西维来司钠通过抑制GAPDH/Siah1途径保护大鼠脊髓损伤

Sivelestat protects the spinal cord injury in rats via inhibiting GAPDH/Siah1 pathway

目的探索西维来司钠(SIV)对脊髓损伤(SCI)的保护作用及其相关分子机制。方法 56只大鼠分为三组:假手术组(Sham组,n=10),SCI组(n=31)和SCI+SIV组(n=15)。SCI+SIV组于SCI手术完成后1 h,利用腹腔注射10 mg/kg SIV,假手术组和SCI组注射等量生理盐水。实验分为二部分:(1)造SCI模型后SCI组取16只大鼠分别利用Western blot检测0 h、12 h、1 d、3 d胞质和胞核甘油醛-3-磷酸脱氢酶(GAPDH)蛋白表达;(2)利用Western blot检测1 d时各组大鼠GAPDH/Siah1,Bax/Bcl-2以及中性粒细胞弹性蛋白酶(NE)和诱导性一氧化氮合酶(iNOS)表达;利用化学定量法检测各组大鼠肿瘤坏死因子-α(TNF-α),白细胞介素1β(IL-1β)及一氧化氮(NO)含量;利用BBB评分评价各组大鼠1 d、7 d、14 d、21 d及28 d下肢运动功能。结果 (1)胞核GAPDH在造模后1 d时表达最高(P <0. 05),而胞质GAPDH在造模后1 d时表达最低(P <0. 05);(2)Western blot结果显示SIV能显著抑制GAPDH/Siah1表达,与SCI组比较有显著差异(P <0. 05);并且SIV能显著抑制NE,iNOS及Bax表达(P <0. 05),提高Bcl-2表达,与SCI组比较有显著差异(P <0. 05);化学定量法提示SIV可显著抑制TNF-α、IL-1β及NO含量,与SCI组比较有显著差异(P <0. 05); BBB评分显示从14 d开始SIV可显著提高大鼠下肢运动功能,与SCI组比较有显著差异(P <0. 05)。结论SIV能显著改善SCI后大鼠下肢运动功能,对SCI产生保护作用,可能是通过抑制GAPDH/Siah1途径发挥作用的。

Objective To explore the effects of sivelestat （SIV） on the spinal cord injury （SCI） in rats and to study its potential mechanism. Methods 56 rats were divided into 3 groups ： the sham group （ n = 10） , SCI group （ n =31 ） and the SCI ＋ SIV group （ n - 15 ）. The SCI ＋ SIV group was injected with 10 mg/kg SIV intraperitoneally 1 h after the completion of SCI surgery. The sham operation group and the SCI group were injected with the same amount of normal saline. Expression of GAPDH protein in cytoplasm and nucleus were detected by western blot at 0 h, 12 h, 1 d and 3 d following SCI. Then, the protein expression of GAPDH/Siahl and Bax/Bcl - 2 were valued by western blot. Meanwhile, protein expression of neutrophil elastase （NE） and inducible nitric oxide synthase （iNOS） were also valued by western blot. Besides, the content of tumor necrosis factor α （TNF -α） , interleukin 1 β （IL- 1 β ） and nitric oxide （NO） were determined by quantitative analysis method. BBB score was performed at 1 d, 7 d, 14 d, 21 d and 28 d, respectively. Results The nuclear GAPDH expression was highest at 1 d after SCI （ P 〈 0.05）, while cytoplasmic GAPDH showed the lowest expression at ld after SCI （ P 〈0.05 ）. Western blot revealed that SIV could significantly suppress the GAPDH/Siahl expression, as compared with SCI group （ P 〈0.05）. Besides, SIV could markedly inhibit the protein expression of NE, iNOS, Bax （ P 〈0.05） , and increase the Bcl - 2 expression （ P 〈 0.05 ）. Meanwhile, SIV could effectively suppress the content of TNF - α, IL - 1 β and NO, as compared with SCI group （ P 〈 0.05）. Moreover, BBB score revealed that SIV could significantly improve the motor function from 14 d, as compared with SCI group （ P 〈 0.05）. Conclusion SIV can significantly alleviate SCI, probably via suppressing the GAPDH/Siah1 pathway.