Selenoprotein R Protects Human Lens Epithelial Cells against D-Galactose-Induced Apoptosis by Regulating Oxidative Stress and Endoplasmic Reticulum Stress
Abstract
:1. Introduction
2. Result
2.1. SelR Gene Silence Effectiveness
2.2. Effect of SelR Gene Knockdown and Na2SeO3 on Cell Viability in d-Galactose-Treated hLE Cells
2.3. Effect of SelR Gene Knockdown and Na2SeO3 on d-Galactose-Induced Cell Apoptosis
2.4. Effect of SelR Gene Knockdown and Na2SeO3 on GPx Activity in d-Galactose-Treated hLE Cells
2.5. Effect of SelR Gene Knockdown and Na2SeO3 on d-Galactose-Induced ROS, MDA, PC and TSH Levels
2.6. Effect of SelR Gene Knockdown on d-Galactose-Induced Mitochondrial Dysfunctions in hLE Cells
2.7. Effect of SelR Gene Knockdown on Cytochrome c Release in d-Galactose-Treated hLE Cells
2.8. Effect of SelR Gene Knockdown on Caspase-3 Activity in d-Galactose-Treated hLE Cells
2.9. Effect of SelR Gene Knockdown on GRP78 Protein Expression in Glactose-Treated hLE Cells
3. Discussion
4. Materials and Methods
4.1. Materials
4.2. Cell Culture
4.3. SelR RNA Interference and Cell Treatment
4.4. Cell Viability Assay
4.5. Isolation of Total RNA and Real-Time PCR
4.6. Western Blot Analysis
4.7. Measurement of GPx Activity
4.8. Measurement of ROS, MDA, PC and TSH
4.9. Detection of Apoptosis
4.10. Assay of Cell Caspase-3 Activity
4.11. Detection of Mitochondrial Membrane Potential (Δψm)
4.12. Release of Cytochrome c from Mitochondria
4.13. Statistical Analysis
5. Conclusions
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
Abbreviations
cDNA | Complementary DNA |
DCFH-DA | 2′,7′-Dichlorodihydro-fluorescein diacetate |
DMEM | Dulbecco’s modified Eagle’s medium |
ECL | Enhanced chemiluminescence |
ER | Endoplasmic reticulum |
FITC | Fluorescein isothiocyanate |
GPx | Glutathione peroxidase |
GRP78 | Glucose-regulated protein |
hLE cells | Human lens epithelial cells |
JC-1 | 5,5′,6,6′-Tetrachloro-1,1′,3,3′-tetraethyl-imidacarbocyanine iodide |
MDA | Malondialdehyde |
MetO | Methionine sulfoxide |
Msr | Methionine sulfoxide reductase |
MTT | 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenylte-trazolium bromide |
NCS | Newborn calf serum |
PBS | Phosphate-buffered saline |
PC | Protein carbonyl |
PI | Propidium iodide |
ROS | Reactive oxygen species |
SelR | Selenoprotein R |
siRNA | Short interfering RNA |
TBS-T | Tris-buffered saline with Tween |
TSH | Total mercapto group |
Δψm | Mitochondrial membrane potential |
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Dai, J.; Liu, H.; Zhou, J.; Huang, K. Selenoprotein R Protects Human Lens Epithelial Cells against D-Galactose-Induced Apoptosis by Regulating Oxidative Stress and Endoplasmic Reticulum Stress. Int. J. Mol. Sci. 2016, 17, 231. https://doi.org/10.3390/ijms17020231
Dai J, Liu H, Zhou J, Huang K. Selenoprotein R Protects Human Lens Epithelial Cells against D-Galactose-Induced Apoptosis by Regulating Oxidative Stress and Endoplasmic Reticulum Stress. International Journal of Molecular Sciences. 2016; 17(2):231. https://doi.org/10.3390/ijms17020231
Chicago/Turabian StyleDai, Jie, Hongmei Liu, Jun Zhou, and Kaixun Huang. 2016. "Selenoprotein R Protects Human Lens Epithelial Cells against D-Galactose-Induced Apoptosis by Regulating Oxidative Stress and Endoplasmic Reticulum Stress" International Journal of Molecular Sciences 17, no. 2: 231. https://doi.org/10.3390/ijms17020231