Nitric oxide production is downregulated during respiratory syncytial virus persistence by constitutive expression of arginase 1

  • Carlos Santiago-Olivares
  • Evelyn Rivera-Toledo
  • Beatriz GómezEmail author
Original Article


Viral persistence alters cellular antiviral activities. Nitric oxide (NO), a highly reactive free radical and a potent antiviral molecule, can inhibit replication of RNA and DNA viruses, but its production and effect during viral persistence are largely unknown. NO synthesis is stimulated in epithelial cells during acute infection with respiratory syncytial virus (RSV) and interferes with viral replication. In this study, we compared the levels of production of NO and expression of its regulatory enzymes, inducible nitric oxide synthase (NOS II) and arginase 1 (Arg-1), during acute and persistent RSV infection in a macrophage cell line to investigate their role in the control and maintenance of viral infection. We observed that NO and NOS II mRNA were induced at higher levels in acutely infected macrophages than in persistently infected macrophages, while the kinetics of NOS II protein expression were similar in both types of infected cultures, except that its disappearance was delayed during acute infection. Thus, NOS II was inducible and expressed at high levels during persistent infection, but production of NO was low relative to acute infection. This was not associated with a lack of enzymatic activity but instead was due to constitutive expression of the Arg-1 enzyme at the mRNA and protein levels, suggesting that arginase restricts availability of L-arginine as a substrate for NOS II to synthesize NO. This hypothesis was supported by showing that arginase enzymatic activity was inhibited in persistently RSV-infected cells by Nω-hydroxy-nor-L-arginine, increasing L-arginine availability in conditioned medium and producing increased levels of nitrites, concurrently with a significant reduction in virus genome replication, implying that Arg-1 overexpression contributes to the maintenance of the RSV genome in the host in persistent infection.



The authors thank Jorge Gaona Bernal and Juan Miranda Ríos for helpful discussions and constructive comments during the course of this work. Also, thanks to Arturo A. Wilkins Rodríguez and Laila Gutiérrez Kobeh of the Research Unit for Translational Medicine, UNAM, for technical assistance with arginase activity determinations, and Isabel Ibarra-González and Marcela Vela-Amieva of the Laboratory of Inborn Errors of Metabolism and Screening of the National Institute of Pediatrics for technical assistance in L-arginine determinations, and finally, to Ana Flisser for English editing and the facilities granted for the culmination of the paper. This paper is part of the fulfillment of the requirements for the PhD degree of CSO within the Posgrado en Ciencias Biológicas of Universidad Nacional Autónoma de México.


This research was supported by grants from the Consejo Nacional de Ciencia y Tecnología, México (Grant 179838), by the Dirección General de Asuntos del Personal Académico, Universidad Nacional Autónoma de México (Grant PAPIIT IN218916), and by the School of Medicine, UNAM.

Compliance with ethical standards

Conflict of interest

The authors declare that they have not conflict of interest.

Supplementary material

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Supplementary material 1 (PDF 5389 kb)


  1. 1.
    Vandini S, Biagi C, Lanari M (2017) Respiratory syncytial virus: the influence of serotype and genotype variability on clinical course of infection. Int J Mol Sci. Google Scholar
  2. 2.
    Piedimonte G (2015) RSV infections: State of the art. Cleve Clin J Med 82:S13–S18. CrossRefGoogle Scholar
  3. 3.
    Falsey AR, McElhaney JE, Beran J et al (2014) Respiratory syncytial virus and other respiratory viral infections in older adults with moderate to severe influenza-like illness. J Infect Dis 209:1873–1881. CrossRefGoogle Scholar
  4. 4.
    Díez-Domingo J, Pérez-Yarza EG, Melero JA et al (2014) Social, economic, and health impact of the respiratory syncytial virus: a systematic search. BMC Infect Dis 14:544. CrossRefGoogle Scholar
  5. 5.
    Rezaee F, Gibson LF, Piktel D et al (2011) Respiratory syncytial virus infection in human bone marrow stromal cells. Am J Respir Cell Mol Biol 45:277–286. CrossRefGoogle Scholar
  6. 6.
    Schwarze J, Schauer U (2004) Enhanced virulence, airway inflammation and impaired lung function induced by respiratory syncytial virus deficient in secreted G protein. Thorax 59:517–521. CrossRefGoogle Scholar
  7. 7.
    Jha A, Jarvis H, Fraser C, Openshaw PJM (2016) Respiratory syncytial virus. SARS MERS Other Viral Lung Infect. Google Scholar
  8. 8.
    Cheung MB, Sampayo-Escobar V, Green R et al (2016) Respiratory syncytial virus-infected mesenchymal stem cells regulate immunity via interferon beta and indoleamine-2,3-dioxygenase. PLoS One 11:1–20. Google Scholar
  9. 9.
    Goritzka M, Makris S, Kausar F et al (2015) Alveolar macrophage-derived type I interferons orchestrate innate immunity to RSV through recruitment of antiviral monocytes. J Exp Med 212:jem.20140825. CrossRefGoogle Scholar
  10. 10.
    Qin L, Hu CP, Feng JT, Xia Q (2011) Activation of lymphocytes induced by bronchial epithelial cells with prolonged RSV infection. PLoS One 6:1–12. Google Scholar
  11. 11.
    Tabarani CM, Bonville CA, Suryadevara M et al (2013) Novel inflammatory markers, clinical risk factors, and virus type associated with severe respiratory syncytial virus infection. Pediatr Infect Dis J 32:437–442. CrossRefGoogle Scholar
  12. 12.
    Kao Y, Piedra P, Larsen G, Colasurdo G (2001) Induction and regulation of nitric oxide synthase in airway epithelial cells by respiratory syncytial virus. Am J Respir Crit Care Med 163:532–539CrossRefGoogle Scholar
  13. 13.
    Song W, Liu G, Bosworth CA et al (2009) Respiratory syncytial virus inhibits lung epithelial Na+ channels by up-regulating inducible nitric-oxide synthase. J Biol Chem 284:7294–7306. CrossRefGoogle Scholar
  14. 14.
    Schindler H, Bogdan C (2001) NO as a signaling molecule: effects on kinases. Int Immunopharmacol 1:1443–1455. CrossRefGoogle Scholar
  15. 15.
    Hiroyuki T, Ryoh T, Masaya O et al (1999) Respiratory syncytial virus infection of human respiratory epithelial cells enhances inducible nitric oxide synthase gene expression. J Leukoc Biol 66:99–104CrossRefGoogle Scholar
  16. 16.
    Aktan F (2004) iNOS-mediated nitric oxide production and its regulation. Life Sci 75:639–653. CrossRefGoogle Scholar
  17. 17.
    Caldwell RB, Toque HA, Narayanan SP, Caldwell RW (2015) Arginase: an old enzyme with new tricks. Trends Pharmacol Sci 36:395–405. CrossRefGoogle Scholar
  18. 18.
    Rath M, Müller I, Kropf P et al (2014) Metabolism via arginase or nitric oxide synthase: two competing arginine pathways in macrophages. Front Immunol 5:1–10. CrossRefGoogle Scholar
  19. 19.
    Dzik JM (2014) Evolutionary roots of arginase expression and regulation. Front Immunol 5:1–11. CrossRefGoogle Scholar
  20. 20.
    Sarmiento RE, Tirado R, Gómez B (2002) Characteristics of a respiratory syncytial virus persistently infected macrophage-like culture. Virus Res 84:45–58. CrossRefGoogle Scholar
  21. 21.
    Rivera-toledo E, Salido-guadarrama I, Rodríguez-dorantes M et al (2017) Conditioned medium from persistently RSV-infected macrophages alters transcriptional profile and inflammatory response of non-infected macrophages. Virus Res 230:29–37. CrossRefGoogle Scholar
  22. 22.
    Rivera-Toledo E, Torres-González L, Gómez B (2015) Respiratory syncytial virus persistence in murine macrophages impairs IFN-β response but not synthesis. Viruses 7:5361–5374. CrossRefGoogle Scholar
  23. 23.
    Gaona J, Santiago-Olivares C, Ortega E, Gómez B (2014) Respiratory syncytial virus persistence in macrophages upregulates fcgamma receptors expression. Viruses 6:624–639. CrossRefGoogle Scholar
  24. 24.
    Ibarra-González I, Rodríguez-Valentín R, Lazcano-Ponce E, Vela-Amieva M (2017) Metabolic screening and metabolomics analysis in the intellectual developmental disorders Mexico study. Salud Publica Mex 59:423–428. CrossRefGoogle Scholar
  25. 25.
    Chowdhury KD, Sen G, Sarkar A, Biswas T (2011) Role of endothelial dysfunction in modulating the plasma redox homeostasis in visceral leishmaniasis. Biochim Biophys Acta Gen Subj 1810:652–665. CrossRefGoogle Scholar
  26. 26.
    Bogdan C (2001) Nitric oxide and the immune response. Nat Immunol 2:907–916. CrossRefGoogle Scholar
  27. 27.
    Majano PL (2003) Does nitric oxide play a pathogenic role in hepatitis C virus infection ? Cell Death Differentiation. Google Scholar
  28. 28.
    Kopincová J, Púzserová A, Bernátová I (2011) Biochemical aspects of nitric oxide synthase feedback regulation by nitric oxide. Interdiscip Toxicol 4:63–68. CrossRefGoogle Scholar
  29. 29.
    Wu G, Morris SM (1998) Arginine metabolism: nitric oxide and beyond. Biochem J 336:1–17. CrossRefGoogle Scholar
  30. 30.
    Kasmi KCEl, Qualls JE, Pesce JT et al (2008) Toll-like receptor-induced arginase 1 in macrophages thwarts effective immunity against intracellular pathogens. Nat Immunol 9:1399–1406. CrossRefGoogle Scholar
  31. 31.
    Stoermer K, Burrack A, Oko L et al (2012) Genetic ablation of arginase 1 in macrophages and neutrophils enhances clearance of an arthritogenic alphavirus. J Immunol 189(8):4047–4059. CrossRefGoogle Scholar
  32. 32.
    Bowen DG, Walker CM (2005) Adaptive immune responses in acute and chronic hepatitis C virus infection. Nature 436:946–952. CrossRefGoogle Scholar
  33. 33.
    Thimme R, Oldach D, Chang K-M et al (2001) Determinants of viral clearance and persistence during acute hepatitis C virus infection. J Exp Med 194:1395–1406. CrossRefGoogle Scholar
  34. 34.
    Cai W, Qin A, Guo P et al (2013) Clinical significance and functional studies of Myeloid-derived suppressor cells in chronic Hepatitis C patients. J Clin Immunol 33:798–808. CrossRefGoogle Scholar
  35. 35.
    Das A, Hoare M, Davies N et al (2008) Functional skewing of the global CD8 T cell population in chronic hepatitis B virus infection. J Exp Med 205:2111–2124. CrossRefGoogle Scholar
  36. 36.
    Sandalova E, Laccabue D, Boni C et al (2012) Increased levels of arginase in patients with acute hepatitis B suppress antiviral T cells. Gastroenterology 143:78-87.e3. CrossRefGoogle Scholar
  37. 37.
    Rodriguez PC, Ochoa AC (2008) Tolerance in cancer: mechanisms and therapeutic perspectives. Immunol Rev 222:180–191. CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Austria, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Departamento de Microbiología y Parasitología, Facultad de MedicinaUniversidad Nacional Autónoma de México, Ciudad UniversitariaMexico CityMexico
  2. 2.Posgrado en Ciencias BiológicasUnidad de Posgrado, Ciudad UniversitariaMexico CityMéxico

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