Biological basis of the HLA-B8,DR3-associated progression of acquired immune deficiency syndrome.

The factors influencing the evolution of human immunodeficiency virus (HIV) infection are not fully known, but the host genotype undoubtedly plays a role in determining the outcome of the disease by affecting the immune response to HIV. The role of the host human leukocyte antigen (HLA) genotype in the regulation of susceptibility to HIV infection and expression has been studied extensively in different major risk groups. Certain HLA alleles and haplotypes, being associated with aberrant immune responses independently from HIV infection, have been reported to facilitate the rapid progression of disorders related to HIV infection. Particularly, the association of rapid acquired immunodeficiency syndrome (AIDS) progression with genes from the HLA-B8,DR3 haplotype has been reported by different research groups. It is well known that this haplotype is associated in all Caucasian populations with a wide variety of diseases with autoimmune features and in healthy subjects with a number of immune system dysfunctions, as a reduced production of T helper (Th)1 type cytokine. HIV infection may act on this genetic background triggering immunopathogenetic mechanisms leading to AIDS with a dominant Th2 profile as a common feature.

Modification of cytokine patterns in subjects bearing the HLA-B8,DR3 phenotype: implications for autoimmunity.

The factors influencing the pathogenesis of autoimmune disease are not fully known, but the host genotype undoubtedly plays a role in determining the outcome of these diseases. The role of the host's major histocompatibility complex (MHC) genotype in the regulation of susceptibility to autoimmune diseases has been extensively studied in different populations, and certain HLA (the human MHC) alleles and haplotypes have been reported to be associated with several autoimmune diseases. In particular, the association with genes from the HLA-B8,DR3 haplotype has been reported by different research groups. This haplotype is associated in all Caucasian populations with a wide variety of diseases with autoimmune features, and in healthy subjects it is associated with a number of immune system dysfunctions. Mainly, peripheral blood mononuclear cells from HLA-B8,DR3-positive and -negative individuals differ in their ability to produce interleukin (IL)-2, IL-5, IL-12 and interferon-gamma upon stimulation with the mitogen phytohaemoagglutinin (PHA), while producing similar amounts of IL4, IL-6 and IL-10. Furthermore, in HLA-B8,DR3-positive subjects tumor necrosis factor alpha secretion is increased both with and without PHA stimulation. Accurate control of the functional repertoire of an immune response is a critical parameter in the 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.

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.

In vitro T cell activation in elderly individuals: failure in CD69 and CD71 expression.

A large number of T cell dysfunctions have been observed in the elderly. The most widely observed is the inability of these cells to proliferate at a level comparable to T cells from young individuals after stimulation by mitogens. To better characterize T cell impairment, we have focused on the in vitro T cell activation, analyzing by flow cytometry the activation molecules CD69 and CD71 on mitogen-stimulated lymphocytes from young and elderly subjects. The results show that the percentages of CD69+ and CD71 + T cells were significantly decreased in cultures from elderly subjects when compared to values obtained culturing cells from young individuals. The differences observed seem not due to differences in CD4 and CD8 rates in the "old' cells that underwent activation, since, following activation, the pattern of CD4 and CD8 phenotypes was the same in both groups of subjects. Signals from CD69 are relevant in controlling cytokine gene expression because its stimulation leads to interleukin-2 production and increases its receptor expression. The interaction of this cytokine with its cellular receptor is an essential requirement for T lymphocytes to express CD71 and to start proliferation. Thus, a key role in the age-associated impairment of T cell activation could be played by an ineffective modulation of CD69 expression suggesting a defect in the signal transduction pathway of the T cell receptor-CD3 complex in elderly.

In vitro impairment of interleukin-5 production in HLA-B8, DR3-positive individuals implications for immunoglobulin A synthesis dysfunction.

Healthy subjects carrying the HLA-B8,DR3 haplotype may show a large number of immune dysfunctions. Concerning T-cell dysfunctions, the most intriguing is a defect of the early phases of T-cell activation, responsible for the impairment of in vitro mitogen-stimulated cytokine production. Regarding B-cell dysfunctions, one the most fascinating topics is the association between this haplotype and IgA deficiency in healthy blood donors. Accordingly, HLA-B8,DR3-positive healthy subjects show significantly lower values of serum IgA than HLA-B8,DR3-negative ones. Because IL-5 is a stimulating factor for the secretion of IgA by committed B cells, we have analyzed the in vitro mitogen-stimulated IL-5 production by MNCs from healthy HLA-B8,DR3-positive individuals to study whether they display an impaired production of IL-5. The results clearly demonstrate that MNCs from HLA-B8,DR3-positive individuals display significant reduction of IL-5 production, suggesting that IgA synthesis dysregulation observed in HLA-B8,DR3-positive subjects could be due to an impairment of IL-5 production.

In vitro cytokine production by HLA-B8,DR3 positive subjects.

It is well known that healthy subjects carrying the HLA-B8,DR3 haplotype may show an impairment of immune system, the T cells being the most affected. To gain insight into the mechanism(s) of the impairment displayed by these subjects, efforts have been centered on the study of in vitro cytokine production because of the pivotal role played by these mediators in the activation and control of several immune functions. The available results indicate that the ability to several immune functions. The available results indicate that the ability to produce interleukin-1 (IL-1), IL-2 and the soluble form of its receptor (sIL-2R) is impaired in HLA-B8,DR3 positive healthy subjects. To better characterize the cytokine production capacity of HLA-B8,DR3 positive subjects, we have investigated the pattern of in vitro production of IL-2, sIL-2R, IL-4. IL-6 and gamma-interferon (gamma-IFN) by mononuclear cells from HLA-B8, DR3 positive subjects after phytohaemoagglutinin stimulation. A significant decrease of IL-2, sIL-2R and gamma-IFN production by HLA-B8,DR3 positive subjects was observed. No significant difference was instead found between the HLA-B8,DR3 positive subjects and the negative ones as regards IL-4 and IL-6 production. We suggest that this imbalanced cytokine production may well account for the pattern of immune response that may observed in HLA-B8,DR3 positive subjects, i.e. a normal or increased humoral response in face of a low T cell immune responsiveness.

Natural killer and lymphokine-activated killer activity in HLA-B8,DR3-positive subjects.

The haplotype HLA-B8,DR3 is over-represented in several autoimmune diseases, implying that genes predisposing people to these disorders are linked to this haplotype. In these diseases, various dysfunctions reflecting an impairment of the immune system have been found. Several reports indicate also that in HLA-B8,DR3-positive healthy subjects similar disorders may be demonstrated. In the present work, we have evaluated NK and LAK activity in these subjects. The study has been performed on monocyte-depleted peripheral blood MNCs by using, the K-562 cell line as a target for NK activity and the HL-60 cell line for as a target LAK activity. LAK cells were obtained by incubating MNCs for 3 days with 100 U/ml of rIL-2. The results of our experiments demonstrate that NK cell activity is significantly decreased in healthy subjects bearing the HLA-B8,DR3 haplotype. Since the number of circulating CD16+ cells is not significantly different between HLA-B8,DR3-positive subjects and negative ones, it is unlikely that this defect is due to a decreased number of NK cells in effector cell preparations. The observation that the treatment with rIL-2 can restore the killer activity of MNCs from these subjects suggests instead that the reduced NK activity may be due at least in part to the imbalance of cytokine network that has been demonstrated in HLA-B8,DR3-positive subjects. Finally, since a decreased NK activity has been reported in the autoimmune diseases linked to this haplotype, our results support the suggestion that immunologic changes observed in autoimmune diseases reflect systemic regulatory disorders that have a genetic basis.

Biological significance of soluble IL-2 receptor.

A NUMBER of receptors for growth factors and differentiation antigens have been found to be secreted or released by cells. Following mononuclear cell (MNC) activation and interleukin-2 receptor (IL-2R) expression, a soluble form of the Alpha;-chain of IL-2R (sIL-2R) is released. The sIL-2R has been shown to be present in the culture supernatants of activated MNCs as well as in normal sera and, in higher amounts, in sera from subjects affected by several diseases including neoplastic, infectious and autoimmune ones, and in sera from transplanted patients suffering allograft rejection. The blood sIL-2R levels depend on the number of producing cells and the number of molecules per cell, so that sIL-2R blood values may represent an index of the number and the functional state of producing cells, both normal and neoplastic. Thus, monitoring of the immune system, mostly T-cells and haematological malignancies might be targets for the measurement of sIL-2R. Since many conditions may influence sIL-2R production, little diagnostic use may result from these measurements. However, since blood sIL-2R levels may correlate with disease progression and/or response to therapy, their measurement may be a useful index of activity and extent of disease. The precise biological role of the soluble form of the IL-2R is still a matter of debate. However, we know that increased sIL-2R levels may be observed in association with several immunological abnormalities and that sIL-2R is able to bind IL-2. It is conceivable then that in these conditions the excess sIL-2R released in vivo by activated lymphoid cells or by neoplastic cells may somehow regulate IL-2-dependent processes. On the other hand, it cannot exclude that sIL-2R is a by-product without biological significance. Finally, it is puzzling that in many conditions in which an increase of blood sIL-2R values has been observed, MNCs display a decreased in vitro capacity to produce sIL-2R. These seemingly contrasting findings are discussed in the light of the data showing that sIL-2R production correlates with IL-2 production.

gamma-Interferon, interleukin-4 and interleukin-6 in vitro production in old subjects.

It is well known that ageing is associated with various alterations of the lymphoid cell functions. Although both B and T cell are affected, the last appear to be more sensitive to ageing process. During the past years, to gain insight into thé mechanism(s) of this impairment, effort has been centered on the helper T cells specifically engaged in the production of interleukin-2 (IL-2) because of the pivotal role played by this cytokine in the activation of several immune functions. The results have demonstrated that the ability to produce IL-2 declines with age. In this paper we report the results of a study performed to determine the influence of age on the capacity to produce gamma-interferon (gamma-IFN), interleukin-4 (IL-4) and interleukin-6 (IL-6). Mononuclear cells from young and old subjects were assessed for cytokine producing capacity in response to phytohaemagglutinin stimulation. A significant decrease of gamma-IFN production by old subjects has been observed. No significant difference was instead observed between the old subjects and the young ones as regards IL-4 and IL-6 production. We suggest that this imbalanced cytokine production may well account for the pattern of immune response which may be observed in elderly, i.e. a normal or increased humoral response in face of a low T cell immune responsiveness.

The effect of age on mitogen responsive T cell precursors in human beings is completely restored by interleukin-2.

It is well known that the function of T lymphocytes is significantly impaired by advancing age. In the present study, attempts have been made to further characterize the T cell impairment of elderly subjects. Thus, we have performed limiting dilution microculture analysis to evaluate the precursor frequency of T lymphocytes responding to a mitogenic stimulus in old and young subjects. Furthermore we have evaluated the activity of recombinant interleukin-2 (rIL-2) on these cells. The results demonstrate that in older subjects the frequency of these precursors is significantly decreased. The in vitro treatment with rIL-2 increased the frequency of mitogen responsive T lymphocyte precursors in both groups so that the difference between the two groups was not significant. Thus present results extend the findings demonstrating that older subjects display an impairment of T cell functions and that IL-2 treatment may correct these alterations. In particular, they confirm the hypothesis that age-associated functional changes are more likely due to diminished numbers of reactive cells, than to a decline in the activity of all cells.

IgD serum levels are influenced by HLA-DR phenotype.

In the present paper we have evaluated IgD serum levels of 84 randomly selected HLA-typed healthy Sicilians. The values were analysed according to age, sex and HLA-DR phenotypes. No correlation between age and IgD serum levels was found in our population since all subjects were in a narrow age range. Furthermore, no significant association was found between IgD serum levels and gender of studied subjects. The evaluation of IgD serum levels according to HLA-DR phenotypes revealed that HLA-DR1 positive subjects displayed significantly higher values. These results are in agreement with previous reports showing that HLA phenotypes may be involved in the control of serum immunoglobulin levels. Furthermore, present data strengthen our suggestion that HLA-DR1 phenotype is related to the 'high responder' immunological profile.

Blood antiphospholipid antibody levels are influenced by age, sex and HLA-B8,DR3 phenotype.

Antiphospholipid antibodies (APA) are known to be associated with a number of seemingly heterogeneous pathological conditions that are part of the antiphospholipid syndrome, formerly called anticardiolipin syndrome. Recent studies on the mechanism of action of these autoantibodies suggest that we are dealing with a new autoimmune syndrome which may occur either in a primary form or in the context of other autoimmune diseases, mainly systemic lupus erythematosus (SLE). Moreover, increased levels of APA have been found in elderly subjects, who are known to display increased frequency of autoimmune phenomena. It is well known that many autoimmune diseases, including SLE, are associated with HLA antigens, particularly with HLA-B8,DR3 phenotype. In our study, APA serum levels were analyzed in 26 old subjects and in 56 young ones. The results demonstrate that HLA-B8,DR3-positive young females display significantly higher levels of APA than HLA-B8,DR3-negative ones. Interestingly, the same is true for elderly subjects on the whole with respect to young individuals. These data are consistent with previous findings demonstrating that HLA-B8,DR3-positive subjects (mainly female) as well as old subjects display (also in the absence of any clinical manifestation), multiple immune dysfunctions that may underlie the predisposition to autoimmunity.

Soluble interleukin-2 receptor release defect in vitro in elderly subjects.

The evidence from several studies indicates that as individuals age, they may display immune dysfunctions, mostly T cell dysfunctions. Recently, a soluble form of the receptor for interleukin-2 (IL-2) (sIL-2R) has been demonstrated in human sera and in vitro stimulated culture supernatants from human T lymphocytes. In the present paper, we report in vitro sIL-2R production from peripheral blood mononuclear cells in elderly subjects. The results show that no difference exists for unstimulated cultures, whereas after mitogen stimulation the elderly subjects showed the lowest values compared with young ones. These findings suggest that sIL-2R may provide a new tool for the study of T lymphocyte dysfunctions in old age.

Immunoglobulin heavy chain allotypes in a sample of Sicilian patients with celiac disease.

The authors have studied the immunoglobulin heavy chain allotypes in 14 Sicilian patients with celiac disease (CD) and in 75 healthy controls. No association has been found between CD and Gm or Km phenotypes. When the role of Gm phenotypes in immune response against gliadin was analyzed, fb-positive CD patients on gluten-free diet for 1 year showed an increased antigliadin IgG response. Comparing the values of fb-positive subjects with those of negative ones, a significant difference was observed. These data, demonstrating that the presence or absence of a phenotype is able to define the breadth of immune response against alpha-gliadin antigen, are indicative of the role played by Ig allotypes in CD.

HLA-B8,DR3 T cell impairment is completely restored by in vitro treatment with interleukin-2.

The activity of recombinant interleukin-2 (rIL-2) on the in vitro lymphocyte proliferative response to phytohemagglutinin mitogen was investigated in healthy HLA-B8,DR3 positive and negative subjects. The response to mitogen, significantly decreased in HLA-B8,DR3 positive subjects, was completely restored by adding rIL-2. Moreover, in HLA-B8,DR3 positive subjects the in vitro treatment with rIL-2 significantly increased the reduced frequency of mitogen responsive T lymphocyte precursors, as assessed by limiting dilution analysis. These data suggest that a decrease in the size of the pool of T cell precursors able to produce IL-2 is responsible for the impairment of T cell function observed in HLA-B8,DR3 positive subjects. Since in autoimmune diseases it is possible to show the same impairment(s) of T cell functions which can be observed in HLA-B8,DR3 positive subjects, these results could be of practical value for the understanding of pathogenetic mechanism(s) of autoimmune diseases and, in case, for therapeutical purposes.

Immunoglobulin heavy-chain allotypes play a role in the clinical history of celiac disease.

We studied the role of the immunoglobulin heavy-chain allotypes in the evolution of celiac disease. Particularly the Gm(fb) phenotype was investigated since this phenotype is able to determine the breadth of immune response against alpha-gliadin antigen. The results show that weight percentiles and intestinal absorption as assessed by xylose test are influenced by Gm(fb) phenotype suggesting that Gm(fb) genes contribute to the determination of clinical aspects and evolution of celiac disease.

Markers of T lymphocyte activation in HLA-B8, DR3 positive individuals.

Many autoimmune diseases are associated in Caucasians with HLA-B8 and/or HLA-DR3 antigens. There is evidence that bearers of these antigens may display significant changes in immune parameters when compared to individuals not having these antigens. Recently, increased numbers of blood activated T lymphocytes have been reported in the majority of these diseases. The increase in activated blood T lymphocytes is paradoxically characterized by an in vitro impairment of T cell activation. Particularly, an inadequate production of interleukins has been observed. We have studied blood levels of activated T cells in HLA-typed, healthy subjects. The results show that the percentage of activated T cells, as recognized by monoclonal antibodies anti-CD25, anti-Ia and anti-MLR3, was more frequent in HLA-B8, DR3 positive individuals. On the other hand, in the 24 h, PHA stimulated cultures IL-2, IFN-gamma and the percentage of T cells CD25 positive were decreased. Thus, there was an apparent discrepancy between the increase of blood activated T cells and the in vitro impaired T cell activation. Since there is evidence that HLA-B8, DR3 positive subjects are genetically low responders, a possible reason for the discrepancy might be their relative inability to remove antigenic stimuli from the body. In this case, the increased number of activated blood T cells may reflect a cellular activation caused by persistent antigenic stimulation.

HLA-B8,DR3 phenotype and lymphocyte responses to phytohaemagglutinin.

Several reports have shown that HLA-B8,DR3 positive subjects may display some changes in immune parameters when compared with HLA-B8,DR3 negative ones and are prone to develop several immunological diseases. In the present study we have analysed the proliferative response to phytohaemagglutin (PHA) in HLA-typed healthy subjects. A twin method was also employed to assess the role of genetic and environmental factors in the regulation of the response to the mitogen. It was not possible to demonstrate any difference in proliferative response to optimal doses of PHA between groups of subjects carrying or not carrying the HLA-B8,DR3 phenotype. When suboptimal responses were studied, however, the results showed that lymphocyte responses were significantly decreased in HLA-B8,DR3 positive subjects compared with the negative ones. Moreover, the experiments performed with twins demonstrated that environmental factors were more important than genetic factors in the proliferative response to mitogen. The fact that the HLA-B8,DR3 phenotype affects the suboptimal response to PHA although environmental factors are more important than genetic factors in the response to the mitogen seems of some interest. However, these results could be consistent with the high incidence of autoimmune disorders among HLA-B8,DR3 positive individuals.