Airway Memory CD4(+) T Cells Mediate Protective Immunity against Emerging Respiratory Coronaviruses.

Two zoonotic coronaviruses (CoVs)-SARS-CoV and MERS-CoV-have crossed species to cause severe human respiratory disease. Here, we showed that induction of airway memory CD4(+) T cells specific for a conserved epitope shared by SARS-CoV and MERS-CoV is a potential strategy for developing pan-coronavirus vaccines. Airway memory CD4(+) T cells differed phenotypically and functionally from lung-derived cells and were crucial for protection against both CoVs in mice. Protection was dependent on interferon-γ and required early induction of robust innate and virus-specific CD8(+) T cell responses. The conserved epitope was also recognized in SARS-CoV- and MERS-CoV-infected human leukocyte antigen DR2 and DR3 transgenic mice, indicating potential relevance in human populations. Additionally, this epitope was cross-protective between human and bat CoVs, the progenitors for many human CoVs. Vaccine strategies that induce airway memory CD4(+) T cells targeting conserved epitopes might have broad applicability in the context of new CoVs and other respiratory virus outbreaks.

Concomitant Disruption of CD4 and CD8 Genes Facilitates the Development of Double Negative αß TCR+ Peripheral T Cells That Respond Robustly to Staphylococcal Superantigen.

Mature peripheral double negative T (DNT) cells expressing αß TCR but lacking CD4/CD8 coreceptors play protective as well as pathogenic roles. To better understand their development and functioning in vivo, we concomitantly inactivated CD4 and CD8 genes in mice with intact MHC class I and class II molecules with the hypothesis that this would enable the development of DNT cells. We also envisaged that these DNT cells could be activated by bacterial superantigens in vivo as activation of T cells by superantigens does not require CD4 and CD8 coreceptors. Because HLA class II molecules present superantigens more efficiently than murine MHC class II molecules, CD4 CD8 double knockout (DKO) mice transgenically expressing HLA-DR3 or HLA-DQ8 molecules were generated. Although thymic cellularity was comparable between wild type (WT) and DKO mice, CD3+ αß TCR+ thymocytes were significantly reduced in DKO mice, implying defects in thymic-positive selection. Splenic CD3+ αß TCR+ cells and Foxp3+ T regulatory cells were present in DKO mice but significantly reduced. However, the in vivo inflammatory responses and immunopathology elicited by acute challenge with the staphylococcal superantigen enterotoxin B were comparable between WT and DKO mice. Choric exposure to staphylococcal enterotoxin B precipitated a lupus-like inflammatory disease with characteristic lympho-monocytic infiltration in lungs, livers, and kidneys, along with production of anti-nuclear Abs in DKO mice as in WT mice. Overall, our results suggest that DNT cells can develop efficiently in vivo and chronic exposure to bacterial superantigens may precipitate a lupus-like autoimmune disease through activation of DNT cells.

Identifying a Small Molecule Blocking Antigen Presentation in Autoimmune Thyroiditis.

We previously showed that an HLA-DR variant containing arginine at position 74 of the DRß1 chain (DRß1-Arg74) is the specific HLA class II variant conferring risk for autoimmune thyroid diseases (AITD). We also identified 5 thyroglobulin (Tg) peptides that bound to DRß1-Arg74. We hypothesized that blocking the binding of these peptides to DRß1-Arg74 could block the continuous T-cell activation in thyroiditis needed to maintain the autoimmune response to the thyroid. The aim of the current study was to identify small molecules that can block T-cell activation by Tg peptides presented within DRß1-Arg74 pockets. We screened a large and diverse library of compounds and identified one compound, cepharanthine that was able to block peptide binding to DRß1-Arg74. We then showed that Tg.2098 is the dominant peptide when inducing experimental autoimmune thyroiditis (EAT) in NOD mice expressing human DRß1-Arg74. Furthermore, cepharanthine blocked T-cell activation by thyroglobulin peptides, in particular Tg.2098 in mice that were induced with EAT. For the first time we identified a small molecule that can block Tg peptide binding and presentation to T-cells in autoimmune thyroiditis. If confirmed cepharanthine could potentially have a role in treating human AITD.

A Central Role for HLA-DR3 in Anti-Smith Antibody Responses and Glomerulonephritis in a Transgenic Mouse Model of Spontaneous Lupus.

MHC, especially HLA-DR3 and HLA-DR2, is one of the most important genetic susceptibility regions for systemic lupus erythematosus. Human studies to understand the role of specific HLA alleles in disease pathogenesis have been hampered by the presence of strong linkage disequilibrium in this region. To overcome this, we produced transgenic mice expressing HLA-DR3 (DRß1*0301) and devoid of endogenous class II (both I-A and I-E genes, AE(0)) on a lupus-prone NZM2328 background (NZM2328.DR3(+)AE(0)). Both NZM2328 and NZM2328.DR3(+)AE(0) mice developed anti-dsDNA and glomerulonephritis, but anti-dsDNA titers were higher in the latter. Although kidney histological scores were similar in NZM2328 and NZM2328.DR3(+)AE(0) mice (7.2 ± 4.3 and 8.6 ± 5.7, respectively, p = 0.48), the onset of severe proteinuria occurred earlier in NZM2328.DR3(+)AE(0) mice compared with NZM2328 mice (median, 5 and 9 mo respectively, p < 0.001). Periarterial lymphoid aggregates, classic wire loop lesions, and occasional crescents were seen only in kidneys from NZM2328.DR3(+)AE(0) mice. Interestingly, NZM2328.DR3(+)AE(0) mice, but not NZM2328 mice, spontaneously developed anti-Smith (Sm) Abs. The anti-Sm Abs were seen in NZM2328.DR3(+)AE(0) mice that were completely devoid of endogenous class II (AE(-/) (-)) but not in mice homozygous (AE(+/+)) or heterozygous (AE(+/-)) for endogenous MHC class II. It appears that only HLA-DR3 molecules can preferentially select SmD-reactive CD4(+) T cells for generation of the spontaneous anti-Sm immune response. Thus, our mouse model unravels a critical role for HLA-DR3 in generating an autoimmune response to SmD and lupus nephritis in the NZM2328 background.

The viral transcription group determines the HLA class I cellular immune response against human respiratory syncytial virus.

The cytotoxic T-lymphocyte-mediated killing of virus-infected cells requires previous recognition of short viral antigenic peptides bound to human leukocyte antigen class I molecules that are exposed on the surface of infected cells. The cytotoxic T-lymphocyte response is critical for the clearance of human respiratory syncytial virus infection. In this study, naturally processed viral human leukocyte antigen class I ligands were identified with mass spectrometry analysis of complex human leukocyte antigen-bound peptide pools isolated from large amounts of human respiratory syncytial virus-infected cells. Acute antiviral T-cell response characterization showed that viral transcription determines both the immunoprevalence and immunodominance of the human leukocyte antigen class I response to human respiratory syncytial virus. These findings have clear implications for antiviral vaccine design.

Systemic inflammatory response elicited by superantigen destabilizes T regulatory cells, rendering them ineffective during toxic shock syndrome.

Life-threatening infections caused by Staphylococcus aureus, particularly the community-acquired methicillin-resistant strains of S. aureus, continue to pose serious problems. Greater virulence and increased pathogenicity of certain S. aureus strains are attributed to higher prevalence of exotoxins. Of these exotoxins, the superantigens (SAg) are likely most pathogenic because of their ability to rapidly and robustly activate the T cells even in extremely small quantities. Therefore, countering SAg-mediated T cell activation using T regulatory cells (Tregs) might be beneficial in diseases such as toxic shock syndrome (TSS). As the normal numbers of endogenous Tregs in a typical host are insufficient, we hypothesized that increasing the Treg numbers by administration of IL-2/anti-IL-2 Ab immune complexes (IL2C) or by adoptive transfer of ex vivo expanded Tregs might be more effective in countering SAg-mediated immune activation. HLA-DR3 transgenic mice that closely recapitulate human TSS were treated with IL2C to increase endogenous Tregs or received ex vivo expanded Tregs. Subsequently, they were challenged with SAg to induce TSS. Analyses of various parameters reflective of TSS (serum cytokine/chemokine levels, multiple organ pathology, and SAg-induced peripheral T cell expansion) indicated that increasing the Tregs failed to mitigate TSS. On the contrary, serum IFN-γ levels were increased in IL2C-treated mice. Exploration into the reasons behind the lack of protective effect of Tregs revealed IL-17 and IFN-γ-dependent loss of Tregs during TSS. In addition, significant upregulation of glucocorticoid-induced TNFR family-related receptor on conventional T cells during TSS could render them resistant to Treg-mediated suppression, contributing to failure of Treg-mediated immune regulation.

DRB1*0402 may influence arthritis by promoting naive CD4+ T-cell differentiation in to regulatory T cells.

HLA-DRB1*0401 expression in humans has been associated with a predisposition to developing rheumatoid arthritis (RA) and collagen-induced arthritis (CIA), while HLA-DRB1*0402 is not associated with susceptibility. Here, we determined if mice transgenic (Tg) for human *0401 have a CD4+ T-cell repertoire that predetermines proinflammatory cytokine production. The data show that both *0401 and *0402 Tg mice can produce TH1/TH17 cytokines, although the kinetics of response may be different. However, in the context of antigen-specific responses in a CIA model, *0402 Tg mice generate a TH2 response that may explain their resistance to developing arthritis. In addition, a significant subset of naïve CD4+ T cells from *0402 Tg mice can be activated in polarizing conditions to differentiate into Treg cells that produce IFN-γ. *0401 Tg mice harbor memory CD4+ T cells that differentiate into IL-17(+) cells in various polarizing conditions. Our data suggest that *0401 Tg mice generate a strong immune response to lipopolysaccharide and may be efficient in clearing infection, and may *0401 have been evolutionarily selected for this ability. Autoimmunity, such as RA, could likely be a bystander effect of the cytokine storm that, along with the presence of low Treg-cell numbers in *0401 Tg mice, causes immune dysregulation.

HLA class II molecules influence susceptibility versus protection in inflammatory diseases by determining the cytokine profile.

The MHC in humans encodes the most polymorphic genes, the HLA genes, which are critical for the immune system to clear infection. This can be attributed to strong selection pressure as populations moved to different parts of the world and encountered new kinds of infections, leading to new HLA class II alleles. HLA genes also have the highest relative risk for autoimmune diseases. Three haplotypes, that is, HLA-DR2DQ6, DR4DQ8, and DR3DQ2, account for HLA association with most autoimmune diseases. We hypothesize that these haplotypes, along with their multiple subtypes, have survived bottlenecks of infectious episodes in human history because of their ability to present pathogenic peptides to activate T cells that secrete cytokines to clear infections. Unfortunately, they also present self-peptides/mimics to activate autoreactive T cells secreting proinflammatory cytokines that cause autoimmune diseases.

HLA class II (DR0401) molecules induce Foxp3+ regulatory T cell suppression of B cells in Plasmodium yoelii strain 17XNL malaria.

Unlike human malaria parasites that induce persistent infection, some rodent malaria parasites, like Plasmodium yoelii strain 17XNL (Py17XNL), induce a transient (self-curing) malaria infection. Cooperation between CD4 T cells and B cells to produce antibodies is thought to be critical for clearance of Py17XNL parasites from the blood, with major histocompatibility complex (MHC) class II molecules being required for activation of CD4 T cells. In order to better understand the correspondence between murine malaria models and human malaria, and in particular the role of MHC (HLA) class II molecules, we studied the ability of humanized mice expressing human HLA class II molecules to clear Py17XNL infection. We showed that humanized mice expressing HLA-DR4 (DR0401) molecules and lacking mouse MHC class II molecules (EA(0)) have impaired production of specific antibodies to Py17XNL and cannot cure the infection. In contrast, mice expressing HLA-DR4 (DR0402), HLA-DQ6 (DQ0601), HLA-DQ8 (DQ0302), or HLA-DR3 (DR0301) molecules in an EA(0) background were able to elicit specific antibodies and self-cure the infection. In a series of experiments, we determined that the inability of humanized DR0401.EA(0) mice to elicit specific antibodies was due to expansion and activation of regulatory CD4(+) Foxp3(+) T cells (Tregs) that suppressed B cells to secrete antibodies through cell-cell interactions. Treg depletion allowed the DR0401.EA(0) mice to elicit specific antibodies and self-cure the infection. Our results demonstrated a differential role of MHC (HLA) class II molecules in supporting antibody responses to Py17XNL malaria and revealed a new mechanism by which malaria parasites stimulate B cell-suppressogenic Tregs that prevent clearance of infection.

Nonequivalence of classical MHC class I loci in ability to direct effective antiviral immunity.

Structural diversity in the peptide binding sites of the redundant classical MHC antigen presenting molecules is strongly selected in humans and mice. Although the encoded antigen presenting molecules overlap in antigen presenting function, differences in polymorphism at the MHC I A, B and C loci in humans and higher primates indicate these loci are not functionally equivalent. The structural basis of these differences is not known. We hypothesize that classical class I loci differ in their ability to direct effective immunity against intracellular pathogens. Using a picornavirus infection model and chimeric H-2 transgenes, we examined locus specific functional determinants distinguishing the ability of class I sister genes to direct effective anti viral immunity. Whereas, parental FVB and transgenic FVB mice expressing the H-2K(b) gene are highly susceptible to persisting Theiler's virus infection within the CNS and subsequent demyelination, mice expressing the D(b) transgene clear the virus and are protected from demyelination. Remarkably, animals expressing a chimeric transgene, comprised primarily of K(b) but encoding the peptide binding domain of D(b), develop a robust anti viral CTL response yet fail to clear virus and develop significant demyelination. Differences in expression of the chimeric K(b)α1α2D(b) gene (low) and D(b) (high) in the CNS of infected mice mirror expression levels of their endogenous H-2(q) counterparts in FVB mice. These findings demonstrate that locus specific elements other than those specifying peptide binding and T cell receptor interaction can determine ability to clear virus infection. This finding provides a basis for understanding locus-specific differences in MHC polymorphism, characterized best in human populations.

The role of HLA-DQ8 beta57 polymorphism in the anti-gluten T-cell response in coeliac disease.

Major histocompatibility complex (MHC) class II alleles HLA-DQ8 and the mouse homologue I-A(g7) lacking a canonical aspartic acid residue at position beta57 are associated with coeliac disease and type I diabetes. However, the role of this single polymorphism in disease initiation and progression remains poorly understood. The lack of Asp 57 creates a positively charged P9 pocket, which confers a preference for negatively charged peptides. Gluten lacks such peptides, but tissue transglutaminase (TG2) introduces negatively charged residues at defined positions into gluten T-cell epitopes by deamidating specific glutamine residues on the basis of their spacing to proline residues. The commonly accepted model, proposing that HLA-DQ8 simply favours binding of negatively charged peptides, does not take into account the fact that TG2 requires inflammation for activation and that T-cell responses against native gluten peptides are found, particularly in children. Here we show that beta57 polymorphism promotes the recruitment of T-cell receptors bearing a negative signature charge in the complementary determining region 3beta (CDR3beta) during the response against native gluten peptides presented by HLA-DQ8 in coeliac disease. These T cells showed a crossreactive and heteroclitic (stronger) response to deamidated gluten peptides. Furthermore, gluten peptide deamidation extended the T-cell-receptor repertoire by relieving the requirement for a charged residue in CDR3beta. Thus, the lack of a negative charge at position beta57 in MHC class II was met by negatively charged residues in the T-cell receptor or in the peptide, the combination of which might explain the role of HLA-DQ8 in amplifying the T-cell response against dietary gluten.

The impact of Staphylococcus aureus-associated molecular patterns on staphylococcal superantigen-induced toxic shock syndrome and pneumonia.

Staphylococcus aureus is capable of causing a spectrum of human illnesses. During serious S. aureus infections, the staphylococcal pathogen-associated molecular patterns (PAMPs) such as peptidoglycan, lipoteichoic acid, and lipoproteins and even intact S. aureus, are believed to act in conjunction with the staphylococcal superantigens (SSAg) to activate the innate and adaptive immune system, respectively, and cause immunopathology. However, recent studies have shown that staphylococcal PAMPs could suppress inflammation by several mechanisms and protect from staphylococcal toxic shock syndrome, a life-threatening systemic disease caused by toxigenic S. aureus. Given the contradictory pro- and anti-inflammatory roles of staphylococcal PAMPs, we examined the effects of S. aureus-derived molecular patterns on immune responses driven by SSAg in vivo using HLA-DR3 and HLA-DQ8 transgenic mice. Our study showed that neither S. aureus-derived peptidoglycans (PGN), lipoteichoic acid (LTA), nor heat-killed Staphylococcus aureus (HKSA) inhibited SSAg-induced T cell proliferation in vitro. They failed to antagonize the immunostimulatory effects of SSAg in vivo as determined by their inability to attenuate systemic cytokine/chemokine response and reduce SSAg-induced T cell expansion. These staphylococcal PAMPs also failed to protect HLA-DR3 as well as HLA-DQ8 transgenic mice from either SSAg-induced toxic shock or pneumonia induced by a SSAg-producing strain of S. aureus.

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

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 V beta 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.

The copolymer P(HEMA-co-SS) binds gluten and reduces immune response in gluten-sensitized mice and human tissues.

BACKGROUND & AIMS: Copolymers of hydroxyethyl methacrylate and styrene sulfonate complex with isolated gliadin (the toxic fraction of gluten) and prevent damage to the intestinal barrier in HLA-HCD4/DQ8 mice. We studied the activity toward gluten and hordein digestion and biologic effects of poly(hydroxyethyl methacrylate-co-styrene sulfonate (P(HEMA-co-SS)). We also investigated the effect of gliadin complex formation in intestinal biopsy specimens from patients with celiac disease. METHODS: We studied the ability of P(HEMA-co-SS) to reduce digestion of wheat gluten and barley hordein into immunotoxic peptides using liquid chromatography-mass spectrometry. The biodistribution and pharmacokinetic profile of orally administered P(HEMA-co-SS) was established in rodents using tritium-labeled polymer. We assessed the capacity of P(HEMA-co-SS) to prevent the immunologic and intestinal effects induced by a gluten-food mixture in gluten-sensitized HLA-HCD4/DQ8 mice after short-term and long-term administration. We measured the effects of gliadin complex formation on cytokine release ex vivo using intestinal biopsy specimens from patients with celiac disease. RESULTS: P(HEMA-co-SS) reduced digestion of wheat gluten and barley hordein in vitro, thereby decreasing formation of toxic peptides associated with celiac disease. After oral administration to rodents, P(HEMA-co-SS) was predominantly excreted in feces, even in the presence of low-grade mucosal inflammation and increased intestinal permeability. In gluten-sensitized mice, P(HEMA-co-SS) reduced paracellular permeability, normalized anti-gliadin immunoglobulin A in intestinal washes, and modulated the systemic immune response to gluten in a food mixture. Furthermore, incubation of P(HEMA-co-SS) with mucosal biopsy specimens from patients with celiac disease showed that secretion of tumor necrosis factor-α was reduced in the presence of partially digested gliadin. CONCLUSIONS: The copolymer P(HEMA-co-SS) reduced digestion of wheat gluten and barley hordein and attenuated the immune response to gluten in a food mixture in rodents. It might be developed to prevent or reduce gluten-induced disorders in humans.

Uveitis-associated epitopes of retinal antigens are pathogenic in the humanized mouse model of uveitis and identify autoaggressive T cells.

Noninfectious uveitis is a leading cause of blindness and thought to involve autoimmune T cell responses to retinal proteins (e.g., retinal arrestin [soluble-Ag (S-Ag)]). There are no known biomarkers for the disease. Susceptibility is associated with HLA, but little is known about susceptible class II alleles or the potentially pathogenic epitopes that they present. Using a humanized HLA-transgenic mouse model of S-Ag-induced autoimmune uveitis, we identified several susceptible and resistant alleles of HLA-DR and -DQ genes and defined pathogenic epitopes of S-Ag presented by the susceptible alleles. The sequences of these epitopes overlap with some previously identified peptides of S-Ag ("M" and "N"), known to elicit memory responses in lymphocytes of uveitis patients. HLA-DR-restricted, S-Ag-specific CD4(+) T cells could be detected in blood and draining lymph nodes of uveitic mice with HLA class II tetramers and transferred the disease to healthy mice. Importantly, tetramer-positive cells were detected in peripheral blood of a uveitis patient. To our knowledge, these findings provide the first tangible evidence that an autoimmune response to retina is causally involved in pathogenesis of human uveitis, demonstrating the feasibility of identifying and isolating retinal Ag-specific T cells from uveitis patients and may facilitate their development as biomarkers for the disease.

Sensitization to gliadin induces moderate enteropathy and insulitis in nonobese diabetic-DQ8 mice.

Celiac disease (CD) is frequently diagnosed in patients with type 1 diabetes (T1D), and T1D patients can exhibit Abs against tissue transglutaminase, the auto-antigen in CD. Thus, gliadin, the trigger in CD, has been suggested to have a role in T1D pathogenesis. The objective of this study was to investigate whether gliadin contributes to enteropathy and insulitis in NOD-DQ8 mice, an animal model that does not spontaneously develop T1D. Gliadin-sensitized NOD-DQ8 mice developed moderate enteropathy, intraepithelial lymphocytosis, and barrier dysfunction, but not insulitis. Administration of anti-CD25 mAbs before gliadin-sensitization induced partial depletion of CD25(+)Foxp3(+) T cells and led to severe insulitis, but did not exacerbate mucosal dysfunction. CD4(+) T cells isolated from pancreatic lymph nodes of mice that developed insulitis showed increased proliferation and proinflammatory cytokines after incubation with gliadin but not with BSA. CD4(+) T cells isolated from nonsensitized controls did not response to gliadin or BSA. In conclusion, gliadin sensitization induced moderate enteropathy in NOD-DQ8 mice. However, insulitis development required gliadin-sensitization and partial systemic depletion of CD25(+)Foxp3(+) T cells. This humanized murine model provides a mechanistic link to explain how the mucosal intolerance to a dietary protein can lead to insulitis in the presence of partial regulatory T cell deficiency.

DQB1*0602 rather than DRB1*1501 confers susceptibility to multiple sclerosis-like disease induced by proteolipid protein (PLP).

BACKGROUND: Multiple sclerosis (MS) is associated with pathogenic autoimmunity primarily focused on major CNS-myelin target antigens including myelin basic protein (MBP), proteolipidprotein (PLP), myelin oligodendrocyte protein (MOG). MS is a complex trait whereby the HLA genes, particularly class-II genes of HLA-DR15 haplotype, dominate the genetic contribution to disease-risk. Due to strong linkage disequilibrium in HLA-II region, it has been hard to establish precisely whether the functionally relevant effect derives from the DRB1*1501, DQA1*0102-DQB1*0602, or DRB5*0101 loci of HLA-DR15 haplotype, their combinations, or their epistatic interactions. Nevertheless, most genetic studies have indicated DRB1*1501 as a primary risk factor in MS. Here, we used 'HLA-humanized' mice to discern the potential relative contribution of DRB1*1501 and DQB1*0602 alleles to susceptibility to "humanized" MS-like disease induced by PLP, one of the most prominent and encephalitogenic target-antigens implicated in human MS. METHODS: The HLA-DRB1*1501- and HLA-DQB1*0602-Tg mice (MHC-II(-/-)), and control non-HLA-DR15-relevant-Tg mice were immunized with a set of overlapping PLP peptides or with recombinant soluble PLP for induction of "humanized" MS-like disease, as well as for ex-vivo analysis of immunogenic/immunodominant HLA-restricted T-cell epitopes and associated cytokine secretion profile. RESULTS: PLP autoimmunity in both HLA-DR15-Tg mice was focused on 139-151 and 175-194 epitopes. Strikingly, however, the HLA-DRB1*1501-transgenics were refractory to disease induction by any of the overlapping PLP peptides, while HLA-DQB1*0602 transgenics were susceptible to disease induction by PLP139-151 and PLP175-194 peptides. Although both transgenics responded to both peptides, the PLP139-151- and PLP175-194-reactive T-cells were directed to Th1/Th17 phenotype in DQB1*0602-Tg mice and towards Th2 in DRB1*1501-Tg mice. CONCLUSIONS: While genome studies map a strong MS susceptibility effect to the region of DRB1*1501, our findings offer a rationale for potential involvement of pathogenic DQ6-associated autoimmunity in MS. Moreover, that DQB1*0602, but not DRB1*1501, determines disease-susceptibility to PLP in HLA-transgenics, suggests a potential differential, functional role for DQB1*0602 as a predisposing allele in MS. This, together with previously demonstrated disease-susceptibility to MBP and MOG in DRB1*1501-transgenics, also suggests a differential role for DRB1*1501 and DQB1*0602 depending on target antigen and imply a potential complex 'genotype/target antigen/phenotype' relationship in MS heterogeneity.

Human leukocyte antigen class II transgenic mouse model unmasks the significant extrahepatic pathology in toxic shock syndrome.

Among the exotoxins produced by Staphylococcus aureus and Streptococcus pyogenes, the superantigens (SAgs) are the most potent T-cell activators known to date. SAgs are implicated in several serious diseases including toxic shock syndrome (TSS), Kawasaki disease, and sepsis. However, the immunopathogenesis of TSS and other diseases involving SAgs are still not completely understood. The commonly used conventional laboratory mouse strains do not respond robustly to SAgs in vivo. Therefore, they must be artificially rendered susceptible to TSS by using sensitizing agents such as d-galactosamine (d-galN), which skews the disease exclusively to the liver and, hence, is not representative of the disease in humans. SAg-induced TSS was characterized using transgenic mice expressing HLA class II molecules that are extremely susceptible to TSS without d-galN. HLA-DR3 transgenic mice recapitulated TSS in humans with extensive multiple-organ inflammation affecting the lung, liver, kidneys, heart, and small intestines. Heavy infiltration with T lymphocytes (both CD4(+) and CD8+), neutrophils, and macrophages was noted. In particular, the pathologic changes in the small intestines were extensive and accompanied by significantly altered absorptive functions of the enterocytes. In contrast to massive liver failure alone in the d-galN sensitization model of TSS, findings of the present study suggest that gut dysfunction might be a key pathogenic event that leads to high morbidity and mortality in humans with TSS.

Two discreet subsets of CD8 T cells modulate PLP(91-110) induced experimental autoimmune encephalomyelitis in HLA-DR3 transgenic mice.

Previously we showed that transgenic mice expressing human HLA-DR3 gene are susceptible to PLP(91-110) induced experimental autoimmune encephalomyelitis (EAE) and can serve as an animal model of multiple sclerosis (MS). HLA-DR3 mice with EAE showed increased number of CD8 T cells indicating their important role in disease pathogenesis. The role of CD8 T cells in MS, an inflammatory demyelinating disease of CNS, has been enigmatic as it has been assigned both regulatory and pathogenic roles. Therefore, to evaluate the role of CD8 T cells, we generated CD8 deficient HLA-DR3 transgenic mice (DR3.CD8(-/-)). Immunization with PLP(91-110) led to more severe EAE in DR3.CD8(-/-) mice compared to HLA-DR3 mice indicating a regulatory role for CD8 T cells. Interestingly, DR3.CD8(-/-) mice with EAE showed decreased CNS pathology compared to DR3 mice thus suggesting a pathogenic role for CD8 T cells. We show that these two subsets of CD8 T cells can be differentiated based on the surface expression of CD122 (IL-2 Rß chain). CD8 T cells expressing CD122 (CD8+CD122+) play a regulatory role while CD8+CD122- T cells act as a pathogenic subset. CD122 expressing CD8 T cells are the regulatory subset of CD8 T cells and regulate the encephalitogenic CD4 T cells through direct modulation of antigen presenting cells and/or through the release of immunoregulatory cytokines such as IL-10, IFNγ and TGFß. We also showed that adoptive transfer of CD8CD122- T cells caused increased spinal cord demyelination indicating that these are pathogenic subset of CD8 T cells. Our study suggests that CD8+ T cells play both regulatory as well as pathogenic role in disease pathogenesis of EAE. A better understanding of these subsets could aid in designing novel therapy for MS patients.

Trans heterodimer between two non-arthritis-associated HLA alleles can predispose to arthritis in humanized mice.

OBJECTIVE: Certain HLA class II alleles are associated with susceptibility to the development of arthritis. However, the development of arthritis in some persons carrying non-rheumatoid arthritis (RA)-associated alleles remains unexplained. An individual who is heterozygous for the DQA1 and DQB1 genes can express the DQ molecule in cis or trans heterodimers. In a cis heterodimer, the α-chain interacts with the ß-chain coded by the same chromosome, while in a trans heterodimer it interacts with the ß-chain on the other chromosome. In this study, we used a humanized mouse model of arthritis in an attempt to determine whether a trans heterodimer of 2 nonassociated alleles, DQB1*0601 and DQB1*0604, can predispose to arthritis. METHODS: DQB1*0601 and *0604 occur in linkage with DQA1*0103 and *0102, respectively. To understand the role of trans heterodimers, we generated DQB1*0604/DQA1*0103-transgenic mice lacking endogenous HLA class II molecules. RESULTS: Severe arthritis developed in the DQB1*0604/A1*0103-trangenic mice, and an antigen-specific response was generated in vitro. DQB1*0604/DQA1*0103 presented type II collagen-derived peptides that were not presented by the arthritis-resistant DQB1*0601 allele, suggesting that trans heterodimer molecules between 2 DQB1 and DQA1 molecules may result in the presentation of unique antigens and susceptibility to the development of arthritis. Molecular modeling of type II collagen peptides showed that DQB1*0604/DQA1*0103 shares a p4 pocket with the arthritis-susceptible DQB1*0302 allele, suggesting a critical role of the p4 and p9 pockets in susceptibility to arthritis. CONCLUSION: These results provide a possible explanation for the parental inheritance of nonsusceptibility alleles in some patients with RA and a mechanism by which they can predispose to the development of arthritis.