Genetics of fat tissue accumulation in pigs: a comparative approach

Fatness traits are important in pig production since they influence meat quality and fattening efficiency. On the other hand, excessive fat accumulation in humans has become a serious health problem due to worldwide spread of obesity. Since the pig is also considered as an animal model for numerous human diseases, including obesity and metabolic syndrome, comparative genomic studies may bring new insights into genetics of fatness/obesity. Input of genetic factors into phenotypic variability of these traits is rather high and the heritability coefficient (h ²) of these traits oscillates around 0.5. Genome scanning revealed the presence of more than 500 QTLs for fatness in the pig genome. In addition to QTL studies, many candidate gene polymorphisms have been analyzed in terms of their associations with pig fatness, including genes encoding leptin (LEP) and its receptor (LEPR), insulin-like growth factor 2 (IGF-2), fatty acid-binding proteins (FABP3 andFABP4), melanocortin receptor type 4 (MC4R), and theFTO (fat mass and obesity-associated) gene. Among them, a confirmed effect on pig fatness was found for a well-known polymorphism of theIGF-2 gene. In humans the strongest association with predisposition to obesity was shown for polymorphism of theFTO gene, while in pigs such an association seems to be doubtful. The development of functional genomics has revealed a large number of genes whose expression is associated with fat accumulation and lipid metabolism, so far not studied extensively in terms of the association of their polymorphism with pig fatness. Recently, epigenomic mechanisms, mainly RNA interference, have been considered as a potential source of information on genetic input into the fat accumulation process. The rather limited progress in studies focused on the identification of gene polymorphism related with fatness traits shows that their genetic background is highly complex.     

Extensive changes in innate immune gene expression in obese Göttingen minipigs do not lead to changes in concentrations of circulating cytokines and acute phase proteins

The usefulness of Göttingen minipigs as models for obesity and obesity‐related pathologies is well established. The low‐grade inflammation associated with obesity involves a range of innate immune factors; however, to our knowledge, the impact of obesity on innate immune factor expression has not been studied in Göttingen minipigs. Therefore, we studied the expression of innate immune genes in liver and adipose tissues as well as serum concentrations of cytokines and acute phase proteins in obese vs. lean Göttingen minipigs. In the liver, of 35 investigated genes, the expression of nine was significantly different in obese pigs (three up‐regulated, six down‐regulated). Of 33 genes in adipose tissues, obesity was associated with changed expression of 12 genes in the visceral adipose tissue (VAT) (three up‐regulated), 11 in the abdominal retroperitoneal adipose tissue (RPAT) (seven of these up‐regulated) and eight in the subcutaneous adipose tissue (SAT) from the neck (five of which were up‐regulated). Obesity‐associated expression changes were observed for three genes in all adipose tissues, namely chemokine (C‐C motif) ligand 3‐like 1 (up‐regulated), CD200 molecule (down‐regulated) and interleukin 1 receptor antagonist (up‐regulated) with interleukin 1 receptor antagonist being the most highly regulated gene in both VAT and RPAT. Looking at patterns of expression across the three types of adipose tissues, obesity was associated with an increased number of acute phase proteins differentially expressed between adipose tissues and a decreased tissue‐specific expression of cytokines and chemokines. In contrast to obese humans, no changes in serum concentrations of haptoglobin, C‐reactive protein, serum amyloid A, tumor necrosis factor‐α and interleukin 6 were found in obese Göttingen minipigs.     

Associations of Variants in FTO and Near MC4R With Obesity Traits in South Asian Indians

Recent genome‐wide association studies show that loci in FTO and melanocortin 4 receptor (MC4R) associate with obesity‐related traits. Outside Western populations the associations between these variants have not always been consistent and in Indians it has been suggested that FTO relates to diabetes without an obvious intermediary obesity phenotype. We investigated the association between genetic variants in FTO (rs9939609) and near MC4R (rs17782313) with obesity‐ and type 2 diabetes (T2DM)‐related traits in a longitudinal birth cohort of 2,151 healthy individuals from the Vellore birth cohort in South India. The FTO locus displayed significant associations with several conventional obesity‐related anthropometric traits. The per allele increase is about 1% for BMI, waist circumference (WC), hip circumference (HC), and waist—hip ratio. Consistent associations were observed for adipose tissue‐specific measurements such as skinfold thickness reinforcing the association with obesity‐related traits. Obesity associations for the MC4R locus were weak or nonsignificant but a signal for height (P < 0.001) was observed. The effect on obesity‐related traits for FTO was seen in adulthood, but not at younger ages. The loci also showed nominal associations with increased blood glucose but these associations were lost on BMI adjustment. The effect of FTO on obesity‐related traits was driven by an urban environmental influence. We conclude that rs9939609 variant in the FTO locus is associated with measures of adiposity and metabolic consequences in South Indians with an enhanced effect associated with urban living. The detection of these associations in Indians is challenging because conventional anthropometric obesity measures work poorly in the Indian “thin‐fat” phenotype.     

Joint effects of porcine leptin and leptin receptor polymorphisms on productivity and quality traits

Leptin signalling plays a fundamental role in growth, fatness and body composition. The aim of this study was to investigate the porcine LEP gene sequence in an Iberian × Landrace experimental cross to identify polymorphisms associated with productivity and quality traits. Because of the documented effects on these traits of the LEPR c.1987C>T polymorphism, the LEP and LEPR c.1987C>T polymorphisms and their interactions have been jointly investigated. The LEP gene sequencing has allowed the identification of 39 polymorphisms, eight of which are novel. Three intronic SNPs, LEP g.1382C>T, LEP g.1387C>T and LEP g.1723A>G, have been genotyped, and association analyses have been carried out. Analyses of LEP g.1387C>T, fully linked to LEP g.1382C>T, have revealed additive effects on live and carcass weights and dominant effects on several backfat thickness measurements. Novel effects of both LEP and LEPR polymorphisms on fatty acid composition in subcutaneous fat have been detected, probably mediated through the effects on fatness. The results reported here suggest that the T alleles of both LEP g.1387C>T and LEPR c.1987C>T, which are fixed in the Iberian pigs, would lead to an increase in growth, fatness and saturated fatty acid content in fat, which could be explained by an increased feed intake.     

BAC transgenic zebrafish reveal hypothalamic enhancer activity around obesity associated SNP rs9939609 within the human FTO gene

Single Nucleotide Polymorphisms in FTO intron 1 have been associated with obesity risk, leading to the hypothesis that FTO is the obesity‐related gene. However, other studies have shown that the FTO gene is part of the regulatory domain of the neighboring IRX3 gene and that enhancers in FTO intron 1 regulate IRX3. While Irx3 activity was shown to be necessary in the hypothalamus for the metabolic function of Irx3 in mouse, no enhancers with hypothalamic activity have been demonstrated in the risk‐associated region within FTO. In order to identify potential enhancers at the human FTO locus in vivo, we tested regulatory activity in FTO intron 1 using BAC transgenesis in zebrafish. A minimal gata2 promoter‐GFP cassette was inserted 1.3 kb upstream of the obesity associated SNP rs9939609 in a human FTO BAC plasmid. In addition to the previously identified expression domains in notochord and kidney, human FTO BAC:GFP transgenic zebrafish larvae expressed GFP in the ventral posterior tuberculum, the posterior hypothalamus and the anterior brainstem, which are also expression domains of zebrafish irx3a. In contrast, an in‐frame insertion of a GFP cassette at the FTO start codon resulted in weak ubiquitous GFP expression indicating that the promoter of FTO does likely not react to enhancers located in the obesity risk‐associated region. genesis 53:640–651, 2015. © 2015 The Authors. genesis Published by Wiley Periodicals, Inc.     

FTO gene variants are associated with growth and carcass traits in cattle

An important aim in animal breeding is the improvement of growth and meat quality traits. Previous studies have demonstrated that genetic variants in the fat mass and obesity associated (FTO) gene have a relatively large effect on human obesity as well as on body composition in rodents and, more recently, in livestock. Here, we examined the effects of the FTO gene variants on growth and carcass traits in the Slovenian population of Simmental (SS) and Brown (SB) cattle. To validate and identify new polymorphisms, we used sequencing, PCR‐RFLP analysis and TaqMan assays in the SS breed and FTO gene variants data from the Illumina BovineSNP50 v1 array for the SB breed. Sequencing of the eight samples of progeny‐tested SS sires detected 108 single nucleotide polymorphisms (SNPs) in the bovine FTO gene. Statistical analyses between growth and carcass traits and 34 FTO polymorphisms revealed significant association of FTO variants with lean meat percentage in both breeds. Additionally, FTO SNPs analyzed in SS cattle were associated with fat percentage, bone weight and live weight at slaughter. The FTO gene can thus be regarded as a candidate gene for the marker‐assisted selection programs in our and possibly other populations of cattle. Future studies in cattle might reveal novel roles for the FTO gene in shaping carcass traits in livestock species as well as body composition control in other mammals.     

Association of leptin and leptin receptor gene polymorphisms with systemic lupus erythematosus in a Chinese population

To explore the association of LEP and leptin receptor (LEPR) gene single‐nucleotide polymorphisms (SNPs) with susceptibility to systemic lupus erythematosus (SLE) in a Chinese population. Four LEP SNPs (rs11761556, rs12706832, rs2071045 and rs2167270) and nine LEPR SNPs (rs10749754, rs1137100, rs1137101, rs13306519, rs8179183, rs1805096, rs3790434, rs3806318 and rs7518632) were genotyped in a cohort of 633 patients with SLE and 559 healthy controls. Genotyping of SNPs was performed with improved multiple ligase detection reaction (iMLDR). No significant differences were detected for the distribution of allele and genotype frequencies of all 13 SNPs between patients with SLE and controls. The genotype effects of recessive, dominant and additive models were also analysed, but no significant evidence for association was detected. However, further analysis in patients with SLE showed that the TT genotype and T allele frequencies of the LEP rs2071045 polymorphism were nominally significantly higher in patients with pericarditis (P = 0.012, P = 0.011, respectively). In LEPR, the GA/AA genotype and A allele frequencies of the rs1137100 polymorphism were both nominally associated with photosensitivity in patients with SLE (P = 0.043, P = 0.018, respectively). Moreover, the genotype and allele distribution of rs3806318 were also nominally associated with photosensitivity in patients with SLE (P = 0.013, P = 0.008, respectively). No significant differences in serum leptin levels were observed in patients with SLE with different genotypes. In summary, LEP and LEPR SNPs are not associated with genetic susceptibility to SLE, but may contribute to some specific clinical phenotype of this disease; further studies are necessary to elucidate the exact role of LEP and LEPR genes in the pathogenesis of SLE.     

Nucleotide variability and haplotype heterogeneity at the porcine fat mass and obesity‐associated (FTO) gene

A large number of studies have confirmed that variants within the fat mass and obesity‐associated (FTO) gene are associated with higher obesity risk in humans. We and others have shown that FTO polymorphisms are associated with fat deposition and related traits in several pig populations, thus confirming the role of this gene in fatness across species. However, some differences observed in different pig populations may be derived, at least in part, from genetic heterogeneity at this locus. Here, we characterise the nucleotide variability and haplotype diversity of the porcine FTO gene in breeds having different predispositions to fat deposition traits. We resequenced 4749 bp of coding and non‐coding regions of the porcine FTO gene in 44 pigs of eight different breeds and identified 27 single nucleotide polymorphisms (SNPs) and four insertions/deletions. A positive Tajima's D‐value (P < 0.10) obtained in Italian Duroc pigs may be compatible with putative balancing selection. From the sequenced pig panel, 20 haplotypes were inferred, some of which clustered according to the breed of origin (Meishan and Italian Duroc). Genetic heterogeneity at this locus could complicate the dissection of the effects of this gene on fat deposition and production traits in pigs. This situation resembles, to some extent, what has been reported in humans, thus making the study of the porcine FTO gene variability especially interesting, as it could be used as a model to understand the complex and elusive role of this gene in human obesity.     

Leptin G-2548A and leptin receptor Q223R gene polymorphisms are not associated with obesity in Romanian subjects

We aimed to investigate whether polymorphisms LEP G-2548A and LEPR Q223R in the human leptin (LEP), and leptin receptor (LEPR) genes are associated with obesity and metabolic traits in a sample of Romanian population. Two hundred and two subjects divided in obese (body mass index, BMI ? 30 kg/m²), and non-obese were included in this study. The polymorphisms were genotyped using polymerase chain reaction (PCR) followed by restriction fragment length polymorphism (RFLP) analysis. The results showed no significant differences in LEP and LEPR genotype and allele frequencies between obese and non-obese subjects. Logistic regression analysis showed that LEP -2548GG genotype presented an increased risk of obesity (p = 0.013, OR = 1.003, 95% CI = 1.000-1.007), after adjusting for age and gender. The association analysis with metabolic syndrome quantitative traits showed that homozygous for LEP -2548G allele had significantly higher leptin levels (17.2 ± 6.6 ng/ml vs. 13.2 ± 4.9 ng/ml, p = 0.011), and carriers of R allele had higher levels of triglycerides (p = 0.017) and glucose (p = 0.040), and enhanced systolic (p = 0.015) and diastolic blood pressure (p = 0.026), after adjustment for age, gender, and BMI. These results indicate that LEP G-2548A and LEPR Q223R SNPs may not be considered as genetic risk factors for obesity in a sample of Romanian population. However, LEP -2548GG genotype appear to be important in regulating leptin levels, whereas the LEPR 223R allele might predispose healthy subjects to develop metabolic disturbances.     

Leptin and leptin receptor genetic variants associate with habitual physical activity and the arm body composition response to resistance training

Purpose

We investigated the influence of Leptin (LEP) and leptin receptor (LEPR) SNPs on habitual physical activity (PA) and body composition response to a unilateral, upper body resistance training (RT) program.

Methods

European-derived American volunteers (men = 111, women = 131, 23.4 ± 5.4 yr, 24.4 ± 4.6 kg·m^− 2) were genotyped for LEP 19 G>A (rs2167270), and LEPR 326 A>G (rs1137100), 668 A>G (rs1137101), 3057 G>A (rs1805096), and 1968 G>C (rs8179183). They completed the Paffenbarger PA Questionnaire. Arm muscle and subcutaneous fat volumes were measured before and after 12 wk of supervised RT with MRI. Multivariate and repeated measures ANCOVA tested differences among phenotypes by genotype and gender with age and body mass index as covariates.

Results

Adults with the LEP 19 GG genotype reported more kcal/wk in vigorous intensity PA (1273.3 ± 176.8, p = 0.017) and sports/recreation (1922.8 ± 226.0, p < 0.04) than A allele carriers (718.0 ± 147.2, 1328.6 ± 188.2, respectively). Those with the LEP 19 GG genotype spent more h/wk in light intensity PA (39.7 ± 1.6) than A allele carriers (35.0 ± 1.4, p = 0.03). In response to RT, adults with the LEPR 668 G allele gained greater arm muscle volume (67,687.05 ± 3186.7 vs. 52,321.87 ± 5125.05 mm³, p = 0.01) and subcutaneous fat volume (10,599.89 ± 3683.57 vs. − 5224.73 ± 5923.98 mm³, p = 0.02) than adults with the LEPR 668 AA genotype, respectively.

Conclusion

LEP19 G>A and LEPR 668 A>G associated with habitual PA and the body composition response to RT. These LEP and LEPR SNPs are located in coding exons likely influencing LEP and LEPR function. Further investigation is needed to confirm our findings and establish mechanisms for LEP and LEPR genotype and PA and body composition associations we observed.     

Leptin and leptin receptor gene polymorphisms and their association with plasma leptin levels and obesity in a multi-ethnic Malaysian suburban population

Background

This study was to investigate the prevalence of single nucleotide polymorphisms (SNPs) in leptin gene LEP (A19G and G2548A) and leptin receptor gene LEPR (K109R and Q223R) and their association with fasting plasma leptin level (PLL) and obesity in a Malaysian suburban population in Kampar, Perak.

Methods

Convenience sampling was performed with informed consents, and the study sample was drawn from patients who were patrons of the Kampar Health Clinic. A total of 408 subjects (mean age, 52.4 ± 13.7 years; 169 men, 239 women; 190 obese, 218 non-obese; 148 Malays, 177 ethnic Chinese, 83 ethnic Indians) participated. Socio-demographic data and anthropometric measurements were taken, and genotyping was performed using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP).

Results

The LEP A19G, G2548A and LEPR K109R, Q223R variant allele frequencies were 0.74, 0.67 and 0.61, 0.79, respectively. The genotype and allele distributions of these gene variants were significantly different among ethnic groups, but not among body mass index (BMI) classes. Subjects with LEPR K109 and Q223 allele had significantly higher systolic blood pressure and adiposity indices after adjustment for ethnicity (higher BMI, total body and subcutaneous fat; lower skeletal muscle percentage). Subjects with LEPR 109R allele had lower PLL than their wild-type allele counterparts. The influence of LEP A19G and G2548A SNPs on blood pressures, anthropometrics, and PLL was not evident. Interestingly, synergistic effect of the LEP and LEPR SNPs was observed as subjects homozygous for all four SNPs studied exhibited significantly higher subcutaneous fat and PLL than those with other genotype combinations.

Conclusions

The LEP and LEPR SNPs in this study may not be an obesity marker among Malaysians in this population, but were associated with ethnicity. Our findings suggest that each of these SNPs contributes to minor but significant variation in obesity-related traits and in combination they display synergistic effects on subcutaneous fat and PLL.     

Contribution of Common Genetic Variants to Obesity and Obesity-Related Traits in Mexican Children and Adults

Background

Several studies have identified multiple obesity-associated loci mainly in European populations. However, their contribution to obesity in other ethnicities such as Mexicans is largely unknown. The aim of this study was to examine 26 obesity-associated single-nucleotide polymorphisms (SNP) in a sample of Mexican mestizos.

Methods

9 SNPs in biological candidate genes showing replications (PPARG, ADRB3, ADRB2, LEPR, GNB3, UCP3, ADIPOQ, UCP2, and NR3C1), and 17 SNPs in or near genes associated with obesity in first, second and third wave GWAS (INSIG2, FTO, MC4R, TMEM18, FAIM2/BCDIN3, BDNF, SH2B1, GNPDA2, NEGR1, KCTD15, SEC16B/RASAL2, NPC1, SFRF10/ETV5, MAF, PRL, MTCH2, and PTER) were genotyped in 1,156 unrelated Mexican-Mestizos including 683 cases (441 obese class I/II and 242 obese class III) and 473 normal-weight controls. In a second stage we selected 12 of the SNPs showing nominal associations with obesity, to seek associations with quantitative obesity-related traits in 3 cohorts including 1,218 Mexican Mestizo children, 945 Mexican Mestizo adults, and 543 Indigenous Mexican adults.

Results

After adjusting for age, sex and admixture, significant associations with obesity were found for 6 genes in the case-control study (ADIPOQ, FTO, TMEM18, INSIG2, FAIM2/BCDIN3 and BDNF). In addition, SH2B1 was associated only with class I/II obesity and MC4R only with class III obesity. SNPs located at or near FAIM2/BCDIN3, TMEM18, INSIG2, GNPDA2 and SEC16B/RASAL2 were significantly associated with BMI and/or WC in the combined analysis of Mexican-mestizo children and adults, and FTO locus was significantly associated with increased BMI in Indigenous Mexican populations.

Conclusions

Our findings replicate the association of 8 obesity-related SNPs with obesity risk in Mexican adults, and confirm the role of some of these SNPs in BMI in Mexican adults and children.     

Polymorphisms in 5��-flanking regions of genes encoding adiponectin, leptin, and resistin are not associated with obesity of Polish children and adolescents

Genes encoding adipokines are important functional candidates for development of obesity. In this study we screened for polymorphism 5??-flanking regions of the adiponectin (ADIPOQ), leptin (LEP) and resistin (RETN) genes in a cohort of Polish obese children and adolescents (n = 243) and a control group of non-obese adults (n = 100). Altogether 13 SNPs (single nucleotide polymorphisms) and 1 InDel (insertion/deletion polymorphism) were found. Among them five polymorphisms, localized in the LEP gene, turned out to be novel, but their distribution was insufficient for association studies. We found no consistent evidence for association between obesity and the SNPs demonstrating minor allele frequency (MAF) above 0.2 (ADIPOQ: ?11377C>G, LEP: ?2548C>T, 19A>G, RETN: ?1300G>A, ?1258C>T, ?420C>G). Comparison of polymorphisms distribution in patients and control group suggested association with ADIPOQ ?11377C>G (Pearson test P = 2.76 × 10^?11), however, we did not observe any effect of this polymorphism on BMI or relative BMI (RBMI) within obese patients (P = 0.41). We conclude that the tested SNPs are not useful markers of childhood and adolescence obesity in Polish population.     

Obesity in Insulin Receptor Substrate‐2–Deficient Mice: Disrupted Control of Arcuate Nucleus Neuropeptides

Objectives : Disturbances in insulin signaling have been shown to induce obesity and/or hyperphagia in brain insulin receptor or insulin receptor substrate‐2 (IRS‐2) knockout (KO) mice. This study aimed to examine the central and peripheral mechanisms underlying the phenotype in IRS‐2 KO mice. Research Methods and Procedures : We measured the histological characterization of adipose tissues, mRNA levels of pro‐opiomelanocortin, agouti‐related protein, and neuropeptide Y in the hypothalamus and uncoupling proteins (UCPs) in peripheral tissues of IRS‐2 KO mice. Results : Female IRS‐2 KO mice showed increased daily food intake. Body weight and adiposity were increased in both sexes, although these differences were more pronounced in female than in male IRS‐2 KO mice. Both male and female IRS‐2 KO mice showed decreased UCP1 mRNA expression in brown adipose tissue with defective thermoregulation, and UCP2 mRNA expression was increased in the white adipose tissue of female knockout mice. Furthermore, arcuate nucleus mRNA expression of pro‐opiomelanocortin, was decreased in both male and female IRS‐2 KO mice, whereas expression of agouti‐related protein and neuropeptide Y were increased in female IRS‐2 KO mice. Discussion : In IRS‐2 KO mice, disrupted control of hypothalamic neuropeptide levels and UCP mRNA expression may contribute to the development of obesity.     

Association of FTO Gene Variants With Adiposity in African‐American Adolescents

The prevalence of obesity continues to increase significantly, with the largest rise in the African‐American adolescents. Genetic contributions to obesity are being identified with the advent of genome‐wide association studies (GWAS). Specifically, variants of the fat mass and obesity associated (FTO) gene have been associated with obesity in populations of European descent. The studies in African Americans have been inconclusive. To further evaluate the association of the FTO gene and adiposity in African Americans, we genotyped 47 single‐nucleotide polymorphisms (SNPs), including seven SNPs previously reported to be significant in the literature in a cohort consisting of 561 non‐Hispanic white and 497 African‐American individuals. Analysis of our data showed 17 SNPs to be associated with BMI Z‐score (BMI‐Z) in our study population. The strongest association was found in the African Americans. The most significant SNP was rs8057044, which was associated with BMI‐Z in the African Americans (P = 0.00054). SNP rs9939609 was found to be significant in the non‐Hispanic white population (P = 0.028). Our data confirm the association between FTO and adiposity suggesting that FTO is a childhood obesity susceptibility gene. Our data also identify a novel SNP of the FTO gene (rs8057044) that is associated with measures of adiposity in the African‐American population.     

The Expression of LEP,LEPR, IGF1 and IL10 in Obesity and the Relationship with microRNAs

Obesity is a multifactorial disease, with epigenetic alterations. Have been described modifications in the expression of some microRNAs, and some proteins related to obesity. The objective was to determine and correlate, in obese patients, the gene expression of LEP, LEPR, IGF1, IL10 and of miR-27a, miR-27b, miR-143 and miR-145. RNA was extracted from biopsies of subcutaneous fat, liver and visceral fat of 15 obese subjects submitted to bariatric surgery and of 15 non-obese subjects submitted to cholecystectomy for cDNA synthesis and for RT-PCR. The microRNAs were chosen using the TargetScan software. An increased expression of LEP and IGF1 was detected in the subcutaneous fat of the obese group compared to control, while the expression of IGF1 was higher in the control group than in the obese one. MiRNA-27a had a higher expression in the omentum of the obese patients and there was also a correlation in the expression of miRNA-145 and LEPR in the omentum of this group.