title: Antigen stimulation of peripheral blood mononuclear cells from Mycobacterium bovis infected cattle yields evidence for a novel gene expression program.
abstract: BACKGROUND
  Bovine tuberculosis (BTB) caused by Mycobacterium bovis continues to cause substantial losses to global agriculture and has significant repercussions for human health. The advent of high throughput genomics has facilitated large scale gene expression analyses that present a novel opportunity for revealing the molecular mechanisms underlying mycobacterial infection. Using this approach, we have previously shown that innate immune genes in peripheral blood mononuclear cells (PBMC) from BTB-infected animals are repressed in vivo in the absence of exogenous antigen stimulation. In the present study, we hypothesized that the PBMC from BTB-infected cattle would display a distinct gene expression program resulting from exposure to M. bovis. A functional genomics approach was used to examine the immune response of BTB-infected (n = 6) and healthy control (n = 6) cattle to stimulation with bovine tuberculin (purified protein derivative - PPD-b) in vitro. PBMC were harvested before, and at 3 h and 12 h post in vitro stimulation with bovine tuberculin. Gene expression changes were catalogued within each group using a reference hybridization design and a targeted immunospecific cDNA microarray platform (BOTL-5) with 4,800 spot features representing 1,391 genes.
  
  
  RESULTS
  250 gene spot features were significantly differentially expressed in BTB-infected animals at 3 h post-stimulation contrasting with only 88 gene spot features in the non-infected control animals (P < or = 0.05). At 12 h post-stimulation, 56 and 80 gene spot features were differentially expressed in both groups respectively. The results provided evidence of a proinflammatory gene expression profile in PBMC from BTB-infected animals in response to antigen stimulation. Furthermore, a common panel of eighteen genes, including transcription factors were significantly expressed in opposite directions in both groups. Real-time quantitative reverse transcription PCR (qRT-PCR) demonstrated that many innate immune genes, including components of the TLR pathway and cytokines were differentially expressed in BTB-infected (n = 8) versus control animals (n = 8) after stimulation with bovine tuberculin.
  
  
  CONCLUSIONS
  The PBMC from BTB-infected animals exhibit different transcriptional profiles compared with PBMC from healthy control animals in response to M. bovis antigen stimulation, providing evidence of a novel gene expression program due to M. bovis exposure.
authors.affiliation: Comparative Immunology Group, School of Biochemistry and Immunology, Trinity College Dublin, Dublin 2, Ireland. kieran.meade@tcd.ie
authors.forename: Kieran G
authors.initials: KG
authors.lastname: Meade
authors.forename: Eamonn
authors.initials: E
authors.lastname: Gormley
authors.forename: Cliona
authors.initials: C
authors.lastname: O'Farrelly
authors.forename: Stephen D
authors.initials: SD
authors.lastname: Park
authors.forename: Eamon
authors.initials: E
authors.lastname: Costello
authors.forename: Joseph
authors.initials: J
authors.lastname: Keane
authors.forename: Yingdong
authors.initials: Y
authors.lastname: Zhao
authors.forename: David E
authors.initials: DE
authors.lastname: MacHugh
mesh_terms: D000818:Animals; D002417:Cattle; D015870:Gene Expression / Q000187:drug effects / Q000235:genetics; D020869:Gene Expression Profiling; D054884:Host-Pathogen Interactions; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000378:metabolism / Q000382:microbiology; D009163:Mycobacterium bovis / Q000502:physiology*; D020411:Oligonucleotide Array Sequence Analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014373:Tuberculin / Q000494:pharmacology; D014380:Tuberculosis, Bovine / Q000235:genetics* / Q000382:microbiology*
chemical_list: D014373:Tuberculin