Marco Capizzi

Zinc Transporter 8 Antibodies Complement GAD and IA-2 Antibodies in the Identification and Characterization of Adult-Onset Autoimmune Diabetes: Non Insulin Requiring Autoimmune Diabetes (NIRAD) 4

OBJECTIVE Zinc transporter 8 (ZnT8) is an islet β-cell secretory granule membrane protein recently identified as an autoantibody antigen in type 1 diabetes. The aim of this study was to determine the prevalence and role of antibodies to ZnT8 (ZnT8As) in adult-onset diabetes. RESEARCH DESIGN AND METHODS ZnT8As were measured by a radioimmunoprecipitation assay using recombinant ZnT8 COOH-terminal or NH2-terminal proteins in 193 patients with adult-onset autoimmune diabetes having antibodies to either GAD (GADAs) or IA-2 (IA-2As) and in 1,056 antibody-negative patients with type 2 diabetes from the Non Insulin Requiring Autoimmune Diabetes (NIRAD) study. RESULTS ZnT8As-COOH were detected in 18.6% patients with autoimmune diabetes and 1.4% with type 2 diabetes. ZnT8As-NH2 were rare. ZnT8As were associated with younger age and a high GADA titer. The use of GADAs, IA-2As, and ZnT8As in combination allowed a stratification of clinical phenotype, with younger age of onset of diabetes and characteristics of more severe insulin deficiency (higher fasting glucose and A1C, lower BMI, total cholesterol, and triglycerides) in patients with all three markers, with progressive attenuation in patients with two, one, and no antibodies (all Ptrend < 0.001). Autoantibody titers, association with high-risk HLA genotypes, and prevalence of thyroid peroxidase antibodies followed the same trend (all P < 0.001). CONCLUSIONS ZnT8As are detectable in a proportion of patients with adult-onset autoimmune diabetes and seem to be a valuable marker to differentiate clinical phenotypes.

The Protein Tyrosine Phosphatase Nonreceptor 22 (PTPN22) Is Associated With High GAD Antibody Titer in Latent Autoimmune Diabetes in Adults Non Insulin Requiring Autoimmune Diabetes (NIRAD) Study 3

OBJECTIVE—We previously demonstrated the presence of two different populations among individuals with adult-onset autoimmune diabetes: those having either a high titer or a low titer of antibodies to GAD (GADAs). Protein tyrosine phosphatase nonreceptor type 22 (PTPN22) has been identified as a new susceptibility gene for type 1 diabetes and other autoimmune diseases. The aim of the present study was to evaluate whether the phenotypic heterogeneity of adult-onset autoimmune diabetes based on the GADA titer is associated with the PTPN22 C1858T polymorphism. RESEARCH DESIGN AND METHODS—Analysis for the C1858T polymorphism using the TaqMan assay was performed in 250 subjects with adult-onset autoimmune diabetes, divided into two subgroups with low (≤32 arbitrary units) or high (>32 arbitrary units) GADA titers and 450 subjects with classic type 2 diabetes (from the Non Insulin Requiring Autoimmune Diabetes [NIRAD] Study cohort of 5,330 subjects with adult-onset diabetes) and in 558 subjects with juvenile-onset type 1 diabetes and 545 normoglycemic subjects. RESULTS—Genotype, allele, and phenotype distributions of the PTPN22 C1858T variant revealed similar frequencies in autoimmune diabetes with high GADA titer and juvenile-onset type 1 diabetes. An increase in TT and CT genotypes was observed in individuals with a high GADA titer compared with a low GADA titer, those with type 2 diabetes, and control subjects (P < 0.002 for all comparisons). The PTPN22 1858T allele and phenotype frequencies were increased in high GADA titer compared with a low GADA titer, type 2 diabetic, and control subjects (P < 0.001 for all comparisons, odds ratio 2.6). CONCLUSIONS—In adult-onset autoimmune diabetes, the PTPN22 1858T variant is associated only with a high GADA titer, providing evidence of a genetic background to clinical heterogeneity identified by GADA titer.

Wrist Circumference Is a Clinical Marker of Insulin Resistance in Overweight and Obese Children and Adolescents

Background— Excess fat is one of the main determinants of insulin resistance, representing the metabolic basis for developing future cardiovascular disease. The aim of the current study was to find an easy-to-detect clinical marker of insulin resistance which can be used to identify young subjects at increased risk of cardiovascular disease. Methods and Results— Four-hundred and seventy-seven overweight/obese children and adolescents (mean age 10.31±2.80 years) were consecutively enrolled. Standard deviation score body mass index, fasting biochemical parameters, and homeostasis model assessment of insulin resistance were evaluated. Statistical differences were investigated using multiple linear regression analysis. Manual measure of wrist circumference was evaluated in all children and adolescents. Fifty-one subjects, randomly selected, underwent nuclear magnetic resonance imaging of the wrist to evaluate transversal wrist area at the Lister tubercle level. A statistically significant association was found between manual measure of wrist circumference and insulin levels or homeostasis model assessment of insulin resistance (&bgr;=0.34 and 0.35, respectively; P<10−5 for both comparisons). These associations were more significant than those between SD score body mass index and insulin levels or homeostasis model assessment of insulin resistance (&bgr;=0.12 and 0.10, respectively; P⩽0.02 for both comparisons). Nuclear magnetic resonance imaging acquisition clarified that the association between wrist circumference and insulin levels or homeostasis model assessment of insulin resistance reflected the association with bone tissue-related areas (P⩽0.01 for both) but not with the adipose tissue ones (P>0.05), explaining 20% and 17% of the variances of the 2 parameters. Conclusions— Our findings suggest a close relationship among wrist circumference, its bone component, and insulin resistance in overweight/obese children and adolescents, opening new perspectives in the prediction of cardiovascular disease.

PPAR-γ2 Pro12Ala Variant Is Associated with Greater Insulin Sensitivity in Childhood Obesity

Several genetic variants of peroxisome proliferator-activated receptor-γ2 (PPAR-γ2), a molecule known to be involved in transcription of target genes, have been identified. Pro12Ala, a missense mutation in exon 2 of the gene, is highly prevalent in Caucasian populations. Conflicting conclusions about the association between this mutation and complex traits such as obesity, insulin sensitivity, and T2DM have been reported. We have investigated the association of PPAR-γ2 Pro12Ala polymorphism with measures of insulin sensitivity in a population of Italian obese children (n = 200; mean age, 10.38 ± 2.8 y) in whom clinical and biochemical analyses were performed. To estimate the insulin sensitivity status, the homeostasis model assessment of insulin resistance (HOMA-IR) was calculated in all subjects. The effect of the Pro12Ala polymorphism on quantitative variables was tested using multiple linear regression analysis. The frequency of Ala carriers was 17%, similar to that reported in other adult Caucasian populations. The X12Ala (either Pro12Ala or Ala12Ala) genotype was associated with significantly lower fasting insulin levels compared with Pro/Pro (p = 0.008). Consistent with this finding, significantly lower HOMA-IR was observed in X12Ala carriers (p = 0.023). In conclusion, our observations demonstrate that the X12Ala variant is significantly associated with greater insulin sensitivity in childhood obesity. Because obesity is one of the most important risk factors for cardiovascular diseases and type 2 diabetes, obese children, who are presumably at a higher risk, may be protected from these diseases by the phenotypic effect of the Ala 12 allele on insulin resistance.

The Protein Tyrosine Phosphatase Non Receptor 22 (PTPN22) is associated with high GAD antibody titre in latent autoimmune diabetes in adults (LADA) (NIRAD study 3)

OBJECTIVE—We previously demonstrated the presence of two different populations among individuals with adult-onset autoimmune diabetes: those having either a high titer or a low titer of antibodies to GAD (GADAs). Protein tyrosine phosphatase nonreceptor type 22 (PTPN22) has been identified as a new susceptibility gene for type 1 diabetes and other autoimmune diseases. The aim of the present study was to evaluate whether the phenotypic heterogeneity of adult-onset autoimmune diabetes based on the GADA titer is associated with the PTPN22 C1858T polymorphism. RESEARCH DESIGN AND METHODS—Analysis for the C1858T polymorphism using the TaqMan assay was performed in 250 subjects with adult-onset autoimmune diabetes, divided into two subgroups with low (≤32 arbitrary units) or high (>32 arbitrary units) GADA titers and 450 subjects with classic type 2 diabetes (from the Non Insulin Requiring Autoimmune Diabetes [NIRAD] Study cohort of 5,330 subjects with adult-onset diabetes) and in 558 subjects with juvenile-onset type 1 diabetes and 545 normoglycemic subjects. RESULTS—Genotype, allele, and phenotype distributions of the PTPN22 C1858T variant revealed similar frequencies in autoimmune diabetes with high GADA titer and juvenile-onset type 1 diabetes. An increase in TT and CT genotypes was observed in individuals with a high GADA titer compared with a low GADA titer, those with type 2 diabetes, and control subjects (P < 0.002 for all comparisons). The PTPN22 1858T allele and phenotype frequencies were increased in high GADA titer compared with a low GADA titer, type 2 diabetic, and control subjects (P < 0.001 for all comparisons, odds ratio 2.6). CONCLUSIONS—In adult-onset autoimmune diabetes, the PTPN22 1858T variant is associated only with a high GADA titer, providing evidence of a genetic background to clinical heterogeneity identified by GADA titer.

GADA Titer-Related Risk for Organ-Specific Autoimmunity in LADA Subjects Subdivided according to Gender (NIRAD Study 6)

CONTEXT Latent autoimmune diabetes in adults (LADA) includes a heterogeneous population wherein, based on glutamic acid decarboxylase antibody (GADA) titer, different subgroups of subjects can be identified. OBJECTIVE The aim of the present study was to evaluate GADA titer-related risk for β-cell and other organ-specific autoimmunity in LADA subjects. METHODS Adult-onset autoimmune diabetes subjects (n=236) and type 2 diabetes (T2DM) subjects (n=450) were characterized for protein tyrosine phosphatase (IA-2IC and IA-2(256-760)), zinc transporter 8 (ZnT8), thyroid peroxidase, (TPO), steroid 21-hydroxylase (21-OH), tissue transglutaminase (tTG), and antiparietal cell (APC) antibodies. RESULTS High GADA titer compared to low GADA titer showed a significantly higher prevalence of IA-2IC, IA-2(256-760), ZnT8, TPO, and APC antibodies (P≤0.04 for all comparison). 21-OH antibodies were detected in 3.4% of high GADA titer. A significant decreasing trend was observed from high GADA to low GADA and to T2DM subjects for IA-2(256-760), ZnT8, TPO, tTG, and APC antibodies (P for trend≤0.001). TPO was the only antibody showing a different prevalence between gender; low GADA titer and T2DM female patients had a higher frequency of TPO antibody compared to males (P=0.0004 and P=0.0006, respectively), where the presence of high GADA titer conferred an odds ratio of 8.6 for TPO compared to low GADA titer. After subdividing high and low GADA titer subjects according to the number of antibodies, we observed that 73.3% of high GADA titer subjects were positive for at least one or more antibodies, compared to 38.3% of low GADA titer (P<0.0001). CONCLUSIONS In LADA subjects, high GADA titer was associated with a profile of more severe autoimmunity and, in male gender, specifically predisposed to thyroid autoimmunity. A regular screening for other antibodies is recommended in LADA patients according to GADA titer and gender.

The PTPN22 1858T Gene Variant in Type 1 Diabetes Is Associated With Reduced Residual β-Cell Function and Worse Metabolic Control

OBJECTIVE Evidence has been reported for a new susceptible locus for type 1 diabetes, the protein tyrosine phosphatase nonreceptor type 2 (PTPN22), which encodes a lymphoid-specific phosphatase. The aim of the study was to evaluate the influence of the C1858T variant of the PTPN22 gene on beta-cell function as measured by C-peptide levels from time of disease diagnosis through 12 months follow-up in a prospective series of 120 consecutive type 1 diabetic subjects. RESEARCH DESIGN AND METHODS The C1858T polymorphism was genotyped using TaqMan. Fasting C-peptide, A1C, and insulin requirements were determined at diagnosis and every 3 months for 12 months; their change during follow-up was analyzed using the general linear model repeated-measures procedure. RESULTS Fasting C-peptide levels were significantly lower and A1C levels were significantly higher in subjects carrying the PTPN22 1858T variant than in subjects homozygous for C1858 from time of disease diagnosis through 12 months of intensive insulin therapy follow-up (P = 0.008 and P = 0.01, respectively). These findings were independent of age at onset, sex, and HLA risk groups. The trend in C-peptide and A1C levels in the 12-month period did not differ significantly between subjects with or without the 1858T variant. Insulin dose was similar in the 1858T carriers and noncarriers. CONCLUSIONS Type 1 diabetic subjects carrying the 1858T variant show significantly lower beta-cell function and worse metabolic control at diagnosis and throughout the study period than subjects homozygous for C1858; these differences remain unchanged over the course of the first year after diagnosis.

Association of TCF7L2 gene variants with low GAD autoantibody titre in LADA subjects (NIRAD Study 5).

Diabet. Med. 27, 701–704 (2010)

Low-risk HLA genotype in Type 1 diabetes is associated with less destruction of pancreatic B-cells 12 months after diagnosis.

Aims  The role of human leukocyte antigen (HLA) genes in the susceptibility to Type 1 diabetes (T1DM) is well known. However, we do not know whether the degree of pancreatic B‐cell destruction depends on different HLA genetic risk. The aim of this study was to analyse the influence of DRB1* and DQB1* genes on the rate of pancreatic B‐cell loss in a prospective series of 120 consecutive newly diagnosed T1DM subjects in the first 12 months after diagnosis.

Temporal Trends of HLA, CTLA-4 and PTPN22 Genotype Frequencies among Type 1 Diabetes in Continental Italy

The incidence of type 1 diabetes has, progressively, increased worldwide over the last decades and also in Continental Italian population. Previous studies performed in northern European countries, showed, alongside a general increase in the disease incidence, a decreasing frequency of the highest risk HLA genotype in type 1 diabetes populations, thus emphasizing the role of environmental factors. The aim of the study was to evaluate whether a decreasing trend of high risk HLA, CTLA-4 and PTPN22 genotypes would be present in type 1 diabetes subjects of Continental Italy, a country considered at low incidence of the disease compared to northern European populations. N = 765 type 1 diabetes patients diagnosed from 1980 to 2012 in Lazio region were included. For HLA, CTLA4 and PTPN22 temporal trend evaluation, subjects were subdivided into groups of years according to age at diagnosis. All subjects were typed for HLA-DRB1 and DQB1 by a reverse line blot. The CT60 polymorphism of the CTLA4 and C1858T of the PTPN22 gene were genotyped using ABI PRISM 7900HT (n = 419 and n = 364 respectively). HLA genotypes were divided in high, moderate and low risk categories. The proportion of the HLA risk categories was not statistically different over the three decades in subjects with age of onset <15 years and ≥15 years. The genotype distribution of CT60 polymorphism of CTLA4 gene did not show any change in the frequencies during time. The analysis of the PTPN22 C1858T variant revealed, instead, that the frequency of CT+TT susceptibility genotypes decreased during time (23.9% vs 13.6%, p = 0.017). We can hypothesize that the pressure of the diabetogenic environment could be milder and therefore not sufficient to reduce the need of a strong genetic background (HLA) “to precipitate” diabetes; the increased pressure of the environment could have, instead, some effects on minor susceptibility genes in our population.