Susceptibility of HLA-B27 transgenic mice to Yersinia enterocolitica infection

The majority of patients with reactive arthritis have the major histocompatibility complex class I gene HLA-B27. The development of arthritis in these patients often occurs following infection with one of several enteric bacteria, including Yersinia enterocolitica. In this study, transgenic mice expressing HLA-B27 and their negative full sibs were infected intravenously with Yersinia enterocolitica 0:8 WA in an attempt to develop an experimental model of reactive arthritis. To date, no reactive arthritis has been observed; however, a significantly higher incidence of paralysis was observed in the HLA-B27⁺ transgenic mice. Injection of 10⁵ organisms induced hind limb paralysis in 8 out of 30 of the HLA-B27 transgenic mice (27%) and in only 1 of the 24 negative siblings (4%). Paralysis occurred in 14 out of 30 HLA-B27⁺ mice (47%) at a dose of 10⁴ organisms. Only 2 of the 25 negative siblings (8%) were affected at this dose. Paraspinal abscesses were found in all of the paralyzed animals. At the 10⁴ dose most of the HLA-B27⁺ mice (70%) succumbled to the disease within 4 weeks, while the mortality in their B27⁻ full sibs was less than 10%. Thus, HLA-B27 transgenic mice have higher mortality and morbidity from infection with Y. enterocolitica 0:8 WA than corresponding HLA-B27⁻ littermates.     

Description of a mouse monoclonal anti-HLA-B27 antibody HLA-ABC-m3

The production and characterization of a new anti-HLA-B27 monoclonal antibody HLA-ABC-m3 is described. This cytotoxic IgG2a antibody binds protein A and is able to precipitate cell surface molecules of 43,000 and 12,000 daltons corresponding to the HLA heavy chain and β2-microglobulin. Population testing revealed that the HLA-ABC-m3 antibody reacted with the peripheral blood lymphocytes of 47/47 individuals conventionally typed as HLA-B27⁺ and with 5/105 HLA-B27⁻ individuals. These five extra reactions were with individuals expressing the cross-reactive HLA-B7 alloantigen, although the affinity of the monoclonal antibody for B27 heterozygous individuals (approx 10⁹ M⁻¹) was tenfold greater than with B7 individuals (approx 10⁸ M⁻¹). In addition, HLA-ABC-m3 reactivity segregated with HLA-B27 in two families. This monoclonal antibody should be of value in the investigation of the role of HLA-B27 in disease.     

CD8 is involved in both class I- and class II—induced proliferation of a cytolytic T-cell clone with dual specificity for HLA-B27 and HLA-DR2 antigens

A CD3⁺ CD4⁻ CD8⁺ cytolytic T-lymphocyte (CTL) clone, CTL 47, could be induced to proliferate in the presence of exogenous interleukin 2 by either HLA-B27.1⁺ or HLA-DR2⁺ cells. B27.1-induced proliferation was strongly and equally inhibited by an anti-B27 and by an anti-CD8 monoclonal antibody (MoAb). DR2-induced proliferation was inhibited by the same anti-CD8 MoAb less efficiently and with a different time course than anti-class II blocking, only being significant when the antibody was added ab initio or very early during the assay. These results indicate that CD8 is essential for class I—induced proliferation but that it also enhances class II—induced stimulation of this CTL clone. It is proposed that the necessary role of CD8 in class I—induced proliferation is related to its interaction with the same class I molecule bound by the T-cell receptor. The accessory role in class II—induced proliferation would be due to an additive effect on the avidity of cell adhesion, resulting from interaction of CD8 with the class I antigens on the stimulator cell, or perhaps to a regulatory role of CD8 as a transducer of early signals for T-cell activation.     

Human suppressor T cells: Induction, differentiation, and regulatory functions

T-cell-mediated suppression of human immune responses involves a complex interaction between distinct lymphocyte subsets with suppressor-inducer and suppressor-effector functions. Recent studies with subset-specific monoclonal antibodies have defined a characteristic phenotype of suppressor-inducer cells (CD4⁺ Leu8⁺ 2H4⁺ 4B4⁻) that can be distinguished from that of helper cells for antibody synthesis (CD4⁺ Leu8⁻ 2H4⁻ 4B4⁺). Similarly, suppressor-effector cells (CD8⁺ CD11⁺ Tp44⁻ can typically be defined as a subset separable from cytotoxic T cells (CD8⁺ CD11⁻ Tp44⁺). Both antigen-specific and nonspecific interactions are important in suppressor T-cell activation and function. Soluble signals required for differentiation of CD8⁺ suppressor cells include an indomethacin-sensitive monocyte product and interferon gamma. In contrast, proliferation of the CD8⁺ suppressor cell subset depends on stimulations first by a product of CD4⁺ Leu8⁺ cells, T suppressor cell growth factor, and second by interleukin 2. Although the molecular basis of antigen-specific interactions between CD4⁺ and CD8⁺ cells in suppressor cell generation has not been defined, it may involve both conventional, presumably MHC-restricted, interactions between antigen and antigen receptors, as well as anti-idiotypic interactions of suppressor-effectors with determinants on suppressor-inducer receptors. Progress in elucidating requirements for activation, growth, and differentiation of suppressor cells should facilitate long-term culture of such cells and lead to clearer understanding of mechanism of suppressor-cell mediated immunoregulation.     

Some disease-associated ancestral haplotypes carry a polymorphism of TNF

We describe here an Nco I restriction fragment length polymorphism of tumor necrosis factor carried by the 8.1 (HLA-A1,B8,BfS,C4AQ0,C4B1,DR3) and the 44.1 (HLA-B44,BfS,C4A3,C4BQ0,DR4) ancestral haplotypes associated with complications of rheumatoid arthritis. By examining multiple examples of these and other ancestral haplotypes it was seen that 8.1 and 44.1 ancestral haplotypes yield fragments of approximately 5.5 kb while many other ancestral haplotypes carry fragments of approximately 10.5 kb. The polymorphism is associated with the ancestral haplotype rather than the HLA-B or -DR allele defined by conventional serology.     

Cell mediated immunity changes in ageing, relative importance of cell subpopulation switches and of nutritional factors

Decreased T-cell functions with ageing have been extensively described. This review focuses on recent data on changes in T-cell subpopulations related to ageing and their consequences on T-cell proliferation. Increase of immature T cells CD2⁺ CD3⁻ is an ageing phenomenon related to T-cell declining proliferation. Recently it was shown that increase of immature T cells was due to an increase in different subtypes of the CD2⁺ CD3⁻ population, double-negative CD2⁺ CD4⁻ CD8⁻ and double-positive CD2⁺ CD4⁺ CD8⁺ subpopulations, the former being associated with nutritional deficit, the latter with associated diseases. Other authors have focused on decreases of naive T cells with parallel increase of memory T cells; such a switch is also relevant to declining T-cell proliferation. This review focuses on two major factors which influence immune ageing: nutritional parameters and antigen exposure.     

The genetic basis for the association of the 8.1 ancestral haplotype (A1, B8, DR3) with multiple immunopathological diseases

Summary: An individual's major histocompatibility complex (MHC) ancestral haplotype (AH) is the dearest single determinant of susceptibility to MHC associated immunopathological disease, as it defines the alleles carried at all loci in the MHC. However, the direct effects of any of the 150–200 genes that constitute the MHC are difficult to determine since recombination only occurs at defined hotspots. This review concerns the 8.1 AH (HLA-A1, C7, B8, C4AQ0, C4B1, DR3, DQ2), which is carried by most Caucasians with HLA-B8. It is associated with accelerated human immunodeficiency virus (HIV) disease, and susceptibility to insulin-dependent diabetes mellitns (IDDM), systemic lupus erythematosus, dermatitis herpetiformis, common variable immunodeficiency and IgA deficiency, myasthenia gravis and several other conditions. We have mapped susceptibility genes for HIV, IDDM and myasthenia gravis co the central MHC between HLA-B and the tumour necrosis factor or complement genes. Here we consider which of the remaining 8.1-associated diseases are more closely associated with HLA-DR3 and/or DQ2. Several candidate genes in the central MHC have the potential to modulate immune or inflammatory responses in an antigen-independent manner, as is seen in studies of cultured cells from healthy carriers of the 8.1 AH. Hence these genes may act as a common co-factor in the diverse immunopathological conditions associated with the 8.1 AH.     

Characterization of CD28CD4 T cells in living kidney transplant patients with long-term allograft acceptance

CD28⁻CD4⁺ T-cell subpopulation is expanded in kidney allograft patients with long graft survival. To seek for the roles of CD28⁻CD4⁺ T cells in the long-term acceptance of kidney allografts, we characterized this population by analyzing cell surface molecules, TCR V_β repertoire, mixed lymphocyte reaction (MLR), and cytokine production. The number of CD28⁻CD4⁺ T cells increased correlatively with time after transplantation in this group of patients. The CD28⁻CD4⁺ T cells did not express detectable levels of CD25, CD69, V24, or CTLA-4 but expressed heterogeneous amounts of CD45 RA on the surface. Freshly sorted CD28⁻CD4⁺ T cells revealed a restricted V_β repertoire, whereas the V_β usage of CD28⁺CD4⁺ T cells from the same patients was much diversified. Expression levels of TGF-β and IFNγ gene were significantly higher in the CD28⁻ CD4⁺ T cells than in the CD28⁺CD4⁺ T cells from the kidney allograft patients. These findings suggest that an oligoclonal CD28⁻ CD4⁺ T-cell population is continuously activated in patients with long allograft survival, which may be linked with the long-term acceptance.     

Impairment of B and T cell maturation in gut associated lymphoid tissues due to malnutrition during lactation

Previously we found that malnutrition during lactation in rats produces an impairment in the immune response to cholera toxin. In this report we found that malnutrition during lactation provokes in 28-day-old rats an increase of Thy1⁺ cμ⁺ cells in gut associated lymphoid tissues concomitantly with a decrease of sIgA⁺ B cells. No differences were found in the percentages of the IgM⁺B cell populations. Furthermore, no differences were found in the Peyer's patch (PP) and mesenteric lymph node (MLN) T cell subsets in weaning rats when compared to controls. However, after 1 week of refeeding a higher percentage of the Thy1⁺ cμ⁻ subset together with a lower percentage of CD5⁺, CD4⁺, and CD8⁺ T cells, were found in malnourished rats when compared to controls. The above results may indicate that B-cell maturation is delayed in malnourished rats at two stages of differentiation: (a) in the passage of pre-B cells (Thy1⁺ cμ⁺) to immature B cells (sμ⁺), and (b) in the switch from sμ⁺ B cells to s⁺ B cells. The decrease of CD5⁺, CD4⁺, and CD8⁺T cells together with and increase of the Thy1⁺ cμ⁻ subset in gutassociated lymphoid tissues (GALT) may indicate that T-cell maturation is also delayed. Results obtained at weaning may be due to an engraftment by maternal milk-derived lymphocytes in the pups.     

Heterogeneity in terms of interleukin 4-dependent regulation of Fce receptor/CD23 expression on chronic B-lymphocytic leukemia cells

To discriminate the stages of maturation arrest of leukemic B cells, we have investigated the cell surface expression of FcεR_1l (H107 antigen) on leukemic B cells from 6 patients with chronic type B-lymphocytic leukemia(B-CLL) by a double staining method combined with cytoflorometry, and their production of soluble FceR_ll ⁺ by an ELISA technique. FceR_ll was expressed onμ⁺/δ⁻ cells of case 5 as well as on μ⁺/δ⁺cells of cases 1,2 and 4, but not on μ⁺/δ⁺cells in cases 3 and 6. The cultivation of leukemic cells with IL-4 not only increased the percentage of FceR_ll⁺cells but also enhanced the production of soluble Fcer_llin most cases. However, IL-4 had no effects on μ⁺/δ⁻/Fcεr_ll⁺ cells of cases 5, which appeared to correspond to a rather late     

Does a central MHC gene in linkage disequilibrium with HLA-DRB1*0401 affect susceptibility to type 1 diabetes?

Subtypes of HLA-DR4 are associated with susceptibility or protection against type 1 diabetes (T1DM). We addressed whether this reflects linkage disequilibrium with the true susceptibility locus by studying broader MHC haplotypes marked by alleles of HLA-B, IKBL (adjacent to TNFA) and complement C4. The study used a largely Caucasian cohort from Western Australia. HLA-DRB1*0401 and HLA-DRB1*0405 marked susceptibility to T1DM. In Caucasians, DRB1*0401 occurs predominantly in the 44.1 ancestral haplotype (AH; HLA-A2,B44, DRB1*0401,DQB1*0301) and the 62.1AH (HLA-A2,B15(62),DRB1*0401,DQB1*0302). HLA-B15 marked susceptibility and HLA-B44 marked with resistance to T1DM in patients and controls preselected for HLA-DRB1*0401. A gene between TNFA and HLA-B on the 8.1AH (HLA-A1,B8,;DR3,DQ2) modifies the effects of the class II alleles. Here, alleles characteristic of the 62.1AH (C4B3, IKBL+446*T and HLA-A2,B15) were screened in donors preselected for HLA-DRB1*0401. C4B3 was associated with diabetes, consistent with a diabetes gene telomeric of MHC class II. However, increases in carriage of IKBL+446*T and HLA-A2,B15 were marginal, as too few control subjects were available with the diabetogenic alleles. However, with these tools, selection of HLA-DRB1*0401, DQB1*0302 donors who are positive and negative for C4B3 will allow bidirectional mapping of diabetes genes in the central MHC.     

Periodontal attachment loss in HIV-infected patients is associated with the major histocompatibility complex 8.1 haplotype (HLA-A1,B8,DR3)

Periodontal attachment loss is mediated by overproduction of tumour necrosis factor (TNF) and interleukin (IL)-1, and appears to have a genetic component. The 8.1 major histocompatibility complex (MHC) ancestral haplotype (HLA-A1,B8,TNFA-308(2),DR3) is associated with elevated TNF production and predisposes carriers to several autoimmune/immunopathological disorders, including rapid progression of HIV disease, but not early onset periodontal disease in healthy individuals. Rather a high proportion of subjects with severe periodontal disease carry allele 2 at IL-1A-889 and IL-1B+3953. We predicted that genetic associations may be different or clearer in HIV patients, as they often show elevated production of TNF and IL-1 and periodontal attachment loss. Hence periodontal parameters and IL-1 polymorphisms were assessed in HIV-positive subjects expressing HLA-B8 with or without other markers of the 8.1 haplotype. Of 16 HLA-B8 subjects, 13 demonstrated elevated probing pocket depth and clinical attachment loss. The difference was statistically significant and did not correlate with smoking, age, CD4 T-cell counts, HIV viral load or levels of dental plaque. As TNFA-308 (allele 2) was present in four non-B8 subjects who had minimal attachment loss, it may not mediate the effect of the 8.1 haplotype. Moreover, polymorphisms at IL-1A-889 and IL-1B+3953 did not significantly affect periodontal parameters. Thus a central MHC gene characteristic of the 8.1 haplotype was the clearest determinant of periodontal attachment loss in HIV-infected individuals.     

ANCESTRAL HAPLOTYPES CARRY HAPLOTYPIC AND HAPLOSPECIFIC POLYMORPHISMS OF BAT1: POSSIBLE RELEVANCE TO AUTOIMMUNE DISEASE

The human BAT1 gene, located in the central MHC region (–170kb centrometric of HLA-B), is polymorphic and the polymorphism correlates with MHC ancestral haplotypes. Allelic RFLP patterns have been assigned to several ancestral haplotypes and have been shown to be ‘haplotypic’ (i.e. found on all examples of the same ancestral haplotype) and in some cases ‘haplospecific’ (i.e. unique to one ancestral haplotype). The relevance of the BAT1 polymorphism to susceptibility to Myasthenia Gravis (MG) has been investigated. The frequency of the BAT1 B allelic pattern is increased in patients with MG (n= 16) compared to an equal number of control subjects. The increase is due to the association between MG and the 8.1 ancestral haplotype (HLA A1, Cw 7, B8, BfS, C4AQ0, C4B1, DR3, DQw2).     

Ancestral haplotypes carry haplotypic and haplospecific polymorphisms of BAT1: possible relevance to autoimmune disease.

The human BAT1 gene, located in the central MHC region (approximately 170 kb centrometric of HLA-B), is polymorphic and the polymorphism correlates with MHC ancestral haplotypes. Allelic RFLP patterns have been assigned to several ancestral haplotypes and have been shown to be 'haplotypic' (i.e. found on all examples of the same ancestral haplotype) and in some cases 'haplospecific' (i.e. unique to one ancestral haplotype). The relevance of the BAT1 polymorphism to susceptibility to Myasthenia Gravis (MG) has been investigated. The frequency of the BAT1 B allelic pattern is increased in patients with MG (n = 16) compared to an equal number of control subjects. The increase is due to the association between MG and the 8.1 ancestral haplotype (HLA A1, Cw7, B8, BfS, C4AQ0, C4B1, DR3, DQw2).     

HLA-DQ1 haplotype ruin the effect of cis complementation of DQA10501-DQB10201 in association with trichosanthin-induced immunosuppression

CD8 cell-mediator (M+) or non-mediator (M−) are distinguishable for healthy subjects according to whether their CD8 T cells keep the down-regulatory function in Trichosanthin (Tk)-induced immunosuppression. Tk is a plant protein of 247 amino acid residues purified from a Chinese medicinal herb. The M+ phenotype has been shown in our previous work to be strongly associated with HLA-DQ2. By genotyping with PCR-based techniques, the essential alleles of the DQ2 were identified as DQA1^*0501 and DQB1^*0201, which were either in cis (DR3) or in trans (DR5, DR7) position. A more detailed examination of the HLA association pattern with M+/M− in 42 Chinese candidates, however, revealed another two points of interest. 1) The cis complementation did not work if another DQA1^*01- or DQA1^*02-related haplotype (e.g. DRB1^*0101-DQA1^*0101-DQB1^*0501) were combined. The later seemed to behave like a ‘negative’ factor superimposed on the ‘positive’ role of DQA1^*0501-DQB1^*0201 haplotype in heterozygous condition. 2) In addition to DQA1^*0501, the DQB1^*0201 was actually able to combine all available DQA1 alleles except DQA1^*01 family to form the trans complementation. Again, the DQ1 haplotype acted negatively. It is thus likely that the cis and trans complementary association of DQA1^*0501-DQB1^*0201 could only be detected conditionally or only appeared as a special case in the Tk-induced immunosuppression.     

A family of T-cell receptor molecules expressed on T-cell clones with different specificities for allomajor histocompatibility antigens

A monoclonal antibody, 3D6, identifies a public idiotope or allotope on the human T-cell receptor for antigen, since it not only reacts with the tumor line HPB-ALL, against which it has been raised, but also with 3–13% of peripheral blood T lymphocytes of normal donors. 3D6⁺ cells have been isolated from an allogeneic mixed lymphocyte culture and cloned by limiting dilution. In this way, allospecific clones were obtained both of the T4⁺ T8⁻ and the T4⁻ T8⁺ phenotype, which included proliferative as well as cytotoxic cells. Within a panel of 20 cytotoxic clones, different specificities for both class I and class II MHC antigens were found. The clones were tested for their reactivities with four additional anti-T-cell receptor antibodies raised against HPB-ALL. Two of these, 1C1 and 1C2, reacted with all 3D6⁺ clones. By means of two other antibodies, 2D4 and 65, the 3D6⁺ receptor family could be divided into four structurally distinct subfamilies. Biochemical analysis suggested that the 1C1, 1C2, 2D4, and 3D6 antibodies define epitopes on the beta chain of the receptor. Isoelectric focusing of receptor molecules isolated from cytotoxic clones with different specificites indicated that there are extensive structural differences in both alpha and beta chains of the receptors. No correlation could be found between the antigenic specificity of a clone and the structure of its receptor in this analysis. It is postulated that the 1C1, 1C2, and 3D6 epitopes may be encoded by a particular germline V beta segment, in analogy with similar, previously described findings in both the human and the murine system.     

Common HLA-B8-DR3 haplotype in Northern India is different from that found in Europe

The aim of this study was to determine whether a common diabetic haplotype, including human leukocyte antigen (HLA)-B8 and HLA-DR3, in Northern India is the same haplotype as the European HLA-B8-DR3 haplotype. DNA samples from Northern Indian subjects selected on the basis of HLA-B8 and HLA-DR3 were tested for microsatellite and single nucleotide polymorphism alleles throughout the major histocompatibility complex (MHC). It was found that the Indian samples represent a conserved haplotype in which all alleles were shared by Indian subjects with HLA-B8 and HLA-DR3, but were different to those that are characteristic of the European 8.1 ancestral haplotype. The Indian and European haplotypes share HLA-B*0801, HLA-DRB1*0301 and HLA-DQB1*02 but differ for subtypes of HLA-Cw*07 and HLA-DRB3 and all central MHC alleles tested. In contrast, Indian subjects selected on the basis of HLA-B58 ( 1-17) and HLA-DR3 shared the same alleles at other MHC loci as have been described in the common Chinese haplotype with HLA-B58/17 and HLA-DR3. A third haplotype, HLA-B50/21 and HLA-DR3, was also found to be highly conserved but shares little in common with the other two HLA-DR3-containing Indian haplotypes.     

C-kit positive porcine bone marrow progenitor cells identified and enriched using recombinant stem cell factor

Porcine haematological studies have been hampered by the lack of monoclonal antibodies against porcine CD34 or CD117 expressed on haematological progenitors. The present report describes the enumeration, phenotyping and isolation of porcine haematopoietic progenitor cells expressing stem cell factor (SCF, c-kit ligand) receptor (c-kit, CD117). Recombinant porcine (rp) SCF and granulocyte-macrophage colony-stimulating factor (GM-CSF) were expressed in the mammalian HEK293 cell-based expression system. Both were biologically active and induced the proliferation of the human erythroleukemic cell line TF-1, as well as of porcine bone marrow haematopoietic cells (BMHC), in a concentration-dependent manner. The effect of rpSCF on BMHC proliferation was synergistic with rpGM-CSF. Furthermore, rpSCF had a synergistic effect on the generation of BMHC-derived dendritic cells (DC) induced by GM-CSF and TNF-. RpSCF was expressed with a 6-histidine epitope, permitting both its purification and immunological detection. Binding studies with BMHC demonstrated ligation of SCF to 4–11% of BMHC. These cells represented the SWC3^low/−SWC8⁻ BMHC subset, with characteristics of immature proliferative progenitor BMHC. In contrast, no expression was noted on the SWC3⁺SWC8⁻ monocytic, the SWC3⁺SWC8⁺ granulocytic or the SWC3⁻SWC8⁺ B cell lineage cells. Using magnetic or fluorescence-activated cell sorting, SCF-ligating BMHC were enriched for pluripotent progenitor cells. In this manner, porcine haematological studies can be pursued in a detailed manner not before possible.     

Major histocompatibility complex genes influence the outcome of HIV infection: Ancestral haplotypes with C4 null alleles explain diverse HLA associations

Several alleles at multiple HLA loci have been found to be associated with infection with human immunodeficiency virus (HIV): HLA A1; B8, B35; Cw7, Cw4; DR1, DR3 and DQ1, are associated with particular disease manifestations and/or disease progression. Furthermore, in a pilot study we have shown an increase in the frequency of C4 null alleles and suggested that all the reported HLA alleles could reflect association with a limited number of ancestral haplotypes (AHs).

On this occasion, we studied 122 Caucasoid patients classified according to Centers for Disease Control (CDC) criteria. The control group consisted of 67 seronegative homosexual or bisexual males at risk of developing HIV infection.

C4 null alleles were unequivocally present in 58% of patients in CDC IV compared with 33% of the seronegative subjects (x² = 5.65, p < 0.05). Furthermore, C4 null alleles could be excluded in only 8% and 16% of CDC III and IV, respectively, but in 30% of the seronegative subjects. An increased frequency of three AHs largely accounted for the increases in C4 null and HLA alleles.

To examine the role of specific AHs we undertook a longitudinal analysis of a subgroup of 26 patients who seroconverted under observation. Seventeen of these patients were followed for 32 to 63 months. All seven patients with the 8.1 AH (A1, CW7, B8, BfS, C4AQ0, C4B1, DR3, DQ2) developed low CD4 lymphocyte counts (<450 × 10⁶/l) compared with only 2 of 10 patients without this haplotype (p < 0.002). All three deaths occurred in patients with the 8.1 AH. The acquired immunodeficiency syndrome developed in three further cases with either 8.1- or B35- bearing (35.x) haplotypes. Sequential CD4/8 ratios showed an early and progressive decline in individuals with 8.1 or 35.x. Since the 8.1 and 35.x AHs contain deletions of the central major histocompatibility complex (MHC) genes, we suggest that the genes affecting HIV infection and progression are within the central MHC region.