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Role of reactive oxygen species-mitochondrial pathway in sodium arsenite-induced apoptosis of L-02 cells / 中华地方病学杂志
Chinese Journal of Endemiology ; (12): 4-9, 2020.
Article in Chinese | WPRIM | ID: wpr-866053
ABSTRACT

Objective:

To explore the mechanism of apoptosis induced by sodium arsenite (NaAsO 2) in human hepatic cells (L-02) through reactive oxygen species (ROS) accumulation and mitochondrial dysfunction, and provide experimental evidence for the mechanism of arsenic poisoning.

Methods:

L-02 cells were divided into control group, NaAsO 2 group (10 μmol/L NaAsO 2), N-acetylcysteine (NAC) group (5 mmol/L NAC), and NaAsO 2 + NAC group (10 μmol/L NaAsO 2, 5 mmol/L NAC), and were cultured in vitro for 24 h. The intracellular ROS level, mitochondrial membrane potential depolarization ratio and cell apoptosis rate were measured by dichlorodihydrofluorescein diacetate (DCFH-DA) fluorescence probe, JC-1 staining and Annexin V-FITC/PI double staining, respectively; the mRNA and the protein of Caspase 3, cytochrome C (Cyt-C) and cytochrome C oxidaseⅣ (COXⅣ) were detected by real time fluorescence quantitative PCR (qRT-PCR) and Western blotting, respectively.

Results:

There were statistically significant differences in intracellular ROS levels (3 857 392.33 ± 44 928.39, 4 515 288.00 ± 32 660.64, 3 670 150.67 ± 101 987.69, 4 035 235.67 ± 99 995.30), mitochondrial membrane potential depolarization ratios (2.16 ± 0.54, 7.95 ± 0.52, 2.70 ± 0.29, 1.01 ± 0.23) and total apoptosis rates (1.45 ± 0.03, 4.27 ± 0.17, 1.87 ± 0.12, 2.52 ± 0.35) between groups ( F = 62.62, 159.81, 112.70, P < 0.05). There were statistically significant differences in Caspase 3, Cyt-C, COXⅣ mRNA expression levels ( F = 9.20, 7.33, 14.87, P < 0.05) and in cleaved-Caspase 3, Cyt-C, COXⅣ protein expression levels( F = 31.42, 8.01, 83.30, P < 0.05) between groups. Compared with the control group, the intracellular ROS level, mitochondrial membrane potential depolarization ratio and total apoptosis rate were significantly increased ( P < 0.05); Caspase3, Cyt-C mRNA and protein expression levels were significantly increased ( P < 0.05), and COXⅣ mRNA and cleaved-Caspase 3, Cyt-C protein expression levels were significantly decreased ( P < 0.05) in NaAsO 2 group. Compared with the NaAsO 2 group, the intracellular ROS level, mitochondrial membrane potential depolarization ratio and total apoptosis rate of NaAsO 2 + NAC group were significantly decreased ( P < 0.05); the Caspase3, Cyt-C mRNA and cleaved-Caspase 3, Cyt-C protein expression levels were significantly decreased ( P < 0.05), the COX Ⅳ mRNA and protein expression levels were significantly increased ( P < 0.05).

Conclusions:

NaAsO 2 stimulates L-02 cells to produce excessive ROS, which induces mitochondrial depolarization and further triggers mitochondrial damage, resulting in increased release of Cyt-C and activation of the mitochondrial apoptosis pathway that Caspase 3 protein induces apoptosis in L-02 cells, which may be one of the main mechanisms of arsenic-induced liver injury.
Full text: Available Index: WPRIM (Western Pacific) Language: Chinese Journal: Chinese Journal of Endemiology Year: 2020 Type: Article

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Full text: Available Index: WPRIM (Western Pacific) Language: Chinese Journal: Chinese Journal of Endemiology Year: 2020 Type: Article