Use of Oral Anticoagulation and Diabetes Do Not Inhibit the Angiogenic Potential of Hypoxia Preconditioned Blood-Derived Secretomes.

Patients suffering from tissue ischemia, who would greatly benefit from angiogenesis-promoting therapies such as hypoxia preconditioned blood-derived secretomes commonly receive oral anticoagulation (OA) and/or have diabetes mellitus (DM). In this study, we investigated the effect of OA administration on the in vitro angiogenic potential of hypoxia preconditioned plasma (HPP) and serum (HPS), prepared from nondiabetic/diabetic subjects who did not receive OA (n = 5) or were treated with acetylsalicylic acid (ASA, n = 8), ASA + clopidogrel (n = 10), or nonvitamin K antagonist oral anticoagulants (n = 7) for longer than six months. The effect of DM was differentially assessed by comparing HPP/HPS obtained from nondiabetic (n = 8) and diabetic (n = 16) subjects who had not received OA in the past six months. The concentration of key proangiogenic (vascular endothelial growth factor or VEGF) and antiangiogenic (thrombospondin-1 or TSP-1 and platelet factor-4 or PF-4) protein factors in HPP/HPS was analyzed via ELISA, while their ability to induce microvessel formations was examined in endothelial cell cultures. We found that OA use significantly reduced VEGF levels in HPP, but not HPS, compared to non-OA controls. While HPP and HPS TSP-1 levels remained largely unchanged as a result of OA usage, HPS PF-4 levels were significantly reduced in samples obtained from OA-treated subjects. Neither OA administration nor DM appeared to significantly reduce the ability of HPP or HPS to induce microvessel formations in vitro. These findings indicate that OA administration does not limit the angiogenic potential of hypoxia preconditioned blood-derived secretomes, and therefore, it does not prohibit the application of these therapies for supporting tissue vascularization and wound healing in healthy or diabetic subjects.

Flow cytometry for near-patient testing in premature neonates reveals variation in platelet function: a novel approach to guide platelet transfusion.

BACKGROUND: Neonatal haemorrhaging is often co-observed with thrombocytopenia; however, no evidence of a causal relationship with low platelet count has been reported. Regardless, the administration of a platelet transfusion is often based upon this parameter. Accurate measurement of platelet function in small volumes of adult blood samples by flow cytometry is well established and we propose that the use of the same technology could provide complementary information to guide the administration of platelet transfusions in premature neonates. METHODS: In 28 neonates born at 27-41 weeks gestation, platelet function after stimulation agonists was measured using fibrinogen binding and P-selectin expression (a marker of degranulation). RESULTS: Platelets of neonates with gestation of ≤36 weeks (n = 20) showed reduced fibrinogen binding and degranulation with ADP, and reduced degranulation with CRP-XL. Degranulation Scores of 7837 ± 5548, 22,408 ± 5301 and 53,131 ± 12,102 (mean ± SEM) identified significant differences between three groups: <29, 29-36 and >36 weeks gestation). Fibrinogen binding and degranulation responses to ADP were significantly reduced in suspected septic neonates (n = 6) and the Fibrinogen Binding scores clearly separated the septic and healthy group (88.2 ± 10.3 vs 38.6 ± 12.2, P = 0.03). CONCLUSIONS: Flow cytometric measurement of platelet function identified clinically different neonatal groups and may eventually contribute to assessment of neonates requiring platelet transfusion.

Remission of congenital hyperinsulinism following conservative treatment: an exploratory study in patients with KATP channel mutations.

BACKGROUND: During conservative treatment, congenital hyperinsulinism (CHI) can resolve spontaneously. This study describes the hormonal and metabolic profiles in three patients with ABCC8/KCNJ11 mutations in clinical remission. METHODS: An age-adapted fasting and oral glucose tolerance test (OGTT) were performed. RESULTS: All patients (aged 6-9 years) tolerated age-adapted fasting durations (20, respectively 24 h), without reaching glucose concentrations ≤2.5 mmol/L, nor developing hypoglycemia-related symptoms. Nevertheless, insulin concentrations from all patients exceeded the 90th reference percentile at the end of the fasting test (range: 4.2-15.8 mU/L). During the OGTT, one patient (patient 2; BMI: 23.4 kg/m2; age: 7 years) reached a glucose concentration of 11.4 mmol/L after 2 h (concomitant insulin concentration: 148.3 mU/L). CONCLUSIONS: The insulin concentration profiles in CHI patients in apparent clinical remission range from almost complete normalization to persistent, yet attenuated, hypersecretion. The hyperglycemia, detected during the OGTT, must be further monitored.

Long-term medical treatment in congenital hyperinsulinism: a descriptive analysis in a large cohort of patients from different clinical centers.

BACKGROUND: Up to now, only limited data on long-term medical treatment in congenital hyperinsulinism (CHI) is available. Moreover, most of the drugs used in CHI are therefore not approved. We aimed to assemble more objective information on medical treatment in CHI with regard to type and duration, dosage as well as side effects. METHODS: We searched MEDLINE (from 1947) and EMBASE (from 1988) using the OVID interface for relevant data to evaluate medical treatment in a large cohort of patients with CHI from different clinical centers. Randomized, controlled trials were not available. We evaluated case reports and case series. No language restrictions were made. RESULTS: A total number of 619 patients were medically treated and information regarding conservative treatment was available. Drugs used were diazoxide (in 84% of patients), somatostatin analogues (16%), calcium channel antagonists (4%) and glucagon (1%). Mean dose of diazoxide was 12.5 (±4.3) mg/kg ⋅ d (range 2-60 mg/kg ⋅ d), mean duration of diazoxide treatment until remission was 57 months. Side effects of diazoxide were usually not severe. The causal relation between diazoxide and severe side effects, e.g. heart failure (3.7%) remains doubtful. Mean dose of octreotide was 14.9 (±7.5) µg/kg ⋅ d (range 2.3-50 µg/kg ⋅ d), of lanreotide 67.3 (±39.8) mg ⋅ month (range 10-120 mg ⋅ month). Mean duration of treatment with somatostatin analogues until remission was 49 months. Frequent side effects included tachyphylaxis and mild gastrointestinal symptoms. The risk of persistent growth deceleration was low (<5%). CONCLUSIONS: Severe side effects are rare and a causal relation remains disputable. We conclude that long-term conservative treatment of CHI is feasible.

The Fibrin Matrix Regulates Angiogenic Responses within the Hemostatic Microenvironment through Biochemical Control.

Conceptually, premature initiation of post-wound angiogenesis could interfere with hemostasis, as it relies on fibrinolysis. The mechanisms facilitating orchestration of these events remain poorly understood, however, likely due to limitations in discerning the individual contribution of cells and extracellular matrix. Here, we designed an in vitro Hemostatic-Components-Model (HCM) to investigate the role of the fibrin matrix as protein factor-carrier, independent of its cell-scaffold function. After characterizing the proteomic profile of HCM-harvested matrix releasates, we demonstrate that the key pro-/anti-angiogenic factors, VEGF and PF4, are differentially bound by the matrix. Changing matrix fibrin mass consequently alters the balance of releasate factor concentrations, with differential effects on basic endothelial cell (EC) behaviors. While increasing mass, and releasate VEGF levels, promoted EC chemotactic migration, it progressively inhibited tube formation, a response that was dependent on PF4. These results indicate that the clot's matrix component initially serves as biochemical anti-angiogenic barrier, suggesting that post-hemostatic angiogenesis follows fibrinolysis-mediated angiogenic disinhibition. Beyond their significance towards understanding the spatiotemporal regulation of wound healing, our findings could inform the study of other pathophysiological processes in which coagulation and angiogenesis are prominent features, such as cardiovascular and malignant disease.

The relation between awareness of personal resources and metabolic control in children and adolescents with type 1 diabetes.

INTRODUCTION: The study aims to elucidate whether awareness of personal resources, such as positive attributions and beliefs or social support, affects metabolic control in children and adolescents with type 1 diabetes. In addition, it will be determined to what extent metabolic control is influenced by concordance between children and parents regarding awareness of resources and the parents' ability to adopt their children's perspective. Also, the children's wishes particularly in relation to their illness will be investigated, as well as the kind of advice they would offer to fellow patients. METHODS: Seventy-eight children/adolescents with type 1 diabetes completed the Essen Resource Inventory for Children and Adolescents including personal, social, structural, and migration-specific resources. In addition, children/adolescents and their parents completed a systemic-oriented, diabetes-specific resource questionnaire in order to explore the parents' ability to adopt their children's perspective. RESULTS: Resources such as body awareness and open-minded attitude to the disease were associated with metabolic control. Particularly, resources associated to a migration background were found to be inversely correlated with hemoglobin A1c (HbA1c) value. Moreover, it was shown that the parents' ability to adopt their children's perspective was associated with improved metabolic control. Children advising fellow patients to accept the disease showed the best HbA1c value. DISCUSSION: This data identified specific modifiable factors related to metabolic control that can be addressed during counseling of pediatric patients. Also the parents' ability for adopting their child's perspective was identified as a relevant factor which should be considered during clinical counseling of young type 1 diabetes patients.

Health behaviour in children and adolescents with type 1 diabetes compared to a representative reference population.

OBJECTIVE: We provide a population-based overview of health behaviours of children and adolescents with type 1 diabetes in comparison to the general population, and analyse their relevance for glycaemic control and self-rated health status. METHODS: Data from questionnaires of 11- to 17-year-old children and adolescents with diabetes (n = 629) were compared to a representative sample (n = 6,813). RESULTS: Children and adolescents with type 1 diabetes had a significantly increased odds of infrequent physical activity (adjusted OR 1.56), short overall duration of physical activity per week (OR 1.55, difference -1.3 hours/week), and high daily computer use (OR 2.51). They had a lower odds of active and passive smoking (OR 0.31 and OR 0.29), and high daily television time (OR 0.68). The odds of an at least good and excellent self-rated health status was increased with intense physical activity, and decreased with active smoking and prolonged daily use of computer and television. Active smoking and prolonged daily use of computer were associated with higher HbA1c. CONCLUSIONS: Children and adolescents with type 1 diabetes showed a different profile of health behaviour. Their overall health may improve if their education stresses specifically frequent physical activity with longer overall duration and less frequent television or computer use.

Carbohydrate intake in relation to BMI, HbA1c and lipid profile in children and adolescents with type 1 diabetes.

BACKGROUND & AIMS: To compare reported and recommended carbohydrate intake in children and adolescents with type 1 diabetes (T1D) and to explore associations with BMI, HbA1c and lipid profile. METHODS: A cross-sectional observational study of reported carbohydrate intake in 46,010 patients with T1D aged 1-18 years from 332 diabetes centres in Germany and Austria in comparison to age-matched healthy children and adolescents. RESULTS: The median reported carbohydrate intake in T1D patients was markedly lower than in healthy children. It varied between 56% and 90% of recommended amounts across the paediatric age range with younger patients showing levels closer to recommend. Lower carbohydrate intake was associated with higher BMI-SDS (p < 0.001), particularly during adolescence, higher total cholesterol (p < 0.001), higher LDL-cholesterol (p = 0.005) and lower HbA1c (p < 0.001). CONCLUSIONS: The methodologically simple measure of reported carbohydrate intake may be a valuable addition to the information gathered on paediatric patients with T1D in an outpatient setting. Children and adolescents with T1D appear to restrict their consumption of carbohydrates, which may have adverse effects on BMI and the lipid profile, particularly if there is a compensatory increased fat intake. Health care providers should therefore advise patients and parents of the recommended age-dependent levels of carbohydrate intake.

Insulin sensitivity assessed by stable isotopes with oral glucose administration: validation with euglycaemic clamp.

Methods of determining insulin sensitivity that use an oral challenge of glucose are preferred to those using intravenous administration since the measurement is made in conditions more akin to normal physiology. One previously reported protocol (ODILE) studies glucose uptake in isolation from absorption and endogenous production by the intravenous administration of tracer approximately forty-five minutes after the oral dose is given. However, this methodology has not been validated against other accredited procedures. This study utilizes the euglycemic hyperinsulinemic clamp in order to validate the ODILE method.

Baseline IGF-I levels determine insulin secretion and insulin sensitivity during the first year on growth hormone therapy in children born small for gestational age. Results from a North European Multicentre Study (NESGAS).

OBJECTIVE: Developmental programming alters growth and metabolic outcome in children born small for gestational age (SGA). We explored insulin and glucose metabolism in SGA children treated with a fixed GH dose over 1 year. METHODS: In the North European Small for Gestational Age Study (NESGAS), 110 short SGA children received GH at 67 µg/kg/day for 1 year. Insulin secretion was assessed by acute insulin response (AIR), insulin sensitivity (IS) by HOMA and disposition index (DI) by insulin secretion adjusted for IS. RESULTS: First-year GH therapy led to increases in height and IGF-I standard deviation score (SDS), and reductions in IS (p < 0.0001). Compensatory increases in AIR (p < 0.0001) were insufficient and resulted in reduced DI (p = 0.032). Children in the highest IGF-I SDS tertile at baseline were the least insulin sensitive at baseline (p = 0.024) and 1 year (p = 0.006). IGF-I responses after 1 year were positively related to AIR (r = 0.30, p = 0.007) and DI (r = 0.29, p = 0.005). CONCLUSION: In SGA children treated with a high GH dose for 1 year, baseline IGF-I levels were related to IS whilst gains in height and IGF-I responses were associated with insulin secretion. Defining heterogeneity in IGF-I in SGA children may be useful in predicting growth and metabolic response.

Lipoatrophy is associated with an increased risk of Hashimoto's thyroiditis and coeliac disease in female patients with type 1 diabetes.

BACKGROUND/AIMS: Lipoatrophy (LA) is a rare, possibly under-recognised side effect of insulin treatment of unclear aetiology. The aim of this study was to describe the characteristics of patients with type 1 diabetes (T1D) who have LA and to explore the relationship between LA and other autoimmune diseases based on the hypothesis that additional autoimmune phenomena are more prevalent in T1D patients with LA. METHODS: This was a cross-sectional observational study of T1D patients with LA in comparison to T1D patients without LA who are registered with the Diabetes Patienten-Verlaufsdokumentationssystem database of 241,650 patients in Germany and Austria. RESULTS: Hashimoto's thyroiditis and coeliac disease were more prevalent in patients with LA (p < 0.001 for both). LA was associated with an increased risk of Hashimoto's thyroiditis and coeliac disease in female patients [odds ratio (OR) 2.5, p = 0.003, and OR 3.1, p = 0.02, respectively]. This relationship persisted after adjustment for current age, duration of diabetes and calendar year of treatment (OR 2.7, p = 0.002, and OR 3.5, p = 0.01, respectively). CONCLUSION: These findings support the hypothesis that an immune complex-mediated inflammatory process may be important in the development of LA.

Higher fasting plasma free fatty acid levels are associated with lower insulin secretion in children and adults and a higher incidence of type 2 diabetes.

CONTEXT: There are limited data in humans on the association between fasting free fatty acid (FFA) levels and pancreatic ß-cell function. OBJECTIVE: Our objective was to examine this association in children and adults with normal glucose tolerance and to explore fasting FFA levels in relation to subsequent risk of impaired glucose tolerance (IGT) and type 2 diabetes (T2D). DESIGN: We measured FFA, glucose, and insulin levels after an overnight fast and 30 min after an oral glucose load in 797 children aged 8 yr in the Avon Longitudinal Study of Parents and Children and 770 adults aged 44-71 yr in the Medical Research Council Ely Study. We calculated the homeostasis model assessment to estimate fasting insulin sensitivity, the insulinogenic index to estimate insulin secretion, and the disposition index to assess insulin secretion corrected for insulin sensitivity. RESULTS: Higher fasting FFA levels were associated with lower insulin secretion in children (boys, P = 0.03; girls, P = 0.001) and adults (men, P = 0.03, women, P = 0.04). Associations with insulin sensitivity were more variable, but after adjustment for insulin sensitivity, higher fasting FFA levels remained associated with lower insulin secretion (disposition index). Compared with adults in the lowest tertile of fasting FFA levels, those in the middle and highest tertiles had a 3-fold higher incidence of IGT or T2D over the following 5-8 yr. CONCLUSIONS: Higher fasting FFA levels were consistently associated with lower insulin secretion in children and adults with normal glucose tolerance. Furthermore, higher fasting FFA levels were prospectively associated with a greater risk of subsequent IGT and T2D.

Non-enhanced ECG-gated respiratory-triggered 3-D steady-state free-precession MR angiography with slab-selective inversion: initial experience in visualisation of renal arteries in free-breathing children without renal artery abnormality.

BACKGROUND: ECG-gated non-enhanced balanced steady-state free precession (bSSFP) MR angiography requires neither breath-holding nor administration of contrast material. OBJECTIVE: To investigate the image quality of free-breathing ECG-gated non-enhanced bSSFP MR angiography of renal arteries in children. MATERIALS AND METHODS: Fourteen boys and seven girls (mean age, 9.7 years; range, 7 weeks-17 years) with no history of renovascular disease were included. MRI was performed at 1.5 T. Subjective image quality of axial and coronal maximum-intensity-projection reconstructions of four segments (I, aorta and renal artery ostium; II, main renal artery; III, segmental branches; IV, intrarenal vessels) was evaluated using a 4-point scale (4 = excellent, 3 = good, 2 = acceptable, 1 = non-diagnostic). RESULTS: Image quality was excellent for segments I (mean ± SD, 3.9 ± 0.3) and II (4.0 ± 0.1), good for segment III (3.4 ± 0.9) and acceptable for segment IV (2.3 ± 1.1 ). Mean image quality did not differ between sedated and non-sedated children. CONCLUSION: bSSFP MR angiography enables visualisation of renal arteries in children.

Effects of growth hormone and free fatty acids on insulin sensitivity in patients with type 1 diabetes.

CONTEXT: Because GH stimulates lipolysis, an increase in circulating free fatty acid levels, as opposed to a direct effect of high GH levels, could underlie the development of insulin resistance in type 1 diabetes (T1D). Our aim was to explore the relative contributions of GH and free fatty acids to the development of insulin resistance in patients with T1D. PATIENTS: Seven (four females, three males) nonobese patients with T1D aged 21-30 yr were studied on four occasions in random order. On each visit, overnight endogenous GH production was suppressed by octreotide. Three 1-h pulses of recombinant human GH (rhGH) or placebo were administered on two visits each. Acipimox, an antilipolytic drug, or a placebo were ingested every 4 h on two visits each. Stable glucose and glycerol isotopes were used to assess glucose and glycerol turnover. The overnight protocol was concluded by a two-step hyperinsulinemic euglycemic clamp on each visit. MAIN OUTCOME: rhGH administration led to increases in the insulin infusion rate required to maintain euglycemia overnight (P = 0.008), elevated basal endogenous glucose production (P = 0.007), decreased basal peripheral glucose uptake (P = 0.03), and reduced glucose uptake during step 1 of the clamp (P < 0.0001). Coadministration of rhGH and acipimox reversed these effects and suppression of lipolysis in the absence of GH replacement led to further increases in insulin sensitivity. RESULTS: GH pulses were associated with an increase in endogenous glucose production and decreased rates of peripheral glucose uptake, which was entirely reversed by acipimox. Therefore, GH-driven decreases in insulin sensitivity are mainly determined by the effect of GH on lipolysis.

Intramyocellular lipid levels are associated with peripheral, but not hepatic, insulin sensitivity in normal healthy subjects.

Increased levels of IMCL (intramyocellular lipid) have been shown to be associated with reduced steady-state glucose infusion rates during a hyperinsulinaemic-euglycaemic clamp (M-value). The aim of the present study was to explore how IMCL levels relate to the insulin-mediated suppression of endogenous glucose production [hepatic SI (insulin sensitivity)] and increase in glucose disposal (peripheral SI). In the present study, 11 healthy young adults (7 male, 4 female; aged 21-31 years) undertook, in random order, an hyperinsulinaemic-euglycaemic clamp combined with stable glucose isotope enrichment to measure peripheral and hepatic SI, a 1H-MRS (proton-magnetic resonance spectroscopy) scan to determine IMCL levels and a DXA (dual-energy X-ray absorptiometry) scan to assess body composition. IMCL levels (range, 3.2-10.7) were associated with whole-body fat mass (r=0.787, P=0.004), fat mass corrected for height (r=0.822, P=0.002) and percentage of central fat mass (r=0.694, P=0.02), but were not related to whole-body FFM (fat-free mass; r=-0.472, P=0.1). IMCL levels correlated closely with the M-value (r=-0.727, P=0.01) and FFM-corrected peripheral SI (r=-0.675, P=0.02), but were not related to hepatic SI adjusted for body weight (r=0.08, P=0.8). The results of the present study suggest that IMCL accumulation may be a sensitive marker for attenuations in peripheral, but not hepatic, SI in normal populations. Given the close relationship of IMCL levels to whole-body and central abdominal fat mass, relative increases in the flux of lipids from adipose tissue to the intramyocellular compartment may be an integral part of the mechanisms underlying reductions in SI.

Insulin administration and rate of glucose appearance in people with type 1 diabetes.

OBJECTIVE: To assess whether prandial insulin, in addition to basal insulin, has an effect on the rate of glucose appearance from a meal in people with type 1 diabetes. RESEARCH DESIGN AND METHODS: The rate of glucose appearance from a mixed meal (Ra(meal)) was investigated in six adult (aged 24 +/- 2 years), lean (BMI 23.6 +/- 1.5 kg/m(2)) subjects with well-controlled type 1 diabetes (duration 7.9 +/- 6.9 years, A1C 7.6 +/- 0.9%) with/without prandial insulin. Actrapid was infused to maintain euglycemia before meals were consumed. Subjects consumed two identical meals on separate occasions, and Ra(meal) was measured using a dual isotope method. [6,6-(2)H(2)]glucose was incorporated into the meal (0.081 g/kg body wt), and a primed constant/variable rate infusion of [1,2,3,4,5,6,6-(2)H(2)]glucose was administered. In the tests with prandial insulin, an additional bolus dose of Actrapid was given 20 min before the meal at 0.1 units/kg body wt. RESULTS: Insulin concentration with prandial insulin was significantly higher than during basal insulin studies (119 +/- 16 vs. 66 +/- 15 pmol/l, P = 0.03 by paired t test). Despite differences in insulin concentration, there were no differences in total glucose appearance (3,398 +/- 197 vs. 3,307 +/- 343 micromol/kg) or time taken for 25% (33.1 +/- 3.3 vs. 31.7 +/- 3.5 min), 50% (54.6 +/- 3.5 vs. 54.1 +/- 4.7 min), and 75% (82.9 +/- 7.1 vs. 82.8 +/- 5.8 min) of total glucose appearance. The fraction of the glucose dose appearing in the circulation was the same for basal (73 +/- 8%) and prandial (75 +/- 4%) study days. CONCLUSIONS: These results suggest that meal glucose appearance is independent of prandial insulin concentration in people with type 1 diabetes.

Insulin resistance is an intrinsic defect independent of fat mass in women with Turner's syndrome.

BACKGROUND/AIMS: Turner's syndrome (TS) is associated with increased insulin resistance and adiposity, which might be associated with type 2 diabetes in later life. We aimed to determine whether the defect in insulin sensitivity is a primary intrinsic defect in TS or dependent on variation in body composition. METHODS: Sixteen women with TS not on growth hormone replacement but receiving oestrogen replacement therapy [age (mean +/- SD): 30.2 +/- 8.5 years; height-corrected fat-free mass: 26.1 +/- 3.1 kg/height] and a control group of 16 normal healthy women (age: 30.1 +/- 8.2 years; height-corrected fat-free mass: 25.9 +/- 2.4 kg/height) were studied. Fasting blood samples were obtained for measurement of glucose, insulin, IGF-I, IGFBP-1, IGFBP-3 and lipid levels. The hyperinsulinaemic euglycaemic clamp was performed to assess peripheral insulin sensitivity (M value), and the Homeostasis Model Assessment (HOMA-S) was used to estimate fasting insulin sensitivity. Body composition was assessed using a dual-energy X-ray absorptiometry scan. RESULTS: Fasting insulin sensitivity (HOMA-S 103.2 +/- 78.6 vs. 193.9 +/- 93.5, p = 0.006) was lower in TS subjects compared to controls as was whole-body insulin sensitivity (M value 2.9 +/- 1.9 vs. 5.5 +/- 2.6 mg/kg/min, p = 0.003). In a multiple regression analysis the Turner karyotype was significantly related to insulin sensitivity (p = 0.008) independent of any differences in fat-free mass and percent whole-body fat mass. CONCLUSION: The increased insulin resistance in women with TS is independent of measures of body composition and may represent an intrinsic defect related to their chromosomal abnormality.