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  1. NTU Theses and Dissertations Repository
  2. 醫學院
  3. 微生物學科所
請用此 Handle URI 來引用此文件: http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/35367
完整後設資料紀錄
DC 欄位值語言
dc.contributor.advisor蕭寧馨
dc.contributor.authorLi-Lin wangen
dc.contributor.author王麗琳zh_TW
dc.date.accessioned2021-06-13T06:49:50Z-
dc.date.available2007-07-11
dc.date.copyright2005-08-01
dc.date.issued2005
dc.date.submitted2005-07-28
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陳翠英 (2005) 以 Caco-2 細胞株探討維生素 C 影響鐵吸收之新作用機制並評估某些市售食品之鐵生物利用率。 國立台灣大學微生物與生化學研究所碩士論文。
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dc.identifier.urihttp://tdr.lib.ntu.edu.tw/jspui/handle/123456789/35367-
dc.description.abstract已知小腸細胞鐵還原酵素 ferric reductase 是影響鐵吸收率的重要因子。本實驗目的在於利用 Caco-2 細胞模式,探討 ferric reductase 反應的電子來源及影響其活性之成份,並評估這些成份對細胞攝鐵能力之影響。 Caco-2 細胞在滿盤後 13-18天進行攝鐵實驗,以 100 M FeCl3 提供鐵源,處理 6 小時之後,測量細胞 ferric reductase 酵素活性及鐵蛋白含量,作為攝鐵能力之指標。培養槽有單槽與雙槽兩種,使用雙槽培養皿時,細胞接種於內槽形成具有 tight junction 之單槽,鐵源均添加於上層,維生素 C 等實驗成份則依實驗設計而添加於上層或下層。
單槽實驗結果顯示,以不添加任何維生素 C 為控制組, 1 mM 還原型維生素 C (AA) 可使 ferric reductase 活性升高 12 倍,使細胞鐵蛋白含量升高 10 倍,其效應與劑量有正相關性。 1 mM 氧化型維生素 C (DeHA) 會使酵素活性升高 3-4 倍,並使鐵蛋白含量增高 5-6 倍。 AA 對酵素活性的影響約為 DeHA 的 4 倍,對鐵蛋白的影響則約為 1-2 倍。 以 DeHA 處理細胞時,若同時利用 BSO 降低 GSH 含量, 可使酵素活性及鐵蛋白含量分別降低 60% 及 50%;若利用 BCNU 降低細胞 GSH 含量,也可使酵素活性及鐵蛋白含量則分別降低 60% 及 40% ;可見 GSH 與鐵之吸收有關,小腸細胞會利用細胞內 GSH 還原氧化型維生素 C 而增加 ferric reductase 活性與促進鐵吸收。以 quercetin 或單寧酸處理細胞時,都會使 ferric reductase 活性上升,但鐵蛋白含量卻低於控制組,表示此兩種物質會藉由化學作用抑制鐵質進入細胞。
雙槽實驗結果顯示, AA 無論添加於上層或下層,均可使 ferric reductase 活性與鐵蛋白含量顯著高於未添加者; DeHA 添加於上層也可使酵素活性與鐵蛋白含量顯著高於未添加者,但添加於下層則無效應。添加於下層之 AA 無法拮抗 quercetin 或單寧酸對鐵吸收的抑制作用。針對 carbonyl Fe、FeCl3、Fe-NTA 三種鐵劑,下層未添加 AA 時,鐵蛋白含量為 Fe-NTA > FeCl3 > carbonyl Fe;添加 AA 時, carbonyl Fe 與 FeCl3 處理組之鐵蛋白含量顯著增加,以 FeCl3 顯著高於 carbonyl Fe ,但 Fe-NTA 與前述兩者並無顯著差異;顯示利用 Caco-2 細胞評估食物鐵可利用率時,細胞維生素 C 含量不足可能成為限制因素而有低估三價鐵之利用率的疑慮。
zh_TW
dc.description.abstractStudies have implicated duodenal ferric reductase activity as an important factor in regulation of intestinal iron absorption. Caco-2 cell model was used to study the electron source of ferric reductase and the effect of ascorbic acid on accuracy of iron bioavailability estimation. In the iron uptake experiments, Caco-2 cells were conducted after 13-18d post-confluence and treated with 100 M of FeCl3 for six hours. Cellular ferric reductase activity was assayed to be an indicator of reducing ability and ferritin level was assayed to be an indicator of iron status in Caco-2 cells.
Compared to the control group, ferric reductase activity and ferritin levels showed respective 3- to 4-fold and 5- to 6-fold increase when dehydroascorbic acid (DeHA) was added. When DeHA + BSO or DeHA + BCNU were added, both ferric reductase activity decrease 60%, and ferritin levels showed respective 50% and 40% decrease compared with the group which only added DeHA. This study demonstrated that cellular vitamin C mediated iron uptake enzymatically in addition to extracellular reduction. Cellular GSH level can also affect the ability of cellular iron uptake. Ferric reductase activity was very high when cells were treated with quercetin or tannic acid, but the ferritin level was lower than control group both.
To assess the effect of ascorbic acid on accuracy of iron bioavailability estimation, cells were seeded in insert plate and ascorbic acid were added in basolateral site. The ferric reductase activity and ferritin level were both significantly higher in the group that was added ascorbic acid in basolateral site. This result indicates the importance of electron source on estimation of iron bioavailability.
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dc.description.tableofcontents總目錄
中文摘要
I
英文摘要
III

圖目錄
iv
縮寫對照表
vi

第一章
文獻回顧 1
第一節
鐵的恆定之調控 1
第二節
小腸細胞鐵吸收之分子機制 4
第三節
維生素 C 的運送與再利用 8
第四節
飲食中與鐵吸收相關之因子 12
第五節
Caco-2 細胞模式 15
第六節
研究目的與內容 19

第二章
材料與方法 21
第一節
Caco-2 細胞實驗 21
第二節
攝鐵實驗用藥品及步驟 23
第三節
Caco-2 細胞存活率測定-MTT 染色法 26
第四節 溶液中之鐵劑其鐵形式分佈測定 27
第五節
Caco-2 細胞 Ferric reductase 活性測定 29
第六節
Caco-2 細胞內鐵蛋白定量 31
第七節
蛋白質定量 32
第八節
統計分析 33

第三章
結果 34
第一節
還原型維生素 C 對細胞 ferric reductase 活性之影響 34
第二節
分化時間對細胞 ferric reductase 活性之影響 35
第三節
氧化型維生素 C 對細胞攝鐵能力的影響 37
第四節
GSH 對於 ferric reductase 及鐵蛋白含量之影響 39
第五節
Quercetin 對 ferric reductase 及鐵蛋白含量之影響 41
第六節
Tannic acid 對 ferric reductase 及鐵蛋白含量之影響 42
第七節
利用 Caco-2 細胞評估不同型式鐵劑之生物利用率 43

第四章
討論 59
第一節
Caco-2 細胞 ferric reductase 與鐵蛋白含量間的關係 59
第二節
還原型維生素 C 對 Caco-2 細胞 ferric reductase 活性及鐵蛋白含量之影響 60
第三節
還原型維生素 C 對 Caco-2 細胞 ferric reductase 活性及鐵蛋白含量之影響 61
第四節
Glutathione 的含量對 Caco-2 細胞 ferric reductase 活性及鐵蛋白含量之影響 62
第五節
Phenolic compound 對 Caco-2 細胞 ferric reductase 活性及鐵蛋白含量之影響 63
第七節
以 Caco-2 細胞作為評估鐵生物利用率之模式的建立 65

第五章 總結 67

第六章 參考文獻 69

















圖目錄
圖 1-1
小腸鐵吸收機制 8
圖 1-2
還原型維生素 C 及氧化型維生素 C 的結構 10
圖 1-3
細胞內抗氧化機制的串聯 12
圖 1-4
Galloyl group、tannic acid 及 quercetin 的結構式 15
圖 1-5
結合體外消化與 Caco-2 細胞用來評估鐵生物利用率之體外模式 18
圖 1-6
本論文建立之利用 Caco-2 細胞評估鐵生物利用率的模式 20

圖 3-1
不同濃度的還原型維生素 C 對 Caco-2 細胞 Ferric reductase 活性(A)及鐵蛋白含量 (B) 的影響 45
圖 3-2
還原型維生素 C 對 Caco-2 細胞 Ferric reductase 活性 (A) 及鐵蛋白含量 (B) 的影響 46
圖 3-3
Caco-2 細胞分化程度與 ferric reductase 活性之影響 47
圖 3-4
不同濃度的氧化型維生素 C 對 Caco-2 細胞 Ferric reductase 活性 (A)及鐵蛋白含量 (B) 的影響 48
圖 3-5
不同來源的氧化型維生素 C 對 Caco-2 細胞 Ferric reductase 活性 (A)及鐵蛋白含量 (B) 的影響 49
圖 3-6
氧化型維生素 C 對 Caco-2 細胞 ferric reductase 活性 (A) 及鐵蛋白含量 (B) 的影響 50
圖 3-7
不同濃度 BSO (A) 或 BCNU (B) 處理對 Caco-2 細胞存活率之影響 51
圖 3-8
BSO 對 Caco-2 細胞 Ferric reductase 活性 (A) 及鐵蛋白含量 (B) 的影響 52
圖 3-9
BCNU 對 Caco-2 細胞 ferric reductase 活性 (A) 及鐵蛋白含量 (B) 的影響 53
圖 3-10
Quercetin 對 Caco-2 細胞 ferric reductase 活性 (A) 及鐵蛋白含量 (B) 的影響-1 54
圖 3-11
Quercetin 對 Caco-2 細胞 ferric reductase 活性 (A) 及鐵蛋白含量 (B) 的影響-2 55
圖 3-12
單寧酸對於 Caco-2 細胞 ferric reductase 活性 (A) 及鐵蛋白含量 (B) 的影響-1 56
圖 3-13
單寧酸對 Caco-2 細胞 ferric reductase 活性 (A) 及鐵蛋白含量 (B) 的影響-2 57
圖 3-14
不同鐵劑對 Caco-2 細胞 ferric reductase 活性 (A) 及鐵蛋白含量 (B) 的影響 58
dc.language.isozh-TW
dc.subjectCaco-2 細胞zh_TW
dc.subject氧化型維生素 Czh_TW
dc.subjectferric reductasezh_TW
dc.subject鐵生物利用率zh_TW
dc.subjectGluthionineen
dc.subjectiron bioavailabilityen
dc.subjectCaco-2 cellsen
dc.subjectDehydroascorbic aciden
dc.title以 Caco-2 細胞模式探討 ferric reductase 的電子來源與影響攝鐵能力之成份zh_TW
dc.titleInvestigation of the electron source for ferric reductase and compounds affecting iron absorption in a Caco-2 Cell Modelen
dc.typeThesis
dc.date.schoolyear93-2
dc.description.degree碩士
dc.contributor.oralexamcommittee江孟燦,駱菲莉,何素珍,沈立言
dc.subject.keywordCaco-2 細胞,氧化型維生素 C,ferric reductase,鐵生物利用率,zh_TW
dc.subject.keywordCaco-2 cells,Dehydroascorbic acid,Gluthionine,iron bioavailability,en
dc.relation.page77
dc.rights.note有償授權
dc.date.accepted2005-07-28
dc.contributor.author-college生命科學院zh_TW
dc.contributor.author-dept微生物與生化學研究所zh_TW
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