Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan.
Faculty of Pharmaceutical Sciences, Osaka University, Osaka, Japan.
Toxicol Lett. 2022 Nov 1;370:1-6. doi: 10.1016/j.toxlet.2022.09.005. Epub 2022 Sep 12.
Exposure of humans to aflatoxin B1 (AFB1) via ingestion of contaminated agricultural products is a major concern for human health throughout the world because epoxidized AFB1, biotransformed from AFB1 by hepatic CYP3A4, is strongly hepatotoxic and hepatocarcinogenic. Intestinal epithelial cells serve as a physical and physiological barrier against xenobiotics via their intercellular tight junction (TJ) seals and the metabolizing enzyme CYP3A4. However, the effect of AFB1 on the intestinal barrier remains unclear. Here, we investigated the influence of AFB1 on these physical and physiological intestinal barriers by means of an in vitro human intestinal model utilizing doxycycline-inducible CYP3A4-expressing Caco-2 cells, in which CYP3A4 activity is comparable to that in the adult human intestine. Cellular toxicity of AFB1 in induced Caco-2 cells (i.e., cells in which expression of CYP3A4 is induced by doxycycline) was approximately 5 times that of uninduced Caco-2 cells. Exposure to 16 µM AFB1 did not decrease the transepithelial electric resistance (TEER; a measure of TJ barrier integrity) in monolayers of uninduced Caco-2 cells to 95.8 % of that in vehicle-treated cells; in contrast, in induced Caco-2 cells, TEER was reduced to 28.8 %. Exposure to 16 µM AFB1 increased paracellular permeation of 4- and 20-kDa dextrans (paracellular permeation markers) through monolayers of induced Caco-2 cells to 5.4 and 5.2 times that through uninduced Caco-2 cells. These results together show that ingested AFB1 can modulate the intestinal barrier, and that inducible CYP3A4-expressing Caco-2 cells are a promising tool for evaluating the safety of food contaminants in the human intestine.
人类通过摄入受污染的农产品接触黄曲霉毒素 B1（AFB1）是全世界关注的主要健康问题，因为通过肝 CYP3A4 生物转化的环氧化 AFB1 具有很强的肝毒性和致癌性。肠上皮细胞通过细胞间紧密连接（TJ）密封和代谢酶 CYP3A4 作为对抗异源生物的物理和生理屏障。然而，AFB1 对肠道屏障的影响尚不清楚。在这里，我们通过利用强力霉素诱导表达 CYP3A4 的 Caco-2 细胞的体外人肠模型研究了 AFB1 对这些物理和生理肠屏障的影响，其中 CYP3A4 活性与成人肠道中的活性相当。AFB1 在诱导的 Caco-2 细胞（即通过强力霉素诱导表达 CYP3A4 的细胞）中的细胞毒性大约是未诱导的 Caco-2 细胞的 5 倍。暴露于 16μM AFB1 并未将未诱导的 Caco-2 细胞单层中的跨上皮电阻（TEER；TJ 屏障完整性的度量）降低至对照处理细胞的 95.8%；相比之下，在诱导的 Caco-2 细胞中，TEER 降低至 28.8%。暴露于 16μM AFB1 增加了 4-和 20kDa 葡聚糖（旁细胞渗透标志物）通过诱导的 Caco-2 细胞单层的旁细胞渗透，至未诱导的 Caco-2 细胞的 5.4 和 5.2 倍。这些结果共同表明，摄入的 AFB1 可以调节肠道屏障，并且诱导表达 CYP3A4 的 Caco-2 细胞是评估人类肠道中食物污染物安全性的有前途的工具。
