Mechanism of nitric oxide-dependent killing of Mycobacterium bovis BCG in human alveolar macrophages

Abstract

We demonstrated that products of the L-arginine-dependent pathway of human alveolar macrophages (AM) effectively kill the Mycobacterium bovis bacillus Calmette-Guérin (BCG) in vitro. The formation of products was triggered by inoculation with BCG itself. Many reports have shown that activated rodent AM could produce an amount of nitric oxide (NO) sufficient to suppress the growth of mycobacteria. However, there have been no definitive results as to whether human AM might have the NO-dependent mechanism for the killing of mycobacteria. Therefore, we have undertaken some experiments to answer this question. Immunofluorescence assays showed an increased production of inducible nitric oxide synthase (iNOS) and peroxynitrite in BCGinoculated AM from patients with pulmonary fibrosis. Reverse transcriptase-PCR also revealed the higher expression of iNOS-coding mRNA. Colony assays demonstrated that these human AM effectively killed BCG in their cytoplasm. However, treatment of AM with N G-monomethyl-L-arginine monoacetate resulted in markedly reduced killing activity. These results clearly show that BCG-induced NO and its reactive product with the oxygen radical peroxynitrite could play an important role in the killing of BCG in human AM.

Induction of nitric oxide release from the human alveolar epithelial cell line A549: an in vitro correlate of innate immune response to Mycobacterium tuberculosis

“…The annealing temperature for iNOS was 54°and for IL-8 and b-actin it was 65°and 66°, respectively. 35,36 The RT-PCR products were run and visualized on 1AE5% agarose gel with ethidium bromide along with the 100 base pair marker (MBI fermentas; Genetix Biotech Asia Pvt Ltd, New Delhi, India).…”

Antigen‐specific lymphocytes enhance nitric oxide production in Mycobacterium bovis BCG‐infected bovine macrophages

“…3 However, it has been reported that M. bovis BCG is killed by a NO-dependent mechanism in alveolar macrophages isolated from patients with pulmonary ®brosis. 15 In the bovine system, iNOS activity and NO production has been measured in alveolar macrophages as well as bone marrow, and peripheral blood monocyte-derived macrophages. 12,16,17 Pre-treatment of alveolar macrophages with IFN-c has been shown to limit intracellular growth of M. bovis BCG.…”

Kinetic studies of bioactive products nitric oxide and 8-iso-PGF2αinBurkholderia pseudomalleiinfected human macrophages, and their role in the intracellular survival of these organisms

“…Nitric oxide synthase is known to play an essential role in controlling intracellular persistence and multiplication of B. pseudomallei via the production of nitric oxide [6]. Researchers have shown that suppression of iNOS expression contributed to the progression of infection [15] and enhanced expression of iNOS was demonstrated to be useful in killing intracellular pathogens like Salmonella [16] and Mycobacterium [17].…”