College of Materials Science and Chemical Engineering, Zhejiang University, 38 Zheda Road, Zhejiang, PR China.
Food Chem Toxicol. 2010 May;48(5):1145-51. doi: 10.1016/j.fct.2010.02.001. Epub 2010 Feb 4.
Ethanol-induced liver injury has been extensively reported in clinic, but still lacks an efficient in vitro platform for investigating its hepatotoxicity and protectants. This study aimed to establish a methodology on the culture conditions regarding the sealability against evaporation of ethanol, culture medium and 2D/3D culture of hepatocytes. Based on the experimental findings, it was indicated that the ethanol evaporation from culture plates was a severe problem reducing its toxicity in hepatocyte. According to the detected ethanol toxic response marked by reduced cell viability, 3D cultured hepatocytes in gel entrapment were suggested to be better than 2D hepatocyte in monolayer, but the cultures in either William's Medium E or DMEM exhibited comparable sensitivity to ethanol toxicity. Subsequently, 3D cultured hepatocytes with Parafilm sealing were systematically illustrated to well reflect the ethanol-induced lipid accumulation, reactive oxygen species/malondialdehyde generation, glutathione depletion and cytochrome 2E1 induction. Finally, such hepatocyte models were proposed as a platform for screening of herbal component against ethanol hepatotoxicity. Nano-silibinin, for the first time, found to perform significant protection against ethanol-induced hepatotoxicity while silibinin in normal particles could not inhibit such toxicity. This protection of nano-silibinin might relate to its improved bioavailability compared to normal insoluble silibinin and could act as an anti-oxidative and anti-steatosis agent against ethanol-induced hepatotoxicity.
乙醇诱导的肝损伤在临床上已有广泛报道，但仍缺乏有效的体外研究平台来研究其肝毒性和保护剂。本研究旨在建立一种关于培养条件的方法学，以解决乙醇、培养基的蒸发密封问题和肝细胞的 2D/3D 培养。根据实验结果，表明培养板中乙醇的蒸发是一个严重的问题，会降低其对肝细胞的毒性。根据细胞活力降低等乙醇毒性反应的检测结果，建议采用凝胶包埋的 3D 培养肝细胞优于单层 2D 肝细胞，但无论是在威廉氏培养基 E 中还是在 DMEM 中培养，细胞对乙醇毒性的敏感性相当。随后，采用 Parafilm 密封的 3D 培养肝细胞系统地说明其能很好地反映乙醇诱导的脂质积累、活性氧/丙二醛生成、谷胱甘肽耗竭和细胞色素 2E1 诱导。最后，将这些肝细胞模型作为一种筛选草药成分对抗乙醇肝毒性的平台。首次发现纳米水飞蓟素对乙醇诱导的肝毒性具有显著的保护作用，而普通颗粒状水飞蓟素则不能抑制这种毒性。纳米水飞蓟素的这种保护作用可能与其改善的生物利用度有关，与不溶性水飞蓟素相比，纳米水飞蓟素可作为一种抗氧化和抗脂肪变性剂，对抗乙醇诱导的肝毒性。
