putative target in treating neuropsychiatric disorders with sensorimotor gating deficits, such as schizophrenia and METH psychosis.

  3. Arjunolic acid, a peroxisome proliferator-activated receptor α agonist, regresses cardiac fibrosis by inhibiting non-canonical TGF-β signaling.

    PubMed

    Bansal, Trisha; Chatterjee, Emeli; Singh, Jasdeep; Ray, Arjun; Kundu, Bishwajit; Thankamani, V; Sengupta, Shantanu; Sarkar, Sagartirtha

    2017-10-06

    Cardiac hypertrophy and associated heart fibrosis remain a major cause of death worldwide. Phytochemicals have gained attention as alternative therapeutics for managing cardiovascular diseases. These include the extract from the plant Terminalia arjuna, which is a popular cardioprotectant and may prevent or slow progression of pathological hypertrophy to heart failure. Here, we investigated the mode of action of a principal bioactive T. arjuna compound, arjunolic acid (AA), in ameliorating hemodynamic load-induced cardiac fibrosis and identified its intracellular target. Our data revealed that AA significantly represses collagen expression and improves cardiac function during hypertrophy. We found that AA binds to and stabilizes the ligand-binding domain of peroxisome proliferator-activated receptor α (PPARα) and increases its expression during cardiac hypertrophy. PPARα knockdown during AA treatment in hypertrophy samples, including angiotensin II-treated adult cardiac fibroblasts and renal artery-ligated rat heart, suggests that AA-driven cardioprotection primarily arises from PPARα agonism. Moreover, AA-induced PPARα up-regulation leads to repression of TGF-β signaling, specifically by inhibiting TGF-β-activated kinase1 (TAK1) phosphorylation. We observed that PPARα directly interacts with TAK1, predominantly via PPARα N-terminal transactivation domain (AF-1) thereby masking the TAK1 kinase domain. The AA-induced PPARα-bound TAK1 level thereby shows inverse correlation with the phosphorylation level of TAK1 and subsequent reduction in p38 MAPK and NF-κBp65 activation, ultimately culminating in amelioration of excess collagen synthesis in cardiac hypertrophy. In conclusion, our findings unravel the mechanism of AA action in regressing hypertrophy-associated cardiac fibrosis by assigning a role of AA as a PPARα agonist that inactivates non-canonical TGF-β signaling. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  4. The polyphenol-rich extract from grape seeds inhibits platelet signaling pathways triggered by both proteolytic and non-proteolytic agonists.

    PubMed

    Olas, Beata; Wachowicz, Barbara; Stochmal, Anna; Oleszek, Wiesław

    2012-01-01

    Mechanisms involved in the reduction of blood platelet functions by various plant extract, including the grape seeds extract (rich in phenolic compounds, a mixture of about 95% oligomeric phenols; GSE) are still unclear. In the literature there are few papers describing studies on the effects of GSE on selected element of hemostasis. The aim of our study was to establish and compare the influence of GSE (at final dose of 0.625-50 µg/ml) and resveratrol (3,4',5 - trihydroxystilben), a phenolic compound synthesized in grapes and vegetables and presents in wine, which has been supposed to be beneficial for the prevention of cardiovascular events, on different steps of platelet activation. We measured the effects of GSE and resveratrol on platelet aggregation, the surface expression of P-selectin, platelet microparticle formation (PMP), and superoxide anion radicals ([Formula: see text]) production in blood platelets stimulated by TRAP and thrombin. P-selectin expression and PMP formation were measured by a flow cytometer. In gel-filtered platelets activated by thrombin or TRAP and treated with different concentrations of GSE (1.25-50 µg/ml) a significant decrease of P-selectin expression, PMP formation and platelet aggregation was observed. GSE caused also a dose-dependent reduction of [Formula: see text] produced in platelets activated by TRAP or thrombin. Our present results indicate that GSE inhibits platelet signaling pathways trigged by both proteolytic (thrombin) and non-proteolytic agonist (TRAP). In the comparative studies, GSE was found to be more effective antiplatelet factor, than the solution of pure resveratrol. Thus, the polyphenol-rich extract from grape seeds can be useful as the protecting factor against cardiovascular diseases.

  5. Clonidine, an alpha-2 adrenoceptor agonist relieves mechanical allodynia in oxaliplatin-induced neuropathic mice; potentiation by spinal p38 MAPK inhibition without motor dysfunction and hypotension.

    PubMed

    Yeo, Ji-Hee; Yoon, Seo-Yeon; Kim, Sol-Ji; Oh, Seog-Bae; Lee, Jang-Hern; Beitz, Alvin J; Roh, Dae-Hyun

    2016-05-15

    Cancer chemotherapy with platinum-based antineoplastic agents including oxaliplatin frequently results in a debilitating and painful peripheral neuropathy. We evaluated the antinociceptive effects of the alpha-2 adrenoceptor agonist, clonidine on oxaliplatin-induced neuropathic pain. Specifically, we determined if (i) the intraperitoneal (i.p.) injection of clonidine reduces mechanical allodynia in mice with an oxaliplatin-induced neuropathy and (ii) concurrent inhibition of p38 mitogen-activated protein kinase (MAPK) activity by the p38 MAPK inhibitor SB203580 enhances clonidine's antiallodynic effect. Clonidine (0.01-0.1 mg kg(-1), i.p.), with or without SB203580(1-10 nmol, intrathecal) was administered two weeks after oxaliplatin injection(10 mg kg(-1), i.p.) to mice. Mechanical withdrawal threshold, motor coordination and blood pressure were measured. Postmortem expression of p38 MAPK and ERK as well as their phosphorylated forms(p-p38 and p-ERK) were quantified 30 min or 4 hr after drug injection in the spinal cord dorsal horn of treated and control mice. Clonidine dose-dependently reduced oxaliplatin-induced mechanical allodynia and spinal p-p38 MAPK expression, but not p-ERK. At 0.1 mg kg(-1), clonidine also impaired motor coordination and decreased blood pressure. A 10 nmol dose of SB203580 alone significantly reduced mechanical allodynia and p-p38 MAPK expression, while a subeffective dose(3 nmol) potentiated the antiallodynic effect of 0.03 mg kg(-1) clonidine and reduced the increased p-p38 MAPK. Coadministration of SB203580 and 0.03 mg kg(-1) clonidine decreased allodynia similar to that of 0.10 mg kg(-1) clonidine, but without significant motor or vascular effects. These findings demonstrate that clonidine treatment reduces oxaliplatin-induced mechanical allodynia. The concurrent administration of SB203580 reduces the dosage requirements for clonidine, thereby alleviating allodynia without producing undesirable motor or cardiovascular effects.

  6. Hepatic mitochondrial DNA/Toll-like receptor 9/MicroRNA-223 forms a negative feedback loop to limit neutrophil overactivation and acetaminophen hepatotoxicity in mice.

    PubMed

    He, Yong; Feng, Dechun; Li, Man; Gao, Yanhang; Ramirez, Teresa; Cao, Haixia; Kim, Seung-Jin; Yang, Yang; Cai, Yan; Ju, Cynthia; Wang, Hua; Li, Jun; Gao, Bin

    2017-07-01

    Acetaminophen (APAP) overdose is a leading cause of acute liver failure worldwide, in which mitochondrial DNA (mtDNA) released by damaged hepatocytes activates neutrophils through binding of Toll-like receptor 9 (TLR9), further aggravating liver injury. Here, we demonstrated that mtDNA/TLR9 also activates a negative feedback pathway through induction of microRNA-223 (miR-223) to limit neutrophil overactivation and liver injury. After injection of APAP in mice, levels of miR-223, the most abundant miRNAs in neutrophils, were highly elevated in neutrophils. Disruption of the miR-223 gene exacerbated APAP-induced hepatic neutrophil infiltration, oxidative stress, and injury and enhanced TLR9 ligand-mediated activation of proinflammatory mediators in neutrophils. An additional deletion of the intercellular adhesion molecule 1 (ICAM-1) gene ameliorated APAP-induced neutrophil infiltration and liver injury in miR-223 knockout mice. In vitro experiments revealed that miR-223-deficient neutrophils were more susceptible to TLR9 agonist-mediated induction of proinflammatory mediators and nuclear factor kappa B (NF-κB) signaling, whereas overexpression of miR-223 attenuated these effects in neutrophils. Moreover, inhibition of TLR9 signaling by either treatment with a TLR9 inhibitor or by disruption of TLR9 gene partially, but significantly, suppressed miR-223 expression in neutrophils post-APAP injection. In contrast, activation of TLR9 up-regulated miR-223 expression in neutrophils in vivo and in vitro. Mechanistically, activation of TLR9 up-regulated miR-223 by enhancing NF-κB binding on miR-223 promoter, whereas miR-223 attenuated TLR9/NF-κB-mediated inflammation by targeting IκB kinase α expression. Collectively, up-regulation of miR-223 plays a key role in terminating the acute neutrophilic response and is a therapeutic target for treatment of APAP-induced liver failure. (Hepatology 2017;66:220-234). © 2017 by the American Association for the Study of Liver

  7. The Toll-like receptor 9 signalling pathway regulates MR1-mediated bacterial antigen presentation in B cells.

    PubMed

    Liu, Jianyun; Brutkiewicz, Randy R

    2017-10-01

    Mucosal-associated invariant T (MAIT) cells are conserved T cells that express a semi-invariant T-cell receptor (Vα7.2 in humans and Vα19 in mice). The development of MAIT cells requires the antigen-presenting MHC-related protein 1 (MR1), as well as commensal bacteria. The mechanisms that regulate the functional expression of MR1 molecules and their loading with bacterial antigen in antigen-presenting cells are largely unknown. We have found that treating B cells with the Toll-like receptor 9 (TLR9) agonist CpG increases MR1 surface expression. Interestingly, activation of TLR9 by CpG-A (but not CpG-B) enhances MR1 surface expression. This is limited to B cells and not other types of cells such as monocytes, T or natural killer cells. Knocking-down TLR9 expression by short hairpin RNA reduces MR1 surface expression and MR1-mediated bacterial antigen presentation. CpG-A triggers early endosomal TLR9 activation, whereas CpG-B is responsible for late endosomal/lysosomal activation of TLR9. Consistently, blocking endoplasmic reticulum to Golgi protein transport, rather than lysosomal acidification, suppressed MR1 antigen presentation. Overall, our results indicate that early endosomal TLR9 activation is important for MR1-mediated bacterial antigen presentation. © 2017 John Wiley & Sons Ltd.

  8. Toll-Like Receptor 9 Activation Rescues Impaired Antibody Response in Needle-free Intradermal DNA Vaccination

    PubMed Central

    Arunachalam, Prabhu S.; Mishra, Ria; Badarinath, Krithika; Selvam, Deepak; Payeli, Sravan K.; Stout, Richard R.; Ranga, Udaykumar

    2016-01-01

    The delivery of plasmid DNA to the skin can target distinct subsets of dermal dendritic cells to confer a superior immune response. The needle-free immunization technology offers a reliable, safe and efficient means to administer intradermal (ID) injections. We report here that the ID injection of DNA vectors using an NF device (NF-ID) elicits a superior cell-mediated immune response, at much lesser DNA dosage, comparable in magnitude to the traditional intramuscular immunization. However, the humoral response is significantly impaired, possibly at the stage of B cell isotype switching. We found that the NF-ID administration deposits the DNA primarily on the epidermis resulting in a rapid loss of the DNA as well as the synthesized antigen due to the faster regeneration rate of the skin layers. Therefore, despite the immune-rich nature of the skin, the NF-ID immunization of DNA vectors may be limited by the impaired humoral response. Additional booster injections are required to augment the antibody response. As an alternative and a viable solution, we rescued the IgG response by coadministration of a Toll-like receptor 9 agonist, among other adjuvants examined. Our work has important implication for the optimization of the emerging needle-free technology for ID immunization. PMID:27658623

  9. Inhibition of plaque progression and promotion of plaque stability by glucagon-like peptide-1 receptor agonist: Serial in vivo findings from iMap-IVUS in Watanabe heritable hyperlipidemic rabbits.

    PubMed

    Sudo, Mitsumasa; Li, Yuxin; Hiro, Takafumi; Takayama, Tadateru; Mitsumata, Masako; Shiomi, Masashi; Sugitani, Masahiko; Matsumoto, Taro; Hao, Hiroyuki; Hirayama, Atsushi

    2017-10-01

    Glucagon-like peptide-1 (GLP-1) is thought to inhibit development of aortic atherosclerosis and plaque formation. However, whether GLP-1 stabilizes fully developed atherosclerotic plaque or alters the complicated plaque composition remains unclarified. Ten Watanabe heritable hyperlipidemic (WHHL) rabbits were divided into GLP-1 receptor agonist treatment group and control group. After confirmation of atherosclerotic plaques in brachiocephalic arteries by iMap intravascular ultrasound (iMAP-IVUS), GLP-1 receptor agonist lixisenatide was administered to WHHL rabbits at 30 nmoL/kg/day for 12 weeks by osmotic pump. An equal volume of normal saline was administered in a control group. After evaluation by iMAP-IVUS at 12 weeks, brachiocephalic arteries were harvested for pathological histological analysis. iMAP-IVUS analysis revealed larger fibrotic plaque components and smaller necrotic and calcified plaque components in the GLP-1 group than in the control group; %fibrotic area: 66.30 ± 2.06% vs. 75.14 ± 2.62%, p < 0.01, %necrotic area: 23.25 ± 1.87% vs. 16.17 ± 2.27%, p = 0.02, %calcified area: 2.15 ± 0.24% vs. 1.00 ± 0.18%, p < 0.01), indicating that GLP-1 receptor agonist might modify plaque composition and increase plaque stability. Histological analysis confirmed that GLP-1 receptor agonist treatment improved smooth muscle cell (SMC)-rich plaque with increased fibrotic content. Furthermore, plaque macrophage infiltration and calcification were significantly reduced by GLP-1 receptor agonist treatment; %SMC area: 6.93 ± 0.31% vs. 8.14 ± 0.48%, p = 0.02; %macrophage area: 9.11 ± 0.80% vs. 6.19 ± 0.85%, p < 0.01; %fibrotic area: 54.75 ± 1.63% vs. 69.60 ± 2.12%, p = 0.02; %calcified area: 3.25 ± 0.67% vs. 0.75 ± 0.15%, p = 0.02). GLP-1 receptor agonist inhibited plaque progression and promoted plaque stabilization by inhibiting plaque growth and modifying plaque composition. Copyright © 2017 Elsevier B.V. All

  10. Inhibition of TNF-α, IL-1α, and IL-1β by Pretreatment of Human Monocyte-Derived Macrophages with Menaquinone-7 and Cell Activation with TLR Agonists In Vitro.

    PubMed

    Pan, Min-Hsiung; Maresz, Katarzyna; Lee, Pei-Sheng; Wu, Jia-Ching; Ho, Chi-Tang; Popko, Janusz; Mehta, Dilip S; Stohs, Sidney J; Badmaev, Vladimir

    2016-07-01

    Circulatory markers of low-grade inflammation such as tumor necrosis factor-alpha (TNF-α), interleukin-1 alpha (IL-1α), and interleukin-1 beta (IL-1β) positively correlate with endothelial damage, atheroma formation, cardiovascular disease, and aging. The natural vitamin K2-menaquinone-7 (MK-7) added to the cell culture of human monocyte-derived macrophages (hMDMs) at the same time as toll-like receptor (TLR) agonists did not influence the production of TNF-α. When the cells were pretreated up to 6 h with MK-7 before treatment with TLR agonists, MK-7 did not inhibit significantly the production of TNF-α after the TLR activation. However, 30 h pretreatment of hMDMs with at least 10 μM of MK-7 effectively and dose dependently inhibited the proinflammatory function of hMDMs. Pretreatment of hMDMs with 10 μM of MK-7 for 30 h resulted in 20% inhibition of TNF-α production after lipopolysaccharide (LPS) activation (P < .05) and 43% inhibition after macrophage-activating lipopeptide (MALP) activation (P < .001). Pathogen-associated molecular pattern (PMPP) activation was inhibited by 20% with MK-7 pretreatment; however, this inhibition was not statistically significant. The 30 h pretreatment of a THP-1-differentiated monocyte cell line with MK-7 resulted in a dose-dependent downregulation of TNFα, IL-1α, and IL-1β gene expression as evaluated by RNA semiquantitative reverse transcription polymerase chain reaction (RT-PCR). MK-7 is able to modulate immune and inflammatory reactions in the dose-response inhibition of TNF-α, IL-1α, and IL-1β gene expression and protein production by the healthy hMDMs in vitro.

  11. A new class of peroxisome proliferator-activated receptor gamma (PPARgamma) agonists that inhibit growth of breast cancer cells: 1,1-Bis(3'-indolyl)-1-(p-substituted phenyl)methanes.

    PubMed

    Qin, Chunhua; Morrow, Derek; Stewart, Jessica; Spencer, Kyle; Porter, Weston; Smith, Roger; Phillips, Timothy; Abdelrahim, Maen; Samudio, Ismael; Safe, Stephen

    2004-03-01

    1,1-Bis(3'-indolyl)-1-(p-trifluoromethylphenyl)methane (DIM-C-pPhCF(3)) and several p-substituted phenyl analogues have been investigated as a new class of peroxisome proliferator-activated receptor gamma (PPARgamma) agonists. Structure-activity studies in PPARgamma-dependent transactivation assays in MCF-7 breast cancer cells show that 5-20 micro M concentrations of compounds containing p-trifluoromethyl, t-butyl, cyano, dimethylamino, and phenyl groups were active, whereas p-methyl, hydrogen, methoxy, hydroxyl, or halogen groups were inactive as PPARgamma agonists. Induction of PPARgamma-dependent transactivation by 15-deoxy-Delta12,14-prostaglandin J2 (PGJ2) and DIM-C-pPhCF(3) was inhibited in MCF-7 cells cotreated with the PPARgamma-specific antagonist N-(4'-aminopyridyl)-2-chloro-5-nitrobenzamide. In mammalian two-hybrid assays, DIM-C-pPhCF(3) and PGJ2 (5-20 micro M) induced interactions of PPARgamma with steroid receptor coactivator (SRC) 1, SRC2 (TIFII), and thyroid hormone receptor-associated protein 220 but not with SRC3 (AIB1). In contrast, DIM-C-pPhCF(3), but not PGJ2, induced interactions of PPARgamma with PPARgamma coactivator-1. C-substituted diindolylmethanes inhibit carcinogen-induced rat mammary tumor growth, induce differentiation in 3T3-L1 preadipocytes, inhibit MCF-7 cell growth and G(0)/G(1)-S phase progression, induce apoptosis, and down-regulate cyclin D1 protein and estrogen receptor alpha in breast cancer cells. These compounds are a novel class of synthetic PPARgamma agonists that induce responses in MCF-7 cells similar to those observed for PGJ2.

  12. An EP2 Agonist Facilitates NMDA-Induced Outward Currents and Inhibits Dendritic Beading through Activation of BK Channels in Mouse Cortical Neurons

    PubMed Central

    Hayashi, Yoshinori; Morinaga, Saori; Liu, Xia; Zhang, Jing; Wu, Zhou; Yokoyama, Takeshi; Nakanishi, Hiroshi

    2016-01-01

    Prostaglandin E2 (PGE2), a major metabolite of arachidonic acid produced by cyclooxygenase pathways, exerts its bioactive responses by activating four E-prostanoid receptor subtypes, EP1, EP2, EP3, and EP4. PGE2 enables modulating N-methyl-D-aspartate (NMDA) receptor-mediated responses. However, the effect of E-prostanoid receptor agonists on large-conductance Ca2+-activated K+ (BK) channels, which are functionally coupled with NMDA receptors, remains unclear. Here, we showed that EP2 receptor-mediated signaling pathways increased NMDA-induced outward currents (INMDA-OUT), which are associated with the BK channel activation. Patch-clamp recordings from the acutely dissociated mouse cortical neurons revealed that an EP2 receptor agonist activated INMDA-OUT, whereas an EP3 receptor agonist reduced it. Agonists of EP1 or EP4 receptors showed no significant effects on INMDA-OUT. A direct perfusion of 3,5′-cyclic adenosine monophosphate (cAMP) through the patch pipette facilitated INMDA-OUT, which was abolished by the presence of protein kinase A (PKA) inhibitor. Furthermore, facilitation of INMDA-OUT caused by an EP2 receptor agonist was significantly suppressed by PKA inhibitor. Finally, the activation of BK channels through EP2 receptors facilitated the recovery phase of NMDA-induced dendritic beading in the primary cultured cortical neurons. These results suggest that a direct activation of BK channels by EP2 receptor-mediated signaling pathways plays neuroprotective roles in cortical neurons. PMID:27298516

  13. CpG oligodeoxynucleotides induce strong up-regulation of interleukin 33 via Toll-like receptor 9.

    PubMed

    Shimosato, Takeshi; Fujimoto, Megumi; Tohno, Masanori; Sato, Takashi; Tateo, Mariko; Otani, Hajime; Kitazawa, Haruki

    2010-03-26

    We previously reported the strong immunostimulatory effects of a CpG oligodeoxynucleotide (ODN), designated MsST, from the lacZ gene of Streptococcus (S.) thermophilus ATCC19258. Here we show that 24h of stimulation with MsST in mouse splenocytes and peritoneal macrophages strongly induces expression of interleukin (IL)-33, a cytokine in the IL-1 superfamily. Other IL-1 superfamily members, including IL-1alpha, IL-1beta and IL-18, are down-regulated after 24h of stimulation of MsST. We also found that MsST-induced IL-33 mRNA expression is inhibited by the suppressive ODN A151, which can inhibit Toll-like receptor 9 (TLR9)-mediated responses. This is the first report to show that IL-33 can be induced by CpG ODNs. The strong induction of IL-33 by MsST suggests that it may be a potential therapeutic ODN for the treatment of inflammatory disease. The presence of a strong CpG ODN in S. thermophilus also suggests that the bacterium may be a good candidate as a starter culture for the development of new physiologically functional foods.

  14. Natural Killer Cell-Dependent Anti-Fibrotic Pathway in Liver Injury via Toll-Like Receptor-9

    PubMed Central

    Abu-Tair, Lina; Axelrod, Jonathan H.; Doron, Sarit; Ovadya, Yossi; Krizhanovsky, Valery; Galun, Eithan

    2013-01-01

    The toll-like receptor-9 (TLR9) agonist cytosine phosphate guanine (CpG), activates hepatic stellate cells (HSCs) and mediates fibrosis. We investigated the TLR9 effects on lymphocyte/HSCs interactions. Liver fibrosis was induced in wild-type (WT) mice by intra-peritoneal carbon-tetrachloride (CCl4) induction for 6 weeks. Fibrotic groups were intravenously treated by a vehicle versus CpG along last 2 weeks. Compared to vehicle-treated fibrotic WT, the in-vivo CpG-treatment significantly attenuated hepatic fibrosis and inflammation, associated with decreased CD8 and increased NK liver cells. In-vitro, co-cultures with vehicle-treated fibrotic NK cells increased HSCs proliferation (P<0.001) while their CpG-treated counterparts achieved a significant decrease. To investigate the role of lymphocytes, TLR9-/- mice induced-hepatic fibrosis were used. Although TLR9-/- mice manifested lower fibrotic profile as compared to their wild-type (WT) counterparts, senescence (SA-β-Gal activity) in the liver and ALT serum levels were significantly greater. In an adoptive transfer model; irradiated WT and TLR9-/- recipients were reconstituted with naïve WT or TLR9-/- lymphocytes. The adoptive transfer of TLR9-/- versus WT lymphocytes led to increased fibrosis of WT recipients. TLR9-/- fibrotic recipients reconstituted with TLR9-/- or WT lymphocytes showed no changes in hepatic fibrosis severity or ALT serum levels. TLR9 activation had inconsistent effects on lymphocytes and HSCs. The net balance of TLR9 activation in WT, displayed significant anti-fibrotic activity, accompanied by CD8 suppression and increased NK-cells, activity and adherence to HSCs. The pro-fibrotic and pro-inflammatory properties of TLR9-/- lymphocytes fail to activate HSCs with an early senescence in TLR9-/- mice.  PMID:24340043

  15. Inhibition of neutrophil migration in mice by mouse formyl peptide receptors 1 and 2 dual agonist: indication of cross-desensitization in vivo

    PubMed Central

    Sogawa, Yoshitaka; Ohyama, Takao; Maeda, Hiroaki; Hirahara, Kazuki

    2011-01-01

    It has been reported that the stimulation of neutrophils with N-formyl-Met-Leu-Phe (fMLF), an agonist for formyl peptide receptor (Fpr) 1, renders cells unresponsive to other chemoattractants in vitro. This is known as cross-desensitization, but its functional relevance in neutrophil migration in vivo has not been investigated. Here, we show that precedent stimulation of mouse neutrophils with compound 43, a non-peptidyl agonist for mouse Fpr1 and Fpr2, rendered the cells unresponsive to a second stimulation with C5a, leukotriene B4, or keratinocyte-derived cytokine (KC) in calcium mobilization and chemotaxis assays in vitro. The expression of chemokine (C-X-C motif) receptor 2 (CXCR2) on the surface of neutrophils was concomitantly diminished by stimulating the cells with the compound. Moreover, oral administration of the compound to mice before they were exposed to lipopolysaccharide (LPS) aerosol resulted in a dose-dependent reduction in the neutrophil count in bronchoalveolar lavage fluid. The expression of CXCR2 on blood neutrophils was also reduced in the compound-administered mice. The recipient mice that underwent adoptive transfer of fluorescence-labelled neutrophils that had been incubated with the compound showed a substantial decrease in neutrophil counts in bronchoalveolar lavage fluid after they were exposed to LPS, when compared with the control mice to which vehicle-treated neutrophils had been transferred. These results are consistent with the idea that the agonist for Fpr1 and Fpr2 induced cross-desensitization in neutrophils and attenuated neutrophil migration into the airways. Our results also revealed the unpredicted effect of an Fpr1 and Fpr2 dual agonist, which may act as a functional antagonist for multiple chemoattractant receptors in vivo. PMID:21039475

  16. TLR9 agonist acts by different mechanisms synergizing with bevacizumab in sensitive and cetuximab-resistant colon cancer xenografts.

    PubMed

    Damiano, Vincenzo; Caputo, Rosa; Garofalo, Sonia; Bianco, Roberto; Rosa, Roberta; Merola, Gerardina; Gelardi, Teresa; Racioppi, Luigi; Fontanini, Gabriella; De Placido, Sabino; Kandimalla, Ekambar R; Agrawal, Sudhir; Ciardiello, Fortunato; Tortora, Giampaolo

    2007-07-24

    Synthetic agonists of Toll-like receptor 9 (TLR9), a class of agents that induce specific immune response, exhibit antitumor activity and are currently being investigated in cancer patients. Intriguingly, their mechanisms of action on tumor growth and angiogenesis are still incompletely understood. We recently discovered that a synthetic agonist of TLR9, immune modulatory oligonucleotide (IMO), acts by impairing epidermal growth factor receptor (EGFR) signaling and potently synergizes with anti-EGFR antibody cetuximab in GEO human colon cancer xenografts, whereas it is ineffective in VEGF-overexpressing cetuximab-resistant GEO cetuximab-resistant (GEO-CR) tumors. VEGF is activated by EGFR, and its overexpression causes resistance to EGFR inhibitors. Therefore, we used IMO and the anti-VEGF antibody bevacizumab as tools to study IMO's role on EGFR and angiogenesis and to explore its therapeutic potential in GEO, LS174T, and GEO-CR cancer xenografts. We found that IMO enhances the antibody-dependent cell-mediated cytotoxicity (ADCC) activity of cetuximab, that bevacizumab has no ADCC, and IMO is unable to enhance it. Nevertheless, the IMO-plus-bevacizumab combination synergistically inhibits the growth of GEO and LS174T as well as of GEO-CR tumors, preceded by inhibition of signaling protein expression, microvessel formation, and human, but not murine, VEGF secretion. Moreover, IMO inhibited the growth, adhesion, migration, and capillary formation of VEGF-stimulated endothelial cells. The antitumor activity was irrespective of the TLR9 expression on tumor cells. These studies demonstrate that synthetic agonists of TLR9 interfere with growth and angiogenesis also by EGFR- and ADCC-independent mechanisms affecting endothelial cell functions and provide a strong rationale to combine IMO with bevacizumab and EGFR inhibitory drugs in colon cancer patients.

  17. TLR9 agonist acts by different mechanisms synergizing with bevacizumab in sensitive and cetuximab-resistant colon cancer xenografts

    PubMed Central

    Damiano, Vincenzo; Caputo, Rosa; Garofalo, Sonia; Bianco, Roberto; Rosa, Roberta; Merola, Gerardina; Gelardi, Teresa; Racioppi, Luigi; Fontanini, Gabriella; De Placido, Sabino; Kandimalla, Ekambar R.; Agrawal, Sudhir; Ciardiello, Fortunato; Tortora, Giampaolo

    2007-01-01

    Synthetic agonists of Toll-like receptor 9 (TLR9), a class of agents that induce specific immune response, exhibit antitumor activity and are currently being investigated in cancer patients. Intriguingly, their mechanisms of action on tumor growth and angiogenesis are still incompletely understood. We recently discovered that a synthetic agonist of TLR9, immune modulatory oligonucleotide (IMO), acts by impairing epidermal growth factor receptor (EGFR) signaling and potently synergizes with anti-EGFR antibody cetuximab in GEO human colon cancer xenografts, whereas it is ineffective in VEGF-overexpressing cetuximab-resistant GEO cetuximab-resistant (GEO-CR) tumors. VEGF is activated by EGFR, and its overexpression causes resistance to EGFR inhibitors. Therefore, we used IMO and the anti-VEGF antibody bevacizumab as tools to study IMO's role on EGFR and angiogenesis and to explore its therapeutic potential in GEO, LS174T, and GEO-CR cancer xenografts. We found that IMO enhances the antibody-dependent cell-mediated cytotoxicity (ADCC) activity of cetuximab, that bevacizumab has no ADCC, and IMO is unable to enhance it. Nevertheless, the IMO-plus-bevacizumab combination synergistically inhibits the growth of GEO and LS174T as well as of GEO-CR tumors, preceded by inhibition of signaling protein expression, microvessel formation, and human, but not murine, VEGF secretion. Moreover, IMO inhibited the growth, adhesion, migration, and capillary formation of VEGF-stimulated endothelial cells. The antitumor activity was irrespective of the TLR9 expression on tumor cells. These studies demonstrate that synthetic agonists of TLR9 interfere with growth and angiogenesis also by EGFR- and ADCC-independent mechanisms affecting endothelial cell functions and provide a strong rationale to combine IMO with bevacizumab and EGFR inhibitory drugs in colon cancer patients. PMID:17636117

  18. The protective effect of the anti-Toll-like receptor 9 antibody against acute cytokine storm caused by immunostimulatory DNA

    PubMed Central

    Murakami, Yusuke; Fukui, Ryutaro; Motoi, Yuji; Shibata, Takuma; Saitoh, Shin-Ichiroh; Sato, Ryota; Miyake, Kensuke

    2017-01-01

    Toll-like Receptor 9 (TLR9) is an innate immune receptor recognizing microbial DNA. TLR9 is also activated by self-derived DNA, such as mitochondrial DNA, in a variety of inflammatory diseases. We show here that TLR9 activation in vivo is controlled by an anti-TLR9 monoclonal Ab (mAb). A newly established mAb, named NaR9, clearly detects endogenous TLR9 expressed in primary immune cells. The mAb inhibited TLR9-dependent cytokine production in vitro by bone marrow-derived macrophages and conventional dendritic cells. Furthermore, NaR9 treatment rescued mice from fulminant hepatitis caused by administering the TLR9 ligand CpGB and D-(+)-galactosamine. The production of proinflammatory cytokines induced by CpGB and D-(+)-galactosamine was significantly impaired by the mAb. These results suggest that a mAb is a promising tool for therapeutic intervention in TLR9-dependent inflammatory diseases. PMID:28266597

  19. Grancalcin (GCA) modulates Toll-like receptor 9 (TLR9) mediated signaling through its direct interaction with TLR9.

    PubMed

    Kim, Tae Whan; Hong, Seunghee; Talukder, Amjad H; Pascual, Virginia; Liu, Yong-Jun

    2016-03-01

    Toll-like receptors (TLRs) are playing important roles in stimulating the innate immune response and intensifying adaptive immune response against invading pathogens. Appropriate regulation of TLR activation is important to maintain a balance between preventing tumor activation and inhibiting autoimmunity. Toll-like receptor 9 (TLR9) senses microbial DNA in the endosomes of plasmacytoid dendritic cells and triggers myeloid differentiation primary response gene 88 (MyD88) dependent nuclear factor kappa B (NF-κB) pathways and type I interferon (IFN) responses. However, mechanisms of how TLR9 signals are mediated and which molecules are involved in controlling TLR9 functions remain poorly understood. Here, we report that penta EF-hand protein grancalcin (GCA) interacts and binds with TLR9 in a yeast two-hybrid system and an overexpression system. Using siRNA-mediated knockdown experiments, we also revealed that GCA positively regulates type I IFN production, cytokine/chemokine production through nuclear localization of interferon regulatory factor 7 (IRF7), NF-κB activation, and mitogen-activated protein kinase (MAPK) activation in plasmacytoid dendritic cells. Our results indicate that heterodimerization of GCA and TLR9 is important for TLR9-mediated downstream signaling and might serve to fine tune processes against viral infection. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Toll-like receptor 9 mediates paraquat-induced acute lung injury: an in vitro and in vivo study.

    PubMed

    Shen, Haitao; Wu, Na; Wang, Yu; Zhang, Lichun; Hu, Xiao; Chen, Zhiguang; Zhao, Min

    2017-06-01

    This study aimed to investigate the role of Toll-like receptor 9 in paraquat-induced acute lung injury (ALI). For in vivo study,C57BL mice were randomly assigned into the vehicle control group, paraquat group, paraquat + TLR9 antagonist (ODN2088) group, and TLR9 antagonist (ODN2088) group (n=36 per group). After paraquat 30mg/kg ip for 2, 24 and 48h, serum samples and lung tissues were collected to evaluate ALI and TLR9 signaling by lung injury score, protein levels of TLR9, MyD88, p-IRAK4, p-p65, and serum TNF-α and IL-1β levels. As for in vitro research A549 cells were randomly divided into the control group, paraquat group, paraquat + TLR9 siRNA group, and TLR9 siRNA group. After paraquat treatment for 24h, the cells and supernatant were collected to measureTLR9, TNF-α, IL-1 mRNA expression, and detect activation of NF-κB, caspase-3. In vivo, the lung injury score, the TLR9, MyD88, p-IRAK4 and p-p65 protein levels, and cytokines TNF-α and IL-1β levels in paraquat group were significantly higher than that in the control group;TLR9 blocker ODN2088 pretreatment attenuated lung injury, inhibited MyD88 and NF-κB activation, and reduced TNF-α and IL-1β in serum. In vitro result shows that the gene silencing of TLR9 reduced the mRNA expression of TLR9, TNF-α and IL-1, inhibited NF-κB and caspase-3 activation, attenuated cell apoptosis. TLR9 mediates paraquat-induced ALI, antagonizing TLR9 or silencing TLR9gene may attenuate paraquat-induced ALI. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. Toll-like receptor 9 gene polymorphism in chronic and aggressive periodontitis patients

    PubMed Central

    Ashok, Nipun; Warad, Shivaraj; Kalburgi, Nagaraj Balasaheb; Bilichodmath, Shivaprasad; Prabhakaran, Prabath Singh Valiyaparambil; Tarakji, Bassel

    2014-01-01

    Aim: Periodontitis is a multifactorial disease, with microbial dental plaque as the primary etiological factor. However, the manifestation and progression of periodontitis is influenced by a wide variety of other determinants and factors such as social and behavioral factors, systemic factors, microbial composition of dental plaque, genetic, and many other emerging risk factors. The aim of this study was to analyze genetic polymorphisms in the toll-like receptor 9 (TLR9) gene at - 1237C/T and its association with chronic and generalized aggressive periodontitis (GAgP) in an Indian population. Materials and Methods: This study was carried out on 90 subjects, which included 30 GAgP and 30 chronic periodontitis patients and 30 healthy controls. Within the limitations of our study, only 30 subjects were included in each group due to the low prevalence of GAgP patients. Blood samples were drawn from the subjects and analyzed for TLR9 genetic polymorphism at - 1237C/T by using polymerase chain reaction-restriction fragment length polymorphism method. Results: No significant difference was found in genotype and allele frequency of TLR9 genetic polymorphism (- 1237C/T) in generalized aggressive and chronic periodontitis patients and healthy controls. Conclusion: Toll-like receptor 9 genetic polymorphism at - 1237C/T may not be associated with GAgP and chronic periodontitis patients in Indian population. PMID:25624628

  2. Inhibition of mammary tumor growth in rats and mice by administration of agonistic and antagonistic analogs of luteinizing hormone-releasing hormone.

    PubMed Central

    Redding, T W; Schally, A V

    1983-01-01

    Experiments were undertaken with estrogen-dependent mammary carcinomas in rats and mice to determine the antitumor activities of agonistic and antagonistic analogs of luteinizing hormone-releasing hormone (LH-RH). Chronic administration of the agonist [D-Trp6]LH-RH or of antagonist 1 ( [NAc-D-p-Cl-Phe1,2-Phe3,D-Arg6-D-Ala10]LH-RH) at doses of 25 and 50 micrograms/day, respectively, for 21 days to mice bearing the MXT mammary carcinoma significantly decreased tumor weight and volume. The weight of the ovaries and serum progesterone levels in mice treated with [D-Trp6]LH-RH or antagonist 1 were also significantly reduced. In rats bearing the MT/W9A mammary adenocarcinoma, chronic administration of [D-Trp6]LH-RH at a dose of 25 micrograms twice a day or of antagonist 2 ( [NAc-D-p-Cl-Phe1,2,D-Trp3,D-Arg6,D-Ala10]LH-RH) at a dose of 50 micrograms twice a day for 28 days significantly decreased tumor weight and volume. Chronic treatment with either [D-Trp6]LH-RH or antagonist 2 markedly diminished the weight of the ovaries and serum levels of both estrogen and progesterone. Serum luteinizing hormone was significantly decreased in rats treated with antagonist 2 but not in rats treated with [D-Trp6]LH-RH. There was a significant drop in serum prolactin levels in rats treated with [D-Trp6]LH-RH but not in those receiving antagonist 2. Regression of mammary tumors in rats and mice in response to chronic administration of [D-Trp6]LH-RH and the two antagonistic analogs of LH-RH suggests that these compounds should be considered for the development of a new hormone therapy for breast cancer in women. PMID:6219395

  3. Inflammatory stimuli inhibit glucocorticoid-dependent transactivation in human pulmonary epithelial cells: rescue by long-acting beta2-adrenoceptor agonists.

    PubMed

    Rider, Christopher F; King, Elizabeth M; Holden, Neil S; Giembycz, Mark A; Newton, Robert

    2011-09-01

    By repressing inflammatory gene expression, glucocorticoids are the most effective treatment for chronic inflammatory diseases such as asthma. However, in some patients with severe disease, or who smoke or suffer from chronic obstructive pulmonary disease, glucocorticoids are poorly effective. Although many investigators focus on defects in the repression of inflammatory gene expression, glucocorticoids also induce (transactivate) the expression of numerous genes to elicit anti-inflammatory effects. Using human bronchial epithelial (BEAS-2B) and pulmonary (A549) cells, we show that cytokines [tumor necrosis factor α (TNFα) and interleukin 1β], mitogens [fetal calf serum (FCS) and phorbol ester], cigarette smoke, and a G(q)-linked G protein-coupled receptor agonist attenuate simple glucocorticoid response element (GRE)-dependent transcription. With TNFα and FCS, this effect was not overcome by increasing concentrations of dexamethasone, budesonide, or fluticasone propionate. Thus, the maximal ability of the glucocorticoid to promote GRE-dependent transcription was reduced, and this was shown additionally for the glucocorticoid-induced gene p57(KIP2). The long-acting β(2)-adrenoceptor agonists (LABAs) formoterol fumarate and salmeterol xinafoate enhanced simple GRE-dependent transcription to a level that could not be achieved by glucocorticoid alone. In the presence of TNFα or FCS, which repressed glucocorticoid responsiveness, these LABAs restored glucocorticoid-dependent transcription to levels that were achieved by glucocorticoid alone. Given the existence of genes, such as p57(KIP2), which may mediate anti-inflammatory actions of glucocorticoids, we propose that repression of transactivation represents a mechanism for glucocorticoid resistance and for understanding the clinical benefit of LABAs as an add-on therapy in asthma and chronic obstructive pulmonary disease.

  4. Retinoic acid receptor agonist Am80 inhibits CXCL2 production from microglial BV-2 cells via attenuation of NF-κB signaling.

    PubMed

    Takaoka, Yuichiro; Takahashi, Moeka; Kurauchi, Yuki; Hisatsune, Akinori; Seki, Takahiro; Shudo, Koichi; Katsuki, Hiroshi

    2016-09-01

    Accumulating lines of evidence suggest that retinoic acid receptor agonists such as Am80 exerts anti-inflammatory actions in the central nervous system, although detailed mechanisms of the action remain largely unknown. Our previous findings suggest that Am80 provides therapeutic effect on intracerebral hemorrhage in mice via suppression of expression of chemokine (C-X-C motif) ligand 2 (CXCL2). Here we investigated the mechanisms of inhibitory action of Am80 on expression of CXCL2 and other pro-inflammatory factors in microglial BV-2 cells. Pretreatment with Am80 markedly suppressed lipopolysaccharide (LPS)-induced expression of CXCL2 mRNA and release of CXCL2 protein. Am80 had no effect on LPS-induced activation of p38 mitogen-activated protein kinase and extracellular signal-regulated kinase. On the other hand, Am80 prevented LPS-induced nuclear translocation of p65 subunit of NF-κB complex. In addition, total expression levels of p65 and IκBα proteins, as well as of mRNAs encoding p65 and IκBα, were lowered by Am80. Dependence of CXCL2 expression on NF-κB was confirmed by the effect of an NF-κB inhibitor caffeic acid phenethyl ester that abolished LPS-induced CXCL2 expression. Caffeic acid phenethyl ester also abolished LPS-induced expression of inducible nitric oxide synthase, interleukin-1β and tumor necrosis factor α, which may be relevant to the inhibitory effect of Am80 on expression of these pro-inflammatory factors. We additionally found that Am80 attenuated LPS-induced up-regulation of CD14, a co-receptor for Toll-like receptor 4 (TLR4). These results suggest that inhibitory effect on TLR4 signaling mediated by NF-κB pathway underlies the anti-inflammatory action of retinoic acid receptor agonists in microglia.

  5. PADMA-28, a traditional tibetan herbal preparation inhibits the respiratory burst in human neutrophils, the killing of epithelial cells by mixtures of oxidants and pro-inflammatory agonists and peroxidation of lipids.

    PubMed

    Ginsburg, I; Sadovnik, M; Sallon, S; Milo-Goldzweig, I; Mechoulam, R; Breuer, A; Gibbs, D; Varani, J; Roberts, S; Cleator, E; Singh, N

    1999-01-01

    Both aqueous and methanolic fractions derived from the Tibetan preparation PADMA-28 (a mixture of 22 plants) used as an anti-atherosclerotic agent, and which is non-cytolytic to a variety of mammalian cells, were found to strongly inhibit (1) the killing of epithelial cells in culture induced by 'cocktails' comprising oxidants, membrane perforating agents and proteinases; (2) the generation of luminol-dependent chemiluminescence in human neutrophils stimulated by opsonized bacteria; (3) the peroxidation of intralipid (a preparation rich in phopholipids) induced in the presence of copper; and (4) the activity of neutrophil elastase. It is proposed that PADMA-28 might prove beneficial for the prevention of cell damage induced by synergism among pro-inflammatory agonists which is central in the initiation of tissue destruction in inflammatory and infectious conditions.

  6. Aspirin metabolites are GPR35 agonists.

    PubMed

    Deng, Huayun; Fang, Ye

    2012-07-01

    Aspirin is widely used as an anti-inflammatory, anti-platelet, anti-pyretic, and cancer-preventive agent; however, the molecular mode of action is unlikely due entirely to the inhibition of cyclooxygenases. Here, we report the agonist activity of several aspirin metabolites at GPR35, a poorly characterized orphan G protein-coupled receptor. 2,3,5-Trihydroxybenzoic acid, an aspirin catabolite, was found to be the most potent GPR35 agonist among aspirin metabolites. Salicyluric acid, the main metabolite of aspirin, was also active. These results suggest that the GPR35 agonist activity of certain aspirin metabolites may contribute to the clinical features of aspirin.

  7. GPR55 agonist lysophosphatidylinositol and lysophosphatidylcholine inhibit endothelial cell hyperpolarization via GPR-independent suppression of Na(+)-Ca(2+) exchanger and endoplasmic reticulum Ca(2+) refilling.

    PubMed

    Bondarenko, Alexander I; Montecucco, Fabrizio; Panasiuk, Olga; Sagach, Vadim; Sidoryak, Nataliya; Brandt, Karim J; Mach, François

    2017-02-01

    Lysophosphatidylinositol (LPI) and lysophosphatidylcholine (LPC) are lipid signaling molecules that induce endothelium-dependent vasodilation. In addition, LPC suppresses acetylcholine (Ach)-induced responses. We aimed to determine the influence of LPC and LPI on hyperpolarizing responses in vitro and in situ endothelial cells (EC) and identify the underlying mechanisms. Using patch-clamp method, we show that LPI and LPC inhibit EC hyperpolarization to histamine and suppress Na(+)/Ca(2+) exchanged (NCX) currents in a concentration-dependent manner. The inhibition is non-mode-specific and unaffected by intracellular GDPβS infusion and tempol, a superoxide dismutase mimetic. In excised mouse aorta, LPI strongly inhibits the sustained and the peak endothelial hyperpolarization induced by Ach, but not by SKA-31, an opener of Ca(2+)-dependent K(+) channels of intermediate and small conductance. The hyperpolarizing responses to consecutive histamine applications are strongly reduced by NCX inhibition. In a Ca(2+)-re-addition protocol, bepridil, a NCX inhibitor, and KB-R7943, a blocker of reversed NCX, inhibit the hyperpolarizing responses to Ca(2+)-re-addition following Ca(2+) stores depletion. These finding indicate that LPC and LPI inhibit endothelial hyperpolarization to Ach and histamine independently of G-protein coupled receptors and superoxide anions. Reversed NCX is critical for ER Ca(2+) refilling in EC. The inhibition of NCX by LPI and LPC underlies diminished endothelium-dependent responses and endothelial dysfunction accompanied by increased levels of these lipids in the blood. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. The structural basis for agonist and partial agonist action on a β(1)-adrenergic receptor.

    PubMed

    Warne, Tony; Moukhametzianov, Rouslan; Baker, Jillian G; Nehmé, Rony; Edwards, Patricia C; Leslie, Andrew G W; Schertler, Gebhard F X; Tate, Christopher G

    2011-01-13

    β-adrenergic receptors (βARs) are G-protein-coupled receptors (GPCRs) that activate intracellular G proteins upon binding catecholamine agonist ligands such as adrenaline and noradrenaline. Synthetic ligands have been developed that either activate or inhibit βARs for the treatment of asthma, hypertension or cardiac dysfunction. These ligands are classified as either full agonists, partial agonists or antagonists, depending on whether the cellular response is similar to that of the native ligand, reduced or inhibited, respectively. However, the structural basis for these different ligand efficacies is unknown. Here we present four crystal structures of the thermostabilized turkey (Meleagris gallopavo) β(1)-adrenergic receptor (β(1)AR-m23) bound to the full agonists carmoterol and isoprenaline and the partial agonists salbutamol and dobutamine. In each case, agonist binding induces a 1 Å contraction of the catecholamine-binding pocket relative to the antagonist bound receptor. Full agonists can form hydrogen bonds with two conserved serine residues in transmembrane helix 5 (Ser(5.42) and Ser(5.46)), but partial agonists only interact with Ser(5.42) (superscripts refer to Ballesteros-Weinstein numbering). The structures provide an understanding of the pharmacological differences between different ligand classes, illuminating how GPCRs function and providing a solid foundation for the structure-based design of novel ligands with predictable efficacies.

  9. Inhibition of alpha-synuclein aggregation by multifunctional dopamine agonists assessed by a novel in vitro assay and an in vivo Drosophila synucleinopathy model

    PubMed Central

    Yedlapudi, Deepthi; Joshi, Gnanada S.; Luo, Dan; Todi, Sokol V.; Dutta, Aloke K.

    2016-01-01

    Aggregation of alpha synuclein (α-syn) leading to dopaminergic neuronal death has been recognized as one of the main pathogenic factors in the initiation and progression of Parkinson’s disease (PD). Consequently, α-syn has been targeted for the development of therapeutics for PD. We have developed a novel assay to screen compounds with α-syn modulating properties by mimicking recent findings from in vivo animal studies involving intrastriatal administration of pre-formed fibrils in mice, resulting in increased α-syn pathology accompanying the formation of Lewy-body (LB) type inclusions. We found that in vitro generated α-syn pre-formed fibrils induce seeding of α-syn monomers to produce aggregates in a dose-and time-dependent manner under static conditions in vitro. These aggregates were toxic towards rat pheochromocytoma cells (PC12). Our novel multifunctional dopamine agonists D-519 and D-520 exhibited significant neuroprotection in this assay, while their parent molecules did not. The neuroprotective properties of our compounds were further evaluated in a Drosophila model of synucleinopathy. Both of our compounds showed protective properties in fly eyes against the toxicity caused by α-syn. Thus, our in vitro results on modulation of aggregation and toxicity of α-syn by our novel assay were further validated with the in vivo experiments. PMID:27917933

  10. Inhibition of rotavirus ECwt infection in ICR suckling mice by N-acetylcysteine, peroxisome proliferator-activated receptor gamma agonists and cyclooxygenase-2 inhibitors

    PubMed Central

    Guerrero, Carlos Arturo; Pardo, Paula; Rodriguez, Victor; Guerrero, Rafael; Acosta, Orlando

    2013-01-01

    Live attenuated vaccines have recently been introduced for preventing rotavirus disease in children. However, alternative strategies for prevention and treatment of rotavirus infection are needed mainly in developing countries where low vaccine coverage occurs. In the present work, N-acetylcysteine (NAC), ascorbic acid (AA), some nonsteroidal anti-inflammatory drugs (NSAIDs) and peroxisome proliferator-activated receptor gamma (PPARγ) agonists were tested for their ability to interfere with rotavirus ECwt infectivity as detected by the percentage of viral antigen-positive cells of small intestinal villi isolated from ECwt-infected ICR mice. Administration of 6 mg NAC/kg every 8 h for three days following the first diarrhoeal episode reduced viral infectivity by about 90%. Administration of AA, ibuprofen, diclofenac, pioglitazone or rosiglitazone decreased viral infectivity by about 55%, 90%, 35%, 32% and 25%, respectively. ECwt infection of mice increased expression of cyclooxygenase-2, ERp57, Hsc70, NF-κB, Hsp70, protein disulphide isomerase (PDI) and PPARγ in intestinal villus cells. NAC treatment of ECwt-infected mice reduced Hsc70 and PDI expression to levels similar to those observed in villi from uninfected control mice. The present results suggest that the drugs tested in the present work could be assayed in preventing or treating rotaviral diarrhoea in children and young animals. PMID:24037197

  11. An adenosine A3 receptor agonist inhibits DSS-induced colitis in mice through modulation of the NF-κB signaling pathway

    PubMed Central

    Ren, Tianhua; Tian, Ting; Feng, Xiao; Ye, Shicai; Wang, Hao; Wu, Weiyun; Qiu, Yumei; Yu, Caiyuan; He, Yanting; Zeng, Juncheng; Cen, Junwei; Zhou, Yu

    2015-01-01

    The role of the adenosine A3 receptor (A3AR) in experimental colitis is controversial. The A3AR agonist N6-(3-iodobenzyl)adenosine-5'-N-methyluronamide (IB-MECA) has been shown to have a clinical benefit, although studies in A3AR-deficient mice suggest a pro-inflammatory role. However, there are no studies on the effect of 2-Cl-IB-MECA and the molecular mechanism of action of A3AR in murine colitis models in vivo. Is it the same as that observed in vitro? The interaction between 2-CL-IB-MECA and A3AR in a murine colitis model and the signaling pathways associated with this interaction remain unclear. Here we demonstrate a role for the NF-κB signaling pathway and its effect on modifying the activity of proinflammatory factors in A3AR-mediated biological processes. Our results demonstrated that A3AR activation possessed marked effects on experimental colitis through the NF-κB signaling pathway. PMID:25762375

  12. Intrathecal baclofen, a GABAB receptor agonist, inhibits the expression of p-CREB and NR2B in the spinal dorsal horn in rats with diabetic neuropathic pain.

    PubMed

    Liu, Peng; Guo, Wen-Ya; Zhao, Xiao-Nan; Bai, Hui-Ping; Wang, Qian; Wang, Xiu-Li; Zhang, Ying-Ze

    2014-08-01

    This study aimed to investigate the effect of baclofen, a γ-aminobutyric acid B (GABAB) receptor agonist, on the expression of p-CREB and NR2B in the spinal dorsal horn of rats with diabetic neuropathic pain (DNP). The DNP rats, which were successfully induced with streptozocin, were distributed among 3 groups that were treated with saline (D1 group), baclofen (D2 group), or CGP55845 + baclofen (D3 group) continuously for 4 days. The rats induced with saline and subsequently treated with saline were used as controls (C group). The times for the paw withdrawal threshold and thermal withdrawal latency of the D1 group were lower than those for the C group, and were significantly increased after baclofen treatment, but not when GABA receptor was pre-blocked with CGP55845 (D3 group). Increased protein expression levels of NR2B and p-CREB and mRNA levels of NR2B were found in the D1 group when compared with the controls. Baclofen treatment significantly suppressed their expression, bringing it close to the levels of controls. However, in the D3 group, the expression of p-CREB and NR2B were still significantly higher than that of the controls. Activation of GABAB receptor by baclofen attenuates diabetic neuropathic pain, which may partly be accomplished via down-regulating the expression of p-CREB and NR2B.

  13. Glucagon-like peptide-1 receptor agonist inhibits asymmetric dimethylarginine generation in the kidney of streptozotocin-induced diabetic rats by blocking advanced glycation end product-induced protein arginine methyltranferase-1 expression.

    PubMed

    Ojima, Ayako; Ishibashi, Yuji; Matsui, Takanori; Maeda, Sayaka; Nishino, Yuri; Takeuchi, Masayoshi; Fukami, Kei; Yamagishi, Sho-ichi

    2013-01-01

    Advanced glycation end products (AGEs) and their receptor (RAGE) play a role in diabetic nephropathy. Asymmetric dimethylarginine (ADMA), an endogenous inhibitor of nitric oxide synthase, contributes to diabetic nephropathy. We have found that glucagon-like peptide-1 (GLP-1) inhibits the AGE-induced inflammatory reactions in endothelial cells. However, effects of GLP-1 on the AGE-RAGE-ADMA axis are unknown. This study examined the effects of GLP-1 on reactive oxygen species (ROS) generation, gene expression of protein arginine methyltransfetase-1 (PRMT-1), an enzyme that mainly generates ADMA, and ADMA levels in human proximal tubular cells. Streptozotocin-induced diabetic rats received continuous i.p. infusion of 0.3 μg of vehicle or 1.5 μg of the GLP-1 analog exendin-4 per kilogram of body weight for 2 weeks. We further investigated whether and how exendin-4 treatment reduced ADMA levels and renal damage in streptozotocin-induced diabetic rats. GLP-1 inhibited the AGE-induced RAGE and PRMT-1 gene expression, ROS, and ADMA generation in tubular cells, which were blocked by small-interfering RNAs raised against GLP-1 receptor. Exendin-4 treatment decreased gene expression of Rage, Prmt-1, Icam-1, and Mcp-1 and ADMA level; reduced urinary excretions of 8-hydroxy-2'-deoxyguanosine and albumin; and improved histopathologic changes of the kidney in diabetic rats. Our present study suggests that GLP-1 receptor agonist may inhibit the AGE-RAGE-mediated ADMA generation by suppressing PRMT-1 expression via inhibition of ROS generation, thereby protecting against the development and progression of diabetic nephropathy.

  14. Thymol, a dietary monoterpene phenol abrogates mitochondrial dysfunction in β-adrenergic agonist induced myocardial infarcted rats by inhibiting oxidative stress.

    PubMed

    Nagoor Meeran, M F; Jagadeesh, G S; Selvaraj, P

    2016-01-25

    Mitochondrial dysfunction has been suggested to be one of the important pathological events in isoproterenol (ISO), a synthetic catecholamine and β-adrenergic agonist induced myocardial infarction (MI). In this context, we have evaluated the impact of thymol against ISO induced oxidative stress and calcium uniporter malfunction involved in the pathology of mitochondrial dysfunction in rats. Male albino Wistar rats were pre and co-treated with thymol (7.5 mg/kg body weight) daily for 7 days. Isoproterenol (100 mg/kg body weight) was subcutaneously injected into rats on 6th and 7th day to induce MI. To explore the extent of cardiac mitochondrial damage, the activities/levels of cardiac marker enzymes, mitochondrial lipid peroxidation products, antioxidants, lipids, calcium, adenosine triphosphate and multi marker enzymes were evaluated. Isoproterenol induced myocardial infarcted rats showed a significant increase in the activities of cardiac diagnostic markers, heart mitochondrial lipid peroxidation, lipids, calcium, and a significant decrease in the activities/levels of heart mitochondrial superoxide dismutase, catalase, glutathione peroxidase, reduced glutathione, isocitrate, malate, α-ketoglutarate and NADH-dehydrogenases, cytochrome-C-oxidase, and adenosine triphosphate. Thymol pre and co-treatment showed near normalized effects on all the biochemical parameters studied. Transmission electron microscopic findings and mitochondrial swelling studies confirmed our biochemical findings. The in vitro study also revealed the potent free-radical scavenging activity of thymol. Thus, thymol attenuates the involvement of ISO against oxidative stress and calcium uniporter malfunction associated with mitochondrial dysfunction in rats.

  15. The PPARδ agonist GW0742 inhibits neuroinflammation, but does not restore neurogenesis or prevent early delayed hippocampal-dependent cognitive impairment after whole-brain irradiation.

    PubMed

    Schnegg, Caroline I; Greene-Schloesser, Dana; Kooshki, Mitra; Payne, Valerie S; Hsu, Fang-Chi; Robbins, Mike E

    2013-08-01

    Brain tumor patients often develop cognitive impairment months to years after partial or fractionated whole-brain irradiation (WBI). Studies suggest that neuroinflammation and decreased hippocampal neurogenesis contribute to the pathogenesis of radiation-induced brain injury. In this study, we determined if the peroxisomal proliferator-activated receptor (PPAR) δ agonist GW0742 can prevent radiation-induced brain injury in C57Bl/6 wild-type (WT) and PPARδ knockout (KO) mice. Dietary GW0742 prevented the acute increase in IL-1β mRNA and ERK phosphorylation measured at 3h after a single 10-Gy dose of WBI; it also prevented the increase in the number of activated hippocampal microglia 1 week after WBI. In contrast, dietary GW074 failed to prevent the radiation-induced decrease in hippocampal neurogenesis determined 2 months after WBI in WT mice or to mitigate their hippocampal-dependent spatial memory impairment measured 3 months after WBI using the Barnes maze task. PPARδ KO mice exhibited defects including decreased numbers of astrocytes in the dentate gyrus/hilus of the hippocampus and a failure to exhibit a radiation-induced increase in activated hippocampal microglia. Interestingly, the number of astrocytes in the dentate gyrus/hilus was reduced in WT mice, but not in PPARδ KO mice 2 months after WBI. These results demonstrate that, although dietary GW0742 prevents the increase in inflammatory markers and hippocampal microglial activation in WT mice after WBI, it does not restore hippocampal neurogenesis or prevent early delayed hippocampal-dependent cognitive impairment after WBI. Thus, the exact relationship between radiation-induced neuroinflammation, neurogenesis, and cognitive impairment remains elusive. Copyright © 2013 Elsevier Inc. All rights reserved.

  16. Rosiglitazone, a Peroxisome Proliferator-Activated Receptor (PPAR)-γ Agonist, Attenuates Inflammation Via NF-κB Inhibition in Lipopolysaccharide-Induced Peritonitis.

    PubMed

    Zhang, Yun-Fang; Zou, Xun-Liang; Wu, Jun; Yu, Xue-Qing; Yang, Xiao

    2015-12-01

    We assessed the anti-inflammatory effect of peroxisome proliferator-activated receptor (PPAR)-γ agonist, rosiglitazone, in a lipopolysaccharide (LPS)-induced peritonitis rat model. LPS was intraperitoneally injected into rats to establish peritonitis model. Male Sprague-Dawley (SD) rats were assigned to normal saline (the solvent of LPS), LPS, rosiglitazone plus LPS, and rosiglitazone alone. A simple peritoneal equilibrium test was performed with 20 ml 4.25 % peritoneal dialysis fluid. We measured the leukocyte count in dialysate and ultrafiltration volume. Peritoneal membrane histochemical staining was performed, and peritoneal thickness was assessed. CD40 and intercellular adhesion molecule-1 messenger RNA (ICAM-1 mRNA) levels in rat visceral peritoneum were detected by reverse transcription (RT)-PCR. IL-6 in rat peritoneal dialysis effluent was measured using enzyme-linked immunosorbent assay. The phosphorylation of NF-κB-p65 and IκBα was analyzed by Western blot. LPS administration resulted in increased peritoneal thickness and decreased ultrafiltration volume. Rosiglitazone pretreatment significantly decreased peritoneal thickness. In addition to CD40 and ICAM-1 mRNA expression, the IL-6, p-p65, and p-IκBα protein expressions were enhanced in LPS-administered animals. Rosiglitazone pretreatment significantly decreased ICAM-1 mRNA upregulation, secretion of IL-6 protein, and phosphorylation of NF-κB-p65 and IκBα without decreasing CD40 mRNA expression. Rosiglitazone has a protective effect in peritonitis, simultaneously decreasing NF-κB phosphorylation, suggesting that NF-κB signaling pathway mediated peritoneal inflammation induced by LPS. PPAR-γ might be considered a potential therapeutic target against peritonitis.

  17. Peroxisome proliferator-activated receptor-gamma-independent inhibition of macrophage activation by the non-thiazolidinedione agonist L-796,449. Comparison with the effects of 15-deoxy-delta(12,14)-prostaglandin J(2).

    PubMed

    Castrillo, A; Mojena, M; Hortelano, S; Boscá, L

    2001-09-07

    The effects of L-796,449 (3-chloro-4-(3-(3-phenyl-7-propylbenzofuran-6-yloxy)propylthio)phenylacetic acid; referred to henceforth as compound G), a thiazolidinedione-unrelated peroxisome proliferator activated-receptor-gamma (PPAR-gamma) agonist, on early signaling in lipopolysaccharide-activated RAW 264.7 macrophages were analyzed and compared with those elicited by 15-deoxy-Delta(12,14)-prostaglandin J(2) and the thiazolidinedione rosiglitazone. Compound G inhibited the activation of nuclear factor kappa B through the impairment of the targeting and degradation of I kappa B proteins and promoted a redistribution of I kappa B alpha and I kappa B beta in the nucleus of activated cells. Compound G inhibited I kappa B kinase (IKK) activity both in vivo and in vitro, suggesting a direct mechanism of interaction between this molecule and the IKK complex. The effect of compound G on IKK activity was independent of PPAR-gamma engagement because RAW 264.7 cells expressed negligible levels of this nuclear receptor, and rosiglitazone failed to mimic these actions. Moreover, treatment of activated macrophages with compound G enhanced the synthesis of superoxide anion, which, in combination with the NO produced under activation conditions, triggered apoptosis through the intracellular synthesis of peroxynitrite. These results suggest that compound G might contribute to the resolution of inflammation by favoring the induction of apoptosis through mechanisms independent of PPAR-gamma engagement.

  18. β2 Agonists.

    PubMed

    Billington, Charlotte K; Penn, Raymond B; Hall, Ian P

    2017-01-01

    History suggests β agonists, the cognate ligand of the β2 adrenoceptor, have been used as bronchodilators for around 5,000 years, and β agonists remain today the frontline treatment for asthma and chronic obstructive pulmonary disease (COPD). The β agonists used clinically today are the products of significant expenditure and over 100 year's intensive research aimed at minimizing side effects and enhancing therapeutic usefulness. The respiratory physician now has a therapeutic toolbox of long acting β agonists to prophylactically manage bronchoconstriction, and short acting β agonists to relieve acute exacerbations. Despite constituting the cornerstone of asthma and COPD therapy, these drugs are not perfect; significant safety issues have led to a black box warning advising that long acting β agonists should not be used alone in patients with asthma. In addition there are a significant proportion of patients whose asthma remains uncontrolled. In this chapter we discuss the evolution of β agonist use and how the understanding of β agonist actions on their principal target tissue, airway smooth muscle, has led to greater understanding of how these drugs can be further modified and improved in the future. Research into the genetics of the β2 adrenoceptor will also be discussed, as will the implications of individual DNA profiles on the clinical outcomes of β agonist use (pharmacogenetics). Finally we comment on what the future may hold for the use of β agonists in respiratory disease.

  19. A peroxisome proliferator-activated receptor-gamma agonist and the p53 rescue drug CP-31398 inhibit the spontaneous immortalization of breast epithelial cells.

    PubMed

    Herbert, Brittney-Shea; Pearce, Virginia P; Hynan, Linda S; LaRue, Denise M; Wright, Woodring E; Kopelovich, Levy; Shay, Jerry W

    2003-04-15

    Cell immortalization is a critical and rate-limiting step in cancer progression. Agents that inhibit cell immortalization may have utility for novel molecular chemopreventive strategies. Preimmortal breast epithelial cells derived from a patient with the Li-Fraumeni Syndrome (LFS) can spontaneously immortalize in vitro at a measurable and reproducible frequency. In the present study, these cells were treated in vitro with low (nM) concentrations of potential and otherwise clinically validated chemopreventive agents, including several nonsteroidal anti-inflammatory drugs, rosiglitazone maleate, and the p53 rescue drug CP-31398. Rosiglitazone maleate (P < 0.05) and CP-31398 (P < 0.05) significantly inhibited the frequency of spontaneous immortalization of LFS breast epithelial cells compared with untreated controls. Nonsteroidal anti-inflammatory drugs, including specific cyclooxengenase-2 inhibitors, only moderately inhibited the spontaneous immortalization of preimmortal LFS breast epithelial cells. The significant effects of the p53 rescue drug CP-31398 correlated with the increase in cellular death induced by telomere shortening-induced DNA damage signals, including increases in p53 and p21 protein levels. Because immortalization is one step in cancer progression, these studies show the potential usefulness of a cell-based model system to screen the effects of known and potentially novel chemopreventive agents, using cell immortalization as an end point.

  20. Gonadotropin-releasing hormone (GnRH) agonist triptorelin inhibits estradiol-induced serum response element (SRE) activation and c-fos expression in human endometrial, ovarian and breast cancer cells.

    PubMed

    Gründker, Carsten; Günthert, Andreas R; Hellriegel, Martin; Emons, Günter

    2004-11-01

    The majority of human endometrial (>80%), ovarian (>80%) and breast (>50%) cancers express GnRH receptors. Their spontaneous and epidermal growth-factor-induced proliferation is dose- and time-dependently reduced by treatment with GnRH and its agonists. In this study, we demonstrate that the GnRH agonist triptorelin inhibits estradiol (E2)-induced cancer cell proliferation. The proliferation of quiescent estrogen receptor alpha (ER alpha)-/ER beta-positive, but not of ER alpha-negative/ER beta-positive endometrial, ovarian and breast cancer cell lines, was significantly stimulated (P<0.001) (ANOVA) after treatment with E2 (10(-8) M). This effect was time- and dose-dependently antagonized by simultaneous treatment with triptorelin. The inhibitory effect was maximal at 10(-5) M concentration of triptorelin (P<0.001). In addition, we could show that, in ER alpha-/ER beta-positive cell lines, E2 induces activation of serum response element (SRE) and expression of the immediate early-response gene c-fos. These effects were blocked by triptorelin (P<0.001). E2-induced activation of estrogen-response element (ERE) was not affected by triptorelin. The transcriptional activation of SRE by E2 is due to ER alpha activation of the mitogen-activated protein kinase (MAPK) pathway. This pathway is impeded by GnRH, resulting in a reduction of E2-induced SRE activation and, in consequence, a reduction of E2-induced c-fos expression. This causes downregulation of E2-induced cancer cell proliferation.

  1. Dihydropyridine Ca/sup + +/ channel agonists enhance /sup 45/Ca/sup + +/ uptake by rabbit aortic smooth muscle cells (RASMC) at 15-50 mM K/sup +/. Nifedipine and D-600 inhibit such effects

    SciTech Connect

    Papaioannou, S.; Knodle, S.

    1986-03-01

    RASMC were prepared, subcultured to passage number22 and characterized morphologically and for /sup 45/Ca/sup + +/ uptake. The initial rate of /sup 45/Ca/sup + +/ uptake in 50 mM K/sup +/ was three times the rate in 5 mM K/sup +/. Steady state /sup 45/Ca/sup + +/ uptake increased with K/sup +/ concentration in a dose-dependent manner. Threshold was at approx. 15 mM K/sup +/. At 25-50 mM K/sup +/ the maximum K/sup +/-induced Ca/sup + +/ uptake was 1.3 nmol Ca/sup + +//mg protein or 0.6 nmol Ca/sup + +//10/sup 6/ cells. The three dihydropyridine agonists (+/-) Bay K 8644, (+/-) CGP 28392 and (+) 202-791 enhanced the /sup 45/Ca/sup + +/ uptake at K/sup +/ greater than or equal to 15 nM. At the /sup 45/Ca/sup + +/ uptake threshold of 15 mM each agonist potentiated /sup 45/Ca/sup + +/ uptake in a dose-dependent manner. Responses were antagonized competitively by nife-dipine and non-competitively by (+/-) D-600. The (-) 202-791 inhibited K/sup +/-induced /sup 45/Ca/sup + +/ uptake (IC/sub 50/ = 4.0 x 10/sup -9/ M). Based on the agreement with literature data on contraction, electrophysiological and /sup 45/Ca/sup + +/ uptake studies using intact vascular tissues, it is concluded that the RASMC possess voltage-dependent Ca/sup + +/ channels functionally similar to intact vascular muscle. These cells are a suitable model system for pharmacological studies of Ca/sup + +/ channels and for characterization of Ca/sup + +/ channel modulators.

  2. AdipoRon, an adiponectin receptor agonist, attenuates PDGF-induced VSMC proliferation through inhibition of mTOR signaling independent of AMPK: Implications toward suppression of neointimal hyperplasia.

    PubMed

    Fairaq, Arwa; Shawky, Noha M; Osman, Islam; Pichavaram, Prahalathan; Segar, Lakshman

    2017-02-22

    Hypoadiponectinemia is associated with an increased risk of coronary artery disease. Although adiponectin replenishment mitigates neointimal hyperplasia and atherosclerosis in mouse models, adiponectin therapy has been hampered in a clinical setting due to its large molecular size. Recent studies demonstrate that AdipoRon (a small-molecule adiponectin receptor agonist) improves glycemic control in type 2 diabetic mice and attenuates postischemic cardiac injury in adiponectin-deficient mice, in part, through activation of AMP-activated protein kinase (AMPK). To date, it remains unknown as to whether AdipoRon regulates vascular smooth muscle cell (VSMC) proliferation, which plays a major role in neointima formation. In the present study, oral administration of AdipoRon (50mg/kg) in C57BL/6J mice significantly diminished arterial injury-induced neointima formation by ∼57%. Under in vitro conditions, AdipoRon treatment led to significant inhibition of platelet-derived growth factor (PDGF)-induced VSMC proliferation, DNA synthesis, and cyclin D1 expression. While AdipoRon induced a rapid and sustained activation of AMPK, it also diminished basal and PDGF-induced phosphorylation of mTOR and its downstream targets, including p70S6K/S6 and 4E-BP1. However, siRNA-mediated AMPK downregulation showed persistent inhibition of p70S6K/S6 and 4E-BP1 phosphorylation, indicating AMPK-independent effects for AdipoRon inhibition of mTOR signaling. In addition, AdipoRon treatment resulted in a sustained and transient decrease in PDGF-induced phosphorylation of Akt and ERK, respectively. Furthermore, PDGF receptor-β tyrosine phosphorylation, which controls the phosphorylation state of Akt and ERK, was diminished upon AdipoRon treatment. Together, the present findings suggest that orally-administered AdipoRon has the potential to limit restenosis after angioplasty by targeting mTOR signaling independent of AMPK activation.

  3. PPAR{alpha} agonist fenofibrate protects the kidney from hypertensive injury in spontaneously hypertensive rats via inhibition of oxidative stress and MAPK activity

    SciTech Connect

    Hou, Xiaoyang; Shen, Ying H.; Li, Chuanbao; Wang, Fei; Zhang, Cheng; Bu, Peili; Zhang, Yun

    2010-04-09

    Oxidative stress has been shown to play an important role in the development of hypertensive renal injury. Peroxisome proliferator-activated receptors {alpha} (PPAR{alpha}) has antioxidant effect. In this study, we demonstrated that fenofibrate significantly reduced proteinuria, inflammatory cell recruitment and extracellular matrix (ECM) proteins deposition in the kidney of SHRs without apparent effect on blood pressure. To investigate the mechanisms involved, we found that fenofibrate treatment markedly reduced oxidative stress accompanied by reduced activity of renal NAD(P)H oxidase, increased activity of Cu/Zn SOD, and decreased phosphorylation of p38MAPK and JNK in the kidney of SHRs. Taken together, fenofibrate treatment can protect against hypertensive renal injury without affecting blood pressure by inhibiting inflammation and fibrosis via suppression of oxidative stress and MAPK activity.

  4. Poly (I:C), an agonist of toll-like receptor-3, inhibits replication of the Chikungunya virus in BEAS-2B cells

    PubMed Central

    2012-01-01

    Background Double-stranded RNA (dsRNA) and its mimic, polyinosinic acid: polycytidylic acid [Poly (I:C)], are recognized by toll-like receptor 3 (TLR3) and induce interferon (IFN)-β in many cell types. Poly (I:C) is the most potent IFN inducer. In in vivo mouse studies, intraperitoneal injection of Poly (I:C) elicited IFN-α/β production and natural killer (NK) cells activation. The TLR3 pathway is suggested to contribute to innate immune responses against many viruses, including influenza virus, respiratory syncytial virus, herpes simplex virus 2, and murine cytomegalovirus. In Chikungunya virus (CHIKV) infection, the viruses are cleared within 7–10 days postinfection before adaptive immune responses emerge. The innate immune response is important for CHIKV clearance. Results The effects of Poly (I:C) on the replication of CHIKV in human bronchial epithelial cells, BEAS-2B, were studied. Poly (I:C) suppressed cytopathic effects (CPE) induced by CHIKV infection in BEAS-2B cells in the presence of Poly (I:C) and inhibited the replication of CHIKV in the cells. The virus titers of Poly (I:C)-treated cells were much lower compared with those of untreated cells. CHIKV infection and Poly (I:C) treatment of BEAS-2B cells induced the production of IFN-β and increased the expression of anti-viral genes, including IFN-α, IFN-β, MxA, and OAS. Both Poly (I:C) and CHIKV infection upregulate the expression of TLR3 in BEAS-2B cells. Conclusions CHIKV is sensitive to innate immune response induced by Poly (I:C). The inhibition of CHIKV replication by Poly (I:C) may be through the induction of TLR3, which triggers the production of IFNs and other anti-viral genes. The innate immune response is important to clear CHIKV in infected cells. PMID:22698190

  5. Abrogation of CC chemokine receptor 9 ameliorates ventricular remodeling in mice after myocardial infarction

    PubMed Central

    Huang, Yan; Wang, Dandan; Wang, Xin; Zhang, Yijie; Liu, Tao; Chen, Yuting; Tang, Yanhong; Wang, Teng; Hu, Dan; Huang, Congxin

    2016-01-01

    CC chemokine receptor 9 (CCR9), which is a unique receptor for CC chemokine ligand (CCL25), is mainly expressed on lymphocytes, dendritic cells (DCs) and monocytes/macrophages. CCR9 mediates the chemotaxis of inflammatory cells and participates in the pathological progression of inflammatory diseases. However, the role of CCR9 in the pathological process of myocardial infarction (MI) remains unexplored; inflammation plays a key role in this process. Here, we used CCR9 knockout mice to determine the functional significance of CCR9 in regulating post-MI cardiac remodeling and its underlying mechanism. MI was induced by surgical ligation of the left anterior descending coronary artery in CCR9 knockout mice and their CCR9+/+ littermates. Our results showed that the CCR9 expression levels were up-regulated in the hearts of the MI mice. Abrogation of CCR9 improved the post-MI survival rate and left ventricular (LV) dysfunction and decreased the infarct size. In addition, the CCR9 knockout mice exhibited attenuated inflammation, apoptosis, structural and electrical remodeling compared with the CCR9+/+ MI mice. Mechanistically, CCR9 mainly regulated the pathological response by interfering with the NF-κB and MAPK signaling pathways. In conclusion, the data reveal that CCR9 serves as a novel modulator of pathological progression following MI through NF-κB and MAPK signaling. PMID:27585634

  6. Alpha 2-adrenoceptor agonist-mediated inhibition of [3H]noradrenaline release from rat hippocampus is reduced by 4-aminopyridine, but that caused by an adenosine analogue or omega-conotoxin is not.

    PubMed

    Hu, P S; Fredholm, B B

    1989-07-01

    The inhibitory effect of an adenosine analogue, R-PIA, and an alpha 2-adrenoceptor agonist, UK 14,304, on [3H]NA efflux from field-stimulated rat hippocampal slices was examined. The effect of 0.1 microM UK 14,304 was mimicked by 30 nM omega-conotoxin and by 10 microM cadmium chloride, inhibitors of N- and L-type Ca2+ channels. R-PIA (1 microM) had no effect per se, but caused a clear-cut inhibition after blockade of the pre-synaptic alpha 2-receptor by yohimbine. 4-Aminopyridine (4-AP) caused a dose-dependent increase in evoked transmitter release. At 30 microM 4-AP did not affect the actions of omega-conotoxin or cadmium chloride. The pre-synaptic effect of R-PIA was similarly unaffected by 30 microM 4-AP. The pre-synaptic effect of UK 14,304 was virtually abolished by 4-AP (30 microM). The effect of UK 14,304 (0.1 microM) could be partly restored by reducing the Ca2+ concentration during treatment with 4-AP (22% inhibition compared to 42% with normal Ca2+). The magnitude of increase in evoked [3H]NA efflux by yohimbine (1 microM) was decreased by 4-AP in a concentration-dependent manner from 142% increase in controls to 21% at 100 microM 4-AP. The present results indicate that NA release is reduced by somewhat different mechanisms by pre-synaptic alpha 2- and adenosine A1-receptors. Furthermore, the results indicate that pre-synaptic A1-receptors on hippocampal NA neurons do not primarily regulate 4-AP-dependent potassium channels, but they might act directly on a Ca2+ conductance.

  7. Innate Immunity Stimulation via Toll-Like Receptor 9 Ameliorates Vascular Amyloid Pathology in Tg-SwDI Mice with Associated Cognitive Benefits.

    PubMed

    Scholtzova, Henrieta; Do, Eileen; Dhakal, Shleshma; Sun, Yanjie; Liu, Shan; Mehta, Pankaj D; Wisniewski, Thomas

    2017-01-25

    Alzheimer's disease (AD) is characterized by the presence of parenchymal amyloid-β (Aβ) plaques, cerebral amyloid angiopathy (CAA) and neurofibrillary tangles. Currently there are no effective treatments for AD. Immunotherapeutic approaches under development are hampered by complications related to ineffectual clearance of CAA. Genome-wide association studies have demonstrated the importance of microglia in AD pathogenesis. Microglia are the primary innate immune cells of the brain. Depending on their activation state and environment, microglia can be beneficial or detrimental. In our prior work, we showed that stimulation of innate immunity with Toll-like receptor 9 agonist, class B CpG (cytosine-phosphate-guanine) oligodeoxynucleotides (ODNs), can reduce amyloid and tau pathologies without causing toxicity in Tg2576 and 3xTg-AD mouse models. However, these transgenic mice have relatively little CAA. In the current study, we evaluated the therapeutic profile of CpG ODN in a triple transgenic mouse model, Tg-SwDI, with abundant vascular amyloid, in association with low levels of parenchymal amyloid deposits. Peripheral administration of CpG ODN, both before and after the development of CAA, negated short-term memory deficits, as assessed by object-recognition tests, and was effective at improving spatial and working memory evaluated using a radial arm maze. These findings were associated with significant reductions of CAA pathology lacking adverse effects. Together, our extensive evidence suggests that this innovative immunomodulation may be a safe approach to ameliorate all hallmarks of AD pathology, supporting the potential clinical applicability of CpG ODN. Recent genetic studies have underscored the emerging role of microglia in Alzheimer's disease (AD) pathogenesis. Microglia lose their amyloid-β-clearing capabilities with age and as AD progresses. Therefore, the ability to modulate microglia profiles offers a promising therapeutic avenue for reducing AD

  8. Chronic exposure to indoxacarb and pulmonary expression of toll-like receptor-9 in mice

    PubMed Central

    Kaur, Sandeep; Mukhopadhyay, C. S.; Sethi, R. S.

    2016-01-01

    Aim: Chronic exposure to indoxacarb and pulmonary expression of toll-like receptor 9 (TLR-9) in mice. Materials and Methods: In this study, healthy male Swiss albino mice (n=30) aging 8-10 weeks were used to evaluate TLR-9 expression in lungs of mice following indoxacarb exposure with and without lipopolysaccharide (LPS). Indoxacarb was administered orally dissolved in groundnut oil at 4 and 2 mg/kg/day for 90 days. On day 91, five animals from each group were challenged with LPS/normal saline solution at 80 µg/animal. The lung tissues were processed for real time and immunohistochemical studies. Results: LPS resulted increase in fold change m-RNA expression level of TLR-9 as compare to control, while indoxacarb (4 mg/kg) alone and in combination with LPS resulted 16.21-fold change and 29.4-fold change increase in expression of TLR-9 m-RNA, respectively, as compared to control. Similarly, indoxacarb (2 mg/kg) alone or in combination with LPS also altered TLR-9 expression. Further at protein level control group showed minimal expression of TLR-9 in lungs as compare to other groups, however, LPS group showed intense positive staining in bronchial epithelium as well as in alveolar septal cells. Indoxacarb at both doses individually showed strong immuno-positive reaction as compare to control, however when combined with LPS resulted intense staining in airway epithelium as compare to control. Conclusion: Chronic oral administration of indoxacarb for 90 days (4 and 2 mg/kg) alters expression of TLR-9 at m-RNA and protein level and co-exposure with LPS exhibited synergistic effect. PMID:27956782

  9. Structural basis of CpG and inhibitory DNA recognition by Toll-like receptor 9.

    PubMed

    Ohto, Umeharu; Shibata, Takuma; Tanji, Hiromi; Ishida, Hanako; Krayukhina, Elena; Uchiyama, Susumu; Miyake, Kensuke; Shimizu, Toshiyuki

    2015-04-30

    Innate immunity serves as the first line of defence against invading pathogens such as bacteria and viruses. Toll-like receptors (TLRs) are examples of innate immune receptors, which sense specific molecular patterns from pathogens and activate immune responses. TLR9 recognizes bacterial and viral DNA containing the cytosine-phosphate-guanine (CpG) dideoxynucleotide motif. The molecular basis by which CpG-containing DNA (CpG-DNA) elicits immunostimulatory activity via TLR9 remains to be elucidated. Here we show the crystal structures of three forms of TLR9: unliganded, bound to agonistic CpG-DNA, and bound to inhibitory DNA (iDNA). Agonistic-CpG-DNA-bound TLR9 formed a symmetric TLR9-CpG-DNA complex with 2:2 stoichiometry, whereas iDNA-bound TLR9 was a monomer. CpG-DNA was recognized by both protomers in the dimer, in particular by the amino-terminal fragment (LRRNT-LRR10) from one protomer and the carboxy-terminal fragment (LRR20-LRR22) from the other. The iDNA, which formed a stem-loop structure suitable for binding by intramolecular base pairing, bound to the concave surface from LRR2-LRR10. This structure serves as an important basis for improving our understanding of the functional mechanisms of TLR9.

  10. A review of the role of CpG oligodeoxynucleotides as toll-like receptor 9 agonists in prophylactic and therapeutic vaccine development in infectious diseases.

    PubMed

    Gupta, Kaveri; Cooper, Curtis

    2008-01-01

    This article reviews the biology of Toll-like receptors, the current understanding of the mechanism by which CpG oligodeoxynucleotides (ODNs) perturb immune function and the published literature describing their evaluation in the development of vaccines in humans. The role of these molecules as immune modulators in HCV treatment is also considered. There has been considerable research evaluating the role of CpG ODNs as an adjuvant and immune modulator in hepatitis B, hepatitis C and influenza. The safety and immunogenicity of the 1018 ISS compound in combination with Engerix-B was assessed in 99 healthy, adult seronegative volunteers. One month following the first immunization dose, 78.7% in the rHBsAg plus 1018 ISS group versus 11.8% in the Engerix-B group achieved protective titres. One hundred percent of rHBsAg plus 1018 ISS and 18.0% of hepatitis B vaccine-alone recipients were seroprotected 1 week following the second dose of study vaccine. After all doses of vaccine had been administered, seroprotection rates were 100% and 64%, respectively (p < 0.001). CPG 7909 was co-administered with Engerix-B in 56 healthy adults. After the second injection (week 6 time point), seroprotection was achieved in 100% of CPG 7909 recipients (0.5 mg 13/13; 1.0 mg 12/12; 0.125 mg 12/12) compared with 55% (6/11) of control participants (p = 0.0003). Twelve months post prime, all subjects who had received the full course of vaccination maintained seroprotective anti-HBs titres. The safety and immunogenicity of Engerix B plus CPG 7909 was assessed in HIV seropositive patients. All CPG 7909 recipients (n = 19) and 17/19 (89%) control subjects achieved seroprotection by 2 weeks after the third and final injection (10 weeks). Seroprotective titres remained in all CPG 7909 recipients at 48 weeks (100%) versus 12/19 (63%) for controls (p = 0.008). This cohort of HIV-infected patients was followed at 6-month intervals for up to 60 months after enrolment. The difference in seroprotection (> or =10 mIU/L) and GMT between study arms remained significant (p < 0.05) at all time points from month 24 to month 60. There is great potential for CpG ODN as vaccine adjuvants and as therapeutic immune modulators. The use of these molecules as a hepatitis B vaccine adjuvant is most promising.

  11. Serotonergic agonists behave as partial agonists at the dopamine D2 receptor.

    PubMed

    Rinken, A; Ferré, S; Terasmaa, A; Owman, C; Fuxe, K

    1999-02-25

    RAT dopamine D2short receptors expressed in CHO cells were characterized by activation of [35S]GTPgammaS binding. There were no significant differences between the maximal effects seen in activation of [35S]GTPgammaS binding caused by dopaminergic agonists, but the effects of 5-HT, 8OH-DPAT and 5-methoxytryptamine amounted to 47 +/- 7%, 43 +/- 5% and 70 +/- 7% of the dopamine effect, respectively. The dopaminergic antagonist (+)butaclamol inhibited activations of both types of ligands with equal potency (pA2 = 8.9 +/- 0.1), indicating that only one type of receptor is involved. In competition with [3H]raclopride binding, dopaminergic agonists showed 53 +/- 2% of the binding sites in the GTP-dependent high-affinity state, whereas 5-HT showed only 20 +/- 3%. Taken together, the results indicate that serotonergic agonists behave as typical partial agonists for D2 receptors with potential antiparkinsonian activity.

  12. Toll-like receptor 9 prevents cardiac rupture after myocardial infarction in mice independently of inflammation.

    PubMed

    Omiya, Shigemiki; Omori, Yosuke; Taneike, Manabu; Protti, Andrea; Yamaguchi, Osamu; Akira, Shizuo; Shah, Ajay M; Nishida, Kazuhiko; Otsu, Kinya

    2016-12-01

    We have reported that the Toll-like receptor 9 (TLR9) signaling pathway plays an important role in the development of pressure overload-induced inflammatory responses and heart failure. However, its role in cardiac remodeling after myocardial infarction has not been elucidated. TLR9-deficient and control C57Bl/6 wild-type mice were subjected to left coronary artery ligation. The survival rate 14 days postoperation was significantly lower in TLR9-deficient mice than that in wild-type mice with evidence of cardiac rupture in all dead mice. Cardiac magnetic resonance imaging showed no difference in infarct size and left ventricular wall thickness and function between TLR9-deficient and wild-type mice. There were no differences in the number of infiltrating inflammatory cells and the levels of inflammatory cytokine mRNA in infarct hearts between TLR9-deficient and wild-type mice. The number of α-smooth muscle actin (αSMA)-positive myofibroblasts and αSMA/Ki67-double-positive proliferative myofibroblasts was increased in the infarct and border areas in infarct hearts compared with those in sham-operated hearts in wild-type mice, but not in TLR9-deficient mice. The class B CpG oligonucleotide increased the phosphorylation level of NF-κB and the number of αSMA-positive and αSMA/Ki67-double-positive cells and these increases were attenuated by BAY1-7082, an NF-κB inhibitor, in cardiac fibroblasts isolated from wild-type hearts. The CpG oligonucleotide showed no effect on NF-κB activation or the number of αSMA-positive and αSMA/Ki67-double-positive cells in cardiac fibroblasts from TLR9-deficient hearts. Although the TLR9 signaling pathway is not involved in the acute inflammatory response in infarct hearts, it ameliorates cardiac rupture possibly by promoting proliferation and differentiation of cardiac fibroblasts. Copyright © 2016 the American Physiological Society.

  13. Toll-Like Receptor 9 Alternatively Spliced Isoform Negatively Regulates TLR9 Signaling in Teleost Fish

    PubMed Central

    Chen, Nai-Yu; Nagarajan, Govindarajulu; Chiou, Pinwen Peter

    2015-01-01

    Toll-like receptor 9 (TLR9) recognizes and binds unmethylated CpG motifs in DNA, which are found in the genomes of bacteria and DNA viruses. In fish, Tlr9 is highly diverse, with the number of introns ranging from 0 to 4. A fish Tlr9 gene containing two introns has been reported to express two alternatively spliced isoforms, namely gTLR9A (full-length) and gTLR9B (with a truncated Cʹ-terminal signal transducing domain), whose regulation and function remain unclear. Here, we report a unique regulatory mechanism of gTLR9 signaling in orange-spotted grouper (Epinephelus coioides), whose gTlr9 sequence also contains two introns. We demonstrated that the grouper gTlr9 gene indeed has the capacity to produce two gTLR9 isoforms via alternative RNA splicing. We found that gTLR9B could function as a negative regulator to suppress gTLR9 signaling as demonstrated by the suppression of downstream gene expression. Following stimulation with CpG oligodeoxynucleotide (ODN), gTLR9A and gTLR9B were observed to translocate into endosomes and co-localize with ODN and the adaptor protein gMyD88. Both gTLR9A and gTLR9B could interact with gMyD88; however, gTLR9B could not interact with downstream IRAK4 and TRAF6. Further analysis of the expression profile of gTlr9A and gTlr9B upon immune-stimulation revealed that the two isoforms were differentially regulated in a time-dependent manner. Overall, these data suggest that fish TLR9B functions as a negative regulator, and that its temporal expression is mediated by alternative RNA splicing. This has not been observed in mammalian TLR9s and might have been acquired relatively recently in the evolution of fish. PMID:25955250

  14. Downregulation of Toll-Like Receptor 9 Expression by Beta Human Papillomavirus 38 and Implications for Cell Cycle Control

    PubMed Central

    Pacini, Laura; Savini, Claudia; Ghittoni, Raffaella; Saidj, Djamel; Lamartine, Jerome; Hasan, Uzma A.; Accardi, Rosita

    2015-01-01

    ABSTRACT Innate immunity is the first line of host defense against infections. Many oncogenic viruses can deregulate several immune-related pathways to guarantee the persistence of the infection. Here, we show that the cutaneous human papillomavirus 38 (HPV38) E6 and E7 oncoproteins suppress the expression of the double-stranded DNA sensor Toll-like receptor 9 (TLR9) in human foreskin keratinocytes (HFK), a key mediator of the antiviral innate immune host response. In particular, HPV38 E7 induces TLR9 mRNA downregulation by promoting accumulation of ΔNp73α, an antagonist of p53 and p73. Inhibition of ΔNp73α expression by antisense oligonucleotide in HPV38 E6/E7 HFK strongly rescues mRNA levels of TLR9, highlighting a key role of ΔNp73α in this event. Chromatin immunoprecipitation experiments showed that ΔNp73α is part of a negative transcriptional regulatory complex with IκB kinase beta (IKKβ) that binds to a NF-κB responsive element within the TLR9 promoter. In addition, the Polycomb protein enhancer of zeste homolog 2 (EZH2), responsible for gene expression silencing, is also recruited into the complex, leading to histone 3 trimethylation at lysine 27 (H3K27me3) in the same region of the TLR9 promoter. Ectopic expression of TLR9 in HPV38 E6/E7 cells resulted in an accumulation of the cell cycle inhibitors p21WAF1 and p27Kip1, decreased CDK2-associated kinase activity, and inhibition of cellular proliferation. In summary, our data show that HPV38, similarly to other viruses with well-known oncogenic activity, can downregulate TLR9 expression. In addition, they highlight a new role for TLR9 in cell cycle regulation. IMPORTANCE The mucosal high-risk HPV types have been clearly associated with human carcinogenesis. Emerging lines of evidence suggest the involvement of certain cutaneous HPV types in development of skin squamous cell carcinoma, although this association is still under debate. Oncogenic viruses have evolved different strategies to hijack the

  15. Sustained activation of toll-like receptor 9 induces an invasive phenotype in lung fibroblasts: possible implications in idiopathic pulmonary fibrosis.

    PubMed

    Kirillov, Varvara; Siler, Jonathan T; Ramadass, Mahalakshmi; Ge, Lingyin; Davis, James; Grant, Geraldine; Nathan, Steven D; Jarai, Gabor; Trujillo, Glenda

    2015-04-01

    Idiopathic pulmonary fibrosis (IPF) is characterized by excessive scarring of the lung parenchyma, resulting in a steady decline of lung function and ultimately respiratory failure. The disease course of IPF is extremely variable, with some patients exhibiting stability of symptoms for prolonged periods of time, whereas others exhibit rapid progression and loss of lung function. Viral infections have been implicated in IPF and linked to disease severity; however, whether they directly contribute to progression is unclear. We previously classified patients as rapid and slow progressors on the basis of clinical features and expression of the pathogen recognition receptor, Toll-like receptor 9 (TLR9). Activation of TLR9 in vivo exacerbated IPF in mice and induced differentiation of myofibroblasts in vitro, but the mechanism of TLR9 up-regulation and progression of fibrosis are unknown. Herein, we investigate whether transforming growth factor (TGF)-β, a pleiotropic cytokine central to IPF pathogenesis, regulates TLR9 in lung myofibroblasts. Results showed induction of TLR9 expression by TGF-β in lung myofibroblasts and a distinct profibrotic myofibroblast phenotype driven by stimulation with the TLR9 agonist, CpG-DNA. Chronic TLR9 stimulation resulted in stably differentiated α-smooth muscle actin(+)/platelet-derived growth factor receptor α(+)/CD44(+)/matrix metalloproteinase-14(+)/matrix metalloproteinase-2(+) myofibroblasts, which secrete inflammatory cytokines, invade Matrigel toward platelet-derived growth factor, and resist hypoxia-induced apoptosis. These results suggest a mechanism by which TGF-β and TLR9 responses in myofibroblasts collaborate to drive rapid progression of IPF.

  16. A TLR9 agonist enhances the anti-tumor immunity of peptide and lipopeptide vaccines via different mechanisms

    PubMed Central

    Song, Ying-Chyi; Liu, Shih-Jen

    2015-01-01

    The toll-like receptor 9 (TLR9) agonists CpG oligodeoxynucleotides (CpG ODNs) have been recognized as promising adjuvants for vaccines against infectious diseases and cancer. However, the role of TLR9 signaling in the regulation of antigen uptake and presentation is not well understood. Therefore, to investigate the effects of TLR9 signaling, this study used synthetic peptides (IDG) and lipopeptides (lipoIDG), which are internalized by dendritic cells (DCs) via endocytosis-dependent and endocytosis-independent pathways, respectively. Our data demonstrated that the internalization of lipoIDG and IDG by bone marrow-derived dendritic cells (BMDCs) was not enhanced in the presence of CpG ODNs; however, CpG ODNs prolonged the co-localization of IDG with CpG ODNs in early endosomes. Surprisingly, CpG ODNs enhanced CD8+ T cell responses, and the anti-tumor effects of IDG immunization were stronger than those of lipoIDG immunization. LipoIDG admixed with CpG ODNs induced low levels of CD8+ T cells and partially inhibit tumor growth. Our findings suggest that CpG ODNs increase the retention of antigens in early endosomes, which is important for eliciting anti-tumor immunity. These results will facilitate the application of CpG adjuvants in the design of different vaccines. PMID:26215533

  17. Epithelial toll-like receptor 9 signaling in colorectal inflammation and cancer: Clinico-pathogenic aspects

    PubMed Central

    Fűri, István; Sipos, Ferenc; Germann, Tiana M; Kalmár, Alexandra; Tulassay, Zsolt; Molnár, Béla; Műzes, Györgyi

    2013-01-01

    Toll-like receptors (TLRs) recognize specific motifs which are frequently present in bacteria, fungi, prokaryotes and viruses. Amongst TLRs, TLR9 can be activated by such bacterial or viral DNA fragments, immunoglobulin-DNA complexes or synthetic oligonucleotides, which all contain unmethylated cytosine-guanine nucleotide sequences (CpGs). Emerging data indicate that TLR9 signaling has a role in, and may influence, colorectal carcinogenesis and colonic inflammation. CpGs are classified into three groups according to their influence on both the antigen-specific humoral- and cellular immunity, and the production of type 1 interferons and proinflammatory cytokines. TLR9 activation via CpGs may serve as a new therapeutic target for several cancerous and various inflammatory conditions. Due to its probable anti-cancer effects, the application possibilities of TLR9-signaling modulation may be extremely diverse even in colorectal tumors. In this review we aimed to summarize the current knowledge about TLR-signaling in the pathogenesis and therapy of inflammatory bowel diseases and colorectal cancer. Due to the species-specific differences in TLR9 expression, however, one must be careful in translating the animal model data into the human system, because of the differences between CpG-oligodeoxynucleotide-responsive cells. TLR9 agonist DNA-based immunomodulatory sequences could also represent a promising therapeutic alternative in systemic inflammatory conditions and chronic colonic inflammations as their side effects are not significant. PMID:23864774

  18. Toll-Like Receptor 9-Activation during Onset of Myocardial Ischemia Does Not Influence Infarct Extension

    PubMed Central

    Ohm, Ingrid Kristine; Gao, Erhe; Belland Olsen, Maria; Alfsnes, Katrine; Bliksøen, Marte; Øgaard, Jonas; Ranheim, Trine; Nymo, Ståle Haugset; Holmen, Yangchen Dhondup; Aukrust, Pål; Yndestad, Arne; Vinge, Leif Erik

    2014-01-01

    Aim Myocardial infarction (MI) remains a major cause of death and disability worldwide, despite available reperfusion therapies. Inflammatory signaling is considered nodal in defining final infarct size. Activation of the innate immune receptor toll-like receptors (TLR) 9 prior to ischemia and reperfusion (I/R) reduces infarct size, but the consequence of TLR9 activation timed to the onset of ischemia is not known. Methods and Results The TLR9-agonist; CpG B was injected i.p. in C57BL/6 mice immediately after induction of ischemia (30 minutes). Final infarct size, as well as area-at-risk, was measured after 24 hours of reperfusion. CpG B injection resulted in a significant increase in circulating granulocytes and monocytes both in sham and I/R mice. Paradoxically, clear evidence of reduced cardiac infiltration of both monocytes and granulocytes could be demonstrated in I/R mice treated with CpG B (immunocytochemistry, myeloperoxidase activity and mRNA expression patterns). In addition, systemic TLR9 activation elicited significant alterations of cardiac inflammatory genes. Despite these biochemical and cellular changes, there was no difference in infarct size between vehicle and CpG B treated I/R mice. Conclusion Systemic TLR9-stimulation upon onset of ischemia and subsequent reperfusion does not alter final infarct size despite causing clear alterations of both systemic and cardiac inflammatory parameters. Our results question the clinical usefulness of TLR9 activation during cardiac I/R. PMID:25126943

  19. Toll-like receptor 9-activation during onset of myocardial ischemia does not influence infarct extension.

    PubMed

    Ohm, Ingrid Kristine; Gao, Erhe; Belland Olsen, Maria; Alfsnes, Katrine; Bliksøen, Marte; Øgaard, Jonas; Ranheim, Trine; Nymo, Ståle Haugset; Holmen, Yangchen Dhondup; Aukrust, Pål; Yndestad, Arne; Vinge, Leif Erik

    2014-01-01

    Myocardial infarction (MI) remains a major cause of death and disability worldwide, despite available reperfusion therapies. Inflammatory signaling is considered nodal in defining final infarct size. Activation of the innate immune receptor toll-like receptors (TLR) 9 prior to ischemia and reperfusion (I/R) reduces infarct size, but the consequence of TLR9 activation timed to the onset of ischemia is not known. The TLR9-agonist; CpG B was injected i.p. in C57BL/6 mice immediately after induction of ischemia (30 minutes). Final infarct size, as well as area-at-risk, was measured after 24 hours of reperfusion. CpG B injection resulted in a significant increase in circulating granulocytes and monocytes both in sham and I/R mice. Paradoxically, clear evidence of reduced cardiac infiltration of both monocytes and granulocytes could be demonstrated in I/R mice treated with CpG B (immunocytochemistry, myeloperoxidase activity and mRNA expression patterns). In addition, systemic TLR9 activation elicited significant alterations of cardiac inflammatory genes. Despite these biochemical and cellular changes, there was no difference in infarct size between vehicle and CpG B treated I/R mice. Systemic TLR9-stimulation upon onset of ischemia and subsequent reperfusion does not alter final infarct size despite causing clear alterations of both systemic and cardiac inflammatory parameters. Our results question the clinical usefulness of TLR9 activation during cardiac I/R.

  20. Small Molecule Fluoride Toxicity Agonists

    PubMed Central

    Nelson1, James W.; Plummer, Mark S.; Blount, Kenneth F.; Ames, Tyler D.; Breaker, Ronald R.

    2015-01-01

    SUMMARY Fluoride is a ubiquitous anion that inhibits a wide variety of metabolic processes. Here we report the identification of a series of compounds that enhance fluoride toxicity in Escherichia coli and Streptococcus mutans. These molecules were isolated by using a high-throughput screen (HTS) for compounds that increase intracellular fluoride levels as determined via a fluoride riboswitch-reporter fusion construct. A series of derivatives were synthesized to examine structure-activity relationships, leading to the identification of compounds with improved activity. Thus, we demonstrate that small molecule fluoride toxicity agonists can be identified by HTS from existing chemical libraries by exploiting a natural fluoride riboswitch. In addition, our findings suggest that some molecules might be further optimized to function as binary antibacterial agents when combined with fluoride. PMID:25910244

  1. Small molecule fluoride toxicity agonists.

    PubMed

    Nelson, James W; Plummer, Mark S; Blount, Kenneth F; Ames, Tyler D; Breaker, Ronald R

    2015-04-23

    Fluoride is a ubiquitous anion that inhibits a wide variety of metabolic processes. Here, we report the identification of a series of compounds that enhance fluoride toxicity in Escherichia coli and Streptococcus mutans. These molecules were isolated by using a high-throughput screen (HTS) for compounds that increase intracellular fluoride levels as determined via a fluoride riboswitch reporter fusion construct. A series of derivatives were synthesized to examine structure-activity relationships, leading to the identification of compounds with improved activity. Thus, we demonstrate that small molecule fluoride toxicity agonists can be identified by HTS from existing chemical libraries by exploiting a natural fluoride riboswitch. In addition, our findings suggest that some molecules might be further optimized to function as binary antibacterial agents when combined with fluoride.

  2. [Melatonin receptor agonist].

    PubMed

    Uchiyama, Makoto

    2015-06-01

    Melatonin is a hormone secreted by the pineal gland and is involved in the regulation of human sleep-wake cycle and circadian rhythms. The melatonin MT1 and MT2 receptors located in the suprachiasmatic nucleus in the hypothalamus play a pivotal role in the sleep-wake regulation. Based on the fact that MT1 receptors are involved in human sleep onset process, melatonin receptor agonists have been developed to treat insomnia. In this article, we first reviewed functions of melatonin receptors with special reference to MT1 and MT2, and properties and clinical application of melatonin receptor agonists as hypnotics.

  3. Investigational melatonin receptor agonists.

    PubMed

    Hardeland, Rüdiger

    2010-06-01

    Melatonin is a major chronobiological regulator involved in circadian phasing, sleep, and numerous other functions including cyto-/neuroprotection, immune modulation, and energy metabolism. The suitability of melatonin as a drug is limited because of its short half-life. Therefore, various indolic and non-indolic melatonergic agonists have been developed. Frequent health problems such as sleep disturbances, neuropsychiatric disorders related to circadian dysphasing, and metabolic diseases associated with insulin resistance are targeted by melatonergic agonists. Various synthetic melatonergic drugs are compared with regard to receptor affinities, selectivity, effects on sleep, endogenous melatonin, circadian phase and insulin-related metabolism. The chemical design of melatonin receptor agonists is discussed in relation to consequences for receptor affinity, selectivity, metabolism, and spectrum of effects. Melatonergic agonists are suitable for phase-shifting circadian rhythms, and may be used for treating disorders related to circadian dysfunction including sleep difficulties. Facilitation of sleep onset is a general property, whereas promotion of sleep maintenance is demonstrable but not always fully sufficient. Details are especially available for tasimelteon. Support of insulin sensitivity may become a new area of application for compounds such as NEU-P11. Some drugs acting additionally as serotonergic antagonists display antidepressant properties.

  4. Fates of endocytosed somatostatin sst2 receptors and associated agonists.

    PubMed Central

    Koenig, J A; Kaur, R; Dodgeon, I; Edwardson, J M; Humphrey, P P

    1998-01-01

    Somatostatin agonists are rapidly and efficiently internalized with the somatostatin sst2 receptor. The fate of internalized agonists and receptors is of critical importance because the rate of ligand recycling back to the cell surface can limit the amount of radioligand accumulated inside the cells, whereas receptor recycling might be of vital importance in providing the cell surface with dephosphorylated, resensitized receptors. Furthermore the accumulation of radioisotope-conjugated somatostatin agonists inside cancer cells resulting from receptor-mediated internalization has been used as a treatment for cancers that overexpress somatostatin receptors. In the present study, radio-iodinated agonists at the sst2 somatostatin receptor were employed to allow quantitative analysis of the fate of endocytosed agonist. After endocytosis, recycling back to the cell surface was the main pathway for both 125I-labelled somatostatin-14 (SRIF-14) and the more stable agonist 125I-labelled cyclo(N-Me-Ala-Tyr-d-Trp-Lys-Abu-Phe) (BIM-23027; Abu stands for aminobutyric acid), accounting for 75-85% of internalized ligand when re-endocytosis of radioligand was prevented. We have shown that there is a dynamic cycling of both somatostatin agonist ligands and receptors between the cell surface and internal compartments both during agonist treatment and after surface-bound agonist has been removed, unless steps are taken to prevent the re-activation of receptors by recycled agonist. Internalization leads to increased degradation of 125I-labelled SRIF-14 but not 125I-labelled BIM-23027. The concentration of recycled agonist accumulating in the extracellular medium was sufficient to re-activate the receptor, as measured both by the inhibition of forskolin-stimulated adenylate cyclase and the recovery of surface receptor number after internalization. PMID:9820803

  5. Beta-Adrenergic Agonists

    PubMed Central

    Barisione, Giovanni; Baroffio, Michele; Crimi, Emanuele; Brusasco, Vito

    2010-01-01

    Inhaled β2-adrenoceptor (β2-AR) agonists are considered essential bronchodilator drugs in the treatment of bronchial asthma, both as symptoms-relievers and, in combination with inhaled corticosteroids, as disease-controllers. In this article, we first review the basic mechanisms by which the β2-adrenergic system contributes to the control of airway smooth muscle tone. Then, we go on describing the structural characteristics of β2-AR and the molecular basis of G-protein-coupled receptor signaling and mechanisms of its desensitization/ dysfunction. In particular, phosphorylation mediated by protein kinase A and β-adrenergic receptor kinase are examined in detail. Finally, we discuss the pivotal role of inhaled β2-AR agonists in the treatment of asthma and the concerns about their safety that have been recently raised. PMID:27713285

  6. Melatonin agonists and insomnia.

    PubMed

    Ferguson, Sally A; Rajaratnam, Shantha M W; Dawson, Drew

    2010-02-01

    The ability of melatonin to shift biological rhythms is well known. As a result, melatonin has been used in the treatment of various circadian rhythm sleep disorders, such as advanced and delayed sleep phase disorders, jet lag and shiftwork disorder. The current evidence for melatonin being efficacious in the treatment of primary insomnia is less compelling. The development of agents that are selective for melatonin receptors provides opportunity to further elucidate the actions of melatonin and its receptors and to develop novel treatments for specific types of sleep disorders. The agonists reviewed here - ramelteon, tasimelteon and agomelatine - all appear to be efficacious in the treatment of circadian rhythm sleep disorders and some types of insomnia. However, further studies are required to understand the mechanisms of action, particularly for insomnia. Clinical application of the agonists requires a good understanding of their phase-dependent properties. Long-term effects of melatonin should be evaluated in large-scale, independent randomized controlled trials.

  7. Toll-like receptor 9 deficiency impacts sensory and motor behaviors.

    PubMed

    Khariv, Veronika; Pang, Kevin; Servatius, Richard J; David, Brian T; Goodus, Matthew T; Beck, Kevin D; Heary, Robert F; Elkabes, Stella

    2013-08-01

    Toll-like receptors (TLRs) mediate the induction of the innate immune system in response to pathogens, injury and disease. However, they also play non-immune roles and are expressed in the central nervous system (CNS) during prenatal and postnatal stages including adulthood. Little is known about their roles in the CNS in the absence of pathology. Several members of the TLR family have been implicated in the development of neural and cognitive function although the contribution of TLR9 to these processes has not been well defined. The current studies were undertaken to determine whether developmental TLR9 deficiency affects motor, sensory or cognitive functions. We report that TLR9 deficient (TLR9(-/-)) mice show a hyper-responsive sensory and motor phenotype compared to wild type (TLR9(+/+)) controls. This is indicated by hypersensitivity to thermal stimuli in the hot plate paw withdrawal test, enhanced motor-responsivity under anxious conditions in the open field test and greater sensorimotor reactivity in the acoustic startle response. Prepulse inhibition (PPI) of the acoustic startle response was also enhanced, which indicates abnormal sensorimotor gating. In addition, subtle, but significant, gait abnormalities were noted in the TLR9(-/-) mice on the horizontal balance beam test with higher foot slip numbers than TLR9(+/+) controls. In contrast, spatial learning and memory, assessed by the Morris water maze, was similar in the TLR9(-/-) and TLR9(+/+) mice. These findings support the notion that TLR9 is important for the appropriate development of sensory and motor behaviors.

  8. Facilitation and inhibition of male rat ejaculatory behaviour by the respective 5-HT1A and 5-HT1B receptor agonists 8-OH-DPAT and anpirtoline, as evidenced by use of the corresponding new and selective receptor antagonists NAD-299 and NAS-181

    PubMed Central

    Hillegaart, Viveka; Ahlenius, Sven

    1998-01-01

    Ejaculatory problems and anorgasmia are well-known side-effects of the SSRI antidepressants, and a pharmacologically induced increase in serotonergic neurotransmission inhibits ejaculatory behaviour in the rat. In the present study the role of 5-HT1A and 5-HT1B receptors in the mediation of male rat ejaculatory behaviour was examined by use of selective agonists and antagonists acting at these 5-HT receptor subtypes.The 5-HT1A receptor agonist 8-OH-DPAT (0.25–4.00 μmol kg−1 s.c.) produced an expected facilitation of the male rat ejaculatory behaviour, and this effect was fully antagonized by pretreatment with the new selective 5-HT1A receptor antagonist (R)-3-N,N-dicyclobutylamino-8-fluoro-3,4-dihydro-2H-1-benzopyran-5-carboxamide hydrogen (2R,3R) tartrate monohydrate (NAD-299) (1.0 μmol kg−1 s.c.). NAD-299 by itself (0.75–3.00 μmol kg−1 s.c.) did not affect the male rat ejaculatory behaviour.The 5-HT1B receptor agonist anpirtoline (0.25–4.00 μmol kg−1 s.c.) produced a dose-dependent inhibition of the male rat ejaculatory behaviour, and this effect was fully antagonized by pretreatment with the 5-HT1B receptor antagonist isamoltane (16 μmol kg−1 s.c.) as well as by the new and selective antagonist (R)-(+)-2-(3-morpholinomethyl-2H-chromene-8-yl)oxymethylmorpholino methansulphonate (NAS-181) (16 μmol kg−1 s.c.). Isamoltane (1.0–16.0 μmol kg−1 s.c.) and NAD-181 (1.0–16.0 μmol kg−1 s.c.) had no, or weakly facilitatory effects on the male rat ejaculatory behaviour. The non-selective 5-HT1 receptor antagonist (−)-pindolol (8 μmol kg−1 s.c.), did not antagonize the inhibition produced by anpirtoline.The present results demonstrate opposite effects, facilitation and inhibition, of male rat ejaculatory behaviour by stimulation of 5-HT1A and 5-HT1B receptors, respectively, suggesting that the SSRI-induced inhibition of male ejaculatory dysfunction is due to 5-HT1B receptor stimulation. PMID:9886765

  9. Synthetic RORγ agonists regulate multiple pathways to enhance antitumor immunity

    PubMed Central

    Hu, Xiao; Liu, Xikui; Moisan, Jacques; Wang, Yahong; Lesch, Charles A.; Spooner, Chauncey; Morgan, Rodney W.; Zawidzka, Elizabeth M.; Mertz, David; Bousley, Dick; Majchrzak, Kinga; Kryczek, Ilona; Taylor, Clarke; Van Huis, Chad; Skalitzky, Don; Hurd, Alexander; Aicher, Thomas D.; Toogood, Peter L.; Glick, Gary D.; Paulos, Chrystal M.; Zou, Weiping; Carter, Laura L.

    2016-01-01

    ABSTRACT RORγt is the key transcription factor controlling the development and function of CD4+ Th17 and CD8+ Tc17 cells. Across a range of human tumors, about 15% of the CD4+ T cell fraction in tumor-infiltrating lymphocytes are RORγ+ cells. To evaluate the role of RORγ in antitumor immunity, we have identified synthetic, small molecule agonists that selectively activate RORγ to a greater extent than the endogenous agonist desmosterol. These RORγ agonists enhance effector function of Type 17 cells by increasing the production of cytokines/chemokines such as IL-17A and GM-CSF, augmenting expression of co-stimulatory receptors like CD137, CD226, and improving survival and cytotoxic activity. RORγ agonists also attenuate immunosuppressive mechanisms by curtailing Treg formation, diminishing CD39 and CD73 expression, and decreasing levels of co-inhibitory receptors including PD-1 and TIGIT on tumor-reactive lymphocytes. The effects of RORγ agonists were not observed in RORγ−/− T cells, underscoring the selective on-target activity of the compounds. In vitro treatment of tumor-specific T cells with RORγ agonists, followed by adoptive transfer to tumor-bearing mice is highly effective at controlling tumor growth while improving T cell survival and maintaining enhanced IL-17A and reduced PD-1 in vivo. The in vitro effects of RORγ agonists translate into single agent, immune system-dependent, antitumor efficacy when compounds are administered orally in syngeneic tumor models. RORγ agonists integrate multiple antitumor mechanisms into a single therapeutic that both increases immune activation and decreases immune suppression resulting in robust inhibition of tumor growth. Thus, RORγ agonists represent a novel immunotherapy approach for cancer. PMID:28123897

  10. Reproductive pharmacology of LHRH and agonists in females and males.

    PubMed

    Corbin, A; Bex, F J

    1980-06-01

    This report reviews research supporting the anti-reproductive pharmacologic characteristics of LHRH (luteinizing hormone releasing hormone) and its agonist analogues, and their relevance to fertility regulation in the clinic. Approximately 1000 derivatives of LHRH have been synthesized since 1971. LHRH and agonistic derivatives have the ability to induce the release of pituitary LH and FSH (follicle stimulating hormone), and ovulation in a variety of animal models. These agents have been shown to produce predictable postcoital contraceptive effects, such activity and potency having been related to its basic agonist properties. This class of peptides also have the ability to 1) retard puberty; 2) disrupt the estrous cycle (delay onset of estrus and mating); 3) induce premature ovulation; 4) induce luteolysis; 5) cause ovarian and uterine regression; 6) reduce fecundity in inseminated animals; and 7) inhibit ovarian/uterine stimulation which occurs with human chorionic gonadotropin. These effects are reversible because once treatment is withdrawn, normal breeding processes resume quickly. Several LHRH agonists are also being tapped for use as a potential luteal phase-inhibiting/menses-inducing approach to contraception. In the male, however, the agonists cannot function as contraceptives due to the inappropriate dissociation between testosterone production and spermatogenesis. The antireproductive mechanisms of LHRH agonists can be traced to the 1) hypersecretion of LH; 2) dysphasic FSH and distorted prolactin secretion; 3) decrease in gonadal LH, FSH and prolactin receptors; and 4) inhibition of steroidogenesis and eventual disruption of the reproductive continuum. They may also be useful as anti-tumor agents in steroid-dependent mammary gland and prostatic neoplasms. Toxicologic, pathologic and ancillary pharmacologic studies involving varied dosing regimens show encouraging potential of selected agonists as contraceptive agents with no related side effects.

  11. The sigma-1 receptor agonist 4-phenyl-1-(4-phenylbutyl) piperidine (PPBP) protects against newborn excitotoxic brain injury by stabilizing the mitochondrial membrane potential in vitro and inhibiting microglial activation in vivo.

    PubMed

    Wegleiter, Karina; Hermann, Martin; Posod, Anna; Wechselberger, Karina; Stanika, Ruslan I; Obermair, Gerald J; Kiechl-Kohlendorfer, Ursula; Urbanek, Martina; Griesmaier, Elke

    2014-11-01

    Premature birth represents a clinical situation of risk for brain injury. The diversity of pathophysiological processes complicates efforts to find effective therapeutic strategies. Excitotoxicity is one important factor in the pathogenesis of preterm brain injury. The observation that sigma-1 receptor agonists possess neuroprotective potential, at least partly mediated by a variety of anti-excitotoxic mechanisms, has generated great interest in targeting those receptors to counteract brain injury. The objective of this study was to evaluate the effect of the highly specific sigma-1 receptor agonist, 4-phenyl-1-(4-phenylbutyl) piperidine (PPBP) to protect against excitotoxic developmental brain injury in vivo and in vitro. Primary hippocampal neurons were pre-treated with PPBP before glutamate was applied and subsequently analyzed for cell death (PI/calcein AM), mitochondrial activity (TMRM) and morphology of the neuronal network (WGA) using confocal microscopy. Using an established neonatal mouse model we also determined whether systemic injection of PPBP significantly attenuates excitotoxic brain injury. PPBP significantly reduced neuronal cell death in primary hippocampal neurons exposed to glutamate. Neurons treated with PPBP showed a less pronounced loss of mitochondrial membrane potential and fewer morphological changes after glutamate exposure. A single intraperitoneal injection of PPBP given one hour after the excitotoxic insult significantly reduced microglial cell activation and lesion size in cortical gray and white matter. The present study provides strong support for the consideration of sigma-1 receptor agonists as a candidate therapy for the reduction of neonatal excitotoxic brain lesions and might offer a novel target to counteract developmental brain injury.

  12. Agonist-induced activation releases peroxisome proliferator-activated receptor beta/delta from its inhibition by palmitate-induced nuclear factor-kappaB in skeletal muscle cells.

    PubMed

    Jové, Mireia; Laguna, Juan C; Vázquez-Carrera, Manuel

    2005-05-01

    The mechanisms by which elevated levels of free fatty acids cause insulin resistance are not well understood, but there is a strong correlation between insulin resistance and intramyocellular lipid accumulation in skeletal muscle. In addition, accumulating evidence suggests a link between inflammation and type 2 diabetes. The aim of this work was to study whether the exposure of skeletal muscle cells to palmitate affected peroxisome proliferator-activated receptor (PPAR) beta/delta activity. Here, we report that exposure of C2C12 skeletal muscle cells to 0.75 mM palmitate reduced (74%, P<0.01) the mRNA levels of the PPARbeta/delta-target gene pyruvatedehydrogenase kinase 4 (PDK-4), which is involved in fatty acid utilization. This reduction was not observed in the presence of the PPARbeta/delta agonist L-165041. This drug prevented palmitate-induced nuclear factor (NF)-kappaB activation. Increased NF-kappaB activity after palmitate exposure was associated with enhanced protein-protein interaction between PPARbeta/delta and p65. Interestingly, treatment with the PPARbeta/delta agonist L-165041 completely abolished this interaction. These results indicate that palmitate may reduce fatty acid utilization in skeletal muscle cells by reducing PPARbeta/delta signaling through increased NF-kappaB activity.

  13. Defective Toll-like receptor 9-mediated cytokine production in B cells from Bruton's tyrosine kinase-deficient mice

    PubMed Central

    Hasan, Maroof; Lopez-Herrera, Gabriela; Blomberg, K Emelie M; Lindvall, Jessica M; Berglöf, Anna; Smith, C I Edvard; Vargas, Leonardo

    2008-01-01

    Bruton's tyrosine kinase (Btk), a member of the Tec family of tyrosine kinases, plays an important role in the differentiation and activation of B cells. Mutations affecting Btk cause immunodeficiency in both humans and mice. In this study we set out to investigate the potential role of Btk in Toll-like receptor 9 (TLR9) activation and the production of pro-inflammatory cytokines such as interleukin (IL)-6, tumour necrosis factor (TNF)-α and IL-12p40. Our data show that Btk-deficient B cells respond more efficiently to CpG-DNA stimulation, producing significantly higher levels of pro-inflammatory cytokines but lower levels of the inhibitory cytokine IL-10. The quantitative reverse transcription–polymerase chain reaction (RT-PCR) analysis presented in this work shows that mRNA production of one of the important new members of the IL-12 family, IL-27, was significantly increased in Btk-deficient B cells after CpG-DNA stimulation. In this study, we demonstrate significant differences in CpG responsiveness between transitional 1 (T1) and T2 B cells for survival and maturation. Furthermore, TLR9 expression, measured both as protein and as mRNA, was increased in Btk-defective cells, especially after TLR9 stimulation. Collectively, these data provide evidence in support of the theory that Btk regulates both TLR9 activation and expression in mouse splenic B cells. PMID:17725607

  14. Expression of Toll-like Receptor 9 in nose, peripheral blood and bone marrow during symptomatic allergic rhinitis

    PubMed Central

    Fransson, Mattias; Benson, Mikael; Erjefält, Jonas S; Jansson, Lennart; Uddman, Rolf; Björnsson, Sven; Cardell, Lars-Olaf; Adner, Mikael

    2007-01-01

    Background Allergic rhinitis is an inflammatory disease of the upper airway mucosa that also affects leukocytes in bone marrow and peripheral blood. Toll-like receptor 9 (TLR9) is a receptor for unmethylated CpG dinucleotides found in bacterial and viral DNA. The present study was designed to examine the expression of TLR9 in the nasal mucosa and in leukocytes derived from different cellular compartments during symptomatic allergic rhinitis. Methods The study was based on 32 patients with seasonal allergic rhinitis and 18 healthy subjects, serving as controls. Nasal biopsies were obtained before and after allergen challenge. Bone marrow, peripheral blood and nasal lavage fluid were sampled outside and during pollen season. The expression of TLR9 in tissues and cells was analyzed using immunohistochemistry and flow cytometry, respectively. Results TLR9 was found in several cell types in the nasal mucosa and in different leukocyte subpopulations derived from bone marrow, peripheral blood and nasal lavage fluid. The leukocyte expression was generally higher in bone marrow than in peripheral blood, and not affected by symptomatic allergic rhinitis. Conclusion The widespread expression of TLR9 in the nasal mucosa along with its rich representation in leukocytes in different compartments, demonstrate the possibility for cells involved in allergic airway inflammation to directly interact with bacterial and viral DNA. PMID:17328813

  15. Phospholipid Scramblase 1 regulates Toll-like receptor 9-mediated type I interferon production in plasmacytoid dendritic cells

    PubMed Central

    Talukder, Amjad H; Bao, Musheng; Kim, Tae Whan; Facchinetti, Valeria; Hanabuchi, Shino; Bover, Laura; Zal, Tomasz; Liu, Yong-Jun

    2012-01-01

    Toll-like receptor 9 (TLR9) senses microbial DNA in the endosomes of plasmacytoid dendritic cells (pDCs) and triggers MyD88-dependent type I interferon (IFN) responses. To better understand TLR9 biology in pDCs, we established a yeast two-hybrid library for the identification of TLR9-interacting proteins. Here, we report that an IFN-inducible protein, phospholipid scramblase 1 (PLSCR1), interacts with TLR9 in pDCs. Knockdown of PLSCR1 expression by siRNA in human pDC cell line led to a 60-70% reduction of IFN-α responses following CpG-ODN (oligodeoxynucleotide) stimulation. Primary pDCs from PLSCR1-deficient mice produced lower amount of type 1 IFN than pDCs from the wild-type mice in response to CpG-ODN, herpes simplex virus and influenza A virus. Following CpG-A stimulation, there were much lower amounts of TLR9 in the early endosomes together with CpG-A in pDCs from PLSCR1-deficient mice. Our study demonstrates that PLSCR1 is a TLR9-interacting protein that plays an important role in pDC's type 1 IFN responses by regulating TLR9 trafficking to the endosomal compartment. PMID:22453241

  16. Binding Mode of CpG Oligodeoxynucleotides to Nanoparticles Regulates Bifurcated Cytokine induction via Toll-like Receptor 9

    NASA Astrophysics Data System (ADS)

    Chinnathambi, Shanmugavel; Chen, Song; Ganesan, Singaravelu; Hanagata, Nobutaka

    2012-07-01

    The interaction of cytosine-phosphate-guanine oligodeoxynucleotides (CpG ODNs) with Toll-like receptor 9 (TLR9) activates the immune system. Multimeric class A CpG ODNs induce interferon-α (IFN-α) and, to a lesser extent, interleukin-6. By contrast, monomeric class B CpG ODNs induce interleukin-6 but not IFN-α. This difference suggests that the multimerization of CpG ODN molecules is a key factor in IFN-α induction. We multimerized class B CpG ODN2006x3-PD molecules that consist entirely of a phosphodiester backbone onto quantum dot silicon nanoparticles with various binding modes. Herein, we present the binding mode-dependent bifurcation of cytokine induction and discuss its possible mechanism of CpG ODN and TLR9 interaction. Our discoveries also suggest that nanoparticles play roles in not only delivery of CpG ODNs but also control of CpG ODN activity.

  17. Histamine H3-receptor inverse agonists as novel antipsychotics.

    PubMed

    Ito, Chihiro

    2009-06-01

    Schizophrenia (SZ) that is resistant to treatment with dopamine (DA) D2 antagonists may involve changes other than those in the dopaminergic system. Recently, histamine (HA), which regulates arousal and cognitive functions, has been suggested to act as a neurotransmitter in the central nervous system. Four HA receptors-H1, H2, H3, and H4-have been identified. Our recent basic and clinical studies revealed that brain HA improved the symptoms of SZ. The H3 receptor is primarily localized in the central nervous system, and it acts not only as a presynaptic autoreceptor that modulates the HA release but also as a presynaptic heteroreceptor that regulates the release of other neurotransmitters such as monoamines and amino acids. H3-receptor inverse agonists have been considered to improve cognitive functions. Many atypical antipsychotics are H3-receptor antagonists. Imidazole-containing H3-receptor inverse agonists inhibit not only cytochrome P450 but also hERG potassium channels (encoded by the human ether-a-go-go-related gene). Several imidazole H3-receptor inverse agonists also have high affinity for H4 receptors, which are expressed at high levels in mast cells and leukocytes. Clozapine is an H4-receptor agonist; this agonist activity may be related to the serious side effect of agranulocytosis caused by clozapine. Therefore, selective non-imidazole H3-receptor inverse agonists can be considered as novel antipsychotics that may improve refractory SZ.

  18. D-Cycloserine: Agonist turned antagonist.

    PubMed

    Lanthorn, T H

    1994-10-01

    D-Cycloserine can enhance activation of the NMDA receptor complex and could enhance the induction of long-term potentiation (LTP). In animals and humans, D-cycloserine can enhance performance in learning and memory tasks. This enhancing effect can disappear during repeated administration. The enhancing effects are also lost when higher doses are used, and replaced by behavioral and biochemical effects like those produced by NMDA antagonists. It has been reported that NMDA agonists, applied before or after tetanic stimulation, can block the induction of LTP. This may be the result of feedback inhibition of second messenger pathways stimulated by receptor activation. This may explain the antagonist-like effects of glycine partial agonists like D-cycloserine. In clinical trials of D-cycloserine in age-associated memory impairment (AAMI) and Alzheimer's disease, chronic treatment provided few positive effects on learning and memory. This may be due to inhibition of second messenger pathways following chronic stimulation of the receptor complex.

  19. Agonist pharmacology of two Drosophila GABA receptor splice variants.

    PubMed Central

    Hosie, A. M.; Sattelle, D. B.

    1996-01-01

    1. The Drosophila melanogaster gamma-aminobutyric acid (GABA) receptor subunits, RDLac and DRC 17-1-2, form functional homo-oligomeric receptors when heterologously expressed in Xenopus laevis oocytes. The subunits differ in only 17 amino acids, principally in regions of the N-terminal domain which determine agonist pharmacology in vertebrate ionotropic neurotransmitter receptors. A range of conformationally restricted GABA analogues were tested on the two homo-oligomers and their agonists pharmacology compared with that of insect and vertebrate iontropic GABA receptors. 2. The actions of GABA, isoguvacine and isonipecotic acid on RDLac and DRC 17-1-2 homo-oligomers were compared, by use of two-electrode voltage-clamp. All three compounds were full agonists of both receptors, but were 4-6 fold less potent agonists of DRC 17-1-2 homo-oligomers than of RDLac. However, the relative potencies of these agonists on each receptor were very similar. 3. A more complete agonist profile was established for RDLac homo-oligomers. The most potent agonists of these receptors were GABA, muscimol and trans-aminocrotonic acid (TACA), which were approximately equipotent. RDLac homo-oligomers were fully activated by a range of GABA analogues, with the order of potency: GABA > ZAPA ((Z)-3-[(aminoiminomethyl)thio]prop-2-enoic acid) > isoguvacine > imidazole-4-acetic acid > or = isonipecotic acid > or = cis-aminocrotonic acid (CACA) > beta-alanine. 3-Aminopropane sulphonic acid (3-APS), a partial agonist of RDLac homo-oligomers, was the weakest agonist tested and 100 fold less potent than GABA. 4. SR95531, an antagonist of vertebrate GABAA receptors, competitively inhibited the GABA responses of RDLac homo-oligomers, which have previously been found to insensitive to bicuculline. However, its potency (IC50 500 microM) was much reduced when compared to GABAA receptors. 5. The agonist pharmacology of Drosophila RDLac homo-oligomers exhibits aspects of the characteristic pharmacology of

  20. Switching cannabinoid response from CB(2) agonists to FAAH inhibitors.

    PubMed

    Tourteau, Aurélien; Leleu-Chavain, Natascha; Body-Malapel, Mathilde; Andrzejak, Virginie; Barczyk, Amélie; Djouina, Madjid; Rigo, Benoit; Desreumaux, Pierre; Chavatte, Philippe; Millet, Régis

    2014-03-01

    A series of 3-carboxamido-5-aryl-isoxazoles designed as CB2 agonists were evaluated as FAAH inhibitors. The pharmacological results led to identify structure-activity relationships enabling to switch cannabinoid response from CB2 agonists to FAAH inhibitors. Two compounds were selected for their FAAH and/or CB2 activity, and evaluated in a colitis model for their anti-inflammatory activity. Results showed that compounds 10 and 11 inhibit the development of DSS-induced acute colitis in mice and then, are interesting leads to explore new drug candidates for IBD.

  1. Herpes simplex virus type-1 induces IFN-alpha production via Toll-like receptor 9-dependent and -independent pathways.

    PubMed

    Hochrein, Hubertus; Schlatter, Beatrix; O'Keeffe, Meredith; Wagner, Cornelia; Schmitz, Frank; Schiemann, Matthias; Bauer, Stefan; Suter, Mark; Wagner, Hermann

    2004-08-03

    Type I IFN production in response to the DNA virus herpes simplex virus type-1 (HSV-1) is essential in controlling viral replication. We investigated whether plasmacytoid dendritic cells (pDC) were the major tissue source of IFN-alpha, and whether the production of IFN-alpha in response to HSV-1 depended on Toll-like receptor 9 (TLR9). Total spleen cells or bone marrow (BM) cells, or fractions thereof, including highly purified pDC, from WT, TLR9, and MyD88 knockout mice were stimulated with known ligands for TLR9 or active HSV-1. pDC freshly isolated from both spleen and BM were the major source of IFN-alpha in response to oligodeoxynucleotides containing CpG motifs, but in response to HSV-1 the majority of IFN-alpha was produced by other cell types. Moreover, IFN-alpha production by non-pDC was independent of TLR9. The tissue source determined whether pDC responded to HSV-1 in a strictly TLR9-dependent fashion. Freshly isolated BM pDC or pDC derived from culture of BM precursors with FMS-like tyrosine kinase-3 ligand, produced IFN-alpha in the absence of functional TLR9, whereas spleen pDC did not. Heat treatment of HSV-1 abolished maturation and IFN-alpha production from all TLR9-deficient DC but not WT DC. Thus pDC and non-pDC produce IFN-alpha in response to HSV-1 via both TLR9-independent and -dependent pathways.

  2. Recognition of CpG oligodeoxynucleotides by human Toll-like receptor 9 and subsequent cytokine induction.

    PubMed

    Suwarti, Suwarti; Yamazaki, Tomohiko; Svetlana, Chechetka; Hanagata, Nobutaka

    2013-01-25

    Toll-like receptor 9 (TLR9) recognizes a synthetic ligand, oligodeoxynucleotide (ODN) containing cytosine-phosphate-guanine (CpG). Activation of TLR9 by CpG ODN induces a signal transduction cascade that plays a pivotal role in first-line immune defense in the human body. The three-dimensional structure of TLR9 has not yet been reported, and the ligand-binding mechanism of TLR9 is still poorly understood; therefore, the mechanism of human TLR9 (hTLR9) ligand binding needs to be elucidated. In this study, we constructed several hTLR9 mutants, including truncated mutants and single mutants in the predicted CpG ODN-binding site. We used these mutants to analyze the role of potential important regions of hTLR9 in receptor signaling induced by phosphorothioate (PTO)-modified CpG ODN and CpG ODNs only consist entirely of a phosphodiester (PD) backbone, CpG ODN2006x3-PD that we developed. We found truncated mutants of hTLR9 lost the signaling activity, indicating that both the C- and N-termini of the extracellular domain (ECD) are necessary for the function of hTLR9. We identified residues, His505, Gln510, His530, and Tyr554, in the C-terminal of hTLR9-ECD that are essential for hTLR9 activation. These residues might form positive charged clusters with which negatively charged CpG ODN could interact. Furthermore, we observed ODN-PD induced interleukin-6 (IL-6) through TLR9 in a CpG-sequence-dependent manner in human peripheral blood mononuclear cells and B cells, whereas ODN-PTO induced IL-6 in a CpG-sequence-independent manner. These finding are relevant for the mechanism of hTLR9 activation by CpG ODNs.

  3. Bitter taste receptor agonists alter mitochondrial function and induce autophagy in airway smooth muscle cells.

    PubMed

    Pan, Shi; Sharma, Pawan; Shah, Sushrut D; Deshpande, Deepak A

    2017-07-01

    Airway remodeling, including increased airway smooth muscle (ASM) mass, is a hallmark feature of asthma and COPD. We previously identified the expression of bitter taste receptors (TAS2Rs) on human ASM cells and demonstrated that known TAS2R agonists could promote ASM relaxation and bronchodilation and inhibit mitogen-induced ASM growth. In this study, we explored cellular mechanisms mediating the antimitogenic effect of TAS2R agonists on human ASM cells. Pretreatment of ASM cells with TAS2R agonists chloroquine and quinine resulted in inhibition of cell survival, which was largely reversed by bafilomycin A1, an autophagy inhibitor. Transmission electron microscope studies demonstrated the presence of double-membrane autophagosomes and deformed mitochondria. In ASM cells, TAS2R agonists decreased mitochondrial membrane potential and increased mitochondrial ROS and mitochondrial fragmentation. Inhibiting dynamin-like protein 1 (DLP1) reversed TAS2R agonist-induced mitochondrial membrane potential change and attenuated mitochondrial fragmentation and cell death. Furthermore, the expression of mitochondrial protein BCL2/adenovirus E1B 19-kDa protein-interacting protein 3 (Bnip3) and mitochondrial localization of DLP1 were significantly upregulated by TAS2R agonists. More importantly, inhibiting Bnip3 mitochondrial localization by dominant-negative Bnip3 significantly attenuated cell death induced by TAS2R agonist. Collectively the TAS2R agonists chloroquine and quinine modulate mitochondrial structure and function, resulting in ASM cell death. Furthermore, Bnip3 plays a central role in TAS2R agonist-induced ASM functional changes via a mitochondrial pathway. These findings further establish the cellular mechanisms of antimitogenic effects of TAS2R agonists and identify a novel class of receptors and pathways that can be targeted to mitigate airway remodeling as well as bronchoconstriction in obstructive airway diseases. Copyright © 2017 the American Physiological

  4. Agonist-activated ion channels

    PubMed Central

    Colquhoun, David

    2006-01-01

    This paper looks at ion channels as an example of the pharmacologist's stock in trade, the action of an agonist on a receptor to produce a response. Looked at in this way, ion channels have been helpful because they are still the only system which is simple enough for quantitative investigation of transduction mechanisms. A short history is given of attempts to elucidate what happens between the time when agonist first binds, and the time when the channel opens. PMID:16402101

  5. Toll-like receptor 9 expression is associated with breast cancer sensitivity to the growth inhibitory effects of bisphosphonates in vitro and in vivo

    PubMed Central

    Sandholm, Jouko; Lehtimäki, Jaakko; Ishizu, Tamiko; Velu, Sadanandan E.; Clark, Jeremy; Härkönen, Pirkko; Jukkola-Vuorinen, Arja; Schrey, Aleksi; Harris, Kevin W.; Tuomela, Johanna M.; Selander, Katri S.

    2016-01-01

    Bisphosphonates are standard treatments for bone metastases. When given in the adjuvant setting, they reduce breast cancer mortality and recurrence in bone but only among post-menopausal patients. Optimal drug use would require biomarker-based patient selection. Such biomarkers are not yet in clinical use. Based on the similarities in inflammatory responses to bisphosphonates and Toll-like receptor (TLR) agonists, we hypothesized that TLR9 expression may affect bisphosphonate responses in cells. We compared bisphosphonate effects in breast cancer cell lines with low or high TLR9 expression. We discovered that cells with decreased TLR9 expression are significantly more sensitive to the growth-inhibitory effects of bisphosphonates in vitro and in vivo. Furthermore, cancer growth-promoting effects seen with some bisphosphonates in some control shRNA cells were not detected in TLR9 shRNA cells. These differences were not associated with inhibition of Rap1A prenylation or p38 phosphorylation, which are known markers for bisphosphonate activity. However, TLR9 shRNA cells exhibited increased sensitivity to ApppI, a metabolite that accumulates in cells after bisphosphonate treatment. We conclude that decreased TLR9-expression sensitizes breast cancer cells to the growth inhibitory effects of bisphosphonates. Our results suggest that TLR9 should be studied as a potential biomarker for adjuvant bisphosphonate sensitivity among breast cancer patients. PMID:27888633

  6. Benzodiazepine agonist and inverse agonist actions on GABAA receptor-operated chloride channels. II. Chronic effects of ethanol

    SciTech Connect

    Buck, K.J.; Harris, R.A. )

    1990-05-01

    Mice were made tolerant to and dependent on ethanol by administration of a liquid diet. Gamma-aminobutyric acid (GABA) receptor-dependent uptake of 36Cl- by mouse cortical microsacs was used to study the actions of benzodiazepine (BZ) agonists and inverse agonists. Chronic exposure to ethanol attenuated the ability of a BZ agonist, flunitrazepam, to augment muscimol-stimulated uptake of 36Cl- and enhanced the actions of BZ inverse agonists, Ro15-4513 (ethyl-8-azido-5,6-dihydro-5-methyl-6-oxo-4H-imidazo(1,4)-benzodiazepine - 3-carboxylate) and DMCM (methyl-6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate), to inhibit GABAA receptor-operated chloride channels. Augmentation of chloride flux by pentobarbital was not reduced by chronic ethanol exposure. Attenuation of flunitrazepam efficacy was transient and returned to control levels within 6 to 24 hr after withdrawal from ethanol, but increased sensitivity to Ro15-4513 was observed as long as 8 days after withdrawal. Chronic exposure to ethanol did not alter (3H)SR 95531 (2-(3'-carbethoxy-2'propyl)-3-amino-6-p-methoxyphenylpyridazinium bromide) binding to low-affinity GABAA receptors or muscimol stimulation of chloride flux; and did not alter (3H)Ro15-4513 or (3H)flunitrazepam binding to central BZ receptors or allosteric modulation of this binding by muscimol (i.e., muscimol-shift). These results suggest that chronic exposure to ethanol reduces coupling between BZ agonist sites and the chloride channel, and may be responsible for the development of cross-tolerance between ethanol and BZ agonists. In contrast, coupling between BZ inverse agonist sites and the chloride channel is increased.

  7. Agonist-trafficking and hallucinogens.

    PubMed

    González-Maeso, Javier; Sealfon, Stuart C

    2009-01-01

    Seven transmembrane domain receptors, also termed G protein-coupled receptors (GPCRs), represent the most common molecular target for therapeutic drugs. The generally accepted pharmacological model for GPCR activation is the ternary complex model, in which GPCRs exist in a dynamic equilibrium between the active and inactive conformational states. However, the demonstration that different agonists sometimes elicit a different relative activation of two signaling pathways downstream of the same receptor has led to a revision of the ternary complex model. According to this agonist- trafficking model, agonists stabilize distinct activated receptor conformations that preferentially activate specific signaling pathways. Hallucinogenic drugs and non-hallucinogenic drugs represent an attractive experimental system with which to study agonist-trafficking of receptor signaling. Thus many of the behavioral responses induced by hallucinogenic drugs, such as lysergic acid diethylamide (LSD), psilocybin or mescaline, depend on activation of serotonin 5-HT(2A) receptors (5-HT2ARs). In contrast, this neuropsychological state in humans is not induced by closely related chemicals, such as lisuride or ergotamine, despite their similar in vitro activity at the 5-HT2AR. In this review, we summarize the current knowledge, as well as unresolved questions, regarding agonist-trafficking and the mechanism of action of hallucinogenic drugs.

  8. [Alpha 2-adrenoceptor agonists for the treatment of chronic pain].

    PubMed

    Kulka, P J

    1996-04-25

    The antinociceptive effect of alpha(2)-adrenoceptor agonists is mediated by activation of descending inhibiting noradrenergic systems, which modulates 'wide-dynamic-range' neurones. Furthermore, they inhibit the liberation of substance P and endorphines and activate serotoninergic neurones. Despite this variety of antinociceptive actions, there is still little experience with alpha(2)-adrenoceptor agonists as therapeutic agents for use in chronic pain syndromes. Studies in animals and patients have shown that the transdermal, epidural and intravenous administration of the alpha(2)-adrenoceptor agonist clonidine reduces pain intensity in neuropathic pain syndromes for periods varying from some hours up to 1 month. Patients suffering from lancinating or sharp pain respond best to this therapy. Topically applied clonidine (200-300 microg) relieves hyperalgesia in sympathetically maintained pain. Epidural administration of 300 microg clonidine dissolved in 5 ml NaCl 0.9 % has also been shown to be effective. In patients suffering from cancer pain tolerant to opioids, pain control has proved possible again with combinations of opioids and clonidine. In isolated cases clonidine has been administered epidurally at a dose of 1500 microg/day for almost 5 months without evidence for any histotoxic property of clonidine. Side effects often observed during administration of alpha(2)-adrenoceptor agonists are dry mouth, sedation, hypotension and bradycardia. Therapeutic interventions are usually not required.

  9. Electrophysiological perspectives on the therapeutic use of nicotinic acetylcholine receptor partial agonists.

    PubMed

    Papke, Roger L; Trocmé-Thibierge, Caryn; Guendisch, Daniela; Al Rubaiy, Shehd Abdullah Abbas; Bloom, Stephen A

    2011-05-01

    Partial agonist therapies rely variously on two hypotheses: the partial agonists have their effects through chronic low-level receptor activation or the partial agonists work by decreasing the effects of endogenous or exogenous full agonists. The relative significance of these activities probably depends on whether acute or chronic effects are considered. We studied nicotinic acetylcholine receptors (nAChRs) expressed in Xenopus laevis oocytes to test a model for the acute interactions between acetylcholine (ACh) and weak partial agonists. Data were best-fit to a basic competition model that included an additional factor for noncompetitive inhibition. Partial agonist effects were compared with the nAChR antagonist bupropion in prolonged bath application experiments that were designed to mimic prolonged drug exposure typical of therapeutic drug delivery. A primary effect of prolonged application of nicotine was to decrease the response of all nAChR subtypes to acute applications of ACh. In addition, nicotine, cytisine, and varenicline produced detectable steady-state activation of α4β2* [(α4)(2)(β2)(3), (α4)(3)(β2)(2), and (α4)(2)(β2)(2)α5)] receptor subtypes that was not seen with other test compounds. Partial agonists produced no detectable steady-state activation of α7 nAChR, but seemed to show small potentiation of ACh-evoked responses; however, "run-up" of α7 ACh responses was also sometimes observed under control conditions. Potential off-target effects of the partial agonists therefore included the modulation of α7 responses by α4β2 partial agonists and decreases in α4β2* responses by α7-selective agonists. These data indicate the dual effects expected for α4β2* partial agonists and provide models and insights for utility of partial agonists in therapeutic development.

  10. Agonists for the Chemokine Receptor CXCR4

    PubMed Central

    2011-01-01

    The development of agonists for the chemokine receptor CXCR4 could provide promising therapeutic candidates. On the basis of previously forwarded two site model of chemokine–receptor interactions, we hypothesized that linking the agonistic N-terminus of SDF-1 to the T140 backbone would yield new high-affinity agonists of CXCR4. We developed chimeras with the agonistic SDF-1 N-terminus grafted to a T140 side chain and tested their binding affinity and chemotactic agonist activity. While chimeras with the peptide grafted onto position 12 of T140 remained high-affinity antagonists, those bearing the peptide on position 14 were in part agonists. One chimera was a full CXCR4 agonist with 25 nM affinity, and several chimeras showed low nanomolar affinities with partial agonist activity. Our results confirmed that we have developed high-affinity agonists of CXCR4. PMID:21841963

  11. Targeting toll-like receptor 9 with CpG oligodeoxynucleotides enhances tumor response to fractionated radiotherapy.

    PubMed

    Mason, Kathryn A; Ariga, Hisanori; Neal, Robert; Valdecanas, David; Hunter, Nancy; Krieg, Arthur M; Whisnant, John K; Milas, Luka

    2005-01-01

    Synthetic oligodeoxynucleotides containing unmethylated CpG motifs detected by Toll-like receptor 9 of dendritic cells and B cells have potent immunomodulatory effects. CpG oligodeoxynucleotides induce cytokines, activate natural killer cells, and elicit T-cell responses leading to antitumor effects, including improved efficacy of chemotherapeutic agents and, as we reported recently, synergy between CpG oligodeoxynucleotide 1826 and single-dose radiotherapy of an immunogenic mouse fibrosarcoma. The present study extends this finding to the fractionated radiotherapy of the fibrosarcoma tumor and assesses the ability of CpG oligodeoxynucleotide 1826 to increase the radioresponse of a tumor (nonimmunogenic fibrosarcoma). The experiments used a murine immunogenic fibrosarcoma tumor, fibrosarcoma growing in the leg of mice, and response to radiotherapy was assessed by tumor growth delay and tumor cure rate (TCD50, radiation dose yielding 50% tumor cure). Multiple s.c. peritumoral or i.t. administrations of CpG oligodeoxynucleotide 1826 at a dose of 100 microg per mouse were given when established tumors were 6 mm in diameter. Local tumor irradiation was initiated when tumors grew to 8 mm in diameter; radiation was delivered in 1 to 9 Gy fractions given twice daily separated by 6 to 7 hours for 5 consecutive days to achieve a total dose of 10 to 90 Gy. CpG oligodeoxynucleotide 1826, given as a single agent, had only a small antitumor effect, but it dramatically enhanced fibrosarcoma response to radiotherapy. Although 83.1 (79.2-90.0) Gy total dose were needed to achieve tumor cures in 50% of mice treated with radiotherapy alone, only 23.0 (11.5-32.7) Gy total dose were needed in mice treated with both CpG oligodeoxynucleotide 1826 and radiotherapy. The magnitude of potentiation of tumor radioresponse at the TCD50 level was by a factor of 3.61, a much higher value than that (a factor of 1.93) that we reported for single-dose radiotherapy. Mice cured of their tumors by

  12. Human herpesvirus 6A infection in CD46 transgenic mice: viral persistence in the brain and increased production of proinflammatory chemokines via Toll-like receptor 9.

    PubMed

    Reynaud, Joséphine M; Jégou, Jean-François; Welsch, Jérémy C; Horvat, Branka

    2014-05-01

    Human herpesvirus 6 (HHV-6) is widely spread in the human population and has been associated with several neuroinflammatory diseases, including multiple sclerosis. To develop a small-animal model of HHV-6 infection, we analyzed the susceptibility of several lines of transgenic mice expressing human CD46, identified as a receptor for HHV-6. We showed that HHV-6A (GS) infection results in the expression of viral transcripts in primary brain glial cultures from CD46-expressing mice, while HHV-6B (Z29) infection was inefficient. HHV-6A DNA persisted for up to 9 months in the brain of CD46-expressing mice but not in the nontransgenic littermates, whereas HHV-6B DNA levels decreased rapidly after infection in all mice. Persistence in the brain was observed with infectious but not heat-inactivated HHV-6A. Immunohistological studies revealed the presence of infiltrating lymphocytes in periventricular areas of the brain of HHV-6A-infected mice. Furthermore, HHV-6A stimulated the production of a panel of proinflammatory chemokines in primary brain glial cultures, including CCL2, CCL5, and CXCL10, and induced the expression of CCL5 in the brains of HHV-6A-infected mice. HHV-6A-induced production of chemokines in the primary glial cultures was dependent on the stimulation of toll-like receptor 9 (TLR9). Finally, HHV-6A induced signaling through human TLR9 as well, extending observations from the murine model to human infection. Altogether, this study presents a first murine model for HHV-6A-induced brain infection and suggests a role for TLR9 in the HHV-6A-initiated production of proinflammatory chemokines in the brain, opening novel perspectives for the study of virus-associated neuropathology. HHV-6 infection has been related to neuroinflammatory diseases; however, the lack of a suitable small-animal infection model has considerably hampered further studies of HHV-6-induced neuropathogenesis. In this study, we have characterized a new model for HHV-6 infection in mice

  13. Toll-like receptor 9-mediated protection of enterovirus 71 infection in mice is due to the release of danger-associated molecular patterns.

    PubMed

    Hsiao, Hung-Bo; Chou, Ai-Hsiang; Lin, Su-I; Chen, I-Hua; Lien, Shu-Pei; Liu, Chia-Chyi; Chong, Pele; Liu, Shih-Jen

    2014-10-01

    Enterovirus 71 (EV71), a positive-stranded RNA virus, is the major cause of hand, foot, and mouth disease (HFMD) with severe neurological symptoms. Antiviral type I interferon (alpha/beta interferon [IFN-α/β]) responses initiated from innate receptor signaling are inhibited by EV71-encoded proteases. It is less well understood whether EV71-induced apoptosis provides a signal to activate type I interferon responses as a host defensive mechanism. In this report, we found that EV71 alone cannot activate Toll-like receptor 9 (TLR9) signaling, but supernatant from EV71-infected cells is capable of activating TLR9. We hypothesized that TLR9-activating signaling from plasmacytoid dendritic cells (pDCs) may contribute to host defense mechanisms. To test our hypothesis, Flt3 ligand-cultured DCs (Flt3L-DCs) from both wild-type (WT) and TLR9 knockout (TLR9KO) mice were infected with EV71. More viral particles were produced in TLR9KO mice than by WT mice. In contrast, alpha interferon (IFN-α), monocyte chemotactic protein 1 (MCP-1), tumor necrosis factor-alpha (TNF-α), IFN-γ, interleukin 6 (IL-6), and IL-10 levels were increased in Flt3L-DCs from WT mice infected with EV71 compared with TLR9KO mice. Seven-day-old TLR9KO mice infected with a non-mouse-adapted EV71 strain developed neurological lesion-related symptoms, including hind-limb paralysis, slowness, ataxia, and lethargy, but WT mice did not present with these symptoms. Lung, brain, small intestine, forelimb, and hind-limb tissues collected from TLR9KO mice exhibited significantly higher viral loads than equivalent tissues collected from WT mice. Histopathologic damage was observed in brain, small intestine, forelimb, and hind-limb tissues collected from TLR9KO mice infected with EV71. Our findings demonstrate that TLR9 is an important host defense molecule during EV71 infection. Importance: The host innate immune system is equipped with pattern recognition receptors (PRRs), which are useful for defending the host

  14. Dopamine agonist therapy in hyperprolactinemia.

    PubMed

    Webster, J

    1999-12-01

    Introduction of the dopamine agonist bromocriptine heralded a major advance in the management of hyperprolactinemic disorders. Although its side effects of nausea, dizziness and headache and its short elimination half-life are limiting factors, its efficacy established it as a reference compound against the activity of which several dopamine agonists, like pergolide, lysuride, metergoline, terguride and dihydroergocristine, fell by the wayside. More recently, two new agents, cabergoline and quinagolide, have been introduced and appear to offer considerable advantages over bromocriptine. Cabergoline, an ergoline D2 agonist, has a long plasma half-life that enables once- or twice-weekly administration. Quinagolide, in contrast, is a nonergot D2 agonist with an elimination half-life intermediate between those of bromocriptine and cabergoline, allowing the drug to be administered once daily. Comparative studies indicate that cabergoline is clearly superior to bromocriptine in efficacy (prolactin suppression, restoration of gonadal function) and in tolerability. In similar studies, quinagolide appeared to have similar efficacy and superior tolerability to that of bromocriptine. Results of a small crossover study indicate that cabergoline is better tolerated, with a trend toward activity superior to that of quinagolide. In hyperprolactinemic men and in women not seeking to become pregnant, cabergoline may be regarded as the treatment of choice.

  15. CB(1) receptor allosteric modulators display both agonist and signaling pathway specificity.

    PubMed

    Baillie, Gemma L; Horswill, James G; Anavi-Goffer, Sharon; Reggio, Patricia H; Bolognini, Daniele; Abood, Mary E; McAllister, Sean; Strange, Phillip G; Stephens, Gary J; Pertwee, Roger G; Ross, Ruth A

    2013-02-01

    We have previously identified allosteric modulators of the cannabinoid CB(1) receptor (Org 27569, PSNCBAM-1) that display a contradictory pharmacological profile: increasing the specific binding of the CB(1) receptor agonist [(3)H]CP55940 but producing a decrease in CB(1) receptor agonist efficacy. Here we investigated the effect one or both compounds in a broad range of signaling endpoints linked to CB(1) receptor activation. We assessed the effect of these compounds on CB(1) receptor agonist-induced [(35)S]GTPγS binding, inhibition, and stimulation of forskolin-stimulated cAMP production, phosphorylation of extracellular signal-regulated kinases (ERK), and β-arrestin recruitment. We also investigated the effect of these allosteric modulators on CB(1) agonist binding kinetics. Both compounds display ligand dependence, being significantly more potent as modulators of CP55940 signaling as compared with WIN55212 and having little effect on [(3)H]WIN55212 binding. Org 27569 displays biased antagonism whereby it inhibits: agonist-induced guanosine 5'-O-(3-[(35)S]thio)triphosphate ([(35)S]GTPγS) binding, simulation (Gα(s)-mediated), and inhibition (Gα(i)-mediated) of cAMP production and β-arrestin recruitment. In contrast, it acts as an enhancer of agonist-induced ERK phosphorylation. Alone, the compound can act also as an allosteric agonist, increasing cAMP production and ERK phosphorylation. We find that in both saturation and kinetic-binding experiments, the Org 27569 and PSNCBAM-1 appeared to influence only orthosteric ligand maximum occupancy rather than affinity. The data indicate that the allosteric modulators share a common mechanism whereby they increase available high-affinity CB(1) agonist binding sites. The receptor conformation stabilized by the allosterics appears to induce signaling and also selectively traffics orthosteric agonist signaling via the ERK phosphorylation pathway.

  16. Ascorbic acid enables reversible dopamine receptor /sup 3/H-agonist binding

    SciTech Connect

    Leff, S.; Sibley, D.R.; Hamblin, M.; Creese, I.

    1981-11-16

    The effects of ascorbic acid on dopaminergic /sup 3/H-agonist receptor binding were studied in membrane homogenates of bovine anterior pituitary and caudate, and rat striatum. In all tissues virtually no stereospecific binding (defined using 1uM (+)butaclamol) of the /sup 3/H-agonists N-propylnorapomorphine (NPA), apomorphine, or dopamine could be demonstrated in the absence of ascorbic acid. Although levels of total /sup 3/H-agonist binding were three to five times greater in the absence than in the presence of 0.1% ascorbic acid, the increased binding was entirely non-stereospecific. Greater amounts of dopamine-inhibitable /sup 3/H-NPA binding could be demonstrated in the absence of 0.1% ascorbic acid, but this measure of ''specific binding'' was demonstrated not to represent dopamine receptor binding since several other catecholamines and catechol were equipotent with dopamine and more potent than the dopamine agonist (+/-)amino-6,7-dihydroxy-1,2,3,4-tetrahydronapthalene (ADTN) in inhibiting this binding. High levels of dopamine-displaceable /sup 3/H-agonist binding were detected in fresh and boiled homogenates of cerebellum, an area of brain which receives no dopaminergic innervation, further demonstrating the non-specific nature of /sup 3/H-agonist binding in the absence of ascorbic acid. These studies emphasize that under typical assay conditions ascorbic acid is required in order to demonstrate reversible and specific /sup 3/H-agonist binding to dopamine receptors.

  17. Evaluation of peroxisome proliferator-activated receptor agonists on interleukin-5-induced eosinophil differentiation

    PubMed Central

    Smith, Steven G; Hill, Mike; Oliveria, John-Paul; Watson, Brittany M; Baatjes, Adrian J; Dua, Benny; Howie, Karen; Campbell, Heather; Watson, Rick M; Sehmi, Roma; Gauvreau, Gail M

    2014-01-01

    Peroxisome proliferator-activated receptor (PPAR) agonists have been suggested as novel therapeutics for the treatment of inflammatory lung disease, such as allergic asthma. Treatment with PPAR agonists has been shown to inhibit airway eosinophilia in murine models of allergic asthma, which can occur through several mechanisms including attenuated generation of chemoattractants (e.g. eotaxin) and decreased eosinophil migrational responses. In addition, studies report that PPAR agonists can inhibit the differentiation of several cell types. To date, no studies have examined the effects of PPAR agonists on interleukin-5 (IL-5) -induced eosinophil differentiation from haemopoietic progenitor cells. Non-adherent mononuclear cells or CD34+ cells isolated from the peripheral blood of allergic subjects were grown for 2 weeks in Methocult® cultures with IL-5 (10 ng/ml) and IL-3 (25 ng/ml) in the presence of 1–1000 nm PPARα agonist (GW9578), PPARβ/δ agonist (GW501516), PPARγ agonist (rosiglitazone) or diluent. The number of eosinophil/basophil colony-forming units (Eo/B CFU) was quantified by light microscopy. The signalling mechanism involved was assessed by phosphoflow. Blood-extracted CD34+ cells cultured with IL-5 or IL-5 + IL-3 formed Eo/B CFU, which were significantly inhibited by rosiglitazone (100 nm, P < 0·01) but not GW9578 or GW501516. In addition, rosglitazone significantly inhibited IL-5-induced phosphorylation of extracellular signal-regulated kinase 1/2. We observed an inhibitory effect of rosiglitazone on eosinophil differentiation in vitro, mediated by attenuation of the extracellular signal-regulated kinase 1/2 signalling pathway. These findings indicate that the PPARγ agonist can attenuate tissue eosinophilia by interfering with local differentiative responses. PMID:24628018

  18. Bitter Taste Receptor Agonists Mitigate Features of Allergic Asthma in Mice

    PubMed Central

    Sharma, Pawan; Yi, Roslyn; Nayak, Ajay P.; Wang, Nadan; Tang, Francesca; Knight, Morgan J.; Pan, Shi; Oliver, Brian; Deshpande, Deepak A.

    2017-01-01

    Asthma is characterized by airway inflammation, mucus secretion, remodeling and hyperresponsiveness (AHR). Recent research has established the bronchodilatory effect of bitter taste receptor (TAS2R) agonists in various models. Comprehensive pre-clinical studies aimed at establishing effectiveness of TAS2R agonists in disease models are lacking. Here we aimed to determine the effect of TAS2R agonists on features of asthma. Further, we elucidated a mechanism by which TAS2R agonists mitigate features of asthma. Asthma was induced in mice using intranasal house dust mite or aerosol ova-albumin challenge, and chloroquine or quinine were tested in both prophylactic and treatment models. Allergen challenge resulted in airway inflammation as evidenced by increased immune cells infiltration and release of cytokines and chemokines in the lungs, which were significantly attenuated in TAS2R agonists treated mice. TAS2R agonists attenuated features of airway remodeling including smooth muscle mass, extracellular matrix deposition and pro-fibrotic signaling, and also prevented mucus accumulation and development of AHR in mice. Mechanistic studies using human neutrophils demonstrated that inhibition of immune cell chemotaxis is a key mechanism by which TAS2R agonists blocked allergic airway inflammation and exerted anti-asthma effects. Our comprehensive studies establish the effectiveness of TAS2R agonists in mitigating multiple features of allergic asthma. PMID:28397820

  19. Bitter Taste Receptor Agonists Mitigate Features of Allergic Asthma in Mice.

    PubMed

    Sharma, Pawan; Yi, Roslyn; Nayak, Ajay P; Wang, Nadan; Tang, Francesca; Knight, Morgan J; Pan, Shi; Oliver, Brian; Deshpande, Deepak A

    2017-04-11

    Asthma is characterized by airway inflammation, mucus secretion, remodeling and hyperresponsiveness (AHR). Recent research has established the bronchodilatory effect of bitter taste receptor (TAS2R) agonists in various models. Comprehensive pre-clinical studies aimed at establishing effectiveness of TAS2R agonists in disease models are lacking. Here we aimed to determine the effect of TAS2R agonists on features of asthma. Further, we elucidated a mechanism by which TAS2R agonists mitigate features of asthma. Asthma was induced in mice using intranasal house dust mite or aerosol ova-albumin challenge, and chloroquine or quinine were tested in both prophylactic and treatment models. Allergen challenge resulted in airway inflammation as evidenced by increased immune cells infiltration and release of cytokines and chemokines in the lungs, which were significantly attenuated in TAS2R agonists treated mice. TAS2R agonists attenuated features of airway remodeling including smooth muscle mass, extracellular matrix deposition and pro-fibrotic signaling, and also prevented mucus accumulation and development of AHR in mice. Mechanistic studies using human neutrophils demonstrated that inhibition of immune cell chemotaxis is a key mechanism by which TAS2R agonists blocked allergic airway inflammation and exerted anti-asthma effects. Our comprehensive studies establish the effectiveness of TAS2R agonists in mitigating multiple features of allergic asthma.

  20. Modulation of [3H]diazepam binding in rat cortical membranes by GABAA agonists.

    PubMed

    Wong, E H; Iversen, L L

    1985-04-01

    GABAA receptor agonists modulate [3H]diazepam binding in rat cortical membranes with different efficacies. At 23 degrees C, the relative potencies for enhancement of [3H]diazepam binding by agonists parallel their potencies in inhibiting [3H]gamma-aminobutyric acid [( 3H]GABA) binding. The agonist concentrations needed for enhancement of [3H]diazepam binding are up to 35 times higher than for [3H]GABA binding and correspond closely to the concentrations required for displacement of [3H]bicuculline methochloride (BMC) binding. The maximum enhancement of [3H]diazepam varied among agonists: muscimol = GABA greater than isoguvacine greater than 3-aminopropane sulphonic acid (3APS) = imidazoleacetic acid (IAA) greater than 4,5,6,7-tetrahydroisoxazolo (4,5,6)-pyridin-3-ol (THIP) = taurine greater than piperidine 4-sulphonic acid (P4S). At 37 degrees C, the potencies of agonists remained unchanged, but isoguvacine, 3 APS, and THIP acquired efficacies similar to GABA, whereas IAA, taurine, and P4S maintained their partial agonist profiles. At both temperatures the agonist-induced enhancement of [3H]diazepam binding was reversible by bicuculline methobromide and by the steroid GABA antagonist RU 5135. These results stress the importance of studying receptor-receptor interaction under near-physiological conditions and offer an in vitro assay that may predict the agonist status of putative GABA receptor ligands.

  1. Telmisartan, a possible PPAR-δ agonist, reduces TNF-α-stimulated VEGF-C production by inhibiting the p38MAPK/HSP27 pathway in human proximal renal tubular cells.

    PubMed

    Kimura, Hideki; Mikami, Daisuke; Kamiyama, Kazuko; Sugimoto, Hidehiro; Kasuno, Kenji; Takahashi, Naoki; Yoshida, Haruyoshi; Iwano, Masayuki

    2014-11-14

    Vascular endothelial growth factor-C (VEGF-C) is a main inducer of inflammation-associated lymphangiogenesis in various inflammatory disorders including chronic progressive kidney diseases, for which angiotensin II receptor type 1 blockers (ARBs) are widely used as the main treatment. Although proximal renal tubular cells may affect the formation of lymphatic vessels in the interstitial area by producing VEGF-C, the molecular mechanisms of VEGF-C production and its manipulation by ARB have not yet been examined in human proximal renal tubular epithelial cells (HPTECs). In the present study, TNF-α dose-dependently induced the production of VEGF-C in HPTECs. The TNF-α-induced production of VEGF-C was mediated by the phosphorylation of p38MAPK and HSP27, but not by that of ERK or NFkB. Telmisartan, an ARB that can activate the peroxisome proliferator-activated receptor (PPAR), served as a PPAR-δ activator and reduced the TNF-α-stimulated production of VEGF-C. This reduction was partially attributed to a PPAR-δ-dependent decrease in p38MAPK phosphorylation. Our results indicate that TNF-α induced the production of VEGF-C in HPTECs by activating p38MAPK/HSP27, and this was partially inhibited by telmisartan in a PPAR-δ dependent manner. These results provide a novel insight into inflammation-associated lymphangiogenesis.

  2. The thyroid hormone receptor (TR) beta-selective agonist GC-1 inhibits proliferation but induces differentiation and TR beta mRNA expression in mouse and rat osteoblast-like cells.

    PubMed

    Beber, Eduardo H; Capelo, Luciane P; Fonseca, Tatiana L; Costa, Cristiane C; Lotfi, Claudimara F; Scanlan, Thomas S; Gouveia, Cecilia H A

    2009-04-01

    Previous studies showed anabolic effects of GC-1, a triiodothyronine (T3) analogue that is selective for both binding and activation functions of thyroid hormone receptor (TR) beta1 over TRalpha1, on bone tissue in vivo. The aim of this study was to investigate the responsiveness of rat (ROS17/2.8) and mouse (MC3T3-E1) osteoblast-like cells to GC-1. As expected, T3 inhibited cellular proliferation and stimulated mRNA expression of osteocalcin or alkaline phosphatase in both cell lineages. Whereas equimolar doses of T3 and GC-1 equally affected these parameters in ROS17/2.8 cells, the effects of GC-1 were more modest compared to those of T3 in MC3T3-E1 cells. Interestingly, we showed that there is higher expression of TRalpha1 than TRbeta1 mRNA in rat (approximately 20-90%) and mouse (approximately 90-98%) cell lineages and that this difference is even higher in mouse cells, which highlights the importance of TRalpha1 to bone physiology and may partially explain the modest effects of GC-1 in comparison with T3 in MC3T3-E1 cells. Nevertheless, we showed that TRbeta1 mRNA expression increases (approximately 2.8- to 4.3-fold) as osteoblastic cells undergo maturation, suggesting a key role of TRbeta1 in mediating T3 effects in the bone forming cells, especially in mature osteoblasts. It is noteworthy that T3 and GC-1 induced TRbeta1 mRNA expression to a similar extent in both cell lineages (approximately 2- to 4-fold), indicating that both ligands may modulate the responsiveness of osteoblasts to T3. Taken together, these data show that TRbeta selective T3 analogues have the potential to directly induce the differentiation and activity of osteoblasts.

  3. Telmisartan, a possible PPAR-δ agonist, reduces TNF-α-stimulated VEGF-C production by inhibiting the p38MAPK/HSP27 pathway in human proximal renal tubular cells

    SciTech Connect

    Kimura, Hideki; Mikami, Daisuke; Kamiyama, Kazuko; Sugimoto, Hidehiro; Kasuno, Kenji; Takahashi, Naoki; Yoshida, Haruyoshi; Iwano, Masayuki

    2014-11-14

    Highlights: • TNF-α increased VEGF-C expression by enhancing phosphorylation of p38MAPK and HSP27. • Telmisartan decreased TNF-α-stimulated expression of VEGF-C. • Telmisartan suppressed TNF-α-induced phosphorylation of p38MAPK and HSP27. • Telmisartan activated endogenous PPAR-δ protein. • Telmisartan suppressed p38MAPK phosphorylation in a PPAR-δ-dependent manner. - Abstract: Vascular endothelial growth factor-C (VEGF-C) is a main inducer of inflammation-associated lymphangiogenesis in various inflammatory disorders including chronic progressive kidney diseases, for which angiotensin II receptor type 1 blockers (ARBs) are widely used as the main treatment. Although proximal renal tubular cells may affect the formation of lymphatic vessels in the interstitial area by producing VEGF-C, the molecular mechanisms of VEGF-C production and its manipulation by ARB have not yet been examined in human proximal renal tubular epithelial cells (HPTECs). In the present study, TNF-α dose-dependently induced the production of VEGF-C in HPTECs. The TNF-α-induced production of VEGF-C was mediated by the phosphorylation of p38MAPK and HSP27, but not by that of ERK or NFkB. Telmisartan, an ARB that can activate the peroxisome proliferator-activated receptor (PPAR), served as a PPAR-δ activator and reduced the TNF-α-stimulated production of VEGF-C. This reduction was partially attributed to a PPAR-δ-dependent decrease in p38MAPK phosphorylation. Our results indicate that TNF-α induced the production of VEGF-C in HPTECs by activating p38MAPK/HSP27, and this was partially inhibited by telmisartan in a PPAR-δ dependent manner. These results provide a novel insight into inflammation-associated lymphangiogenesis.

  4. Matrix metalloproteinase-13 is regulated by toll-like receptor-9 in colorectal cancer cells and mediates cellular migration

    PubMed Central

    RATH, TIMO; STÖCKLE, JULIA; RODERFELD, MARTIN; TSCHUSCHNER, ANNETTE; GRAF, JÜRGEN; ROEB, ELKE

    2011-01-01

    Matrix metalloproteinases (MMPs) are associated with cancer cell invasion and metastasis, and are currently the most prominent proteases associated with tumorigenesis. In particular, abundant expression of MMP-13 in colorectal cancer (CRC) is correlated with poor survival and the existence of distant metastasis. As suggested by recent in vitro studies, MMP-13 expression is regulated in a toll-like receptor (TLR)-9-dependent manner. In this study, we quantified the expression of MMP-13, TLR-9 and second messengers of the TLR signal transduction in CRC cells compared to colonic fibroblasts by RT-PCR. Furthermore, the effects of a selective TLR-9 stimulation on the expression of MMP-13 in CRC cells and colonic fibroblasts were analyzed. MMP-13 and TLR-9 as well as associated second messengers were simultaneously up-regulated in LS174 and SW620 cells compared to fibroblasts. Selective TLR-9 agonism with CpG oligonucleotides led to a significant increase in MMP-13 gene expression after 12 h of incubation in LS174 cells and after 12 and 24 h in SW620 cells, but not when using GpC oligonucleotides as a control substance. By contrast, MMP-13 gene expression remained unchanged in colonic fibroblasts following treatment with CpG or GpC oligonucleotides. The effects of selective MMP-13 inhibition on cellular migration were analyzed in Boyden chamber experiments. In the presence of 10 and 20 μM of the specific MMP-13 inhibitor, CL-82198, migration of the LS174 cells was significantly reduced by 55 and 52%, respectively, compared to untreated cells. In conclusion, the results of this study provide evidence of the TLR-9-dependent regulation of MMP-13 in CRC cells, but not in colonic fibroblasts. Since the specific inhibition of MMP-13 significantly reduces the migration of LS174 cells, selective MMP-13 inhibition may be a promising therapeutic strategy in CRC. PMID:22866107

  5. Antimitogenic effect of bitter taste receptor agonists on airway smooth muscle cells.

    PubMed

    Sharma, Pawan; Panebra, Alfredo; Pera, Tonio; Tiegs, Brian C; Hershfeld, Alena; Kenyon, Lawrence C; Deshpande, Deepak A

    2016-02-15

    Airway remodeling is a hallmark feature of asthma and chronic obstructive pulmonary disease. Clinical studies and animal models have demonstrated increased airway smooth muscle (ASM) mass, and ASM thickness is correlated with severity of the disease. Current medications control inflammation and reverse airway obstruction effectively but have limited effect on remodeling. Recently we identified the expression of bitter taste receptors (TAS2R) on ASM cells, and activation with known TAS2R agonists resulted in ASM relaxation and bronchodilation. These studies suggest that TAS2R can be used as new therapeutic targets in the treatment of obstructive lung diseases. To further establish their effectiveness, in this study we aimed to determine the effects of TAS2R agonists on ASM growth and promitogenic signaling. Pretreatment of healthy and asthmatic human ASM cells with TAS2R agonists resulted in a dose-dependent inhibition of ASM proliferation. The antimitogenic effect of TAS2R ligands was not dependent on activation of protein kinase A, protein kinase C, or high/intermediate-conductance calcium-activated K(+) channels. Immunoblot analyses revealed that TAS2R agonists inhibit growth factor-activated protein kinase B phosphorylation without affecting the availability of phosphatidylinositol 3,4,5-trisphosphate, suggesting TAS2R agonists block signaling downstream of phosphatidylinositol 3-kinase. Furthermore, the antimitogenic effect of TAS2R agonists involved inhibition of induced transcription factors (activator protein-1, signal transducer and activator of transcription-3, E2 factor, nuclear factor of activated T cells) and inhibition of expression of multiple cell cycle regulatory genes, suggesting a direct inhibition of cell cycle progression. Collectively, these findings establish the antimitogenic effect of TAS2R agonists and identify a novel class of receptors and signaling pathways that can be targeted to reduce or prevent airway remodeling as well as

  6. PPAR Agonists: I. Role of Receptor Subunits in Alcohol Consumption in Male and Female Mice.

    PubMed

    Blednov, Yuri A; Black, Mendy; Benavidez, Jillian M; Stamatakis, Eleni E; Harris, R Adron

    2016-03-01

    Several peroxisome proliferator-activated receptor (PPAR) agonists reduce voluntary alcohol consumption in rodent models, and evidence suggests that PPARα and γ subunits play an important role in this effect. To define the subunit dependence of this action, we tested selective PPARα and α/γ agonists and antagonists in addition to null mutant mice lacking PPARα. The effects of fenofibrate (PPARα agonist) and tesaglitazar (PPARα/γ agonist) on continuous and intermittent 2-bottle choice drinking tests were examined in male and female wild-type mice and in male mice lacking PPARα. We compared the ability of MK886 (PPARα antagonist) and GW9662 (PPARγ antagonist) to inhibit the effects of fenofibrate and tesaglitazar in wild-type mice. The estrogen receptor antagonist, tamoxifen, can inhibit PPARγ-dependent transcription and was also studied in male and female mice. Fenofibrate and tesaglitazar reduced ethanol (EtOH) consumption and preference in wild-type mice, but these effects were not observed in mice lacking PPARα. MK886 inhibited the action of fenofibrate, but not tesaglitazer, while GW9662 did not inhibit either agonist. The PPAR agonists were more effective in male mice compared to females, and drinking in the continuous 2-bottle choice test was more sensitive to fenofibrate and tesaglitazar compared to drinking in the intermittent access test. Tamoxifen also reduced EtOH consumption in male mice and this action was inhibited by GW9662, but not MK886, suggesting that it acts by activation of PPARγ. Our study using selective PPAR agonists, antagonists, and null mutant mice indicates a key role for PPARα in mediating reduced EtOH consumption by fenofibrate and tesaglitazar. Copyright © 2016 by the Research Society on Alcoholism.

  7. Antimitogenic effect of bitter taste receptor agonists on airway smooth muscle cells

    PubMed Central

    Sharma, Pawan; Panebra, Alfredo; Pera, Tonio; Tiegs, Brian C.; Hershfeld, Alena; Kenyon, Lawrence C.

    2015-01-01

    Airway remodeling is a hallmark feature of asthma and chronic obstructive pulmonary disease. Clinical studies and animal models have demonstrated increased airway smooth muscle (ASM) mass, and ASM thickness is correlated with severity of the disease. Current medications control inflammation and reverse airway obstruction effectively but have limited effect on remodeling. Recently we identified the expression of bitter taste receptors (TAS2R) on ASM cells, and activation with known TAS2R agonists resulted in ASM relaxation and bronchodilation. These studies suggest that TAS2R can be used as new therapeutic targets in the treatment of obstructive lung diseases. To further establish their effectiveness, in this study we aimed to determine the effects of TAS2R agonists on ASM growth and promitogenic signaling. Pretreatment of healthy and asthmatic human ASM cells with TAS2R agonists resulted in a dose-dependent inhibition of ASM proliferation. The antimitogenic effect of TAS2R ligands was not dependent on activation of protein kinase A, protein kinase C, or high/intermediate-conductance calcium-activated K+ channels. Immunoblot analyses revealed that TAS2R agonists inhibit growth factor-activated protein kinase B phosphorylation without affecting the availability of phosphatidylinositol 3,4,5-trisphosphate, suggesting TAS2R agonists block signaling downstream of phosphatidylinositol 3-kinase. Furthermore, the antimitogenic effect of TAS2R agonists involved inhibition of induced transcription factors (activator protein-1, signal transducer and activator of transcription-3, E2 factor, nuclear factor of activated T cells) and inhibition of expression of multiple cell cycle regulatory genes, suggesting a direct inhibition of cell cycle progression. Collectively, these findings establish the antimitogenic effect of TAS2R agonists and identify a novel class of receptors and signaling pathways that can be targeted to reduce or prevent airway remodeling as well as

  8. PPARγ agonists induce adipocyte differentiation by modulating the expression of Lipin-1, which acts as a PPARγ phosphatase.

    PubMed

    Kim, Jina; Lee, Yu-Jin; Kim, Jung Min; Lee, So Young; Bae, Myung-Ae; Ahn, Jin Hee; Han, Dong Cho; Kwon, Byoung-Mog

    2016-12-01

    PPARγ agonists induced obesity in animal models as a side effect. Microarray experiments reveal that PPARγ agonist upregulates the expression of lipin-1 and this upregulation is correlated with the activity of the agonists. Lipin-1 induced by PPARγ agonists decreased the levels of PPARγ and ERK1/2 phosphorylation through direct interaction with these proteins in 3T3-L1 cells. In PPARγ agonist-treated 3T3-L1 preadipocytes, the knockdown of lipin-1 expression by small interfering RNA inhibited the adipogenesis that was induced by PPARγ agonists. In contrast, PPARγ2 expression was increased, and lipid droplets were accumulated in lipin-1-overexpressing 3T3-L1 adipocytes. Rosiglitazone (RGZ), a strong PPARγ agonist, synergistically promoted PPARγ dephosphorylation and adipogenesis in lipin-1-overexpressing 3T3-L1 preadipocytes. Therefore, lipin-1 has dual functions as a transcriptional cofactor and phosphatidate phosphatase (PAP) in the differentiation of preadipocyte cells induced by strong PPARγ agonists. In addition, the adipogenesis of 3T3-L1 cells was markedly upregulated by diacylglycerol (DAG), which was produced by lipin-1. Therefore, lipin-1 induction by PPARγ agonists might be an important factor in understanding the biological mechanism of the agonists' adverse effects, and this information may be valuable in the development of type-2 diabetes mellitus (T2DM) therapeutics with reduced adverse effects and greater tolerability. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Characterization of a novel bivalent morphinan possessing kappa agonist and micro agonist/antagonist properties.

    PubMed

    Mathews, Jennifer L; Peng, Xuemei; Xiong, Wennan; Zhang, Ao; Negus, S Stevens; Neumeyer, John L; Bidlack, Jean M

    2005-11-01

    Previous research has shown that compounds with mixed kappa and mu activity may have utility for the treatment of cocaine abuse and dependence. The present study characterizes the pharmacological profile of a bivalent morphinan that was shown to be a kappa opioid receptor agonist and a mu opioid receptor agonist/antagonist. MCL-145 [bis(N-cyclobutylmethylmorphinan) fumarate] is related to the morphinan cyclorphan and its N-cyclobutylmethyl derivative MCL-101 [3-hydroxy-N-cyclobutylmethyl morphinan S-(+)-mandelate]. MCL-145 consists of two morphinans connected by a spacer at the 3-hydroxy position. This compound had K(i) values of 0.078 and 0.20 nM for the kappa and mu opioid receptors, respectively, using radioligand binding assays as shown by Neumeyer et al. in 2003. In the guanosine 5'-O -(3-[(35) S]thiotriphosphate) binding assay, MCL-145 produced an E(max) value of 80% for the kappa opioid receptor and 42% for the mu opioid receptor. The EC(50) values obtained for this compound were 4.3 and 3.1 nM for the kappa and mu opioid receptors, respectively. In vivo MCL-145 produced a full dose-response curve in the 55 degrees C warm water tail-flick test and was equipotent to morphine. The agonist properties of MCL-145 were antagonized by the mu-selective antagonist beta-funaltrexamine and the kappa-selective antagonist nor-binaltorphimine. MCL-145 also acted as a mu antagonist, as measured by the inhibition of morphine-induced antinociception.

  10. Gremlin: vexing VEGF receptor agonist.

    PubMed

    Claesson-Welsh, Lena

    2010-11-04

    Gremlins are mischievous creatures in English folklore, believed to be the cause of otherwise unexplainable breakdowns (the word gremlins is derived from the Old English "gremian" or "gremman," "to vex"). Gremlin (or Gremlin-1) is also the designation of a secreted protein that is known to regulate bone formation during development. In this issue of Blood, Mitola et al report the novel role of Gremlin as a VEGFR2 agonist and the function of the Gremlin protein seems vexing indeed.

  11. Novel diazabicycloalkane delta opioid agonists.

    PubMed

    Loriga, Giovanni; Lazzari, Paolo; Manca, Ilaria; Ruiu, Stefania; Falzoi, Matteo; Murineddu, Gabriele; Bottazzi, Mirko Emilio Heiner; Pinna, Giovanni; Pinna, Gérard Aimè

    2015-09-01

    Here we report the investigation of diazabicycloalkane cores as potential new scaffolds for the development of novel analogues of the previously reported diazatricyclodecane selective delta (δ) opioid agonists, as conformationally constrained homologues of the reference δ agonist (+)-4-[(αR)-α((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide (SNC80). In particular, we have simplified the diazatricyclodecane motif of δ opioid agonist prototype 1a with bridged bicyclic cores. 3,6-diazabicyclo[3.1.1]heptane, 3,8-diazabicyclo[3.2.1]octane, 3,9-diazabicyclo[3.3.1]nonane, 3,9-diazabicyclo[4.2.1]nonane, and 3,10-diazabicyclo[4.3.1]decane were adopted as core motifs of the novel derivatives. The compounds were synthesized and biologically assayed as racemic (3-5) or diastereoisomeric (6,7) mixtures. All the novel compounds 3-7 showed δ agonism behaviour and remarkable affinity to δ receptors. Amongst the novel derivatives, 3,8-diazabicyclo[3.2.1]octane based compound 4 evidenced improved δ affinity and selectivity relative to SNC80. Published by Elsevier Ltd.

  12. Dihydropyrrolo[2,3-d]pyrimidines: Selective Toll-Like Receptor 9 Antagonists from Scaffold Morphing Efforts

    PubMed Central

    2014-01-01

    Toll-like receptors (TLRs) play important roles in the innate immune system. In fact, recognition of endogenous immune complexes containing self-nucleic acids as pathogen- or damage-associated molecular patterns contributes to certain autoimmune diseases, and inhibition of these recognition signals is expected to have therapeutic value. We identified dihydropyrrolo[2,3-d]pyrimidines as novel selective TLR9 antagonists with high aqueous solubility. A structure–activity relationship study of a known TLR9 antagonist led to the promising compound 18, which showed potent TLR9 antagonistic activity, sufficient aqueous solubility for parenteral formulation, and druggable properties. Compound 18 suppressed the production of the proinflammatory cytokine IL-6 in CpG-induced mouse model. It is therefore believed that compound 18 has great potential in the treatment of TLR9-mediated systemic uncontrollable inflammatory response like sepsis. PMID:25408837

  13. CB1 Receptor Allosteric Modulators Display Both Agonist and Signaling Pathway Specificity

    PubMed Central

    Baillie, Gemma L.; Horswill, James G.; Anavi-Goffer, Sharon; Reggio, Patricia H.; Bolognini, Daniele; Abood, Mary E.; McAllister, Sean; Strange, Phillip G.; Stephens, Gary J.; Pertwee, Roger G.

    2013-01-01

    We have previously identified allosteric modulators of the cannabinoid CB1 receptor (Org 27569, PSNCBAM-1) that display a contradictory pharmacological profile: increasing the specific binding of the CB1 receptor agonist [3H]CP55940 but producing a decrease in CB1 receptor agonist efficacy. Here we investigated the effect one or both compounds in a broad range of signaling endpoints linked to CB1 receptor activation. We assessed the effect of these compounds on CB1 receptor agonist–induced [35S]GTPγS binding, inhibition, and stimulation of forskolin-stimulated cAMP production, phosphorylation of extracellular signal-regulated kinases (ERK), and β-arrestin recruitment. We also investigated the effect of these allosteric modulators on CB1 agonist binding kinetics. Both compounds display ligand dependence, being significantly more potent as modulators of CP55940 signaling as compared with WIN55212 and having little effect on [3H]WIN55212 binding. Org 27569 displays biased antagonism whereby it inhibits: agonist-induced guanosine 5′-O-(3-[35S]thio)triphosphate ([35S]GTPγS) binding, simulation (Gαs-mediated), and inhibition (Gαi-mediated) of cAMP production and β-arrestin recruitment. In contrast, it acts as an enhancer of agonist-induced ERK phosphorylation. Alone, the compound can act also as an allosteric agonist, increasing cAMP production and ERK phosphorylation. We find that in both saturation and kinetic-binding experiments, the Org 27569 and PSNCBAM-1 appeared to influence only orthosteric ligand maximum occupancy rather than affinity. The data indicate that the allosteric modulators share a common mechanism whereby they increase available high-affinity CB1 agonist binding sites. The receptor conformation stabilized by the allosterics appears to induce signaling and also selectively traffics orthosteric agonist signaling via the ERK phosphorylation pathway. PMID:23160940

  14. Novel nonsecosteroidal VDR agonists with phenyl-pyrrolyl pentane skeleton.

    PubMed

    Shen, Wei; Xue, Jingwei; Zhao, Zekai; Zhang, Can

    2013-11-01

    In order to find the vitamin D receptor (VDR) ligand whose VDR agonistic activity is separated from the calcemic activity sufficiently, novel nonsecosteroidal analogs with phenyl-pyrrolyl pentane skeleton were synthesized and evaluated for the VDR binding affinity, antiproliferative activity in vitro and serum calcium raising ability in vivo (tacalcitol used as control). Among them, several compounds showed varying degrees of VDR agonistic and growth inhibition activities of the tested cell lines. The most effective compound 2g (EC₅₀: 1.06 nM) exhibited stronger VDR agonistic activity than tacalcitol (EC₅₀: 7.05 nM), inhibited the proliferations of HaCaT and MCF-7 cells with IC₅₀ of 2.06 μM and 0.307 μM (tacalcitol: 2.07 μM and 0.057 μM) and showed no significant effect on serum calcium. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

  15. Contact- and agonist-regulated microvesiculation of human platelets.

    PubMed

    Zhang, Yanjun; Liu, Xiao; Liu, Li; Zaske, Ana-Maria; Zhou, Zhou; Fu, Yuanyuan; Yang, Xi; Conyers, Jodie L; Li, Min; Dong, Jing-fei; Zhang, Jianning

    2013-08-01

    After exposure to an agonist, platelets are activated and become aggregated. They also shed membrane microparticles that participate in the pathogenesis of thrombosis, hyper-coagulation and inflammation. However, microvesiculation can potentially disrupt the integrity of platelet aggregation by shedding the membrane receptors and phosphatidylserine critical for forming and stabilising a platelet clot. We tested the hypothesis that adhesion and microvesiculation are functions of different subsets of platelets at the time of haemostasis by real-time monitoring of agonist-induced morphological changes and microvesiculation of human platelets.We identified two types of platelets that are adherent to fibrinogen: a high density bubble shape (HDBS) and low-density spread shape (LDSS). Adenosine diphosphate (ADP) predominantly induced HDBS platelets to vesiculate, whereas LDSS platelets were highly resistant to such vesiculation. Thrombin-receptor activating peptide (TRAP) stabilised platelets against microvesiculation by promoting a rapid HDBS-to-LDSS morphological transition. These activities of ADP and TRAP were reversed for platelets in suspension, independent of an engagement integrin αIIbβ3. As the result of membrane contact, LDSS platelets inhibited the microvesiculation of HDBS platelets in response to ADP. Aspirin and clopidogrel inhibited ADP-induced microvesiculation through different mechanisms. These results suggest that platelet aggregation and microvesiculation occur in different subsets of platelets and are differently regulated by agonists, platelet-platelets and platelet-fibrinogen interactions.

  16. GABAergic involvement in motor effects of an adenosine A(2A) receptor agonist in mice.

    PubMed

    Khisti, R T; Chopde, C T; Abraham, E

    2000-04-03

    Adenosine A(2A) agonists are known to induce catalepsy and inhibit dopamine mediated motor hyperactivity. An antagonistic interaction between adenosine A(2A) and dopamine D(2) receptors is known to regulate GABA-mediated neurotransmission in striatopallidal neurons. Stimulation of adenosine A(2A) and dopamine D(2) receptors has been shown to increase and inhibit GABA release respectively in pallidal GABAergic neurons. However, the role of GABAergic neurotransmission in the motor effects of adenosine A(2A) receptors is not yet known. Therefore in the present study the effect of GABAergic agents on adenosine A(2A) receptor agonist (NECA- or CGS 21680) induced catalepsy and inhibition of amphetamine elicited motor hyperactivity was examined. Pretreatment with GABA, the GABA(A) agonist muscimol or the GABA(B) agonist baclofen potentiated whereas the GABA(A) antagonist bicuculline attenuated NECA- or CGS 21680-induced catalepsy. However, the GABA(B) antagonists phaclophen and delta-aminovaleric acid had no effect. Administration of NECA or CGS 21680 not only reduced spontaneous locomotor activity but also antagonized amphetamine elicited motor hyperactivity. These effects of NECA and CGS 21680 were potentiated by GABA or muscimol and antagonized by bicuculline. These findings provide behavioral evidence for the role of GABA in the motor effects of adenosine A(2A) receptor agonists. Activation of adenosine A(2A) receptors increases GABA release which could reduce dopaminergic tone and induce catalepsy or inhibit amphetamine mediated motor hyperactivity.

  17. Regulation of membrane cholecystokinin-2 receptor by agonists enables classification of partial agonists as biased agonists.

    PubMed

    Magnan, Rémi; Masri, Bernard; Escrieut, Chantal; Foucaud, Magali; Cordelier, Pierre; Fourmy, Daniel

    2011-02-25

    Given the importance of G-protein-coupled receptors as pharmacological targets in medicine, efforts directed at understanding the molecular mechanism by which pharmacological compounds regulate their presence at the cell surface is of paramount importance. In this context, using confocal microscopy and bioluminescence resonance energy transfer, we have investigated internalization and intracellular trafficking of the cholecystokinin-2 receptor (CCK2R) in response to both natural and synthetic ligands with different pharmacological features. We found that CCK and gastrin, which are full agonists on CCK2R-induced inositol phosphate production, rapidly and abundantly stimulate internalization. Internalized CCK2R did not rapidly recycle to plasma membrane but instead was directed to late endosomes/lysosomes. CCK2R endocytosis involves clathrin-coated pits and dynamin and high affinity and prolonged binding of β-arrestin1 or -2. Partial agonists and antagonists on CCK2R-induced inositol phosphate formation and ERK1/2 phosphorylation did not stimulate CCK2R internalization or β-arrestin recruitment to the CCK2R but blocked full agonist-induced internalization and β-arrestin recruitment. The extreme C-terminal region of the CCK2R (and more precisely phosphorylatable residues Ser(437)-Xaa(438)-Thr(439)-Thr(440)-Xaa(441)-Ser(442)-Thr(443)) were critical for β-arrestin recruitment. However, this region and β-arrestins were dispensable for CCK2R internalization. In conclusion, this study allowed us to classify the human CCK2R as a member of class B G-protein-coupled receptors with regard to its endocytosis features and identified biased agonists of the CCK2R. These new important insights will allow us to investigate the role of internalized CCK2R·β-arrestin complexes in cancers expressing this receptor and to develop new diagnosis and therapeutic strategies targeting this receptor.

  18. Phase 2 trial of erlotinib with or without PF-3512676 (CPG 7909, a Toll-like receptor 9 agonist) in patients with advanced recurrent EGFR-positive non-small cell lung cancer

    PubMed Central

    Belani, Chandra P; Nemunaitis, John J; Chachoua, Abraham; Eisenberg, Peter D; Raez, Luiz E; Cuevas, J Daniel; Mather, Cecile B; Benner, Rebecca J; Meech, Sandra J

    2013-01-01

    This phase 2 study assessed PF-3512676 plus erlotinib in patients with epidermal growth factor receptor-positive advanced non-small cell lung cancer after prior chemotherapy failure. Patients were randomized 1:1 to PF-3512676 (0.20 mg/kg injected subcutaneously once weekly) plus erlotinib (150 mg daily) or erlotinib alone. The primary objective was to estimate progression-free survival (PFS). Patients received PF-3512676 plus erlotinib (n = 18) or erlotinib alone (n = 21). The study was halted because an unplanned interim analysis indicated that large improvement in PFS with addition of PF-3512676 would be unlikely. In the PF-3512676-plus-erlotinib and erlotinib-alone arms, median PFS was 1.6 and 1.7 mo (hazard ratio, 1.00; 95% confidence interval, 0.5–2.0; P = 0.9335), respectively. Salient grade ≥ 3 adverse events in PF-3512676-plus-erlotinib and erlotinib-alone arms were diarrhea (5/0), dyspnea (5/6), fatigue (4/1), other flu-like symptoms (2/0), anemia (2/1), and lymphocytopenia (based on laboratory values, 1/4). Adding PF-3512676 to erlotinib did not show potential for increased progression-free survival over erlotinib alone in patients with advanced recurrent epidermal growth factor receptor-positive non-small cell lung cancer. PMID:23792641

  19. Phase 2 trial of erlotinib with or without PF-3512676 (CPG 7909, a Toll-like receptor 9 agonist) in patients with advanced recurrent EGFR-positive non-small cell lung cancer.

    PubMed

    Belani, Chandra P; Nemunaitis, John J; Chachoua, Abraham; Eisenberg, Peter D; Raez, Luiz E; Cuevas, J Daniel; Mather, Cecile B; Benner, Rebecca J; Meech, Sandra J

    2013-07-01

    This phase 2 study assessed PF-3512676 plus erlotinib in patients with epidermal growth factor receptor-positive advanced non-small cell lung cancer after prior chemotherapy failure. Patients were randomized 1:1 to PF-3512676 (0.20 mg/kg injected subcutaneously once weekly) plus erlotinib (150 mg daily) or erlotinib alone. The primary objective was to estimate progression-free survival (PFS). Patients received PF-3512676 plus erlotinib (n = 18) or erlotinib alone (n = 21). The study was halted because an unplanned interim analysis indicated that large improvement in PFS with addition of PF-3512676 would be unlikely. In the PF-3512676-plus-erlotinib and erlotinib-alone arms, median PFS was 1.6 and 1.7 mo (hazard ratio, 1.00; 95% confidence interval, 0.5-2.0; P = 0.9335), respectively. Salient grade ≥ 3 adverse events in PF-3512676-plus-erlotinib and erlotinib-alone arms were diarrhea (5/0), dyspnea (5/6), fatigue (4/1), other flu-like symptoms (2/0), anemia (2/1), and lymphocytopenia (based on laboratory values, 1/4). Adding PF-3512676 to erlotinib did not show potential for increased progression-free survival over erlotinib alone in patients with advanced recurrent epidermal growth factor receptor-positive non-small cell lung cancer.

  20. The glycine transport inhibitor sarcosine is an NMDA receptor co-agonist that differs from glycine

    PubMed Central

    Zhang, Hai Xia; Hyrc, Krzysztof; Thio, Liu Lin

    2009-01-01

    Sarcosine is an amino acid involved in one-carbon metabolism and a promising therapy for schizophrenia because it enhances NMDA receptor (NMDAR) function by inhibiting glycine uptake. The structural similarity between sarcosine and glycine led us to hypothesize that sarcosine is also an agonist like glycine. We examined this possibility using whole-cell recordings from cultured embryonic mouse hippocampal neurons. We found that sarcosine is an NMDAR co-agonist at the glycine binding site. However, sarcosine differed from glycine because less NMDAR desensitization occurred with sarcosine than with glycine as the co-agonist. This finding led us to examine whether the physiological effects of NMDAR activation with these two co-agonists are the same. The difference in desensitization probably accounts for rises in intracellular Ca2+, as assessed by the fluorescent indicator fura-FF, being larger when NMDAR activation occurred with sarcosine than with glycine. In addition, Ca2+-activated K+ currents following NMDAR activation were larger with sarcosine than with glycine. Compared to glycine, NMDAR-mediated autaptic currents decayed faster with sarcosine suggesting that NMDAR deactivation also differs with these two co-agonists. Despite these differences, NMDAR-dependent neuronal death as assessed by propidium iodide was similar with both co-agonists. The same was true for neuronal bursting. Thus, sarcosine may enhance NMDAR function by more than one mechanism and may have different effects from other NMDAR co-agonists. PMID:19433577

  1. Synergy of interleukin 10 and toll-like receptor 9 signalling in B cell proliferation: Implications for lymphoma pathogenesis.

    PubMed

    Feist, Maren; Kemper, Judith; Taruttis, Franziska; Rehberg, Thorsten; Engelmann, Julia C; Gronwald, Wolfram; Hummel, Michael; Spang, Rainer; Kube, Dieter

    2017-03-01

    A network of autocrine and paracrine signals defines B cell homeostasis and is thought to be involved in transformation processes. Investigating interactions of these microenvironmental factors and their relation to proto-oncogenes as c-Myc (MYC) is fundamental to understand the biology of B cell lymphoma. Therefore, B cells with conditional MYC expression were stimulated with CD40L, insulin-like growth factor 1, α-IgM, Interleukin-10 (IL10) and CpG alone or in combination. The impact of forty different interventions on cell proliferation was investigated in MYC deprived cells and calculated by linear regression. Combination of CpG and IL10 led to a strong synergistic activation of cell proliferation (S-phase/doubling of total cell number) comparable to cells with high MYC expression. A synergistic up-regulation of CDK4, CDK6 and CCND3 expression by IL10 and CpG treatment was causal for this proliferative effect as shown by qRT-PCR analysis and inhibition of the CDK4/6 complex by PD0332991. Furthermore, treatment of stimulated MYC deprived cells with MLN120b, ACHP, Pyridone 6 or Ruxolitinib showed that IL10/CpG induced proliferation and CDK4 expression were JAK/STAT3 and IKK/NF-κB dependent. This was further supported by STAT3 and p65/RELA knockdown experiments, showing strongest effects on cell proliferation and CDK4 expression after double knockdown. Additionally, chromatin immunoprecipitation revealed a dual binding of STAT3 and p65 to the proximal promotor of CDK4 after IL10/CpG treatment. Therefore, the observed synergism of IL10R and TLR9 signalling was able to induce proliferation in a comparable way as aberrant MYC and might play a role in B cell homeostasis or transformation. © 2016 UICC.

  2. [Effects of agonists and antagonists of benzodiazepine, GABA and NMDA receptors, on caffeine-induced seizures in mice].

    PubMed

    Inano, S

    1992-08-01

    In mice, tonic convulsive seizure induced by intravenous administration of caffeine (adenosine A1, A2 receptors antagonist) was significantly potentiated by any one of L-PIA (adenosine A1 receptor agonist), NECA (adenosine A2 receptor agonist) and 2-ClAd (adenosine A1, A2 receptors agonist). The caffeine-induced seizure was unaffected by diazepam (benzodiazepine receptor agonist), but was inhibited by Ro 15-1788 (antagonist or partial agonist). beta-DMCM (antagonist or inverse agonist) increased the seizure. Muscimol (GABA-a receptor agonist), baclofen (GABA-b receptor agonist) and AOAA (GABA transaminase inhibitor) did not show significant effect on caffeine-induced convulsion. Bicuculline (GABA-a receptor antagonist) and picrotoxin (chloride channel blocker) significantly potentiated the convulsion at the doses which did not induce it. Caffeine-induced convulsion was potentiated by NMDA with its non-convulsive dose. CPP (competitive NMDA receptor antagonist) and MK-801 (non-competitive NMDA receptor antagonist) significantly inhibited the seizures. These results suggest that caffeine-induced seizure is not caused by blockade of adenosine receptors. Caffeine may act to beta-carboline sensitive benzodiazepine receptor (Type 1) which has no linkage with GABA-a receptor. Furthermore, it is implied that caffeine plays some role at NMDA receptor calcium ion channel complex.

  3. Kappa Opioid Receptor Agonist and Brain Ischemia

    PubMed Central

    Chunhua, Chen; Chunhua, Xi; Megumi, Sugita; Renyu, Liu

    2014-01-01

    Opioid receptors, especially Kappa opioid receptor (KOR) play an important role in the pathophysiological process of cerebral ischemia reperfusion injury. Previously accepted KOR agonists activity has included anti-nociception, cardiovascular, anti-pruritic, diuretic, and antitussive effects, while compelling evidence from various ischemic animal models indicate that KOR agonist have neuroprotective effects through various mechanisms. In this review, we aimed to demonstrate the property of KOR agonist and its role in global and focal cerebral ischemia. Based on current preclinical research, the KOR agonists may be useful as a neuroprotective agent. The recent discovery of salvinorin A, highly selective non-opioid KOR agonist, offers a new tool to study the role of KOR in brain HI injury and the protective effects of KOR agonist. The unique pharmacological profile of salvinorin A along with the long history of human usage provides its high candidacy as a potential alternative medication for brain HI injury. PMID:25574482

  4. Kappa Opioid Receptor Agonist and Brain Ischemia.

    PubMed

    Chunhua, Chen; Chunhua, Xi; Megumi, Sugita; Renyu, Liu

    2014-01-01

    Opioid receptors, especially Kappa opioid receptor (KOR) play an important role in the pathophysiological process of cerebral ischemia reperfusion injury. Previously accepted KOR agonists activity has included anti-nociception, cardiovascular, anti-pruritic, diuretic, and antitussive effects, while compelling evidence from various ischemic animal models indicate that KOR agonist have neuroprotective effects through various mechanisms. In this review, we aimed to demonstrate the property of KOR agonist and its role in global and focal cerebral ischemia. Based on current preclinical research, the KOR agonists may be useful as a neuroprotective agent. The recent discovery of salvinorin A, highly selective non-opioid KOR agonist, offers a new tool to study the role of KOR in brain HI injury and the protective effects of KOR agonist. The unique pharmacological profile of salvinorin A along with the long history of human usage provides its high candidacy as a potential alternative medication for brain HI injury.

  5. Alpha2-adrenergic receptor agonists as analgesics.

    PubMed

    Boyd, R E

    2001-08-01

    Alpha2-adrenergic receptor agonists are analgesic agents, and the alpha2-adrenergic agonist clonidine has been used in clinical studies for regional analgesia after intrathecal administration. We review here recent developments concerning the structure activity relationships of a new class of potent alpha2-adrenergic agonists and their use as analgesic agents. The effect of structure upon cardiovascular side-effects is also monitored, such as the prolongation of the QT portion of the cardiac action potential.

  6. The role of MAPK in CD4{sup +} T cells toll-like receptor 9-mediated signaling following HHV-6 infection

    SciTech Connect

    Chi, Jing; Wang, Fang; Li, Lingyun; Feng, Dongju; Qin, Jian; Xie, Fangyi; Zhou, Feng; Chen, Yun; Wang, Jinfeng; Yao, Kun

    2012-01-05

    Human herpesvirus-6 (HHV-6) is an important immunosuppressive and immunomodulatory virus that primarily infects immune cells (mainly CD4{sup +} T cells) and strongly suppresses the proliferation of infected cells. Toll-like receptors are pattern-recognition receptors essential for the development of an appropriate innate immune defense against infection. To understand the role of CD4{sup +} T cells in the innate response to HHV-6 infection and the involvement of TLRs, we used an in vitro infection model and observed that the infection of CD4{sup +} T cells resulted in the activation of JNK/SAPK via up-regulation of toll-like receptor 9 (TLR9). Associated with JNK activation, annexin V-PI staining indicated that HHV-6A was a strong inducer of apoptosis. Apoptotic response associated cytokines, IL-6 and TNF-{alpha} also induced by HHV-6A infection.

  7. Effect of topical betaxolol on the acute rise of aqueous flare induced by highly selective agonists for prostaglandin E2 receptor subtypes in pigmented rabbits.

    PubMed

    Yanagisawa, Shuichiro; Hayasaka, Seiji; Zhang, Xue-Yun; Hayasaka, Yoriko; Nagaki, Yasunori; Kitagawa, Kiyotaka

    2002-01-01

    To evaluate the role of topical betaxolol on experimental ocular inflammation in rabbits. Transcorneal diffusion of highly selective agonists for prostaglandin E2 receptor subtypes (EP), 25 microg/ml, with the use of a glass cylinder, was performed to produce aqueous flare elevation in pigmented rabbits. Betaxolol was topically administered before EP agonist application. Aqueous flare was measured with a laser flare cell meter. Performing topical instillation of 0.5% betaxolol 4 times inhibited 52 +/- 9% of EP2-agonist (ONO-AE1-259-01)-induced aqueous flare elevation. The inhibition of flare elevation was dependent on the number of betaxolol instillations. Betaxolol did not suppress the elevation induced by an EP4 agonist (ONO-AE1-392). Betaxolol inhibited EP2-agonist-induced aqueous flare elevation in pigmented rabbits. Copyright 2002 S. Karger AG, Basel

  8. Optimized combination therapy using bortezomib, TRAIL and TLR agonists in established breast tumors.

    PubMed

    Lee, Sujin; Yagita, Hideo; Sayers, Thomas J; Celis, Esteban

    2010-07-01

    TNF-related apoptosis-inducing ligand (TRAIL) is a member of the TNF family of cytokines, which can induce apoptosis in various tumor cells by engaging the receptors, DR4 and DR5. Bortezomib (Velcade) is a proteasome inhibitor that has been approved for patients with multiple myeloma. There is some experimental evidence in preclinical models that bortezomib can enhance the susceptibility of tumors to TRAIL-induced apoptosis. In this study, we investigated the effects of TRAIL-induced death using an agonistic antibody to the TRAIL receptor DR5 (alpha-DR5) in combination with bortezomib administered to mice previously injected with breast cancer cells (TUBO). This combination had some beneficial therapeutic effect, which was significantly enhanced by the co-administration of a Toll-like receptor 9 agonist (CpG). In contrast, single agent treatments had little effect on tumor growth. In addition, we evaluated the effect of combination with alpha-DR5, bortezomib, and CpG in the prevention/treatment of spontaneous mammary tumors in Balb-neuT mice. In this model, which is more difficult to treat, we observed dramatic antitumor effects of alpha-DR5, bortezomib and CpG combination therapy. Since such a mouse model more accurately reflects the immunological tolerance that exists in human cancer, our results strongly suggest that these combination strategies could be directly applied to the therapy for cancer patients.

  9. Dopamine receptor agonists, partial agonists and psychostimulant addiction.

    PubMed

    Pulvirenti, L; Koob, G F

    1994-10-01

    Despite the epidemic growth of psychostimulant addiction over the past years, few pharmacological means of intervention are available to date for clinical treatment. This is of importance since the withdrawal syndrome that follows abstinence from drugs such as cocaine and the amphetamines is characterized, among other symptoms, by intense craving for the abused drug, and this is considered a critical factor leading into relapse of drug use. In this article, Luigi Pulvirenti and George Koob focus on the modulatory role shown by drugs acting at the dopamine receptor on the various phases of psychostimulant dependence in preclinical models and in human studies, and suggest that a class of compounds with partial agonist properties at the dopamine receptor may have therapeutic potential.

  10. Myeloid cell leukaemia 1 has a vital role in retinoic acid-mediated protection of Toll-like receptor 9-stimulated B cells from spontaneous and DNA damage-induced apoptosis.

    PubMed

    Holm, Kristine L; Indrevaer, Randi L; Myklebust, June Helen; Kolstad, Arne; Moskaug, Jan Øivind; Naderi, Elin H; Blomhoff, Heidi K

    2016-09-01

    Vitamin A is an essential anti-infective agent with pleiotropic effects on cells of the immune system. The goal of the present study was to unravel the impact of the vitamin A metabolite retinoic acid (RA) on B-cell survival related both to normal B-cell homeostasis and to the detrimental effects imposed by DNA-damaging agents. By combining RA with Toll-like receptor 9 (TLR9) ligands, we show that RA prevents spontaneous, irradiation- and doxorubicin-induced apoptosis of human B cells in an RA receptor-dependent manner. RA-mediated survival involved up-regulation of the anti-apoptotic protein myeloid cell leukemia 1 (MCL1) at the transcriptional level, and knock down of MCL1 by small interfering RNA partially reversed the effects of RA. To ensure that the combination of TLR9-ligands and RA would not promote the survival of malignant B cells, the combined effects of stimulation with RA and TLR9 ligands was assessed on cells from patients with B-cell malignancies. In contrast to the effects on normal B cells, the combination of TLR9 stimulation and RA neither enhanced the MCL1 levels nor inhibited the death of malignant B cells challenged by DNA-damaging agents. Taken together, the present results reveal a vital role of MCL1 in RA-mediated survival of normal B cells. Moreover, the findings suggest that RA in combination with TLR9 ligands might be useful adjuvants in the treatment of B-cell malignancies by selectively protecting normal and not malignant B cells from DNA-damage-induced cell death. © 2016 John Wiley & Sons Ltd.

  11. Alternative Splicing of Toll-Like Receptor 9 Transcript in Teleost Fish Grouper Is Regulated by NF-κB Signaling via Phosphorylation of the C-Terminal Domain of the RPB1 Subunit of RNA Polymerase II

    PubMed Central

    Lee, Frank Fang-Yao; Hui, Cho-Fat; Chang, Tien-Hsien; Chiou, Pinwen Peter

    2016-01-01

    Similar to its mammalian counterparts, teleost Toll-like receptor 9 (TLR9) recognizes unmethylated CpG DNA presented in the genome of bacteria or DNA viruses and initiates signaling pathway(s) for immune responses. We have previously shown that the TLR9 pathway in grouper, an economically important teleost, can be debilitated by an inhibitory gTLR9B isoform, whose production is mediated by RNA alternative splicing. However, how does grouper TLR9 (gTLR9) signaling impinge on the RNA splicing machinery to produce gTlr9B is unknown. Here we show that the gTlr9 alternative splicing is regulated through ligand-induced phosphorylation of the C-terminal domain (CTD) of the largest subunit of RNA polymerase II (Pol II). We first observed that ligand-activated NF- κB pathway biased the production of the gTlr9B isoform. Because NF- κB is known to recruit p-TEFb kinase, which phosphorylates the Pol II CTD at Ser2 residues, we examined p-TEFb’s role in alternative splicing. We found that promoting p-TEFb kinase activity significantly favored the production of the gTlr9B isoform, whereas inhibiting p-TEFb yielded an opposite result. We further showed that p-TEFb-mediated production of the gTlr9B isoform down-regulates its own immune responses, suggesting a self-limiting mechanism. Taken together, our data indicate a feedback mechanism of the gTLR9 signaling pathway to regulate the alternative splicing machinery, which in turn produces an inhibitor to the pathway. PMID:27658294

  12. A Small Molecule Agonist of an Integrin,αLβ2*S

    PubMed Central

    Yang, Wei; Carman, Christopher V.; Kim, Minsoo; Salas, Azucena; Shimaoka, Motomu; Springer, Timothy A.

    2006-01-01

    The binding of integrin αLβ2 to its ligand intercellular adhesion molecule-1 is required for immune responses and leukocyte trafficking. Small molecule antagonists of αLβ2 are under intense investigation as potential anti-inflammatory drugs. We describe for the first time a small molecule integrin agonist. A previously described α/βI allosteric inhibitor, compound 4, functions as an agonist of αLβ2 in Ca2+ and Mg2+ and as an antagonist in Mn2+. We have characterized the mechanism of activation and its competitive and noncompetitive inhibition by different compounds. Although it stimulates ligand binding, compound 4 nonetheless inhibits lymphocyte transendothelial migration. Agonism by compound 4 results in accumulation of αLβ2 in the uropod, extreme uropod elongation, and defective de-adhesion. Small molecule integrin agonists open up novel therapeutic possibilities. PMID:17023419

  13. Dopamine agonists for cocaine dependence.

    PubMed

    Soares, B G; Lima, M S; Reisser, A A; Farrell, M

    2001-01-01

    Cocaine is a major drug of abuse. Cocaine dependence is a common and serious condition, which has become nowadays a substantial public health problem. There is a wide and well documented range of consequences associated to chronic use of this drug, such as medical, psychological and social problems, including the spread of infectious diseases (e.g. AIDS, hepatitis and tuberculosis), crime, violence and neonatal drug exposure. Therapeutic management of the cocaine addicts includes an initial period of abstinence from the drug. During this phase the subjects may experience, besides the intense craving for cocaine, symptoms such as depression, fatigue, irritability, anorexia, and sleep disturbances. It was demonstrated that the acute use of cocaine may enhance dopamine transmission and chronically it decreases dopamine concentrations in the brain. Pharmacological treatment that affects dopamine could theoretically reduce these symptoms and contribute to a more successful therapeutic approach. To evaluate the efficacy and acceptability of dopamine agonists for treating cocaine dependence. We searched: The Cochrane Controlled Trials Register (Cochrane Library, issue 4, 2000), MEDLINE (from 1966 - 2000), EMBASE (from 1980 - 2000), LILACS (from 1982 - 2000), PsycLIT (from 1974 - 2000), Biological Abstracts (1982 to 2000). Reference searching; personal communication; conference abstracts; unpublished trials from pharmaceutical industry; book chapters on treatment of cocaine dependence. The inclusion criteria for all randomised controlled trials were that they should focus on the use of dopamine agonists on the treatment of cocaine dependence. Trials including patients with additional diagnosis such as opiate dependence were also eligible. The reviewers extracted the data independently and Relative Risks, weighted mean difference and number needed to treat were estimated. The reviewers assumed that people who died or dropped out had no improvement and tested the sensitivity

  14. Dopamine agonists for cocaine dependence.

    PubMed

    Soares, B G O; Lima, M S; Reisser, A A P; Farrell, M

    2003-01-01

    Cocaine dependence is a common and serious condition, which has become nowadays a substantial public health problem. There is a wide and well documented range of consequences associated to chronic use of this drug, such as medical, psychological and social problems, including the spread of infectious diseases (e.g. AIDS, hepatitis and tuberculosis), crime, violence and neonatal drug exposure. Therapeutic management of the cocaine addicts includes an initial period of abstinence from the drug. During this phase the subjects may experience, besides the intense craving for cocaine, symptoms such as depression, fatigue, irritability, anorexia, and sleep disturbances. It was demonstrated that the acute use of cocaine may enhance dopamine transmission and chronically it decreases dopamine concentrations in the brain. Pharmacological treatment that affects dopamine could theoretically reduce these symptoms and contribute to a more successful therapeutic approach. To evaluate the efficacy and acceptability of dopamine agonists for treating cocaine dependence. Electronic searches of Cochrane Library, EMBASE, MEDLINE, PsycLIT, Biological Abstracts and LILACS; reference searching; personal communication; conference abstracts; unpublished trials from pharmaceutical industry; book chapters on treatment of cocaine dependence, was performed for the primary version of this review in 2001. Another search of the electronic databases was done in December of 2002 for this update. The specialised register of trials of the Cochrane Group on Drugs and Alcohol was searched until February 2003. The inclusion criteria for all randomised controlled trials were that they should focus on the use of dopamine agonists on the treatment of cocaine dependence. The reviewers extracted the data independently and Relative Risks, weighted mean difference and number needed to treat were estimated. The reviewers assumed that people who died or dropped out had no improvement and tested the sensitivity of

  15. Purines, a new class of agonists in salivary glands?

    PubMed

    Dehaye, J P; Moran, A; Marino, A

    1999-05-01

    The response of rat submandibular glands to extracellular purines was tested. In crude cellular suspensions, ATP increased the [Ca2+]i mostly by promoting uptake of extracellular calcium. ATP caused the pHi to drop, a response blocked by chloride channel inhibitors. ATP also inhibited the basal and isoproterenol-stimulated activity of the Na+ -K+ -2Cl-cotransporter. These effects were reproduced by benzoyl-ATP, an agonist of ionotropic purinoceptors. In pure ductal suspensions, ATP activated a metabotropic P2Y1 purinergic receptor coupled to phospholipase C and opened a non-specific cation channel coupled to a P2X7 receptor. Activation of these receptors stimulated a Ca2+ -dependent and a Ca2+ -independent phospholipase A2, the latter resulting in kallikrein secretion. We conclude that purinergic agonists can modulate the activity of both acinar and ductal phases of secretion. Activation of metabotropic receptors coupled to phospholipase C could lead to responses resembling those to muscarinic or adrenergic agonists. Activation of ionotropic receptors could stimulate new intracellular responses also involved in secretory function.

  16. Beta-agonists and animal welfare

    USDA-ARS?s Scientific Manuscript database

    The use of beta-agonists in animal feed is a high profile topic within the U.S. as consumers and activist groups continue to question its safety. The only beta-agonist currently available for use in swine is ractopamine hydrochloride (RAC). This is available as Paylean™ (Elanco Animal Health – FDA a...

  17. Physostigmine, galanthamine and codeine act as 'noncompetitive nicotinic receptor agonists' on clonal rat pheochromocytoma cells.

    PubMed

    Storch, A; Schrattenholz, A; Cooper, J C; Abdel Ghani, E M; Gutbrod, O; Weber, K H; Reinhardt, S; Lobron, C; Hermsen, B; Soskiç, V

    1995-08-15

    The acetylcholine esterase inhibitor (-)-physostigmine has been shown to act as agonist on nicotinic acetylcholine receptors from muscle and brain, by binding to sites on the alpha-polypeptide that are distinct from those for the natural transmitter acetylcholine (Schröder et al., 1994). In the present report we show that (-)-physostigmine, galanthamine, and the morphine derivative codeine activate single-channel currents in outside-out patches excised from clonal rat pheochromocytoma (PC12) cells. Although several lines of evidence demonstrate that the three alkaloids act on the same channels as acetylcholine, the competitive nicotinic antagonist methyllycaconitine only inhibited channel activation by acetylcholine but not by (-)-physostigmine, galanthamine or codeine. In contrast, the monoclonal antibody FK1, which competitively inhibits (-)-physostigmine binding to nicotinic acetylcholine receptors, did not affect channel activation by acetylcholine but inhibited activation by (-)-physostigmine, galanthamine and codeine. The three alkaloids therefore act via binding sites distinct from those for acetylcholine, in a 'noncompetitive' fashion. The potency of (-)-physostigmine and related compounds to act as a noncompetitive agonist is unrelated to the level of acetylcholine esterase inhibition induced by these drugs. (-)-Physostigmine, galanthamine and codeine do not evoke sizable whole-cell currents, which is due to the combined effects of low open-channel probability, slow onset and slow inactivation of response. In contrast, they sensitize PC12 cell nicotinic receptors in their submaximal response to acetylcholine. While the abundance of nicotinic acetylcholine receptor isoforms expressed in PC12 cells excludes identification of specific nicotinic acetylcholine receptor subtypes that interact with noncompetitive agonists, the identical patterns of single-channel current amplitudes observed with acetylcholine and with noncompetitive agonists suggested that all

  18. Agonist-Specific Recruitment of Arrestin Isoforms Differentially Modify Delta Opioid Receptor Function

    PubMed Central

    Perroy, Julie; Walwyn, Wendy M.; Smith, Monique L.; Vicente-Sanchez, Ana; Segura, Laura; Bana, Alia; Kieffer, Brigitte L.; Evans, Christopher J.

    2016-01-01

    Ligand-specific recruitment of arrestins facilitates functional selectivity of G-protein-coupled receptor signaling. Here, we describe agonist-selective recruitment of different arrestin isoforms to the delta opioid receptor in mice. A high-internalizing delta opioid receptor agonist (SNC80) preferentially recruited arrestin 2 and, in arrestin 2 knock-outs (KOs), we observed a significant increase in the potency of SNC80 to inhibit mechanical hyperalgesia and decreased acute tolerance. In contrast, the low-internalizing delta agonists (ARM390, JNJ20788560) preferentially recruited arrestin 3 with unaltered behavioral effects in arrestin 2 KOs. Surprisingly, arrestin 3 KO revealed an acute tolerance to these low-internalizing agonists, an effect never observed in wild-type animals. Furthermore, we examined delta opioid receptor–Ca2+ channel coupling in dorsal root ganglia desensitized by ARM390 and the rate of resensitization was correspondingly decreased in arrestin 3 KOs. Live-cell imaging in HEK293 cells revealed that delta opioid receptors are in pre-engaged complexes with arrestin 3 at the cell membrane and that ARM390 strengthens this membrane interaction. The disruption of these complexes in arrestin 3 KOs likely accounts for the altered responses to low-internalizing agonists. Together, our results show agonist-selective recruitment of arrestin isoforms and reveal a novel endogenous role of arrestin 3 as a facilitator of resensitization and an inhibitor of tolerance mechanisms. SIGNIFICANCE STATEMENT Agonists that bind to the same receptor can produce highly distinct signaling events and arrestins are a major mediator of this ligand bias. Here, we demonstrate that delta opioid receptor agonists differentially recruit arrestin isoforms. We found that the high-internalizing agonist SNC80 preferentially recruits arrestin 2 and knock-out (KO) of this protein results in increased efficacy of SNC80. In contrast, low-internalizing agonists (ARM390 and JNJ20788560

  19. Agonist-Specific Recruitment of Arrestin Isoforms Differentially Modify Delta Opioid Receptor Function.

    PubMed

    Pradhan, Amynah A; Perroy, Julie; Walwyn, Wendy M; Smith, Monique L; Vicente-Sanchez, Ana; Segura, Laura; Bana, Alia; Kieffer, Brigitte L; Evans, Christopher J

    2016-03-23

    Ligand-specific recruitment of arrestins facilitates functional selectivity of G-protein-coupled receptor signaling. Here, we describe agonist-selective recruitment of different arrestin isoforms to the delta opioid receptor in mice. A high-internalizing delta opioid receptor agonist (SNC80) preferentially recruited arrestin 2 and, in arrestin 2 knock-outs (KOs), we observed a significant increase in the potency of SNC80 to inhibit mechanical hyperalgesia and decreased acute tolerance. In contrast, the low-internalizing delta agonists (ARM390, JNJ20788560) preferentially recruited arrestin 3 with unaltered behavioral effects in arrestin 2 KOs. Surprisingly, arrestin 3 KO revealed an acute tolerance to these low-internalizing agonists, an effect never observed in wild-type animals. Furthermore, we examined delta opioid receptor-Ca(2+)channel coupling in dorsal root ganglia desensitized by ARM390 and the rate of resensitization was correspondingly decreased in arrestin 3 KOs. Live-cell imaging in HEK293 cells revealed that delta opioid receptors are in pre-engaged complexes with arrestin 3 at the cell membrane and that ARM390 strengthens this membrane interaction. The disruption of these complexes in arrestin 3 KOs likely accounts for the altered responses to low-internalizing agonists. Together, our results show agonist-selective recruitment of arrestin isoforms and reveal a novel endogenous role of arrestin 3 as a facilitator of resensitization and an inhibitor of tolerance mechanisms. Agonists that bind to the same receptor can produce highly distinct signaling events and arrestins are a major mediator of this ligand bias. Here, we demonstrate that delta opioid receptor agonists differentially recruit arrestin isoforms. We found that the high-internalizing agonist SNC80 preferentially recruits arrestin 2 and knock-out (KO) of this protein results in increased efficacy of SNC80. In contrast, low-internalizing agonists (ARM390 and JNJ20788560) preferentially recruit

  20. Thermodynamic analysis of antagonist and agonist interactions with dopamine receptors.

    PubMed

    Duarte, E P; Oliveira, C R; Carvalho, A P

    1988-03-01

    The binding of [3H]spiperone to dopamine D-2 receptors and its inhibition by antagonists and agonists were examined in microsomes derived from the sheep caudate nucleus, at temperatures between 37 and 1 degree C, and the thermodynamic parameters of the binding were evaluated. The affinity of the receptor for the antagonists, spiperone and (+)-butaclamol, decreased as the incubation temperature decreased; the affinity for haloperidol did not further decrease at temperatures below 15 degrees C. The binding of the antagonists was associated with very large increases in entropy, as expected for hydrophobic interactions. The enthalpy and entropy changes associated with haloperidol binding were dependent on temperature, in contrast to those associated with spiperone and (+)-butaclamol. The magnitude of the entropy increase associated with the specific binding of the antagonists did not correlate with the degree of lipophilicity of these drugs. The data suggest that, in addition to hydrophobic forces, other forces are also involved in the antagonist-dopamine receptor interactions, and that a conformational change of the receptor could occur when the antagonist binds. Agonist binding data are consistent with a two-state model of the receptor, a high-affinity state (RH) and a low-affinity state (RL). The affinity of dopamine binding to the RH decreased with decreasing temperatures below 20 degrees C, whereas the affinity for the RL increased at low temperatures. In contrast, the affinity of apomorphine for both states of receptor decreased as the temperature decreased from 30 to 8 degrees C. A clear distinction between the energetics of high-affinity and low-affinity agonist binding was observed. The formation of the high-affinity complex was associated with larger increases in enthalpy and entropy than the interaction with the low-affinity state was. The results suggest that the interaction of the receptor with the G-proteins, induced or stabilized by the binding of

  1. The actions of prolonged exposure to cholinergic agonists on isolated bladder strips from the rat.

    PubMed

    Gillespie, James I; Rouget, Celine; Palea, Stefano; Korstanje, Cees

    2015-07-01

    The present study was done to explore the cholinergic systems operating in the wall of the isolated rat bladder. In a first set of experiments, bladder strips in vitro were subjected to cumulative concentration-response curve (CRC) to non-selective muscarine agonist carbachol or the partially M2>M3 selective agonist arecaidine to establish optimal concentration to be used thereafter. In a second set of experiments, the effects of drugs (solifenacin, isoproterenol, and mirabegron) were tested on urinary bladder contraction induced by the non-selective muscarinergic agonist carbachol. For both agonists, the contractile responses are qualitatively similar: an initial transient rise in tension followed by complex bursts of high-frequency small 'micro'-contractions superposed on a tonic contraction, with immediate transient 'rebound' contraction after the agonist is washed from the preparation. This rebound contraction is greater with carbachol than arecaidine. Components of the responses to cholinergic stimulation, notably the micro-contractions, were found to be differently stimulated and inhibited by the M3>M2 selective antagonist solifenacin and by the β-adrenoceptor agonists isoprenaline and mirabegron. A physiological role for the muscarinic dependent phasic contractions and the micro-anatomical elements that might be involved are not known but may be related to non-voiding activity observed during filling cystometry in conscious animals related to afferent discharge and possibly sensation. Furthermore, suggestions for the potential impact of these findings and design of further studies in relation to bladder physiology, pharmacology, and pathology are discussed.

  2. Analysis of the agonist activity of fenoldopam (SKF 82526) at the vascular 5-HT2 receptor.

    PubMed Central

    Christie, M. I.; Harper, D.; Smith, G. W.

    1992-01-01

    1. The 5-HT2 receptor agonist activity of fenoldopam (SKF 82526) was characterized in the rabbit isolated aorta preparation. 2. Fenoldopam was an agonist at the vascular 5-HT2 receptor with lower affinity and efficacy than the naturally occurring agonist 5-hydroxytryptamine (5-HT). Fenoldopam had an affinity (pKA) of 5.84 +/- 0.04 and efficacy (tau) of 0.57 +/- 0.04, whereas 5-HT had a pKA of 6.65 +/- 0.12 and tau of 2.66 +/- 0.41. 3. The constrictor effects of fenoldopam and 5-HT were competitively antagonized by the 5-HT2 antagonist, ketanserin, with pKB values of 8.81 +/- 0.11 and 8.83 +/- 0.10 respectively. 4. Prior incubation with fenoldopam produced a concentration-related rightward shift of a subsequent 5-HT concentration-response curve. This inhibition was specific for 5-HT since constrictor responses to angiotensin II were unaffected. 5. This study indicates that the D1 receptor agonist, fenoldopam, acts as an agonist at the vascular 5-HT2 receptor, but with an affinity and efficacy less than that of the naturally occurring agonist, 5-HT. PMID:1361397

  3. In Vitro Effects of Beta-2 Agonists on Skeletal Muscle Differentiation, Hypertrophy, and Atrophy

    PubMed Central

    2012-01-01

    Background Beta-2 agonists are widely used in the treatment of asthma and chronic obstructive pulmonary disease for their effect on airway smooth muscle relaxation. They also act on skeletal muscle, although their reported ergogenic effect is controversial. Aim To evaluate the in vitro effects of short-acting and long-acting beta-2 agonists on adrenergic receptor (ADR) expression, hypertrophy, and atrophy markers, in a skeletal muscle cell line. Methods The C2C12 cell line was used as a model of skeletal muscle differentiation. ADR messenger RNA expression was evaluated in proliferating myoblasts, committed cells, and differentiated myotubes, in basal conditions and after treatment with 10-6 M clenbuterol, salbutamol, salmeterol, and formoterol. Effect of beta-2 agonists on gene and protein expression of hypertrophy and atrophy markers was assessed in differentiated myotubes. Results Our study shows that beta-2 ADR messenger RNA was expressed and progressively increased during cell differentiation. Beta-2 agonist treatment did not affect its expression. Skeletal muscle hypertrophy markers (fast and slow myosin, myogenin) were not modulated by any of the beta-2 agonists evaluated. However, clenbuterol induced a significant, dose-dependent downregulation of skeletal muscle atrophy genes (atrogin-1, MuRF-1, and cathepsin L). Conclusions The reported ergogenic effect of beta-2 agonists, if any, should be considered as drug-specific and not class-specific and that of clenbuterol is mediated by the inhibition of the atrophic pathway. PMID:23283108

  4. Combined sodium ion sensitivity in agonist binding and internalization of vasopressin V1b receptors.

    PubMed

    Koshimizu, Taka-Aki; Kashiwazaki, Aki; Taniguchi, Junichi

    2016-05-03

    Reducing Na(+) in the extracellular environment may lead to two beneficial effects for increasing agonist binding to cell surface G-protein coupled receptors (GPCRs): reduction of Na(+)-mediated binding block and reduce of receptor internalization. However, such combined effects have not been explored. We used Chinese Hamster Ovary cells expressing vasopressin V1b receptors as a model to explore Na(+) sensitivity in agonist binding and receptor internalization. Under basal conditions, a large fraction of V1b receptors is located intracellularly, and a small fraction is in the plasma membrane. Decreases in external Na(+) increased cell surface [(3)H]AVP binding and decreased receptor internalization. Substitution of Na(+) by Cs(+) or NH4(+) inhibited agonist binding. To suppress receptor internalization, the concentration of NaCl, but not of CsCl, had to be less than 50 mM, due to the high sensitivity of the internalization machinery to Na(+) over Cs(+). Iso-osmotic supplementation of glucose or NH4Cl maintained internalization of the V1b receptor, even in a low-NaCl environment. Moreover, iodide ions, which acted as a counter anion, inhibited V1b agonist binding. In summary, we found external ionic conditions that could increase the presence of high-affinity state receptors at the cell surface with minimum internalization during agonist stimulations.

  5. Combined sodium ion sensitivity in agonist binding and internalization of vasopressin V1b receptors

    PubMed Central

    Koshimizu, Taka-aki; Kashiwazaki, Aki; Taniguchi, Junichi

    2016-01-01

    Reducing Na+ in the extracellular environment may lead to two beneficial effects for increasing agonist binding to cell surface G-protein coupled receptors (GPCRs): reduction of Na+-mediated binding block and reduce of receptor internalization. However, such combined effects have not been explored. We used Chinese Hamster Ovary cells expressing vasopressin V1b receptors as a model to explore Na+ sensitivity in agonist binding and receptor internalization. Under basal conditions, a large fraction of V1b receptors is located intracellularly, and a small fraction is in the plasma membrane. Decreases in external Na+ increased cell surface [3H]AVP binding and decreased receptor internalization. Substitution of Na+ by Cs+ or NH4+ inhibited agonist binding. To suppress receptor internalization, the concentration of NaCl, but not of CsCl, had to be less than 50 mM, due to the high sensitivity of the internalization machinery to Na+ over Cs+. Iso-osmotic supplementation of glucose or NH4Cl maintained internalization of the V1b receptor, even in a low-NaCl environment. Moreover, iodide ions, which acted as a counter anion, inhibited V1b agonist binding. In summary, we found external ionic conditions that could increase the presence of high-affinity state receptors at the cell surface with minimum internalization during agonist stimulations. PMID:27138239

  6. Evaluation of anti-diabetic effect and gall bladder function with 2-thio-5-thiomethyl substituted imidazoles as TGR5 receptor agonists.

    PubMed

    Zhang, Xuqing; Sui, Zhihua; Kauffman, Jack; Hou, Cuifen; Chen, Cailin; Du, Fuyong; Kirchner, Thomas; Liang, Yin; Johnson, Dana; Murray, William V; Demarest, Keith

    2017-04-15

    A novel series of 2-thio-5-thiomethyl substituted imidazoles was discovered to be potent TGR5 agonists that possessed glucose-lowering effects while inhibiting gall bladder emptying in mice. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Therapeutic utility of natural estrogen receptor beta agonists on ovarian cancer.

    PubMed

    Liu, Jinyou; Viswanadhapalli, Suryavathi; Garcia, Lauren; Zhou, Mei; Nair, Binoj C; Kost, Edward; Rao Tekmal, Rajeshwar; Li, Rong; Rao, Manjeet K; Curiel, Tyler; Vadlamudi, Ratna K; Sareddy, Gangadhara R

    2017-07-25

    Ovarian cancer is the deadliest of all gynecologic cancers. Despite success with initial chemotherapy, the majority of patients relapse with an incurable disease. Development of chemotherapy resistance is a major factor for poor long-term survival in ovarian cancer. The biological effects of estrogens are mediated by estrogen receptor alpha (ERα) and estrogen receptor beta (ERβ). Emerging evidence suggests that ovarian cancer cells express ERβ that functions as a tumor suppressor; however, the clinical utility of ERβ agonists in ovarian cancer remains elusive. We tested the utility of two natural ERβ agonists liquiritigenin (Liq), which is isolated from Glycyrrhiza uralensis and S-equol, which is isolated from soy isoflavone daidzein, for treating ovarian cancer. Both natural ERβ ligands had significant growth inhibition in cell viability and survival assays, reduced migration and invasion, and promoted apoptosis. Further, ERβ agonists showed tumor suppressive functions in therapy-resistant ovarian cancer model cells and sensitized ovarian cancer cells to cisplatin and paclitaxel treatment. Global RNA-Seq analysis revealed that ERβ agonists modulate several tumor suppressive pathways, including downregulation of the NF-κB pathway. Immunoprecipitation assays revealed that ERβ interacts with p65 subunit of NF-κB and ERβ overexpression reduced the expression of NF-κB target genes. In xenograft assays, ERβ agonists reduced tumor growth and promoted apoptosis. Collectively, our findings demonstrated that natural ERβ agonists have the potential to significantly inhibit ovarian cancer cell growth by anti-inflammatory and pro-apoptotic actions, and natural ERβ agonists represent novel therapeutic agents for the management of ovarian cancer.

  8. Investigation of the mechanism of agonist and inverse agonist action at D2 dopamine receptors.

    PubMed

    Roberts, David J; Lin, Hong; Strange, Philip G

    2004-05-01

    This study investigated, for the D2 dopamine receptor, the relation between the ability of agonists and inverse agonists to stabilise different states of the receptor and their relative efficacies. Ki values for agonists were determined in competition versus the binding of the antagonist [3H]spiperone. Competition data were fitted best by a two-binding site model (with the exception of bromocriptine, for which a one-binding site model provided the best fit) and agonist affinities for the higher (Kh) (G protein-coupled) and lower affinity (Kl) (G protein-uncoupled) sites determined. Ki values for agonists were also determined in competition versus the binding of the agonist [3H]N-propylnorapomorphine (NPA) to provide a second estimate of Kh. Maximal agonist effects (Emax) and their potencies (EC50) were determined from concentration-response curves for agonist stimulation of guanosine-5'-O-(3-[32S]thiotriphosphate) ([35S]GTPgammaS) binding. The ability of agonists to stabilise the G protein-coupled state of the receptor (Kl/Kh determined from ligand-binding assays) did not correlate with either of two measures of relative efficacy (relative Emax, Kl/EC50) of agonists determined in [35S]GTPgammaS-binding assays, when the data for all of the compounds tested were analysed. For a subset of compounds, however, there was a relation between Kl/Kh and Emax. Competition-binding data versus [3H]spiperone and [3H]NPA for a range of inverse agonists were fitted best by a one-binding site model. Ki values for the inverse agonists tested were slightly lower in competition versus [3H]NPA compared to [3H]spiperone. These data do not provide support for the idea that inverse agonists act by binding preferentially to the ground state of the receptor.

  9. Muscarinic Receptor Agonists and Antagonists: Effects on Cancer

    PubMed Central

    2012-01-01

    Many epithelial and endothelial cells express a cholinergic autocrine loop in which acetylcholine acts as a growth factor to stimulate cell growth. Cancers derived from these tissues similarly express a cholinergic autocrine loop and ACh secreted by the cancer or neighboring cells interacts with M3 muscarinic receptors expressed on the cancer cells to stimulate tumor growth. Primary proliferative pathways involve MAPK and Akt activation. The ability of muscarinic agonists to stimulate, and M3 antagonists to inhibit tumor growth has clearly been demonstrated for lung and colon cancer. The ability of muscarinic agonists to stimulate growth has been shown for melanoma, pancreatic, breast, ovarian, prostate and brain cancers, suggesting that M3 antagonists will also inhibit growth of these tumors as well. As yet no clinical trials have proven the efficacy of M3 antagonists as cancer therapeutics, though the widespread clinical use and low toxicity of M3 antagonists support the potential role of these drugs as adjuvants to current cancer therapies. PMID:22222710

  10. The pharmacologic basis for clinical differences among GLP-1 receptor agonists and DPP-4 inhibitors.

    PubMed

    Morales, Javier

    2011-11-01

    The incretin system plays an important role in glucose homeostasis, largely through the actions of glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1). Unlike GIP, the actions of GLP-1 are preserved in patients with type 2 diabetes mellitus, which has led to the development of injectable GLP-1 receptor (GLP-1R) agonists and oral dipeptidyl peptidase-4 (DPP-4) inhibitors. GLP-1R agonists-which can be dosed to pharmacologic levels-act directly upon the GLP-1R. In contrast, DPP-4 inhibitors work indirectly by inhibiting the enzymatic inactivation of native GLP-1, resulting in a modest increase in endogenous GLP-1 levels. GLP-1R agonists generally lower the fasting and postprandial glucose levels more than DPP-4 inhibitors, resulting in a greater mean reduction in glycated hemoglobin level with GLP-1R agonists (0.4%-1.7%) compared with DPP-4 inhibitors (0.4%-1.0%). GLP-1R agonists also promote satiety and reduce total caloric intake, generally resulting in a mean weight loss of 1 to 4 kg over several months in most patients, whereas DPP-4 inhbitors are weight-neutral overall. GLP-1R agonists and DPP-4 inhibitors are generally safe and well tolerated. The glucose-dependent manner of stimulation of insulin release and inhibition of glucagon secretion by both GLP-1R agonists and DPP-4 inhibitors contribute to the low incidence of hypoglycemia. Although transient nausea occurs in 26% to 28% of patients treated with GLP-1R agonists but not DPP-4 inhibitors, this can be reduced by using a dose-escalation strategy. Other adverse events (AEs) associated with GLP-1R agonists include diarrhea, headache, and dizziness. The main AEs associated with DPP-4 inhibitors include upper respiratory tract infection, nasopharyngitis, and headache. Overall, compared with other therapies for type 2 diabetes mellitus with similar efficacy, incretin-based agents have low risk of hypoglycemia and weight gain. However, GLP-1R agonists demonstrate greater

  11. Tyrphostin analogs are GPR35 agonists.

    PubMed

    Deng, Huayun; Hu, Haibei; Fang, Ye

    2011-06-23

    GPR35 is an orphan G protein-coupled receptor that is not well-characterized. Here we employ dynamic mass redistribution (DMR) assays to discover new GPR35 agonists. DMR assays identified tyrphostin analogs as GPR35 agonists, which were confirmed with receptor internalization, Tango β-arrestin translocation, and extracellular-signal-regulated kinase phosphorylation assays. These agonists provide pharmacological tools to study the biology and function of GPR35. Copyright © 2011 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  12. Effect of the α2 -receptor agonists medetomidine, detomidine, xylazine and romifidine on the ketamine metabolism in equines assessed with enantioselective capillary electrophoresis.

    PubMed

    Sandbaumhüter, Friederike A; Theurillat, Regula; Bettschart-Wolfensberger, Regula; Thormann, Wolfgang

    2017-03-02

    The combination of ketamine and an α2 -receptor agonist is often used in veterinary medicine. Four different α2 -receptor agonists, medetomidine, detomidine, xylazine and romifidine, which differ in their chemical structure and thus in selectivity for the α2 -receptor and in the sedative and analgesic potency, are typically employed during surgery of equines. Recovery following anesthesia with ketamine and an α2 -receptor agonist is dependent on the α2 -receptor agonist. This prompted us to investigate i) the inhibition characteristics for the N-demethylation of ketamine to norketamine and ii) the formation of the ketamine metabolites norketamine, 6-hydroxynorketamine (6HNK) and 5,6-dehydronorketamine (DHNK) in presence of the four α2 -receptor agonists and equine liver microsomes. Samples were analyzed with enantioselective capillary electrophoresis using highly sulfated γ-cyclodextrin as chiral selector. All four α2 -receptor agonists have an impact on the ketamine metabolism. Medetomidine was found to be the strongest inhibitor, followed by detomidine, whereas xylazine and romifidine showed almost no effect on the ketamine N-demethylation in the inhibition studies with a short incubation period of the reaction mixture. After prolonged incubation, inhibition with xylazine and romifidine was also observed. The formation of 6HNK and DHNK is affected by all selected α2 -receptor agonists. With medetomidine, levels of these metabolites are reduced compared to the case without an α2 -receptor agonist. For detomidine, xylazine and romifidine, the opposite was found. This article is protected by copyright. All rights reserved.

  13. Inhibitory effects of peroxisome proliferator-activated receptor γ agonists on collagen IV production in podocytes.

    PubMed

    Li, Yanjiao; Shen, Yachen; Li, Min; Su, Dongming; Xu, Weifeng; Liang, Xiubin; Li, Rongshan

    2015-07-01

    Peroxisome proliferator-activated receptor-γ (PPAR-γ) agonists have beneficial effects on the kidney diseases through preventing microalbuminuria and glomerulosclerosis. However, the mechanisms underlying these effects remain to be fully understood. In this study, we investigate the effects of PPAR-γ agonist, rosiglitazone (Rosi) and pioglitazone (Pio), on collagen IV production in mouse podocytes. The endogenous expression of PPAR-γ was found in the primary podocytes and can be upregulated by Rosi and Pio, respectively, detected by RT-PCR and Western blot. PPAR-γ agonist markedly blunted the increasing of collagen IV expression and extraction in podocytes induced by TGF-β. In contrast, adding PPAR-γ antagonist, GW9662, to podocytes largely prevented the inhibition of collagen IV expression from Pio treatment. Our data also showed that phosphorylation of Smad2/3 enhanced by TGF-β in a time-dependent manner was significantly attenuated by adding Pio. The promoter region of collagen IV gene contains one putative consensus sequence of Smad-binding element (SBE) by promoter analysis, Rosi and Pio significantly ameliorated TGF-β-induced SBE4-luciferase activity. In conclusion, PPAR-γ activation by its agonist, Rosi or Pio, in vitro directly inhibits collagen IV expression and synthesis in primary mouse podocytes. The suppression of collagen IV production was related to the inhibition of TGF-β-driven phosphorylation of Smad2/3 and decreased response activity of SBEs of collagen IV in PPAR-γ agonist-treated mouse podocytes. This represents a novel mechanistic support regarding PPAR-γ agonists as podocyte protective agents.

  14. Effect of dopamine receptor agonists on sensory nerve activity: possible therapeutic targets for the treatment of asthma and COPD

    PubMed Central

    Birrell, Mark A; Crispino, Natascia; Hele, David J; Patel, Hema J; Yacoub, Magdi H; Barnes, Peter J; Belvisi, Maria G

    2002-01-01

    Sensory nerves regulate central and local reflexes such as airway plasma leakage, and cough and their function may be enhanced during inflammation. Evidence suggests that dopamine receptor agonists may inhibit sensory nerve-mediated responses.In this study dopamine inhibited vagal sensory nerve induced microvascular leakage in the rat. In order to characterize the receptor involved rat vagus preparations were utilized. Quinagolide (D2/3 agonist), ropinirole (D2/3/4 agonist), SKF 38393 (D1/5 agonist), AR-C68397AA (Viozan™) (dual D2/B2 agonist) and dopamine inhibited hypertonic saline induced depolarization by approximately 50%. Data suggests that AR-C68397AA and quinagolide also inhibited depolarization of the human vagus.The quinagolide response was blocked by sulpiride (D2/3 antagonist) but not SCH 23390 (D1/5 antagonist); ropinirole was partially blocked by sulpiride, totally blocked by spiperone (at a concentration that blocks all dopamine receptors) but not by SCH 23390. The response to SKF 38393 was not blocked by sulpiride but was by SCH 23390. The inhibition evoked by AR-C68397AA was only partially blocked by SCH 23390 but not by sulpiride or spiperone whereas dopamine was blocked by spiperone. The effect of dopamine was not stimulus-specific as it inhibited capsaicin-induced depolarization of the rat vagus in a spiperone sensitive manner.In conclusion, dopamine receptor ligands inhibit depolarization of the rat and human vagus. These data suggest that dopamine receptor agonists may be of therapeutic benefit in the treatment of symptoms such as cough and mucus secretion which are evident in respiratory diseases such as asthma and chronic obstructive pulmonary disease. PMID:12055141

  15. Effect of dopamine receptor agonists on sensory nerve activity: possible therapeutic targets for the treatment of asthma and COPD.

    PubMed

    Birrell, Mark A; Crispino, Natascia; Hele, David J; Patel, Hema J; Yacoub, Magdi H; Barnes, Peter J; Belvisi, Maria G

    2002-06-01

    Sensory nerves regulate central and local reflexes such as airway plasma leakage, and cough and their function may be enhanced during inflammation. Evidence suggests that dopamine receptor agonists may inhibit sensory nerve-mediated responses. In this study dopamine inhibited vagal sensory nerve induced microvascular leakage in the rat. In order to characterize the receptor involved rat vagus preparations were utilized. Quinagolide (D(2/3) agonist), ropinirole (D(2/3/4) agonist), SKF 38393 (D(1/5) agonist), AR-C68397AA (Viozan) (dual D(2)/B(2) agonist) and dopamine inhibited hypertonic saline induced depolarization by approximately 50%. Data suggests that AR-C68397AA and quinagolide also inhibited depolarization of the human vagus. The quinagolide response was blocked by sulpiride (D(2/3) antagonist) but not SCH 23390 (D(1/5) antagonist); ropinirole was partially blocked by sulpiride, totally blocked by spiperone (at a concentration that blocks all dopamine receptors) but not by SCH 23390. The response to SKF 38393 was not blocked by sulpiride but was by SCH 23390. The inhibition evoked by AR-C68397AA was only partially blocked by SCH 23390 but not by sulpiride or spiperone whereas dopamine was blocked by spiperone. The effect of dopamine was not stimulus-specific as it inhibited capsaicin-induced depolarization of the rat vagus in a spiperone sensitive manner. In conclusion, dopamine receptor ligands inhibit depolarization of the rat and human vagus. These data suggest that dopamine receptor agonists may be of therapeutic benefit in the treatment of symptoms such as cough and mucus secretion which are evident in respiratory diseases such as asthma and chronic obstructive pulmonary disease.

  16. Toll-Like Receptor 9 Enhances Bacterial Clearance and Limits Lung Consolidation in Murine Pneumonia Caused by Methicillin-Resistant Staphylococcus aureus

    PubMed Central

    van der Meer, Anne Jan; Achouiti, Achmed; van der Ende, Arie; Soussan, Aicha Ait; Florquin, Sandrine; de Vos, Alex; Zeerleder, Sacha S; van der Poll, Tom

    2016-01-01

    Methicillin-resistant Staphylococcus aureus (MRSA) is an important pathogen in pneumonia associated with severe lung damage. Tissue injury causes release of damage-associated molecular patterns (DAMPs), which may perpetuate inflammation. DNA has been implicated as a DAMP that activates inflammation through Toll-like receptor 9 (TLR9). The aim of this study was to evaluate the role of TLR9 in MRSA pneumonia. Wild-type (Wt) and TLR9 knockout (tlr9−/−) mice were infected intranasally with MRSA USA300 (BK 11540) (5E7 CFU) and euthanized at 6, 24, 48 or 72 h for analyses. MRSA pneumonia was associated with profound release of cell-free host DNA in the airways, as reflected by increases in nucleosome and DNA levels in bronchoalveolar lavage fluid (BALF), accompanied by transient detection of pathogen DNA in MRSA-free BALF supernatants. In BALF, as compared with Wt mice, tlr9−/− mice showed reduced tumor necrosis factor α and IL-6 levels at 6 h and reduced bacterial clearance at 6 and 24 h postinfection. Furthermore, tlr9−/− mice exhibited a greater influx of neutrophils in BALF and increased lung consolidation at 24 and 48 h. This study demonstrates the release of host- and pathogen-derived TLR9 ligands (DNA) into the alveolar space after infection with MRSA via the airways and suggests that TLR9 has proinflammatory effects during MRSA pneumonia associated with enhanced bacterial clearance and limitation of lung consolidation. PMID:27508882

  17. Correlation of Surface Toll-Like Receptor 9 Expression with IL-17 Production in Neutrophils during Septic Peritonitis in Mice Induced by E. coli.

    PubMed

    Ren, Yunjia; Hua, Li; Meng, Xiuping; Xiao, Yue; Hao, Xu; Guo, Sheng; Zhao, Peiyan; Wang, Luowei; Dong, Boqi; Yu, Yongli; Wang, Liying

    2016-01-01

    IL-17 is a proinflammatory cytokine produced by various immune cells. Polymorphonuclear neutrophils (PMNs) are the first line of defense in bacterial infection and express surface Toll-like receptor 9 (sTLR9). To study the relationship of sTLR9 and IL-17 in PMNs during bacterial infection, we infected mice with E. coli intraperitoneally to establish a septic peritonitis model for studying the PMNs response in peritoneal cavity. We found that PMNs and some of "giant cells" were massively accumulated in the peritoneal cavity of mice with fatal septic peritonitis induced by E. coli. Kinetically, the CD11b(+) PMNs were increased from 20-40% at 18 hours to >80% at 72 hours after infection. After E. coli infection, sTLR9 expression on CD11b(+) and CD11b(-) PMNs and macrophages in the PLCs were increased at early stage and deceased at late stage; IL-17 expression was also increased in CD11b(+) PMNs, CD11b(-) PMNs, macrophages, and CD3(+) T cells. Using experiments of in vitro blockage, qRT-PCR and cell sorting, we confirmed that PMNs in the PLCs did increase their IL-17 expression during E. coli infection. Interestingly, sTLR9(-)CD11b(+)Ly6G(+) PMNs, not sTLR9(+)CD11b(+)Ly6G(+) PMNs, were found to be able to increase their IL-17 expression. Together, the data may help understand novel roles of PMNs in septic peritonitis.

  18. Stimulation of Toll-like receptor 9 by chronic intraventricular unmethylated cytosine-guanine DNA infusion causes neuroinflammation and impaired spatial memory.

    PubMed

    Tauber, Simone C; Ebert, Sandra; Weishaupt, Jochen H; Reich, Arno; Nau, Roland; Gerber, Joachim

    2009-10-01

    Bacterial DNA contains a high frequency of unmethylated cytosine-guanine (CpG) motifs that have strong immunostimulatory properties; they are recognized by mammalian Toll-like receptor 9 (TLR9). Because accumulating data suggest that chronic inflammatory processes are involved in the pathogenesis of neurodegenerative diseases, we hypothesized that inflammatory responses stimulated by CpG DNA might contribute to neurodegeneration and brain dysfunction. To assess the effects of continuous CpG DNA exposure in the brain, C57BL/6 (n = 21) and TLR9-deficient mice (n = 15) were given intracerebroventricular infusions of CpG DNA or saline for 28 days. Spatial memory assessed weekly by Morris water maze demonstrated impairment in CpG-treated wild-type mice but not in TLR9-deficient or control-treated mice. Motor function was not affected. Immunohistochemical analysis revealed marked microglial activation and acute axonal damage surrounding the ventricles, ependymal disruption, and reactive astrogliosis within the hippocampal formation in the CpG-treated wild-type but not TLR9-deficient mice or saline-infused controls. These results suggest that the unfavorable effects of CpG DNA are dependent on TLR9 signaling and that exposure to bacterial DNA may contribute to impaired neural function, neuroinflammation, and subsequent neurodegeneration.

  19. Chemical communication in scarab beetles: reciprocal behavioral agonist-antagonist activities of chiral pheromones.

    PubMed Central

    Leal, W S

    1996-01-01

    A novel mechanism of reciprocal behavioral agonist-antagonist activities of enantiomeric pheromones plays a pivotal role in overcoming the signal-to-noise problem derived from the use of a single-constituent pheromone system in scarab beetles. Female Anomala osakana produce (S, Z)-5-(+)-(1-decenyl)oxacyclopentan-2-one, which is highly attractive to males; the response is completely inhibited even by 5% of its antipode. These two enantiomers have reverse roles in the Popillia japonica sex pheromone system. Chiral GC-electroantennographic detector experiments suggest that A. osakana and P. japonica have both R and S receptors that are responsible for behavioral agonist and antagonist responses. PMID:8901541

  20. [Safety of beta-agonists in asthma].

    PubMed

    Oscanoa, Teodoro J

    2014-01-01

    Beta 2 agonist bronchodilators (β2A) are very important part in the pharmacotherapy of bronchial asthma, a disease that progresses in the world in an epidemic way. The β2A are prescribed to millions of people around the world, therefore the safety aspects is of public interest. Short-Acting β2 Agonists (SABAs), such as albuterol inhaler, according to current evidence, confirming its safety when used as a quick-relief or rescue medication. The long-acting β2 agonists (LABAs) The long-acting bronchodilators β2A (Long acting β2 Agonists or LABAs) are used associated with inhaled corticosteroids as controller drugs for asthma exacerbationsaccess, for safety reasons LABAs are not recommended for use as monotherapy.

  1. The β2-adrenoceptor agonist formoterol stimulates mitochondrial biogenesis.

    PubMed

    Wills, Lauren P; Trager, Richard E; Beeson, Gyda C; Lindsey, Christopher C; Peterson, Yuri K; Beeson, Craig C; Schnellmann, Rick G

    2012-07-01

    Mitochondrial dysfunction is a common mediator of disease and organ injury. Although recent studies show that inducing mitochondrial biogenesis (MB) stimulates cell repair and regeneration, only a limited number of chemicals are known to induce MB. To examine the impact of the β-adrenoceptor (β-AR) signaling pathway on MB, primary renal proximal tubule cells (RPTC) and adult feline cardiomyocytes were exposed for 24 h to multiple β-AR agonists: isoproterenol (nonselective β-AR agonist), (±)-(R*,R*)-[4-[2-[[2-(3-chlorophenyl)-2-hydroxyethyl]amino]propyl]phenoxy] acetic acid sodium hydrate (BRL 37344) (selective β(3)-AR agonist), and formoterol (selective β(2)-AR agonist). The Seahorse Biosciences (North Billerica, MA) extracellular flux analyzer was used to quantify carbonylcyanide p-trifluoromethoxyphenylhydrazone (FCCP)-uncoupled oxygen consumption rate (OCR), a marker of maximal electron transport chain activity. Isoproterenol and BRL 37244 did not alter mitochondrial respiration at any of the concentrations examined. Formoterol exposure resulted in increases in both FCCP-uncoupled OCR and mitochondrial DNA (mtDNA) copy number. The effect of formoterol on OCR in RPTC was inhibited by the β-AR antagonist propranolol and the β(2)-AR inverse agonist 3-(isopropylamino)-1-[(7-methyl-4-indanyl)oxy]butan-2-ol hydrochloride (ICI-118,551). Mice exposed to formoterol for 24 or 72 h exhibited increases in kidney and heart mtDNA copy number, peroxisome proliferator-activated receptor γ coactivator 1α, and multiple genes involved in the mitochondrial electron transport chain (F0 subunit 6 of transmembrane F-type ATP synthase, NADH dehydrogenase subunit 1, NADH dehydrogenase subunit 6, and NADH dehydrogenase [ubiquinone] 1β subcomplex subunit 8). Cheminformatic modeling, virtual chemical library screening, and experimental validation identified nisoxetine from the Sigma Library of Pharmacologically Active Compounds and two compounds from the ChemBridge DIVERSet

  2. Negative cooperativity in binding of muscarinic receptor agonists and GDP as a measure of agonist efficacy

    PubMed Central

    Jakubík, J; Janíčková, H; El-Fakahany, EE; Doležal, V

    2011-01-01

    BACKGROUND AND PURPOSE Conventional determination of agonist efficacy at G-protein coupled receptors is measured by stimulation of guanosine-5′-γ−thiotriphosphate (GTPγS) binding. We analysed the role of guanosine diphosphate (GDP) in the process of activation of the M2 muscarinic acetylcholine receptor and provide evidence that negative cooperativity between agonist and GDP binding is an alternative measure of agonist efficacy. EXPERIMENTAL APPROACH Filtration and scintillation proximity assays measured equilibrium binding as well as binding kinetics of [35S]GTPγS and [3H]GDP to a mixture of G-proteins as well as individual classes of G-proteins upon binding of structurally different agonists to the M2 muscarinic acetylcholine receptor. KEY RESULTS Agonists displayed biphasic competition curves with the antagonist [3H]-N-methylscopolamine. GTPγS (1 µM) changed the competition curves to monophasic with low affinity and 50 µM GDP produced a similar effect. Depletion of membrane-bound GDP increased the proportion of agonist high-affinity sites. Carbachol accelerated the dissociation of [3H]GDP from membranes. The inverse agonist N-methylscopolamine slowed GDP dissociation and GTPγS binding without changing affinity for GDP. Carbachol affected both GDP association with and dissociation from Gi/o G-proteins but only its dissociation from Gs/olf G-proteins. CONCLUSIONS AND IMPLICATIONS These findings suggest the existence of a low-affinity agonist-receptor conformation complexed with GDP-liganded G-protein. Also the negative cooperativity between GDP and agonist binding at the receptor/G-protein complex determines agonist efficacy. GDP binding reveals differences in action of agonists versus inverse agonists as well as differences in activation of Gi/o versus Gs/olf G-proteins that are not identified by conventional GTPγS binding. PMID:20958290

  3. Negative cooperativity in binding of muscarinic receptor agonists and GDP as a measure of agonist efficacy.

    PubMed

    Jakubík, J; Janíčková, H; El-Fakahany, E E; Doležal, V

    2011-03-01

    Conventional determination of agonist efficacy at G-protein coupled receptors is measured by stimulation of guanosine-5'-γ-thiotriphosphate (GTPγS) binding. We analysed the role of guanosine diphosphate (GDP) in the process of activation of the M₂ muscarinic acetylcholine receptor and provide evidence that negative cooperativity between agonist and GDP binding is an alternative measure of agonist efficacy. Filtration and scintillation proximity assays measured equilibrium binding as well as binding kinetics of [³⁵S]GTPγS and [³H]GDP to a mixture of G-proteins as well as individual classes of G-proteins upon binding of structurally different agonists to the M₂ muscarinic acetylcholine receptor. Agonists displayed biphasic competition curves with the antagonist [³H]-N-methylscopolamine. GTPγS (1 µM) changed the competition curves to monophasic with low affinity and 50 µM GDP produced a similar effect. Depletion of membrane-bound GDP increased the proportion of agonist high-affinity sites. Carbachol accelerated the dissociation of [³H]GDP from membranes. The inverse agonist N-methylscopolamine slowed GDP dissociation and GTPγS binding without changing affinity for GDP. Carbachol affected both GDP association with and dissociation from G(i/o) G-proteins but only its dissociation from G(s/olf) G-proteins. These findings suggest the existence of a low-affinity agonist-receptor conformation complexed with GDP-liganded G-protein. Also the negative cooperativity between GDP and agonist binding at the receptor/G-protein complex determines agonist efficacy. GDP binding reveals differences in action of agonists versus inverse agonists as well as differences in activation of G(i/o) versus G(s/olf) G-proteins that are not identified by conventional GTPγS binding. © 2011 The Authors. British Journal of Pharmacology © 2011 The British Pharmacological Society.

  4. PPAR{gamma} agonists prevent TGF{beta}1/Smad3-signaling in human hepatic stellate cells

    SciTech Connect

    Zhao Caiyan; Chen, Wei; Yang Liu; Chen Lihong; Stimpson, Stephen A.; Diehl, Anna Mae . E-mail: annamae.diehl@duke.edu

    2006-11-17

    PPAR{gamma} agonists inhibit liver fibrosis, but the mechanisms involved are uncertain. We hypothesized that PPAR{gamma} agonists inhibit transforming growth factor (TGF){beta}1-activation of TGF{beta} receptor (TGF{beta}R)-1 signaling in quiescent stellate cells, thereby abrogating Smad3-dependent induction of extracellular matrix (ECM) genes, such as PAI-1 and collagen-1{alpha}I. To test this, human HSC were cultured to induce a quiescent phenotype, characterized by lipid accumulation and PPAR{gamma} expression and transcriptional activity. These adipocytic HSC were then treated with TGF{beta}1 {+-} a TGF{beta}R-1 kinase inhibitor (SB431542) or a PPAR{gamma} agonist (GW7845). TGF{beta}1 caused dose- and time-dependent increases in Smad3 phosphorylation, followed by induction of collagen and PAI-1 expression. Like the TGF{beta}R-1 kinase inhibitor, the PPAR{gamma} agonist caused dose-dependent inhibition of all of these responses without effecting HSC proliferation or viability. Thus, the anti-fibrotic actions of PPAR{gamma} agonists reflect their ability to inhibit TGF{beta}1-TGF{beta}R1 signaling that initiates ECM gene expression in quiescent HSC.

  5. Dopaminergic agonists in Parkinson's disease.

    PubMed

    Alonso Cánovas, A; Luquin Piudo, R; García Ruiz-Espiga, P; Burguera, J A; Campos Arillo, V; Castro, A; Linazasoro, G; López Del Val, J; Vela, L; Martínez Castrillo, J C

    2014-05-01

    Non-ergoline dopamine agonists (DA) are effective treatments for Parkinson's disease (PD). This review presents the pharmacology, evidence of efficacy and safety profile of pramipexole, ropinirole, and rotigotine, and practical recommendations are given regarding their use in clinical practice. Extended-release formulations of pramipexole and ropinirole and transdermal continuous delivery rotigotine patches are currently available; these may contribute to stabilising of plasma levels. In early PD, the three drugs significantly improve disability scales, delay time to dyskinesia and allow a later introduction of levodopa. In late PD they reduced total 'off'-time, improved Unified Parkinson's Disease Rating Scale (UPDRS) in both 'on' and 'off' state and allowed a reduction in total levodopa dosage. A significant improvement in quality of life scales has also been demonstrated. Extended-release formulations have proved to be non-inferior to the immediate release formulations and are better tolerated (ropinirole). Despite a generally good safety profile, serious adverse events, such as impulse control disorder and sleep attacks, need to be routinely monitored. Although combination therapy has not been addressed in scientific literature, certain combinations, such as apomorphine and another DA, may be helpful. Switching from one DA to another is feasible and safe, although in the first days an overlap of dopaminergic side effects may occur. When treatment with DA is stopped abruptly, dopamine withdrawal syndrome may present. Suspending any DA, especially pramipexole, has been linked to onset of apathy, which may be severe. New non-ergotine DAs are a valuable option for the treatment of both early and late PD. Despite their good safety profile, serious adverse effects may appear; these effects may have a pathoplastic effect on the course of PD and need to be monitored. Copyright © 2011 Sociedad Española de Neurología. Published by Elsevier Espana. All rights reserved.

  6. PPAR Agonists and Cardiovascular Disease in Diabetes

    PubMed Central

    Calkin, Anna C.; Thomas, Merlin C.

    2008-01-01

    Peroxisome proliferators activated receptors (PPARs) are ligand-activated nuclear transcription factors that play important roles in lipid and glucose homeostasis. To the extent that PPAR agonists improve diabetic dyslipidaemia and insulin resistance, these agents have been considered to reduce cardiovascular risk. However, data from murine models suggests that PPAR agonists also have independent anti-atherosclerotic actions, including the suppression of vascular inflammation, oxidative stress, and activation of the renin angiotensin system. Many of these potentially anti-atherosclerotic effects are thought to be mediated by transrepression of nuclear factor-kB, STAT, and activator protein-1 dependent pathways. In recent clinical trials, PPARα agonists have been shown to be effective in the primary prevention of cardiovascular events, while their cardiovascular benefit in patients with established cardiovascular disease remains equivocal. However, the use of PPARγ agonists, and more recently dual PPARα/γ coagonists, has been associated with an excess in cardiovascular events, possibly reflecting unrecognised fluid retention with potent agonists of the PPARγ receptor. Newer pan agonists, which retain their anti-atherosclerotic activity without weight gain, may provide one solution to this problem. However, the complex biologic effects of the PPARs may mean that only vascular targeted agents or pure transrepressors will realise the goal of preventing atherosclerotic vascular disease. PMID:18288280

  7. Long-term studies of dopamine agonists.

    PubMed

    Hubble, Jean P

    2002-02-26

    Dopamine agonists have long been used as adjunctive therapy for the treatment of Parkinson's disease (PD). In more recent years these drugs have also been proved safe and effective as initial therapy in lieu of levodopa in the treatment of PD. Long-term levodopa therapy is associated with motor complications, including fluctuating response patterns and dyskinesia. By initially introducing a dopamine agonist as symptomatic drug therapy, it may be possible to postpone the use of levodopa and delay or prevent the development of motor complications. Recently, four clinical trials have explored this hypothesis by comparing the long-term response and side effects of levodopa with dopamine agonist therapy. The drugs studied have included ropinirole, pramipexole, cabergoline, and pergolide. In each of these projects, the occurrence of motor complications, such as wearing off and dyskinesia, was significantly less in the subjects assigned to initiation of therapy with a dopamine agonist. The addition of levodopa could be postponed by many months or even several years. Therefore, these long-term studies of dopamine agonists support the initiation of a dopamine agonist instead of levodopa in an effort to postpone levodopa-related motor complications. This therapeutic approach may be particularly appropriate in PD patients with a long treatment horizon on the basis of age and general good health. The extension phase of the long-term study comparing pramipexole with levodopa is ongoing, and follow-up information may help to establish the value of this treatment strategy.

  8. Design and Synthesis of Dopaminergic Agonists.

    PubMed

    Matute, Maria Soledad; Matute, Rosa; Merino, Pedro

    2016-01-01

    The use of dopaminergic agonists is key in the treatment of Parkinson's disease and related central nervous system (CNS) neurodegenerative disorders. Despite there are a number of commercialized dopaminergic agonists that are currently being used successfully in the first stages of the disease, they often fail to provide sustained clinical benefit for a long period due to the appearance of side-effects such as augmentation, sleepiness, nausea, hypothension, and compulsive behaviors among others. New dopaminergic agonists with less side effects are being developed. These novel compounds offer an alternative when the disease progresses and patients fail to respond to standard dopaminergic treatments or side-effects increased. Chemistry, and in particular chemical synthesis, has played a major role in bringing synthetic dopaminergic agonists to the clinic and continues to be crucial for the development of new and necessary drugs for long-term treatments with less undesired side effects. A number of structural modifications of parent compounds have led to enhanced agonism but also partial agonism or even antagonism of one or more dopamine receptors. In some cases, these activities are accompanied by agonist effect at serotonin receptors which suggests a potential clinical application in the treatment of schizophrenia In this review, chemical synthesis of dopaminergic agents, their affinity, and the corresponding agonist/antagonist effects will be highlighted.

  9. Adverse Effects of GLP-1 Receptor Agonists

    PubMed Central

    Filippatos, Theodosios D.; Panagiotopoulou, Thalia V.; Elisaf, Moses S.

    2014-01-01

    Glucagon-like peptide-1 (GLP-1) receptor agonists are a class of injective anti-diabetic drugs that improve glycemic control and many other atherosclerosis-related parameters in patients with type 2 diabetes (T2D). However, the use of this relatively new class of drugs may be associated with certain adverse effects. Concerns have been expressed regarding the effects of these drugs on pancreatic and thyroid tissue, since animal studies and analyses of drug databases indicate an association of GLP-1 receptor agonists with pancreatitis, pancreatic cancer, and thyroid cancer. However, several meta-analyses failed to confirm a cause-effect relation between GLP-1 receptor agonists and the development of these adverse effects. One benefit of GLP-1 receptor agonists is that they do not cause hypoglycemia when combined with metformin or thiazolidinediones, but the dose of concomitant sulphonylurea or insulin may have to be decreased to reduce the risk of hypoglycemic episodes. On the other hand, several case reports have linked the use of these drugs, mainly exenatide, with the occurrence of acute kidney injury, primarily through hemodynamic derangement due to nausea, vomiting, and diarrhea. The most common symptoms associated with the use of GLP-1 receptor agonists are gastrointestinal symptoms, mainly nausea. Other common adverse effects include injection site reactions, headache, and nasopharyngitis, but these effects do not usually result in discontinuation of the drug. Current evidence shows that GLP-1 receptor agonists have no negative effects on the cardiovascular risk of patients with T2D. Thus, GLP-1 receptor agonists appear to have a favorable safety profile, but ongoing trials will further assess their cardiovascular effects. The aim of this review is to analyze critically the available data regarding adverse events of GLP-1 receptor agonists in different anatomic systems published in Pubmed and Scopus. Whenever possible, certain differences between GLP-1

  10. Correlation of Surface Toll-Like Receptor 9 Expression with IL-17 Production in Neutrophils during Septic Peritonitis in Mice Induced by E. coli

    PubMed Central

    Ren, Yunjia; Hao, Xu; Zhao, Peiyan; Yu, Yongli; Wang, Liying

    2016-01-01

    IL-17 is a proinflammatory cytokine produced by various immune cells. Polymorphonuclear neutrophils (PMNs) are the first line of defense in bacterial infection and express surface Toll-like receptor 9 (sTLR9). To study the relationship of sTLR9 and IL-17 in PMNs during bacterial infection, we infected mice with E. coli intraperitoneally to establish a septic peritonitis model for studying the PMNs response in peritoneal cavity. We found that PMNs and some of “giant cells” were massively accumulated in the peritoneal cavity of mice with fatal septic peritonitis induced by E. coli. Kinetically, the CD11b+ PMNs were increased from 20–40% at 18 hours to >80% at 72 hours after infection. After E. coli infection, sTLR9 expression on CD11b+ and CD11b− PMNs and macrophages in the PLCs were increased at early stage and deceased at late stage; IL-17 expression was also increased in CD11b+ PMNs, CD11b− PMNs, macrophages, and CD3+ T cells. Using experiments of in vitro blockage, qRT-PCR and cell sorting, we confirmed that PMNs in the PLCs did increase their IL-17 expression during E. coli infection. Interestingly, sTLR9−CD11b+Ly6G+ PMNs, not sTLR9+CD11b+Ly6G+ PMNs, were found to be able to increase their IL-17 expression. Together, the data may help understand novel roles of PMNs in septic peritonitis. PMID:27057095

  11. Interaction of a radiolabeled agonist with cardiac muscarinic cholinergic receptors

    SciTech Connect

    Harden, T.K.; Meeker, R.B.; Martin, M.W.

    1983-12-01

    The interaction of a radiolabeled muscarinic cholinergic receptor agonist, (methyl-/sup 3/H)oxotremorine acetate ((/sup 3/H)OXO), with a washed membrane preparation derived from rat heart, has been studied. In binding assays at 4 degrees C, the rate constants for association and dissociation of (/sup 3/H)OXO were 2 X 10(7) M-1 min-1 and 5 X 10(-3) min-1, respectively, Saturation binding isotherms indicated that binding was to a single population of sites with a Kd of approximately 300 pM. The density of (/sup 3/H)OXO binding sites (90-100 fmol/mg of protein) was approximately 75% of that determined for the radiolabeled receptor antagonist (/sup 3/H)quinuclidinyl benzilate. Both muscarinic receptor agonists and antagonists inhibited the binding of (/sup 3/H)OXO with high affinity and Hill slopes of approximately one. Guanine nucleotides completely inhibited the binding of (/sup 3/H)OXO. This effect was on the maximum binding (Bmax) of (/sup 3/H)OXO with no change occurring in the Kd; the order of potency for five nucleotides was guanosine 5'-O-(3-thio-triphosphate) greater than 5'-guanylylimidodiphosphate greater than GTP greater than or equal to guanosine/diphosphate greater than GMP. The (/sup 3/H)OXO-induced interaction of muscarinic receptors with a guanine nucleotide binding protein was stable to solubilization. That is, membrane receptors that were prelabeled with (/sup 3/H)OXO could be solubilized with digitonin, and the addition of guanine nucleotides to the soluble, (/sup 3/H)OXO-labeled complex resulted in dissociation of (/sup 3/H)OXO from the receptor. Pretreatment of membranes with relatively low concentrations of N-ethylmaleimide inhibited (/sup 3/H)OXO binding by 85% with no change in the Kd of (/sup 3/H)OXO, and with no effect on (/sup 3/H)quinuclidinyl benzilate binding.

  12. Differential opioid agonist regulation of the mouse mu opioid receptor.

    PubMed

    Blake, A D; Bot, G; Freeman, J C; Reisine, T

    1997-01-10

    Mu opioid receptors mediate the analgesia induced by morphine. Prolonged use of morphine causes tolerance development and dependence. To investigate the molecular basis of tolerance and dependence, the cloned mouse mu opioid receptor with an amino-terminal epitope tag was stably expressed in human embryonic kidney (HEK) 293 cells, and the effects of prolonged opioid agonist treatment on receptor regulation were examined. In HEK 293 cells the expressed mu receptor showed high affinity, specific, saturable binding of radioligands and a pertussis toxin-sensitive inhibition of adenylyl cyclase. Pretreatment (1 h, 3 h, or overnight) of cells with 1 microM morphine or [D-Ala2MePhe4,Gly(ol)5]enkephalin (DAMGO) resulted in no apparent receptor desensitization, as assessed by opioid inhibition of forskolin-stimulated cAMP levels. In contrast, the morphine and DAMGO pretreatments (3 h) resulted in a 3-4-fold compensatory increase in forskolin-stimulated cAMP accumulation. The opioid agonists methadone and buprenorphine are used in the treatment of addiction because of a markedly lower abuse potential. Pretreatment of mu receptor-expressing HEK 293 cells with methadone or buprenorphine abolished the ability of opioids to inhibit adenylyl cyclase. No compensatory increase in forskolin-stimulated cAMP accumulation was found with methadone or buprenorphine; these opioids blocked the compensatory effects observed with morphine and DAMGO. Taken together, these results indicate that methadone and buprenorphine interact differently with the mouse mu receptor than either morphine or DAMGO. The ability of methadone and buprenorphine to desensitize the mu receptor and block the compensatory rise in forskolin-stimulated cAMP accumulation may be an underlying mechanism by which these agents are effective in the treatment of morphine addiction.

  13. Co-administration of delta- and mu-opioid receptor agonists promotes peripheral opioid receptor function

    PubMed Central

    Schramm, Cicely L.; Honda, Christopher N.

    2010-01-01

    Enhancement of peripheral opioid analgesia following tissue injury or inflammation in animal models is well-documented, but clinical results of peripheral opioid therapy remain inconsistent. Previous studies in the central nervous system have shown that co-administration of μ- and δ-opioid receptor agonists can enhance analgesic outcomes; however, less is known about the functional consequences of opioid receptor interactions in the periphery. The present study examines the effects of intraplantar injection of the μ- and δ-opioid receptor agonists, morphine and deltorphin, alone and in combination on behavioral tests of nociception in naïve rats and on potassium-evoked release of CGRP from sciatic nerves of naïve rats. Neither drug alone affected nociceptive behaviors or CGRP release. Two separate measures of mechanical nociceptive sensitivity remained unchanged after co-administration of the two drugs. In contrast, when deltorphin was co-injected with morphine, dose-dependent and peripherally-restricted increases in paw withdrawal latencies to radiant heat were observed. Similarly, concentration-dependent inhibition of CGRP release was observed when deltorphin and morphine were administered in sequence prior to potassium stimulation. However, no inhibition was observed when morphine was administered prior to deltorphin. All combined opioid effects were blocked by co-application of antagonists. Deltorphin exposure also enhanced the in vivo and in vitro effects of another μ-opioid receptor agonist, DAMGO. Together, these results suggest that under normal conditions, δ-opioid receptor agonists enhance the effect of μ-opioid receptor agonists in the periphery, and local co-administration of δ- and μ-opioid receptor agonists may improve results of peripheral opioid therapy for the treatment of pain. PMID:20970925

  14. Development of CINPA1 analogs as novel and potent inverse agonists of constitutive androstane receptor.

    PubMed

    Lin, Wenwei; Yang, Lei; Chai, Sergio C; Lu, Yan; Chen, Taosheng

    2016-01-27

    Constitutive androstane receptor (CAR, NR1I3) and pregnane X receptor (PXR, NR1I2) are master regulators of endobiotic and xenobiotic metabolism and disposition. Because CAR is constitutively active in certain cellular contexts, inhibiting CAR might reduce drug-induced hepatotoxicity and resensitize drug-resistant cancer cells to chemotherapeutic drugs. We recently reported a novel CAR inhibitor/inverse agonist CINPA1 (11). Here, we have obtained or designed 54 analogs of CINPA1 and used a time-resolved fluorescence resonance energy transfer (TR-FRET) assay to evaluate their CAR inhibition potency. Many of the 54 analogs showed CAR inverse agonistic activities higher than those of CINPA1, which has an IC50 value of 687 nM. Among them, 72 has an IC50 value of 11.7 nM, which is about 59-fold more potent than CINPA1 and over 10-fold more potent than clotrimazole (an IC50 value of 126.9 nM), the most potent CAR inverse agonist in a biochemical assay previously reported by others. Docking studies provide a molecular explanation of the structure-activity relationship (SAR) observed experimentally. To our knowledge, this effort is the first chemistry endeavor in designing and identifying potent CAR inverse agonists based on a novel chemical scaffold, leading to 72 as the most potent CAR inverse agonist so far. The 54 chemicals presented are novel and unique tools for characterizing CAR's function, and the SAR information gained from these 54 analogs could guide future efforts to develop improved CAR inverse agonists.

  15. β2-Adrenoceptor agonists as novel, safe and potentially effective therapies for Amyotrophic lateral sclerosis (ALS).

    PubMed

    Bartus, Raymond T; Bétourné, Alexandre; Basile, Anthony; Peterson, Bethany L; Glass, Jonathan; Boulis, Nicholas M

    2016-01-01

    Amyotrophic lateral sclerosis (ALS) is a chronic and progressive neuromuscular disease for which no cure exists and better treatment options are desperately needed. We hypothesize that currently approved β2-adrenoceptor agonists may effectively treat the symptoms and possibly slow the progression of ALS. Although β2-agonists are primarily used to treat asthma, pharmacologic data from animal models of neuromuscular diseases suggest that these agents may have pharmacologic effects of benefit in treating ALS. These include inhibiting protein degradation, stimulating protein synthesis, inducing neurotrophic factor synthesis and release, positively modulating microglial and systemic immune function, maintaining the structural and functional integrity of motor endplates, and improving energy metabolism. Moreover, stimulation of β2-adrenoceptors can activate a range of downstream signaling events in many different cell types that could account for the diverse array of effects of these agents. The evidence supporting the possible therapeutic benefits of β2-agonists is briefly reviewed, followed by a more detailed review of clinical trials testing the efficacy of β-agonists in a variety of human neuromuscular maladies. The weight of evidence of the potential benefits from treating these diseases supports the hypothesis that β2-agonists may be efficacious in ALS. Finally, ways to monitor and manage the side effects that may arise with chronic administration of β2-agonists are evaluated. In sum, effective, safe and orally-active β2-agonists may provide a novel and convenient means to reduce the symptoms of ALS and possibly delay disease progression, affording a unique opportunity to repurpose these approved drugs for treating ALS, and rapidly transforming the management of this serious, unmet medical need.

  16. Occurrence of aryl hydrocarbon receptor agonists and genotoxic compounds in the river systems in Southern Taiwan.

    PubMed

    Chou, Pei-Hsin; Liu, Tong-Cun; Ko, Fung-Chi; Liao, Mong-Wei; Yeh, Hsiao-Mei; Yang, Tse-Han; Wu, Chun-Ting; Chen, Chien-Hsun; Tsai, Tsung-Ya

    2014-07-01

    Water and sediment samples from river systems located in Southern Taiwan were investigated for the presence of aryl hydrocarbon receptor (AhR) agonists and genotoxicants by a combination of recombinant cell assays and gas chromatography-mass spectrometry analysis. AhR agonist activity and genotoxic response were frequently detected in samples collected during different seasons. In particular, dry-season water and sediment samples from Erren River showed strong AhR agonist activity (201-1423 ng L(-1) and 1374-5631 ng g(-1) β-naphthoflavone equivalents) and high genotoxic potential. Although no significant correlation was found between AhR agonist activity and genotoxicity, potential genotoxicants in sample extracts were suggested to be causative agents for yeast growth inhibition in the AhR-responsive reporter gene assay. After high performance liquid chromatography fractionation, AhR agonist candidates were detected in several fractions of Erren River water and sediment extracts, while possible genotoxicants were only found in water extracts. In addition, polycyclic aromatic hydrocarbons, the typical contaminants showing high AhR binding affinity, were only minor contributors to the AhR agonist activity detected in Erren River sediment extracts. Our findings displayed the usefulness of bioassays in evaluating the extent of environmental contamination, which may be helpful in reducing the chances of false-negative results obtained from chemical analysis of conventional contaminants. Further research will be undertaken to identify major candidates for xenobiotic AhR agonists and genotoxicants to better protect the aquatic environments in Taiwan.

  17. Neuroprotective effects of KR-62980, a new PPARγ agonist, against chemical ischemia-reperfusion in SK-N-SH cells.

    PubMed

    Kim, Ki Young; Cho, Hyun Sill; Lee, Su Hee; Ahn, Jin Hee; Cheon, Hyae Gyeong

    2011-02-04

    PPARγ agonists exert neuroprotective effects against various types of brain injuries. In the present study, we investigated the effects of KR-62980, a new PPARγ agonist, and rosiglitazone on the neuronal cell death induced by chemical ischemia-reperfusion in SK-N-SH cells and their underlying molecular mechanisms. Both agonists inhibited chemical ischemia-reperfusion-induced cell death, and the effects were associated with anti-apoptotic action. KR-62980 and rosiglitazone suppressed NO and ROS formation, and N-acetyl-N-acetoxy-4-chlorobenzenesulfonamide, an NO generator, reversed the protective effects of the agonists on cell viability. In the agonist-induced anti-apoptotic process, PTEN expression was suppressed in parallel with increased Akt and ERK phosphorylation, whereas PD98059 (an ERK inhibitor) or wortmannin (a PI-3K inhibitor) abolished the cell survival by KR-62980 and rosiglitazone. All of the effects of KR-62980 and rosiglitazone appeared to be PPARγ-dependent because the effects were reversed by bisphenol A diglycidyl ether, a PPARγ antagonist, or by PPARγ knockdown. Our results demonstrate that two PPARγ agonists, KR-62980 and rosiglitazone, inhibited chemical ischemia-reperfusion-induced neuronal cell death by PPARγ-mediated anti-apoptotic and anti-oxidant mechanisms related to PTEN suppression and ERK phosphorylation. Copyright © 2010 Elsevier B.V. All rights reserved.

  18. Thiazolidinediones are Partial Agonists for the Glucocorticoid Receptor

    PubMed Central

    Matthews, L; Berry, A; Tersigni, M; D’Acquisto, F; Ianaro, A; Ray, D

    2014-01-01

    Although thiazolidinediones were designed as specific PPARγ-ligands there is evidence for some off-target effects mediated by a non-PPARγ mechanism. Previously we have shown that Rosiglitazone has anti-inflammatory actions not explicable by activation of PPARγ, but possibly by the glucocorticoid receptor (GR). Rosiglitazone induces nuclear translocation both of GR-GFP, and endogenous GR in HeLa and U20S cells but with slower kinetics than Dexamethasone. Rosiglitazone also induces GR phosphorylation (Ser211), a GR ligand-binding specific effect. Rosiglitazone drives luciferase expression from a simple GRE containing reporter gene in a GR-dependent manner (EC50 4μM), with a similar amplitude response to the partial GR agonist RU486. Rosiglitazone also inhibits Dexamethasone driven reporter gene activity (IC50 2.9μM) in a similar fashion to RU486, suggesting partial agonist activity. Importantly we demonstrate a similar effect in PPARγ-null cells suggesting both GR-dependence and PPARγ-independence. Rosiglitazone also activates a GAL4-GR chimera, driving a UAS promoter, demonstrating DNA template sequence independence, and furthermore enhanced SRC1-GR interaction, measured by a mammalian two-hybrid assay. Both Ciglitazone and Pioglitazone, structurally related to Rosiglitazone, show similar effects on the GR. The antiproliferative effect of Rosiglitazone is increased in U20S cells that overexpress GR, suggesting a biologically important GR-dependent component of Rosiglitazone action. Rosiglitazone is a partial GR agonist, affecting GR activation and trafficking to influence engagement of target genes and affect cell function. This novel mode of action may explain some off-target effects observed in vivo. Additionally, antagonism of glucocorticoid action may contribute to the anti-diabetic actions of Rosiglitazone. PMID:18801908

  19. Receptors and Channels Targeted by Synthetic Cannabinoid Receptor Agonists and Antagonists

    PubMed Central

    Pertwee, R.G.

    2010-01-01

    It is widely accepted that non-endogenous compounds that target CB1 and/or CB2 receptors possess therapeutic potential for the clinical management of an ever growing number of disorders. Just a few of these disorders are already treated with Δ9-tetrahydrocannabinol or nabilone, both CB1/CB2 receptor agonists, and there is now considerable interest in expanding the clinical applications of such agonists and also in exploiting CB2-selective agonists, peripherally restricted CB1/CB2 receptor agonists and CB1/CB2 antagonists and inverse agonists as medicines. Already, numerous cannabinoid receptor ligands have been developed and their interactions with CB1 and CB2 receptors well characterized. This review describes what is currently known about the ability of such compounds to bind to, activate, inhibit or block non-CB1, non-CB2 G protein-coupled receptors such as GPR55, transmitter gated channels, ion channels and nuclear receptors in an orthosteric or allosteric manner. It begins with a brief description of how each of these ligands interacts with CB1 and/or CB2 receptors. PMID:20166927

  20. Reconstitution of high-affinity opioid agonist binding in brain membranes

    SciTech Connect

    Remmers, A.E.; Medzihradsky, F. )

    1991-03-15

    In synaptosomal membranes from rat brain cortex, the {mu} selective agonist ({sup 3}H)dihydromorphine in the absence of sodium, and the nonselective antagonist ({sup 3}H)naltrexone in the presence of sodium, bound to two populations of opioid receptor sites with K{sub d} values of 0.69 and 8.7 nM for dihydromorphine, and 0.34 and 5.5 nM for naltrexone. The addition of 5 {mu}M guanosine 5{prime}-({gamma}-thio)triphosphate (GTP({gamma}S)) strongly reduced high-affinity agonist but not antagonist binding. Exposure of the membranes to high pH reduced the number of GTP({gamma}-{sup 35}S) binding sites by 90% and low K{sub m}, opioid-sensitive GTPase activity by 95%. In these membranes, high-affinity agonist binding was abolished and modulation of residual binding by GTP({gamma}S) was diminished. Alkali treatment of the glioma cell membranes prior to fusion inhibited most of the low K{sub m} GTPase activity and prevented the reconstitution of agonist binding. The results show that high-affinity opioid agonist binding reflects the ligand-occupied receptor - guanine nucleotide binding protein complex.

  1. Computational Prediction and Biochemical Analyses of New Inverse Agonists for the CB1 Receptor

    PubMed Central

    2015-01-01

    Human cannabinoid type 1 (CB1) G-protein coupled receptor is a potential therapeutic target for obesity. The previously predicted and experimentally validated ensemble of ligand-free conformations of CB1 [Scott, C. E. et al. Protein Sci.2013, 22, 101−11323184890; Ahn, K. H. et al. Proteins2013, 81, 1304–131723408552] are used here to predict the binding sites for known CB1-selective inverse agonists including rimonabant and its seven known derivatives. This binding pocket, which differs significantly from previously published models, is used to identify 16 novel compounds expected to be CB1 inverse agonists by exploiting potential new interactions. We show experimentally that two of these compounds exhibit inverse agonist properties including inhibition of basal and agonist-induced G-protein coupling activity, as well as an enhanced level of CB1 cell surface localization. This demonstrates the utility of using the predicted binding sites for an ensemble of CB1 receptor structures for designing new CB1 inverse agonists. PMID:26633590

  2. Identification of dual PPARα/γ agonists and their effects on lipid metabolism.

    PubMed

    Gao, Quanqing; Hanh, Jacky; Váradi, Linda; Cairns, Rose; Sjöström, Helena; Liao, Vivian W Y; Wood, Peta; Balaban, Seher; Ong, Jennifer Ai; Lin, Hsuan-Yu Jennifer; Lai, Felcia; Hoy, Andrew J; Grewal, Thomas; Groundwater, Paul W; Hibbs, David E

    2015-12-15

    The three peroxisome proliferator-activated receptor (PPAR) isoforms; PPARα, PPARγ and PPARδ, play central roles in lipid metabolism and glucose homeostasis. Dual PPARα/γ agonists, which stimulate both PPARα and PPARγ isoforms to similar extents, are gaining popularity as it is believed that they are able to ameliorate the unwanted side effects of selective PPARα and PPARγ agonists; and may also be used to treat dyslipidemia and type 2 diabetes mellitus simultaneously. In this study, virtual screening of natural product libraries, using both structure-based and ligand-based drug discovery approaches, identified ten potential dual PPARα/γ agonist lead compounds (9-13 and 16-20). In vitro assays confirmed these compounds to show no statistically significant toxicity to cells, with the exception of compound 12 which inhibited cell growth to 74.5%±3.5 and 54.1%±3.7 at 50μM and 100μM, respectively. In support of their potential as dual PPARα/γ agonists, all ten compounds upregulated the expression of cholesterol transporters ABCA1 and ABCG1 in THP-1 macrophages, with indoline derivative 16 producing the greatest elevation (2.3-fold; 3.3-fold, respectively). Furthermore, comparable to the activity of established PPARα and PPARγ agonists, compound 16 stimulated triacylglycerol accumulation during 3T3-L1 adipocyte differentiation as well as fatty acid β-oxidation in HuH7 hepatocytes. Copyright © 2015 Elsevier Ltd. All rights reserved.

  3. 2-Dialkynyl derivatives of (N)-methanocarba nucleosides: 'Clickable' A(3) adenosine receptor-selective agonists.

    PubMed

    Tosh, Dilip K; Chinn, Moshe; Yoo, Lena S; Kang, Dong Wook; Luecke, Hans; Gao, Zhan-Guo; Jacobson, Kenneth A

    2010-01-15

    We modified a series of (N)-methanocarba nucleoside 5'-uronamides to contain dialkyne groups on an extended adenine C2 substituent, as synthetic intermediates leading to potent and selective A(3) adenosine receptor (AR) agonists. The proximal alkyne was intended to promote receptor recognition, and the distal alkyne reacted with azides to form triazole derivatives (click cycloaddition). Click chemistry was utilized to couple an octadiynyl A(3)AR agonist to azido-containing fluorescent, chemically reactive, biotinylated, and other moieties with retention of selective binding to the A(3)AR. A bifunctional thiol-reactive crosslinking reagent was introduced. The most potent and selective novel compound was a 1-adamantyl derivative (K(i) 6.5nM), although some of the click products had K(i) values in the range of 200-400nM. Other potent, selective derivatives (K(i) at A(3)AR innM) were intended as possible receptor affinity labels: 3-nitro-4-fluorophenyl (10.6), alpha-bromophenacyl (9.6), thiol-reactive isothiazolone (102), and arylisothiocyanate (37.5) derivatives. The maximal functional effects in inhibition of forskolin-stimulated cAMP were measured, indicating that this class of click adducts varied from partial to full A(3)AR agonist compared to other widely used agonists. Thus, this strategy provides a general chemical approach to linking potent and selective A(3)AR agonists to reporter groups of diverse structure and to carrier moieties.

  4. Inhaled P2Y2 receptor agonists as a treatment for patients with Cystic Fibrosis lung disease.

    PubMed

    Kellerman, Don; Evans, Richard; Mathews, Dave; Shaffer, Christy

    2002-12-05

    P2Y(2) receptor agonists are a new class of compounds that are being evaluated as a treatment for the pulmonary manifestations of Cystic Fibrosis (CF). Results of preclinical research suggest that these compounds inhibit sodium absorption, restore chloride conductance and rehydrate the CF airway surface. In addition, P2Y(2) receptor agonists have been shown to enhance ciliary beat frequency and increase mucociliary clearance in animals and subjects with impaired mucociliary clearance. The normalization of airway surface liquid and enhancement of lung clearance is expected to provide a clinical benefit to CF patients. A number of P2Y(2) agonist compounds have been evaluated in healthy subjects and patients with CF. Most recently, INS37217, a metabolically stable and potent P2Y(2) agonist has been developed and studies have shown it to be well-tolerated when given via inhalation. This compound is currently being evaluated in children and adults with CF lung disease.

  5. Anti-inflammatory properties of a dual PPARgamma/alpha agonist muraglitazar in in vitro and in vivo models

    PubMed Central

    2013-01-01

    Introduction Peroxisome proliferator-activated receptor (PPAR) agonists are widely used drugs in the treatment of diabetes and dyslipidemia. In addition to their metabolic effects, PPAR isoforms PPARα and PPARγ are also involved in the regulation of immune responses and inflammation. In the present study, we investigated the effects of a dual PPARγ/α agonist muraglitazar on inflammatory gene expression in activated macrophages and on carrageenan-induced inflammation in the mouse. Methods J774 murine macrophages were activated by lipopolysaccharide (LPS) and treated with dual PPARγ/α agonist muraglitazar, PPARγ agonist GW1929 or PPARα agonist fenofibrate. The effects of PPAR agonists on cytokine production and the activation of inducible nitric oxide synthase (iNOS) pathway were investigated by ELISA, Griess method, Western blotting and quantitative RT-PCR. Nuclear translocation, DNA-binding activity and reporter gene assays were used to assess the activity of nuclear factor kappa B (NF-kB) transcription factor. Carrageenan-induced paw oedema was used as an in vivo model of acute inflammation. Results Muraglitazar as well as PPARγ agonist GW1929 and PPARα agonist fenofibrate inhibited LPS-induced iNOS expression and NO production in activated macrophages in a dose-dependent manner. Inhibition of iNOS expression by muraglitazar included both transcriptional and post-transcriptional components; the former being shared by GW1929 and the latter by fenofibrate. All tested PPAR agonists also inhibited IL-6 production, while TNFα production was reduced by muraglitazar and GW1929, but not by fenofibrate. Interestingly, the anti-inflammatory properties of muraglitazar were also translated in vivo. This was evidenced by the finding that muraglitazar inhibited carrageenan-induced paw inflammation in a dose-dependent manner in mice as did iNOS inhibitor L-NIL and anti-inflammatory steroid dexamethasone. Conclusions These results show that muraglitazar has anti

  6. Agonists and antagonists for P2 receptors

    PubMed Central

    Jacobson, Kenneth A.; Costanzi, Stefano; Joshi, Bhalchandra V.; Besada, Pedro; Shin, Dae Hong; Ko, Hyojin; Ivanov, Andrei A.; Mamedova, Liaman

    2015-01-01

    Recent work has identified nucleotide agonists selective for P2Y1, P2Y2 and P2Y6 receptors and nucleotide antagonists selective for P2Y1, P2Y12 and P2X1 receptors. Selective non-nucleotide antagonists have been reported for P2Y1, P2Y2, P2Y6, P2Y12, P2Y13, P2X2/3/P2X3 and P2X7 receptors. For example, the dinucleotide INS 37217 (Up4dC) potently activates the P2Y2 receptor, and the non-nucleotide antagonist A-317491 is selective for P2X2/3/P2X3 receptors. Nucleotide analogues in which the ribose moiety is substituted by a variety of novel ring systems, including conformation-ally locked moieties, have been synthesized as ligands for P2Y receptors. The focus on conformational factors of the ribose-like moiety allows the inclusion of general modifications that lead to enhanced potency and selectivity. At P2Y1,2,4,11 receptors, there is a preference for the North conformation as indicated with (N)-methanocarba analogues. The P2Y1 antagonist MRS2500 inhibited ADP-induced human platelet aggregation with an IC50 of 0.95 nM. MRS2365, an (N)-methanocarba analogue of 2-MeSADP, displayed potency (EC50) of 0.4 nM at the P2Y1 receptor, with >10 000-fold selectivity in comparison to P2Y12 and P2Y13 receptors. At P2Y6 receptors there is a dramatic preference for the South conformation. Three-dimensional structures of P2Y receptors have been deduced from structure activity relationships (SAR), mutagenesis and modelling studies. Detailed three-dimensional structures of P2X receptors have not yet been proposed. PMID:16805423

  7. Peroxidative Metabolism of β2-Agonists Salbutamol and Fenoterol and Their Analogs

    PubMed Central

    Reszka, Krzysztof J.; McGraw, Dennis W.; Britigan, Bradley E.

    2009-01-01

    Phenolic β2-adrenoreceptor agonists salbutamol, fenoterol and terbutaline relax smooth muscle cells that relieve acute airway bronchospasm associated with asthma. Why their use sometimes fails to relieve bronchospasm, and why the drugs appear to be less effective in patients with severe asthma exacerbations, remains unclear. We show that in the presence of hydrogen peroxide, both myeloperoxidase, secreted by activated neutrophils present in inflamed airways, and lactoperoxidase, which is naturally present in the respiratory system, catalyze oxidation of these β2-agonists. Azide, cyanide, thiocyanate, ascorbate, glutathione, and methimazole inhibited this process, while methionine was without effect. Inhibition by ascorbate and glutathione was associated with their oxidation to corresponding radical species by the agonists’-derived phenoxyl radicals. Using electron paramagnetic resonance (EPR), we detected free radical metabolites from β2-agonists by spin trapping with 2-methyl-2-nitrosopropane (MNP). Formation of these radicals was inhibited by pharmacologically-relevant concentrations of methimazole and dapsone. In alkaline buffers radicals from fenoterol and its structural analog, metaproteronol, were detected by direct EPR. Analysis of these spectra suggests that oxidation of fenoterol and metaproterenol, but not terbutaline, causes their transformation through intramolecular cyclization by addition of their amino nitrogen to the aromatic ring. Together, these results indicate that phenolic β2-agonists function as substrates for airway peroxidases and that the resulting products differ in their structural and functional properties from their parent compounds. They also suggest that these transformations can be modulated by pharmacological approaches using appropriate peroxidase inhibitors or alternative substrates. These processes may affect therapeutic efficacy and also play a role in adverse reactions of the β2-agonists. PMID:19462961

  8. Effects of RXR Agonists on Cell Proliferation/Apoptosis and ACTH Secretion/Pomc Expression

    PubMed Central

    Saito-Hakoda, Akiko; Uruno, Akira; Yokoyama, Atsushi; Shimizu, Kyoko; Parvin, Rehana; Kudo, Masataka; Saito-Ito, Takako; Sato, Ikuko; Kogure, Naotaka; Suzuki, Dai; Shimada, Hiroki; Yoshikawa, Takeo; Fujiwara, Ikuma; Kagechika, Hiroyuki; Iwasaki, Yasumasa; Kure, Shigeo; Ito, Sadayoshi; Sugawara, Akira

    2015-01-01

    Various retinoid X receptor (RXR) agonists have recently been developed, and some of them have shown anti-tumor effects both in vivo and in vitro. However, there has been no report showing the effects of RXR agonists on Cushing’s disease, which is caused by excessive ACTH secretion in a corticotroph tumor of the pituitary gland. Therefore, we examined the effects of synthetic RXR pan-agonists HX630 and PA024 on the proliferation, apoptosis, ACTH secretion, and pro-opiomelanocortin (Pomc) gene expression of murine pituitary corticotroph tumor AtT20 cells. We demonstrated that both RXR agonists induced apoptosis dose-dependently in AtT20 cells, and inhibited their proliferation at their higher doses. Microarray analysis identified a significant gene network associated with caspase 3 induced by high dose HX630. On the other hand, HX630, but not PA024, inhibited Pomc transcription, Pomc mRNA expression, and ACTH secretion dose-dependently. Furthermore, we provide new evidence that HX630 negatively regulates the Pomc promoter activity at the transcriptional level due to the suppression of the transcription factor Nur77 and Nurr1 mRNA expression and the reduction of Nur77/Nurr1 heterodimer recruiting to the Pomc promoter region. We also demonstrated that the HX630-mediated suppression of the Pomc gene expression was exerted via RXRα. Furthermore, HX630 inhibited tumor growth and decreased Pomc mRNA expression in corticotroph tumor cells in female nude mice in vivo. Thus, these results indicate that RXR agonists, especially HX630, could be a new therapeutic candidate for Cushing’s disease. PMID:26714014

  9. Long-Acting Beta Agonists Enhance Allergic Airway Disease.

    PubMed

    Knight, John M; Mak, Garbo; Shaw, Joanne; Porter, Paul; McDermott, Catherine; Roberts, Luz; You, Ran; Yuan, Xiaoyi; Millien, Valentine O; Qian, Yuping; Song, Li-Zhen; Frazier, Vincent; Kim, Choel; Kim, Jeong Joo; Bond, Richard A; Milner, Joshua D; Zhang, Yuan; Mandal, Pijus K; Luong, Amber; Kheradmand, Farrah; McMurray, John S; Corry, David B

    2015-01-01

    Asthma is one of the most common of medical illnesses and is treated in part by drugs that activate the beta-2-adrenoceptor (β2-AR) to dilate obstructed airways. Such drugs include long acting beta agonists (LABAs) that are paradoxically linked to excess asthma-related mortality. Here we show that LABAs such as salmeterol and structurally related β2-AR drugs such as formoterol and carvedilol, but not short-acting agonists (SABAs) such as albuterol, promote exaggerated asthma-like allergic airway disease and enhanced airway constriction in mice. We demonstrate that salmeterol aberrantly promotes activation of the allergic disease-related transcription factor signal transducer and activator of transcription 6 (STAT6) in multiple mouse and human cells. A novel inhibitor of STAT6, PM-242H, inhibited initiation of allergic disease induced by airway fungal challenge, reversed established allergic airway disease in mice, and blocked salmeterol-dependent enhanced allergic airway disease. Thus, structurally related β2-AR ligands aberrantly activate STAT6 and promote allergic airway disease. This untoward pharmacological property likely explains adverse outcomes observed with LABAs, which may be overcome by agents that antagonize STAT6.

  10. Long-Acting Beta Agonists Enhance Allergic Airway Disease

    PubMed Central

    Knight, John M.; Mak, Garbo; Shaw, Joanne; Porter, Paul; McDermott, Catherine; Roberts, Luz; You, Ran; Yuan, Xiaoyi; Millien, Valentine O.; Qian, Yuping; Song, Li-Zhen; Frazier, Vincent; Kim, Choel; Kim, Jeong Joo; Bond, Richard A.; Milner, Joshua D.; Zhang, Yuan; Mandal, Pijus K.; Luong, Amber; Kheradmand, Farrah

    2015-01-01

    Asthma is one of the most common of medical illnesses and is treated in part by drugs that activate the beta-2-adrenoceptor (β2-AR) to dilate obstructed airways. Such drugs include long acting beta agonists (LABAs) that are paradoxically linked to excess asthma-related mortality. Here we show that LABAs such as salmeterol and structurally related β2-AR drugs such as formoterol and carvedilol, but not short-acting agonists (SABAs) such as albuterol, promote exaggerated asthma-like allergic airway disease and enhanced airway constriction in mice. We demonstrate that salmeterol aberrantly promotes activation of the allergic disease-related transcription factor signal transducer and activator of transcription 6 (STAT6) in multiple mouse and human cells. A novel inhibitor of STAT6, PM-242H, inhibited initiation of allergic disease induced by airway fungal challenge, reversed established allergic airway disease in mice, and blocked salmeterol-dependent enhanced allergic airway disease. Thus, structurally related β2-AR ligands aberrantly activate STAT6 and promote allergic airway disease. This untoward pharmacological property likely explains adverse outcomes observed with LABAs, which may be overcome by agents that antagonize STAT6. PMID:26605551

  11. Cannabidiol, a novel inverse agonist for GPR12.

    PubMed

    Brown, Kevin J; Laun, Alyssa S; Song, Zhao-Hui

    2017-09-06

    GPR12 is a constitutively active, Gs protein-coupled receptor that currently has no confirmed endogenous ligands. GPR12 may be involved in physiological processes such as maintenance of oocyte meiotic arrest and brain development, as well as pathological conditions such as metastatic cancer. In this study, the potential effects of various classes of cannabinoids on GPR12 were tested using a cAMP accumulation assay. Our data demonstrate that cannabidiol (CBD), a major non-psychoactive phytocannabinoid, acted as an inverse agonist to inhibit cAMP accumulation stimulated by the constitutively active GPR12. Thus, GPR12 is a novel molecular target for CBD. The structure-activity relationship studies of CBD indicate that both the free hydroxyl and the pentyl side chain are crucial for the effects of CBD on GPR12. Furthermore, studies using cholera toxin, which blocks Gs protein and pertussis toxin, which blocks Gi protein, revealed that Gs, but not Gi is involved in the inverse agonism of CBD on GPR12. CBD is a promising novel therapeutic agent for cancer, and GPR12 has been shown to alter viscoelasticity of metastatic cancer cells. Since we have demonstrated that CBD is an inverse agonist for GPR12, this provides novel mechanism of action for CBD, and an initial chemical scaffold upon which highly potent and efficacious agents acting on GPR12 may be developed with the ultimate goal of blocking cancer metastasis. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Cold Suppresses Agonist-induced Activation of TRPV1

    PubMed Central

    Chung, M.-K.; Wang, S.

    2011-01-01

    Cold therapy is frequently used to reduce pain and edema following acute injury or surgery such as tooth extraction. However, the neurobiological mechanisms of cold therapy are not completely understood. Transient receptor potential vanilloid 1 (TRPV1) is a capsaicin- and heat-gated nociceptive ion channel implicated in thermosensation and pathological pain under conditions of inflammation or injury. Although capsaicin-induced nociception, neuropeptide release, and ionic currents are suppressed by cold, it is not known if cold suppresses agonist-induced activation of recombinant TRPV1. We demonstrate that cold strongly suppressed the activation of recombinant TRPV1 by multiple agonists and capsaicin-evoked currents in trigeminal ganglia neurons under normal and phosphorylated conditions. Cold-induced suppression was partially impaired in a TRPV1 mutant that lacked heat-mediated activation and potentiation. These results suggest that cold-induced suppression of TRPV1 may share a common molecular basis with heat-induced potentiation, and that allosteric inhibition may contribute, in part, to the cold-induced suppression. We also show that combination of cold and a specific antagonist of TRPV1 can produce an additive suppression. Our results provide a mechanistic basis for cold therapy and may enhance anti-nociceptive approaches that target TRPV1 for managing pain under inflammation and tissue injury, including that from tooth extraction. PMID:21666106

  13. Cold suppresses agonist-induced activation of TRPV1.

    PubMed

    Chung, M-K; Wang, S

    2011-09-01

    Cold therapy is frequently used to reduce pain and edema following acute injury or surgery such as tooth extraction. However, the neurobiological mechanisms of cold therapy are not completely understood. Transient receptor potential vanilloid 1 (TRPV1) is a capsaicin- and heat-gated nociceptive ion channel implicated in thermosensation and pathological pain under conditions of inflammation or injury. Although capsaicin-induced nociception, neuropeptide release, and ionic currents are suppressed by cold, it is not known if cold suppresses agonist-induced activation of recombinant TRPV1. We demonstrate that cold strongly suppressed the activation of recombinant TRPV1 by multiple agonists and capsaicin-evoked currents in trigeminal ganglia neurons under normal and phosphorylated conditions. Cold-induced suppression was partially impaired in a TRPV1 mutant that lacked heat-mediated activation and potentiation. These results suggest that cold-induced suppression of TRPV1 may share a common molecular basis with heat-induced potentiation, and that allosteric inhibition may contribute, in part, to the cold-induced suppression. We also show that combination of cold and a specific antagonist of TRPV1 can produce an additive suppression. Our results provide a mechanistic basis for cold therapy and may enhance anti-nociceptive approaches that target TRPV1 for managing pain under inflammation and tissue injury, including that from tooth extraction.

  14. The Glycine Transport Inhibitor Sarcosine Is an Inhibitory Glycine Receptor Agonist

    PubMed Central

    Zhang, Hai Xia; Lyons-Warren, Ariel; Thio, Liu Lin

    2009-01-01

    Summary Sarcosine is an endogenous amino acid that is a competitive inhibitor of the type I glycine transporter (GlyT1), an N-methyl-D-aspartate receptor (NMDAR) co-agonist, and an important intermediate in one-carbon metabolism. Its therapeutic potential for schizophrenia further underscores its clinical importance. The structural similarity between sarcosine and glycine and sarcosine's ability to serve as an NMDAR co-agonist led us to examine whether sarcosine is also an agonist at the inhibitory glycine receptor (GlyR). We examined this possibility using whole-cell recordings from cultured embryonic mouse hippocampal neurons and found that sarcosine evoked a dose-dependent, strychnine sensitive, Cl- current that cross-inhibited glycine currents. Sarcosine evoked this current with Li+ in the extracellular solution to block GlyT1, in neurons treated with the essentially irreversible GlyT1 inhibitor N[3-(4′-fluorophenyl)-3-(4′-phenylphenoxy)propyl]sarcosine (NFPS), and in neurons plated in the absence of glia. These results indicate that the sarcosine currents did not result from GlyT1 inhibition or heteroexchange. We conclude that sarcosine is a GlyR agonist. PMID:19619564

  15. Peroxisome Proliferator–Activated Receptor-γ Agonists Prevent In Vivo Remodeling of Human Artery Induced by Alloreactive T Cells

    PubMed Central

    Tobiasova, Zuzana; Zhang, Lufeng; Yi, Tai; Qin, Linfeng; Manes, Thomas D.; Kulkarni, Sanjay; Lorber, Marc I.; Rodriguez, Frederick C.; Choi, Je-Min; Tellides, George; Pober, Jordan S.; Kawikova, Ivana; Bothwell, Alfred L.M.

    2012-01-01

    Background Ligands activating the transcription factor peroxisome proliferator–activated receptor-γ (PPARγ) have antiinflammatory effects. Vascular rejection induced by allogeneic T cells can be responsible for acute and chronic graft loss. Studies in rodents suggest that PPARγ agonists may inhibit graft vascular rejection, but human T-cell responses to allogeneic vascular cells differ from those in rodents, and the effects of PPARγ in human transplantation are unknown. Methods and Results We tested the effects of PPARγ agonists on human vascular graft rejection using a model in which human artery is interposed into the abdominal aorta of immunodeficient mice, followed by adoptive transfer of allogeneic (to the artery donor) human peripheral blood mononuclear cells. Interferon-γ–dependent rejection ensues within 4 weeks, characterized by intimal thickening, T-cell infiltrates, and vascular cell activation, a response resembling clinical intimal arteritis. The PPARγ agonists 15-deoxy-prostaglandin-J2, ciglitazone, and pioglitazone reduced intimal expansion, intimal infiltration of CD45RO+ memory T cells, and plasma levels of inflammatory cytokines. The PPARγ antagonist GW9662 reversed the protective effects of PPARγ agonists, confirming the involvement of PPARγ-mediated pathways. In vitro, pioglitazone inhibited both alloantigen-induced proliferation and superantigen-induced transendothelial migration of memory T cells, indicating the potential mechanisms of PPARγ effects. Conclusion Our results suggest that PPARγ agonists inhibit allogeneic human memory T cell responses and may be useful for the treatment of vascular graft rejection. PMID:21690493

  16. Estrogen agonist/antagonist properties of dibenzyl phthalate (DBzP) based on in vitro and in vivo assays.

    PubMed

    Zhang, Zhaobin; Hu, Ying; Zhao, Liang; Li, Jun; Bai, Huicheng; Zhu, Desheng; Hu, Jianying

    2011-11-10

    The most commonly used phthalates have been banned or restricted for use as plasticizers in toys in some countries because of their endocrine-disrupting properties. Dibenzyl phthalate (DBzP) has been proposed as a possible alternative for the banned/restricted phthalates. In this study, the estrogen agonist/antagonist properties of DBzP were predicted by molecular docking and confirmed by yeast estrogen screen (YES) and immature mouse uterotrophic assays. The YES assay results showed a dose-dependent increase in DBzP estrogen agonist activity from 10⁻⁶ to 10⁻⁴ M, and at concentrations from 1.95×10⁻⁶ M to higher, DBzP significantly inhibited the agonist activity of 10⁻⁹ M 17β-estradiol (E₂), inhibiting 10⁻⁹ M E₂ by 74.5% at its maximum effectiveness. The in vivo estrogen agonist/antagonist activities of DBzP were demonstrated in immature mouse uterotrophic assays. The antagonist activity of DBzP inhibited E₂-induced uterine growth promoted at 40 and 400 μg/kg bw (body weight) (P<0.05). In addition, we also analyzed the estrogen agonist/antagonist potentials of benzyl butyl phthalate (BBP) by YES, and found both were weaker than those of DBzP, suggesting DBzP would be more toxic than BBP and should not be used as an alternative plasticizer.

  17. Salvinorin A, an active component of the hallucinogenic sage salvia divinorum is a highly efficacious kappa-opioid receptor agonist: structural and functional considerations.

    PubMed

    Chavkin, Charles; Sud, Sumit; Jin, Wenzhen; Stewart, Jeremy; Zjawiony, Jordan K; Siebert, Daniel J; Toth, Beth Ann; Hufeisen, Sandra J; Roth, Bryan L

    2004-03-01

    The diterpene salvinorin A from Salvia divinorum has recently been reported to be a high-affinity and selective kappa-opioid receptor agonist (Roth et al., 2002). Salvinorin A and selected derivatives were found to be potent and efficacious agonists in several measures of agonist activity using cloned human kappa-opioid receptors expressed in human embryonic kidney-293 cells. Thus, salvinorin A, salvinorinyl-2-propionate, and salvinorinyl-2-heptanoate were found to be either full (salvinorin A) or partial (2-propionate, 2-heptanoate) agonists for inhibition of forskolin-stimulated cAMP production. Additional studies of agonist potency and efficacy of salvinorin A, performed by cotransfecting either the chimeric G proteins Gaq-i5 or the universal G protein Ga16 and quantification of agonist-evoked intracellular calcium mobilization, affirmed that salvinorin A was a potent and effective kappa-opioid agonist. Results from structure-function studies suggested that the nature of the substituent at the 2-position of salvinorin A was critical for kappa-opioid receptor binding and activation. Because issues of receptor reserve complicate estimates of agonist efficacy and potency, we also examined the agonist actions of salvinorin A by measuring potassium conductance through G protein-gated K(+) channels coexpressed in Xenopus oocytes, a system in which receptor reserve is minimal. Salvinorin A was found to be a full agonist, being significantly more efficacious than (trans)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl] benzeneacetamide methane-sulfonate hydrate (U50488) or (trans)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl] benzeneacetamide methane-sulfonate hydrate (U69593) (two standard kappa-opioid agonists) and similar in efficacy to dynorphin A (the naturally occurring peptide ligand for kappa-opioid receptors). Salvinorin A thus represents the first known naturally occurring non-nitrogenous full agonist at kappa-opioid receptors.

  18. Local Delivery of the Toll-Like Receptor 9 Ligand CpG Downregulates Host Immune and Inflammatory Responses, Ameliorating Established Leishmania (Viannia) panamensis Chronic Infection.

    PubMed

    Ehrlich, Allison K; Fernández, Olga L; Rodriguez-Pinto, Daniel; Castilho, Tiago M; Corral Caridad, Maria J; Goldsmith-Pestana, Karen; Saravia, Nancy Gore; McMahon-Pratt, Diane

    2017-03-01

    Infection by Leishmania (Viannia) panamensis, the predominant etiologic agent for cutaneous leishmaniasis in Colombia, is characterized by a chronic mixed inflammatory response. Current treatment options are plagued by toxicity, lengthy treatment regimens, and growing evidence of drug resistance. Immunotherapy, modulating the immune system to mount a protective response, may provide an alternate therapeutic approach. We investigated the ability of the Toll-like receptor 9 (TLR9) ligand CpG to modulate established disease in the L (V) panamensis mouse model. Treatment of established infection with a high dose (50 μg) of CpG ameliorated disease and lowered parasite burden. Interestingly, immediately after treatment there was a significant increase in transforming growth factor β (TGF-β) and concomitantly an increase in T regulatory cell (Treg) function. Although a general reduction in cell-mediated immune cytokine and chemokine (gamma interferon [IFN-γ], interleukin 10 [IL-10], IL-13, IL-6, granulocyte-macrophage colony-stimulating factor [GM-CSF], IL-4, and MIP-1α) responses of the treated mice was observed, certain chemokines (RANTES, monocyte chemoattractant protein 1[MCP-1], and IP-10) were increased. Further, in peripheral blood mononuclear cells (PBMCs) from patients with cutaneous leishmaniasis, CpG treatment similarly exhibited a dose-response effect on the production of IFN-γ, IL-17, IL-10, and IL-13, with reductions observed at higher doses. To further understand the underlying mechanisms and cell populations driving the CpG mediated response, we examined the ex vivo dose effects mediated by the TLR9(+) cell populations (dendritic cells, macrophages, and B cells) found to accumulate labeled CpG in vivo Notably, B cells altered the production of IL-17, IL-13, and IFN-γ, supporting a role for B cells functioning as antigen-presenting cells (APCs) and/or regulatory cells during infection. Interestingly, B cells have been previously demonstrated as a

  19. CpG-ODN 7909 increases radiation sensitivity of radiation-resistant human lung adenocarcinoma cell line by overexpression of Toll-like receptor 9.

    PubMed

    Yan, Li; Xu, Guoxiong; Qiao, Tiankui; Chen, Wei; Yuan, Sujuan; Li, Xuan

    2013-09-01

    Radioresistance is one of the main reasons for the failure of radiotherapy in lung cancer. The aim of this study was to establish a radiation-resistant lung cancer cell line, to evaluate whether CpG oligodeoxyribonucleotide (CpG-ODN) 7909 could increase its radiosensitivity and to explore the relevant mechanisms. The radioresistant cell line, referred to as R-A549, was generated by reduplicative fractionated irradiation from the human lung adenocarcinoma cell line A549. The radioresistance of R-A549 cells were confirmed by the Cell Counting Kit-8 (CCK-8), cell viability assay, and clonogenic assay. Cell growth kinetics, morphological feature, and radiosensitivity were compared between the original A549 cells and R-A549 cells treated with or without CpG-ODN 7909 or radiation. To further explore the potential mechanisms of radiosensitivity, the cell cycle distributions and the expression of Toll-like receptor 9 (TLR-9) were examined by Western blot and flow cytometry. The R-A549 cell line was generated and its radioresistance was further confirmed. CpG-ODN 7909 was found to increase much more radiosensitivity of R-A549 cells under combined treatments with CpG-ODN 7909 and radiation compared with its control group without any treatments. They presented their respective D0 1.33 ± 0.20 Gy versus 1.76 ± 0.25 Gy with N 3.44 ± 1.01 versus 4.96 ± 0.32. Further, there was a larger cell population of R-A549 cells under combined treatment in the G2/M phase compared with the control group after treatment with CpG-ODN7909 or radiation alone at 24 and 48 hour. The expression level of TLR-9 in R-A549 cells was found higher than in A549 cells. These results suggested that CpG-ODN 7909 increased the radiosensitivity of R-A549 cells, which might be mediated via the upregulated TLR-9 and prolonged cell cycle arrest in the G2/M phase compared with A549 cells.

  20. Local Delivery of the Toll-Like Receptor 9 Ligand CpG Downregulates Host Immune and Inflammatory Responses, Ameliorating Established Leishmania (Viannia) panamensis Chronic Infection

    PubMed Central

    Fernández, Olga L.; Rodriguez-Pinto, Daniel; Castilho, Tiago M.; Corral Caridad, Maria J.; Goldsmith-Pestana, Karen; Saravia, Nancy Gore; McMahon-Pratt, Diane

    2017-01-01

    ABSTRACT Infection by Leishmania (Viannia) panamensis, the predominant etiologic agent for cutaneous leishmaniasis in Colombia, is characterized by a chronic mixed inflammatory response. Current treatment options are plagued by toxicity, lengthy treatment regimens, and growing evidence of drug resistance. Immunotherapy, modulating the immune system to mount a protective response, may provide an alternate therapeutic approach. We investigated the ability of the Toll-like receptor 9 (TLR9) ligand CpG to modulate established disease in the L. (V.) panamensis mouse model. Treatment of established infection with a high dose (50 μg) of CpG ameliorated disease and lowered parasite burden. Interestingly, immediately after treatment there was a significant increase in transforming growth factor β (TGF-β) and concomitantly an increase in T regulatory cell (Treg) function. Although a general reduction in cell-mediated immune cytokine and chemokine (gamma interferon [IFN-γ], interleukin 10 [IL-10], IL-13, IL-6, granulocyte-macrophage colony-stimulating factor [GM-CSF], IL-4, and MIP-1α) responses of the treated mice was observed, certain chemokines (RANTES, monocyte chemoattractant protein 1[MCP-1], and IP-10) were increased. Further, in peripheral blood mononuclear cells (PBMCs) from patients with cutaneous leishmaniasis, CpG treatment similarly exhibited a dose-response effect on the production of IFN-γ, IL-17, IL-10, and IL-13, with reductions observed at higher doses. To further understand the underlying mechanisms and cell populations driving the CpG mediated response, we examined the ex vivo dose effects mediated by the TLR9+ cell populations (dendritic cells, macrophages, and B cells) found to accumulate labeled CpG in vivo. Notably, B cells altered the production of IL-17, IL-13, and IFN-γ, supporting a role for B cells functioning as antigen-presenting cells (APCs) and/or regulatory cells during infection. Interestingly, B cells have been previously

  1. Phase II study of a TLR-9 agonist (1018 ISS) with rituximab in patients with relapsed or refractory follicular lymphoma.

    PubMed

    Friedberg, Jonathan W; Kelly, Jennifer L; Neuberg, Donna; Peterson, Derick R; Kutok, Jeffery L; Salloum, Rabih; Brenn, Thomas; Fisher, David C; Ronan, Elizabeth; Dalton, Virginia; Rich, Lynn; Marquis, Diana; Sims, Paul; Rothberg, Paul G; Liesveld, Jane; Fisher, Richard I; Coffman, Robert; Mosmann, Tim; Freedman, Arnold S

    2009-08-01

    Toll-like receptor-9 (TLR-9) agonists have pleotropic effects on both the innate and adaptive immune systems, including increased antigen expression, enhanced antibody-dependent cell-mediated cytotoxicity (ADCC) and T helper cell type 1 shift in the immune response. We combined a TLR-9 agonist (1018 ISS, 0.2 mg/kg sc weekly x 4 beginning day 8) with standard rituximab (375 mg/m(2) weekly x 4) in patients (n = 23) with relapsed/refractory, histologically confirmed follicular lymphoma, and evaluated immunological changes following the combination. Treatment was well-tolerated with no significant adverse events attributable to therapy. Clinical responses were observed in 48% of patients; the overall median progression-free survival was 9 months. Biologically relevant increases in ADCC and circulating CD-3 positive T cells were observed in 35% and 39% of patients, respectively. Forty-five percent of patients had increased T cells and dendritic cells in skin biopsies of 1018 ISS injection sites 24 h post-therapy. Pre- and post-biopsies of tumour tissue demonstrated an infiltration of CD8(+) T cells and macrophages following treatment. This group of patients had favourable clinical outcome despite adverse prognostic factors. This study is the first to histologically confirm perturbation of the local immune microenvironment following systemic biological therapy of follicular lymphoma.

  2. Phase II study of a TLR-9 agonist (1018 ISS) with rituximab in patients with relapsed or refractory follicular lymphoma

    PubMed Central

    Friedberg, Jonathan W; Kelly, Jennifer L; Neuberg, Donna; Peterson, Derick R.; Kutok, Jeffery L; Salloum, Rabih; Brenn, Thomas; Fisher, David C; Ronan, Elizabeth; Dalton, Virginia; Rich, Lynn; Marquis, Diana; Sims, Paul; Rothberg, Paul G.; Liesveld, Jane; Fisher, Richard I; Coffman, Robert; Mosmann, Tim; Freedman, Arnold S

    2009-01-01

    Summary Toll-Like Receptor-9 (TLR-9) agonists have pleotropic effects on both the innate and adaptive immune systems, including increased antigen expression, enhanced antibody-dependent cell-mediated cytotoxicity (ADCC) and T helper cell type 1 shift in the immune response. We combined a TLR-9 agonist (1018 ISS, 0.2 mg/kg sc weekly × 4 beginning day 8) with standard rituximab (375mg/m2 weekly × 4) in patients (n=23) with relapsed/refractory, histologically confirmed follicular lymphoma, and evaluated immunological changes following the combination. Treatment was well-tolerated with no significant adverse events attributable to therapy. Clinical responses were observed in 48% of patients; the overall median progression-free survival was 9 months. Biologically relevant increases in ADCC and circulating CD-3 positive T cells were observed in 35% and 39% of patients, respectively. 45% of patients had increased T cells and dendritic cells in skin biopsies of 1018 ISS injection sites 24 h post-therapy. Pre-and post-biopsies of tumour tissue demonstrated an infiltration of CD8+ T cells and macrophages following treatment. This group of patients had favourable clinical outcome despite adverse prognostic factors. This study is the first to histologically confirm perturbation of the local immune microenvironment following systemic biological therapy of follicular lymphoma. PMID:19519691

  3. A behavioural and biochemical study in rats of 5-hydroxytryptamine receptor agonists and antagonists, with observations on structure-activity requirements for the agonists

    PubMed Central

    Green, A.R.; Hall, J.E.; Rees, A.R.

    1981-01-01

    1 The effect of the putative 5-hydroxytryptamine (5-HT) receptor antagonists, methysergide, methergoline, mianserin, cyproheptadine, cinanserin (all at 10 mg/kg), methiothepin (5 mg/kg) and (-)-propranolol (20 mg/kg) on the behavioural responses to tranylcypromine (10 mg/kg) followed 30 min later by L-tryptophan (100 mg/kg) was examined. 2 Methysergide, methergoline, methiothepin and (-)-propranolol inhibited head weaving, forepaw treading and hind-limb abduction. Methysergide and methergoline increased reactivity. In contrast, cypropheptadine, cinanserin and mianserin had no effects on the behaviour. 3 Similar findings were obtained when the behaviours were elicited by administration of tranylcypromine (10 mg/kg) followed by the putative 5-HT receptor agonist, 5-methoxy-N,N-dimethyltryptamine (5-MeODMT) (2 mg/kg). 4 When the behaviours were elicited by the putative 5-HT receptor agonist, quipazine (50 mg/kg), all the drugs effectively inhibited head weaving and forepaw treading. 5 When the dose of cypropheptadine was doubled to 20 mg/kg an inhibition of the tranylcypromine/L-tryptophan induced behaviours was seen. 6 Methiothepin produced a marked inhibition of apomorphine-induced locomotor activity whilst all the others enhanced this response, suggesting that only methiothepin inhibits the 5-HT behaviours by dopamine antagonism and that the increased reactivity seen following tranylcypromine/L-tryptophan after pretreatment with methysergide or methergoline might be due to enhanced dopamine function. 7 Pretreatment with p-chlorophenylalanine resulted in enhanced behavioural responses to both 5-MeODMT and quipazine. 8 Both methergoline and methiothepin decreased the rate of 5-HT synthesis in whole brain but not spinal cord and methergoline decreased spinal cord 5-HIAA concentration. None of the other drugs had any significant effects on the concentration of 5-HT, 5-HIAA or 5-HT synthesis rate in brain or spinal cord. 9 Experiments with compounds structurally related

  4. Saralasin and Sarile Are AT2 Receptor Agonists

    PubMed Central

    2014-01-01

    Saralasin and sarile, extensively studied over the past 40 years as angiotensin II (Ang II) receptor blockers, induce neurite outgrowth in a NG108-15 cell assay to a similar extent as the endogenous Ang II. In their undifferentiated state, these cells express mainly the AT2 receptor. The neurite outgrowth was inhibited by preincubation with the AT2 receptor selective antagonist PD 123,319, which suggests that the observed outgrowth was mediated by the AT2 receptor. Neither saralasin nor sarile reduced the neurite outgrowth induced by Ang II proving that the two octapeptides do not act as antagonists at the AT2 receptor and may be considered as AT2 receptor agonists. PMID:25313325

  5. Tricyclic Spirolactones as Modular TRPV1 Synthetic Agonists.

    PubMed

    Mostinski, Yelena; Noy, Gilad; Kumar, Rakesh; Tsvelikhovsky, Dmitry; Priel, Avi

    2017-08-16

    TRPV1 is a prominent signal integrator of the pain system, known to be activated by vanilloids, a family of endogenous and exogenous pain-evoking molecules, through the vanilloid-binding site (VBS). The extensive preclinical profiling of small molecule inhibitors provides intriguing evidence that TRPV1 inhibition can be a useful therapeutic approach. However, the dissimilarity of chemical species that activate TRPV1 creates a major obstacle to understanding the molecular mechanism of pain induction, which is viewed as a pivotal trait of the somatosensory system. Here, we establish the existence of a unique family of synthetic agonists that interface with TRPV1 through the VBS, containing none of the molecular domains previously believed to be required for this interaction. The overarching value obtained from our inquiry is the novel advancement of the existing TRPV1 activation model. These findings uncover new potential in the area of pain treatment, providing a novel synthetic platform.

  6. Muscimol as an ionotropic GABA receptor agonist.

    PubMed

    Johnston, Graham A R

    2014-10-01

    Muscimol, a psychoactive isoxazole from Amanita muscaria and related mushrooms, has proved to be a remarkably selective agonist at ionotropic receptors for the inhibitory neurotransmitter GABA. This historic overview highlights the discovery and development of muscimol and related compounds as a GABA agonist by Danish and Australian neurochemists. Muscimol is widely used as a ligand to probe GABA receptors and was the lead compound in the development of a range of GABAergic agents including nipecotic acid, tiagabine, 4,5,6,7-tetrahydroisoxazolo(5,4-c)pyridin-3-ol, (Gaboxadol(®)) and 4-PIOL.

  7. The CRTH2 agonist Pyl A prevents lipopolysaccharide-induced fetal death but induces preterm labour

    PubMed Central

    Sykes, Lynne; Herbert, Bronwen R; MacIntyre, David A; Hunte, Emma; Ponnampalam, Sathana; Johnson, Mark R; Teoh, Tiong G; Bennett, Phillip R

    2013-01-01

    We have previously demonstrated that the anti-inflammatory prostaglandin 15-deoxy-Δ 12,14-prostaglandin J2 (15dPGJ2) delays inflammation-induced preterm labour in the mouse and improves pup survival through the inhibition of nuclear factor-κB (NF-κB) by a mechanism yet to be elucidated. 15dPGJ2 is an agonist of the second prostaglandin D2 receptor, chemoattractant receptor homologous to the T helper 2 cell (CRTH2). In human T helper cells CRTH2 agonists induce the production of the anti-inflammatory interleukins IL-10 and IL-4. We hypothesized that CRTH2 is involved in the protective effect of 15dPGJ2 in inflammation-induced preterm labour in the murine model. We therefore studied the effects of a specific small molecule CRTH2 agonist on preterm labour and pup survival. An intrauterine injection of lipopolysaccharide (LPS) was administered to CD1 mice at embryonic day 16, ± CRTH2 agonist/vehicle controls. Mice were killed at 4.5 hr to assess fetal wellbeing and to harvest myometrium and pup brain for analysis of NF-κB, and T helper type 1/2 interleukins. To examine the effects of the CRTH2 agonist on LPS-induced preterm labour, mice were allowed to labour spontaneously. Direct effects of the CRTH2 agonist on uterine contractility were examined ex vivo on contracting myometrial strips. The CRTH2 agonist increased fetal survival from 20 to 100% in LPS-treated mice, and inhibited circular muscle contractility ex vivo. However, it augmented LPS-induced labour and significantly increased myometrial NF-κB, IL-1β, KC-GRO, interferon-γ and tumour necrosis factor-α. This suggests that the action of 15dPGJ2 is not via CRTH2 and therefore small molecule CRTH2 agonists are not likely to be beneficial for the prevention of inflammation-induced preterm labour. PMID:23374103

  8. Regulation of noradrenaline release from rat occipital cortex tissue chops by alpha 2-adrenergic agonists.

    PubMed

    Ong, M L; Ball, S G; Vaughan, P F

    1991-04-01

    Noradrenaline (NA) and the alpha 2-adrenergic agonists clonidine, BHT-920, and UK 14304-18 inhibit potassium-evoked release of [3H]NA from rat occipital cortex tissue chops with similar potencies. NA (10(-5) M) was most effective as up to 85% inhibition could be observed compared with 75%, 55%, and 35% for UK 14304-18, clonidine, and BHT-920, respectively, all at 10(-5) M. Potassium-evoked release was enhanced by both forskolin (10(-5) M) and 1 mM dibutyryl cyclic AMP. Pretreatment of tissue chops with 1 mM dibutyryl cyclic AMP in the presence of 3-isobutyl-1-methylxanthine partially reversed the alpha 2-adrenergic agonist inhibition of NA release. No reversal of inhibition was observed following pretreatment with 10(-5) M forskolin. The effects of clonidine, BHT-920, UK-14308-18, and NA on cyclic AMP formation stimulated by (a) forskolin, (b) isoprenaline, (c) adenosine, (d) potassium, and (e) NA were examined. Only cAMP formation stimulated by NA was inhibited by these alpha 2-adrenergic agonists. These results suggest that only a small fraction of adenylate cyclase in rat occipital cortex is coupled to alpha 2-adrenergic receptors. These results are discussed in relation to recent findings that several alpha 2-adrenergic receptor subtypes occur, not all of which are coupled to the inhibition of adenylate cyclase, and that alpha 2-adrenergic receptors inhibit NA release in rat occipital cortex by a mechanism that does not involve decreasing cyclic AMP levels.

  9. Immune Response Modulation of Conjugated Agonists with Changing Linker Length.

    PubMed

    Ryu, Keun Ah; Slowinska, Katarzyna; Moore, Troy; Esser-Kahn, Aaron

    2016-12-16

    We report immune response modulation with linked Toll-like receptor (TLR) agonists. Conjugating two agonists of synergistic TLRs induce an increase in immune activity compared to equal molarity of soluble agonists. Additionally, varying the distance between the agonists by changing the linker length alters the level of macrophage NF-κB activity as well as primary bone marrow derived dendritic cell IL-6 production. This modulation is effected by the size of the agonists and the pairing of the stimulated TLRs. The sensitivity of linker-length-dependent immune activity of conjugated agonists provides the potential for developing application specific therapeutics.

  10. Mechanical stress activates NMDA receptors in the absence of agonists

    PubMed Central

    Maneshi, Mohammad Mehdi; Maki, Bruce; Gnanasambandam, Radhakrishnan; Belin, Sophie; Popescu, Gabriela K.; Sachs, Frederick; Hua, Susan Z.

    2017-01-01

    While studying the physiological response of primary rat astrocytes to fluid shear stress in a model of traumatic brain injury (TBI), we found that shear stress induced Ca2+ entry. The influx was inhibited by MK-801, a specific pore blocker of N-Methyl-D-aspartic acid receptor (NMDAR) channels, and this occurred in the absence of agonists. Other NMDA open channel blockers ketamine and memantine showed a similar effect. The competitive glutamate antagonists AP5 and GluN2B-selective inhibitor ifenprodil reduced NMDA-activated currents, but had no effect on the mechanically induced Ca2+ influx. Extracellular Mg2+ at 2 mM did not significantly affect the shear induced Ca2+ influx, but at 10 mM it produced significant inhibition. Patch clamp experiments showed mechanical activation of NMDAR and inhibition by MK-801. The mechanical sensitivity of NMDARs may play a role in the normal physiology of fluid flow in the glymphatic system and it has obvious relevance to TBI. PMID:28045032

  11. Design, synthesis, and biological evaluation of new 5-HT4 receptor agonists: application as amyloid cascade modulators and potential therapeutic utility in Alzheimer's disease.

    PubMed

    Russo, Olivier; Cachard-Chastel, Marthe; Rivière, Céline; Giner, Mireille; Soulier, Jean-Louis; Berthouze, Magali; Richard, Tristan; Monti, Jean-Pierre; Sicsic, Sames; Lezoualc'h, Frank; Berque-Bestel, Isabelle

    2009-04-23

    Serotonin 5-HT(4) receptor (5-HT(4)R) agonists are of particular interest for the treatment of Alzheimer's disease because of their ability to ameliorate cognitive deficits and to modulate production of amyloid beta-protein (Abeta). However, despite the range of 5-HT(4)R agonists synthesized to date, potent and selective 5-HT(4)R agonists are still lacking. In the present study, two libraries of molecules based on the scaffold of ML10302, a highly specific and partial 5-HT(4)R agonist, were efficiently prepared by parallel supported synthesis and their binding affinities and agonist activities evaluated. Furthermore, we showed that, in vivo, the two best candidates exhibited neuroprotective activity by increasing the level of the soluble form of the amyloid precursor protein (sAPPalpha) in the cortex and hippocampus of mice. Interestingly, one of these compounds could also inhibit Abeta fibril formation in vitro.

  12. Suppression by an RAR-γ Agonist of Collagen Degradation Mediated by Corneal Fibroblasts.

    PubMed

    Kimura, Kazuhiro; Zhou, Hongyan; Orita, Tomoko; Kobayashi, Masaaki; Nishida, Teruo; Sonoda, Koh-Hei

    2017-04-01

    To examine the role of retinoic acid receptor (RAR) isoforms in interleukin-1β (IL-1β)-induced collagen degradation by corneal fibroblasts. Primary rabbit corneal fibroblasts embedded in a three-dimensional collagen gel were incubated with or without all-trans retinoic acid (ATRA), the RAR-α agonist Am580, the RAR-β agonist AC55649, or the RAR-γ agonist R667. Collagen degradation was determined by measurement of hydroxyproline produced in acid hydrolysates of culture supernatants. Matrix metalloproteinase (MMP) expression was evaluated by immunoblot analysis and gelatin zymography. The phosphorylation of mitogen-activated protein kinases (MAPKs) and the endogenous nuclear factor (NF)-κB inhibitor IκB-α was examined by immunoblot analysis. Cell proliferation was measured with a bromodeoxyuridine incorporation assay, and cell viability was determined by measurement of the release of lactate dehydrogenase. Interleukin-1β-induced collagen degradation by corneal fibroblasts was inhibited by ATRA, Am580, and R667 in a concentration-dependent manner but was unaffected by AC55649, with the inhibitory effects of ATRA and R667 being markedly greater than that of Am580. The IL-1β-induced production of MMP-1, MMP-2, MMP-3, and MMP-9 by corneal fibroblasts was also inhibited by R667 in a concentration-dependent manner. R667 inhibited the IL-1β-induced phosphorylation of IκB-α but not that of MAPKs. R667 had no effect on the proliferation or viability of corneal fibroblasts. The RAR-γ agonist R667 suppressed MMP production and thereby inhibited collagen degradation by corneal fibroblasts exposed to the proinflammatory cytokine IL-1β. These effects of R667 may be mediated by the NF-κB signaling pathway.

  13. Melatonin and its agonists: an update.

    PubMed

    Arendt, Josephine; Rajaratnam, Shantha M W

    2008-10-01

    The pineal hormone melatonin is able to shift the timing of circadian rhythms, including the sleep-wake cycle, and to promote sleep. Melatonin agonists with similar properties have therapeutic potential for the treatment of circadian rhythm sleep disorders. Depression is specifically targeted by agomelatine, which is also a serotonin-2C (5-HT(2C)) antagonist.

  14. Multiple tyrosine metabolites are GPR35 agonists.

    PubMed

    Deng, Huayun; Hu, Haibei; Fang, Ye

    2012-01-01

    Both kynurenic acid and 2-acyl lysophosphatidic acid have been postulated to be the endogenous agonists of GPR35. However, controversy remains whether alternative endogenous agonists exist. The molecular targets accounted for many nongenomic actions of thyroid hormones are mostly unknown. Here we report the agonist activity of multiple tyrosine metabolites at the GPR35. Tyrosine metabolism intermediates that contain carboxylic acid and/or catechol functional groups were first selected. Whole cell dynamic mass redistribution (DMR) assays enabled by label-free optical biosensor were then used to characterize their agonist activity in native HT-29. Molecular assays including β-arrestin translocation, ERK phosphorylation and receptor internalization confirmed that GPR35 functions as a receptor for 5,6-dihydroxyindole-2-carboxylic acid, 3,3',5'-triiodothyronine, 3,3',5-triiodothyronine, gentisate, rosmarinate, and 3-nitrotyrosine. These results suggest that multiple tyrosine metabolites are alternative endogenous ligands of GPR35, and GPR35 may represent a druggable target for treating certain diseases associated with abnormality of tyrosine metabolism.

  15. Corepressors of agonist-bound nuclear receptors

    SciTech Connect

    Gurevich, Igor; Aneskievich, Brian J.

    2007-09-15

    Nuclear receptors (NRs) rely on coregulator proteins to modulate transcription of target genes. NR coregulators can be broadly subdivided into coactivators which potentiate transcription and corepressors which silence gene expression. The prevailing view of coregulator action holds that in the absence of agonist the receptor interacts with a corepressor via the corepressor nuclear receptor (CoRNR, 'corner') box motifs within the corepressor. Upon agonist binding, a conformational change in the receptor causes the shedding of corepressor and the binding of a coactivator which interacts with the receptor via NR boxes within the coregulator. This view was challenged with the discovery of RIP140 which acts as a NR corepressor in the presence of agonist and utilizes NR boxes. Since then a number of other corepressors of agonist-bound NRs have been discovered. Among them are LCoR, PRAME, REA, MTA1, NSD1, and COPR1 Although they exhibit a great diversity of structure, mechanism of repression and pathophysiological function, these corepressors frequently have one or more NR boxes and often recruit histone deacetylases to exert their repressive effects. This review highlights these more recently discovered corepressors and addresses their potential functions in transcription regulation, disease pharmacologic responses and xenobiotic metabolism.

  16. Direct antiatherosclerotic effects of PPAR agonists.

    PubMed

    Jandeleit-Dahm, Karin A M; Calkin, Anna; Tikellis, Chris; Thomas, Merlin

    2009-02-01

    Peroxisome proliferator activated receptors (PPARs) are ligand-dependent transcription factors that mediate a range of important metabolic functions by transactivation, transrepression or corepression of various gene targets. PPAR agonists also have direct antiatherosclerotic effects, independent of their metabolic effects on glucose and lipid homeostasis. The purpose of this review is to evaluate the currently available evidence for a direct vasculoprotective effect of PPAR agonists. Current studies have emphasized PPAR-mediated effects on inflammatory and immune responses, oxidative stress, the renin-angiotensin system and modulation of plaque composition. Furthermore, it has become evident that the relative activation of the different PPAR isoforms and the contribution of transactivation of target genes against transrepression of transcription factors need to be considered when assessing the vasculoprotective effects of PPAR agonists. It is anticipated that the antiatherosclerotic effects of PPAR agonists observed in experimental studies will translate into reduced cardiovascular events. This promise is yet to be realized in short-to-medium term studies. Given the central role of the PPAR in gene regulation, particularly in metabolic states, it is possible that more targeted modulation of PPAR signalling may hold many rewards for the prevention of atherosclerosis.

  17. Reciprocity of agonistic support in ravens.

    PubMed

    Fraser, Orlaith N; Bugnyar, Thomas

    2012-01-01

    Cooperative behaviour through reciprocation or interchange of valuable services in primates has received considerable attention, especially regarding the timeframe of reciprocation and its ensuing cognitive implications. Much less, however, is known about reciprocity in other animals, particularly birds. We investigated patterns of agonistic support (defined as a third party intervening in an ongoing conflict to attack one of the conflict participants, thus supporting the other) in a group of 13 captive ravens, Corvus corax. We found support for long-term, but not short-term, reciprocation of agonistic support. Ravens were more likely to support individuals who preened them, kin and dominant group members. These results suggest that ravens do not reciprocate on a calculated tit-for-tat basis, but aid individuals from whom reciprocated support would be most useful and those with whom they share a good relationship. Additionally, dyadic levels of agonistic support and consolation (postconflict affiliation from a bystander to the victim) correlated strongly with each other, but we found no evidence to suggest that receiving agonistic support influences the victim's likelihood of receiving support (consolation) after the conflict ends. Our findings are consistent with an emotionally mediated form of reciprocity in ravens and provide additional support for convergent cognitive evolution in birds and mammals.

  18. Reciprocity of agonistic support in ravens

    PubMed Central

    Fraser, Orlaith N.; Bugnyar, Thomas

    2012-01-01

    Cooperative behaviour through reciprocation or interchange of valuable services in primates has received considerable attention, especially regarding the timeframe of reciprocation and its ensuing cognitive implications. Much less, however, is known about reciprocity in other animals, particularly birds. We investigated patterns of agonistic support (defined as a third party intervening in an ongoing conflict to attack one of the conflict participants, thus supporting the other) in a group of 13 captive ravens, Corvus corax. We found support for long-term, but not short-term, reciprocation of agonistic support. Ravens were more likely to support individuals who preened them, kin and dominant group members. These results suggest that ravens do not reciprocate on a calculated tit-for-tat basis, but aid individuals from whom reciprocated support would be most useful and those with whom they share a good relationship. Additionally, dyadic levels of agonistic support and consolation (postconflict affiliation from a bystander to the victim) correlated strongly with each other, but we found no evidence to suggest that receiving agonistic support influences the victim’s likelihood of receiving support (consolation) after the conflict ends. Our findings are consistent with an emotionally mediated form of reciprocity in ravens and provide additional support for convergent cognitive evolution in birds and mammals. PMID:22298910

  19. Multiple tyrosine metabolites are GPR35 agonists

    PubMed Central

    Deng, Huayun; Hu, Haibei; Fang, Ye

    2012-01-01

    Both kynurenic acid and 2-acyl lysophosphatidic acid have been postulated to be the endogenous agonists of GPR35. However, controversy remains whether alternative endogenous agonists exist. The molecular targets accounted for many nongenomic actions of thyroid hormones are mostly unknown. Here we report the agonist activity of multiple tyrosine metabolites at the GPR35. Tyrosine metabolism intermediates that contain carboxylic acid and/or catechol functional groups were first selected. Whole cell dynamic mass redistribution (DMR) assays enabled by label-free optical biosensor were then used to characterize their agonist activity in native HT-29. Molecular assays including β-arrestin translocation, ERK phosphorylation and receptor internalization confirmed that GPR35 functions as a receptor for 5,6-dihydroxyindole-2-carboxylic acid, 3,3′,5′-triiodothyronine, 3,3′,5-triiodothyronine, gentisate, rosmarinate, and 3-nitrotyrosine. These results suggest that multiple tyrosine metabolites are alternative endogenous ligands of GPR35, and GPR35 may represent a druggable target for treating certain diseases associated with abnormality of tyrosine metabolism. PMID:22523636

  20. Effectiveness of the PPARγ agonist, GW570, in liver fibrosis.

    PubMed

    Yang, Liu; Stimpson, Stephen A; Chen, Lihong; Wallace Harrington, W; Rockey, Don C

    2010-12-01

    The peroxisome proliferator-activated receptors (PPARs) are well established to be important in modulating the fibrogenic response to liver injury. PPARγ plays a role in hepatic fibrosis, presumably by virtue of its expression in hepatic stellate cells, which are key effectors of fibrosis. In this study, we evaluated whether the potent nonthiozolidinedione PPARγ agonist, GW570, had effects on isolated stellate cells and hepatic fibrosis in vivo. Liver fibrosis and stellate cell activation were induced in vivo by either bile duct ligation (BDL) or administration of carbon tetrachloride (CCl(4)). Primary cultures of stellate cells isolated from normal rats were exposed to GW570. The PPARγ agonist was also given to male Sprague-Dawley rats before or during injury to test its ability to ameliorate fibrosis. Fibrosis biomarkers including total collagen, hydroxyproline, collagen I α1 and smooth muscle α actin were measured. GW570 had potent effects on isolated stellate cells, both simulating PPARγ mediated gene transcription, as well as inhibiting collagen I α1 mRNA and protein expression and smooth muscle α actin protein abundance, consistent with suppression of stellate cell activation. In BDL liver injury, a daily dose of 10 mg/kg per day of GW570 inhibited collagen I α1 mRNA, while concentrations of 1 also inhibited fibrosis as measured by hydroxyproline and total collagen content. Lower doses of GW570 (0.1-1.0 mg/kg per day) did not significantly abrogate whole liver collagen or hydroxyproline content in this model. In a CCl(4) model, 0.1-1.0 mg/kg per day GW570 reduced expression of smooth muscle α actin, but did not affect whole liver collagen or hydroxyproline content. Finally, we found that GW570 had anti-inflammatory effects on Kupffer cells as well as in vivo during CCl(4) injury. PPARγ receptor agonism with the nonthiozolidinedione, GW570, inhibited stellate cell activation in vitro and in vivo, and abrogated the fibrogenic response to injury in a

  1. Dopamine agonists, anti-progestins, anti-androgens, long-term-release GnRH agonists and anti-estrogens in canine reproduction: a review.

    PubMed

    Gobello, C

    2006-10-01

    Over the last 10 years, new drugs have been applied to canine reproduction, widening the spectrum of therapeutic possibilities for diseases that were previously surgically treated, and facilitating better control of the estrous cycle and fertility. Some are not approved for use in dogs; their use is experimental and further clinical trials are necessary. Dopamine agonists such as cabergoline, bromocriptine or metergoline are ergoderivative alkaloids that exert an anti-prolactinergic effect via stimulation of D2 pituitary receptors or inhibition of central serotoninergic ones. Their main indication is suppression of lactation. Anti-prolactinergic compounds have also been successfully used for pregnancy termination and shortening of interestrous intervals. Anti-progestins, (e.g. mifepristone and aglepristone) are synthetic steroids that bind with high affinity to progesterone (P4) receptors, preventing P4 from exerting its biological effects. Anti-progestins have been indicated in P4-dependent conditions, such as pregnancy termination, induction of parturition and the medical treatment of pyometra. Several groups of drugs have been described to have anti-androgenic properties through different mechanisms of action: progestins, receptor binding anti-androgens (e.g. flutamide), competitive enzyme inhibitors (e.g. finasteride), aromatase inhibitors, and GnRH agonists. Their main application is medical treatment of benign prostatic hyperplasia. Long-term release formulations of GnRH agonists (e.g. leuprolide or deslorelin acetate) postponed puberty and reversibly suppressed reproductive function in male and female dogs for periods exceeding 1 year. Anti-estrogens (e.g. clomiphene and tamoxifen citrate) are synthetic non-steroidal type I anti-estrogenic compounds that competitively block estrogen receptors with a combined antagonist-agonistic effect. In dogs, their action is more agonistic than antagonistic.

  2. Toll-like receptor 7 agonists are potent and rapid bronchodilators in guinea pigs

    PubMed Central

    Kaufman, Elad H.; Fryer, Allison D.; Jacoby, David B.

    2011-01-01

    Background Respiratory tract viral infections result in asthma exacerbations. Toll-like receptor (TLR) 7 is a receptor for viral single-stranded RNA and is expressed at high levels in the lungs. Objective Because TLR7 polymorphisms are associated with asthma, we examined the effects of TLR7 agonists in guinea pig airways. Methods We induced bronchoconstriction in guinea pigs in vivo by means of electrical stimulation of the vagus nerve or intravenous administration of acetylcholine and measured the effect of a TLR7 agonist administered intravenously. We induced contraction of airway smooth muscle in segments of isolated guinea pig tracheas in vitro and measured the effect of TLR7 agonists, antagonists, and pharmacologic inhibitors of associated signaling pathways administered directly to the bath. Results TLR7 agonists acutely inhibited bronchoconstriction in vivo and relaxed contraction of airway smooth muscle in vitro within minutes of administration. Airway relaxation induced by the TLR7 agonist R837 (imiquimod) was partially blocked with a TLR7 antagonist and was also blocked by inhibitors of large-conductance, calcium-activated potassium channels; prostaglandin synthesis; and nitric oxide generation. Another TLR7 agonist, 21-mer single-stranded phosphorothioated polyuridylic acid (PolyUs), mediated relaxation that was completely blocked by a TLR7 antagonist. Conclusions These data demonstrate a novel protective mechanism to limit bronchoconstriction and maintain airflow during respiratory tract viral infections. The fast time frame is inconsistent with canonical TLR7 signaling. R837 mediates bronchodilation by means of TLR7-dependent and TLR7-independent mechanisms, whereas PolyUs does so through only the TLR7-dependent mechanism. TLR7-independent mechanisms involve prostaglandins and large-conductance, calcium-activated potassium channels, whereas TLR7-dependent mechanisms involve nitric oxide. TLR7 is an attractive therapeutic target for its ability to

  3. Potentiation of cytotoxic chemotherapy by growth hormone-releasing hormone agonists

    PubMed Central

    Jaszberenyi, Miklos; Rick, Ferenc G.; Popovics, Petra; Block, Norman L.; Zarandi, Marta; Cai, Ren-Zhi; Vidaurre, Irving; Szalontay, Luca; Jayakumar, Arumugam R.; Schally, Andrew V.

    2014-01-01

    The dismal prognosis of malignant brain tumors drives the development of new treatment modalities. In view of the multiple activities of growth hormone-releasing hormone (GHRH), we hypothesized that pretreatment with a GHRH agonist, JI-34, might increase the susceptibility of U-87 MG glioblastoma multiforme (GBM) cells to subsequent treatment with the cytotoxic drug, doxorubicin (DOX). This concept was corroborated by our findings, in vivo, showing that the combination of the GHRH agonist, JI-34, and DOX inhibited the growth of GBM tumors, transplanted into nude mice, more than DOX alone. In vitro, the pretreatment of GBM cells with JI-34 potentiated inhibitory effects of DOX on cell proliferation, diminished cell size and viability, and promoted apoptotic processes, as shown by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide proliferation assay, ApoLive-Glo multiplex assay, and cell volumetric assay. Proteomic studies further revealed that the pretreatment with GHRH agonist evoked differentiation decreasing the expression of the neuroectodermal stem cell antigen, nestin, and up-regulating the glial maturation marker, GFAP. The GHRH agonist also reduced the release of humoral regulators of glial growth, such as FGF basic and TGFβ. Proteomic and gene-expression (RT-PCR) studies confirmed the strong proapoptotic activity (increase in p53, decrease in v-myc and Bcl-2) and anti-invasive potential (decrease in integrin α3) of the combination of GHRH agonist and DOX. These findings indicate that the GHRH agonists can potentiate the anticancer activity of the traditional chemotherapeutic drug, DOX, by multiple mechanisms including the induction of differentiation of cancer cells. PMID:24379381

  4. Recovery of brain biomarkers following peroxisome proliferator-activated receptor agonist neuroprotective treatment before ischemic stroke

    PubMed Central

    2014-01-01

    Background Lipid lowering agent such as agonists of peroxisome proliferator-activated receptors (PPAR) are suggested as neuroprotective agents and may protect from the sequelae of brain ischemic stroke. Although the demonstration is not clearly established in human, the underlying molecular mechanism may be of interest for future therapeutic purposes. To this end, we have used our well established rodent model of ischemia-reperfusion pre-treated or not with fenofibrate or atorvastatin and performed a differential proteomics analyses of the brain and analysed the protein markers which levels returned to “normal” following pre-treatments with PPARα agonists. Results In order to identify potential therapeutic targets positively modulated by pre-treatment with the PPARα agonists, two-dimensional gel electrophoresis proteome profiles between control, ischemia-reperfusion and pre-treated or not, were compared. The polypeptide which expression was altered following ischemia – reperfusion but whose levels remain unchanged after pre-treatment were characterized by mass spectrometry and further investigated by Western-blotting and immunohistochemistry. A series of 28 polypeptides were characterized among which the protein disulfide isomerase reduction – a protein instrumental to the unfolded protein response system - was shown to be reduced following PPARα agonists treatment while it was strongly increased in ischemia-reperfusion. Conclusions Pre-treatment with PPARα agonist or atorvastatin show potential neuroprotective effects by inhibiting the PDI overexpression in conjunction with the preservation of other neuronal markers, several of which are associated with the regulation of protein homeostasis, signal transduction and maintenance of synaptic plasticity. This proteomic study therefore suggests that neuroprotective effect of PPARα agonists supposes the preservation of the expression of several proteins essential for the maintenance of protein homeostasis

  5. Studies on the pharmacology of the novel histamine H3 receptor agonist Sch 50971.

    PubMed

    Hey, J A; Aslanian, R; Bolser, D C; Chapman, R W; Egan, R W; Rizzo, C A; Shih, N Y; Fernandez, X; McLeod, R L; West, R; Kreutner, W

    1998-09-01

    Experiments were performed to characterize the pharmacology of Sch 50971 ((+)-trans-4-(4(R)-methyl-3(R)-pyrolidinyl)-1H-imidazole dihydrochloride, CAS 167610-28-8), a novel histamine H3 receptor agonist. The activity of Sch 50971 was compared with that of (R)-alpha-methylhistamine (CAS 75614-87-8), a potent and moderately selective agonist of histamine H3 receptors, in a series of in vitro and in vivo assays. Sch 50971 is a high affinity, selective H3 receptor agonist in vitro and in vivo. Sch 50971 inhibits [3H]-N-alpha-methylhistamine (CAS 673-50-7) binding to the histamine H3 receptor in human brain (Ki = 5.0 nmol/l) and guinea pig brain (Ki = 2.5 nmol/l). Sch 50971 also inhibits electric field stimulated guinea pig ileum contractions (pD2 = 7.47) and decreases [3H]-norepinephrine (CAS 51-41-2) release (pD2 = 7.48) from guinea pig pulmonary artery by activation of presynaptic inhibitory H3 receptors. The in vitro effects of Sch 50971 are antagonized by low concentrations of a selective H3 antagonist, thioperamide (CAS 106243-16-7). Sch 50971 has low affinity (IC50's > 10 mumol/l) for histamine H1, dopamine D1 and D2, serotonin 5-HT2 and muscarinic cholinergic receptors. It also does not exhibit histamine H2-antagonist activity. In guinea pigs and cats, Sch 50971 exhibits in vivo H3 agonist activity. Sch 50971 inhibits sympathetic hypertension evoked by stimulation of the medulla oblongata in anesthetized guinea pigs (ED30 = 0.3 mg/kg i.v., ED30 = 1.0 mg/kg i.d.). Sch 50971 also inhibits the effects of sympathetic nerve stimulation on nasal resistance in cats. In these assays, Sch 50971 exhibits an efficacy and potency comparable to H3-agonist (R)-alpha-methylhistamine. However, under in vivo conditions, Sch 50971 does not exhibit histamine H1-mediated responses that are seen with (R)-alpha-methylhistamine at doses close to those that produce H3 effects. Therefore, Sch 50971 is a novel, potent and selective agonist of histamine H3 receptors with an improved in

  6. FXR agonist activity of conformationally constrained analogs of GW 4064

    SciTech Connect

    Akwabi-Ameyaw, Adwoa; Bass, Jonathan Y.; Caldwell, Richard D.; Caravella, Justin A.; Chen, Lihong; Creech, Katrina L.; Deaton, David N.; Madauss, Kevin P.; Marr, Harry B.; McFadyen, Robert B.; Miller, Aaron B.; Navas, III, Frank; Parks, Derek J.; Spearing, Paul K.; Todd, Dan; Williams, Shawn P.; Wisely, G. Bruce

    2010-09-27

    Two series of conformationally constrained analogs of the FXR agonist GW 4064 1 were prepared. Replacement of the metabolically labile stilbene with either benzothiophene or naphthalene rings led to the identification of potent full agonists 2a and 2g.

  7. Bradykinin type 2 receptor -9/-9 genotype is associated with triceps brachii muscle hypertrophy following strength training in young healthy men

    PubMed Central

    2012-01-01

    Background Bradykinin type 2 receptor (B2BRK) genotype was reported to be associated with changes in the left-ventricular mass as a response to aerobic training, as well as in the regulation of the skeletal muscle performance in both athletes and non-athletes. However, there are no reports on the effect of B2BRK 9-bp polymorphism on the response of the skeletal muscle to strength training, and our aim was to determine the relationship between the B2BRK SNP and triceps brachii functional and morphological adaptation to programmed physical activity in young adults. Methods In this 6-week pretest-posttest exercise intervention study, twenty nine healthy young men (21.5 ± 2.7 y, BMI 24.2 ± 3.5 kg/m2) were put on a 6-week exercise protocol using an isoacceleration dynamometer (5 times a week, 5 daily sets with 10 maximal elbow extensions, 1 minute rest between sets). Triceps brachii muscle volumes were assessed by using magnetic resonance imaging before and after the strength training. Bradykinin type 2 receptor 9 base pair polymorphism was determined for all participants. Results Following the elbow extensors training, an average increase in the volume of both triceps brachii was 5.4 ± 3.4% (from 929.5 ± 146.8 cm3 pre-training to 977.6 ± 140.9 cm3 after training, p<0.001). Triceps brachii volume increase was significantly larger in individuals homozygous for −9 allele compared to individuals with one or two +9 alleles (−9/-9, 8.5 ± 3.8%; vs. -9/+9 and +9/+9 combined, 4.7 ± 4.5%, p < 0.05). Mean increases in endurance strength in response to training were 48.4 ± 20.2%, but the increases were not dependent on B2BRK genotype (−9/-9, 50.2 ± 19.2%; vs. -9/+9 and +9/+9 combined, 46.8 ± 20.7%, p > 0.05). Conclusions We found that muscle morphological response to targeted training – hypertrophy – is related to polymorphisms of B2BRK. However, no significant influence of different B2BRK genotypes on functional muscle properties after strength training in

  8. Bradykinin type 2 receptor -9/-9 genotype is associated with triceps brachii muscle hypertrophy following strength training in young healthy men.

    PubMed

    Popadic Gacesa, Jelena Z; Momcilovic, Milica; Veselinovic, Igor; Brodie, David A; Grujic, Nikola G

    2012-11-06

    Bradykinin type 2 receptor (B2BRK) genotype was reported to be associated with changes in the left-ventricular mass as a response to aerobic training, as well as in the regulation of the skeletal muscle performance in both athletes and non-athletes. However, there are no reports on the effect of B2BRK 9-bp polymorphism on the response of the skeletal muscle to strength training, and our aim was to determine the relationship between the B2BRK SNP and triceps brachii functional and morphological adaptation to programmed physical activity in young adults. In this 6-week pretest-posttest exercise intervention study, twenty nine healthy young men (21.5 ± 2.7 y, BMI 24.2 ± 3.5 kg/m(2)) were put on a 6-week exercise protocol using an isoacceleration dynamometer (5 times a week, 5 daily sets with 10 maximal elbow extensions, 1 minute rest between sets). Triceps brachii muscle volumes were assessed by using magnetic resonance imaging before and after the strength training. Bradykinin type 2 receptor 9 base pair polymorphism was determined for all participants. Following the elbow extensors training, an average increase in the volume of both triceps brachii was 5.4 ± 3.4% (from 929.5 ± 146.8 cm(3) pre-training to 977.6 ± 140.9 cm(3) after training, p<0.001). Triceps brachii volume increase was significantly larger in individuals homozygous for -9 allele compared to individuals with one or two +9 alleles (-9/-9, 8.5 ± 3.8%; vs. -9/+9 and +9/+9 combined, 4.7 ± 4.5%, p < 0.05). Mean increases in endurance strength in response to training were 48.4 ± 20.2%, but the increases were not dependent on B2BRK genotype (-9/-9, 50.2 ± 19.2%; vs. -9/+9 and +9/+9 combined, 46.8 ± 20.7%, p > 0.05). We found that muscle morphological response to targeted training - hypertrophy - is related to polymorphisms of B2BRK. However, no significant influence of different B2BRK genotypes on functional muscle properties after strength training in young healthy non athletes was found. This

  9. Heterologous Expression in Remodeled C. elegans: A Platform for Monoaminergic Agonist Identification and Anthelmintic Screening

    PubMed Central

    Law, Wenjing; Wuescher, Leah M.; Ortega, Amanda; Hapiak, Vera M.; Komuniecki, Patricia R.; Komuniecki, Richard

    2015-01-01

    Monoamines, such as 5-HT and tyramine (TA), paralyze both free-living and parasitic nematodes when applied exogenously and serotonergic agonists have been used to clear Haemonchus contortus infections in vivo. Since nematode cell lines are not available and animal screening options are limited, we have developed a screening platform to identify monoamine receptor agonists. Key receptors were expressed heterologously in chimeric, genetically-engineered Caenorhabditis elegans, at sites likely to yield robust phenotypes upon agonist stimulation. This approach potentially preserves the unique pharmacologies of the receptors, while including nematode-specific accessory proteins and the nematode cuticle. Importantly, the sensitivity of monoamine-dependent paralysis could be increased dramatically by hypotonic incubation or the use of bus mutants with increased cuticular permeabilities. We have demonstrated that the monoamine-dependent inhibition of key interneurons, cholinergic motor neurons or body wall muscle inhibited locomotion and caused paralysis. Specifically, 5-HT paralyzed C. elegans 5-HT receptor null animals expressing either nematode, insect or human orthologues of a key Gαo-coupled 5-HT1-like receptor in the cholinergic motor neurons. Importantly, 8-OH-DPAT and PAPP, 5-HT receptor agonists, differentially paralyzed the transgenic animals, with 8-OH-DPAT paralyzing mutant animals expressing the human receptor at concentrations well below those affecting its C. elegans or insect orthologues. Similarly, 5-HT and TA paralyzed C. elegans 5-HT or TA receptor null animals, respectively, expressing either C. elegans or H. contortus 5-HT or TA-gated Cl- channels in either C. elegans cholinergic motor neurons or body wall muscles. Together, these data suggest that this heterologous, ectopic expression screening approach will be useful for the identification of agonists for key monoamine receptors from parasites and could have broad application for the identification

  10. Heterologous Expression in Remodeled C. elegans: A Platform for Monoaminergic Agonist Identification and Anthelmintic Screening.

    PubMed

    Law, Wenjing; Wuescher, Leah M; Ortega, Amanda; Hapiak, Vera M; Komuniecki, Patricia R; Komuniecki, Richard

    2015-04-01

    Monoamines, such as 5-HT and tyramine (TA), paralyze both free-living and parasitic nematodes when applied exogenously and serotonergic agonists have been used to clear Haemonchus contortus infections in vivo. Since nematode cell lines are not available and animal screening options are limited, we have developed a screening platform to identify monoamine receptor agonists. Key receptors were expressed heterologously in chimeric, genetically-engineered Caenorhabditis elegans, at sites likely to yield robust phenotypes upon agonist stimulation. This approach potentially preserves the unique pharmacologies of the receptors, while including nematode-specific accessory proteins and the nematode cuticle. Importantly, the sensitivity of monoamine-dependent paralysis could be increased dramatically by hypotonic incubation or the use of bus mutants with increased cuticular permeabilities. We have demonstrated that the monoamine-dependent inhibition of key interneurons, cholinergic motor neurons or body wall muscle inhibited locomotion and caused paralysis. Specifically, 5-HT paralyzed C. elegans 5-HT receptor null animals expressing either nematode, insect or human orthologues of a key Gαo-coupled 5-HT1-like receptor in the cholinergic motor neurons. Importantly, 8-OH-DPAT and PAPP, 5-HT receptor agonists, differentially paralyzed the transgenic animals, with 8-OH-DPAT paralyzing mutant animals expressing the human receptor at concentrations well below those affecting its C. elegans or insect orthologues. Similarly, 5-HT and TA paralyzed C. elegans 5-HT or TA receptor null animals, respectively, expressing either C. elegans or H. contortus 5-HT or TA-gated Cl- channels in either C. elegans cholinergic motor neurons or body wall muscles. Together, these data suggest that this heterologous, ectopic expression screening approach will be useful for the identification of agonists for key monoamine receptors from parasites and could have broad application for the identification

  11. Antidiabetic properties of the histamine H3 receptor protean agonist proxyfan.

    PubMed

    Henry, Melanie B; Zheng, Shuqin; Duan, Chenxia; Patel, Bhuneshwari; Vassileva, Galya; Sondey, Christopher; Lachowicz, Jean; Hwa, Joyce J

    2011-03-01

    Proxyfan is a histamine H3 receptor protean agonist that can produce a spectrum of pharmacological effects including agonist, inverse agonist, and antagonist. We have discovered that proxyfan (10 mg/kg orally) significantly improved glucose excursion after an ip glucose tolerance test in either lean or high-fat/cholesterol diet-induced obese mice. It also reduced plasma glucose levels comparable to that of metformin (300 mg/kg orally) in a nongenetic type 2 diabetes mouse model. The dose-dependent decrease in glucose excursion correlated with inhibition of ex vivo H3 receptor binding in the cerebral cortex. In addition, glucose levels were significantly reduced compared with vehicle-treated mice after intracerebroventricular administration of proxyfan, suggesting the involvement of central H3 receptors. Proxyfan-induced reduction of glucose excursion was not observed in the H3 receptor knockout mice, suggesting that proxyfan mediates this effect through H3 receptors. Proxyfan reduced glucose excursion by significantly increasing plasma insulin levels in a glucose-independent manner. However, no difference in insulin sensitivity was observed in proxyfan-treated mice. The H1 receptor antagonist chlorpheniramine and the H2 receptor antagonist zolantidine had modest effects on glucose excursion, and neither inhibited the glucose excursion reduced by proxyfan. The H3 receptor antagonist/inverse agonist, thioperamide, had weaker effects on glucose excursion compared with proxyfan, whereas the H3 receptor agonist imetit did not affect glucose excursion. In conclusion, these findings demonstrate, for the first time, that manipulation of central histamine H3 receptor by proxyfan can significantly improve glucose excursion by increasing plasma insulin levels via a glucose-independent mechanism.

  12. Distinct activities of GABA agonists at synaptic- and extrasynaptic-type GABAA receptors

    PubMed Central

    Mortensen, Martin; Ebert, Bjarke; Wafford, Keith; Smart, Trevor G

    2010-01-01

    The activation characteristics of synaptic and extrasynaptic GABAA receptors are important for shaping the profile of phasic and tonic inhibition in the central nervous system, which will critically impact on the activity of neuronal networks. Here, we study in isolation the activity of three agonists, GABA, muscimol and 4,5,6,7-tetrahydoisoxazolo[5,4-c]pyridin-3(2H)-one (THIP), to further understand the activation profiles of α1β3γ2, α4β3γ2 and α4β3δ receptors that typify synaptic- and extrasynaptic-type receptors expressed in the hippocampus and thalamus. The agonists display an order of potency that is invariant between the three receptors, which is reliant mostly on the agonist dissociation constant. At δ subunit-containing extrasynaptic-type GABAA receptors, both THIP and muscimol additionally exhibited, to different degrees, superagonist behaviour. By comparing whole-cell and single channel currents induced by the agonists, we provide a molecular explanation for their different activation profiles. For THIP at high concentrations, the unusual superagonist behaviour on α4β3δ receptors is a consequence of its ability to increase the duration of longer channel openings and their frequency, resulting in longer burst durations. By contrast, for muscimol, moderate superagonist behaviour was caused by reduced desensitisation of the extrasynaptic-type receptors. The ability to specifically increase the efficacy of receptor activation, by selected exogenous agonists over that obtained with the natural transmitter, may prove to be of therapeutic benefit under circumstances when synaptic inhibition is compromised or dysfunctional. PMID:20176630

  13. Differentiation of δ, μ, and κ opioid receptor agonists based on pharmacophore development and computed physicochemical properties

    NASA Astrophysics Data System (ADS)

    Filizola, Marta; Villar, Hugo O.; Loew, Gilda H.

    2001-04-01

    Compounds that bind with significant affinity to the opioid receptor types, δ, μ, and κ, with different combinations of activation and inhibition at these three receptors could be promising behaviorally selective agents. Working on this hypothesis, the chemical moieties common to three different sets of opioid receptor agonists with significant affinity for each of the three receptor types δ, μ, or κ were identified. Using a distance analysis approach, common geometric arrangements of these chemical moieties were found for selected δ, μ, or κ opioid agonists. The chemical and geometric commonalities among agonists at each opioid receptor type were then compared with a non-specific opioid recognition pharmacophore recently developed. The comparison provided identification of the additional requirements for activation of δ, μ, and κ opioid receptors. The distance analysis approach was able to clearly discriminate κ-agonists, while global molecular properties for all compounds were calculated to identify additional requirements for activation of δ and μ receptors. Comparisons of the combined geometric and physicochemical properties calculated for each of the three sets of agonists allowed the determination of unique requirements for activation of each of the three opioid receptors. These results can be used to improve the activation selectivity of known opioid agonists and as a guide for the identification of novel selective opioid ligands with potential therapeutic usefulness.

  14. Structural insights for the design of new PPARgamma partial agonists with high binding affinity and low transactivation activity

    NASA Astrophysics Data System (ADS)

    Guasch, Laura; Sala, Esther; Valls, Cristina; Blay, Mayte; Mulero, Miquel; Arola, Lluís; Pujadas, Gerard; Garcia-Vallvé, Santiago

    2011-08-01

    Peroxisome Proliferator-Activated Receptor γ (PPARγ) full agonists are molecules with powerful insulin-sensitizing action that are used as antidiabetic drugs. Unfortunately, these compounds also present various side effects. Recent results suggest that effective PPARγ agonists should show a low transactivation activity but a high binding affinity to inhibit phosphorylation at Ser273. We use several structure activity relationship studies of synthetic PPARγ agonists to explore the different binding features of full and partial PPARγ agonists with the aim of differentiating the features needed for binding and those needed for the transactivation activity of PPARγ. Our results suggest that effective partial agonists should have a hydrophobic moiety and an acceptor site with an appropriate conformation to interact with arm II and establish a hydrogen bond with Ser342 or an equivalent residue at arm III. Despite the fact that interactions with arm I increase the binding affinity, this region should be avoided in order to not increase the transactivation activity of potential PPARγ partial agonists.