Effect of combined PARP-1 and Caspase-3 inhibitors on neurocyte apoptosis after spinal cord injury in rats

English Keywords:Spinal cord injury  PARP-1 inhibitor  Caspase-3 inhibitor  Apoptosis-inducing factor  Rat

English Abstract:

  【Abstract】 Objectives: To discuss the influence of combined PARP-1 inhibitor 3-aminobenzamide(3-AB) and Caspase-3 inhibitor Z-DEVD-FMK on the spinal neurocyte apoptosis after spinal cord injury in rats. Methods: 120 healthy adult SD rats were randomly divided into sham operation group(group A), model group(group B), PARP-1 inhibitor group(group C) and the combination of 3-AB and Z-DEVD-FMK group(group D) with 30 rats in each group. Then the rats of each group were divided into 1, 3, 7 days. Spinal cord injury models were established by Allen′s method. The BBB locomotor testing score was used to evaluate the rats′ locomotor function of hindlimbs in four groups after operation. The immunostaining was performed to detect the expressions of PARP-1, apoptosis-inducing factor(AIF), Caspase-3 and Bcl-2 in all groups. Western blotting was used to measure the expressions of PARP-1 and Caspase-3 protein after spinal cord injury, and real-time quantitative PCR was further used to detect the expressions of PARP-1, AIF, Caspase-3 and Bcl-2 mRNA in all groups following spinal cord injury. The level of cell apoptosis was examined by the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling(TUNEL) assay. Results: The BBB scores of rats in group D and C were significantly higher than that in group B at 7 days after SCI. Furthermore, the score in group D was higher than that in group C(P＜0.05). Immunohistochemistry results and Western blotting results showed that, from 1d to 7d after SCI, the expressions of PARP-1, AIF and Caspase-3 increased in group B, while the expressions of Bcl-2 gradually weakened(P＜0.05). Compared with group B, the expression of PARP-1, AIF and Caspase-3 in group D and C after SCI decreased significantly, and the expression in group D was lower than that in group C(P＜0.05). While compared with group B, the expression of Bcl-2 in group D and C obviously increased, and the expression in group D was higher than that in group C(P＜0.05). The results of real-time quantitative PCR corroborated the immunohistochemistry and Western blotting detection results. TUNEL result showed that, the apoptotic cells in group B peaked at 3d and decreased at 7d after SCI(P＜0.05). The apoptotic index of group D and C significantly decreased than that of group B, and the rate in group D was lower than that in group C(P＜0.05). Conclusions: Combined PARP-1 inhibitor 3-AB and Caspase-3 inhibitor Z-DEVD-FMK can reduce the apoptosis of cells after SCI, which may be related to the inhibition of the expression of PARP-1, AIF and Caspase-3, and the up-regulation of the expressions of Bcl-2.