ABSTRACT
Objective:
To observe the
role of nuclear factor erythroid 2-related factor 2 (Nrf2) in regulating
apoptosis during malignant transformation of
human bronchial
epithelial cells (HBE
cells) induced by
sodium arsenite (NaAsO 2).
Methods:
HBE
cells were treated with 0.0 and 1.0 μmol/L NaAsO 2, which were
control group and
arsenic exposed group respectively. HBE
cells were treated with 1.0 μmol/L NaAsO 2 for 43 passages to establish a malignant transformation model. The dynamic changes of indexes in different passages (0, 1st, 8th, 15th, 22nd, 29th, 36th, and 43rd) after exposure to NaAsO 2 were monitored, including the
apoptosis rate detected by
flow cytometry and
apoptosis-related
proteins and Nrf2
protein detected by
Western blotting. Nrf2
siRNA was transfected into malignant transformed HBE
cells (T-HBE
cells) to silence Nrf2. The silencing effect of Nrf2
protein was verified. And, the
apoptosis rate and
apoptosis-related
proteins were detected.
Results:
With the increase of
arsenic exposure, the
apoptosis rates of HBE
cells decreased (0, 1, 8, 15, 22, 29, 36 and 43 passages were 0.370 ± 0.029, 0.443 ± 0.069, 0.357 ± 0.046, 0.330 ± 0.016, 0.273 ± 0.050, 0.160 ± 0.024, 0.110 ± 0.022, 0.097 ± 0.012, respectively, Ftrend = 22.981, P < 0.05). Compared with the 0 passage
cells, the
apoptosis rates of the 22nd, 29th, 36th and 43rd passages in the arsenite group were lower. The differences between them were statistically significant ( P < 0.05). With the increase of
arsenic exposure, the expressions of
pro-apoptotic proteins caspase-3, cleaved-
caspase-3, C/EBP-homologous
protein (CHOP) and
B-cell lymphoma-2 (Bcl-2) associated X
protein (Bax) showed downward
trends ( Ftrend = 22.356, 3.738, 6.130, 8.061, P < 0.05), while the
anti-apoptotic proteins myeloid cell leukemia 1
protein (Mcl-1) and Bcl-2 showed upward
trends ( Ftrend = 58.201, 7.691, P < 0.05). Compared with the 0 passage and the
control group of the same passage, from the 22nd passage of
caspase-3, cleaved-
caspase-3, from the 15th passage of CHOP, Mcl-1, and Bcl-2, from the 29th passage of Bax in the arsenite group, the differences of
protein were statistically significant ( P < 0.05). However, there were no significant differences in
caspase-8, cleaved-
caspase-8,
caspase-12 and cleaved-
caspase-12 protein expressions in the
arsenic group ( P > 0.05). Compared with the 0 passage and the
control group of the same passage, from the 8th passage of Nrf2
proteins in the arsenite group, the differences of expressions were statistically significant ( P < 0.05). Compared with T-HBE
cells transfected with Con
siRNA (control), the
apoptosis rate of T-HBE
cells transfected with Nrf2
siRNA was higher ( P < 0.05). Compared with T-HBE
cells transfected with Con
siRNA, the expression levels of Nrf2, Bcl-2 and Mcl-1 in T-HBE
cells transfected with Nrf2
siRNA were lower ( P < 0.05), while the expression levels of cleaved-
caspase-3/
caspase-3,
caspase-3, cleaved-
caspase-3, CHOP, and Bax were higher ( P < 0.05).
Conclusion:
Nrf2 may regulate mitochondrial apoptotic pathway through Bcl-2, Mcl-1 and Bax, and
endoplasmic reticulum apoptotic pathway through CHOP, so as to inhibit the
apoptosis of HBE
cells and participate in the process of malignant transformation of HBE
cells induced by NaAsO 2.