【作者】 张馨；

【导师】 陆付耳；

【作者基本信息】 华中科技大学 ， 中西医结合临床， 2009， 博士

【摘要】 糖尿病一词描述的是一种多病因的代谢性疾病，特点是慢性高血糖，伴随因胰岛素分泌和／或作用缺陷引起的糖、脂肪和蛋白质代谢紊乱。2型糖尿病患者占糖尿病病人总数的90％左右，其发病率随着社会经济的发展和生活方式的变化呈逐年上升趋势，成为仅次于心脑血管病和肿瘤的严重威胁人类健康的第三大非传染性疾病。据国际糖尿病联盟(International Diabetes Federation，IDF)统计，20世纪90年代全球糖尿病患者约为1.00亿人，到了2007年该数字已经迅速增长到2.46亿，预计到2025年全球将有3.80亿人受到糖尿病的困扰。目前，我国已经成为仅次于印度的糖尿病第二大国，患者将近4 000万人。人口老龄化进程的加速和居民生活方式的改变使得我国的糖尿病患病率、致残率和死亡率呈现明显的上升趋势。2型糖尿病是一种异质性疾病，其发病中心环节是胰岛素抵抗(IR)和胰岛β细胞功能衰竭。lR是2型糖尿病的主要特征并贯穿2型糖尿病发生、发展的始终，但仅有IR不足以导致2型糖尿病。纵向研究表明，显著高血糖的发生与胰岛β细胞分泌功能下降有关，β细胞功能缺陷是2型糖尿病发病的必要条件，胰岛β细胞衰竭是其发展的结果，也是2型糖尿病病情加重的驱动力量。早期降低血糖浓度，可逆转胰岛β细胞功能减退，使胰岛素分泌恢复正常。因此，加强2型糖尿病发病机理的研究，针对其病因，寻找既能保护胰岛β细胞功能，又能改善IR的药物，是当前糖尿病治疗领域研究的热点课题。桑树是一种优良的药用植物。桑叶、桑椹、桑枝和桑白皮都是传统的中药材，现代药理学研究表明上述药材均具有降血糖、调血脂、抗炎、抗氧化等作用，特别是在对糖尿病及其并发症的预防和治疗方面具有药性平和、作用广泛和毒副作用小等特点。桑精胶囊系导师的经验方，由桑叶、桑椹、桑枝和桑白皮等药物组成，临床上应用其治疗2型糖尿病疗效显著。本研究旨在探讨桑精胶囊及其拆方的抗2型糖尿病的效应及作用机制。首先，本研究采用尾静脉注射小剂量STZ溶液辅以高脂高热卡饲料喂养的方法复制出具有肥胖以及葡萄糖和脂质代谢紊乱等特征，与人类2型糖尿病相类似的动物模型。在此基础上，研究了桑精胶囊及其拆方对该模型动物糖脂代谢的影响，并从抗炎、抗氧化和改善胰岛素信号转导等方面探究其可能的作用机制。本研究发现，桑精胶囊能有效地降低血糖、提高胰岛素敏感性和胰岛β细胞基础状态下的分泌功能、纠正血脂紊乱、改善IR。桑精胶囊的抗糖尿病效应可能与其有效地提高机体抗氧化能力，保护胰岛形态及功能，调节脂肪和骨骼肌组织中胰岛素信号转导通路中的相关蛋白含量及其活性而改善胰岛素信号转导有关。为进一步明确桑精胶囊复方中治疗2型糖尿病的主要作用药物，阐明其药物配伍的合理性，本实验对桑精胶囊进行了拆方研究，观察比较桑叶、桑椹、桑枝和桑白皮抗大鼠2型糖尿病的效应，探讨其可能的作用机制。结果表明，上述诸药都能降血糖、调血脂，其作用机制可能与平衡血清促炎细胞因子和抗炎细胞因子水平、提高抗氧化酶活性，改善炎症，从而保护胰腺组织形态和功能有关，但各味药作用机制不尽相同，各有侧重，以桑叶的作用较为明显、全面，从一个侧面说明了桑精胶囊复方配伍的合理性和科学性。第一部分桑精胶囊对小剂量STZ加高脂高热卡饲料诱导2型糖尿病大鼠糖脂代谢影响的研究【目的】建立小剂量STZ加高脂高热卡饲料诱导的2型糖尿病大鼠模型，观察桑精胶囊对2型糖尿病大鼠葡萄糖和脂质代谢的影响，以明确该方治疗2型糖尿病的效应。【方法】采用尾静脉注射小剂量链脲佐菌素(Streptozotocin，STZ，25mg／kg)加高脂高热卡饲料喂养的方法建立2型糖尿病大鼠模型，将成模动物随机分为模型组、二甲双胍组和桑精胶囊组，另设正常对照组。药物干预治疗8周后，各组动物进行口服糖耐量试验(Oral glucose tolerance test，OGTT)并检测各组动物体重变化、空腹血糖(Fasting bloodglucose，FBG)、空腹胰岛素(Fasting insulin，FINS)、甘油三酯(Triglyceride，TG)、总胆固醇(Total Cholesterol，TC)、高密度脂蛋白胆固醇(High Density lipoprotein cholesterol，HDL-C)、低密度脂蛋白胆固醇(Low Density Lipoprotein Cholesterol，LDL-C)、载脂蛋白AⅠ(Apolipoprotein AⅠ，ApoAⅠ)、载脂蛋白B(Apolipoprotein B，ApoB)和游离脂肪酸(Free Fatty Acid，FFA)水平。【结果】尾静脉注射小剂量STZ溶液辅以高脂高热卡饲料喂养的方法可成功复制出2型糖尿病动物模型，实验动物具有肥胖以及葡萄糖和脂质代谢紊乱等特征。桑精胶囊可显著提高实验动物胰岛素敏感性和胰岛β细胞在基础状态下的分泌功能，降低血清TC、TG、LDL-C、Apo B和FFA水平，同时提高HDL-C和ApoAⅠ水平，并显示出明显的减肥作用。【结论】桑精胶囊对2型糖尿病模型动物具有较好的减肥、降糖、调脂作用。第二部分桑精胶囊抗小剂量STZ加高脂高热卡饲料诱导大鼠2型糖尿病的研究【目的】建立小剂量STZ加高脂高热卡饲料诱导的2型糖尿病大鼠模型，观察桑精胶囊抗模型大鼠2型糖尿病的效应，探讨该方可能的作用机制。【方法】以尾静脉注射小剂量STZ加高脂高热卡饲料喂养的方法建立2型糖尿病大鼠模型，将成模动物随机分为模型组、二甲双胍组和桑精胶囊组，另设正常对照组。药物干预治疗8周后，检测各组犬鼠糖代谢相关指标、血清抗氧化酶活性指标、胰腺组织Bax、Bcl-2蛋白表达和靶组织中胰岛素信号转导通路关键蛋白表达。【结果】桑精胶囊可明显降低2型糖尿病模型动物的空腹血糖和胰岛素水平，降低血清丙二醛(Malonaldehyde，MDA)、一氧化氮(Nitric Oxide，NO)水平，提高超氧化物歧化酶(Superoxide Dismutase，SOD)、谷胱甘肽过氧化物酶(Glutathione Peroxidase，GSH-Px)活性；保护实验动物胰岛形态，降低胰岛组织Bax蛋白表达，增加Bcl-2蛋白表达从而保护胰岛：抑制实验动物靶组织胰岛素受体底物-1(Insulin ReceptorSubstrate-1，IRS-1)Ser307磷酸化，提高IRS-1和磷脂酰肌醇-3激酶(Phosphatidylinositol-3-kinase，PI-3K)蛋白表达，促进葡萄糖转运蛋白4(GlucoseTransporter 4，GLUT4)转位至细胞膜上，增加细胞膜GLUT4含量，改善糖尿病大鼠靶组织胰岛素信号转导，提高外周组织对葡萄糖的利用率，从而改善胰岛素抵抗。【结论】桑精胶囊对2型糖尿病模型动物具有良好的治疗效应，其作用机制可能与其保护胰岛功能、增强机体抗氧化能力以及改善胰岛素信号转导有关。第三部分桑精胶囊拆方抗小剂量STZ加高脂高热卡饲料诱导大鼠2型糖尿病的研究【目的】建立小剂量STZ加高脂高热卡饲料诱导的2型糖尿病大鼠模型，研究桑精胶囊拆方抗模型大鼠2型糖尿病效应的差异性及其可能的作用机制。【方法】以尾静脉注射小剂量STZ加高脂高热卡饲料喂养的方法建立2型糖尿病大鼠模型，将造模动物随机分为模型组、二甲双胍组、桑叶组、桑椹组、桑枝组和桑白皮组，另设正常对照组。药物干预治疗8周后，比较各组药物对糖尿病大鼠体重、葡萄糖和脂质代谢参数、炎症相关细胞因子、氧化应激相关指标以及胰腺组织Bax、Bcl-2蛋白表达的影响。【结果】桑叶、桑椹、桑枝和桑白皮均可明显改善实验动物葡萄糖和脂质代谢紊乱，提高实验动物胰岛素敏感性和胰岛β细胞在基础状念下的分泌功能，以桑叶降糖、降TG作用最明显，桑枝降低FFA作用突出。上述诸药均可提高2型糖尿病大鼠血清抗炎细胞因子水平、降低促炎细胞因子水平，以桑枝和桑白皮作用较为突出。在提高2型糖尿病大鼠抗氧化能力方面，桑叶和桑椹作用最佳。桑叶具有较好保护2型糖尿病大鼠胰岛形态和功能、抑制胰岛β细胞凋亡的作用。【结论】桑叶、桑椹、桑枝和桑白皮都能降血糖、调血脂，其作用机制可能与平衡实验动物血清促炎细胞因子和抗炎细胞因子水平、提高抗氧化酶活性，改善其炎症，从而保护胰腺组织形态和功能，延缓β细胞功能衰退有关。但各味药作用机制不尽相同，各有侧重，以上结果从一个侧面说明桑精胶囊复方配伍的合理性和科学性。更多还原

【Abstract】 The term Diabetes Mellitus describes a metabolic disorder of multiple aetiologiescharacterized by chronic hyperglycemia with disturbances of carbohydrate, fat and proteinmetabolism, resulting from defects in insulin secretion, insulin action, or both. At present,the prevalence of diabetes mellitus is increasing, which is a serious disease damaginghuman being’s health. Type 2 diabetes mellitus (T2DM) is the most prevalent form ofdiabetes accounting for about 90% of all diabetes cases. And its incidence rate with thesocio-economic development and lifestyle changes was an upward trend year after year,becoming second only to cardio-cerebral vascular diseases and tumors of a serious threat tohuman health, the third largest non-communicable diseases. According to the InternationalDiabetes Federation (IDF)’s statistics, there were 100 milli(?)n diabetic patients about theworld in the 20th century, however, the case number has rapidly increased to 246 million by2007, is expected 380 million diabetic people will be plagued by diabetes in the world bythe year 2025. At present, China has become second largest country of diabetes second onlyto India, patients with nearly 40 million. With the aging of population and the changes inlifestyle, our country’s prevalence, disability and mortality of diabetes showed a clearupward trend.Type 2 diabetes is a heterogeneous disease, its development requires the presence of two fundamental defects: insulin resistance (IR) and pancreaticβ-cell failure. Insulinresistance, characterized by reduced responsiveness to the normal circulating insulin intarget tissues, is now considered as a fundamental aspect of etiology of type 2 diabetesmellitus, only IR alone is not sufficient to lead to T2DM. Longitudinal studies have shownthat significant hyperglycemia with pancreaticβ-cell function decline in the secretion,β-cell dysfunction in T2DM is a necessary condition. Pancreaticβ-cell failure is the resultof the development of T2DM as well as the driving force in T2DM condition worsened.Exploring the molecular mechanism of insulin resistance and searching for drugs thatprotectedβcells and improved insulin resistance has appeared to be a focus in globalmedical research.At present, many scientists focus on investigating the mechanism of traditional Chinesemedicinal herbs and its components in treating diabetes, and expect it can bring a thought indeveloping new drugs with more safe and effective for treating diabetes. Mulberry,Mulberry Leaf, Ramulus Mori and Mori Cortex are commonly used Chinese medicinalherbs. Modern pharmacological research showed that they can be used to treat type 2diabetes mellitus and have many advantages especially in diabetes and its complicationssuch as mild, broad and durable role and few side effects features and so on. SangjingCapsule is constituted of Mulberry, Mulberry Leaf, Ramulus Mori and Mori Cortex whichaccording to the clinical experience of Prof. Lu Fu-er. This research aimed to exploreantidiabetic effect of Sangjing Capsule on animals with type 2 diabetes mellitus and itspossible mechanism.The rats with T2DM were induced by the intravenous injection of low-dosestreptozotocin (STZ, 25mg/kg) and high sucrose-fat diet. The development of this animalmodel was similar to that of human suffered from type 2 diabetes mellitus. In this animalmodel, the low dose streptozotocin injection could damage pancreaticβ-cells slightly andlead to glucose intolerance and serum insulin reduction. On this basis, a high sucrose-fatdiet was followed to induce obesity and led to dyslipidemia, insulin resistance, which elevates serum insulin level in feedback.These experimental results showed that, Sangjing Capsule could decrease bloodglucose, correct impaired glucose tolerance, improve dyslipidemia, and ameliorate insulinresistance of type 2 diabetes mellitus. Furthermore, it is suggested by the present resultsthat the therapeutic effects of Sangjing Capsule on T2DM might be related to its ability toscavenge free radical, restore pancreatic islets viability and increase IRS-1, PI-3K andGLUT4 protein expression and-improve insulin signal transduction in adipose and skeletalmuscle tissues of T2DM rats.To investigate which herb plays the major role in Sangjing Capsule, Mulberry,Mulberry Leaf, Ramulus Mori and Mori Cortex were chosen as study objects. The resultsshowed that all of these herbs not only have hypoglycemic property, but also modulate theblood lipids. Its mechanism may be related to the balance of serum pro-inflammatorycytokines and anti-inflammatory cytokines levels and raise the activity of antioxidantenzymes, thereby protecting pancreatic tissue morphology and functions. But itsmechanism is different, each with its own characteristic. The results prove the reasonabilityof Sangjing Capsule in composition manner from this regard. SectionⅠAmeliorative Effect of Sang]ing Capsule on blood glucoseand lipid metabolism in T2DM rats induced by low-dose STZ andhigh sucrose-fat diet【OBJECTIVE】To investigate the effects of Sangjing Capsule on blood glucose and lipid metabolismof experimental T2DM rats.【METHODS】The rats with T2DM were induced by the intravenous injection of low-dosestreptozotocin (STZ, 25mg/kg) and high sucrose-fat diet. Then modeled diabetic animalswere divided into model, metformin and Sangjing Capsule group. Normal rats fed withcommon chow were designated as normal group. Eight weeks later, the oral glucosetolerance test (OGTT) was performed in all animals, the changes of murine body weight,fasting blood glucose (FBG), fasting insulin (FINS), Triglyceride (TG), Total Cholesterol(TC), High Density Lipoprotein Cholesterol (HDL-C), Low Density LipoproteinCholesterol(LDL-C), Apolipoprotein A I (ApoA I), Apolipoprotein B(ApoB), Free FattyAcid (FFA) were routinely determined.【RESULTS】Compared with the model group, the result of OGTT in Sangjing Capsule group wasimproved. The levels of the body weights and FBG, FINS, TG, TC, LDL-C, ApoB and FFAdecreased, while HDL-C and ApoA I protein increased obviously.【CONCLUSION】Sangjing Capsule not only has hypoglycemic property, but also modulates the plasmalipids. SectionⅡEffect of Sangjing Capsule on T2DM rats induced bylow-dose STZ and high sucrose-fat diet and its possible mechanism【OBJECTIVE】To investigate the antidiabetoc effect of Sangjing Capsule on the rats with T2DMinduced by low-dose STZ and high sucrose-fat diet, and to explore its possible mechanism.【METHODS】The rats with T2DM were induced by the intravenous injection of low-dose ofstreptozotocin (STZ, 25mg/kg) and high sucrose-fat diet. Then modeled diabetic animalswere divided into model, metformin and Sangjing Capsule group. Normal rats fed withcommon chow were designated as normal group. Eight weeks later, the effects of SangjingCapsule on glucose metabolic parameters, factors related to oxidative stress, anti-injuryfactors of pancreatic islets and some key proteins in insulin signal transduction pathwaywere observed.【RESULTS】Intragastric administration of Sangjing Capsule significantly decreased FBG and FINSlevels in diabetic rats. Furthermore, Sangjing Capsule treatment significantly blocked theincrease of MDA and NO levels, increased SOD and GSH-px levels in diabetic rats.Histopathological scores showed that Sangjing Capsule had restored the damage ofpancreas tissues in rats with T2DM. Decreased IRS-1, PI 3-kinase p85 and GLUT4 proteinexpression in diabetes were significantly improved by Sangjing Capsule treatment. Theincreased serine phosphorylation level of IRS-1 (pSer307) was significantlydown-regulated in adipose and skeletal muscle tissues of T2DM rats treated with SangjingCapsule.【CONCLUSION】It is suggested that Sangjing Capsule could significantly inhibit the progression ofdiabetes induced by low-dose STZ and high sucrose-fat diet, and its inhibitory effect ondiabetes might be associated with its hypoglycemic effec and its ability to scavenge freeradical, restore pancreatic islets viability and enhance insulin signaling. SectionⅢComparison of Effects of Sangjing Capsule’s SeparatedRecipes on T2DM Rats Induced by Low-dose STZ and HighSucrose-fat Diet and its Possible Mechanism【OBJECTIVE】To investigate the antidiabetic effect of Sangjing Capsule’s separated recipes on therats with T2DM induced by low-dose STZ and high sucrose-fat diet, and to explore itspossible mechanism.【METHODS】The rats with T2DM were induced by the intravenous injection of low-dose ofstreptozotocin (STZ, 25mg/kg) and high sucrose-fat diet. Then modeled diabetic animalswere divided into into model, metformin, Mulberry leaf, Mulberry, Ramulus Mori and MoriCortex group. Normal rats fed with common chow were designated as normal group. Eightweeks later, the changes of rat body weight, and the effects of Mulberry, Mulberry leaf,Ramulus Mori and Mori Cortex on glucose and lipid metabolic parameters, factors relatedto inflammation and oxidative stress and anti-injury factors of pancreatic islets wereobserved.【RESULTS】Compared with the model group, the result of OGTT in every herbs group wasimproved. The levels of the body weights and FBG, FINS, TG, TC, LDL-C, ApoB and FFAdecreased, while HDL-C and ApoA I protein increased obviously also. Furthermore,Sangjing Capsule treatment significantly blocked the increase of CRP, TNF-α, IL-1β, IL-6,ICAM-1, VCAM-1, MDA and NO levels, increased IL-4, IL-10, SOD and GSH-px levels in diabetic rats. Histopathological scores showed that Sangjing Capsule had restored thefunction of pancreas tissues in rats with T2DM.【CONCLUSION】The results showed that all of these herbs not only have hypoglycemic property, butalso modulate the blood lipids. Its mechanism may be related to the balance of serumpro-inflammatory cytokines and anti-inflammatory cytokines levels and raise the activity ofantioxidant enzymes, thereby protecting pancreatic tissue morphology and functions. Butits mechanism is different, each with its own figures.更多还原