Miyuki Onishi

Genetic variants at the 9p21 locus contribute to atherosclerosis through modulation of ANRIL and CDKN2A/B

UNLABELLED Genome-wide association studies (GWAS) have identified genetic variants contributing to the risk of cardiovascular disease (CVD) at the chromosome 9p21 locus. The CVD-associated region is adjacent to the two cyclin dependent kinase inhibitors (CDKN)2A and 2B and the last exons of the non-coding RNA, ANRIL. It is still not clear which of or how these transcripts are involved in the pathogenesis of atherosclerosis. OBJECTIVE We assessed the hypothesis that 9p21 locus polymorphisms influence the expression of the transcripts in the region (ANRIL, CDKN2A/B) and that these transcripts contribute to atherogenesis through the modulation of proliferation in VSMC. METHODS We genotyped 18 SNPs (r(2)<0.8 and MAF>0.05) across the region of interest: CDKN2A/B and ANRIL, encompassing the CVD-associated region. RNA and DNA were extracted from the blood of 57 volunteers (69-72 years old). Carotid ultrasound was performed in 56 subjects. CDKN2A/B and ANRIL (exons 1-2 and 17-18) expression was measured employing RT-PCR. Gene expression and cell growth were evaluated in cultured VSMC after the siRNA-mediated knock-down of ANRIL. RESULTS The risk alleles for atherosclerosis-related phenotypes were consistently associated with a lower expression of ANRIL when evaluating exons 1-2. Common carotid artery stenosis was associated with a significantly lower (P<0.01) expression of ANRIL (exons 1-2). ANRIL knock-down in VSMC caused significant variation in expression of CDKN2A/B (P<0.05) and reduction of cell growth (P<0.05) in vitro. CONCLUSION Disease-associated SNPs at the 9p21 locus predominantly affect the expression of ANRIL. Overall, our results suggest that several CVD-associated SNPs in the 9p21 locus affect the expression of ANRIL, which, in turn modulate cell growth, possibly via CDKN2A/B regulation.

The impact of visit-to-visit variability in blood pressure on renal function.

Hypertension is an important risk factor for cardiovascular diseases such as chronic kidney disease. It is still not fully understood how blood pressure impacts the kidneys. In this study, we aimed to establish the significance of visit-to-visit variability in blood pressure for renal function. We analyzed 143 consecutive patients undergoing renal Doppler ultrasonography in our hospital ward and measured blood pressure at outpatient visits six or more times. We analyzed the correlation between visit-to-visit variability in blood pressure and multiple clinical parameters, including albuminuria and resistive index evaluated by renal Doppler ultrasonography, which is thought to be a good indicator of renal vascular resistance. Subjects with higher variability in systolic blood pressure showed a significantly higher prevalence rate of clinical albuminuria and microalbuminuria, and showed significantly higher resistive index. Stepwise multiple regression analysis showed that variability in systolic blood pressure was a significant risk factor for higher resistive index, independent of other renal risk factors. Visit-to-visit variability in blood pressure correlates significantly with renal function and renal arteriosclerotic change. This parameter could provide additional information about renal arteriosclerotic change independent of estimated glomerular filtration rate and albuminuria, and should be considered a therapeutic target for renal protection.

Deletion allele of the angiotensin-converting enzyme gene as a risk factor for pneumonia in elderly patients ☆

PURPOSE Aspiration due to an age-related reduction in cough is a major cause of pneumonia in elderly persons. Because the insertion/deletion (I/D) polymorphism of the angiotensin-converting enzyme (ACE) gene (ACE) has been associated with the cough reflex, we studied whether this genetic polymorphism was also associated with the risk of pneumonia. SUBJECTS AND METHODS We studied 1011 elderly inpatients (221 men and 790 women, mean [+/- SD] age of 82 +/- 7 years) in a long-term care hospital. The association between the ACE I/D polymorphism and the incidence of pneumonia (defined using specific criteria that included radiographic abnormalities) was assessed during an 8-month period that excluded the winter. Data were analyzed using proportional hazards models, with adjustment for age, sex, and other potential confounders. RESULTS During follow-up, 87 cases (9%) of pneumonia occurred, 38 of which were fatal. The ACE DD allele (vs. ID + II) was associated with an increased risk of pneumonia (relative risk [RR] = 2.9; 95% confidence interval [CI]: 1.7 to 4.8, P < 0.001) and fatal pneumonia [RR = 4.4; 95% CI: 2.1 to 9.0; P < 0.0001). CONCLUSIONS The ACE D allele is an independent risk factor for pneumonia in elderly persons.

Usefulness of the resistive index in renal Doppler ultrasonography as an indicator of vascular damage in patients with risks of atherosclerosis

BACKGROUND Chronic kidney disease (CKD) is caused by various risk factors of cardiovascular disease (CVD). The estimated glomerular filtration rate (eGFR) is commonly used for the evaluation of the renal function in patients with CKD; however, it is difficult to assess the pathogenesis of CKD and predict the renal prognosis accurately using only eGFR. The resistive index (RI) in renal Doppler ultrasonography (RDU) is thought to be a good indicator of renal vascular resistance caused by atherosclerosis. In the present study, we investigated whether RI could be used to evaluate the pathogenesis of renal damage and predict the renal prognosis and investigated the correlation between RI and blood pressure (BP) fluctuations in patients with or without hypertension. METHODS The total study population included 194 patients (mean age: 66.2 years), who underwent RDU in our hospital ward between February 2009 and July 2010. We investigated the correlation between RI and multiple clinical parameters, including ambulatory blood pressure monitoring (ABPM). RESULTS RI significantly correlated with age, eGFR, diastolic BP, pulse pressure and level of albuminuria. Patients with diabetes mellitus (DM) showed a significantly higher RI than patients without DM, although their eGFR was similar; thus, DM might accelerate renal vascular damage and RI could detect earlier changes of vascular damage proceeding the time eGFR is reduced. Regarding ABPM, patients with a larger morning surge [systolic blood pressure (SBP) in the early morning--lowest SBP during sleep] showed a significantly higher RI. CONCLUSIONS The present study indicated that RI might be very useful for the evaluation of very early renal damage more effectively than eGFR and that diurnal BP change might be partly due to the progression of atherosclerotic change in the kidney evaluated by RI.

Alteration of vascular function is an important factor in the correlation between visit-to-visit blood pressure variability and cardiovascular disease.

Objective: To assess how visit-to-visit variability of SBP correlates with systemic atherosclerotic change and various prognoses. Background: Visit-to-visit SBP variability correlates with cardiovascular events. However, the mechanisms underlying the impact of visit-to-visit SBP variability on prognoses are poorly understood. Methods and results: A total of 485 patients with essential hypertension from the Non-Invasive Atherosclerotic Evaluation in Hypertension (NOAH) study cohort were included. We analyzed the correlation between visit-to-visit SBP variability and multiple clinical parameters. Next, we prospectively examined the correlation of SBP variability and frequency of cardiovascular disease (CVD) and total mortality. Patients with higher SBP variability exhibited significantly higher rates of statin use, as well as higher pulse wave velocity (PWV), left-ventricular mass index (LVMI), plaque score, and resistive index of the common carotid artery; these patients also exhibited lower estimated glomerular filtration rate. Kaplan–Meier analysis demonstrated that patients with higher SBP variability have a significantly higher incidence of CVD and mortality rate. The hazard ratio of SBP variability for incidence of CVD was greatly diminished after adjustment for intima–media thickness, plaque score, and resistive index, and was slightly diminished after adjustment for PWV and LVMI. Visit-to-visit SBP variability remained an independent risk factor for mortality after adjustment. Conclusion: Visit-to-visit SBP variability correlates significantly with systemic atherosclerotic change, incidence of CVD, and mortality rate. Altered arterial functions, such as macrovascular atherosclerosis and vascular resistance, are responsible for the correlations between visit-to-visit SBP variability and incidence of CVD.

Differences between daytime and nighttime blood pressure variability regarding systemic atherosclerotic change and renal function

Recently, new parameters related to hypertension, such as variability in blood pressure and ambulatory arterial stiffness index (AASI), were demonstrated to correlate with arteriosclerotic change. In this study, we investigated the correlation between circadian variability in blood pressure/AASI and renal function. We also investigated differences in the clinical impact of 24 h, daytime and nighttime blood pressure variability on renal and systemic atherosclerotic changes. We analyzed data from 120 patients who underwent renal Doppler ultrasonography (RDU) and ambulatory blood pressure monitoring (ABPM) at our hospital ward, and investigated the correlation between circadian variability in blood pressure/AASI and renal function, including resistive index (RI) evaluated with RDU, which is thought to be a good indicator of renal vascular resistance. Subjects with higher circadian variability in systolic blood pressure (SBP) had significantly higher RI. Daytime variability in SBP correlated more strongly with RI than nighttime variability. Meanwhile, only nighttime variability, but not daytime variability, in SBP was related to carotid atherosclerosis. Similarly, AASI was significantly correlated with RI. Circadian variability in SBP and AASI were both significantly correlated with renal function. Daytime SBP s.d. was especially more strongly correlated with renal vascular resistance, and nighttime SBP s.d. was significantly correlated with intima-media thickness (IMT) and plaque score. These results indicate that evaluating both daytime and nighttime blood pressure variability enables an assessment of pathological conditions in hypertensive patients to prevent cardiovascular diseases.

Serum uric acid is an independent risk factor for cardiovascular disease and mortality in hypertensive patients.

The purpose of the study was to investigate the association of serum uric acid (UA) levels in hypertensive patients with the prognosis for cardiovascular disease (CVD) and mortality. This hospital-based cohort study included 669 patients with essential hypertension. A questionnaire was used to identify patients in whom hypertensive complications had occurred, as well as causes of death. The primary end point of this study was new onset of stroke or CVD (new onset of angina pectoris, myocardial infarction or heart failure). We evaluated the baseline characteristics of patients, including UA levels, and assessed whether UA levels could be used to predict stroke and CVD. We also classified subjects into four groups according to the serum UA levels. During a mean follow-up period of 7.1±0.1 years, 71 strokes, 58 cases of CVD and 64 deaths were recorded. Kaplan–Meier analysis revealed that subjects in the high UA group had a higher frequency of stroke and CVD (P=0.0120) and total mortality (P=0.0021). A Cox proportional hazard model determined that, after adjusting for traditional risk factors, serum UA levels were predictive of CVD (relative risk=1.30; P=0.0073), stroke and CVD (relative risk=1.19; P=0.0141), mortality (relative risk=1.23; P=0.0353) and stroke CVD and mortality (relative risk=1.19; P=0.0083), but not stroke (P=0.4268). The significant correlations were particularly marked in women. Serum UA levels may be an independent risk factor for stroke and CVD in patients with essential hypertension, particularly women.

β -Adrenergic receptor gene polymorphism is a genetic risk factor for cardiovascular disease: a cohort study with hypertensive patients

Single-nucleotide polymorphisms (SNPs) of the β-adrenergic receptor (βADR) subtypes are related to hypertension and obesity. This hospital-based cohort study with hypertensive patients evaluated five βADR SNPs in association with cardiovascular events. The cohort included 357 hypertensive patients (male=181; mean age=61.5±11.8 years) seen between January 1998 and June 2004. The SNPs (Ser49Gly and Arg389Gly for β1ADR; Gly16Arg and Glu27Gln for β2ADR; Trp64Arg for β3ADR) were identified by PCR. We used Kaplan–Meier curves to assess the prognostic effect of these SNPs on cardiovascular disease (CVD). The SNP frequencies were Ser/Ser:Ser/Gly:Gly/Gly=243:104:10; Arg/Arg:Arg/Gly:Gly/Gly=256:95:6; Gly/Gly:Gly/Arg:Arg/Arg=71:201:85; Gln/Gln:Glu/Gln=308:49; and Trp/Trp:Trp/Arg:Arg/Arg=265:89:3. A total of 17 stroke and 15 coronary artery disease cases were recorded. By Kaplan–Meier analysis, the Ser/Ser SNP in Ser49Gly (P=0.0398), the Glu/Gln SNP in Glu27Gln (P=0.0390) and the Trp/Trp SNP in Trp64Arg (P=0.0132) were associated with lower event-free CVD survival (log-rank, Mantel–Cox model). A Cox proportional hazards model revealed that only the Trp/Trp SNP (P=0.0321) and age (P=0.0186) were independently related to lower event-free survival for CVD, adjusted for gender, diabetes, dyslipidemia, blood pressure, body mass index, medication and hypertensive complications. Combination Kaplan–Meier analysis of these three positive SNPs indicated a higher frequency of CVD among patients with the combination of Ser/Ser in Ser49Gly of β1, Glu/Gln in Glu27Gln of β2 and Trp/Trp in Trp64Arg of β3 (P=0.0209). These three SNPs, especially the Trp64Arg SNP of β3ADR, might be risk factors for CVD in hypertensive patients.

The combination of chronic kidney disease and increased arterial stiffness is a predictor for stroke and cardiovascular disease in hypertensive patients

To clarify the clinical utility of pulse wave velocity (PWV) and chronic kidney disease (CKD) in hypertension, we analyzed the prognostic impact of PWV and CKD on cerebrocardiovascular disease in hypertensive patients. This study consisted of 531 patients with essential hypertension (male/female=292/239, mean age=61.7±12.3, mean follow-up=7.0±3.0 years) and was performed between January 1998 and June 2004. We used questionnaires to assess stroke (n=57), cardiovascular diseases (CVDs; myocardial infarction, angina and congestive heart failure; n=44) and death (n=53) as primary end points. At baseline, we evaluated the carotid–femoral PWV (9.1±1.8 m s−1), the glomerular filtration rate and urinary protein excretions. We divided these subjects into those in the highest quartile of PWV and other subjects and into CKD (n=149) and non-CKD (n=458). We evaluated the prognostic influences of PWV and CKD with Kaplan–Meier analysis and Cox’s proportional hazard model. PWV in CKD (9.6±1.9 m s−1) was higher than in non-CKD (8.8±1.6 m s−1; P<0.0001), and creatinine was slightly decreased in the highest PWV group (1.09±0.35 mg dl−1, P<0.0001). On the basis of Kaplan–Meier analysis, the highest PWV group (PWV>10.1 m s−1; P=0.0003) and the CKD group (P=0.0005) showed significantly higher proportions of stroke and CVD events. In addition, the highest PWV group showed the highest percentage of stroke (P=0.0007), and the CKD group showed the highest proportion of CVD (P<00001). High PWV and CKD were independent predictors for stroke and CVD (P=0.0332) by Cox’s proportional hazard model. These data suggest that increased aortic stiffness and CKD may be predictors for stroke and cardiovascular events in hypertensive patients.

Carotid plaque score and intima media thickness as predictors of stroke and mortality in hypertensive patients.

The mean intima media thickness (IMT) and plaque score from carotid ultrasonography are both widely used to evaluate macrovascular atherosclerotic change. The present study sought to examine which parameter more effectively predicts patient prognosis. This hospital-based cohort study included 356 patients with essential hypertension (mean age: 62.4±0.6). We investigated how the mean IMT and plaque score correlated with various parameters, including pulse wave velocity (PWV), and we assessed the ability of the mean IMT and plaque score to predict cardiovascular events and total mortality. The mean IMT and plaque score significantly correlated with systemic atherosclerotic change, target organ damage, age and PWV. Subjects with a higher mean IMT and subjects with higher plaque scores showed higher frequencies of stroke and total mortality. In addition, subjects with marginal thickening of the intima media (mean⩾0.7) showed a significantly higher frequency of stroke than subjects with a mean IMT of <0.7. After adjustment for traditional risk factors, plaque score was significantly and independently predictive of stroke, and the predictive ability of the plaque score for the onset of stroke was equivalent to that of PWV. The mean IMT and plaque score showed a nonsignificant trend of higher risk of mortality after adjustment for traditional risk factors. The mean IMT and plaque score were significantly correlated with systemic atherosclerotic change. We revealed that plaque score predicted the onset of stroke more accurately than the mean IMT, and the accuracy of this prediction was equivalent to that from PWV in hypertensive patients. We also showed that marginal thickening of the intima media (as measured by mean IMT) may be a predictor of stroke.