The First Dongguan Affiliated Hospital, Guangdong Medical University, Dongguan 523808, China; Dongguan Key Laboratory of Environmental Medicine, School of Public Health, Guangdong Medical University, Dongguan 523808, China.
Dongguan Key Laboratory of Environmental Medicine, School of Public Health, Guangdong Medical University, Dongguan 523808, China.
Toxicol Lett. 2023 Jan 15;373:132-140. doi: 10.1016/j.toxlet.2022.11.014. Epub 2022 Nov 25.
Aflatoxin B1 (AFB1) is a human procarcinogen known to be activated by cytochrome P450 (CYP) 1A2 and 3A4. In a previous study AFB1 caused chromosomal rearrangement in a yeast strain genetically engineered for stably expressing human CYP1B1. Yet, further verification of the effect of AFB1 in human cells, a potential role of the aryl hydrocarbon receptor (AhR), and CYP1B1-catalyzed AFB1 metabolism remain unidentified. In this study, a human hepatocyte (L-02) line and a human lymphoblastoid (TK6) cell line were genetically engineered for the expression of human CYP1B1, producing L-02-hCYP1B1 and TK6-hCYP1B1, respectively. They were exposed to AFB1 and analyzed for the formation of micronucleus and elevation of γ-H2AX (indicating double-strand DNA breaks); the metabolites formed by CYP1B1 from AFB1 after incubation of AFB1 with human CYP1B1 isoenzyme microsomes were determined by LC-MS. The results showed significantly more potent induction of micronucleus by AFB1 in L-02-hCYP1B1 and TK6-hCYP1B1 than in the parental (L-02 and TK6) cells, and the effects were reduced by (E)- 2,3',4,5'-tetramethoxystilbene, a specific CYP1B1 inhibitor. In the AFB1- CYP1B1 microsomes incubations AFM1, a known stable metabolite of AFB1, was detected. Moreover, in L-02 and TK6 cells, AFB1 apparently increased the protein levels of AhR, ANRT and CYP1B1, and caused the nuclear translocation of AhR and ARNT, the latter effect being blocked by BAY-218 (an inhibitor of AhR). In conclusion, this study indicates that human CYP1B1 is capable of metabolically activating AFB1 through the AhR signaling pathway.
黄曲霉毒素 B1（AFB1）是一种已知的人类前致癌物，可被细胞色素 P450（CYP）1A2 和 3A4 激活。在之前的一项研究中，AFB1 导致了一株经过基因工程改造以稳定表达人 CYP1B1 的酵母菌株的染色体重排。然而，AFB1 在人细胞中的作用、芳香烃受体（AhR）的潜在作用以及 CYP1B1 催化的 AFB1 代谢物仍未得到进一步验证。在这项研究中，一条人肝细胞（L-02）系和一条人淋巴母细胞（TK6）系被基因工程改造以表达人 CYP1B1，分别产生 L-02-hCYP1B1 和 TK6-hCYP1B1。它们被暴露于 AFB1 中，并分析微核的形成和γ-H2AX 的升高（表示双链 DNA 断裂）；通过 LC-MS 确定 CYP1B1 从 AFB1 孵育后的 AFB1 代谢物。结果表明，AFB1 在 L-02-hCYP1B1 和 TK6-hCYP1B1 中的微核诱导作用明显强于亲本（L-02 和 TK6）细胞，并且（E）-2,3',4,5'-四甲氧基二苯乙烯的作用降低，一种特异性 CYP1B1 抑制剂。在 AFB1-CYP1B1 微粒体孵育中，检测到 AFM1，一种已知的 AFB1 稳定代谢物。此外，在 L-02 和 TK6 细胞中，AFB1 明显增加了 AhR、ANRT 和 CYP1B1 的蛋白水平，并导致 AhR 和 ARNT 的核转位，后一效应被 BAY-218（AhR 抑制剂）阻断。总之，这项研究表明，人 CYP1B1 能够通过 AhR 信号通路代谢激活 AFB1。
