【作者】 陈长青；

【导师】 赖小平；

【作者基本信息】 广州中医药大学 ， 中药学， 2008， 博士

【摘要】 糖尿病(diabetes mellitus)是由于遗传和环境因素相互作用,导致胰岛素绝对或相对分泌不足以及靶组织细胞对胰岛素敏感性降低,引起蛋白质、脂肪、水和电解质等一系列代谢紊乱综合征。糖尿病患者长期血糖升高可致器官组织损害,引起脏器功能障碍以致功能衰竭。由于糖尿病的严重危害性及发病率的逐年增高,再加上治疗需要高额的费用,使得预治糖尿病已成为世界各国面临的严峻挑战;也使抗糖尿病药物具有巨大的市场前景,抗糖尿病新药的研发亦成为全球热点之一。由于1型糖尿病发病率较低,不到5%,而且主要依赖胰岛素治疗,所以本研究主要针对2型糖尿病。对2型糖尿病的认识半个多世纪以来一直局限在胰岛素分泌不足上,对其治疗自然也就主要集中在促进胰岛细胞对胰岛素的分泌上;药物的研发亦主要集中在胰岛素促泌剂上,导致治疗手段长期相对匮乏。胰岛素抵抗(Insulin resistance,IR)是胰岛素的外周靶组织对胰岛素的敏感性与反应性降低,从而导致胰岛素难以产生正常的生理效应。研究表明,IR是2型糖尿病发病的关键驱动因素,也是一系列代谢异常,并导致心血管等并发症的核心,是产生糖尿病、冠心病、高血压等疾病的“共同土壤”。因此,针对IR的治疗是治疗2型糖尿病的关键。由于对2型糖尿病发病机理的认识发生了根本性的转变,从对胰岛细胞本身的分泌功能的重视,转移到对胰岛素抵抗和胰外作用的重视上来,导致药物的研发速度不断加快,通过改善胰岛素抵抗来治疗2型糖尿病的新药便不断问世。糖尿病属中医的“消渴病”。中医对消渴病的病机认识,自古迄今大都认为是阴津亏损,燥热内盛;医生按照肺、胃、肾三消论治。但自当代施今墨、祝谌予氏创立气阴两虚和瘀血阻络病机学说以来,已被大多数临床医生所接受,极大地推动了对消渴病病机的研究。随后痰、湿、毒致病等各种病机学说被不断地提出,极大地丰富了对消渴病发病机理的认识,使得临床疗效不断提高。由于对消渴病病因病机认识的不断深化,使得近十余年来抗糖尿病中药新药品种从无到有、增长迅猛。据季绍良报道,1985年～1992年未见有申报批准治疗糖尿病的中药新药;1992年～1999年4月,共批准14个;而1999年～2002年已批准临床研究的达到约30个。但是,目前临床上抗糖尿病药物仍然是以西药为主,中药仍处于辅助地位。究其原因:一是中药新药研发依然沿袭西药新药研发的思路,主要精力都集中在有效成分提纯和药理作用研究上;二是新药处方设计缺乏中医病因病机理论和中药配伍理论的指导,药物筛选局限在古方、验方上;三是新药研发者与临床联系不紧密,研发思路落后于临床实践,对糖尿病的认识仍然局限在阴虚燥热、气阴两虚的病机上,一些病机新理论如:瘀血、痰湿、热毒等,则还没有进入抗糖尿病新药研发者的视野。因此,充分发挥中医药理论的指导作用,针对瘀血、痰湿和热毒等病机新理论,有目标地扩大药物筛选范围,研发疗效确切的抗糖尿病中药新药就是我们的当务之急。现代中医病机理论认为,气阴两虚——阴虚燥热——阴阳两虚是2型糖尿病发生、发展的基本规律,而“瘀血阻络”则贯穿2型糖尿病的始终。气为血帅,气虚致血行无力;而“阴虚必血滞”,血流缓慢易凝滞血瘀。临床研究表明,2型糖尿病血瘀证与胰岛素抵抗密切相关。临床研究还表明,活血化瘀法对降低血糖和防治并发症均具有显著疗效。本研究在胰岛素抵抗——“共同土壤说”和瘀血阻络——“共同病机说”的理论启发下,选择复方丹参片类制剂——即具有活血化瘀功效的、常用于治疗冠心病的小复方制剂——作为切入点,针对糖尿病胰岛素抵抗和瘀血阻络这个关键环节,首先对其进行抗糖尿病作用的临床和实验验证,探讨不同类的活血化瘀药物降血糖的可能性,以期能够有目标地拓展抗糖尿病有效部位的筛选范围;然后对其进行拆方研究,筛选有效部位,观察其降低血糖和防治糖尿病慢性并发症的效果,并探讨其作用机制,为下一步进行有效部位重新组方打下基础;并探索一条抗糖尿病中药新药筛选的新途径。具体研究内容及结果如下:西医学认为,胰岛素抵抗是2型糖尿病发生发展主要因素,2型糖尿病和冠心病有共同的发病土壤——胰岛素抵抗;中医学认为,瘀血既是糖尿病的病理产物,又是糖尿病发生发展的动因,2型糖尿病和冠心病都有共同的的发病机理——瘀血阻络。据此,我们推测凡是具有活血化淤作用的能够治疗冠心病的中药制剂都有可能用于治疗胰岛素抵抗的2型糖尿病。首先,我们对活血化瘀法治疗冠心病的代表性中成药——复方丹参片和复方丹参注射液,进行2型糖尿病的临床疗效对照研究,结果表明:复方丹参片和复方丹参注射液均具有一定的降低血糖、增加胰岛素敏感性、改善临床症状的作用;与对照组比较,在消除临床症状、提高胰岛素敏感性方面更胜一筹。其次,我们根据复方丹参片类制剂(即具有活血化瘀功效的、常用于治疗冠心病的小复方制剂)药味少、活性部位相对明确、研究较为简便的特点,选择了复方丹参滴丸、速效救心丸、地奥心血康胶囊以及复方丹参片与罗格列酮作比较,进行了降血糖的实验验证,发现它们都显示出一定的降血糖效果,同时,都能够提高胰岛素敏感指数;其中复方丹参片效果最好,这为我们下一步的筛选指明了方向。本实验结果充分证明,在中西医理论的指导下,我们可以缩小新药研发的筛选范围,使无目标的撒网式筛选变成有目标的猎取式筛选,取得事半功倍的效果。第三,我们根据复方丹参片中主要成分性质不同,将其拆分成水溶性部位和脂溶性部位,并进行抗糖尿病的实验研究,发现复方丹参片水溶性部位、脂溶性部位均具有降低血糖、改善胰岛素抵抗的作用,但复方丹参片水溶性部位作用相对较强。第四、我们将复方丹参片水溶性部位进行进一步拆分和重新组万,并进行抗糖尿病的实验研究,结果显示:除三七皂苷组之外,丹酚酸组、丹酚酸+三七皂苷组、丹酚酸+三七皂苷+冰片组血糖均显著下降。其中,丹酚酸+三七皂苷+冰片的血糖下降幅度最大。我们还发现了丹酚酸类这个具有显著降血糖作用的新部位。第五、我们对复方丹参片有效部位拆方进行了抗糖尿病作用机理探讨,发现它们能够抑制超氧化物MDA,修复胰腺组织;降低血浆FFA、TNF—α,增强胰岛素受体的敏感性;提高ISR-1、IRS-2和PI3K的表达水平,减轻胰岛素受体后的抵抗;并且降低TG、改善脂肪肝,减少肝糖原的输出;通过以上多环节、多途径、多靶点的作用来达到控制血糖、治疗2型糖尿病的目的。还发现了丹酚酸+三七皂苷+冰片组这一新组方在改善胰岛素抵抗、减轻胰岛细胞损伤、改善脂代谢、抑制脂肪肝等多方面都有明显作用和独特优势,值得进一步的开发研究。第六,我们的研究还发现,复方丹参片可能通过减少糖尿病大鼠糖基化产物、降低血脂、抑制组织过氧化产物的产生及组织增生来改善2型糖尿病大鼠慢性并发症。病理组织学检查显示,复方丹参片抑制模型大鼠眼球小血管血栓形成的作用极为显著,可改善眼球血液循环;能减轻模型大鼠动脉硬化或病变的程度;减轻模型动物坐骨神经损伤程度;能抑制模型动物肾脏系膜内浆液浸润、肾小管基膜增厚,减轻肾脏病变程度。本研究的创新之处主要有三点:一是尝试运用中医瘀血阻络的糖尿病病机理论指导,并成功地将复方丹参片类制剂移植到2型糖尿病的治疗上,发现了复方丹参类制剂——复方丹参滴丸、速效救心丸、地奥心血康胶囊都有一定的降血糖作用,有效地缩小了抗糖尿病中药的筛选范围,初步验证了中医瘀血阻络病机理论对抗糖尿病中药筛选的指导意义。二是发现了丹参中的丹酚酸类部位具有明显的降低2型糖尿病模型大鼠血糖的作用。丹酚酸类的降血糖功效在以往的文献中尚未见报道,这是一个令人鼓舞的新发现,说明如果有理论的指导,筛选将更有针对性,筛选的效率会更高。三是我们还发现丹酚酸+三七皂苷+冰片组这一新组方通过改善胰岛素抵抗、减轻胰岛细胞损伤、改善脂代谢、抑制脂肪肝等多个方面作用,达到抗糖尿病的目的,值得进一步的开发研究。总之,本实验研究对运用中医病机理论指导抗糖尿病药物筛选进行了有益尝试,发现了一些好的苗头,值得我们进行深入探讨,不断总结经验,以期建立一种新的中药新药筛选与研发模式。更多还原

【Abstract】 Diabetes is a series of metabolic disorder syndrome in protein, fat, water and electrolytes, which due to the interaction of genetic and environmental factors, and causes absolutely or relatively to shortage of insulin secretion and sensitivity decrease of target cells to insulin. Diabetic patients with long-term elevated blood sugar can cause organ damage, which leads to function failure due to the viscera dysfunction. Because of the serious harmfulness of diabetes and the incidence rate increases year by year, coupling with the need for the high cost, which has become the world to face a serious challenge to prevent and therapy diabetes. Likewise, anti-diabetes drugs also have great market prospects, which makes anti-diabetes drugs researched and developed to become one of the focus in the world.As the incidence of type 1 diabetes is low, less than 5 percent, and mainly relies on insulin therapy, this study is for type 2 diabetes primarily. The knowledge about type 2 diabetes has been confined to the lack of insulin secretion more than half a century, which causes the treatment for it has been focused on promoting insulin secretion agent naturally. Drugs research and development has been focused on promoting insulin secretion agent, leading to the relative lack of long-term treatment.Insulin resistance (Insulin resistance, IR) is the peripheral insulin target tissues to insulin sensitivity and responsiveness decreases, leading to it is difficult for insulin to produce the normal physiological effects. Research shows that IR is the key driving factor leading to the incidence of type 2 diabetes. It is also a range of metabolic disorders, leading to cardiovascular complications such as the core. It is the "common soil" where diabetes, coronary heart disease and hypertension happen. Therefore, the treatment for IR is the key of treating type 2 diabetes.Since the awareness about the pathogenesis of type 2 diabetes have changed fundamentally, the attention of which transits from the islet cells their own secretion to the action of IR and outside the pancreas. It leads to drug research and development continuing to accelerate, which causes that it is constantly available to improve insulin resistance to the treatment of type 2 diabetes drugs.Diabetes belongs to Chinese medicine "wasting-thirst". The awareness about the pathogenesis of "wasting-thirst" has been considered to be the deficiency of body fluid that causes dryness-heat being excessive in the interior. Doctors take the way of treatment for diabetes in accordance with the lung, stomach and kidney. However, since the deficiency of both vital energy and yin and stagnation of blood theory has been founded by Shi jinmo and zhu chenyu this contemporary, which has been accepted by the majority of clinicians. It drives enormously to have research on the pathogenesis of diabetes. Subsequently, sputum, humid, and harm as such pathogenesis theories were constantly raised, greatly enriching the awareness on the pathogenesis of diabetes, which makes therapeutic effect continue to increase. The awareness for pathogeny and pathogenesis of "wasting-thirst" has deepened constantly, which makes anti-diabetes drugs varieties of Chinese medicine from scratch to rapid growth in nearly past 10 years. According to the report by Ji shoaling, the approval of new Chinese medicine treatment of diabetes drug have not been declared from 1985 to 1992; 14 had been approved from 1992 to April 1999; while about 30 had been approved to have clinical research from 1999 to 2002. However, the current clinical anti-diabetes drug is still mainly western medicine, Chinese medicine is still in secondary status. The reasons: First, traditional Chinese medicine research and development of new drugs continues to follow the thinking of western medicine research, the main efforts are concentrated on active ingredients purification and research on the pharmacological effects; Second, the design of new Chinese medicine prescription lacks guiding of pathogeny and pathogenesis theory of Chinese medicine and Chinese medicine compatibility theories, drugs selecting confined to the ancient prescription and experimental prescription. Third, the researchers of new drugs are not closely associated with the clinic, the ideas of research and development drop behind the clinical practice. The awareness of diabetes is still confined to the pathogenesis of deficiency of yin feverish sensation and deficiency of both vital energy and yin. Some new theories of pathogenesis such as blood stasis, phlegmatic hygrosis, pyretic toxicity have not enter into the the vision of the anti-diabetes drug researchers. Therefore, it is urgent to take full advantage of the guiding of Chinese medicine theory, for the new pathogenesis theory of blood stasis, phlegmatic hygrosis, pyretic toxicity, expanding the scope of drugs selected with aim, researching and developmenting certain effect of the anti-diabetes drug Chinese medicine.The pathogenesis of modern traditional Chinese medicine theory considers that deficiency of both vital energy yin-deficiency of yin feverish sensation-simultaneous asthenia of yin and yang is the basic rule for the occurrence and development of the type 2 diabetes, while the blocking collaterals of blood stasis traverses from the beginning and the end of type 2 diabetes. Qi is the commander of blood, the deficiency of qi causing it incapable to circulate, while deficiency of yin must cause blood stasis, slow flow blood easily leads to blood stagnation. Clinical studies show that blood stasis of type 2 diabetes is closely related to insulin resistance. They also show that it is significantly effective to lower blood sugar and prevent complications for activating blood to resolve stasis treatment.This study is inspired by the theory of insulin resistance- the "common soil" and blood stasis blocks collaterals-"common pathogenesis theory", selecting compound Danshen tablet generic medicine, that is, it has the effect of activating blood to resolve stasis and the small compound preparation which is often used to therapy coronary heart disease - as a point of penetration. Facing the key link of insulin resistance and blood stasis blocks collaterals, first of all its anti-diabetic role of the clinical and experimental verification, discussing the possibility of lowering blood sugar about different types of activating blood to resolve stasis drugs, expecting to develop the selected scope of anti-diabetic effective parts with aim. Then demolition research has been done and the effective parts are selected. Observing the effect of its lowering blood sugar and prevent chronic complications of diabetes, discussing its mechanism, which lays the foundation to form medicine over again about effective parts for the next step. To explore a new way of selecting anti-diabetes Chinese medicine.Modern medicine considers that insulin resistance is the major factor which causes type 2 diabetes to start and develop. Type 2 diabetes and coronary heart disease have a common soil - insulin resistance. Traditional Chinese Medicine considers that blood stasis is not only the pathology production of diabetes, but also is the agent of the occurrence and development of diabetes. Type 2 diabetes and coronary heart disease have a common pathogenesis- blood stasis blocks collaterals. Accordingly, we speculate that any Chinese medicine formulations relieving blood stasis role can therapy coronary heart disease, which may also be used for the treatment of type 2 diabetes caused by insulin resistance.First of all, we have a control research on the clinical efficacy of type 2 diabetes about the representative Chinese medicines to therapy coronary heart disease which can relieve blood stasis-compound Danshen tablet and compound Danshen injection. The results indicate that compound Danshen tablet and compound Danshen injection can lower blood sugar, increase insulin sensitivity and improve the clinical symptoms in a way. They are superior to the control group in the elimination of clinical symptoms and improvement of insulin sensitivity.Secondly, due to these Chinese medicine are made up of a few ingredients, relatively clear active parts, so we choose Fufang danshen diwan, Suxiao jiuxin wan, Diao xinxuekang capsule and Fufang danshen tablet to compare with Rosiglitazone, and have experimental verification of lowering blood sugar, which showed that they all show some certain effect of lowering blood sugar. At the same time, they all can improve insulin sensitivity index. Among them, Fufangdanshen tablet is the best, which points out the direction t for our selecting of the next step. The results of this study fully demonstrate that under the guidance of traditional Chinese medicine, we can take the effect of getting twice the result with half the effort, by narrowing the selection scope of new drug research and development, which makes no goal-selecting of the nets become the hunt for selecting with aim.Third, due to the major components of a different nature in Fufang danshen tablet, it is split into water-soluble part and fat-soluble part, the experimental anti-diabetes research is done. It is found that both water-soluble part and fat-soluble part of Fufang danshen tablet can lower blood sugar and improve insulin resistance, but the action of water-soluble part is relatively stronger than that fat-soluble part.Fourth, water-soluble part of Fufang danshen tablet is further split into and re-group and the experimental research is conducted on the anti-diabetes. The results show that blood sugar decreases significantly in the salvianolic acid group, the salvianolic acid+ notoginsenoside group and the salvianolic acid+ notoginsenoside+ borneol group in addition to notoginsenoside group. Among them, blood sugar of the salvianolic acid+ notoginsenoside+ borneol group decreases most severely. We also find that the new section of the salvianolic acid which can lower blood sugar significantly.Fifth, the mechanism of anti-diabetes is discussed by splitting the effective parts of Fufang danshen tablet. It is found that they can depress superoxide dismutase MDA and repair pancreatic tissue. They can reduce plasma FFA, TNF-α, enhance the sensitivity of insulin receptors. They can improve the expression of ISR-1, IRS-2 and PI3K and reduce the insulin resistance. They also lower TG, improve fatty liver and reduce the output of liver glycogen. It is the aim to control blood sugar and therapy type 2 diabetes by much more links, multi-channel and multi-role target. It is also found that the new group medicine of the salvianolic acid+ notoginsenoside+ borneol group has obvious role and unique advantages in improving insulin resistance, reducing the islet cell damage and improving fat metabolism and depressing fatty liver, which is worth the further development of research.Sixth, the study also found that maybe Fufang danshen tablet improves the proliferation of type 2 diabetic rats’ chronic complications by reducing the glycosylation product, lowering blood fat and depressing organized suppression of the product produced. Pathological examination reveals that the action of depressing the model rats’ small vascular thrombosis in the eyeball is extremely significant for Fufang danshen tablet, which can improve blood circulation of the eyeball. It can reduce the extent of atherosclerosis or disease. It can alleviate the extent of sciatic nerve injury. It can depress model rats’ kidney serous membrane of the invasion and tubular-film thickness to reduce the extent of kidney disease.The main innovation of this study have four points: First, using the blood stasis blocks collaterals of diabetes pathogenesis theory of traditional Chinese medicine as guidance, which is successful in transplanted Fuangdanshen tablet to the type 2 diabetes therapy, which is effective to reduce the selecting scope of anti-diabetes Chinese medicine. The second is that the new section of the salvianolic acid which can lower blood sugar significantly is found. Third, the invention of this new set of anti-diabetes, which is the salvianolic acid+notoginsenoside+borneol group. Fourth, it was found that the Fufang Danshen category-the lowering blood sugar effect of Fufang danshen diwan, Suxiao Jiuxin pill, Diao xinxuekang capsule.In short, this study makes a useful attempt to guide the anti-diabetes Chinese medicine selecting by the pathogenesis theory of traditional Chinese medicine, some good signs are found, which is worth going deep into discussion, summing up constantly to establish a new traditional Chinese medicine drug selecting and research and development pattern.更多还原