Fat Mass- And Obesity-associated mRNA Expression Research Articles

Abstract 9526: Fto Demethylase Mediates Altered N6-methyladenosine (m6a)-rna Modification in Human Failing Heart and Stressed-Induced Rat Cardiomyocytes

Background: N6-methyladenosine (m6A) methylation is one of the important posttranscriptional modification of RNA, which affects RNA splicing, translation and stability. These modifications are dynamically regulated by writers, readers and erasers, and their alteration affects m6A modification and have shown to play key regulatory role in numerous biological processes. Fat mass- and obesity-associated (FTO) gene, known as a nucleic acid demethylase removes the methyl group from RNA and has been shown to be associated with progression of heart diseases. In this study, we determined m6A methylation in human failing heart, and evaluated if FTO regulates m6A-methylation of transcripts related to inflammation in cultured rat cardiomyocytes under various stress conditions. Methods and Results: m6A RNA-methylation was increased in human failing hearts as compared to heart samples from non-failing subjects. Interestingly, failing hearts were associated with significant decrease in FTO mRNA and protein expression. Furthermore, cultured rat cardiomyoblasts (H9C2) cells exposed to stressors (hypoxia or isoproterenol) for 24hrs showed significant decrease of FTO and an associated increase in m6A-RNA methylation (dot blot analysis). To further test if stress-induced increase in m6A-RNA modification is mediated through FTO, we knocked-down FTO, treated with isoproterenol for 24hrs and evaluated m6A methylation status of RNA. Interestingly, dot blot analysis showed significant increase in m6A methylation levels and proinflammatory cytokines (IL6 and TNFa) in FTO knockdown cells as compared to scramble, both at basal level and after exposure to stress. Furthermore, we evaluated the effect of FTO on methylation of RNA transcripts that encode inflammatory cytokines. MeRIP assay (Methylated RNA immunoprecipitation using m6A antibody and analysis of m6A-transcript by qRT-PCR) demonstrated that the gene expression of proinflammatory cytokines, IL6 and TNF-alpha was significantly increased in FTO knockdown cells as compared to scramble control cells . Conclusion: This data suggests stress-induced loss of FTO in the heart might mediate m6A-RNA methylation and therefore potential leading to upregulation of inflammatory response in the myocardium.

FTO mRNA expression in the lower quartile is associated with bad prognosis in clear cell renal cell carcinoma based on TCGA data mining

Fat mass and obesity associated (FTO) is a protein-coding gene, also known as the obesity gene. It has been reported previously to be associated with a variety of malignant cancers, such as breast, thyroid and acute myeloid leukemia. The aim of the present study was to investigate the FTO mRNA expression in human clear cell renal cell carcinoma and its clinical value. FTO mRNA expression and its prognostic value were investigated by bioinformatic analysis of the data from The Cancer Genome Atlas (TCGA, https://cancergenome.nih.gov/). The Kaplan-Meier analysis showed that FTO mRNA expression in the lower quartile is significantly associated with poor survival in clear cell renal cell carcinoma patients (P < 0.0001). This study indicated that higher FTO mRNA expression may have a protective role and it may be a vital molecular marker in the prognosis of clear cell renal cell carcinoma patients.

Birth weight is associated with placental fat mass- and obesity-associated gene expression and promoter methylation in a Chinese population

Objective: To explore the relationship between birth weight and fat mass- and obesity-associated (FTO) gene expression and promoter methylation status in the Chinese population.Methods: Seventy-five neonates and their mothers were recruited from Yuying Children’s Hospital of Wenzhou Medical University. Subjects were divided into three groups by birth weight: low (<3500 g, n = 20), medium (3500–3999 g, n = 30) and high (≥4000 g, n = 25). Placental FTO transcript levels and promoter methylation were determined by quantitative PCR and Sequenom MassARRAY®.Results: Placental FTO mRNA expression was significantly increased in the high- and medium-weight groups compared to the low-weight group (p = 0.023). Methylation rates of CpG11 sites were significantly decreased in high-birth weight newborns (p = 0.018). Multiple linear regressions showed placental FTO mRNA, maternal pre-pregnancy body mass index (BMI) and CpG11 methylation rate were independently associated with increased fetal birth weight. Additionally, FTO mRNA expression was negatively associated with CpG6.7.8.9 methylation in mothers that underwent C-section.Conclusions: High placental FTO expression is associated with increased birth weight in Chinese neonates, and FTO promoter methylation level at a specific CpG site is negatively associated with birth weight. Further work is needed to determine the functionality of this CpG site in placentas.

Decreased N(6)-methyladenosine in peripheral blood RNA from diabetic patients is associated with FTO expression rather than ALKBH5.

N(6)-methyladenosine (m(6)A) modification plays a fundamental role in the epigenetic regulation of the mammalian transcriptome. m(6)A can be demethylated by fat mass- and obesity-associated (FTO) protein and α-ketoglutarate-dependent dioxygenase alkB homolog 5 (ALKBH5) protein. However, the importance of m(6)A alteration in type 2 diabetes mellitus (T2DM) has not been explored. The objective of the study was to investigate whether m(6)A content was reduced in T2DM patients and whether m(6)A content was correlated with the mRNA expression levels of the FTO and ALKBH5 genes. In this case-control study, peripheral blood samples were obtained from 88 T2DM patients and 92 healthy controls. For the diabetic animal model experiment, blood samples were obtained from seven diabetic and eight nondiabetic rats. A sensitive liquid chromatography-electrospray ionization-tandem mass spectrometry method was developed for the determination of the m(6)A content in RNA, quantitative real-time PCR was used to examine the mRNA expression levels of the FTO and ALKBH5 genes, and high-resolution melting and DNA sequencing were used to detect FTO single-nucleotide polymorphisms. Our results showed that the m(6)A contents in the RNA from T2DM patients and diabetic rats were significantly lower compared with the control groups (P = 2.6 × 10(-24) for T2DM patients; P = .001 for diabetic rats, respectively), and T2DM can be characterized by the content of m(6)A. The mRNA expression level of FTO was significantly higher in T2DM patients than that of the controls (P = .0007) and was associated with the risk of T2DM (odds ratio 2.797, 95% confidence interval 1.452-5.389, P = .002). Moreover, the m(6)A contents were correlated with FTO mRNA expression. These data suggest that the increased mRNA expression of FTO could be responsible for the reduction of m(6)A in T2DM, which may further increase the risk of complications of T2DM. Low m(6)A should be investigated further as a novel potential biomarker of T2DM.

Placental expression of the obesity-associated gene FTO is reduced by fetal growth restriction but not by macrosomia in rats and humans.

Genetic variants in the FTO (fat mass- and obesity-associated) gene have the highest association of all obesity-associated genes. Its placental expression was shown to relate to birth weight, suggesting that it may participate in the control of fetal weight gain. To gain more insight into the implication of FTO in fetal growth, we measured its placental expression in samples including extremes of abnormal fetal growth, such as after intrauterine growth restriction (IUGR) or macrosomia in both rats and humans. In rats, fetal growth was modulated by maternal nutritional modifications. In humans, placental villi were collected from pathological pregnancies (i.e. with IUGR or fetal macrosomia). Placental FTO mRNA expression was reduced by IUGR but was not significantly affected by macrosomia in either rats or humans. Our data suggest that placental FTO may participate in interactions between the in utero environment and the control of fetal growth under IUGR conditions by modulating epigenetic processes.

Chicken FTO gene: Tissue-specific expression, brain distribution, breed difference and effect of fasting

Fat mass and obesity-associated (FTO) gene is widely expressed in central and peripheral tissues of mammals, and exhibits a range of functions, especially in energy balance. However, basic knowledge of FTO in the chicken is lacking. Therefore, we studied the tissue distribution, age and breed dependent changes, brain localization, as well as the impact of fasting on FTO mRNA expression in the chicken. FTO mRNA was expressed in all the tissues studied, and generally, with high expression in hypothalamus, liver, visceral fat and cerebellum. However it exhibited breed-specific patterns: in broilers, the highest expression was seen in the liver, while in layers, hypothalamus and cerebellum showed relatively higher FTO mRNA expression. One-week-old broilers expressed markedly higher FTO mRNA in liver compared with the layers of the same age (P<0.01), while the breed difference was reversed in visceral fat and cerebellum (P<0.05). Compared with newly hatched chicks (one week of age), adult layers expressed higher FTO mRNA in liver and visceral fat, while adult broilers showed higher expression in hypothalamus and cerebellum. In situ hybridization demonstrated distribution of FTO mRNA in paraventricularis magnocellularis (PVN), nucleus ventromedialis hypothalami (VMN), nucleus lateralis hypothalami (LHy), nucleus dorsomedialis hypothalami (DMN) of the hypothalamus and nucleus habenularis medialis (HM) and stratum cellulare externum (SCE) of the thalamus. Breed-specific expression of FTO mRNA was shown in PVN, but not in VMN, with higher abundance in broilers compared to layers. The decrease in FTO mRNA levels after 24h of fasting was seen only in VMN of layer chickens. These results may provide some intriguing hints for further investigation of FTO function in the chicken.

Early hypothalamic FTO overexpression in response to maternal obesity--potential contribution to postweaning hyperphagia.

BackgroundIntrauterine and postnatal overnutrition program hyperphagia, adiposity and glucose intolerance in offspring. Single-nucleotide polymorphisms (SNPs) of the fat mass and obesity associated (FTO) gene have been linked to increased risk of obesity. FTO is highly expressed in hypothalamic regions critical for energy balance and hyperphagic phenotypes were linked with FTO SNPs. As nutrition during fetal development can influence the expression of genes involved in metabolic function, we investigated the impact of maternal obesity on FTO.MethodsFemale Sprague Dawley rats were exposed to chow or high fat diet (HFD) for 5 weeks before mating, throughout gestation and lactation. On postnatal day 1 (PND1), some litters were adjusted to 3 pups (vs. 12 control) to induce postnatal overnutrition. At PND20, rats were weaned onto chow or HFD for 15 weeks. FTO mRNA expression in the hypothalamus and liver, as well as hepatic markers of lipid metabolism were measured.ResultsAt weaning, hypothalamic FTO mRNA expression was increased significantly in offspring of obese mothers and FTO was correlated with both visceral and epididymal fat mass (P<0.05); body weight approached significance (P = 0.07). Hepatic FTO and Fatty Acid Synthase mRNA expression were decreased by maternal obesity. At 18 weeks, FTO mRNA expression did not differ between groups; however body weight was significantly correlated with hypothalamic FTO. Postnatal HFD feeding significantly reduced hepatic Carnitine Palmitoyltransferase-1a but did not affect the expression of other hepatic markers investigated. FTO was not affected by chronic HFD feeding.SignificanceMaternal obesity significantly impacted FTO expression in both hypothalamus and liver at weaning. Early overexpression of hypothalamic FTO correlated with increased adiposity and later food intake of siblings exposed to HFD suggesting upregulation of FTO may contribute to subsequent hyperphagia, in line with some human data. No effect of maternal obesity was observed on FTO in adulthood.

Involvement of Leptin Receptor Long Isoform (LepRb)-STAT3 Signaling Pathway in Brain Fat Mass- and Obesity-Associated (FTO) Downregulation during Energy Restriction

Obesity is an important risk factor for cardiovascular disease, diabetes and certain cancers. The fat mass- and obesity-associated (FTO) gene is tightly associated with the pathophysiology of obesity, whereas the exact role of FTO remains poorly understood. Here, we investigated the alternations of FTO mRNA and protein expression in the peripheral metabolic tissues and the brain upon energy restriction (ER) and explored the involvement of the leptin signaling pathway in FTO regulation under ER status. ER decreased the FTO mRNA and protein expression in hypothalamus and brainstem but not in periphery. Using double-immunofluorescence staining, FTO was found to be colocalized with the leptin receptor long isoform (LepRb) in arcuate nucleus of hypothalamus and the nucleus of the solitary tract. In LepRb mutant db/db mice, the FTO downregulation in brain and body weight reduction induced by ER were completely abolished. The enhanced phosphorylation of signal transducer and activator of transcription 3 (STAT3) induced by ER was also impaired in db/db mice. Moreover, leptin directly activated the STAT3 signaling pathway and downregulated FTO in in vitro arcuate nucleus of hypothalamus cultures and in vivo wild-type mice but not db/db mice. Thus, our results provide the first evidence that the LepRb-STAT3 signaling pathway is involved in the brain FTO downregulation during ER.

Involvement of leptin receptor long isoform (LepRb)-STAT3 signaling pathway in brain fat mass- and obesity-associated (FTO) downregulation during energy restriction.

Obesity is an important risk factor for cardiovascular disease, diabetes and certain cancers. The fat mass- and obesity-associated (FTO) gene is tightly associated with the pathophysiology of obesity, whereas the exact role of FTO remains poorly understood. Here, we investigated the alternations of FTO mRNA and protein expression in the peripheral metabolic tissues and the brain upon energy restriction (ER) and explored the involvement of the leptin signaling pathway in FTO regulation under ER status. ER decreased the FTO mRNA and protein expression in hypothalamus and brainstem but not in periphery. Using double-immunofluorescence staining, FTO was found to be colocalized with the leptin receptor long isoform (LepRb) in arcuate nucleus of hypothalamus and the nucleus of the solitary tract. In LepRb mutant db/db mice, the FTO downregulation in brain and body weight reduction induced by ER were completely abolished. The enhanced phosphorylation of signal transducer and activator of transcription 3 (STAT3) induced by ER was also impaired in db/db mice. Moreover, leptin directly activated the STAT3 signaling pathway and downregulated FTO in in vitro arcuate nucleus of hypothalamus cultures and in vivo wild-type mice but not db/db mice. Thus, our results provide the first evidence that the LepRb-STAT3 signaling pathway is involved in the brain FTO downregulation during ER.

Anti-inflammatory Profile of FTO Gene Expression in Adipose Tissues From Morbidly Obese Women

Background: The fat mass and obesity associated (FTO) gene has been found to contribute to the risk of obesity in humans, but the function and regulation of FTO mRNA expression in adipose tissues remain to be clarified. Our aims were to assess the FTO gene expression in subcutaneous and visceral adipose tissues from morbidly obese women and its relation with obesity, insulin resistance indices, and most importantly, to obesity-related inflammatory markers. Methods: Paired subcutaneous and visceral fat were excised from 33 morbidly obese women and 12 control women who underwent bariatric surgery by laparoscopic gastric by-pass and elective surgery respectively. Adipose tissue mRNA expression was determined by real time RT-PCR. Results: FTO mRNA expression in visceral adipose tissue (VAT) was significantly higher than in subcutaneous adipose tissue (SAT) from obese but not control patients. SAT FTO expression was reduced in obese women compared to control subjects. It correlated negatively with BMI and insulin resistance indices. FTO expression in SAT was positively related to both circulating and mRNA levels of adiponectin, to adiponectin receptor and to PPAR-Δexpression, but negatively with IL-6 gene expression and with circulating levels of leptin. FTO in VAT was also positively correlated with adiponectin, adiponectin receptor and PPAR-Δ mRNA expression. Conclusion: FTO expression in subcutaneous adipose tissue negatively correlates with obesity and insulin resistance. On the other hand, FTO presents a positive association with the expression of adiponectin, an anti-inflammatory adipokine, and with PPAR-Δ in both adipose tissues. Taken together, our results suggest that FTO is associated with an anti-inflammatory behaviour in morbid obesity.

Regulation and Function of FTO mRNA Expression in Human Skeletal Muscle and Subcutaneous Adipose Tissue

OBJECTIVECommon variants in FTO (the fat mass– and obesity-associated gene) associate with obesity and type 2 diabetes. The regulation and biological function of FTO mRNA expression in target tissue is unknown. We investigated the genetic and nongenetic regulation of FTO mRNA in skeletal muscle and adipose tissue and their influence on in vivo glucose and fat metabolism.RESEARCH DESIGN AND METHODSThe FTO rs9939609 polymorphism was genotyped in two twin cohorts: 1) 298 elderly twins aged 62–83 years with glucose tolerance ranging from normal to type 2 diabetes and 2) 196 young (25–32 years) and elderly (58–66 years) nondiabetic twins examined by a hyperinsulinemic-euglycemic clamp including indirect calorimetry. FTO mRNA expression was determined in subcutaneous adipose tissue (n = 226) and skeletal muscle biopsies (n = 158).RESULTSHeritability of FTO expression in both tissues was low, and FTO expression was not influenced by FTO rs9939609 genotype. FTO mRNA expression in skeletal muscle was regulated by age and sex, whereas age and BMI were predictors of adipose tissue FTO mRNA expression. FTO mRNA expression in adipose tissue was associated with an atherogenic lipid profile. In skeletal muscle, FTO mRNA expression was negatively associated to fat and positively to glucose oxidation rates as well as positively correlated with expression of genes involved in oxidative phosphorylation including PGC1α.CONCLUSIONSThe heritability of FTO expression in adipose tissue and skeletal muscle is low and not influenced by obesity-associated FTO genotype. The age-dependent decline in FTO expression is associated with peripheral defects of glucose and fat metabolism.