Abstract
Mycobacterium avium subsp. paratuberculosis (MAP) is the causative pathogen of chronic granulomatous enteropathy (Johne's disease) in animals, and has been focused on its association with various autoimmune diseases in humans, including Crohn's disease. The discovery of novel mycobacterial antigens and exploring their role in host immunity can contribute to the advancement of effective defense strategies including vaccines and diagnostic tools. In a preliminary study, we identified cellular extract proteins of MAP that strongly react with the blood of patients with Crohn's disease. In particular, MAP1981c, a putative nucleic acid-binding protein, showed high expression levels and strong reactivity to IgG and IgM in the sera of patients. Here, we investigated the immunological features of MAP1981c and focused on its interaction with dendritic cells (DCs), confirming its immunomodulatory ability. MAP1981c was shown to recognize Toll-like receptor (TLR) 4, and induce DC maturation and activation by increasing the expression of co-stimulatory (CD80 and CD86) and MHC class I/II molecules and the secretion of pro-inflammatory cytokines (IL-6, IL-1β, and TNF-α) in DCs. This DC activation by MAP1981c was mediated by downstream signaling of TLR4 via MyD88- and TRIF-, MAP kinase-, and NF-κB-dependent signaling pathways. In addition, MAP1981c-treated DCs activated naïve T cells and induced the differentiation of CD4+ and CD8+ T cells to express T-bet, IFN-γ, and/or IL-2, but not GATA-3 and IL-4, thus indicating that MAP1981c contributes to Th1-type immune responses both in vitro and in vivo. Taken together, these results suggest that MAP1981c is a novel immunocompetent antigen that induces DC maturation and a Th1-biased response upon DC activation, suggesting that MAP1981c can be an effective vaccine and diagnostic target.
Highlights
Mycobacterium avium subsp. paratuberculosis (MAP) causes Johne’s disease, which is a form of chronic gastroenteritis in ruminants (Motiwala et al, 2006)
The predicted molecular mass of MAP1981c is ∼26.95 kDa. We confirmed this prediction by expressing recombinant MAP1981c for 24 h and the culture supernatants were analyzed for the amounts of TNF-α, IL-6, IL-1β, L-12p70, and IL-10 by using quantitative ELISA. (B) Dot plots of intracellular IL-12p70 and IL-10 in CD11c+ dendritic cells (DCs)
The induction of cytokine production by MAP1981c treatment was significantly reduced in MyD88 K/O, and Toll/IL-1R domain-containing adaptor inducing IFNβ (TRIF) K/O-derived DCs compared with that in WT-DCs (Figure 6C). These results indicate that toll-like receptor 4 (TLR4)/MyD88/TRIFmediated signals are key players in MAP1981c-induced DC maturation
Summary
Mycobacterium avium subsp. paratuberculosis (MAP) causes Johne’s disease, which is a form of chronic gastroenteritis in ruminants (Motiwala et al, 2006). Paratuberculosis (MAP) causes Johne’s disease, which is a form of chronic gastroenteritis in ruminants (Motiwala et al, 2006). This disease is difficult to eradicate once it occurs in herds because of the shedding of MAP through feces and milk and the propagation of the infection during the latency period (Harris and Barletta, 2001). The association of this pathogen with human diseases has been found by isolating MAP or detecting antibodies specific to MAP from the tissue samples of patients (Naser et al, 2000a,b, 2004; Sechi et al, 2005; D’amore et al, 2010; Dow and Ellingson, 2010; Cossu et al, 2011). Even though it has been suggested that MAP is a possible causative agent of Crohn’s disease and/or other autoimmune diseases, this hypothesis has not been fully validated yet, mainly because of the slow growth and the difficulty in culturing this pathogen (Chamberlin et al, 2001)
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