Lina Zhang

Polymorphism of tumor necrosis factor alpha (TNF-alpha) gene promoter, circulating TNF-alpha level, and cardiovascular risk factor for ischemic stroke

BackgroundTumor necrosis factor-α (TNF-α) is one of the most typical pro-inflammatory cytokines with both beneficial and destructive properties for the central nervous system. Increasing evidences have demonstrated the important role of TNF-α in the development of ischemic stroke, but studies examining the possible association with stroke or direct functional effects of polymorphisms in TNF-α have been contradictory.FindingsIn this study, a 2-kb length of the proximal promoter of the TNF-α was screened and four polymorphisms were investigated in the case–control study. Our data confirmed the association between -308G/A variant with stroke in 1,388 stroke patients and 1,027 controls and replicated in an independent population of 961 stroke patients and 821 controls (odds ratio (OR)u2009=u20091.34, 95% confidence interval (CI) =1.02 to 1.77 and ORu2009=u20091.56, 95% CIu2009=u20091.09 to 2.23, respectively). To reconcile the association between polymorphisms and stroke and to give a comprehensive picture of the genetic architecture of this important gene, we performed a meta-analysis of 15 published studies in an Asian population. Our results demonstrated an association between rs1800629 and ischemic stroke (ORu2009=u20091.43, 95% CIu2009=u20091.21 to 1.69). Another meta-analysis results of 14 studies demonstrated that ischemic stroke patients have higher serum TNF-α level than the control subjects (standardized mean difference (SMD) = 2.33, 95% CI = 1.85 to 2.81). In vitro evaluation of potential interaction between variants of the TNF-α gene (−308G/A, -857C/T, and -1031T/C) demonstrated that these three polymorphisms could interact together to determine the overall activity of the TNF-α gene.ConclusionsThese findings strongly implicate the involvement of TNF-α in the pathogenesis of stroke.

A functional variant in APOA5/A4/C3/A1 gene cluster contributes to elevated triglycerides and severity of CAD by interfering with microRNA 3201 binding efficiency.

BACKGROUNDnRecent genome-wide association studies identified the APOA5/A4/C3/A1 gene cluster polymorphisms influencing triglyceride level and risk of coronary artery disease (CAD).nnnOBJECTIVESnThe purposes of this study were to fine-map triglyceride association signals in the APOA5/A4/C3/A1 gene cluster and then explore the clinical relevance in CAD and potential underlying mechanisms.nnnMETHODSnWe resequenced the APOA5/A4/C3/A1 gene cluster in 200 patients with extremely high triglyceride levels (≥10 mm/l) and 200 healthy control subjects who were ethnically matched and genotyped 20 genetic markers among 4,991 participants with Chinese Han ethnicity. Subsequently, 8 risk markers were investigated in 917 early-onset and 1,149 late-onset CAD patients, respectively. The molecular mechanism was explored.nnnRESULTSnBy resequencing, a number of newly and potentially functional variants were identified, and both the common and rare variants have remarkable cumulative effects on hypertriglyceridemia risk. Of note, gene dosage of rs2266788 demonstrated a robust association with triglyceride level (p = 1.39 × 10(-19)), modified Gensini scores (p = 1.67 × 10(-3)), and numbers of vascular lesions in CAD patients (odds ratio: 1.96, 95% confidence interval: 1.31 to 2.14, p = 8.96 × 10(-4)). Functional study demonstrated that the rs2266788 C allele destroyed microRNA 3201 binding to the 3 UTR of APOA5, resulting in prolonging the half-life of APOA5 messenger RNA and increasing its expression levels.nnnCONCLUSIONSnGenetic variants in APOA5/A4/C3/A1 gene cluster play an important role in the regulation of plasma triglyceride levels by an increased APOA5 concentration and contribute to the severity of CAD.

Decreased Peripheral Mitochondrial DNA Copy Number is Associated with the Risk of Heart Failure and Long-term Outcomes.

AbstractMitochondrial DNA (mtDNA) copy number variation (CNV), which reflects the oxidant-induced cell damage, has been observed in a wide range of human diseases. However, whether it correlates with heart failure, which is closely related to oxidative stress, has never been elucidated before. We aimed to systematically investigate the associations between leukocyte mtDNA CNV and heart failure risk and prognosis.A total of 1700 hospitalized patients with heart failure and 1700 age- and sex-matched community population were consecutively enrolled in this observational study, as well as 1638 (96.4%) patients were followed prospectively for a median of 17 months (12–24 months). The relative mtDNA copy number of leukocyte of peripheral blood or cardiac tissue was measured in triplicate by quantitative real-time PCR method.Patients with heart failure possessed much lower relative mtDNA copy number compared with control subjects (median 0.83, interquartile range [IQR] 0.60–1.16 vs median 1.00, IQR 0.47–2.20; Pu200a<u200a0.001), especially for the patients with ischemic etiology (median, 0.77 for ischemic and 0.91 for non-ischemic, Pu200a<u200a0.001). Patients with lower mtDNA copy number exhibited 1.7 times higher risk of heart failure (odds ratio 1.71, 95% confidence interval [CI] 1.48–1.97, Pu200a<u200a0.001). Long-term follow-up (median of 17 months) showed that decreased mtDNA copy number was significant associated with both increased cardiovascular deaths (hazard ratio [HR] 1.58, 95% CI 1.16–2.16, Pu200a=u200a0.004) and cardiovascular rehospitalization (HR 1.48, 95% CI 1.21–1.82, Pu200a<u200a0.001). After adjusting for the conventional risk factors and medications, lower mtDNA copy numbers were still significantly associated with 50% higher cardiovascular mortality (Pu200a=u200a0.035).In conclusion, mtDNA copy number depletion is an independent risk factor for heart failure and predicts higher cardiovascular mortality in patients with heart failure.

Comparison of High-Resolution Melting Analysis, TaqMan Allelic Discrimination Assay, and Sanger Sequencing for Clopidogrel Efficacy Genotyping in Routine Molecular Diagnostics

Clopidogrel, as a routine antiplatelet drug, is widely used in patients to reduce cardiovascular events following percutaneous coronary intervention. Because of genetic variation, patients undergoing percutaneous coronary intervention show differing responses to clopidogrel therapy. Recently, five single nucleotide polymorphisms (SNPs) within CYP2C19 (rs4244285, rs4986893, rs12248560), ABCB1 (rs1045642), and ITGB3 (rs5918) were identified that contribute prominently to variability in response to clopidogrel. Given that Sanger sequencing is labor intensive and time consuming, rapid genotyping methods for SNP detection are urgently required before clopidogrel therapy. Accordingly, we developed a high-resolution melting analysis (HRMA) and TaqMan allelic discrimination assay (TaqMan) to genotype those five SNPs, and compared these two assays with Sanger sequencing on accuracy of genotyping as well as operational characteristics. These two assays showed high accuracy (0.995, 95% CI 0.991 to 0.998 for HRMA; 0.997, 95% CI 0.994 to 0.999 for TaqMan, respectively), sensitivity (0.996, 95% CI 0.989 to 1.002 for HRMA; 0.998, 95% CI 0.993 to 1.002 for TaqMan, respectively), and specificity (0.995, 95% CI 0.991 to 0.999 for HRMA; 0.996, 95% CI 0.993 to 1.000 for TaqMan, respectively). Our study indicates that HRMA and TaqMan are easier to operate and obviously faster than Sanger sequencing. In conclusion, HRMA and TaqMan are rapid, convenient, and reliable assays forxa0clopidogrel efficacy genotyping.

Association Analysis of Polymorphisms in ROCK2 with Cardiovascular Disease in a Chinese Population

Background Rho-kinase (ROCK) has been shown to play an important role in cardiovascular disease such as coronary artery disease (CAD) and hypertension. Recently, common variants of ROCK2 have been reported to influence blood pressure, but the relationship between common ROCK2 variants and cardiovascular disease has not been extensively studied in the Chinese population. Methods To derive a more precise estimation of their relationship, we screened for the common variants by direct sequencing of all exons of ROCK2, and then we performed genetic association analyses in a CAD case–control study, including a total of 1344 cases and 1267 ethnically and geographically matched controls. Results Unconditional logistic regression showed that no significant association between common variants in the coding region of ROCK2 and CAD was observed in our study (for rs978906, ORu200a=u200a0.92, 95% CI 0.72–1.20 and Pu200a=u200a0.63; for rs2230774, ORu200a=u200a0.90, 95% CI 0.70–1.16 and Pu200a=u200a0.47; for rs56304104, ORu200a=u200a0.97, 95% CI 0.70–1.31 and Pu200a=u200a0.83; respectively). Conclusions The relationship between the ROCK2 polymorphisms and cardiovascular disease risk cannot be entirely discounted and warrants further evaluation in a large population.

Development of a high resolution melting method for genotyping of risk HLA-DQA1 and PLA2R1 alleles and ethnic distribution of these risk alleles.

Recent studies have demonstrated that alleles at single nucleotide polymorphisms (SNPs) rs2187668 and rs4664308 within genes HLA-DQA1 and PLA2R1, respectively, had a significant impact on the susceptibility to idiopathic membranous nephropathy (IMN). Analysis of the two genomic loci could identify alleles for individuals at risk for IMN. Conventional methods for genotyping are labor intensive, expensive or time consuming. High resolution melting (HRM) is a new technique for genotyping and has the advantages of simplicity, speed, high sensitivity and low cost. Here, we describe genotyping of SNPs rs2187668 and rs4664308 using HRM. In this study, we identified polymorphisms of rs2187668 and rs4664308 in 480 healthy unrelated Chinese volunteers of two ethnic groups from three different geographical areas in China. The two genomic loci were genotyped by HRM using a saturating fluorescent dye SYTO® 9 on 7900 HT and RG 6000 instruments, and were further confirmed by direct DNA sequencing. Three different SNP genotypes were sufficiently distinguished by HRM with mean sensitivity of 98.8% and mean error rate of 1.9%. In addition, the allele frequencies varied greatly based on ethnic or geographic origins. In conclusion, HRM is a rapid, cost efficient, sensitive, suitable technique for genotyping, and simple enough to be readily implemented in a diagnostic laboratory. We believe this will be a valuable technique for determining the genotype of rs2187668 and rs4664308 and for assessing individual susceptibility to IMN.

Genotyping on ALDH2: Comparison of Four Different Technologies

Objectives This study aimed to compare the accuracy and performance of four genotyping methods for detecting single nucleotide polymorphisms (SNPs) in aldehyde dehydrogenase-2 (ALDH2), which is the principal enzyme involved in alcohol metabolism. Design and Methods We genotyped rs671 of ALDH2 in 96 coronary heart disease (CHD) patients with four methods including high resolution melting analysis (HRM), TaqMan allelic discrimination assay (TaqMan), allele-specific PCR (AS-PCR) and pyrosequencing. Meanwhile, we compared the accuracy and performance of these methods. Results All selected patients were successfully genotyped with referred methods. The results of these four assays showed 100% concordant results and had 100% accuracy as verified by Sanger sequencing. Conclusions All of the referred methods can be used for genotyping ALDH2 rs671 with the same accuracy compared to Sanger sequencing. In small size of clinical samples, HRM and AS-PCR outperform over others due to their lower cost and less hands-on operation, which are suitable for clinical application.

Lack of causal relationship between leukocyte telomere length and coronary heart disease

OBJECTIVEnTo investigate the association between genetic variation in telomerase RNA component (TERC) and leukocyte telomere length (LTL) with risk of coronary heart disease (CHD).nnnMETHODS AND RESULTSnAn analysis of LTL was conducted, focusing on two SNPs in 2 community-based cohort populations comprising 3500 Chinese Han individuals. In addition, LTL ratio was determined in a case-control setting involving 4351 participants: 2211 healthy individuals and 2140 CHD patients. The association between LTL and the presence and extent of cardiovascular and cerebrovascular lesions were tested. Results confirmed the association of rs12696304 and rs16847897 with LTL in the Chinese Han population (P=1.63×10(-6) and P=1.44×10(-7), respectively). However, these SNPs confer a moderate risk for CHD but did not achieve significant threshold after multiple corrections. Decreased LTL ratio was associated with CHD (odds ratio [OR], 1.13; 95% confidence interval [CI], 1.02-1.34; P<0.01). In addition, the LTL ratio in CHD patients was related to numbers of vascular disease lesions.nnnCONCLUSIONSnOur results do not support a causal role of LTL for the development of CHD. However, LTL may be related to complex conditions associated with cardiovascular and cerebrovascular disease manifestations.

Effects of early age at natural menopause on coronary heart disease and stroke in Chinese women

BACKGROUNDnMenopause is identified as a risk factor for cardiovascular disease because of the change of estrogen. The objective of the study was to explore the associations between early age at natural menopause (menopause at an age≤45years) and the presence of CHD and stroke.nnnMETHODSnThe study subjects were from the first follow-up survey of the Dongfeng-Tongji cohort study. A total of 16,515 postmenopausal women were included for the analysis. Logistic regression models were used to examine the associations between age at natural menopause (≤45, 45-52, >52years) and the presence of CHD and stroke adjusted for sociodemographic characteristics, lifestyle, reproductive history and metabolic factors.nnnRESULTSnIn the fully adjusted model, for each 1-year delay in menopausal age, the prevalence of CHD and stroke was reduced by 3% (OR, 0.97; 95% CI, 0.95-0.98) and 5% (OR, 0.95; 95% CI, 0.92-0.98), respectively. Women with early menopause (≤45years) had a higher prevalence of CHD (OR, 1.33; 95% CI, 1.13-1.57) compared with those with menopause at ages 45-52years. Similarly, women with early menopause (≤45years) was associated with higher prevalence of stroke (OR, 1.69; 95% CI, 1.25-2.30) compared with those with menopause at ages 45-52years.nnnCONCLUSIONSnEarly age at natural menopause is significantly associated with the presence of CHD and stroke among Chinese women.

Gene variants in responsiveness to clopidogrel have no impact on clinical outcomes in Chinese patients undergoing percutaneous coronary intervention — A multicenter study

BACKGROUNDnGene variants contribute to variability in individual responsiveness to clopidogrel and influence cardiovascular outcomes in Caucasian patients with acute coronary syndrome (ACS). However, limited data is available in Asian populations.nnnMETHODSnWe resequenced 14 genes in metabolizing and activity pathway of clopidogrel in 138 patients with ACS and prospectively assessed the modulating effects of 13 variants possibly related to clopidogrel efficacy on one-year cardiovascular event occurrence in 5820 ACS patients after percutaneous coronary intervention (PCI). In addition, platelet aggregation rate was measured in 1084 participants and plasma levels of active metabolite were determined in 15 patients to test whether increasing clopidogrel maintenance doses increases active metabolite exposure.nnnRESULTSnNo significant associations were found between any of the tested variants and risk of cardiovascular events (P>0.05), although CYP2C19*2 carriers had slightly higher on-treatment platelet aggregation rate and lower active metabolite exposure compared with that of non-carriers (Median [IQR] 51.49 [35.43-66.75] vs. 49.05 [32.36-63.38], P=0.012) (means±SD AUC, 22.84±5.00 vs. 35.05±12.34, P=0.008). Switching from 75mg daily clopidogrel to 150mg daily fully overcomes low exposure to clopidogrel active metabolite in CYP2C19*2 carriers (means±SD AUC, 32.35±8.65 vs. 35.05±12.34, P=0.314).nnnCONCLUSIONnDifferent from Caucasian populations, genetic variants have no significant influence on clinical outcomes and have much milder effects on inhibition of platelet and active clopidogrel metabolite levels in Chinese patients with ACS after PCI, an effect which could be overcome with a dose escalation to 150mg daily.