糖尿病是一種以不正常的高血糖為主要特徵的長期性的糖代謝紊亂疾病。二型糖尿病是常見的糖尿病類型，多於九成的糖尿病病人患有此種類型。各種引起糖尿病的病因最終都會導致血糖過高，並且最終會引起有關眼睛，腎臟，神經和血管系統的併發癥。迄今，糖尿病正影響著大約世界6%的人口，而現在患病率依然在逐年增加。在香港，由於高能量的食和缺乏運動，越來越多的老年人和青年人正在遭受著糖尿病的困擾。糖尿病不是一種致命性的疾病，但是如果沒有採取好的治療控制措施，糖尿病最終會引起一些併發癥，這些併發癥最終會使糖尿病患者走向死亡。高血糖癥不僅是糖尿病的主要特徵，而且也是引起各種糖尿病併發癥的重要因素，在二型糖尿病的治療當中，根據各種病理因素，市場上已經研製出了很多西藥來治療糖尿病。然而，它們都有一些副作用的限制。因此，我們需要通過綜合治療和通過新的途徑研製新的製劑來控制血糖水平，保護病人遠離長期併發癥的困擾。如今，腎臟在血糖平衡中的重要角色已經被很好的認知。 在過去的二十年裡， 通過減少血糖在腎臟的重吸收來增加尿液中血糖的排出，從而達到降低體內血糖水平的方法已經被提出并認為是治療糖尿病的一直新的途徑。 在腎臟中，鈉葡萄糖共轉運體2（SGLT 2）主要負責葡萄糖的重吸收，因此，鈉葡萄糖共轉運體2（SGLT 2）抑製劑被認為是一種有潛質的新型的治療糖尿病的製劑。然而，市場上至今沒有成功研製這種製劑。达格列嗪（dapagliflozin），作為一種最有潛質的鈉葡萄糖共轉運體2（SGLT 2）抑製劑，依然處於臨床三期實驗。至今，對具有鈉葡萄糖共轉運體2（SGLT 2）抑製作用的天然產物和傳統中醫藥的信息報導非常少。中醫中藥的治療理念強調整體治療，從此點看來，爲了使糖尿病患者遠離長期的糖尿病併發癥的困擾，中醫中藥可能比西藥更有優勢。 / 因此，本研究的目的是尋找那些具有體外能專門抑制鈉葡萄糖共轉運體2（SGLT 2）並且體內能通過增加尿糖排出來降低血糖水平的抗糖尿天然產物或傳統中藥。從文獻分析中找到了經常用於治療糖尿病的11種中藥和兩種天然產物。 / 試管實驗確立了五味子醇提物和丹皮酚對表達了人的鈉葡萄糖共轉運體2（SGLT 2）基因的COS 7細胞鏈中鈉葡萄糖共轉運體2對¹⁴C-α-甲基- D-葡萄糖苷的吸收作用具有很強的抑制作用。 / 生物活性引導的片段分析確立了五味子醇提物中的活性片段--乙酸乙酯：甲醇（4:6）（F8）片段具有明顯的專門抑制鈉葡萄糖共轉運體2的作用。本實驗也對F8進行了高效液相色譜和液質聯用色譜分析。五味子中三種常見的化合物：五味子甲素，五味子乙素和五味子醇甲存在于F8中，但濃度都很低。試管實驗顯示，這三種常見化合物均無抑制鈉葡萄糖共轉運體2的作用。因此得出結論，這三種常見的五味子化合物不是F8中有效的抑制鈉葡萄糖共轉運體2的活性成份。 / 本實驗也利用動物實驗調查了丹皮酚的抗糖尿作用。糖尿病大鼠被餵食了三個星期的丹皮酚，基礎血糖實驗和尿糖排出實驗均無陽性結果。 / Diabetes Mellitus (DM) is a chronic disorder of glucose metabolism characterized by abnormally high blood glucose level. Type 2 DM is the common form of diabetes which accounts for more than 90% of all DM cases. All causes of diabetes ultimately lead to hyperglycemia, and it can cause the late complications involving the eyes, kidneys, nerves and blood vessels, which are harmful to health. DM is now affecting about 6% population of the world, and the prevalence is still increasing quickly year by year. In Hong Kong, more and more elderly and youth are suffering from diabetes because of lacking of exercise and high energy diet. DM is not a fatal disease, but if no good action is taken, it can finally cause some kinds of complications, which can lead the patients to the end of their lives. Hyperglycemia is the major characteristics of diabetes, and it is also an important factor which induces all kinds of diabetic complications. In the therapy of type 2 diabetes, a lot of western medicine have been developed in the market according to various pathological causes. However, they have limitations such as existence of side effects. Therefore, combination therapy and development of new agents with novel mechanisms should be required to control the glycemic level and protect the patients from the long-term complications. Nowadays, the significance of the kidney's role in glucose homeostasis is well recognized. Glucose excretion with urine by reducing the renal glucose reabsorption to attenuate the glycemic level has been considered as a new mechanism to treat diabetes since the past two decades. Inhibitors on sodium glucose co-transporters 2 (SGLT 2) which are responsible for the glucose reabsorption in kidney are considered as a kind of new agents that have a potential on the treatment of diabetes. However, there is still no such kind of drug developed in the market, since the most potential one, dapagliflozin, is still on Phase III clinical trial. So far, only few information is found on natural products/traditional Chinese medicines (TCMs) that possess SGLT inhibitory action. Regarding the protection of patients from long-term complications, Chinese medicine which consider the body as a whole, may have advantages over western drugs. / Therefore, the aim of this study is to search for anti-diabetic TCM/natural products which specifically inhibit the activity of SGLT2 in vitro and attenuate plasma glucose level in vivo via increasing glucose excretion through urination. From literature review, 11 TCMs and 2 natural products frequently used in treating DM were selected for screening. / Using hSGLT 1 and hSGLT 2-expressed COS-7 cell lines as a model, in vitro study demonstrated that Fructus Schisandrae chinensis (ethanolic extract) and paeonol posses the most potent inhibitory effect on SGLT 2 in the in vitro ¹⁴C-α-methyl-D-glucopyranoside (¹⁴C-AMG) uptake assay. / The purification of active fraction(s) in ethanolic extract of Schisandrae chinensis fructus was carried out using the bioassay-guided fractionation assay. The ethyl acetate-methanol (4:6) fraction (F8) was selected with significant specific inhibitory effect on SGLT 2. UPLC and LC/MS-MS profiles of F8 were also given in this study. The concentrations of three common compounds of Fructus Shisansrae chinensis: deoxyschisandrin, schisandrin B (γ-schisandrin) and schisandrin were shown very low concentration in F8, the results of uptake assay showed none of these three compounds have inhibitory effects on SGLT 2. It is concluded that these three common compounds in Schisandrae chinensis fructus are not the effective ingredients in F8 which can specifically inhibit SGLT 2. / The anti-diabetic effects of paeonol in treating type 2 DM was investigated in animal study. Paeonol (200 and 300 mg/mL) was given to the type 2 diabetic rat model - Zucker Diabetic Fatty (ZDF) rats for three weeks, the results showed no positive effects on the basal glycaemia test and urinary glucose excretion test. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Qu, Yue. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 141-153). / Abstracts also in Chinese. / TABLE OF CONTENTS / ABSTRACT --- p.iv / 摘要 --- p.vii / ACKNOWLEDGEMENT --- p.ix / LIST OF ABBREVIATIONS --- p.x / LIST OF TABLES --- p.xiii / LIST OF FIGURES --- p.xiv / TABLE OF CONTENTS --- p.1 / Chapter CHAPTER 1 --- INTRODUCTION --- p.8 / Chapter 1.1 --- Definition, diagnosis, classification and epidemiology of Diabetes Mellitus --- p.8 / Chapter 1.1.1 --- Definition of Diabetes Mellitus --- p.8 / Chapter 1.1.2 --- Diagnosis of Diabetes Mellitus --- p.8 / Chapter 1.1.3 --- Classification of Diabetes Mellitus --- p.9 / Chapter 1.1.4 --- Prevalence of Diabetes Mellitus --- p.11 / Chapter 1.2 --- Glucose Homeostasis and Diabetes Mellitus --- p.12 / Chapter 1.2.1 --- General Description --- p.12 / Chapter 1.2.2 --- Kidney's role in Glucose Homeostasis --- p.14 / Chapter 1.2.2.1 --- Gluconeogenesis in the Kidney --- p.15 / Chapter 1.2.2.2 --- Glucose Reabsorption in the Kidney --- p.15 / Chapter 1.2.2.3 --- Renal glucose transporters --- p.17 / Chapter 1.2.2.4 --- Disorders with abnormal renal glucose transport --- p.19 / Chapter 1.3 --- Etiology of Diabetes Mellitus --- p.20 / Chapter 1.3.1 --- Pancreatic β cell dysfunction --- p.21 / Chapter 1.3.2 --- Insulin resistance --- p.21 / Chapter 1.4 --- Diabetic complications --- p.23 / Chapter 1.5 --- Treatment of type 2 Diabetes Mellitus --- p.25 / Chapter 1.5.1 --- Conventional therapy of type 2 Diabetes Mellitus --- p.25 / Chapter 1.5.2 --- New mechanism for the treatment of type 2 Diabetes Mellitus - Inhibition of glucose reabsorption by glucose transporters in Kidney --- p.29 / Chapter 1.6 --- Traditional Chinese Medicine for Diabetes Mellitus --- p.30 / Chapter 1.7 --- Project objective --- p.33 / Chapter CHAPTER 2 --- TRADITIONAL CHINESE HERBAL MATERIALS AND NATURAL PRODUCTS --- p.36 / Chapter 2.1 --- Materials --- p.36 / Chapter 2.2 --- General description and anti-diabetic effects of selected herbs/natural products --- p.38 / Chapter 2.3 --- Extraction Method --- p.45 / Chapter CHAPTER 3 --- IN VITRO STUDIES OF THE INHIBITORY EFFECT OF SELECTED TRADITIONAL CHINESE HERBS AND NATURAL PRODUCTS ON SODIUM GLUCOSE COTRANSPORTERS (SGLT) --- p.48 / Chapter 3.1 --- Introduction --- p.48 / Chapter 3.2 --- Materials --- p.49 / Chapter 3.3 --- Methods and Methods --- p.52 / Chapter 3.3.1 --- In vitro model for screening of SGLT inhibitor --- p.52 / Chapter 3.3.1.1 --- Preparation of hSGLT1 and hSGLT2 Plasmid --- p.52 / Chapter 3.3.1.2 --- Transient Transfection of SGLT1 or SGLT2 clone --- p.53 / Chapter 3.3.1.3 --- Detection of mRNA expression level by Reverse Transcription-Polymerase Chain Reaction (RT-PCR) --- p.54 / Chapter 3.3.1.4 --- Development of SGLT1 or SGLT2 stable cell lines --- p.56 / Chapter 3.3.1.5 --- Results --- p.56 / Chapter 3.3.2 --- Cell proliferation assay (MTT assay) --- p.57 / Chapter 3.3.2.1 --- Methods --- p.57 / Chapter 3.3.2.2 --- Results --- p.58 / Chapter 3.3.3 --- Uptake Assay of ¹⁴C-α-methyl-D-glucopyranoside (¹⁴C-AMG) in cultured COS-7 cells expressing SGLT1 or SGLT2 --- p.63 / Chapter 3.3.3.1 --- Methods --- p.63 / Chapter 3.3.3.2 --- Screening Results of Effective Chinese Herbs/Natural Products --- p.64 / Chapter 3.4 --- Discussion --- p.83 / Chapter CHAPTER 4 --- FRACTIONATION OF SCHISANDRAE CHINENSIS FRUCTUS --- p.86 / Chapter 4.1 --- Introduction --- p.86 / Chapter 4.2 --- Organic Extraction of Schisandrae Chinensis Fructus --- p.86 / Chapter 4.2.1 --- Material and Methods --- p.86 / Chapter 4.2.2 --- Result --- p.86 / Chapter 4.3 --- Bioassay-guided Fractionation of Ethanolic Extract of Schisandrae Chinensis Fructus --- p.87 / Chapter 4.3.1 --- Materials --- p.87 / Chapter 4.3.2 --- Methods --- p.87 / Chapter 4.3.2 --- Results --- p.89 / Chapter 4.4 --- ¹⁴C-α-methyl-D-glucopyranoside (¹⁴C-AMG) Uptake Assay of fractions in cultured COS-7 cells expressing SGLT1 or SGLT2 --- p.92 / Chapter 4.4.1 --- Methods --- p.92 / Chapter 4.4.2 --- Results --- p.93 / Chapter 4.5 --- Characterization of F8 of Schisandrae chinensis fructus using Ultra Performance Liquid Chromatography (UPLC) --- p.98 / Chapter 4.5.1 --- Introduction --- p.98 / Chapter 4.5.2 --- Materials and Methods --- p.98 / Chapter 4.5.3 --- UPLC chromatograms --- p.99 / Chapter 4.6 --- Characterization of F8 using Liquid Chromatography/Mass Spectrometry-Mass Spectrometry (LC/MS-MS) --- p.101 / Chapter 4.6.1. --- Materials --- p.101 / Chapter 4.6.2 --- Methods --- p.102 / Chapter 4.6.3 --- Results --- p.103 / Chapter 4.7 --- ¹⁴C-α-methyl-D-glucopyranoside (¹⁴C-AMG) Uptake Assay of three chemical standards in cultured COS-7 cells expressing SGLT1 or SLGT2 --- p.108 / Chapter 4.7.1 --- Methods --- p.108 / Chapter 4.7.2 --- Results --- p.108 / Chapter 4.8 --- Discussion --- p.111 / Chapter CHAPTER 5 --- IN VIVO STUDIES OF THE ANTI-DIABETIC EFFECT OF SELECTED TRADITIONAL CHINESE HERBS AND NATURAL PRODUCTS IN TYPE 2 DIABETIC RAT MODEL --- p.114 / Chapter 5.1 --- Introduction --- p.114 / Chapter 5.1.1 --- Diabetic Animal Models --- p.114 / Chapter 5.2 --- In vivo Study Tests --- p.117 / Chapter 5.2.1 --- Introduction --- p.117 / Chapter 5.2.2 --- Animals --- p.117 / Chapter 5.2.3 --- Methods --- p.118 / Chapter 5.2.4 --- Results --- p.120 / Chapter 5.3 --- Discussion --- p.125 / Chapter CHAPTER 6 --- GENERAL DISCUSSION --- p.128 / Chapter 6.1 --- Importance of SGLT --- p.128 / Chapter 6.2 --- Current developed SGLT 2 Inhibitors --- p.130 / Chapter 6.3 --- Importance and Treatment of DM by TCMs --- p.132 / Chapter 6.4 --- Screening and Developing drugs from Traditional Chinese medicinal plants --- p.134 / Chapter 6.5 --- Limitations and Improvements --- p.136 / Chapter 6.6 --- Future Works --- p.137 / Chapter 6.7 --- Conclusions --- p.139 / REFERENCES --- p.141
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_328707 |
| Date | January 2012 |
| Contributors | Qu, Yue., Chinese University of Hong Kong Graduate School. Division of Chinese Medicine. |
| Source Sets | The Chinese University of Hong Kong |
| Language | English, Chinese |
| Detected Language | English |
| Type | Text, bibliography |
| Format | electronic resource, electronic resource, remote, 1 online resource (xvii, 156 leaves) : ill. (some col.) |
| Coverage | China |
| Rights | Use of this resource is governed by the terms and conditions of the Creative Commons “Attribution-NonCommercial-NoDerivatives 4.0 International” License (http://creativecommons.org/licenses/by-nc-nd/4.0/) |
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