Mycobacterium bovis is the causative agent of most cases of bovine tuberculosis. The identification of bTB biomarkers in specific stages of the disease will contribute to a better understanding of the immunopathology associated with tuberculosis and will enable their use in disease diagnosis and prognosis. The aim of this study was to evaluate the gene expression profile induced after specific stimulation of bovine peripheral blood mononuclear cells from cattle infected with M. bovis using the Affymetrix？ GeneChip？ Bovine Genome Array. A total of 172 genes showed differential expression profile that was statistically significant with log2-fold change >2.5 and <？2.5. Twenty-four out of these genes were upregulated and 148 were downregulated in bovine peripheral blood mononuclear cells of M. bovis-infected cattle. The highest differentially-expressed genes were related to immune and inflammatory responses, apoptosis, endocytosis, cellular trafficking and genes encoding proteins involved in cellular matrix degradation. Microarray results were confirmed in another group of infected cattle by RT-qPCR for the CD14, IL-1R, THBS1, MMP9 and FYVE genes. This study confirms previous findings that have shown that M. bovis infection in cattle results in the downregulation of immune response-related genes. Moreover, it validates the use of microarray platforms in combination with RT-qPCR to identify biomarkers of bovine tuberculosis. In addition, we propose CD14, IL-1R, THBS1, MMP9 and FYVE as potential biomarkers of bovine tuberculosis.
References
[1]
Biet F, Boschiroli ML, Thorel MF, Guilloteau LA (2005) Zoonotic aspects of Mycobacterium bovis and Mycobacterium avium-intracellulare complex (MAC). Vet Res 36: 411–436.
[2]
Corner LA (1994) Post mortem diagnosis of Mycobacterium bovis infection in cattle. Vet Microbiol 40: 53–63.
[3]
Neill SD, Pollock JM, Bryson DB, Hanna J (1994) Pathogenesis of Mycobacterium bovis infection in cattle. Vet Microbiol 40: 41–52.
[4]
Whipple DL, Bolin CA, Miller JM (1996) Distribution of lesions in cattle infected with Mycobacterium bovis. J Vet Diagn Invest 8: 351–354.
[5]
McDonough KA, Kress Y, Bloom BR (1993) Pathogenesis of tuberculosis: interaction of Mycobacterium tuberculosis with macrophages. Infect Immun 61: 2763–2773.
[6]
Buddle BM (2010) Tuberculosis vaccines for cattle: the way forward. Expert Rev Vaccines 9: 1121–1124.
[7]
Gentleman R, Morgan M, Huber W (2008) Gene Set Enrichment Analysis In: Bioconductor Case Studies; R. Gentleman MMaWHE, editor.
[8]
Rhodes SG, Buddle BM, Hewinson RG, Vordermeier HM (2000) Bovine tuberculosis: immune responses in the peripheral blood and at the site of active disease. Immunology 99: 195–202.
[9]
Meade KG, Gormley E, Doyle MB, Fitzsimons T, O’Farrelly C, et al. (2007) Innate gene repression associated with Mycobacterium bovis infection in cattle: toward a gene signature of disease. BMC Genomics 8: 400.
[10]
Killick KE, Browne JA, Park SD, Magee DA, Martin I, et al. (2011) Genome-wide transcriptional profiling of peripheral blood leukocytes from cattle infected with Mycobacterium bovis reveals suppression of host immune genes. BMC Genomics 12: 611.
[11]
Van Lint P, Libert C (2007) Chemokine and cytokine processing by matrix metalloproteinases and its effect on leukocyte migration and inflammation. J Leukoc Biol 82: 1375–1381.
[12]
Matsuura E, Umehara F, Hashiguchi T, Fujimoto N, Okada Y, et al. (2000) Marked increase of matrix metalloproteinase 9 in cerebrospinal fluid of patients with fungal or tuberculous meningoencephalitis. J Neurol Sci 173: 45–52.
[13]
Hoheisel G, Sack U, Hui DS, Huse K, Chan KS, et al. (2001) Occurrence of matrix metalloproteinases and tissue inhibitors of metalloproteinases in tuberculous pleuritis. Tuberculosis (Edinb) 81: 203–209.
[14]
Price NM, Gilman RH, Uddin J, Recavarren S, Friedland JS (2003) Unopposed matrix metalloproteinase-9 expression in human tuberculous granuloma and the role of TNF-alpha-dependent monocyte networks. J Immunol 171: 5579–5586.
[15]
Taylor JL, Hattle JM, Dreitz SA, Troudt JM, Izzo LS, et al. (2006) Role for matrix metalloproteinase 9 in granuloma formation during pulmonary Mycobacterium tuberculosis infection. Infect Immun 74: 6135–6144.
[16]
Volkman HE, Pozos TC, Zheng J, Davis JM, Rawls JF, et al. (2010) Tuberculous granuloma induction via interaction of a bacterial secreted protein with host epithelium. Science 327: 466–469.
[17]
Bowdish DM, Sakamoto K, Kim MJ, Kroos M, Mukhopadhyay S, et al. (2009) MARCO, TLR2, and CD14 are required for macrophage cytokine responses to mycobacterial trehalose dimycolate and Mycobacterium tuberculosis. PLoS Pathog 5: e1000474.
[18]
Smit van Dixhoorn MG, Munir R, Sussman G, Stad R, de Haan M, et al. (2008) Gene expression profiling of suppressor mechanisms in tuberculosis. Mol Immunol 45: 1573–1586.
[19]
Simonsen A, Lippe R, Christoforidis S, Gaullier JM, Brech A, et al. (1998) EEA1 links PI(3)K function to Rab5 regulation of endosome fusion. Nature 394: 494–498.
[20]
Leevers SJ, Vanhaesebroeck B, Waterfield MD (1999) Signalling through phosphoinositide 3-kinases: the lipids take centre stage. Curr Opin Cell Biol 11: 219–225.
[21]
Magee DA, Taraktsoglou M, Killick KE, Nalpas NC, Browne JA, et al. (2012) Global gene expression and systems biology analysis of bovine monocyte-derived macrophages in response to in vitro challenge with Mycobacterium bovis. PLoS ONE 7: e32034.
[22]
Aranday-Cortes E, Hogarth PJ, Kaveh DA, Whelan AO, Villarreal-Ramos B, et al. (2012) Transcriptional profiling of disease-induced host responses in bovine tuberculosis and the identification of potential diagnostic biomarkers. PLoS ONE 7: e30626.
[23]
Berry MP, Graham CM, McNab FW, Xu Z, Bloch SA, et al. (2010) An interferon-inducible neutrophil-driven blood transcriptional signature in human tuberculosis. Nature 466: 973–977.
[24]
Maertzdorf J, Ota M, Repsilber D, Mollenkopf HJ, Weiner J, et al. (2011) Functional correlations of pathogenesis-driven gene expression signatures in tuberculosis. PLoS One 6: e26938.
[25]
Schroder K, Hertzog PJ, Ravasi T, Hume DA (2004) Interferon-gamma: an overview of signals, mechanisms and functions. J Leukoc Biol 75: 163–189.
[26]
Wedlock DN, Denis M, Painter GF, Ainge GD, Vordermeier HM, et al. (2008) Enhanced protection against bovine tuberculosis after coadministration of Mycobacterium bovis BCG with a Mycobacterial protein vaccine-adjuvant combination but not after coadministration of adjuvant alone. Clin Vaccine Immunol 15: 765–772.
[27]
Brettschneider J, Collin F, Bolstad BM, Speed TP (2008) Quality Assessment for Short Oligonucleotide Microarray Data. Technometrics 50: 241–264.
[28]
Gautier L, Cope L, Bolstad BM, Irizarry RA (2004) affy–analysis of Affymetrix GeneChip data at the probe level. Bioinformatics 20: 307–315.
[29]
Smyth GK (2005) Limma?: Linear Models for Microarray Data. 397–420 p.
[30]
Blanco FC, Schierloh P, Bianco MV, Caimi K, Meikle V, et al. (2009) Study of the immunological profile towards Mycobacterium bovis antigens in naturally infected cattle. Microbiol Immunol 53: 460–467.
[31]
Pfaffl MW, Horgan GW, Dempfle L (2002) Relative expression software tool (REST) for group-wise comparison and statistical analysis of relative expression results in real-time PCR. Nucleic Acids Res 30: e36.
