Pulmonary tuberculosis (TB) is characterized by oxidative stress and lung tissue destruction by matrix metalloproteinases. The relationship between these distinct processes and the implications for TB diagnosis and clinical disease staging are poorly understood. Our work in this area has focused on the interplay between the anti-oxidant enzyme heme oxygenase-1 (HO-1) and matrix metalloproteinases (MMPs) in human and experimental TB infection. As described in last years report and now published on-line (see Scientific Advance), Bruno Andrade and our clinical colleagues in India, Brazil and the NIH demonstrated that patients with TB (but not healthy controls or individuals with latent TB infection) express either very high levels of HO-1 or of MMP-1. They also showed that production of the two enzymes is cross-regulated in human macrophages infected with Mtb in vitro. Together their studies argued that combined measurement of HO-1 and MMP-1 offers a strategy both for discrimination of active TB from other lung diseases and for clinical staging of TB patients. In this years work (supported in part by a joint US-South Africa UO-1 grant) we have extended this analysis to a larger, better characterized archived cohort in Cape Town and to a new treatment cohort being initiated as a major component of the UO-1 project.
The aim will be to both further validate our initial findings in a geographically distinct African patient group and to more definitively explore the relationship of biomarker expression with clinical disease. At the same time we have initiated new studies with Mtb experimental animal models to examine the function of HO-1 and MMP-1 in both host resistance and pathology. In collaboration with Chuck Scanga and Joanne Flynn (University of Pittsburgh) we measured the two markers in plasma of Mtb infected cynomolgus macaques and in the case of HO-1 observed a close association of enzyme expression and infection progression which was reduced upon treatment. A less consistent correlation with active infection was observed when MMP-1 expression was measured in the same monkeys, nor did combined assay of HO-1 and MMP-1 reveal any disease correlations in the limited number of animals studied so far. We also initiated experiments in the mouse model to assess the role of HO-1 in MTb induced host resistance. Interestingly, administration of an HO-1 inhibitor (SnPP) was found to induce a highly significant reduction in bacterial load in aerosol infected MTb infected mice and to enhance the effects of chemotherapy when given together with a standard TB drug regimen. This effect was observed only when the inhibitor was administered to established infections as the drug failed to block new infection. Together these experiments suggest that HO-1 might be a useful target for host-directed therapy of tuberculosis. A major ongoing project in the lab has been to investigate the role of the host immune response in chemotherapy of tuberculosis and in particular the involvement of T cell dependent immunity. Studies by Diego Costa using T cell deficient TCR alpha knock-out mice indicated that standard TB drug treatment with isoniaizid, rifampicin and pyrazinamide (HRZ) causes a similar reduction in pulmonary bacterial load whether or not T cells are present with only a minor delay in final bacillary clearance. Similarly, attempts to enhance treatment by simultaneous administration of transgenic CD4+ T cells recognizing important TB antigens so far yielded negative results. In parallel experiments, we tested the effects of adjunct administration of IL-12, a cytokine required for both acute control of TB infection and the Th1 response to the pathogen on the outcome of chemotherapy. Again, this immunotherapeutic intervention failed to significantly effect drug induced bacterial control in the lungs but in agreement with previously published work in the lab on Mycobacterium avium infection significantly reduced disseminated infection measure in spleen. These results taken together with other experiments examining the immune dependence of chemotherapy in the mouse model suggest that drug induced clearance Mtb in the lungs during the early intensive phase is not significantly affected by either the endogenous cellular immune response or by exogenous administration of cytokines, or for that matter, TB effector cells themselves.
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