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研究生:颯雅
研究生(外文):Zayabileg Altansukh
論文名稱:液化澱粉芽孢桿菌用於產生富含GABA的水稻以增強小鼠皮膚中膠原蛋白表達的可能機制
論文名稱(外文):Possible mechanisms employed by Bacillus amyloliquefaciens to generate GABA-rich rice for enhancement of collagen expression in mouse skin
指導教授:黃俊銘黃俊銘引用關係
指導教授(外文):Chun-Ming Huang
學位類別:碩士
校院名稱:國立中央大學
系所名稱:生醫科學與工程學系
學門:工程學門
學類:生醫工程學類
論文出版年:2019
畢業學年度:107
語文別:英文
論文頁數:42
中文關鍵詞:白飯液化澱粉芽孢桿菌γ-氨基丁酸第一型膠原蛋白
外文關鍵詞:RiceBacillus amyloliquefaciens (B. amyloliquefaciens)Gammaaminobutyric acid (GABA)Collagen type 1
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摘要
皮膚是人類最大的器官,包括衰老過程中所造成的皺紋在內的幾種皮膚狀
況可能與膠原蛋白的降解有關。纖維細胞是產生膠原蛋白的主要細胞。第
1 型膠原蛋白(Col1)是人體內最豐富的膠原蛋白,為了創造一種可以刺
激皮膚中膠原蛋白生成的方式,將水稻與生活在土壤和人類皮膚中的解澱
粉芽孢桿菌一起培養。 我們的研究結果表明,水稻與解澱粉芽孢桿菌細菌
的孵育上調了γ-氨基丁酸(GABA)的水平。 局部應用 GABA 或富含
GABA 的水稻顯著增加了小鼠皮膚中 Col1 的產生。 此外,通過小干擾核
糖核酸(siRNA)體內敲除 GABA 受體亞單位β-3(GABRB3)顯著降低了
小鼠皮膚中 Col1 的產生,表明 GABRB3 介導 GABA 誘導的皮膚中 Col1 的
產生。
Abstract
Skin is the largest organ in humans. Several skin conditions including wrinkles
during aging could be associated with degradation of the collagen. Fibroblast is
the main cell that produces collagen. Collagen type 1 (Col1) is the most
abundant collagen in the human body. With an aim to create a modality that can
stimulate collagen production in skin, rice was incubated with a Bacillus
amyloliquefaciens (B. amyloliquefaciens) bacteria which live in both soil and
human skin. Our results demonstrate that incubation of rice with B.
amyloliquefaciens bacteria up-regulated the level of Gamma-aminobutyric acid
(GABA). Topical application of GABA or GABA-rich rice significantly
increased the production of Col1 in mouse skin. Furthermore, in vivo knockout
of GABA receptor subunit beta-3 (GABRB3) by small interfering ribonucleic
(siRNA) markedly reduced Col1 production in the mouse skin, indicating that
GABRB3 mediates the GABA-induced Col1 production in skin.
Contents
Chinese abstract...................................................................................................i
English abstract...................................................................................................ii
Acknowledge.......................................................................................................iii
Contents...............................................................................................................iv
List of figures.......................................................................................................v
List of tables........................................................................................................vi
List of abbreviations..........................................................................................vii
Chapter 1: Introduction......................................................................................1
1.1 Skin...................................................................................................1
1.2 Wrinkle.............................................................................................2
1.3 Collagen............................................................................................4
1.4 Gamma-Aminobutyric acid..............................................................6
1.5 Nutrition of rice................................................................................7
1.6 Bacillus amyloliquefaciens...............................................................9
Chapter 2: Materials and methods..................................................................11
2.1 Materials.........................................................................................11
2.2 Inoculation and Bacterial culture....................................................13
2.3 Rice plant preparation.....................................................................13
2.4 Extraction and purification of IAA.................................................13
2.5 Quantification of GABA content in rice grain................................14
2.6 Animal experiment..........................................................................14
2.6.1 Chemical administrations and behavioral analysis...............14
2.7 Protein extraction and measurement...............................................15
2.7.1 Preparation of lysate from tissues.........................................15
2.7.2 Determination of protein concentration................................15
2.8 Immuniblotting analysis..................................................................15
2.8.1 Polyacrylamide gel electrophoresis......................................15
2.8.2 Western blotting...................................................................16
2.8.3 Immunostaining....................................................................16
2.8.4 Densitometry analyze...........................................................17
2.9 Statistical analysis...........................................................................17
Chapter 3: Results.............................................................................................18
3.1 Enhancement of collagen formation in the mouse skin by
commercial GABA.........................................................................18
3.2 GABA concentration of rice..........................................................19
3.3 GABA concentration of rice in calcium phosphate agar plates...20
3.4 Quantitative Analysis of Indole-3-acetic acid...............................21
3.5 Enhancement of collagen formation in the mouse skin by GABArich rice...........................................................................................22
3.6 Enhancement of the collagen formation in mouse skin by GABArich rice (using CaP agar plates).....................................................23
3.7 The expression of collagen 1 in the GABRB3 receptor blocked by
siRNA in mouse skin......................................................................24
Chapter 4: Discussion and Conclusion............................................................25
References..........................................................................................................27
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