IFN-gamma regulates the isotypes of Ig secreted during in vivo humoral immune responses

The lymphokine IFN-gamma has been shown in vitro to stimulate IgG2a secretion and inhibit IgG1 and IgE secretion by LPS-activated B lymphocytes. To determine whether IFN-gamma has a similar isotype regulatory role in vivo, we studied the abilities of rIFN-gamma and a mAb to IFN-gamma to modify the isotypes of Ig secreted in mice injected with a goat antibody to mouse IgD, which by itself induces large increases in levels of serum IgG1 and IgE and a relatively small increase in serum IgG2a. Multiple injections of IFN-gamma substantially inhibited production of IgG1 and IgE, and stimulated production of IgG2a in affinity purified goat antibody specific for mouse IgD-treated mice; anti-IFN-gamma antibody blocked the effects of IFN-gamma and in fact enhanced IgG1 and IgE secretion and inhibited the IgG2a response in these mice. The role of IFN-gamma in the selection of isotypes of Ig produced in response to injection of mice with the bacterium Brucella abortus (BA) was also studied, because killed, fixed BA are known to stimulate IFN secretion and a predominantly IgG2a antibody response. Anti-IFN-gamma antibody strongly suppressed IgG2a secretion and stimulated IgG1, but not IgE, secretion in BA-immunized mice. BA suppressed IgG1 and IgE secretion and enhanced IgG2a secretion in affinity purified goat antibody specific for mouse IgD-injected mice; treatment of these mice with anti-IFN-gamma antibody reversed the effects of BA on IgG1 and IgG2a secretion, but not the suppressive effect of BA on IgE secretion. These observations demonstrate that IFN-gamma has an important and perhaps unique physiologic role in the stimulation of IgG2a secretion and in the suppression of secretion of IgG1, whereas bacterial antigens can suppress IgE secretion by other mechanisms in addition to IFN-gamma secretion.     

IgE production by normal human B cells induced by alloreactive T cell clones is mediated by IL-4 and suppressed by IFN-gamma

Seven T cell clones were established from mixed leukocyte cultures in which PBMC from two healthy donors and from one patient suffering from the hyper-IgE syndrome were stimulated by the irradiated EBV-transformed B cell lines JY or UD53. Five of seven T cell clones, after activation by co-cultivation with JY or UD53 cells, induced a low degree of IgE production by normal blood B cells. In one experiment in which the normal B cells could activate the T cell clones directly, IgE production was also observed in the absence of the specific stimulator cells. IgE production was also obtained with supernatants of the T cell clones collected 4 to 5 days after activation by their specific stimulator cells. In addition, the supernatants induced IgG, IgA, and IgM synthesis. All seven clones produced variable concentrations of IL-4 and IFN-gamma. The clones FA-28 and BG-39, which failed to induce IgE synthesis, produced, compared with the other clones tested, relatively high quantities of IFN-gamma (4700 and 2500 pg/ml, respectively). These high levels of IFN-gamma accounted for the lack of induction of IgE synthesis, because in the presence of a polyclonal anti-IFN-gamma antiserum, supernatants of FA-10 and BG-39 induced significant IgE production. In addition, the low degree of IgE production induced by supernatants of two other T cell clones (FA28 and BG24) was 15- and 3-fold enhanced, respectively, in the presence of the anti-IFN-gamma antiserum. IgE synthesis by normal B cells was also induced by rIL-4, indicating that IL-4 present in T cell clone supernatants was responsible for induction of IgE production. This notion was supported by the finding that IgE production induced by supernatant of BG-24 was strongly inhibited by a polyclonal anti-IL-4 antiserum. In contrast, IgG and IgA production induced by supernatant of BG-24 were not significantly affected by the anti-IL-4 antiserum. Only a slight inhibition of IgM synthesis was observed. Collectively, our results indicate that both recombinant and naturally produced IL-4 induce normal human B cells to synthesize IgE. However, final IgE production induced by T cell clone supernatants is the net result of the inducing and suppressive effects of IL-4 and IFN-gamma respectively, that are secreted simultaneously by the T cell clones upon activation.     

B cell stimulatory factor-1 enhances the IgE response of lipopolysaccharide-activated B cells

Supernatants from some mouse helper T cell (TH) lines contain an activity that can enhance IgE production by lipopolysaccharide (LPS)-stimulated B cells by at least two orders of magnitude. During purification, this activity could not be resolved from B cell stimulatory factor-1 (BSF-1). Highly purified BSF-1 from a different source, the T lymphoma cell line EL-4, enhanced IgE production to the same extent as TH supernatants, which suggests that BSF-1 is responsible for this increase in IgE production. Monoclonal antibody to BSF-1 totally inhibits the IgE-enhancing activity of a TH supernatant, lending further support to this conclusion. The effects of BSF-1 on LPS-stimulated B cells are specific for IgE and, as previously reported, IgG1 and IgG3, because the levels of IgM, IgG2a, IgG2b, and IgA in the cultures change relatively little when BSF-1 is added.     

Role of porin of Shigella dysenteriae type 1 in modulation of lipopolysaccharide mediated nitric oxide and interleukin-1 release by murine peritoneal macrophages

The ability of Shigella dysenteriae type 1 porin to induce the release of nitric oxide (NO) and interleukin-1 (IL-1) from peritoneal macrophages of mouse and to regulate lipopolysaccharide (LPS) and gamma interferon (IFN-gamma) mediated release of the two proinflammatory mediators was investigated. Porin released nitrite when added to macrophage cultures. A maximum of 3.2-fold nitrite release by macrophages was observed with 100 ng ml(-1) of porin. The nitrite release of LPS was enhanced significantly by lower concentrations of porin, whereas the effect of IFN-gamma was enhanced by porin at higher concentrations. Polysaccharide (PS) moiety of LPS stimulated the nitrite release of elicited macrophages by 1.6-fold compared to untreated control. It also enhanced the stimulatory effect of 1 and 10 ng ml(-1) of porin by 1.3-fold. Lipid A (LPA) moiety of LPS did not release nitrite, nor did it increase the porin mediated nitrite production. Porin treated 24 h old macrophage culture supernatants were applied for ConA activated thymocyte proliferation as a measure for determination of IL-1 release. Sixty percent depletion of thymocyte proliferation was observed when the porin treated macrophage supernatants were absorbed with anti-IL-1 antibody. A maximum of 5.5-fold increase of thymocyte proliferation over control was found with 1 and 10 ng ml(-1) of porin. One or 10 ng ml(-1) of porin and LPS augmented the thymocyte growth, 1.5-fold beyond that obtained by porin and 1.8-/1. 7-fold more than that obtained by LPS, alone. Similarly, porin and IFN-gamma co-stimulated the cell growth also. PS enhanced the thymocyte proliferation by 5-fold. It also enhanced the thymocyte growth by co-stimulating 1.4-fold the effect observed by 1 or 10 ng ml(-1) of porin alone. LPA could not participate in the cell proliferating activity nor did it enhance the stimulatory effect of porin. Therefore, both nitrite release and thymocyte proliferation by LPS could be substituted by PS only. The tight association of the two bacterial outer membrane components, porin and LPS, could be a necessary co-signal for boosting the release of the two proinflammatory mediators, namely NO and IL-1, which may be associated with the inflammatory response of the colon during Shigella invasion.     

Evidence for differential induction of helper T cell subsets during Trichinella spiralis infection

The H-2-compatible mouse strains, AKR and B10.BR, exhibit disparate responses to infection with the parasitic nematode Trichinella spiralis. The resistant AKR mice expel intestinal adult worms faster than susceptible B10.BR mice. We tested antibody and lymphokine responses in these strains. With respect to antibody responses, the B10.BR mice had 3- to 10-fold more serum IgE and T. spiralis-specific IgG1 and IgA than AKR mice. The B10.BR mice also had greater numbers of IgG and IgA plaque-forming cells than AKR mice. In contrast, AKR mice produced T. spiralis-specific IgG2a, whereas the B10.BR mice did not. The antibody response kinetics of these strains were similar. We also analyzed lymphokine secretion after restimulating lymphocytes in vitro with T. spiralis Ag. The AKR mesenteric lymph node cells produced more IFN-gamma and less IL-4 than the B10.BR mesenteric lymph node cells. The B10.BR splenocytes produced more IL-4 than the AKR splenocytes, although splenocyte IFN-gamma production was not different. The kinetics of IL-4 production also differed between the two strains. In summary, resistant AKR mice produced more IFN-gamma and T. spiralis-specific IgG2a than susceptible B10.BR mice, which produced more IL-4, IgE, and T. spiralis-specific IgG1. Our results are consistent with differential activation of Th cell subsets in T. spiralis-infected AKR and B10.BR mice.     

Immunological determinants of ventilatory changes induced in mice by dermal sensitization and respiratory challenge with toluene diisocyanate

The objective of the study was to characterize better the immunologic mechanisms underlying a previously developed animal model of chemical-induced asthma. BALB/c and severe combined immunodeficiency disease (SCID) mice received toluene diisocyanate (TDI) or vehicle on each ear on day 1 and/or day 7. On day 10, they were intranasally challenged with TDI or vehicle. Ventilatory function was monitored by whole body plethysmography for 40 min after challenge. Reactivity to methacholine was measured 23 h later: enhanced pause and actual resistance measurements. Pulmonary inflammation was assessed 1, 6, and 24 h after challenge by bronchoalveolar lavage (BAL). Tumor necrosis factor-alpha and macrophage inflammatory protein (MIP)-2 levels were measured in BAL. Immunological parameters included total IgE, IgG1, and IgG2a in serum, lymphocyte populations in auricular and cervical lymph nodes, and IL-4 and IFN-gamma levels in supernatants of lymph node cells, cultured with or without concanavalin A. Ventilatory changes suggestive of airway obstruction and increased methacholine reactivity were observed in all TDI-sensitized and TDI intranasally instilled mice, except in SCID mice. A neutrophil influx, accompanied by an increase in MIP-2 levels, was found in BAL of all responding groups 6 and 24 h after intranasal challenge. In BALB/c mice an increased level of CD19+ B cells was found in the auricular lymph nodes. IL-4 and IFN-gamma levels were increased in supernatants of concanavalin A-stimulated auricular lymph node cells from BALB/c mice completely treated with TDI. These results indicate that our model is dependent on the presence of lymphocytes, but it is not characterized by a preferential stimulation of Th1 or Th2 lymphocytes.     

IFN-gamma stimulates IgG2a secretion by murine B cells stimulated with bacterial lipopolysaccharide

rIFN-gamma strikingly enhances the secretion of IgG2a by murine splenic B cells stimulated with bacterial LPS in vitro and concomitantly suppresses the production of IgG3, IgG1, IgG2b, and IgE while sparing IgM secretion. IFN-gamma stimulates highly purified B cell populations to secrete IgG2a, strongly suggesting that it acts directly on B cells. It increases the frequency of precursors of IgG2a-expressing soft agar colonies and enhances the number of IgG2a+ cells in colonies indicating that it both increases the frequency of precursors of IgG2a+ cells and enhances the number of IgG2a+ daughter cells emerging from each precursor. IFN-gamma completes its action within the first 24 to 48 h of a 6-day culture with LPS and its addition cannot be delayed beyond the first 48 h. Preincubation of resting B cells in the presence of IFN-gamma leads to a time dependent increase, up to 42 h, in IgG2a secretion upon subsequent addition of LPS. IFN-gamma can exert this action on resting B cells that have been selected for absence of membrane IgG expression by cell sorting. The promotion of IgG2a secretion appears to be a specific property of IFN-gamma in that IFN-alpha, IFN-beta, IL-1, IL-2, IL-3, IL-4, IL-5, granulocyte-macrophage-CSF, granulocyte-CSF, and CSF-1 fail to enhance IgG2a secretion by LPS-stimulated B cells.     

The polyclonal and antigen-specific IgE and IgG subclass response of mice injected with ovalbumin in alum or complete Freund's adjuvant

BALB/c mice were injected ip with 1 microgram ovalbumin (OVA) in alum or complete Freund's adjuvant (cFA) and the changes of the IgE and IgG subclass serum levels and isotypes of the anti-OVA specific antibodies determined by radioimmunoassays. By Day 10, OVA in alum had induced a 5- to 10-fold increase of the IgE serum level and an initial decrease of the IgG subclass levels which subsequently increased to two to threefold over the preinjection level. OVA in cFA induced a gradual twofold increase of the IgE serum level, a rapid fourfold increase of the IgG2a level occurring by Day 7, and a gradual two to threefold increase of the other IgG subclasses. Over 90% of the anti-OVA antibodies were of the IgGl isotype with both adjuvants; OVA in alum induced slightly more IgGl anti-OVA antibodies than cFA. In contrast, the OVA in alum injected mice formed significantly more (5- to 10-fold) IgE anti-OVA antibodies than the cFA-injected mice. OVA in alum also induced a large nonspecific increase of the IgE serum level because only approximately 40% of the increase observed on Day 14 was absorbable with OVA, whereas approximately 90% the IgE increase in cFA injected mice was absorbable with OVA. The data demonstrate that mice form mainly IgGl and IgE antibodies to OVA irrespective of the adjuvant. The low specific and lack of nonspecific IgE formation by mice injected with OVA in cFA may be the result of cFA-induced interferon-gamma (IFN-gamma) production because IFN-gamma has been shown to stimulate IgG2a and inhibit IgE secretion in vitro.     

Distinct modulatory effects of LPS and CpG on IL-18-dependent IFN-gamma synthesis

Innate cellular production of IFN-gamma is suppressed after repeated exposure to LPS, whereas CpG-containing DNA potentiates IFN-gamma production. We compared the modulatory effects of LPS and CpG on specific cellular and cytokine responses necessary for NK-cell dependent IFN-gamma synthesis. C3H/HeN mice pretreated with LPS for 2 days generated 5-fold less circulating IL-12 p70 and IFN-gamma in response to subsequent LPS challenge than did challenged control mice. In contrast, CpG-pretreated mice produced 10-fold more circulating IFN-gamma without similar changes in IL-12 p70 levels, but with 10-fold increases in serum IL-18 relative to LPS-challenged control or endotoxin-tolerant mice. The role of IL-18 in CpG-induced immune potentiation was studied in splenocyte cultures from control, LPS-conditioned, or CpG-conditioned mice. These cultures produced similar amounts of IFN-gamma in response to rIL-12 and rIL-18. However, only CpG-conditioned cells produced IFN-gamma when cultured with LPS or CpG, and production was ablated in the presence of anti-IL-18R Ab. Anti-IL-18R Ab also reduced in vivo IFN-gamma production by >2-fold in CpG-pretreated mice. Finally, combined pretreatment of mice with LPS and CpG suppressed the production of circulating IFN-gamma, IL-12 p70, and IL-18 after subsequent LPS challenge. We conclude that CpG potentiates innate IFN-gamma production from NK cells by increasing IL-18 availability, but that the suppressive effects of LPS on innate cellular immunity dominate during combined LPS and CpG pretreatment. Multiple Toll-like receptor engagement in vivo during infection can result in functional polarization of innate immunity dominated by a specific Toll-like receptor response.     

The IgE and IgG subclass responses of mice to four helminth parasites

To investigate whether the formation of IgE is linked in vivo to an IgG subclass, mice were infected with four helminth parasites, Nippostrongylus brasiliensis (Nbr), Mesocestoides corti, Taenia crassiceps and Trichinella spiralis, and the changes in the serum levels of the different Ig isotypes as well as the antibody response to M. corti and T. crassiceps antigen extracts were determined by radioimmunoassays. All four parasites induced a concomitant increase of the IgE and IgG1 serum levels and usually a decrease of the IgG2a level. They also induced an increase of the IgM level but had little effect on the IgG2b, IgG3, and IgA serum levels. The specific antibodies to an M. corti antigen extract were mainly of the IgG1 subclass, whereas it was of both IgG1 and IgG2a subclasses to T. crassiceps. Injections of dead M. corti induced an increase of all IgG subclasses and similar levels of IgG1 and IgG2a anti-parasite antibodies. Subcutaneous instead of intraperitoneal infection with T. crassiceps induced higher IgG2a than IgG1 levels and 10-fold lower IgE levels than the natural ip infection; however, despite the greater IgG2a polyclonal response, anti-parasite antibodies were predominantly of the IgG1 subclass. The data demonstrate that natural infection with four different helminth parasites induces a concomitant polyclonal IgG1 and IgE response. These in vivo observations corroborate the recent in vitro findings demonstrating that interleukin-4 induces lipopolysaccharide-activated murine B cells to secrete both IgG1 and IgE, suggesting that the regulation of these two isotypes is linked.     

Immunomodulatory role of IL-4 on the secretion of Ig by human B cells

The effect of IL-4 on the production of Ig by human B cells was examined. Highly purified B cells were stimulated with Staphylococcus aureus (SA) and IL-4 alone or in combination with various other cytokines and the supernatants assayed for Ig by isotype-specific ELISA. IL-4 (10 to 100 U/ml) did not support Ig secretion by SA-stimulated blood, spleen, or lymph node B cells, whereas IL-2 supported the production of all isotypes including IgE. Moreover, IL-4 suppressed the production of all isotypes of Ig by B cells stimulated with SA and IL-2 including IgG1, IgG2, and IgE. IL-4-mediated suppression was partially reversed by IFN-gamma or -alpha and low m.w. B cell growth factor. TNF-alpha and IL-6 did not reverse the IL-4-induced suppression of Ig production. The inhibitory action of IL-4 on Ig production appeared to depend on the polyclonal activator used to stimulate the B cells. Thus, Ig secretion by B cells activated by LPS and supported by IL-2 was not inhibited by IL-4. Whereas IL-4 alone supported minimal Ig production by LPS-activated B cells, it augmented production of all Ig isotypes in cultures stimulated with LPS and supported by IL-2. IFN-gamma further enhanced production of Ig in these cultures. When the effect of IL-4 on the responsiveness of B cells preactivated with SA and IL-2 was examined, it was found not to inhibit but rather to promote Ig production modestly. A direct effect of IL-4 on the terminal differentiation of B cells was demonstrated using B lymphoblastoid cell lines. IL-4 was able to enhance the Ig secreted by an IgA-secreting hybridoma, 219 and by SKW6-CL-4, an IL-6-responsive IgM-secreting EBV transformed B cell line. These results indicate that IL-4 exerts a number of immunoregulatory actions on human B cell differentiation. It interferes with the activation of B cells by SA and IL-2, but promotes the differentiation of preactivated B cells, B cell lines, and B cells activated by LPS without apparent isotype specificity.     

Kinetics of interleukin-4 induction and interferon-gamma inhibition of IgE secretion by Epstein-Barr virus-infected human peripheral blood B cells.

Interleukin-4 (IL-4) acts directly on purified human peripheral blood B cells cultured in the presence of Epstein-Barr virus (EBV) to induce IgE secretion and to enhance the secretion of IgG and IgM. Interferon-gamma (IFN-gamma) inhibits IgE secretion in this system, without affecting the secretion of the other Ig isotypes. To identify the time period during which EBV-infected B cells can be induced by IL-4 to secrete IgE, we have studied the effects of delayed addition of IL-4, or the termination of IL-4 stimulation by wash out or by neutralization with anti-IL-4 antibodies, on the induction of an IgE response. To induce a maximal IgE response, IL-4 had to be added to cultures of B cells plus EBV no later than 2 days after the initiation of culture, and had to remain present through the tenth day of culture. These two time points correspond to the initiation of detectable DNA synthesis (Days 3 to 4) and the earliest detectable Ig secretion (Days 10 to 12) by EBV-stimulated B cells. No IgE response was induced if the period during which EBV-stimulated B cells were cultured with IL-4 was less than 4 days, or if IL-4 were added later than the tenth day of culture, regardless of how long the culture was continued beyond that time. In contrast, IL-4 considerably enhanced IgG and IgM secretion and B cell CD23 expression, even if it was added after the tenth day of culture. IFN-gamma strongly inhibited the IgE response of B cells cultured with IL-4 plus EBV if added within 6 days of the initiation of culture, but had little effect on the generation of IgM or IgG responses made by these cells, regardless of the time of addition. Neither IL-4 nor IFN-gamma affected ongoing IgE secretion by an established, IgE-secreting, EBV-transformed cell line. These observations suggest that: (i) IL-4 first becomes able to induce EBV-activated B cells to secrete IgE as these cells begin to synthesize DNA, must stimulate B cells for at least 4 days to induce IgE secretion, and loses its ability to induce IgE secretion as these cells differentiate into Ig-secreting cells; (ii) the ability of IFN-gamma to suppress an IgE response is limited to this same time period; and (iii) IL-4 enhancement of CD23 expression and IgM and IgG secretion are independent of IL-4 induction of an IgE response.     

Interferon-gamma induces enhanced expression of Ia and H-2 antigens on B lymphoid, macrophage, and myeloid cell lines

The levels of class II major histocompatibility complex (MHC) antigens (la antigens) on cells of a cultured B lymphoma line (WEHI-279) were significantly increased after 24 hr incubation with medium conditioned by concanavalin A-stimulated mouse or rat spleen cells, or by an azobenzenearsonate- (ABA) specific T cell clone that had been stimulated with ABA-coupled spleen cells or concanavalin A. The levels and properties of the la-inducing activity correlated with those of interferon-gamma (IFN-gamma) measured by inhibition of virus plaque formation. Both the la-inducing activity and the IFN-gamma from the T cell clone had an apparent m.w. of 40,000 determined by gel filtration, were sensitive to treatment with trypsin or exposure to pH 2, but were stable to heat (56 degrees C, 1 hr). The induction of la antigens on WEHI-279 cells was dose-dependent, and the maximum response occurred at a concentration corresponding to 1 to 2 U/ml of antiviral activity. This T cell-derived IFN-gamma-like molecule also increased the expression of cell surface la antigens on another B cell line (WEHI-231), and cell lines of macrophage (J774) and myeloid (WEHI-3B and WEHI-265) origin. Furthermore, in all cases the levels of class I MHC (H-2K or H-2D) antigens were also increased. Similar patterns of induction of Ia and H-2 antigens were obtained with supernatants containing IFN-gamma produced by a monkey cell line (COS) that had been transfected with a plasmid bearing the cloned murine IFN-gamma gene. This activity was sensitive to pH 2 and was not present in the supernatant from COS cells that were not transfected with the murine IFN-gamma gene. These results established that IFN-gamma is the T cell-derived molecule that induces the enhanced expression of Ia and H-2 antigens on B cells and macrophages. A major physiologic role of IFN-gamma may be to regulate immune function through the enhanced expression of MHC antigens.     

Suppression of a polyclonal B-cell response by supernatants from the murine autologous mixed lymphocyte reaction

Previously, we have demonstrated that supernatants from autologous mixed lymphocyte (AMLR) cultures contain helper factors which can mediate the development of a cytotoxic T-cell response to hapten modified self. In the current study, the effect of AMLR supernatants on the humoral response was explored. BALB/C splenic non-T cells produced a large polyclonal antibody response to lipopolysaccharide (LPS), as measured in a Protein A SRBC plaque assay. Surprisingly, syngeneic AMLR supernatants suppressed the LPS-induced generation of plaque-forming cells. The presence of T cells in the stimulated cultures did not affect suppressor activity. The decreased response was not the result of a shift in kinetics, as maximal activity was observed on Day 4, whether or not AMLR supernatants were added. The AMLR culture supernatants were most effective in suppressing the plaque-forming cell response when added at the initiation of culture. AMLR supernatants added after 24 hr of culture resulted in only about 50% of maximum suppression. Supernatants added at 48 or 72 hr had no effect. Interferon-gamma (IFN-gamma) has been detected in AMLR culture supernatant and has been reported to suppress the development of plaque-forming cells in response to LPS. However, it is unlikely the suppressive activity observed in these studies is due to IFN-gamma. Dialysis of the AMLR culture supernatant against a pH 2 buffer for 24 hr or incubation at 70, 80, or 90 degrees C for 10 min, treatments that inactivate IFN-gamma, enhanced suppression. These results suggest that in addition to cytotoxic-T-cell helper factors, the cellular interactions in the AMLR induces the production of a stable mediator(s) which is able to directly suppress B cells at an early stage of their development into plasma cells.     

Inhibition by Trypanosoma cruzi of interferon-gamma production by mitogen-stimulated mouse spleen cells

Infection by Trypanosoma cruzi is accompanied by severe immunosuppression during the acute period. As part of our studies, to define the alterations caused by Trypanosoma cruzi in lymphocyte function, we examined in this work the interferon-gamma (IFN-gamma)-producing capacity of mitogen-stimulated mouse spleen and human peripheral blood mononuclear cells in the presence or absence of blood forms of the parasite. Co-culture of phytohaemagglutinin- or concanavalin A-stimulated spleen cells from normal mice with T. cruzi significantly decreased the levels of IFN-gamma activity found in the supernatants at 48 or 72 h. In contrast, human peripheral blood mononuclear cells, though suppressed by T. cruzi in their capacity to proliferate upon mitogenic stimulation, showed no significant decrease in IFN-gamma production. The addition of exogenous IFN-gamma did not reverse the suppressive effect of T. cruzi on either mouse or human cells. These results revealed, for the first time, the ability of T. cruzi to impair IFN-gamma production by activated mouse lymphocytes. The lack of restoration by exogenous IFN-gamma suggested that the reduced levels of this lymphokine were not, at least by themselves, the causative factor of reduced lymphoproliferation.     

IFN-gamma enhances secretion of IgG2a from IgG2a-committed LPS-stimulated murine B cells: implications for the role of IFN-gamma in class switching

IFN-gamma is a pleiotropic lymphokine that influences the isotypes of immunoglobulin secreted by B cells. IFN-gamma inhibits the secretion of IgG3, IgG2b, IgG1, and IgE, and enhances the secretion of IgG2a. We have examined the mechanism of IFN-gamma-mediated enhancement of IgG2a secretion in sorted populations of B cells and find that IFN-gamma reproducibly stimulates a twofold increase in the precursor frequency of IgG2a-secreting cells in the sIgG2a+ population. Additionally, we find that IFN-gamma does not induce an increase in the clone size of IgG2a-secreting cells. IFN-gamma stimulates a twofold increase in the precursor frequency of IgG2a-secreting cells from sIgG- and unsorted B cells which can be attributed to an increase in IgG2a secretion from IgG2a-committed cells in these populations. Hence, under the culture conditions utilized in these studies. IFN-gamma enhances IgG2a secretion from IgG2a-committed cells and does not induce a class switch.     

Gene expression and production of the monokine induced by IFN-gamma (MIG), IFN-inducible T cell alpha chemoattractant (I-TAC), and IFN-gamma-inducible protein-10 (IP-10) chemokines by human neutrophils

Monokine induced by IFN-gamma (MIG), IFN-inducible T cell alpha chemoattractant (I-TAC), and IFN-gamma-inducible protein of 10 kDa (IP-10) are related members of the CXC chemokine subfamily that bind to a common receptor, CXCR3, and that are produced by different cell types in response to IFN-gamma. We have recently reported that human polymorphonuclear neutrophils (PMN) have the capacity to release IP-10. Herein, we show that PMN also have the ability to produce MIG and to express I-TAC mRNA in response to IFN-gamma in combination with either TNF-alpha or LPS. While IFN-gamma, alone or in association with agonists such as fMLP, IL-8, granulocyte (G)-CSF and granulocyte-macrophage (GM)-CSF, failed to influence MIG, IP-10, and I-TAC gene expression, IFN-alpha, in combination with TNF-alpha, LPS, or IL-1beta, resulted in a considerable induction of IP-10 release by neutrophils. Furthermore, IL-10 and IL-4 significantly suppressed the expression of MIG, IP-10, and I-TAC mRNA and the extracellular production of MIG and IP-10 in neutrophils stimulated with IFN-gamma plus either LPS or TNF-alpha. Finally, supernatants harvested from stimulated PMN induced migration and rapid integrin-dependent adhesion of CXCR3-expressing lymphocytes; these activities were significantly reduced by neutralizing anti-MIG and anti-IP-10 Abs, suggesting that they were mediated by MIG and IP-10 present in the supernatants. Since MIG, IP-10, and I-TAC are potent chemoattractants for NK cells and Th1 lymphocytes, the ability of neutrophils to produce these chemokines might contribute not only to the progression and evolution of the inflammatory response, but also to the regulation of the immune response.