綠茶表沒食子酸酯型唲茶素酸酯對胰島素刺激前脂肪細胞增生的抑制

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劉咸君(Hsien-Chun Liu) 查詢紙本館藏

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生命科學系

論文名稱

綠茶表沒食子酸酯型唲茶素酸酯對胰島素刺激前脂肪細胞增生的抑制
(Inhibitory effect of green tea (-)-epigallocatechin gallate on insulin-stimulated mitogenesis of 3T3-L1 preadipocytes)

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摘要(中)

肥胖是一種常見的疾病，與癌症，糖尿病，高血壓以及心血管疾病都有相關。造成肥胖的原因是脂肪細胞數目的增加，或脂肪細胞內三酸甘油酯的堆積(泛稱脂肪細胞分化)使然。根據研究指出，內分泌與營養都會影響到肥胖的調節作用，其中胰島素在細胞分化時期會刺激細胞進行mitotic clonal expansion及adipogenesis，並促使細胞進行脂肪生成和抑制脂肪分解作用。然而綠茶唲茶素(亦稱維生素P)，尤其是綠茶表沒食子酸酯型唲茶素酸酯(英文名稱和簡稱分別為 (-)-epigallocatechin gallate和EGCG)，已被證實可以抑制脂肪細胞的增生與分化，並可促使前脂肪細胞和脂肪細胞的apoptosis，以及促進脂肪氧化作用，因此可視為預防肥胖的保健物質。有鑑於此，EGCG是否可調節胰島素刺激前脂肪細胞的增生仍然不清楚。在本論文中，使用3T3-L1前脂肪細胞株做為研究素材，我發現胰島素濃度在1-1000 nM，細胞數目和BrdU攝入皆有增加的情形，可以得知胰島素會刺激前脂肪細胞的增生。而胰島素的有效濃度EC50-100 在2- 4天的處理為100 nM。此外，由於胰島素可分別增加胰島素接受器、IRS-1和 Erk三種蛋白的磷酸化，可見胰島素分別增加胰島素接受器、IRS-1和MEK-1的活性。然而前處理EGCG (10-50 μM)於前脂肪細胞可以明顯抑制胰島素所促進的細胞數目、BrdU攝入、胰島素接受器的活性、IRS-1的活性、 MEK-1的活性及胰島素接受器與IRS-1兩者之間的結合作用。以上種種的跡象顯示，EGCG可透過阻礙胰島素的訊息傳導路徑而抑制前脂肪細胞的增生。

摘要(英)

Obesity, a common disease resulting from mitogenesis and lipid accumulation (so-called adipogenesis) of fat cells, is associated with the risks of cancers, diabetes, hypertension, and cardiovascular disease. Development of obesity can be regulated by endocrine and nutritional cues based on a variety of laboratory studies. Insulin (INS) was reported to stimulate mitotic clonal expansion and adipogenesis during fat cell differentiation in additional to stimulating lipogenesis and inhibiting lipolysis. In contrast, green tea polyphenols (the so-called vitamin P), especially the catechin, (-)-epigallocatechin gallate (EGCG), was found to inhibit proliferation and differentiation of fat cells, to induce the apoptosis of preadipocytes and adipocytes, to stimulate fat oxidation, and to be proposed as an obesity chemopreventative. But, determining whether EGCG acts to regulate the stimulatory effect of INS on the mitogenesis of fat cells is still not clear. In this thesis using 3T3-L1 preadipocytes as the research cell model, I found that INS at 1-1000 nM could stimulate the growth of 3T3-L1 preadipocytes as indicated by increased cell number and BrdU incorporation. The EC50-100 of INS was about 100 nM after 2~4 days of treatment. Also, INS stimulated activities of INS receptor (INSR), insulin receptor substrate 1 (IRS-1), and MEK1, respectively, as indicated by increased phosphorylation of INSR, IRS-1, and Erk proteins. However, pretreatment of preadipocytes with EGCG (10-50 μM) resulted in significant decreases in the INS-induced increases in their cell number, BrdU incorporation, activities of INSR, IRS-1, and MEK-1, and the association of INSR with IRS-1. These data suggest that EGCG can inhibit preadipocyte mitogenesis by reduction of INS-mediated signaling in these cells.

關鍵字(中)

★ 綠茶表沒食子酸酯型唲茶素酸酯
★ 前脂肪細胞
★ 胰島素

關鍵字(英)

★ (-)-epigallocatechin gallate
★ insulin
★ 3T3-L1 preadipocytes

論文目次

中文摘要 I
Abstract II
縮寫與全名對照 III
目錄 IV
圖目錄 VI
第一章：緒論 1
一、胰島素(Insulin) 1
二、胰島素接受器(insulin receptor,簡寫為INSR) 1
三、胰島素訊息傳導途徑 2
四、胰島素與前脂肪細胞 3
五、綠茶唲茶素與前脂肪細胞 4
六、動機與目的 5
第二章：材料與方法 6
一.實驗材料 6
二.實驗方法 7
1. 細胞培養 7
2. 細胞處理 7
3. 細胞計數方法 7
4. DNA合成5-bromo-2’-deoxyuridine (BrdU)試驗 8
5. 蛋白質濃度定量 8
6. 抑制劑(PD98059, LY294002, U0126, SB203580）的處理 9
7. 西方點墨法 (Western blotting) 9
8. 免疫沉析法 (Immunoprecipitation) 10
9. 人類胰島素的配製 10
10.生統分析 11
第三章：結果 12
一、胰島素刺激前脂肪細胞的增生 12
二、胰島素刺激前脂肪細胞的增生與MEK-1活性有關 12
三、胰島素刺激前脂肪細胞的增生受到綠茶唲茶素EGCG抑制 13
四、綠茶唲茶素EGCG對胰島素刺激MEK-1活性的影響 13
五、不同種類綠茶唲茶素影響胰島素刺激前脂肪細胞的生長和MEK-1活性 14
六、胰島素刺激前脂肪細胞的胰島素接受器和下游IRS-1蛋白的活性 15
七、綠茶唲茶素EGCG抑制胰島素刺激前脂肪細胞的胰島素接受器和下游IRS-1蛋白的活性 15
八、綠茶唲茶素EGCG抑制胰島素刺激前脂肪細胞之胰島素接受器和下游IRS-1之間的結合能力 16
第四章：討論 17
第五章：結論與展望 20
第六章：參考文獻 21
第七章：附錄 38

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指導教授

高永旭(Yung-Hsi Kao)

審核日期

2006-1-24

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