Jürgen Graessler

Detailed Physiologic Characterization Reveals Diverse Mechanisms for Novel Genetic Loci Regulating Glucose and Insulin Metabolism in Humans

OBJECTIVE Recent genome-wide association studies have revealed loci associated with glucose and insulin-related traits. We aimed to characterize 19 such loci using detailed measures of insulin processing, secretion, and sensitivity to help elucidate their role in regulation of glucose control, insulin secretion and/or action. RESEARCH DESIGN AND METHODS We investigated associations of loci identified by the Meta-Analyses of Glucose and Insulin-related traits Consortium (MAGIC) with circulating proinsulin, measures of insulin secretion and sensitivity from oral glucose tolerance tests (OGTTs), euglycemic clamps, insulin suppression tests, or frequently sampled intravenous glucose tolerance tests in nondiabetic humans (n = 29,084). RESULTS The glucose-raising allele in MADD was associated with abnormal insulin processing (a dramatic effect on higher proinsulin levels, but no association with insulinogenic index) at extremely persuasive levels of statistical significance (P = 2.1 × 10−71). Defects in insulin processing and insulin secretion were seen in glucose-raising allele carriers at TCF7L2, SCL30A8, GIPR, and C2CD4B. Abnormalities in early insulin secretion were suggested in glucose-raising allele carriers at MTNR1B, GCK, FADS1, DGKB, and PROX1 (lower insulinogenic index; no association with proinsulin or insulin sensitivity). Two loci previously associated with fasting insulin (GCKR and IGF1) were associated with OGTT-derived insulin sensitivity indices in a consistent direction. CONCLUSIONS Genetic loci identified through their effect on hyperglycemia and/or hyperinsulinemia demonstrate considerable heterogeneity in associations with measures of insulin processing, secretion, and sensitivity. Our findings emphasize the importance of detailed physiological characterization of such loci for improved understanding of pathways associated with alterations in glucose homeostasis and eventually type 2 diabetes.

Impact of type 2 diabetes susceptibility variants on quantitative glycemic traits reveals mechanistic heterogeneity

Patients with established type 2 diabetes display both β-cell dysfunction and insulin resistance. To define fundamental processes leading to the diabetic state, we examined the relationship between type 2 diabetes risk variants at 37 established susceptibility loci, and indices of proinsulin processing, insulin secretion, and insulin sensitivity. We included data from up to 58,614 nondiabetic subjects with basal measures and 17,327 with dynamic measures. We used additive genetic models with adjustment for sex, age, and BMI, followed by fixed-effects, inverse-variance meta-analyses. Cluster analyses grouped risk loci into five major categories based on their relationship to these continuous glycemic phenotypes. The first cluster (PPARG, KLF14, IRS1, GCKR) was characterized by primary effects on insulin sensitivity. The second cluster (MTNR1B, GCK) featured risk alleles associated with reduced insulin secretion and fasting hyperglycemia. ARAP1 constituted a third cluster characterized by defects in insulin processing. A fourth cluster (TCF7L2, SLC30A8, HHEX/IDE, CDKAL1, CDKN2A/2B) was defined by loci influencing insulin processing and secretion without a detectable change in fasting glucose levels. The final group contained 20 risk loci with no clear-cut associations to continuous glycemic traits. By assembling extensive data on continuous glycemic traits, we have exposed the diverse mechanisms whereby type 2 diabetes risk variants impact disease predisposition.

Comparison of circulating phagocyte oxidative activity measured by chemiluminescence in whole blood and isolated polymorphonuclear leukocytes

The generation of reactive oxygen species (ROS) was measured by chemiluminescence (CL) in whole blood and isolated polymorphonuclear leukocytes (PMNLs) in 113 subjects. There were no differences in ROS production in males and females, smokers and non-smokers and no age dependency. A significant correlation was found between whole blood CL parameters and between parameters derived from isolated cells, but there was no correlation when these parameters were compared using isolated PMNLs and whole blood. Variations in serum lipid levels did not account for the variability of CL parameters in blood or PMNLs. Both methodologies had the same validity in demonstrating priming of phagocytes. Zymosan-induced CL was more sensitive in detecting distinct functional states of phagocytes than FMLP-induced CL.

b-adrenergic modulation of FMLP- and zymosan-induced intracellular and extracellular oxidant production by polymorphonuclear leukocytes

Evaluation of catecholamine modulation of PMNL extracellular and intracellular oxidant production may reflect beneficial and harmful effects of b-adrenergic agonists in various disease states. We investigated the kinetics and potency of adrenaline-mediated inhibition of oxidant generation in FMLP- and zymosan-stimulated PMNLs. In FMLP-stimulated cells, the short-term burst of oxidant generation was inhibited by adrenaline in a dose-dependent fashion. Intra- and extracellular chemiluminescence and extracellular superoxide anion and hydrogen peroxide generation showed similar IC50 values for adrenaline (1.3-3.0 ï 10-8 M) indicating that both extracellular and intracellular events were inhibited with the same potency. In contrast, intracellular oxidant production evoked by the phagocytosis of zymosan was only minimally affected by 3 ï 10-5 - 3 ï 10-12 M adrenaline. Extracellular inhibition of oxidant production was also apparent in zymosan-stimulated cells. In conclusion, adrenalines ability to depress extracellular generation of oxygen metabolites while retaining prolonged intracellular oxidant production for phagocytosis supports its beneficial role as selectively targeted physiological protector.

Distinct effects of LDL apheresis by hemoperfusion (DALI) and heparin-induced extracorporeal precipitation (HELP) on leukocyte respiratory burst activity of patients with familial hypercholesterolemia

Hypercholesterolemia and oxidative stress are major risk factors in atherogenesis. In the last years, lipid apheresis has been established as an effective clinical therapy by lowering not only elevated plasma low‐density lipoprotein (LDL) levels but also by reducing the incidence of cardiovascular events. The aim of the present study was to investigate peripheral leukocyte oxidant generation in patients with familial hypercholesterolemia (FH) undergoing regular LDL apheresis. The activity state of leukocytes was estimated prior to, immediately after, and 2 days after LDL apheresis carried out by two distinct techniques: hemoperfusion with the DALI system and heparin‐induced extracorporeal LDL precipitation (HELP). Oxidant generating activity was measured by chemiluminescence (CL) in whole blood and isolated polymorphonuclear leukocytes (PMNL). The results of our study show increased baseline respiratory burst activities in FH patients as compared to healthy controls. Apheresis with the HELP system was followed by increases in leukocyte count, zymosan‐induced whole blood CL, and plasma PMNL elastase levels. The DALI technique caused no changes in leukocyte count and elastase levels and decreased whole blood CL activity. Two days after lipid removal the observed changes returned to pre‐apheresis levels. Leukocyte activity parameters before and after apheresis did not correlate with the corresponding plasma levels of triglycerides, total cholesterol, and LDL cholesterol, suggesting that different handling in the framework of both apheresis techniques rather than lipid profile changes during therapy accounted for leukocyte activity modulation. J. Clin. Apheresis 15:249–255, 2000.

Laboratory stress in normotensives, borderline hypertensives and essential hypertensives is associated with priming of phagocytic cells

The present study was designed to investigate the effect of standardized laboratory physical and mental stress on phagocytic generation of reactive oxygen species (ROS) and hormonal responses in normotensives (NT), borderline hypertensives (BH) and essential hypertensives (EH). BH were characterized by increased neurohormonal activation in the prestress situation as indicated by significant higher noradrenaline and ACTH values. Although physical stress resulted in a marked activation of the sympatho-adrenal system in all groups, the BH group revealed a strong tendency of more pronounced sympatho-adrenal response. The intracellular and extracellular production of ROS was quantified using chemiluminescence (CL) assays. Phagocytic cells of NT, BH and EH responded to stress with a significantly enhanced zymosan-stimulated luminol- and lucigenin-amplified CL, predominantly localized intracellularly. In the BH group, but not in NT and EH, significant stress-related increases of extracellularly and intracellularly generated oxidative metabolites of isolated granulocytes after stimulation with the chemotactic peptide FMLP were found. Baseline values of all measured CL parameters did not differ between NT, BH and EH. The results show that stress is followed by priming of phagocytes for enhanced oxygen radical generation. In addition to stress-related activation of intracellular ROS production granulocytes of BH showed enhanced release of oxygen metabolites into the extracellular space.