Allelic distribution and the effect of haplotype combination for HLA type II loci in the celiac disease population of the Valencian community (Spain)

The association between human leukocyte antigen (HLA) class II antigens and celiac disease (CD) was analyzed in a Spanish population. No association with DRB1*04 and DQB1*0302 was noted. The main associated haplotype (70.8%) was DRB1*03–DQB1*0201–DQA1*0501(DR3–DQ2), followed by DRB1*07–DQB1*0202–DQA1*0201 (DR7–DQ2) haplotype, which is associated with DRB1*11–DQB1*0301–DQA1*0505 (DR11–DQ7). The combinations of DR3–DQ2 with DR7–DQ2, and DR7–DQ2 with DR11–DQ7, present a twofold risk compared with each haplotype in homozygosis. An independence test in DR3-DQ2 haplotype found that association with CD was attributable to the whole haplotype, but for DR7-DQ2 was secondary to DQB1/DQA1. There is no need of a double gene dosage to increase the risk. CD-associated alleles typing demonstrates a very high negative predictive value to exclude CD in risk groups.     

HLA-DR and HLA-DQ polymorphism in human thyroglobulin-induced autoimmune thyroiditis: DR3 and DQ8 transgenic mice are susceptible

In contrast to H2-based susceptibility to experimental autoimmune thyroiditis (EAT) induced with thyroglobulin (Tg), human leukocyte antigen (HLA) association with Hashimoto's thyroiditis, the human counterpart, is less clear, and determining association is further complicated by DR/DQ linkage disequilibrium. Previously, we addressed the controversial implication of HLA-DR genes by introducing HLA-DRA/DRB1*0301 (DR3) transgene into endogenous class II negative H2Ab(0) mice. EAT induction with either human (h) or mouse (m) Tg demonstrated the permissiveness of DR3 molecules for shared Tg epitopes. Here, we examined the participation of HLA-DQ genes by introducing DQA1*0301/DQB1*0302 (DQ8) transgene into class II negative Ab(0) or class I and II negative beta(2)m((-/-)) Ab(0) mice. About 50% and 80% of HLA-DQ8(+) Ab(0) and beta(2)m(-) Ab(0) mice, respectively, developed moderate EAT after hTg immunization, but only minimal response to mTg. The hTg presentation to hTg-primed cells was blocked by anti-DQ mAb in vitro. By contrast, HLA-DRB1*1502 (DR2) and *0401 (DR4) transgenes contributed little to hTg induction. Similarly, DQA1*0103/DQB1*0601 or DQA1*0103/DQB1*0602 (DQ6) transgenic Ab(0) mice were unresponsive to hTg induction and carried no detectable influence in DQ8/DQ6 double transgenic mice. Thus, both HLA-DR and -DQ polymorphism exists for hTg in autoimmune thyroiditis. The use of defined single or double transgenic mice obviates the complications seen in polygenic human studies.     

Coexpression of susceptible and resistant HLA class II transgenes in murine experimental autoimmune thyroiditis: DQ8 molecules downregulate DR3-mediated thyroiditis

Experimental autoimmune thyroiditis (EAT) can be induced in genetically susceptible mice by immunization with the self antigen, thyroglobulin (Tg). Since susceptibility is linked to H2 class II molecules, we have generated human leukocyte antigen (HLA) class II transgenic mice to study potential HLA associations with Hashimoto's thyroiditis. DR3 (HLA-DRA/DRB1*0301) and DQ8 (HLA-DQA1*0301/DQB1*0302) transgenes were introduced into class II-negative Ab(0)/B10 and Ab(0) nonobese diabetic (Ab(0)/NOD) mice. Previous work had shown that DR3 transgenic mice were susceptible to both mouse Tg and human Tg-induced EAT, whereas DQ8 transgenic mice were moderately susceptible only to human Tg induction. In this report, we examined the effect of DQ8 transgene on mouse Tg- and human Tg-induced EAT in double transgenic DR3/DQ8 mice. After mouse Tg induction, thyroiditis in DR3(+)DQ8(+) Ab(0)/B10 mice was significantly less severe than in DR3(+) mice but more severe than in DQ8(+) mice. No difference in thyroiditis was observed between DR3(+) and DR3(+)DQ8(+) mice in another background strain, Ab(0)/NOD. However, after immunization with human Tg, DQ8 coexpression downregulated thyroiditis severity, compared to DR3(+) mice, whereas thyroiditis was more extensive than in DQ8(+) mice. Thus, depending on the background strain and the Tg used to induce disease, the presence of the DQ8 transgene can reduce thyroiditis mediated by DR3 molecules.     

Additional association of intra-MHC genes,MICA and D6S273, with Addison's disease

Intra-MHC sequences including MHC class I chain-related genes (MICAs), D6S273 and D6S2223 are associated with autoimmune diseases in addition to HLA class II. In the current study, we ascertained the haplotypes of 57 Caucasian patients with Addison's disease composed of these genetic markers and compared them either with 72 general population controls or with 105 child controls carrying Addison's disease high-risk DR3-DQ2/DR4-DQ8 genotypes. The MICA-A5.1/A5.1 genotype as well as HLA DR3/4 especially with DRB1*0404 were the main susceptibility markers. The homozygous MICA-A5.1/A5.1 genotype was significantly more frequent in the patients with Addison's disease (61%) than in the healthy controls (6%). The MICA-A5.1 allele was increased on both the DR3 and DR4 haplotypes, independent of DQ and DRB1 subtyping, in the patients with Addison's disease compared with the controls. Furthermore, the D6S273*140 allele on the DR3 haplotype and the D6S273*134 allele on the DR4 haplotype in the DR3/4 heterozygotes influenced susceptibility relative to the DR3/4 controls. The risk for Addison's disease was increased for the DR3-D6S273*140-MICA-A5.1/DRB1*0404-D6S273*134-MICA-A5.1 genotypes compared with that conferred by the DR3/4 controls. Susceptibility to Addison's disease is influenced by the genes around MICA and D6S273 for both the HLA DR3-DQ2 and DR4-DQ8 haplotypes.     

New revelations in susceptibility to autoimmune thyroiditis by the use of H2 and HLA class II transgenic models

H2 and HLA transgenes were utilized to clarify the role of class II genes in susceptibility to experimental autoimmune thyroiditis (EAT), a model for Hashimoto's thyroiditis (HT). Susceptibility was transferred by H2 class II transgenes to a resistant haplotype and by HLA-DRA/DRB1*0301 (DR3) transgene into class II-negative Ab0 mice. Mice with a HLA-DRB1*1502 (DR2) transgene remain resistant to mouse thyroglobulin (mTg)-induced EAT, illustrating the role of HLA-DRB1 polymorphism. A role for HLA-DQ polymorphism was shown with hTg-induced EAT in HLA-DQ*0301/DQB1*0302 (DQ8), but not HLA-DQ*0103/DQB1*0601 (DQ6), transgenic mice. Yet, both DQ8+ and DQ6+ mice were unresponsive to mTg. Single transgenes obviate the problems from DR/DQ linkage disequilibrium and may distinguish the degree of susceptibility and the response to shared or specific epitopes. The introduction of conserved Eak transgene into Ab0 mice reveals a new role for H2E molecules in EAT. Without H2A molecules, EalphaEbetab molecules and T cells respond to hTg or pTg with severe thyroiditis, but not to mTg, thus distinguishing self from nonself. However, IAb genes in resistant mice ameliorate Eak transgene-mediated thyroiditis, similar to the effect of Eak transgene on IAs-mediated EAT. Studies in HLA DQ/DR double transgenic mice simulating human haplotypes could reveal HLA class II gene interactions in HT.     

HLA-DQB1 alleles may influence the surface expression of DQ molecules in lymphomononuclear cells of type 1 diabetes mellitus patients

To evaluate the expression of human leucocyte antigen (HLA) class II (DR and DQ) molecules on lymphomononuclear cells involved in the pathogenesis of type 1 diabetes, we studied 20 patients and 20 controls matched to patients for age, sex and HLA class II profile. The coexpression of HLA and CD3, CD4, CD8, CD19 and CD14 molecules was evaluated by flow cytometry. HLA-DRB1, -DQA1 and -DQB1 alleles were assigned using amplified DNA hybridized with sequence-specific primers. The fluorescence intensity of HLA-DR and -DQ molecules observed on the surface of the lymphomononuclear cells of patients did not differ significantly from controls. Patients presented decreased percentage of double-positive CD4(+)/DQ(+) cells and increased percentage of CD19(+)/DR(+) cells, irrespective of the HLA class II profile; however, the more dramatic alteration of the lymphomononuclear phenotype profile was observed for patients possessing the HLA-DQB1*0201 allele. These patients exhibited decreased percentage of CD3(+), CD4(+), CD8(+), CD19(+) and CD14(+) cells bearing HLA-DQ molecules and decreased fluorescence intensity for HLA-DQ molecules on CD19(+) cells compared to patients without the DQB1*0201 allele. Although type 1 diabetes patients shared CD4/DQ or CD19/DR phenotype abnormalities, patients typed as DQB1*0201 presented additional abnormalities in terms of DQ expression and cell phenotypes bearing DQ molecules.     

Role of HLA class II genes in susceptibility and resistance to multiple sclerosis: studies using HLA transgenic mice

Multiple sclerosis (MS), an inflammatory and demyelinating autoimmune disease of CNS has both, a genetic and an environmental predisposition. Among all the genetic factors associated with MS susceptibility, HLA class II haplotypes such as DR2/DQ6, DR3/DQ2, and DR4/DQ8 show the strongest association. Although a direct role of HLA-DR alleles in MS have been confirmed, it has been difficult to understand the contribution of HLA-DQ alleles in disease pathogenesis, due to strong linkage disequilibrium. Population studies have indicated that DQ alleles may play a modulatory role in the progression of MS. To better understand the mechanism by which HLA-DR and -DQ genes contribute to susceptibility and resistance to MS, we utilized single and double transgenic mice expressing HLA class II gene(s) lacking endogenous mouse class II genes. HLA class II transgenic mice have helped us in identifying immunodominant epitopes of PLP in context of various HLA-DR and -DQ molecules. We have shown that HLA-DR3 transgenic mice were susceptible to PLP_91–110 induced experimental autoimmune encephalomyelitis (EAE), while DQ6 (DQB1*0601) and DQ8 (DQB1*0302) transgenic mice were resistant. Surprisingly DQ6/DR3 double transgenic mice were resistant while DQ8/DR3 mice showed higher disease incidence and severity than DR3 mice. The protective effect of DQ6 in DQ6/DR3 mice was mediated by IFNγ, while the disease exacerbating effect of DQ8 molecule was mediated by IL-17. Further, we have observed that myelin-specific antibodies play an important role in PLP_91–110 induced EAE in HLA-DR3DQ8 transgenic mice. Based on these observations, we hypothesize that epistatic interaction between HLA-DR and -DQ genes play an important role in predisposition to MS and our HLA transgenic mouse model provides a novel tool to study the effect of linkage disequilibrium in MS.     

Autoimmunity versus tolerance: analysis using HLAtransgenic mice

On the basis of our extensive studies on collagen induced arthritis in HLA class II transgenic mice, we proposed a hypothesis to explain role of shared epitope in rheumatoid arthritis (RA) association. According to our hypothesis, complementation between both DQ and DR molecules is required for susceptibility or protection from disease. While certain DQ alleles predispose individuals to RA, DRB1 molecule can modulate disease by shaping T-cell repertoire in the thymus by providing self-peptides and presented by DQ molecules. Using Aβo.DQ8 transgenic mice, we tested ability of peptides derived from HV3 of DR molecules, implicated in RA positively or negatively, to activate T cells. While the peptides derived from RA susceptible DR molecule were poor binders and poor in activating T cells, the peptides derived from RA resistant DR molecules were high affinity binders and efficient T-cell activators. Our experiments suggest that high affinity DR peptides could induce tolerance to autoimmunity while the low affinity peptides could be permissive to autoimmunity. Using peptide from DRB1*0402 molecule, known to be associated with resistance to RA, prior to induction of collagen induced arthritis prevents the onset of disease. Thus, self-peptides derived from HLA molecules could potentially generate tolerance or autoimmunity depending on their binding affinity with HLA molecules.     

HLA class II haplotype associations with inflammatory bowel disease in Jewish (Ashkenazi) and non-Jewish caucasian populations

Ulcerative colitis (UC) and Crohn's disease (CD) are the clinical entities comprising idiopathic inflammatory bowel disease (IBD). Previous studies on the association of IBD and human leukocyte antigen (HLA) class II genes suggested a role for HLA in this disease. Here we present HLA class II (DRB1, DQB1, DQA1, DPB1) allele and haplotype distributions determined using the polymerase chain reaction and sequence-specific oligonucleotide probe methods. A total of 578 UC and CD Caucasian patients and controls from Jewish (Ashkenazi) and non-Jewish populations was examined. Our previously reported association of DR1-DQ5 with CD was attributable to DRB1*0103. A dramatic association with IBD and the highly unusual DRB1*0103-DQA1*0501-DQB1*0301 haplotype (OR = 6.6, p = 0.036) was found. The more common DR1 haplotype, DRB1*0103-DQA1*0101-DQB1*0501, was also associated with IBD (OR = 3.1, p = 0.014), a result suggesting that interaction between DR and DQ may determine the extent of disease risk. Our previously reported association of DR2 with UC was attributable to DRB1*1502 (OR = 2.6, p = 0.006). At the DPB1 locus, a significant association of DPB1*0401 with CD was observed for the combined populations (OR = 1.85, p = 0.007). These observations indicate that some class II alleles and haplotypes confer susceptibility to both UC and CD, implying common immunogenetic mechanisms of pathogenesis, while others confer risk to only one of these diseases, and illustrate the value of DNA HLA typing in disease susceptibility analyses.     

Autoimmunity versus tolerance: analysis using transgenic mice

On the basis of our extensive studies on collagen induced arthritis in HLA class II transgenic mice, we proposed a hypothesis to explain role of shared epitope in rheumatoid arthritis (RA) association. According to our hypothesis, complementation between both DQ and DR molecules is required for susceptibility or protection from disease. While certain DQ alleles predispose individuals to RA, DRB1 molecule can modulate disease by shaping T-cell repertoire in the thymus by providing self-peptides and presented by DQ molecules. Using A beta o.DQ8 transgenic mice, we tested ability of peptides derived from HV3 of DR molecules, implicated in RA positively or negatively, to activate T cells. While the peptides derived from RA susceptible DR molecule were poor binders and poor in activating T cells, the peptides derived from RA resistant DR molecules were high affinity binders and efficient T-cell activators. Our experiments suggest that high affinity DR peptides could induce tolerance to autoimmunity while the low affinity peptides could be permissive to autoimmunity. Using peptide from DRB1*0402 molecule, known to be associated with resistance to RA, prior to induction of collagen induced arthritis prevents the onset of disease. Thus, self-peptides derived from HLA molecules could potentially generate tolerance or autoimmunity depending on their binding affinity with HLA molecules.     

Multiple sclerosis associated amino acids of polymorphic regions relevant for the HLA antigen binding are confined to HLA-DR2

Among the candidate genes for multiple sclerosis (MS), the strongest influence is conferred by human leucocyte antigen (HLA) class II genes, in particular the DR2, DQ6, Dw2 haplotype (DRB1*1501, DQA1*0102, DQB1*0602). Similar to other autoimmune diseases, it is not clear yet how the presence of a specific HLA-DR or -DQ molecule translates into an increased disease susceptibility. Previous observations by us and others imply a HLA-DR2 dependent propensity of antigen-specific T-cell lines to produce increased amounts of TNF-alpha/beta as one mechanism how DR2 could contribute to susceptibility. In this article, we investigated the distribution of polymorphic stretches of the DRB1, DQA1, and DQB1 chains known to be relevant for antigen binding, in 66 unrelated patients with relapsing remitting MS and 210 unrelated controls. We found a significant association with disease for the appearance of proline at position 11, arginine at position 13, and alanine at position 71 of HLA-DRbeta1. Surprisingly, we identified only residues preferentially expressed in the MS group that were related to HLA-DR2. Thus, the contribution of HLA class II to the pathogenesis of MS is not mediated by allele-overlapping antigen binding sites, but is confined to the disease associated HLA allele.     

HLA class II transgenic mice as models of human diseases

Summary: Predisposition co develop Various autoimmune disorders has been associated with certain HLA class II molecules but there is a lack of information on che pathophysiological rule of HLA genes in conferring susceptibility Various experimental animal models of autoimmune disease have been studied to address the role of immune response genes. To study the interactions involved between class II molecules (DQ and DR) and define the immunologic mechanisms in various diseases, we generated HLA-DR and DQ transgenic mice that lacked endogenous class II molecules. The HLA molecules in these mice arc expressed on the cell surface and can positively select CD4+ T cells expressing Various Vβ T-cell receptors (TCR). A peripheral tolerance is maintained co transgenic HLA molecules thus indicating that these molecules act as self, Mouse co stimulatory and accessory molecules can interact with the HLA-peptide-TCR complex leading to efficient T-cell activation. In this review, we describe immunogenetic models for human diseases using these transgenic mice. Our studies show that HLA class II transgene-restricted T cells recognize the immunodominant antigens and peptide epitopes, similar to HLA class II-restricted human T cells. Thus these mice provide powerful tools to understand the role of HLA class II molecules in predisposition and onset of human diseases and to develop immunotherapy and vaccines.     

Induction of CTL response by a minimal epitope vaccine in HLA A*0201/DR1 transgenic mice: dependence on HLA class II restricted T(H) response

CTL play a pivotal role in the immune response during viral infections. In this study, the HLA class II restricted T(H) requirement for optimal in vivo induction of HLA class I restricted CTL responses has been investigated. Towards this goal, transgenic mice expressing both HLA class I (A*0201 or A2.1) and class II (DRB1*0101 or DR1) molecules have been derived. Immunization of these mice with an HLA A*0201-restricted and CMV-specific CTL epitope (pp65(495-503)), and either of three different tetanus toxin-derived MHC class II-binding T(H) epitopes, resulted in a vigorous CTL response. CTL specific for the pp65(495-503) epitope were dramatically enhanced in mice expressing both the HLA-DR1 and HLA-A*0201 transgenes. Notably, preinjection of three TT peptides (TT(639-652), TT(830-843), and TT(947-967)) increased the capability of HLA A*0201/DR1 Tg mice to respond to subsequent immunization with the T(H) + CTL peptide mixture. These results indicate that the use of HLA A*0201/DR1 Tg mice constitute a versatile model system (in lieu of immunizing humans) for the study of both HLA class I and class II restricted T-cell responses. These studies provide a rational model for the design and assessment of new minimal-epitope vaccines based on their in vivo induction of a pathogen-specific CTL response.     

CD4+T cells restricted by DQ not by DR support CD8+T cells to grow

Much attention has been paid whether there are any differences in regulating the human immune response between HLA-DR and -DQ molecules encoded by the genes within the HLA class II multigene family. Previous studies have suggested that HLA DQ molecules control low responsiveness through activating CD4 T cells which generate CD8 positive T cells, whereas HLA -DR molecules control high responsiveness through activating CD4 helper T cells. To examine this model we investigated the streptococcal cell wall antigen (SCW) specific T cell lines restricted by either DR or DQ molecule. To identify the restricting molecules, L cell transfectants expressing DQw1, DR2AB1 or DR2AB5 from Dw12 haplotype or DQw4, DR4 or DRw53 from DW15 haplotype were used. 1. From individuals with Dw12 which is a low responder haplotype to SCW, T cell clones specific to SCW and restricted by HLA-DQw1 or DR2 were identified, whereas from individuals with Dw15 which is a high responder haplotype, only DR4 or DRw53 restricted T cell clones were identified and DQw4 restricted T cells were never observed. 2. SCW specific CD4 T cells restricted by DQw1 were able to support the growth of CD8 positive cells, whereas those restricted by DR4 could not do so. 3. The CD8 T cells also required autologous antigen presenting cells and SCW to grow, and they completely blocked the immune response to SCW in vitro. These observations clearly demonstrated the distinct function of HLA-DQ and -DR molecules in regulating the human immune response to SCW.     

To B or not to B: role of B cells in pathogenesis of arthritis in HLA transgenic mice

Population studies have shown that amongst all the genetic factors linked with autoimmune disease development, MHC class II genes are the most significant. Experimental autoimmune arthritis resembling human rheumatoid arthritis (RA) can be induced in susceptible strains of mice following immunization with type II collagen (CIA). We generated transgenic mice lacking endogenous class II molecules and expressing various HLA genes including RA-associated, HLA-DRB1*0401 and HLA-DQ8, and RA-resistant, DRB1*0402, genes. The HLA molecules in these mice are expressed on the cell surface and can positively select CD4+ T cells expressing various Vβ T cell receptors. Endogenous class II invariant chain is required for proper functioning of the class II transgene. Arthritis development in transgenic mice is CD4+ and B cells dependent. Studies in humanized mice showed that B cells are required as antigen presenting cells in addition to antibody producing cells for the development of CIA. The transgenic mice expressing *0401 and *0401/DQ8 genes developed sex-biased arthritis with predominantly females being affected, similar to that of human RA. Further, the transgenic mice produced autoantibodies like rheumatoid factor and anti-cyclic antibodies. Antigen presentation by B cells leads to a sex-specific immune response in DRB1*0401 mice suggesting a role of B cells and HLA-DR in rendering susceptibility to develop arthritis in females.     

Identification of T cell epitopes on human proteolipid protein and induction of experimental autoimmune encephalomyelitis in HLA class II-transgenic mice

Multiple sclerosis (MS) is an immune-mediated demyelinating disease of the CNS that is associated with HLA class II molecules HLA-DR2, -DR3 and -DR4. Previously, it has been difficult to analyze the role of individual HLA molecules in disease pathogenesis due to heterogeneity of MHC genes, linkage disequilibrium, influence of non-MHC genes and contribution of environment. To overcome some of these problems, we have generated HLA-transgenic (tg) mice to investigate function and interaction of these molecules in disease pathogenesis. To investigate the role of individual HLA class II genes in immune responses to human proteolipid protein (PLP), a candidate autoantigen in MS, mice expressing HLA genes DR2, DR3, DR4 (DRB1*0401 and DRB1*0402), DQ6 and DQ8, lacking endogenous class II molecules were immunized with overlapping peptides of PLP. In all tg mice, the majority of the dominant T cell epitopes were clustered mainly to three region; amino acids 31-70, 91-120 and 178-228, of the PLP molecules. We also identified an encephalitogenic epitope PLP(91-110) that induced clinical EAE in HLA-DR3 tg mice. These tg mice had inflammatory infiltrates classically associated with EAE and showed a Th1 cytokine profile. This humanized mouse model of MS will be valuable in deciphering the role of HLA molecules and autoantigens in MS.     

HLA class II phenotype controls diversification of the autoantibody response in primary Sjögren's syndrome (pSS)

The coexistence of anti-La (SS-B) and anti-Ro (SS-A) autoantibodies in pSS is probably explained by intermolecular spreading of autoimmunity toward different components of the La/Ro ribonucleoprotein (RNP). In order to evaluate the role of the HLA class II phenotype in controlling diversification of this autoantibody response, 80 patients with pSS were typed by polymerase chain reaction sequence-specific oligonucleotide (PCR-SSO) at the HLA class II loci DRB1, DQA1 and DQB1. Serum samples were examined for anti-La and anti-Ro by counterimmunoelectrophoresis and by ELISA using purified recombinant La and 60-kD Ro proteins. Patient sera were classified according to the extent of diversification of the anti-La, anti-Ro response including the presence or absence of precipitating anti-La antibodies. Immunogenic characteristics of these stratified groups were then studied. All patients with pSS, with or without autoantibodies to Ro and La, were found to have at least one of the HLA-DRB1 types DR2, DR3 or DR5. The HLA DR3-DQA1*0501-DQB1*02 (DR3-DQ2) haplotype was primarily associated with a diversified La/Ro RNP response containing precipitating autoantibodies to La (P < 0.001); whereas the haplotype HLA DR2-DQA1*0102-DQB1*0602 (DR2-DQ1) was associated with a less diversified La/Ro RNP response containing non-precipitating (restricted epitope) anti-La autoantibodies (P < 0.001). Anti-La-positive patients lacking both HLA-DR2 and HLA-DR3 all expressed the HLA-DQA1*0501 allele, which was present at increasing frequency with greater diversification of the anti-La/Ro autoantibody response. The association of distinct HLA haplotypes with different degrees of autoantibody diversification in patients with pSS suggests a model of HLA-restricted presentation of La/Ro peptide determinants to autoreactive helper T cells. We propose that non-precipitating anti-La responses are driven by limited intermolecular help from DR2-DQ1-restricted T helper cells recognizing Ro determinants. On the other hand, we speculate that the more diversified, precipitating anti-La responses obtain more efficient cognate T help from DR3-DQ2-restricted T helper cells recognizing La determinants, where HLA-DQA1*0501 may be a critical determinant for antigen presentation.     

Role of HLA class II genes in susceptibility/resistance to inflammatory arthritis: studies with humanized mice

Summary: Predisposition to develop rheumatoid arthritis (RA) has been associated with certain human leukocyte antigen (HLA) class II molecules, although the mechanism is still unknown. Various experimental animal models of inflammatory arthritis have been studied to address the role of major histocompatibility complex (MHC) genes in pathogenesis. We have generated transgenic mice expressing HLA class II molecules (DR and DQ) lacking complete endogenous class II molecules to study the interactions involved between class II molecules (DQ and DR) and to define the immunologic mechanisms in inflammatory arthritis. The HLA transgene can positively select CD4⁺ T cells expressing various Vβ T-cell receptors, and a peripheral tolerance is maintained to transgenic HLA molecules. The expression of HLA molecules on various cells in these mice is similar to that known in humans. In this review, we describe collagen-induced arthritis as a model for human inflammatory arthritis using these transgenic mice. The transgenic mice carrying RA-susceptible haplotype develop gender-biased inflammatory arthritis with clinical and histopathological similarities to RA. Our studies show that polymorphism of HLA class II genes determine the predisposition to rheumatoid/inflammatory arthritis and the epistatic interactions between HLA-DQ and HLA-DR molecules dictate the severity, progression, and modulation of the disease.     

Characteristics of HLA class I and class II polymorphisms in Rwandan women

OBJECTIVE: To define HLA class I and class II polymorphisms in Rwandans. METHODS: PCR-based HLA genotyping techniques were used to resolve variants of HLA-A, B, and C to their 2- or 4-digit allelic specificities, and those of DRB1 and DQB1 to their 4- or 5-digit alleles. RESULTS: Frequencies of 14 A, 8 C, and 14 B specificities and of 13 DRB1 and 8 DQB1 alleles were >/=0.02 in a group of 280 Rwandan women. These major HLA factors produced 6 haplotypes extending across the class I and class II regions: A*01-Cw*04-B* 4501-DRB1*1503-DQB1*0602 (A1-Cw4-B12- DR15 - DQ6), A * 01 - Cw * 04 - B * 4901 -DRB1 * 1302-DQB1*0604 (A1-Cw4-B21-DR13-DQ6), A*30 - Cw*04 - B*15 - DRB1*1101 - DQB1*0301 (A19-Cw4-B15-DR11-DQ7), A*68-Cw*07-B* 4901-DRB1*1302-DQB1*0604(A28-Cw7-B21- DR13 - DQ6), A*30 - Cw*07 - B*5703 - DRB1* 1303-DQB1*0301(A19 - Cw7 - B17 - DR13 - DQ7), and A*74-Cw*07-B*4901-DRB1*1302-DQB1* 0604 (A19-Cw7-B21-DR13-DQ6), respectively. Collectively, these extended haplotypes accounted for about 19% of the total. Other apparent class I-class II haplotypes (e.g., Cw*17-B*42-DRB1*0302-DQB1*0402, Cw*06- B*58-DRB1*1102-DQB1*0301, and Cw*03- B*15-DRB1*03011-DQB1*0201) did not extend to the telomeric HLA-A locus, and other 3-locus class I haplotypes (e.g., A*68-Cw*04-B*15, A*74-Cw*04-B*15, and A*23-Cw*07-B*4901) completely or partially failed to link with any specific class II alleles. DISCUSSION: Frequent recombinations appeared to occur between the three evolutionarily conserved HLA blocks carrying the class I and class II loci. The HLA class I profile seen in Rwandans was not directly comparable with those known in the literature, although the class II profile appeared to resemble those in several African populations. These data provide additional evidence for the extensive genetic diversity in Africans.     

Functional difference between HLA-DR and HLA-DQ molecules in mixed lymphocyte reaction

In order to analyze the functional differences between HLA-DR and HLA-DQ molecules, we have established transfectants expressing HLA class II molecules. We investigated the contribution of these molecules in mixed lymphocyte reaction (MLR) using these transfectants. 1) The genomic clones encoding for DR alpha, DR beta, DQ alpha, and DQ beta from HLA-Dw 15 haplotype were isolated. These genes were introduced into murine L cells and two kinds of stable transfectants expressing either of HLA-DR4 and HLA-DQw4 were established. Expression of HLA class II molecules on transfectants was confirmed by FACS analysis using monoclonal antibodies specific for HLA class II molecules. 2) Primary MLR against class II transfectants and blocking experiments showed that DR molecules function as dominant stimulator molecules in allo MLR, whereas DQ molecules as well as DR molecules stimulate equally auto MLR. 3) We also determined the clone size of MLR reactive CD4+ T cells by the limiting dilution analysis. Frequencies of allo DR, auto DQ, and allo DQ reactive CD4+ T cells was estimated to be almost equal, but frequency of auto DR reactive CD4+ T cells was estimated to be far low. These results suggest the relatively high occurrence of auto DQ reactive clones which contribute significantly to auto MLR. These auto DQ reactive clones may not be eliminated as efficiently as DR reactive clones, because of lower expression of DQ molecules than DR molecules on bone marrow derived cells.