近年來，腸促胰島素類藥物胰高血糖素樣肽－1受體（GLP-1R）激動劑（如liraglutide，exenatide）和二肽基肽酶-4（DPP-4）抑制劑（如sitagliptin，vildagliptin）在2型糖尿病治療中得到廣泛應用。然而，2型糖尿病患者中腸促胰島素效應嚴重受損。研究表明，2型糖尿病患者的腸促胰島素激素（GLP-1和GIP）分泌並不顯著降低，因此腸促胰島素效應受損主要是由於2型糖尿病患者中β細胞對腸促胰島素激素反應性降低。GIP在2型糖尿病患者中刺激胰島素分泌的功能幾乎完全消失。相比較，GLP-1刺激胰島素分泌功能在2型糖尿病患者中部分保留。2型糖尿病中腸促胰島素效應受損的具體機制目前仍不清楚。本論文主要從2型糖尿病患者的腸促胰島素效應受損的機理及對腸促胰島素類藥物療效的影響上進行相關研究。 / 我們較早的研究發現高血糖能降低胰島β細胞GLP-1R受體的表達，從而損傷胰島β細胞GLP-1R信號通路是2型糖尿中腸促胰島素受損的部分原因。由於高血脂和高血糖都是糖尿病的主要特徵，我進一步研究了高血脂對β細胞的腸促胰島素信號通路的影響。體外實驗表明，棕櫚酸能降低胰島β細胞中GLP-1R的表達，並且抑制了GLP-1刺激的cAMP產生及CREB的磷酸化；在β細胞中外源性表達GLP-1R能部分恢復棕櫚酸損傷的GLP-1刺激cAMP產生及CREB的磷酸化。此外，在db/db小鼠和HFD誘導的肥胖及糖尿病小鼠模型中，降脂藥bezafibrate和niacin 能顯著提高腸促胰島素類藥物sitagliptin 和exendin-4的降糖效果，並且伴隨著對胰島形態結構的改善以及增加胰島β細胞質量。 / 另一方面，2型糖尿病患者胰島β細胞的功能和品質持續性的減少。其中慢性的高血糖和高血脂是兩個主要因素。臨床研究發現sitagliptin的降糖效果隨著糖尿病持續的時間增加而降低，而具體機理並不清楚。我們以前研究發現高血糖損傷胰島β細胞GLP-1R的表達及其信號通路是2型糖尿病腸促胰島素效應受損的重要原因。我進一步探討了exendin-4在STZ-HFD 誘導的較輕程度糖尿病（MH）小鼠（相對較輕的高血糖以及β細胞質量減少）和重度糖尿病(SH)小鼠（嚴重高血糖以及β細胞質量減少）的治療效果。研究發現， exendin-4只在MH小鼠中顯著降低血糖，改善糖耐量，恢復正常胰島結構及增加β細胞質量。而在兩組小鼠中exendin-4 都能降低體重，增加胰腺重量，但對食都沒影響。儘管exendin-4能顯著降低MH小鼠中胰島素耐量實驗葡萄糖水平，但HORM-IR無顯著差異。此外，exendin-4處理對胰島素刺激的肝臟及肌肉組織中磷酸化AKT和GSK水準在兩組小鼠中都無明顯改變。 / 總之，我的研究強調了血糖血脂的控制在2型糖尿病患者中的重要作用。我也發現高血糖、高血脂導致2型糖尿病患者β細胞功能持續受損的同時也損傷了GLP-1R信號通路，以至腸促胰島素類藥物療效的降低。我們的研究對腸促胰島素類藥物在2型糖尿病患者中的合理使用提供了重要資訊。 / Incretin-based drugs, such as glucagon-like peptide-1 (GLP-1) receptor agonists (e.g. liraglutide and exenatide) and dipeptidyl peptidase-4 (DPP-4) inhibitors (e.g. sitagliptin and vildagliptin), which inhibit degrading intact GLP-1, have been a novel therapeutics for the treatment of type 2 diabetes. Type 2 diabetes mellitus (T2DM) is associated with reduced incretin effects. The underlying mechanism, however, is not well understood. / Our previous studies showed that hyperglycemia downregulates glucagon-like peptide-1 (GLP-1) receptor (GLP-1R) which potentially contributes to the impaired incretin responses in cells. Whereas the effects of hyperlipidemia, another common clinical feature of T2DM, on GLP-1 response is largely unknown. In this study, I investigated the effects of free fatty acids (FFA) on incretin receptor signalings, and examined the glucose-lowering efficacy of incretin-based drugs in combination with lipid-lowering agents. I found that palmitate treatment decreased GLP-1R expression in rodent insulinoma cell lines, which was associated with impaired GLP-1 stimulated cAMP production and phosphorylation of CREB. Over-expression of GLP-1R restored the cAMP production and the phosphorylation of CREB. Treatment with bezafibrate or niacin in combination with des-fluoro-sitagliptin or exendin-4 produced more robust glycemic control associated with improved pancreatic islet morphology and islet cell function in db/db mice and HFD-fed mice. / On the other hand, chronic hyperglycemia and hyperlipidemia can cause a progressive deterioration in pancreatic beta-cell function and mass in patients with type 2 diabetes mellitus. It has been reported that the efficacy of incretin-based therapeutics is attenuated with the duration of diabetes. In our previous study, we have shown that hyperglycemia downregulates GLP-1 receptor which in turn may contribute to the impaired incretin effect in type 2 diabetes. In this study, I further investigated the efficacy of GLP-1 based drug exendin-4 in a rodent model of type 2 diabetes with different degrees of hyperglycemia and reduction of beta cell mass. I found that in moderate hyperglycemia (MH) group but not in severe hyperglycemia (SH) group, exendin-4 treatment significantly reduced fed glucose levels and plasma lipid profiles, improved glucose tolerance and glucose stimulated insulin secretion, and was associated with restored islets morphology and increased beta cell mass. Exendin-4 significantly decreased body weight gain and increased pancreatic mass both in MH and SH group. Although glucose levels were significantly reduced in MH group with exentin-4 treatment during insulin tolerance test, exendin-4 had no effects on HORM-IR, food intake, and insulin stimulated p-AKT/p-GSK in liver and muscle in both MH and SH group. / In summary, my findings highlight the importance of comprehensive lipid and glycemic control in type 2 diabetes mellitus. I found that factors including hyperglycemia and hyperlipidemia that cause progressive decline in beta cell failure impaired beta cell GLP-1R signaling as well as the efficacy of incretin-based therapies. These results add to our knowledge regarding the mechanism of incretin-based therapy in improving glycemic control in type 2 diabetic patients and provide new insights in designing treatment strategies including incretin-based therapy for type 2 diabetic patients. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Kang, Zhanfang. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 103-123). / Abstract also in Chinese. / 論文摘要 --- p.viii / Impaired Incretin Effects in Type 2 Diabetes: Mechanism and Therapeutic Implication --- p.1 / DECLARATION --- p.i / ACKNOWLEGEMENTS --- p.ii / ABBREVIATIONS --- p.iii / PUBLICATIONS AND AWARDS --- p.v / Chapter 1 --- Abstract --- p.vi / Chapter 2 --- Chapter : Introduction --- p.1 / Chapter 2.1 --- The history of incretin discovery --- p.1 / Chapter 2.2 --- The incretin hormones:GLP-1 and GIP --- p.1 / Chapter 2.3 --- Gene structure and regulation of incretin hormone gene expression --- p.2 / Chapter 2.3.1 --- Gene structure and regulation of GLP-1 gene expression --- p.2 / Chapter 2.3.2 --- Gene structure and regulation of GIP gene expression --- p.5 / Chapter 2.4 --- Secretion and metabolism of GLP-1 and GIP --- p.5 / Chapter 2.4.1 --- Regulation of GLP-1 and GIP secretion --- p.5 / Chapter 2.4.2 --- Degradation of GLP-1 and GIP by DPP-4 enzyme --- p.8 / Chapter 2.5 --- GLP-1 and GIP receptor --- p.10 / Chapter 2.6 --- biological actions of GLP-1 and GIP --- p.11 / Chapter 2.6.1 --- Actions of GLP-1 in peripheral tissues --- p.12 / Chapter 2.6.2 --- Actions of GIP in peripheral tissues --- p.17 / Chapter 2.7 --- Impaired incretin effect in type 2 diabetes patients --- p.17 / Chapter 2.7.1 --- Secretion of incretin hormones in patients with type 2 diabetes --- p.18 / Chapter 2.7.2 --- Impaired the responsiveness to GLP-1 and GIP in pancreatic beta cells --- p.19 / Chapter 2.8 --- Incretin-based drugs --- p.19 / Chapter 2.9 --- Type 2 diabetes and beta cell failure --- p.21 / Chapter 2.9.1 --- Natural history of type 2 diabetes --- p.21 / Chapter 2.9.2 --- Decline of beta cell function and mass in type 2 diabetes --- p.22 / Chapter 2.9.3 --- Factors for progressive loss of beta cell function and mass --- p.24 / Chapter 3 --- Chapter: Pharmacological reduction of free fatty acids restores the efficacy of incretin-based therapies in diabetic mouse models through beta cell GLP-1 receptor signalings --- p.30 / Chapter 3.1 --- Summary --- p.30 / Chapter 3.2 --- Introduction --- p.32 / Chapter 3.3 --- Materials and Methods --- p.35 / Chapter 3.3.1 --- Chemicals and Reagents --- p.35 / Chapter 3.3.2 --- Preparation of 8 mM sodium palmitate solution with 10.5% BSA --- p.35 / Chapter 3.3.3 --- Construct of an adenoviral vector for expressing mouse GLP-1R --- p.36 / Chapter 3.3.4 --- Cell culture and treatment --- p.37 / Chapter 3.3.5 --- RNA extraction and quantitative RT-PCR --- p.37 / Chapter 3.3.6 --- Analysis of phosphorylation of CREB --- p.38 / Chapter 3.3.7 --- Measurement of insulin secretion --- p.39 / Chapter 3.3.8 --- RT-PCR analysis of mouse GLP-1R expression --- p.39 / Chapter 3.3.9 --- Measurement of cAMP production --- p.40 / Chapter 3.3.10 --- Animals and experimental protocols --- p.40 / Chapter 3.3.11 --- Oral glucose tolerance test (OGTT) and insulin tolerance test (ITT) --- p.41 / Chapter 3.3.12 --- Acute glucose-lowering actions of Ex-4 and D-GIP in db/db Mice --- p.41 / Chapter 3.3.13 --- Lipid levels measurement --- p.42 / Chapter 3.3.14 --- Histological analysis --- p.42 / Chapter 3.3.15 --- Statistical analysis --- p.42 / Chapter 3.4 --- Results --- p.43 / Chapter 3.4.1 --- Reduced expression of GLP-1R in palmitate-treated b cells and islets of hyperlipedemic db/db mice. --- p.43 / Chapter 3.4.2 --- Palmitate impairs GLP-1 stimulated cAMP production and p-CREB in rodent insulinoma cell lines --- p.45 / Chapter 3.4.3 --- Palmitate reduced GLP-1 and GIP stimulated insulin secretion in rat INS-1E cells --- p.46 / Chapter 3.4.4 --- Mouse GLP-1R mRNA is expressed in rat INS-1E cells after infected with Ad-GLP-1R --- p.47 / Chapter 3.4.5 --- Expression of exogenous GLP-1R restores GLP-1 stimulated cAMP production and p-CREB in palmitate-treated rodent insulinoma cell lines --- p.48 / Chapter 3.4.6 --- Lipid lowering enhances the efficacy of DPP-4 inhibitor des-fluoro-sitagliptin in db/db mice --- p.50 / Chapter 3.4.7 --- Lipid-lowering enhances the efficacy of DPP-4 inhibitor des-flouro-sitagliptin in HFD-fed mice --- p.56 / Chapter 3.4.8 --- Lipid lowering enhances the efficacy of an agonist to GLP-1R (Ex-4) but not to GIPR (D-GIP) in db/db mice --- p.59 / Chapter 3.5 --- Discussion --- p.67 / Chapter 4 --- Chapter : Further study on the impairment of incretin effect by hyperglycemia in a rodent model of type 2 diabetes --- p.71 / Chapter 4.1 --- Summary --- p.71 / Chapter 4.2 --- Introduction --- p.73 / Chapter 4.3 --- Materials and Methods --- p.76 / Chapter 4.3.1 --- Animals and treatment --- p.76 / Chapter 4.3.2 --- Oral glucose tolerance test and insulin tolerance test --- p.76 / Chapter 4.3.3 --- Plasma parameters --- p.77 / Chapter 4.3.4 --- Histological staining and quantification of beta cell mass --- p.77 / Chapter 4.3.5 --- Insulin signaling analysis --- p.78 / Chapter 4.3.6 --- Western blot analysis --- p.78 / Chapter 4.3.7 --- Statistical analysis --- p.79 / Chapter 4.4 --- Results --- p.80 / Chapter 4.4.1 --- Exendin-4 reduced fed glucose levels in MH mice but not in SH mice --- p.80 / Chapter 4.4.2 --- Exendin-4 reduced body weight gain and did not affect food intake in both MH mice and SH mice --- p.81 / Chapter 4.4.3 --- Exendin-4 improved glucose tolerance and glucose stimulated insulin secretion in MH mice but not in SH mice --- p.83 / Chapter 4.4.4 --- Effects of exendin-4 on insulin sensitivity in MH and SH mice --- p.84 / Chapter 4.4.5 --- Effects of exendin-4 on lipid profiles in MH and SH mice --- p.86 / Chapter 4.4.6 --- Effects of exendin-4 on tissues weight in MH and SH mice --- p.87 / Chapter 4.4.7 --- Pancreatic islet morphology and analysis of beta cell mass --- p.88 / Chapter 4.4.8 --- Exendin-4 had no significant effects on insulin signaling pathway in liver and muscle --- p.91 / Chapter 4.5 --- Discussion --- p.94 / Chapter 5 --- Chapter : Summary --- p.100 / Chapter 6 --- References --- p.103
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_328462 |
| Date | January 2012 |
| Contributors | Kang, Zhanfang., Chinese University of Hong Kong Graduate School. Division of Medical Sciences. |
| 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 (xiv, 123 leaves) : ill. (some col.) |
| 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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