人羊膜间充质干细胞移植对脊髓损伤大鼠诱导型一氧化氮合酶表达的影响

Effects of human amniotic mesenchymal stem cells transplantation on the expression of inducible nitric oxide synthase after spinal cord injury in rats

目的探讨大鼠脊髓损伤(spinal cord injury,SCI)后移植人羊膜间充质干细胞(human amniotic mesenchymal stem cells,hAMSCs)对诱导型一氧化氮合酶(inducible nitric oxide synthase,iNOS)表达的影响.方法将成年雌性SD大鼠36只,体质量250~300 g,随机分为假手术组(Sham组)、对照组(SCI组)和细胞移植组(hAMSCs组);采用改良Allen's方法建立大鼠脊髓损伤模型,Sham组仅行椎板切除术,SCI组于脊髓损伤部位注射磷酸盐缓冲液,hAMSCs组于脊髓损伤部位注射hAMSCs悬液;各组大鼠于术后第4天取其尾静脉血后麻醉处死,采集脊髓组织样本,采用免疫蛋白印记、免疫组化检测脊髓中iNOS蛋白表达,采用硝酸还原酶法测定血液NO的浓度.结果脊髓损伤后iNOS的表达存在于脊髓灰质前脚、后脚及中央管周围的神经细胞、星型胶质细胞和小胶质细胞胞质内.与Sham组比较,SCI组和hAMSCs组的iNOS蛋白表达明显升高(P<0.05);与SCI组比较,hAMSCs组的iNOS蛋白表达低(P<0.05).与Sham组比较,SCI组和hAMSCs组血液NO含量高(μmol/L:42.86±5.37 vs.167.76±18.47,42.86±5.37 vs.128.09±13.79,P<0.05);且SCI组含量高于hAMSCs组.结论大鼠急性脊髓损伤后移植人羊膜间充质干细胞能够降低诱导型一氧化氮合酶的表达,进而减少伤后NO的生成.

Objective To investigate the effects of human amniotic mesenchymal stem cells(hAMSCs)transplantation on the expression of inducible nitric oxide synthase(iINOS)after spinal cord injury(SCI)in rats.Methods 36 adult female SD rats(250-300 g)were randomly divided into sham operation group,SCI group and hAMSCs group.After SCI models were established by modified Allen's method,only laminectomy was performed in sham group,the rats of SCI group were injected with hosphate buffered saline,and the rats of hAMSCs group were injected with hAMSCs.Samples of caudal vein blood and spinal cord tissue were taken on the 4th day.The expression of iNOS protein in spinal cord was detected by W estern blot and immunohistochemistry.The concentration of NO in blood was measured by nitrate reductase method.Results The expression of iNOS was found in the cytoplasm of nerve cells,astrocytesand microglia in the anterior and posterior horn of spinal cord and around the central canal.Compared with sham group,iNOS protein expression in SCI group and hAMSCs group increased significantly(P<0.05),and iNOS protein expression in hAMSCs group was lower than that in SCI group(P<0.05).Compared with sham group,NO content in blood was higher in SCI group and hAMSCs group(μmol/L:42.86±5.37 vs.167.76±18.47;42.86±5.37 vs.128.09±13.79,P<0.05),and the NO content in SCI group was higher than that in hAMSCs group(μmoVL:167.76±18.47 vs.128.09±13.79,P<0.05).Conclusion Transplantation of hAMSCs after acute SCI in the rats can reduce the expression of iNOS and the NO content in blood after injury.