T-cell markers in type I diabetes: progress, prospects and realistic expectations.
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Antibody Formation↗
Biomedical subjects
M Peakman
Publications and source records attributed to M Peakman.
Spontaneous T-cell proliferation in the non-obese diabetic mouse to a peptide from the unique class II MHC molecule, I-Ag7, which is also protective against the development of autoimmune diabetes.
AIMS/HYPOTHESIS: Major histocompatibility complex class II molecules present antigenic peptides to T-cells and have an important role in T-cell thymic education. The mechanism by which major histocompatibility complex alleles confer a high genetic risk for autoimmune diabetes is not known. One hypothesis is that during positive thymic selection, the peripheral T-cell repertoire is modelled by major histocompatibility complex-restricted presentation of self major histocompatibility complex molecule-derived peptides, some of which mimic tissue autoantigens. The sequence similarity between a known T-cell epitope of glutamic acid decarboxylase-65, 509:VPPSLRTLED and the non-obese diabetic mouse class II major histocompatibility complex molecule I-Ag7 86:VPTSLRRLEQ is consistent with this. METHODS: We measured spontaneous proliferation of peripheral T-cells from non-obese diabetic mice and other, non-diabetes-prone strains, to the I-Ag7 86-101 and glutamic acid decarboxylase-65(509-524) peptides, binding of these peptides to intact I-Ag7 and assessed the effect of tolerance induction on diabetes development, by injecting young non-obese diabetic mice with high doses of peptide. RESULTS: T-cells from non-obese diabetic, but not other mice strains, spontaneously proliferate to the I-Ag7 86-101 and glutamic acid decarboxylase-65(509-524) peptides, but not control peptides. Both test peptides bind I-Ag7. Tolerance induction prolongs diabetes-free survival in non-obese diabetic mice when either the I-Ag7 86-101 or glutamic acid decarboxylase-65(509-524) peptide, but not control peptide, is used. CONCLUSION/INTERPRETATION: A peptide from the unique class II major histocompatibility complex, diabetes-susceptibility molecule, I-Ag7, presented by I-Ag7 is a target of T-cell responses in diabetes-prone nonobese diabetic mice and tolerance induction against the peptide offers appreciable protection against the development of diabetes.
Amino Acid Sequence↗
Prevalence of autoantibodies to autonomic nervous tissue structures in Type 1 diabetes mellitus.
AIMS: The pathogenesis of diabetic autonomic neuropathy is multifactorial, but recent studies have suggested a link between the presence of autoantibodies to nervous tissue structures and severe, symptomatic autonomic neuropathy. The present study was designed to examine the true prevalence of these autoantibodies in a large clinic-based population of Type 1 diabetic patients compared to nondiabetic controls. METHODS: The presence of complement fixing autoantibodies to vagus nerve (CF-VN), sympathetic ganglion (CF-SG) and adrenal medulla (CF-ADM) was assessed by immunofluorescence in a large cohort of patients (n = 394) of varying duration of Type 1 DM (median 28 years, range 6 months to 73 years) and 160 age and sex-matched nondiabetic control subjects. RESULTS: All three autoantibodies were frequently detected in Type 1 DM (CF-VN, 22.1%; CF-SG, 30.7%; CF-ADM, 13.2%) but only rarely in healthy control subjects (4.4%, 4.4% and 3.1%, respectively; P < 0.0005 for all). There was no association between any of the autoantibodies and retinopathy (fundoscopy), peripheral somatic neuropathy (biothesiometry) or nephropathy (urinary albumin-creatinine ratio). CONCLUSIONS: Our results on this large cohort establish the extensive presence of autonomic nervous tissue autoantibodies in Type 1 DM. Their role in reflecting, causing or predicting autonomic neuropathy remains to be determined.
Adolescent↗
Neutrophil response to Neisseria meningitidis: inhibition of adhesion molecule expression and phagocytosis by recombinant bactericidal/permeability-increasing protein (rBPI21).
Polymorphonuclear neutrophil (PMNL) activation enhances microbial clearance but also contributes to the vascular damage and multiorgan failure associated with severe meningococcal sepsis. By use of a whole blood model of meningococcal bacteremia, loss of PMNL L-selectin and up-regulation of CD11b was observed in response to Neisseria meningitidis serogroups B and C, which is followed by opsonophagocytosis. PMNL priming with either Escherichia coli lipopolysaccharide (LPS) or FMLP prior to meningococcal challenge resulted in enhancement of both PMNL L-selectin shedding (1.5- to 4-fold) and phagocytosis (2- to 3-fold). Blockade of meningococcal LPS lipid A with recombinant bactericidal/permeability-increasing protein (rBPI21) resulted in partial inhibition of the PMNL activation and phagocytosis response to N. meningitidis. The effect of rBPI21 was reversed by excess E. coli LPS or FMLP. It is proposed that PMNL priming by N. meningitidis results in an exaggerated activation and phagocytosis response to the organism.
Adult↗
Naturally processed and presented epitopes of the islet cell autoantigen IA-2 eluted from HLA-DR4.
During immune responses, antigen-presenting cells (APCs) process antigens and present peptide epitopes complexed with human leukocyte antigen (HLA) molecules. CD4 cells recognize these naturally processed and presented epitopes (NPPEs) bound to HLA class II molecules. Epitope identification is important for developing diagnostic and therapeutic tools for immune-mediated diseases and providing insight into their etiology, but current approaches overlook effects of natural processing on epitope selection. We have developed a technique to identify NPPEs using mass spectrometry (MS) after antigen is targeted onto APCs using a lectin-based antigen delivery system (ADS). We applied the technique to identify NPPEs of the intracellular domain of the type 1 diabetes mellitus-associated (type 1 DM-associated) autoantigen insulinoma-associated-2 (IA-2ic), presented by HLA-DR4 (0401). IA-2ic-derived NPPEs eluted from HLA-DR4 constitute 6 sets of peptides nested around distinct core regions. Synthetic peptides based on these regions bind HLA-DR4 and elicit primary T-cell proliferation frequently in HLA-DR4-positive type 1 DM patients, but rarely in non-HLA-DR4 patients, and in none of the HLA-DR4 nondiabetic controls we tested. This flexible, direct approach identifies an HLA allele-specific map of NPPEs for any antigen, presented by any HLA class II molecule. This method should enable a greater understanding of epitope selection and lead to the generation of sensitive and specific reagents for detecting autoreactive T cells.
Adolescent↗
Human leukocyte antigen-DQB1* genotypes encoding aspartate at position 57 are associated with 3beta-hydroxysteroid dehydrogenase autoimmunity in premature ovarian failure.
Premature ovarian failure (POF) has an autoimmune pathogenesis in a significant proportion of cases. Autoantibodies to the steroid cell enzyme, 3beta-hydroxysteroid dehydrogenase (3betaHSD) are present in one fifth of patients and may identify an autoimmune subgroup. As autoimmune diseases are associated with alleles of the human leukocyte antigen (HLA) genes, we examined the distribution of HLA-DRB1 and -DQB1 genotypes in 118 women with POF, of whom 21% had 3betaHSD autoantibodies, and 134 racially matched control subjects. Two HLA-DQB1 alleles, 0301 and 0603, were associated with 3betaHSD autoantibody positivity (P = 0.04 and P = 0.006, respectively). As the DQB1*0301 and -0603 genes share an identical codon at position 57 (aspartate, Asp), we analyzed the frequency of DQbeta-Asp57 encoding DQB1 genes in our series. Eighteen of 21 POF patients with 3betaHSD autoantibodies had DQbeta-Asp57-encoding genotypes (haplotype frequency, 27 of 42; 64%) compared with 92 of 134 control subjects (haplotype frequency, 109 of 268; 41%; P = 0.004), and 9 of 21 (43%) cases were homozygous for codon 57 genotypes compared with 17 of 134 (13%) control subjects (P = 0.0006). These probability values were not significant after correction for multiple testing, and these trends will therefore require confirmation in larger cohorts. HLA class II molecules present antigenic peptides to CD4+ T lymphocytes. DQbeta57 occupies a key site at the boundary of the peptide binding groove, with a major impact on peptide binding. Our preliminary demonstration of an association between POF, 3betaHSD autoimmunity, and a distinctive HLA-DQ molecule supports the hypothesis that autoantibodies to this steroid cell enzyme may be markers of autoimmune ovarian failure and suggests that presentation of autoantigenic or external peptides to T lymphocytes by HLA-DQ molecules with Asp57-beta-chains is important in the pathogenesis of this disease.
3-Hydroxysteroid Dehydrogenases↗
Autonomic function and autoantibodies to autonomic nervous structures, glutamic acid decarboxylase and islet tyrosine phosphatase in adolescent patients with IDDM.
Recent studies have linked autoimmunity to nervous tissue structures and diabetic autonomic neuropathy, but data on the early stage of IDDM and on the natural history of this association are not available. For this reason, we investigated autonomic nervous function, and the presence of autoantibodies to sympathetic and parasympathetic nervous structures, to glutamic acid decarboxylase (GAD) and tyrosine phosphatase (IA-2/ICA512) in 85 adolescents with insulin-dependent diabetes mellitus (IDDM) (mean age 14.7+/-1.6 yr, mean duration of diabetes 6.8+/-3.5 yr), and 45 age and sex-matched healthy subjects. Nervous tissues autoantibodies were detected using an indirect immunofluorescent complement-fixation technique, with monkey adrenal gland, rabbit cervical ganglia and vagus nerve as substrates. GAD and IA-2/ICA512 autoantibodies were detected by radioimmunoprecipitation assay. Seven patients (8%) had anti-vagus nerve autoantibodies, 7 other patients (8%) had anti-cervical ganglia autoantibodies, while all controls were negative (P < 0.05). Anti-adrenal medulla antibodies were detected in 16 patients (19%) and in 2 control subjects (P<0.02). None of the patients had autonomic symptoms. When patients were divided according to the presence or absence of autoantibodies, values of the cardiovascular tests (deep breathing, 30:15 ratio, Valsalva ratio) were similar in the two groups and similar to those in healthy subjects. However, when considered together, patients positive for one or more autoantibody showed a trend for lower values of deep breathing test and 30:15 ratio test, compared with healthy control subjects, which failed to reach conventional significance values (P=0.17 and P=0.07, respectively). No correlation was found between cardiovascular parameters and metabolic control or diabetes duration. There was no association between autoimmunity to nervous tissue structures and presence of GAD and IA-2/ICA512 Ab, and no correlation between these two autoantibodies and values of cardiovascular tests. Our data indicate that autonomic dysfunction is not a characteristic of young diabetic patients, but that autoantibodies against autonomic nervous structures are present during the first 1 to 15 yr of diabetes. GAD and tyrosine phosphatase appear to be excluded as target autoantigens within autonomic structures. Follow-up studies are required to evaluate future autonomic dysfunction and symptoms in these patients, and to establish whether the subtle autonomic dysfunction detected and/or the nervous tissue autoantibodies, are predictive of the development of this complication.
Adolescent↗
Enhanced T cell proliferation and increased responder frequency following delivery of antigen to the antigen-presenting cell; B cell dependency and use in detection of autoreactive T cells.
Reported frequencies of peripheral blood autoantigen-specific cells in autoimmune diseases are typically low, which could be due to true scarcity or to limitations of in vitro assays. In the present study, antigens were targeted to the antigen-presenting cell (APC) to enhance T cell proliferation, using an antigen delivery system (ADS), consisting of biotinylated anti-IgG, streptavidin and biotinylated antigen. This was able to bind B cells and monocytes and was internalized within 24 hours. T cell proliferation to tetanus toxoid was at least doubled using the ADS compared to conventional assay with antigen in simple solution. To evaluate the ADS in an autoimmune disease, we determined T cell responses to the insulin-dependent diabetes mellitus (IDDM)-associated autoantigen IA-2ic in patients with recent-onset IDDM. When IA-2ic was available conventionally in solution, proliferation was poor, but significantly higher in IDDM patients than control subjects. However, the ADS significantly enhanced proliferation by a mean 3-fold for all subjects, while maintaining the significant difference between IDDM patients and healthy controls. Increases in T cell proliferation via the ADS were due to the recruitment of approximately 3 times the number of CD4 + T cells stimulated in conventional assays. B cell depletion abolished enhancement suggesting that the ADS operates through recruitment of B cells as APCs. This flexible modification of the T cell assay offers greatly enhanced sensitivity for determining the frequency of antigen and autoantigen-reactive T cells.
Adult↗
Recent advances. Immunology.
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Allergy and Immunology↗
Autoimmunity in diabetic autonomic neuropathy: does the immune system get on your nerves?
Symptomatic autonomic neuropathy is a devastating occasional complication of diabetes mellitus, especially Type 1. Although the full-blown clinical syndrome is not common, dysfunction of the autonomic nerves is detectable in up to 40% of Type 1 diabetic patients but its aetiopathogenesis is poorly understood. There is evidence to suggest that the damage to the autonomic nerves may be immune-mediated. This evidence is reviewed in the following article.
Autoimmunity↗
Combined analysis of GAD65 and ICA512(IA-2) autoantibodies in organ and non-organ-specific autoimmune diseases confers high specificity for insulin-dependent diabetes mellitus.
There is evidence that insulin-dependent diabetes mellitus (IDDM) may develop in association with other non-beta-cell-specific autoimmune diseases. We aimed to assess whether autoantibodies to the islet cell antigens glutamic acid decarboxylase (Mr 65,000 isoform) (GAD65) and ICA512(IA-2), present alone or in combination, are limited to IDDM or also occur in other organ- or non-organ-specific autoimmune disorders. We determined the frequency of these autoantibodies by radioimmunoassay in 199 sera from patients with autoimmune thyroid diseases (AITD), rheumatoid arthritis (RA), systemic lupus erythematosus (SLE) and primary biliary cirrhosis (PBC), and compared the results with those from 507 newly diagnosed patients with IDDM and 280 healthy controls. ICA512(IA-2) autoantibodies were detected exclusively in AITD with concurrent IDDM, but not in other autoimmune diseases without IDDM, whereas GAD65 autoantibodies exceeded the limit of normal in 67.7% (21 of 31) of patients with AITD who also had IDDM and in 5.5% (three of 55) of patients with PBC. The frequency of either GAD65 and/or ICA512(IA-2) autoantibodies was significantly higher in patients with AITD who also had IDDM (27 of 31, 87.1%) than in those with AITD alone (one of 53, 1.9%; P<10(-6)), but was not significantly different from those patients with newly diagnosed IDDM (418 of 507, 82.4%). Neither patients with organ- or non-organ-specific autoimmune diseases without IDDM nor healthy controls had autoantibodies against both GAD65 and ICA512(IA-2). Despite the fact that one of the two autoantibodies was occasionally detected in patients with non-beta-cell-specific autoimmune diseases without IDDM, combined determination of GAD65 and ICA512(IA-2) autoantibodies specifically identified IDDM in the majority of patients with AITD. In conclusion, because of the strong association of IDDM with AITD, testing for multiple islet autoanti-bodies could be useful as a predictive marker for risk of progression to IDDM onset amongst patients with autoimmune thyroid disorders.
Adolescent↗
Cytokine overproduction in healthy first degree relatives of patients with IDDM.
Healthy family members of patients with insulin-dependent diabetes mellitus (IDDM) are known to share a number of immunological abnormalities with their affected relatives. Since monocyte and type 1 T-cell-derived cytokines contribute to the pathogenesis of IDDM, we studied the production of these cytokines in the healthy first degree relatives of 29 children with IDDM. We report that circulating tumour necrosis factor-alpha (TNF-alpha) and soluble interleukin-2 (sIL-2) receptor were present in increased amounts in non-diabetic family members at levels similar to those found in the diabetic children (duration of disease 3 months-5 years). Furthermore, marked hypersecretion of IL-1alpha and TNF-alpha by mitogen-stimulated peripheral blood mononuclear cells was found in both diabetic and healthy family members. Abnormalities of cytokine production in healthy relatives did not correlate with the presence of islet cell antibodies or with HLA DR type. These data indicate that healthy family members of patients with IDDM exhibit overproduction of a number of cytokines that have been implicated in diabetogenesis.
Adolescent↗
Irradiated NC adenocarcinoma cells transduced with both B7.1 and interleukin-2 induce CD4+-mediated rejection of established tumors.
Previous studies have shown that expression of the immune co-stimulator B7.1 reduces the tumorigenicity of some, but not all, malignant cell lines. However, B7.1-expressing tumor cells are not very effective in inducing the rejection of established tumors. This may in part be due to induction of anergy in the potentially reactive T cells. Previous studies have shown that IL-2 can reverse the anergic state both in vitro and in vivo. Therefore, we have examined the effect of retrovirus-mediated delivery and expression of murine B7.1 and interleukin-2 on tumor formation and rejection of established MHC class I+/II- NC adenocarcinomas. Neither the expression of B7.1 nor IL-2 alone had a significant effect on NC tumorigenicity. In contrast, combined expression of B7.1 and IL-2 substantially decreased the tumorigenicity of these cells in the immunecompetent syngeneic hosts. T-cell depletion studies show this to be dependent primarily on the activation of CD4+ cells. Furthermore, distant subcutaneous injection of irradiated NC/IL-2/B7.1 can induce, much more effectively than NC/B7.1 or NC/IL-2, the rejection of small NC tumors, and prevent the recurrence of large surgically resected tumors. Together, these results suggest that tumor cells genetically modified to express B7.1 and IL-2 can induce the immune-mediated rejection of established class II- tumors by a mechanism involving CD4+ cells.
Adenocarcinoma↗
Reproducibility and persistence of neural and adrenal autoantibodies in diabetic autonomic neuropathy.
The presence of autoantibodies to autonomic nervous tissue structures is a feature of patients with symptomatic diabetic autonomic neuropathy. It has not been established whether these autoantibodies cause, contribute to or simply reflect nervous tissue damage. Serum samples were tested for the presence of complement-fixing autoantibodies to adrenal medulla, vagus nerve, and sympathetic ganglion cells, to demonstrate: (a) reproducibility of the technique, (b) persistence of the antibodies, and (c) whether or not they occur in patients with non-insulin-dependent (Type 2) diabetes (NIDDM) with neuropathy. Examination of 37 samples, by different observers 2 years apart, revealed a high degree of concordance of both positive and negative results, demonstrating the method of testing to be highly reproducible. Re-testing of 37 patients (by analysing a second blood sample) between 0.5 and 2.7 years (mean 1.7 years) after their first test also demonstrated that antibodies, once present, normally persist; and that most patients initially negative remained so. Of 17 neuropathic NIDDM patients, 16 were negative for all three antibodies, indicating their rarity in this group of patients.
Adrenal Medulla↗
Clinical improvement in chronic fatigue syndrome is not associated with lymphocyte subsets of function or activation.
The relationship between markers of immune function and chronic fatigue syndrome (CFS) is controversial. To examine the relationship directly, 43 subjects with CFS entering a randomized controlled trial of a nonpharmacological treatment for CFS gave samples for immunological analysis before and after treatment. Percentage levels of total CD3+ T cells, CD4 T cells, CD8 T cells, and activated subsets did not differ between CFS subjects and controls. Naive (CD45RA+ RO-) and memory (CD45RA- RO+) T cells did not differ between subjects and controls. Natural killer cells (CD16+/CD56+/CD3-) were significantly increased in CFS patients compared to controls, as was the percentage of CD11b+ CD8 cells. There were no correlations between any immune variable and measures of clinical status, with the exception of a weak correlation between total CD4 T cells and fatigue. There was a positive correlation between memory CD4 and CD8 T cells and depression scores and a negative correlation between naive CD4 T cells and depression. No immune measures changed during the course of the study, and there was no link between clinical improvement as a result of the treatment program and immune status. Immune measures did not predict response or lack of response to treatment. In conclusion, we have been unable to replicate previous findings of immune activation in CFS and unable to find any important associations between clinical status, treatment response, and immunological status.
Adult↗
T lymphocyte subpopulations in anorexia nervosa and refeeding.
Several studies have addressed the question of the effects of starvation on immune function and changes in lymphocyte subsets. Patients with anorexia nervosa are severely malnourished, but there have been few studies of immune parameters in this group. For this reason, phenotypic markers of T cell function and activation were studied in 20 severely underweight patients with anorexia nervosa and again after a period of refeeding. The most significant finding was a reduction in the percentage and absolute number of CD8+ T cells in patients with anorexia, the result of a marked reduction in memory (CD45RO+RA-) CD8 cells. A tendency for recovery in numbers of this subset was seen after refeeding. A decreased memory:maive cell ratio was also seen among CD4 cells, but was less marked. Subtle abnormalities in activated CD4 and CD8 cells were also found in the patient group at the initial sampling, but did not follow any clear pattern. These findings indicate that starvation in anorexic patients is accompanied by a large change in memory CD8 T cells. It may be speculated that this relates to the perceived lack of symptomatic common viral infections in underweight anorexic patients and their return with the recovery of weight.
Adolescent↗
Lymphocyte vaccination protects prediabetic non-obese diabetic mice from developing diabetes mellitus.
In the therapeutic manoeuvre termed "lymphocyte vaccination", activated lymphocytes capable of transferring an autoimmune disease are instead attenuated and given in vaccine form. We have previously shown that such a therapy administered to non-obese diabetic (NOD) mice at 6 weeks of age prevents diabetes mellitus. To assess whether this therapy has potential clinical relevance, in the present study lymphocyte vaccination was applied in NOD mice in 3 weekly doses commencing in the immediate prediabetic period (age 12 weeks), when insulitis is advanced and diabetes incipient. Of 30 NOD mice receiving active vaccine (composed of attenuated lymphocytes from diabetic NOD mice) 13 (43.3%) remained non-diabetic to the age of 30 weeks, in comparison with 2 of 30 (6.7%; p < 0.01) mice receiving a control vaccine (composed of attenuated lymphocytes from non-diabetic NOD/B10 mice) and 5 of 26 (19.2%; p < 0.01) mice receiving saline carrier alone. Moreover, in an additional group of 10 NOD mice receiving active vaccine weekly between 12 and 30 weeks, 8 remained diabetes free at the end of the treatment. The most notable effect of the vaccine was that the delay in diabetes onset was accompanied by a reduction in insulitis and in some cases a complete absence of infiltrating lymphocytes at 30 weeks of age. Immunocytochemistry indicated that when present, islet infiltrating lymphocytes in non-diabetic mice that received active vaccine showed significantly reduced staining for interferon-gamma, compared with the infiltrate seen in diabetic mice receiving the control vaccine or saline. This study demonstrates that the rapid progression to diabetes typically seen in 12-week-old NOD mice can be delayed by lymphocyte vaccination, supporting the possibility that a vaccine composed of attenuated autologous peripheral blood lymphocytes could be effective in high risk first degree relatives of patients with insulin dependent diabetes mellitus.
Animals↗
Expression of glutamic acid decarboxylase in nervous tissue structures targeted by autoantibodies in patients with diabetic autonomic neuropathy.
We have previously identified an association between symptomatic diabetic autonomic neuropathy (DAN) and autoantibodies to sympathetic and parasympathetic nervous structures. The antigens identified by these autoantibodies are not known, but glutamic acid decarboxylase (GAD) has been suggested as a candidate target, since anti-GAD autoantibodies are present in patients with long-term diabetes and GAD is expressed in a variety of cell types and structures in the nervous system. The aim of this study was to examine GAD expression in sympathetic ganglia and vagus nerve and to compare the distribution of GAD within these tissues with that of anti-sympathetic ganglia and anti-vagus nerve autoantibodies from patients with DAN, using single and double indirect immunofluorescence on tissue sections. The monoclonal antibody GAD-6, specific for GAD65, gave a granular, peripheral, cytoplasmic staining pattern in sympathetic ganglion cells. Dual immunofluorescence demonstrated that serum from a patient with anti-sympathetic ganglion autoantibodies stained the same cells, but homogeneously throughout the cytoplasm. In the vagus nerve, patient's serum stained the fibres only; GAD-6 stained the cytoplasm of parasympathetic ganglion cells but only occasional fibres. In addition, GAD enzymatic activity was detectable in both sympathetic ganglia and vagus nerve. Incubation of sera or GAD-6 overnight with a crude homogenate of human brain as an antigen source abolished staining of the nervous tissues by GAD-6, but not by patients' sera. The different localisation of GAD and the autoantigens targeted by patients' sera indicates that GAD is not the target of the autoantibodies characteristic of DAN. Moreover, absorption studies using human brain homogenate suggest that the targets of anti-sympathetic ganglion and anti-vagus nerve autoantibodies are absent or represented only at low levels in the central nervous system and may be confined to the periphery.
Autoantibodies↗