香椿與乳漿蛋白濃縮物對過氧化氫引起癌細胞株氧化性傷害影響之研究

Effects of Toona sinensis and whey protein concentrate on H2O2-induced oxidative damage of cancer cell lines

蔡妤姍(Yu-Shan Tsai)

指導教授 ： 蔡麗玉 ； 侯明鋒

高雄醫學大學/醫學院/醫學研究所/碩士(2003年)

摘要

越來越多的證據顯示，自由基在人類許多疾病的發生、發展中有著十分重要的角色。現代醫學發現癌症的發生與自由基的侵襲有很直接的關係，因此，如何預防免於受到自由基的傷害是一項重要課題。本研究的目的在於評估香椿(Toona sinensis)與乳漿蛋白濃縮物(whey protein concentrate)對於過氧化氫引起肺癌及乳癌細胞株氧化性傷害之影響。在本研究中，以人類肺癌及乳癌細胞株當作實驗的材料，分別以不同濃度過氧化氫及不同時間作用，評估細胞存活率，求得50 % 細胞死亡(lethal concentration 50 %，LC50 )之過氧化氫作用濃度與作用時間，作為後續過氧化氫引起氧化傷害之作用模式。之後利用此模式，分別評估香椿與乳漿濃縮蛋白對於過氧化氫引起癌細胞株氧化性傷害的影響。先將肺癌及乳癌細胞株培養於不同濃度的香椿與乳漿濃縮蛋白的培養基24小時，再個別給予30 mM/15 mM 過氧化氫溶液，作用2小時後，觀察其細胞存活率(cell viability)之變化、細胞毒性(cytotoxicity)、超氧陰離子歧化脢(superoxide dismutase，以下簡稱SOD)與穀胱甘肽還原脢(glutathine reductase，以下簡稱GRx)抗氧化酵素之活性變化、抗氧化物榖胱甘肽濃度(glutathione，以下簡稱GSH)、羰基化蛋白質(protein carbonyl)以及丙酮醛產物(malondialdehyde，以下簡稱MDA)的產量變化的情形。結果顯示:對肺癌細胞而言，給予0.01和0.05 mg/mL香椿與1和10 mg/mL乳漿蛋白濃縮物，可加強過氧化氫引起之氧化傷害，使細胞存活率下降，細胞毒性、protein carbonyl及MDA的產量增加，促使GRx活性增加，但SOD活性與GSH濃度則下降；對乳癌細胞而言，給予0.01和0.05 mg/mL香椿與10 mg/mL乳漿蛋白濃縮物，可加強過氧化氫引起之氧化傷害，使細胞密度下降，細胞毒性及MDA的產量增加，促使SOD與GRx活性及GSH濃度增加，減低羰基化蛋白質的產量。但當給予1 mg/mL乳漿蛋白濃縮物下，則會降低GSH濃度及GRx的活性。
歸納以上研究結果，我們建立一個過氧化氫作用的體外評估模式，並發現給予香椿與乳漿蛋白濃縮物，伴隨過氧化氫引起之氧化傷害時，皆會加強肺癌與乳癌細胞株的毒殺作用，但藉由抗氧化酵素活性及抗氧化物之測定結果，推測二細胞株本身之抗氧化物含量不同而對於香椿與乳漿蛋白濃縮物具有不同的反應機制。

關鍵字

氧化性傷害 ； 香椿 ； 乳漿蛋白濃縮物 ； 超氧陰離子歧化脢 ； 羰基化蛋白質 ； 榖胱甘肽

並列摘要

More and more evidences show that free radical plays important roles on human diseases. In recent medical studies, there are direct relationships between incidences of cancer and the invasion of free radical. Therefore, it is important to know how to prevent damage of free radical. In this study, we evaluated the effects of Toona sinensis and whey protein concentrate (WPC) on hydrogen peroxide(H2O2)-induced oxidative stress in lung cancer and breast cancer cell lines. To study lethal conctration of 50 % (LC50) model and used for evaluating damage effects, we used human lung-cancer and breast-cancer cell lines in different concentration of H2O2 and time. After the addition of different levels of Toona sinensis and WPC for 24 hours, the cell culture were treated with 30 mM/15 mM H2O2 for 2 hours separately, and the cell viability, the percentage of cytotoxicity, the activity of superoxide dismutase(SOD) and glutathione reductase(GRx) , glutathione(GSH), protein carbonyl and malondialdehyde(MDA) levels were assayed. Result showed that the addition of 0.01, 0.05 mg/mL Toona sinensis and 1, 10 mg/mL WPC to lung cancer cell line, it could enhance the H2O2 -induced cytotoxicity to A549 cell, decrease cell viability, SOD activity and GSH level, and elevate the protein carbonyl, MDA level and GRx activity. On the other hand, in breast cancer cell line, Toona sinensis and WPC enhanced H2O2-induced oxidative damage, decrease the cell viability and protein carbonyl, elevate the SOD, GRx activity and GSH, MDA level. When addition of 1 mg/mL WPC, GSH level and GRx activity were decreased.
In conclusion, we establish an in vitro model to study the H2O2—induced oxidative damage of A549 and MCF7 cell lines. During oxidative damage, higher concentration of Toona sinensis and WPC could enhance cytotoxicity of lung and breast cancer cell lines. Results of the antioxidant enzyme activity and antioxidant level, we suggest that two cancer cell lines have different responses to the addition of Toona sinensis or WPC.

並列關鍵字

oxidative damage ； Toona sinensis ； whey protein concentrate ； superoxide dismutase ； protein carbonyl ； glutathione

參考文獻

捌?參考文獻

1. Sykes JA, Mc Cormauk FX, O’Brien TJ: A preliminary study

of the Superoxide dismutase content of some human tumors.

Cancer Res 1978; 38: 2759-2762.

2. Serra JA, Marshoff ER, Dominguez RO, de Lustig ES, Famulari