Effects on survival of shRNA mediated APE/Ref1 gene silencing in rat spiral ganglion cells in oxidative stress / 中华耳鼻咽喉头颈外科杂志
Chinese Journal of Otorhinolaryngology Head and Neck Surgery
; (12): 145-150, 2014.
Article
in Zh
| WPRIM
| ID: wpr-302988
Responsible library:
WPRO
ABSTRACT
<p><b>OBJECTIVE</b>To investigate the effects of reducing APE/Ref1 expression in the cultures of rat spiral ganglion cells with oxidative damage induced by H(2)O(2).</p><p><b>METHODS</b>Primary cultured rat spiral ganglion cells were infected with small interfering RNA to APE/Ref1 (Ape1siRNA) for 72 h, followed by treating with H(2)O(2) (0, 10, 25, 50, 100 and 300 µmol/L) for 1 h , and then cultured in normal medium for 24 h. Western blot were used to detect the level of APE/Ref1 protein and phosphorylation of histone protein H2AX in the infected cells. The caspase3 activation was tested by spectrophotometric method . The cell viability was determined by MTT and the apoptosis of spiral ganglion cells was determined by terminal-deoxynucleotidyl transferase mediated nick and labeling (TUNEL).</p><p><b>RESULTS</b>Western blot showed that infection with Ape1siRNA resulted in APE/Ref1 reduced expression in the spiral ganglion cells. Exposing spiral ganglion cultures with reduced expression of APE/Ref1 to H(2)O(2) (50, 100, 300 µmol/L) for 1 h resulted in increasing in the phosphorylation of histone protein H2AX. The reduction in APE/Ref1 significantly reduced cell viability in cultures 24 h after 1 h expression to 50-300 µmol/L H(2)O(2). The apoptosis of cells and caspase 3 activity was detected significantly improved.</p><p><b>CONCLUSIONS</b>The induced of APE/Ref1 results in significantly decrease in spiral ganglion cells viability in oxidative stress. The repairing function of APE/Ref1 is necessary for optimal levels of neuronal rat spiral ganglion cells survival.</p>
Full text:
1
Database:
WPRIM
Main subject:
Oxidation-Reduction
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Cells, Cultured
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Spiral Ganglion
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Rats, Sprague-Dawley
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Oxidative Stress
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Gene Silencing
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Cell Biology
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RNA, Small Interfering
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DNA-(Apurinic or Apyrimidinic Site) Lyase
/
Genetics
Limits:
Animals
Language:
Zh
Journal:
Chinese Journal of Otorhinolaryngology Head and Neck Surgery
Year:
2014
Document type:
Article