A Low ω-6 to ω-3 PUFA Ratio (n-6:n-3 PUFA) Diet to Treat Fatty Liver Disease in Obese Youth.

BACKGROUND: Recent literature suggests that the Western diet's imbalance between high ω-6 (n-6) and low ω-3 (n-3) PUFA intake contributes to fatty liver disease in obese youth. OBJECTIVES: We tested whether 12 wk of a low n-6:n-3 PUFA ratio (4:1) normocaloric diet mitigates fatty liver and whether the patatin-like containing domain phospholipase 3 (PNPLA3) rs738409 variant affects the response. METHODS: In a single-arm unblinded study, obese youth 9-19 y of age with nonalcoholic fatty liver disease were treated with a normocaloric low n-6:n-3 PUFA ratio diet for 12 wk. The primary outcome was change in hepatic fat fraction (HFF%), measured by abdominal MRI. Metabolic parameters included alanine aminotransferase (ALT), lipids, measures of insulin sensitivity, and plasma oxidized linoleic acid metabolites (OXLAMs). Outcomes were also analyzed by PNPLA3 rs738409 genotype. Wilcoxon's signed rank test, the Mann-Whitney U test, and covariance pattern modeling were used. RESULTS: Twenty obese adolescents (median age: 13.3 y; IQR: 10.5-16.4 y) were enrolled and 17 completed the study. After 12 wk of dietary intervention, HFF% decreased by 25.8% (P = 0.009) despite stable weight. We observed a 34.4% reduction in ALT (P = 0.001), 21.9% reduction in triglycerides (P = 0.046), 3.28% reduction in LDL cholesterol (P = 0.071), and a 26.3% improvement in whole body insulin sensitivity (P = 0.032). The OXLAMs 9-hydroxy-octadecandienoic acid (9-HODE) (P = 0.011), 13-HODE (P = 0.007), and 9-oxo-octadecadienoic acid (9-oxoODE) (P = 0.024) decreased after 12 wk. HFF% declined in both the not-at-risk (CC/CG) and at-risk (GG) PNPLA3 rs738409 genotype groups, with significant (P = 0.016) HFF% reduction in the GG group. Changes in 9-HODE (P = 0.023), 9-oxoODE (P = 0.009), and 13-oxoODE (P = 0.003) differed between the 2 genotype groups over time. CONCLUSIONS: These data suggest that, independently of weight loss, a low n-6:n-3 PUFA diet ameliorates the metabolic phenotype of adolescents with fatty liver disease and that response to this diet is modulated by the PNPLA3 rs738409 genotype.This trial was registered at clinicaltrials.gov as NCT01556113.

Intrahepatic fat, irrespective of ethnicity, is associated with reduced endogenous insulin clearance and hepatic insulin resistance in obese youths: A cross-sectional and longitudinal study from the Yale Pediatric NAFLD cohort.

AIM: To evaluate whether intrahepatic fat accumulation contributes to impaired insulin clearance and hepatic insulin resistance across different ethnic groups. METHODS: The intrahepatic fat content (HFF%) was quantified by magnetic resonance imaging in a multi-ethnic cohort of 632 obese youths aged 7-18 years at baseline and after a 2-year follow-up. Insulin secretion rate (ISR), endogenous insulin clearance (EIC) and hepatic insulin resistance index (HIRI) were estimated by modelling glucose, insulin and C-peptide data during 3-hour, 9-point oral glucose tolerance tests. RESULTS: African American youths exhibited the lowest HFF% and a prevalence of non-alcoholic fatty liver disease (NAFLD) less than half of that shown by Caucasians and Hispanics. Furthermore, African Americans had lower EIC and glucose-stimulated ISR, despite similar HIRI and plasma insulin levels, compared with Caucasians and Hispanics. EIC and HIRI were markedly reduced in individuals with NAFLD and declined across group-specific HFF% tertiles in all ethnic groups. Consistently, the HFF% correlated with EIC and HIRI, irrespective of the ethnic background, after adjustment for age, sex, ethnicity, adiposity, waist-hip ratio, pubertal status and plasma glucose levels. An increased HFF% at follow-up was associated with decreased EIC and increased HIRI across all groups. CONCLUSIONS: Intrahepatic lipid accumulation is associated with reduced insulin clearance and hepatic insulin sensitivity in obese youths, irrespective of their ethnic background.

A low visceral fat proportion, independent of total body fat mass, protects obese adolescent girls against fatty liver and glucose dysregulation: a longitudinal study.

BACKGROUND: The relative proportion of visceral fat (VAT) to subcutaneous fat (SAT) has been described as a major determinant of insulin resistance (IR). Our study sought to evaluate the effect of body fat distribution on glucose metabolism and intrahepatic fat content over time in a multiethnic cohort of obese adolescents. SUBJECTS/METHODS: We examined markers of glucose metabolism by oral glucose tolerance test, and body fat distribution by abdominal MRI at baseline and after 19.2 ± 11.4 months in a cohort of 151 obese adolescents (88 girls, 63 boys; mean age 13.3 ± 3.4 years; mean BMI z-score 2.15 ± 0.70). Hepatic fat content was assessed by fast-gradient MRI in a subset of 93 subjects. We used the median value of VAT/(VAT + SAT) ratio within each gender at baseline to stratify our sample into high and low ratio groups (median value 0.0972 in girls and 0.118 in boys). RESULTS: Female subjects tended to remain in their VAT/(VAT + SAT) category over time (change over follow-up P = 0.14 among girls, and P = 0.04 among boys). Baseline VAT/(VAT + SAT) strongly predicted the hepatic fat content, fasting insulin, 2-h glucose, and whole-body insulin sensitivity index at follow-up among girls, but not in boys. CONCLUSIONS: The VAT/(VAT + SAT) ratio is a major determinant of impaired glucose metabolism and hepatic fat accumulation over time, and its effects are more pronounced in girls than in boys.

Metabolic Features of Nonalcoholic Fatty Liver (NAFL) in Obese Adolescents: Findings From a Multiethnic Cohort.

We conducted a prospective study in a large, multiethnic cohort of obese adolescents to characterize clinical and genetic features associated with pediatric nonalcoholic fatty liver (NAFL), the most common cause of chronic liver disease in youth. A total of 503 obese adolescents were enrolled, including 191 (38.0%) whites, 134 (26.6%) blacks, and 178 (35.4%) Hispanics. Participants underwent abdominal magnetic resonance imaging (MRI) to quantify hepatic fat fraction (HFF), an oral glucose tolerance test (OGTT) to assess glucose tolerance and insulin sensitivity, and the genotyping of three single-nucleotide polymorphisms (SNPs) associated with nonalcoholic fatty liver disease (NAFLD) (patatin-like phospholipase domain-containing protein 3 [PNPLA3] rs738409, glucokinase regulatory protein [GCKR] rs1260326, and transmembrane 6 superfamily member 2 [TM6SF2] rs58542926). Assessments were repeated in 133 subjects after a 2-year follow-up. Prevalence of nonalcoholic fatty liver (NAFL) was 41.6% (209 patients) and ranged widely among ethnicities, being 42.9% in whites, 15.7% in blacks, and 59.6% in Hispanics (P < 0.0001). Among adolescents with NAFL, blacks showed the highest prevalence of altered glucose homeostasis (66%; P = 0.0003). Risk factors for NAFL incidence were white or Hispanic ethnicity (P = 0.021), high fasting C-peptide levels (P = 0.0006), and weight gain (P = 0.0006), whereas baseline HFF (P = 0.004) and weight loss (P = 0.032) predicted resolution of NAFL at follow-up. Adding either gene variant to these variables improved significantly the model predictive performance. CONCLUSION: Black obese adolescents are relatively protected from liver steatosis, but are more susceptible to the deleterious effects of NAFL on glucose metabolism. The combination of ethnicity/race with markers of insulin resistance and genetic factors might help identify obese youth at risk for developing NAFL.

The rs626283 Variant in the MBOAT7 Gene is Associated with Insulin Resistance and Fatty Liver in Caucasian Obese Youth.

OBJECTIVES: Non alcoholic fatty liver disease (NAFLD) is a leading cause of liver damage in childhood, its occurrence is influenced by genetic and environmental factors. Recently, the rs626283 polymorphism in the MBOAT7 gene has been found to be associated with alcoholic liver disease and NAFLD in adults. METHODS: In a multiethnic cohort of obese children and adolescents we genotyped the rs626283 polymorphism in the MBOAT7 gene, evaluated insulin sensitivity by an oral glucose tolerance test, and measured the intra-hepatic fat content (HFF%) by magnetic resonance imaging. RESULTS: In Caucasian youth, the minor allele (C) was associated with HFF% in (P=0.003), fasting insulin (P=0.03), area under the curve of glucose (P=0.03), and lower degree of whole-body insulin sensitivity (P=0.01) independent of age, gender, and body mass index z-score. A partial correlation showed that the association between the rs626283 variant and insulin resistance was driven by the presence of hepatic steatosis (P=0.009). However, there was no association between the rs626283 and hepatic steatosis among Hispanic and African American children and youth. The association between the rs626283 in the MBOAT7 gene among Caucasians was independent of the PNPLA3 rs738409, GCKR 1260326, and TM6SF2 rs58542926 (P=0.01). The four polymorphisms combined explained~19% of the HFF% in Caucasian obese children and adolescents. CONCLUSIONS: The rs626283 variant in the MBOAT7 gene is associated with NAFLD and may affect glucose metabolism by modulating intra-hepatic fat content in Caucasian obese children and adolescents.

Impact of Severe Obesity on Cardiovascular Risk Factors in Youth.

OBJECTIVE: To compare cardiovascular risk factor clustering (CVRFC) in severely obese youth with those with lower degrees of obesity. STUDY DESIGN: We divided a childhood obesity clinic derived cohort into the degrees of obesity (class I, II, and III) and added a "class IV" category corresponding to >160% of the 95th centile of body mass index (BMI) for age and sex. In a cross-sectional analysis, we investigated the presence of CVRFC in 2244 participants; in 621 who were followed longitudinally, we investigated the determinants of endpoint CVRFC. RESULTS: Class IV obesity was associated with increased risk for CVRFC compared with overweight (OR = 17.26, P < .001) at a similar magnitude to class III obesity (OR = 17.26, P < .001). Male children were at greater risk for presence of CVRFC (OR = 1.57, P = .03) compared with female children. Adiponectin (OR = 0.90, P < .001) and leptin levels (OR = 0.98, P = .008) were protective, independent of degree of obesity. Baseline class IV obesity was associated with increased risk compared with overweight of having CVRFC at follow-up (OR = 5.76, P = .001), to a similar extent as class III obesity (OR = 5.36, P = .001). Changes in the degree of obesity were significant predictors of CVRFC on follow-up (OR = 1.04, P < .01 per percent BMI change). CONCLUSIONS: The metabolic risk associated with obesity in childhood is conferred prior to reaching class IV obesity. An individualized risk stratification approach in children with severe obesity should be based on presence of complications rather than simple BMI cutoffs. TRIAL REGISTRATION: ClinicalTrials.gov NCT01967849.

Role of TM6SF2 rs58542926 in the pathogenesis of nonalcoholic pediatric fatty liver disease: A multiethnic study.

UNLABELLED: We assessed the association between the single-nucleotide polymorphism (SNP) rs58542926 in the transmembrane 6 superfamily member 2 (TM6SF2) gene and fatty liver disease in obese youth. We genotyped the TM6SF2 rs58542926 SNP in a multiethnic cohort of 957 obese children and adolescents (42% Caucasians, 28% African Americans, 30% Hispanics). All underwent an oral glucose tolerance test, a liver panel, and a lipid profile. Of them, 454 children underwent a magnetic resonance imaging study to assess hepatic fat content and 11 underwent liver biopsy to assess the degree of disease severity. The minor allele of the rs58542926 SNP was associated with high hepatic fat content in Caucasians and African Americans (all P < 0.05), with high alanine aminotransferase levels in Hispanics (P < 0.05) and a more favorable lipoprotein profile (lower low-density lipoprotein, small dense low-density lipoprotein, and very small low-density lipoprotein) in Caucasians and Hispanics (all P < 0.05). The liver biopsy showed a higher prevalence of fibrosis (P = 0.04) and a higher nonalcoholic fatty liver disease activity score (P = 0.05) in subjects carrying the minor allele than in those homozygous for the common allele. Moreover, we observed a joint effect among the TM6SF2 rs58542926, the PNPLA3 rs738409, and the GCKR rs1260326 SNPs in determining intrahepatic fat accumulation (P < 0.05). CONCLUSION: The rs58542926 SNP in the TM6SF2 gene is associated with pediatric nonalcoholic fatty liver disease but may confer protection against cardiovascular risk.

A low disposition index in adolescent offspring of mothers with gestational diabetes: a risk marker for the development of impaired glucose tolerance in youth.

AIMS/HYPOTHESIS: With the increase in gestational diabetes mellitus (GDM), there is a growing need to understand the effects of intrauterine glucose exposure on the newborn at birth and later in life. The risk of developing impaired glucose tolerance (IGT) in individuals exposed to diabetes in utero has not been adequately investigated. METHODS: We studied 255 obese adolescents with normal glucose tolerance. All of them were investigated for in utero exposure to GDM and underwent an OGTT, which was repeated after approximately 2.8 years. RESULTS: 210 (82.3%) participants were not exposed to GDM (NGDM group), and 45 (17.7%) were exposed to GDM (EGDM group). In the NGDM group, only 8.6% (n = 18) developed either IGT or type 2 diabetes compared with 31.1% (n = 14) of the EGDM group who developed either IGT or type 2 diabetes (p < 0.001). Exposure to GDM was the most significant predictor of developing IGT or type 2 diabetes (OR 5.75, 95% CI 2.19, 15.07, p < 0.001). At baseline and at follow-up, the EGDM group showed a reduction in beta cell function determined by the oral disposition index (p = 0.03 and p = 0.01, respectively), and, at follow-up, they also displayed a reduction in insulin sensitivity compared with the NGDM group (p = 0.05). CONCLUSIONS/INTERPRETATION: Obese youth exposed in utero to GDM show early inability of the beta cell to compensate adequately in response to decreasing levels of insulin sensitivity.

Variant in the glucokinase regulatory protein (GCKR) gene is associated with fatty liver in obese children and adolescents.

UNLABELLED: Recently, the single nucleotide polymorphism (SNP) identified as rs1260326, in the glucokinase regulatory protein (GCKR), was associated with hypertriglyceridemia in adults. Because accumulation of triglycerides in hepatocytes represents the hallmark of steatosis, we aimed to investigate whether this variant might be associated with fatty liver (hepatic fat content, HFF%). Moreover, because recently rs738409 in the PNPLA3 and rs2854116 in the APOC3 were associated with fatty liver, we explored how the GCKR SNP and these two variants jointly influence hepatosteatosis. We studied 455 obese children and adolescents (181 Caucasians, 139 African Americans, and 135 Hispanics). All underwent an oral glucose tolerance test and fasting lipoprotein subclasses measurement by proton nuclear magnetic resonance. A subset of 142 children underwent a fast gradient magnetic resonance imaging to measure the HFF%. The rs1260326 was associated with elevated triglycerides (Caucasians P = 0.00014; African Americans P = 0.00417), large very low-density lipoprotein (VLDL) (Caucasians P = 0.001; African Americans, P = 0.03), and with fatty liver (Caucasians P = 0.034; African Americans P = 0.00002; and Hispanics P = 0.016). The PNPLA3, but not the APOC3 rs2854116 SNP, was associated with fatty liver but not with triglyceride levels. There was a joint effect between the PNPLA3 and GCKR SNPs, explaining 32% of HFF% variance in Caucasians (P = 0.00161), 39.0% in African Americans (P = 0.00000496), and 15% in Hispanics (P = 0.00342). CONCLUSION: The rs1260326 in GCKR is associated with hepatic fat accumulation along with large VLDL and triglyceride levels. GCKR and PNPLA3 act together to convey susceptibility to fatty liver in obese youths.

Incretin effect determines glucose trajectory and insulin sensitivity in youths with obesity.

In youths with obesity, the gut hormone potentiation of insulin secretion - the incretin effect - is blunted. We explored the longitudinal impact of the incretin effect during pubertal transition on ß cell function and insulin sensitivity. Youths with obesity and 2-hour glucose level ≥ 120 mg/dL underwent a 3-hour oral glucose-tolerance test (OGTT) and an isoglycemic i.v. glucose infusion to quantify the incretin effect. After 2 years, 30 of 39 participants had a repeated OGTT and were stratified into 3 tertiles according to the baseline incretin effect. The high-incretin effect group demonstrated a longitudinal increase in ß cell function (disposition index, minimal model [DIMM]), with greater insulin sensitivity at follow-up and stable insulin secretion (φtotal). A lower incretin effect at baseline was associated with higher 1-hour and 2-hour glucose level at follow-up. The high-incretin effect group displayed a greater increase of GLP-17-36 than the moderate- and low-incretin group at baseline, while such a difference did not persist after 2 years. Glucagon suppression was reduced at follow-up in those with low-baseline incretin in respect to the high-incretin group. The incretin effect during pubertal transition affected the longitudinal trajectory of ß cell function and weight in youths with obesity.

Growth differentiation factor 15 (GDF15) is associated with non-alcoholic fatty liver disease (NAFLD) in youth with overweight or obesity.

OBJECTIVE: Growth differentiation factor 15 (GDF15) has been associated with food intake and weight regulation in response to metabolic stress. In animal models, it has been noted that it may play a role in the progression of non-alcoholic fatty liver disease (NAFLD), the leading cause of chronic liver disease in children. DESIGN: In the current study, we explored the association of circulating plasma concentrations of GDF15 with NAFLD in youth with overweight/obesity, and whether changes in plasma concentrations in GDF15 parallel the changes in intrahepatic fat content (HFF%) over time. METHODS: Plasma GDF15 concentrations were measured by ELISA in 175 youth with overweight/obesity who underwent an oral glucose tolerance test (OGTT) and magnetic resonance imaging (MRI) to assess intrahepatic, visceral, and subcutaneous fat. Baseline fasting GDF15 concentrations were measured in twenty-two overweight/obese youth who progressed (n = 11) or regressed (n = 11) in HFF% by more than 30% of original over a 2-year period. RESULTS: Youth with NAFLD had significantly higher plasma concentrations of GDF15 than those without NAFLD, independent of age, sex, ethnicity, BMI z-score (BMIz), and visceral fat (P = 0.002). During the OGTT, there was a decline in plasma GDF15 concentrations from 0 to 60 min, but GDF15 concentrations returned to basal levels by the end of the study. There was a statistically significant association between change in HFF% and change in GDF15 (P = 0.008; r2 = 0.288) over ~2 years of follow-up. CONCLUSIONS: These data suggest that plasma GDF15 concentrations change with change in intrahepatic fat content in youth with overweight/obesity and may serve as a biomarker for NAFLD in children.

A common variant in the patatin-like phospholipase 3 gene (PNPLA3) is associated with fatty liver disease in obese children and adolescents.

UNLABELLED: The genetic factors associated with susceptibility to nonalcoholic fatty liver disease (NAFLD) in pediatric obesity remain largely unknown. Recently, a nonsynonymous single-nucleotide polymorphism (rs738409), in the patatin-like phospholipase 3 gene (PNPLA3) has been associated with hepatic steatosis in adults. In a multiethnic group of 85 obese youths, we genotyped the PNLPA3 single-nucleotide polymorphism, measured hepatic fat content by magnetic resonance imaging and insulin sensitivity by the insulin clamp. Because PNPLA3 might affect adipogenesis/lipogenesis, we explored the putative association with the distribution of adipose cell size and the expression of some adipogenic/lipogenic genes in a subset of subjects who underwent a subcutaneous fat biopsy. Steatosis was present in 41% of Caucasians, 23% of African Americans, and 66% of Hispanics. The frequency of PNPLA3(rs738409) G allele was 0.324 in Caucasians, 0.183 in African Americans, and 0.483 in Hispanics. The prevalence of the G allele was higher in subjects showing hepatic steatosis. Surprisingly, subjects carrying the G allele showed comparable hepatic glucose production rates, peripheral glucose disposal rate, and glycerol turnover as the CC homozygotes. Carriers of the G allele showed smaller adipocytes than those with CC genotype (P = 0.005). Although the expression of PNPLA3, PNPLA2, PPARγ2(peroxisome proliferator-activated receptor gamma 2), SREBP1c(sterol regulatory element binding protein 1c), and ACACA(acetyl coenzyme A carboxylase) was not different between genotypes, carriers of the G allele showed lower leptin (LEP)(P = 0.03) and sirtuin 1 (SIRT1) expression (P = 0.04). CONCLUSION: A common variant of the PNPLA3 gene confers susceptibility to hepatic steatosis in obese youths without increasing the level of hepatic and peripheral insulin resistance. The rs738409 PNPLA3 G allele is associated with morphological changes in adipocyte cell size.

CIDEA expression in SAT from adolescent girls with obesity and unfavorable patterns of abdominal fat distribution.

OBJECTIVE: This study investigated whether variations in cell death-inducing DNA fragmentation factor alpha subunit-like effector A (CIDEA) mRNA expression and protein levels are modulated by the pattern of abdominal fat distribution in adolescent girls with obesity. METHODS: This study recruited 35 adolescent girls with obesity and characterized their abdominal fat distribution by magnetic resonance imaging. Participants had only a periumbilical/abdominal (n = 14) or a paired abdominal and gluteal subcutaneous adipose tissue (SAT) biopsy (n = 21). CIDEA expression was determined by reverse transcription-polymerase chain reaction, CIDEA protein level by Western blot, and the turnover of adipose lipids and adipocytes by 2 H2 O labeling. In six girls, a second abdominal SAT biopsy was performed (after ~34.2 months) to explore the weight gain effect on CIDEA expression in abdominal SAT. RESULTS: CIDEA expression decreased in abdominal SAT from participants with high visceral adipose tissue (VAT)/(VAT+SAT); CIDEA inversely correlated with number of small adipocytes, with the increase in preadipocyte proliferation, and with adipogenesis. A strong inverse correlation was found between CIDEA protein level with the newly synthetized glycerol (r = -0.839, p = 0.0047). Following weight gain, an increase in adipocytes' cell diameter with a decrease in CIDEA expression and RNA-sequencing transcriptomic profile typical of adipocyte dysfunction was observed. CONCLUSIONS: Reduced expression of CIDEA in girls with high VAT/(VAT+SAT) is associated with adipocyte hypertrophy and insulin resistance.

Glucose dysregulation and hepatic steatosis in obese adolescents: is there a link?

UNLABELLED: Fatty liver is increasingly common in obese adolescents. We determined its association with glucose dysregulation in 118 (37M/81F) obese adolescents of similar age and percent total fat. Fast-magnetic resonance imaging (MRI) and simple MRI were used to quantify hepatic fat content and abdominal fat distribution. All subjects had a standard oral glucose tolerance test. Insulin sensitivity was estimated by the Matsuda Index and homeostasis model assessment of insulin resistance. Baseline total and high molecular weight (HMW)-adiponectin and interleukin (IL)-6 levels were measured. The cohort was stratified according to tertiles of hepatic fat content. Whereas age and %fat were comparable across tertiles, ethnicity differed in that fewer Blacks and more Whites and Hispanics were in the moderate and high category of hepatic fat fraction (HFF). Visceral and the visceral-to-subcutaneous fat ratio increased and insulin sensitivity decreased across tertiles. Two-hour plasma glucose rose with increasing hepatic steatosis (P < 0.008). 73.7% of the subjects in the high HFF had the metabolic syndrome compared to 19.5% and 30.6%, respectively, in the low and moderate categories. Both total and HMW-adiponectin decreased, and IL-6 increased with increasing hepatic steatosis. CONCLUSION: In obese adolescents, independent of total fat, increasing severity of fatty liver is associated with glucose dysregulation, metabolic syndrome, and with a proinflammatory milieu.

A Reduced Incretin Effect Mediated by the rs7903146 Variant in the TCF7L2 Gene Is an Early Marker of ß-Cell Dysfunction in Obese Youth.

OBJECTIVE: The risk genotype for the common variant rs7903146 of the transcription factor 7-like-2 (TCF7L2) gene has been found to affect the incretin response in healthy and obese adults; however, whether a similar functional defect is also present in obese adolescents remains unexplored. Herein, we examined the functional effect of the rs7903146 variant in the TCF7L2 gene on the incretin effect and determined its translational metabolic manifestation by performing deep phenotyping of the incretin system, ß-cell function relative to insulin sensitivity, the gastrointestinal-induced glucose disposal (GIGD) in obese youth with normal and impaired glucose tolerance. RESEARCH DESIGN AND METHODS: Thirty-nine obese adolescents without diabetes (median age 15 [25th, 75th percentile 14, 18] years; BMI 37 [33, 43] kg/m2) were genotyped for the rs7903146 variant of TCF7L2 and underwent a 3-h oral glucose tolerance test (OGTT) followed by an isoglycemic intravenous glucose infusion (iso-intravenous glucose tolerance test [IVGTT]) to match the plasma glucose concentrations during the OGTT and a hyperglycemic clamp with arginine stimulation. The incretin effect was measured as 100 * (AUC-SROGTT - AUC-SRiso-IVGTT) / AUC-SROGTT, where AUC-SR = area under the curve of C-peptide secretion rate. Participants were grouped into tertiles according to the percentage incretin effect (high, moderate, and low) to describe their metabolic phenotype. RESULTS: The presence of T risk allele for TCF7L2 was associated with a markedly reduced incretin effect compared with the wild-type genotype (0.3% [-7.2, 14] vs. 37.8% [12.5, 52.4], P < 0.002). When the cohort was stratified by incretin effect, the high, moderate, and low incretin effect groups did not differ with respect to anthropometric features, while the low incretin effect group exhibited higher 1-h glucose (P = 0.015) and a reduced disposition index, insulin sensitivity, and insulin clearance compared with the high incretin effect group. GIGD was reduced in the low incretin effect group (P = 0.001). The three groups did not differ with respect to intravenous glucose-induced insulin secretion and arginine response during the hyperglycemic clamp. CONCLUSIONS: A reduced incretin effect and its association with the TCF7L2 variant rs7903146 identify an early metabolic phenotype in obese youth without diabetes, featuring a higher plasma glucose peak at 1 h; lower insulin secretion, sensitivity, and clearance; and GIGD.

Effect of Gut Microbiota and PNPLA3 rs738409 Variant on Nonalcoholic Fatty Liver Disease (NAFLD) in Obese Youth.

CONTEXT: Nonalcoholic fatty liver disease (NAFLD) is the most common cause of liver disease, affecting approximately 3 in 10 obese children worldwide. OBJECTIVE: We aimed to investigate the potential relationship between gut microbiota and NAFLD in obese youth, while considering the role of PNPLA3 rs738409, a strong genetic contributor to NAFLD. DESIGN: In this cross-sectional study, participants completed an abdominal magnetic resonance imaging to measure hepatic fat fraction (HFF), oral glucose tolerance test, and PNPLA3 rs738409 genotyping. Fecal samples were collected to analyze the V4 region of the 16S rRNA gene for intestinal bacteria characterization. SETTING: Yale Pediatric Obesity Clinic. PARTICIPANTS: Obese youth (body mass index >95th percentile) with NAFLD (HFF ≥5.5%; n = 44) and without NAFLD (HFF <5.5%; n = 29). MAIN OUTCOME MEASURE: Shannon-Wiener diversity index values and proportional bacterial abundance by NAFLD status and PNPLA3 genotype. RESULTS: Subjects with NAFLD had decreased bacterial alpha-diversity compared with those without NAFLD (P = 0.013). Subjects with NAFLD showed a higher Firmicutes to Bacteroidetes (F/B) ratio (P = 0.019) and lower abundance of Bacteroidetes (P = 0.010), Prevotella (P = 0.019), Gemmiger (P = 0.003), and Oscillospira (P = 0.036). F/B ratio, Bacteroidetes, Gemmiger, and Oscillospira were associated with HFF when controlling for group variations. We also observed an additive effect on HFF by PNPLA3 rs738409 and Gemmiger, and PNPLA3 rs738409 and Oscillospira. CONCLUSIONS: Obese youth with NAFLD have a different gut microbiota composition than those without NAFLD. These differences were still statistically significant when controlling for factors associated with NAFLD, including PNPLA3 rs738409.

Metabolic and Genetic Determinants of Glucose Shape After Oral Challenge in Obese Youths: A Longitudinal Study.

CONTEXT: The time-to-glucose-peak following the oral glucose tolerance test (OGTT) is a highly reproducible marker for diabetes risk. In obese youths, we lack evidence for the mechanisms underlying the effects of the TCF7L2 rs7903146 variant on glucose peak. METHODS: We analyzed the metabolic phenotype and the genotype for the TCF7L2 rs7903146 in 630 obese youths with normal (NGT) and impaired (IGT) glucose tolerance. Participants underwent a 3-hour, 9-point OGTT to estimate, using the oral minimal model, the disposition index (DI), the static (φstatic) and dynamic (φdynamic) components ß-cell responsiveness and insulin sensitivity (SI). In a subgroup (n = 241) longitudinally followed for 2 years, we estimated the effect of time-to-glucose-peak on glucose tolerance change. RESULTS: Participants were grouped into early (<30 minutes) and late (≥30 minutes) glucose peakers. A delayed glucose peak was featured by a decline in φstatic (P < .001) in the absence of a difference in φdynamic. The prevalence of T-risk allele for TCF7L2 rs7903146 variant significantly increased in the late peak group. A lower DI was correlated with higher glucose concentration at 1 and 2 hours, whereas SI was inversely associated with 1-hour glucose. Glucose peak <30 minutes was protective toward worsening of glucose tolerance overtime (odds ratio 0.35 [0.15-0.82]; P = .015), with no subjects progressing to NGT or persisting IGT, in contrast to the 40% of progressor in those with late glucose peak. CONCLUSION: The prevalence of T-risk allele for the TCF7L2 rs7903146 prevailed in the late time-to-glucose peak group, which in turn is associated with impaired ß-cell responsiveness to glucose (φ), thereby predisposing to prediabetes and diabetes in obese youths.

The omentum of obese girls harbors small adipocytes and browning transcripts.

Severe obesity (SO) affects about 6% of youth in the United States, augmenting the risks for cardiovascular disease and type 2 diabetes. Herein, we obtained paired omental adipose tissue (omVAT) and abdominal subcutaneous adipose tissue (SAT) biopsies from girls with SO undergoing sleeve gastrectomy (SG), to test whether differences in cellular and transcriptomic profiles between omVAT and SAT depots affect insulin sensitivity differently. Following weight loss, these analyses were repeated in a subgroup of subjects having a second SAT biopsy. We found that omVAT displayed smaller adipocytes compared with SAT, increased lipolysis through adipose triglyceride lipase phosphorylation, reduced inflammation, and increased expression of browning/beiging markers. Contrary to omVAT, SAT adipocyte diameter correlated with insulin resistance. Following SG, both weight and insulin sensitivity improved markedly in all subjects. SAT adipocytes' size became smaller, showing increased lipolysis through perilipin 1 phosphorylation, decreased inflammation, and increased expression in browning/beiging markers. In summary, in adolescent girls with SO, both omVAT and SAT depots showed distinct cellular and transcriptomic profiles. Following weight loss, the SAT depot changed its cellular morphology and transcriptomic profiles into more favorable ones. These changes in the SAT depot may play a fundamental role in the resolution of insulin resistance.

Lower Insulin Clearance Parallels a Reduced Insulin Sensitivity in Obese Youths and Is Associated With a Decline in ß-Cell Function Over Time.

We examined the relationship between insulin clearance, insulin sensitivity, and ß-cell function and the longitudinal effect of insulin clearance on ß-cell function in lean and obese insulin-sensitive and insulin-resistant adolescents. A hyperinsulinemic-euglycemic and a hyperglycemic clamp were performed in 110 youths to quantify hepatic and peripheral clearance, insulin sensitivity, and ß-cell function (disposition index, DIh-clamp). Participants underwent an oral glucose tolerance test at baseline and after 2 years to assess glucose tolerance and oral ß-cell function (oDIcpep) and were sorted into four groups (lean and obese normal glucose tolerance, insulin sensitive, insulin resistant, and impaired glucose tolerance). Insulin sensitivity was defined based on the median of insulin stimulated glucose disposal (M) measured during the hyperinsulinemic-euglycemic clamp. Lean and obese insulin-sensitive participants did not differ with respect to hepatic and peripheral clearance or for insulin sensitivity. Insulin sensitivity was linearly correlated with whole-body insulin clearance. Hepatic insulin extraction at baseline acted as an independent determinant of ß-cell function at follow-up. The decline in insulin sensitivity, even in the absence of an impairment of glucose tolerance, is associated with lowering of hepatic insulin clearance in obese youth, which in turn may contribute to the decline in ß-cell function over time.

Relation of the degree of obesity in childhood to adipose tissue insulin resistance.

AIMS: In this study, we investigated whether adipose tissue insulin resistance (IR) is affected by the degree of obesity during the fasting and post-prandial state, independent of glucose tolerance among obese children and adolescents. We also tested whether systemic subclinical inflammation is associated with adipose tissue IR. METHODS: Subjects were recruited to the Yale Pathophysiology of Type 2 Diabetes in Youth Study (NCT01967849). An oral glucose-tolerance test was performed to establish glucose-tolerance status and blood samples were drawn for measurement of free fatty acids (FFAs), to calculate the area under the curve (AUC) of FFA. Adipose tissue insulin resistance was calculated as the product of insulin and FFA concentrations. RESULTS: In total, 671 children and adolescents (58.6% females) were included with a mean age of 13.3(2.7) years and BMI Z score of 2.45(0.31). The degree of obesity emerged as an independent predictor of both fasting and post-prandial adipose IR, p < 0.0001. Higher degree of obesity was associated with greater AUC FFA (lower suppression) compared to lower degree of obesity, p = 0.01. Furthermore, higher levels of IL-6 were positively associated with post-prandial adipose tissue IR, p = 0.02. CONCLUSIONS: The degree of obesity in childhood and adolescence is strongly associated with adipose tissue IR independent of glucose tolerance. This is reflected not only in calculated indices of adipose IR but also in lower suppression of FFAs during the OGTT regardless of glucose tolerance or fasting adipose tissue IR. Furthermore, markers of subclinical inflammation such as IL-6 are associated with adipose tissue IR, independent of other factors.