The TUDID Study - Background and Design of a Prospective Cohort.

Prevalence of both type 1 and type 2 diabetes mellitus is growing worldwide and one major cause for morbidity and mortality. However, not every patient develops diabetes-related complications, but causes for the individual susceptibility are still not fully understood. As a platform to address this, we initiated the TUDID (TUebingen DIabetes Database) study, a prospective, monocentric, observational study that includes adults with diabetes mellitus who are treated in the inpatient clinic of a University Hospital in southern Germany. Besides a thorough clinical examination and extensive laboratory tests (with integrated biobanking), major study focuses are the kidneys, the eyes, the vasculature as well as cognition and mood where standardized investigations for early stages for diabetes complications are performed. Analyses of the data generated by this precise characterization of diabetes-related complications will contribute to our understanding of the development and course of such complications, and thus facilitate the implementation of tailored treatment options that can reduce the risk and severity of diabetes-related complications.

Carotid Intima-Media Thickness is a Predictor of Subclinical Myocardial Damage in Men with Type 2 Diabetes Mellitus.

BACKGROUND: Type 2 diabetes mellitus (T2DM) promotes the development of atherosclerosis and is a major risk factor for cardiovascular disease. High-sensitivity cardiac troponin I (hs-cTnI) assays fundamentally improved the diagnosis of myocardial injury and even enable the prediction of future cardiovascular events in the general population. However, data about the association of hs-cTnI with cardiovascular risk factors and carotid intima media thickness (cIMT) as a marker of atherosclerosis are limited, especially in patients with T2DM. METHODS: In this cross-sectional study we analyzed clinical and laboratory parameters of 234 patients (43% women) with T2DM and a median age of 65 years (interquartile range: 57-71). The median duration of diabetes mellitus was 10 years (6-17). Anthropometric data, blood pressure, glycemic parameters and lipid profiles were determined. Hs-cTnI plasma concentrations were measured on an ADVIA Centaur XPT immunoassay analyzer and cIMT was evaluated by high-resolution ultrasound. RESULTS: Hs-cTnI plasma concentrations were below the gender-specific 99th percentile in 93% of T2DM patients with a median concentration of 4.0 ng/l (interquartile range: 2.0-10.0). Hs-cTnI was significantly associated with gender, renal function and C-reactive protein in the entire study cohort. Gender-specific analyses revealed cIMT and renal function to be significantly associated with hs-cTnI in men. Contrary, only age was significantly associated with hs-cTnI in women. CONCLUSION: In a real-world clinical setting in patients with T2DM, cIMT is a predictor of subclinical myocardial damage in men, but not in women.

Reduced insulin clearance is linked to subclinical atherosclerosis in individuals at risk for type 2 diabetes mellitus.

Hyperglycemia and insulin resistance contribute to vascular damage and are regulated by different pathophysiological processes. The aim of the study was to systematically investigate the relative contributions of multiple fasting state- and oral glucose tolerance test (oGTT)-derived glycemic traits to carotid intima-media thickness (cIMT), a surrogate parameter of subclinical atherosclerosis, in individuals with increased risk for type 2 diabetes mellitus (T2D). 667 volunteers (417 women and 250 men, mean age 44.1 years), who were free of cardiovascular disease (CVD), were included in this cross-sectional study. Glucose tolerance, insulin sensitivity, insulin secretion and insulin clearance were assessed by frequently sampled 75 g oGTT. CIMT was measured by high-resolution ultrasound. Insulin clearance was associated with cIMT in univariate analysis (ßst = - 0.17, p < 0.0001) and in a stepwise regression analysis on 15 variables possibly affecting cIMT, age (r2 = 0.3923, p < 0.0001), insulin clearance (r2 = 0.4564, p < 0.0001), systolic blood pressure (r2 = 0.4733, p < 0.0001), body mass index (BMI) (r2 = 0.4804, p = 0.002), gender (r2 = 0.4831, p = 0.013), and fasting insulin clearance (r2 = 0.4857, p = 0.030) turned out to be significant determinants of cIMT. In a cross-validated model resulting from this analysis, insulin clearance was found to be an independent determinant of cIMT (ßst = - 0.16, p < 0.0001) even after adjusting for traditional CVD risk factors. Reduced insulin clearance may be an early marker of damage on the vasculature, independent of classical CVD risk factors. Reduced insulin clearance should be considered with regard to vascular insulin resistance.

Acquired arteriovenous fistula of a finger.

Arteriovenous fistulae are defined as congenital or acquired abnormal direct communications between an artery and a vein leading to abnormal blood circulation. This report describes an unusual manifestation of an acquired peripheral arteriovenous fistula with a high shunt volume of 410 ml/min following a fracture of the 5th finger.

Visceral Adiposity Index as an Independent Marker of Subclinical Atherosclerosis in Individuals Prone to Diabetes Mellitus.

AIM: The visceral adiposity index (VAI) has been proposed as an estimate of visceral adipose tissue (VAT) mass and as an indicator of VAT dysfunction. Both parameters are associated with cardiometabolic risk, including insulin resistance. In this study, we investigated whether VAI is associated with subclinical atherosclerosis in subjects who were free of cardiovascular disease but were at risk of developing diabetes mellitus. METHODS: A total of 731 adults with a median age of 47 years old without diabetes mellitus were included in this cross-sectional study. The anthropometric data, blood pressure, and lipid profiles of 398 women and 333 men were measured. All subjects underwent an oral glucose tolerance test, and carotid intima-media thickness (cIMT) was evaluated by ultrasound. Insulin resistance was estimated using the homeostatic model assessment of insulin resistance (HOMA-IR). RESULTS: VAI and HOMA-IR (ßst=0.44, p＜0.0001), VAI and cIMT (ßst=0.17, p＜0.0001), and HOMA-IR and cIMT (ßst=0.09, p=0.0127) were correlated with each other. After adjusting for cofounding variables, VAI is still correlated with HOMA-IR (ßst=0.42, p＜0.0001). Furthermore, VAI (ßst=0.07, p=0.0392) but not HOMA-IR (ßst=0.03, p=0.37) was correlated with cIMT independently of other established cardiovascular risk factors. CONCLUSION: The calculation of VAI may provide a better estimation of subclinical atherosclerosis than the calculation of HOMA-IR.

Genome-wide analysis of PDX1 target genes in human pancreatic progenitors.

OBJECTIVE: Homozygous loss-of-function mutations in the gene coding for the homeobox transcription factor (TF) PDX1 leads to pancreatic agenesis, whereas heterozygous mutations can cause Maturity-Onset Diabetes of the Young 4 (MODY4). Although the function of Pdx1 is well studied in pre-clinical models during insulin-producing ß-cell development and homeostasis, it remains elusive how this TF controls human pancreas development by regulating a downstream transcriptional program. Also, comparative studies of PDX1 binding patterns in pancreatic progenitors and adult ß-cells have not been conducted so far. Furthermore, many studies reported the association between single nucleotide polymorphisms (SNPs) and T2DM, and it has been shown that islet enhancers are enriched in T2DM-associated SNPs. Whether regions, harboring T2DM-associated SNPs are PDX1 bound and active at the pancreatic progenitor stage has not been reported so far. METHODS: In this study, we have generated a novel induced pluripotent stem cell (iPSC) line that efficiently differentiates into human pancreatic progenitors (PPs). Furthermore, PDX1 and H3K27ac chromatin immunoprecipitation sequencing (ChIP-seq) was used to identify PDX1 transcriptional targets and active enhancer and promoter regions. To address potential differences in the function of PDX1 during development and adulthood, we compared PDX1 binding profiles from PPs and adult islets. Moreover, combining ChIP-seq and GWAS meta-analysis data we identified T2DM-associated SNPs in PDX1 binding sites and active chromatin regions. RESULTS: ChIP-seq for PDX1 revealed a total of 8088 PDX1-bound regions that map to 5664 genes in iPSC-derived PPs. The PDX1 target regions include important pancreatic TFs, such as PDX1 itself, RFX6, HNF1B, and MEIS1, which were activated during the differentiation process as revealed by the active chromatin mark H3K27ac and mRNA expression profiling, suggesting that auto-regulatory feedback regulation maintains PDX1 expression and initiates a pancreatic TF program. Remarkably, we identified several PDX1 target genes that have not been reported in the literature in human so far, including RFX3, required for ciliogenesis and endocrine differentiation in mouse, and the ligand of the Notch receptor DLL1, which is important for endocrine induction and tip-trunk patterning. The comparison of PDX1 profiles from PPs and adult human islets identified sets of stage-specific target genes, associated with early pancreas development and adult ß-cell function, respectively. Furthermore, we found an enrichment of T2DM-associated SNPs in active chromatin regions from iPSC-derived PPs. Two of these SNPs fall into PDX1 occupied sites that are located in the intronic regions of TCF7L2 and HNF1B. Both of these genes are key transcriptional regulators of endocrine induction and mutations in cis-regulatory regions predispose to diabetes. CONCLUSIONS: Our data provide stage-specific target genes of PDX1 during in vitro differentiation of stem cells into pancreatic progenitors that could be useful to identify pathways and molecular targets that predispose for diabetes. In addition, we show that T2DM-associated SNPs are enriched in active chromatin regions at the pancreatic progenitor stage, suggesting that the susceptibility to T2DM might originate from imperfect execution of a ß-cell developmental program.

Periaortic Adipose Tissue Compared With Peribrachial Adipose Tissue Mass as Markers and Possible Modulators of Cardiometabolic Risk.

Increased perivascular fat mass contributes to cardiometabolic risk (CMR). High peribrachial adipose tissue (PBAT) associates with insulin resistance independently of established CMR parameters. It is unknown to what extent periaortic adipose tissue (PAAT) may have a similar impact. In 95 participants, precise quantification of total adipose tissue, PBAT, PAAT, visceral adipose tissue (VAT), and liver fat (LF) content was performed by whole-body magnetic resonance imaging. Insulin sensitivity was determined by oral glucose tolerance test and carotid intima-media thickness (cIMT) by high-resolution ultrasound. In univariate analyses, PAAT correlated with PBAT (ß = .65, P < .0001). A negative correlation of PAAT (ß = -.35, P = .0002) and PBAT (ß = -.43, P < .0001) with insulin sensitivity was observed. While in a stepwise forward regression analysis the relationship of PAAT with insulin sensitivity was no longer significant after adjustment for VAT, LF content, and other CMR factors ( P = 0.42), PBAT still correlated with insulin sensitivity ( r2 = .35, P = .01). The association between PAAT and cIMT (ß = .49, P < .0001) remained significant after adjustment for these variables ( r2 = .42, P = .0001). Although PAAT and PBAT strongly correlate, PAAT is not associated with insulin resistance, but with cIMT. Therefore, PAAT and PBAT may act differently as possible modulators of insulin resistance and subclinical atherosclerosis.

Generation of a human induced pluripotent stem cell (iPSC) line from a patient carrying a P33T mutation in the PDX1 gene.

Homozygous loss-of-function mutations in the gene coding for the homeobox transcription factor PDX1 leads to pancreatic agenesis, whereas certain heterozygous point mutations are associated with Maturity-Onset Diabetes of the Young 4 (MODY4) and Type 2 Diabetes Mellitus (T2DM). To understand the pathomechanism of MODY4 and T2DM, we have generated iPSCs from a woman with a P33T heterozygous mutation in the transactivation domain of PDX1. The resulting PDX1 P33T iPSCs generated by episomal reprogramming are integration-free, have a normal karyotype and are pluripotent in vitro and in vivo. Taken together, this iPSC line will be useful to study diabetes pathomechanisms.

Generation of a human induced pluripotent stem cell (iPSC) line from a patient with family history of diabetes carrying a C18R mutation in the PDX1 gene.

Homozygous loss-of-function mutations in the gene coding for the homeobox transcription factor PDX1 leads to pancreatic agenesis, whereas certain heterozygous point mutations are associated with Maturity-Onset Diabetes of the Young 4 (MODY4) and Type 2 Diabetes Mellitus (T2DM). To understand the pathomechanism of MODY4 and T2DM, we have generated iPSCs from a woman with a C18R heterozygous mutation in the transactivation domain of PDX1. The resulting PDX1 C18R iPSCs generated by episomal reprogramming are integration-free, have a normal karyotype and are pluripotent in vitro and in vivo. Taken together, this iPSC line will be useful to study diabetes pathomechanisms.

Variation in the Phosphoinositide 3-Kinase Gamma Gene Affects Plasma HDL-Cholesterol without Modification of Metabolic or Inflammatory Markers.

OBJECTIVE: Phosphoinositide 3-kinase γ (PI3Kγ) is a G-protein-coupled receptor-activated lipid kinase mainly expressed in leukocytes and cells of the cardiovascular system. PI3Kγ plays an important signaling role in inflammatory processes. Since subclinical inflammation is a hallmark of atherosclerosis, obesity-related insulin resistance, and pancreatic ß-cell failure, we asked whether common genetic variation in the PI3Kγ gene (PIK3CG) contributes to body fat content/distribution, serum adipokine/cytokine concentrations, alterations in plasma lipid profiles, insulin sensitivity, insulin release, and glucose homeostasis. STUDY DESIGN: Using a tagging single nucleotide polymorphism (SNP) approach, we analyzed genotype-phenotype associations in 2,068 German subjects genotyped for 10 PIK3CG SNPs and characterized by oral glucose tolerance tests. In subgroups, data from hyperinsulinaemic-euglycaemic clamps, magnetic resonance spectroscopy of the liver, whole-body magnetic resonance imaging, and intravenous glucose tolerance tests were available, and peripheral blood mononuclear cells (PBMCs) were used for gene expression analysis. RESULTS: After appropriate adjustment, none of the PIK3CG tagging SNPs was significantly associated with body fat content/distribution, adipokine/cytokine concentrations, insulin sensitivity, insulin secretion, or blood glucose concentrations (p>0.0127, all; Bonferroni-corrected α-level: 0.0051). However, six non-linked SNPs displayed at least nominal associations with plasma HDL-cholesterol concentrations, two of them (rs4288294 and rs116697954) reaching the level of study-wide significance (p = 0.0003 and p = 0.0004, respectively). More precisely, rs4288294 and rs116697954 influenced HDL2-, but not HDL3-, cholesterol. With respect to the SNPs' in vivo functionality, rs4288294 was significantly associated with PIK3CG mRNA expression in PBMCs. CONCLUSIONS: We could demonstrate that common genetic variation in the PIK3CG locus, possibly via altered PIK3CG gene expression, determines plasma HDL-cholesterol concentrations. Since HDL2-, but not HDL3-, cholesterol is influenced by PIK3CG variants, PI3Kγ may play a role in HDL clearance rather than in HDL biogenesis. Even though the molecular pathways connecting PI3Kγ and HDL metabolism remain to be further elucidated, this finding could add a novel aspect to the pathophysiological role of PI3Kγ in atherogenesis.

Peroxisome proliferator-activated receptor gamma (PPARG) modulates free fatty acid receptor 1 (FFAR1) dependent insulin secretion in humans.

Genetic variation in FFAR1 modulates insulin secretion dependent on non-esterified fatty acid (NEFA) concentrations. We previously demonstrated lower insulin secretion in minor allele carriers of PPARG Pro12Ala in high-NEFA environment, but the mode of action could not been revealed. We tested if this effect is mediated by FFAR1 in humans. Subjects with increased risk of diabetes who underwent oral glucose tolerance tests were genotyped for 7 tagging SNPs in FFAR1 and PPARG Pro12Ala. The FFAR1 SNPs rs12462800 and rs10422744 demonstrated interactions with PPARG on insulin secretion. FFAR1 rs12462800 (p = 0.0006) and rs10422744 (p = 0.001) were associated with reduced insulin secretion in participants concomitantly carrying the PPARG minor allele and having high fasting FFA. These results suggest that the minor allele of the PPARG SNP exposes its carriers to modulatory effects of FFAR1 on insulin secretion. This subphenotype may define altered responsiveness to FFAR1-agonists, and should be investigated in further studies.