anti-TNP Response Research Articles

Affinity maturation occurs in channel catfish (Ictalurus punctaus) following immunization with a T-cell dependent antigen

Affinity maturation of the antibody response, a process of antibody affinity increasing over response, is one of the key features of the mammalian immune system. However, the process is incompletely understood in teleost, including channel catfish (Ictalurus punctaus). In this study, IgM affinity maturation in channel catfish was investigated by estimating the kinetics of antibody affinity using ELISA and ELISPOT assays. Fish were immunized with a T-cell dependent antigen (TNP-KLH), and individual serum IgM antibody titers and affinities, and IgM+ antibody-secreting cells (ASCs) in peripheral blood were analyzed over a period of 14 weeks. A detectable serum anti-TNP response developed by 2-weeks post-immunization, and the maximal antibody production was observed by 6-weeks post-immunization. The average affinity of anti-TNP serum antibody increased consistently and reached the maximum by 10-weeks post-immunization. The increase of antibody affinity beyond the point of optimal antibody titer revealed that the affinity maturation of IgM antibody response occurred in channel catfish. Dissection of dynamics of individual affinity subpopulations indicated that a significant proportion of low affinity subpopulations appeared at early response, and high affinity subpopulations appeared predominantly at later, resulting in a 100-fold increase in affinity over response. Additional, TNP+ IgM+ ASCs was detected by 2-weeks post-immunization and achieved the maximal number by 6-weeks post-immunization. Using an inhibition ELISPOT assay, the findings of a consistent increase in the average affinity of secreted IgM antibody by peripheral blood ASCs, as the immune response progressed, confirmed the occurrence of the affinity maturation. Taken together, the results of this study indicated that affinity maturation occurred in channel catfish following immunization with a TD antigen TNP-KLH.

SUPPRESSION OF IgE ANTIBODY RESPONSE IN MICE BY A POLYSACCHARIDE, AZ9, PRODUCED BY KLEBSIELLA OXYTOCA STRAIN TNM3

ABSTRACTA soil bacterium, Klebsiella oxytoca TNM3 was found to produce a polysaccharide named AZ9 that shows suppressive effects on IgE antibody response in mice. When mice were administered with 50∼ 100 mg/kg AZ9 subcutaneousely for 4 consecutive days after immunization with trinitrophenyl (TNP)-keyhole limpet hemocyanin, anti-TNP IgE production was significantly suppressed, while the level of anti-TNP IgM was affected marginally. In AZ9-administered mice, IL-4 secretion from splenic cells was reduced to ∼30% of the untreated control. Thus, AZ9 suppression of IgE production may be due to attenuating effects on the Th2-type response. Although oral administration of AZ9 alone had no effects on IgE production, ovalbumin (OVA)-induced oral tolerance of anti-TNP IgE response to TNP-OVA was markedly augmented when a suboptimal dose of OVA was administered orally in combination with AZ9. Collectively, our data suggest that AZ9 has beneficial suppressive effects on IgE-dependent allergic responses.

Duck antibody responses to keyhole limpet haemocyanin, human immunoglobulin G and the trinitrophenyl hapten. Evidence of affinity maturation

White Pekin ducks (three in each of four groups) received trinitrofluorobenzene (TNP) conjugated to keyhole limpet haemocyanin (KLH) or human immunoglobulin G (HIgG) either incorporated into adjuvants (complete Freund's adjuvant (CFA), then incomplete Freund's adjuvant (IFA) followed by intravenous (i.v.) injection) or by repeated i.v. injection. A rabbit received TNP-KLH with CFA, IFA, then i.v. Antibody (Ab) responses to the carrier proteins and to the TNP hapten were monitored by enzyme-linked immunosorbent assay (ELISA), optimized for use with duck Abs, and the response to TNP was also assessed in an ELISA designed to detect shifts in Ab affinity (Ka). The rabbit mounted good Ab responses to KLH and TNP, the Ka of the anti-TNP response increasing from 105.99 to 108.87 l/mol (758-fold) during the experiment. Ducks showed weak Ab responses to KLH and HIgG, with weakest responses among ducks receiving antigen by the i.v. route exclusively. The duck anti-TNP responses were vigorous in all cases. However, following i.v. administration of antigens (Ags), the anti-TNP responses were transient, being strong by 7 days but diminishing to background levels by 14 to 28 days after each inoculation. The duck Ab responses to TNP displayed affinity maturation. In ducks receiving Ags with adjuvants, Ka increased by levels comparable with or even exceeding those in the rabbit. Affinity increases were less, but nevertheless apparent, following i.v. administration of Ag. These results show that, contrary to expectation, the duck Ab response experiences affinity maturation. They also point to the transience of the duck Ab response to selected haptens/epitopes when Ag is not delivered in adjuvant. These findings have clinical implications for field responses to pathogens and vaccines.

Hapten-induced model of murine inflammatory bowel disease: mucosa immune responses and protection by tolerance.

We report here a murine model for experimental chronic colitis where administration of trinitrobenzene sulfonic acid (TNBS) in 50% ethanol induced inflammation of large intestine in susceptible (C3H/HeJ and BALB/c) but not resistant (C57BL/6 and DBA/2) mouse strains. We queried whether mucosal trinitrophenyl (TNP)-specific B cell responses were induced in mice with TNBS-induced colitis, and if induction of tolerance to TNBS by oral administration of this hapten protected mice from development of colitis. Isotypes and subclasses of polyclonal and TNP-specific Ab-forming cells (AFC) were assessed in mucosal and peripheral lymphoid tissues of C3H/HeJ mice with TNBS-induced colitis. Increased numbers of IgA- and IgG-secreting cells were found in the inflamed colon lamina propria. Inflamed colonic tissue also contained high frequencies of IgG anti-TNP AFC (predominantly of IgG1, IgG2a, and IgG2b subclasses); however, anti-TNP responses in noninflamed mucosal tissues of mice with colitis exhibited dominant IgA and IgM with low IgG anti-TNP responses. CD4+ T cells stimulated with TNP-splenocytes produced more IFN-gamma and less IL-4, suggesting a Th1-type response. Oral administration of TNBS before induction of colitis markedly decreased mucosal anti-TNP responses and completely inhibited anti-TNP IgG2a and IgG2b responses. Control mice did not show inhibition of anti-TNP AFC responses or TNBS-induced colitis. Intracolonic sensitization of susceptible C3H/HeJ mice with TNBS induces a localized IgG anti-TNP B cell response in the inflamed tissue, whereas prior oral administration of TNBS results in unresponsiveness to this agent and protects mice from development of TNBS-induced colitis.

Cross-reactivity of IgG and IgM secreting B cells in normal and autoimmune mice.

Antibodies from animals with lupus react with a diverse array of foreign and self antigens. This has been attributed to the activation of large numbers of mono-specific B cells, although it could result from the stimulation of a smaller population of highly cross-reactive lymphocytes. To examine this issue, a chamber ELIspot assay was used to compare the cross-reactivity of Ig-secreting B cells from autoimmune vs normal mice. The cross-reactivity of IgM-secreting lymphocytes was significantly greater than that of IgG-secreting cells in normal BALB/c and DBA/2 mice. For example, 5-23% of IgM but only 0.1%-0.4% of cells secreting IgG antibodies against DNA or TNP-BSA were cross-reactive. When normal animals were immunized and boosted with TNP-KLH, the resultant IgG anti-TNP response was essentially mono-specific. There was no significant difference in the proportion of lymphocytes producing cross-reactive IgM antibodies in normal vs autoimmune mice. In contrast, the frequency of cross-reactive IgG-secreting B cells in lupus-prone MRL-lpr/lpr and (NZB x NZW)F1 animals was at least 15 times higher than in normal mice. These observations indicate that the repertoire of IgG-secreting cells in active lupus differs qualitatively from that seen spontaneously or following antigenic stimulation in normal animals.

Lipopolysaccharide from Brucella abortus behaves as a T-cell-independent type 1 carrier in murine antigen-specific antibody responses.

In order to determine the carrier nature of lipopolysaccharide from Brucella abortus (LPS-BA) in evoking humoral responses, normal and immunodeficient mice were immunized with trinitrophenyl (TNP)-conjugated LPS-BA (TNP-LPS-BA) and the responses were compared with those to known T-dependent and T-independent antigens. TNP-LPS-BA, like T-independent type 1 (TI-1) antigens such as TNP-BA and TNP-LPS from Escherichia coli (TNP-LPS-EC), generated anti-TNP responses in BALB/c, athymic BALB/c nu/nu, and CBA/N mice. In contrast, N-2,4-dinitrophenyl-beta-alanylglycylglycyl-substituted keyhole limpet hemocyanin, a typical T-dependent antigen, was not immunogenic in athymic mice, and TNP-Ficoll (T-independent type 2) was ineffective in eliciting humoral responses in CBA/N mice. These results indicate that LPS from B. abortus acts as a TI-1 carrier in generating antibody responses. In C3H/HeJ mice, TNP-LPS-BA generated higher-titer immunoglobulin G1 (IgG1), IgG2a, and IgG2b anti-TNP antibodies than TNP-LPS-EC. Compared with those from BALB/c mice, pure resting B cells isolated from C3H/HeJ mice exhibited a 30-fold lower proliferative response to LPS-EC, whereas the LPS-BA response was reduced to a lesser extent (5-fold). This suggests that the disparity observed in antibody titers was due to different abilities of LPS from B. abortus and E. coli to stimulate C3H/HeJ B cells. The ability of LPS from B. abortus to act as a carrier in generating humoral immune responses indicates that LPS-BA can be substituted for whole B. abortus organisms in vaccine development.

Antigen-specific but not polyclonal IgE response in murine B cells cocultured with Th2 clone is refractory to suppression by IL4-depletion.

We established culture conditions allowing the induction of antitrinitrophenyl (TNP) IgE response by the coculture of unprimed C3H B cells with conalbumin (CA)-specific type 2 helper T cell clone, D10.G4.1 in the presence of TNP-CA. With the help of this culture system, the role of Th2 cells and interleukin 4 (IL4) derived from them in the induction of antigen-specific IgE response was analyzed. A maximum level of anti-TNP IgE was secreted at 0.5-1 micrograms/ml TNP-CA. When the antigen was increased to 100 micrograms/ml, anti-TNP IgE production was abolished, while antigen-nonspecific (polyclonal) IgE was produced concomitantly. Anti-TNP IgE synthesis was not significantly suppressed in the presence of monoclonal anti-IL4 (11B11), nor was the response further enhanced by the addition of recombinant IL4. In contrast, polyclonal IgE response was abolished by 11B11. In order to deplete endogenous IL4 more strictly, we employed an antisense DNA for IL4 mRNA that can effectively inhibit IL4 production from D10 cells. Even in the presence of both 11B11 and the antisense DNA, it was found that anti-TNP IgE response was not suppressed significantly. These results suggest that antigen-specific IgE response induced in B cells cocultured with established Th2 cells does not depend on IL4 in contrast to polyclonal IgE response.

In vitro antigen-specific IgE response is refractory to suppression by interferon-gamma.

Although interferon (IFN)-gamma has been shown to be involved in the down-regulation of polyclonal IgE response in murine B cells that were activated by lipopolysaccharide (LPS) and interleukin 4 (IL4), effects of IFN-gamma on antigen-specific IgE responses have not been fully investigated. We have developed the following culture systems for inducing antigen-specific IgE responses in murine lymphocytes, and examined the effects of IFN-gamma on the following responses in vitro. (1) Anti-trinitrophenyl (TNP) IgE response induced by the stimulation with TNP-keyhole limpet hemocyanin (KLH) of BALB/c spleen cells that had been primed in vivo with the same antigen. (2) Anti-TNP IgE response induced by the coculture of unprimed C3H B cells with conalbumin (CA)-specific helper T cell clone, D10.G4.1, in the presence of TNP-CA. The former anti-TNP IgE response was not suppressed, and the latter suppressed only partially (less than 30%) by the addition of 100-200 U/ml IFN-gamma. In contrast, polyclonal IgE response in murine B cells that were stimulated by LPS and IL4 was abolished by 10 U/ml IFN-gamma. These results indicate that IgE production from antigen-stimulated B cells, in contrast to those activated polyclonally, are refractory to direct suppression by IFN-gamma.

Dextran-conjugated anti-Ig antibodies as a model for T cell-independent type 2 antigen-mediated stimulation of Ig secretion in vitro. I. Lymphokine dependence.

We have previously demonstrated that dextran-conjugated anti-IgD antibodies (anti-delta-dex) stimulate high levels of B cell proliferation at concentrations that are 1000-fold lower than that required by unconjugated anti-Ig. We now show that anti-delta-dex may provide a suitable model to study Ig secretion stimulated by soluble T cell-independent type 2 Ag exemplified by TNP-Ficoll. Thus, both TNP-Ficoll and anti-delta-dex stimulate low to undetectable levels of Ig secretion when cultured with resting B cells. Addition of IL-5 or IL-2 stimulated enhanced anti-TNP responses in the presence of TNP-Ficoll, or induced polyclonal Ig secretion in the presence of anti-delta-dex. Both TNP-Ficoll and anti-delta-dex conjugates stimulated Ig production by Percoll-separated low density (partially activated) B cells in the absence of added lymphokines. These findings point to the similarities in the activation requirements of TNP-Ficoll and anti-delta-dex and suggest that dextran-anti-Ig conjugates, which can induce B cell activation irrespective of Ag specificity, may provide a useful model for studying various parameters that characterize the responses to soluble TI type 2 Ag.

Influence of the Ah locus on the humoral immunotoxicity of 2,3,7,8-tetrachlorodibenzo- p-dioxin: Evidence for Ah-receptor-dependent and Ah-receptor-independent mechanisms of immunosuppression

There are conflicting reports in the literature regarding the role of the Ah locus in 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) immunotoxicity. The present studies have utilized two congenic strains of C57Bl 6 mice that differ only at this locus to assess its influence on TCDD-induced suppression of antibody responses. Mice were given a single oral dose of TCDD 2 days prior to challenge with sheep red blood cells (SRBC) or trinitrophenyl-lipopolysaccharide (TNP-LPS). The subsequent dose-dependent effects of TCDD on the amount of antibody produced by splenic plasma cells were measured using the hemolytic antibody isotope release assay. In addition, the relative importance of the Ah genotype of lymphoid versus nonlymphoid tissue was examined in adoptive transfer experiments. Aryl hydrocarbon hydroxylase (AHH) activity was significantly induced in Ah bb mice by a dose of 0.5 μg/kg TCDD and maximally induced by a dose of 2 μg/kg. Ah dd mice required 10-fold higher doses of TCDD to induce comparable levels of AHH. The degree of thymic involution and liver hypertrophy induced by TCDD was also influenced by the Ah genotype of the animals. Both Ah bb and Ah dd mice exhibited dose-dependent suppression of the anti-TNP response following TCDD exposure. The ID50 was 7.0 μg/kg in Ah bb mice and 30.8 μg/kg in Ah dd mice. Suppression of the antibody response to SRBC was also dependent on the Ah locus. The ID50 in Ah bb mice was 0.6 μg/kg TCDD. However, an apparent biphasic dose response for suppression of the anti-SRBC response in Ah dd mice suggested the involvement of an Ah-independent component of suppression as well. In adoptive transfer studies, lymphocytes were identified as an Ah-dependent component of the response. The Ah-independent component of the response was not identified, and could be either lymphoid or nonlymphoid in nature. The possibility that T helper cells represent the Ah-independent component is discussed.

Primary antibody responses to thymus-independent antigens in the lungs and hilar lymph nodes of mice

B lymphocytes from the pulmonary lymphoid tissues were stimulated with a variety of thymus-independent (TI) antigens by intratracheal (i.t.) immunization. Immune responses in the lungs and hilar lymph nodes (HLN), which are part of the localized lymphoid tissue, as well as in the spleen, the systemic lymphoid organ, were studied. Thus, primary i.t. immunization of mice with the TI-1 antigen trinitrophenyl-lipopolysaccharide (TNP-LPS) elicited both antigen-specific and polyclonal plaque-forming cell responses from HLN, lung, and splenic B lymphocytes. These responses appeared as early as 3 days after immunization and declined by day 7. Similar immunization with another TI-1 antigen, TNP-Brucella abortus, resulted in anti-TNP responses in both pulmonary and systemic lymphoid tissues, although the kinetics of the antibody response were different than those to TNP-LPS. Interestingly an i.t. immunization with a TI-2 antigen, TNP-Ficoll, failed to induce an anti-TNP PFC response from HLN and lung B cells, although there was good antibody formation from splenic B cells. Antibody response to TNP-Ficoll was restored in pulmonary tissues when mice were immunized with TNP-Ficoll mixed with unconjugated B. abortus. In conclusion, our results indicate that TI-1 and TI-2 antigens differ in their ability to induce antibody responses in the pulmonary lymphoid tissues. The inability of TNP-Ficoll to elicit an antibody response in pulmonary lymphoid tissues has significance in the development of vaccines containing bacterial polysaccharides.

Humoral immunity in the ewe 1. The influence of adjuvancy and immunogenic substitution on immune reactivity following immunisation with protein antigens

The nature of antigen and presence of adjuvant are major factors which influence the level of immune reactivity following immunisation. This study examines the quantity and isotype of antibody produced in ewes immunised with different proteins in combination with different adjuvants. Results using indirect ELISA assays show that animals immunised with keyhole limpet haemocyanin (KLH) in adjuvant produced lower levels ( P < 0.05) of immunoglobulin M (IgM) and had increased levels and persistence of immunoglobulin G1 (IgG1) compared with ewes immunised with antigen in saline ( P < 0.005). The anti-KLH immunoglobulin G2 (IgG2) titre was significantly higher in animals given oil-emulsion adjuvant than all other groups ( P < 0.005). Ewes were also immunised with bovine serum albumin (BSA) or BSA haptenated with trinitrophenyl (TNP) for the primary injection and carrier BSA alone for the secondary inoculation. Chemical haptenation of BSA antigen reduced primary IgM ( P < 0.005), IgG1 ( P < 0.005) and IgG2 ( P < 0.005) levels compared with animals immunised with pure BSA. There was an increased secondary anti-BSA IgM response in all animals first immunised with TNP-BSA ( P < 0.05). The class of anti-hapten antibody produced to TNP determinants was influenced by the degree of TNP haptenation of the carrier BSA. Mid-range molar ratios of TNP produced the strongest IgM, IgG1 and IgG2 anti-TNP responses compared with all other groups ( P < 0.01).

Effects of interleukin 4 on an antigen-specific IgE response in vitro in murine lymphocytes

The present study shows that interleukin 4 (IL4) can exert either stimulatory or inhibitory effects on an antigen-specific IgE response in vitro. Spleen cells from BALB/c mice that had been primed with trinitrophenyl keyhole limpet hemocyanin (TNP-KLH) were cultured with the same antigen for 2 days, washed, transferred to an antigen-free culture medium, and then cultured for 4 days. Anti-TNP IgE antibodies secreted into the medium were determined by an enzyme immunoassay developed in our laboratory. Anti-TNP IgE response was elicited by TNP-KLH in cultures of the spleen cells from mice that had been primed twice with the antigen at an interval of 3 weeks, but not in cells from animals that had received only a single injection of the antigen. When the latter cells were cultured with the antigen in the presence of mouse recombinant IL4, a considerable level of the antigen-specific IgE response was induced. In contrast, the IgE response elicited in the cells of mice that had been primed twice with the antigen was markedly down-regulated by added IL4. These observations suggest a novel function of IL4 in the regulation of IgE antibody responses.

Total lymphoid irradiation reduces IgG autoantibody production and enhances specific antibody responses in NZB/NZW F1 mice

Thymus-independent primary antibody responses were studied in young and old (9 months) untreated and TLI-treated NZB/NZW and BALB/c mice. Untreated old NZB/NZW mice had a low primary response to Brucella abortus (BA) as compared to that of young NZB/NZW and BALB/c mice. However, TLI treatment resulted in a 130-fold increase in the IgG anti-BA primary antibody response at day 21 postimmunization, achieving similar levels to those of young NZB/NZW or nonautoimmune BALB/c mice. Anti-TNP responses to trinitrophenylated BA or Ficoll were masked by high background levels of anti-TNP antibodies. Despite the increase in the anti-BA response, spontaneous immunoglobulin secretion and autoantibody levels were markedly decreased after TLI in old NZB/NZW mice.

Self versus non-self carrier in primary and secondary anti-TNP B cell response: I. specificity of help

Humoral anti-hapten responses are supposed to require carrier-specific help. Yet, «TNPspecific» help can be activated with TNP coupled to syngeneic lymphocytes. To further clarify the role of hapten-specific vs. carrier-specific helper T cells (T H) in the humoral anti-TNP response, BALB/c mice were immunized with TNP coupled to cellular or soluble self or nonself carriers, we analyzed primary and secondary B cell responses as well as the apparent specificity of help. TNP bound to syngeneic red blood cells (sRBC), syngeneic albumin (sA) and syngeneic polyclonal or monoclonal immunoglobulin (s1g/smIg) and TNP coupled to non-self carriers initiated equivalent primary anti-TNP responses. On the other hand, a secondary anti-TNP response was only obtained with heterologous carriers or with smIg. Even with heterologous carriers or with smIg, the magnitude of the secondary anti-TNP response exceeded only slightly the amplitudes of a primary anti-TNP response. Furthermore, if animals were challenged shortly after priming, they appeared un/-hyporesponsive. In vitro analysis revealed that in the primary as well as in the secondary anti-hapten response, TNP-specific T H were involved, i.e., the primary response to s1g, sA and sRBC was exclusively due to TNP-specific help, the response after priming with TNP-smIg or TNPheterologous carrier was due to the additive effects of carrier- and TNP-specific help. Since «carrier-specific» help with smIg was independent of the antibody specificity as well as of the Ig class, we suppose that sm1g activated idiotype-specific TH, which functioned like «carrier» T H. Two mechanisms were responsible for the low magnitude of the secondary anti-hapten response: competition for carrier-specific help (using heterologous carriers), antibody (AB)-dependent suppression.

Antigen-induced and polyclonal B-cell responses in human peripheral blood lymphocyte cultures

This work was designed to delineate the anti-hapten antibody (Ab) response induced by trinitrophenol-polyacrylamide (TNP-PAA) beads from the nonspecific B-cell response which concomitantly occurs in human peripheral blood mononuclear cell (PBMC) cultures. Indeed human PBMC produce consistent amounts of immunoglobulins when cultured at high cell density in the presence of fetal bovine serum, regardless of the presence of antigen. In contrast, the stimulation of such cultures by TNP-PAA leads to an Ab response characterized by the following: cells secreting anti-hapten Ab at a high rate (detected by a plaque-forming cel (PFC) assay); a 10–30 times enhancement in the number of hapten-specific binding cells (detected by a rosetteforming cell (RFC) assay); the production of anti-TNP IgM Ab (detected by an ELISA assay). The anti-TNP response is specifically triggered by the particulate antigen, as shown by the following: (i) The TNP-PAA antigen induces a clear-cut increase in the amount of anti-TNP Ab whereas it only marginally increases that of total IgM. (ii) The anti-TNP Ab response is specifically abolished when anti-TNP RFC are depleted from the PBMC preparation before the initiation of the cultures. (iii) The anti-TNP Ab response is specifically abolished when PBMC are triggered by TNP-PAA in the concomitant presence of a soluble TNP-protein conjugate. These results demonstrate the ability of polymeric antigens to specifically activate human peripheral blood B cells.

Monoclonal antibodies to the human C3b/C4b receptor (CR1) enhance specific B cell differentiation.

The addition of monoclonal antibodies against the human C3b/C4b receptor (CR1) to cultures of peripheral blood lymphocytes in the presence of suboptimal amounts of TNP bound to polyacrylamide beads enhanced by 150 to 400% the specific anti-TNP response, as measured by a plaque-forming cell assay on day 7. Anti-CR1 antibodies similarly enhanced the anti-fluorescein antibody response. Enhancement only occurred in cultures performed in the presence of the relevant antigen. No enhancing effect on the anti-TNP response was observed on addition to cultures of monoclonal antibodies directed against other surface antigens of B cells or an anti-T cell antibody of the same subclass as that of anti-CR1 antibodies. Anti-CR1 antibodies alone did not induce nonspecific B cell proliferation and did not provide B cells with a first signal for proliferation in the presence of a source of B cell growth factors. Anti-CR1 antibodies did not enhance the nonspecific proliferative response of B cells to growth factors derived from PHA-stimulated T cells, semi-purified BCGF 20 KD, BCGF 50 KD, or recombinant IL 2 in the presence of anti-mu. In this respect, the effect of anti-CR1 antibodies differs from that of anti-CR2 antibodies which interact with early stages of B cell activation. In contrast, anti-CR1 antibodies enhanced specific differentiation of antigen-activated B cells in the absence of T cells when soluble T cell factors were provided. Similar results were obtained by using either of two sources of differentiation factors, the MLA-144 supernatant or a 30 to 15 KD fraction from PHA-stimulated T cells. These results indicate that triggering of CR1 on B cells positively regulates the specific antibody response to low doses of antigen by enhancing B cell differentiation whether T cell help is provided by intact T cells or by T cell-derived differentiation factors.

Enhancement of a human antibody response in vitro mediated by interaction of interferon-alpha with T lymphocytes.

This study describes the effects of human recombinant IFN-alpha 2 on antibody production in vitro. Whereas the inclusion of IFN-alpha 2 in cultures for 7 days had a relatively minor effect on pokeweed mitogen (PWM)-induced antibody production, it resulted in a dose-related enhancement of a hapten-specific primary antibody response. Comparison of PWM and IFN-induced [3H]thymidine uptake indicated that the observed IFN activation was not polyclonal. Pretreatment of T cells with IFN for 1 hr before recombination with untreated autologous B lymphocytes increased the anti-TNP response four-fold, whereas similar pretreatment of B lymphocytes had no effect. Furthermore, 2000 R x-irradiation of T cells before coculture with autologous B lymphocytes and IFN abrogated the TNP-specific response. These results indicate that IFN modulates TNP-specific antibody production via a radiosensitive T-helper function. Further subfractionation by panning suggests that the enhancement is mediated by the Leu-3a+ helper/inducer T cell subset. Evidence that a 1-hr exposure to IFN was sufficient to modulate antibody production prompted the examination of T cells for possible receptor mechanisms. Scatchard analysis of 125I-IFN-alpha 2 binding revealed approximately 65 high affinity IFN receptors per cell with an apparent dissociation constant (Kd) of 4.4 X 10(-10) M. This paper is the first demonstration of the role of T cells in mediating the effects of recombinant IFN-alpha 2 on human primary antibody responses in vitro. These data further suggest that the observed modulation of hapten-specific antibody production in vitro by IFN may involve the binding of IFN to specific cellular receptors expressed by T lymphocytes.

Immunoresponses to Neisseria meningitidis epitopes: Immunodulation by meningococcus B acts on more than one meningococcal surface epitope

Neisseria meningitidis group B strain M986 (serotypes 2a, 7) (NMB) elicits a specific primary antiphosphorylcholine immune response in mice but not a secondary response. The ability of other serotype and serogroup meningococci to induce similar primary responses in mice was studied, as was the immunogenicity of trinitrophenyl coupled NMB (TNP-NMB) in primary and secondary antitrinitrophenyl responses. Except for NMB, all other strains tested (three serogroup B and one serogroup A meningococcal strains) were found to be very poor phosphorylcholine immunogens. TNP-NMB, however, though proving to be a very good TNP antigen, was only a weak phosphorylcholine antigen. Priming NBF1 female mice with TNP-NMB one month or more before challenging them with the same antigen induced a strong depression of anti-TNP response in the subsequent challenge. However, this effect was not observed with Xid NBF1 male mice. Furthermore, priming mice with NMB weakly affected the anti-TNP response, but greatly depressed the antiphosphorylcholine response, after TNP-NMB challenge. In addition, whereas apparently only one TNP-specific B cell subpopulation was responding in unprimed mice challenged simultaneously with TNP-NMB and TNP-Ficoll (non-additive response), priming mice with NMB appeared to facilitate the independent activation of two different TNP-specific B cell subpopulations (additive response).

Recombinant DNA produced human IL-2, injected in vivo, will substitute for carrier priming of helper function in the South African clawed toad, Xenopus laevis

The studies included in this report tested the ability of a recombinant DNA produced human lymphokine (rIL-2) to serve as a substitute for the carrier priming required for the in vivo generation of anti-hapten responses, in young adults of Xenopus laevis, the South African clawed toad. Heterologous erythrocyte carriers were used. Immune reactivity was monitored with hapten-specific antigen binding cells (ABC) in spleen cell suspensions, 8 days after injection i.p. of trinitrophenylated sheep erythrocytes (TNP-SRBC) or rIL-2 or the two together. No substantial anti-hapten responses were generated by either TNP-SRBC or rIL-2 alone. However, a greatly (9x) enhanced anti-TNP response was produced after co-injection. By varying the time between injection of the rIL-2 and the TNP-SRBC challenge, it was found that the effectiveness of this mediator would last for up to three hours in this animal. When rIL-2 was co-injected with an optimal priming dose of SRBC, two days prior to TNP-SRBC challenge, it did not enhance the level of helper function generated by carrier priming. Therefore, it is likely that the rIL-2 acts on the same cells affected by carrier priming which is maximally efficient in this regard. Thus, cells, which participate in the generation of helper function in this primitive species, are sensitive to human rIL-2. The universality of an immune regulatory molecule of this kind is considered.