三(2-氯乙基)磷酸酯诱导肝细胞线粒体毒性的研究

张文娟; 张佑健; 王志远; 殷文军; 徐甜; 郑红燕; 熊伟; 袁晶

doi:10.13213/j.cnki.jeom.2015.15156

三(2-氯乙基)磷酸酯诱导肝细胞线粒体毒性的研究

Tris (2-chloroethyl) phosphate Induced Mitochondrial Toxicity in Hepatocytes

摘要

摘要:
目的研究三(2-氯乙基)磷酸酯tris(2-chloroethyl)phosphate, TCEP对肝细胞线粒体功能的影响。

方法用0.00(溶剂对照组)、3.12、12.50、50.00 和200.00 mg/L TCEP 分别处理人正常肝细胞(L02 细胞)和人肝癌细胞(HepG2 细胞)24 h 和48 h。测定细胞活力、线粒体内活性氧水平、线粒体DNA拷贝数、线粒体膜电位和胞内游离钙(Ca²⁺)水平以及胞内ATP 的浓度。

结果与相应溶剂对照组相比, 在24 h, 200.00 mg/L TCEP 处理组L02 细胞活力降低(P <0.05), ≥ 12.50 mg/L TCEP 处理组HepG2 细胞活力降低(P < 0.05);在48 h, 50.00 mg/L 和200.00 mg/L TCEP 处理组两种细胞活力降低(P < 0.05)。所有TCEP 处理组两种细胞的线粒体DNA拷贝数均减少(P < 0.05 或P < 0.01)。在24 h, 所有TCEP处理组两种细胞的线粒体膜电位均下降(P < 0.05 或P < 0.01)。在24 h, 200.00 mg/L TCEP 处理组两种细胞胞内游离Ca²⁺ 水平均升高(P < 0.01);在48 h, 50.00 mg/L和200.00 mg/L TCEP处理组两种细胞胞内游离Ca²⁺ 均升高(P < 0.01)。在24 h 和48 h, 200.00 mg/L TCEP 处理组L02 胞内ATP 水平降低(P < 0.05);在24 h, 50.00 mg/L 和200.00 mg/L TCEP 处理组HepG2 胞内ATP 浓度下降(P < 0.05 或P < 0.01)。

结论一定浓度的TCEP 可致肝细胞线粒体损伤和线粒体功能指标异常, 提示TCEP 有肝细胞线粒体毒性。

Abstract:
Objective To evaluate effects of tris(2-chloroethyl)phosphate (TCEP) on mitochondrial functions in liver cells.

Methods At 24 and 48 h after L02 and HepG2 cells were treated with 0.00 (solvent control group), 3.12, 12.50, 50.00, 200.00 mg/L TCEP, we detected cell viability, mitochondrial reactive oxygen species (mtROS) levels, mitochondrial DNA copy numbrane, mitochondrial membrane potential, intracellular free Ca²⁺ levels, and intracellular ATP level.

Results Compared with the control group, TCEP decreased the cell viabilities of L02 cells in the 200 mg/L TCEP group (P < 0.05) and the cell viabilities of HepG2 cells in the ≥ 12.50 mg/L TCEP groups (P < 0.05) at 24 h; decreased the cell viabilities of L02 and HepG2 cells in the 50.00 and 200.00 mg/L TCEP groups at 48 h (P < 0.05); reduced the mitochondrial DNA number of L02 and HepG2 cells in all treatment groups (P < 0.05 or P < 0.01); decreased the mitochondrial membrane potential of L02 or HepG2 cells in all treatment groups at 24 h (P < 0.05 or P < 0.01); increased the intracellular free Ca²⁺ concentrations of L02 and HepG2 cells only in the 200.00 mg/L TCEP group at 24 h and in the 50.00 and 200.00 mg/L groups at 48 h (P < 0.01); decreased the intracellular ATP levels of L02 cells only in the 200.00 mg/L TCEP group at 24 and 48 h (P < 0.05) and in the 50.00 and 200.00 mg/L TCEP groups of HepG2 cells at 24 and 48 h (P < 0.05 or P < 0.01).

Conclusion A certain concentrations of TCEP could cause mitochondrial damage and abnormal changes in indices reflecting mitochondrial dysfunctions, implying that TCEP could induce mitochondrial toxicity in hepatocytes.

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