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Kirk M Druey - One of the best experts on this subject based on the ideXlab platform.
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R4 Regulator of G Protein Signaling (RGS) Proteins in Inflammation and Immunity
The AAPS Journal, 2016Co-Authors: Eunice C. Chan, Kirk M DrueyAbstract:G protein-coupled receptors (GPCRs) have important functions in both innate and adaptive immunity, with the capacity to bridge interactions between the two arms of the host responses to pathogens through direct recognition of secreted microbial products or the by-products of host cells damaged by pathogen exposure. In the mid-1990s, a large group of intracellular proteins was discovered, the regulator of G protein signaling (RGS) family, whose main, but not exclusive, function appears to be to constrain the intensity and duration of GPCR signaling. The R4/B subfamily—the focus of this review—includes RGS1–5, 8, 13, 16, 18, and 21, which are the smallest RGS proteins in size, with the exception of RGS3. Prominent roles in the trafficking of B and T lymphocytes and macrophages have been described for RGS1, RGS13, and RGS16, while RGS18 appears to control platelet and osteoclast functions. Additional G protein independent functions of RGS13 have been uncovered in gene expression in B lymphocytes and mast cell-mediated allergic reactions. In this review, we discuss potential physiological roles of this RGS protein subfamily, primarily in leukocytes having central roles in immune and inflammatory responses. We also discuss approaches to target RGS proteins therapeutically, which represents a virtually untapped strategy to combat exaggerated immune responses leading to inflammation.
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CXCL12 regulates platelet activation via the regulator of G-protein signaling 16.
Biochimica et biophysica acta, 2015Co-Authors: Zubair A. Karim, Kirk M Druey, Fatima Z. Alshbool, Hari Priya Vemana, Christine Conlon, Fadi T. KhasawnehAbstract:The regulators of G protein signaling (RGS) protein superfamily negatively controls G protein-coupled receptor signal transduction pathways. One of the members of this family, RGS16, is highly expressed in megakaryocytes and platelets. Studies of its function in platelet and megakaryocyte biology have been limited, in part, due to lack of pharmacological inhibitors. For example, RGS16 overexpression inhibited CXC chemokine receptor 4 (CXCR4)-mediated megakaryocyte migration. More recent studies showed that the chemokine stromal cell-derived factor (SDF1α or CXCL12) regulates platelet function via CXCR4. Based on these considerations, the present study investigated the capacity of RGS16 to regulate CXCL12-dependent platelet function, using the RGS16 knockout mouse model (RGS16(-/-)). RGS16-deficient platelets had increased protease activated receptor 4 and collagen-induced aggregation, as well as increased CXCL12-dependent agonist-induced aggregation, dense and alpha granule secretion, integrin αIIbβ3 activation and phosphatidylserine exposure compared to those from WT littermates. CXCL12 alone did not stimulate aggregation or secretion in either RGS16-deficient or WT platelets. Furthermore, platelets from RGS16(-/-) mice displayed enhanced phosphorylation of ERK and Akt following CXCL12 stimulation relative to controls. Finally, we also found that PKCδ is involved in regulating CXCL12-dependent activation of ERK and Akt, in the RGS16-deficient platelets. Collectively, our findings provide the first evidence that RGS16 plays an important role in platelet function by modulating CXCL12-dependent platelet activation.
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RGS16 attenuates pulmonary th2 th17 inflammatory responses
Journal of Immunology, 2012Co-Authors: Sucharita Shankar, Mark S. Wilson, Jeffrey A. Divietro, Zhihui Xie, Thomas A. Wynn, Margaret M Mentinkkane, Kirk M DrueyAbstract:The regulators of G protein signaling (RGS) protein superfamily negatively controls G protein-coupled receptor signal transduction pathways. RGS16 is enriched in activated/effector T lymphocytes. In this paper, we show that RGS16 constrains pulmonary inflammation by regulating chemokine-induced T cell trafficking in response to challenge with Schistosoma mansoni. Naive RGS16−/− mice were “primed” for inflammation by accumulation of CCR10+ T cells in the lung. Upon pathogen exposure, these mice developed more robust granulomatous lung fibrosis than wild-type counterparts. Distinct Th2 or putative Th17 subsets expressing CCR4 or CCR10 accumulated more rapidly in RGS16−/− lungs following challenge and produced proinflammatory cytokines IL-13 and IL-17B. CCR4+RGS16−/− Th2 cells migrated excessively to CCL17 and localized aberrantly in challenged lungs. T lymphocytes were partially excluded from lung granulomas in RGS16−/− mice, instead forming peribronchial/perivascular aggregates. Thus, RGS16-mediated confinement of T cells to Schistosome granulomas mitigates widespread cytokine-mediated pulmonary inflammation.
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RGS16 Attenuates Pulmonary Th2/Th17 Inflammatory Responses
Journal of immunology (Baltimore Md. : 1950), 2012Co-Authors: Sucharita P. Shankar, Mark S. Wilson, Jeffrey A. Divietro, Margaret M. Mentink-kane, Zhihui Xie, Thomas A. Wynn, Kirk M DrueyAbstract:The regulators of G protein signaling (RGS) protein superfamily negatively controls G protein-coupled receptor signal transduction pathways. RGS16 is enriched in activated/effector T lymphocytes. In this paper, we show that RGS16 constrains pulmonary inflammation by regulating chemokine-induced T cell trafficking in response to challenge with Schistosoma mansoni. Naive RGS16−/− mice were “primed” for inflammation by accumulation of CCR10+ T cells in the lung. Upon pathogen exposure, these mice developed more robust granulomatous lung fibrosis than wild-type counterparts. Distinct Th2 or putative Th17 subsets expressing CCR4 or CCR10 accumulated more rapidly in RGS16−/− lungs following challenge and produced proinflammatory cytokines IL-13 and IL-17B. CCR4+RGS16−/− Th2 cells migrated excessively to CCL17 and localized aberrantly in challenged lungs. T lymphocytes were partially excluded from lung granulomas in RGS16−/− mice, instead forming peribronchial/perivascular aggregates. Thus, RGS16-mediated confinement of T cells to Schistosome granulomas mitigates widespread cytokine-mediated pulmonary inflammation.
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RGS16 inhibits breast cancer cell growth by mitigating phosphatidylinositol 3-kinase signaling.
The Journal of biological chemistry, 2009Co-Authors: Genqing Liang, Zhihui Xie, Geetanjali Bansal, Kirk M DrueyAbstract:Aberrant activity of the phosphatidylinositol 3-kinase (PI3K) pathway supports growth of many tumors including those of breast, lung, and prostate. Resistance of breast cancer cells to targeted chemotherapies including tyrosine kinase inhibitors (TKI) has been linked to persistent PI3K activity, which may in part be due to increased membrane expression of epidermal growth factor (EGF) receptors (HER2 and HER3). Recently we found that proteins of the RGS (regulator of G protein signaling) family suppress PI3K activity downstream of the receptor by sequestering its p85α subunit from signaling complexes. Because a substantial percentage of breast tumors have RGS16 mutations and reduced RGS16 protein expression, we investigated the link between regulation of PI3K activity by RGS16 and breast cancer cell growth. RGS16 overexpression in MCF7 breast cancer cells inhibited EGF-induced proliferation and Akt phosphorylation, whereas shRNA-mediated extinction of RGS16 augmented cell growth and resistance to TKI treatment. Exposure to TKI also reduced RGS16 expression in MCF7 and BT474 cell lines. RGS16 bound the amino-terminal SH2 and inter-SH2 domains of p85α and inhibited its interaction with the EGF receptor-associated adapter protein Gab1. These results suggest that the loss of RGS16 in some breast tumors enhances PI3K signaling elicited by growth factors and thereby promotes proliferation and TKI evasion downstream of HER activation.
Astrid E. Alewijnse - One of the best experts on this subject based on the ideXlab platform.
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Sphingosine-1-phosphate regulates RGS2 and RGS16 mRNA expression in vascular smooth muscle cells
European Journal of Pharmacology, 2009Co-Authors: Mariëlle C. Hendriks-balk, Najat Hajji, Pieter B. Van Loenen, Martin C. Michel, Stephan L. M. Peters, Astrid E. AlewijnseAbstract:Abstract Regulator of G protein signalling (RGS) protein expression is altered under growth promoting conditions in vascular smooth muscle cells (VSMCs). Since sphingosine-1-phosphate (S1P) is an important growth stimulatory factor, we investigated whether stimulation of VSMCs with S1P results in alterations in mRNA expression levels of several RGS proteins and which signalling components are involved. VSMCs were stimulated with S1P and mRNA expression levels of RGS2, RGS3, RGS4, RGS5 and RGS16 were measured by real-time polymerase chain reaction. S1P caused a time-dependent up-regulation of RGS2 and RGS16 mRNA expression. FTY720-P, a S1P1/S1P3–5 agonist, did not regulate RGS2 mRNA levels although it did up-regulate RGS16 mRNA expression. Pertussis toxin treatment revealed that the S1P-induced RGS16 expression was Gi/o-dependent whereas up-regulation of RGS2 mRNA was not. Phosphatidylinositol 3-kinase, protein kinase C and mitogen-activated protein kinase kinase apparently were not involved in the S1P-induced up-regulation of both RGS proteins. The present study demonstrates that S1P induces RGS2 and RGS16 mRNA expression but uses distinct S1P receptor subtypes and signalling pathways to regulate expression of these RGS proteins.
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LPS differentially affects vasoconstrictor responses: a potential role for RGS16?
Journal of physiology and biochemistry, 2009Co-Authors: Mariëlle C. Hendriks-balk, Najat Hajji, Astrid E. Alewijnse, M. Tjon-atsoi, Stephan L. M. PetersAbstract:The profound hypotension in septic shock patients is difficult to treat as it is accompanied by depressed constrictor responses to α1-adrenoceptor agonists. Bacterial lipopolysaccharide (LPS) is the main trigger for most of the cardiovascular alterations occurring in septic shock. In this study we investigated the effects of LPS exposure on vascular contractility in general and the role of Regulator of G protein Signalling (RGS) proteins in the LPS-induced vascular alterations. Exposure of rat aortic rings to various LPS concentrations (3, 10, 30 μg/ml) for 22 hours differentially affected agonist-induced contractile responses at four distinct G-protein coupled receptors (α1-adrenoceptors, angiotensin II, serotonin and endothelin-1 receptors). While the endothelin-1-induced contraction was unaffected by LPS pre-treatment, phenylephrine- and angiotensin II-induced contraction were significantly reduced whereas serotonin-induced contraction was significantly enhanced. Concomitantly, LPS treatment increased the RGS16 mRNA expression both in aortic rings and cultured vascular smooth muscle cells (VSMCs) but not that of RGS2, RGS3, RGS4 or RGS5. The significant increase in RGS16 mRNA expression in VSMCs by LPS was time- and concentration-dependent but independent of increased inducible NO synthase (iNOS) activity. The changes in RGS16 mRNA might contribute to the differential regulation of the contractile responses to vasoconstrictors upon LPS exposure.
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S1P receptor signalling and RGS proteins; expression and function in vascular smooth muscle cells and transfected CHO cells.
European Journal of Pharmacology, 2008Co-Authors: Mariëlle C. Hendriks-balk, Najat Hajji, Pieter B. Van Loenen, Martin C. Michel, Stephan L. M. Peters, Astrid E. AlewijnseAbstract:Abstract Sphingosine-1-phosphate (S1P) signalling via G protein-coupled receptors is important for the regulation of cell function and differentiation. Specific Regulators of G protein Signalling (RGS) proteins modulate the function of these receptors in many cell types including vascular smooth muscle cells (VSMCs). Therefore, we investigated the role of altered expression levels of RGS proteins in S1P receptor function in VSMCs and transfected CHO cells. The mRNA expression of the S1P 1 receptor, RGS4 and RGS16 were down-regulated in VSMCs during phenotypic modulation induced by culturing, whereas mRNA levels of RGS2, RGS3, S1P 2 and S1P 3 receptors were unchanged. Interestingly, the expression level of RGS5 was transiently up-regulated. Despite major alterations in RGS levels, S1P-induced calcium elevation in VSMCs was not altered. Co-transfection of RGS2, RGS3, RGS4, RGS5 and RGS16 into CHO-Flp-In cells stably expressing the S1P 1 or S1P 3 receptor did not modify S1P-induced inhibition of cAMP accumulation to a major extent. Similar results were obtained with SEW2871, a selective S1P 1 receptor agonist. However, the inhibition of cAMP accumulation by the agonist FTY720-P via the S1P 1 receptor was significantly decreased by co-transfection with RGS5. These results indicate that mRNA of the S1P 1 receptor, RGS4, RGS5 and RGS16 is differentially regulated during phenotypic modulation. However, major alterations in RGS protein expression have only limited effect on S1P receptor function.
Mariëlle C. Hendriks-balk - One of the best experts on this subject based on the ideXlab platform.
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Sphingosine-1-phosphate regulates RGS2 and RGS16 mRNA expression in vascular smooth muscle cells
European Journal of Pharmacology, 2009Co-Authors: Mariëlle C. Hendriks-balk, Najat Hajji, Pieter B. Van Loenen, Martin C. Michel, Stephan L. M. Peters, Astrid E. AlewijnseAbstract:Abstract Regulator of G protein signalling (RGS) protein expression is altered under growth promoting conditions in vascular smooth muscle cells (VSMCs). Since sphingosine-1-phosphate (S1P) is an important growth stimulatory factor, we investigated whether stimulation of VSMCs with S1P results in alterations in mRNA expression levels of several RGS proteins and which signalling components are involved. VSMCs were stimulated with S1P and mRNA expression levels of RGS2, RGS3, RGS4, RGS5 and RGS16 were measured by real-time polymerase chain reaction. S1P caused a time-dependent up-regulation of RGS2 and RGS16 mRNA expression. FTY720-P, a S1P1/S1P3–5 agonist, did not regulate RGS2 mRNA levels although it did up-regulate RGS16 mRNA expression. Pertussis toxin treatment revealed that the S1P-induced RGS16 expression was Gi/o-dependent whereas up-regulation of RGS2 mRNA was not. Phosphatidylinositol 3-kinase, protein kinase C and mitogen-activated protein kinase kinase apparently were not involved in the S1P-induced up-regulation of both RGS proteins. The present study demonstrates that S1P induces RGS2 and RGS16 mRNA expression but uses distinct S1P receptor subtypes and signalling pathways to regulate expression of these RGS proteins.
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LPS differentially affects vasoconstrictor responses: a potential role for RGS16?
Journal of physiology and biochemistry, 2009Co-Authors: Mariëlle C. Hendriks-balk, Najat Hajji, Astrid E. Alewijnse, M. Tjon-atsoi, Stephan L. M. PetersAbstract:The profound hypotension in septic shock patients is difficult to treat as it is accompanied by depressed constrictor responses to α1-adrenoceptor agonists. Bacterial lipopolysaccharide (LPS) is the main trigger for most of the cardiovascular alterations occurring in septic shock. In this study we investigated the effects of LPS exposure on vascular contractility in general and the role of Regulator of G protein Signalling (RGS) proteins in the LPS-induced vascular alterations. Exposure of rat aortic rings to various LPS concentrations (3, 10, 30 μg/ml) for 22 hours differentially affected agonist-induced contractile responses at four distinct G-protein coupled receptors (α1-adrenoceptors, angiotensin II, serotonin and endothelin-1 receptors). While the endothelin-1-induced contraction was unaffected by LPS pre-treatment, phenylephrine- and angiotensin II-induced contraction were significantly reduced whereas serotonin-induced contraction was significantly enhanced. Concomitantly, LPS treatment increased the RGS16 mRNA expression both in aortic rings and cultured vascular smooth muscle cells (VSMCs) but not that of RGS2, RGS3, RGS4 or RGS5. The significant increase in RGS16 mRNA expression in VSMCs by LPS was time- and concentration-dependent but independent of increased inducible NO synthase (iNOS) activity. The changes in RGS16 mRNA might contribute to the differential regulation of the contractile responses to vasoconstrictors upon LPS exposure.
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S1P receptor signalling and RGS proteins; expression and function in vascular smooth muscle cells and transfected CHO cells.
European Journal of Pharmacology, 2008Co-Authors: Mariëlle C. Hendriks-balk, Najat Hajji, Pieter B. Van Loenen, Martin C. Michel, Stephan L. M. Peters, Astrid E. AlewijnseAbstract:Abstract Sphingosine-1-phosphate (S1P) signalling via G protein-coupled receptors is important for the regulation of cell function and differentiation. Specific Regulators of G protein Signalling (RGS) proteins modulate the function of these receptors in many cell types including vascular smooth muscle cells (VSMCs). Therefore, we investigated the role of altered expression levels of RGS proteins in S1P receptor function in VSMCs and transfected CHO cells. The mRNA expression of the S1P 1 receptor, RGS4 and RGS16 were down-regulated in VSMCs during phenotypic modulation induced by culturing, whereas mRNA levels of RGS2, RGS3, S1P 2 and S1P 3 receptors were unchanged. Interestingly, the expression level of RGS5 was transiently up-regulated. Despite major alterations in RGS levels, S1P-induced calcium elevation in VSMCs was not altered. Co-transfection of RGS2, RGS3, RGS4, RGS5 and RGS16 into CHO-Flp-In cells stably expressing the S1P 1 or S1P 3 receptor did not modify S1P-induced inhibition of cAMP accumulation to a major extent. Similar results were obtained with SEW2871, a selective S1P 1 receptor agonist. However, the inhibition of cAMP accumulation by the agonist FTY720-P via the S1P 1 receptor was significantly decreased by co-transfection with RGS5. These results indicate that mRNA of the S1P 1 receptor, RGS4, RGS5 and RGS16 is differentially regulated during phenotypic modulation. However, major alterations in RGS protein expression have only limited effect on S1P receptor function.
Li Wang - One of the best experts on this subject based on the ideXlab platform.
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Expression, purification and characterization of yellow grouper Epinephelus awoara regulator of G protein signaling 16 protein
Fish physiology and biochemistry, 2011Co-Authors: Li WangAbstract:Regulators of G-protein signaling (RGS) proteins are a family of proteins, which accelerate GTPase-activity intrinsic to the alpha subunits of heterotrimeric G-proteins and play crucial roles in the physiological control of G-protein signaling. Here, yellow grouper RGS16 protein was expressed in Escherichia coli and purified by Ni–NTA affinity chromatography. The expression level of the fusion protein was up to 30% of the total cellular protein.Western blotting analysis showed that a band with the molecular mass of about 21 Kda was detected. The purified recombinant protein was used to prepare polyclonal antibody, and antiserum obtained was highly specific with the titer of over 1:32,000. Additionally, RGS16 protein was expressed in the Tn-5B1-4 insect cells. Western blotting analysis revealed that the expressed protein had immunoreactivity.
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Isolation and characterization of a regulator of G protein signaling (RGS16) gene homologue in yellow grouper Epinephelus awoara
Aquaculture, 2007Co-Authors: Li WangAbstract:Abstract Regulators of G protein signaling (RGS) are GTPase-activating proteins (GAP) which act as modulators of G protein-coupled receptors. We isolated a RGS16 homologue in yellow grouper (Epinephelus awoara) spleen using suppression subtractive hybridization and RACE-PCR. The nucleotide sequence of yellow grouper RGS16 full-length cDNA was 700 bp and contained an open reading frame of 537 bp, encoding a putative protein of 178 amino acids. The encoded protein shows 47–61% identities to other homologues. RT-PCR analysis demonstrated that RGS16 was expressed in yellow grouper spleen and up-regulated in kidney heart, liver, and anterior kidney by lipopolysaccharide. This study will help towards validating the specific function of RGS in marine fish.
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Promotion of glioma C6 cells proliferation by over-expressed RGS16
Journal of the Fourth Military Medical University, 2002Co-Authors: Feng Zhang, Guangsheng Chen, Bicheng Zhang, Li Wang, Sheng-cai LinAbstract:AIM To study the effect of RGS16 on the biological characteristics of glioma C6 cells. METHODS pCMV-5-RGS16 was transfected into C6 cells by lipofectin. The morphological and adhesive changes of the cells were observed under an inverted microscope. Proliferation of C6 cells was measured by ~3H-thymidine (~3H-TdR) assay after gradient transfections of pCMV5-RGS16 and pCMV5. Expression of RGS16 was examined by immunocytochemical method both before and after the transfection. Flow cytometry was adopted to measure changes in the fraction number of the cell cycle phase and to detect whether RGS16 could induce apoptosis of C6 cell. RESULTS 21 hours after the transfection of pCMV5-RGS16 approximately 30% of C6 cell grew round and 13% expressed RGS16, 36 h latter the positive relationship between the proliferation of C6 cells and the gradient transfections of pCMV5-RGS16 was displayed by ~3H-TdR assay. Flow cytometry showed that the fraction number of G1 phase of C6 cells reduced by 10% and that of S phase accumulated by 14% and RGS16 could not induce apoptosis of C6 cells. CONCLUSION RGS16 might promote the proliferation of C6 cells.
Stephan L. M. Peters - One of the best experts on this subject based on the ideXlab platform.
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Sphingosine-1-phosphate regulates RGS2 and RGS16 mRNA expression in vascular smooth muscle cells
European Journal of Pharmacology, 2009Co-Authors: Mariëlle C. Hendriks-balk, Najat Hajji, Pieter B. Van Loenen, Martin C. Michel, Stephan L. M. Peters, Astrid E. AlewijnseAbstract:Abstract Regulator of G protein signalling (RGS) protein expression is altered under growth promoting conditions in vascular smooth muscle cells (VSMCs). Since sphingosine-1-phosphate (S1P) is an important growth stimulatory factor, we investigated whether stimulation of VSMCs with S1P results in alterations in mRNA expression levels of several RGS proteins and which signalling components are involved. VSMCs were stimulated with S1P and mRNA expression levels of RGS2, RGS3, RGS4, RGS5 and RGS16 were measured by real-time polymerase chain reaction. S1P caused a time-dependent up-regulation of RGS2 and RGS16 mRNA expression. FTY720-P, a S1P1/S1P3–5 agonist, did not regulate RGS2 mRNA levels although it did up-regulate RGS16 mRNA expression. Pertussis toxin treatment revealed that the S1P-induced RGS16 expression was Gi/o-dependent whereas up-regulation of RGS2 mRNA was not. Phosphatidylinositol 3-kinase, protein kinase C and mitogen-activated protein kinase kinase apparently were not involved in the S1P-induced up-regulation of both RGS proteins. The present study demonstrates that S1P induces RGS2 and RGS16 mRNA expression but uses distinct S1P receptor subtypes and signalling pathways to regulate expression of these RGS proteins.
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LPS differentially affects vasoconstrictor responses: a potential role for RGS16?
Journal of physiology and biochemistry, 2009Co-Authors: Mariëlle C. Hendriks-balk, Najat Hajji, Astrid E. Alewijnse, M. Tjon-atsoi, Stephan L. M. PetersAbstract:The profound hypotension in septic shock patients is difficult to treat as it is accompanied by depressed constrictor responses to α1-adrenoceptor agonists. Bacterial lipopolysaccharide (LPS) is the main trigger for most of the cardiovascular alterations occurring in septic shock. In this study we investigated the effects of LPS exposure on vascular contractility in general and the role of Regulator of G protein Signalling (RGS) proteins in the LPS-induced vascular alterations. Exposure of rat aortic rings to various LPS concentrations (3, 10, 30 μg/ml) for 22 hours differentially affected agonist-induced contractile responses at four distinct G-protein coupled receptors (α1-adrenoceptors, angiotensin II, serotonin and endothelin-1 receptors). While the endothelin-1-induced contraction was unaffected by LPS pre-treatment, phenylephrine- and angiotensin II-induced contraction were significantly reduced whereas serotonin-induced contraction was significantly enhanced. Concomitantly, LPS treatment increased the RGS16 mRNA expression both in aortic rings and cultured vascular smooth muscle cells (VSMCs) but not that of RGS2, RGS3, RGS4 or RGS5. The significant increase in RGS16 mRNA expression in VSMCs by LPS was time- and concentration-dependent but independent of increased inducible NO synthase (iNOS) activity. The changes in RGS16 mRNA might contribute to the differential regulation of the contractile responses to vasoconstrictors upon LPS exposure.
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S1P receptor signalling and RGS proteins; expression and function in vascular smooth muscle cells and transfected CHO cells.
European Journal of Pharmacology, 2008Co-Authors: Mariëlle C. Hendriks-balk, Najat Hajji, Pieter B. Van Loenen, Martin C. Michel, Stephan L. M. Peters, Astrid E. AlewijnseAbstract:Abstract Sphingosine-1-phosphate (S1P) signalling via G protein-coupled receptors is important for the regulation of cell function and differentiation. Specific Regulators of G protein Signalling (RGS) proteins modulate the function of these receptors in many cell types including vascular smooth muscle cells (VSMCs). Therefore, we investigated the role of altered expression levels of RGS proteins in S1P receptor function in VSMCs and transfected CHO cells. The mRNA expression of the S1P 1 receptor, RGS4 and RGS16 were down-regulated in VSMCs during phenotypic modulation induced by culturing, whereas mRNA levels of RGS2, RGS3, S1P 2 and S1P 3 receptors were unchanged. Interestingly, the expression level of RGS5 was transiently up-regulated. Despite major alterations in RGS levels, S1P-induced calcium elevation in VSMCs was not altered. Co-transfection of RGS2, RGS3, RGS4, RGS5 and RGS16 into CHO-Flp-In cells stably expressing the S1P 1 or S1P 3 receptor did not modify S1P-induced inhibition of cAMP accumulation to a major extent. Similar results were obtained with SEW2871, a selective S1P 1 receptor agonist. However, the inhibition of cAMP accumulation by the agonist FTY720-P via the S1P 1 receptor was significantly decreased by co-transfection with RGS5. These results indicate that mRNA of the S1P 1 receptor, RGS4, RGS5 and RGS16 is differentially regulated during phenotypic modulation. However, major alterations in RGS protein expression have only limited effect on S1P receptor function.