【作者】 刘海鹏；

【导师】 徐希平；

【作者基本信息】 中国科学技术大学 ， 人类遗传学， 2009， 博士

【摘要】 原发性高血压是由多种遗传和环境因素交互作用引起的一类复杂疾病。我国现有高血压患者2亿人,用于高血压的医疗费达366亿元。临床研究表明,平均降低收缩压10mm Hg、舒张压5mm Hg,脑卒中发生危险下降40%-50%,冠心病发生危险下降15%-20%,心力衰竭发生危险减少50%。因此,有效降低高血压患者的血压水平,对预防其并发症具有十分重要的作用。高血压药物治疗中出现相当程度的个体差异。MATH试验显示,单独使用硝苯地平缓释剂治疗高血压的有效率为76%。造成这种药物治疗个体差异的原因还不完全清楚,目前认为有其遗传学基础。研究表明,除了编码药物靶标和主要代谢酶的基因以外,参与调解血压的候选基因可能也对降压药物个体间疗效差异产生影响。交感神经系统在原发性高血压发病过程中起到重要作用,其主要作用途径是通过儿茶酚胺类物质作用于G蛋白偶联α和β肾上腺素受体。研究发现,高血压患者的心血管组织会出现交感神经功能上调,并出现不同类型的肾上腺素受体分布及密度的平衡发生变化。在中老年人群中,高血压常和下尿路综合征(良性前列腺增生是其中男性患者的主要诱因)并存,增加了治疗的难度,并使患者的生活质量进一步下降,个人和社会负担进一步加重。中国人群中65岁以上男性同时患有高血压和中重度下尿路症状的患者达42.7%。高血压可能增加患者下尿路症状的发病风险。目前临床上这两种疾病状态通常分别由心血管内科和泌尿科分别进行治疗,往往顾此失彼,达不到满意的疗效和明显改善患者生活质量的目的。因此,研究影响抗高血压药物治疗的遗传因素,以及为临床综合治疗高血压及其合并的下尿路综合征提供个体化治疗依据,具有重要的临床价值和公共卫生学意义。本研究首先开展肾上腺素受体基因多态性与L型钙通道阻断剂硝苯地平降血压疗效的相关性研究,在其主要发现的基础上进一步开展L型钙通道阻断剂氨氯地平联合α1肾上腺素受体拮抗剂特拉唑嗪降血压和改善下尿路功能紊乱作用的动物试验探索,并设计随机、对照、双盲临床试验,对L型钙通道阻断剂氨氯地平联合α1肾上腺素受体拮抗剂特拉唑嗪降血压和改善下尿路综合征的疗效和安全性进一步验证。1、肾上腺素受体多态性与硝苯地平降压疗效的相关性研究目的:研究α1A肾上腺素受体Arg347Cys和β2肾上腺素受体Arg16Gly和Gln27Glu多态性是否影响硝苯地平控释片的降血压疗效。方法:符合入选标准的中国安徽高血压患者(收缩压140-200mmHg和舒张压＜115mmHg或收缩压＜200mmHg和舒张压90-115mmHg)接受硝苯地平控释片30 mg口服治疗15天,每天1次。采用Taqman方法检测α1A受体基因T1475C(Arg347Cys)多态性和β2受体基因A265G(Arg16Gly)及C298G(Gln27Glu)基因多态性。采用高效液相方法紫外条件测定第16天硝苯地平稳态血药浓度。采用多元线性回归方法分析447名患者基因多态性与硝苯地平控释片降压疗效的关系。结果:在校正了重要的协变量后,与携带α1A受体基因Arg347Arg基因型的患者比较,携带347Cys等位子(Arg347Cys/Cys347Cys)的患者收缩压降低更为显著(adjustedβ(se)=4.1(1.5),P=0.006),但是舒张压的下降没有达到统计学上的显著性水平(adjustedβ(se)=1.2(0.8),P=0.101)。然而,不管校正重要的协变量前后,没有发现β2受体基因两个多态性位点与血压降低水平的相关性。结论及提示:我们的研究发现,α1A受体基因Arg347Cys多态性可能对预测中国高血压患者硝苯地平控释片的降压反应具有一定价值。这一发现提示L型钙通道可能与α肾上腺素受体在血压调节方面存在交互作用,而L型钙通道阻断剂与α肾上腺素受体拮抗剂在血压控制方面也可能存在机制的互补。2、L型钙通道阻断剂联合α1受体拮抗剂对高血压大鼠血压及对下尿路功能紊乱大鼠尿流动力学改善作用试验目的:基于研究1的发现,同时通过文献调研发现,高血压和下尿路综合征常同时发生于老年人群中,L型钙通道对调节下尿路功能也有重要作用,L型钙通道阻断剂对改善某些下尿路症状,如膀胱过度活动症状也有作用,而α1肾上腺素受体拮抗剂同时具有抗高血压和改善下尿路综合征两方面的适应症。本部分动物研究在以上发现和提示的基础上进一步探索L型钙通道阻断剂单独或者与α肾上腺素受体拮抗剂联用是否会同时有效控制这两种疾病状态。包括:研究L型钙通道阻断剂氨氯地平与α1受体拮抗剂特拉唑嗪组合对二肾一夹型高血压模型大鼠的降血压作用;研究氨氯地平单用,或者联合特拉唑嗪,对良性前列腺增生大鼠和逼尿肌不稳定大鼠尿动力学的改善作用。方法:1、两肾一夹法造肾性高血压大鼠模型。成模大鼠按照体重和血压半随机分组,即:模型组、氨氯地平组(0.5 mg/kg)、特拉唑嗪组(0.4 mg/kg)、氨氯地平联合特拉唑嗪组(0.5+0.4 mg/kg)。另设假手术对照组(行手术但不结扎肾动脉)。各组连续灌胃给药28天,模型组和假手术对照组给予等容生理盐水。采用尾动脉无创血压测定仪测量给药前及给药后第1、2、3、4周各组大鼠血压变化,观察氨氯地平+特拉唑嗪对高血压大鼠血压的影响;2、雄性Sprague-Dawley大鼠皮下注射睾酮(0.5 mg/每只,连续21天)诱导产生良性前列腺增生模型,雌性Sprague-Dawley大鼠膀胱出口不完全结扎6周诱导产生逼尿肌不稳定模型。模型大鼠随机分组后分别给予不同剂量的氨氯地平、特拉唑嗪或者氨氯地平联合特拉唑嗪,连续14天。末次给药后,双腔套管经膀胱顶部插入膀胱中,生理记录仪记录大鼠尿动力学指标,并测定膀胱指数和前列腺指数,观察药物对上述指标的改善作用。结果:1、与高血压模型组大鼠比较,氨氯地平组和特拉唑嗪组收缩压和舒张压均显著下降(P＜0.05或P＜0.01);氨氯地平联合特拉唑嗪组与氨氯地平或特拉唑嗪组比较,收缩压和舒张压下降更为显著(P＜0.05或P＜0.01);2、与良性前列腺增生模型组大鼠比较,氨氯地平0.5,1和3 mg/kg显著降低良性前列腺增生大鼠的膀胱指数,排尿压阈值和排尿压峰值,并显著其延长排尿间隔时间。氨氯地平0.5 mg/kg联合特拉唑嗪0.4 mg/kg对良性前列腺增生大鼠和逼尿肌不稳定大鼠尿动力学的改善最为显著,与两药单独使用比较,进一步降低了其膀胱指数,排尿压阈值和排尿压峰值,并进一步延长了其排尿间隔时间,与特拉唑嗪1 mg/kg作用相当。与模型组比较,氨氯地平0.5 mg/kg联合特拉唑嗪0.4 mg&g显著降低了逼尿肌不稳定大鼠的非排尿性膀胱收缩频率(2.0/4.5,P＜0.01)。结论及提示:1、氨氯地平联合特拉唑嗪可有效降低高血压模型大鼠血压,并显著优于两药单独使用;2、氨氯地平可改善良性前列腺增生和逼尿肌不稳定大鼠的下尿路功能紊乱症状。这一研究结果支持L型钙通道阻断剂与α1受体拮抗剂联合降低高血压的加性效应,并提示氨氯地平和特拉唑嗪低剂量的联合使用具有治疗下尿路综合征的临床潜力,并可能特别适用于具有膀胱过度活动的患者。3、L型钙通道阻断剂联合α1受体拮抗剂降血压和改善下尿路综合征的随机、对照、双盲临床试验目的:高血压和下尿路综合征常同时发生于老年男性中。为进一步验证研究2的发现,我们继续将氨氯地平和特拉唑嗪作为研究用药,通过本临床实验来研究高血压合并下尿路综合征时L型钙通道阻断剂联合α1受体拮抗剂的疗效与安全性。方法:355名患有1-2期原发性高血压合并下尿路综合征[定义为国际前列腺症状评分,International Prostate Symptom Score(IPSS)≥10]的男性患者随机分入2 mg特拉唑嗪治疗组(n=117)、5 mg氨氯地平治疗组(n=119)或5 mg氨氯地平加2 mg特拉唑嗪治疗组(n=119),各组每天服药一次,共28天。本研究的主要疗效终点是国际前列腺症状评分的总评分和亚评分以及血压变化。采用意向性治疗(intention to treat,ITT)人群数据集进行分析。本试验已在ClinicalTrials.gov网站注册(No.NCT00693199)。结果:治疗28天后,与特拉唑嗪或氨氯地平组比较,氨氯地平联合特拉唑嗪组达到最佳的血压降低水平(25.2±15.7 mmHg/11.9±16.3 mmHg或21.8±13.9 mmHg,P＜0.01或P＜0.05),与特拉唑嗪组比较,血压控制率显著升高(73.1%/36.8%,P＜0.001)。与特拉唑嗪组比较,氨氯地平联合特拉唑嗪组改善刺激性症状亚评分有效率更高(53.8%/39.3%,P＜0.05);与氨氯地平组比较,联合用药组改善生活质量评分作用更为显著(1.4±1.1/1.1±1.1,P＜0.05),改善生活质量评分有效率也更高(47.1%/33.6%,P＜0.05)。三个治疗组的患者均对药物耐受良好。结论及提示:本次的4周随机对照双盲试验显示,中国男性高血压合并下尿路综合征患者中,较低剂量的氨氯地平联合特拉唑嗪可能是一种安全和有效的同时控制这两种疾病的药物疗法,特别是对具有突出的膀胱过度活动的患者,印证了研究2的发现。更多还原

【Abstract】 It is widely recognized that essential hypertension is a complex trait resulting from the interaction of environmental and genetic factors.It affects about 0.2 billion individuals in China and in 2008,a total of 0.266 billion Renmingbi were expended on the treatment of hypertension.Previous clinical trials have demonstrated that an average reduction of 10 mm Hg in systolic blood pressure or 5 mm Hg in diastolic blood pressure is associated with a 40-50%reduction in stroke,15-20%in coronary artery disease,and 50%in congestive heart failure.Thus,the effective reduction in blood pressure plays an important role to prevent it’s related complications.However, considerable inter-individual variation was observed in the treatment of hypertension. MATH trial showed that responsive rate of nifedipine GITS on hypertension is 76%. The mechanisms underlying such inter-individual variation are yet unclear.One of the possible explanation is the individuals have different genetic background. Besides those encoding the target proteins and the key metabolic enzymes,genes involving in the regulation of blood pressure may also contribute to such inter-individual variation in antihypertensive treatment.The sympathetic nervous system plays an important role in the pathogenesis of essential hypertension,mainly through the catecholamines acting on G protein-coupledαandβadrenoceptors.A chronic increase in the sympathetic functions as well as the alterations in the balance of adrenoceptors in cardiovascular tissues is found in many hypertensive patients.In old population,hypertension and lower urinary tract symptoms are the common comorbid conditions,both of which may be caused by increased overactivity of the sympathetic system.This makes it more difficult to treat these patients properly and efficiently.Furthermore,it will reduce the quality of life of those patients and increase the economical burden individually and socially.In Chinese male population aged more than 65 years,42.7%suffer from hypertension and lower urinary tract symptoms simultaneously.Hypertension may increase the risk of developing lower urinary tract symptoms.Given the high comorbidity of hypertension and LUTS,it is important to consider and investigate therapeutic regiment that can effectively and safely manage both conditions in a certain patient, rather than treating the two conditions separately.Based on these evidences,it is valuable to study the genetic factors underlying the antihypertensive treatment and to find more ways and personalized medications for effectively treating hypertension and the co-existing lower urinary tract symptoms.This study aimed to 1) evaluate whether the polymorphisms in theα1A andβ2 adrenoceptor genes were associated with the antihypertensive effects of nifedipine in Chinese population;2) investigate the efficiacy of the L-type calcium channel blocker amlodipine combined withα1 adrenergic antagonist terazosin on the simultaneous treatment of hypertension and lower urinary tract disorder in model rats;3) by applying a randomized,double-blind clinical trial,evaluate the efficacy and safety of those two drugs used in combination on the treatment of hypertension and lower urinary tract symptoms.1.Association between Arg347Cys polymorphism ofα1A-adrenoceptor gene, Arg16Giy and Gln27Glu polymorphisms of Beta 2 adrenoceptor gene and blood pressure lowering effect of nifedipine[Aims]:To investigate whether polymorphisms in theα1A andβ2 adrenoceptor genes influence antihypertensive effect of nifedipine GITS.[Methods]:Hypertensive patients,aged 35-60 years old,not taking antihypertensive medications for 4 weeks prior to this study,received daily treatment with an oral dosage of 30 mg nifedipine GITS for 16 days.Genotypes of the Arg347Cys polymorphism in theα1A-adrenoceptor gene,Arg16Gly and Gln27Glu polymorphisms in theβ2-adrenoceptor gene were determined by TaqMan SNP genotyping assay.The 16^th day steady state plasma concentration of nifedipine was measured using high performance liquid chromatography with ultraviolet detection.Multivariate linear regression was performed in a total of 447 patients to evaluate the impacts of these polymorphisms on the blood pressure response to nifedipine GITS.[Results]:1) Patients carrying the 347Cys allele of theα1A-adrenoceptor gene had a greater systolic blood pressure reduction than those carrying two Arg347 alleles of theα1A-adrenoceptor gene（27.3±15.5mmHg versus 32.5±14.0mmHg,P=0.006）.2) Diastolic blood pressure reduction was not associated with the Arg347Cys polymorphism in theα1A-adrenoceptor gene;3) No significant associations were observed between blood pressure reduction and two polymorphisms in theβ2-adrenoceptor gene.[Conclusion]:The Arg347Cys polymorphism in theα1A-adrenoceptor gene may predict the blood pressure response to nifedipine GITS in Chinese hypertensive patients. 2.Effects of L-type calcium channel blocker amlodipine or combined withα1 adrenergic antagonist terazosin on hypertension and lower urinary tract disorder in rat models[Aims]:To investigate the blood pressure lowering effects of amlodipine combined with terazosin in hypertension rat model and to investigate the effects of amlodipine, alone or combined with terazosin on urodynamics in rats with benign prostatic hyperplasia（BPH） and in female rats with detrusor instability（DI）.[Method]:Two kidney 1 clip operation was conducted to induce hypertensive rat model.Model rats were semi-randomly assigned to four groups according to their body weight and blood pressure level:Model control group,amlodipine group（0.5 mg/kg）,terazosin group（0.4 mg/kg） and amlodipine plus terazosin group（0.5 + 0.4 mg/kg）.All the rats were intragastrically administered with assigned drugs for 28 days. Non-invasive blood pressure measurement was conducted in week 1,2,3 and 4. Rat models of BPH were induced by subcutaneous injection of testosterone（0.5 mg per rat for 21 days）.Female rat models of DI were induced by partial bladder outlet ligation for 6 weeks.Model rats were intragastrically administered with assigned drugs（amlodipine,terazosin or combination of these two drugs） for 14 days in the three experiments.Continuous cystometry was performed under urethane anesthesia. [Results]:1) Systolic blood pressure（SBP） and diastolic blood pressure（DBP） in model rats were significantly increased at each timepoint.SBP and DBP in amlodipine and terazosin group were all significantly decreased.Compared with the two monotherapy group,the amlodipine plus terazosin group reached the greatest reduction on SBP and DBP.2) Amlodipine 0.5,1 and 3 mg/kg significantly decreased bladder index（BI）,threshold pressure（TP）,micturition pressure（MP） and significantly increased inter-micturition duration（IMD） in BPH rats.Moreover, amlodipine 0.5 mg/kg plus terazosin 0.4 mg/kg obtained the greatest improvements in all urodynamic parameters and reported greater decreased BI,TP,MP and greater increased IMD compared to either of the two drugs used alone in BPH and DI rats and showed similar efficacy compared to terazosin 1 mg/kg.The combination treatment also reached greater decreased nonvoiding bladder contraction（NVC） in DI rats.[Conclusions]:1) Amlodipine plus terazosin could efficiently decrease blood pressure in model rats,which showed more advantage on blood pressure lowering effects than amlodipine alone or terazosin alone.2) Amlodipine or combined with terazosin may have a potential for alleviating lower urinary tract symptoms（LUTS）. Meanwhile,the combination therapy appears to be more suitable for LUTS with predominant irritative symptoms.3.Efficacy of combined amlodipine and terazosin therapy in male hypertensive patients with lower urinary tract symptoms:a randomized,double-blind clinical trial[Aims]:Based on the findings in the second part of the study,this trial was to investigate the therapeutic efficacy and safety of amlodipine alone or in combination with terazosin for the presence of hypertension.[Methods]:A total of 355 patients with Stage 1 or 2 hypertension and LUTS（as defined by an International Prostate Symptom Score of≥10） were randomly assigned to receive 2 mg of terazosin（n=117）,5 mg of amlodipine（n=119）,or 5 mg of amlodipine plus 2 mg of terazosin（n=119） once daily for a total of 28 days.The primary outcomes were a reduction in the total and subscores of the International Prostate Symptom Score and blood pressure.Analyses were performed by intention to treat.This trial is registered with ClinicalTrials.gov（No.NCT00693199）.[Results]:At day 28 of the trial,the amlodipine plus terazosin group achieved the greatest blood pressure control compared with either the terazosin group（P＜0.01） or amlodipine group（P＜0.05）.The amlodipine plus terazosin group also demonstrated comparable efficacy in lowering the total International Prostate Symptom Score and significant improvement in the presence of overactive bladder compared with the terazosin group（P＜0.05） and significant improvement in quality of life compared with the amlodipine group（P＜0.05）.All 3 treatment regimens were well tolerated by the study patients.[Conclusions]:The results of this 4-week,double-blind,randomized trial have demonstrated that in Chinese male hypertensive patients with LUTS,low-dose amlodipine plus terazosin therapy appears to be a safe and effective combination therapy to control both conditions,especially for those with predominant overactive bladder symptoms.更多还原