“调肝肾、祛痰瘀”治法方药逆转SHR血管重构的作用机理研究
|
摘要
第一部分理论探讨
一、血管重构与原发性高血压
血管对血压始终都发挥着自身的重要调节作用。它能产生许多活性物质，调节血管的结构和功能。因此，血管本身障碍及如何纠正这一障碍是高血压发病机制及其防治中的一个重要问题，而其关键又是纠正血管平滑肌细胞(Vascular Smooth Muscle Cell，VSMC)结构和功能的改变。
与高血压伴随的血管结构和功能的变化，即是血管重构(Vascular Remodeling，VR)主要包括：①血管壁增厚，尤其是中层肥厚，导致壁与腔之间的比例明显加大，血流阻力和血管对缩血管物质的反应性增强；②阻力血管变得稀少，尤其是直径12-25μm的阻力血管数目明显减少，随之产生一系列血管功能异常。许多研究证实：高血压阻力血管结构变化与VSMC增生和肥大有关。VSMC结构的变化及血管壁肥厚在不同类型的高血压表现都较为相似，即通过VSMC生长和血管壁“重塑”共同作用的结果。
高血压时VSMC通过自分泌、旁分泌和细胞内分泌等多种方式分泌多种生物活性物质调节自身生长，如血小板源生长因子(Platelet-derived growth factor，PDGF)和转移生长因子—β_1(transforming growth factorβ_1，TGF-β_1)，这在高血压血管肥厚的发生发展中起重要作用。大量实验证明钙，尤其是VSMC内游离Ca~(2+)，在维持细胞正常功能和许多心血管疾病的发生、发展中具有十分重要的作用。血管平滑肌细胞内游离钙的增多，被认为是原发性高血压发生机制的最后共同途径。当细胞内Ca~(2+)浓度升高后，Ca~(2+)与其细胞内受体钙调素(Calmodin，CaM)结合生成复合物，后者通过一系列磷酸化过程引起平滑肌细胞收缩，并可促进多种癌基因的表达，如c-fos、c-myc、c-myb等，从而引起血管平滑肌肌细胞的增殖和肥厚。
二、逆转血管重构的研究意义及研究概况
血管重构与高血压、动脉粥样硬化和冠心病等多种心血管的疾病的发病有密切的关系，可加速疾病的进程。尽管目前血管重构的机制尚未完全明确，但其有关机制以及逆转心血管重构的研究已成为研究的热点。
目前大多数合成药物往往由于副作用，或是由于联合用药时的拮抗作用，或是由于长期用药所带来的经济负担，仍然处于不断研究探索当中；而中药复方所表现出的整体调节作用，避免了化学药物针对单一的病理环节，从而显示出良好的应用开发前景。本课题组长期以来一直从事中医“调肝肾，祛痰瘀”治法方药防治高血压及其并发症的研究，我们既往的研究工作表明“调肝肾，祛痰瘀”治法方药对高血压病具有良好的整体调节作用。
第二部分实验研究
一、材料和方法
(一)药物
调肝肾、祛痰瘀中药复方(简称“调方”)，其组成为：熟地25g、龟版30g(打碎、先煎)、丹参15g、三七5g、钩藤15g(后下)、全瓜萎15g、怀牛膝15g。所用中药均由本校第一附属医院中药房提供，经药剂科鉴定为纯正药材。常规煎煮取汁后浓缩至含生药1.06g／ml。药液常温冷却后，置4℃冰箱内保存备用。
(二)动物分组及给药
1、整体疗效学研究
亲代自发性高血压大鼠(Spontaneously Hypertensive Rat，SHR)由北京维通利华实验动物有限公司提供，合格证号为0048303，12周龄，清洁级，其中雄性17只，体重250±25g，雌性34只，体重160±25g。SHR按体重及雄、雌1：2的比例随机分为4组。在测定尾动脉收缩压后同笼配对繁育，每笼1雄2雌，每组4笼。
从6周起观察血压测量值；至12周取材，观察血管中膜厚度及与管腔比值、细胞的超微结构。
2、离体培养VSMC重构机制探讨
SHR VSMC原代培养采用贴块法，常规方法制备调方含药血清及正常对照血清，噻唑蓝(MTT)染色法筛选适合VSMC生长的调方含药血清浓度，从钙系统、PDGF—A、TGF—β_1角度探讨调方逆转血管重构的机制。
二、结果
(一)调肝肾、祛痰瘀复方发病前不同时段给药逆转SHR血管重构的整体疗效学影响
本文采用SHR模型研究高血压血管重构的变化，揭示高血压和血管重构在发病前不同时段的关系，由实验的结果可以看出调肝肾、祛痰瘀治法及其组方于发病前和(或)发病早期实施干预，能有效地延缓SHR早期的血压上升，在6周龄后显示出明显的控制血压上升的效果；第6周时S1—4和S1—10组的血压低于另两组，提示“酿乳”这一间接给药方式的效应，其药代动力学有待进一步研究。通过细胞形态计量分析发现，S1—10组对于改善VSM中膜层厚度和内膜损伤与S空白组比较有非常显著性差异，S1—4对于改善VSM中膜层厚度与S空白组比较有显著性差异，而S5—10与S空白组比较无显著性差异。而且，VSMC有超微结果损害，突出表现为胞质内线粒体密度不均匀，形态肿胀，嵴结构紊乱或消失，线粒体是细胞内的重要钙离子库，这种改变提示细胞内钙代谢失调可能是高血压的细胞损害的重要机制。提示调方具有良好的逆转SHR血管重构的作用，早期干预(1—4周)对于血管平滑肌中膜的形成、生长可能有决定性意义。
(二)调肝肾、祛痰瘀复方含药血清对VSMC内钙系统的影响
本实验观察到，各组大鼠血管平滑肌钙离子水平、钙调素水平的变化成时段依赖性，第10周和第12周取材的VSMC内游离[Ca~(2+)]i较第1周和第5周取材组明显增高，可能与血管活性物质和生长因子可能促进细胞膜钙离子通道的开放增加了细胞对钙的摄取有关；第1周和第5周取材组[Ca~(2+)]i与S12比较有非常显著性差异；使用含药血清后，各中药血清组[Ca~(2+)]i均较正常血清组低，虽无统计学意义，但提示调方含药血清有降低胞内游离钙浓度的趋势，使VSMC增殖肥大作用明显减弱，与实验三的结果相结合来看，可能与具有拮抗生长因子促进细胞膜钙离子通道主动转运钙的作用有关。各组VSMC Ca~(2+)—ATP活性变化成时段依赖性，即S1＞S5＞S10＞S12，S1、S5、10中药血清组VSMC Ca~(2+)—ATP活性与S12比较有非常显著性差异，提示Ca~(2+)—ATP活性随高血压的发生、发展有密切关系；使用含药血清后，各中药血清组VSMC Ca~(2+)—ATP活性均较正常血清组高，提示调方含药血清可上调VSMC Ca~(2+)—ATP活性，其机制可能与降低胞内高Ca~(2+)浓度可以进一步上调Ca~(2+)活性有关。
(三)调肝肾、祛痰瘀复方对VSMC内PDGF—A、TGF—β_1的影响
本研究发现：PDGF—A链和TGF—β_1表达在S12，S10_空和S5，S1具有显著差异，S12＞S10_空＞S10_药＞S5_空＞S5_药＞S1，在S1各组中PDGF-A和TGF—β_1链几乎无表达，S5各组中仅有微量表达，而在S10两组中比较，含药血清组的表达又少于空白组，S10_空与S12的表达量差异不明显。这一结果提示：①这种自身PDGF—A链和TGF—β_1的高表达，可能在自发性高血压大鼠血管平滑肌细胞增殖肥大、血管功能异常中具有重要作用。②含药血清不同时段给药对PDGF—A链和TGF—β_1表达的抑制作用不同，提示早期干预有重要意义。
第三部分研究结论
一、研究显示，调肝肾、祛痰瘀治法及其方药在高血压发病前不同时段给药，有确切的延缓血压上升的作用，而且作用效果平稳、持久、缓和，以全程给药组作用最佳，从而部分揭示了这一临床组方思路的可行性与合理性。
二、调方能改善VSM中膜厚度和VSMC超微结构，提示调方能逆转高血压血管重构现象，具有一定的血管保护作用，且早期干预(1—4周)对于血管平滑肌中膜的形成、生长可能有决定性意义。
三、调方含药血清有降低Ca~(2+)浓度、CaM活性的趋势，提高钙泵的活性，并能上调钙泵基因的表达，抑制CaM基因的表达，提示其降压作用机制和逆转血管重构的机制可能是通过钙系统起作用。
四、调方含药血清能抑制PDGF-A、TGFβ1蛋白表达，提示其对VSMC的增殖有抑制作用。
Part One Theory Studies
1. Vascular remodeling (VR) and Essential hypertension (EH)
Blood vessal exet important regulate function on blood pressure. It canproduce many vasoactive substance and regulate the structure andfunction. So, it's per se handicap and how to rectify it is an important problemon mechanism and prevention and cure of EH, and rectify the changes on vascularsmooth muscle cell's structure and function is the key of the problem.
The changes on vessal's structure and function with EH is called vascularremodeling, including:①vas wall incrassation, especially on middle-level,lead the proportion between wall and cavity increased, the resistance of bloodstream and the reaction to the shrink substance is tone up;②resistance vasis decreased, especially on dia 12-25μm's vas, and bring a series of functionabnormity with it. Many researches are proved: the changes on vas structurehave the relationship with the hyperplasia and hypertrophy of VSMC. VSMC'sstruction changes and vas wall incrassation have the resembled represent indifferent type hypertension, namely, the result of VSMC growth and vasremodeling.
VSMC rely on multi-mode excreting biologic active substance to regulateself growth, such as self- excreting. Platelet-derived growth factor (PDGF)and transforming growth factorβ_1(TGF-β_1) have important effect on theevolution of vas plump. Many experiments have proved calcium, especially themanifold dissociate Ca~(2+) in VSMC, is the ultimately together way of themechanism on EH. When cellular Ca~(2+) concentration increased, Ca~(2+)与can combinewith calmodin (CaM) create compound, the latter arose VSMC shrinking through phosphated, and can accelerate express of many cancer gene, such as c-fos、c-myc、c-myb, consequently bring the multiplication and plumped of VSMC.
2. The research meaning and overview of reverse vascular remodeling
Vascular remodeling has the consanguineous relation with manyangiocardiopathy, and it can speed the disease process. Although the mechanismof vascular remodeling is still unknown, but the related study and reversevascular remodeling have become the hot spot in the science of the currentresearch.
Now most of the biosynthesis preparation can only aim directly at onepath-link The formula of traditional Chinese medicine have the holo-regulationfunction, so its manifests favorable prospect of implication andexploitation. Our research team have been studying by regulating the functionof liver and kidney and removing sputum and blood stasis therapy and formulato prevent and cure the EH and complication, we found that RR-therapy andformula have the favorable holo-regulation function on EH.
Part Two Experimental Studies
1. Materials and Methods
1.1 Herbs
Composition and dosage of RR-formula: raidx rehmanniae preparata 25g、carapax et plastrum testudins 30g (break into pieces、fore-decoct)、radixsalviae miltiorrhizae 15g、radix notoginseng 5g、ramulus uncariae cum uncis15g (post-decoct)、fructus trichosanthis 15g、radix achyranthis bidentatae15g. The herbs are bought from Guangzhou University of Traditional ChineseMedicine affiliated first hospital TCM pharmacy and appraised as pure crudeherbs by the drug department, The concentration of crude herbs is 1.06g/mlafter being fried and concentrated and preserved in refrigerator at 4centi-degree after being cooled under the common temperature.
1.2 The subgroup and administration of rats
1.2.1 The study of whole curative effect
Spontaneously Hypertensive Rat, offered by Beijing Vitalriver laboratoryanimals limited liability company (certification number:0048303), 12week,grade SPF, male aggregately 17, 250±25g in weight, female aggregately 34,160±25g in weight。According to weight and gender divided into four groups:filial generation 1-4 week administration, filial generation 5-10 week administration, filial generation 1-10 week administration, filial generationmodel group: without any administration.
From 6 week, observe blood pressure; till 12weeks, kill all rats, observethickness of vas mid-level and the ratio to cavity、cell's ultrastructure.
1.2.2 The mechanism discuss by part from body cultivate VSMC
SHR primary VSMCwere cultured by tissue culture method, general methodpreparation RR-formuta pastille serum and normal comparison serum, MTT dyeingfiltrate serum concentration that fit the growth of VSMC. Discuss the mechanismof reverse vascular remodeling from calcium system、PDGF—A and TGF—β_1.
2. Results
2.1 The influence of RR-formula that administration at different period oftime before invasion reverse vascular remodeling
SHR were chosen for our research. From the result we can find: the earlytreatment with RR-therapy and formula depressed the development ofhypertension, show in evidence effect after 6 week; S1-4 and S1-10 group'spressure lower than the other groups at 6week, clue to the effect of "brewmilk", but the drug metabolism dynamics need more research. By cell's shapemeasure, we find that herbs can lessened VSM medial thickness of aortae andreduce intima's injury, S1-10 group is best, S1-4 group take second place.Besides, herbs can ameliorate abnormal ultrastructural signs of VSMC from SHRincluding heterogeneous population of nuclei, swelling of mitochondria and tubular systems as well as decreased Ca~(2+). The result clue on that the earlytreatment has finality meaning to the form and growth on VSMC.
2.2 The influence of RR-formula pastille serum to calium system on VSMC
We found that the changes of Ca~(2+) and CaM level are increased with timein each group. In S10 and S12 group, [Ca~(2+)]i increased evidently than S1 andS5 group, the result maybe related with growth factor. After use pastille serum,[Ca~(2+)]i were all lower than normal comparison serum, the result hint thatpastille serum can lower the [Ca~(2+)]i and weaken VSMC multiplication andhypertrophy. Combine with experiment 4, maybe related with rivalry Ca~(2+)initiative transfer. The changes of Ca~(2+)-ATP activity decreased with time ineach group. Pastille serum in S1、S5 and S10 groups compared with S12 group,Ca~(2+)-ATP activity were higher than S12 group significantly. The resultsuggested that Ca~(2+)-ATP activity have affinity with EH's occurrence anddevelopment, it's mechanism maybe related with a fall in the concentrationof Ca~(2+).
2.3 The influence of RR-formula pastille serum to PDGF-A and TGF-β_1, on VSMC
We found that the express of PDGF-A and TGF-β_1 were higher in S12 and S10grouop than S5 and S1 group significantly. The result clue on:①The highexpression maybe important in VSMC's multiplication and vas disfunction inSHR;②Pastille serum on different period of time have differentrestrainability, that hint administration in forepart is important.
Part Three Conclusions
1 The research show that the treatment with RR-therapy and formula on differentperiod of time before invasion can depressed the development of hypertensionin some degrees, and the effect of S1-10group is the best. It partially openedout the feasibility and rationality for clinic thought of formula andadministration.
2 RR-formula can lessened VSM medial thickness and ameliorate abnormalultrastructural signs of VSMC from SHR. The result clue on that RR-formulacan reverse vascular remodeling in some degrees and the early treatment hasfinality meaning to the form and growth on VSMC.
3 RR-formula pastille serum can decrease [Ca~(2+)]i and CaM level, increase theactivity of Ca~(2+)-ATP, at the same time, it can up-regulate the expression ofCa~(2+)-ATP gene and restrain CaM gene. It clue on that the mechanism maybe related with calcium system.
4 RR-formula pastille serum can repression the expression of PDGF-A、TGFβ_1protein, it can repression multiplication of VSMC.
一、血管重构与原发性高血压
血管对血压始终都发挥着自身的重要调节作用。它能产生许多活性物质，调节血管的结构和功能。因此，血管本身障碍及如何纠正这一障碍是高血压发病机制及其防治中的一个重要问题，而其关键又是纠正血管平滑肌细胞(Vascular Smooth Muscle Cell，VSMC)结构和功能的改变。
与高血压伴随的血管结构和功能的变化，即是血管重构(Vascular Remodeling，VR)主要包括：①血管壁增厚，尤其是中层肥厚，导致壁与腔之间的比例明显加大，血流阻力和血管对缩血管物质的反应性增强；②阻力血管变得稀少，尤其是直径12-25μm的阻力血管数目明显减少，随之产生一系列血管功能异常。许多研究证实：高血压阻力血管结构变化与VSMC增生和肥大有关。VSMC结构的变化及血管壁肥厚在不同类型的高血压表现都较为相似，即通过VSMC生长和血管壁“重塑”共同作用的结果。
高血压时VSMC通过自分泌、旁分泌和细胞内分泌等多种方式分泌多种生物活性物质调节自身生长，如血小板源生长因子(Platelet-derived growth factor，PDGF)和转移生长因子—β_1(transforming growth factorβ_1，TGF-β_1)，这在高血压血管肥厚的发生发展中起重要作用。大量实验证明钙，尤其是VSMC内游离Ca~(2+)，在维持细胞正常功能和许多心血管疾病的发生、发展中具有十分重要的作用。血管平滑肌细胞内游离钙的增多，被认为是原发性高血压发生机制的最后共同途径。当细胞内Ca~(2+)浓度升高后，Ca~(2+)与其细胞内受体钙调素(Calmodin，CaM)结合生成复合物，后者通过一系列磷酸化过程引起平滑肌细胞收缩，并可促进多种癌基因的表达，如c-fos、c-myc、c-myb等，从而引起血管平滑肌肌细胞的增殖和肥厚。
二、逆转血管重构的研究意义及研究概况
血管重构与高血压、动脉粥样硬化和冠心病等多种心血管的疾病的发病有密切的关系，可加速疾病的进程。尽管目前血管重构的机制尚未完全明确，但其有关机制以及逆转心血管重构的研究已成为研究的热点。
目前大多数合成药物往往由于副作用，或是由于联合用药时的拮抗作用，或是由于长期用药所带来的经济负担，仍然处于不断研究探索当中；而中药复方所表现出的整体调节作用，避免了化学药物针对单一的病理环节，从而显示出良好的应用开发前景。本课题组长期以来一直从事中医“调肝肾，祛痰瘀”治法方药防治高血压及其并发症的研究，我们既往的研究工作表明“调肝肾，祛痰瘀”治法方药对高血压病具有良好的整体调节作用。
第二部分实验研究
一、材料和方法
(一)药物
调肝肾、祛痰瘀中药复方(简称“调方”)，其组成为：熟地25g、龟版30g(打碎、先煎)、丹参15g、三七5g、钩藤15g(后下)、全瓜萎15g、怀牛膝15g。所用中药均由本校第一附属医院中药房提供，经药剂科鉴定为纯正药材。常规煎煮取汁后浓缩至含生药1.06g／ml。药液常温冷却后，置4℃冰箱内保存备用。
(二)动物分组及给药
1、整体疗效学研究
亲代自发性高血压大鼠(Spontaneously Hypertensive Rat，SHR)由北京维通利华实验动物有限公司提供，合格证号为0048303，12周龄，清洁级，其中雄性17只，体重250±25g，雌性34只，体重160±25g。SHR按体重及雄、雌1：2的比例随机分为4组。在测定尾动脉收缩压后同笼配对繁育，每笼1雄2雌，每组4笼。
从6周起观察血压测量值；至12周取材，观察血管中膜厚度及与管腔比值、细胞的超微结构。
2、离体培养VSMC重构机制探讨
SHR VSMC原代培养采用贴块法，常规方法制备调方含药血清及正常对照血清，噻唑蓝(MTT)染色法筛选适合VSMC生长的调方含药血清浓度，从钙系统、PDGF—A、TGF—β_1角度探讨调方逆转血管重构的机制。
二、结果
(一)调肝肾、祛痰瘀复方发病前不同时段给药逆转SHR血管重构的整体疗效学影响
本文采用SHR模型研究高血压血管重构的变化，揭示高血压和血管重构在发病前不同时段的关系，由实验的结果可以看出调肝肾、祛痰瘀治法及其组方于发病前和(或)发病早期实施干预，能有效地延缓SHR早期的血压上升，在6周龄后显示出明显的控制血压上升的效果；第6周时S1—4和S1—10组的血压低于另两组，提示“酿乳”这一间接给药方式的效应，其药代动力学有待进一步研究。通过细胞形态计量分析发现，S1—10组对于改善VSM中膜层厚度和内膜损伤与S空白组比较有非常显著性差异，S1—4对于改善VSM中膜层厚度与S空白组比较有显著性差异，而S5—10与S空白组比较无显著性差异。而且，VSMC有超微结果损害，突出表现为胞质内线粒体密度不均匀，形态肿胀，嵴结构紊乱或消失，线粒体是细胞内的重要钙离子库，这种改变提示细胞内钙代谢失调可能是高血压的细胞损害的重要机制。提示调方具有良好的逆转SHR血管重构的作用，早期干预(1—4周)对于血管平滑肌中膜的形成、生长可能有决定性意义。
(二)调肝肾、祛痰瘀复方含药血清对VSMC内钙系统的影响
本实验观察到，各组大鼠血管平滑肌钙离子水平、钙调素水平的变化成时段依赖性，第10周和第12周取材的VSMC内游离[Ca~(2+)]i较第1周和第5周取材组明显增高，可能与血管活性物质和生长因子可能促进细胞膜钙离子通道的开放增加了细胞对钙的摄取有关；第1周和第5周取材组[Ca~(2+)]i与S12比较有非常显著性差异；使用含药血清后，各中药血清组[Ca~(2+)]i均较正常血清组低，虽无统计学意义，但提示调方含药血清有降低胞内游离钙浓度的趋势，使VSMC增殖肥大作用明显减弱，与实验三的结果相结合来看，可能与具有拮抗生长因子促进细胞膜钙离子通道主动转运钙的作用有关。各组VSMC Ca~(2+)—ATP活性变化成时段依赖性，即S1＞S5＞S10＞S12，S1、S5、10中药血清组VSMC Ca~(2+)—ATP活性与S12比较有非常显著性差异，提示Ca~(2+)—ATP活性随高血压的发生、发展有密切关系；使用含药血清后，各中药血清组VSMC Ca~(2+)—ATP活性均较正常血清组高，提示调方含药血清可上调VSMC Ca~(2+)—ATP活性，其机制可能与降低胞内高Ca~(2+)浓度可以进一步上调Ca~(2+)活性有关。
(三)调肝肾、祛痰瘀复方对VSMC内PDGF—A、TGF—β_1的影响
本研究发现：PDGF—A链和TGF—β_1表达在S12，S10_空和S5，S1具有显著差异，S12＞S10_空＞S10_药＞S5_空＞S5_药＞S1，在S1各组中PDGF-A和TGF—β_1链几乎无表达，S5各组中仅有微量表达，而在S10两组中比较，含药血清组的表达又少于空白组，S10_空与S12的表达量差异不明显。这一结果提示：①这种自身PDGF—A链和TGF—β_1的高表达，可能在自发性高血压大鼠血管平滑肌细胞增殖肥大、血管功能异常中具有重要作用。②含药血清不同时段给药对PDGF—A链和TGF—β_1表达的抑制作用不同，提示早期干预有重要意义。
第三部分研究结论
一、研究显示，调肝肾、祛痰瘀治法及其方药在高血压发病前不同时段给药，有确切的延缓血压上升的作用，而且作用效果平稳、持久、缓和，以全程给药组作用最佳，从而部分揭示了这一临床组方思路的可行性与合理性。
二、调方能改善VSM中膜厚度和VSMC超微结构，提示调方能逆转高血压血管重构现象，具有一定的血管保护作用，且早期干预(1—4周)对于血管平滑肌中膜的形成、生长可能有决定性意义。
三、调方含药血清有降低Ca~(2+)浓度、CaM活性的趋势，提高钙泵的活性，并能上调钙泵基因的表达，抑制CaM基因的表达，提示其降压作用机制和逆转血管重构的机制可能是通过钙系统起作用。
四、调方含药血清能抑制PDGF-A、TGFβ1蛋白表达，提示其对VSMC的增殖有抑制作用。
Part One Theory Studies
1. Vascular remodeling (VR) and Essential hypertension (EH)
Blood vessal exet important regulate function on blood pressure. It canproduce many vasoactive substance and regulate the structure andfunction. So, it's per se handicap and how to rectify it is an important problemon mechanism and prevention and cure of EH, and rectify the changes on vascularsmooth muscle cell's structure and function is the key of the problem.
The changes on vessal's structure and function with EH is called vascularremodeling, including:①vas wall incrassation, especially on middle-level,lead the proportion between wall and cavity increased, the resistance of bloodstream and the reaction to the shrink substance is tone up;②resistance vasis decreased, especially on dia 12-25μm's vas, and bring a series of functionabnormity with it. Many researches are proved: the changes on vas structurehave the relationship with the hyperplasia and hypertrophy of VSMC. VSMC'sstruction changes and vas wall incrassation have the resembled represent indifferent type hypertension, namely, the result of VSMC growth and vasremodeling.
VSMC rely on multi-mode excreting biologic active substance to regulateself growth, such as self- excreting. Platelet-derived growth factor (PDGF)and transforming growth factorβ_1(TGF-β_1) have important effect on theevolution of vas plump. Many experiments have proved calcium, especially themanifold dissociate Ca~(2+) in VSMC, is the ultimately together way of themechanism on EH. When cellular Ca~(2+) concentration increased, Ca~(2+)与can combinewith calmodin (CaM) create compound, the latter arose VSMC shrinking through phosphated, and can accelerate express of many cancer gene, such as c-fos、c-myc、c-myb, consequently bring the multiplication and plumped of VSMC.
2. The research meaning and overview of reverse vascular remodeling
Vascular remodeling has the consanguineous relation with manyangiocardiopathy, and it can speed the disease process. Although the mechanismof vascular remodeling is still unknown, but the related study and reversevascular remodeling have become the hot spot in the science of the currentresearch.
Now most of the biosynthesis preparation can only aim directly at onepath-link The formula of traditional Chinese medicine have the holo-regulationfunction, so its manifests favorable prospect of implication andexploitation. Our research team have been studying by regulating the functionof liver and kidney and removing sputum and blood stasis therapy and formulato prevent and cure the EH and complication, we found that RR-therapy andformula have the favorable holo-regulation function on EH.
Part Two Experimental Studies
1. Materials and Methods
1.1 Herbs
Composition and dosage of RR-formula: raidx rehmanniae preparata 25g、carapax et plastrum testudins 30g (break into pieces、fore-decoct)、radixsalviae miltiorrhizae 15g、radix notoginseng 5g、ramulus uncariae cum uncis15g (post-decoct)、fructus trichosanthis 15g、radix achyranthis bidentatae15g. The herbs are bought from Guangzhou University of Traditional ChineseMedicine affiliated first hospital TCM pharmacy and appraised as pure crudeherbs by the drug department, The concentration of crude herbs is 1.06g/mlafter being fried and concentrated and preserved in refrigerator at 4centi-degree after being cooled under the common temperature.
1.2 The subgroup and administration of rats
1.2.1 The study of whole curative effect
Spontaneously Hypertensive Rat, offered by Beijing Vitalriver laboratoryanimals limited liability company (certification number:0048303), 12week,grade SPF, male aggregately 17, 250±25g in weight, female aggregately 34,160±25g in weight。According to weight and gender divided into four groups:filial generation 1-4 week administration, filial generation 5-10 week administration, filial generation 1-10 week administration, filial generationmodel group: without any administration.
From 6 week, observe blood pressure; till 12weeks, kill all rats, observethickness of vas mid-level and the ratio to cavity、cell's ultrastructure.
1.2.2 The mechanism discuss by part from body cultivate VSMC
SHR primary VSMCwere cultured by tissue culture method, general methodpreparation RR-formuta pastille serum and normal comparison serum, MTT dyeingfiltrate serum concentration that fit the growth of VSMC. Discuss the mechanismof reverse vascular remodeling from calcium system、PDGF—A and TGF—β_1.
2. Results
2.1 The influence of RR-formula that administration at different period oftime before invasion reverse vascular remodeling
SHR were chosen for our research. From the result we can find: the earlytreatment with RR-therapy and formula depressed the development ofhypertension, show in evidence effect after 6 week; S1-4 and S1-10 group'spressure lower than the other groups at 6week, clue to the effect of "brewmilk", but the drug metabolism dynamics need more research. By cell's shapemeasure, we find that herbs can lessened VSM medial thickness of aortae andreduce intima's injury, S1-10 group is best, S1-4 group take second place.Besides, herbs can ameliorate abnormal ultrastructural signs of VSMC from SHRincluding heterogeneous population of nuclei, swelling of mitochondria and tubular systems as well as decreased Ca~(2+). The result clue on that the earlytreatment has finality meaning to the form and growth on VSMC.
2.2 The influence of RR-formula pastille serum to calium system on VSMC
We found that the changes of Ca~(2+) and CaM level are increased with timein each group. In S10 and S12 group, [Ca~(2+)]i increased evidently than S1 andS5 group, the result maybe related with growth factor. After use pastille serum,[Ca~(2+)]i were all lower than normal comparison serum, the result hint thatpastille serum can lower the [Ca~(2+)]i and weaken VSMC multiplication andhypertrophy. Combine with experiment 4, maybe related with rivalry Ca~(2+)initiative transfer. The changes of Ca~(2+)-ATP activity decreased with time ineach group. Pastille serum in S1、S5 and S10 groups compared with S12 group,Ca~(2+)-ATP activity were higher than S12 group significantly. The resultsuggested that Ca~(2+)-ATP activity have affinity with EH's occurrence anddevelopment, it's mechanism maybe related with a fall in the concentrationof Ca~(2+).
2.3 The influence of RR-formula pastille serum to PDGF-A and TGF-β_1, on VSMC
We found that the express of PDGF-A and TGF-β_1 were higher in S12 and S10grouop than S5 and S1 group significantly. The result clue on:①The highexpression maybe important in VSMC's multiplication and vas disfunction inSHR;②Pastille serum on different period of time have differentrestrainability, that hint administration in forepart is important.
Part Three Conclusions
1 The research show that the treatment with RR-therapy and formula on differentperiod of time before invasion can depressed the development of hypertensionin some degrees, and the effect of S1-10group is the best. It partially openedout the feasibility and rationality for clinic thought of formula andadministration.
2 RR-formula can lessened VSM medial thickness and ameliorate abnormalultrastructural signs of VSMC from SHR. The result clue on that RR-formulacan reverse vascular remodeling in some degrees and the early treatment hasfinality meaning to the form and growth on VSMC.
3 RR-formula pastille serum can decrease [Ca~(2+)]i and CaM level, increase theactivity of Ca~(2+)-ATP, at the same time, it can up-regulate the expression ofCa~(2+)-ATP gene and restrain CaM gene. It clue on that the mechanism maybe related with calcium system.
4 RR-formula pastille serum can repression the expression of PDGF-A、TGFβ_1protein, it can repression multiplication of VSMC.
引文
1．孙宁玲，徐成斌．今日高血压，北京：中国医药科技出版社，2000，第一版：47-58
2. Nakayama M, Fukuda N, Watanabe Y, et al. Low dose of eicosapentaenoic acid inhibits the exaggerated growth of vascular smooth muscle cells from spontaneously hypertensive rats through suppression of transforming growth factor-beta. J Hypertens, 1999; 17 (10): 1421-30.
3. Sabri A, Levy BI, Poitevin P, et al. Differential roles of AT1 and AT2 receptor subtypes in vascular trophic and phenotypie changes in response to stimulation with angiotensin Ⅱ. Arterioscler Thromb Vase Biol, 1997; 17 (2): 257-64.
4. Heagerty AM, Aalkjaer C, Bund S J, et al. Small artery structure in hypertension. Dual processes of remodeling and growth. Hypertension, 1993; 21 (4): 391-7
5. Schiffrin EL, Hayoz D. How to assess vascular remodelling in small and medium-sized muscular arteries in humans. J Hypertension, 1997; 15 (6): 517-584.
6. Dickhout JG, Lee RM. Structural and functional analysis of small arteries from young spontaneously hypertensiverats. Hypertension, 1997; 29 (3): 781-9.
7. Dzau VJ, Gibbons GH, Modshita R, et al. New perspectives in hypertension research. Potentials of vascular biology. Hypertension, 1994; 23 (6 Pt 2): 1132-40.
8. Lee RM, Adams MA, Friberg P, et al. Debate: vascular remodelling. J Hypertens, 1997; 15 (4): 333-7.
9. Komer PI, Angus JA. Vascular remodeling. Hypertension, 1997; 29 (4): 1065-6.
10. Bohr DF, Dominiczak AF, Webb Re. Pathophysiology of the vasculatttre in hypertension. Hypertension, 1991, 18 (5 supple): Ⅲ69-75
11. Rizzoni D, Castellano M, Porteri E, et al. Vascular structural and functional alterations before and after the development of hypertension in SHR. Am J Hypertens, 1994; 7 (2): 193-200.
12. Saltis J, Agrotis A and Bobik A. Differences in growth characteristics of vascular smooth muscle from spontaneously hypertensive and Wistar-Kyoto rats are growth factor dependent. J Hypertens, 1993; 11 (6): 629-37
13. Bussy C, Boutouyrie P, Lacolley P, et al. Intrinsic stiffness of the carotid arterial wall material in essential hypertensives. Hypertension, 2000; 35 (5): 1049-54
14. Intengan and Schiffrin. Structure and mechanical properties of resistance arteries in hypertension: role of adhesion molecules and extracellular matrix determinants. Hypertension, 2000; 36 (3): 312-8.
15. Touyz RM. Molecular and cellular mechanisms regulating vascular function and structure—implications in the pathogenesis of hypertension. Can J Cardiol, 2000; 16 (9): 1137-46.
16. Hadrava V, Kruppa U, Russo RC, et al. Vascular smooth muscle cell proliferation and its therapeutic modulation in hypertension. Am Heart J, 1991; 122 (4 Pt 2): 1198-203.
17．汪浩川，刘秉文．生长因子、癌基因与动脉平滑肌细胞增殖．生理科学进展，1994；5(1)：48-53
18. Wang DH, Prewitt RL, Beebe SJ. Regulation of PDGF-A: a possible mechanism for angiotension Ⅱ-induced vascular growth. Am J Physiol, 1995; 269(1 Pt 2): H356-364.
19. Parker SB, Wade SS, Prewitt RL. Pressure mediates angiotension Ⅱ-induced arterial hypertrophy and PDGF-A expression. Hypertension, 1998; 32 (3): 452-458.
20. Parker SB, DobrianAD, Wade SS, et al. AT (1) receptor inhibition does not reduce arterial wall hypertrophy or PDGF-A expression in renal hypertension. Am J Physiol Heart Circ Physiol, 2000; 278 (2): H613-22
21. Eghbali M, TomeK R, Sukhatme VP, et al. Differential effects of transforming growth factor beta-1 and phorbol myristate acetate on cardiac fibroblasts. Regulation of fibrillar collagen mRNAs and expression of early transcription factor. Circ Res, 1991; 69 (2): 483-490
22. Villarreal FJ, Dillmann WH. Cardiac hypertrophy-induced changes in mRNA levels for TGF-beta 1, fibronectin, andcollagen.Am J Physiol, 1992; 262 (6Pt2): H1861-6
23. Youn TJ, Kim HS, Oh BH, et al. Ventricular remodeling and transforming growth factor-beta 1 mRNA expression after nontransmural myocardial infarction in rats: effects of angiotensin converting enzyme inhibition and angiotensin Ⅱ type 1 receptor blockade. Basic Res Cardiol, 1999; 94 (4): 246-53.
24．彭龙云，高修仁，麦炜颐，等．高血压大鼠心肌TGF-β1和MMP2蛋白表达及RAS阻断后的变化．国际心血管病学杂志，2002；12(4)：303-306(英文)
25．高修仁，彭龙云，曾群英，等．ARB及ACEI联合应用对高血压大鼠心室重塑的影响．中华临床医药杂志，2003；1(4)：24-27
26. Agrotis A, Saltis J, Bobik A, et al. Transforming growth factor—TGF-β1 gene activation and growth of smooth muscle from hypertensive rats. Hypertension, 1994; 23 (5): 593-599
27. Griffin SA, Brown WC, MacPherson F, et al. Angiotensin Ⅱ causes vascular hypertrophy in part by a non-pressor mechanism. Hypertension, 1991; 17 (5): 626-35.
28. Paquet JL, Baudouin-Legros M, Brunelle G, et al. Angiotensin Ⅱ-indueed proliferation of aortic myocytes in spontaneously hypertensive rats. J Hypertens, 1990; 8 (6): 565-72
29. Postnov IuV. The role of mitochondrial calcium overload and energy deficiency in pathogenesis of arterial hypertension. Arkh Patol, 2001; 63 (3): 3-10.
30．文允镒．钙、钙调素与高血压关系的研究进展[J]．高血压杂志，1994；2(S1)：59-62
31．文允镒，张志明，胡蓓．钙与钙调素[M]．北京：化工出版社，1989，第一版：100-106，235-236
32．许顺滨，文允镒，陈孟勤．川芎嗪对正常及高血压大鼠动脉平滑肌Ca～(2+)内流的动力学影响[J]．中国应用生理学杂志，1989；1：26
33. Lu KP, Means AR. Regulation of the cell cycle by calcium and calmodulin. Endocr Rev, 1993; 14 (1): 40-58.
34. Wen YY, Jiang L, Chen M, et al. The changes of calmodulin and cAMP contents in tissues of renal hypertensive rats. Proc Chin Acad Med Sci Peking Union Med Coll, 1987; 2 (1): 22-7
35．靳琦．王琦“辨体—辨病—辨证诊疗模式”的理论要素与临床应用[J]．北京中医药大学学报2006；29(1)：41—45
36．卢焯明，潘毅，梁颖瑜，等．调肝肾、祛痰瘀治法提前干预对SHR血压及血管活性物质的影响[J]．中国中医药信息杂志，2004；11(9)：780—782
37．王琦．调治亚健康状态是中医学在21世纪对人类的新贡献[J]．北京中医药大学学报，2001：24(2)：1—4
38. Watanabe T, Barker TA, Berk BC. Angiotensin Ⅱ and the endothelium: diverse signals and effects. Hypertension, 2005; 45 (2): 163-9.
39. Chobanian AV, Bakris GL, Black HR, et al. The Seventh Report of the Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure: the JNC 7 report. JAMA, 2003; 289 (19): 2560-72.
40. Asmar RG, London GM, O'Rourke ME, et al. Improvement in blood pressure, artedal stiffness and wave reflections with a very-low-dose perindopdl/indapamide combination in hypertensive patient: a comparison with atenolol. Hypertension, 2001; 38 (4): 922-6.
41. Jorgensen B, Simonsen S, Endresen K, et al. Restenosis and clinical outcome in patients treated with amlodipine after angioplasty: results from the Coronary AngioPlasty Amlodipine REStenosis Study (CAPARES). J Am Coll Cardiol, 2000, 35 (3): 592-9.
42．付文波．他汀类药物对高血压病心血管重建作用的研究现状．心脏杂志，2006；18(5)：588
43．秦晋梅，李建军．他汀类药物的抗炎作用[J]．中国动脉硬化杂志，2004；12(3)：363-365
44．覃军，李爱民，何作云．他汀类药物对血管平滑肌细胞的直接作用及其意义[J]．中国动脉硬化杂志，2000：8(4)：373-375．
45．谢良地．HMG-CoA还原酶抑制剂的非调脂作用及机制(上)[J]．高血压杂志，2004，12(3)：265-269
46．张子新，谭庆，宋丽新，等．西洛他唑对家兔血管成形术后内膜增殖和血管重构的影响．中国医科大学学报，2005，34(5)：420-421
47．汪新良，何兆初，曾昭华，等．安体舒通对感觉神经损伤性盐敏感性高血压大鼠血管重构及RAAS的影响．中国心血管病研究杂志，2006，4(10)：773-776
48. Dumitrascu R, Weissmann N, Ghofrani HA, et al. Activation of soluble guanylate cyclase reverses experimental pulmonary hypertension and vascular remodeling. Circulation, 2006; 113 (2): 286-95.
49. Pascarella L, Schonbein GW, Bergan JJ. Microcirculation and venous ulcers: a review. Ann Vase Surg, 2005; 19 (6): 921-7.
50．曹守沛，钱春发，金妙文．中西医结合治疗急性心肌梗死疗效分析[J]．实用中医内科杂志，2005；19(4)：345-346
51．史载祥．后再灌注时代难题的中西医结合治疗思考[J]．中西医结合心脑血管病杂志，2005：3(1)：2-5
52．王振涛，王硕仁，赵明镜，等．活血、益气方药对心梗后左心衰大鼠血TNF和血小板聚集影响的对比研究[J]．中国中医药信息杂志，2002；9(11)：20-22
53. Ji XY, Tan BK, Zhu YZ. Salvia miltiorrhiza and ischemie diseases. Acta Pharmacol Sin, 2000; 21 (12): 1089-94
54. Sun J, Huang SH, Tan BK, et al. Effects of purified herbal extract of Salvia miltiorrhiza on ischemic rat myocardium after acute myocardial infarction. Life Sci, 2005; 76 (24): 2849-60.
55. Wang B, Zhang JD, Feng J-B, et al. Effect of traditional Chinese medicine Qin-Dan-Jiang-Ya-Tang on remodeled vascular phenotype and osteopontin in spontaneous hypertensive rats. J Ethnopharmacol, 2007:110 (1): 176-82
56．严灿，邓中炎，潘毅，等．活血祛痰法逆转高血压左室肥厚的实验研究．中国中医基础杂志，1999；5(4)：25-27
57．李建平，严灿，潘毅，等．活血祛痰方药预防自发性高血压大鼠心肌纤维化实验研究．中医杂志，2000；41(5)：301-302
58．李剑，潘毅，严灿，等．滋补肝肾、活血祛痰方药对原发性高血压大鼠左心室肥厚的影响．山西中医，2003；19(2)：54-56
59．卢焯明，潘毅，梁颖瑜，等．调肝肾、祛痰瘀治法提前干预对SHR血压及血管活性物质的影响[J]．中国中医药信息杂志，2004;;11(9)：780-782
1．王培林．遗传病学，北京：人民卫生出版社，2000，第1版：315，318
2．方厚华．医学实验模型动物，北京：军事医学科学出版社，2002，第1版：28-30
3．谢雍．“母体疗法”治疗婴幼儿泄泻．四川中医，1990；8(2)：19-20
4．王纪民．哺乳幼儿病宜母子同治．四川中医，1989；7(7)：6-7
5．渡边贤治．汉方经母乳治疗婴儿皮疹有效7例及汉方药的乳汁移行．日本东洋医学杂志，1996；46(6)：135-137
6．刘湘华．肝阳上亢证患者个性、情绪特征及血浆精氨酸加压素水平的初步研究．湖南医科大学学报，1998：23(1)：31
7．叶任高．内科学，北京：人民卫生出版社，2002，第5版：258-261
8．孙宁玲，徐成斌．今日高血压，北京：中国医药科技出版社，2000，第1版：125-131，152-154，160，166-172，149，431，596-599
9．邓中炎，徐志伟，陈群，等．中医基础理论体系现代研究，广州：广东科技出版社，2002，第一版：7，88，193
10．王本祥．现代中药药理与临床，天津，天津科技翻译出版公司，2004，第1版：
11．侯家玉．中药药理学，中国中医药出版社，2002，第1版：
12．李军，张丽萍，张振凌，等．地黄研究进展．河南中医学院学报，2005；121 (20)：79-83
13．曾艳，贾正平，张汝学，等．地黄化学成分及药理研究进展．中成药，2006；28(4)：609-611
14．管家齐，郭艳丽，吴海良，等．地黄药理研究近况．中药材，2006；24(5)：380-382
15．孙苏亚，王锦，刘铮，等．龟板提取液对去势大鼠骨质疏松的作用．中药药理与临床，1998：14(4)：20
16．黎晖，周健洪，陈东风，等．龟板对大鼠骨髓间充质干细胞向成骨分化的影响．中药新药与临床药理，2005；16(3)：159-161
17．朱莉芬，李美珠，钟佳新，等．龟鹿补肾口服液的药理研究．中成药，1993；15(2)；24
18．李以菊，孙泽玲．丹参的临床应用进展．实用医技杂志，2006，8；13(16)：2903-2905
19．付辛芳，刘晓红．丹参的药理作用与临床应用研究进展．中国药业，2006；15(1)：76-77
20．李艳杰，孙玉华．丹参素药理作用研究进展．延边大学医学学报，2006；29(1)：73-75
21．书海，李青旺，郑雪莉，等．牛膝正丁醇萃取物对小鼠繁殖性能的影响．西北农林科技大学学报，2006；34(6)：31-34
22．曹明俐，张小军，王振生，等．牛膝的药理作用研究进展．中医药信息，2002；18(3)：20-21
23．谭德讲，邱海强，袁惠南．田七洋参冲剂的抗应激作用和免疫促进作用．2000，7：11(4)：242-243
24．屠婕红，余菁，陈伟光．瓜蒌的化学成分和药理作用研究概况．中国药师，2004；7(7)：562-563
25．孙兴龙，段崇英．瓜蒌的药理作用及临床应用．中国药师，2003；6(10)：655-656
26．刘佳，富志军．钩藤的研究概况．海峡药学，2006；18(5)：90-93
27．莫志贤，许丹丹．钩藤临床运用的新进展．时珍国医国药，2006；17(5)：684-685
28．何小琦．滋肾养肝活血化瘀法治疗糖尿病并高血压60例临床观察．实用医学杂 志，1998：14(1)：69
1．蔡文琴，现代实用细胞与分子生物学实验技术．北京：人民军医出版社，2003，第一版：13—15
2. Carson DA, Ribeiro JM. Apoptosis and disease. Lancet, 1993; 341 (8855): 1251-4.
3．刘树民，李玉洁，罗明媚．中药毒性研究及思路探讨[J]．世界科学技术—中医药现代化，2003；5(2)：21-23
4．高月，吕秋军，刘永学，等．中药复方物质基础的研究[J]．中国新药杂志，2000；9(5)：307-308
5．马骏，唐灿，李增强，等．中药血清药理学的方法学研究——在中药复方药物动力学研究中的应用[J]．中药药理与临床，1999：15(3)：44-46
6．杨彦芳，王玉芹．中药复方血清药理学方法规范化探讨[J]．中国中西医结合杂志，2000；20(5)：380-382
7．周明眉，杨奎，姜远平，等．中药血清药理学的方法学研究——采血时间的确定及时效关系研究[J]．中药药理与临床，1999；15(1)：43-44
8．周明眉，杨奎，姜远平，等．中药血清药理学的方法学研究——含药血清低温保存和血清灭活的影响[J]．中药药理与临床，1999；15(2)：44-46
9．李仪奎，吴健宇．中药血清药理实验方法给药的通法方案．中国药理学通报，1999；15(6)：569-570
1．丁文勇，赵宝昌，韩旭．钙调素的环核苷酸磷酸二酯酶检测法．大连医科大学学报，1999：21(1)：3-6
2. Dolphin AC. Mechanisms of modulation of voltage-dependent calcium channels by G proteins. J Physiol, 1998; 506 (Pt 1): 3-11.
3. Marks AR. Intracellular calcium-release channels: regulators of cell life and death. Am J Physiol, 1997; 272 (2 Pt 2): H597-605.
4. Tostes RC, Storm DS, Chi DH, et al. Intracellular calcium stores and oscillatory contractions in arteries from genetically hypertensive rats. Hypertens Res, 1996; 19 (2): 103-11.
5. Morimoto S, Takeda K, Oguni A, et al. Reduction of white coat effect by cilnidipine in essential hypertension. Am J Hypertens, 2001; 14 (10): 1053-7.
6. Onose Y, Oki T, Yamada H, et al. Effect of cilnidipine on left ventricular diastolic function in hypertensive patients as assessed by pulsed Doppler echocardiography and pulsed tissue Doppler imaging. Jpn Circ J, 2001; 65 (4): 305-9.
7. Resnick LM. Physiologic rationale for calcium antagonist therapy in essential hypertension. Ethn Dis, 1998; 8 (1): 111-9.
8. Vincent M, Samani NJ, Gauguier D, et al. A pharmacogenetic approach to blood pressure in Lyon hypertensive rats. A chromosome 2 locus influences the response to a calcium antagonist. J Clin Invest, 1997; 100 (8): 2000-6.
9. Kosch M, Hausberg M, Barenbrock M, et al. Increased membraneous calcium concentrations in primary hypertension: a causal link to pathogenesis? J Hum Hypertens, 2001; 15 (1): 37-40.
10. Cicco G, Carbonara MC, Stingi GD, et al. Cytosolic calcium and hemorheological patterns during arterial hypertension. Clin Hemorheol Microcirc, 2001; 24 (1): 25-31
11. Sugden PH. Signaling pathways activated by vasoactive peptides in the cardiac myocyte and their role in myocardial pathologies. J Card Fail, 2002; 8 (6 Suppl): S359-69.
12. Kosch M, Hausberg M, Westermann G, et al. Alterations in calcium and magnesium content of red cell membranes in patients with primary hypertension. Am J Hypertens, 2001; 14 (3): 254-8.
13. Ohkusa T, Hisamatsu Y, Yano M, et at. Altered cardiac mechanism and sarcoplasmic reticulum function in pressure overload-induced cardiac hypertrophy in rats. J Mol Cell Cardiol, 1997; 29 (1): 45-54.
14. Jiang M, Xu A, Tokmakejian S, et al. Thyroid hormone-induced overexpression of functional ryanodine receptors in the rabbit heart. Am J Physiol Heart Circ Physiol, 2000; 278 (5): H1429-38.
15. Rannou F, Dambrin G, Marry I, et al. Expression of the cardiac ryanodine receptor in the compensated phase of hypertrophy in rat heart. Cardiovasc Res, 1996; 32 (2): 258-65.
16. Assayag P, Charlemagne D, de Leiris J, et al. Senescent heart compared with pressure overload-induced hypertrophy. Hypertension, 1997; 29 (1 Pt 1): 15-21.
17. Arai M, Alpert NR, MacLennan DH, et al. Alterations in sarcoplasmic reticulum gene expression in human heart failure. A possible mechanism for alterations in systolic and diastolic properties of the failing myocardium. Circ Res, 1993; 72 (2): 463-9.
18. Anger M, Lompre AM, Vallot O, et al. Cellular distribution of Ca2+ pumps and Ca2+ release channels in rat cardiac hypertrophy induced by aortic stenosis, Circulation, 1998; 98 (22): 2477-2486
19. Rivet-Bastide M, Vandecasteele G, Hatem S, et al. cGMP-stimulated cyclic nucleotide phosphodiesterase regulates the basal calcium current in human atrial myocytes. J Clin Invest, 1997, 99: 2710-2718.
20. Boixel C, Tessier S, Pansard Y, et al. Tyrosine kinase and protein kinase C regulate L-type Ca(2+) current cooperatively in human atrial myocytes. Am J Physiol Heart CircPhysiol, 2000; 278 (2): H670-6.
21. Mogami H, Kojima I. Stimulation of calcium entry is prerequisite for DNA synthesis induced by platelet-derived growth factor in vascular smooth muscle cells. Biochem Biophys Res Commun, 1993; 196 (2): 650-8.
22. Short AD, Bian J, Ghosh TK, et al. Intracellular Ca2+ pool content is linked to control of cell growth. Proc Natl Acad Sci U S A, 1993; 90 (11): 4986-90.
23. Dolmetsch RE, Lewis RS, Goodnow CC, et al. Differential activation of transcription factors induced by Ca2+ response amplitude and duration. Nature, 1997; 386 (6627): 855-8
24. Bers DM, Perez-Reyes E. Ca channels in cardiac myocytes: structure and function in Ca influx and intracellular Ca release. Cardiovasc Res, 1999; 42 (2): 339-60.
25．韩济生，神经科学原理，北京：北京医科大学出版社，1993年，第一版：222-227
26．杨钢，内分泌生理与病理生理学，天津：天津科学技术出版社，1996年，第一版：59-67
27．张均田，现代药理实验方法，北京：北京医科大学中国协和医科大学联合出版社，1998年，第一版：472-473
28. Chafouless J G, Dedmn J R, Means A R. Redioimmunoassay of cal modulin. Methods Enzymol, 1983; 84: 138
29. Strobl J S, Peterson U A, Woodfork K A. A survey of human breast cancer sensitivity to growth inhidition by calmodulin antagonists in tissue culture. Biochem Pharmacol, 1994; 47: 2157
30. Brooks-Frederich K M, Cianciarulo F L, Rittling S R, et al. Cell cycledependent regulation of Ca′'in young and senescent WI-38 cells. Exp Cell Res, 1993; 205:412
31. Wallace R W, Tallant E A, Cheung W Y. Assay of calmidulin by Ca′ '—dependent phisphodiesterase. Methods Enzymol, 1983: 102:39
32．孙宁玲，徐成斌．今日高血压，北京：中国医药科技出版社，2000年，第1版：56
1. Heldin CH and Westermark B. Role of platelet-derived growth factor in vivo (2nd ed.). New York: Plenum, 1996; p. 249-273.
2. Nakayama M, Fukuda N, Watanabe Y, et al. Low dose of eicosapentaenoic acid inhibits the exaggerated growth of vascular smooth muscle cells from spontaneously hypertensive rats through suppression of transforming growth factor-beta. J Hypertens, 1999; 17 (10): 1421-30.
3. Sabri A, Levy BI, Poitevin P, et al. Differential roles of AT1 and AT2 receptor subtypes in vascular trophic and phenotypie changes in response to stimulation with angiotensin Ⅱ. Arterioscler Thromb Vase Biol, 1997; 17 (2): 257-64.
4. Heagerty AM, Aalkjaer C, Bund S J, et al. Small artery structure in hypertension. Dual processes of remodeling and growth. Hypertension, 1993; 21 (4): 391-7
5. Schiffrin EL, Hayoz D. How to assess vascular remodelling in small and medium-sized muscular arteries in humans. J Hypertension, 1997; 15 (6): 517-584.
6. Dickhout JG, Lee RM. Structural and functional analysis of small arteries from young spontaneously hypertensiverats. Hypertension, 1997; 29 (3): 781-9.
7. Dzau VJ, Gibbons GH, Modshita R, et al. New perspectives in hypertension research. Potentials of vascular biology. Hypertension, 1994; 23 (6 Pt 2): 1132-40.
8. Lee RM, Adams MA, Friberg P, et al. Debate: vascular remodelling. J Hypertens, 1997; 15 (4): 333-7.
9. Komer PI, Angus JA. Vascular remodeling. Hypertension, 1997; 29 (4): 1065-6.
10. Bohr DF, Dominiczak AF, Webb Re. Pathophysiology of the vasculatttre in hypertension. Hypertension, 1991, 18 (5 supple): Ⅲ69-75
11. Rizzoni D, Castellano M, Porteri E, et al. Vascular structural and functional alterations before and after the development of hypertension in SHR. Am J Hypertens, 1994; 7 (2): 193-200.
12. Saltis J, Agrotis A and Bobik A. Differences in growth characteristics of vascular smooth muscle from spontaneously hypertensive and Wistar-Kyoto rats are growth factor dependent. J Hypertens, 1993; 11 (6): 629-37
13. Bussy C, Boutouyrie P, Lacolley P, et al. Intrinsic stiffness of the carotid arterial wall material in essential hypertensives. Hypertension, 2000; 35 (5): 1049-54
14. Intengan and Schiffrin. Structure and mechanical properties of resistance arteries in hypertension: role of adhesion molecules and extracellular matrix determinants. Hypertension, 2000; 36 (3): 312-8.
15. Touyz RM. Molecular and cellular mechanisms regulating vascular function and structure—implications in the pathogenesis of hypertension. Can J Cardiol, 2000; 16 (9): 1137-46.
16. Hadrava V, Kruppa U, Russo RC, et al. Vascular smooth muscle cell proliferation and its therapeutic modulation in hypertension. Am Heart J, 1991; 122 (4 Pt 2): 1198-203.
17．汪浩川，刘秉文．生长因子、癌基因与动脉平滑肌细胞增殖．生理科学进展，1994；5(1)：48-53
18. Wang DH, Prewitt RL, Beebe SJ. Regulation of PDGF-A: a possible mechanism for angiotension Ⅱ-induced vascular growth. Am J Physiol, 1995; 269(1 Pt 2): H356-364.
19. Parker SB, Wade SS, Prewitt RL. Pressure mediates angiotension Ⅱ-induced arterial hypertrophy and PDGF-A expression. Hypertension, 1998; 32 (3): 452-458.
20. Parker SB, DobrianAD, Wade SS, et al. AT (1) receptor inhibition does not reduce arterial wall hypertrophy or PDGF-A expression in renal hypertension. Am J Physiol Heart Circ Physiol, 2000; 278 (2): H613-22
21. Eghbali M, TomeK R, Sukhatme VP, et al. Differential effects of transforming growth factor beta-1 and phorbol myristate acetate on cardiac fibroblasts. Regulation of fibrillar collagen mRNAs and expression of early transcription factor. Circ Res, 1991; 69 (2): 483-490
22. Villarreal FJ, Dillmann WH. Cardiac hypertrophy-induced changes in mRNA levels for TGF-beta 1, fibronectin, andcollagen.Am J Physiol, 1992; 262 (6Pt2): H1861-6
23. Youn TJ, Kim HS, Oh BH, et al. Ventricular remodeling and transforming growth factor-beta 1 mRNA expression after nontransmural myocardial infarction in rats: effects of angiotensin converting enzyme inhibition and angiotensin Ⅱ type 1 receptor blockade. Basic Res Cardiol, 1999; 94 (4): 246-53.
24．彭龙云，高修仁，麦炜颐，等．高血压大鼠心肌TGF-β1和MMP2蛋白表达及RAS阻断后的变化．国际心血管病学杂志，2002；12(4)：303-306(英文)
25．高修仁，彭龙云，曾群英，等．ARB及ACEI联合应用对高血压大鼠心室重塑的影响．中华临床医药杂志，2003；1(4)：24-27
26. Agrotis A, Saltis J, Bobik A, et al. Transforming growth factor—TGF-β1 gene activation and growth of smooth muscle from hypertensive rats. Hypertension, 1994; 23 (5): 593-599
27. Griffin SA, Brown WC, MacPherson F, et al. Angiotensin Ⅱ causes vascular hypertrophy in part by a non-pressor mechanism. Hypertension, 1991; 17 (5): 626-35.
28. Paquet JL, Baudouin-Legros M, Brunelle G, et al. Angiotensin Ⅱ-indueed proliferation of aortic myocytes in spontaneously hypertensive rats. J Hypertens, 1990; 8 (6): 565-72
29. Postnov IuV. The role of mitochondrial calcium overload and energy deficiency in pathogenesis of arterial hypertension. Arkh Patol, 2001; 63 (3): 3-10.
30．文允镒．钙、钙调素与高血压关系的研究进展[J]．高血压杂志，1994；2(S1)：59-62
31．文允镒，张志明，胡蓓．钙与钙调素[M]．北京：化工出版社，1989，第一版：100-106，235-236
32．许顺滨，文允镒，陈孟勤．川芎嗪对正常及高血压大鼠动脉平滑肌Ca～(2+)内流的动力学影响[J]．中国应用生理学杂志，1989；1：26
33. Lu KP, Means AR. Regulation of the cell cycle by calcium and calmodulin. Endocr Rev, 1993; 14 (1): 40-58.
34. Wen YY, Jiang L, Chen M, et al. The changes of calmodulin and cAMP contents in tissues of renal hypertensive rats. Proc Chin Acad Med Sci Peking Union Med Coll, 1987; 2 (1): 22-7
35．靳琦．王琦“辨体—辨病—辨证诊疗模式”的理论要素与临床应用[J]．北京中医药大学学报2006；29(1)：41—45
36．卢焯明，潘毅，梁颖瑜，等．调肝肾、祛痰瘀治法提前干预对SHR血压及血管活性物质的影响[J]．中国中医药信息杂志，2004；11(9)：780—782
37．王琦．调治亚健康状态是中医学在21世纪对人类的新贡献[J]．北京中医药大学学报，2001：24(2)：1—4
38. Watanabe T, Barker TA, Berk BC. Angiotensin Ⅱ and the endothelium: diverse signals and effects. Hypertension, 2005; 45 (2): 163-9.
39. Chobanian AV, Bakris GL, Black HR, et al. The Seventh Report of the Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure: the JNC 7 report. JAMA, 2003; 289 (19): 2560-72.
40. Asmar RG, London GM, O'Rourke ME, et al. Improvement in blood pressure, artedal stiffness and wave reflections with a very-low-dose perindopdl/indapamide combination in hypertensive patient: a comparison with atenolol. Hypertension, 2001; 38 (4): 922-6.
41. Jorgensen B, Simonsen S, Endresen K, et al. Restenosis and clinical outcome in patients treated with amlodipine after angioplasty: results from the Coronary AngioPlasty Amlodipine REStenosis Study (CAPARES). J Am Coll Cardiol, 2000, 35 (3): 592-9.
42．付文波．他汀类药物对高血压病心血管重建作用的研究现状．心脏杂志，2006；18(5)：588
43．秦晋梅，李建军．他汀类药物的抗炎作用[J]．中国动脉硬化杂志，2004；12(3)：363-365
44．覃军，李爱民，何作云．他汀类药物对血管平滑肌细胞的直接作用及其意义[J]．中国动脉硬化杂志，2000：8(4)：373-375．
45．谢良地．HMG-CoA还原酶抑制剂的非调脂作用及机制(上)[J]．高血压杂志，2004，12(3)：265-269
46．张子新，谭庆，宋丽新，等．西洛他唑对家兔血管成形术后内膜增殖和血管重构的影响．中国医科大学学报，2005，34(5)：420-421
47．汪新良，何兆初，曾昭华，等．安体舒通对感觉神经损伤性盐敏感性高血压大鼠血管重构及RAAS的影响．中国心血管病研究杂志，2006，4(10)：773-776
48. Dumitrascu R, Weissmann N, Ghofrani HA, et al. Activation of soluble guanylate cyclase reverses experimental pulmonary hypertension and vascular remodeling. Circulation, 2006; 113 (2): 286-95.
49. Pascarella L, Schonbein GW, Bergan JJ. Microcirculation and venous ulcers: a review. Ann Vase Surg, 2005; 19 (6): 921-7.
50．曹守沛，钱春发，金妙文．中西医结合治疗急性心肌梗死疗效分析[J]．实用中医内科杂志，2005；19(4)：345-346
51．史载祥．后再灌注时代难题的中西医结合治疗思考[J]．中西医结合心脑血管病杂志，2005：3(1)：2-5
52．王振涛，王硕仁，赵明镜，等．活血、益气方药对心梗后左心衰大鼠血TNF和血小板聚集影响的对比研究[J]．中国中医药信息杂志，2002；9(11)：20-22
53. Ji XY, Tan BK, Zhu YZ. Salvia miltiorrhiza and ischemie diseases. Acta Pharmacol Sin, 2000; 21 (12): 1089-94
54. Sun J, Huang SH, Tan BK, et al. Effects of purified herbal extract of Salvia miltiorrhiza on ischemic rat myocardium after acute myocardial infarction. Life Sci, 2005; 76 (24): 2849-60.
55. Wang B, Zhang JD, Feng J-B, et al. Effect of traditional Chinese medicine Qin-Dan-Jiang-Ya-Tang on remodeled vascular phenotype and osteopontin in spontaneous hypertensive rats. J Ethnopharmacol, 2007:110 (1): 176-82
56．严灿，邓中炎，潘毅，等．活血祛痰法逆转高血压左室肥厚的实验研究．中国中医基础杂志，1999；5(4)：25-27
57．李建平，严灿，潘毅，等．活血祛痰方药预防自发性高血压大鼠心肌纤维化实验研究．中医杂志，2000；41(5)：301-302
58．李剑，潘毅，严灿，等．滋补肝肾、活血祛痰方药对原发性高血压大鼠左心室肥厚的影响．山西中医，2003；19(2)：54-56
59．卢焯明，潘毅，梁颖瑜，等．调肝肾、祛痰瘀治法提前干预对SHR血压及血管活性物质的影响[J]．中国中医药信息杂志，2004;;11(9)：780-782
1．王培林．遗传病学，北京：人民卫生出版社，2000，第1版：315，318
2．方厚华．医学实验模型动物，北京：军事医学科学出版社，2002，第1版：28-30
3．谢雍．“母体疗法”治疗婴幼儿泄泻．四川中医，1990；8(2)：19-20
4．王纪民．哺乳幼儿病宜母子同治．四川中医，1989；7(7)：6-7
5．渡边贤治．汉方经母乳治疗婴儿皮疹有效7例及汉方药的乳汁移行．日本东洋医学杂志，1996；46(6)：135-137
6．刘湘华．肝阳上亢证患者个性、情绪特征及血浆精氨酸加压素水平的初步研究．湖南医科大学学报，1998：23(1)：31
7．叶任高．内科学，北京：人民卫生出版社，2002，第5版：258-261
8．孙宁玲，徐成斌．今日高血压，北京：中国医药科技出版社，2000，第1版：125-131，152-154，160，166-172，149，431，596-599
9．邓中炎，徐志伟，陈群，等．中医基础理论体系现代研究，广州：广东科技出版社，2002，第一版：7，88，193
10．王本祥．现代中药药理与临床，天津，天津科技翻译出版公司，2004，第1版：
11．侯家玉．中药药理学，中国中医药出版社，2002，第1版：
12．李军，张丽萍，张振凌，等．地黄研究进展．河南中医学院学报，2005；121 (20)：79-83
13．曾艳，贾正平，张汝学，等．地黄化学成分及药理研究进展．中成药，2006；28(4)：609-611
14．管家齐，郭艳丽，吴海良，等．地黄药理研究近况．中药材，2006；24(5)：380-382
15．孙苏亚，王锦，刘铮，等．龟板提取液对去势大鼠骨质疏松的作用．中药药理与临床，1998：14(4)：20
16．黎晖，周健洪，陈东风，等．龟板对大鼠骨髓间充质干细胞向成骨分化的影响．中药新药与临床药理，2005；16(3)：159-161
17．朱莉芬，李美珠，钟佳新，等．龟鹿补肾口服液的药理研究．中成药，1993；15(2)；24
18．李以菊，孙泽玲．丹参的临床应用进展．实用医技杂志，2006，8；13(16)：2903-2905
19．付辛芳，刘晓红．丹参的药理作用与临床应用研究进展．中国药业，2006；15(1)：76-77
20．李艳杰，孙玉华．丹参素药理作用研究进展．延边大学医学学报，2006；29(1)：73-75
21．书海，李青旺，郑雪莉，等．牛膝正丁醇萃取物对小鼠繁殖性能的影响．西北农林科技大学学报，2006；34(6)：31-34
22．曹明俐，张小军，王振生，等．牛膝的药理作用研究进展．中医药信息，2002；18(3)：20-21
23．谭德讲，邱海强，袁惠南．田七洋参冲剂的抗应激作用和免疫促进作用．2000，7：11(4)：242-243
24．屠婕红，余菁，陈伟光．瓜蒌的化学成分和药理作用研究概况．中国药师，2004；7(7)：562-563
25．孙兴龙，段崇英．瓜蒌的药理作用及临床应用．中国药师，2003；6(10)：655-656
26．刘佳，富志军．钩藤的研究概况．海峡药学，2006；18(5)：90-93
27．莫志贤，许丹丹．钩藤临床运用的新进展．时珍国医国药，2006；17(5)：684-685
28．何小琦．滋肾养肝活血化瘀法治疗糖尿病并高血压60例临床观察．实用医学杂 志，1998：14(1)：69
1．蔡文琴，现代实用细胞与分子生物学实验技术．北京：人民军医出版社，2003，第一版：13—15
2. Carson DA, Ribeiro JM. Apoptosis and disease. Lancet, 1993; 341 (8855): 1251-4.
3．刘树民，李玉洁，罗明媚．中药毒性研究及思路探讨[J]．世界科学技术—中医药现代化，2003；5(2)：21-23
4．高月，吕秋军，刘永学，等．中药复方物质基础的研究[J]．中国新药杂志，2000；9(5)：307-308
5．马骏，唐灿，李增强，等．中药血清药理学的方法学研究——在中药复方药物动力学研究中的应用[J]．中药药理与临床，1999：15(3)：44-46
6．杨彦芳，王玉芹．中药复方血清药理学方法规范化探讨[J]．中国中西医结合杂志，2000；20(5)：380-382
7．周明眉，杨奎，姜远平，等．中药血清药理学的方法学研究——采血时间的确定及时效关系研究[J]．中药药理与临床，1999；15(1)：43-44
8．周明眉，杨奎，姜远平，等．中药血清药理学的方法学研究——含药血清低温保存和血清灭活的影响[J]．中药药理与临床，1999；15(2)：44-46
9．李仪奎，吴健宇．中药血清药理实验方法给药的通法方案．中国药理学通报，1999；15(6)：569-570
1．丁文勇，赵宝昌，韩旭．钙调素的环核苷酸磷酸二酯酶检测法．大连医科大学学报，1999：21(1)：3-6
2. Dolphin AC. Mechanisms of modulation of voltage-dependent calcium channels by G proteins. J Physiol, 1998; 506 (Pt 1): 3-11.
3. Marks AR. Intracellular calcium-release channels: regulators of cell life and death. Am J Physiol, 1997; 272 (2 Pt 2): H597-605.
4. Tostes RC, Storm DS, Chi DH, et al. Intracellular calcium stores and oscillatory contractions in arteries from genetically hypertensive rats. Hypertens Res, 1996; 19 (2): 103-11.
5. Morimoto S, Takeda K, Oguni A, et al. Reduction of white coat effect by cilnidipine in essential hypertension. Am J Hypertens, 2001; 14 (10): 1053-7.
6. Onose Y, Oki T, Yamada H, et al. Effect of cilnidipine on left ventricular diastolic function in hypertensive patients as assessed by pulsed Doppler echocardiography and pulsed tissue Doppler imaging. Jpn Circ J, 2001; 65 (4): 305-9.
7. Resnick LM. Physiologic rationale for calcium antagonist therapy in essential hypertension. Ethn Dis, 1998; 8 (1): 111-9.
8. Vincent M, Samani NJ, Gauguier D, et al. A pharmacogenetic approach to blood pressure in Lyon hypertensive rats. A chromosome 2 locus influences the response to a calcium antagonist. J Clin Invest, 1997; 100 (8): 2000-6.
9. Kosch M, Hausberg M, Barenbrock M, et al. Increased membraneous calcium concentrations in primary hypertension: a causal link to pathogenesis? J Hum Hypertens, 2001; 15 (1): 37-40.
10. Cicco G, Carbonara MC, Stingi GD, et al. Cytosolic calcium and hemorheological patterns during arterial hypertension. Clin Hemorheol Microcirc, 2001; 24 (1): 25-31
11. Sugden PH. Signaling pathways activated by vasoactive peptides in the cardiac myocyte and their role in myocardial pathologies. J Card Fail, 2002; 8 (6 Suppl): S359-69.
12. Kosch M, Hausberg M, Westermann G, et al. Alterations in calcium and magnesium content of red cell membranes in patients with primary hypertension. Am J Hypertens, 2001; 14 (3): 254-8.
13. Ohkusa T, Hisamatsu Y, Yano M, et at. Altered cardiac mechanism and sarcoplasmic reticulum function in pressure overload-induced cardiac hypertrophy in rats. J Mol Cell Cardiol, 1997; 29 (1): 45-54.
14. Jiang M, Xu A, Tokmakejian S, et al. Thyroid hormone-induced overexpression of functional ryanodine receptors in the rabbit heart. Am J Physiol Heart Circ Physiol, 2000; 278 (5): H1429-38.
15. Rannou F, Dambrin G, Marry I, et al. Expression of the cardiac ryanodine receptor in the compensated phase of hypertrophy in rat heart. Cardiovasc Res, 1996; 32 (2): 258-65.
16. Assayag P, Charlemagne D, de Leiris J, et al. Senescent heart compared with pressure overload-induced hypertrophy. Hypertension, 1997; 29 (1 Pt 1): 15-21.
17. Arai M, Alpert NR, MacLennan DH, et al. Alterations in sarcoplasmic reticulum gene expression in human heart failure. A possible mechanism for alterations in systolic and diastolic properties of the failing myocardium. Circ Res, 1993; 72 (2): 463-9.
18. Anger M, Lompre AM, Vallot O, et al. Cellular distribution of Ca2+ pumps and Ca2+ release channels in rat cardiac hypertrophy induced by aortic stenosis, Circulation, 1998; 98 (22): 2477-2486
19. Rivet-Bastide M, Vandecasteele G, Hatem S, et al. cGMP-stimulated cyclic nucleotide phosphodiesterase regulates the basal calcium current in human atrial myocytes. J Clin Invest, 1997, 99: 2710-2718.
20. Boixel C, Tessier S, Pansard Y, et al. Tyrosine kinase and protein kinase C regulate L-type Ca(2+) current cooperatively in human atrial myocytes. Am J Physiol Heart CircPhysiol, 2000; 278 (2): H670-6.
21. Mogami H, Kojima I. Stimulation of calcium entry is prerequisite for DNA synthesis induced by platelet-derived growth factor in vascular smooth muscle cells. Biochem Biophys Res Commun, 1993; 196 (2): 650-8.
22. Short AD, Bian J, Ghosh TK, et al. Intracellular Ca2+ pool content is linked to control of cell growth. Proc Natl Acad Sci U S A, 1993; 90 (11): 4986-90.
23. Dolmetsch RE, Lewis RS, Goodnow CC, et al. Differential activation of transcription factors induced by Ca2+ response amplitude and duration. Nature, 1997; 386 (6627): 855-8
24. Bers DM, Perez-Reyes E. Ca channels in cardiac myocytes: structure and function in Ca influx and intracellular Ca release. Cardiovasc Res, 1999; 42 (2): 339-60.
25．韩济生，神经科学原理，北京：北京医科大学出版社，1993年，第一版：222-227
26．杨钢，内分泌生理与病理生理学，天津：天津科学技术出版社，1996年，第一版：59-67
27．张均田，现代药理实验方法，北京：北京医科大学中国协和医科大学联合出版社，1998年，第一版：472-473
28. Chafouless J G, Dedmn J R, Means A R. Redioimmunoassay of cal modulin. Methods Enzymol, 1983; 84: 138
29. Strobl J S, Peterson U A, Woodfork K A. A survey of human breast cancer sensitivity to growth inhidition by calmodulin antagonists in tissue culture. Biochem Pharmacol, 1994; 47: 2157
30. Brooks-Frederich K M, Cianciarulo F L, Rittling S R, et al. Cell cycledependent regulation of Ca′'in young and senescent WI-38 cells. Exp Cell Res, 1993; 205:412
31. Wallace R W, Tallant E A, Cheung W Y. Assay of calmidulin by Ca′ '—dependent phisphodiesterase. Methods Enzymol, 1983: 102:39
32．孙宁玲，徐成斌．今日高血压，北京：中国医药科技出版社，2000年，第1版：56
1. Heldin CH and Westermark B. Role of platelet-derived growth factor in vivo (2nd ed.). New York: Plenum, 1996; p. 249-273.