Virulence of Mycobacterium tuberculosis affects interleukin-8, monocyte chemoattractant protein-1 and interleukin-10 production by human mononuclear phagocytes.

Microbial virulence and cytokine-mediated immune responses to Mycobacterium tuberculosis infection are important determinants of the pathogenesis of human tuberculosis. To determine the interrelationship between mycobacterial virulence and cytokine induction, human monocytes and monocyte-derived macrophages were infected with attenuated (H37Ra) and virulent (H37Rv and CH306) strains of M. tuberculosis and the amount of proinflammatory [interleukin (IL)-8 and monocyte chemoattractant protein (MCP)- 1] and inhibitory (IL- 10) cytokines was measured in the culture supernatants by enzyme-linked immunosorbent assay (ELISA). Infection with live bacilli induced de novo synthesis of IL-8, MCP-1 and IL-10, since cytokine release was abolished when cells were preincubated with the protein synthesis inhibitor cycloheximide. A differential production of antiinflammatory and inhibitory cytokines was observed. The amount of IL-8 and MCP-1 release was inversely related to strain virulence, the attenuated H37Ra strain being more prone than virulent strains to induce secretion of chemokines. In contrast, virulent strains induced greater amounts of the inhibitory cytokine IL-10. Efficient upregulation of IL-10 synthesis, but not of chemokines, required infection of cells with live bacilli, since heat killing of organisms or challenge with soluble mycobacterial products completely abrogated the effect. Moreover, cells infected with virulent strains produced IL-10 even at a very low bacillus-to-cell ratio and secreted IL-10 continuously during the 96 h that followed infection. The results suggest that the degree of virulence affects host cell responses to M. tuberculosis infection. Continued production of IL-10 may be one of the means by which M. tuberculosis downregulates acute local inflammatory reactions, favoring the development of tuberculosis.

Diazepam-binding inhibitor-derived peptides induce intracellular calcium changes and modulate human neutrophil function.

We studied the effects of two diazepam-binding inhibitor (DBI)-derived peptides, triakontatetraneuropeptide (DBI 17-50, TTN) and eiksoneuropeptide (DBI 51-70, ENP), on cytosolic free Ca2+ concentrations ([Ca2+]i), chemotaxis, superoxide anion (O2-) generation, and phagocytosis in human neutrophils. Both TTN and ENP induced a rapid and transient rise of [Ca2+]i. The effect of TTN depended on the presence of extracellular Ca2+, whereas the effect of ENP also persisted after extracellular Ca2+ chelation. TTN induced neutrophil chemotaxis, stimulated O2- generation, and enhanced phagocytosis. ENP did not affect cell migration and oxidative metabolism but enhanced phagocytosis. Both peptides modulated N-formyl-methionyl-leucyl-phenylalanine- and phorbol myristate acetate-induced O2- generation. Because neutrophils express benzodiazepine receptors of the peripheral type (pBRs) and DBI-derived peptides may interact with such receptors, we investigated the possible role of pBRs in TTN- or ENP-induced effects. The synthetic pBR ligand RO 5-4864 increased [Ca2+]i through extracellular Ca2+ influx and this effect was prevented by the pBR antagonist PK-11195. RO 5-4864, however, was ineffective on neutrophil migration and O2- generation and only slightly affected phagocytosis. Moreover, PK-11195 delayed the [Ca2+]i rise induced by TTN but did not significantly affect its extent, and had no effect on the [Ca2+]i rise induced by ENP. We conclude that DBI-derived peptides induce [Ca2+]i changes and modulate neutrophil function mainly through pBR-independent pathways. In view of the wide cell and tissue distribution of DBI in the brain and in peripheral organs, modulation of neutrophil function by DBI-derived peptides may be relevant for both the neuroimmune network and the development and regulation of the inflammatory processes.

Mycobacterial lipoarabinomannan affects human polymorphonuclear and mononuclear phagocyte functions differently.

BACKGROUND AND OBJECTIVE: The role of mycobacterial lipoarabinomannan (LAM) in regulating the granulomatous response and its effects on cells involved in early responses to tuberculosis have not been clearly defined. The aim of this study was to acquire further evidence about the mechanisms by which LAM takes part in the host response to mycobacterial infections. DESIGN AND METHODS: We compared the in vitro ability of mannosylated LAM (ManLAM) and LAM lacking the terminal mannosyl units (AraLAM) to induce distinct responses in human polymorphonuclear (PMNs) and mononuclear phagocytes [both monocytes and 48-hr monocyte-derived macrophages (MDMs)]. The responses examined were chemotaxis, transient changes in free cytosolic calcium, phagocytosis and metabolic activation. RESULTS: AraLAM and ManLAM affected mononuclear, but not polymorphonuclear, phagocyte functions. Both forms of LAM were chemotactic for monocytes and MDMs. The LAM-induced chemotactic response required new protein synthesis, did not induce a rise in cytosolic free calcium levels and was partially inhibited (about 50%) by genistein, but not by calphostin C or PD 98059. Lastly, at physiologic doses ManLAM significantly reduced phagocytosis of M. tuberculosis and zymosan particles by MDMs. INTERPRETATION AND CONCLUSIONS: Different phagocytic cells can exhibit variable responses to AraLAM and ManLAM. Moreover, LAMs affect cell functions through different mechanisms. Protein synthesis and activation of protein tyrosine kinases are important intermediates in the signal transduction pathway of the chemotactic response of mononuclear phagocytes to AraLAM and ManLAM; whereas ManLAM-induced inhibition of macrophage phagocytic ability could depend on the binding of macrophage mannose receptors and/or the insertion of this molecule into cellular plasma membrane. Together these data highlight the danger of making generalizations regarding the activity of LAMs on immune defenses.