1.Cell apoptosis in atrophic skeletal muscle induced by immoblization in rabbits--an experimental study using TUNEL.
Xuhong JIN ; Ninghou QU ; Yong HE ; Ying GUO ; Chun WANG ; Changyi LU ; Chen LI ; Zhaolan WEI ; Jieying CHEN
Journal of Biomedical Engineering 2004;21(4):628-635
This experiment was designed to explore the correlation between the mechanism of immobilization-induced skeletal muscle atrophy and the apoptosis of muscular cells. The models of skeletal muscle atrophy induced by immobilization for different length of time were established according to Sievanen II methods. 24 rabbits, each of them having one hind leg fixed by the tubal plaster and the other one free as control, were randomly divided into four groups depending on time of fixation (3, 7, 14, and 28 days respectively). The animals were sacrificed by the end of fixation. TdT-mediated d-UTP nick end labeling (TUNEL) was used to investigate the apoptotic muscle cells in the animal's bone. By comparing the apoptotic muscle cells with the morphology of the skeletal muscle, the correlation between cell apoptosis and skeletal muscle atrophy were analyzed. Apoptotic muscle cells did appear after immobilization in the atrophied skeletal muscle. In various groups, some cells with false positive stained TUNEL were found in the atrophic muscle, which could be distinguished from apoptotic cells by their characteristics. In conclusion, cell apoptosis participates in the process of skeletal muscle atrophy induced by immobilization; the amount of apoptotic cells is strongly associated with the time of immobilization, its peak appears on the 14th day of immobilization; the distribution of apoptotic skeletal muscle cell varies with the time of fixation. The severity of skeletal muscle atrophy is associated with the degree of the muscle cell apoptosis.
Animals
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Apoptosis
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physiology
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Immobilization
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In Situ Nick-End Labeling
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Muscle, Skeletal
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pathology
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Muscular Atrophy
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etiology
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Rabbits
2.Effects on survival of shRNA mediated APE/Ref1 gene silencing in rat spiral ganglion cells in oxidative stress.
Zhendong JIANG ; Cheng ZHONG ; Taijun LI ; Zhaolan XIANG ; Xueyuan ZHANG
Chinese Journal of Otorhinolaryngology Head and Neck Surgery 2014;49(2):145-150
OBJECTIVETo 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).
METHODSPrimary 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).
RESULTSWestern 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.
CONCLUSIONSThe 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.
Animals ; Cells, Cultured ; DNA-(Apurinic or Apyrimidinic Site) Lyase ; genetics ; Gene Silencing ; Hydrogen Peroxide ; Oxidation-Reduction ; Oxidative Stress ; RNA, Small Interfering ; Rats ; Rats, Sprague-Dawley ; Spiral Ganglion ; cytology ; metabolism