A human leucocyte antigen-DR1 transgene confers susceptibility to experimental allergic encephalomyelitis elicited by an epitope of myelin basic protein.

Much evidence now indicates that human leucocyte antigen (HLA) class I and class II transgenic (Tg) mice can be of value in analysing HLA-restricted presentation of T-cell epitopes relevant to experimental models of autoimmune diseases. One area where this has been applied is the characterization of myelin epitopes presented by HLA class II molecules in experimental model of multiple sclerosis (experimental allergic encephalomyelitis (EAE)). As a first step towards humanized disease models in HLA Tg mice, we have analysed immune response of lymph node cells of HLA-DR1 Tg mice immunized with the human myelin basic protein (MBP) peptides 13-33, 87-106 and 139-154 bound by HLA-DR1. We report here that HLA-DR1 Tg mice display a hierarchy of response in vivo and in vitro to MBP epitopes depending on the binding affinity to DRB*0101 molecule. In fact, the 13-33 epitope induced a strong T helper 1 (Th1) response accompanied by high T-cell precursor frequency and caused mild EAE, while the two other epitopes gave poor (139-154) or no disease (87-106), and these data correlate with in vitro Th1 response. These data could prove a useful tool in understanding the role played by different MBP epitopes in EAE.

Induction and tolerization of anti-male CD8+ cytotoxic T lymphocytes by in vivo immunization with an H-Y-derived peptide.

We have analyzed the immune response induced by a 9mer synthetic peptide derived from the male histocompatibility antigen H-Y and containing Db-binding motifs in C57BL/6 mice. In this study we report that a single, subcutaneous injection of the peptide emulsified in IFA gave rise to the development of male-specific CD8+ T cells which displayed H-Y-specific proliferative response in vitro and showed a Tc1-type pattern of cytokine production (i.e. they secreted IFN-gamma and IL-2, but not IL-4 and IL-10). Development of a strong cytotoxic activity required in vitro stimulation with specific peptide and IL-2: under these culture conditions, we were able to generate potent CD8+ CTLs that lysed both male cells and peptide-pulsed female cells. Continuous administration of soluble peptide, delivered over a 7-day period by a mini-osmotic pump implanted subcutaneously, inhibited proliferative and cytotoxic responses and IFN-gamma production in lymph node cells from C57BL/6 mice subsequently primed with peptide in adjuvant. This decreased responses were associated with a strong increase in the secretion of IL-4 by antigen-specific CD8+ T lymphocytes. Subcutaneous administration of the H-Y-peptide in adjuvant significantly accelerates rejection of male skin graft, while continuous administration of peptide in soluble form did not modify the time course of rejection.

Gluten stimulation induces an in vitro expansion of peripheral blood T gamma delta cells from HLA-DQ2-positive subjects of families of patients with celiac disease.

The intestinal gluten sensitivity formally known as celiac disease (CD) is characterized by an evident involvement of local immune response and it is associated with the expression of HLA-DQ2 allele. The major role in the disease seems to be played by the T lymphocyte population bearing gamma delta T cell receptor (T gamma delta cells) which are increased both in peripheral blood and intestinal mucosae of celiac patients. In this paper data on the effects of in vitro gluten stimulation on lymphocytes expressing the T gamma delta phenotype are reported. Gluten seems to be able to induce the expansion of the T gamma delta cell population both in CD patients and their HLA-DQ2-positive asymptomatic relatives, in spite of the absence of clinical evidence of the disease. In addition, the evaluation of gluten-induced cytokine production shows that interleukin-4 could be implied in the early phases of pathogenesis of CD.

Development of hapten-induced IL-4-producing CD4+ T lymphocytes requires early IL-4 production by alphabeta T lymphocytes carrying invariant V(alpha)14 TCR alpha chains.

This paper investigates the mechanisms responsible for the generation of IL-4-producing CD4+ T cells during contact sensitization with the hapten trinitrochlorobenzene (TNCB). Lymph node cells taken 1 day after immunization spontaneously released IL-4 while lymph node cells taken 2 and 3 days after immunization did not produce IL-4. A second wave of IL-4 production that was both antigen-specific and MHC class II (I-A)-restricted was observed 4 days after immunization. The spontaneous release of IL-4 at day 1 was due to the alphabeta+ double-negative (CD4- CD8-) T lymphocytes that also expressed NK1.1 and showed V(alpha)14 rearrangement, while alphabeta+ CD4+ T lymphocytes were the source of the antigen-specific IL-4 production at day 4. Early IL-4 production was required for the development of IL-4-producing CD4+ T cells as mice injected with anti-V(alpha)14 or anti-IL-4 mAb produced little IL-4 and IL-10, while production of IFN-gamma was increased approximately 2-fold. These results indicate that the development of IL-4-producing CD4+ T lymphocytes in the TNCB system requires early production of IL-4 by alphabeta+ double-negative cells carrying invariant V(alpha)14 TCR alpha chain.

The effect of cyclosporin A, FK506 and rapamycin on the murine contact sensitivity reaction.

We have evaluated the effects of three potent immunosuppressive agents, cyclosporin A (CsA), FK506 and rapamycin, on the murine contact sensitivity (CS) reaction to the hapten trinitrochlorobenzene. Development of CS reaction requires participation of three distinct T cell subsets: alphabeta+, CD4+ T lymphocytes, which are the classical effector cell of the CS reaction, gammadelta+ T lymphocytes, and alphabeta+, double-negative (CD4- CD8-) T lymphocytes that express the B220 molecule and produce IL-4. We found that all three drugs inhibit the development of the CS reaction, but they affect different target cells. In fact, rapamycin and FK-506 block both alphabeta+, CD4+ and gammadelta+ T lymphocytes, while CsA inhibits only the alphabeta+, CD4+ T lymphocyte. None of the three drugs exerted any inhibitory activity on the alphabeta+, double-negative (CD4- CD8-) T lymphocytes. Hapten-immune lymph node cells from mice treated in vivo with CsA or FK506 failed to proliferate and to produce IL-2 when re-exposed to the specific antigen in vitro. In contrast, immune lymph node cells from mice that had been treated in vivo with rapamycin gave optimal antigen-specific proliferation and IL-2 production in vitro. The implications of these observations are discussed in relation to the use of these immunosuppressive agents for prevention of allograft rejection.

Major histocompatibility complex regulation of cytokine production.

This review describes the phenomenon of the major histocompatibility complex (MHC) control of cytokine production both in experimental animals and in humans. H-2 (mouse MHC) regulates which type of cytokine is selectively produced in response to the hapten trinitrophenyl (TNP). T cells from TNP-immune H-2k mice produce interferon-gamma (IFN-gamma), interleukin-2 (IL-2), IL-3, IL-5, tumor necrosis factor-alpha (TNF-alpha), IL-10, and very low levels of IL-4 on reexposure to the specific antigen in vitro. By contrast, T cells from H-2d mice produce IL-3, TNF-alpha, IL-10, and IL-4 but very low levels of IL-2, IL-5 and IFN-gamma. As MHC-congenic matched strains (BALB/k and BALB/c) are used, this makes it unlikely that non-MHC genes influence the class of response observed. A similar pattern of haplotype regulation of cytokine production is observed in humans. In fact, peripheral blood mononuclear cells from HLA-B8,DR3-positive and negative individuals differ in their ability to produce IL-2, IL-5, and IFN-gamma on stimulation with the mitogen phytohemagglutinin while producing similar amounts of IL-4, IL-6, and IL-10. The following main considerations emerge from these observations. The MHC/peptide complex generated after antigen immunization, indicates which class of cytokine production is preferentially induced and, therefore, the outcome of the immune response. Furthermore, MHC genotype may affect cytokine production (and then immune responses) by completely different mechanism(s), that is, by an antigen-nonspecific control that does not depend on the ability of MHC molecules to bind in different ways the different peptides. Accurate control of the functional repertoire of an immune response is a critical parameter in response to infections as well as in immunopathology. MHC control of the class of the immune response at the level of cytokine production is a sophisticated way in which this occurs. This control might be involved in adaptive immune responses to infections as well as in immunopathology.

Control mechanisms of cell-mediated reactions.

One of the most relevant aspects of tumor adoptive immunotherapy is provided by clinical trails on transfer of cytotoxic cells (LAK and TIL). However, LAK cell therapy is effective in a small number (16-22%) of only certain tumors, while therapy with TIL cells is efficient in about 40% of melanomas. Several possibilities have been raised to explain the low efficacy of cytotoxic cells in tumor therapy, amongst which are the poor immunogenicity of tumor and tumor-induced immunodepression. Furthermore, the possibility that cytotoxic cells do not reach the tumor site in adequate numbers has to be considered. We have developed an experimental system to study the ability of antigen-specific T cells to reach the target antigen in the tissues. The results obtained demonstrate that gamma delta cells and IL-4 are required to allow tissue localization of antigen-specific alpha beta cells, thus indicating that their ability to exert certain effect or functions requires cooperation by other cells types. These results may be relevant to the understanding of the mechanisms leading to localization of immunologically active cells at a tumor site.

IL-5 enhances in vitro and in vivo antigen-specific IgA production in MHC genetically determined low IL-5 responder mice.

Lymphonode cells from BALB/k mice, but not from BALB/c mice, immunized with picryl chloride (PCl) produce IL-5 when stimulated with the specific antigen in vitro and this correlates with picryl-specific IgA levels in vivo, which are 6 to 10 times higher in BALB/k mice. B lymphocytes from BALB/k mice cultured with PCl-immune T cells from BALB/k produce in vivo anti-PCl-IgA, while B lymphocytes from BALB/c mice, cultured with T cells from BALB/c mice, fail to produce appreciable amounts of anti-PCl IgA, unless IL-5 is added to cultures. B lymphocytes from both strains of mice produce similar amounts of total IgA antibodies when stimulated in vitro with lipopolysaccharide. In vivo administration of IL-5 to BALB/c mice increases significantly PCl-specific IgA levels to those observed in BALB/k mice and a dose-response analysis reveals that 500 units of IL-5 was the minimal effective dose, although a small increase in PCl-specific IgA levels was observed with 100 units of IL-5. Total IgA levels were increased in both strains of mice following in vivo injection of IL-5, but no significant difference in the values was observed. Our results therefore indicate that IL-5 in vivo enhances antigen-specific IgA production in MHC-determined low IL-5 responder mice and suggest an explanation for IgA deficiency in humans.

Major histocompatibility complex control of the class of the immune response to the hapten trinitrophenyl.

This paper investigates major histocompatibility complex (MHC) regulation of the class of the immune response given in vitro and in vivo following immunization of the congenic BALB/k (H-2k) and BALB/c (H-2d) mice with the hapten trinitrophenyl (TNP). TNP-immune lymph node cells from BALB/k mice produced high levels of interferon-gamma (IFN-gamma), interleukin-5 (IL-5) and IL-2 when stimulated with TNP-antigen-presenting cells (APC) in vitro, while TNP-immune lymph node cells from BALB/c mice produced very low levels of these cytokines. No significant difference was found in antigen-specific production of IL-3, IL-4 and tumour necrosis factor-alpha (TNF-alpha). There was a strong correlation between the pattern of cytokine production in vitro and the secondary antibody production in vivo. Sera from BALB/k mice had anti-TNP IgG2a, IgG2b and IgG3 levels threefold greater, and anti-TNP IgA levels eightfold greater, than BALB/c mice. The level of specific IgG1 and IgE was only marginally raised in BALB/k mice. In contrast to these strain differences in cytokine and antibody production, there was no difference in two measures of cellular immunity: contact sensitivity in vivo and antigen-specific lymphocyte response in vitro. Our results suggest that there is a good correlation between the production of cytokines in vitro and antibody response in vivo, but not with measures of cellular immunity. Moreover, this MHC control of the class of the immune response to TNP does not fit into the T-helper type-1 (Th1)-Th2 paradigm.

Major histocompatibility complex regulation of interleukin-5 production in the mouse.

Lymph node cells of CBA (H-2k), but not of BALB/c (H-2d) mice immunized epicutaneously with picryl chloride secrete interleukin (IL)-5 when stimulated with the specific antigen in vitro. The low IL-5 production in BALB/c mice persists when either picryl chloride or the unrelated antigen oxazolone are used, when the amount of antigen in vitro is varied and when a secondary response is studied. The difference in IL-5 production maps to the major histocompatibility complex (MHC) in the congenic BALB/b, BALB/c and BALB/k mice. Furthermore, lymph node cells from (k x d) F1 mice produce IL-5 when stimulated by antigen presented on H-2k but not on H-2d antigen-presenting cells. Finally, the low IL-5 production in vitro in BALB/c mice is correlated with low picryl-specific IgA levels in vivo, which otherwise are ten times greater in CBA and BALB/k mice. The influence of MHC on IL-5 production and IgA secretion in the mouse might be a possible basis for the association of MHC with IgA deficiency in humans.

Dose-related effect of beclomethasone dipropionate on airway responsiveness in asthma.

The effects of twice daily inhaled beclomethasone dipropionate (BDP) at two dose levels (500 and 1,000 micrograms daily) on the airway responsiveness to inhaled histamine was evaluated by a randomized, single-blind, cross-over study in 10 patients with stable asthma. The 12-week study began with a 3-week run-in period of baseline treatment, which was continued unchanged throughout the study, and the two treatment periods were separated by a 3-week placebo period. Patients attended the laboratory every 3 weeks for spirometry and histamine inhalation tests to determine the provocative concentration of histamine causing a 20% fall in forced expiratory volume in 1 s (PC20 of FEV1). There was a similar significant improvement (p less than 0.05) in mean FEV1 after both treatments. There was no significant change in PC20 after treatment with 500 micrograms daily, the geometric mean being 0.587 mg/ml after the placebo period and 0.860 mg/ml after BDP treatment. There was a significant improvement in PC20 (1.930 mg/ml) after treatment with 1,000 micrograms BDP daily in comparison with the placebo and treatment periods with 500 micrograms BDP daily (p less than 0.001). These results suggest that higher doses than usual of inhaled BDP must be used to control airway responsiveness in asthmatics.

Effect of ketotifen on the bronchodilation induced by salbutamol.

Twelve subjects with stable asthma each inhaled two puffs (200 micrograms) of salbutamol on 2 separate days 3 h after double-blind oral administration of ketotifen (two 1-mg capsules) or identical placebo. FEV1 was recorded before and at intervals for 4 h after inhalation of salbutamol. Overall, the FEV1 was significantly greater during the 4-hour period after premedication with ketotifen (p less than 0.02) and the difference between the effect of placebo and ketotifen was statistically significant at 120, 180 and 240 min after salbutamol (p less than 0.05).