The adaptive immune system is an antigen-specific structure that discriminates non-self molecules through the recognition of peptide antigens using receptor interactions between T-cells and antigen-presenting cells (APCs). This highly specific system uses receptor interaction between T-cells and APCs to discriminate self from nonself. Adaptive immunity establishes long-term immunological memory responses that trigger clonal expansion of T lymphocytes, which in turn cross-talk to B-cells to produce antigen-specific antibodies. The components of adaptive immunity are T and B lymphocytes, each with their own structurally unique cell receptors, which are somatically generated during thymic cell development. The adaptive immune system depends on the ability to assemble rearranged genes for both the T-cell receptor (TCR) and the immunoglobulin gene. This ability results from two genes known as RAG-1 and RAG-2 and their gene products that encode a recombinase involved in somatic recombination. The adaptive immune system allows T- and B-cells to generate an enormously diverse response to different pathogens. Both the naive T- and B-cell receptor repertoire are generated by interaction with self-ligands, such as the major histocompatibility complex (MHC), which in turn can signal to T- and B-cells to mature and survive. T-cells that are selected on self-ligands and sustained on self-ligands are termed “autoreactive T-cells.”
T-cells secrete large quantities of cytokines in response to antigen-specific activation and, based on their cytokine secretion profiles, are defined as T-helper type 1 (TH
2, or TH
17 (). TH
1 cells mature in response to interleukin (IL)-12 and produce interferon (IFN)-γ, which enhances cellular immunity and is important for intracellular defense, autoimmunity, and anti-tumor response. TH
2 cells develop in response to IL-4 and produce IL-4, IL-5, and IL-13, which enhance humoral immunity and are important for extracellular defense. IL-2 is essential for transforming growth factor-β–mediated induction of Foxp3+
regulatory T-cells (Tregs) and for the survival of Foxp3+
Tregs in the periphery (3
). Interest in Tregs has been heightened by evidence that anti-CD3 monoclonal antibody treatment reverses hyperglycemia in newly diagnosed NOD mice, and perhaps also in humans, as a result of the induction of regulatory T-cells (5
). Tregs can be expanded in vitro and in vivo and could be harnessed therapeutically to treat type 1 diabetes or facilitate tolerance to an allogeneic graft (7
). An exhaustive review of Tregs in type 1 diabetes is provided in this issue of Diabetes
FIG. 1. CD4+ T-cells have been subdivided into different subsets on the basis of their cytokine production and their functions. TH1, TH2, and Tregs have been well characterized with respect to factors influencing their development. Recently, another subset (more ...)
-17 subset of T helper cells was identified on the basis of its ability to produce IL-17A, IL-17F, and IL-22. TH
-17 cells were first recognized during assessment of the involvement of IL-23 in autoimmune disease. IL-23 is a member of the IL-6 family, and the nuclear receptor RORt acts as a key transcription factor in this lineage-commitment process (8
) (). TH
-17 cells provide protection in certain infections but have also been linked to a number of autoimmune diseases, a function previously assigned to TH
1 cells and IFN-γ. TH
-17 cells seem to mediate pathology in uveitis, multiple sclerosis, experimental autoimmune encephalomyelitis (EAE) psoriasis, and rheumatoid arthritis (9
), but it is not entirely clear if the TH
-17 pathway is influenced in type 1 diabetes. A recent study in NOD mice indicated that IFN-γ induced by an adjuvant-free antigen can reverse hyperglycemia, possibly as a result of suppression of pathogenic IL-17–producing cells (10
). However, additional research is necessary to establish to what extent TH
-17 cells play a role in type 1 diabetes pathogenesis.
The HLA complex.
Type 1 diabetes is a complex heterogeneous disease for which there is a small number of genes with large effects (i.e., HLA) and a large number of genes with small effects (11
). There are probably many genetic forms of type 1 diabetes, and most forms are influenced by genes within the HLA region on chromosome 6p21 (IDDM1
). Certain combinations of HLA alleles are found to be associated with each other on the same chromosome with a frequency greater than expected, and, consequently, they are not randomly distributed within the general population. This phenomenon is known as linkage disequilibrium, and it is quantified by the difference between the observed and the expected frequencies of certain combinations of alleles. It is the combination of these alleles on single chromosomes (haplotypes) and combinations of both chromosomes (one from each parent: genotype) that predominantly determines diabetes risk.
The principal genes localized within the MHC code for human leukocyte antigens, or HLA, two molecular classes of cell surface glycoproteins differing in structure, function, and tissue distribution. The genes that encode class I MHC consist of HLA-A, -B, and -C, whereas class II molecules are encoded by the DR, DQ, and DP genes.
APCs are cell populations specialized to take up pathogens and other antigens, present them to lymphocytes, and provide signals that stimulate the proliferation and differentiation of lymphocytes, generally T lymphocytes (). The main type of APC that gives rise to T-cell responses is the dendritic cell. Other cells functioning as APCs include macrophages, B-cells (12
), and, recently, stellate cells (Ito cells), which appear to be new actors in antigen presentation (13
). T-cell activation requires a sustained interaction between a naïve T-cell's TCR and the MCH-peptide complex on an antigen presenting (signal 1, signal 2, signal 3; ).
FIG. 2. Exogenous antigens are taken into the APCs, cleaved into peptides, and coupled with MHC molecules for presentation and recognition by naïve T-cells. T-cells bear on their surface unique receptors created by genetic recombinatorial processes. Signal (more ...)
Susceptibility to type 1 diabetes is conferred by specific HLA DR/DQ alleles (e.g., DRB1*03-DQB1*0201 [DR3] or DRB1*04-DQB1*0302 [DR4]) (14
). Each allele is simply given a number that represents a unique amino acid sequence. Each sequence binds only certain peptides and thus helps direct targeting of the immune system. The genotype associated with the highest risk for type 1 diabetes is the DR3/4-DQ8 (DQ8 is DQA1*0301, DQB1*0302) heterozygous genotype. In addition, HLA alleles such as DQB1*0602 are associated with dominant protection from type 1 diabetes in multiple populations (16
). There is a different disease risk for each MHC genotype, and although it is possible that only a single peptide epitope will relate to disease with multiple MHC genotypes, this remains to be experimentally evaluated.
Aly et al. (17
) provided evidence that risk for islet autoimmunity drastically increased in DR3/4-DQ2/DQ8 siblings who shared both HLA haplotypes identical by descent with their diabetic proband sibling (63% by age 7 years, and 85% by age 15 years) compared with siblings who did not share both HLA haplotypes with their diabetic proband sibling. These data suggest that HLA genotyping at birth may identify individuals at very high risk of developing type 1 diabetes before the occurrence of clear signs of islet autoimmunity and, eventually, overt disease.
Class I MHC molecules are expressed in virtually all nucleated cells, whereas class II molecule expression is restricted to B lymphocytes, dendritic cells, macrophages, and activated T lymphocytes. Peptides presented via MHC class I structures interact with CD8+
T-cells, while those peptides presented via MHC class II structures interact with CD4+
T-cells. Once a processed peptide is recognized through an MHC-TCR interaction, a cascade of signaling events occurs dependent upon the class of MHC structure recognized and the T-cell type that is activated. Once activated, CD4+
T-cells promote T-cell activation and differentiation, along with the ability to signal B-cells to generate an antibody response. CD8+
T-cells, when activated, produce inflammatory mediators and can directly target the destruction of specific peptide-presenting cells. Specific HLA class I alleles also influence diabetes risk, after correcting for linkage disequilibrium with DR and DQ alleles (18
Similar to other autoimmune disorders, in type 1 diabetes, CD4+
T-cells contribute to immune-mediated β-cell destruction. Accumulating evidence indicates that multiple islet antigens, including insulin, are targeted by MHC class II–restricted autoreactive T-cells (20
). In particular, a high degree of T-cell clonal expansion was observed in pancreatic lymph nodes from two long-term diabetic patients but not from control subjects. The oligoclonally expanded T-cells from diabetic subjects with DR4, which is a notorious susceptibility allele for type 1 diabetes, recognized the insulin A 1–15 epitope restricted by DR4. These experiments indicated that clonally expanded, autoreactive T-cells could be cloned out directly from the pancreatic draining lymph nodes of NOD mice. Although intriguing, there are a number of caveats to be considered in the interpretation of the abovementioned data. The two diabetic subjects had high glycemic levels and had longstanding insulin-dependent diabetes. Thus, it is possible that daily administration of exogenous insulin could initiate and sustain a systemic anti-insulin T-cell response. There is a need to have access to pancreatic and lymphoid tissue from cadaveric donors with signs of autoimmunity before disease onset to uncover the role for T-cell responses against islet autoantigens in disease pathogenesis.
By using MHC class II tetramers to probe the TCR specificity and avidity of GAD65-reactive T-cell clones isolated from patients with type 1 diabetes, Reijonen et al. (22
) identified high-avidity CD4+
T-cells from patients’ peripheral blood. The presence of autoreactive T-cells with potential preferential usage of TCR to diabetes-related autoantigens may serve as both a potential marker for disease progression and a target for immune manipulation in autoimmune diabetes.
Class I–restricted T-cells may also play a critical role in the development of autoimmune diabetes, as suggested by the observation that, in newly diagnosed diabetic patients, islet-infiltrating CD8+
T-cells represent the prevalent cell type of insulitis. The autoantigens targeted by autoreactive CD8+
T-cells in NOD mice appear to be insulin (23
) and the islet-specific glucose-6-phosphatase catalytic subunit–related protein (IGRIP) (24
Another intriguing observation provided evidence that peptide 10–18 of the insulin B-chain is related with recurrence of autoimmunity and loss of β-cell function in islet-grafted type 1 diabetes recipients (25
). Other investigations indicated that islet destruction is caused by autoreactive T-cells, whereas the tolerant nondiabetic state is characterized by autoreactive T-cells that secrete the immune suppressive cytokine, IL-10 (26
How much of type 1 diabetes pathoetiology is genetic and how much is environmental?
Perhaps the most convincing advancement in our knowledge of the genetics of type 1 diabetes derives from the discoveries of an autosomal recessive mutation on chromosome 21 causing the autoimmune polyendocrine syndrome type 1 (APS-I) (27
) and an X chromosome mutation leading to an X-linked autoimmunity-allergic dysregulation syndrome (XLAAD; also termed IPEX) (28
). The autoimmune polyendocrine syndrome type 1 is a rare syndrome with a relatively high incidence in Finland and Sardinia and among Iranian Jews, and it is characterized by type 1 diabetes, mucocutaneous candidiasis, hypoparathyroidism, Addison's disease, and hepatitis. This disease is caused by a mutation of the autoimmune regulator (AIRE) gene, which encodes a transcription factor. The gene product of AIRE is expressed in the thymus, and it might play an important role in maintaining self-tolerance to peripheral antigens such as insulin and other tissue-specific self-antigens in normal individuals (29
). Loss-of-function mutations in the AIRE gene in autoimmune polyendocrine syndrome type 1 patients and, for the most part, in mutant mice lead to progressive autoimmune destruction of many tissues, including the pancreatic islets, adrenal cortex, parathyroid glands, and gonads.
The syndrome XLAAD is associated with severe neonatal autoimmunity, which is characterized by mononuclear infiltration of multiple organs including pancreatic β-cells. The causative gene, FOXP3 (Foxp3 in mice), and its protein product that encodes a transcription repressor are specifically expressed in CD25+CD4+ T-cells in the thymus and in the periphery. Lack of such regulatory T-cells lead to overwhelming autoimmunity in humans and mice. This is an important syndrome to diagnose because bone marrow transplantation is an effective therapeutic approach restoring regulatory T-cells in these patients and, possibly, preventing type 1 diabetes.
The mechanisms by which class II genes influence susceptibility to or protection from type 1 diabetes have been a subject of endless discussions. The crystal structure of DQ8 and I-Ag7
revealed important similarities between these two MHC class II molecules, and this implies that antigen presentation may occur in a comparable fashion in both humans and NOD mice. As a matter of fact, both DQ8 and IA-g7 bind similar sets of peptides, including those representing immunodominant epitopes in NOD mice. Interestingly, in a transgenic NOD mouse model, the expression of an I-Aβ (the equivalent to the human class II DQB allele) transgene carrying Asp 57 instead of Ser 57 prevents these mice from developing diabetes (30
Brown et al. (31
) characterized the structure of the crystallized HLA class II molecule. One hypothesis is that effective antigen binding depends on the conformation of the antigen binding site on the DQ dimer. It has been postulated that a substitution of an amino acid residue at these positions of the DQ molecule leads to conformational changes of the antigen-binding site and, consequently, to a modification of the affinity of the class II molecule for the “diabetogenic” peptide(s). As support for this hypothesis, it is known that Asp-57 is involved in hydrogen and salt bonding with both the peptide main chain and the DRα Arg-76 side chain. There are several highly diabetogenic class II DQ molecules with aspartic acid at position 57, and thus it is the complete amino acid sequence rather that any single amino acid residue that is relevant (32
Autoimmunity is thought to result from an imbalance between the two functionally opposite processes, namely tolerance induction and immune responsiveness, each of which is dependent on the presence of MHC class I and class II molecules with appropriate structures (dictated by the genes encoding them) that are able to present antigenic peptides. In genetically susceptible individuals, certain class II molecules may poorly present self-peptides because of inefficiencies in the peptide-MHC structural interaction of these molecules, thereby leading to inadequate negative selection of T-cell populations that could later become activated to elicit an islet-specific destructive autoimmune response. Nepom and Kwok (33
) explained the molecular basis of HLA-DQ associations with type 1 diabetes exactly on this basis. Paradoxically, some self-peptides that normally negatively select T-cells are likely to lead to positive selection when the MHC molecule is, for example, the HLA-DQ3.2. There are many non-MHC genes associated with type 1 diabetes, including polymorphisms influencing thymic insulin expression and T-cell receptor signaling (34
), with essentially all related to immune function.
Environmental factors such as congenital rubella and enteroviruses (particularly Coxsackie B virus) have been related to type 1 diabetes pathogenesis. The presence of a viral infection can lead to immune cell activation through numerous mechanisms. Viruses may directly alter a host cell that may be lysed, releasing self-peptides and fragments of the host cell into the extracellular milieu, whereby they may be processed and presented via APCs. Upon reacting to a viral infection, the immune system may process and present a homologous viral protein in such a manner that the epitope targeted by the immune system can interact with both self-antigens and viral proteins. This process is termed “molecular mimicry.” GAD, a well-defined autoantigen in type 1 diabetes, shares similarities with the P2-C viral sequence of the Coxsackie B virus and the major outer capsid protein of Rotavirus (35
). Viral infections or immunostimulators such as poly I:C, which is used to stimulate viral infections, can trigger islet autoimmunity by activating the innate immune system alone, as demonstrated in the Kilham Rat virus–induced autoimmune diabetes model (37
Recent observations suggest that, in the Aire-deficient mice model, which causes a number of autoimmune diseases including autoimmune diabetes, the stochastic genesis of pathogenic T-cells can initiate autoimmune disease without the need for environmental stimulation, underlining the importance of Aire-dependent thymic deletion rather than an environmental triggering event (38
Overall studies on viral elements in the pathogenesis of type 1 diabetes have been conflicting and have failed to prove conclusively that any of the environmental factors has an undisputable role in the development of type 1 diabetes in genetically and nongenetically susceptible individuals. To date, clear conclusions are limited because most of the studies were not adequately powered to detect differences in exposure and disease associations, had inaccurate exposure estimates, and had confounding exposures.