Insulin secretion and insulin sensitivity in diabetic subgroups: studies in the prediabetic and diabetic state

Aims/hypothesis. To evaluate insulin sensitivity and insulin secretion in prediabetic and diabetic subjects with mutations in MODY1 (HNF-4α) and MODY3 (HNF-1α) genes, in subjects with GAD antibodies, latent autoimmune diabetes in adults and in subjects with the common form of Type II (non-insulin-dependent) diabetes mellitus. Methods. Insulin secretion was measured as the incremental 30-min insulin (I30) and insulin glucose ratio (I:G30) during OGTT whereas insulin sensitivity was measured as the insulin sensitivity index during OGTT in 131 carriers of MODY mutations [NGT = 38, IFG/IGT = 21, diabetes mellitus (DM) = 72], in 293 subjects with GADA (NGT = 47, IFG/IGT = 29, DM = 217) and in 2961 subjects with a family history of the common form of Type II diabetes but without MODY mutations or GADA (NGT = 1360, IFG/IGT = 857, DM = 744). A subgroup of the subjects underwent a euglycaemic clamp (n = 210) and intravenous glucose tolerance test (n = 337) for the estimation of insulin sensitivity and first-phase insulin secretion. Results. Non-diabetic subjects with MODY mutations had pronounced impaired insulin secretion (I30, I:G30) compared with the two other groups (p = 0.005). Normal or non-diabetic glucose tolerance was maintained by enhanced insulin sensitivity compared with the other two groups (p < 0.05 and p < 0.005). In contrast to patients with Type II diabetes and with adult latent autoimmune diabetes, MODY patients showed only a modest deterioration in insulin sensitivity at onset of diabetes. The 2-h glucose values inversely correlated with insulin sensitivity in subjects with GADA (r = –0.447, p < 0.001) and subjects from Type II diabetic families (r = –0.426, p < 0.001), whereas no such relation was observed in subjects with MODY mutations (r = 0.151, p = NS). There were no statistically significant differences in insulin secretion or insulin sensitivity between subjects with GADA or subjects with a family history of Type II diabetes, either at the NGT or the IFG/IGT stage. Conclusion/interpretation. Glucose-tolerant carriers of MODY mutations are characterised by a severe impairment in insulin secretion. Enhanced insulin sensitivity is the most likely explanation for the normal glucose tolerance. Whereas subjects with positive GADA or Type II diabetes have impaired insulin sensitivity with increasing glucose concentrations, MODY mutation carriers seem to be protected from the effect of glucose toxicity. [Diabetologia (2000) 43: 1476–1483] Received: 23 March 2000 and in revised form: 29 August 2000     

Oxidative stress could precede endothelial dysfunction and insulin resistance in Indian Mauritians with impaired glucose metabolism

Aims/hypothesis. To measure oxidative stress, endothelial dysfunction and insulin resistance in Indian Mauritians at different stages of development of Type II (non-insulin-dependent) diabetes mellitus. Methods. Plasma total 8-epi-PGF₂α, an indicator of oxidative stress, was determined in age-matched subjects with normal glucose metabolism (n = 39), impaired glucose tolerance (n = 14), newly diagnosed diabetes (n = 8) and established diabetes (n = 14). Plasma glucose and insulin were measured at baseline and 2 h following an oral glucose tolerance test. Endothelial function was assessed by non-invasive digital pulse wave photoplethysmography. Results. Plasma 8-epi-PGF₂α increased in subjects with impaired glucose tolerance (p < 0.05) compared with control subjects, and was even higher in newly diagnosed diabetic patients (p < 0.01) and established (p < 0.01) diabetic patients. A tendency towards reduced endothelial function in subjects with impaired glucose tolerance became significant in patients with newly diagnosed and established diabetes (p < 0.01), and was correlated with 8-epi-PGF₂α (r = 0.36, p < 0.01). Insulin resistance (homeostasis model assessment) did not change in subjects with impaired glucose tolerance compared with control subjects, but increased in newly diagnosed (p < 0.01) and established (p < 0.001) diabetic subjects. The 8-epi-PGF₂α was correlated with fasting glucose (r = 0.50, p < 0.001), triglycerides (r = 0.40, p < 0.001) and insulin resistance (r = 0.35, p < 0.001). Conclusion/interpretation. Oxidant stress is an early event in the evolution of Type II diabetes and could precede the development of endothelial dysfunction and insulin resistance. [Diabetologia (2001) 44: 706–712 Received: 1 November 2001 and in revised form: 14 February 2001     

Diabetic nephropathy is associated with low-grade inflammation in Type 1 diabetic patients

Aims/hypothesis Increased concentrations of C-reactive protein and interleukin-6, a finding suggestive of the presence of inflammation, have been observed in Type 2 diabetes. In such patients, C-reactive protein was predictive of diabetic nephropathy. Studies on low-grade inflammatory markers and nephropathy in Type 1 diabetic patients have shown conflicting results. Therefore we studied whether low-grade inflammation is associated with diabetic nephropathy in Type 1 diabetic patients.Methods We divided 194 Type 1 diabetic patients into three groups from the Finnish Diabetic Nephropathy Study based upon their albumin excretion rate. Patients with normoalbuminuria (n=67) had no antihypertensive medication or signs of cardiovascular disease, while patients with microalbuminuria (n=64) or macroalbuminuria (n=63) were all treated with an angiotensin-converting enzyme inhibitor, a drug that could attenuate low-grade inflammation. As a measure of insulin sensitivity we used estimated glucose disposal rate. C-reactive protein was measured by radioimmunoassay and interleukin-6 by high sensitivity enzyme immunoassay.Results C-reactive protein was higher in micro- and macroalbuminuric patients compared to normoalbuminuric patients (normoalbuminuria 2.0±1.7, microalbuminuria 2.6±1.7, macroalbuminuria 2.9±2.5 mg/l; p=0.016), while interleukin-6 increased in parallel with the severity of the renal disease (1.9±1.5, 2.9±3.3, 3.6±3.1 ng/l; p<0.0001). In multiple regression analysis albumin excretion rate was the only variable independently associated with C-reactive protein (p=0.03), whereas albumin excretion rate (p=0.0003), HDL-cholesterol (p=0.0135) and duration of diabetes (p=0.0176) were independently associated with interleukin-6.Conclusions/interpretation Low-grade inflammatory markers are associated with diabetic nephropathy in Type 1 diabetic patients. The predictive value needs to be assessed.Abbreviations DN diabetic nephropathy - CRP C-reactive protein - eGDR estimated glucose disposal rate - FinnDiane finnish diabetic nephropathy study - MDRD modification of diet in renal disease     

Serum Immunoglobulin Concentrations in Diabetic Patients

The relationship between glycated haemoglobin (an index of long-term diabetic control), fructosamine (an index of intermediate-term diabetic control), and serum IgA, IgG, and IgM was studied in 110 diabetic patients (41 Type 1 and 69 Type 2) and compared with 111 healthy non-diabetic subjects. Significant increases in serum IgA (by 82.7%, p < 0.001) and IgG (by 35.2%, p< 0.001) concentrations were observed whereas the concentration of IgM was significantly decreased (by 46.7%, p < 0.001) in diabetic patients compared with non-diabetic subjects. Using Spearman's rank correlations, IgA correlated with fructosamine (r = 0.77, p < 0.001), HbA₁ (r = 0.76, p < 0.001), and albumin (r = ?0.58, p < 0.001) for the entire population sample but only fructosamine (r = 0.19, p < 0.05) and HbA₁ (r = 0.28, p < 0.001) correlated with IgA in diabetic patients, respectively. It is concluded that abnormal levels of IgA, IgG, and IgM are very common in diabetic patients in whom serum IgA concentrations are influenced by the degree of glycaemic control. Whether changes in IgA and other immunoglobulins are implicated in the pathogenesis of diabetic complications (such as susceptibility to infection) deserve further study.     

Psychological Factors and their Relationship to Diabetes Control

Thirty-nine Type 1 diabetic patients were asked to complete an Eysenck Personality Questionnaire (EPQ). A significant relationship was found between neuroticism scores and glycosylated haemoglobin concentrations (r = 0.43, p < 0.01) and also fructosamine (r = 0.45, p < 0.01). Patients with glycosylated haemoglobin concentrations greater than or equal to 10 (n = 13) had significantly higher neuroticism scores than patients with glycosylated haemoglobin results less than or equal to 8 (n = 11) (p < 0.01).     

Enhanced P-selectin expression and increased soluble CD40 Ligand in patients with Type 1 diabetes mellitus and microangiopathy: evidence for platelet hyperactivity and chronic inflammation

Aims/hypothesis Platelet activation, endothelial dysfunction and inflammation may be involved in early stages of diabetic microangiopathy. We therefore investigated patients with Type 1 diabetes mellitus, without (n=19) and with (n=20) microangiopathy, matched for glycaemic control and duration of disease, and matched with healthy control subjects (n=27).Methods Platelet activation was measured as platelet P-selectin expression using whole blood flow cytometry and as soluble P-selectin by immunoassay. Von Willebrand factor antigen in plasma, serum soluble E-selectin, CD40 ligand (sCD40L) and C-reactive protein (CRP) served as markers for endothelial function and inflammation.Results Thrombin-induced platelet P-selectin expression was enhanced, and soluble P-selectin and sCD40L concentrations were increased in patients with microangiopathy compared with the control subjects (p<0.01 for both) and with patients without microangiopathy (p<0.05 for P-selectin expression and sP-selectin), whereas all three parameters were similar in patients without microangiopathy and in the control subjects. CRP and soluble E-selectin were increased in patients with microangiopathy, compared with the control subjects (p<0.01 and p<0.05), whereas von Willebrand factor did not differ between the groups.Conclusions/interpretation Microangiopathy in Type 1 diabetes is associated with platelet hyperactivity, endothelial dysfunction and low-grade inflammation, indicating an increased risk for cardiovascular disease.Abbreviations sP-selectin Soluble P-selectin - sCD40L soluble CD40 Ligand - CRP C-reactive protein     

Bone mineral density, collagen type 1 α 1 genotypes and bone turnover in premenopausal women with diabetes mellitus

Summary Osteopenia is a recognised complication of diabetes mellitus which could be due to abnormal bone turnover or disturbances in the calcium/parathyroid hormone/vitamin D axis or both. Genetic factors also play an important part in determining bone mass although this has not been studied in diabetes. Recently a polymorphism of the collagen type 1 α 1 (COL1A1) gene has been shown to be associated with low bone mass in British women. To identify subjects with diabetes who may be at risk of developing osteoporosis and fractures, we analysed bone mineral density in relation to the biochemical markers of bone turnover, calcium homeostasis and the COL1A1 genotype in a group of premenopausal women with Type I (insulin-dependent) diabetes mellitus (n = 31), Type II (non-insulin dependent) diabetes mellitus (n = 21) and control subjects (n = 20). Bone mineral density was lower at the femoral neck in the subjects with Type I diabetes (p = 0.08) as were serum 25-hydroxyvitamin D compared with control subjects (p = 0.023) and this was negatively correlated with serum collagen type 1 C-terminal propeptide (r = –0.56, p < 0.001). Bone mineral density in Type II diabetes was not different from control subjects, after correction for body mass index. Bone resorption was, however, raised in the Type II diabetic subjects as reflected by the higher urinary deoxypyridinoline values (p = 0.016) and lower collagen type 1 C-terminal propeptide:deoxypyridinoline ratio (p = 0.04). In the whole group studied, subjects with the COL1A1 ’s' genotype had lower bone mineral density at the femoral neck (p = 0.01) which was partly attributable to a lower body mass index. Following multiple regression analysis body mass index and collagen type 1 C-terminal propeptide concentrations remained determinants of bone mass at all three sites, whereas genotype appeared to be a predictor of bone mass at the femoral neck only. We conclude that measurement of these variables could prove useful in firstly identifying those diabetic women at risk of osteoporosis and secondly guiding therapeutic intervention. [Diabetologia (1998) 41: 1314–1320] Received: 2nd March 1998 and in revised form: 27 April 1998     

The relation of proinsulin and insulin to insulin sensitivity and acute insulin response in subjects with newly diagnosed Type II diabetes: the Insulin Resistance Atherosclerosis Study

Abstract Aims/hypothesis. Proinsulin concentrations are increased relative to insulin concentrations in subjects with Type II (non-insulin-dependent) diabetes mellitus. This could be secondary to hyperglycaemia or insulin resistance or due to a defect in insulin secretion. Methods. We investigated the association between fasting insulin, intact proinsulin and the intact proinsulin: insulin ratio with insulin sensitivity, estimated by a frequently sampled intravenous glucose tolerance test and the minimal model and with acute insulin response (AIR) in 182 newly diagnosed Type II diabetic subjects aged 40 to 69 years. None of the subjects was receiving hypoglycaemic medication. Results. Insulin sensitivity correlated inversely with fasting insulin (r _s = –0.42) and intact proinsulin (r _s = –0.32) (p < 0.001). The intact proinsulin:insulin ratio was not correlated with insulin sensitivity. AIR correlated positively with intact proinsulin (r _s = 0.23) and inversely with the intact proinsulin:insulin ratio (r _s = –0.29, p < 0.001). Fasting glucose correlated positively with intact proinsulin (r _s = 0.34) and the intact proinsulin:insulin ratio (r _s = 0.24, p < 0.001). The intact proinsulin:insulin ratio increased by decreasing AIR (quartiles of AIR from high to low: 7.8, 8.2, 9.7 and 12.1 %, p < 0.001). This association was independent of age, sex, ethnicity, body mass index, fasting glucose, and insulin sensitivity. Conclusion/interpretation. Insulin resistance (low insulin sensitivity) was not related to the intact proinsulin:insulin ratio in subjects with Type II diabetes. In contrast, both low AIR and high fasting glucose concentrations were associated with a disproportionate increase in proinsulin concentration. These results suggest that increased intact proinsulin:insulin ratio is a marker of a defect in insulin secretion in Type II diabetic subjects. [Diabetologia (1999) 42: 1060–1066] Received: 25 February 1999 and in revised form: 12 April 1999     

Genetic and immunological characteristics of Type I diabetes mellitus in an Indo-Aryan population

Abstract Aims/hypothesis. Our aim was to characterise the genetic and immunological features associated with Type I (insulin-dependent) diabetes mellitus in a cohort of Indo-Aryan children resident in the United Kingdom.?Methods. Children with Type I diabetes (n = 53), unaffected first-degree relatives (n = 146) and unrelated healthy control children (n = 54) were typed for alleles of the HLA-DRB1, HLA-DQA1 and HLA-DQB1 genes. Islet cell antibodies and antibodies to glutamic acid decarboxylase, protein tyrosine phosphatase-2 (IA-2ic) and insulin were measured in the diabetic and control children.?Results. The DRB1^*03.DQA1*05.DQB1^*02 haplotype was positively associated with the disease, occurring in 78 % of diabetic children compared with 22.6 % of healthy children (p _c < 2.4 × 10^–5). In simplex families, this haplotype was transmitted more frequently to the diabetic children than to their unaffected siblings (p < 1 × 10^–4). The DRB1^*04.DQA1* 03.DQB1^*0302 haplotype was also transmitted preferentially to the diabetic probands (p < 0.025) but was not associated with disease in the case control study. Islet-related autoantibodies were detected in 89.6 % of diabetic patients compared with 11.8 % of control children (p < 1 × 10^–6). Although protein tyrosine phosphatase-2 autoantibodies were detected more frequently among DRB1^*04-positive diabetic patients compared with patients lacking this allele, the overall frequency of these autoantibodies was lower than observed in Europid diabetic subjects. This could reflect the absence of a disease association with DRB1*04 in the Indo-Aryan cohort.?Conclusion/interpretation. Type I diabetes in our Indo-Aryan cohort is similar to the disease observed in Anglo-Europeans but has important immunogenetic differences. The low frequency of protein tyrosine phosphatase-2 autoantibodies among the Indo-Aryan diabetic children could have important implications for the design of future strategies for disease prediction in this population. [Diabetologia (2000) 43: 450–456] Received: 2 September 1999 and in revised form: 8 November 1999     

Genetic influence of the R/Q353 genotype on factor VII activity is overwhelmed by environmental factors in Chinese patients with Type II (non-insulin-dependent) diabetes mellitus

Summary We studied the effects of genetic and environmental influences on factor VII coagulant activity (VIIc) in Chinese diabetic patients (263 with Type II [non-insulin-dependent] diabetes mellitus, 78 with Type I [insulin-dependent] diabetes mellitus) and 143 normal control subjects. VIIc was measured by a one-stage biological assay. The R/Q353 or Msp1 polymorphism at codon 353 of the factor VII gene was detected after Msp1 digestion of polymerase chain reaction-amplified genomic DNA. In both diabetic and control subjects the allele frequencies of the R (M1) and Q (M2) alleles were 0.96 and 0.04; the corresponding reported frequencies in Caucasians being 0.90 and 0.10: VIIc were 21 % lower in Chinese control subjects and Type I diabetic patients with R/Q, compared with R/R subjects (p < 0.001 and p < 0.05). The corresponding difference was 4 % for Type II diabetc patients (p = NS). Type II diabetic patients had higher mean VIIc levels than control subjects and Type I diabetic patients (p < 0.01); they were also older, and had higher serum creatinine and triglyceride (all p < 0.01). They also had higher VIIc levels than an age-matched older control group (p < 0.01; n = 182) in whom the genotype effect was clearly seen. On stepwise linear regression analysis, the significant independent determinants of VIIc were serum triglyceride (contributing 20 % and 25 % to variance in control subjects and diabetic patients), the R/Q353 genotype (contributing to 12 % of the variance in control subjects but only 1 % in diabetic patients), age and total cholesterol in all subjects, and in the diabetic patients female sex, urinary albumin excretion rate and serum creatinine. VIIc was higher in diabetic patients with macroangiopathy and retinopathy (both p < 0.0001). We conclude that compared with Caucasians, the Q allele frequency is significantly lower in these Chinese subjects. Plasma VIIc is determined by both genetic and environmental influences such that in Chinese Type II diabetic patients, the effect of environmental factors predominates, almost negating the influence of the R/Q353 genotype. High VIIc may contribute to the increased cardiovascular risk in Type II diabetic patients. [Diabetologia (1998) 41: 760–766] Received: 24 October 1997 and in final revised form: 28 January 1998     

Short-term moderate sodium restriction induces relative hyperfiltration in normotensive normoalbuminuric Type I diabetes mellitus

Aims/hypothesis. Type I (insulin-dependent) diabetes mellitus is associated with an increased extracellular volume. Sodium restriction might seem a logical form of treatment but data on its renal effects is conflicting. We therefore studied the effects of sodium restriction on renal haemodynamics in uncomplicated Type I diabetes mellitus. Methods. Uncomplicated Type I diabetic patients (n = 24) and matched control subjects (n = 24) were studied twice in random order: after a week of 50 mmol or after 200 mmol sodium intake, respectively. The diabetic patients were studied under normoglycaemic clamp conditions. Glomerular filtration rate and effective renal plasma flow were measured as the clearances of iothalamate and hippuran, respectively. Results. During liberal sodium intake, glomerular filtration, effective renal plasma flow and filtration fraction were similar between the diabetic patients and the control subjects. Sodium restriction decreased the effective renal plasma flow in both groups, whereas glomerular filtration rate only decreased in the control subjects. Consequently, in the diabetic patients, the filtration fraction was increased on low sodium (4.1 ± 8.4 %, p < 0.05 vs liberal sodium). As a consequence, filtration fraction (24.0 ± 2.6 vs 22.1 ± 2.0 %, p < 0.05) and glomerular filtration (119 ± 14 vs 110 ± 13 ml/min, p < 0.05) were higher in the diabetic patients than in the control subjects during sodium restriction. Conclusion/interpretation. Short-term moderate sodium restriction induces relative hyperfiltration in uncomplicated Type I diabetes. This could indicate an increased intraglomerular pressure. Sodium restriction could be an unfavourable preventive approach in diabetes mellitus but its long-term effects are not known. [Diabetologia (2002) 45: ▪–▪] Received: 17 September 2001 and in revised form: 7 November 2001     

Morphometric documentation of abnormal intramyocellular fat storage and reduced glycogen in obese patients with Type II diabetes

Aims/hypothesis. Insulin resistance of skeletal muscle has been associated with increased lipid availability. This study aimed to estimate volume fractions of intramyocellular triglyceride droplets and glycogen granules in skeletal muscle using electron microscopy and furthermore, relate these findings to insulin sensitivity and the level of circulating lipids. Methods. We compared 11 obese patients with Type II (non-insulin-dependent) diabetes mellitus and 11 obese normoglycaemic subjects matched for age and sex. Glucose metabolism was determined using the euglycaemic hyperinsulinaemic clamp technique (40 mU · m^–2· min^–1) coupled with indirect calorimetry and tritiated glucose. On the second day, using an automatic procedure, a fasting muscle biopsy was carried out and processed for electron microscopy. Volume fractions of intramyocellular structures were estimated by pointcounting on photographic pictures in a blinded manner. Results. Insulin-stimulated total glucose disposal rate was lower in the Type II diabetic subjects compared with the obese normoglycaemic subjects (4.96 ± 049 vs 10.35 ± 0.89 mg · min^–1· kg ffm^–1, p < 0.001) as was glucose storage (2.03 ± 0.50 vs 6.59 ± 0.83, p < 0.001). The electron microscopy study revealed that the diabetic subjects had higher intramyocellular amounts of triglyceride (1.43 ± 0.21 vs 0.39 ± 0.07 %, p < 0.001) and lower amounts of glycogen (3.53 ± 0.33 vs 6.94 ± 0.54 %, p < 0.001). Mitochondrial volume was identical indicating equal aerobic capacity. The fractional intramyocellular lipid volume was found to be positively associated with fasting NEFA (r = 0.63, p = < 0.05 and r = 0.79, p = < 0.05) and triglyceride (r = 0.74, p = 0.01 and r = 0.62, p < 0.05) in the obese diabetic and normoglycaemic cohorts respectively. Intramyocellular lipid content was negatively correlated to insulin sensitivity (r = –0.71, p < 0.02) in the obese diabetic group whereas no significant association was found in the obese normoglycaemic group. Conclusion/interpretation. This study shows that fat accumulates intramyocellulary while glycogen stores are simultaneously reduced in obese subjects with Type II (non-insulin-dependent) diabetes mellitus. Quantitatively, a major component of the excessive lipid accumulation could be secondary in origin, related to the diabetic state in itself, although a contribution from the altered insulin action cascade of obesity and diabetes cannot be excluded. In both groups significant positive relations were found between circulating and intramyocellular lipid. [Diabetologia (2001) 44: 824–833] Received: 6 December 2000 and in revised form: 16 March 2001     

Glucosaminyl N-deacetylase in cultured fibroblasts: comparison of patients with and without diabetic nephropathy,and identification of a possible mechanism for diabetes-induced N-deacetylase inhibition

Summary Impaired heparan sulphate biosynthesis through diabetes-induced inhibition of glucosaminyl N-deacetylase may have a central role in the development of diabetic nephropathy, and genetic differences in the vulnerability of the N-deacetylase could influence the risk of developing nephropathy. We studied N-deacetylase activity in fibroblast cultures from Type 1 (insulin-dependent) diabetic patients with (n = 14) or without (n = 13) diabetic nephropathy, together with non-diabetic control subjects (n = 7). No difference in N-deacetylase activity was found (p = 0.13), and no inhibition of N-deacetylase was found in cultures grown at 25 mmol/l glucose. N-deacetylase activity was inversely correlated to growth rate (r=−0.59,p=0.0008), and in patients with nephropathy a negative correlation between HbA_1C and fibroblast N-deacetylase activity (r=−0.72,p = 0.012) was found. Cell-cycle analysis revealed an increased fraction of S-phase cells in patients with nephropathy (28%(21–52%)) compared to healthy control subjects (17% (9–24%)),p=0.0008, but not between patients with and without nephropathy (latter group 26%(11–43%)),p = 0.43. Forskolin, an activator of protein kinase A, specifically decreased N-deacetylase activity, whereas activation of protein kinase C produced a combined reduction in N-deacetylase activity and total protein synthesis. In conclusion, no constitutive defects in N-deacetylase activity were found in fibroblasts from these patients. Further studies should consider possible associations between fibroblast characteristics and pre-biopsy environmental parameters related to cellular memory phenomena. Finally, activation of protein kinase A provides a potential general pathway for regulating N-deacetylase activity.     

Relationship of skeletal muscle glucose 6-phosphate to glucose disposal rate and glycogen synthase activity in insulin-resistant and non-insulin-dependent diabetic rhesus monkeys

Summary Reduced insulin action on skeletal muscle glycogen synthase activity and reduced whole-body insulin-mediated glucose disposal rates in insulin-resistant subjects may be associated with an alteration in muscle glucose transport (or phosphorylation) or with a defect distal to glucose 6-phosphate. To examine this issue we determined the glucose 6-phosphate concentration and glycogen synthase activity in muscle samples obtained under basal and euglycaemic hyperinsulinaemic clamp conditions in 27 rhesus monkeys (Macaca mulatta). They ranged from metabolically normal (n =11) to insulin-resistant (n =8) to overtly diabetic (non-insulin-dependent) (n =8). The glucose 6-phosphate measured under insulin-stimulated conditions was inversely correlated to insulin-stimulated glycogen synthase independent activity (r = –0.54, p<0.005), the change in glycogen synthase independent activity (insulin-stimulated minus basal) (r = –0.58, p<0.002) and to whole-body insulin-mediated glucose disposal rate (r = –0.60, p<0.002). The insulin-resistant and diabetic monkeys had significantly higher insulin-stimulated glucose 6-phosphate concentrations (0.57±0.11 and 0.62±0.11 nmol/mg dry weight, respectively) compared to the normal monkeys (0.29±0.05 nmol/mg dry weight) (p's <0.05). We conclude that under euglycaemic/hyperinsulinaemic conditions, a defect distal to glucose 6-phosphate is a major contributor to reduced whole-body insulin-mediated glucose disposal rates and to reduced insulin action on glycogen synthase in insulin-resistant and diabetic monkeys. [Diabetologia (1994) 37: 127–133] Received: 28 May 1993 and in revised form: 13 August 1993     

Production and metabolic clearance of glucose under basal conditions in Type II (non-insulin-dependent) diabetes mellitus

Aims/hypothesis: The pathogenesis of fasting hyperglycaemia in Type II (non-insulin-dependent) diabetes mellitus has yet to be clarified. Rates of glucose production (R _a), utilization and metabolic clearance rate were therefore measured during an extended fast, in control subjects and in Type II diabetic patients. Methods: Nine subjects with newly-diagnosed or diet-treated diabetes and seven control subjects matched for age and weight (BMI 36.0 ± 2.4 and 35.3 ± 3.1 kg/m² respectively) underwent an overnight fast followed by a 10-h unprimed infusion of [6-³H]glucose. Plasma tracer concentrations were fitted by a single-compartment model. Results: The metabolic clearance rate was near-constant [61.7 + 2.4 ml/(min-m²)] in diabetic patients and [75.5 ± 3.3 ml/(min-m²)] in control subjects (p < 0.05). It was correlated to the glucose concentrations both at t = 0 (r = –0.752, p = 0.0008) and t = 10 h (r = –0.675, p = 0.004). The calculated volume of distribution was 17.3 ± 1.4 l (18.2 % weight, diabetes), 19.6 ± 2.4 l (18.4 % weight, control). Glycaemia fell from 10.7 ± 0.8 mmol/l to 6.5 ± 0.3 mmol/l by 10 h (p < 0.05) in diabetes and from 5.6 ± 0.6 to 4.8 ± 0.1 mmol/l in control subjects (p < 0.05). The rate of glucose production decreased in parallel, from 563 ± 33 to 363 ± 23 μmol/(min-m²) (p < 0.05) in diabetes from 419 ± 20 to 347 ± 32 μmol/(min-m²) in control subjects. Initial R _a was higher in diabetic patients than in control subjects (p < 0.05) and was highly correlated to glycaemia (r = 0.836, p = 0.0001). By 10 h, R _a had converged in diabetic patients and control subjects and all correlation with glycaemia was lost (r = 0.0017, p = 0.95). Conclusions/interpretation: In relatively early diabetes, the more “labile” portion of fasting hyperglycaemia, which subsequently decreased, was closely related to the simultaneously decreasing R _a. The 25 % increase in glucose concentrations which persisted as stabilized R _a, resulted from about a 20 % lower metabolic clearance rate. [Diabetologia (2001) 44: 983–991] Received: 28 February 2001 and in revised form: 17 April 2001     

Plasma phospholipid transfer protein activity is related to insulin resistance: impaired acute lowering by insulin in obese Type II diabetic patients

Summary Cholesteryl ester transfer protein (CETP) and phospholipid transfer protein (PLTP) have important functions in high density lipoprotein (HDL) metabolism. We determined the association of plasma CETP and PLTP activities (measured with exogenous substrate assays) with insulin resistance, plasma triglycerides (TG) and non-esterified fatty acids (NEFA), and assessed the lipid transfer protein response to insulin during a 6–7 h hyperinsulinaemic euglycaemic clamp in non-obese and obese healthy subjects and patients with Type II (non-insulin-dependent) diabetes mellitus (n = 8 per group). Plasma PLTP activity was higher in obese healthy subjects and obese Type II diabetic patients compared with non-obese healthy subjects (p < 0.05 to 0.01) and was correlated with insulin resistance, plasma TG and NEFA (p = 0.02 to < 0.01). In non-obese healthy subjects, insulin decreased plasma TG and increased the HDL cholesteryl ester (CE)/TG ratio (p < 0.01 compared with saline infusion). Plasma PLTP activity fell by 14 % at the end of the clamp (p < 0.01 compared with saline) but CETP activity did not change. The decreases in plasma NEFA, TG and PLTP activity and the rise in HDL CE/TG were smaller in obese Type II diabetic patients than in non-obese healthy subjects (p < 0.01 for all). Baseline HDL CE/TG was negatively correlated with plasma TG (p < 0.001, n = 32) and PLTP activity (p < 0.01) but not with CETP activity. Likewise, the rise in HDL CE/TG during the clamp was related to the fall in plasma TG (p < 0.001) and in PLTP activity (p < 0.02). It is concluded that plasma PLTP, but not CETP, is regulated by insulin in an acute setting. High plasma PLTP activity is associated with insulin resistance in conjunction with altered NEFA and triglyceride metabolism. High plasma TG and PLTP activity have coordinate effects on HDL metabolism. [Diabetologia (1998) 41: 929–934] Received: 12 January 1998 and in revised form: 9 April 1998     

Diabetic retinopathy risk correlates with intracellular concentrations of the glycoxidation product Ne-(carboxymethyl) lysine independently of glycohaemoglobin concentrations

Aims/hypothesis. We investigated whether either the amount of diabetes-induced intracellular oxidative stress or the concentration of hyperglycaemia-induced advanced glycation endproducts is associated with the risk of diabetic retinopathy. Methods. We measured concentrations of the glycoxidation product N^e-(carboxymethyl)lysine and two non-oxidation-dependent advanced glycation endproducts (methylglyoxal-derived and 3-deoxyglucosone-derived) in CD45RA⁺ T-cells from 21 Type I (insulin-dependent) diabetic patients with and without diabetic retinopathy and from age-matched non-diabetic control subjects. Results. Intracellular concentrations of both oxidation-dependent N^e-(carboxymethyl)lysine and oxidation-independent advanced glycation endproducts were increased in memory T-cells from diabetic patients. N^e-(carboxymethyl)lysine: diabetic median-24 176 arbitrary units/mg protein (95 % confidence interval 18 690–34 099 arbitrary units/mg protein); nondiabetic-9088 arbitrary units/mg protein (confidence interval 6994–10 696 arbitrary units/mg protein; p < 0.0001). Methylglyoxal-derived advanced glycation end products: diabetic-5430 arbitrary units/mg protein (confidence interval 3458–13 610); nondiabetic-271 arbitrary units/mg protein (confidence interval 61–760 arbitrary units/mg protein; p < 0.0001). 3-Deoxyglucosone-derived advanced glycation end products: diabetic-8070 arbitrary units/mg protein (confidence interval 7049–16 551 arbitrary units/mg protein); nondiabetic-1479 arbitrary units/mg protein (confidence interval 1169–3170; p < 0.0001). Only N^e-(carboxymethyl)lysine concentrations, however, inversely correlated with the duration of retinopathy-free diabetes (r = –0.51; p < 0.02). Diabetes-dependent N^e-(carboxymethyl)lysine accumulation did not correlate with age, diabetes duration, or averaged glycohaemoglobin concentrations. In vitro experiments wih menadione and lymphocytes confirmed that N^e-(carboxymethyl)lysine concentrations reflect intracellular oxidative stress. Conclusion/interpretation. Monitoring intracellular concentrations of increased oxidative stress in long-lived CD45RA⁺ lymphocytes by markers such as N^e-(carboxymethyl)lysine possibly identifies a subgroup of patients at high risk for microvascular complications. [Diabetologia (1999) 42: 603–607] Received: 9 November 1998 and in revised form: 18 December 1998     

Relationship between non-enzymatic glycosylation and changes in serum insulin-like growth factor-1 (IGF-1) and IGF-binding protein-3 levels in patients with type 2 diabetes mellitus

The possible occurrence of increased non-enzymatic glycosylation of serum insulin-like growth factor binding protein-3 (IGFBP-3) in vivo and the changes that would simultaneously occur in serum levels of IGFBP-3 and insulin-like growth factor-1 (IGF-I) were investigated. We measured levels of IGF-I and IGFBP-3 and the degree of glycation of total serum protein and IGFBP-3, in serum samples obtained from patients with poorly controlled non-insulin-dependent diabetes (type 2) and from age-matched non-diabetic controls. Type 2 diabetic patients had significantly higher glycated serum protein (GlyP) levels. GlyP significantly correlated with age in the control (r = 0.315, P<0.05) but not in the type 2 diabetes group. Control and diabetic subjects had comparable serum IGF-I levels and in both groups IGF-I levels tended to decrease with age (r = –0.567, P<0.001 and r = –0.465, P<0.05 for control and type 2 diabetic subjects, respectively). In the type 2 diabetes group, IGF-I levels showed a negative correlation with serum GlyP values (r = –0.476, P<0.05). Type 2 diabetic and control patients had comparable serum IGFBP-3 levels, which were significantly higher in diabetic patients in the older age subgroups. A negative correlation was found between IGFBP-3 levels and age in the control (r = –0.705, P<0.001) and in the type 2 diabetes groups (r = –0.463, P<0.05). A significant negative correlation was found between IGFBP-3 levels and GlyP in control (r = –0.449, P<0.002) but not in type 2 diabetic subjects. The mean glycated IGFBP-3 (GlyIGFBP-3) levels were higher in the oldest type 2 diabetic patients. In these patients, GlyIGFBP-3 was negatively associated with IGF-I levels (r = –0.447, P<0.05). The IGF-I/IGFBP-3 molar ratio was significantly reduced in the 46–60-year-old type 2 diabetic group, whereas the IGF-I/IGFBP-3 ratio was positively and significantly correlated with GlyP levels only in the control group (r = 0.489, P<0.01). Our results show that: a) increased non-enzymatic glycosylation of IGFBP-3 occurs in vivo; and b) this effect is accompanied by an increase in IGFBP-3 levels. These results suggest that the IGF-I/IGFBP-3 system is another target for the metabolic derangements of type 2 diabetes. Its alterations might play a role in diabetic complications. Received: 22 September 1997 / Accepted in revised form: 30 April 1998     

Haptoglobin phenotype and diabetic nephropathy

Abstract Aims/hypothesis. To determine if the haptoglobin 2 allele is associated with an increased risk for the development of diabetic nephropathy. Methods. This study included 110 consecutive normotensive subjects with Type I (insulin-dependent) diabetes mellitus and Type II (non-insulin-dependent) diabetes mellitus seen in two outpatient clinics in Israel. Diabetes duration was greater than 10 years for Type I diabetes and more than 5 years for Type II diabetic subjects. Microalbuminuria was defined as urinary protein excretion of 30 to 300 mg/24 h, and macroalbuminuria was defined as urinary protein excretion of greater than 300 mg/24 h. Serum was taken from subjects for haptoglobin typing by gel electrophoresis. Results. Of the participating subjects 54 had Type I and 56 had Type II diabetes. None (0/18) of the subjects homozygous for the haptoglobin 1 allele (1–1) showed any sign of diabetic nephropathy, as compared with 34 % (19/55) of subjects homozygous for the haptoglobin 2 allele (2–2) and 27 % (10/37) of heterozygous subjects (2–1) (p < 0.04). Of the subjects 29 showed macroalbuminuria. The risk of developing macroalbuminuria was found to be greater in subjects with two haptoglobin 2 alleles (22 %) (12/55) as compared with one haptoglobin 2 allele (8 %) (3/37) or no haptoglobin 2 alleles (0 %) (0/18) (p < 0.03). Conclusion/interpretation. By showing a graded risk relation to the number of haptoglobin 2 alleles in Type I and Type II diabetic subjects, these studies further support our hypothesis that the haptoglobin phenotype is a major susceptibility gene for the development of diabetic nephropathy. [Diabetologia (2001) 44: 602–604] Received in revised from: 5 January 2001     

Physical training decreases plasma thrombomodulin in Type I and Type II diabetic patients

Aims/hypothesis. Endothelial damage is an early step in the pathogenesis of atherosclerosis and its improvement through physical training can contribute to the known reduction of cardiovascular risk associated with exercise. An increase in some endothelium-dependent haemostatic parameters, considered as markers of endothelial damage, has been observed in diabetic patients. Methods. The effect of a three-month physical exercise programme on thrombomodulin, tissue factor pathway inhibitor, plasminogen activator inhibitor, tissue-type plasminogen activator and von-Willebrand factor was evaluated in 14 well-controlled patients with Type I (insulin-dependent) diabetes mellitus and 13 patients with Type II (non-insulin-dependent) diabetes mellitus (HbA_{1 c} 6.5 ± 0.8 and 7.4 ± 0.8 %, respectively). A matched control group was also studied. Results. Thrombomodulin at baseline was higher in both Type I and Type II diabetic patients than in their respective matched control subjects (50.0 ± 16 vs 31.1 ± 8.7 μg/l, p < 0.05; 51.0 ± 10 vs 28.5 ± 11 μg/l, p < 0.05, respectively). After the exercise programme, thrombomodulin plasma concentrations had decreased (p < 0.05) in both groups of patients, with final thrombomodulin values being similar to those observed in their control groups (38.2 ± 11 μg/l for Type I and 34.6 ± 12 μg/l for Type II patients). The thrombomodulin decrement correlated with baseline thrombomodulin and VO_2max increase in Type I diabetic patients. A decrease in tissue factor pathway inhibitor was also observed in Type II diabetic patients. Conclusion/interpretation. We conclude that the normalisation of plasma thrombomodulin concentrations in Type I and Type II diabetic patients after physical training might reflect the improvement in endothelial function associated with physical exercise. [Diabetologia (2001) 44: 693–699]