Modified Vaccinia virus Ankara but not vaccinia virus induces chemokine expression in cells of the monocyte/macrophage lineage
© Lehmann et al.; licensee BioMed Central. 2015
Received: 6 October 2014
Accepted: 29 January 2015
Published: 12 February 2015
The orthopoxvirus strain Modified Vaccinia virus Ankara (MVA) rapidly induces innate immune responses. Previously, we demonstrated that CCL2 and CCR1 are important players in MVA induced recruitment of leukocytes to the lung. Alveolar macrophages are sentinel cells in the lung, which are likely amongst the first cells of the immune system to encounter and respond to virus during respiratory infection. Therefore we examined the potential of the murine alveolar macrophage MH-S cell line as a model to study chemokine expression during infection with MVA and vaccinia virus (VACV) strain Western Reserve (WR).
MVA but not VACV infected MH-S cells increased the expression of the CXCR2 acting chemokine CXCL2. MH-S cells constitutively produced CCL2 and CCR1 acting chemokines CCL3, CCL5 and CCL9. Consequently, supernatants of mock treated and virus infected MH-S cells induced chemotaxis of murine promyelocyte MPRO cells and human monocytic THP-1 cells at the same level. However, supernatants of MVA infected MH-S cells significantly increased chemotaxis of the CCR2 deficient human monocytic cell line U-937. Chemotaxis of all three cell types was inhibited by J 113863, a CCR1 antagonist. Additionally, we show that MVA but not VACV WR infection of THP-1 cells induces expression of C-C motif and C-X-C motif chemokines and generates a chemotactic activity for monocytes, which was J 113863 sensitive.
These results extend our previous findings, demonstrating that MVA but not VACV WR induces chemokine production in alveolar macrophages and monocytes, which can induce recruitment of monocytes in a CCR1 dependent manner.
Highly attenuated, replication deficient vaccinia strains such as Modified Vaccinia virus Ankara (MVA) are increasingly used as viral vectors for the development of new vaccines. Intranasal inoculation is an attractive route of vaccination for delivery of MVA to mucosal tissue in the respiratory tract, which was found to be both safe and immunogenic in a phase I clinical trial . A distinguishing feature of MVA is its ability to activate a robust innate immune response [2-4]. We recently showed that MVA triggers the production of chemokines in primary lung fibroblasts and bone marrow derived macrophages (BMDM) which induce chemotaxis of neutrophils .
Alveolar macrophages (AM) are permanent residents of the lung which are capable of exerting both pro- and anti-inflammatory immune responses. Due to their presence on the luminal surface of the respiratory system AM are likely amongst the first cells to encounter virus after respiratory infection. Indeed, it has been shown that AM help to control VACV infection, and their depletion exacerbates the host inflammatory response to viral infection, which is probably due to the increased viral burden . However little is known about the early immune responses that is elicited in AM upon encounter with VACV or highly attenuated strains such as MVA. Activation of AM is tightly regulated by the local microenvironment, cell-cell interactions and soluble mediators that inhibit activation of pro-inflammatory signalling pathways, and removal of these signals leads to spontaneous activation of AM . Consequently, our attempts to investigate cytokine expression in AM ex-vivo were largely unsuccessful due primarily to strong cytokine production in non-infected cells and rapid cell death of infected cells after isolation by bronchoalveolar lavage (data not shown). Therefore we tested the immortalized murine alveolar macrophage MH-S cell line  as a model system for MVA induced chemokine expression.
Recently, we showed that CCR1 plays an important role in the recruitment of CD11b+Ly6Chi inflammatory monocytes into the lung . Monocytes express the chemokine receptor CCR2 , and CCL2, a ligand for this receptor, has been shown to be important for MVA induced monocyte migration in vitro and in vivo . However, monocytes are a heterogeneous population, and some subsets do not express CCR2 . Moreover, CCR1 expression is up-regulated on differentiating monocytes, leading to increased sensitivity to CCR1 ligands whilst simultaneously decreasing sensitivity to CCR2 ligands . Consequently, we asked whether CCR1 also plays a role in mediating MVA triggered monocyte chemotaxis.
To avoid the masking effect of CCL2 when investigating MVA for its potential to induce a CCR1 dependent chemotactic activity in cells of the monocyte/macrophage lineage we took advantage of the human monocytic cell line U-937, which, under our cell culture conditions, did not express CCR2 in contrast to human monocytic THP-1 cells (Figure 3A and B) . Firstly, we tested whether supernatants from MH-S cells are also capable of inducing chemotaxis of THP-1 cells. This was possible since human chemokine receptors CCR1 and CCR2 can also be activated by the relevant murine ligands [18-20]. As with MPRO cells, supernatants from mock and virus infected MH-S cells increased chemotaxis of THP-1 at a similar level, which was decreased when THP-1 cells were pre-incubated with J 113863 (Figure 3D, left panel). Supernatants from MVA infected MH-S cells increased the chemotaxis of U-937 cells compared to supernatants from mock infected and VACV WR infected MH-S cells. Importantly, this chemotaxis of U-937 cells was significantly reduced when U-937 cells were pre-incubated with J 113863 (Figure 3D, right panel). Of note, J 113863 also acts on human CCR3 but this receptor was not expressed in either cell line (Figure 3B). Interestingly, despite the constitutive expression of CCR1 ligands by MH-S cells, levels of chemotaxis of U-937 towards culture supernatants of mock-infected cells were only slightly increased as compared to medium control (Figure 3D). This potentially indicates that other mechanisms induced by MVA may be required for optimal induction of cell migration.
In summary, we demonstrated that MH-S cells can be used as model system to study MVA infection in alveolar macrophages but due to the high constitutive production of C-C motif chemokines only limited conclusions can be drawn from functional studies where the involvement of these proteins is relevant. Nevertheless, the results presented here confirm our recent findings in primary murine lung fibroblasts and bone marrow derived macrophages showing additionally that MVA, but not VACV WR, induces chemokine expression in cells of the monocyte/macrophage lineage that is capable of inducing chemotaxis of neutrophils and monocytes.
This study was supported by the German Center for Infection Research (DZIF; TTU 07.803). MPRO cells were kindly provided by Dr. Adelheid Cerwenka (Deutsches Krebsforschungszentrum) and MH-S cells were kindly provided by Stephanie Konrad (Medical School Hannover).
- Satti I, Meyer J, Harris SA, Thomas ZR, Griffiths K, Antrobus RD, et al. Safety and immunogenicity of a candidate tuberculosis vaccine MVA85A delivered by aerosol in BCG-vaccinated healthy adults: a phase 1, double-blind, randomised controlled trial. Lancet Infect Dis. 2014;14:939–46.View ArticlePubMed CentralPubMedGoogle Scholar
- Royo S, Sainz Jr B, Hernandez-Jimenez E, Reyburn H, Lopez-Collazo E, Guerra S. Differential induction of apoptosis, interferon signaling, and phagocytosis in macrophages infected with a panel of attenuated and nonattenuated poxviruses. J Virol. 2014;88:5511–23.View ArticlePubMed CentralPubMedGoogle Scholar
- Dai P, Wang W, Cao H, Avogadri F, Dai L, Drexler I, et al. Modified vaccinia virus Ankara triggers type I IFN production in murine conventional dendritic cells via a cGAS/STING-mediated cytosolic DNA-sensing pathway. PLoS Pathog. 2014;10:e1003989.View ArticlePubMed CentralPubMedGoogle Scholar
- Price PJ, Torres-Dominguez LE, Brandmuller C, Sutter G, Lehmann MH. Modified Vaccinia virus Ankara: innate immune activation and induction of cellular signalling. Vaccine. 2013;31:4231–4.View ArticlePubMedGoogle Scholar
- Price PJ, Luckow B, Torres-Dominguez LE, Brandmuller C, Zorn J, Kirschning CJ, et al. Chemokine (C-C Motif) Receptor 1 Is Required for Efficient Recruitment of Neutrophils during Respiratory Infection with Modified Vaccinia Virus Ankara. J Virol. 2014;88:10840–50.View ArticlePubMed CentralPubMedGoogle Scholar
- Rivera R, Hutchens M, Luker KE, Sonstein J, Curtis JL, Luker GD. Murine alveolar macrophages limit replication of vaccinia virus. Virology. 2007;363:48–58.View ArticlePubMed CentralPubMedGoogle Scholar
- Snelgrove RJ, Goulding J, Didierlaurent AM, Lyonga D, Vekaria S, Edwards L, et al. A critical function for CD200 in lung immune homeostasis and the severity of influenza infection. Nat Immunol. 2008;9:1074–83.View ArticlePubMedGoogle Scholar
- Mbawuike IN, Herscowitz HB. MH-S, a murine alveolar macrophage cell line: morphological, cytochemical, and functional characteristics. J Leukoc Biol. 1989;46:119–27.PubMedGoogle Scholar
- Staib C, Lowel M, Erfle V, Sutter G. Improved host range selection for recombinant modified vaccinia virus Ankara. Biotechniques. 2003;34:694–6. 698, 700.PubMedGoogle Scholar
- Proost P, De Wolf-Peeters C, Conings R, Opdenakker G, Billiau A, Van Damme J. Identification of a novel granulocyte chemotactic protein (GCP-2) from human tumor cells. In vitro and in vivo comparison with natural forms of GRO, IP-10, and IL-8. J Immunol. 1993;150:1000–10.PubMedGoogle Scholar
- Mack M, Cihak J, Simonis C, Luckow B, Proudfoot AE, Plachy J, et al. Expression and characterization of the chemokine receptors CCR2 and CCR5 in mice. J Immunol. 2001;166:4697–704.View ArticlePubMedGoogle Scholar
- Lehmann MH, Kastenmuller W, Kandemir JD, Brandt F, Suezer Y, Sutter G. Modified vaccinia virus ankara triggers chemotaxis of monocytes and early respiratory immigration of leukocytes by induction of CCL2 expression. J Virol. 2009;83:2540–52.View ArticlePubMed CentralPubMedGoogle Scholar
- Gordon S, Taylor PR. Monocyte and macrophage heterogeneity. Nat Rev Immunol. 2005;5:953–64.View ArticlePubMedGoogle Scholar
- Kaufmann A, Salentin R, Gemsa D, Sprenger H. Increase of CCR1 and CCR5 expression and enhanced functional response to MIP-1 alpha during differentiation of human monocytes to macrophages. J Leukoc Biol. 2001;69:248–52.PubMedGoogle Scholar
- Meng A, Zhang X, Shi Y. Role of p38 MAPK and STAT3 in lipopolysaccharide-stimulated mouse alveolar macrophages. Exp Ther Med. 2014;8:1772–6.PubMed CentralPubMedGoogle Scholar
- Berkman N, John M, Roesems G, Jose PJ, Barnes PJ, Chung KF. Inhibition of macrophage inflammatory protein-1 alpha expression by IL-10. Differential sensitivities in human blood monocytes and alveolar macrophages. J Immunol. 1995;155:4412–8.PubMedGoogle Scholar
- Lehmann MH, Masanetz S, Kramer S, Erfle V. HIV-1 Nef upregulates CCL2/MCP-1 expression in astrocytes in a myristoylation- and calmodulin-dependent manner. J Cell Sci. 2006;119:4520–30.View ArticlePubMedGoogle Scholar
- Gladue RP, Cole SH, Roach ML, Tylaska LA, Nelson RT, Shepard RM, et al. The human specific CCR1 antagonist CP-481,715 inhibits cell infiltration and inflammatory responses in human CCR1 transgenic mice. J Immunol. 2006;176:3141–8.View ArticlePubMedGoogle Scholar
- Sarafi MN, Garcia-Zepeda EA, MacLean JA, Charo IF, Luster AD. Murine monocyte chemoattractant protein (MCP)-5: a novel CC chemokine that is a structural and functional homologue of human MCP-1. J Exp Med. 1997;185:99–109.View ArticlePubMed CentralPubMedGoogle Scholar
- Schall TJ, Simpson NJ, Mak JY. Molecular cloning and expression of the murine RANTES cytokine: structural and functional conservation between mouse and man. Eur J Immunol. 1992;22:1477–81.View ArticlePubMedGoogle Scholar
- Price PJ, Banki Z, Scheideler A, Stoiber H, Verschoor A, Sutter G, et al. Complement Component C5 Recruits Neutrophils in the Absence of C3 during Respiratory Infection with Modified Vaccinia Virus Ankara. J Immunol. 2015;194:1164–8.View ArticlePubMedGoogle Scholar
- Proost P, Menten P, Struyf S, Schutyser E, De Meester I, Van Damme J. Cleavage by CD26/dipeptidyl peptidase IV converts the chemokine LD78beta into a most efficient monocyte attractant and CCR1 agonist. Blood. 2000;96:1674–80.PubMedGoogle Scholar
- De Buck M, Gouwy M, Proost P, Struyf S, Van Damme J. Identification and characterization of MIP-1alpha/CCL3 isoform 2 from bovine serum as a potent monocyte/dendritic cell chemoattractant. Biochem Pharmacol. 2013;85:789–97.View ArticlePubMedGoogle Scholar
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.