Antonio Petrone

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 common PPAR-gamma2 Pro12Ala variant is associated with greater insulin sensitivity.

Several genetic variants of peroxisome proliferator-activated receptor-γ2 (PPAR-γ2) have been identified, among which Pro12Ala, a missense mutation in exon 2, is highly prevalent in Caucasian populations. Up to now, conflicting results with regard to the association between this mutation and complex traits, such as obesity, insulin sensitivity and Type 2 diabetes, have been reported. We investigated the influence of the Pro12Ala polymorphism of PPAR-γ2 on insulin sensitivity in a large Italian population sample, n=1215, in whom extensive clinical and biochemical analyses were performed. To estimate the insulin sensitivity status, the homeostasis model assessment of insulin resistance (HOMA-IR) was calculated; in the obese/overweight subjects an oral glucose tolerance test (OGTT) was also performed and the Matsuda insulin sensitivity index (ISI) calculated. The insulin secretion index (homeostasis model assessment of percent β-cell function, HOMA-β%) was utilized to evaluate β-cell function. The effect of the Pro12Ala polymorphism on quantitative variables was tested using multiple linear regression analysis. X12Ala (either Pro12Ala or Ala12Ala) genotype was associated with significantly lower fasting insulin levels compared to Pro/Pro (P=0.01 after correction for multiple comparisons) in all subjects. Consistent with this finding, significantly lower HOMA-IR was observed in X12Ala carriers (P=0.013 after correction for multiple comparisons) in all cohort. Moreover, no significant interaction effect was observed between body mass index and X12Ala polymorphism and between gender and X12Ala polymorphism in modulating insulin sensitivity. Our observations substantially extend previous findings and demonstrated that X12Ala variant is significantly associated with greater insulin sensitivity.

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.

The promoter region of the adiponectin gene is a determinant in modulating insulin sensitivity in childhood obesity

We investigated the association of the −11,391G>A, −11,377G>C, +45T>G, and +276G>T adiponectin single‐nucleotide polymorphisms (SNPs) and expected haplotypes with the insulin resistance (IR) state in overweight/obese children; by using the haplotype background analysis, we also assessed the effect of each SNP independently. GG genotype at the −11,391 locus was associated with higher fasting insulin levels and homeostasis model assessment‐IR index and lower adiponectin levels compared with GA + AA genotypes (p = 0.01, 0.002, and 0.03, respectively). Those heterozygous and homozygous for G allele at the −11,377 locus showed higher fasting glucose (p = 0.001 for both), fasting insulin (p = 0.001 for both), homeostasis model assessment‐IR index (p < 0.001 for both), and triglyceride levels (p = 0.02 and 0.03, respectively) and lower adiponectin levels (p = 0.002 and 0.02, respectively) compared with C homozygotes. The +45G carriers showed higher fasting and 2‐hour glucose levels (p = 0.01 for both) and lower adiponectin levels (p = 0.02) compared with non‐carriers. Haplotype analysis suggested that, considering the same haplotypic background, each of the three polymorphisms exerted an independent effect on investigated parameters. The −11,391G>A, −11,377C>G, and +45T>G SNPs are associated with IR syndrome in overweight/obese children; they independently influence the investigated variables. The effect of +45T>G SNP seems to be marginal compared with the promoter SNPs. The GGT haplotype is associated with the highest degree of IR.

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.

Residual insulin secretion at diagnosis of type 1 diabetes is independently associated with both, age of onset and HLA genotype.

We investigated whether residual insulin secretion and metabolic derangement at diagnosis of type 1 diabetes (T1DM) are influenced by human leukocyte antigens (HLA) class II genes.

Genetic prediction of type 1 diabetes in a population with low frequency of HLA risk genotypes and low incidence of the disease (the DIABFIN study)

To develop a sensitive, specific screening strategy for predicting genetic risk for type 1 diabetes mellitus (T1DM) in the low‐incidence continental Italian population, and to define with this tool, a cohort of high‐to‐moderate risk infants for an immunological follow‐up study aimed at identifying environmental risk factors for T1DM.

A new variation in the promoter region, the −604 C>T, and the Leu72Met polymorphism of the ghrelin gene are associated with protection to insulin resistance

Objective:Previous studies suggested that polymorphisms in the coding region of the preproghrelin were involved in the etiology of obesity and might modulate glucose-induced insulin secretion. We evaluated the association of a new variation, −604C>T, in the promoter region of the ghrelin gene, of Leu72Met (247C>A) and of Gln90Leu (265A>T), all haplotype-tagging single nucleotide polymorphisms (SNPs), with measures of insulin sensitivity in 1420 adult individuals.Research methods:The three SNPs were genotyped using ABI PRISM 7900 HT Sequence Detection System. We used multiple linear regression analysis for quantitative traits and THESIAS software for haplotype analysis.Results:We observed a protective effect exerted by Met72 variant of Leu72Met SNP on insulin resistance parameters; a significant decreasing trend from Leu/Leu to Leu/Met and to Met/Met homozygous subjects in triglycerides, fasting insulin levels and HOMA-IR index (P=0.02, 0.01 and 0.003, respectively), and, consistently, an increase in ghrelin levels (P=0.003) was found. A significant decrease from CC to TC and to TT genotypes in insulin levels and HOMA-IR index was also detected (P=0.00l for both), but only in subjects homozygous for Leu72, where the protective effect of Met72 was not present. The haplotype analysis results supported the data obtained by the evaluation of each single SNP, showing the highest value of insulin levels and HOMA-IR index in the −604c247c haplotype intermediate value in −604T247C and lowest value in −604C247A.Conclusion:Our observations suggest a protective role of the Met72 variant and of −604 T allele in modulating insulin resistance. These SNPs or an unknown functional variant in linkage disequilibrium could increase ghrelin levels and probably insulin sensitivity.

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.