零價鐵(ZVI)是廣泛用於淨化受污染土壤和地下水環境的材料，然而其對生命系統的影響很少被討論。先前實驗發現含有ZVI的奈米粒子在治療劑量範圍下對癌細胞具有明顯細胞毒性而正常細胞則無，且癌細胞和健康細胞間總鐵攝取量無顯著差異。奈米粒子僅在癌細胞中能快速降解並釋放鐵離子到胞內，而在正常細胞內則保留大部分為奈米粒子的形式，釋放出來的鐵離子比例很低。本研究藉由穿透式電子顯微鏡觀察口腔癌細胞(OECM-1)、抗藥性細胞株(R3)及健康細胞(HNOK)分別以ZVI @ Au NPs作用1、8、24小時後，從細胞內純化的奈米粒子結構的變化。從穿透式電子顯微鏡的影像可發現口腔癌細胞株曝露在ZVI@Au奈米粒子下，隨著時間奈米粒子會出現明顯的結構改變：從完整的圓形奈米粒子經細胞吞噬後，逐漸出現中空化的現象，甚至結構破裂的情況，然而在正常健康細胞吞噬後，奈米粒子的結構仍然保持完整。另外從元素分析的結果發現，在口腔癌細胞株吞噬後的奈米粒子的ZVI核心其鐵元素比例大幅下降，只剩下外殼的金元素訊號，顯示ZVI@Au在進入細胞後其不穩定的ZVI核心受到侵蝕轉變成鐵離子而流失，而穩定的金原子外殼不易被轉化成金離子則明顯保留其外殼的形狀；然而，正常健康細胞內的ZVI@Au奈米粒子幾乎完好無損。進一步發現加入ZVI@Au奈米粒子後的細胞內結構變化與鐵離子釋放和氧自由基的產生與代謝機制有關。我們觀察到氧自由基僅在ZVI@Au處理的OEC-M1中顯著增加，但在R3或hNOK中沒有顯著增加。抗性細胞株R3雖然仍可觀察到ZVI核心侵蝕，但R3的氧自由基代謝基因xCT，AKR1B10和AKR1C3的高表達，是讓細胞免受由ZVI @ Au暴露導致的氧化反應刺激所影響，這與hNOK中的分子機制截然不同。簡而言之， ZVI @ Au可作為ZVI的癌症選擇性和控制釋放遞送系統，本研究揭示了細胞如何通過穀胱甘肽(GSH)和煙酰胺腺嘌呤二核苷酸(NAD)代謝系統平衡ZVI攻擊的分子機制。我們相信ZVI @ Au可用作癌症選擇性鐵庫，用於未來的癌症治療，做為精準奈米醫療的重要分子標靶藥物。

In previous study, we found that ZVI containing nanoparticles (NPs) are cytotoxic to cancer cell but do not significantly affect normal cells under the proposed therapeutic dosage range. To further explore the underlying mechanisms of ZVI-based NPs in cancer-selective cytotoxicity, the aqueous synthesis method to fabricate ZVI@Au NPs and integrated approaches of structural biology and molecular biology was applied in this study. These NPs presented different structural changes in the time points of 1, 8, and 24 hours among oral cancer line (OEC-M1), ZVI resistant OEC-M1 line (R3) and healthy normal keratinocyte (HNOK) under TEM. NPs in OECM-1 and R3 showed a rapid degradation of the NPs within the first hour treatment, leaving a hollow sphere of Au shell when observed under energy dispersive X-ray spectroscopy mapping, whereas the NPs within hNOK remained almost intact. The reactive oxygen species (ROS) dramatically increased in ZVI@gold treated OEC-M1 but not in R3 or in hNOK. Through molecular biology approaches, the ZVI resistant cells, R3, still presented significant degradation of ZVI core components. The expression of xCT, AKR1B10, and AKR1C3 were elevated in R3 in response to ZVI@Au treatment compared to OEC-M1, suggesting the important roles of these genes in protecting cancer cells from ZVI@Au induced cell death. However, in hNOK, GSR and AKR1C3 expression were suppressed by ZVI@Au, while other anti-ferroptosis genes such as xCT, GPx-4, NRF2, AKR1B1, AKR1B10, and AKR1C1 were induced to highly express thus to protect hNOK from further damage. In summary, ZVI@Au may serve as a cancer-selective and controlled-release system of ZVI. This research shed a light on the molecular mechanisms for how cells balanced the ZVI challenge through glutathione and nicotinamide adenine dinucleotide metabolic system. We believe that ZVI@Au could be applied as a new class of cancer-selective iron reservoirs for advanced cancer therapy.

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