2012313_加减一阴煎对血管内皮细胞和平滑肌细胞增殖影响

2012313_加减一阴煎对血管内皮细胞和平滑肌细胞增殖影响 加减一阴煎对血管内皮细胞和平滑肌细胞增殖的影响 中山大学医学部临床医学系2001级 郑跃 陈惠国 罗杰鸿 摘要 目的:探讨中药制剂加减一阴煎对培养的血管内皮细胞和平滑肌细胞增殖的影响及其机制。方法:以中药复方水煎液对新西兰大白兔灌胃一周～在无菌条件下取血制备血清药～在培养血管内皮和平滑肌细胞中加入不同剂量的血清药～通过细胞计数研究药血清对它们增殖的影响及其机制。结果:与空白组和纯血清组对比～平滑肌细胞数目减少～内皮细胞数目大幅度增加～并有剂量依赖性。结论:加减一阴煎对血管平滑肌细胞增殖有一定的抑制作用～而对血管内皮细胞有触增殖的作用。 关键词 加减一阴煎 血管平滑肌细胞 血管内皮细胞 血清药 研究表明许多植物中含有植物雌激素，如异黄酮、β,谷甾醇等，它们既有拟雌激素样的作用，同时还具有心血管保护作用，以明代《景岳全书》中名方一阴煎为主方的加减一阴煎主要由生地、熟地、白芍、麦冬、黄精、知母、地骨皮、牛膝和炙甘草组成，滋阴补血，活血逐瘀的功效，用于治疗治疗妇女的月经不调的疾病，现代有人将它用于中药人工周期来 [1]治疗卵巢功能早衰继发性闭经，表明加减一阴煎具有恢复卵巢功能的作用，具有拟雌激素样的作用。而加减一阴煎是否具有心血管保护作用还未见报道。因此本实验从从中医养阴补血理论出发，运用生理、生化和分子生物学的方法从细胞水平探讨加减一阴煎抗AS的机制，使之能对绝经期妇女AS防治有所帮助。 材料与方法 一 材料 1( 动物: 健康成年新西兰大白兔(2.0-2.5kg)和SD大鼠(150-250g)均由中山大学医学部动物实验中心提供。小牛主动脉内皮细胞，由广州市奶牛研究所提供。 2( 主要药物和试剂: 加减一阴煎(主要由生地、熟地、白芍、麦冬、黄精、知母、地骨皮、牛膝和炙甘草组成)， 胰蛋白酶，血管紧张素? 、PBS溶液、无血清的DMEN和含血清含DMEM培养液、 3. 主要实验仪器: CO培养箱(德国HERAEUAS)二台，Revco超低温冰箱一台，冷冻离心机(德国2 integrated energy, chemicals and textile Yibin city, are the three core pillars of the industry. In 2014, the wuliangye brand value to 73.58 billion yuan, the city's liquor industry slip to stabilise. Promoting deep development of integrated energy, advanced equipment manufacturing industry, changning district, shale gas production capacity reached 277 million cubic metres, built the country's first independent high-yield wells and pipelines in the first section, the lead in factory production and supply to the population. 2.1-3 GDP growth figure 2.1-4 Yibin, Yibin city, Yibin city, fiscal revenue growth 2.1.4 topography terrain overall is Southwest, North-Eastern State. Low mountains and hills in the city landscape as the main ridge-and-Valley, pingba small fragmented nature picture for "water and the second land of the seven hills". 236 meters to 2000 meters above sea level in the city, low mountain, 46.6% hills 45.3%, pingba only 8.1%. 2.1.5 development of Yibin landscapes and distinctive feature in the center of the city, with limitations, and spatial structure of typical zonal group, 2012-cities in building with an area of about 76.2km2. From city-building situation, "old town-the South Bank" Center construction is lagging behind, disintegration of the old city is slow, optimization and upgrading, quality public service resources are still heavily concentrated in the old town together. Southbank Centre has not been formed, functions of the service area space is missing. Meanwhile, peripheral group centres service was weak and inadequate accounting for city development, "suspicious" pattern could not be formed. As regards transport, with the outward expansion of cities, cities have been expanding, centripetal city traffic organization has not changed, integrated energy, chemicals and textile Yibin city, are the three core pillars of the industry. In 2014, the wuliangye brand value to 73.58 billion yuan, the city's liquor industry slip to stabilise. Promoting deep development of integrated energy, advanced equipment manufacturing industry, changning district, shale gas production capacity reached 277 million cubic metres, built the country's first independent high-yield wells and pipelines in the first section, the lead in factory production and supply to the population. 2.1-3 GDP growth figure 2.1-4 Yibin, Yibin city, Yibin city, fiscal revenue growth 2.1.4 topography terrain overall is Southwest, North-Eastern State. Low mountains and hills in the city landscape as the main ridge-and-Valley, pingba small fragmented nature picture for "water and the second land of the seven hills". 236 meters to 2000 meters above sea level in the city, low mountain, 46.6% hills 45.3%, pingba only 8.1%. 2.1.5 development of Yibin landscapes and distinctive feature in the center of the city, with limitations, and spatial structure of typical zonal group, 2012-cities in building with an area of about 76.2km2. From city-building situation, "old town-the South Bank" Center construction is lagging behind, disintegration of the old city is slow, optimization and upgrading, quality public service resources are still heavily concentrated in the old town together. Southbank Centre has not been formed, functions of the service area space is missing. Meanwhile, peripheral group centres service was weak and inadequate accounting for city development, "suspicious" pattern could not be formed. As regards transport, with the outward expansion of cities, cities have been expanding, centripetal city traffic organization has not changed, EPPENDORF)一台，超净工作台二台，倒置显微镜一台，各种水浴箱，天平，自动煎药煲和普通冰箱 二 方法 (一) 药血清的制备 方剂成分的含量:生地15g、熟地20g、白芍15g、麦冬15g、黄精15g、知母15g、牛膝15g 、丹参15g、川芎15g、地骨皮12g、和炙甘草6g。每剂药加水750ml并煎至250ml。选择健康成年新西兰大白兔，每天用中药复方水煎液定时灌胃二次，每次60ml，持续一周. 最后一次灌胃后2小时，在无菌局麻条件下行兔颈总动脉插管放血，离心取血清，56?、30min灭活补体，4?保存备用。对照组灌等量生理盐水，同法制备对照血清。 (二)研究药血清影响VSMC增殖的方法 1.VSMC的培养 利用组织块贴壁法进行VSMC培养.取150-250g大鼠,颈椎脱臼法处死,迅速打开胸腔,无菌条件下取出胸主动脉,放入无菌的PBS平衡液中,清除血块和外周结缔组织,去除内皮, 3o用眼科剪剪成1mm组织块,均匀分散于培养瓶中,放入37C二氧化碳培养箱孵育,待组织块贴壁后,加入含20%小牛血清的无酚红的DMEN液中继续培养, 七天后细胞从组织块边沿长出, o待细胞张满瓶底后可传代.倒去培养液,用PBS洗涤,加入0.125%的胰蛋白酶于37C消化细胞,倒置显微镜下观察,待细胞收缩变圆,立即倾倒消化液,加入含10%小牛血清的无酚红的DMEN液终止反应,1:2的比例进行分瓶传代.实验用4-6代细胞. 2. 加入药血清培养和细胞计数 5 VSMC以4X10个细胞/ml密度0.5ml接种于24孔板,当细胞长至亚融合状态,用无血清 -7DMEM液培养24h;加入不同浓度的药血清及对照血清,1h后加入终浓度为10mol/L的血管紧 o张素?,培养18h ,倒去培养液,用PBS洗涤,加入0.125%的胰蛋白酶于37C消化细胞,倒去胰酶,加入0.5mlPBS,吹打使全部细胞悬浮,用滴管滴入计数板,计数. 每孔细胞数目=每计数板细胞×5000 3. 实验分组 分为三组:空白组?;对照组?(每孔加10-50ul的对照血清);药血清组(分别加:10ul、20ul、30ul、50ul、80ul、100ul、120ul和150ul药血清)。每组设4个复管. (三) 研究药血清影响血管内皮增殖的方法 1.新生小牛主动脉内皮细胞(BAECs)的培养和细胞计数 integrated energy, chemicals and textile Yibin city, are the three core pillars of the industry. In 2014, the wuliangye brand value to 73.58 billion yuan, the city's liquor industry slip to stabilise. Promoting deep development of integrated energy, advanced equipment manufacturing industry, changning district, shale gas production capacity reached 277 million cubic metres, built the country's first independent high-yield wells and pipelines in the first section, the lead in factory production and supply to the population. 2.1-3 GDP growth figure 2.1-4 Yibin, Yibin city, Yibin city, fiscal revenue growth 2.1.4 topography terrain overall is Southwest, North-Eastern State. Low mountains and hills in the city landscape as the main ridge-and-Valley, pingba small fragmented nature picture for "water and the second land of the seven hills". 236 meters to 2000 meters above sea level in the city, low mountain, 46.6% hills 45.3%, pingba only 8.1%. 2.1.5 development of Yibin landscapes and distinctive feature in the center of the city, with limitations, and spatial structure of typical zonal group, 2012-cities in building with an area of about 76.2km2. From city-building situation, "old town-the South Bank" Center construction is lagging behind, disintegration of the old city is slow, optimization and upgrading, quality public service resources are still heavily concentrated in the old town together. Southbank Centre has not been formed, functions of the service area space is missing. Meanwhile, peripheral group centres service was weak and inadequate accounting for city development, "suspicious" pattern could not be formed. As regards transport, with the outward expansion of cities, cities have been expanding, centripetal city traffic organization has not changed, integrated energy, chemicals and textile Yibin city, are the three core pillars of the industry. In 2014, the wuliangye brand value to 73.58 billion yuan, the city's liquor industry slip to stabilise. Promoting deep development of integrated energy, advanced equipment manufacturing industry, changning district, shale gas production capacity reached 277 million cubic metres, built the country's first independent high-yield wells and pipelines in the first section, the lead in factory production and supply to the population. 2.1-3 GDP growth figure 2.1-4 Yibin, Yibin city, Yibin city, fiscal revenue growth 2.1.4 topography terrain overall is Southwest, North-Eastern State. Low mountains and hills in the city landscape as the main ridge-and-Valley, pingba small fragmented nature picture for "water and the second land of the seven hills". 236 meters to 2000 meters above sea level in the city, low mountain, 46.6% hills 45.3%, pingba only 8.1%. 2.1.5 development of Yibin landscapes and distinctive feature in the center of the city, with limitations, and spatial structure of typical zonal group, 2012-cities in building with an area of about 76.2km2. From city-building situation, "old town-the South Bank" Center construction is lagging behind, disintegration of the old city is slow, optimization and upgrading, quality public service resources are still heavily concentrated in the old town together. Southbank Centre has not been formed, functions of the service area space is missing. Meanwhile, peripheral group centres service was weak and inadequate accounting for city development, "suspicious" pattern could not be formed. As regards transport, with the outward expansion of cities, cities have been expanding, centripetal city traffic organization has not changed, 在无菌条件下取新生小牛胸主动脉,按常规方法分离,培养BAECs,原代BAECs呈铺路石状,?因子染色阳性.实验采用传代培养第3-5代的细胞,培养液为含100ml/L小牛血清的无酚红的DMEN,处理前24h换用无血清加入转铁蛋白等补充营养的无酚红的DMEN. 每孔细胞数目=每计数板细胞×5000 2.实验分组 5血管内皮细胞以3×10个细胞/ml密度 0.5ml接种于24孔板，分为三大组:空白组?;对照组?(每孔加不同量的对照血清);药血清组?(10ul、20ul、30ul、50ul、80ul、100ul、120ul和150ul八个药血清)。每组设6个复管。 四 统计学处理: 所有数据均以mean?SD表示,组间及组内均数的比较用方差分析(？检验)。 结果 一 药血清对VSMC增殖的影响: 图一示与空白组和对照组比较，不同量的药血清对VSMC增殖有一定的抑制作用，并有一定的剂量依赖性，抑制作用的最适浓度为80ul。当药血清量>80ul时，其抑制作用呈降低趋势。 平滑肌细胞计数结果 70 60 50 40 30 20 10细胞数(×5000) 0 空白对照1020405080100120150 药血清量(ul) 图一:药血清对VSMC增殖的影响(10ul药血清组P>0.05,其余组P<0.05) 二 药血清对血管内皮细胞增殖的影响: 图二示与对照组相比，药血清对血管内皮细胞有明显的触增殖作用，有一定的剂量依赖性，在30ul增殖最强。{触增殖率=(每个剂量的计数结果,空白组计数结果)/空白组计数结果} integrated energy, chemicals and textile Yibin city, are the three core pillars of the industry. In 2014, the wuliangye brand value to 73.58 billion yuan, the city's liquor industry slip to stabilise. Promoting deep development of integrated energy, advanced equipment manufacturing industry, changning district, shale gas production capacity reached 277 million cubic metres, built the country's first independent high-yield wells and pipelines in the first section, the lead in factory production and supply to the population. 2.1-3 GDP growth figure 2.1-4 Yibin, Yibin city, Yibin city, fiscal revenue growth 2.1.4 topography terrain overall is Southwest, North-Eastern State. Low mountains and hills in the city landscape as the main ridge-and-Valley, pingba small fragmented nature picture for "water and the second land of the seven hills". 236 meters to 2000 meters above sea level in the city, low mountain, 46.6% hills 45.3%, pingba only 8.1%. 2.1.5 development of Yibin landscapes and distinctive feature in the center of the city, with limitations, and spatial structure of typical zonal group, 2012-cities in building with an area of about 76.2km2. From city-building situation, "old town-the South Bank" Center construction is lagging behind, disintegration of the old city is slow, optimization and upgrading, quality public service resources are still heavily concentrated in the old town together. Southbank Centre has not been formed, functions of the service area space is missing. Meanwhile, peripheral group centres service was weak and inadequate accounting for city development, "suspicious" pattern could not be formed. As regards transport, with the outward expansion of cities, cities have been expanding, centripetal city traffic organization has not changed, integrated energy, chemicals and textile Yibin city, are the three core pillars of the industry. In 2014, the wuliangye brand value to 73.58 billion yuan, the city's liquor industry slip to stabilise. Promoting deep development of integrated energy, advanced equipment manufacturing industry, changning district, shale gas production capacity reached 277 million cubic metres, built the country's first independent high-yield wells and pipelines in the first section, the lead in factory production and supply to the population. 2.1-3 GDP growth figure 2.1-4 Yibin, Yibin city, Yibin city, fiscal revenue growth 2.1.4 topography terrain overall is Southwest, North-Eastern State. Low mountains and hills in the city landscape as the main ridge-and-Valley, pingba small fragmented nature picture for "water and the second land of the seven hills". 236 meters to 2000 meters above sea level in the city, low mountain, 46.6% hills 45.3%, pingba only 8.1%. 2.1.5 development of Yibin landscapes and distinctive feature in the center of the city, with limitations, and spatial structure of typical zonal group, 2012-cities in building with an area of about 76.2km2. From city-building situation, "old town-the South Bank" Center construction is lagging behind, disintegration of the old city is slow, optimization and upgrading, quality public service resources are still heavily concentrated in the old town together. Southbank Centre has not been formed, functions of the service area space is missing. Meanwhile, peripheral group centres service was weak and inadequate accounting for city development, "suspicious" pattern could not be formed. As regards transport, with the outward expansion of cities, cities have been expanding, centripetal city traffic organization has not changed, 中药触发内皮增殖 对照组 对照组 药血清组 药血清组 44(×10) 触增率 (×10) 触增率 10ul 13.33?1.08 0.82 16.58?4.34 1.26 20ul 23.00?1.70 2.14 38.17?7.10 4.23 30ul 23.67?2.27 2.23 33.00?3.83 3.50 50ul 10.92?1.46 0.49 33.75?4.22 3.62 80ul 18.25?1.72 0.92 37.67?4.27 2.96 120ul 24.92?1.39 1.62 41.58?7.61 3.38 150ul 24.58?3.01 1.59 48.42?2.22 4.10 中药触发内皮增殖 7 6 5 4药血 对照3增殖率 2 1 0 1020305080120150 加入血清(ul) 图二 中药触发内皮增殖 讨论 研究表明雌激素具有广泛的心血管保护作用雌激素可保护内皮功能和抑制血管平滑肌[1，8]细胞的增殖，临床上使用雌激素替代疗法可有效的减少绝经期妇女的AS的发病率。但雌 激素的长期使用但也会带来如子宫内膜癌和乳腺癌等严重副作用。如何在发挥雌激素的保护 作用同时，减少它的副作用是急需解决的问题。目前临床上用于治疗AS的中药方剂主要集 integrated energy, chemicals and textile Yibin city, are the three core pillars of the industry. In 2014, the wuliangye brand value to 73.58 billion yuan, the city's liquor industry slip to stabilise. Promoting deep development of integrated energy, advanced equipment manufacturing industry, changning district, shale gas production capacity reached 277 million cubic metres, built the country's first independent high-yield wells and pipelines in the first section, the lead in factory production and supply to the population. 2.1-3 GDP growth figure 2.1-4 Yibin, Yibin city, Yibin city, fiscal revenue growth 2.1.4 topography terrain overall is Southwest, North-Eastern State. Low mountains and hills in the city landscape as the main ridge-and-Valley, pingba small fragmented nature picture for "water and the second land of the seven hills". 236 meters to 2000 meters above sea level in the city, low mountain, 46.6% hills 45.3%, pingba only 8.1%. 2.1.5 development of Yibin landscapes and distinctive feature in the center of the city, with limitations, and spatial structure of typical zonal group, 2012-cities in building with an area of about 76.2km2. From city-building situation, "old town-the South Bank" Center construction is lagging behind, disintegration of the old city is slow, optimization and upgrading, quality public service resources are still heavily concentrated in the old town together. Southbank Centre has not been formed, functions of the service area space is missing. Meanwhile, peripheral group centres service was weak and inadequate accounting for city development, "suspicious" pattern could not be formed. As regards transport, with the outward expansion of cities, cities have been expanding, centripetal city traffic organization has not changed, integrated energy, chemicals and textile Yibin city, are the three core pillars of the industry. In 2014, the wuliangye brand value to 73.58 billion yuan, the city's liquor industry slip to stabilise. Promoting deep development of integrated energy, advanced equipment manufacturing industry, changning district, shale gas production capacity reached 277 million cubic metres, built the country's first independent high-yield wells and pipelines in the first section, the lead in factory production and supply to the population. 2.1-3 GDP growth figure 2.1-4 Yibin, Yibin city, Yibin city, fiscal revenue growth 2.1.4 topography terrain overall is Southwest, North-Eastern State. Low mountains and hills in the city landscape as the main ridge-and-Valley, pingba small fragmented nature picture for "water and the second land of the seven hills". 236 meters to 2000 meters above sea level in the city, low mountain, 46.6% hills 45.3%, pingba only 8.1%. 2.1.5 development of Yibin landscapes and distinctive feature in the center of the city, with limitations, and spatial structure of typical zonal group, 2012-cities in building with an area of about 76.2km2. From city-building situation, "old town-the South Bank" Center construction is lagging behind, disintegration of the old city is slow, optimization and upgrading, quality public service resources are still heavily concentrated in the old town together. Southbank Centre has not been formed, functions of the service area space is missing. Meanwhile, peripheral group centres service was weak and inadequate accounting for city development, "suspicious" pattern could not be formed. As regards transport, with the outward expansion of cities, cities have been expanding, centripetal city traffic organization has not changed, [2]中在活血化瘀药和里气、补气药上，而对养阴补血方面研究甚少。对于广大绝经期妇女而言，往往有阴虚内热之症，加减一阴煎主要通过其滋阴补血的功效来治疗治疗妇女的月经不 [3]调的疾病，现代有人将它用于中药人工周期来治疗卵巢功能早衰继发性闭经 ，这种类似雌激素的作用对临床上使用雌激素替代疗法来抗AS是一个很好的补充，可通过与加减一阴煎的合用来减少雌激素的用量，这对缓解雌激素的副作用大有裨益，有助于广大绝经期妇女AS的防治。同时通过对加减一阴煎的作用机制的深入研究，有望寻找到一种新的有效的抗AS药物，使之对广大男性患者也有效。 大量研究证实，血管内皮细胞(VECs)和血管平滑肌细胞(VSMC)的增殖、迁移是AS [9]形成过程中的关键环节， 血管内皮细胞在血管创伤修复、止血、血栓形成等一系列生理 [4]病理过程中起重要作用，VECs的损伤和VSMC的异常增殖导致血管腔狭窄和痉挛是多种疾病(高血压、AS、等)发生发展的基础，加减一阴煎在兔体内经过生物转化后，在血清中产生有效成分，能促进血管内皮细胞增殖，维持内皮的完整性，又能抑制VSMC增殖，可见其具有血管的保护作用，但其对平滑肌增殖的抑制和对血管内皮的触增殖作用的途径和机制尚不清楚，有待进一步的研究。 在抑制平滑肌增殖实验中，当药血清量>80ul时，其抑制增殖的作用降低。其原因可能是由于80ul血清已经发挥最大的抑制增殖作用，同时由于血清本身对平滑肌细胞有营养增殖作用(由表二和图二，血清对照组结果表明血清也有血管触增殖作用)，抵消了药血清有效成分的抑制作用。 本实验采用药血清法研究体外培养细胞的增值。复方中药体外实验主要有两种方法，一种是将中药粗提物直接添加(如水煎液)作为作用细胞的处理因素，另一种是制成含药血清 [5]来作用细胞用于作用机理的研究。两者方法各有优缺点，大致认为直接添加有三方面缺点:a.缺乏复方的体内代谢物或机体反应物等实际有效成分，引起假阴性;b.机体非吸收物质的体外作用，造成假阳性;c.粗提物理化性质如杂质、酸碱度、渗透压、pH 值等对离体系的干扰。而含药血清法具有两大优势:a.体现复方药物的体内生化转化效应;b.克服药物本身理化性质对实验的直接干扰。当然，含药血清也有其无法克服的缺点:药物血清材料可来自各种不同生理病理状态的个体，会出现含药血清差异;而且经过体内反应，药物含量进一步被稀释，成分更加复杂，增加了质控难度。血清中部分物质已是药物作用机体细胞的反应，如果细胞存在反馈机制，血清二次重复作用于细胞，将可能掩盖药物血清的实际作用。含药 ,6，7,血清对离体系统同样也有干扰，如血清本身对体外酶活性的影响等。但与直接加药法相integrated energy, chemicals and textile Yibin city, are the three core pillars of the industry. In 2014, the wuliangye brand value to 73.58 billion yuan, the city's liquor industry slip to stabilise. Promoting deep development of integrated energy, advanced equipment manufacturing industry, changning district, shale gas production capacity reached 277 million cubic metres, built the country's first independent high-yield wells and pipelines in the first section, the lead in factory production and supply to the population. 2.1-3 GDP growth figure 2.1-4 Yibin, Yibin city, Yibin city, fiscal revenue growth 2.1.4 topography terrain overall is Southwest, North-Eastern State. Low mountains and hills in the city landscape as the main ridge-and-Valley, pingba small fragmented nature picture for "water and the second land of the seven hills". 236 meters to 2000 meters above sea level in the city, low mountain, 46.6% hills 45.3%, pingba only 8.1%. 2.1.5 development of Yibin landscapes and distinctive feature in the center of the city, with limitations, and spatial structure of typical zonal group, 2012-cities in building with an area of about 76.2km2. From city-building situation, "old town-the South Bank" Center construction is lagging behind, disintegration of the old city is slow, optimization and upgrading, quality public service resources are still heavily concentrated in the old town together. Southbank Centre has not been formed, functions of the service area space is missing. Meanwhile, peripheral group centres service was weak and inadequate accounting for city development, "suspicious" pattern could not be formed. As regards transport, with the outward expansion of cities, cities have been expanding, centripetal city traffic organization has not changed, integrated energy, chemicals and textile Yibin city, are the three core pillars of the industry. In 2014, the wuliangye brand value to 73.58 billion yuan, the city's liquor industry slip to stabilise. Promoting deep development of integrated energy, advanced equipment manufacturing industry, changning district, shale gas production capacity reached 277 million cubic metres, built the country's first independent high-yield wells and pipelines in the first section, the lead in factory production and supply to the population. 2.1-3 GDP growth figure 2.1-4 Yibin, Yibin city, Yibin city, fiscal revenue growth 2.1.4 topography terrain overall is Southwest, North-Eastern State. Low mountains and hills in the city landscape as the main ridge-and-Valley, pingba small fragmented nature picture for "water and the second land of the seven hills". 236 meters to 2000 meters above sea level in the city, low mountain, 46.6% hills 45.3%, pingba only 8.1%. 2.1.5 development of Yibin landscapes and distinctive feature in the center of the city, with limitations, and spatial structure of typical zonal group, 2012-cities in building with an area of about 76.2km2. From city-building situation, "old town-the South Bank" Center construction is lagging behind, disintegration of the old city is slow, optimization and upgrading, quality public service resources are still heavily concentrated in the old town together. Southbank Centre has not been formed, functions of the service area space is missing. Meanwhile, peripheral group centres service was weak and inadequate accounting for city development, "suspicious" pattern could not be formed. As regards transport, with the outward expansion of cities, cities have been expanding, centripetal city traffic organization has not changed, 比，血清药研究仍有很大的优越性 。 参考文献 1(谈智、王庭槐、杨丹等，17β-雌二醇抑制内皮素诱导的血管平滑肌细胞增殖 作用。中国应用生理学杂志，2001，17(3):74,77。 2(钱振维，活血化瘀与冠心病，全国胸痹病(冠心病)学术研讨会论文集.1992.17. 3(陈国姿、徐虹，中药人工周期法治疗卵巢功能早衰继发性闭经，广东医学， 1999， 4(高笑等，中药对血管内皮细胞的保护作用的研究概况，天津药学 2003年4 月，14卷2期 5. 刘成海. 中药复方体外药理研究思考,中药新药与临床药理，2000，21(1): 53,56 6. 李仪奎.中药药血清理学实验方法的若干问题,J,.中药新药与临床药理， 1999，10(2):95. 7(贺玉琢.日本汉方药“药血清理学”、“药血清化学”的研究概况,J,.国外医 学*中医中药分册，1998，20(5):3. 8(Alkihiko S, kimio M, Yasuhi I, et al. Effect of 17β-estrodiol and progresterone on growth-factor induced proliferation and migration in human female aortic smooth muscle cells in vitro. Cardiovasc Research. 1996,32:516 5282。 9(Ross R.,et al. Cell biology of atherosclerosis.Annu.Rev.physiol. 1995,57:791-804. integrated energy, chemicals and textile Yibin city, are the three core pillars of the industry. In 2014, the wuliangye brand value to 73.58 billion yuan, the city's liquor industry slip to stabilise. Promoting deep development of integrated energy, advanced equipment manufacturing industry, changning district, shale gas production capacity reached 277 million cubic metres, built the country's first independent high-yield wells and pipelines in the first section, the lead in factory production and supply to the population. 2.1-3 GDP growth figure 2.1-4 Yibin, Yibin city, Yibin city, fiscal revenue growth 2.1.4 topography terrain overall is Southwest, North-Eastern State. Low mountains and hills in the city landscape as the main ridge-and-Valley, pingba small fragmented nature picture for "water and the second land of the seven hills". 236 meters to 2000 meters above sea level in the city, low mountain, 46.6% hills 45.3%, pingba only 8.1%. 2.1.5 development of Yibin landscapes and distinctive feature in the center of the city, with limitations, and spatial structure of typical zonal group, 2012-cities in building with an area of about 76.2km2. From city-building situation, "old town-the South Bank" Center construction is lagging behind, disintegration of the old city is slow, optimization and upgrading, quality public service resources are still heavily concentrated in the old town together. Southbank Centre has not been formed, functions of the service area space is missing. Meanwhile, peripheral group centres service was weak and inadequate accounting for city development, "suspicious" pattern could not be formed. As regards transport, with the outward expansion of cities, cities have been expanding, centripetal city traffic organization has not changed, integrated energy, chemicals and textile Yibin city, are the three core pillars of the industry. In 2014, the wuliangye brand value to 73.58 billion yuan, the city's liquor industry slip to stabilise. Promoting deep development of integrated energy, advanced equipment manufacturing industry, changning district, shale gas production capacity reached 277 million cubic metres, built the country's first independent high-yield wells and pipelines in the first section, the lead in factory production and supply to the population. 2.1-3 GDP growth figure 2.1-4 Yibin, Yibin city, Yibin city, fiscal revenue growth 2.1.4 topography terrain overall is Southwest, North-Eastern State. Low mountains and hills in the city landscape as the main ridge-and-Valley, pingba small fragmented nature picture for "water and the second land of the seven hills". 236 meters to 2000 meters above sea level in the city, low mountain, 46.6% hills 45.3%, pingba only 8.1%. 2.1.5 development of Yibin landscapes and distinctive feature in the center of the city, with limitations, and spatial structure of typical zonal group, 2012-cities in building with an area of about 76.2km2. From city-building situation, "old town-the South Bank" Center construction is lagging behind, disintegration of the old city is slow, optimization and upgrading, quality public service resources are still heavily concentrated in the old town together. Southbank Centre has not been formed, functions of the service area space is missing. Meanwhile, peripheral group centres service was weak and inadequate accounting for city development, "suspicious" pattern could not be formed. As regards transport, with the outward expansion of cities, cities have been expanding, centripetal city traffic organization has not changed,