Insulin influences the nitric oxide cyclic nucleotide pathway in cultured human smooth muscle cells from corpus cavernosum by rapidly activating a constitutive nitric oxide synthase
We have evaluated, in cultured human cavernosal smooth muscle cells, the expression and activity of calcium-dependent constitutive nitric oxide synthase (cNOS) and the ability of insulin to induce nitric oxide (NO) production and to increase intracellular cyclic nucleotides guanosine 3',5'...
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| Veröffentlicht in: | European journal of endocrinology 2002-11, Vol.147 (5), p.689-700 |
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| creator | ANFOSSI, Giovanni MASSUCCO, Paola TROVATI, Mariella MATTIELLO, Luigi BALBO, Alessandra RUSSO, Isabella DORONZO, Gabriella ROLLE, Luigi GHIGO, Dario FONTANA, Dario BOSIA, Amalia |
| description | We have evaluated, in cultured human cavernosal smooth muscle cells, the expression and activity of calcium-dependent constitutive nitric oxide synthase (cNOS) and the ability of insulin to induce nitric oxide (NO) production and to increase intracellular cyclic nucleotides guanosine 3',5'-cyclic monophosphate (cGMP) and adenosine 3',5'-cyclic monophosphate (cAMP).
cNOS mRNA was detected by RT-PCR amplification, cNOS protein by immunofluorescence, cNOS activity as l-[3H]-citrulline production from l-[3H]-arginine and cyclic nucleotides by radioimmunoassay.
cNOS mRNA and cNOS protein were found in cultured cells; cNOS activity was increased by 5-min exposure to 1 micro mol/l calcium ionophore ionomycin (from 0.1094+/-0.0229 to 0.2685+/-0.0560 pmol/min per mg cell protein, P=0.011) and to 2 nmol/l insulin (from 0.1214+/-0.0149 to 0.2045+/-0.0290 pmol/min per mg cell protein, P=0.041). Insulin increased both cGMP and cAMP in a dose- and time-dependent manner (i.e. with 2 nmol/l insulin, cGMP rose from 2.71+/-0.10 to 6.80+/-0.40 pmol/10(6) cells at 30 min, P=0.0001; cAMP from 1.26+/-0.06 to 3.02+/-0.30 pmol/10(6) cells at 60 min, P=0.0001). NOS inhibitor N(G)-monomethyl-l-arginine and phosphatidylinositol 3-kinase (PI 3-kinase) inhibitors wortmannin and LY 294002 blunted these effects of insulin. The action of insulin on cyclic nucleotides persisted in the presence of phosphodiesterase inhibition, guanylate cyclase activation by NO donors and adenylate cyclase activation by Iloprost or forskolin.
Human cavernosal smooth muscle cells, by expressing cNOS activity, are a source of NO and not only its target; in these cells, insulin rapidly activates cNOS through a PI 3-kinase pathway, with a consequent increase of both cyclic nucleotides, thus directly influencing the mechanisms involved in penile vascular tone and interplaying with classical haemodynamic mediators. |
| doi_str_mv | 10.1530/eje.0.1470689 |
| format | Article |
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cNOS mRNA was detected by RT-PCR amplification, cNOS protein by immunofluorescence, cNOS activity as l-[3H]-citrulline production from l-[3H]-arginine and cyclic nucleotides by radioimmunoassay.
cNOS mRNA and cNOS protein were found in cultured cells; cNOS activity was increased by 5-min exposure to 1 micro mol/l calcium ionophore ionomycin (from 0.1094+/-0.0229 to 0.2685+/-0.0560 pmol/min per mg cell protein, P=0.011) and to 2 nmol/l insulin (from 0.1214+/-0.0149 to 0.2045+/-0.0290 pmol/min per mg cell protein, P=0.041). Insulin increased both cGMP and cAMP in a dose- and time-dependent manner (i.e. with 2 nmol/l insulin, cGMP rose from 2.71+/-0.10 to 6.80+/-0.40 pmol/10(6) cells at 30 min, P=0.0001; cAMP from 1.26+/-0.06 to 3.02+/-0.30 pmol/10(6) cells at 60 min, P=0.0001). NOS inhibitor N(G)-monomethyl-l-arginine and phosphatidylinositol 3-kinase (PI 3-kinase) inhibitors wortmannin and LY 294002 blunted these effects of insulin. The action of insulin on cyclic nucleotides persisted in the presence of phosphodiesterase inhibition, guanylate cyclase activation by NO donors and adenylate cyclase activation by Iloprost or forskolin.
Human cavernosal smooth muscle cells, by expressing cNOS activity, are a source of NO and not only its target; in these cells, insulin rapidly activates cNOS through a PI 3-kinase pathway, with a consequent increase of both cyclic nucleotides, thus directly influencing the mechanisms involved in penile vascular tone and interplaying with classical haemodynamic mediators.</description><identifier>ISSN: 0804-4643</identifier><identifier>EISSN: 1479-683X</identifier><identifier>DOI: 10.1530/eje.0.1470689</identifier><identifier>PMID: 12444902</identifier><language>eng</language><publisher>Colchester: Portland Press</publisher><subject>Adult ; Biological and medical sciences ; Calcium - physiology ; Cell Division ; Cells, Cultured ; Cyclic AMP - metabolism ; Cyclic GMP - metabolism ; Dose-Response Relationship, Drug ; Enzyme Activation - physiology ; Fundamental and applied biological sciences. Psychology ; Guanylate Cyclase - metabolism ; Hormone metabolism and regulation ; Humans ; Insulin - administration & dosage ; Insulin - pharmacology ; Insulin - physiology ; Male ; Mammalian male genital system ; Muscle, Smooth - cytology ; Muscle, Smooth - drug effects ; Muscle, Smooth - metabolism ; Nitric Oxide - biosynthesis ; Nitric Oxide - metabolism ; Nitric Oxide Donors - pharmacology ; Nitric Oxide Synthase - genetics ; Nitric Oxide Synthase - metabolism ; Nucleotides, Cyclic - metabolism ; Penis - cytology ; Penis - drug effects ; Penis - metabolism ; Phosphatidylinositol 3-Kinases - metabolism ; Phosphodiesterase Inhibitors - pharmacology ; RNA, Messenger - metabolism ; Signal Transduction - physiology ; Time Factors ; Vertebrates: reproduction</subject><ispartof>European journal of endocrinology, 2002-11, Vol.147 (5), p.689-700</ispartof><rights>2003 INIST-CNRS</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c387t-5d2a508774a3014917376ef316b6d82a5af462a8265586be175d8951df2e507f3</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,776,780,27901,27902</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=14038948$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/12444902$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>ANFOSSI, Giovanni</creatorcontrib><creatorcontrib>MASSUCCO, Paola</creatorcontrib><creatorcontrib>TROVATI, Mariella</creatorcontrib><creatorcontrib>MATTIELLO, Luigi</creatorcontrib><creatorcontrib>BALBO, Alessandra</creatorcontrib><creatorcontrib>RUSSO, Isabella</creatorcontrib><creatorcontrib>DORONZO, Gabriella</creatorcontrib><creatorcontrib>ROLLE, Luigi</creatorcontrib><creatorcontrib>GHIGO, Dario</creatorcontrib><creatorcontrib>FONTANA, Dario</creatorcontrib><creatorcontrib>BOSIA, Amalia</creatorcontrib><title>Insulin influences the nitric oxide cyclic nucleotide pathway in cultured human smooth muscle cells from corpus cavernosum by rapidly activating a constitutive nitric oxide synthase</title><title>European journal of endocrinology</title><addtitle>Eur J Endocrinol</addtitle><description>We have evaluated, in cultured human cavernosal smooth muscle cells, the expression and activity of calcium-dependent constitutive nitric oxide synthase (cNOS) and the ability of insulin to induce nitric oxide (NO) production and to increase intracellular cyclic nucleotides guanosine 3',5'-cyclic monophosphate (cGMP) and adenosine 3',5'-cyclic monophosphate (cAMP).
cNOS mRNA was detected by RT-PCR amplification, cNOS protein by immunofluorescence, cNOS activity as l-[3H]-citrulline production from l-[3H]-arginine and cyclic nucleotides by radioimmunoassay.
cNOS mRNA and cNOS protein were found in cultured cells; cNOS activity was increased by 5-min exposure to 1 micro mol/l calcium ionophore ionomycin (from 0.1094+/-0.0229 to 0.2685+/-0.0560 pmol/min per mg cell protein, P=0.011) and to 2 nmol/l insulin (from 0.1214+/-0.0149 to 0.2045+/-0.0290 pmol/min per mg cell protein, P=0.041). Insulin increased both cGMP and cAMP in a dose- and time-dependent manner (i.e. with 2 nmol/l insulin, cGMP rose from 2.71+/-0.10 to 6.80+/-0.40 pmol/10(6) cells at 30 min, P=0.0001; cAMP from 1.26+/-0.06 to 3.02+/-0.30 pmol/10(6) cells at 60 min, P=0.0001). NOS inhibitor N(G)-monomethyl-l-arginine and phosphatidylinositol 3-kinase (PI 3-kinase) inhibitors wortmannin and LY 294002 blunted these effects of insulin. The action of insulin on cyclic nucleotides persisted in the presence of phosphodiesterase inhibition, guanylate cyclase activation by NO donors and adenylate cyclase activation by Iloprost or forskolin.
Human cavernosal smooth muscle cells, by expressing cNOS activity, are a source of NO and not only its target; in these cells, insulin rapidly activates cNOS through a PI 3-kinase pathway, with a consequent increase of both cyclic nucleotides, thus directly influencing the mechanisms involved in penile vascular tone and interplaying with classical haemodynamic mediators.</description><subject>Adult</subject><subject>Biological and medical sciences</subject><subject>Calcium - physiology</subject><subject>Cell Division</subject><subject>Cells, Cultured</subject><subject>Cyclic AMP - metabolism</subject><subject>Cyclic GMP - metabolism</subject><subject>Dose-Response Relationship, Drug</subject><subject>Enzyme Activation - physiology</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Guanylate Cyclase - metabolism</subject><subject>Hormone metabolism and regulation</subject><subject>Humans</subject><subject>Insulin - administration & dosage</subject><subject>Insulin - pharmacology</subject><subject>Insulin - physiology</subject><subject>Male</subject><subject>Mammalian male genital system</subject><subject>Muscle, Smooth - cytology</subject><subject>Muscle, Smooth - drug effects</subject><subject>Muscle, Smooth - metabolism</subject><subject>Nitric Oxide - biosynthesis</subject><subject>Nitric Oxide - metabolism</subject><subject>Nitric Oxide Donors - pharmacology</subject><subject>Nitric Oxide Synthase - genetics</subject><subject>Nitric Oxide Synthase - metabolism</subject><subject>Nucleotides, Cyclic - metabolism</subject><subject>Penis - cytology</subject><subject>Penis - drug effects</subject><subject>Penis - metabolism</subject><subject>Phosphatidylinositol 3-Kinases - metabolism</subject><subject>Phosphodiesterase Inhibitors - pharmacology</subject><subject>RNA, Messenger - metabolism</subject><subject>Signal Transduction - physiology</subject><subject>Time Factors</subject><subject>Vertebrates: reproduction</subject><issn>0804-4643</issn><issn>1479-683X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2002</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpdkU1v1DAQhi0EokvhyBX5ArcUO3Zi54gqPipV4gISt8jrTIgrx148dkt-GP8Pr3alSj3NOzPPjGb0EvKWsyveCfYR7uCqSqlYr4dnZFfV0PRa_HpOdkwz2cheigvyCvGOMV41e0kueCulHFi7I_9uAhbvAnVh9gWCBaR5ARpcTs7S-NdNQO1mfU1CsR5iPlYOJi8PZqtT1BafS4KJLmU1geIaY17oWrDC1IL3SOcUV2pjOhSk1txDChHLSvcbTebgJr9RY7O7N9mF39RUMmB2udTSk0NwC3kxCK_Ji9l4hDfneEl-fvn84_pbc_v96831p9vGCq1y002t6ZhWShrBuBy4EqqHWfB-30-69sws-9botu863e-Bq27SQ8enuYWOqVlckg-nvYcU_xTAPK4Ojz-ZALHgqFrVCs11BZsTaFNETDCPh-RWk7aRs_Ho01h9Gqs8-VT5d-fFZb_C9EifjanA-zNg0Bo_JxOsw0dOMqEHqcV_LoCgTg</recordid><startdate>20021101</startdate><enddate>20021101</enddate><creator>ANFOSSI, Giovanni</creator><creator>MASSUCCO, Paola</creator><creator>TROVATI, Mariella</creator><creator>MATTIELLO, Luigi</creator><creator>BALBO, Alessandra</creator><creator>RUSSO, Isabella</creator><creator>DORONZO, Gabriella</creator><creator>ROLLE, Luigi</creator><creator>GHIGO, Dario</creator><creator>FONTANA, Dario</creator><creator>BOSIA, Amalia</creator><general>Portland Press</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope></search><sort><creationdate>20021101</creationdate><title>Insulin influences the nitric oxide cyclic nucleotide pathway in cultured human smooth muscle cells from corpus cavernosum by rapidly activating a constitutive nitric oxide synthase</title><author>ANFOSSI, Giovanni ; MASSUCCO, Paola ; TROVATI, Mariella ; MATTIELLO, Luigi ; BALBO, Alessandra ; RUSSO, Isabella ; DORONZO, Gabriella ; ROLLE, Luigi ; GHIGO, Dario ; FONTANA, Dario ; BOSIA, Amalia</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c387t-5d2a508774a3014917376ef316b6d82a5af462a8265586be175d8951df2e507f3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2002</creationdate><topic>Adult</topic><topic>Biological and medical sciences</topic><topic>Calcium - physiology</topic><topic>Cell Division</topic><topic>Cells, Cultured</topic><topic>Cyclic AMP - metabolism</topic><topic>Cyclic GMP - metabolism</topic><topic>Dose-Response Relationship, Drug</topic><topic>Enzyme Activation - physiology</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Guanylate Cyclase - metabolism</topic><topic>Hormone metabolism and regulation</topic><topic>Humans</topic><topic>Insulin - administration & dosage</topic><topic>Insulin - pharmacology</topic><topic>Insulin - physiology</topic><topic>Male</topic><topic>Mammalian male genital system</topic><topic>Muscle, Smooth - cytology</topic><topic>Muscle, Smooth - drug effects</topic><topic>Muscle, Smooth - metabolism</topic><topic>Nitric Oxide - biosynthesis</topic><topic>Nitric Oxide - metabolism</topic><topic>Nitric Oxide Donors - pharmacology</topic><topic>Nitric Oxide Synthase - genetics</topic><topic>Nitric Oxide Synthase - metabolism</topic><topic>Nucleotides, Cyclic - metabolism</topic><topic>Penis - cytology</topic><topic>Penis - drug effects</topic><topic>Penis - metabolism</topic><topic>Phosphatidylinositol 3-Kinases - metabolism</topic><topic>Phosphodiesterase Inhibitors - pharmacology</topic><topic>RNA, Messenger - metabolism</topic><topic>Signal Transduction - physiology</topic><topic>Time Factors</topic><topic>Vertebrates: reproduction</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>ANFOSSI, Giovanni</creatorcontrib><creatorcontrib>MASSUCCO, Paola</creatorcontrib><creatorcontrib>TROVATI, Mariella</creatorcontrib><creatorcontrib>MATTIELLO, Luigi</creatorcontrib><creatorcontrib>BALBO, Alessandra</creatorcontrib><creatorcontrib>RUSSO, Isabella</creatorcontrib><creatorcontrib>DORONZO, Gabriella</creatorcontrib><creatorcontrib>ROLLE, Luigi</creatorcontrib><creatorcontrib>GHIGO, Dario</creatorcontrib><creatorcontrib>FONTANA, Dario</creatorcontrib><creatorcontrib>BOSIA, Amalia</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>European journal of endocrinology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>ANFOSSI, Giovanni</au><au>MASSUCCO, Paola</au><au>TROVATI, Mariella</au><au>MATTIELLO, Luigi</au><au>BALBO, Alessandra</au><au>RUSSO, Isabella</au><au>DORONZO, Gabriella</au><au>ROLLE, Luigi</au><au>GHIGO, Dario</au><au>FONTANA, Dario</au><au>BOSIA, Amalia</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Insulin influences the nitric oxide cyclic nucleotide pathway in cultured human smooth muscle cells from corpus cavernosum by rapidly activating a constitutive nitric oxide synthase</atitle><jtitle>European journal of endocrinology</jtitle><addtitle>Eur J Endocrinol</addtitle><date>2002-11-01</date><risdate>2002</risdate><volume>147</volume><issue>5</issue><spage>689</spage><epage>700</epage><pages>689-700</pages><issn>0804-4643</issn><eissn>1479-683X</eissn><abstract>We have evaluated, in cultured human cavernosal smooth muscle cells, the expression and activity of calcium-dependent constitutive nitric oxide synthase (cNOS) and the ability of insulin to induce nitric oxide (NO) production and to increase intracellular cyclic nucleotides guanosine 3',5'-cyclic monophosphate (cGMP) and adenosine 3',5'-cyclic monophosphate (cAMP).
cNOS mRNA was detected by RT-PCR amplification, cNOS protein by immunofluorescence, cNOS activity as l-[3H]-citrulline production from l-[3H]-arginine and cyclic nucleotides by radioimmunoassay.
cNOS mRNA and cNOS protein were found in cultured cells; cNOS activity was increased by 5-min exposure to 1 micro mol/l calcium ionophore ionomycin (from 0.1094+/-0.0229 to 0.2685+/-0.0560 pmol/min per mg cell protein, P=0.011) and to 2 nmol/l insulin (from 0.1214+/-0.0149 to 0.2045+/-0.0290 pmol/min per mg cell protein, P=0.041). Insulin increased both cGMP and cAMP in a dose- and time-dependent manner (i.e. with 2 nmol/l insulin, cGMP rose from 2.71+/-0.10 to 6.80+/-0.40 pmol/10(6) cells at 30 min, P=0.0001; cAMP from 1.26+/-0.06 to 3.02+/-0.30 pmol/10(6) cells at 60 min, P=0.0001). NOS inhibitor N(G)-monomethyl-l-arginine and phosphatidylinositol 3-kinase (PI 3-kinase) inhibitors wortmannin and LY 294002 blunted these effects of insulin. The action of insulin on cyclic nucleotides persisted in the presence of phosphodiesterase inhibition, guanylate cyclase activation by NO donors and adenylate cyclase activation by Iloprost or forskolin.
Human cavernosal smooth muscle cells, by expressing cNOS activity, are a source of NO and not only its target; in these cells, insulin rapidly activates cNOS through a PI 3-kinase pathway, with a consequent increase of both cyclic nucleotides, thus directly influencing the mechanisms involved in penile vascular tone and interplaying with classical haemodynamic mediators.</abstract><cop>Colchester</cop><pub>Portland Press</pub><pmid>12444902</pmid><doi>10.1530/eje.0.1470689</doi><tpages>12</tpages><oa>free_for_read</oa></addata></record> |
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| subjects | Adult Biological and medical sciences Calcium - physiology Cell Division Cells, Cultured Cyclic AMP - metabolism Cyclic GMP - metabolism Dose-Response Relationship, Drug Enzyme Activation - physiology Fundamental and applied biological sciences. Psychology Guanylate Cyclase - metabolism Hormone metabolism and regulation Humans Insulin - administration & dosage Insulin - pharmacology Insulin - physiology Male Mammalian male genital system Muscle, Smooth - cytology Muscle, Smooth - drug effects Muscle, Smooth - metabolism Nitric Oxide - biosynthesis Nitric Oxide - metabolism Nitric Oxide Donors - pharmacology Nitric Oxide Synthase - genetics Nitric Oxide Synthase - metabolism Nucleotides, Cyclic - metabolism Penis - cytology Penis - drug effects Penis - metabolism Phosphatidylinositol 3-Kinases - metabolism Phosphodiesterase Inhibitors - pharmacology RNA, Messenger - metabolism Signal Transduction - physiology Time Factors Vertebrates: reproduction |
| title | Insulin influences the nitric oxide cyclic nucleotide pathway in cultured human smooth muscle cells from corpus cavernosum by rapidly activating a constitutive nitric oxide synthase |
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