Veena Taneja

Loss of Sex and Age Driven Differences in the Gut Microbiome Characterize Arthritis-Susceptible *0401 Mice but Not Arthritis-Resistant *0402 Mice

Background HLA-DRB1*0401 is associated with susceptibility, while HLA-DRB1*0402 is associated with resistance to developing rheumatoid arthritis (RA) and collagen-induced arthritis in humans and transgenic mice respectively. The influence of gut-joint axis has been suggested in RA, though not yet proven. Methodology/Principal Findings We have used HLA transgenic mice carrying arthritis susceptible and -resistant HLA-DR genes to explore if genetic factors and their interaction with gut flora gut can be used to predict susceptibility to develop arthritis. Pyrosequencing of the 16S rRNA gene from the fecal microbiomes of DRB1*0401 and DRB1*0402 transgenic mice revealed that the guts of *0401 mice is dominated by a Clostridium-like bacterium, whereas the guts of *0402 mice are enriched for members of the Porphyromonadaceae family and Bifidobacteria. DRB1*0402 mice harbor a dynamic sex and age-influenced gut microbiome while DRB1*0401 mice did not show age and sex differences in gut microbiome even though they had altered gut permeability. Cytokine transcripts, measured by rtPCR, in jejuna showed differential TH17 regulatory network gene transcripts in *0401 and *0402 mice. Conclusions/Significance We have demonstrated for the first time that HLA genes in association with the gut microbiome may determine the immune environment and that the gut microbiome might be a potential biomarker as well as contributor for susceptibility to arthritis. Identification of pathogenic commensal bacteria would provide new understanding of disease pathogenesis, thereby leading to novel approaches for therapy.

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.

An expansion of rare lineage intestinal microbes characterizes rheumatoid arthritis.

BackgroundThe adaptive immune response in rheumatoid arthritis (RA) is influenced by an interaction between host genetics and environment, particularly the host microbiome. Association of the gut microbiota with various diseases has been reported, though the specific components of the microbiota that affect the host response leading to disease remain unknown. However, there is limited information on the role of gut microbiota in RA. In this study we aimed to define a microbial and metabolite profile that could predict disease status. In addition, we aimed to generate a humanized model of arthritis to confirm the RA-associated microbe.MethodsTo identify an RA biomarker profile, the 16S ribosomal DNA of fecal samples from RA patients, first-degree relatives (to rule out environment/background as confounding factors), and random healthy non-RA controls were sequenced. Analysis of metabolites and their association with specific taxa was performed to investigate a potential mechanistic link. The role of an RA-associated microbe was confirmed using a human epithelial cell line and a humanized mouse model of arthritis.ResultsPatients with RA exhibited decreased gut microbial diversity compared with controls, which correlated with disease duration and autoantibody levels. A taxon-level analysis suggested an expansion of rare taxa, Actinobacteria, with a decrease in abundant taxa in patients with RA compared with controls. Prediction models based on the random forests algorithm suggested that three genera, Collinsella, Eggerthella, and Faecalibacterium, segregated with RA. The abundance of Collinsella correlated strongly with high levels of alpha-aminoadipic acid and asparagine as well as production of the proinflammatory cytokine IL-17A. A role for Collinsella in altering gut permeability and disease severity was confirmed in experimental arthritis.ConclusionsThese observations suggest dysbiosis in RA patients resulting from the abundance of certain rare bacterial lineages. A correlation between the intestinal microbiota and metabolic signatures could determine a predictive profile for disease causation and progression.

HLA transgenic mice as humanized mouse models of disease and immunity.

The application of molecular genetics to human disease in the past decade has been helpful in elucidating the genetic influences involved in the induction and pathogenesis of various autoimmune diseases. Among the numerous genes studied for their role in disease development, polymorphisms within HLA class I and class II loci play a significant role in predisposition to disease. HLA molecules are encoded by genes on the short arm of chromosome 6. Crystal structures of MHC molecules show a peptide binding cleft which contains the variable region of MHC molecules. Genetic polymorphism at the MHC determines the specificity and affinity of peptide binding and T cell recognition. HLA molecules play a pivotal role in T cell repertoire selection in the thymus and antigen presentation in the periphery. Analysis of T cell responses in humans has involved the use of T cell lines or clones in vitro from naturally primed individuals. On the other hand, MHC restriction, mapping of epitope recognition, and T cell function in murine systems have been determined by in vivo studies. Various experimental animal models of autoimmune diseases have been studied, contributing greatly to our basic understanding of the disease. For example, type II collagen‐induced arthritis (CIA) 1 in mice and rats has been used as an experimental model for RA. Even though the model differs in the manner polyarthritis is induced, disease expression is broadly similar to RA, with the occurrence of symmetrical peripheral polyarthritis and systemic inflammation. Although growing knowledge of the functions of T and B cells in the immune response has shed light on their role in disease induction, the pathogenic mechanisms of most autoimmune diseases remain unresolved. There are many unanswered questions as to how tolerance to self is usually maintained, because autoreactive T cells can be found in normal as well as diseased individuals. How is tolerance broken in autoimmunity? Do specific autoantigens trigger the immune system to mount tissue-destructive responses? These questions need to be solved for most of the autoimmune diseases. Although the strongest MHC association with an autoimmune disease is between HLA class I B27 and spondyloarthropathies, class II alleles are implicated in most other cases. For example, RA is strongly associated with alleles of the DRB1 locus, whereas diabetes shows a stronger association with DQB1 alleles. HLA-DR and -DQ alleles are inherited en bloc and are known to occur in linkage disequilibrium. To better understand the role of HLA molecules in autoimmune diseases, transgenic animals expressing human HLA genes associated with disease have been developed. The generation of transgenic mice expressing functional HLA molecules has been an important step toward the creation of an in vivo model for enhancing our understanding of the function of human molecules in disease induction and predisposition. The potential value of HLA transgenic animals as a “humanized” disease model was first illustrated in the HLA-B27

HLA Class II Transgenic Mice Mimic Human Inflammatory Diseases

Population studies have shown that among all the genetic factors linked with autoimmune disease development, MHC class II genes on chromosome 6 accounts for majority of familial clustering in the common autoimmune diseases. Despite the highly polymorphic nature of HLA class II genes, majority of autoimmune diseases are linked to a limited set of class II-DR or -DQ alleles. Thus a more detailed study of these HLA-DR and -DQ alleles were needed to understand their role in genetic predisposition and pathogenesis of autoimmune diseases. Although in vitro studies using class-II restricted CD4 T cells and purified class II molecules have helped us in understanding some aspects of HLA class-II association with disease, it is difficult to study the role of class II genes in vivo because of heterogeneity of human population, complexity of MHC, and strong linkage disequilibrium among different class II genes. To overcome this problem, we pioneered the generation of HLA-class II transgenic mice to study role of these molecule in inflammatory disease. These HLA class II transgenic mice were used to develop novel in vivo disease model for common autoimmune diseases such as rheumatoid arthritis, multiple sclerosis, insulin-dependent diabetes mellitus, myasthenia gravis, celiac disease, autoimmune relapsing polychondritis, autoimmune myocarditis, thyroiditis, uveitis, as well as other inflammatory disease such as allergy, tuberculosis and toxic shock syndrome. As the T-cell repertoire in these humanized HLA transgenic mice are shaped by human class II molecules, they show the same HLA restriction as humans, implicate potential triggering mechanism and autoantigens, and identify similar antigenic epitopes seen in human. This review describes the value of these humanized transgenic mice in deciphering role of HLA class II molecules in immunopathogenesis of inflammatory diseases.

Lessons from animal models for human autoimmune diseases.

A variety of different animal models are used to study autoimmune disease. What have we learned?

Arthritis susceptibility and the gut microbiome

Rheumatoid arthritis (RA) is an autoimmune disease with unknown etiology though both genetic and environmental factors have been suggested to be involved in its pathogenesis. While infections and other environmental factors (e.g. smoking) have been studied extensively and show some association, a direct link between all the factors has been difficult to prove. With the recent advances in technology, it has become possible to sequence the commensals that are residing in our gut. The gut microbiome may provide the missing link to this puzzle and help solve the mystery of many leaky gut syndromes. The gut commensals are involved in maintaining host immune homeostasis and function suggesting that they might be critical in altering the immune system, which leads to autoimmune diseases like RA. Mouse models support the role of the gut microbiota in predisposition to RA. If that is true, the power of gut‐derived commensal can be harnessed to our benefit by generating a biomarker profile along with genetic factors to define individuals at risk and by altering the gut microbial composition using various means.

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.

Delineating the Role of the HLA-DR4 “Shared Epitope” in Susceptibility versus Resistance to Develop Arthritis

In humans, HLA-DR alleles sharing amino acids at the third hypervariable region with DRB1*0401(shared epitope) are associated with a predisposition to rheumatoid arthritis, whereas DRB1*0402 is not associated with such a predisposition. Both DRB1*0402 and DRB1*0401 occur in linkage with DQ8 (DQB1*0302). We have previously shown that transgenic (Tg) mice expressing HLA-DRB1*0401 develop collagen-induced arthritis. To delineate the role of “shared epitope” and gene complementation between DR and DQ in arthritis, we generated DRB1*0402, DRB1*0401.DQ8, and DRB1*0402.DQ8 Tg mice lacking endogenous class II molecules, AE°. DRB1*0402 mice are resistant to develop arthritis. In double-Tg mice, the DRB1*0401 gene contributes to the development of collagen-induced arthritis, whereas DRB1*0402 prevents the disease. Humoral response to type II collagen is not defective in resistant mice, although cellular response to type II collagen is lower in *0402 mice compared with *0401 mice. *0402 mice have lower numbers of T cells in thymus compared with *0401 mice, suggesting that the protective effect could be due to deletion of autoreactive T cells. Additionally, DRB1*0402 mice have a higher number of regulatory T cells and show increased activation-induced cell death, which might contribute toward protection. In DRB1*0401.DQ8 mice, activated CD4+ T cells express class II genes and can present DR4- and DQ8-restricted peptides in vitro, suggesting a role of class II+ CD4 T cells locally in the joints. The data suggest that polymorphism in DRB1 genes determines predisposition to develop arthritis by shaping the T cell repertoire in thymus and activating autoreactive or regulatory T cells.

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.