TCA循环中间体通过调节Nrf2/NQO1信号轴减轻邻苯二甲酸二酯（2-乙基己基）诱导的胆汁淤积性肝损伤

TCA Cycle Intermediate Mitigates Di(2-ethylhexyl) Phthalate-Induced Cholestatic Liver Injury Through Modulation of the Nrf2/NQO1 Signalling Axis

TCA循环中间体通过调节Nrf2/NQO1信号轴减轻邻苯二甲酸二酯（2-乙基己基）诱导的胆汁淤积性肝损伤

Authors: Yue Jiang, Fang Xie, Xutao Ling, Jiayi Zhang, Yun Yu, Qianqian Huang, Lun Zhang, Lu Ye, Wenkang Tao, Mengzhen Hou, Cheng Zhang, Jianqing Wang

Source: Basic & Clinical Pharmacology & Toxicology

PMID: 40370325

DOI: 10.1111/bcpt.70047 

Abstract

As a commonly used phthalate compound, di(2-ethylhexyl) phthalate (DEHP) has been shown to disrupt the tricarboxylic acid (TCA) cycle and aggravate tissue damage. However, whether the TCA cycle is involved in cholestatic liver injury (CLI) induced by DEHP and the protective effect of dimethyl fumarate (DMF), which is used to supplement TCA intermediate metabolites, remained unclear. Here, mice were randomized into five groups (n = 6/group): (1) Control, (2) DEHP (200 mg/kg/day), (3) DMF (100 mg/kg/day), (4) DEHP + DMF (30 mg/kg/day) and (5) DEHP + DMF (100 mg/kg/day). Our data demonstrated that DEHP exposure upregulated total bile acid (TBA) levels and broke the TCA cycle, resulting in reduced fumaric acid and malic acid. However, we further supplemented fumaric acid with DMF and found that DMF effectively reversed the high levels of TBA, alkaline phosphatase (ALP) and glutamyl transpeptidase (GGT) induced by DEHP in mice. Meanwhile, pathological results in the liver showed that DMF improved bile duct cell damage, inflammatory cell infiltration, collagen deposition and necrosis caused by DEHP. In addition, we found that DEHP elevated the level of interleukin (IL)-1β, IL-6, TNF-α and MDA and decreased the level of SOD in the mouse liver, which was effectively reversed by DMF treatment. Besides, DMF upregulated the expression of Nrf2 and NQO1 in the liver of DEHP-exposed mice. For in vitro validation, AML-12 cells were treated with (1) Control, (2) DEHP (250 μM), (3) DEHP + DMF (10 μM), (4) DEHP + DMF (25 μM) and (5) DEHP + DMF (50 μM). DEHP exposure increased the expression of IL-1β, IL-6 and TNF-α, which was mitigated by DMF, while ML385, an Nrf2 inhibitor, could counteract the anti-inflammatory effects of DMF. These findings indicate that DEHP broke the TCA cycle of the mouse liver, and DMF supplementation protects against DEHP-induced CLI by activating the Nrf2/NQO1 pathway.

Keywords: TCA cycle; cholestasis liver injury; di(2‐ethylhexyl) phthalate; dimethyl fumarate; inflammation.

摘要

作为一种常用的邻苯二甲酸酯化合物，二（2-乙基己基）邻苯二甲酸酯（DEHP）已被证实会扰乱三羧酸（TCA）循环，并加剧组织损伤。然而，关于 TCA 循环是否参与由 DEHP 引发的胆汁淤积性肝损伤（CLI）以及用于补充 TCA 中间代谢物的二甲基富马酸（DMF）的保护作用，目前仍不清楚。在此，我们将小鼠随机分为五组（每组 6 只）：（1）对照组，（2）DEHP 组（200 毫克/千克/天），（3）DMF 组（100 毫克/千克/天），（4）DEHP + DMF 组（30 毫克/千克/天），（5）DEHP + DMF 组（100 毫克/千克/天）。我们的数据表明，DEHP 暴露会上调总胆汁酸（TBA）水平，并破坏 TCA 循环，导致富马酸和苹果酸水平降低。然而，我们进一步用 DMF 补充富马酸，并发现 DMF 能有效逆转 DEHP 引起的小鼠体内高 TBA、碱性磷酸酶（ALP）和谷氨酰转肽酶（GGT）的水平。同时，肝脏的病理结果表明，DMF 改善了由 DEHP 引起的胆管细胞损伤、炎症细胞浸润、胶原沉积和坏死情况。此外，我们还发现，DEHP 会提高小鼠肝脏中白细胞介素（IL）-1β、IL-6、TNF-α 和丙二醛（MDA）的水平，并降低超氧化物歧化酶（SOD）的水平，而这些变化通过 DMF 治疗得以有效逆转。此外，DMF 还上调了 DEHP 暴露小鼠肝脏中 Nrf2 和 NQO1 的表达。对于体外验证，对 AML-12 细胞进行了以下处理：（1）对照组，（2）DEHP（250 μM），（3）DEHP + DMF（10 μM），（4）DEHP + DMF（25 μM）和（5）DEHP + DMF（50 μM）。DEHP 暴露会增加 IL-1β、IL-6 和 TNF-α 的表达，而 DMF 可以缓解这一现象，而 Nrf2 抑制剂 ML385 则可以抵消 DMF 的抗炎作用。这些发现表明，DEHP 扰乱了小鼠肝脏的 TCA 循环，而补充 DMF 可通过激活 Nrf2/NQO1 通路来防止 DEHP 引起的胆汁淤积性肝损伤。

关键词：TCA 循环；胆汁淤积性肝损伤；二（2-乙基己基）邻苯二甲酸酯；二甲基富马酸；炎症。