ホモ接合体性家族性高コレステロール血症における低比重リポ蛋白受容体遺伝子変異に関する研究

取得学位 : 博士（医学）, 学位授与番号 : 医博甲第1110号, 学位授与年月日：平成6年3月25日,学位授与年：199

Non-synonymous single-nucleotide variations of the human oxytocin receptor gene and autism spectrum disorders: a case-control study in a Japanese population and functional analysis.

Background: The human oxytocin receptor (hOXTR) is implicated in the etiology of autism spectrum disorders (ASDs) and is a potential target for therapeutic intervention. Several studies have reported single-nucleotide polymorphisms (SNPs) of the OXTR gene associated with ASDs. These SNPs, however, reside outside the protein-coding region. Not much is known about genetic variations that cause amino acid substitutions that alter receptor functions. Methods. Variations in the OXTR gene were analyzed in 132 ASD patients at Kanazawa University Hospital in Japan and 248 unrelated healthy Japanese volunteers by re-sequencing and real-time polymerase chain reaction-based genotyping. Functional changes in variant OXTRs were assessed by radioligand binding assay and measurements of intracellular free calcium concentrations ([Ca§ssup§2+§esup§]§ssub§i§esub§) and inositol 1,4,5-trisphosphate (IP§ssub§3§esub§) levels. Results: Six subjects (4.5%) in the ASD group and two in the control group (0.8%) were identified as heterozygotes carrying the R376G variation (rs35062132; c.1126C>G); one individual from the ASD group (0.8%) and three members of the control group (1.2%) were found to be carrying R376C (c.1126C>T). The C/G genotype significantly correlated with an increased risk of ASDs (odds ratio (OR) = 5.83; 95% confidence interval (CI) = 1.16 to 29.33; P = 0.024, Fisher\u27s exact test). Consistently, the G allele showed a correlation with an increased likelihood of ASDs (OR = 5.73; 95% CI = 1.15 to 28.61; P = 0.024, Fisher\u27s exact test). The frequencies of the C/T genotype and the T allele in the ASD and control groups did not differ significantly. We also examined changes in agonist-induced cellular responses mediated by the variant receptors hOXTR-376G and hOXTR-376C. OXT-induced receptor internalization and recycling were faster in hOXTR-376G-expressing HEK-293 cells than in cells expressing hOXTR-376R or hOXTR-376C. In addition, the elevation in [Ca§ssup§2+§esup§]§ssub§i§esub§ and IP§ssub§3§esub§ formation decreased in the cells expressing hOXTR-376G and hOXTR-376C tagged with enhanced green fluorescent protein (EGFP), in comparison with the cells expressing the common-type hOXTR-376R tagged with EGFP. Conclusions: These results suggest that the rare genetic variation rs35062132 might contribute to the pathogenesis of ASDs, and could provide a molecular basis of individual differences in OXTR-mediated modulation of social behavior. © 2013 Ma et al.; licensee BioMed Central Ltd

Decreased ADP-Ribosyl Cyclase Activity in Peripheral Blood Mononuclear Cells from Diabetic Patients with Nephropathy

Aims/hypothesis. ADP-ribosyl-cyclase activity (ADPRCA) of CD38 and other ectoenzymes mainly generate cyclic adenosine 5’diphosphate-(ADP-) ribose (cADPR) as a second messenger in various mammalian cells, including pancreatic beta cells and peripheral blood mononuclear cells (PBMCs). Since PBMCs contribute to the pathogenesis of diabetic nephropathy, ADPRCA of PBMCs could serve as a clinical prognostic marker for diabetic nephropathy. This study aimed to investigate the connection between ADPRCA in PBMCs and diabetic complications. Methods. PBMCs from 60 diabetic patients (10 for type 1 and 50 for type 2) and 15 nondiabetic controls were fluorometrically measured for ADPRCA based on the conversion of nicotinamide guanine dinucleotide (NGD+) into cyclic GDP-ribose. Results. ADPRCA negatively correlated with the level of HbA1c (P = .040, R2 = .073), although ADPRCA showed no significant correlation with gender, age, BMI, blood pressure, level of fasting plasma glucose and lipid levels, as well as type, duration, or medication of diabetes. Interestingly, patients with nephropathy, but not other complications, presented significantly lower ADPRCA than those without nephropathy (P = .0198) and diabetes (P = .0332). ANCOVA analysis adjusted for HbA1c showed no significant correlation between ADPRCA and nephropathy. However, logistic regression analyses revealed that determinants for nephropathy were systolic blood pressure and ADPRCA, not HbA1c. Conclusion/interpretation. Decreased ADPRCA significantly correlated with diabetic nephropathy. ADPRCA in PBMCs would be an important marker associated with diabetic nephropathy

Effect of common methylenetetrahydrofolate reductase gene mutation on coronary artery disease in familial hypercholesterolemia

Familial hypercholesterolemia (FH) is an autosomal dominant disorder characterized by primary hypercholesterolemia and premature coronary artery disease (CAD). However, the development of CAD in FH shows considerable interindividual variations. Elevated levels of plasma homocysteine have been recognized as independent risk factors for CAD. A 5,10-methylenetetrahydrofolate reductase (MTHFR) gene mutation (valine [V] was substituted for alanine [A]) has been reported to be associated with elevated levels of plasma homocysteine in mutant homozygotes (i.e., VV). We studied 199 consecutive male heterozygous FH patients, 99 with and 100 without CAD. In the CAD group, genotype VV and V alleles were significantly more frequent than in the non-CAD group (15% vs 7% in genotypes [p = 0.035] and 0.41 vs 0.30 in alleles [p = 0.017]). The mean ages at onset in the CAD group were 50, 51, and 43 years for genotypes AA, AV, and VV, respectively (p <0.05); the age of onset of CAD in genotype VV was significantly lower than in the other 2 genotypes. Kaplan-Meier survivor curves indicated that the development of CAD was significantly accelerated by MTHFR mutation, probably in a gene dose-dependent manner. Furthermore, only MTHFR genotype VV was shown to be an independent predictor of the early onset of CAD in a stepwise multiple regression analysis. The mean plasma homocysteine levels of genotype VV were significantly higher than those of the other 2 genotypes. Thus, the MTHFR mutation appears to accelerate the onset of CAD through elevation of plasma homocysteine levels in male heterozygous patients with FH

Contribution of visceral adiposity and insulin resistance to metabolic risk factors in Japanese men

金沢大学大学院医学系研究科北陸中央病院内科We investigated the relative impacts of visceral adiposity and insulin resistance on the metabolic risk profile in middle-aged Japanese men. A cross-sectional study was conducted in 636 nondiabetic Japanese men with a mean age of 51.6 years. Visceral adipose tissue (AT) was assessed using computed tomography, and insulin resistance was determined by the homeostasis model assessment of insulin resistance (HOMA-IR). Metabolic risk factors were diagnosed according to the National Cholesterol Education Program Adult Treatment Panel III metabolic syndrome criteria: (1) hypertriglyceridemia, (2) low high-density lipoprotein cholesterol, (3) hypertension, (4) impaired fasting glucose, and (5) impaired glucose tolerance. Visceral AT and HOMA-IR were significantly and positively correlated with each other (r = 0.41, P < .001). Using the 75th percentile value as a cut point, those with isolated large visceral AT showed significantly greater odds ratios for each of the 5 risk factors measured except impaired fasting glucose, whereas those with isolated high HOMA-IR showed significantly greater odds ratios for each of the 5 risk factors except hypertriglyceridemia and impaired glucose tolerance, compared with the control group. The combined group (increased visceral AT and HOMA-IR) had the highest odds ratios for all studied risk factors. On logistic regression analysis using visceral AT and HOMA-IR as continuous independent variables, they were each independently associated with most of the metabolic risk factors and their clustering. In conclusion, neither visceral AT nor HOMA-IR stands out as the sole driving force of the risk profile; each makes a significant contribution to metabolic abnormalities in Japanese men. © 2010 Elsevier Inc. All rights reserved

A study of single nucleotide polymorphisms of the SLC19A1/RFC1 gene in subjects with autism spectrum disorder

Autism Spectrum Disorder (ASD) is a group of neurodevelopmental disorders with complex genetic etiology. Recent studies have indicated that children with ASD may have altered folate or methionine metabolism, suggesting that the folate-methionine cycle may play a key role in the etiology of ASD. SLC19A1, also referred to as reduced folate carrier 1 (RFC1), is a member of the solute carrier group of transporters and is one of the key enzymes in the folate metabolism pathway. Findings from multiple genomic screens suggest the presence of an autism susceptibility locus on chromosome 21q22.3, which includes SLC19A1. Therefore, we performed a case-control study in a Japanese population. In this study, DNA samples obtained from 147 ASD patients at the Kanazawa University Hospital in Japan and 150 unrelated healthy Japanese volunteers were examined by the sequence-specific primer-polymerase chain reaction method pooled with fluorescence correlation spectroscopy. p < 0.05 was considered to represent a statistically significant outcome. Of 13 single nucleotide polymorphisms (SNPs) examined, a significant p-value was obtained for AA genotype of one SNP (rs1023159, OR = 0.39, 95% CI = 0.16-0.91, p = 0.0394; Fisher’s exact test). Despite some conflicting results, our findings supported a role for the polymorphism rs1023159 of the SLC19A1 gene, alone or in combination, as a risk factor for ASD. However, the findings were not consistent after multiple testing corrections. In conclusion, although our results supported a role of the SLC19A1 gene in the etiology of ASD, it was not a significant risk factor for the ASD samples analyzed in this study. © 2016 by the authors; licensee MDPI, Basel, Switzerland

Regulation of Heme Oxygenase-1 Expression and Function by Insulin through PI3-kinase/Akt-2 Pathway but not Nrf2 Activation.

金沢大学附属病院発行後1年より全文公開

Genetic cholesteryl ester transfer protein deficiency caused by two prevalent mutations as a major determinant of increased levels of high density lipoprotein cholesterol

金沢大学大学院医学系研究科病態検査学Genetic determinants of HDL cholesterol (HDL-C) levels in the general population are poorly understood. We previously described plasma cholesteryl ester transfer protein (CETP) deficiency due to an intron 14 G(+1)-to-A mutation(Int14 A) in several families with very high HDL-C levels in Japan. Subjects with HDL-C ≥ 100 mg/dl (n = 130) were screened by PCR single strand conformational polymorphism analysis of the CETP gene. Two other mutations were identified by DNA sequencing or primer-mediated restriction map modification of PCR products: a novel intron 14 splice donor site mutation caused by a T insertion at position +3 from the exon14/intron14 boundary (Int14 T) and a missense mutation (Asp442 to Gly) within exon 15 (D442G). The Int14 T mutation was only found in one family. However, the D442G and Int14 A mutations were highly prevalent in subjects with HDL-C ≥ 60 mg/dl, with combined allele frequencies of 9%, 12%, 21%, and 43% for HDL-C 60-79, 80-99, 100-119, and ≥ 120 mg/dl, respectively. Furthermore, prevalences of the D442G and Int14 A mutations were extremely high in a general sample of Japanese men (n = 236), with heterozygote frequencies of 7% and 2%, respectively. These two mutations accounted for about 10% of the total variance of HDL-C in this population. The phenotype in a genetic compound heterozygote (Int14 T and Int14 A) was similar to that of Int14 A homozygotes (no detectable CETP and markedly increased HDL-C), indicating that the Int14 T produces a null allele. In four D442G homozygotes, mean HDL-C levels (86±26 mg/dl) were lower than in Int14 A homozygotes (158±35 mg/dl), reflecting residual CETP activity in plasma. In 47 D442G heterozygotes, mean HDL-C levels were 91±23 mg/dl, similar to the level in D442G homozygotes, and significantly greater than mean HDL-C levels in Int14 A heterozygotes (69±15 mg/dl). Thus, the D442G mutation acts differently to the null mutations with weaker effects on HDL in the homozygous state and stronger effects in the heterozygotes, suggesting dominant expression of a partially defective allele. CETP deficiency, reflecting two prevalent mutations (D442G and Int14 A), is the first example of a genetic deficiency state which is sufficiently common to explain a significant fraction of the variation in HDL-C in the general population

Clinical Study Impact of Serum Retinol-Binding Protein 4 Levels on Regulation of Remnant-Like Particles Triglyceride in Type 2 Diabetes Mellitus

Background. Although retinol-binding protein 4 (RBP4) associates with insulin resistance and remnant-like particles triglyceride (RLP-TG) elevated in the insulin resistant state, few data exist regarding the relationship between RBP4 and RLP-TG. Subjects and Methods. The study included 92 Japanese type 2 diabetic mellitus (T2DM) male patients (age 60.5 ± 13.6 years, body mass index (BMI) 24.7±4.1 kg/m 2 , waist circumference (WC) 88.4±10.7 cm, and HbA1c (NGSP) 7.2±1.9%). Patients on medications affecting insulin sensitivity, including fibrates, biguanides, and thiazolidinedione, were excluded. Visceral fat area (VFA) and subcutaneous fat area (SFA) were measured by computed tomography. Results. RBP4 levels showed a significant positive correlation with RLP-TG ( = 0.2544 and = 0.0056), TG ( = 0.1852 and = 0.041), RLP-TG/TG ( = 0.23765 and = 0.0241), and age ( = −0.2082 and = 0.0219), although there was no significant correlation with VFA, SFA, adiponectin levels, or homeostasis model of assessment insulin resistance (HOMA-R). Multiple regression analysis revealed that RBP4 was an independent determinant of RLP-TG ( = 0.0193) but was not a determinant of TG. Conclusions. RBP4 correlates positively with serum RLP-TG independent of fat accumulation in T2DM. RBP4 may regulate remnant metabolism independent of glycemic control in T2DM