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Published in: European Journal of Applied Physiology 1/2012

01-01-2012 | Original Article

Exercise-induced angiogenesis correlates with the up-regulated expression of neuronal nitric oxide synthase (nNOS) in human skeletal muscle

Authors: Felicitas A. M. Huber-Abel, Mélanie Gerber, Hans Hoppeler, Oliver Baum

Published in: European Journal of Applied Physiology | Issue 1/2012

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Abstract

The contribution of neuronal nitric oxide synthase (nNOS) to angiogenesis in human skeletal muscle after endurance exercise is controversially discussed. We therefore ascertained whether the expression of nNOS is associated with the capillary density in biopsies of the vastus lateralis (VL) muscle that had been derived from 10 sedentary male subjects before and after moderate training (four 30-min weekly jogging sessions for 6 months, with a heart-rate corresponding to 75% VO2max). In these biopsies, nNOS was predominantly expressed as alpha-isoform with exon-mu and to a lesser extent without exon-mu, as determined by RT-PCR. The mRNA levels of nNOS were quantified by real-time PCR and related to the capillary-to-fibre ratio and the numerical density of capillaries specified by light microscopy. If the VL biopsies of all subjects were co-analysed, mRNA levels of nNOS were non-significantly elevated after training (+34%; P > 0.05). However, only five of the ten subjects exhibited significant (P ≤ 0.05) elevations in the capillary-to-fibre ratio (+25%) and the numerical density of capillaries (+21%) and were thus undergoing angiogenesis. If the VL biopsies of these five subjects alone were evaluated, the mRNA levels of nNOS were significantly up-regulated (+128%; P ≤ 0.05) and correlated positively (r = 0.8; P ≤ 0.01) to angiogenesis. Accordingly, nNOS protein expression in VL biopsies quantified by immunoblotting was significantly increased (+82%; P ≤ 0.05) only in those subjects that underwent angiogenesis. In conclusion, the expression of nNOS at mRNA and protein levels was statistically linked to capillarity after exercise suggesting that nNOS is involved in the angiogenic response to training in human skeletal muscle.
Literature
go back to reference Arany Z, Foo SY, Ma Y, Ruas JL, Bommi-Reddy A, Girnun G, Cooper M, Laznik D, Chinsomboon J, Rangwala SM, Baek KH, Rosenzweig A, Spiegelman BM (2008) HIF-independent regulation of VEGF and angiogenesis by the transcriptional coactivator PGC-1alpha. Nature 451:1008–1012PubMedCrossRef Arany Z, Foo SY, Ma Y, Ruas JL, Bommi-Reddy A, Girnun G, Cooper M, Laznik D, Chinsomboon J, Rangwala SM, Baek KH, Rosenzweig A, Spiegelman BM (2008) HIF-independent regulation of VEGF and angiogenesis by the transcriptional coactivator PGC-1alpha. Nature 451:1008–1012PubMedCrossRef
go back to reference Baum O, Da Silva-Azevedo L, Willerding G, Wöckel A, Planitzer G, Gossrau R, Pries AR, Zakrzewicz A (2004) Endothelial NOS is main mediator for shear stress-dependent angiogenesis in skeletal muscle after prazosin administration. Am J Physiol Heart Circ Physiol 287:H2300–H2308PubMedCrossRef Baum O, Da Silva-Azevedo L, Willerding G, Wöckel A, Planitzer G, Gossrau R, Pries AR, Zakrzewicz A (2004) Endothelial NOS is main mediator for shear stress-dependent angiogenesis in skeletal muscle after prazosin administration. Am J Physiol Heart Circ Physiol 287:H2300–H2308PubMedCrossRef
go back to reference Bouchard C, Rankinen T (2001) Individual differences in response to regular physical activity. Med Sci Sports Exerc 33:446–451CrossRef Bouchard C, Rankinen T (2001) Individual differences in response to regular physical activity. Med Sci Sports Exerc 33:446–451CrossRef
go back to reference Bradley SJ, Kingwell BA, Canny BJ, McConell GK (2007) Skeletal muscle neuronal nitric oxide synthase micro protein is reduced in people with impaired glucose homeostasis and is not normalized by exercise training. Metabolism 56:1405–1411PubMedCrossRef Bradley SJ, Kingwell BA, Canny BJ, McConell GK (2007) Skeletal muscle neuronal nitric oxide synthase micro protein is reduced in people with impaired glucose homeostasis and is not normalized by exercise training. Metabolism 56:1405–1411PubMedCrossRef
go back to reference Bray MS, Hagberg JM, Perusse L, Rankinen T, Roth SM, Wolfarth B, Bouchard C (2009) The human gene map for performance and health-related fitness phenotypes: the 2006–2007 update. Med Sci Sports Exerc 41:35–73PubMed Bray MS, Hagberg JM, Perusse L, Rankinen T, Roth SM, Wolfarth B, Bouchard C (2009) The human gene map for performance and health-related fitness phenotypes: the 2006–2007 update. Med Sci Sports Exerc 41:35–73PubMed
go back to reference Chinsomboon J, Ruas J, Gupta RK, Thom R, Shoag J, Rowe GC, Sawada N, Raghuram S, Arany Z (2009) The transcriptional coactivator PGC-1alpha mediates exercise-induced angiogenesis in skeletal muscle. Proc Natl Acad Sci U S A 106:21401–21406PubMedCrossRef Chinsomboon J, Ruas J, Gupta RK, Thom R, Shoag J, Rowe GC, Sawada N, Raghuram S, Arany Z (2009) The transcriptional coactivator PGC-1alpha mediates exercise-induced angiogenesis in skeletal muscle. Proc Natl Acad Sci U S A 106:21401–21406PubMedCrossRef
go back to reference Copp SW, Hirai DM, Schwagerl PJ, Musch TI, Poole DC (2010) Effects of neuronal nitric oxide synthase inhibition on resting and exercising hindlimb muscle blood flow in the rat. J Physiol 588:1321–1331PubMedCrossRef Copp SW, Hirai DM, Schwagerl PJ, Musch TI, Poole DC (2010) Effects of neuronal nitric oxide synthase inhibition on resting and exercising hindlimb muscle blood flow in the rat. J Physiol 588:1321–1331PubMedCrossRef
go back to reference Da Silva-Azevedo L, Baum O, Zakrzewicz A, Pries AR (2002) Vascular endothelial growth factor is expressed in endothelial cells isolated from skeletal muscles of nitric oxide synthase knockout mice during prazosin-induced angiogenesis. Biochem Biophys Res Commun 297:1270–1276PubMedCrossRef Da Silva-Azevedo L, Baum O, Zakrzewicz A, Pries AR (2002) Vascular endothelial growth factor is expressed in endothelial cells isolated from skeletal muscles of nitric oxide synthase knockout mice during prazosin-induced angiogenesis. Biochem Biophys Res Commun 297:1270–1276PubMedCrossRef
go back to reference Da Silva-Azevedo L, Jahne S, Hoffmann C, Stalder D, Heller M, Pries AR, Zakrzewicz A, Baum O (2009) Up-regulation of the peroxiredoxin-6 related metabolism of reactive oxygen species in skeletal muscle of mice lacking neuronal nitric oxide synthase. J Physiol 587:655–668PubMedCrossRef Da Silva-Azevedo L, Jahne S, Hoffmann C, Stalder D, Heller M, Pries AR, Zakrzewicz A, Baum O (2009) Up-regulation of the peroxiredoxin-6 related metabolism of reactive oxygen species in skeletal muscle of mice lacking neuronal nitric oxide synthase. J Physiol 587:655–668PubMedCrossRef
go back to reference Frandsen U, Hoffner L, Betak A, Saltin B, Bangsbo J, Hellsten Y (2000) Endurance training does not alter the level of neuronal nitric oxide synthase in human skeletal muscle. J Appl Physiol 89:1033–1038PubMed Frandsen U, Hoffner L, Betak A, Saltin B, Bangsbo J, Hellsten Y (2000) Endurance training does not alter the level of neuronal nitric oxide synthase in human skeletal muscle. J Appl Physiol 89:1033–1038PubMed
go back to reference Gavin TP (2009) Basal and exercise-induced regulation of skeletal muscle capillarization. Exerc Sport Sci Rev 37:86–92PubMedCrossRef Gavin TP (2009) Basal and exercise-induced regulation of skeletal muscle capillarization. Exerc Sport Sci Rev 37:86–92PubMedCrossRef
go back to reference Gustafsson T, Rundqvist H, Norrbom J, Rullman E, Jansson E, Sundberg CJ (2007) The influence of physical training on the angiopoietin and VEGF-A systems in human skeletal muscle. J Appl Physiol 103:1012–1020 Gustafsson T, Rundqvist H, Norrbom J, Rullman E, Jansson E, Sundberg CJ (2007) The influence of physical training on the angiopoietin and VEGF-A systems in human skeletal muscle. J Appl Physiol 103:1012–1020
go back to reference Hoppeler H, Howald H, Conley K, Lindstedt SL, Claassen H, Vock P, Weibel ER (1985) Endurance training in humans: aerobic capacity and structure of skeletal muscle. J Appl Physiol 59:320–327PubMed Hoppeler H, Howald H, Conley K, Lindstedt SL, Claassen H, Vock P, Weibel ER (1985) Endurance training in humans: aerobic capacity and structure of skeletal muscle. J Appl Physiol 59:320–327PubMed
go back to reference Hoppeler H, Klossner S, Fluck M (2007) Gene expression in working skeletal muscle. Adv Exp Med Biol 618:245–254PubMedCrossRef Hoppeler H, Klossner S, Fluck M (2007) Gene expression in working skeletal muscle. Adv Exp Med Biol 618:245–254PubMedCrossRef
go back to reference Hudlicka O (1998) Is physiological angiogenesis in skeletal muscle regulated by changes in microcirculation? Microcirculation 5:7–23PubMed Hudlicka O (1998) Is physiological angiogenesis in skeletal muscle regulated by changes in microcirculation? Microcirculation 5:7–23PubMed
go back to reference Hudlicka O, Brown MD (2009) Adaptation of skeletal muscle microvasculature to increased or decreased blood flow: role of shear stress, nitric oxide and vascular endothelial growth factor. J Vasc Res 46:504–512PubMedCrossRef Hudlicka O, Brown MD (2009) Adaptation of skeletal muscle microvasculature to increased or decreased blood flow: role of shear stress, nitric oxide and vascular endothelial growth factor. J Vasc Res 46:504–512PubMedCrossRef
go back to reference Hudlicka O, Brown MD, Silgram H (2000) Inhibition of capillary growth in chronically stimulated rat muscles by N(G)-nitro-l-arginine, nitric oxide synthase inhibitor. Microvasc Res 59:45–51PubMedCrossRef Hudlicka O, Brown MD, Silgram H (2000) Inhibition of capillary growth in chronically stimulated rat muscles by N(G)-nitro-l-arginine, nitric oxide synthase inhibitor. Microvasc Res 59:45–51PubMedCrossRef
go back to reference Jackson MJ, Pye D, Palomero J (2007) The production of reactive oxygen and nitrogen species by skeletal muscle. J Appl Physiol 102:1664–1670PubMedCrossRef Jackson MJ, Pye D, Palomero J (2007) The production of reactive oxygen and nitrogen species by skeletal muscle. J Appl Physiol 102:1664–1670PubMedCrossRef
go back to reference Kobayashi YM, Rader EP, Crawford RW, Iyengar NK, Thedens DR, Faulkner JA, Parikh SV, Weiss RM, Chamberlain JS, Moore SA, Campbell KP (2008) Sarcolemma-localized nNOS is required to maintain activity after mild exercise. Nature 456:511–515PubMedCrossRef Kobayashi YM, Rader EP, Crawford RW, Iyengar NK, Thedens DR, Faulkner JA, Parikh SV, Weiss RM, Chamberlain JS, Moore SA, Campbell KP (2008) Sarcolemma-localized nNOS is required to maintain activity after mild exercise. Nature 456:511–515PubMedCrossRef
go back to reference Kobzik L, Reid MB, Bredt DS, Stamler JS (1994) Nitric oxide in skeletal muscle. Nature 372:546–548PubMedCrossRef Kobzik L, Reid MB, Bredt DS, Stamler JS (1994) Nitric oxide in skeletal muscle. Nature 372:546–548PubMedCrossRef
go back to reference Laine R, de Montellano PR (1998) Neuronal nitric oxide synthase isoforms alpha and mu are closely related calpain-sensitive proteins. Mol Pharmacol 54:305–312PubMed Laine R, de Montellano PR (1998) Neuronal nitric oxide synthase isoforms alpha and mu are closely related calpain-sensitive proteins. Mol Pharmacol 54:305–312PubMed
go back to reference Lau KS, Grange RW, Isotani E, Sarelius IH, Kamm KE, Huang PL, Stull JT (2000) nNOS and eNOS modulate cGMP formation and vascular response in contracting fast-twitch skeletal muscle. Physiol Genomics 2:21–27PubMed Lau KS, Grange RW, Isotani E, Sarelius IH, Kamm KE, Huang PL, Stull JT (2000) nNOS and eNOS modulate cGMP formation and vascular response in contracting fast-twitch skeletal muscle. Physiol Genomics 2:21–27PubMed
go back to reference Leick L, Hellsten Y, Fentz J, Lyngby SS, Wojtaszewski JF, Hidalgo J, Pilegaard H (2009) PGC-1alpha mediates exercise-induced skeletal muscle VEGF expression in mice. Am J Physiol Endocrinol Metab 297:E92–E103PubMedCrossRef Leick L, Hellsten Y, Fentz J, Lyngby SS, Wojtaszewski JF, Hidalgo J, Pilegaard H (2009) PGC-1alpha mediates exercise-induced skeletal muscle VEGF expression in mice. Am J Physiol Endocrinol Metab 297:E92–E103PubMedCrossRef
go back to reference McConell GK, Bradley SJ, Stephens TJ, Canny BJ, Kingwell BA, Lee-Young RS (2007) Skeletal muscle nNOS mu protein content is increased by exercise training in humans. Am J Physiol Regul Integr Comp Physiol 293:R821–R828PubMedCrossRef McConell GK, Bradley SJ, Stephens TJ, Canny BJ, Kingwell BA, Lee-Young RS (2007) Skeletal muscle nNOS mu protein content is increased by exercise training in humans. Am J Physiol Regul Integr Comp Physiol 293:R821–R828PubMedCrossRef
go back to reference Melikian N, Seddon MD, Casadei B, Chowienczyk PJ, Shah AM (2009) Neuronal nitric oxide synthase and human vascular regulation. Trends Cardiovasc Med 19:256–262PubMedCrossRef Melikian N, Seddon MD, Casadei B, Chowienczyk PJ, Shah AM (2009) Neuronal nitric oxide synthase and human vascular regulation. Trends Cardiovasc Med 19:256–262PubMedCrossRef
go back to reference Percival JM, Anderson KN, Gregorevic P, Chamberlain JS, Froehner SC (2008) Functional deficits in nNOSmu-deficient skeletal muscle: myopathy in nNOS knockout mice. PLoS One 3:e3387PubMedCrossRef Percival JM, Anderson KN, Gregorevic P, Chamberlain JS, Froehner SC (2008) Functional deficits in nNOSmu-deficient skeletal muscle: myopathy in nNOS knockout mice. PLoS One 3:e3387PubMedCrossRef
go back to reference Rudnick J, Puttmann B, Tesch PA, Alkner B, Schoser BG, Salanova M, Kirsch K, Gunga HC, Schiffl G, Luck G, Blottner D (2004) Differential expression of nitric oxide synthases (NOS 1–3) in human skeletal muscle following exercise countermeasure during 12 weeks of bed rest. FASEB J 18:1228–1230PubMed Rudnick J, Puttmann B, Tesch PA, Alkner B, Schoser BG, Salanova M, Kirsch K, Gunga HC, Schiffl G, Luck G, Blottner D (2004) Differential expression of nitric oxide synthases (NOS 1–3) in human skeletal muscle following exercise countermeasure during 12 weeks of bed rest. FASEB J 18:1228–1230PubMed
go back to reference Salanova M, Schiffl G, Puttmann B, Schoser BG, Blottner D (2008) Molecular biomarkers monitoring human skeletal muscle fibres and microvasculature following long-term bed rest with and without countermeasures. J Anat 212:306–318PubMedCrossRef Salanova M, Schiffl G, Puttmann B, Schoser BG, Blottner D (2008) Molecular biomarkers monitoring human skeletal muscle fibres and microvasculature following long-term bed rest with and without countermeasures. J Anat 212:306–318PubMedCrossRef
go back to reference Sarelius I, Pohl U (2010) Control of muscle blood flow during exercise: local factors and integrative mechanisms. Acta Physiol (Oxf) 199:349–365CrossRef Sarelius I, Pohl U (2010) Control of muscle blood flow during exercise: local factors and integrative mechanisms. Acta Physiol (Oxf) 199:349–365CrossRef
go back to reference Silvagno F, Xia H, Bredt DS (1996) Neuronal nitric-oxide synthase-mu, an alternatively spliced isoform expressed in differentiated skeletal muscle. J Biol Chem 271:11204–11208PubMedCrossRef Silvagno F, Xia H, Bredt DS (1996) Neuronal nitric-oxide synthase-mu, an alternatively spliced isoform expressed in differentiated skeletal muscle. J Biol Chem 271:11204–11208PubMedCrossRef
go back to reference Stamler JS, Meissner G (2001) Physiology of nitric oxide in skeletal muscle. Physiol Rev 81:209–237PubMed Stamler JS, Meissner G (2001) Physiology of nitric oxide in skeletal muscle. Physiol Rev 81:209–237PubMed
go back to reference Suter E, Hoppeler H, Claassen H, Billeter R, Aebi U, Horber F, Jaeger P, Marti B (1995) Ultrastructural modification of human skeletal muscle tissue with 6-month moderate-intensity exercise training. Int J Sports Med 16:160–166PubMedCrossRef Suter E, Hoppeler H, Claassen H, Billeter R, Aebi U, Horber F, Jaeger P, Marti B (1995) Ultrastructural modification of human skeletal muscle tissue with 6-month moderate-intensity exercise training. Int J Sports Med 16:160–166PubMedCrossRef
go back to reference Timmons JA, Sundberg CJ (2006) Oligonucleotide microarray expression profiling: human skeletal muscle phenotype and aerobic exercise training. IUBMB Life 58:15–24PubMedCrossRef Timmons JA, Sundberg CJ (2006) Oligonucleotide microarray expression profiling: human skeletal muscle phenotype and aerobic exercise training. IUBMB Life 58:15–24PubMedCrossRef
go back to reference Vassilakopoulos T, Deckman G, Kebbewar M, Rallis G, Harfouche R, Hussain SN (2003) Regulation of nitric oxide production in limb and ventilatory muscles during chronic exercise training. Am J Physiol Lung Cell Mol Physiol 284:L452–L457PubMed Vassilakopoulos T, Deckman G, Kebbewar M, Rallis G, Harfouche R, Hussain SN (2003) Regulation of nitric oxide production in limb and ventilatory muscles during chronic exercise training. Am J Physiol Lung Cell Mol Physiol 284:L452–L457PubMed
go back to reference Wang Y, Newton DC, Robb GB, Kau CL, Miller TL, Cheung AH, Hall AV, VanDamme S, Wilcox JN, Marsden PA (1999) RNA diversity has profound effects on the translation of neuronal nitric oxide synthase. Proc Natl Acad Sci U S A 96:12150–12155PubMedCrossRef Wang Y, Newton DC, Robb GB, Kau CL, Miller TL, Cheung AH, Hall AV, VanDamme S, Wilcox JN, Marsden PA (1999) RNA diversity has profound effects on the translation of neuronal nitric oxide synthase. Proc Natl Acad Sci U S A 96:12150–12155PubMedCrossRef
go back to reference Wehling-Henricks M, Oltmann M, Rinaldi C, Myung KH, Tidball JG (2009) Loss of positive allosteric interactions between neuronal nitric oxide synthase and phosphofructokinase contributes to defects in glycolysis and increased fatigability in muscular dystrophy. Hum Mol Genet 18:3439–3451PubMedCrossRef Wehling-Henricks M, Oltmann M, Rinaldi C, Myung KH, Tidball JG (2009) Loss of positive allosteric interactions between neuronal nitric oxide synthase and phosphofructokinase contributes to defects in glycolysis and increased fatigability in muscular dystrophy. Hum Mol Genet 18:3439–3451PubMedCrossRef
go back to reference Williams JL, Cartland D, Hussain A, Egginton S (2006) A differential role for nitric oxide in two forms of physiological angiogenesis in mouse. J Physiol 570:445–454PubMedCrossRef Williams JL, Cartland D, Hussain A, Egginton S (2006) A differential role for nitric oxide in two forms of physiological angiogenesis in mouse. J Physiol 570:445–454PubMedCrossRef
Metadata
Title
Exercise-induced angiogenesis correlates with the up-regulated expression of neuronal nitric oxide synthase (nNOS) in human skeletal muscle
Authors
Felicitas A. M. Huber-Abel
Mélanie Gerber
Hans Hoppeler
Oliver Baum
Publication date
01-01-2012
Publisher
Springer-Verlag
Published in
European Journal of Applied Physiology / Issue 1/2012
Print ISSN: 1439-6319
Electronic ISSN: 1439-6327
DOI
https://doi.org/10.1007/s00421-011-1960-x

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