Naoharu Iwai

Meta-analysis of genome-wide association studies identifies common variants associated with blood pressure variation in East Asians

We conducted a meta-analysis of genome-wide association studies of systolic (SBP) and diastolic (DBP) blood pressure in 19,608 subjects of east Asian ancestry from the AGEN-BP consortium followed up with de novo genotyping (n = 10,518) and further replication (n = 20,247) in east Asian samples. We identified genome-wide significant (P < 5 × 10−8) associations with SBP or DBP, which included variants at four new loci (ST7L-CAPZA1, FIGN-GRB14, ENPEP and NPR3) and a newly discovered variant near TBX3. Among the five newly discovered variants, we obtained significant replication in the independent samples for all of these loci except NPR3. We also confirmed seven loci previously identified in populations of European descent. Moreover, at 12q24.13 near ALDH2, we observed strong association signals (P = 7.9 × 10−31 and P = 1.3 × 10−35 for SBP and DBP, respectively) with ethnic specificity. These findings provide new insights into blood pressure regulation and potential targets for intervention.

Identification of two subtypes in the rat type I angiotensin II receptor

A rat adrenal cDNA library was screened by colony hybridization using a rat cDNA fragment of type I angiotensin II receptor (ATIA) previously isolated from the kidney. Two cDNA clones were identified, designated as AT1B, to have a nucleotide sequence highly homologous to and yet distinct from AT1A. The amino acid sequence of AT1B consists of 359 amino acid residues and has 96% identity with AT1A. No conspicuous difference in the ligand binding characteristics was observed between AT1A and AT1B. The mRNA for AT1B was expressed in many tissues as is the case with AT1A, and most abundantly expressed in the adrenal glands in the Sprague—Dawley rats. The existence of two subtypes in the rat type I anglotensin II receptor might explain the diverse actions of angiotensin II in various tissues.

Plasma MicroRNA 499 as a Biomarker of Acute Myocardial Infarction

BACKGROUND MicroRNAs (miRNAs) are endogenous small RNAs 21-25 nucleotides in length. Recently, we reported that miRNA 208 (miR-208) is produced exclusively in the rat myocardium and that plasma miR-208 is a biomarker of myocardial injury in rats. In the present study, we assessed the hypothesis that plasma concentrations of myocardial-specific miRNAs can be used to diagnose myocardial injury in humans. METHODS We used array analysis of miRNA production in various human tissues to identify heart-specific miRNAs. We assessed the plasma concentrations of miR-499 in 14 individuals with acute coronary syndromes, 15 individuals with congestive heart failure, and 10 individuals without cardiovascular diseases. Plasma miR-499 concentrations were measured with a real-time reverse-transcription PCR method that used an artificial small RNA as an internal calibrator. RESULTS The miRNA array analysis of various human tissues indicated that miR-499 was produced almost exclusively in the heart. Plasma miR-499 concentrations were measurably increased in all individuals with acute myocardial infarction but were below the limit of detection for all individuals in the other patient groups. CONCLUSIONS The plasma concentration of miR-499 may be a useful biomarker of myocardial infarction in humans.

Meta-analysis identifies common variants associated with body mass index in east Asians.

Multiple genetic loci associated with obesity or body mass index (BMI) have been identified through genome-wide association studies conducted predominantly in populations of European ancestry. We performed a meta-analysis of associations between BMI and approximately 2.4 million SNPs in 27,715 east Asians, which was followed by in silico and de novo replication studies in 37,691 and 17,642 additional east Asians, respectively. We identified ten BMI-associated loci at genome-wide significance (P < 5.0 × 10−8), including seven previously identified loci (FTO, SEC16B, MC4R, GIPR-QPCTL, ADCY3-DNAJC27, BDNF and MAP2K5) and three novel loci in or near the CDKAL1, PCSK1 and GP2 genes. Three additional loci nearly reached the genome-wide significance threshold, including two previously identified loci in the GNPDA2 and TFAP2B genes and a newly identified signal near PAX6, all of which were associated with BMI with P < 5.0 × 10−7. Findings from this study may shed light on new pathways involved in obesity and demonstrate the value of conducting genetic studies in non-European populations.

DD genotype of the angiotensin-converting enzyme gene is a risk factor for left ventricular hypertrophy.

BackgroundThe cardiac renin-angiotensin system has been suggested to be involved in the development of left ventricular hypertrophy. In humans, a strong correlation has been found between plasma angiotensin I-converting enzyme (ACE) activity and the insertion/deletion (I/D) polymorphism of the ACE gene, which has been reported to be associated with myocardial infarction, ischemic and idiopathic dilated cardiomyopathy, sudden death in hypertrophic cardiomyopathy, and restenosis after percutaneous transluminal coronary angioplasty. In the present study, we examined the possibility that the genotype of the ACE gene might influence the development of left ventricular hypertrophy. Methods and ResultsThe study population consisted of 268 subjects randomly selected from our outpatient clinic. In 142 subjects, left ventricular mass (LVM) was determined by echocardiogram. The genotype of the ACE gene was determined by the polymerase chain reaction. ANCOVA revealed that the genotype of the ACE gene had no effect on blood pressure. The percentage of the explained variance of LVM with variables including diastolic blood pressure (DBP, P = .0001), body mass index (BMI, P = .0001), sex (P = .0009), and the genotype of the ACE gene (P = .0017) was 34.61%. Significant differences in the effects of the genotype of the ACE gene on LVM were observed between the II and DD (P = .0004) and between the ID and DD (P = .0077) genotypes. The percentage of the explained variance of the LVM/ht ratio with variables including sex (P = .134), age (P = .3655), the genotype of the ACE gene (P = .0014), BMI (P = .0001), and DBP (P = .0001) was 31.25%. Significant differences in the effects of the genotype of the ACE gene on LVM/ht were observed between the II and DD genotypes (P = .0003) and between the ID and DD genotypes (P = .0091). ConclusionsIn addition to BMI and DBP, the genotype of the ACE gene was a significant predictor of LVM and LVM/ht in our study population.

Rat angiotensin II receptor: cDNA sequence and regulation of the gene expression

The nucleotide and amino acid sequences for rat type I angiotensin II receptor were deduced through molecular cloning and sequence analysis of its complementary DNAs. The rat angiotensin II receptor consists of 359 amino acid residues and has a sequence similar to G protein-coupled receptors. The expression of this receptor gene was detected in the adrenal, liver and kidney by Northern blotting. Sodium deprivation positively modulated the expression of the receptor gene in the adrenal. No detectable change was observed in the expression levels of this receptor gene between spontaneously hypertensive rats and Wistar-Kyoto rats in the tissues examined including the adrenal, brain, kidney and liver. Interestingly the expression of this receptor gene was developmentally regulated.

Isolation of preferentially expressed genes in the kidneys of hypertensive rats.

By differential hybridization, three complementary DNAs designated as S3, S2, and SA were isolated, and the corresponding messenger RNAs (mRNAs) were differentially expressed between the kidneys of spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto (WKY) rats. S3 is identical to cytochrome P450 IV A2. SA encoded a protein of 546 amino acid residues, and its carboxyl terminal region had a slight homology to luciferase. No homologous sequence has been reported in S2 sequences. S3 mRNA was about four times more abundantly expressed in the kidneys of 28 -day-old SHR than in those of age-matched WKY rats, but there was no difference at age 16 weeks. A low NaCI diet positively modulated the expression of the S3 gene. S2 mRNA was almost undetectable in the kidneys of 28-day-old WKY rats but was clearly detected in those of age-matched SHR. The expression level of S2 mRNA in the livers of 16-week-old SHR was about five times higher than that of age-matched WKY rats. The expression of S2 mRNA in the livers was modulated by dietary NaCI and captopril. S^ mRNA was more than 10 times more abundantly expressed in the kidneys of SHR than in those of WKY rats from age 4 weeks. With the administration of captopril, the expressions of S^ mRNA in the livers of SHR were positively modulated. Because these three genes are not only differentially expressed between SHR and WKY rats but also related to sodium metabolism or blood pressure control, the identification of these genes may provide important probes to examine the mechanisms of hypertension.

Genetic Polymorphism of CYP11B2 Gene and Hypertension in Japanese

Low-renin hypertension is characterized by a high ratio of aldosterone to plasma renin activity (ALD/PRA), which may suggest inappropriately increased aldosterone biosynthesis. The genes for the enzymes involved in aldosterone synthesis may contribute to low-renin hypertension. We investigated the associations between genetic variations of CYP11B2 (aldosterone synthase) T(-344)C and hypertension in 482 Japanese subjects. Subjects older than 50 years with a blood pressure <140/85 mm Hg were considered normotensive (n=227 subjects), and subjects younger than 65 years old with a BP >160/95 mm Hg were considered hypertensive (n=255 subjects). The frequency of the TC+CC genotypes in the normotensive group was significantly lower than in the hypertensive group. Logistic analysis on 482 subjects revealed that body mass index, gender, and the genotype of CYP11B2 T(-344)C were significantly associated with hypertension. ALD and PRA were assessed in 97 subjects with hypertension, and the TC+CC genotypes were significantly associated with higher ALD/PRA. Sixty-five subjects with hypertension were assessed by 24-hour ambulatory blood pressure monitoring, and the frequency of nondippers (a difference in mean blood pressure of <10% between the daytime [6 AM to 9 PM] and nighttime [9 PM to 6 AM] hours) was significantly higher in subjects with the TC+CC (hetero+homo mutation) genotype than in subjects with the TT (wild-type) genotype. Echocardiographic assessment (n=136) revealed that the ratio of left ventricular end-diastolic dimension to height tended to be higher in subjects with the TC+CC genotype than in subjects with the TT genotype. The present study suggests that the (-344)C allele of the CYP11B2 gene may be a genetic marker for low-renin hypertension in Japanese.

Angiotensinogen Gene and Blood Pressure in the Japanese Population

A molecular variant of the angiotensinogen gene with threonine instead of methionine at position 235 (ie, with M235T polymorphism) has been shown to be associated with essential hypertension in Caucasian populations. The purpose of the present study was to assess whether the M235T polymorphism was associated with essential hypertension in the Japanese population. The study population consisted of 347 subjects selected in our outpatient clinic. The clinical data included in the analyses were sex, age, body mass index, cholesterol level, genotype of the angiotensinogen gene, genotype of the angiotensin-converting enzyme gene, and systolic and diastolic blood pressure. Multiple regression analysis revealed that only body mass index was a predictor of both diastolic and systolic blood pressure in these 347 subjects, but the genotype of the angiotensinogen gene was identified as a predictor of both diastolic and systolic blood pressure in a subpopulation less than 50 years of age. However, in a subpopulation more than 50 years of age, body mass index was the only predictor of both systolic and diastolic blood pressure. Of the 347 subjects, 189 had a technically excellent echocardiogram at the initial observation period. Multiple regression analysis revealed that sex, body mass index, diastolic blood pressure, and genotype of the angiotensin-converting enzyme gene were predictors of left ventricular mass. Although subjects with the TT angiotensinogen genotype had significantly greater left ventricular mass than those with either the TM or the MM genotype, the effects of the genotype of the angiotensinogen gene on left ventricular mass were mainly due to effects on blood pressure.

Two distinct pathways in the down-regulation of type-1 angiotensin II receptor gene in rat glomerular mesangial cells

The mRNA level of the type-1 angiotensin II receptor (AT1) was down-regulated by angiotensin II in cultured rat glomerular mesangial cells. The effect was maximum with 1 microM AII at 6 h, sensitive to cycloheximide, and specific to AT1 since this phenomenon was blocked by DuP753, an AT1 antagonist, but not by type-2 antagonist PD123319. Dibutyryl cAMP, forskolin, and cholera toxin also caused AT1 down-regulation. These effects were not altered by either the protein kinase A inhibitor H-8 or cycloheximide. Calcium ionophore A23187, pertussis toxin, protein kinase C inhibitor staurosporine, or prolonged incubation with phorbol ester were without effect. These results suggest that there are at least two pathways to down-regulate AT1 mRNA; one way is an angiotensin II-induced, protein kinase C-independent, and cycloheximide-sensitive pathway and the other is an angiotensin II-independent, cAMP-induced, and cycloheximide-insensitive pathway.