綠茶表沒食子兒茶素沒食子酸酯經由MicroRNA-let-7a/HMGA2訊息路徑抑制米色前脂肪細胞的生長

以作者查詢圖書館館藏

、以作者查詢臺灣博碩士

、以作者查詢全國書目

、勘誤回報

、線上人數：9

、訪客IP：52.15.236.223

姓名

陳雯婷(Wen-Ting Chen) 查詢紙本館藏

畢業系所

生命科學系

論文名稱

綠茶表沒食子兒茶素沒食子酸酯經由MicroRNA-let-7a/HMGA2訊息路徑抑制米色前脂肪細胞的生長
(Green tea epigallocatechin gallate inhibits beige preadipocyte growth via the microRNA-let-7a/HMGA2 signaling pathways)

相關論文

★ 中華鱉腦垂體甘丙氨激素之研究：cDNA選殖、表現及調控	★ 辛基苯酚對3T3-L1脂肪細胞中resistin的調節作用
★ 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	★ FoxO1 調節抗胰島素激素基因的表現
★ 綠茶表沒食子唲茶素沒食子酸酯受器對於人類乳癌細胞株MCF7生長的影響	★ 綠茶表沒食子酸酯型唲茶素酸酯抑制第一型内皮素作用於脂肪細胞上攝入葡萄糖的訊息機制
★ 綠茶表兒茶素藉由microRNA-494路徑改善橫向主動脈繃紮術誘導型小鼠的心臟疾病	★ 內皮素誘導前脂肪細胞生長的訊息路徑
★ 綠茶對前脂肪細胞生長的影響	★ 綠茶唲茶素對由第一型類胰島素所調節前脂肪細胞生長的影響
★ 綠茶唲茶素對於前脂肪細胞分化的影響	★ Cdk2在綠茶唲茶素調節3T3-L1前脂肪細胞的生長和細胞凋亡扮演著必要性的角色
★ 綠茶唲茶素透過MAPK相關途徑抑制3T3-L1前脂肪細胞的生長	★ 第一型類胰島素生長因子、綠茶唲茶素及雌性素對3T3-L1脂肪細胞中resistin的基因表達有不同的調節效果
★ 綠茶唲茶素對前脂肪細胞內活性氧及榖胱甘肽的影響	★ 胰島素接受器受質在綠茶唲茶素對胰島素刺激前脂肪細胞生長作用中扮演的角色

檔案

[Endnote RIS 格式]

[Bibtex 格式]

[相關文章]

[文章引用]

[完整記錄]

[館藏目錄]

至系統瀏覽論文 (2026-8-1以後開放)

摘要(中)

根據先前研究指出，綠茶表沒食子兒茶素沒食子酸酯 (Epigallocatechin gallate, EGCG) 和 microRNA (miR) 藉由抑制脂肪細胞分裂及脂肪分化過程，進而抑制脂肪細胞生長及脂肪生成，但鮮少研究探討EGCG 藉由調控miR影響脂肪細胞生長。雖然先前實驗結果發現EGCG可能藉由調控miR-let-7a及HMGA2表現影響3T3-L1白色前脂肪細胞生長，但需更進一步實驗驗證，而且目前也無研究指出EGCG是否藉由調控miR路徑影響米色前脂肪細胞生長，故本論文探討EGCG是否影響D12及X9米色前脂肪細胞的生長，以及證實EGCG是否藉由調控miR-let-7a/HMGA2路徑調控3T3-L1白色及D12及X9米色前脂肪細胞的生長。結果發現EGCG會抑制D12及X9米脂細胞的生長，且也會時間性及劑量依賴性地上調miR-let-7a及減少HMGA2基因表現。此外，大量表達miR-let-7a後，3T3-L1、D12及X9細胞生長明顯受到抑制，且HMGA2基因表現顯著性下降。而抑制miR-let-7a表現後，阻擋了EGCG對3T3-L1、D12及X9細胞生長之抑制作用，以及上調miR-let-7a及減少HMGA2基因表現之作用。接著根據雙冷光酵素報導基因檢測實驗結果得知，miR-let-7a可直接作用於HMGA2 3’UTR，但無法作用於mutant HMGA2 3’UTR site，由此可知HMGA2為miR-let-7a之標靶基因。進一步的發現，大量表達HMGA2後，D12及3T3-L1細胞數目明顯增加，且阻擋EGCG對D12及3T3-L1細胞生長及HMGA2基因表現之抑制作用。總之，EGCG 藉由調控miR-let-7a/HMGA2的路徑，進而影響D12及X9米色以及3T3-L1白色前脂肪細胞的生長。本篇研究成果可能有助於解釋 EGCG 介導脂肪細胞活性和脂肪細胞相關疾病的miR 信號傳導機制。

摘要(英)

Green tea epigallocatechin gallate (EGCG) and microRNA (miR) have been reported to inhibit the growth and fat synthesis of fat cell by suppressing mitogenic and adipogenic processes, respectively. Few studies have reported how EGCG affects fat cell growth by regulating miR molecules. Although previous report had shown that EGCG might affect 3T3-L1 white preadipocyte growth by regulating miR-let-7a and HMGA2 expressions, the contention remains to be demonstrated in further studies. No reports were found whether EGCG affects beige preadipocyte growth through the miR pathway. Thus, this study investigated whether EGCG affects beige preadipocyte growth, and demonstrated whether the miR-let-7a/HMGA2 pathways transduce EGCG signals to regulate growths of white and beige preadipocytes. Using D12 and X9 cells, we found that EGCG inhibited beige preadipocyte growth and that EGCG time- and dose-dependently upregulated miR-let-7a mRNA levels and downregulated HMGA2 gene expression. In addition, overexpression of miR-let-7a significantly inhibited growths of D12, X9, and 3T3-L1 cells, and decreased HMGA2 gene expression. Whereas, the miR-let-7a inhibitor antagonized the inhibitory effects of EGCG on cell number and cell viability of all three cell lines. Moreover, miR-let-7a inhibitor reversed the EGCG-increased level of miR-let-7a mRNA and the EGCG-decreased level of HMGA2 gene expression. Further dual-luciferase report assay indicated that miR-let-7a could directly bind to the HMGA2 3′-UTR site but not mutant HMGA2 3′-UTR site, suggesting that HMGA2 is the miR-let-7a target. Also, overexpression of HMGA2 induced increases in both cell number and cell viability and antagonized EGCG-suppressed growth and HMGA2 expression in D12 beige and 3T3-L1 white preadipocytes. We concluded that EGCG inhibits growths of D12 and X9 beige and 3T3-L1 white preadipocytes through modulations of the miR-let-7a and HMGA2 pathways. Results of this study may help explain the miR signaling mechanism through which EGCG mediates fat cell activity and fat cell-associated disease.

關鍵字(中)

★ 綠茶
★ 表沒食子兒茶素沒食子酸酯
★ 米色前脂肪細胞

關鍵字(英)

★ Green tea
★ epigallocatechin gallate
★ beige preadipocyte

論文目次

中文摘要 i
Abstract ii
致謝 iii
目錄 iv
表目錄 vii
圖目錄 viii
縮寫表與全名對照表 x
壹、前言 1
一、MicroRNA小分子核醣核酸 1
二、脂肪細胞 2
三、綠茶 3
四、研究動機及目的 5
貳、材料與方法 7
一、實驗材料 7
二、實驗方法 7
1. 細胞培養 7
2. EGCG配製 7
3. 細胞生長實驗 8
4. 大量表現miR-let-7a (miR-let-7a mimic) 與抑制miR-let-7a表現 (miR-let-7a inihibitor) 之試驗以及大量表現HMGA2之試驗 9
5. 基因分析 10
6. 蛋白質分析-西方墨點法 (Western blot) 12
7. 雙冷光酵素報導基因檢測方法 (Dual-Luciferase Reporter Assay) 14
8. 統計分析 14
參、實驗結果 15
一、 EGCG對D12及X9米色前脂肪細胞生長之影響 15
二、 EGCG對D12及X9米色前脂肪細胞內miR-let-7a及HMGA2表現之影響 16
三、大量表現miR-let-7a對D12及X9米色前脂肪細胞生長及細胞內HMGA2表現之影響 17
四、大量表現miR-let-7a對3T3-L1白色前脂肪細胞生長及細胞內HMGA2表現之影響 18
五、 miR-let-7a抑制劑對D12及X9米色前脂肪細胞生長及細胞內HMGA2表現之影響 19
六、 miR-let-7a抑制劑對3T3-L1白色前脂肪細胞生長及細胞內HMGA2表現之影響 21
七、 EGCG在D12米色前脂肪細胞中上調miR-let-7a作用於HMGA2 22
八、 EGCG在3T3-L1白色前脂肪細胞中上調miR-let-7a作用於HMGA2 23
九、大量表現HMGA2阻擋EGCG對D12米色前脂肪細胞生長及細胞內HMGA2表現之影響 23
十、大量表現HMGA2阻擋EGCG對3T3-L1白色前脂肪細胞生長及細胞內HMGA2表現之影響 24
肆、討論 26
伍、結論 30
陸、參考文獻 31
柒、附錄 65
附圖一、EGCG在不同時間點對 3T3 L1 前白色脂肪細胞內miR let 7a及其標靶基因HMGA2表現的影響 (陳嘉佩, 2020) 66
附圖二、不同時間的EGCG處理對D12及X9米色前脂肪細胞內miR-143基因表現之影響 67
附圖三、以Dual-luciferase Report assay證實HMGA2為miR-let-7a之標靶基因 68
附圖四、不同劑量的EGCG處理抑制D12米色前脂肪細胞內HMGA1基因表現 69
附錄五、使用試劑之配方 70
附錄六、MicroRNA mimic及inhibitor序列 73
附錄七、HMGA2及β-actin完整印跡圖 74
附錄八、Plasmid map 75

參考文獻

陳嘉佩. (2020). 綠茶表沒食子兒茶素沒食子酸酯藉由微核醣核酸-143/蛋白δ同源物1路徑抑制3T3-L1前脂肪細胞的生長.
Ambros, V. (2004). The functions of animal microRNAs. Nature 431, 350-355. Anand, A., & Chada, K. (2000). In vivo modulation of Hmgic reduces obesity. Nature genetics 24, 377-380.
Arce-Cerezo, A., García, M., Rodríguez-Nuevo, A., Crosa-Bonell, M., Enguix, N., Peró, A., Muñoz, S., Roca, C., Ramos, D., & Franckhauser, S. (2015). HMGA1 overexpression in adipose tissue impairs adipogenesis and prevents diet-induced obesity and insulin resistance. Scientific reports 5, 1-14.
Arffa, M., Zapf, M., Kothari, A., Chang, V., Gupta, G., Ding, X., Al-Gayyar, M., Syn, W., Elsherbiny, N., & Kuo, P. (2016). Epigallocatechin-3-gallate upregulates miR-221 to inhibit osteopontin-dependent hepatic fibrosis. PLoS One 11, e0167435.
Argilés, J. M., López-Soriano, F. J., & Busquets, S. (2019). Mediators of cachexia in cancer patients. Nutrition 66, 11-15.
Arner, P., & Kulyté, A. (2015). MicroRNA regulatory networks in human adipose tissue and obesity. Nature Reviews Endocrinology 11, 276.
Ashar, H. R., Chouinard Jr, R. A., Dokur, M., & Chada, K. (2010). In vivo modulation of HMGA2 expression. Biochimica et Biophysica Acta (BBA)-Gene Regulatory Mechanisms 1799, 55-61.
Brown, A. L., Lane, J., Coverly, J., Stocks, J., Jackson, S., Stephen, A., Bluck, L., Coward, A., & Hendrickx, H. (2008). Effects of dietary supplementation with the green tea polyphenol epigallocatechin-3-gallate on insulin resistance and associated metabolic risk factors: randomized controlled trial. British journal of nutrition 101, 886-894.
Bushati, N., & Cohen, S. M. (2007). microRNA functions. Annual Review of Cell and Developmental Biology 23, 175-205.
Camp, H. S., Ren, D., & Leff, T. (2002). Adipogenesis and fat-cell function in obesity and diabetes. Trends in molecular medicine 8, 442-447.
Chacko, S. M., Thambi, P. T., Kuttan, R., & Nishigaki, I. (2010). Beneficial effects of green tea: a literature review. Chinese medicine 5, 1-9.
Chen, Y., Siegel, F., Kipschull, S., Haas, B., Fröhlich, H., Meister, G., & Pfeifer, A. (2013). miR-155 regulates differentiation of brown and beige adipocytes via a bistable circuit. Nature communications 4, 1-13.
Colomer, R., Sarrats, A., Lupu, R., & Puig, T. (2017). Natural polyphenols and their synthetic analogs as emerging anticancer agents. Current drug targets, 18, 147-159.
Dulloo, A., Seydoux, J., Girardier, L., Chantre, P., & Vandermander, J. (2000). Green tea and thermogenesis: interactions between catechin-polyphenols, caffeine and sympathetic activity. International Journal of Obesity 24, 252-258.
Furuyashiki, T., Nagayasu, H., Aoki, Y., Bessho, H., Hashimoto, T., Kanazawa, K., & Ashida, H. (2004). Tea catechin suppresses adipocyte differentiation accompanied by down-regulation of PPARγ2 and C/EBPα in 3T3-L1 cells. Bioscience, biotechnology, and biochemistry 68, 2353-2359.
Goody, D., & Pfeifer, A. (2019). MicroRNAs in brown and beige fat. Biochimica et Biophysica Acta (BBA)-Molecular and Cell Biology of Lipids 1864, 29-36.
Hu, W., Ru, Z., Xiao, W., Xiong, Z., Wang, C., Yuan, C., Zhang, X., & Yang, H. (2018). Adipose tissue browning in cancer-associated cachexia can be attenuated by inhibition of exosome generation. Biochemical and Biophysical Research Communications 506, 122-129.
Hydbring, P., & Badalian-Very, G. (2013). Clinical applications of microRNAs. F1000Research 2.
John, B., Enright, A. J., Aravin, A., Tuschl, T., Sander, C., & Marks, D. S. (2004). Human microRNA targets. PLOS Biology 2, e363.
Johnson, C. D., Esquela-Kerscher, A., Stefani, G., Byrom, M., Kelnar, K., Ovcharenko, D., Wilson, M., Wang, X., Shelton, J., & Shingara, J. (2007). The let-7 microRNA represses cell proliferation pathways in human cells. Cancer research 67, 7713-7722.
Kajimoto, K., Naraba, H., & Iwai, N. (2006). MicroRNA and 3T3-L1 pre-adipocyte differentiation. RNA 12, 1626-1632.
Kao, Y. H., Chang, H. H., Lee, M. J., & Chen, C. L. (2006). Tea, obesity, and diabetes. Molecular nutrition & food research 50, 188-210.
Khan, N., & Mukhtar, H. (2007). Tea polyphenols for health promotion. Life sciences, 81, 519-533.
Kim, Y. J., Hwang, S. H., Cho, H. H., Shin, K. K., Bae, Y. C., & Jung, J. S. (2012). MicroRNA 21 regulates the proliferation of human adipose tissue‐derived mesenchymal stem cells and high‐fat diet‐induced obesity alters microRNA 21 expression in white adipose tissues. Journal of cellular physiology 227, 183-193.
Klaus, S., Pültz, S., Thöne-Reineke, C., & Wolfram, S. (2005). Epigallocatechin gallate attenuates diet-induced obesity in mice by decreasing energy absorption and increasing fat oxidation. International Journal of Obesity 29, 615-623.
Lafontan, M., & Berlan, M. (1993). Fat cell adrenergic receptors and the control of white and brown fat cell function. Journal of lipid research 34, 1057-1091.
Lee, C.-C., Shih, Y.-C., Kang, M.-L., Chang, Y.-C., Chuang, L.-M., Devaraj, R., & Juan, L.-J. (2019). Naa10p Inhibits Beige Adipocyte-Mediated Thermogenesis through N-α-acetylation of Pgc1α. Molecular cell 76, 500-515. e508.
Lee, M. S., Kim, C. T., Kim, I. H., & Kim, Y. (2009). Inhibitory effects of green tea catechin on the lipid accumulation in 3T3‐L1 adipocytes. Phytotherapy research : PTR 23, 1088-1091.
Martos-Rus, C., Katz-Greenberg, G., Lin, Z., Serrano, E., Whitaker-Menezes, D., Domingo-Vidal, M., Roche, M., Ramaswamy, K., Hooper, D. C., & Falkner, B. (2021). Macrophage and adipocyte interaction as a source of inflammation in kidney disease. Scientific reports 11, 1-14.
McGregor, R. A.; Choi, M. S. (2011). microRNAs in the regulation of adipogenesis and obesity. Current molecular medicine 11, 304-316.
Melillo, R. M., Pierantoni, G. M., Scala, S., Battista, S., Fedele, M., Stella, A., De Biasio, M. C., Chiappetta, G., Fidanza, V., & Condorelli, G. (2001). Critical role of the HMGI (Y) proteins in adipocytic cell growth and differentiation. Molecular and cellular biology 21, 2485-2495.
Otton, R., Bolin, A. P., Ferreira, L. T., Marinovic, M. P., Rocha, A. L. S., & Mori, M. A. (2018). Polyphenol-rich green tea extract improves adipose tissue metabolism by down-regulating miR-335 expression and mitigating insulin resistance and inflammation. The Journal of nutritional biochemistry 57, 170-179.
Rasheed, Z., Rasheed, N., & Al-Shaya, O. (2018). Epigallocatechin-3-O-gallate modulates global microRNA expression in interleukin-1β-stimulated human osteoarthritis chondrocytes: potential role of EGCG on negative co-regulation of microRNA-140-3p and ADAMTS5. European journal of nutrition 57, 917-928.
Roush, S., & Slack, F. J. (2008). The let-7 family of microRNAs. Trends in cell biology 18, 505-516.
Slota, J. A., & Booth, S. A. (2019). MicroRNAs in neuroinflammation: implications in disease pathogenesis, biomarker discovery and therapeutic applications. Non-coding RNA 5, 35.
Sun, L., Sun, P., Zhou, Q.-y., Gao, X., & Han, Q. (2016). Long noncoding RNA MALAT1 promotes uveal melanoma cell growth and invasion by silencing of miR-140. American journal of translational research 8, 3939.
Sun, T., Fu, M., Bookout, A. L., Kliewer, S. A., & Mangelsdorf, D. J. (2009). MicroRNA let-7 regulates 3T3-L1 adipogenesis. Molecular endocrinology 23, 925-931.
Tsang, W. P., & Kwok, T. T. (2010). Epigallocatechin gallate up-regulation of miR-16 and induction of apoptosis in human cancer cells. The Journal of nutritional biochemistry 21, 140-146.
Vignali, R., & Marracci, S. (2020). HMGA genes and proteins in development and evolution. International journal of molecular sciences 21, 654.
Wolfram, S., Wang, Y., & Thielecke, F. (2006). Anti‐obesity effects of green tea: from bedside to bench. Molecular nutrition & food research 50, 176-187.
Wu, J., Boström, P., Sparks, L. M., Ye, L., Choi, J. H., Giang, A.-H., Khandekar, M., Virtanen, K. A., Nuutila, P., & Schaart, G. (2012). Beige adipocytes are a distinct type of thermogenic fat cell in mouse and human. Cell 150, 366-376.
Xi, Y., Shen, W., Ma, L., Zhao, M., Zheng, J., Bu, S., Hino, S., & Nakao, M. (2016). HMGA2 promotes adipogenesis by activating C/EBPβ-mediated expression of PPARγ. Biochemical and Biophysical Research Communications 472, 617-623.
Xing, L., Zhang, H., Qi, R., Tsao, R., & Mine, Y. (2019). Recent advances in the understanding of the health benefits and molecular mechanisms associated with green tea polyphenols. Journal of agricultural and food chemistry 67, 1029-1043.
Yamada, S., Tsukamoto, S., Huang, Y., Makio, A., Kumazoe, M., Yamashita, S., & Tachibana, H. (2016). Epigallocatechin-3-O-gallate up-regulates microRNA-let-7b expression by activating 67-kDa laminin receptor signaling in melanoma cells. Scientific reports 6, 1-8.
Zhang, P., Du, J., Wang, L., Niu, L., Zhao, Y., Tang, G., Jiang, Y., Shuai, S., Bai, L., & Li, X. (2018). MicroRNA-143a-3p modulates preadipocyte proliferation and differentiation by targeting MAPK7. Biomedicine & Pharmacotherapy 108, 531-539.

指導教授

高永旭(Yung-Hsi Kao)

審核日期

2021-7-27

推文