Macrophage migration inhibitory factor (MIF) controls cytokine release during respiratory syncytial virus infection in macrophages

  • Gabriela F. de Souza
  • Stéfanie P. Muraro
  • Leonardo D. Santos
  • Ana Paula T. Monteiro
  • Amanda G. da Silva
  • Ana Paula D. de Souza
  • Renato T. Stein
  • Patrícia T. Bozza
  • Bárbara N. PortoEmail author
Original Research Paper


Objective and design

Respiratory syncytial virus (RSV) is the major cause of infection in children up to 2 years old and reinfection is very common among patients. Tissue damage in the lung caused by RSV leads to an immune response and infected cells activate multiple signaling pathways and massive production of inflammatory mediators like macrophage migration inhibitory factor (MIF), a pro-inflammatory cytokine. Therefore, we sought to investigate the role of MIF during RSV infection in macrophages.


We evaluated MIF expression in BALB/c mice-derived macrophages stimulated with different concentrations of RSV by Western blot and real-time PCR. Additionally, different inhibitors of signaling pathways and ROS were used to evaluate their importance for MIF expression. Furthermore, we used a specific MIF inhibitor, ISO-1, to evaluate the role of MIF in viral clearance and in RSV-induced TNF-α, MCP-1 and IL-10 release from macrophages.


We showed that RSV induces MIF expression dependently of ROS, 5-LOX, COX and PI3K activation. Moreover, viral replication is necessary for RSV-triggered MIF expression. Differently, p38 MAPK in only partially needed for RSV-induced MIF expression. In addition, MIF is important for the release of TNF-α, MCP-1 and IL-10 triggered by RSV in macrophages.


In conclusion, we demonstrate that MIF is expressed during RSV infection and controls the release of pro-inflammatory cytokines from macrophages in an in vitro model.


Respiratory syncytial virus MIF Macrophage migration inhibitory factor Macrophages Cytokines Signaling pathways 



This study was supported by CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico—Grant nr. 456282/2014-9 to Bárbara N. Porto and Grant nr. 481366/2011-3 to Renato T. Stein). Gabriela F. de Souza was recipient of a scholarship from CNPq and Stéfanie P. Muraro was recipient of a scholarship from CAPES (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, Brazil).

Author contributions

BNP and GFS conceived and designed the study. GFS, SPM, APTM, LDS and AGS performed the experiments. BNP, GFS and SPM performed the statistical analysis and interpreted the data. BNP, GFS and SPM wrote the manuscript. BNP, APDS, RTS and PTB critically revised the draft. All authors contributed to the manuscript revision and approved the submitted version.

Compliance with ethical standards

Conflict of interest

The authors declare that there is no conflict of interest regarding the publication of this paper.


  1. 1.
    Borchers AT, Chang C, Gershwin ME, et al. Respiratory syncytial virus—a comprehensive review. Clin Rev Allergy Immunol. 2013;45(3):331–79.Google Scholar
  2. 2.
    Johansson C. Respiratory syncytial virus infection: an innate perspective. F100Res. 2016;5:2898.Google Scholar
  3. 3.
    Blanken MO, Rovers MM, Molenaar JM, et al. Respiratory syncytial virus and recurrent wheeze in healthy preterm infants. N Engl J Med. 2013;368(19):1791–9.Google Scholar
  4. 4.
    Glezen WP, Taber LH, Frank AL, et al. Risk of primary infection and reinfection with respiratory syncytial virus. Am J Dis Child. 1986;140(6):543–6.Google Scholar
  5. 5.
    Selwyn BJ, Coordinated Data Group of BOSTID Researchers. The epidemiology of acute respiratory tract infection in young children: comparison of findings from several developing countries. Rev Infect Dis. 1990;12(Suppl 8):S870–88.Google Scholar
  6. 6.
    Garenne M, Ronsmans C, Campbell H. The magnitude of mortality from acute respiratory infections in children under 5 years in developing countries. World Health Stat Q. 1992;45(2–3):180–91.Google Scholar
  7. 7.
    Griffiths C, Drews SJ, Marchant DJ. Respiratory syncytial virus: infection, detection, and new options for prevention and treatment. Clin Microbiol Rev. 2017;30(1):277–319.Google Scholar
  8. 8.
    Lloyd CM, Marsland BJ. Lung homeostasis: influence of age, microbes, and the immune system. Immunity. 2017;46(4):549–61.Google Scholar
  9. 9.
    Takeuchi O, Akira S. Innate immunity to virus infection. Immunol Rev. 2009;227(1):75–86.Google Scholar
  10. 10.
    Kim TH, Lee HK. Innate immune recognition of respiratory syncytial virus infection. BMB Rep. 2014;47(4):184–91.Google Scholar
  11. 11.
    Bueno SM, González PA, Pacheco R, et al. Host immunity during RSV pathogenesis. Int Immunopharmacol. 2008;8(10):1320–9.Google Scholar
  12. 12.
    Noah TL, Becker S. Respiratory syncytial virus-induced cytokine production by a human bronchial epithelial cell line. Am J Physiol. 1993;265(5 Pt1):L472–8.Google Scholar
  13. 13.
    Xie J, Yang L, Tian L, et al. Macrophage migration inhibitor factor upregulates MCP-1 expression in an autocrine manner in hepatocytes during acute mouse liver injury. Sci Rep. 2016;8(6):27665.Google Scholar
  14. 14.
    Sheeran P, Jafri H, Carubelli C, et al. Elevated cytokine concentrations in the nasopharyngeal and tracheal secretions of children with respiratory syncytial virus disease. Pediatr Infect Dis J. 1999;18(2):115–22.Google Scholar
  15. 15.
    Kurt-Jones EA, Popova L, Kwinn L, et al. Pattern recognition receptors TLR4 and CD14 mediate response to respiratory syncytial virus. Nat Immunol. 2000;1(5):398–401.Google Scholar
  16. 16.
    Nguyen TH, Maltby S, Simpson JL, et al. TNF-α and macrophages are critical for respiratory syncytial virus-induced exacerbations in a mouse model of allergic airways disease. J Immunol. 2016;196(9):3547–58.Google Scholar
  17. 17.
    Calandra T, Roger T. Macrophage migration inhibitory factor: a regulator of innate immunity. Nat Rev Immunol. 2003;3(10):791–800.Google Scholar
  18. 18.
    Baugh JA, Bucala R. Macrophage migration inhibitory factor. Crit Care Med. 2002;30(1 Supp):S27–35.Google Scholar
  19. 19.
    Bernhagen J, Mitchell RA, Calandra T, et al. Purification, bioactivity, and secondary structure analysis of mouse and human macrophage migration inhibitory factor (MIF). Biochemistry. 1994;33(47):14144–55.Google Scholar
  20. 20.
    Mitchell RA, Metz CN, Peng T, et al. Sustained mitogen-activated protein kinase (MAPK) and cytoplasmic phospholipase A2 activation by macrophage migration inhibitory factor (MIF). Regulatory role in cell proliferation and glucocorticoid action. J Biol Chem. 1999;274(25):18100–6.Google Scholar
  21. 21.
    Leng L, Metz CN, Fang Y, et al. MIF signal transduction initiated by binding to CD74. J Exp Med. 2003;197(11):1467–76.Google Scholar
  22. 22.
    Chuang YC, Su WH, Lei HY, et al. Macrophage migration inhibitory factor induces autophagy via reactive oxygen species generation. PLoS One. 2012;7(5):e37613.Google Scholar
  23. 23.
    Magalhães ES, Mourao-Sa DS, Vieira-de-Abreu A, et al. Macrophage migration inhibitory factor is essential for allergic asthma but not for Th2 differentiation. Eur J Immunol. 2007;37(4):1097–106.Google Scholar
  24. 24.
    Paiva CN, Arras RH, Magalhães ES, et al. Migration inhibitory factor (MIF) released by macrophages upon recognition of immune complexes is critical to inflammation in Arthus reaction. J Leukoc Biol. 2009;85(5):855–61.Google Scholar
  25. 25.
    de Souza HS, Tortori CA, Lintomen L, et al. Macrophage migration inhibitory factor promotes eosinophil accumulation and tissue remodeling in eosinophilic esophagitis. Mucosal Immunol. 2015;8(5):1154–65.Google Scholar
  26. 26.
    Bozza M, Satoskar AR, Lin G, et al. Targeted disruption of migration inhibitory factor gene reveals its critical role in sepsis. J Exp Med. 1999;189(2):341–6.Google Scholar
  27. 27.
    Calandra T, Echtenacher B, Roy DL, et al. Protection from septic shock by neutralization of macrophage migration inhibitory factor. Nat Med. 2000;6(2):164–70.Google Scholar
  28. 28.
    Suzuki T, Ogata A, Tashiro K, et al. Japanese encephalitis virus up-regulates expression of macrophage migration inhibitory factor (MIF) mRNA in the mouse brain. Biochim Biophys Acta. 2000;1517(1):100–6.Google Scholar
  29. 29.
    Satoskar AR, Bozza M, Sosa MR, et al. Migration-inhibitory factor gene-deficient mice are susceptible to cutaneous Leishmania major infection. Infect Immun. 2001;69(2):906–11.Google Scholar
  30. 30.
    Bacher M, Eickmann M, Schrader J, et al. Human cytomegalovirus-mediated induction of MIF in fibroblasts. Virology. 2002;299(1):32–7.Google Scholar
  31. 31.
    Magalhães ES, Paiva CN, Souza HS, et al. Macrophage migration inhibitory factor is critical to interleukin-5-driven eosinophilopoiesis and tissue eosinophilia triggered by Schistosoma mansoni infection. FASEB J. 2009;23(4):1262–71.Google Scholar
  32. 32.
    Cavalcanti MG, Mesquita JS, Madi K, et al. MIF participates in Toxoplasma gondii-induced pathology following oral infection. PLoS One. 2011;6(9):e25259.Google Scholar
  33. 33.
    Hou XQ, Gao YW, Yang ST, et al. Role of macrophage migration inhibitory factor in influenza H5N1 virus pneumonia. Acta Virol. 2009;53(4):225–31.Google Scholar
  34. 34.
    Arndt U, Wennemuth G, Barth P, et al. Release of macrophage migration inhibitory factor and CXCL8/interleukin-8 from lung epithelial cells rendered necrotic by influenza A virus infection. J Virol. 2002;76(18):9298–306.Google Scholar
  35. 35.
    Akira S, Uematsu S, Takeuchi O. Pathogen recognition and innate immunity. Cell. 2006;124(4):783–801.Google Scholar
  36. 36.
    Okabe Y, Medzhitov R. Tissue biology perspective on macrophages. Nat Immunol. 2016;17(1):9–17.Google Scholar
  37. 37.
    Calandra T, Bernhagen J, Mitchell RA, et al. The macrophage is an important and previously unrecognized source of macrophage migration inhibitory factor. J Exp Med. 1994;179(6):1895–902.Google Scholar
  38. 38.
    Kim JH, Lee J, Bae SJ, et al. NADPH oxidase 4 is required for the generation of macrophage migration inhibitory factor and host defense against Toxoplasma gondii infection. Sci Rep. 2017;7(1):6361.Google Scholar
  39. 39.
    Behera AK, Kumar M, Matsuse H, et al. Respiratory syncytial virus induces the expression of 5-lipoxygenase and endothelin-1 in bronchial epithelial cells. Biochem Biophys Res Commun. 1998;251(3):704–9.Google Scholar
  40. 40.
    Richardson JY, Ottolini MG, Pletneva L, et al. Respiratory syncytial virus (RSV) infection induces cyclooxygenase 2: a potential target for RSV therapy. J Immunol. 2005;174(7):4356–64.Google Scholar
  41. 41.
    Dave M, Islam ABMMK, Jensen RV, et al. Proteomic analysis shows constitutive secretion of MIF and p53-associated activity of COX-2(−/−) lung fibroblasts. Genom Proteom Bioinform. 2017;15(6):339–51.Google Scholar
  42. 42.
    Graham BS. Biological challenges and technological opportunities for respiratory syncytial virus vaccine development. Immunol Rev. 2011;239(1):149–66.Google Scholar
  43. 43.
    Munir S, Hillyer P, Le Nouën C, et al. Respiratory syncytial virus interferon antagonist NS1 protein suppresses and skews the human T lymphocyte response. PLoS Pathog. 2011;7(4):e1001336.Google Scholar
  44. 44.
    de Jong YP, Abadia-Molina AC, Satoskar AR, et al. Development of chronic colitis is dependent on the cytokine MIF. Nat Immunol. 2001;2(11):1061–6.Google Scholar
  45. 45.
    Santos LL, Morand EF. The role of macrophage migration inhibitory factor in the inflammatory immune response and rheumatoid arthritis. Wien Med Wochenschr. 2006;156(1–2):11–8.Google Scholar
  46. 46.
    Kimura K, Nagaki M, Nishihira J, Satake S, Kuwata K, Moriwaki H. Role of macrophage migration inhibitory factor in hepatitis B virus-specific cytotoxic-T-lymphocyte-induced liver injury. Clin Vaccine Immunol. 2006;13(3):415–9.Google Scholar
  47. 47.
    Assunção-Miranda I, Amaral FA, Bozza FA, et al. Contribution of macrophage migration inhibitory factor to the pathogenesis of dengue virus infection. FASEB J. 2010;24(1):218–28.Google Scholar
  48. 48.
    Assunção-Miranda I, Bozza MT, Da Poian AT. Pro-inflammatory response resulting from sindbis virus infection of human macrophages: implications for the pathogenesis of viral arthritis. J Med Virol. 2010;82(1):164–74.Google Scholar
  49. 49.
    Regis EG, Barreto-de-Souza V, Morgado MG, et al. Elevated levels of macrophage migration inhibitory factor (MIF) in the plasma of HIV-1-infected patients and in HIV-1-infected cell cultures: a relevant role on viral replication. Virology. 2010;399(1):31–8.Google Scholar
  50. 50.
    Delaloye J, De Bruin IJ, Darling KE, et al. Increased macrophage migration inhibitory factor (MIF) plasma levels in acute HIV-1 infection. Cytokine. 2012;60(2):338–40.Google Scholar
  51. 51.
    Huang J, Canadien V, Lam GY, et al. Activation of antibacterial autophagy by NADPH oxidases. Proc Natl Acad Sci USA. 2009;106(15):6226–31.Google Scholar
  52. 52.
    Bae YS, Oh H, Rhee SG, et al. Regulation of reactive oxygen species generation in cell signaling. Mol Cells. 2011;32(6):491–509.Google Scholar
  53. 53.
    Park HS, Jung HY, Park EY, et al. Cutting edge: direct interaction of TLR4 with NAD(P)H oxidase 4 isozyme is essential for lipopolysaccharide-induced production of reactive oxygen species and activation of NF-kappa B. J Immunol. 2004;173(6):3589–93.Google Scholar
  54. 54.
    Sun Y, Wang Y, Li JH, et al. Macrophage migration inhibitory factor counter-regulates dexamethasone-induced annexin 1 expression and influences the release of eicosanoids in murine macrophages. Immunology. 2013;140(2):250–8.Google Scholar
  55. 55.
    Wang F, Wu H, Xu S, et al. Macrophage migration inhibitory factor activates cyclooxygenase 2-prostaglandin E2 in cultured spinal microglia. Neurosci Res. 2011;71(3):210–8.Google Scholar
  56. 56.
    Carey MA, Bradbury JA, Seubert JM, et al. Contrasting effects of cyclooxygenase-1 (COX-1) and COX-2 deficiency on the host response to influenza A viral infection. J Immunol. 2005;175(10):6878–84.Google Scholar
  57. 57.
    Lindemans CA, Coffer PJ, Schellens IM, et al. Respiratory syncytial virus inhibits granulocyte apoptosis through a phosphatidylinositol 3-kinase and NF-kappaB-dependent mechanism. J Immunol. 2006;176(9):5529–37.Google Scholar
  58. 58.
    Funchal GA, Jaeger N, Czepielewski RS, et al. Respiratory syncytial virus fusion protein promotes TLR-4-dependent neutrophil extracellular trap formation by human neutrophils. PLoS One. 2015;10(4):e0124082.Google Scholar
  59. 59.
    Boukhvalova MS, Prince GA, Soroush L, et al. The TLR4 agonist, monophosphoryl lipid A, attenuates the cytokine storm associated with respiratory syncytial virus vaccine-enhanced disease. Vaccine. 2006;24(23):5027–35.Google Scholar
  60. 60.
    Dou Y, Zhao Y, Zhang ZY, et al. Respiratory syncytial virus infection induces higher Toll-like receptor-3 expression and TNF-α production than human metapneumovirus infection. PLoS One. 2013;8(9):e73488.Google Scholar
  61. 61.
    Olszewska-Pazdrak B, Casola A, Saito T, et al. Cell-specific expression of RANTES, MCP-1, and MIP-1alpha by lower airway epithelial cells and eosinophils infected with respiratory syncytial virus. J Virol. 1998;72(6):4756–64.Google Scholar
  62. 62.
    Bacher M, Metz CN, Calandra T, et al. An essential regulatory role for macrophage migration inhibitory factor in T-cell activation. Proc Natl Acad Sci USA. 1996;93(15):7849–54.Google Scholar
  63. 63.
    Abe R, Peng T, Sailors J, et al. Regulation of the CTL response by macrophage migration inhibitory factor. J Immunol. 2001;166(2):747–53.Google Scholar
  64. 64.
    Gore Y, Starlets D, Maharshak N, et al. Macrophage migration inhibitory factor induces B cell survival by activation of a CD74–CD44 receptor complex. J Biol Chem. 2008;283(5):2784–92.Google Scholar
  65. 65.
    Chuang TY, Chang HT, Chung KP, et al. High levels of serum macrophage migration inhibitory factor and interleukin 10 are associated with a rapidly fatal outcome in patients with severe sepsis. Int J Infect Dis. 2014;20:13–7.Google Scholar

Copyright information

© Springer Nature Switzerland AG 2019

Authors and Affiliations

  • Gabriela F. de Souza
    • 1
  • Stéfanie P. Muraro
    • 1
  • Leonardo D. Santos
    • 1
  • Ana Paula T. Monteiro
    • 2
  • Amanda G. da Silva
    • 1
  • Ana Paula D. de Souza
    • 1
  • Renato T. Stein
    • 3
  • Patrícia T. Bozza
    • 2
  • Bárbara N. Porto
    • 1
    • 4
    Email author
  1. 1.Laboratory of Clinical and Experimental Immunology, Infant Center, School of MedicinePontifical Catholic University of Rio Grande do Sul (PUCRS)Porto AlegreBrazil
  2. 2.Laboratory of ImmunopharmacologyOswaldo Cruz Institute, Oswaldo Cruz Foundation (FIOCRUZ)Rio de JaneiroBrazil
  3. 3.Laboratory of Pediatric Respirology, Infant Center, School of MedicinePontifical Catholic University of Rio Grande do Sul (PUCRS)Porto AlegreBrazil
  4. 4.Program in Translational MedicineThe Hospital for Sick ChildrenTorontoCanada

Personalised recommendations