Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

HMG-CoA reductase inhibitors inhibit inducible nitric oxide synthase gene expression in macrophages

  • 39 Accesses

  • 63 Citations


The 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitors, statins, are potent inhibitors of cholesterol synthesis and have wide therapeutic use in cardiovascular diseases. Recent evidence, however, suggests that the beneficial effects of statins may extend beyond their action on serum cholesterol levels. In this study, we investigated the effects of lovastatin, pravastatin, atorvastatin and fluvastatin on macrophage formation of nitric oxide (NO) in murine RAW 264.7 cells. Stimulation of macrophages with lipopolysaccharide (LPS) and interferon-γ (IFN-γ) resulted in inducible NO synthase (iNOS) expression, which was accompanied by a large amount of NO formation. At concentrations of 0.1–30 µM, statins can inhibit stimuli-induced NO formation and iNOS induction to different extents. This inhibition occurs at the transcriptional level, and displays potency in the order of lovastatin > atorvastatin > fluvastatin >> pravastatin. We found that LPS-induced IκB kinase and nuclear factor-κB (NF-κB) activation, as well as IFN-γ-induced signal transducer and activator of transcription 1 (STAT1) phosphorylation, were reduced by lovastatin. Moreover, inhibition by lovastatin of NO production and κB activation was reversed by mevalonate, geranylgeranyl pyrophosphate and farnesyl pyrophosphate. All these results suggest that inhibition of iNOS gene expression by statins can be attributed to interference with protein isoprenylation, which mediates both NF-κB and STAT1 activation in the upstream signaling pathways for iNOS gene transcription.

This is a preview of subscription content, log in to check access.


  1. 1

    Alberts AW. Discovery, biochemistry and biology of lovastatin. Am J Cardiol 62:10J-15J;1988.

  2. 2

    Aliev G, Smith MA, Turmaine M, Neal ML, Zimina TV, Friedland RP, Perry G, Lamanna JC, Burnstock G. Atherosclerotic lesions are associated with increased immunoreactivity for inducible nitric oxide synthase and endothelin-1 in thoracic aortic intimal cells of hyperlipidemic Watanabe rabbits. Exp Mol Pathol 71:40–54;2001.

  3. 3

    Amin-Hanjani S, Stagliano NE, Yamada M, Huang PL, Liao JK, Moskowitz MA. Mevastatin, an HMG-CoA reductase inhibitor, reduces stroke damage and upregulates endothelial nitric oxide synthase in mice. Stroke 32:980–986;2001.

  4. 4

    Aviram M, Dankner G, Cogan U, Hochgraf E, Brook JG. Lovastatin inhibits low-density lipoprotein oxidation and alters its fluidity and uptake by macrophages: In vitro and in vivo studies. Metabolism 41:229–235;1992.

  5. 5

    Aviram M, Rosenblat M, Bisgaier CL, Newton RS. Atorvastatin and gemfibrozil metabolites, but not the parent drugs, are potent antioxidants against lipoprotein oxidation. Atherosclerosis 138:271–280;1998.

  6. 6

    Axel DI, Riessen R, Runge H, Viebahn R, Karsch KR. Effects of cerivastatin on human arterial smooth muscle cell proliferation and migration in transfilter cocultures. J Cardiovasc Pharmacol 35:619–629;2000.

  7. 7

    Caron E, Self A, Hall A. The GTPase Rap1 controls functional activation of macrophage integrin αMβ2 by LPS and other inflammatory mediators. Curr Biol 10:974–978;2000.

  8. 8

    Caunt CJ, Kiss-Toth E, Carlotti F, Chapman R, Owarnstrom EE. Ras controls tumor necrosis factor receptor-associated factor (TRAF)6-dependent induction of nuclear factor-κB. J Biol Chem 276:6280–6288;2001.

  9. 9

    Chen BC, Chou CF, Lin WW. Pyrimidinoceptor-mediated potentiation of inducible nitric-oxide synthase induction in J774 macrophages. J Biol Chem 273:29754–29763;1998.

  10. 10

    Chen BC, Lin WW. PKC- and ERK-dependent activation of IκB kinase by lipopolysaccharide in macrophages: Enhancement by P2Y receptor-mediated CaMK activation. Br J Pharmacol 134:1055–1065;2001.

  11. 11

    Chen CW, Chao Y, Chang YH, Hsu MJ, Lin WW. Inhibition of cytokine-induced JAK-STAT signaling pathways by an endonuclease inhibitor aurintricarboxylic acid. Br J Pharmacol 137:1011–1020;2002.

  12. 12

    Chen H, Ikeda U, Shimpo M, Ikeda M, Minota S, Shimada K. Fluvastatin upregulates inducible nitric oxide synthase expression in cytokine-stimulated vascular smooth muscle cells. Hypertension 36:923–928;2000.

  13. 13

    Diaz-Meco MT, Dominguez I, Sana L, Dent P, Lozano J, Municio MM, Berra E, Hay RT, Sturgill TW, Moscat J. PKCζ induces phosphorylation and inactivation of IκBα in vitro. EMBO J 13:2842–2848;1994.

  14. 14

    DiDonato JA, Hayakawa M, Rothwarf DM, Zandi E, Karin M. A cytokine-responsive IκB kinase that activates the transcription factor NF-κB. Nature 388:548–554;1997.

  15. 15

    Endres M, Laufs U, Huang Z, Nakamura T, Huang P, Moskowitz MA, Liao JK. Stroke protection by 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase inhibitors mediated by endothelial nitric oxide synthase. Proc Natl Acad Sci USA 95:8880–8885;1998.

  16. 16

    Finlin BS, Andres DA. Rem is a new member of the Rad- and Gem/Kir Ras-related GTP-binding protein family repressed by lipopolysaccharide stimulation. J Biol Chem 272:21982–21988;1997.

  17. 17

    Fisher JE, Rogers MJ, Halasy JM, Luckman SP, Hughes DE, Masarachia PJ, Wesolowski G, Russell RGG, Rodan GA, Reszka AA. Alendronate mechanism of action: Geranylgeraniol, an intermediate in the mevalonate pathway, prevents inhibition of osteoclast formation, bone resorption, and kinase activation in vitro. Proc Natl Acad Sci USA 96:133–138;1999.

  18. 18

    Frost JA, Swantek JL, Stippec S, Yin MJ, Gaynor R, Cobb JH. Stimulation of NFκB activity by multiple signaling pathways requires PAK1. J Biol Chem 275:19693–19699;2000.

  19. 19

    Ganne F, Vasse M, Beaudeux JL, Peynet J, Francois A, Mixhal Z, Chartier A, Tobelem G, Vannier JP, Soria J, Soria C. Cerivastatin, an inhibitor of HMG-CoA reductase, inhibits urokinase/urokinase-receptor expression and MMP-9 secretion by peripheral blood monocytes. Thromb Haemost 84:680–688;2000.

  20. 20

    Giroux LM, Davignon J, Naruszewicz M. Simvastatin inhibits the oxidation of low-density lipoproteins by activated human monocyte-derived macrophages. Biochim Biophys Acta 1165:335–338;1993.

  21. 21

    Glomset JA, Gelb MH, Farnsworth CC. Prenyl proteins in eukaryotic cells: A new type of membrane anchor. Trends Biochem Sci 15:139–142;1990.

  22. 22

    Glomset JA, Gelb MH, Farnsworth CC. Geranylgeranylated proteins. Biochem Soc Trans 20:479–484;1992.

  23. 23

    Goldstein JL, Brown MS. Regulation of the mevalonate pathway. Nature 343:425–430;1990.

  24. 24

    Gonzalez-Fernandez F, Jimenez A, Lopez-Blaya A, Velasco S, Arriero MM, Celdran A, Rico L, Farre J, Casado S, Lopez-Farre A. Cerivastatin prevents tumor necrosis factor-α-induced downregulation of endothelial nitric oxide synthase: Role of endothelial cytosolic proteins. Atherosclerosis 155:61–70;2001.

  25. 25

    Guha M, O'Connell MA, Pawlinski R, Hollis A, McGovern P, Yan SF, Stern D, Mackman N. Lipopolysaccharide activation of the MEK-ERK1/2 pathway in human monocytic cells mediates tissue factor and tumor necrosis factor α expression by inducing Elk-1 phosphorylation and Egr-1 expression. Blood 98:1429–1439;2001.

  26. 26

    Han BH. Interferon-γ and lipopolysaccharide induce mouse guanylate-binding protein 3 (mGBP3) expression in the murine macrophage cell line RAW264.7. Arch Pharm Res 22:130–136;1999.

  27. 27

    Hausding M, Witteck A, Rodriguez-Pascual F, Von Eichel-Streiber C, Forstermann U, Kleinert H. Inhibition of small G proteins of the Rho family by statins orClostridium difficile toxin B enhances cytokine-mediated induction of NO synthase II. Br J Pharmacol 131:553–561;2000.

  28. 28

    Heitmeier MR, Scarim AL, Corbett JA. Prolonged STAT1 activation is associated with interferon-γ priming for interleukin-1-induced inducible nitric-oxide synthase expression by islets of Langerhans. J Biol Chem 274:29266–29273;1999.

  29. 29

    Henwood JM, Heel RC. Lovastatin. A preliminary review of its pharmacodynamic properties and therapeutic use in hyperlipidaemia. Drugs 36:429–454;1988.

  30. 30

    Hernandez-Perera O, Perez-Sala D, Navarro-Antolin J, Sanchez-Pascuala R, Hernandez G, Diaz C, Lamas S. Effects of the 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors, atorvastatin and simvastatin, on the expression of endothelin-1 and endothelial nitric oxide synthase in vascular endothelial cells. J Clin Invest 101:2711–2719;1998.

  31. 31

    Hrboticky N, Draude G, Hapfelmeier G, Lorenz R, Weber PC. Lovastatin decreases the receptor-mediated degradation of acetylated and oxidized LDLs in human blood monocytes during the early stage of differentiation into macrophages. Arterioscler Thromb Vasc Biol 19:1267–1275;1999.

  32. 32

    Ikeda U, Shimpo M, Ikeda N, Minota S, Shimada K. Lipophilic statins augment inducible nitric oxide synthase expression in cytokine-stimulated cardiac myocytes. J Cardiovasc Pharmacol 38:69–77;2001.

  33. 33

    Izumo N, Fujita T, Nakamuta H, Moida M. Lipophilic statins can be osteogenic by promoting osteoblastic calcification in a Cbfa1- and BMP-2-independent manner. Methods Find Exp Clin Pharmacol 23:389–394;2001.

  34. 34

    Kiener PA, Davis PM, Murray JL, Youssef S, Rankin BM, Kowala M. Stimulation of inflammatory responses in vitro and in vivo by lipophilic HMG-CoA reductase inhibitors. Int Immunopharmacol 1:105–118;2001.

  35. 35

    Koh KK. Effects of statins on vascular wall: Vasomotor function, inflammation, and plaque stability. Cardiovasc Res 47:648–657;2000.

  36. 36

    Kraynack NC, Corey DA, Elmer HL, Kelley TJ. Mechanisms of NOS2 regulation by Rho GTPase signaling in airway epithelial cells. Am J Physiol Lung Cell Mol Physiol 283:L604-L611;2002.

  37. 37

    Kuhlencordt PJ, Chen J, Han F, Astern J, Huang PL. Genetic deficiency of inducible nitric oxide synthase reduces atherosclerosis and lowers plasma lipid peroxides in apolipoprotein E-knockout mice. Circulation 103:3099–3104;2001.

  38. 38

    Kumano T, Mutoh T, Nakagawa H, Kuriyama M. HMG-CoA reductase inhibitor induces a transient activation of high affinity nerve growth factor receptor, Trk, and morphological differentiation with fatal outcome in PC12 cells. Brain Res 859:169–172;2000.

  39. 39

    Kumar B, Cole WC, Prasad KN. Alpha tocopheryl succinate, retinoic acid and polar carotenoids enhanced the growth-inhibitory effect of a cholesterol-lowering drug on immortalized and transformed nerve cells in culture. J Am Coll Nutr 20:628–636;2001.

  40. 40

    Laufs U, La Fata V, Plutzky J, Liao JK. Upregulation of endothelial nitric oxide synthase by HMG CoA reductase inhibitors. Circulation 97:1129–1135;1998.

  41. 41

    Laufs U, Liao JK. Direct vascular effects of HMG-CoA reductase inhibitors. Trends Cardiovasc Med 10:143–148;2000.

  42. 42

    Lea AP, McTavish D. Atorvastatin. A review of its pharmacology and therapeutic potential in the management of hyperlipidaemias. Drugs 53:828–847;1997.

  43. 43

    Lowenstein CJ, Alley EW, Raval P, Snowman AM, Snyder SH, Russell SW, Murphy WJ. Macrophage nitric oxide synthase gene: Two upstream regions mediate induction by interferon γ and lipopolysaccharide. Proc Natl Acad Sci USA 90:9730–9734;1993.

  44. 44

    Luo SF, Wang CC, Chiu CT, Chien CS, Hsiao LD, Lin CH, Yang CM. Lipopolysaccharide enhances bradykinin-induced signal transduction via activation of Ras/Raf/MEK/MAPK in canine tracheal smooth muscle cells. Br J Pharmacol 130:1799–1808;2000.

  45. 45

    Ma PTS, Gil G, Sudhof TC, Bilheimer DW, Goldstein JL, Brown MS. Mevinolin, an inhibitor of cholesterol synthesis, induces mRNA for low density lipoprotein receptor in livers of hamsters and rabbits. Proc Natl Acad Sci USA 83:8370–8374;1986.

  46. 46

    Maltese WA. Posttranslational modification of proteins by isoprenoids in mammalian cells. FASEB J 4:3319–3328;1990.

  47. 47

    Matar P, Rozados VR, Roggero EA, Scharovsky OG. Lovastatin inhibits tumor growth and metastasis development of a rat fibrosarcoma. Cancer Biother Radiopharm 13:387–393;1998.

  48. 48

    Mitin N, Kudla AJ, Konieczny SF, Taparowsky EJ. Dfferential effects of Ras signaling through NFκB on skeletal myogenesis. Oncogene 20:1276–1286;2001.

  49. 49

    Moncada S, Palmer RMJ, Higgs EA. Nitric oxide: Physiology, pathophysiology, and pharmacology. Pharmacol Rev 43:109–142;1991.

  50. 50

    Mundy G, Garrett R, Harris S, Chan J, Chen D, Rossini G, Boyce G, Boyce B, Zhao M, Gutierrez G. Stimulation of bone formation in vitro and in rodents by statins. Science 286:1946–1949;1999.

  51. 51

    Muniyappa R, Xu R, Ram JL, Sowers JR. Inhibition of Rho protein stimulates iNOS expression in rat vascular smooth muscle cells. Am J Physiol Heart Circ Physiol 278:H1762-H1768;2000.

  52. 52

    Mutoh T, Kumano T, Nakagawa H, Kuriyama M. Role of tyrosine phosphorylation of phospholipase Cγ1 in the signaling pathway of HMG-CoA reductase inhibitor-induced cell death of L6 myoblasts. FEBS Lett 446:91–94;1999.

  53. 53

    Nagasawa K, Muraki Y, Katsuda T, Ohnishi N, Yokoyama T. Inhibitory effect of statins on fetal bovine serum-induced proliferation of rat cultured mesangial cells and correlation between their inhibitory effect and transport characteristics. J Pharm Sci 89:1594–1604;2000.

  54. 54

    Nathan C. Inducible nitric oxide synthesis: What difference does it make? J Clin Invest 100:2417–2423;1997.

  55. 55

    Olkkonen VM, Stenmark H. Role of Rab GTPases in membrane traffic. Int Rev Cytol 176:1–85;1997.

  56. 56

    Pahan K, Liu X, McKinney MJ, Wood C, Sheikh FG, Raymond JR. Expression of a dominant-negative mutant of p21ras inhibits induction of nitric oxide synthase and activation of nuclear factor-κB in primary astrocytes. J Neurochem 74:2288–2295;2000.

  57. 57

    Pahan K, Sheikh FG, Namboodiri AMS, Singh I. Lovastatin and phenylacetate inhibit the induction of nitric oxide synthase and cytokines in rat primary astrocytes, microglia, and macrophages. J Clin Invest 100:2671–2679;1997.

  58. 58

    Park WH, Lee YY, Kim ES, Seol JG, Jung CW, Lee CC, Kim BK. Lovastatin-induced inhibition of HL-60 cell proliferation via cell cycle arrest and apoptosis. Anticancer Res 19:3133–3140;1999.

  59. 59

    Perona R, Montaner S, Saniger L, Sanchez-Perez I, Brano R, Lacal JC. Activation of the nuclear factor-κB by Rho, Cdc42, and Rac-1 proteins. Genes Dev 11:463–475;1997.

  60. 60

    Raiteri M, Arnaboldi L, McGeady P, Gelb MH, Verri D, Tagliabue C, Quarato P, Ferraboschi P, Santaniello E, Paoletti R, Fumagalli R, Corsini AJ. Pharmacological control of the mevalonate pathway: Effect on arterial smooth muscle cell proliferation. J Pharmacol Exp Ther 281:1144–1153;1997.

  61. 61

    Riessen R, Axel DI, Fenchel M, Herzog UU, Rossmann H, Karsch KR. Effect of HMG-CoA reductase inhibitors on extracellular matrix expression in human vascular smooth muscle cells. Basic Res Cardiol 94:322–332;1999.

  62. 62

    Sakaeda T, Takara K, Kakumoto M, Ohmoto N, Nakamura T, Iwaki K, Tanigawara Y, Okumura K. Simvastatin and lovastatin, but not pravastatin, interact with MDR1. J Pharm Pharmacol 54:419–423;2002.

  63. 63

    Sakai M, Kobori S, Matsumura T, Biwa T, Sat Y, Takemura T, Hakamata H, Horiuchi S, Shichiri M. HMG-CoA reductase inhibitors suppress macrophage growth induced by oxidized low density lipoprotein. Atherosclerosis 133:51–59;1997.

  64. 64

    Sanlioglu S, Williams CM, Samavati L, Butler NS, Wang G, McCray PB, Ritchie TC, Hunninghake GW, Zandi E, Engelhardt JF. Lipopolysaccharide induces Rac1-dependent reactive oxygen species formation and coordinates tumor necrosis factor-α secretion through IKK regulation of NF-κB. J Biol Chem 276:30188–30198;2001.

  65. 65

    Schmidt A, Caron E, Hall A. Lipopolysaccharide-induced activation of β2-integrin function in macrophages requires IRAK kinase activity, p38 mitogen-activated protein kinase, and Rap1 GTPase. Mol Cell Biol 21:438–448;2001.

  66. 66

    Serajuddin AT, Ranadive SA, Mahone EMJ. Relative lipophilicities, solubilities, and structure-pharmacological considerations of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitors pravastatin, lovastatin, mevastatin, and simvastatin. J Pharm Sci 80:830–834;1991.

  67. 67

    Shepherd J, Cobbe SM, Ford I, Islets CG, Lorimer AR, MacFarlane PW, McKillop JH, Packard CJ. Prevention of coronary heart disease with pravastatin in men with hypercholesterolemia: West of Scotland Coronary Prevention Study Group. N Engl J Med 333:1301–1307;1995.

  68. 68

    Trushin SA, Pennington KN, Algeciras-Schimnich A, Paya CV. Protein kinase C and calcineurin synergize to activate IκB kinase and NF-κB in T lymphocytes. J Biol Chem 274:22923–22931;1999.

  69. 69

    Wagner AH, Schwabe O, Hecker M. Atorvastatin inhibition of cytokine-inducible nitric oxide synthase expression in native endothelial cells in situ. Br J Pharmacol 136:143–149;2002.

  70. 70

    Wang IK, Lin-Shiau SY, Lin JK. Induction of apoptosis by lovastatin through activation of caspase-3 and DNase II in leukaemia HL-60 cells. Pharmacol Toxicol 86:83–91;2000.

  71. 71

    Weber C, Erl W, Weber KSC, Weber PC. HMG-CoA reductase inhibitors decrease CD11b expression and CD11b-dependent adhesion of monocytes to endothelium and reduce increased adhesiveness of monocytes isolated from patients with hypercholesterolemia. J Am Coll Cardiol 30:1212–1217;1997.

  72. 72

    Wiesbauer F, Kaun C, Zorn G, Maurer G, Huber K, Wojta J. HMG-CoA reductase inhibitors affect the fibrinolytic system of human vascular cells in vitro: A comparative study using different statins. Br J Pharmacol 135:284–292;2002.

  73. 73

    Xie QW, Whisnant R, Nathan C. Promoter of the mouse gene encoding calcium-independent nitric oxide synthase confers inducibility by interferon γ and bacterial lipopolysaccharide. J Exp Med 177:1779–1784;1993.

  74. 74

    Yada T, Nakata M, Shiraishi T, Kakei M. Inhibition by simvastatin, but not pravastatin, of glucose-induced cytosolic Ca2+ signaling and insulin secretion due to blockade of L-type Ca2+ channels in rat islet β-cells. Br J Pharmacol 126:1205–1213;1999.

Download references

Author information

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Huang, K., Chen, C., Chen, J. et al. HMG-CoA reductase inhibitors inhibit inducible nitric oxide synthase gene expression in macrophages. J Biomed Sci 10, 396–405 (2003). https://doi.org/10.1007/BF02256431

Download citation

Key Words

  • Inducible nitric oxide synthase
  • Statins
  • HMG-CoA reductase
  • NF-κB
  • STAT1
  • Macrophage