Wu_2015_Zhonghua.Yi.Xue.Za.Zhi_95_2955

OBJECTIVE: To investigate the effect of paraoxonase1 (PON1) overexpression on mouse diaphragmatic muscle cells injury caused by acute dichlorvos poisoning.
METHODS: Mouse diaphragmatic muscle cells were cultured routinely and infected with overexpression lentivirus. Cells were divided into normal control group, DDVP group, LV-GFP + DDVP group, LV-PON1 + DDVP group. Cell viability was determined by CCK-8 assay. Flow cytometry was used to detect cell apoptosis. The mRNA and protein expression of PON1 and Nrf2 in mouse diaphragmatic muscle cells was measured by RT-PCR and Western blot. Enzyme-linked immunosorbent assay was used to determine levels of acetyl cholinesterase (AchE), heme oxygenase 1 (HO-1) and quinone oxidoreductase-1 (NQO-1) in mouse diaphragmatic muscle cells. The activity of superoxide dismutase (SOD) and catalase (CAT) as well as malondialdehyde (MDA) content in cells was measured by chemical colorimetry.
RESULTS: After induced by 0, 80, 160, 320, 640 micromol/L DDVP for 24 hours, the viability of mouse diaphragmatic muscle cells was (100 +/- 3.82)%, (82.13 +/- 2.60)%, (53.57 +/- 5.05)%, (30.77 +/- 3.30)%, (14.20 +/- 2.19)% respectively, changing in a concentration-dependent manner (P < 0.05). After induced by 160 micromol/L DDVP for 0, 6, 12, 24 hours, the viability of mouse diaphragmatic muscle cells was (100.17 +/- 2.74)%, (76.13 +/- 6.01)%, (66.53 +/- 3.55)%, (53.57 +/- 5.05)%, changing in a time-dependent manner (P < 0.05). The PON1 protein level in LV-PON1 group was higher than that of blank control group (0.370 +/- 0.015 vs 0.232 +/- 0.004, 0.197 +/- 0.015 vs 0.037 +/- 0.003, P < 0.05). The cell viability of LV-PON1 group is higher than that of DDVP group at different time point after induction of DDVP (P < 0.05). After induced by DDVP for 24 hours, the cell apoptosis rate and MDA content in LV-PON1 group were lower than those of DDVP group (P < 0.05). While levels of AchE, PON1 and Nrf2 protein expression, SOD and CAT, HO-1 and NQO-1 were higher than those of DDVP group (P < 0.05).
CONCLUSIONS: The overexpression of PON1 could effectively alleviate AchE inhibition by DDVP and induce Nrf2 expression to exert antioxidant effect, thus protected the mouse diaphragmatic muscle cells.