Early IFN-γ Production Research Articles

Interleukin-6 induces early gamma interferon production in the infected lung but is not required for generation of specific immunity to Mycobacterium tuberculosis infection.

Immunity to Mycobacterium tuberculosis is dependent upon the generation of a protective gamma interferon (IFN-gamma)-producing T-cell response. Recent studies have suggested that interleukin-6 (IL-6) is required for the induction of a protective T-cell response and that IL-4 may suppress the induction of IFN-gamma. To evaluate the role of the cytokines IL-6 and IL-4 in the generation of pulmonary immunity to M. tuberculosis, IL-6 and IL-4 knockout mice were infected with M. tuberculosis via aerosol. The absence of IL-6 led to an early increase in bacterial load with a concurrent delay in the induction of IFN-gamma. However, mice were able to contain and control bacterial growth and developed a protective memory response to secondary infection. This demonstrates that while IL-6 is involved in stimulating early IFN-gamma production, it is not essential for the development of protective immunity against M. tuberculosis. In contrast, while the absence of IL-4 resulted in increased IFN-gamma production, this had no significant effect upon bacterial growth.

Dominance of IL-12 over IL-4 in gamma delta T cell differentiation leads to default production of IFN-gamma: failure to down-regulate IL-12 receptor beta 2-chain expression.

Gamma delta T cells secrete Th1- and Th2-like cytokines that help mediate innate and acquired immunity. We have addressed the mechanism whereby murine gamma delta T cells acquire the capacity to differentially produce such cytokines. Splenic gamma delta T cells could be polarized into IFN-gamma- or IL-4-secreting cells in vitro; however, in contrast to CD4+ alpha beta T cells, gamma delta T cells predominantly produced IFN-gamma, even in the presence of IL-4, a finding independent of genetic background. Like CD4+ Th1 cells, IFN-gamma-producing cells expressed the IL-12 receptor beta 2-chain after activation in the presence of IL-12; however, unlike Th2 cells, IL-4-primed gamma delta T cells also expressed this receptor, even in the absence of IFN-gamma and despite the presence of the transcription factor GATA-3. IL-12 also induced IL-4-primed gamma delta T cells to proliferate and to translocate Stat3/Stat4, indicating signaling through the IL-12 receptor. These molecular events can account for the predominant production of IFN-gamma by gamma delta T cells in the presence of IL-12, despite the availability of IL-4. Early and predominant production of IFN-gamma by gamma delta T cells likely is critical for the roles that these cells play in protection against intracellular pathogens and in tumor immunity.

Plasma cytokine response in mice with bacterial infection.

Exposure to microorganisms elicts the production of cytokines. These soluble factors enhance several innate immune functions and regulate the ensuing specific immune response aimed at limiting the spread of infection. This study was undertaken to quantify the plasma levels of pro-inflammatory cytokines during the course of primary Listeria monocytogenes and Campylobacter jejuni infection. Using an in vivo infection the relationship between endogenous cytokines and the bacterial number in the liver of infected animals was examined. C57BL/6 mice were infected by the intraperitoneal route. At different time points we determined the number of colony-forming units of bacteria in the liver of infected animals and paralled these with the plasma levels of interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) measured by enzyme immunoassays. L. monocytogenes infection lasted 10-11 days. IFN-gamma production occurred in the early phase but was more pronounced after day 4, following the appearance of specific immunity. The duration of experimental campylobacteriosis was 15 days. Early IFN-gamma production was not significant but a progressive rise of this cytokine in plasma was seen during the second week post infection. Mice produced measurable amounts of plasma TNF-alpha immediately after being given viable L. monocytogenes, peaking on day 2-3 when the greatest number of bacteria was present in the examined organs. During C. jejuni infection plasma TNF-alpha was produced in a similar manner, but the highest concentrations were found a few days later than in listeriosis, in correlation with the different course of campylobacteriosis. The quantity of IL-6 increased and decreased in concordance with clearance of L monocytogenes and the clinical status of the animals. C. jejuni did not promote the induction of this cytokine. This is to some extent an unusual finding. With respect to the role of IL-6 in Th2 responses and antibody production, the appearance of this cytokine in campylobacteriosis was more expected. During systemic bacterial infection, a network of pro-inflammatory cytokines is activated and blood levels of these cytokines are elevated, albeit inconsistently, with large individual variations and depending on microbial characteristics and structure.

Protection against Lethal Toxoplasmosis in Mice by an Avirulent Strain of Toxoplasma gondii: Stimulation of IFN-γ and TNF-α Response

Haque, S., Franck, J., Dumon, H., Kasper, L. H., and Haque, A. 1999. Protection against lethal toxoplasmosis in mice by an avirulent strain of Toxoplasma gondii: Stimulation of IFN-γ and TNF-α response. Experimental Parasitology93, 231–240. In this study, we examined whether the PTN strain (isolated from an AIDS patient) of Toxoplasma gondii could induce cross-protection in mice against infection with a lethal dose of the PLK strain. Mice were first infected with tachyzoites (5 × 105) of PTN and 5 days later challenged with PLK (1 × 105, LD90) parasites. None of these mice succumbed to infection until day 21 after infection, whereas 100% of the mice given the same dose of PLK infection alone died between 5 and 11 days after infection. The protection was accompanied by an increased expansion of NK cells and CD4 + T cells. This condition was associated by increased production of IFN-γ and an augmented number of IFN-γ-producing cells in the spleen. Further, PTN + PLK-infected mice showed higher production of TNF-α and nitrite compared to PLK-infected mice. Mice infected with the PTN strain had an enhanced capacity to activate the immune system early in infection since they produced higher levels of IFN-γ, TNF-α, and NO than PLK-infected mice. Administration of anti-IFN-γ mAb or anti-asialo GM1 antibody resulted in 100 and 20% mortality, respectively, in PTN-infected mice but no death in PTN + PLK-infected mice. Together, these results suggest that early production of IFN-γ and NK-cell activity is important in protection against PTN infection, whereas in PTN + PLK infection components of adaptive immunity rapidly developed following elaboration of an effective early innate immune response.

Early IFN-γ Production and Innate Immunity DuringListeria monocytogenesInfection in the Absence of NK Cells

AbstractNK cells are believed to play a mandatory role during the early phases of Listeria monocytogenes infection by producing IFN-γ, which is required for the activation of macrophage effector functions. Mice deficient in the common cytokine receptor γ-chain (γc−/−), which completely lack NK cells, were used to examine whether NK cells were essential for resistance to Listeria infection in vivo. Surprisingly, infected γc−/− mice showed normal innate immunity and macrophage responses against sublethal Listeria infection 2 days postinfection. At this time point, γc−/− mice showed increased blood IFN-γ levels compared with those in noninfected controls, demonstrating an NK-independent source of IFN-γ, which explains early resistance. Listeria-infected γc−/− × recombinase-activating gene-2−/− double-deficient mice were unable to produce IFN-γ and were highly susceptible to L. monocytogenes. Since T cells, but not B cells, are major IFN-γ producers, and γc−/− T cells were found to be efficient IFN-γ producers in vitro, we conclude from these results that T cells functionally replace NK cells for the early IFN-γ production that is necessary for activating the innate immune system following infection with L. monocytogenes. This novel observation in listeriosis underscores how the adaptive immune response can maintain and influence innate immunity.

Role of CD80 (B7.1) and CD86 (B7.2) in the Immune Response to an Intracellular Pathogen

The costimulatory ligands CD80 and CD86 play a crucial role in the initiation and maintenance of an immune response. We demonstrate that whereas infection of human monocytes with viable tachyzoites of Toxoplasma gondii resulted in rapid induction of expression of CD80 and up-regulation of expression of CD86, incubation with killed organisms failed to alter the levels of expression of these costimulatory ligands. The T. gondii-mediated changes in levels of expression of these molecules are critical to the T cell response to the parasite. Proliferation of resting T cells in response to parasite-infected cells was dependent on both CD80 and CD86. More importantly, early production of IFN-gamma in response to T. gondii by T cells from T. gondii-seronegative individuals occurred only after stimulation with monocytes that exhibited increased expression of CD80 and CD86 (monocytes infected with viable parasites) and was almost completely ablated by the combination of anti-CD80 plus anti-CD86 mAb. Moreover, proliferation and IFN-gamma production by CD4+ CD45RA+ T cells from unexposed individuals were dependent on both CD80 and CD86. These data indicate that pathogen-monocyte interaction influences the ensuing T cell response.

Interferon-γ Enhances Resolution of Herpes Simplex Virus Type 2 Infection of the Murine Genital Tract

The requirement for interferon-γ (IFNγ) in resolution of an HSV-2 vaginal infection and the cellular sources of this cytokine in the vaginal mucosa were assessed. IFNγ levels in vaginal secretions peaked on Days 2 and 5 following HSV-2 inoculation. Natural killer (NK) cell depletion greatly diminished the early production of IFNγ but had no significant effect on the rate of virus clearance. CD4+T cells were primarily responsible for the second peak of IFNγ levels and neutralization of this IFNγ beginning 3 days after virus inoculation delayed, but did not prevent, virus clearance from the vagina. HSV-2 persisted in mice depleted of both CD4+and CD8+T cells while clearance was delayed in CD4+, but not CD8+T cell-depleted mice, demonstrating the T cell dependence and predominant role of CD4+T cells in resolution of the infection. Together, these data suggest that IFNγ is not essential for virus clearance but plays an important role in enhancing T cell-mediated clearance mechanisms. The implication of these results is that IFNγ produced locally in the genital tract enhances virus clearance and may ultimately be important for reducing the amount of virus available to infect sensory ganglia.

Toxoplasma gondii:Evidence for Interleukin-12-Dependent and -Independent Pathways of Interferon-γ Production Induced by an Attenuated Parasite Strain

Scharton-Kersten, T., Caspar, P., Sher, A., and Denkers, E. Y. 1996.Toxoplasma gondii:Evidence for interleukin-12-dependent and -independent pathways of interferon-γ production induced by an attenuated parasite strain.Experimental Parasitology84,102–114. Immunity in mice infected withToxoplasma gondiiis dependent upon the ability to generate protective levels of the cytokine IFN-γ. In this report, we present evidence that the attenuated vaccine strain, ts-4, induces the latter cytokine by both IL-12-dependent and -independent pathways. In contrast, strain ME49 appears to induce IFN-γ wholly in dependence upon IL-12. Thus, 88% of wild-type C57BL/6 mice treated with anti-IL-12 mAb survive ts-4 infection, unlike similarly treated ME49-infected animals. Moreover, while anti-IL-12 treatment reduced early IFN-γ and nitric oxide production to background levels in ts-4-infected scid animals, the same treatment in infected C57BL/6 mice had no effect on production of the latter mediators. In addition, we found that anti-IL-12 treatment induces 100% mortality in CD4+-deficient MHC class II knockout mice infected with ts-4. Finally, production of nitric oxide (a molecule implicated in parasite control) was abrogated in ts-4-infected scid mice following depletion of IFN-γ producing NK cells. Together, our results suggest that ts-4 induces IL-12-dependent and -independent IFN-γ production in normal mice, but ME49 induces the latter cytokine only in dependence upon IL-12. Our data, furthermore, implicate involvement of T cells in the IL-12-independent component of the IFN-γ response.

Analysis of the Interrelationship between IL-12, TNF-α, and IFN-γ/Production during Murine Listeriosis

IL-12, a recently described cytokine, is an important mediator in the early production of IFN-γ during infection. To evaluate the timing of IL-12 production, and its relationship to TNF-α and IFN-γ production during primary murine listeriosis, we measured cytokine mRNA and protein levels in C57B1/6 mice infected intravenously with Listeria monocytogenes (LM). IL-12 is a disulfide-linked heterodimer containing two chains (designated P35 and P40); however, bioactive cytokine production has been more closely linked with P40 expression. Consequently, we monitored mRNA and protein levels of P40 in the spleen as a marker for IL-12 production in vivo. Splenic P40 mRNA levels (assayed using RNase protection methods) were low in uninfected animals, but increased markedly beginning 15 to 18 hr after LM infection. In sublethally infected animals, P40 mRNA levels remained elevated for 5 days, returning to baseline with the resolution of infection. P40 protein (assayed using an antibody capture ELISA) could be detected in the spleens of LM-infected animals beginning around 18 hr postinfection confirming linkage between P40 mRNA accumulation and the generation of a protein product. In comparing P40 and IFN-γ mRNA levels in vivo, we found in each case that substantial increases in mRNA accumulation did not appear until 15-18 hr postinfection. In comparable studies using BALB/c animals, cytokine production began slightly earlier (between 12 and 15 hr) but once again P40 and IFN-γ mRNA levels increased in a coordinated manner. P40 mRNA (like IFN-γ and TNF-α mRNA) only accumulated in animals infused with live, virulent bacteria. Although we could detect no obvious lag between the time of onset of IL-12 and IFN-γ accumulation in vivo, infusions of anti-IL-12 antibodies markedly reduced IFN-γ expression implying that IL-12 production precedes and directs IFN-γ production. TNF-α production, on the other hand, was not diminished by anti-IL-12 treatment. Our studies demonstrate that IL-12 generation is an essential step in normal IFN-γ production during listeriosis, and suggest that IL-12, once produced, may begin enhancing IFN-γ production in vivo in less than 3 hr.

Enhanced Capacity of Females for Early Interferon-γ Production by Natural Killer-Like Cells Following Stimulation by Staphylococcal Enterotoxin A

Staphylococcal enterotoxin A (SEA) induced the production of interferon-gamma (IFN-gamma) by spleen cells from ICR Swiss mice during the first 24 h of culture. Splenocytes from females produced higher levels of IFN-gamma than did those from males at 8, 12, and 16 h. By 20 h after SEA stimulation, IFN-gamma production by spleen cells from males was similar to that of females. The cell types involved in IFN-gamma production in this SEA/spleen cell system were analyzed by depletion studies. Removal of Thy-1+ cells by panning prevented production of IFN-gamma in the 24 h after SEA stimulation. In vivo depletion of asialo GM1+ (AGM1+) cells prevented production of IFN-gamma through 16 h of culture with SEA, but permitted a modest IFN-gamma response at 20 h that was similar in magnitude in both sexes. Following removal of L3T4+ and Lyt-2+ cells by panning, IFN-gamma production was detected at 12 h after SEA stimulation and maintained through 24 h of culture with cells from females producing higher levels of IFN-gamma. These data suggest that male ICR Swiss mice are deficient in the activity of Thy-1+, AGM1+, L3T4-, and Lyt-2- cells in the early (8-16 h) production of IFN-gamma following SEA stimulation of spleen cells.

Differential interleukin-2 and interferon-γ production by human lymphocyte cultures exceptionally resistant to Epstein-Barr virus immortalization

Epstein-Barr virus (EBV) readily immortalizes human peripheral blood lymphocytes (PBL) in vitro. However, during the past several years, we found that PBL from two exceptional EBV-seropositive healthy adult individuals were refractory to immortalization by EBV. We report here a study aimed at learning about the immunobiological features which differentiate these EBV-resistant (R) PBL from others which are susceptible (S) to EBV immortalization. Results of this investigation indicate that: (a) Following EBV infection, R-PBL produced significantly higher amounts of interferon γ (IFN-γ) than S-PBL. There were however no differences in regard to interferon α production between these two types (R and S) of EBV-infected cultures. (b) R-PBL had a maximal interleukin-2 (IL-2) production from Days 3 to 5 post-EBV infection, while maximal IL-2 production by S-PBL occurred at least 48 hr later, i.e., at Day 7. (c) The percentage of non-B cells expressing the IL-2 receptor was also higher in EBV-infected R-PBL than S-PBL. (d) In contrast, expression of IL-2 receptors after EBV infection was higher on B cells from S-PBL than on B cells from R-PBL. Interestingly, no differences were noted in regard to IL-2 receptor expression between R-PBL and S-PBL treated with mitogens (i.e., phytohemag-glutinin and pokeweed mitogen). (e) Finally, using anti-IL-2 and anti-IFN-γ antibodies in EBV-infected R-PBL cultures, we were able to obtain EBV-induced immortalization of these cultures. Taken together, these results suggest that an early IL-2 synthesis and high IFN-γ production by EBV-infected PBL play an important role against lymphocyte immortalization by EBV.