【作者】 辛颖；

【导师】 惠汝太； 李建军； 何作祥； 陈敬洲；

【作者基本信息】 中国协和医科大学 ， 遗传学， 2009， 博士

【摘要】 第一部分内皮型一氧化氮合酶基因与高血压继发左心室肥厚的关联性研究研究背景左心室肥厚（LVH）是心血管疾病发病率和死亡率的独立预测危险因素。目前已知年龄、性别、绝经、生活方式与糖尿病等均是LVH的重要危险因素。虽然高血压的主要并发症是LVH,但是高血压继发LVH的患者其LVH的程度与高血压病史的长短、血压水平及降压后肥厚逆转的程度并不呈比例,提示有高血压以外的因素参与心肌肥厚进程。研究表明,遗传因素对左心室重量的影响约占非血压因素的60%。内皮型一氧化氮合酶（eNOS）可催化L-精氨酸生成NO,NO/NOS途径参与心肌肥厚的病理、生理过程。在动物模型中,长期给予NOS抑制剂（L-NAME）或者NOS3-/-小鼠均会出现LVH、心肌纤维化和心功能异常;建立NOS3-/-小鼠主动脉弓缩窄（TAC）压力超负荷左心室肥厚的动物模型,向心脏内注射eNOS后可以缓解LVH以及由此导致的心功能异常。此外,eNOS还可以抑制心梗后小鼠的心肌肥厚,保护其心脏的功能。心衰病人心室肌中eNOS基因（NOS3）的表达是增加的。关于NOS3与高血压继发LVH的关联研究目前仅一篇报道,其结果表明NOS3基因多态性与左室肥厚相关联。但是这种关联在其他人群中尚未得到证实。研究目的本研究拟在中国人群中验证NOS3多态性与高血压继发左心室肥厚的关联。对象和方法本研究选择两个独立的高血压继发左室肥厚的病例和对照人群（2179和343人）,采用聚合酶链反应-限制性片段长度多态性（PCR-RFLP）法研究eNOS基因的三个功能性多态性位点（-T786C/rs2070744、eNOS4a/b和+G894T/rs1799983）多态性与原发性高血压继发左室肥厚遗传易感因素的关系。所有入选者均进行心脏超声的检测。研究结果结果显示只有+G894T（Glu298Asp）位点与高血压继发左室肥厚的易感相关（第一个人群:OR=1.67,95%CI:1.19-2.36,P＜0.05;第二个人群:OR=1.41,95%CI:1.01-2.28,P＜0.05）,并呈隐性遗传模式。在两个独立样本中,与携带G等位基因（GT+GG）的相比,携带TT基因型的患者的室间隔厚度、左室后壁厚度、左室重量指数和相对室壁厚度均增加（分别增加16.2%和11.7%、8.3%和7.1%、14.0%和25.1%、13.1%和16.2%）且均有统计学意义（P＜0.01）。结论eNOS基因+G894T多态可能是高血压继发左室肥厚遗传易感性的标记之一。这为高血压继发左室肥厚遗传易感性提供了循证医学依据,同时也为在中国开展+G894T多态性与高血压继发左室肥厚关系的研究提供了基础。第二部分二甲基精氨酸-二甲胺水解酶基因与高血压继发左心室肥厚的关联性研究研究背景左心室肥厚（LVH）是心血管疾病发病率和死亡率的独立预测危险因素。目前已知年龄、性别、绝经、生活方式与糖尿病等均是LVH的重要危险因素。虽然高血压的主要并发症是LVH,但是高血压继发LVH的患者其LVH的程度与高血压病史的长短、血压水平及降压后肥厚逆转的程度并不呈比例,提示有高血压以外的因素参与心肌肥厚进程。研究表明,遗传因素对左心室重量的影响约占非血压因素的60%。非对称性二甲基精氨酸（ADMA）是一种内源性一氧化氮和酶（NOS）的抑制剂,研究显示ADMA与左心质量呈显著正相关,与射血分数呈显著负相关。NO可以调节心肌的生长,ADMA/NO途径对于心肌肥厚的病理过程是重要的。二甲基精氨酸-二甲胺水解酶（DDAH2）存在于表达内皮型一氧化氮合酶的组织中,可以代谢大部分的ADMA。关于DDAH2与高血压继发LVH的关联研究目前还没有相关报道。研究目的本研究拟在大样本人群中研究DDAH2多态性与高血压左心室肥厚的关联性。对象和方法研究对象为河南信阳高血压基地收集的2,274例高血压患者,其中包括1,135例高血压伴LVH患者,1,139例高血压不伴有LVH患者及767例健康对照者。所有入选者均进行心脏超声的检测。使用单倍体型-tagging-SNPs的方法,研究DDAH2基因tagSNPs与原发性高血压继发左室肥厚遗传易感因素的关系。并进一步运用萤光素酶分析研究DDAH2变异对基因表达的影响效应。研究结果本研究发现DDAH2只有一个tag-SNP,是位于启动子区的变异-449C/G（rs805305）,其与其他SNPs（rs70791 6,rs805304）完全连锁。对于-449C/G变异,与携带GG基因型的患者相比,携带C等位基因（GC+CC）的高血压左室肥厚患者的左室重量指数(62.0±1 5.7g/m^2.7vs.59.9±13.1g/m^2.7,P＜0.05)、左室后壁厚度(62.0±15.7g/m^2.7vs.59.9±13.1g/m^2.7,P＜0.05)以及相对室壁厚度（47.9±12.9%vs.46.0±11.0%,P＜0.01）均增加。此外,-449位携带C等位基因（GC+CC）增加高血压左心室肥厚的易感度（OR=1.26,95%CI:1.03-1.53,P＜0.01）。含有SNP-449C的DAH2启动子区明显降低转录活性（49.5%）。结论DDAH2-449C/G变异可能是高血压继发左室肥厚遗传易感性的基因标记之一。这为高血压继发左室肥厚遗传易感性提供了循证医学依据,同时也为在世界上开展-449C/G多态性与高血压继发左室肥厚关系的研究提供了基础。更多还原

【Abstract】 BackgroundLeft ventricular hypertrophy(LVH)is a strong,independent risk factor for cardiovascular morbidity and all-cause mortality.The following factors,age,male,gender, postmenopausal status in women,life style and diabetes,have been known to contribute to the development of LVH.Although hypertension is a major cause of LVH,the degree of LVH does not parallel the level of blood pressure,the duration of hypertension,or reversal of hypertensive LVH by pharmacological treatments.Indeed,genetic factors account for 60%of blood pressure-independent cardiac mass variances,suggesting the important role played by genetic factors in the development of LVH in essential hypertension.Endothelial nitric oxide synthase(eNOS)plays a critical role in the development of ventricular remodeling and cardiac hypertrophy.The purpose of the study was to determine whether three common variants in the eNOS gene(NOS3)are associated with risk of LVH in patients with essential hypertension.In animal models, long-term systemic inhibition of NO production with the NOS inhibitor L-NAME or NOS3-deficient mice have been shown to induce greater LVH,fibrosis and dysfunction, especially concentric LVH.Restoration of eNOS in the heart of NOS3-deficient mice has been shown to attenuate LVH and dysfunction in transverse ascending aorta constriction (TAC)model,and also inhibits hypertrophy in the remote myocardium and preserves cardiac function after myocardial infarction.Furthermore,the expression of NOS3 has been shown to be increased in the ventricular myocardium of failing human hearts, suggesting that the endothelial NO/NOS pathway has an important role in left ventricular remodeling and LVH.ObjectivesThe purpose of the study was to determine whether three common variants in the eNOS gene(NOS3)are associated with risk of left ventricular hypertrophy(LVH)in patients with essential hypertension.MethodsThree NOS3 genetic variants,-T786C(rs2070744),eNOS4a/b and +G894T(rs1799983) were genotyped in two independent case-control studies:the first study consisted of 1,061 hypertensive patients with LVH and 1,118 hypertensive patients without LVH,and the second sample consisted of 120 patients with LVH and 223 patients without LVH. Echocardiographic measurements were obtained in all the hypertensive patients.ResultsOnly the+G894T(Glu298Asp)variant of NOS3 was associated with higher risk of LVH (OR=1.67,95%CI:1.19-2.36,P＜0.01)in the first population;and replicated in the second population(OR=1.41,95%CI:1.01-2.28,P＜0.05)in a recessive model. Compared with carriers of the G allele(GT+GG),patients carrying the TT genotype had increased septal wall thickness(16.2%,P＜0.01;11.7%,P＜0.01,respectively);left ventricular posterior wall thickness(8.3%,P＜0.01;7.1%,P＜0.01,respectively);left ventricular mass index(14.0%,P＜0.01;25.1%,P＜0.01,respectively)and relative wall thickness(13.1%,P＜0.01;16.2%,P＜0.01,respectively)in the first and second populations.ConclusionsOur results support that homozygosity for +G894T(Glu298Asp)in NOS3 is a genetic risk factor for the development of LVH in patients with hypertension. BackgroundLeft ventricular hypertrophy(LVH)is a strong,independent risk factor for cardiovascular morbidity and all-cause mortality.The following factors,age,male,gender, postmenopausal status in women,life style and diabetes,have been known to contribute to the development of LVH.Although hypertension is a major cause of LVH,the degree of LVH does not parallel the level of blood pressure,the duration of hypertension,or reversal of hypertensive LVH by pharmacological treatments.Indeed,genetic factors account for 60%of blood pressure-independent cardiac mass variances,suggesting the important role played by genetic factors in the development of LVH in essential hypertension.Since nitric oxide(NO)modulates the growth of the myocardium, Asymmetric dimethylarginine(ADMA),an endogenous inhibitor of nitric oxide synthase (NOS)has been shown to be significantly related to left-ventricular mass and inversely related to ejection fraction.The bulk of ADMA is degraded by an enzyme named dimethylarginine dimethylaminohydrolase 2(DDAH2),which is found in tissues that express endothelial isoforms of nitric oxide synthase.ADMA/NO pathway is important for the pathogenesis of LVH.ObjectivesThe purpose of the study was to determine whether the common variant in the dimethylarginine dimethylaminohydrolase 2(DDAH2)gene is associated with risk of left ventricular hypertrophy(LVH)in patients with essential hypertension.MethodsThe case-control study consisted of 1,135 hypertensive patients with LVH and 1,139 hypertensive patients without LVH,as well as 767 control subjects.We used a haplotype-tagging single nucleotide polymorphisms(SNPs)approach to identify tag SNPs in DDAH2.The tag SNPs were genotyped in case-control study.The effect of DDAH2 variants on gene expression was studiedby use of luciferase reporter assays. ResultsA promoter variant-449C/G(rs805305)was identified and found to be the only tag-SNP in completely linkage disequilibrium with other SNPs(rs707916,rs805304)in the region containing DDAH2.Compared with the carriers of the GG genotype,the hypertensive patients with LVH carrying the C allele(GC+CC)had an increase in the LV mass index (62.0±15.7 g/m2.7 vs.59.9±13.1 g/m2.7,P＜0.05),posterior wall thickness(12.1±2.9 mm vs.11.4±1.9 mm,P＜0.01),and relative wall thickness(47.9±12.9%vs.46.0±11.0%, P＜0.01).In addition,the C allele(GC±CC)of the -449 locus was associated with higher risk ofLVH(OR=1.26,95%CI:1.03-1.53,P＜0.01)in the study.The SNP-449C-bearing DDAH2 promoter exhibited 49.5%of lower transcription activity than the SNP-449G-bearing promoter.ConclusionsOur results support that the DDAH2 common variant is a genetic risk factor for the development of LVH in patients with essential hypertension.更多还原