Abstract
The aim of the study was to investigate the activity of human neutrophil peptide (HNP)-1 to kill Mycobacterium tuberculosis H37Rv in vitro and ex vivo in the murine macrophage cell line J744A.1 on the basis of colony forming units. Macromolecular biosynthesis was studied by monitoring the incorporation of radioactive precursors into different macromolecules. The binding and localization studies were carried out with radioiodinated HNP-1 whereas the cytotoxicity of HNP-1 to macrophages was determined by trypan blue exclusion assay. A concentration dependent inhibition in the growth of M. tuberculosis H37Rv was observed in the presence of HNP-1. The minimum inhibitory concentration and median inhibitory concentration of HNP-1 were found to be 2.5 microg x mL(-1) and 0.8 microg x mL(-1). Treatment of both in vitro grown and phagocytosed mycobacterial cells with HNP-1 resulted in generalized inhibition in the macromolecular biosynthesis with maximum inhibition in deoxyribonucleic acid and lipid biosynthesis. HNP-1 exhibited equilibrium binding with respect to time and two-thirds of bound radioactivity was shown to be present inside the macrophages. Approximately 50% and 98% killing of intracellular mycobacteria was observed after 3 days of treatment with 5 microg x mL(-1) and 40 microg x mL(-1) of HNP-1, respectively. HNP-1 exhibited low cytotoxicity towards the macrophage cell line at the bactericidal concentration to mycobacteria. From the results of this study, it is concluded that human neutrophil peptide-1 possesses potent bactericidal activity against virulent mycobacteria in vitro as well as mycobacteria replicating within macrophages.
