Furthermore, to investigate whether sMTL-13 is expressed during active infection in vivo, we have performed immuno-staining in pleural biopsies from ATB patients. Figure 2C shows positive staining for sMTL-13 in tissue granulomas from ATB patients. In contrast, as expected no staining was observed in biopsies from negative IgG1 isotype control (Fig. 2D), skin biopsies from M. leprae-infected patients (Fig. 2E), or in tissue granulomas associated with fungal infection (Fig. 2F and data not shown). A hallmark
of mycobacterial infection is the generation of a strong immune response against secreted antigens. A number of antigens secreted by Mtb have been proposed to function as virulence factors and may influence the clinical outcome of TB 11, 12, 29. We therefore investigated whether sMTL-13 is recognized by TB patients during active disease. First, we measured recall Selleckchem ACP-196 responses by means of IFN-γ production of PBMC following exposure to sMTL-13 in vitro. As demonstrated in Fig. 3A, sMTL-13-stimulated PBMC from active TB patients (n=11) display increased production of IFN-γ when compared with BCG-vaccinated purified protein
derivative (PPD)-negative control subjects (n=6). In addition, we have performed ELISA in serum samples from 34 diseased individuals as well as 38 control subjects. As shown in Fig. 3B, recently diagnosed TB patients (either naive of treatment or up to 15 days undergoing early chemotherapy; ATB group) presented high titers of anti-sMTL-13 total IgG Ab. Importantly, www.selleckchem.com/products/Rapamycin.html anti-sMTL-13 IgG titers rapidly decreased during the first months (1–2) of treatment and reached background levels as compared with those from endemic or non-endemic subjects. Moreover, anti-sMTL-13 IgG Ab titers remained at background levels following successful anti-TB chemotherapy (6 months). Furthermore,
receiver operating characteristic (ROC) curves analysis at the optimal cutoff point revealed that anti-sMTL-13 IgG titers display high specificity (90%) as well as sensitivity (93%) for TB diagnosis (Fig. 3C). There was no significant difference between the areas for ESAT-6 (AUC=0.956 (AUC, area under the curve), CI 95%: 0.865–0.985) and sMTL-13 (AUC=0.943, CI 95%: 0.855–0.981). Together, these mTOR inhibitor data suggest that TB patients display adaptive immune responses against sMTL-13 during active disease and anti-sMTL-13 Ab are decreased following therapeutic control of Mtb in vivo. Proteins actively secreted during the in vitro early growth phase of Mtb have been the subject of intensive investigation for their ability to elicit immune responses either in vitro or in vivo30–34. In support of this concept, mice immunized with live but not dead bacilli can induce a protective T-cell response, reinforcing the notion that secreted proteins are among the antigens encountered and presented by the host immune system 35.