Nitric oxide modulates interleukin-1beta and tumour necrosis factor-alpha synthesis, and disease regression by alveolar macrophages in pulmonary tuberculosis

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Abstract
Pretreatment with nitric oxide synthase (NOS) inhibitors profoundly increases mortality, bacterial burden and pathological tissue damage in mice infected with Mycobacterium tuberculosis. Nitric oxide (NO) production is enhanced in alveolar macrophages (AM) of tuberculosis (TB) patients. Interleukin (IL)-1beta and tumour necrosis factor (TNF)-alpha released from AM are involved in the immune response to mycobacterial infection. The aim of the present study was to examine whether NO is implicated in IL-1beta and TNF-alpha synthesis by AM and related to the resolution of disease activity in TB patients. Purified AM were retrieved by bronchoalveolar lavage from TB patients and normal subjects, and cultured in the presence or absence of a NO inhibitor, NG-monomethyl-L-arginine (L-NMMA). The release of IL-1beta and TNF-alpha, and their mRNA expression were determined by enzyme-linked immunosorbent assay (ELISA) and northern analysis, respectively. The level of nitrite released into the culture medium was determined. The rate of disease regression was evaluated by serial chest radiography. The release of nitrite, IL-1beta and TNF-alpha was much greater from AM of TB patients than normal subjects. NG-monomethyl-L-arginine inhibited the production of nitrite as well as IL-1beta and TNF-alpha in TB patients. The mRNA expression for IL-1beta and TNF-alpha was upregulated in TB patients and was depressed by L-NMMA. Immunocytochemistry using a monoclonal antibody against nuclear factor-kappaB (NF-kappaB) subunit p65 showed NF-kappaB was highly expressed and translocated to the nuclei of AM in TB patients, and was inhibited by L-NMMA. An inhibition of NF-kappaB by pyrrolidine dithiocarbamate attenuated IL-1beta and TNF-alpha synthesis. More generation of NO from cultured AM increased the disease regression in TB patients. We conclude that the enhanced NO generation by AM of TB patients may play an autoregulatory role in amplifying the synthesis of pro-inflammatory cytokines, probably through the activation of NF-kappaB. Nitric oxide may also play an important role in resistance to M. tuberculosis infection.