以 Caco-2 細胞模式探討 Hepcidin 對鐵吸收相關分子表現之影響

Effect of Hepcidin On The Expression of Iron-Related Protein in Caco-2 Cells

李慧玲(Heui-Ling Li)

指導教授 ： 蕭寧馨

國立臺灣大學/生命科學院/微生物與生化學研究所/碩士(2004年)

https://doi.org/10.6342/NTU.2004.01688

摘要

Hepcidin 為近年發現肝臟分泌具抗菌性之小分子胜肽，可能為傳遞鐵需求訊息給小腸細胞之傳訊分子，而具有調節鐵吸收之荷爾蒙角色。不過，由於缺乏大量純化 hepcidin 活性蛋白質來源，目前的研究主要集中於 mRNA 的表現層次，難以確認其蛋白質分子的作用機制。因此本篇研究的目的為利用人類大腸癌細胞株 Caco-2 之細胞模式，探討化學合成之 hepcidin 的活性以及與細胞鐵吸收相關分子表現之關係。以化學合成胺基酸序列為 20 與 25 之 hepcidin (Hepc20、Hepc25)，於 cysteine/cystine 緩衝液中進行氧化反應，形成分子內雙硫鍵，用於處理 Caco-2 細胞。在此也建立 Caco-2 細胞鐵吸收相關背景資料，探討鐵營養狀況、細胞分化、培養皿構造及培養血清濃度對鐵吸收相關分子表現之影響，並決定以 Transwell 之方式培養 Caco-2 細胞，於細胞分化 14 ∼ 16 天時，改變 basolateral 端為 2% FBS-DMEM 培養兩天後，分別添加不同濃度之 Hepc20、Hepc25 處理不同時間，以探索化學合成 hepcidin 之作用。以 10

關鍵字

Caco-2 cell ； Hepcidin ； iron transporter

並列摘要

Hepcidin is a small antimicrobial peptide specifically produced in the liver. It has been suggested to be the signal of hepatic iron stores that regulates intestinal iron absorption, and may be an iron-regulatory hormone. Due to lack of purified active hepcidin peptides, present studies mainly focused on regulation of hepcidin mRNA expression, while cellular and molecular mechanisms of hepcidin functions are still lacking. Therefore, the aim of this study was to investigate biological activity of synthetic hepcidin and its effects on iron transporters expression in Caco-2 cells, a human intestinal cell line. Hepc20 and Hepc25, composed of 20 and 25 amino acid residues, respectively, were synthesized by solid-phase peptide synthesis, oxidized in the presence of the cysteine/cystine redox system to molecules containing four disulfide bonds, and purified with reverse-phase HPLC. The effects of iron status, cell differentiation, plate structure and serum concentration on the expression of DMT1, Ireg1, HFE and TfR in Caco-2 cells were first verified. For hepcidin treatment, Caco-2 cells were cultured in 10% FBS-DMEM and grown in 24-mm-diameter Transwell bicameral chambers with 3.0

並列關鍵字

Hepcidin ； Caco-2 cell ； iron transporter

參考文獻

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