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Aims/Introduction Weight reduction therapy is the primary treatment to prevent complications of obesity, such as lifestyle diseases and cardiovascular disease; however, to date, useful methods and genetic factors for predicting the outcomes of weight reduction therapy in obese patients have not been established. Protein tyrosine phosphatase 1B (PTP1B), a negative regulator for insulin and leptin signaling, potentially modulates glucose and energy homeostasis. This study aimed to investigate the contribution of PTPN1 polymorphisms on weight reduction and diabetes in obese Japanese patients. Materials and Methods PTPN1‐tagged single‐nucleotide polymorphisms (SNPs) rs3787348 and rs6067484 were genotyped in 447 obese Japanese patients from the general population. In this prospective cohort study, all obese patients underwent a 3‐month weight reduction therapy with lifestyle modifications, as recommended by guidelines. Results In obese patients (male/female 196/251, age 50 ± 15 years, body mass index [BMI] 32 ± 6 kg/m2), the minor allele appeared at a frequency of 45.5% in rs3787348 SNP of the PTPN1 gene. The T allele of rs3787348 was significantly associated with a higher BMI (P = 0.041 in the additive model). The patients with the T allele in SNP rs3787348 of PTPN1 had significantly smaller reductions in BMI, bodyweight and waist circumference levels during weight reduction therapy (BMI G/G, −1.9 ± 0.2; G/T, −1.5 ± 0.1; T/T, −1.2 ± 0.1; P = 0.001 in the additive model). Conclusions Our findings show that the SNP rs3787348 in PTPN1 was associated with the effects of weight reduction therapy on BMI and waist circumference among obese Japanese patients., Protein tyrosine phosphatase 1B negatively regulates insulin and leptin signaling. The rs3787348 in PTPN1 was associated with the effects of weight reduction therapy. The single‐nucleotide polymorphism of PTPN1 might predict the efficacy of weight reduction in obese patients.

The progression of nonalcoholic fatty liver disease (NAFLD) is affected by epigenetics. We performed differentially methylated region (DMR) and co-methylation analyses to identify DMR networks associated with the progression of NAFLD. DMRs displaying differences in multiple consecutive differentially methylated CpGs between mild and advanced NAFLD were extracted. The average values of topological overlap measures for the CpG matrix combining two different DMRs were calculated and two DMR networks that strongly correlated with the stages of fibrosis were identified. The annotated genes of one network included genes involved in transcriptional regulation, cytoskeleton organization, and cellular proliferation. The annotated genes of the second network were primarily associated with metabolic pathways. The CpG methylation levels in these networks were strongly affected by age and fasting plasma glucose levels, which may be important co-regulatory factors. The methylation status of five DMRs in the second network was reversible following weight loss. Our results suggest that CpG methylation in DMR networks is regulated concomitantly via aging and hyperglycemia and plays important roles in hepatic metabolic dysfunction, fibrosis, and potential tumorigenesis, which occur during the progression of NAFLD. By controlling weight and blood glucose levels, the methylation of DMRs in the second network may be reduced.

Aim Nonalcoholic fatty liver disease (NAFLD) progresses because of the interaction between numerous genes. Thus, we performed weighted gene co-expression network analysis to identify core gene networks and key genes associated with NAFLD progression. Methods We enrolled 39 patients with mild NAFLD (fibrosis stages 0 to 2) and 21 with advanced NAFLD (fibrosis stages 3 or 4). Total RNA was extracted from frozen liver biopsies, and sequenced to capture a large dynamic range of expression levels. Results A total of 1,777 genes differentially expressed between mild and advanced NAFLD (q-value

Background: Retinitis pigmentosa (RP) is a heterogeneous group of disorders that have a high genotypic diversity. Therefore, it is difficult to presume the genotype and/or prognosis from th...

Bardet–Biedl syndrome (BBS), characterized by rod-cone dystrophy, postaxial polydactyly, central obesity, hypogonadism, renal abnormalities, and mental retardation, is a rare autosomal recessive disorder. To date, 21 causative genes have been reported. Here we describe a Japanese BBS patient with a novel compound heterozygous mutation in TTC8. To the best of our knowledge, this is the first description of a BBS patient with a mutation in the TTC8 gene in Japan.

Aim There are a considerable number of patients with non-obese non-alcoholic fatty liver disease (NAFLD). However, the clinical characteristics of non-obese NAFLD is not fully understood. We investigated genetic and other clinical parameters in non-obese and obese NAFLD. Methods The single nucleotide polymorphism rs738409 in the patatin-like phospholipase 3 gene (PNPLA3) was genotyped by the Invader assay in 540 NAFLD patients (134 non-obese and 406 obese) and 1012 control subjects (782 non-obese and 230 obese). All NAFLD patients underwent liver biopsy. Odds ratios were calculated by multiple logistic regression analysis using age, sex, body mass index (BMI), type 2 diabetes mellitus (T2DM) and rs738409 genotype as explanatory variables. Results Non-obese NAFLD subjects had a higher rs738409 GG genotype than obese NAFLD. Multiple logistic regression analysis indicated that the odds ratios of T2DM and rs738409 GG genotype for NAFLD were higher in non-obese than in obese groups. In non-obese NAFLD, rs738409 GG genotype was associated with lobular inflammation, hepatocyte ballooning and NAFLD activity score. In obese NAFLD, BMI and T2DM but not rs738409 GG genotype were associated with severity of histology. Conclusion We demonstrated that the risk factors for the development and progression of NAFLD were different between non-obese and obese patients and that PNPLA3 rs738409 was strongly associated with the development and progression of non-obese NAFLD.

AIM Visceral fat accumulation contributes to the development of metabolic syndrome. As visceral fat accumulation increases, adiponectin levels decrease; therefore, adiponectin provides a link between visceral fat accumulation and metabolic disorders. Genome-wide association studies (GWASs) have identified genetic variations in the cadherin 13 (CDH13) gene that are associated with adiponectin levels. METHODS We investigated whether single nucleotide polymorphisms (SNPs) in CDH13 was associated with adiponectin levels and metabolic syndrome traits independent of the visceral fat area (VFA), as measured using computed tomography (CT) in 945 Japanese individuals. RESULTS We found that three CDH13 SNPs reported by recent GWASs (i.e., rs3865188, rs4783244, and rs12051272) were significantly associated with higher adiponectin levels (P ＜ 1 × 10 (-14)), even after adjustment for VFA. However, these adiponectin-inducing alleles of CDH13 SNPs were significantly associated with traits consistent with deteriorating metabolic symptoms, such as higher fasting insulin, homeostasis model assessment-insulin resistance (HOMA-IR) scores, and triglycerides and lower high-density lipoprotein (HDL)-cholesterol levels, similar to increasing VFA and decreasing adiponectin levels. CONCLUSION These results suggested that CDH13 SNPs cause an adiponectin-resistant status to compensate for increasing adiponectin levels and could result in the deterioration of metabolic syndrome traits.

Aim A genome-wide association study revealed that the single nucleotide polymorphism (SNP) rs738409 in the patatin-like phospholipase 3 gene (PNPLA3) was strongly associated with non-alcoholic fatty liver disease (NAFLD). Recent pilot studies investigated the effects of dipeptidyl peptidase-4 inhibitors on liver function and glucose metabolism in NAFLD with type 2 diabetes mellitus (DM). We herein evaluated the efficacy of alogliptin in NAFLD patients with type 2 DM as well as the relationship between genotypes at rs738409 in PNPLA3 and treatment efficacy. Methods Forty-one biopsy-proven NAFLD patients with type 2 DM treated with 25 mg/day alogliptin were retrospectively enrolled. SNP rs738409 in PNPLA3 was present in all patients. Clinical data were measured before and after the treatment. Results Average hemoglobin A1c (HbA1c) levels mostly remained unchanged. Moreover, significant changes were not noted in the levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) during the follow-up period. A positive correlation was observed between improvements in HbA1c (ΔHbA1c) levels and changes in AST (ΔAST) and ALT (ΔALT) levels (r = 0.325 and 0.439, respectively). Patients with the risk allele (G-allele) showed more positive correlation between ΔHbA1c and changes in transaminase. Furthermore, improvements in the levels of total cholesterol, triglycerides and hyaluronic acid were significantly greater in G-allele patients in the weight loss group. Conclusion The treatment of NAFLD with type 2 DM with alogliptin contributed to the amelioration of NAFLD. Our results suggested that differences in the PNPLA3 risk allele affected the therapeutic effects of this treatment.

Visceral fat accumulation contributes to the development of insulin resistance, leading to metabolic syndrome. Adiponectin provides a link between visceral fat accumulation and insulin resistance. In addition to environmental factors, genetic factors play important roles in visceral fat accumulation and circulating adiponectin levels. Genome-wide association studies (GWASs) have identified genetic variations in the adiponectin, C1Q and collagen domain containing (ADIPOQ) gene that are associated with adiponectin levels. In this study, we investigated whether ADIPOQ single nucleotide polymorphisms (SNPs) were associated with visceral fat accumulation and insulin resistance. We measured the visceral fat area (VFA) by computed tomography (CT) and examined the presence of the insulin resistance-related phenotype (fasting plasma glucose, fasting insulin, and homeostasis model assessment-insulin resistance [HOMA-IR]) in a set of Japanese individuals (731 men and 864 women) who were genotyped for seven ADIPOQ SNPs reported by recent GWASs (namely, rs6810075, rs10937273, rs1648707, rs864265, rs182052, rs17366568, and rs6773957). SNPs associated with the phenotype (P < 0.05) were then evaluated by association analysis using a second set of the study participants (383 men and 510 women). None of the SNPs was associated with body mass index (BMI) or VFA in men or women. However, the adiponectin-decreasing alleles of rs10937273 and rs1648707 were significantly associated with HOMA-IR (P = 0.0030 and P = 0.00074, respectively) in women, independently of BMI. These SNPs were significantly associated with decreased adiponectin levels in women. Our results suggested that rs10937273 and rs1648707 may affect insulin sensitivity by regulating adiponectin production by adipose tissue in women.

Aim The progression of non-alcoholic fatty liver disease (NAFLD) is affected by epigenetics. We undertook co-methylation and differentially methylated region (DMR) analyses to identify the genomic region that is under epigenetic regulation during NAFLD progression. Methods We collected liver biopsy specimens from 60 Japanese patients with NAFLD and classified these into mild (fibrosis stages 0-2) or advanced (fibrosis stages 3-4) NAFLD. We carried out a genome-wide DNA methylation analysis and identified the differentially methylated CpGs between mild and advanced NAFLD. Differentially methylated regions with multiple consecutive differentially methylated CpGs between mild and advanced NAFLD were extracted. Results Co-methylation analysis showed that individual differentially methylated CpG sites were clustered into three modules. The CpG sites clustered in one module were hypomethylated in advanced NAFLD and their annotated genes were enriched for "immune system" function. The CpG sites in another module were hypermethylated and their annotated genes were enriched for "mitochondria" or "lipid particle", and "lipid metabolism" or "oxidoreductase activity". Hypomethylated DMRs included tumorigenesis-related genes (FGFR2, PTGFRN, and ZBTB38), the expressions of which are upregulated in advanced NAFLD. Tumor suppressor MGMT had two DMRs and was downregulated. Conversely, FBLIM1 and CYR61, encoding proteins that reduce cell proliferation, showed hypomethylated DMRs and were upregulated. Expression of the antioxidant gene NQO1 was upregulated, with a hypomethylated DMR. The DMR containing cancer-related MIR21 was hypomethylated in advanced NAFLD. Conclusions Co-methylation and DMR analyses suggest that the NAFLD liver undergoes mitochondrial dysfunction, decreased lipid metabolism, and impaired oxidoreductase activity, and acquires tumorigenic potential at the epigenetic level.

Objective We examined associations between the fat-mass and obesity-associated (FTO) gene (rs9939609) and any weight change over a 5-year period following a 14-week lifestyle intervention among middle-aged Japanese women. Materials/Methods One hundred twenty-eight Japanese women (BMI > 25 kg/m2) participated in a 14-week weight loss intervention between 2004 and 2006. Of the participants, 62 consented to the 5-year follow-up measurement session. Of these women, 47 women who achieved a weight loss of at least 10% from their baseline values during the 14-week intervention were included in the analysis. Body weight, body fat, abdominal fat assessed by CT scans, and metabolic risk factors (i.e., blood pressure, lipids, and glucose) were measured at baseline, post-intervention, and at the 5-year follow-up. Results During the 5-year non-intervention period, increases in body weight, fat mass, total abdominal fat, and subcutaneous abdominal fat were significantly greater in subjects with the homozygous minor allele (AA genotype, n = 4; 8.5%) than in those with the homozygous major allele (TT genotype, n = 31; 66.0%) or heterozygous allele (TA genotype, n = 12; 25.5%). In multiple regression analyses, the variation in rs9939609 was a significant and independent predictor (P < 0.001) for regaining weight during the 5-year follow-up. Conclusions Our data suggest that Japanese women with the risk allele (AA) of rs9939609 may have more difficulty preventing fat gain from reoccurring after weight loss intervention than women with the other genotypes.

Recent studies have suggested that fat mass and obesity-associated gene (FTO) may predispose individuals to develop hypertension. However, the results have been inconsistent. We performed a meta-analysis to investigate the relationship of FTO gene variant with risk of hypertension and influence of body mass index (BMI) on this risk.A systematic literature search in PubMed, Embase and ISI web of science databases was performed to identify eligible published literatures. Pooled odds ratios (ORs) with 95% confidence intervals (CIs) were calculated.A total of seven studies comprising 57,464 hypertensive cases and 41,256 controls met the inclusion criteria and were included in the meta-analysis. The FTO gene variant(s) showed significant association with the risk of hypertension (OR=1.16, 95% CI=1.07-1.25, P0.001) which disappeared on adjustment for BMI (OR=1.04, 95% CI=0.98-1.10, P=0.162). In addition, stratified analysis demonstrated a significant association of the FTO variant with the risk of hypertension in obese subjects (OR=1.10, 95% CI=1.01-1.19, P=0.032) but not in non-obese individuals (OR=1.00, 95% CI=0.97-1.03, P=0.832). Subgroup analysis based on ethnicity showed significant association between FTO variant and hypertension in both European (OR=1.07, 95% CI=1.01-1.14, P=0.028) and Asian populations (OR=1.37, 95% CI=1.23-1.53, P0.001). However, the association remained significant only in Asians (OR=1.17, 95% CI=1.01-1.35, P=0.035) but not in the Europeans (OR=1.02, 95% CI=0.97-1.07, P=0.390) on adjustment for BMI.The present meta-analysis confirms that FTO genotype mediates obesity-related hypertension.

The genomic regions containing PNPLA3, SAMM50 and PARVB are susceptibility loci for the development and progression of nonalcoholic fatty liver disease (NAFLD). In order to search for all common variations in this region, we amplified the genomic DNA of 28 NAFLD patients by long-range PCR, covering the entire susceptibility region and sequenced the DNA using indexed multiplex next-generation sequencing. We found 329 variations, including four novel variations. Fine mapping of variations including insertion/deletions was performed for 540 NAFLD patients (488 with nonalcoholic steatohepatitis (NASH) and 52 with simple steatosis) and 1012 control subjects. HaploView analysis showed that linkage disequilibrium (LD) block 1 and 2 occurred in PNPLA3, block 3 in SAMM50 and block 4 in PARVB. Variations in LD blocks 1-4 were significantly associated with NAFLD as compared with control subjects (P, Published online 13 March 2014

In several genome-wide association studies, nonalcoholic fatty liver disease and alanine aminotransferase susceptibility variants have been identified in several genes, including LYPLAL1, ZP4, GCKR, HSD17B13, PALLD, PPP1R3B, FDFT1, TRIB1, COL13A1, CPN1, ERLIN1, CWF19L1, EFCAB4B, PZP, and NCAN. To investigate the relationship between these genes and nonalcoholic fatty liver disease in the Japanese population, we genotyped 540 patients and 1012 control subjects for 18 variations. We performed logistic regression analyses to characterize the association between the tested variations and nonalcoholic fatty liver disease. Metabolic syndrome and histological traits were also analyzed by linear regression. We also examined GCKR rs780094, TRIB1 rs2954021, and PNPLA3 rs738409 for epistatic effects. The A-allele of rs780094 in GCKR (P = 0.0024) and the A-allele of rs2954021 TRIB1 (P = 4.5×10-5) were significantly associated with nonalcoholic fatty liver disease. GCKR rs780094 was also associated with decreased plasma glucose, and increased triglycerides in the patient and control groups. GCKR rs780094 was also associated with an increased ratio of visceral to subcutaneous fat area in the patients with nonalcoholic fatty liver disease. Variations in GCKR, TRIB1, and PNPLA3 independently influenced nonalcoholic fatty liver disease and had no epistatic effects. Our data suggest variations in GCKR and TRIB1 are involved in the development of nonalcoholic fatty liver disease.

The predominant risk factor of metabolic syndrome is intra-abdominal fat accumulation, which is determined by waist circumference, waist-hip ratio measurements and visceral fat area (VFA); the latter can be accurately measured by performing computed tomography (CT). In addition to environmental factors, genetic factors play an important role in obesity and fat distribution. New genetic loci associated with body mass index (BMI) and adiposity have been identified by genome-wide association studies (GWASs). This study utilized CT to investigate whether single nucleotide polymorphisms (SNPs) that confer susceptibility to higher BMI are associated with VFA, subcutaneous fat area (SFA), and the ratio of VFA to SFA (V/S ratio). We measured the VFA and SFA of 1424 obese Japanese subjects (BMI ≥ 25 kg/m(2), 635 men and 789 women) who were genotyped for 13 single nucleotide polymorphisms (SNPs) reported by recent GWASs, namely, TNNI3K rs1514175, PTBP2 rs1555543, ADCY3 rs713586, IRS1 rs2943650, POC5 rs2112347, NUDT3 rs206936, LINGO2 rs10968576, STK33 rs4929949, MTIF3 rs4771122, SPRY2 rs534870, MAP2K5 rs2241423, QPCTL rs2287019, and ZC3H4 rs3810291. The G-allele of NUDT3 rs206936 was significantly associated with increased BMI (P = 5.3 × 10(-5)) and SFA (P = 0.00039) in the obese Japanese women. After adjustment with BMI, the association between rs206936 and SFA was not observed. This significant association was not observed in the men. The other SNPs analyzed were not significantly associated with BMI, VFA, SFA, or V/S ratio. Our results suggest that NUDT3 rs206936 is associated with BMI in Japanese women.

AIM Visceral fat accumulation plays an integral role in morbidity and mortality rates by increasing the risk of developing metabolic disorders such as type 2 diabetes, dyslipidemia, and hypertension. New genetic loci associated with fat distribution, measured by waist-hip ratios and computed tomography (CT), have recently been identified by genome-wide association studies in European-descent populations. This study used CT to investigate whether single nucleotide polymorphisms (SNPs) that confer susceptibility to fat distribution are associated with visceral fat area (VFA) and subcutaneous fat area (SFA) in the Japanese population. METHODS We measured the VFAs and SFAs of 1424 obese Japanese subjects (BMI≥25 kg/m(2), 635 men and 789 women) that were genotyped at 15 SNPs, namely, TBX15 rs984222, DNM3 rs1011731, LYPLAL1 rs4846567, GRB14 rs10195252, NISCH rs6784615, ADAMTS9 rs6795735, CPEB4 rs6861681, LY86 rs1294421, VEGFA rs6905288, RSPO3 rs9491696, NFE2L3 rs1055144, ITPR2 rs718314, HOXC13 rs1443512, ZNRF3 rs4823006 and THNSL2 rs1659258. RESULTS The G-allele of LYPLAL1 rs4846567 was borderline associated with an increased ratio of VFA to SFA (V/S ratio; p= 0.0020). LYPLAL1 rs4846567 had a stronger effect on the V/S ratio in women (p= 0.0078) than in men (p= 0.12); however, neither result was significant after Bonferroni correction for multiple comparisons. NISCH rs6784615 was nominally associated with increased VFA (p=0.040) and V/S ratio (p= 0.020). The other SNPs analyzed were not significantly associated with body mass index (BMI), VFA, or SFA. CONCLUSION Our results suggest that LYPLAL1 rs4846567 and NISCH rs6784615 may influence fat distribution in the Japanese population.

The predominant risk factor of metabolic syndrome is intra-abdominal fat accumulation, which is determined by waist circumference, waist-hip ratio measurements and visceral fat area (VFA); the latter can be accurately measured by performing computed tomography (CT). In addition to environmental factors, genetic factors play an important role in obesity and fat distribution. New genetic loci associated with body mass index (BMI) and adiposity have been identified by genome-wide association studies (GWASs). This study utilized CT to investigate whether single nucleotide polymorphisms (SNPs) that confer susceptibility to higher BMI are associated with VFA, subcutaneous fat area (SFA), and the ratio of VFA to SFA (V/S ratio). We measured the VFA and SFA of 1424 obese Japanese subjects (BMI ≥ 25 kg/m(2), 635 men and 789 women) who were genotyped for 13 single nucleotide polymorphisms (SNPs) reported by recent GWASs, namely, TNNI3K rs1514175, PTBP2 rs1555543, ADCY3 rs713586, IRS1 rs2943650, POC5 rs2112347, NUDT3 rs206936, LINGO2 rs10968576, STK33 rs4929949, MTIF3 rs4771122, SPRY2 rs534870, MAP2K5 rs2241423, QPCTL rs2287019, and ZC3H4 rs3810291. The G-allele of NUDT3 rs206936 was significantly associated with increased BMI (P = 5.3 × 10(-5)) and SFA (P = 0.00039) in the obese Japanese women. After adjustment with BMI, the association between rs206936 and SFA was not observed. This significant association was not observed in the men. The other SNPs analyzed were not significantly associated with BMI, VFA, SFA, or V/S ratio. Our results suggest that NUDT3 rs206936 is associated with BMI in Japanese women.

Non-alcoholic fatty liver disease (NAFLD) progresses because of the interaction between numerous genes. Thus, we carried out a weighted gene coexpression network analysis to identify core gene networks and key genes associated with NAFLD progression.We enrolled 39 patients with mild NAFLD (fibrosis stages 0-2) and 21 with advanced NAFLD (fibrosis stages 3-4). Total RNA was extracted from frozen liver biopsies, and sequenced to capture a large dynamic range of expression levels.A total of 1777 genes differentially expressed between mild and advanced NAFLD (q-value0.05) clustered into four modules. One module was enriched for genes that encode cell surface or extracellular matrix proteins, and are involved in cell adhesion, proliferation, and signaling. This module formed a scale-free network containing four hub genes (PAPLN, LBH, DPYSL3, and JAG1) overexpressed in advanced NAFLD. PAPLN is a component of the extracellular matrix, LBH and DPYSL3 are reported to be tumor suppressors, and JAG1 is tumorigenic. Another module formed a random network, and was enriched for genes that accumulate in the mitochondria. These genes were downregulated in advanced NAFLD, reflecting impaired mitochondrial function. However, the other two modules did not form unambiguous networks. KEGG analysis indicated that 71 differentially expressed genes were involved in "pathways in cancer". Strikingly, expression of half of all differentially expressed genes was inversely correlated with methylation of CpG sites (q-value0.05). Among clinical parameters, serum type IV collagen 7 s was most strongly associated with the epigenetic status in NAFLD.Newly identified core gene networks suggest that the NAFLD liver undergoes mitochondrial dysfunction and fibrosis, and acquires tumorigenic potential epigenetically. Our data provide novel insights into the pathology and etiology of NAFLD progression, and identify potential targets for diagnosis and treatment.

Visceral fat accumulation has an important role in increasing the morbidity and mortality rates, by increasing the risk of developing several metabolic disorders, such as type 2 diabetes, dyslipidemia and hypertension. New genetic loci that are associated with increased systolic and diastolic blood pressures have been identified by genome-wide association studies in Caucasian populations. This study investigates whether single nucleotide polymorphisms (SNPs) that confer susceptibility to high blood pressure are also associated with visceral fat obesity. We genotyped 1279 Japanese subjects (556 men and 723 women) who underwent computed tomography for measuring the visceral fat area (VFA) and subcutaneous fat area (SFA) at the following SNPs: FGF5 rs16998073, CACNB2 rs11014166, C10orf107 rs1530440, CYP17A1 rs1004467, NT5C2 rs11191548, PLEKHA7 rs381815, ATP2B1 rs2681472 and rs2681492, ARID3B rs6495112, CSK rs1378942, PLCD3 rs12946454, and ZNF652 rs16948048. In an additive model, risk alleles of the CYP17A1 rs1004467 and NT5C2 rs11191548 were found to be significantly associated with reduced SFA (P=0.00011 and 0.0016, respectively). When the analysis was performed separately in men and women, significant associations of rs1004467 (additive model) and rs11191548 (recessive model) with reduced VFA (P=0.0018 and 0.0022, respectively) and SFA (P=0.00039 and 0.00059, respectively) were observed in women, but not in men. Our results suggest that polymorphisms in the CYP17A1 and NT5C2 genes influence a reduction in both visceral and subcutaneous fat mass in Japanese women.

The predominant risk factor of metabolic syndrome is intra-abdominal fat accumulation, which is determined by waist circumference and waist-hip ratio measurements and visceral fat area (VFA) that is measured by computed tomography (CT). There is evidence that waist circumference and waist-hip ratio in the Caucasian population are associated with variations in several genes, including neurexin 3 (NRXN3), transcription factor AP-2β (TFAP2B), methionine sulfoxide reductase A (MSRA), lysophospholipase-like-1 (LYPLAL1), fat mass and obesity associated (FTO) and melanocortin 4 receptor (MC4R) genes. To investigate the relationship between VFA and subcutaneous fat area (SFA) and these genes in the recruited Japanese population, we genotyped 8 single-nucleotide polymorphisms (SNPs) in these 6 genes from 1228 subjects. Multiple regression analysis revealed that gender, age, and rs1558902 and rs1421085 genotypes (additive model) in FTO were significantly associated with body mass index (BMI; P=0.0039 and 0.0039, respectively), SFA (P=0.0027 and 0.0023, respectively) and VFA (P=0.045 and 0.040, respectively). However, SNPs in other genes, namely, NRXN3, TFAP2B, MSRA, LYPLAL1 and MC4R were not significantly associated with BMI, SFA or VFA. Our data suggest that some SNPs, which were identified in genome-wide studies in the Caucasians, also confer susceptibility to fat distribution in the Japanese subjects.

Orosomucoid (ORM), also called alpha-1 acid glycoprotein, is an abundant plasma protein that is an immunomodulator induced by stressful conditions such as infections. In this study, we reveal that Orm is induced selectively in the adipose tissue of obese mice to suppress excess inflammation that otherwise disturbs energy homeostasis. Adipose Orm levels were elevated by metabolic signals, including insulin, high glucose, and free fatty acid, as well as by the proinflammatory cytokine tumor necrosis factor-alpha, which is found in increased levels in the adipose tissue of morbid obese subjects. In both adipocytes and macrophages, ORM suppressed proinflammatory gene expression and pathways such as NF-kappaB and mitogen-activated protein kinase signalings and reactive oxygen species generation. Concomitantly, ORM relieved hyperglycemia-induced insulin resistance as well as tumor necrosis factor-alpha-mediated lipolysis in adipocytes. Accordingly, ORM improved glucose and insulin tolerance in obese and diabetic db/db mice. Taken together, our results suggest that ORM integrates inflammatory and metabolic signals to modulate immune responses to protect adipose tissue from excessive inflammation and thereby from metabolic dysfunction.

There is evidence that the obesity phenotype in the Caucasian populations is associated with variations in several genes, including neuronal growth regulator 1 (NEGR1), SEC16 homolog B (SCE16B), transmembrane protein 18 (TMEM18), ets variant 5 (ETV5), glucosamine-6-phosphate deaminase 2 (GNPDA2), prolactin (PRL), brain-derived neurotrophic factor (BDNF), mitochondrial carrier homolog 2 (MTCH2), Fas apoptotic inhibitory molecule 2 (FAIM2), SH2B adaptor protein 1 (SH2B1), v-maf musculoaponeurotic fibrosarcoma oncogene homolog (MAF), Niemann-Pick disease, type C1 (NPC1), melanocortin 4 receptor (MC4R) and potassium channel tetramerisation domain containing 15 (KCTD15). To investigate the relationship between obesity and these genes in the Japanese population, we genotyped 27 single-nucleotide polymorphisms (SNPs) in 14 genes from obese subjects (n=1129, body mass index (BMI) > or =30 kg m(-2)) and normal-weight control subjects (n=1736, BMI

Several studies indicate that expression of the peroxisome proliferator-activated receptor gamma (PPARG) gene is influenced by calorie restriction. The aim of this study was to investigate whether PPARG gene variations are associated with weight reduction and changes in coronary heart disease (CHD) risk factors in response to a 14-week calorie restriction. In total, 95 middle-aged, Japanese women (BMI>or=25 kg/m2) enrolled as subjects for 14 weeks and attended weekly dietary lectures instructing them on how to consume a nutritionally balanced diet of 1,200 kcal/day. Eight single-nucleotide polymorphisms (SNPs) in the PPARG gene (rs1801282 (Pro/Ala), rs2292101, rs2959272, rs1386835, rs709158, rs1175540, rs1175544, and rs1797912) were analyzed. Body weight decreased significantly (-7.7+/-3.1 kg; -11.3+/-4.4%) during the intervention. Six PPARG SNPs (rs2959272, rs1386835, rs709158, rs1175540, rs1175544, and rs1797912) were significantly associated with the weight reduction, with rs1175544 having the strongest association (P=0.004). No differences across the rs1175544 genotypes were observed in any of the blood analyses or in blood pressure. In a multiple regression analysis, the rs1175544 genotypes accounted for 7% of the total weight reduction variance. These data suggest that one SNP of the PPARG genotype accounted for a significant portion of the total body weight reduction variance in response to a short-term intervention consisting of calorie restriction; however, no relationship was found between these SNPs and the changes in CHD risk factors which accompanied weight loss.

Background: Nonalcoholic fatty liver disease (NAFLD) is one of the most common causes of chronic liver injury in many countries. Genetic factors are important for the development of NAFLD, as well as environmental factors. Recently an angiotensin II type 1 receptor (AGTR1) has been recognized as important in the aetiology of fibrosis in the liver. Objective: In this study we investigated the association between angiotensin II type 1 receptor gene polymorphism (ATGR1) and NAFLD. Methods: One hundred and sixty-seven NAFLD patients [106 with nonalcoholic steatohepatitis (NASH) and 61 with simple steatosis] with a positive diagnosis by liver biopsy and 435 healthy control subjects were recruited in this study. Results: We investigated 12 single nucleotide polymorphisms (SNPs) of the ATGR1 gene, among which rs3772622 showed the lowest P-value of allele frequency model (P=0.0000012) with an odds ratio (95% confidence interval) of 1.95 (1.49–2.55). Five SNPs (rs3772622, rs3772633, rs2276736, rs3772630 and rs3772627) were significantly associated with NAFLD, even when the most conservative Bonferroni's correction was applied. Linkage disequilibrium analysis revealed that SNP rs3772622 and another four SNPs (rs3772633, rs2276736, rs3772630 and rs3772627) were in the same block. We investigated the association between rs3772622 genotypes and the fibrosis index. The results of the analysis revealed an additive increase of the fibrosis index in the patients with the A allele of rs3772622. Conclusions: This is the first report to demonstrate the genetic variations in ATGR1 that may influence the risk of NAFLD and liver fibrosis in NAFLD.

Functional variations in the secretogranin III (SCG3) gene are associated with susceptibility to obesity. SCG3 forms secretory granules with orexin, melanin-concentrating hormone (MCH), neuropeptide Y (NPY), and POMC in the hypothalamus. In this study, we screened proteins for SCG3-binding activity and identified secretogranin II (SCG2) using a yeast two-hybrid system. Immunoprecipitation revealed that SCG2 interacts with SCG3. In situ hybridization and immunohistochemistry indicated that SCG2 was highly expressed in the lateral hypothalamic area, paraventricular nucleus, and arcuate nucleus of the hypothalamus. Double-labeling immunohistochemical analysis demonstrated that SCG2 was expressed in orexin-, MCH-, NPY-, and POMC-expressing neurons. SCG2 was also coexpressed with SCG3. Upon introduction into neuroblastoma cells, SCG2 was expressed in the cytosol and formed granule-like structures with SCG3, orexin, NPY, or POMC. SCG3 bound to POMC; however, it did not bind to orexin, MCH, or NPY. By contrast, SCG2 formed aggregates with orexin, MCH, NPY, and POMC. SCG2 may act as a hormone carrier for orexin, MCH, NPY, and POMC by binding with SCG3, which targets proteins to the secretory granules. SCG2 mRNA levels increased along with those of SCG3, orexin, MCH, and NPY after a 24-h fast, suggesting that the SCG2/SCG3 system may respond in an adaptive manner to acute body weight changes. However, this SCG2/SCG3 system appears to be unresponsive to chronic body weight changes, such as diet-induced obesity or obesity in ob/ob mice. We suggest that SCG2, as well as SCG3, may be a potential regulator of food intake based on its capacity to accumulate appetite-related hormones into secretory granules.

Genetic factors are important in the development of metabolic syndrome. However, the genetic background of metabolic syndrome remains unclear. We screened polymorphisms in 85 obesity-related genes to determine which may be associated with metabolic syndrome. A total of 336 single-nucleotide polymorphisms (SNPs) in 85 genes selected from the JSNP database were genotyped. We conducted case-control association analyses using patients with metabolic syndrome (n=1080) and control individuals (n=528) who had no risk of the metabolic syndrome. Three SNPs in the McKusick-Kaufman syndrome (MKKS) gene were significantly related to metabolic syndrome by case-control association study; rs1545 (odds ratio (OR) adjusted for age and gender, 1.45; 95% confidence interval (CI), 1.21-1.74; P=0.000043 (additive model)); rs1547 (OR, 1.45; 95% CI, 1.21-1.74; P=0.000041); and rs2294901 (OR, 1.46; 95% CI, 1.22-1.75; P=0.000033). We selected five tag SNPs (rs2294901, rs221667, rs6133922, rs6077785 and rs6108572) in the MKKS gene. They were in one linkage disequilibrium (LD) block and rs6133922 (P=0.00042), rs6077785 (P=0.000013) and rs6108572 (P=0.000019) as well as rs2294901 were significantly associated with metabolic syndrome. TGAAA haplotype was protective against the metabolic syndrome (P=0.0074), and CCGTT haplotype was susceptible (P=0.00070) to the metabolic syndrome. Our data suggest that genetic variations at MKKS gene influence the risk of metabolic syndrome.

The single nucleotide polymorphism (SNP) rs7566605 in the upstream region of the insulin-induced gene 2 (INSIG2) is associated with the obesity phenotype in many Caucasian populations. In Japanese, this association with the obesity phenotype is not clear. To investigate the relationship between rs7566605 and obesity in Japanese, we genotyped rs7566605 from severely obese subjects [n = 908, body mass index (BMI) ≥ 30 kg/m2] and normal-weight control subjects (n = 1495, BMI < 25 kg/m2). A case–control association analysis revealed that rs7566605 was significantly associated with obesity in Japanese. The P value in the minor allele recessive mode was 0.00020, and the odds ratio (OR) adjusted for gender and age was 1.61 [95% confidential interval (CI) = 1.24–2.09]. Obesity-associated phenotypes, which included the level of BMI, plasma glucose, hemoglobin A1c, total cholesterol, triglycerides, high-density lipoprotein (HDL) cholesterol, and blood pressure, were not associated with the rs7566605 genotype. Thus, rs7566605 in the upstream region of the INSIG2 gene was found to be associated with obesity, i.e., severe obesity, in Japanese.

Context: Genetic factors are important for the development of obesity. However, the genetic background of obesity still remains unclear. Objective: Our objective was to search for obesity-related genes using a large number of gene-based single-nucleotide polymorphisms (SNPs). Design and Setting: We conducted case-control association analyses using 94 obese patients and 658 controls with 62,663 SNPs selected from the SNP database. SNPs that possessed P ≤ 0.02 were further analyzed using 796 obese and 711 control subjects. One SNP (rs3764220) in the secretogranin III (SCG3) gene showed the lowest P value (P = 0.0000019). We sequenced an approximately 300-kb genomic region around rs3764220 and discovered SNPs for haplotype analyses. SCG3 was the only gene within a haplotype block that contained rs3764220. The functions of SCG3 were studied. Patients: Obese subjects (body mass index ≥ 30 kg/m2, n = 890) and control subjects (general population; n = 658, body mass index ≤ 25kg/m2; n = 711) were recruited for this study. Results: Twelve SNPs in the SCG3 gene including rs3764220 were in almost complete linkage disequilibrium and significantly associated with an obesity phenotype. Two SNPs (rs16964465, rs16964476) affected the transcriptional activity of SCG3, and subjects with the minor allele seemed to be resistant to obesity (odds ratio, 9.23; 95% confidence interval, 2.77–30.80; χ2 = 19.2; P = 0.0000067). SCG3 mRNA and immunoreactivity were detected in the paraventricular nucleus, lateral hypothalamic area, and arcuate nucleus, and the protein coexisted with orexin, melanin-concentrating hormone, neuropeptide Y, and proopiomelanocortin. SCG3 formed a granule-like structure together with these neuropeptides. Conclusions: Genetic variations in the SCG3 gene may influence the risk of obesity through possible regulation of hypothalamic neuropeptide secretion.

Objective: Gap-junctional communication (GJC) plays critical roles in cell growth and differentiation. Several studies have demonstrated the involvement of GJC in myogenesis and osteogenesis; however, the role of GJC in adipogenesis has not been fully studied. Thus, we investigated the role of GJC in adipogenesis. Research Methods and Procedures: 3T3-L1 preadipocytes were differentiated in the presence of gap junction inhibitor, 18-α-glycyrrhetinic acid (AGA), and accumulation of cytoplasmic triglycerides was measured. 3T3-L1 cells were transfected with 100 nM small interfering RNA duplexes targeting connexin (Cx) 43. The mRNA levels of CCAAT/enhancer-binding protein (C/EBP) α, peroxisome proliferator-activated receptor γ, glucose transporter 4, C/EBPβ, and Cx43 were measured by real-time polymerase chain reaction. The protein levels of C/EBPβ were quantitated by Western blotting. The cell proliferation was measured by counting cell numbers, and DNA synthesis was measured by bromodeoxyuridine incorporation. Results: AGA inhibited adipocyte differentiation dose-dependently. The lipid accumulation and the mRNA levels of C/EBPα, peroxisome proliferator-activated receptor γ, and glucose transporter 4 were markedly reduced in AGA-treated adipocytes. The mRNA levels of C/EBPβ did not decrease; however, C/EBPβ [liver-enriched transcriptional activator protein (LAP)] expression and the C/EBPβ (LAP)-to-C/EBP [liver-enriched transcriptional inhibitory protein (LIP)] ratio were reduced by AGA treatment. The increase in both cell number and DNA synthesis, which occurs during mitotic clonal expansion, was reduced by AGA in a dose-dependent fashion. The major component of gap junctions in 3T3-L1 cells was Cx43. Down-regulation of Cx43 using small interfering RNA reduced the expression of C/EBPβ (LAP) and inhibited adipogenesis. Discussion: Our data suggest that GJC plays some important roles in adipogenesis through inhibiting mitotic clonal expansion and modulating C/EBPβ (LAP) expression.

Nonalcoholic steatohepatitis (NASH) is a subset of nonalcoholic fatty liver disease (NAFLD) and sometimes progresses to cirrhosis and liver failure. We analyzed the expression profiles of approximately 50,000 genes and biological pathways in NASH patients in comparison with simple steatosis patients by using the analytical technique of GSEA (Gene Set Enrichment Analysis) by DNA microarrays. Although expressions of various genes were altered, GSEA showed clearly lower expression of nuclear receptors, including the peroxisome proliferator-activated receptor gamma (PPARγ) pathway. In a preliminary study we therefore investigated the therapeutic effect of low-dose pioglitazone (15 mg/day per body for 24 weeks), a synthetic ligand for PPARγ, in 12 NASH patients. A decrease in aminotransferase (ALT) values to within the normal range was observed in 7 (58.3%) of the patients, and because the dose of pioglitazone was lower than that ordinarily used, no side effects, such as fatigue, lower extremity edema, or weight gain, were observed. In conclusion, the results confirmed involvement of the PPARγ pathway in NASH and the therapeutic utility of a PPARγ ligand. Keywords:: life style-related disease, nonalcoholic steatohepatitis (NASH), nonalcoholic fatty liver disease (NAFLD), microarray, Gene Set Enrichment Analysis (GSEA), pioglitazone

There are a considerable number of patients with non-obese non-alcoholic fatty liver disease (NAFLD). However, the clinical characteristics of non-obese NAFLD is not fully understood. We investigated genetic and other clinical parameters in non-obese and obese NAFLD.The single nucleotide polymorphism rs738409 in the patatin-like phospholipase 3 gene (PNPLA3) was genotyped by the Invader assay in 540 NAFLD patients (134 non-obese and 406 obese) and 1012 control subjects (782 non-obese and 230 obese). All NAFLD patients underwent liver biopsy. Odds ratios were calculated by multiple logistic regression analysis using age, sex, body mass index (BMI), type 2 diabetes mellitus (T2DM) and rs738409 genotype as explanatory variables.Non-obese NAFLD subjects had a higher rs738409 GG genotype than obese NAFLD. Multiple logistic regression analysis indicated that the odds ratios of T2DM and rs738409 GG genotype for NAFLD were higher in non-obese than in obese groups. In non-obese NAFLD, rs738409 GG genotype was associated with lobular inflammation, hepatocyte ballooning and NAFLD activity score. In obese NAFLD, BMI and T2DM but not rs738409 GG genotype were associated with severity of histology.We demonstrated that the risk factors for the development and progression of NAFLD were different between non-obese and obese patients and that PNPLA3 rs738409 was strongly associated with the development and progression of non-obese NAFLD.

Visceral fat accumulation contributes to the development of insulin resistance, leading to metabolic syndrome. Adiponectin provides a link between visceral fat accumulation and insulin resistance. In addition to environmental factors, genetic factors play important roles in visceral fat accumulation and circulating adiponectin levels. Genome-wide association studies (GWASs) have identified genetic variations in the adiponectin, C1Q and collagen domain containing (ADIPOQ) gene that are associated with adiponectin levels. In this study, we investigated whether ADIPOQ single nucleotide polymorphisms (SNPs) were associated with visceral fat accumulation and insulin resistance. We measured the visceral fat area (VFA) by computed tomography (CT) and examined the presence of the insulin resistance-related phenotype (fasting plasma glucose, fasting insulin, and homeostasis model assessment-insulin resistance [HOMA-IR]) in a set of Japanese individuals (731 men and 864 women) who were genotyped for seven ADIPOQ SNPs reported by recent GWASs (namely, rs6810075, rs10937273, rs1648707, rs864265, rs182052, rs17366568, and rs6773957). SNPs associated with the phenotype (P0.05) were then evaluated by association analysis using a second set of the study participants (383 men and 510 women). None of the SNPs was associated with body mass index (BMI) or VFA in men or women. However, the adiponectin-decreasing alleles of rs10937273 and rs1648707 were significantly associated with HOMA-IR (P = 0.0030 and P = 0.00074, respectively) in women, independently of BMI. These SNPs were significantly associated with decreased adiponectin levels in women. Our results suggested that rs10937273 and rs1648707 may affect insulin sensitivity by regulating adiponectin production by adipose tissue in women.

A genome-wide association study revealed that the single nucleotide polymorphism (SNP) rs738409 in the patatin-like phospholipase 3 gene (PNPLA3) was strongly associated with non-alcoholic fatty liver disease (NAFLD). Recent pilot studies investigated the effects of dipeptidyl peptidase-4 inhibitors on liver function and glucose metabolism in NAFLD with type 2 diabetes mellitus (DM). We herein evaluated the efficacy of alogliptin in NAFLD patients with type 2 DM as well as the relationship between genotypes at rs738409 in PNPLA3 and treatment efficacy.Forty-one biopsy-proven NAFLD patients with type 2 DM treated with 25 mg/day alogliptin were retrospectively enrolled. SNP rs738409 in PNPLA3 was present in all patients. Clinical data were measured before and after the treatment.Average hemoglobin A1c (HbA1c) levels mostly remained unchanged. Moreover, significant changes were not noted in the levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) during the follow-up period. A positive correlation was observed between improvements in HbA1c (ΔHbA1c) levels and changes in AST (ΔAST) and ALT (ΔALT) levels (r = 0.325 and 0.439, respectively). Patients with the risk allele (G-allele) showed more positive correlation between ΔHbA1c and changes in transaminase. Furthermore, improvements in the levels of total cholesterol, triglycerides and hyaluronic acid were significantly greater in G-allele patients in the weight loss group.The treatment of NAFLD with type 2 DM with alogliptin contributed to the amelioration of NAFLD. Our results suggested that differences in the PNPLA3 risk allele affected the therapeutic effects of this treatment.

Background — Vascular smooth muscle cell proliferation plays an important role in the development of atherosclerosis. We previously reported that adiponectin, an adipocyte-specific plasma protein, accumulated in the human injured artery and suppressed endothelial inflammatory response as well as macrophage-to-foam cell transformation. The present study investigated the effects of adiponectin on proliferation and migration of human aortic smooth muscle cells (HASMCs). Methods and Results — HASMC proliferation was estimated by [ 3 H] thymidine uptake and cell number. Cell migration assay was performed using a Boyden chamber. Physiological concentrations of adiponectin significantly suppressed both proliferation and migration of HASMCs stimulated with platelet-derived growth factor (PDGF)-BB. Adiponectin specifically bound to 125 I-PDGF-BB and significantly inhibited the association of 125 I-PDGF-BB with HASMCs, but no effects were observed on the binding of 125 I-PDGF-AA or 125 I-heparin–binding epidermal growth factor (EGF)–like growth factor (HB-EGF) to HASMCs. Adiponectin strongly and dose-dependently suppressed PDGF-BB–induced p42/44 extracellular signal–related kinase (ERK) phosphorylation and PDGF β-receptor autophosphorylation analyzed by immunoblot. Adiponectin also reduced PDGF-AA–stimulated or HB-EGF–stimulated ERK phosphorylation in a dose-dependent manner without affecting autophosphorylation of PDGF α-receptor or EGF receptor. Conclusions — The adipocyte-derived plasma protein adiponectin strongly suppressed HASMC proliferation and migration through direct binding with PDGF-BB and generally inhibited growth factor–stimulated ERK signal in HASMCs, suggesting that adiponectin acts as a modulator for vascular remodeling.

Background & Aims The pathogenesis of non-alcoholic fatty liver disease (NAFLD) is affected by epigenetic factors as well as by genetic variation. Methods We performed targeted-bisulfite sequencing to determine the levels of DNA methylation of 4 CpG islands (CpG99, CpG71, CpG26, and CpG101) in the regulatory regions of PNPLA3 , SAMM50 , PARVB variant 1, and PARVB variant 2, respectively. We compared the levels of methylation of DNA in the livers of the first and second sets of patients with mild (fibrosis stages 0 and 1) or advanced (fibrosis stages 2 to 4) NAFLD and in those of patients with mild (F0 to F2) or advanced (F3 and F4) chronic hepatitis C infection. The hepatic mRNA levels of PNPLA3 , SAMM50 , and PARVB were measured using qPCR. Results CpG26, which resides in the regulatory region of PARVB variant 1, was markedly hypomethylated in the livers of patients with advanced NAFLD. Conversely, CpG99 in the regulatory region of PNPLA3 was substantially hypermethylated in these patients. These differences in DNA methylation were replicated in a second set of patients with NAFLD or chronic hepatitis C. PNPLA3 mRNA levels in the liver of the same section of a biopsy specimen used for genomic DNA preparation were lower in patients with advanced NAFLD compared with those with mild NAFLD and correlated inversely with CpG99 methylation in liver DNA. Moreover, the levels of CpG99 methylation and PNPLA3 mRNA were affected by the rs738409 genotype. Conclusions Hypomethylation of CpG26 and hypermethylation of CpG99 may contribute to the severity of fibrosis in patients with NAFLD or chronic hepatitis C infection.

The current study demonstrates that aquaporin adipose (AQPap), an adipose-specific glycerol channel (Kishida, K., Kuriyama, H., Funahashi, T., Shimomura, I., Kihara, S., Ouchi, N., Nishida, M., Nishizawa, H., Matsuda, M., Takahashi, M., Hotta, K., Nakamura, T., Yamashita, S., Tochino, Y., and Matsuzawa, Y. (2000) J. Biol. Chem. 275, 20896-20902), is a target gene of peroxisome proliferator-activated receptor (PPAR) gamma. The AQPap mRNA amounts increased following the induction of PPARgamma in the differentiation of 3T3-L1 adipocytes. The AQPap mRNA in the adipose tissue increased when mice were treated with pioglitazone (PGZ), a synthetic PPARgamma ligand, and decreased in PPARgamma(+/-) heterozygous knockout mice. In 3T3-L1 adipocytes, PGZ augmented the AQPap mRNA expression and its promoter activity. Serial deletion of the promoter revealed the putative peroxisome proliferator-activated receptor response element (PPRE) at -93/-77. In 3T3-L1 preadipocytes, the expression of PPARgamma by transfection and PGZ activated the luciferase activity of the promoter containing the PPRE, whereas the PPRE-deleted mutant was not affected. The gel mobility shift assay showed the direct binding of PPARgamma-retinoid X receptor alpha complex to the PPRE. DeltaPPARgamma, which we generated as the dominant negative PPARgamma lacking the activation function-2 domain, suppressed the promoter activity in 3T3-L1 cells, dose-dependently. We conclude that AQPap is a novel adipose-specific target gene of PPARgamma through the binding of PPARgamma-retinoid X receptor complex to the PPRE region in its promoter.

Galectins constitute a family of proteins that bind to beta-galactoside residues and have diverse physiological functions. Here we report on the identification of a galectin-like molecule, galectin-12, in a human adipose tissue cDNA library. The protein contained two potential carbohydrate-recognition domains with the second carbohydrate-recognition domain being less conserved compared with other galectins. In vitro translated galectin-12 bound to a lactosyl-agarose column far less efficiently than galectin-8. Galectin-12 mRNA was predominantly expressed in adipose tissue of human and mouse and in differentiated 3T3-L1 adipocytes. Caloric restriction and treatment of obese animals with troglitazone increased galectin-12 mRNA levels and decreased the average size of the cells in adipose tissue. The induction of galectin-12 expression by the thiazolidinedione, troglitazone, was paralleled by an increase in the number of apoptotic cells in adipose tissue. Immunocytochemical analysis revealed that galectin-12 was localized in the nucleus of adipocytes, and transfection with galectin-12 cDNA induced apoptosis of COS-1 cells. These results suggest that galectin-12, an adipose-expressed galectin-like molecule, may participate in the apoptosis of adipocytes.

Objective: Adipocytes secrete various cytokines and matrix proteins. Several of them precipitate in obesity-associated diseases, including atherosclerosis. In the current study, we have examined the expression of secreted protein, acidic and rich in cysteine (SPARC) in adipose tissue and its significance in obesity and coronary artery disease (CAD). Research Methods and Procedures: The SPARC mRNA expressions both in vivo and in vitro were detected by Northern blot analysis. Plasma SPARC concentrations were measured by enzyme immunosorbent assay. First, we investigated the plasma SPARC levels of 88 unrelated adult Japanese subjects (62 men and 26 women; average age: [± SD] 50 ± 12 years; body mass index [BMI]: 16 to 46 kg/m2). Additionally 31 subjects with CAD diagnosed by coronary angiography (20 men and 11 women) were also investigated. Results: Human adipose tissues expressed abundant SPARC mRNA. SPARC expression in adipose tissues was upregulated in obese db/db mice. Markedly enhanced expression of SPARC mRNA was observed in 3T3-L1 fibroblasts during adipocyte differentiation. Consistent with these results, plasma SPARC levels proved a positive correlation with BMI in humans (r = 0.27; p < 0.01). Interestingly, plasma SPARC concentrations were significantly elevated in age- and BMI-matched subjects with CAD (p < 0.05). Discussion: SPARC was expressed in adipose tissues and its expression was enhanced in obese mice. In human, plasma SPARC levels were elevated in obesity and CAD patients. This elevated SPARC may be involved in the progression of CAD.

Adiponectin is an adipose-specific plasma protein whose plasma concentrations are decreased in obese subjects and type 2 diabetic patients. This protein possesses putative antiatherogenic and anti-inflammatory properties. In the current study, we have analyzed the relationship between adiponectin and insulin resistance in rhesus monkeys (Macaca mulatta), which spontaneously develop obesity and which subsequently frequently progress to overt type 2 diabetes. The plasma levels of adiponectin were decreased in obese and diabetic monkeys as in humans. Prospective longitudinal studies revealed that the plasma levels of adiponectin declined at an early phase of obesity and remained decreased after the development of type 2 diabetes. Hyperinsulinemic-euglycemic clamp studies revealed that the obese monkeys with lower plasma adiponectin showed significantly lower insulin-stimulated peripheral glucose uptake (M rate). The plasma levels of adiponectin were significantly correlated to M rate (r = 0.66, P < 0.001). Longitudinally, the plasma adiponectin decreased in parallel to the progression of insulin resistance. No clear association was found between the plasma levels of adiponectin and its mRNA levels in adipose tissue. These results suggest that reduction in circulating adiponectin may be related to the development of insulin resistance.

Plasma concentrations of adiponectin, a novel adipose-specific protein with putative antiatherogenic and antiinflammatory effects, were found to be decreased in Japanese individuals with obesity, type 2 diabetes, and cardiovascular disease, conditions commonly associated with insulin resistance and hyperinsulinemia. To further characterize the relationship between adiponectinemia and adiposity, insulin sensitivity, insulinemia, and glucose tolerance, we measured plasma adiponectin concentrations, body composition (dual-energy x-ray absorptiometry), insulin sensitivity (M, hyperinsulinemic clamp), and glucose tolerance (75-g oral glucose tolerance test) in 23 Caucasians and 121 Pima Indians, a population with a high propensity for obesity and type 2 diabetes. Plasma adiponectin concentration was negatively correlated with percent body fat (r = -0.43), waist-to-thigh ratio (r = -0.46), fasting plasma insulin concentration (r = -0.63), and 2-h glucose concentration (r = -0.38), and positively correlated with M (r = 0.59) (all P < 0.001); all relations were evident in both ethnic groups. In a multivariate analysis, fasting plasma insulin concentration, M, and waist-to-thigh ratio, but not percent body fat or 2-h glucose concentration, were significant independent determinates of adiponectinemia, explaining 47% of the variance (r(2) = 0.47). Differences in adiponectinemia between Pima Indians and Caucasians (7.2 +/- 2.6 vs. 10.2 +/- 4.3 microg/ml, P < 0.0001) and between Pima Indians with normal, impaired, and diabetic glucose tolerance (7.5 +/- 2.7, 6.1 +/- 2.0, 5.5 +/- 1.6 microg/ml, P < 0.0001) remained significant after adjustment for adiposity, but not after additional adjustment for M or fasting insulin concentration. These results confirm that obesity and type 2 diabetes are associated with low plasma adiponectin concentrations in different ethnic groups and indicate that the degree of hypoadiponectinemia is more closely related to the degree of insulin resistance and hyperinsulinemia than to the degree of adiposity and glucose intolerance.

Background —Excessive lipid accumulation in macrophages plays an important role in the development of atherosclerosis. Recently, we discovered an adipocyte-specific plasma protein, adiponectin, that is decreased in patients with coronary artery disease. We previously demonstrated that adiponectin acts as a modulator for proinflammatory stimuli and inhibits monocyte adhesion to endothelial cells. The present study investigated the effects of adiponectin on lipid accumulation in human monocyte-derived macrophages. Methods and Results —Human monocytes were differentiated into macrophages by incubation in human type AB serum for 7 days, and the effects of adiponectin were investigated at different time intervals. Treatment with physiological concentrations of adiponectin reduced intracellular cholesteryl ester content, as determined using the enzymatic, fluorometric method. The adiponectin-treated macrophages contained fewer lipid droplets stained by oil red O. Adiponectin suppressed the expression of the class A macrophage scavenger receptor (MSR) at both mRNA and protein levels by Northern and immunoblot analyses, respectively, without affecting the expression of CD36, which was quantified by flow cytometry. Adiponectin reduced the class A MSR promoter activity, as measured by luciferase reporter assay. Adiponectin treatment dose-dependently decreased class A MSR ligand binding and uptake activities. The mRNA level of lipoprotein lipase as a marker of macrophage differentiation was decreased by adiponectin treatment, but that of apolipoprotein E was not altered. Adiponectin was detected around macrophages in the human injured aorta by immunohistochemistry. Conclusions —The adipocyte-derived plasma protein adiponectin suppressed macrophage-to-foam cell transformation, suggesting that adiponectin may act as a modulator for macrophage-to-foam cell transformation.

Abstract —Adiponectin is a novel, adipose-specific protein abundantly present in the circulation, and it has antiatherogenic properties. We analyzed the plasma adiponectin concentrations in age- and body mass index (BMI)–matched nondiabetic and type 2 diabetic subjects with and without coronary artery disease (CAD). Plasma levels of adiponectin in the diabetic subjects without CAD were lower than those in nondiabetic subjects (6.6±0.4 versus 7.9±0.5 μg/mL in men, 7.6±0.7 versus 11.7±1.0 μg/mL in women; P P r =−0.18, P r =−0.26, P

Aging has been shown to have an effect on the capacity to differentiate preadipocytes and on the expression of some genes expressed in adipose tissue. The mRNA levels of adipocyte differentiation-related genes were examined in rhesus monkeys (Macaca Mulatta) ranging in age from 7 to 30 years. The effect of aging on the expression of peroxisome proliferator activated receptor gamma (PPARgamma), adipocyte determination- and differentiation-dependent factor 1/sterol regulatory element binding protein 1 (ADD1/SREBP1), CCAAT/enhancer binding protein alpha (C/EBPalpha), lipoprotein lipase (LPL), GLUT4 glucose transporter, and adipsin were examined by slot blot analysis. Significant inverse correlations were observed between age and the mRNA levels of PPARgamma, ADD1/SREBP1, LPL, and GLUT4. The coordinate downregulation of these genes may be linked to the declining fat mass of senescent animals. There was no correlation between age and the mRNA levels of adipsin. The mRNA levels of these genes were not correlated to body weight orfasting plasma insulin. These findings indicate that aging may have an effect on the adipocyte differentiation program and this effect appears to be gene specific.

Objective: We have cloned the rhesus monkey leptin receptor and examined its mRNA expression levels in the adipose tissue of monkeys to investigate the regulation of gene expression of the leptin receptor. Research Methods and Procedures: Monkey leptin receptor cDNA was cloned by reverse transcriptase-polymerase chain reaction (RT-PCR). Tissue distribution of monkey leptin receptor was examined by Northern blot analysis and RT-PCR. The mRNA levels of monkey leptin receptor in adipose tissue of normal (n=10), hyperinsulinemic obese (n = 8), and type 2 diabetic monkeys (n = 8) were measured by quantitative RT-PCR. Results: Monkey leptin receptor cDNA had at least two alternatively spliced isoforms (long and short forms). The long form of the leptin receptor mRNA was expressed relatively highly in liver, adipose tissue, hypothalamus, and choroid plexus, whereas the total leptin receptors were expressed in every tissue examined. The mRNA levels of the long form of the leptin receptor in adipose tissue were not correlated to body weight, fasting plasma insulin, plasma glucose, or plasma leptin levels. The mRNA levels of the long form of the leptin receptor were highly correlated to that of the total leptin receptor (long and short form). Discussion: The long form of leptin receptor mRNA existed in adipose tissue as well as in liver and hypothalamus, suggesting that the leptin receptor in adipose tissue may be functional in adipose tissue. The expression of the leptin receptor mRNA in adipose tissue is not affected by obesity, hyperinsulinemia, or diabetes.