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[The cytotoxicity and expression changes of endoplasmic reticulum related genes induced by MWCNTs with different
Jian Guo1, Hai-yu Nie, Hai-fang Wang
1Department of Occupational and Environmental Health, Peking University School of Public Health, Beijing 100191, China.
Objective:
To compare the cytotoxicity of RAW264.7 cells induced by two types of multi-walled carbon nanotubes (MWCNTs) with different surface modifications (MWCNTs modified by taurine and MWCNTs treated by acid), and explore the role of endoplasmic reticulum (ER) in MWCNTs-induced apoptosis.
Methods:
RAW264.7 cells were exposed to tau-MWCNTs or c-MWCNTs, of which the diameters and impurity contents were the same, at the dose of 0, 1.56, 3.12, 6.25, 12.50, 25.00 μg/cm(2) for 3, 6, 12 and 24 h, respectively. Then the cytotoxicity of MWCNTs was determined by WST-1 assay, and the percentages of apoptosis were analyzed via flow cytometry with AnnexinV-FITC/PI label. Real-time PCR was used to detect mRNA expression levels of CRT, GRP78 and CHOP genes which were related to ER calcium regulation, stress and apoptosis.
Results:
Compared with the c-MWCNTs groups, the cytotoxicity of tau-MWCNTs was lower at the same dose and treatment time. Both the two types of MWCNTs could induce cytotoxicity apparently and statistically, when the cells were treated for only 3 h at the doses of more than 12.50 μg/cm(2) or treated for more than 12 h at the lower dose of 3.12 μg/cm(2) (P<0.05). The results of flow cytometry showed that the two MWCNTs could induce apoptosis of RAW264.7 cells in the dose- and time-dependently manner. The apoptosis rate of the cells treated for 24 h was higher than that for 12 h. And at each dose, the apoptosis rate induced by c-MWCNTs was also higher than that of the water soluble tau-MWCNTs within our study design. However, in this study, the mRNA expression levels of CRT, GRP78 and CHOP mRNA treated with both types of MWCNTs did not show any significant differences compared with the control groups.
Conclusion:
The cytotoxicity of tau-MWCNTs was much lower than that of c-MWCNTs. Unfortunately, we found the expression of genes related to ER had little effects on the apoptosis of RAW264.7 treated with MWCNTs, indicating that the ER pathway might not be the mechanism of MWCNTs-induced apoptosis.
