Catherine Le Stunff

Variation in FTO contributes to childhood obesity and severe adult obesity

We identified a set of SNPs in the first intron of the FTO (fat mass and obesity associated) gene on chromosome 16q12.2 that is consistently strongly associated with early-onset and severe obesity in both adults and children of European ancestry with an experiment-wise P value of 1.67 × 10−26 in 2,900 affected individuals and 5,100 controls. The at-risk haplotype yields a proportion of attributable risk of 22% for common obesity. We conclude that FTO contributes to human obesity and hence may be a target for subsequent functional analyses.

Common nonsynonymous variants in PCSK1 confer risk of obesity.

Mutations in PCSK1 cause monogenic obesity. To assess the contribution of PCSK1 to polygenic obesity risk, we genotyped tag SNPs in a total of 13,659 individuals of European ancestry from eight independent case-control or family-based cohorts. The nonsynonymous variants rs6232, encoding N221D, and rs6234-rs6235, encoding the Q665E-S690T pair, were consistently associated with obesity in adults and children (P = 7.27 × 10−8 and P = 2.31 × 10−12, respectively). Functional analysis showed a significant impairment of the N221D-mutant PC1/3 protein catalytic activity.

The insulin gene VNTR is associated with fasting insulin levels and development of juvenile obesity.

In millions of people, obesity leads to type 2 diabetes (T2D; also known as non-insulin-dependent diabetes mellitus). During the early stages of juvenile obesity, the increase of insulin secretion in proportion to accumulated fat balances insulin resistance and protects patients from hyperglycaemia. After several decades, however, β-cell function deteriorates and T2D develops in approximately 20% of obese patients. In modern societies, obesity has thus become the leading risk factor for T2D (ref. 5). The factors that predispose obese patients to alteration of insulin secretion upon gaining weight remain unknown. To determine which genetic factors predispose obese patients to β-cell dysfunction, and possibly T2D, we studied single-nucleotide polymorphisms (SNPs) in the region of the insulin gene (INS) among 615 obese children. We found that, in the early phase of obesity, alleles of the INS variable number of tandem repeat (VNTR) locus are associated with different effects of body fatness on insulin secretion. Young obese patients homozygous for class I VNTR alleles secrete more insulin than those with other genotypes.

Paternal transmission of the very common class I INS VNTR alleles predisposes to childhood obesity.

To identify some of the genetic factors that contribute to obesity in children of Central European and North African descent, we studied the parental transmission of alleles at the insulin locus to offspring with early-onset obesity. A variable nucleotide tandem repeat (VNTR) polymorphism upstream of the insulin gene (INS) is associated with variations in the expression of INS and the nearby gene encoding insulin-like growth factor 2 (IGF2). We found an excess of paternal transmission of class I VNTR alleles to obese children: children who inherited a class I allele from their father (but not those inheriting it from their mother) had a relative risk of early-onset obesity of 1.8. Due to the frequency of class I alleles in this population, this risk concerns 65–70% of all infants. These results suggest that increased in utero expression of paternal INS or IGF2 due to the class I INS VNTR allele may predispose offspring to postnatal fat deposition.

In vivo resistance of lipolysis to epinephrine. A new feature of childhood onset obesity.

A decreased mobilization of triglycerides may contribute to fat accumulation in adipocytes, leading to obesity. However, this hypothesis remains to be proven. In this study, epinephrine-induced lipid mobilization was investigated in vivo in nine markedly obese children (160+/-5% ideal body weight) aged 12.1+/-0.1 yr during the dynamic phase of fat deposition, compared with six age-matched nonobese children. As an in vivo index of lipolysis, we measured glycerol flux using a nonradioactive tracer dilution approach, and plasma free fatty acid concentrations. In the basal state, the obese children had a 30% lower rate of glycerol release per unit fat mass than the lean children. To study the regulation of lipolysis, epinephrine was infused stepwise at fixed doses of 0.75 and then 1. 50 microg/min in both groups. In lean children, glycerol flux and plasma free fatty acid increased to an average of 249-246% of basal values, respectively, in response to a mean plasma epinephrine of 396+/-41 pg/ml. The corresponding increase was only 55-72% in the obese children, although their mean plasma epinephrine reached 606+/-68 pg/ml. All obese and nonobese children, except an Arg64Trp heterozygote, were homozygotes for Trp at position 64 of their beta3-adrenoreceptor. The resistance of lipolysis to epinephrine showed no relationship with the Arg64 polymorphism of the beta3-adrenoreceptor gene. In summary, in vivo lipolysis, which mostly reflects the mobilization of lipid stores from subcutaneous adipose tissue, shows a decreased sensitivity to epinephrine in childhood onset obesity. Since our study was carried out in obese children during the dynamic phase of fat accumulation, the observed resistance to catecholamines might possibly be causative rather than the result of obesity.

Molecular Genetics of Human Obesity‐Associated MC4R Mutations

Abstract: Heterozygous coding mutations in the melanocortin 4 receptor (MC4R) are implicated in 1 to 6% of early onset or severe adult obesity cases. To better address the problem of the genotype:phenotype relationship within this specific form of obesity, we systematically studied the functional characteristics of 50 different obesity‐associated MC4R mutations. Structure modeling of MC4R indicates that obesity‐associated MC4R mutations are not localized in a single domain of the protein. We developed a flow cytometry‐based assay to compare cell membrane expression of obesity‐associated MC4R mutants. Using this assay, we demonstrate that over 54% of the obesity‐associated MC4R mutations impair the membrane expression of MC4R. All other mutations impair the basal constitutive activity and/or the EC50 for the physiological agonist α‐MSH as measured in a cAMP‐ dependent luciferase assay. The extent of the alterations in receptor activity ranges from a total suppression of MC4R activation in response to α‐MSH to a mild alteration of the basal constitutive activity of the receptor. Since most patients are heterozygous for MC4R mutations, these data indicate that a small decrease in overall MC4R activity can cause obesity, strongly supporting the hypothesis that the MC4R is a critical component of the adipostat in humans.

Endocrine Manifestations of the Rapid-Onset Obesity with Hypoventilation, Hypothalamic, Autonomic Dysregulation, and Neural Tumor Syndrome in Childhood

CONTEXT Rapid-onset obesity with hypoventilation, hypothalamic, autonomic dysregulation, and neural tumor (ROHHADNET) is a newly described syndrome that can cause cardiorespiratory arrests and death. It mimics several endocrine disorders or genetic obesity syndromes during early childhood and is associated with various forms of hypothalamic-pituitary endocrine dysfunctions that have not yet been fully investigated. OBJECTIVE The current report aspires to facilitate the earlier recognition and appropriate treatment of the ROHHADNET syndrome when children present with various endocrine manifestations, such as early obesity, growth failure, pseudo-Cushings syndrome, glucocorticoid insufficiency, congenital hypopituitarism, or adrenal tumors. A more widespread knowledge of the syndrome will help characterize its molecular origin. DESIGN Endocrine studies were performed in six patients admitted for seemingly common early-onset obesity associated with growth failure in five of them. The six patients later showed distinctive features of the ROHHADNET syndrome. RESULTS Abnormalities of the pituitary adrenal axis ranged from a true Cushing-like profile (one of six), to glucocorticoid deficiency with normal ACTH (two of six). Complete GH deficiency with low IGF-I was observed in four of six, hypogonadotropic hypogonadism in four of six, hyperprolactinemia in six of six, and various degrees of TSH/T(4) abnormalities in five of five patients. All had increased natremia without diabetes insipidus. Five children had unilateral macroscopic adrenal ganglioneuroma. Two patients died at 8.5 and 12 yr of age. CONCLUSIONS Various hypothalamic-pituitary endocrine dysfunctions are associated with ROHHADNET, carrying a risk of misdiagnosis until other elements of the syndrome make it more easily recognizable. Given its severity, ROHHADNET syndrome should be considered in all cases of isolated, rapid, and early obesity.

Association analysis indicates that a variant GATA-binding site in the PIK3CB promoter is a Cis-acting expression quantitative trait locus for this gene and attenuates insulin resistance in obese children.

OBJECTIVE—In search of functional polymorphisms associated with the genetics of insulin resistance, we studied a variant in the promoter of PIK3CB, the gene coding for the catalytic p110β subunit of phosphatidylinositol (PI) 3-kinase, a major effector of insulin action. RESEARCH DESIGN AND METHODS—The rs361072 C/T variant was selected among single nucleotide polymorphisms of the PIK3CB region because we suspected that its common C allele (allelic frequency ∼50% in Europeans) could create a GATA-binding motif and was genotyped in five cohorts of obese (n = 1,876) and two cohorts of nonobese (n = 1,490) European children. To estimate insulin resistance in these children, the homeostasis model assessment for insulin resistance (HOMA-IR) index was measured in strict nutritional conditions. GATA-binding and functional effects of rs361072 were explored in transfected cell lines and in lymphocytes from obese children. RESULTS—The rs361072 C/T variant was associated with HOMA-IR in the obese children cohorts (1.7 × 10−12 < P < 2 × 10−4 for C/C vs. T/T using regression analysis). HOMA-IR averaged 3.3 ± 0.1 in C/C and 4.5 ± 0.2 in T/T obese children (P = 4.5 × 10−6 by ANOVA). C/T patients had intermediate values. As shown by the interaction between BMI and genotype (P = 2.1 × 10−9), the association of rs361072 with HOMA-IR depended on BMI and was only marginal in nonobese children (P = 0.04). At the molecular level, the C allele of rs361072 was found to create a GATA-binding site able to increase transcription of PIK3CB. CONCLUSIONS—We postulate that the C allele of rs361072 is a causal variant capable of attenuating insulin resistance in obese children through increased expression of p110β.

In obese and non-obese adults, the cis-regulatory rs361072 promoter variant of PIK3CB is associated with insulin resistance not with type 2 diabetes

We recently reported that rs361072, a promoter C/T variant of p110beta, the catalytic subunit of PI3-kinase, was associated with a protection from insulin resistance (IR) in Caucasian adolescents in proportion of their body mass. We tested if this cis-regulatory QTL is associated with IR and type 2 diabetes in 7885 middle-aged obese and non-obese adults of European ancestry. We genotyped rs361072 in 1139 non-diabetic obese (NDO) European adults, in whom IR was estimated by the HOMA-IR index. We also studied 427 type 2 diabetic obese adults (DO) and 424 diabetic non-obese (DNO) adults to test whether their disease status was associated with a decreased prevalence of the protective variant. The prevalence of rs361072 and association with IR was also examined in 5895 non-obese non-diabetic adults (NDNO). rs361072 was associated with HOMA-IR (p=4.10(-4)) in NDO, so that C/C patients had a 17% decrease of this index (p=0.002). A statistical trend (p=1.1.10(-2)) for the same genotypic differences was also observed in NDNO adults, but of insignificant magnitude (4.2%). The distribution of rs361072 genotype was comparable in NDO, DO, DNO and NDNO individuals. Allele C of rs361072 is associated with a protection from IR in obese and non-obese adults, but has no significant effect, however, on diabetes risk in obese or non-obese Europeans.

Methylation and Transcripts Expression at the Imprinted GNAS Locus in Human Embryonic and Induced Pluripotent Stem Cells and Their Derivatives

Summary Data from the literature indicate that genomic imprint marks are disturbed in human pluripotent stem cells (PSCs). GNAS is an imprinted locus that produces one biallelic (Gsα) and four monoallelic (NESP55, GNAS-AS1, XLsα, and A/B) transcripts due to differential methylation of their promoters (DMR). To document imprinting at the GNAS locus in PSCs, we studied GNAS locus DMR methylation and transcript (NESP55, XLsα, and A/B) expression in human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs) derived from two human fibroblasts and their progenies. Results showed that (1) methylation at the GNAS locus DMRs is DMR and cell line specific, (2) changes in allelic transcript expression can be independent of a change in allele-specific DNA methylation, and (3) interestingly, methylation at A/B DMR is correlated with A/B transcript expression. These results indicate that these models are valuable to study the mechanisms controlling GNAS methylation, factors involved in transcript expression, and possibly mechanisms involved in the pathophysiology of pseudohypoparathyroidism type 1B.