Deficiency of dolichol-phosphate-mannose synthase-1 causes congenital disorder of glycosylation type Ie

Congenital disorders of glycosylation (CDG), formerly known as carbohydrate-deficient glycoprotein syndromes, lead to diseases with variable clinical pictures. We report the delineation of a novel type of CDG identified in 2 children presenting with severe developmental delay, seizures, and dysmorphic features. We detected hypoglycosylation on serum transferrin and cerebrospinal fluid beta-trace protein. Lipid-linked oligosaccharides in the endoplasmic reticulum of patient fibroblasts showed an accumulation of the dolichyl pyrophosphate Man(5)GlcNAc(2) structure, compatible with the reduced dolichol-phosphate-mannose synthase (DolP-Man synthase) activity detected in these patients. Accordingly, 2 mutant alleles of the DolP-Man synthase DPM1 gene, 1 with a 274C>G transversion, the other with a 628delC deletion, were detected in both siblings. Complementation analysis using DPM1-null murine Thy1-deficient cells confirmed the detrimental effect of both mutations on the enzymatic activity. Furthermore, mannose supplementation failed to improve the glycosylation status of DPM1-deficient fibroblast cells, thus precluding a possible therapeutic application of mannose in the patients. Because DPM1 deficiency, like other subtypes of CDG-I, impairs the assembly of N-glycans, this novel glycosylation defect was named CDG-Ie.     

Diagnosis of congenital disorders of glycosylation by capillary zone electrophoresis of serum transferrin

BACKGROUND: Congenital disorders of glycosylation (CDG) are usually diagnosed by isoelectric focusing (IEF) of serum transferrin (Tf). The aim of this study was to evaluate capillary zone electrophoresis (CZE) as a diagnostic alternative to IEF. METHODS: We performed 792 CZE analyses of Tf, using the CEofix(TM)-CDT (carbohydrate-deficient transferrin) assay. Peak identification was based on relative migration times (RMTs) to reduce migration variability. RESULTS: Tf profiles comprised three main groups (A-C). Groups A and B were characterized by one or two dominant tetrasialo-Tf peaks, whereas group C showed a widely variable Tf isoform composition. Group A was composed of four subgroups: a major group with a typical Tf profile (considered as reference group), two minor groups with decreased or moderately increased trisialo-Tf isoform, and a group showing the presence of unknown compounds with RMTs similar to mono- and disialo-Tf. However, these compounds were absent on IEF. Group C contained all profiles from patients with confirmed as well as putative CDG. From the reference group, 99% confidence intervals were calculated for the RMTs of the Tf isoforms, and percentiles representing the Tf isoform distributions were defined. CONCLUSIONS: All patients with abnormal IEF results and confirmed CDG were identified by CZE; thus, this method can be used as a diagnostic alternative to IEF in a manner suitable for automation. Because whole serum is analyzed, it should be kept in mind that CZE profiles can show substances other than Tf.     

Aldose reductase genotypes and cardiorenal complications: an 8-year prospective analysis of 1,074 type 2 diabetic patients

OBJECTIVE—We report the independent risk association of type 2 diabetic nephropathy with the z−2 allele of the 5′-(CA)_n microsatellite and C-106T promoter polymorphisms of the aldose reductase gene (ALR2) using a case-control design. In this expanded cohort, we examined their predictive roles on new onset of cardiorenal complications using a prospective design.RESEARCH DESIGN AND METHODS—In this 8-year prospective cohort of 1,074 type 2 diabetic patients (59% male, median age 61 years; disease duration 7 years) with an observation period of 8,592 person-years, none had clinical evidence of coronary heart disease (CHD) or chronic kidney disease at recruitment. The renal end point was defined as new onset of estimated glomerular filtration rate <60 ml/min per 1.72 m² or hospitalizations with dialysis or death due to renal disease, and CHD was defined as hospitalizations with myocardial infarction, ischemic heart disease, or related deaths.RESULTS—After controlling for baseline risk factors and use of medications, we found that the ALR2 z−2 allele of (CA)_n microsatellite carriers had increased risk of renal (hazard ratio 1.53 [95% CI 1.14–2.05], P = 0.005) or combined cardiorenal (1.31 [1.01–1.72], P = 0.047) end points. Carriers of the ALR2 C-106T polymorphism also had increased risk of renal (1.54 [1.15–2.07], P = 0.004) and cardiorenal (1.49 [1.14–1.95], P = 0.004) end points. Compared with noncarriers, patients with two risk-conferring genotypes had a twofold increased risk of renal (2.41 [1.57–3.70], P < 0.001) and cardiorenal (1.94 [1.29–2.91], P = 0.002) end points.CONCLUSIONS—In Chinese type 2 diabetic patients, genetic polymorphisms of ALR2 independently predicted new onset of renal and cardiorenal end points, with the latter being largely mediated through renal disease.Cardiovascular diseases and end-stage renal disease (ESRD) are the main causes of mortality and morbidity in diabetic patients. The onset of chronic kidney disease (CKD) markedly increases the risk of cardiovascular disease as a result of further perturbation in the metabolic milieu and vascular homeostasis (1). In type 2 diabetes, control of hyperglycemia reduces proteinuria and the rate of decline in renal function (2). In type 1 diabetes, established renal lesions were reversed with restoration of normoglycemia after pancreatic transplantation (3). Aldose reductase (ALR2) converts glucose to sorbitol, especially under hyperglycemic condition. Accumulation of sorbitol can lead to increased oxidative and osmotic stresses, causing cataract, microvascular complications, and myocardial ischemic injury (4).In a case-control cohort consisting of 92 Chinese type 2 diabetic patients without nephropathy despite long disease duration and 121 patients with both retinopathy and nephropathy, the z−2 allele of 5′-(CA)_n microsatellite and promoter C-106T polymorphisms of the ALR2 gene independently conferred a risk of diabetic nephropathy (5). In this expanded consecutive cohort of 1,074 type 2 diabetic patients with documentation of risk factors, complications, and clinical outcomes, we examined the independent and additive effects of these two polymorphisms of the ALR2 gene on cardiorenal end points after an 8-year observational period.     

Two-year pulmonary safety and efficacy of inhaled human insulin (Exubera) in adult patients with type 2 diabetes

OBJECTIVE—The purpose of this study was to evaluate the 2-year pulmonary safety of inhaled human insulin (Exubera [EXU]) in 635 nonsmoking adults with type 2 diabetes.RESEARCH DESIGN AND METHODS—Patients were randomly assigned to receive prandial EXU or subcutaneous insulin (regular or short-acting) plus basal (intermediate- or long-acting) insulin. The primary end points were the annual rate of decline in forced expiratory volume in 1 s (FEV₁) and carbon monoxide diffusing capacity (DL_CO).RESULTS—Small differences in FEV₁ favoring subcutaneous insulin developed during the first 3 months but did not progress. Adjusted treatment group differences in FEV₁ annual rate of change were −0.007 l/year (90% CI −0.021 to 0.006) between months 0 and 24 and 0.000 l/year (−0.016 to 0.016) during months 3–24. Treatment group differences in DL_CO annual rate of change were not significant. Both groups sustained similar reductions in A1C by month 24 (last observation carried forward) (EXU 7.7–7.3% vs. subcutaneous insulin 7.8–7.3%). Reductions in fasting plasma glucose (FPG) were greater with EXU than with subcutaneous insulin (adjusted mean treatment difference −12.4 mg/dl [90% CI −19.7 to −5.0]). Incidence of hypoglycemia was comparable in both groups. Weight increased less with EXU than with subcutaneous insulin (−1.3 kg [−1.9 to −0.7]). Adverse events were comparable, except for a higher incidence of mild cough and dyspnea with EXU.CONCLUSIONS—Two-year prandial EXU therapy showed a small nonprogressive difference in FEV₁ and comparable sustained A1C improvement but lower FPG levels and less weight gain than seen in association with subcutaneous insulin in adults with type 2 diabetes.Inhaled human insulin (Exubera [EXU], insulin human [rDNA origin] inhalation powder) is effective in patients with type 2 diabetes in whom glycemic control is not achieved with diet and exercise (1) and provides better glycemic control in patients whose diabetes remains inadequately controlled with either monoagent or combination oral agent regimens (2–5). In addition, 6-month EXU therapy is at least comparable in efficacy to subcutaneous insulin therapy in type 2 diabetes (6).Previous controlled studies of EXU in type 2 diabetic patients have shown slight but consistent treatment group differences in pulmonary function in favor of comparators, using nonstandardized lung function testing (3–6). These small differences occurred early, did not progress for up to 2 years, and were not clinically meaningful. However, these studies were not designed specifically to examine respiratory safety primary end points.The aim of the present study was to provide a robust evaluation of the long-term pulmonary safety of EXU versus subcutaneous insulin. Adult patients with type 2 diabetes were evaluated by means of validated, highly standardized pulmonary function testing, trained coordinators, and centralized data collection (7–10). A secondary objective was to evaluate the long-term efficacy of EXU versus subcutaneous insulin.     

Screening using serum percentage of carbohydrate-deficient transferrin for congenital disorders of glycosylation in children with suspected metabolic disease

BACKGROUND: Diagnoses of congenital disorders of glycosylation (CDG) are based on clinical suspicion and analysis of transferrin (Tf) isoforms. Here we present our experience of CDG screening in children with a suspected metabolic disease by determination of serum percentage of carbohydrate-deficient transferrin (%CDT) in tandem with isoelectric focusing (IEF) analysis of Tf and alpha(1)-antitrypsin (alpha(1)-AT). METHODS: We performed approximately 8000 serum %CDT determinations using %CDT turbidimetric immunoassay (TIA). In selected samples, IEF analysis of Tf and alpha(1)-AT was carried out on an agarose gel (pH 4-8) using an electrophoresis unit. The isoforms were detected by Western blotting and visualized by color development. We performed neuraminidase digestion of serum to detect polymorphic variants of Tf. RESULTS: We established a cutoff value for serum %CDT of 2.5% in our pediatric population. Sixty-five patients showed consistently high values of serum %CDT. In accordance with Tf and alpha(1)-AT IEF profiles, enzyme assays, and mutation analysis, we made the following diagnoses: 23 CDG-Ia, 1 CDG-Ib, and 1 conserved oligomeric Golgi 1 (COG-1) deficiency. In addition, we identified 13 CDG-Ix non Ia, non-Ib; 3 CDG-Ix; and 9 CDG-IIx cases, albeit requiring further characterization; 9 patients with a secondary cause of hypoglycosylation and 6 with a polymorphic Tf variant were also detected. CONCLUSION: The combined use of CDT immunoassay with IEF of Tf and alpha(1)-AT is a useful 1st-line screening tool for identifying CDG patients with an N-glycosylation defect. Additional molecular investigations must of course be carried out to determine the specific genetic disease.     

Relationship between risk factors and mortality in type 1 diabetic patients in Europe: the EURODIAB Prospective Complications Study (PCS)

OBJECTIVE—The purpose of this study was to examine risk factors for mortality in patients with type 1 diabetes.RESEARCH DESIGN AND METHODS—Baseline risk factors were measured in the EURODIAB Prospective Cohort Study with 2,787 type 1 diabetic patients (51% men and 49% women) recruited from 16 European countries. Mortality data were collected during a 7-year follow-up.RESULTS—There was an annual mortality rate of 5 per 1,000 person-years in patients with type 1 diabetes (mean age at baseline 33 years, range 15–61 years); of the total 2,787 subjects, 102 died. The final multivariable model contained age at baseline (standardized hazard ratio 1.78 [95% CI 1.44–2.20]), A1C (1.18 [0.95–1.46]), waist-to-hip ratio (WHR) (1.32 [1.14–1.52]), pulse pressure (1.33 [1.13–1.58]), and non-HDL cholesterol (1.33 [1.12–1.60]) as risk factors for all-cause mortality. Macroalbuminuria (2.39 [1.19–4.78]) and peripheral (1.88 [1.06–3.35]) and autonomic neuropathy (2.40 [1.32–4.36]) were the most important risk markers for mortality. Similar risk factors were found for all-cause, non-cardiovascular disease (CVD), unknown-cause, and CVD mortality.CONCLUSIONS—Important risk factors for the increased total and non-CVD mortality in type 1 diabetic patients are age, WHR, pulse pressure, and non-HDL cholesterol. Microvascular complications from macroalbuminuria and peripheral and autonomic neuropathy are strong risk markers for future mortality exceeding the effect of the traditional risk factors.The presence of type 1 diabetes is associated with a three- to fourfold increased risk of mortality compared with that of the general population (1,2). It is still not clear what risk factors explain this excess mortality risk. We have shown that risk factors associated with insulin resistance, such as triglyceride, waist-to-hip ratio (WHR), and albuminuria strongly predict cardiovascular disease (CVD) in type 1 diabetes (3). Although CVD is the major cause of death in patients with type 1 diabetes, it only accounts for approximately half of all deaths, and it is therefore important to study the totality of risk and non-CVD causes of mortality in this young population that is particularly vulnerable to premature death. Previous studies have often had too few deaths (4) or have not collected key risk factors at baseline (1,2). The few large cohort studies that have studied this question have produced inconsistent findings (4–6), in part because key common risk factors were not always included. Therefore, the aim of this study was to examine risk factors for all-cause mortality in a large, 7-year prospective cohort study of patients with type 1 diabetes.     

Dietary patterns and risk of incident type 2 diabetes in the Multi-Ethnic Study of Atherosclerosis (MESA)

OBJECTIVE—We characterized dietary patterns and their relation to incident type 2 diabetes in 5,011 participants from the Multi-Ethnic Study of Atherosclerosis (MESA).RESEARCH DESIGN AND METHODS—White, black, Hispanic, and Chinese adults, aged 45–84 years and free of cardiovascular disease and diabetes, completed food frequency questionnaires at baseline (2000–2002). Incident type 2 diabetes was defined at three follow-up exams (2002–2003, 2004–2005, and 2005–2007) as fasting glucose >126 mg/dl, self-reported type 2 diabetes, or use of diabetes medication. Two types of dietary patterns were studied: four empirically derived (principal components analysis) and one author-defined (low-risk food pattern) as the weighted sum of whole grains, vegetables, nuts/seeds, low-fat dairy, coffee (positively weighted), red meat, processed meat, high-fat dairy, and soda (negatively weighted).RESULTS—The empirically derived dietary pattern characterized by high intake of tomatoes, beans, refined grains, high-fat dairy, and red meat was associated with an 18% greater risk (hazard ratio per 1-score SD 1.18 [95% CI 1.06–1.32]; P_trend = 0.004), whereas the empirically derived dietary pattern characterized by high intake of whole grains, fruit, nuts/seeds, green leafy vegetables, and low-fat dairy was associated with a 15% lower diabetes risk (0.85 [0.76–0.95]; P_trend = 0.005). The low-risk food pattern was also inversely associated with diabetes risk (0.87 [0.81–0.99]; P_trend = 0.04). Individual component food groups were not independently associated with diabetes risk. Associations were not modified by sex or race/ethnicity.CONCLUSIONS—Multiple food groups collectively influence type 2 diabetes risk beyond that of the individual food groups themselves.Type 2 diabetes and obesity have reached epidemic proportions in the U.S. and the world. In addition to the role diet plays in preventing obesity and, consequently, type 2 diabetes, diet may also reduce risk of type 2 diabetes independent of changes in body weight. Whole grains (1), nuts/seeds (2), coffee (3), low-fat dairy (4,5), and vegetables (6,7) have been inversely associated with incident type 2 diabetes or related metabolic traits independent of differences in body weight or other measures of adiposity, whereas sugar-sweetened beverages (8,9), red meat (10), processed meats (11), and white potatoes (fried or baked/boiled) (12) have been positively associated.In practice, each nutrient or food is part of a larger pattern consisting of many nutrients and foods, and, thus, characterization of multiple, concurrent dietary exposures has particular relevance to health. Using data-driven techniques, such as principal components analysis (PCA), several epidemiological studies have evaluated associations between dietary patterns and type 2 diabetes (10,11,13–15). Generally, studies show that dietary patterns characterized by high whole grain, fruit/vegetable, and low-fat dairy intake are inversely associated with type 2 diabetes risk. Analogously, dietary patterns characterized by high intake of red or processed meats, refined grains, fried foods, and foods containing high amounts of added sugars are associated with greater type 2 diabetes risk. Studies have been conducted in relatively homogenous populations (predominantly white cohorts) (10,11,13–15). Validation of these findings in racially/ethnically diverse samples is needed.Empirical methods such as PCA do not necessarily maximize the disease-predictive value of each dietary pattern; rather, such methods maximize the amount of variation in dietary intake explained by each dietary pattern. Variation in dietary intake comprises dietary behaviors, taste, and convenience. For this reason, combinations of foods other than those identified by PCA might be more predictive of incident disease. Studies have shown that individual food groups, such as those listed above, are independently associated with incident type 2 diabetes. However, the collective contribution of these foods to type 2 diabetes risk has not been characterized.Using data from the Multi-Ethnic Study of Atherosclerosis (MESA), we evaluated the relationship between type 2 diabetes risk and the following two trends: 1) PCA-derived dietary patterns and 2) a low-risk food pattern score based on the intake of foods previously associated with risk of type 2 diabetes (whole grains, vegetables, low-fat dairy foods, nuts/seeds, and coffee [positively weighted] and red meat, processed meat, high-fat dairy foods, white potatoes, and nondiet soda [negatively weighted]).     

Sprint training increases muscle oxidative metabolism during high-intensity exercise in patients with type 1 diabetes

OBJECTIVE—To investigate sprint-training effects on muscle metabolism during exercise in subjects with (type 1 diabetic group) and without (control group) type 1 diabetes.RESEARCH DESIGN AND METHODS—Eight subjects with type 1 diabetes and seven control subjects, matched for age, BMI, and maximum oxygen uptake (V̇o_2peak), undertook 7 weeks of sprint training. Pretraining, subjects cycled to exhaustion at 130% V̇o_2peak. Posttraining subjects performed an identical test. Vastus lateralis biopsies at rest and immediately after exercise were assayed for metabolites, high-energy phosphates, and enzymes. Arterialized venous blood drawn at rest and after exercise was analyzed for lactate and [H⁺]. Respiratory measures were obtained on separate days during identical tests and during submaximal tests before and after training.RESULTS—Pretraining, maximal resting activities of hexokinase, citrate synthase, and pyruvate dehydrogenase did not differ between groups. Muscle lactate accumulation with exercise was higher in type 1 diabetic than nondiabetic subjects and corresponded to indexes of glycemia (A1C, fasting plasma glucose); however, glycogenolytic and glycolytic rates were similar. Posttraining, at rest, hexokinase activity increased in type 1 diabetic subjects; in both groups, citrate synthase activity increased and pyruvate dehydrogenase activity decreased; during submaximal exercise, fat oxidation was higher; and during intense exercise, peak ventilation and carbon dioxide output, plasma lactate and [H⁺], muscle lactate, glycogenolytic and glycolytic rates, and ATP degradation were lower in both groups.CONCLUSIONS—High-intensity exercise training was well tolerated, reduced metabolic destabilization (of lactate, H⁺, glycogenolysis/glycolysis, and ATP) during intense exercise, and enhanced muscle oxidative metabolism in young adults with type 1 diabetes. The latter may have clinically important health benefits.Repeated bouts of brief, high-intensity exercise are characterized not only by marked metabolic and ionic destabilization in exercising muscle but also by progressively increasing aerobic ATP generation, such that by just the third 30-s bout, 63% of ATP is generated oxidatively (1). It is therefore not entirely surprising that high-intensity (sprint) exercise training results in oxidative adaptations in muscle. These adaptations include reduced glycogenolysis and lower accumulation of muscle lactate and hydrogen ions during high-intensity matched-work exercise (2); increased activity of oxidative enzymes such as citrate synthase (3–5), cytochrome c oxidase (6), and β-hydroxyacyl-CoA dehydrogenase (β-HAD) (4); reduced ATP degradation during intense exercise (2,5,7); and increased peak oxygen consumption (V̇o_2peak) (2–5,8).The only study (9) to examine muscle metabolism during exercise in patients with type 1 diabetes (compared with control subjects) reported lower muscle oxidative capacity and higher glycolytic flux and acidosis. No studies have directly investigated the longitudinal effects of exercise training on muscle metabolism in these patients. Thus, while young adults with type 1 diabetes undertake exercise and sports of various intensities, including high-intensity exercise and training, clinicians have little evidence upon which to provide advice. Based on adaptations in healthy subjects, we hypothesized that high-intensity exercise training in young adults with type 1 diabetes would increase muscle oxidative capacity, attenuate glycogenolytic rate, and reduce myocellular metabolic destabilization during high-intensity exercise.     

Burden and rates of treatment and control of cardiovascular disease risk factors in obesity: the Framingham Heart Study

OBJECTIVE— Obesity is associated with an increased risk for cardiovascular disease (CVD). We sought to determine rates of treatment and control of CVD risk factors among normal weight, overweight, and obese individuals in a community-based cohort.RESEARCH DESIGN AND METHODS— Participants free of CVD (n = 6,801; mean age 49 years; 54% women) from the Framingham Offspring and Third Generation cohorts who attended the seventh Offspring examination (1998–2001) or first Third Generation (2002–2005) examination were studied.RESULTS— Obese participants with hypertension were more likely to receive antihypertensive treatment (62.3%) than normal weight (58.7%) or overweight (59.0%) individuals (P = 0.002), but no differences in hypertension control across BMI subgroups among participants with hypertension were observed (36.7% [normal weight], 37.3% [overweight], and 39.4% [obese]; P = 0.48). Rates of lipid-lowering treatment were higher among obese participants with elevated LDL cholesterol (39.5%) compared with normal weight (34.2%) or overweight (36.4%) participants (P = 0.02), but control rates among those with elevated LDL cholesterol did not differ across BMI categories (26.7% [normal weight], 26.0% [overweight], and 29.2% [obese]; P = 0.11). There were no differences in diabetes treatment among participants with diabetes across BMI groups (69.2% [normal weight], 50.0% [overweight], 55.0% [obese]; P = 0.54), but obese participants with diabetes were less likely to have fasting blood glucose <126 mg/dl (15.7%) compared with normal weight (30.4%) or overweight (20.7%) participants (P = 0.02).CONCLUSIONS— These findings emphasize the suboptimal rates of treatment and control of CVD risk factors among overweight and obese individuals.Obesity affects more than one-third of the adult population in the U.S. Excess weight is associated with multiple cardiovascular disease (CVD) risk factors, including hypertension, dyslipidemia, diabetes, and the metabolic syndrome.Although the incidence and mortality of CVD have declined markedly during the past decades, some studies suggested that the increasing prevalence of obesity and diabetes may have slowed this rate of decline (1). In addition, recent data suggest that the prevalence of chronic kidney disease is increasing, in part because of the increasing rates of diabetes (2). Unfortunately, the efficacy of current therapies for obesity, including lifestyle and pharmacological interventions, is limited (3). Although bariatric surgery is an effective method of weight loss among severely obese individuals, eligibility criteria limit its use to only the most significantly affected patients.Given the current limitations of effective weight loss therapies, minimizing the risk of complications of obesity and diabetes due to CVD risk factors is essential. Few studies have focused on a comprehensive approach to CVD risk factor burden, treatment, and control among obese individuals. Therefore, the aim of this study was to examine the burden of CVD risk factors as well as rates of treatment and control among normal weight, overweight, and obese individuals in an unselected population-based cohort. As abdominal fat accumulation is strongly associated with metabolic and CVD risk factors, and as recent guidelines have emphasized the importance of measuring waist circumference as part of clinical cardiovascular risk assessment, we also studied individuals with and without abdominal obesity.     

Carbohydrate-deficient glycoprotein syndrome: not an N-linked oligosaccharide processing defect, but an abnormality in lipid-linked oligosaccharide biosynthesis?

The carbohydrate-deficient glycoprotein syndrome (CDGS) is a developmental disease associated with an abnormally high isoelectric point of serum transferrin. Carbohydrate analyses of this glycoprotein initially suggested a defect in N-linked oligosaccharide processing, although more recent studies indicate a defect in the attachment of these sugar chains to the protein. We studied both serum glycoproteins and fibroblast-derived [2-3H]mannose-labeled oligosaccharides from CDGS patients and normal controls. While there was a decrease in the glycosylation of serum glycoproteins of affected individuals, differences were not seen in either monosaccharide composition or oligosaccharide structures. The lectin-binding profiles of glycopeptides from [2-3H]-mannose-labeled fibroblasts were likewise indistinguishable. However, the incorporation of [2-3H]mannose into both glycoproteins and the dolichol-linked oligosaccharide precursor was significantly reduced. Thus, at least in some patients, CDGS is not due to a defect in processing of N-linked oligosaccharides, but rather to defective synthesis and transfer of nascent dolichol-linked oligosaccharide precursors. This abnormality could result in both a failure to glycosylate some sites on some proteins, as well as secondary abnormalities in overall glycoprotein processing and/or function.     

Novel Noninvasive Breath Test Method for Screening Individuals at Risk for Diabetes

OBJECTIVE—Diagnosis of pre-diabetes and early-stage diabetes occurs primarily by means of an oral glucose tolerance test (OGTT), which requires invasive blood sampling. The aim of this study was to determine whether differences exist in breath ¹³CO₂ excretion during a ¹³C-labeled OGTT between individuals with normal glucose tolerance (NGT) and individuals with pre-diabetes and early-stage diabetes (PDED) and whether these differences correlated with blood glucose kinetics.RESEARCH DESIGN AND METHODS—Blood and breath samples were collected at baseline and every 30 min for a 10-h period after ingestion of 75 g glucose isotopically labeled with 150 mg [U-¹³C₆]d-glucose.RESULTS—Age (56 ± 5 vs. 47 ± 3 years) and BMI (31 ± 2 vs. 31 ± 2 kg/m²) were not different between individuals with NGT (n = 10) and PDED (n = 7), respectively. Blood glucose concentrations were significantly higher in those with PDED compared with those with NGT from baseline to 4.5 h after glucose ingestion (P ≤ 0.05). Glucose-derived breath ¹³CO₂ was significantly lower in individuals with PDED compared with those with NGT from 1 to 3.5 h after glucose (P ≤ 0.05). Peak breath ¹³CO₂ abundance occurred at 4.5 and 3.5 h in individuals with PDED and NGT, respectively (36.87 ± 3.15 vs. 41.36 ± 1.56‰ delta over baseline).CONCLUSIONS—These results suggest that this novel breath test method may assist in recognition of pre-diabetes or early-stage diabetes in at-risk persons without the need for invasive blood sampling, thus making it an attractive option for large-scale testing of at-risk populations, such as children.Type 2 diabetes and its cluster of disorders (i.e., metabolic syndrome) have become a major public health concern (1,2) and form a predictive risk profile for cardiovascular disease (3–5). Recently, the risk of development of metabolic syndrome has significantly increased in the pediatric population (6). Traditionally, pre-diabetes and early-stage diabetes are assessed in at-risk adult and pediatric populations using a standard oral glucose tolerance test (OGTT) requiring repeated invasive blood sampling over the 2 to 3 h after a glucose load. Postprandially, the mix of fuels used for cellular energy production is dramatically altered when insulin sensitivity is decreased (7–9). The ability to noninvasively detect the shift in fuel source utilization between individuals with normal glucose tolerance (NGT) and those at risk for pre-diabetes and early-stage diabetes (PDED) has considerable clinical utility.It has been demonstrated previously that ¹³CO₂ in breath is as effective as the hyperinsulinemic-euglycemic clamp method for assessing insulin resistance (10). However, the clinical utility of detecting breath CO₂ kinetics using a standard diagnostic infrared ¹³CO₂ breath analyzer between individuals with NGT and those with PDED has not been clearly established. Therefore, the aim of the present study was to evaluate the efficacy of a diagnostic breath analyzer available in many physician offices to characterize the kinetics of glucose-derived CO₂ in breath after administration of a standard OGTT in individuals with NGT and PDED. We compared the breath CO₂ kinetics from individuals with NGT with those from individuals with PDED for 10 h after an OGTT (75-g glucose load) isotopically labeled with [U-¹³C₆]glucose. In addition, breath CO₂ data were correlated with indexes of insulin resistance (whole-body index of insulin sensitivity [WBISI], homeostasis model assessment of insulin resistance [HOMA-IR], and quantitative insulin sensitivity check index [QUICKI]) calculated from measures of blood glucose and plasma insulin. We hypothesized that the standard breath analyzer would be sufficiently sensitive to detect impaired glucose utilization for cellular respiration during a standard OGTT between those with NGT and PDED.     

Nitric oxide bioavailability and its potential relevance to the variation in susceptibility to the renal and vascular complications in patients with type 2 diabetes

OBJECTIVE—We compared the renal and systemic vascular (renovascular) response to a reduction of bioavailable nitric oxide (NO) in type 2 diabetic patients without nephropathy and of African and Caucasian heritage.RESEARCH DESIGN AND METHODS—Under euglycemic conditions, renal blood flow was determined by a constant infusion of paraminohippurate and changes in blood pressure and renal vascular resistance estimated before and after an infusion of l-Ng-monomethyl-l-arginine.RESULTS—In the African-heritage group, there was a significant fall in renal blood flow (Δ−46.0 ml/min per 1.73 m²; P < 0.05) and rise in systolic blood pressure (Δ10.0 mmHg [95% CI 2.3–17.9]; P = 0.017), which correlated with an increase in renal vascular resistance (r² = 0.77; P = 0.004).CONCLUSIONS—The renal vasoconstrictive response associated with NO synthase inhibition in this study may be of relevance to the observed vulnerability to renal injury in patients of African heritage.The bioavailability of nitric oxide (NO) is central to the regulation of renovascular function and is reduced in established hypertension and diabetic nephropathy (1–3). Studies in rodents suggest that a deficiency of NO is an important susceptibility factor in the development of diabetes-related renal injury (4). It is unknown whether the differences in vulnerability to renal injury in diabetic patients of African heritage (5) versus Caucasians is related to NO bioactivity.     

The Diabeo software enabling individualized insulin dose adjustments combined with telemedicine support improves HbA1c in poorly controlled type 1 diabetic patients: a 6-month, randomized, open-label, parallel-group, multicenter trial (TeleDiab 1 Study)

OBJECTIVE

To demonstrate that Diabeo software enabling individualized insulin dose adjustments combined with telemedicine support significantly improves HbA_1c in poorly controlled type 1 diabetic patients.

RESEARCH DESIGN AND METHODS

In a six-month open-label parallel-group, multicenter study, adult patients (n = 180) with type 1 diabetes (>1 year), on a basal-bolus insulin regimen (>6 months), with HbA_1c ≥8%, were randomized to usual quarterly follow-up (G1), home use of a smartphone recommending insulin doses with quarterly visits (G2), or use of the smartphone with short teleconsultations every 2 weeks but no visit until point end (G3).

RESULTS

Six-month mean HbA_1c in G3 (8.41 ± 1.04%) was lower than in G1 (9.10 ± 1.16%; P = 0.0019). G2 displayed intermediate results (8.63 ± 1.07%). The Diabeo system gave a 0.91% (0.60; 1.21) improvement in HbA_1c over controls and a 0.67% (0.35; 0.99) reduction when used without teleconsultation. There was no difference in the frequency of hypoglycemic episodes or in medical time spent for hospital or telephone consultations. However, patients in G1 and G2 spent nearly 5 h more than G3 patients attending hospital visits.

CONCLUSIONS

The Diabeo system gives a substantial improvement to metabolic control in chronic, poorly controlled type 1 diabetic patients without requiring more medical time and at a lower overall cost for the patient than usual care.HbA_1c remains unsatisfactory in many patients of type 1 diabetes with levels consistently above 8.0%, despite close monitoring and participation in educational programs, with the American Diabetes Association (ADA) recommended therapy for type 1 diabetes (basal and prandial analog insulin, either with multiple daily injections [MDI] or insulin pump) (1–3). Prandial insulin has to be adjusted according to carbohydrate intake, premeal blood glucose, and anticipated physical activity (4). The main reasons for these unsatisfactory results are: patient has difficulty in coping with the constraints of the disease; difficulty applying the complex determination of the required amount of prandial insulin, leading to incorrect insulin doses and thus to hypo- or, more often, hyperglycemia; finally, patients may find it difficult to comply with scheduled doctor visits since they need to take a day off for their medical visit. Another difficulty arises from the patient’s paper diary, currently the usual tool of data communication between patients and diabetologists. It is often perceived as a boring document and therefore may be poorly filled in. In such cases, the diabetologist has limited information to recommend the appropriate insulin dose. The blood glucose measurements stored in the memory of the patient’s meter are of little use without reliable information on the time and content of meals, physical activity, or insulin doses injected.We have created the Diabeo system to overcome some of these hurdles. Diabeo is a software uploaded onto smartphones with internet connection that provides to the patient 1) bolus calculators using validated algorithms, taking into account carbohydrate intake, premeal blood glucose, and anticipated physical activity reported by the patient; 2) plasma glucose targets; 3) automatic algorithms for the adjustment of carbohydrate ratio and basal insulin or pump basal rates when the postprandial or fasting plasma glucose levels are off target; 4) data transmission to medical staff computers, through General Packet Radio Service and secured websites, to allow easy telemonitoring and teleconsultations. One pilot single-center study has demonstrated the feasibility, safety and accuracy of Diabeo (5). The aim of this study was to evaluate the efficiency of the Diabeo system in improving metabolic control of chronic, poorly controlled type 1 diabetes.     

Dietary energy density predicts the risk of incident type 2 diabetes: the European Prospective Investigation of Cancer (EPIC)-Norfolk Study

OBJECTIVE—Accumulating evidence suggests that energy-dense foods predispose to obesity and that such foods may also be associated with an increased risk of type 2 diabetes, but there is limited evidence. Our aim was to investigate whether there is an independent association between dietary energy density and incidence of diabetes.RESEARCH DESIGN AND METHODS—The European Prospective Investigation of Cancer (EPIC)-Norfolk Cohort Study was a population-based prospective study of individuals aged 40–79 years at baseline. We calculated energy density for overall diet (all solids and drinks) using food frequency questionnaires. During 12 years of follow-up, we documented 725 new-onset cases of diabetes among 21,919 participants without diabetes, cancer, or cardiovascular disease at baseline.RESULTS—Baseline energy density (adjusted for age, sex, and baseline BMI) was higher in those who developed type 2 diabetes (mean 3.08 kJ/g [95% CI 3.03–3.13]) than in those who remained nondiabetic (3.01 kJ/g [3.00–3.02]) (P = 0.012). Energy density was positively associated with incident diabetes (odds ratio 1.21 per unit increase [95% CI 1.06–1.38]) adjusted for known risk factors. There was a 60% higher risk of diabetes (1.60 [1.19–2.16]) in the highest quintile of energy density (range 3.55–7.97 kJ/g) compared with the lowest quintile (1.04–2.43 kJ/g) in adjusted analysis.CONCLUSIONS—This is the first large population-based prospective study to report that an energy-dense diet may be associated with increased risk of development of diabetes, independent of baseline obesity. The potential public health impact of a low–energy-dense diet on reducing the risk of diabetes deserves further study.Dietary energy density (DED) is defined as the amount of energy able to be metabolized per unit weight or volume of food (1). Thus, assuming energy expenditure is held constant, high energy density of a given volume of food consumed would lead to increased energy intake and weight gain, as demonstrated by both short-term and long-term intervention trials (reviewed by Yao and Roberts [1]). Energy-dense foods were also reported to be associated with body fatness in children: for instance, at age 7 years a 1 unit (1 kJ/g) rise in DED increased the odds of excess adiposity at 9 years by 36% (odds ratio [OR] 1.36 [95% CI 1.09–1.69]) (2). Mendoza et al. (3) also found that DED was independently associated with elevated fasting insulin and the metabolic syndrome in a cross-sectional setting. It is plausible that such foods may also be associated with an increased risk of type 2 diabetes, but there is limited evidence. In the Finnish Diabetes Prevention Study (DPS), overweight men and women with impaired glucose tolerance receiving standard care or intensive dietary and exercise counseling showed a positive, but not significant, association between energy density and the risk of developing diabetes after 3 years of follow-up (hazard ratio 1.74 [95% CI 0.89–3.37]) (4). To the best of our knowledge, this is the only published study regarding DED and the risk of diabetes. Therefore, the purpose of our study was to investigate the association of DED with new-onset diabetes in a population-based cohort study including both men and women, appropriately adjusted for a comprehensive range of lifestyle factors, social factors, and dietary factors.     

Evolutionary conservation of human ketodeoxynonulosonic acid production is independent of sialoglycan biosynthesis

Human metabolic incorporation of nonhuman sialic acid (Sia) N-glycolylneuraminic acid into endogenous glycans generates inflammation via preexisting antibodies, which likely contributes to red meat–induced atherosclerosis acceleration. Exploring whether this mechanism affects atherosclerosis in end-stage renal disease (ESRD), we instead found serum accumulation of 2-keto-3-deoxy-d-glycero-d-galacto-2-nonulosonic acid (Kdn), a Sia prominently expressed in cold-blooded vertebrates. In patients with ESRD, levels of the Kdn precursor mannose also increased, but within a normal range. Mannose ingestion by healthy volunteers raised the levels of urinary mannose and Kdn. Kdn production pathways remained conserved in mammals but were diminished by an M42T substitution in a key biosynthetic enzyme, N-acetylneuraminate synthase. Remarkably, reversion to the ancestral methionine then occurred independently in 2 lineages, including humans. However, mammalian glycan databases contain no Kdn-glycans. We hypothesize that the potential toxicity of excess mannose in mammals is partly buffered by conversion to free Kdn. Thus, mammals probably conserve Kdn biosynthesis and modulate it in a lineage-specific manner, not for glycosylation, but to control physiological mannose intermediates and metabolites. However, human cells can be forced to express Kdn-glycans via genetic mutations enhancing Kdn utilization, or by transfection with fish enzymes producing cytidine monophosphate–Kdn (CMP-Kdn). Antibodies against Kdn-glycans occur in pooled human immunoglobulins. Pathological conditions that elevate Kdn levels could therefore result in antibody-mediated inflammatory pathologies.     

Weight change in diabetes and glycemic and blood pressure control

OBJECTIVE—Weight loss in type 2 diabetes is undisputedly important, and data from community settings are limited. We evaluated weight change and resulting glycemic and blood pressure control in type 2 diabetic patients at an HMO.RESEARCH DESIGN AND METHODS—Using electronic medical records, this retrospective cohort study identified 2,574 patients aged 21–75 years who received a new diagnosis of type 2 diabetes between 1997 and 2002. We estimated 3-year weight trajectories using growth curve analyses, grouped similar trajectories into four categories using cluster analysis, compared category characteristics, and predicted year-4 above-goal A1C and blood pressure by group.RESULTS—The weight-trajectory groups were defined as higher stable weight (n = 418; 16.2%), lower stable weight (n = 1,542; 59.9%), weight gain (n = 300; 11.7%), and weight loss (n = 314; 12.2%). The latter had a mean weight loss of 10.7 kg (−9.8%; P < 0.001) by 18 months, with near-complete regain by 36 months. After adjusting for age, sex, baseline control, and related medication use, those with higher stable weight, lower stable weight, or weight-gain patterns were more likely than those who lost weight to have above-goal A1C (odds ratio [OR] 1.66 [95% CI 1.12–2.47], 1.52 [1.08–2.14], and 1.77 [1.15–2.72], respectively). Those with higher stable weight or weight-gain patterns were more likely than those who lost weight to have above-goal blood pressure (1.83 [1.31–2.57] and 1.47 [1.03–2.10], respectively).CONCLUSIONS—A weight-loss pattern after new diagnosis of type 2 diabetes predicted improved glycemic and blood pressure control despite weight regain. The initial period postdiagnosis may be a critical time to apply weight-loss treatments to improve risk factor control.Almost all adults with diabetes are overweight; more than half are obese (1). Obesity is associated with worse blood glucose and other cardiovascular risk factor control (2). Results from the Look AHEAD trial show that weight loss in diabetes improves glycemic control, reduces blood pressure, and improves blood lipids (3). Observational studies also support a likely link between weight loss and reduced mortality in people with diabetes (2).Limited data describe the extent to which weight loss, as well as resulting levels of glycemic and blood pressure control, is achieved in community-living people with type 2 diabetes (4,5). Most weight information on these subjects comes from research volunteers (4,5). Prior studies of health effects of weight change have been plagued by confounding of low weight by disease burden and by difficulty separating intentional from unintentional weight loss (6,7).This study used electronic medical records data to evaluate weight trajectories in the initial years following a new type 2 diabetes diagnosis, associated demographic and comorbidity factors, and resulting glycemic and blood pressure control. The initial period after a diabetes diagnosis is of particular interest because this may be a time of heightened patient and clinician interest in patient behavior change (8).     

The use of lipid-linked oligosaccharides (neoglycolipids) in the identification of carbohydrate receptors for microbial pathogens

Specific oligosaccharide chains on the host cell surface act as receptors for many microbial pathogens. Identification of receptor structures is an important step in the understanding of the pathogenesis of infection. Glycolipid receptors have been identified by direct binding assays. However, technical difficulties have prevented demonstration of bacterial binding to the oligosaccharides of glycoproteins; these have been identified mainly by inhibition assays. By a novel technique developed in our laboratory, oligosaccharides released from glycoproteins are linked to lipids to form neoglycolipids. These can be used in bacterial binding assays. The feasibility of this approach has been demonstrated using type 1 fimbriated Escherichia coli binding specifically to neoglycolipids rich in mannose residues. The application of the method has resulted in a demonstration of a new type of adhesive specificity for E coli and differences in the binding specificities of E coli and Pseudomonas aeruginosa. Further application of this technique by generating oligosaccharides purified from mucus glycoproteins from patients with cystic fibrosis to use in binding assays with P aeruginosa is currently being undertaken. The basic knowledge gained by this approach may in time see the development of novel therapy in the form of receptor blocking agents.     

Association between p.Leu54Met polymorphism at the paraoxonase-1 gene and plantar fascia thickness in young subjects with type 1 diabetes

OBJECTIVE— In type 1 diabetes, plantar fascia, a collagen-rich tissue, is susceptible to glycation and oxidation. Paraoxonase-1 (PON1) is an HDL-bound antioxidant enzyme. PON1 polymorphisms have been associated with susceptibility to macro- and microvascular complications. We investigated the relationship between plantar fascia thickness (PFT) and PON1 gene variants, p.Leu54Met, p.Gln192Arg, and c.-107C>T, in type 1 diabetes.RESEARCH DESIGN AND METHODS—This was a cross-sectional study of 331 adolescents with type 1 diabetes (162 male and 169 female). PFT was assessed by ultrasound, PON1 was assessed by genotyping with PCR and restriction fragment–length polymorphism, and serum PON1 activity was assessed by rates of hydrolysis of paraoxon and phenylacetate.RESULTS—Median (interquartile range) age was 15.4 (13.5–17.3) years, and diabetes duration was 7.6 (4.9–10.6) years. The distribution of p.Leu54Met genotypes was LL 135 (40.8%), ML 149 (45%), and MM 47 (14.2%). PFT was abnormal (>1.7 mm) in 159 adolescents (48%). In multivariate analysis, predictors of abnormal PFT were ML/LL versus MM p.Leu54Met polymorphism (odds ratio 3.84 [95% CI 1.49–9.82], P = 0.005); BMI (percentile) (1.02 [1.01–1.03], P = 0.007); systolic blood pressure (percentile) (1.01 [1.00–1.02], P = 0.03); and male sex (3.29 [1.98–5.46], P < 0.001).CONCLUSIONS—Thickening of the plantar aponeurosis occurs predominantly in overweight and male adolescents with type 1 diabetes. The MM genotype at PON1 p.Leu54Met is associated with a reduced risk of abnormal PFT.Paraoxonase-1 (PON1) is a calcium-dependent HDL-associated enzyme that protects LDLs from oxidation. In type 1 diabetic patients, the serum paraoxonase concentration is lower and HDL is a less efficient antioxidant than in healthy individuals (1). Oxidized LDL is implicated in the pathogenesis of atherosclerosis, diabetic retinopathy, and nephropathy (2). Variations in lipoprotein-related enzymes and genotypes may also promote diabetic microvascular damage (3).Soft-tissue thickening is associated with chronic hyperglycemia and is hypothesized to be due to collagen glycation (4). With use of ultrasound techniques to measure plantar aponeurosis, a collagen-rich tissue, researchers demonstrated previously that people with diabetes have increased plantar fascia thickness (PFT) (5). Recently, this group reported that increased PFT predicted the development of microvascular complications in adolescents with type 1 diabetes and proposed abnormal PFT as a putative marker of soft-tissue glycation (6).The PON gene cluster maps to chromosome 7q21-22 and influences gene expression and serum activity. There is an established link between PON1 and macrovascular disease (7) and emerging evidence linking PON1 to microvascular complications (8,9). In this study we investigated whether the variants c.-107C>T at the promoter region and p.Leu54Met and p.Gln192Arg at the coding regions of PON1 are associated with PFT in type 1 diabetes.     

Analysis of microRNA processing machinery gene (DROSHA,DICER1, RAN,and XPO5) variants association with end-stage renal disease

BackgroundMicroRNA (miRNA) processing machinery gene variant was associated with several diseases. We aimed to explore for the first time the association of machinery gene (DROSHA rs10719A/G; DICER1 rs3742330A/G; RAN rs14035C/T; and XPO5 rs11077T/G) variants with the susceptibility and phenotype of end‐stage renal disease (ESRD).MethodA total of 281 participants (98 ESRD patients and 183 healthy volunteers) were enrolled. Real‐Time TaqMan allelic discrimination assay was applied for the genotyping of the specified variants. Multiple logistic regression models, univariate, multivariate, and principal component analyses were carried out.ResultsCarrying one DICER1 rs3742330*G allele conferred protection against developing ESRD [heterozygote comparison: OR = 0.30, 95% CI = 0.15‐0.62, dominant model: OR = 0.35, 95% CI = 0.17‐0.70]. Similarly, for XPO5 rs11077T/G, homozygote and heterozygote carriers of G variant were less likely to develop ESRD [homozygote comparison: adjusted OR = 0.23, 95% CI = 0.11‐0.50, and heterozygote comparison: OR = 0.50, 95% CI = 0.22‐0.92, and allelic model: OR = 0.46, 95% CI = 0.34‐0.70]. RAN rs14035*TT subjects were 5 times more likely to develop ESRD while being heterozygote (CT) have twice the risk [homozygote comparison: 5.18, 95% CI = 2.28‐11.8, heterozygote comparison: OR = 2.12, 95% CI = 1.10‐409]. Subgroup analysis also detected DICER1 rs3742330*A, XPO5 rs11077*T, and RAN rs14035*T as genetic risk determinants for ESRD development in both sex and age categories. Two genotype combinations of DROSHA/DICER1/XPO5/RAN were associated with increased susceptibility to ESRD [A‐A‐T‐T: OR = 9.49, 95%CI = 2.48‐36.31 (P = .001) and G‐A‐T‐T: OR = 5.92, 95%CI = 1.77‐19.83 (P = .004), respectively].ConclusionVariants and combined genotypes of DICER1 rs3742330, XPO5 rs11077, and RAN rs14035 might be associated with ESRD development in the study population. Integrating molecular analysis in ESRD risk stratification is warranted.     

Activities of HMR 3004 (RU 64004) and HMR 3647 (RU 66647) Compared to Those of Erythromycin, Azithromycin, Clarithromycin, Roxithromycin, and Eight Other Antimicrobial Agents against Unusual Aerobic and Anaerobic Human and Animal Bite Pathogens Isolated from Skin and Soft Tissue Infections in Humans

The activities of HMR 3004 and HMR 3647 and comparator agents, especially macrolides, were determined by the agar dilution method against 262 aerobic and 120 anaerobic strains isolated from skin and soft tissue infections associated with human and animal bite wounds. HMR 3004 and HMR 3647 were active against almost all aerobic and fastidious facultative isolates (MIC at which 90% of the isolates are inhibited [MIC₉₀], ≤0.5 and 1 μg/ml, respectively) and against all anaerobes [Bacteroides tectum, Porphyromonas macacae (salivosa), Prevotella heparinolytica, Porphyromonas sp., Prevotella sp., and peptostreptococci] at ≤0.25 and ≤0.5 μg/ml, respectively, except Fusobacterium nucleatum (HMR 3004, MIC₉₀ = 16 μg/ml; HMR 3647, MIC₉₀ = 8 μg/ml) and other Fusobacterium species (MIC₉₀, 1 and 2 μg/ml, respectively). In general, HMR 3004 and HMR 3647 were more active than any of the macrolides tested. Azithromycin was more active than clarithromycin against all Pasteurella species, including Pasteurella multocida subsp. multocida, Eikenella corrodens, and Fusobacterium species, while clarithromycin was more active than azithromycin against Corynebacterium species, Weeksella zoohelcum, B. tectum, and P. heparinolytica.