Mutations in the Corneal Endothelial Dystrophy-Associated Gene SLC4A11 Render the Cells More Vulnerable to Oxidative Insults
| Autor: | Venkata Pulla Rao Vendra, D. C. Praneetha, Sanhita Roy |
|---|---|
| Rok vydání: | 2015 |
| Předmět: |
Corneal endothelium
Mitochondrial Diseases NF-E2-Related Factor 2 Mutant Anion Transport Proteins Mutation Missense Apoptosis Oxidative phosphorylation medicine.disease_cause Real-Time Polymerase Chain Reaction Transfection Antiporters tert-Butylhydroperoxide medicine NAD(P)H Dehydrogenase (Quinone) Humans Cellular localization chemistry.chemical_classification Corneal Dystrophies Hereditary Reactive oxygen species Microscopy Confocal HEK 293 cells Fuchs' Endothelial Dystrophy Flow Cytometry Cell biology Ophthalmology Oxidative Stress HEK293 Cells chemistry Gene Expression Regulation Colorimetry Reactive Oxygen Species Oxidative stress Heme Oxygenase-1 |
| Zdroj: | Cornea. 34(6) |
| ISSN: | 1536-4798 |
| Popis: | PURPOSE To investigate the effect of mutations in SLC4A11 on cellular localization of the protein, mitochondrial function, and apoptosis due to oxidative stress. Mutations in SLC4A11 have been associated with 2 different forms of corneal endothelial dystrophy that lead to degeneration of the corneal endothelium, causing opacity of the cornea and gradual vision loss. METHODS HEK 293 cells were transfected with wild-type SLC4A11 or mutants, Ser213Leu, Arg233Cys, Gly418Asp, and Thr584Lys, and exposed to oxidative stress. Cellular localization of the proteins was detected by confocal microscopy, whereas mitochondrial dysfunction, reactive oxygen species (ROS) generation, and apoptosis were analyzed by flow cytometry and a colorimetric assay. Expressions of antioxidant genes were quantitated by real-time polymerase chain reaction. RESULTS Although wild-type SLC4A11 was localized on the cell membrane, mutant proteins were found diffused in the cytoplasm. Mutations in SLC4A11 caused an increase in generation of ROS and mitochondrial dysfunction due to oxidative stress. NRF2, HO-1, and NQO expression decreased significantly, and a higher rate of apoptosis was detected in cells with mutant proteins under oxidative stress. CONCLUSIONS Our data suggest that mutations in SLC4A11 cause retention of the protein in the cytoplasm and generate increased reactive oxygen species. We found that cells containing mutant SLC4A11 are more vulnerable to oxidative and mitochondrial damage, less able to overcome oxidative stress through the expression of sufficient levels of antioxidant genes, and are more prone to apoptotic death. |
| Databáze: | OpenAIRE |
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