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 previous 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. In particular, our collaborative studies in India, Brazil and the NIH clinical center identified HO-1 as a biomarker that distinguishes patients with active TB disease from those with latent or successfully treated Mtb infection. In the past year we have re-focussed our work on human TB to a newly established collaboration with colleagues at the University of Cape Town (funded in part by a joint US-South Africa UO-1 grant). An important component of this project is to extend our analysis of host inflammatory markers to a larger, better characterized treatment cohort. Because of delays in obtaining approval for our clinical protocol, Bruno Andrade (now participating as a collaborator from his new lab in Brazil) teamed up with Neesha Rockwood (an MD, Ph.D student in Cape Town) and performed an extensive analysis of over 30 inflammation associated biomarkers in archived serum and plasma samples collected prior to and 2 and 5-6 months following anti-TB treatment in a South African cohort consisting of 133 TB and HIV/ TB patients. Levels of HO-1, in common with 29 other markers, were found to decrease following chemotherapy but HO-1 (in contrast to CRP and ferritin) did not on its own predict successful treatment outcome. The data also revealed that HO-1 levels discriminate between HIV/TB patients with low (<100) vs high (>100) CD4+ T cell counts. These preliminary associations will be re-evaluated in the more extensive UO-1 sponsored post-treatment study that will be initiated on a new cohort of patients during the next few months and which will include more detailed clinical parameters. HO-1 expression is upregulated in both experimental and human Mycobacterium tuberculosis (Mtb) infection and as noted above is a biomarker of active disease in patients. Whether the enzyme plays a protective versus pathogenic role in tuberculosis has been the subject of debate. In experiments described initially in last years report, we addressed this controversy, by administering tin protoporphyrin IX (SnPPIX), a well-characterized HO-1 enzymatic inhibitor, to mice during acute Mtb infection. These SnPPIX-treated animals displayed a substantial reduction in pulmonary bacterial loads comparable to that achieved following conventional antibiotic therapy. Moreover, when administered adjunctively with anti-mycobacterial drugs the HO-1 inhibitor markedly enhanced and accelerated pathogen clearance. In experiments performed during the present report period we determined that both the pulmonary induction of HO-1 expression and the efficacy of SnPPIX treatment in reducing bacterial burden are dependent on the presence of host T lymphocytes. Although Mtb expresses its own heme-cleaving homolog, we found that SnPPIX fails to inhibit its enzymatic activity or significantly restrict bacterial growth in liquid culture. Together, these findings (submitted for publication) reveal mammalian HO-1 as a target for host directed mono-and adjunctive therapy of Mtb infection and identify the immune response as a critical regulator of this function. 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. Previous studies by Diego Costa using T cell deficient TCR alpha knock-out mice infected with low numbers ( 100 cfu) of Mtb bacilli 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. However in follow-up experiments performed this year he repeated these experiments using higher infectious doses of bacteria and at 10.000 cfu/mouse observed that HRZ could no longer effectively bacterial loads in T cell deficient mice while at a dose of 1000 cfu only a partial effect of chemotherapy was observed in the latter animals. These new findings reveal an unappreciated role for T cells in TB chemotherapy at high bacterial loads.
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