SUMMARYWe evaluated the use of recombinant human interleukin-6 (rhIL-6) and a monoclonal antibody speci®c for interferon-c (IFN-c) as co-adjuvants in a subunit vaccine against tuberculosis consisting of the culture ®ltrate proteins of Mycobacterium tuberculosis (ST-CF) emulsi®ed in the adjuvant dimethyl-dioctadecylammonium bromide (DDA). Both the addition of rhIL-6 and the neutralization of IFN-c resulted in an increased T helper type 1 (Th1) response characterized by enhanced IFN-c production and cell proliferation. Nevertheless, this did not result in the enhancement of protection against either an intravenous or an aerosol M. tuberculosis challenge. Our data stress the need to identify further correlates of protection in addition to IFN-c production to screen vaccines against tuberculosis infection.

We examined the role of cytokines in the development of gamma interferon (IFN-γ)-secreting protective T cells following immunization with a culture filtrate subunit vaccine againstMycobacterium tuberculosis containing the adjuvant dimethyldioctadecylammonium bromide (DDA). Depletion of either interleukin-6 (IL-6) or IL-12 with specific neutralizing antibodies during vaccination reduced the priming of T cells for antigen-specific proliferation and IFN-γ secretion. Such reduction was also observed in IL-6 gene-disrupted mice as compared to wild-type animals. IL-6 was found to play a role in the initial differentiation of Th1 cells but not in their expansion. The defect found after IL-6 depletion or in IL-6-knockout mice was compensated by the inclusion of recombinant mouse IL-12 in the vaccine. The induction of protective immunity against an intravenous or an aerosol challenge with live, virulentM. tuberculosis was markedly reduced by neutralizing either IL-6 or IL-12 during immunization with the vaccine. Likewise, the effects of IL-6 neutralization were partially reversed by including IL-12 in the vaccine. Our data point to an important role of IL-6 and IL-12 in the generation of cell-mediated immunity to tuberculosis.

Summary We investigated the role of interleukin‐6 (IL‐6) in the development of the immune response to a subunit vaccine against tuberculosis consisting of the culture filtrate proteins of Mycobacterium tuberculosis emulsified in the adjuvant dimethyldioctadecylammonium bromide (DDA). C57Bl/6 mice immunized with this vaccine developed a strong T helper 1 (Th1) response characterized by an increased production of interferon‐γ (IFN‐γ) secreted by CD4+ T cells. Neutralization of IL‐6 during in vivo priming resulted in marked reduction in the ability of T cells to secrete IFN‐γ and IL‐2 and to proliferate. IL‐6 gene‐disrupted mice primed with the vaccine showed a decrease in the number of IFN‐γ‐producing cells and an increase in IL‐4‐secreting cells as compared to control mice. In contrast, neutralization of IL‐6 during a boost of the vaccine in previously primed mice did not affect the development of IFN‐γ‐producing cells but still increased the number of IL‐4‐producing cells. Our work shows that IL‐6 plays a major role in the priming but not in the later expression of a Th1 response to a tuberculosis vaccine.

Listeria monocytogenes mutants defective in the actA gene, the plcB gene, and the inlA and inlB genes were less virulent when injected intravenously into BALB/c mice. The growth of these strains as well as of the virulent wild-type strains was increased by treating mice with a neutrophil-specific depleting monoclonal antibody, RB6-8C5. Histologic examination of the livers of the treated animals showed intrahepatocytic proliferation of the listeriae in all cases. Our data show that more than one pathway exists that allows L. monocytogenes to invade parenchymal cells. One pathway most likely involves the actA and plcB gene products, and a second one probably involves the internalins.

After infection with a low-virulence strain of Mycobacterium avium, C57BL/6 and C57BL/10 mice had clear differences in the control of the infection in their livers and spleens. This difference in susceptibility was not associated with differences in the H-2 complex. It was dependent on the activity of CD4 ؉ T cells but unrelated to the ability of these cells to secrete gamma interferon or to the development of delayed-type hypersensitivity responses at 3 weeks of infection. It was associated with lower total numbers of CD4 ؉ cells present in infected spleens and was related to an earlier induction of protective T cells, as measured by adoptive-transfer assays. These data further strengthen the notion of gamma-interferon-independent mechanisms of protection against mycobacteria.Mycobacterium avium is an opportunistic pathogen in humans affecting most frequently immunocompromised patients, namely, patients with AIDS (9). This intracellular mycobacterium proliferates inside macrophages and is controlled by as yet poorly understood effector mechanisms of these phagocytes, namely, after their activation by cytokines produced during innate and acquired immune responses (1). One of the major genetic determinants of resistance to infection by M. avium was identified in mice as the Nramp1 gene (formerly known as the Bcg/Ity/Lsh locus) (2). This gene is expressed mostly in macrophages and imparts an innate capacity to these cells to control M. avium infection, as well as other mycobacterial, Salmonella, and protozoal infections, even in the absence of external modulation by other immune cells and the cytokines produced by them (8). Although the mechanism of action of the encoded transmembrane polypeptide is still not clarified, it is possible that it is involved in the transport of iron out of the phagosome, thereby leading to the deprivation of such nutrients and consequent mycobacteriostasis (3, 6). Alternatively, it may work via the acidification of the mycobacterium-bearing phagosome (7).It has been suggested by Orme and colleagues (13) that additional genetic determinants affecting the in vivo growth of M. avium are present in mice. Preliminary observations in our laboratory showed that strains of mice sharing the D169 mutant allele of Nramp1 (i.e., the susceptibility allele of the Bcg gene) differed in the control of the infection induced by an AIDS isolate of M. avium. We study here two such strains, C57BL/6 and C57BL/10, that differ in their susceptibilities to M. avium and show that the efficiency in mobilizing protective T cells against the infection by M. avium differs between these two strains. MATERIALS AND METHODSAnimals. Female C57BL/6J and C57BL/10ScSnOlaHsd mice were used when they were 6 to 8 weeks old. C57BL/6J mice were obtained from the Gulbenkian Institute (Oeiras, Portugal) and C57BL/10ScSnOlaHsd and B10.D2/nOlaHsd mice were purchased from Harlan (Oxon, United Kingdom). Mice were bred under similar conditions in our facilities and given sterile food and acidified drinking water. T-cell-depleted mice...