Institute for Oncology and Radiology of Serbia, Belgrade, Serbia.
VINČA Institute of Nuclear Sciences, National Institute of the Republic of Serbia, University of Belgrade, Mike Petrovića Alasa 12-14, 11000 Belgrade, Serbia.
J Trace Elem Med Biol. 2023 Dec;80:127286. doi: 10.1016/j.jtemb.2023.127286. Epub 2023 Aug 22.
Although luteolin has been confirmed as potent anticancer agent, its potential application as therapeutic is limited by its water solubility. To overcome this shortcoming nanoparticle technology approach was applied. Owing to their proven low toxicity and the possibility to be easily functionalized gold nanoparticles (AuNP) were the nanosystem of choice used in this study. Novel luteolin capped gold nanoparticles (AuNPL) were synthesized and their anticancer effect towards human cervical adenocarcinoma HeLa cells was investigated in vitro.
AuNPL were synthesized by reducing chloroauric acid by trisodium citrate with subsequent addition of luteoline during synthesis and their physicochemical characterization was done. AuNPL cytotoxicity against HeLa, human malignant melanoma A375, and normal human keratinocytes HaCaT cells was tested by MTT cell survival assay, and their IC values were determined. The capability of AuNPL to induce cell cycle arrest and apoptosis in HeLa cells were demonstrated by flow cytometry. The antioxidant activity of AuNPL was assessed by DPPH and ABTS scavenging assays. Cytoprotective properties of AuNPL towards HaCaT cells were examined by measuring the physiological and HO induced intracellular reactive oxygen species (ROS) levels using flow cytometry. Also, genotoxicity of AuNPL in HaCaT cells was investigated by the single cell alkaline comet assay.
Spherical AuNPL, stable in aqueous solution up to six months at 4 °C were obtained in the synthesis. The selectivity in the cytotoxic action of AuNPL on HeLa and A375 cancer cells compared with their cytotoxicity on normal keratinocytes HaCaT was observed. AuNPL exerted their cytotoxic activity against HeLa cells through accumulation of the cells in the subG1 phase of the cell cycle, inducing the apoptotic cell death mediated by the activation of caspase-3 - 8, and - 9. AuNPL antioxidative potential was confirmed by DPPH and ABTS scavenging assays. IC concentration of AuNPL exerted cytoprotective effect against HaCaT cells by the significant reduction of the physiological intracellular ROS level. Additionally, AuNPL were shown as more cytoprotective towards HaCaT cells then luteolin due to the more successful elimination of HO induced intracellular ROS. Moreover, nontoxic concentrations of AuNPL did not cause considerable DNA damage of HaCaT cells, indicating low genotoxicity of the nanoparticles.
Synthesized AuNPL showed selective cytotoxic activity against HeLa cells, while being nontoxic and cytoprotective against HaCaT cells. The observed findings encourage further investigation of AuNPL as a promising novel anticancer agent.
尽管木犀草素已被证实为有效的抗癌剂，但由于其水溶性较差，其作为治疗药物的应用受到限制。为了克服这一缺点，采用了纳米技术方法。由于其低毒性已得到证实，并且易于功能化，因此金纳米粒子（AuNP）是本研究中选择的纳米系统。合成了新型木犀草素包覆的金纳米粒子（AuNPL），并在体外研究了其对人宫颈腺癌 HeLa 细胞的抗癌作用。
通过三氯化金被柠檬酸钠还原来合成 AuNPL，然后在合成过程中加入木犀草素，对其进行理化特性表征。通过 MTT 细胞存活测定法检测 AuNPL 对 HeLa、人恶性黑色素瘤 A375 和正常人角质形成细胞 HaCaT 的细胞毒性，并确定其 IC 值。通过流式细胞术证明 AuNPL 诱导 HeLa 细胞周期停滞和凋亡的能力。通过 DPPH 和 ABTS 清除试验评估 AuNPL 的抗氧化活性。通过流式细胞术测量生理和 HO 诱导的细胞内活性氧（ROS）水平来检测 AuNPL 对 HaCaT 细胞的细胞保护作用。还通过单细胞碱性彗星试验研究了 AuNPL 在 HaCaT 细胞中的遗传毒性。
在合成中获得了稳定的球形 AuNPL，在 4°C 下在水溶液中可稳定保存长达六个月。观察到 AuNPL 对 HeLa 和 A375 癌细胞的细胞毒性选择性高于其对正常角质形成细胞 HaCaT 的细胞毒性。AuNPL 通过将细胞积累在细胞周期的 subG1 期，激活 caspase-3-8 和 -9，诱导细胞凋亡死亡，对 HeLa 细胞发挥细胞毒性作用。DPPH 和 ABTS 清除试验证实了 AuNPL 的抗氧化潜力。AuNPL 的 IC 浓度通过显著降低生理细胞内 ROS 水平对 HaCaT 细胞发挥细胞保护作用。此外，由于 AuNPL 更成功地消除了 HO 诱导的细胞内 ROS，因此 AuNPL 对 HaCaT 细胞的细胞保护作用比木犀草素更强。此外，非毒性浓度的 AuNPL 不会对 HaCaT 细胞造成明显的 DNA 损伤，表明纳米粒子的遗传毒性较低。
合成的 AuNPL 对 HeLa 细胞表现出选择性的细胞毒性，而对 HaCaT 细胞则无毒且具有细胞保护作用。观察到的结果鼓励进一步研究 AuNPL 作为一种有前途的新型抗癌剂。
