CD11b+Gr-1low cells that accumulate in M.leprae-induced granulomas of the footpad skin of nude mice have the characteristics of monocytic-myeloid-derived suppressor cells.

CD11b+Gr-1low cells that are increased in the lungs of a Mycobacterium (M) tuberculosis-infection mouse model have the characteristics of monocytic (M)-myeloid-derived suppressor cells (MDSCs) and harbor M.tuberculosis. Interestingly, a high number of M-MDSCs have also been observed in skin lesions of patients with lepromatous leprosy. We hypothesized that CD11b+Gr-1low cells might be involved in the pathogenesis of leprosy, as they are in tuberculosis. In the current study, we investigated the issue of whether CD11b+Gr-1low cells accumulate in Mycobacterium (M) leprae-induced granulomas of the footpad skin of nude mice. Our results show that CD11b+Gr-1low cells began to accumulate in the 7-month-old M.leprae-induced granulomas and were replaced by other leukocytes, including CD11b+Gr-1high over time during M.leprae infections. CD11b + Gr-1low cells expressed the surface markers of M-MDSC, Ly6Chigh and Ly6Glow. In addition, CD11b+Gr-1low cells have the nuclei of a mononuclear cell type and expressed higher levels of arginase 1 (Arg1) and inducible NO synthetase (iNOS). Furthermore, they showed a higher infection rate by M.leprae. Taken together, our results indicate that the inoculation with M.leprae induced an accumulation of CD11b + Gr-1low at a relatively early stage, 7-month-old M.leprae-induced granulomas, and that CD11b+Gr-1low have the characteristics of M-MDSC and may act as a reservoir for M.leprae.

Lipoarabinomannan from Mycobacterium indicus pranii shows immunostimulatory activity and induces autophagy in macrophages.

Mycobacterium indicus pranii (MIP) known for its immunotherapeutic potential against leprosy and tuberculosis is undergoing various clinical trials and also simultaneously being studied in animal models to get insight into the mechanistic details contributing to its protective efficacy as a vaccine candidate. Studies have shown potential immunomodulatory properties of MIP, the most significant being the ability to induce strong Th1 type of response, enhanced expression of pro-inflammatory cytokines, activation of APCs and lymphocytes, elicitation of M.tb specific poly-functional T cells. All of these form crucial components of host-immune response during M.tb infection. Also, MIP was found to be potent inducer of autophagy in macrophages which resulted in enhanced clearance of M.tb from MIP and M.tb co-infected cells. Hence, we further examined the component/s of MIP responsible for autophagy induction. Interestingly, we found that MIP lipids and DNA were able to induce autophagy but not the protein fraction. LAM being one of the crucial components of mycobacterial cell-wall lipids and possessing the ability of immunomodulation; we isolated LAM from MIP and did a comparative study with M.tb-LAM. Stimulation with MIP-LAM resulted in significantly high secretion of pro-inflammatory cytokines and displayed high autophagy inducing potential in macrophages as compared to M.tb-LAM. Treatment with MIP-LAM enhanced the co-localization of M.tb within the phago-lysosomes and increased the clearance of M.tb from the infected macrophages. This study describes LAM to be a crucial component of MIP which has significant contribution to its immunotherapeutic efficacy against TB.

Early secreted antigen ESAT-6 of Mycobacterium Tuberculosis promotes apoptosis of macrophages via targeting the microRNA155-SOCS1 interaction.

BACKGROUND: The early secreted antigenic target 6-kDa protein (ESAT-6) of Mycobacterium tuberculosis (Mtb) not only acts as a key player for virulence but also exhibits a strong immunotherapeutic potential against Mtb. However, little is known about the molecular basis for its potential in immunotherapy. The present study was designed to unravel the role of miRNA-155 in ESAT-6-mediated enhancement of host immunity and apoptosis in macrophages. METHODS: Lentivirus-mediated miR-155 sponge and miR-155 and SOCS1 overexpression vectors were developed in macrophages. TLR2- or p65-specific siRNA knockdown was employed to silence TLR2 or p65. Quantitative polymerase chain reaction and western blotting analyses were performed to determine mRNA and protein expression levels, respectively. Macrophage apoptosis was analyzed by flow cytometry. RESULTS: ESAT-6 significantly increased miR-155 expression, which was dependent on TLR2/NF-κB activation in macrophages. Induced expression of miRNA-155 was required for the ESAT-6-mediated protective immune response and macrophage apoptosis. ESAT-6 promoted macrophage apoptosis by targeting the miR-155-SOCS1 pathway. The differential expression levels of TLR2, BIC, and SOCS1 were involved in regulating the immune response in human peripheral blood mononuclear cells of patients with active tuberculosis (TB) and latent TB (LTB). CONCLUSION: ESAT-6 promotes apoptosis of macrophages via targeting the miRNA155-SOCS1 interaction.

Isolation of a distinct Mycobacterium tuberculosis mannose-capped lipoarabinomannan isoform responsible for recognition by CD1b-restricted T cells.

Mannose-capped lipoarabinomannan (ManLAM) is a complex lipoglycan abundantly present in the Mycobacterium tuberculosis cell envelope. Many biological properties have been ascribed to ManLAM, from directly interacting with the host and participating in the intracellular survival of M. tuberculosis, to triggering innate and adaptive immune responses, including the activation of CD1b-restricted T cells. Due to its structural complexity, ManLAM is considered a heterogeneous population of molecules which may explain its different biological properties. The presence of various modifications such as fatty acids, succinates, lactates, phosphoinositides and methylthioxylose in ManLAM have proven to correlate directly with its biological activity and may potentially be involved in the interactions between CD1b and the T cell population. To further delineate the specific ManLAM epitopes involved in CD1b-restricted T cell recognition, and their potential roles in mediating immune responses in M. tuberculosis infection, we established a method to resolve ManLAM into eight different isoforms based on their different isoelectric values. Our results show that a ManLAM isoform with an isoelectric value of 5.8 was the most potent in stimulating the production of interferon-γ in different CD1b-restricted T-cell lines. Compositional analyses of these isoforms of ManLAM revealed a direct relationship between the overall charge of the ManLAM molecule and its capacity to be presented to T cells via the CD1 compartment.

Aspects of genetic susceptibility to human infectious diseases.

Host genetic factors play a major role in determining differential susceptibility to major infectious diseases of humans, such as malaria, HIV/AIDS, tuberculosis, and invasive pneumococcal disease. Progress in identifying the relevant genetic loci has come from a variety of approaches. Most convincing associations have been identified by case-control studies assessing biologically plausible candidate genes. All six of the genes that have a major effect on infectious disease susceptibility in humans have been identified in this way. However, recently genome-wide linkage analysis of affected sibling pairs has identified susceptibility loci for chronic infections such as leprosy and chronic hepatitis B virus persistence. Other approaches used successfully have included assessment in humans of the homologues of susceptibility genes mapped and identified in murine models. However, the great majority of susceptibility loci remain to be identified and the advent of large-scale genome-wide association scans offers a new approach to defining many of these.

The importance of Toll-like receptor 2 polymorphisms in severe infections.

Toll-like receptor 2 (TLR2) is a member of the TLR family, which plays a central role in the innate immune response to a wide variety of microorganisms. Animal studies have shown that TLR2-knockout mice are more susceptible to septicemia due to Staphylococcus aureus and Listeria monocytogenes, meningitis due to Streptococcus pneumoniae, and infection with Mycobacterium tuberculosis, suggesting that functional TLR2 polymorphisms may impair host response to a certain spectrum of microbial pathogens. In humans, 2 polymorphisms in the exon part of TLR2, which attenuate receptor signaling, enhance the risk of acute severe infections, tuberculosis, and leprosy. Because gram-positive bacteria have became the first cause of severe infections, including septic shock, knowledge of the role that alteration or lack of TLR2 function plays in the pathogenesis of infectious diseases could contribute to the design of new therapeutic strategies, including prevention, pharmacological intervention, and vaccine development.

Cortisol metabolism, cortisol sensitivity and the pathogenesis of leprosy reactions.

The concentration of cortisol in a tissue is regulated by a reversible enzyme 'shuttle' that can deactivate cortisol by converting it to cortisone, or activate cortisone by converting it to cortisol. The activity of this shuttle, and the direction in which it operates, is regulated by numerous factors including cytokines. This results in large swings in the effective cortisol concentration in sites of inflammation at different phases of an inflammatory response. Thus changes in local cortisol concentration can be largely independent of circulating cortisol levels. The relevant shuttle enzymes are present in skin, blood vessels and nervous tissue, and inhibition of the enzymes in skin enhances the local anti-inflammatory effect of cortisol. We therefore suggest that changes in the activity or direction of action of the shuttle in leprosy lesions may predispose to reactions, requiring exogenous steroid supplements to regain control of the inflammation.

Evidence for a highly conserved, immunoreactive lipid in Mycobacterium tuberculosis.

A survey of the fractionated lipids of Mycobacterium tuberculosis H37Rv was conducted using rabbit antiserum raised to homologous and heterologous mycobacteria. One minor, highly apolar lipid was resolved by Florisil column chromatography, which reacted preferentially to anti-M. tuberculosis H37Rv rabbit antibodies. Other chromatographic properties, i.e., thin-layer chromatographic mobility and staining properties, suggested an analog of the phenolic glycolipid of Mycobacterium leprae. Preliminary results in the application of the glycolipid to tuberculous populations in northeast Thailand suggest a usefulness in screening for tuberculosis.

Detection of mycolic acid in leprous tissue.

A method is described for the extraction of mycolic acid from lepromatous leprosy tissues and from tuberculous tissues and its identification by thin layer chromatography. Verification of the mycolic acid extracted and separated by thin layer chromatography was accomplished by infrared spectroscopy. Although the presence of mycolic acid in tissues is not, in itself, diagnostic for leprosy, its demonstration in biopsy tissues might be a useful additional qualitative test if used in conjunction with clinical evidence of the disease.