摘要目的 探讨乌司他丁对周围神经损伤后脊髓神经元凋亡及CCAAT增强子结合蛋白(CHOP)表达的影响. 方法 将225只健康雄性SPF级C57BL/6J小鼠按随机数字表法分为3组:假手术组、周围神经损伤组和乌司他丁组,每组75只.后2组制备单侧坐骨神经损伤模型,造模成功后乌司他丁组腹腔注射乌司他丁溶液0.2 mL(10 000 U/kg),假手术组和周围神经损伤组给予等容量生理盐水,1次/d,连续3d.分别于术后1、3、7、14、28 d取小鼠脊髓L4-6节段,采用HE染色观察组织病理学变化,采用TUNEL法检测神经元凋亡并计算神经元凋亡指数(AD,采用Western blotting检测CHOP、Bcl-2、Bax、活化型Caspase-3蛋白表达并计算Bcl-2/Bax比值,采用RT-PCR检测CHOP mRNA表达. 结果 HE染色显示周围神经损伤组脊髓损伤较假手术组明显加重,而乌司他丁组脊髓损伤较周围神经损伤组明显减轻.术后1、3、7、14、28 d时,与假手术组比较,周围神经损伤组和乌司他丁组神经元AI明显升高,Bcl-2蛋白表达明显降低,活化型Caspase-3及Bax蛋白表达明显升高,Bcl-2/Bax比值明显降低,CHOP蛋白及mRNA表达明显升高,差异均有统计学意义(P＜0.05);与周围神经损伤组比较,乌司他丁组神经元AI明显降低,Bcl-2蛋白表达明显升高,活化型Caspase-3及Bax蛋白表达明显降低,Bcl-2/Bax比值明显升高,CHOP蛋白及mRNA表达明显降低,差异均有统计学意义(P＜0.05). 结论 乌司他丁对周围神经损伤后脊髓损伤的保护作用与其降低CHOP表达、抑制脊髓神经元凋亡有关.

abstractsObjective To evaluate the effect of ulinastatin on neuron apoptosis and CCAAT/enhancer-binding protein (CHOP) expression of spinal cord after peripheral nerve injury.Methods A total of 225 healthy male SPF C57BL/6J mice were divided into three groups by using a random number table:sham-operated group,peripheral nerve injury group and ulinastatin group (n=75).The models of unilateral sciatic nerve transection were established in the latter two groups.After the models being established,intraperitoneal injection ofulinastatin 0.2 mL (10 000 U/kg) was performed once daily for 3 consecutive d in ulinastatin group,and the equal volume of normal saline was given once daily for 3 consecutive d in sham-operated group and peripheral nerve injury group.One,3,7,14 and 28 d after surgery,L4-6 spinal cord segments were removed for pathological examination by HE staining,and for detection of neuron apoptosis and apoptotic index (AI) by TUNEL method;the expressions of CHOP,Bcl-2,Bax and cleaved caspase-3 proteins were determined by Western blotting and the ratio of Bcl-2/Bax was calculated,and the CHOP mRNA expression was detected by RT-PCR.Results HE staining showed that the injury of spinal cord in peripheral nerve injury group was more aggravated as compared with that in the sham-operated group,and the injury of spinal cord in ulinastatin group was more alleviated as compared with that in the peripheral nerve injury group.One,3,7,14 and 28 d after surgery,AI was significantly higher,Bcl-2 protein expression was down-regulated,cleaved caspase-3 and Bax protein expressions were up-regulated,Bcl-2/Bax ratio was lower,and CHOP protein or mRNA expressions were up-regulated in the peripheral nerve injury group and ulinastatin group as compared with those in the sham-operated group,with statistically significant differences (P＜0.05).As compared with those in the peripheral nerve injury group,AI was significantly lower,Bcl-2 protein expression was up-regulated,cleaved caspase-3 and Bax protein expressions were down-regulated,Bcl-2/Bax ratio was higher,and CHOP protein or mRNA expressions were down-regulated in ulinastatin group,with statistically significant differences (P＜0.05).Conclusion The mechanism by which ulinastatin protects spinal cord injury after peripheral nerve injury is related to down-regulation of CHOP expression and suppression of neuron apoptosis of spinal cord.