Metformin improves blood glucose by increasing incretins independent of changes in gluconeogenesis in youth with type 2 diabetes.

AIMS/HYPOTHESIS: Metformin is the only approved oral agent for youth with type 2 diabetes but its mechanism of action remains controversial. Recent data in adults suggest a primary role for the enteroinsular pathway, but there are no data in youth, in whom metformin efficacy is only ~50%. Our objectives were to compare incretin concentrations and rates of glucose production and gluconeogenesis in youth with type 2 diabetes before and after short-term metformin therapy compared with peers with normal glucose tolerance (NGT). METHODS: This is a case-control observational study in youth with type 2 diabetes who were not on metformin (n = 18) compared with youth with NGT (n = 10) who were evaluated with a 2 day protocol. A 75 g OGTT was administered to measure intact glucagon-like 1 peptide (iGLP-1), gastric inhibitory polypeptide (GIP) and peptide YY (PYY). Insulinogenic index (IGI) and whole-body insulin sensitivity were calculated using glucose and insulin levels from the OGTT. Basal rates of gluconeogenesis (2H2O), glucose production ([6,6-2H2]glucose) and whole-body lipolysis ([2H5]glycerol) were measured after an overnight fast on study day 2. Youth with type 2 diabetes (n = 9) were subsequently evaluated with an identical 2 day protocol after 3 months on the metformin study. RESULTS: Compared with individuals with NGT, those with type 2 diabetes had higher fasting (7.8 ± 2.5 vs 5.1 ± 0.3 mmol/l, mean ± SD p = 0.002) and 2 h glucose concentrations (13.8 ± 4.5 vs 5.9 ± 0.9 mmol/l, p = 0.001), higher rates of absolute gluconeogenesis (10.0 ± 1.7 vs 7.2 ± 1.1 µmol [kg fat-free mass (FFM)]-1 min-1, p < 0.001) and whole-body lipolysis (5.2 ± 0.9 vs 4.0 ± 1.4 µmol kgFFM-1 min-1, p < 0.01), but lower fasting iGLP-1 concentrations (0.5 ± 0.5 vs 1.3 ± 0.7 pmol/l, p < 0.01). Metformin decreased 2 h glucose (pre metformin 11.4 ± 2.8 vs post metformin 9.9 ± 1.9 mmol/l, p = 0.04) and was associated with ~20-50% increase in IGI (median [25th-75th percentile] pre 1.39 [0.89-1.47] vs post 1.43 [0.88-2.70], p = 0.04), fasting iGLP-1 (pre 0.3 ± 0.2 vs post 1.0 ± 0.7 pmol/l, p = 0.02), 2 h iGLP (pre 0.4 ± 0.2 vs post 1.2 ± 0.9 pmol/l, p = 0.06), fasting PYY (pre 6.3 ± 2.2 vs post 10.5 ± 4.3 pmol/l, p < 0.01) and 2 h PYY (pre 6.6 ± 2.9 vs post 9.0 ± 4.0 pmol/l, p < 0.01). There was no change in BMI, insulin sensitivity or GIP concentrations pre vs post metformin. There were no differences pre vs post metformin in rates of glucose production (15.0 ± 3.9 vs 14.9 ± 2.2 µmol kgFFM-1 min-1, p = 0.84), absolute gluconeogenesis (9.9 ± 1.8 vs 9.7 ± 1.7 µmol kgFFM-1 min-1, p = 0.76) or whole-body lipolysis (5.0 ± 0.7 vs 5.3 ± 1.3 µmol kgFFM-1 min-1, p = 0.20). Post metformin iGLP-1 and PYY concentrations in youth with type 2 diabetes were comparable to levels in youth with NGT. CONCLUSIONS/INTERPRETATION: Overall, the improved postprandial blood glucose levels and increase in incretins observed in the absence of changes in insulin sensitivity and gluconeogenesis, support an enteroinsular mechanistic pathway in youth with type 2 diabetes treated with short-term metformin.

Free Vitamin D: Relationship to Insulin Sensitivity and Vascular Health in Youth.

OBJECTIVE: To evaluate the relationship of free 25 hydroxy vitamin D [free 25(OH)D] or bioavailable vitamin D (BioD) concentrations to insulin sensitivity and cardiovascular disease risk markers in normal weight and overweight youth. STUDY DESIGN: Cross-sectional study of 79 adolescents 15.4 ± 0.2 years, 18 normal weight, 30 overweight, and 31 overweight with prediabetes who underwent peripheral arterial tonometry, dual-energy x-ray absorptiometry, and hyperinsulinemic-euglycemic clamp in subset (n = 71) for determination of reactive hyperemia index (RHI), body composition, and insulin sensitivity. 25(OH)D and vitamin D binding protein were measured; free 25(OH)D and BioD were calculated. RESULTS: Across tertiles of free 25(OH)D concentrations (4.0 ± 0.2, 7.5 ± 0.3, and 17.0 ± 2.1 pg/mL, P < .001), the group in the lowest tertile had significantly higher percent body fat (37.8 ± 1.1, 35.2 ± 1.5 and 25.3 ± 2.1%, P < .001), lower insulin sensitivity (4.4 ± 0.4, 6.7 ± 1.2, and 8.2 ± 0.9 mg/kg fat-free mass/minute per µu/mL, P = .03), lower RHI (1.42 ± 0.06, 1.54 ± 0.06, and 1.77 ± 0.09, P = .002), higher high-sensitivity C-reactive protein (3.4 ± 0.6, 1.7 ± 0.3, and 1.6 ± 0.4 mg/L, P = .015) compared with the second and third tertiles, respectively. Free 25(OH)D levels were inversely related to percent body fat and high-sensitivity C-reactive protein, and positively related to RHI and insulin sensitivity. The relationships of free 25(OH)D to RHI and to insulin sensitivity were no longer significant after adjusting for %body fat. Similar relationships were observed for BioD. CONCLUSIONS: Youth with low free 25(OH)D or BioD concentrations have lower insulin sensitivity and worse endothelial function and inflammatory biomarkers compared with those with more sufficient 25(OH)D. However, the effects of vitamin D on these biomarkers may not be independent of the effect of adiposity.

Candy twists as an alternative to the glucola beverage in gestational diabetes mellitus screening.

OBJECTIVE: Screening for gestational diabetes mellitus commonly uses an oral glucose challenge test with a 50-g glucola beverage and subsequent venous puncture. However, up to 30% of pregnant women report significant side-effects, and the beverage is costly. We hypothesized that equivalent glucose loads could be achieved from a popular candy twist (Twizzlers; The Hershey Company, Hershey, PA) and tested it as cost-effective, tolerable alternative with a test of equivalency. STUDY DESIGN: The glucose equivalent of the 50-g glucola was calculated as 10 candy twists. We initially used a triple crossover design in nonpregnant patients whereby each subject served as her own control; this ensured the safety and equivalency of this load before using it among pregnant subjects. We then recruited pregnant women with an abnormal screening at 1 hour (glucose challenge test) in a double crossover design study. Subjects consumed 10 candy twists with a 1-hour venous blood glucose assessment. All subjects subsequently completed the confirmatory 3-hour glucose tolerance test. Sensitivity, specificity, positive predictive values, negative predictive values, false-referral rates, and detection rates were calculated. RESULTS: At ≥130 mg/dL, the sensitivity (100%) was the same for candy twists and glucola. However, the false-referral rate (82% vs 90%), positive predictive value (18% vs 10%), and detection rate (18% vs 10%) were improved for candy twists when compared with the 50-g glucola beverage. CONCLUSION: Our results indicate that strawberry-flavored candy twists are potentially an equally effective screening test, compared with the gold standard glucola beverage but lead to fewer false-positive screens and therefore could be a cost-effective alternative.

Distinct Amino Acid Profile Characterizes Youth With or at Risk for Type 2 Diabetes.

Branched-chain amino acids (BCAAs) and aromatic AAs (AAAs) are associated with increased risk for type 2 diabetes in adults. Studies in youth show conflicting results. We hypothesized that an AA metabolomic signature can be defined to identify youth at risk for ß-cell failure and the development of type 2 diabetes. We performed targeted AA metabolomics analysis on 127 adolescents (65 girls; 15.5 [SD ±1.9] years old, Tanner stage II-V) with normal weight or obesity across the spectrum of glycemia, with assessment of AA concentrations by mass spectrometry, at fasting, and steady state of a hyperinsulinemic-euglycemic clamp, with determination of insulin sensitivity (IS) per fat-free mass (FFM). We measured insulin secretion during a 2-h hyperglycemic clamp and calculated the disposition index per FFM (DIFFM), a measure of ß-cell function. Our results showed that concentration of glycine (Gly) and the glutamine (Gln)-to-glutamate (Glu) ratio were lower, whereas BCAA, tyrosine, and lysine (Lys) concentrations were higher in the groups with obesity and dysglycemia compared with those with normal weight. Gly and Gln-to-Glu ratio were positively related to IS and DIFFM, with opposite relationships observed for BCAAs, AAAs, and Lys. We conclude that a metabolic signature of low Gly concentration and low Gln-to-Glu ratio, and elevated BCAAs, AAAs, and Lys concentrations may constitute a biomarker to identify youth at risk for ß-cell failure.

Westerlund and Narayan predictability test: Step-by-step approach using COVID-19 and oil price data.

In this note, we provide a step-by-step approach of Westerlund and Narayan (WN, 2012, 2015) predictability test using COVID-19 and oil price data. This is an important exercise because the WN model addresses three salient features of time series data, namely persistency, endogeneity and heteroskedasticity. We consider COVID-19 and oil price data as predictors of stock market returns for four Asian countries to demonstrate the applicability of the WN (2012, 2015) predictability approach.•This note demonstrates a step-by-step approach of the WN (2012, 2015) predictability test.•WN model accommodates three salient features of time-series data, namely persistency, endogeneity, and heteroskedasticity.•COVID-19 and oil price does not significantly predict stock returns of Japan, Russia, and Singapore (except in the case of South Korea).

Metabolic flexibility across the spectrum of glycemic regulation in youth.

BACKGROUNDMetabolic flexibility (MF) refers to the relative ability to utilize lipid and carbohydrate substrates and to transition between them. It is not clear whether MF is impaired in obese youth and what the determining factors are.METHODSWe investigated the determinants of MF (increased respiratory exchange ratio [ΔRER] under insulin-stimulated conditions) in pubertal youth (n = 104; 15.6 ± 1.8 years) with obesity across the spectrum of glucose tolerance compared with normal weight (NW) controls, including body composition (fat-free mass [FFM], %body fat), visceral adipose fat (VAT) (MRI), glycemia, and insulin sensitivity (IS) [3-hour hyperinsulinemic-euglycemic clamp with measurement of lipolysis ([2H5] glycerol), free fatty acids (FFAs), and RER (indirect calorimetry)].RESULTSYouth with prediabetes and type 2 diabetes had lower ΔRER and oxidative and nonoxidative glucose disposal compared with NW, with no significant difference in ΔRER between NW and obese with normal glucose tolerance. In multiple regression analysis, ISFFM (ß = 0.4, P = 0.004), percentage suppression of FFAs (r = 0.26, P = 0.007), and race/ethnicity (ß = -0.23, P = 0.02) contributed to the variance in ΔRER (R2 = 0.30, P < 0.001) independent of percentage body fat (or VAT), sex, Tanner stage, and hemoglobin A1c.ConclusionMF is defective at the extreme of the metabolic phenotype in obese youth with dysglycemia related to a defect in IS limiting substrate utilization.FUNDINGUSDA/ARS Project Number 3092-51000-057.

Measurement of gluconeogenesis using glucose fragments and mass spectrometry after ingestion of deuterium oxide.

We report a new method to measure the fraction of glucose derived from gluconeogenesis using gas chromatography-mass spectrometry and positive chemical ionization. After ingestion of deuterium oxide by subjects, glucose derived from gluconeogenesis is labeled with deuterium. Our calculations of gluconeogenesis are based on measurements of the average enrichment of deuterium on carbon 1, 3, 4, 5, and 6 of glucose and the deuterium enrichment in body water. In a sample from an adult volunteer after ingestion of deuterium oxide, fractional gluconeogenesis using the "average deuterium enrichment method" was 48.3 +/- 0.5% (mean +/- SD) and that with the C-5 hexamethylenetetramine (HMT) method by Landau et al. (Landau BR, Wahren J, Chandramouli V, Schumann WC, Ekberg K, Kalhan SC; J Clin Invest 98: 378-385, 1996) was 46.9 +/- 5.4%. The coefficient of variation of 10 replicate analyses using the new method was 1.0% compared with 11.5% for the C-5 HMT method. In samples derived from an infant receiving total parenteral nutrition, fractional gluconeogenesis was 13.3 +/- 0.3% using the new method and 13.7 +/- 0.8% using the C-5 HMT method. Fractional gluconeogenesis measured in six adult volunteers after 66 h of continuous fasting was 83.7 +/- 2.3% using the new method and 84.2 +/- 5.0% using the C-5 HMT method. In conclusion, the average deuterium enrichment method is simple, highly reproducible, and cost effective. Furthermore, it requires only small blood sample volumes. With the use of an additional tracer, glucose rate of appearance can also be measured during the same analysis. Thus the new method makes measurements of gluconeogenesis available and affordable to large numbers of investigators under conditions of low and high fractional gluconeogenesis ( approximately 10 to approximately 90) in all subject populations.

Drug pricing for a novel treatment for wet macular degeneration: using incremental cost-effectiveness ratios to ensure societal value.

OBJECTIVE: Health economic models can assist policy-makers in determining the value of novel treatments from the viewpoint of society. In this context, value is defined as the benefit of treatment, given its cost. A new treatment for wet age-related macular degeneration (AMD), juxtascleral administration of anecortave acetate, 15 mg for depot suspension (Retaane), is now in a late-phase clinical trial. In a theoretical analysis, we sought to determine the cost at which this treatment might offer economic value to society, using incremental cost-effectiveness ratios (ICERs). METHODS: A series of 1-year cost-utility models was created for the investigational treatment and standard treatment (photodynamic therapy [PDT] with verteporfin [Visudyne]). Value to society was defined in terms of theoretical associated ICERs (in US dollars): $100,000 per quality-adjusted life-year (QALY), $50,000/QALY, $20,000/QALY and $0/QALY, the point of economic indifference. Models were created from the societal perspective and included a patient-derived utility assessment involving regression equations to estimate time trade-off preferences, event probabilities derived from a randomized clinical trial comparing the safety and efficacy of anecortave administration and PDT with verteporfin, decision analysis and relevant costing information. RESULTS: An ICER of $100,000/QALY would be associated with an anecortave cost of $3022/vial, an ICER of $50,000/QALY with an anecortave cost of $2986/vial and an ICER of $20,000/QALY with an anecortave cost of $2964/vial. The point of economic indifference between anecortave administration and standard therapy would occur with an anecortave cost of $2950/vial. INTERPRETATION: In theory, an anecortave cost of $2986/vial is associated with an ICER of $50,000/QALY, the threshold used by many health technology assessment and reimbursement agencies.

Dielectrically Addressable Microspheres Engineered Using Self-Assembled Monolayers.

We have used self-assembled monolayer techniques to produce a new class of microspheres with specifically engineered dielectric properties to enable their dielectrophoretic manipulation and identification in microsystems. Dielectrophoresis is an electrokinetic phenomenon that exploits frequency-dependent polarizability differences between a particle and its suspending medium to drive the movement of the particle toward or away from the high-field regions of an inhomogeneous electric field. While dielectrophoretic methods have been used extensively for cell manipulation, separation, and identification, we wished to extend the applicability of dielectrophoresis to molecular analysis by developing a panel of dielectric microspheres or "handles". Dielectric shell theory was used to model the dielectrophoretic response for a biomimetic particle composed of a thin insulating shell over a conductive interior. We specifically sought to modulate the specific capacitance, and thereby the dielectric properties, of the particle by controlling the thickness of the insulating layer. Such a structure was fabricated by covering a gold-coated polystyrene core particle with self-assembled monolayers of alkanethiol and phospholipid. To test the prediction that the carbon chain length of these layers should dictate the dielectric properties of the particles, we constructed a panel of six microsphere types with shell compositions ranging from a C(9) alkanethiol monolayer to a C(32) hybrid bilayer membrane. These microsphere populations were distinguishable and manipulatable by dielectrophoresis in a characteristic, frequency-dependent manner as predicted by theory. Experimentally derived specific membrane capacitance values were inversely related to the insulating shell thickness and agreed with published capacitance values for planar layers of similar thicknesses. These proof of principle studies are the first to demonstrate that the dielectric properties of particles can be specifically engineered to allow their dielectrophoretic manipulation and are a first step toward the development of bead-based dielectrophoretic microsystems for multiplexed molecular separation and analysis.

Functional reassembly of the Escherichia coli maltose transporter following purification of a MalF-MalG subassembly.

Taking advantage of a chaperone-like function of MalK, a stable complex of MalF-MalG could be solubilized from the Escherichia coli membrane and purified in high yield in the absence of MalK. This MalF-MalG complex was competent for efficient reassembly of a functional MalFGK(2) maltose transporter complex both in detergent solution and in proteoliposomes.