瑞芬太尼联合脐血间充质干细胞移植对脊髓损伤大鼠电生理及后肢功能的影响

摘要

目的 检测脐血间充质干细胞(UCMSC)移植联合瑞芬太尼对大鼠脊髓损伤电生理的影响及后肢功能变化.方法 83只成年Wistar大鼠,建立脊髓损伤模型,造模成功的80只大鼠运用随机数字表法分为4组.①UCMSC移植组,经尾静脉泵注等体积UCMSC细胞液;②对照组,尾静脉注入培养液;③瑞芬太尼组,瑞芬太尼注射液(2 mL·kg-1·h-1)尾静脉泵注4 h;④联合组,尾静脉注射UCMSC细胞后,经尾静脉泵注瑞芬太尼注射液(2 mL·kg-1·h-1)持续4h.依次于移植前及移植后1周、2周、4周、6周通过大鼠脊髓损伤(BBB)评分、改良Tarlov评分、斜板试验进行运动功能评定.移植后4周取材行荧光显微镜观测PKH-26标记的UCMSC存活及病理切片HE染色及分布情况.第4周进行辣根过氧化物酶(HRP)示踪分析神经纤维的再生情况,行运动诱发电位(MEP)和体感诱发电位(SEP)分析大鼠神经电生理恢复情况.结果 造模后,大鼠下肢运动功能评价联合组优于UCMSC移植组及瑞芬太尼组,UCMSC移植组和瑞芬太尼组优于对照组.瑞芬太尼组和UCMSC移植组损伤区可见少量神经轴索样的结构,该脊髓空洞比较小,联合组可见较多的神经轴索样结构,未见脊髓空洞.HE染色,对照组可见脊髓组织缺失及脊髓空洞形成,无神经轴索通过.移植后4周HRP阳性神经纤维数和PKH-26阳性细胞,对照组最少,联合组最多,UCMSC移植组和瑞芬太尼组次之,组间差异均有统计学意义(P＜0.05).MEP潜伏期,对照组＞瑞芬太尼组与UCMSC移植组＞联合组,且各组间差异均有统计学意义(P＜0.05);SEP潜伏期,对照组＜瑞芬太尼组与UCMSC移植组＜联合组,且各组间差异均有统计学意义(P＜0.05).结论 瑞芬太尼对脊髓损伤起到神经保护的作用,UCMSC移植的同时泵注瑞芬太尼能够促进脊髓损伤大鼠神经突触的再生,改善大鼠电生理功能及肢体运动功能.%Objective To determine the effects of cord blood mesenchymal stem cells (UCMSC) transplantation combined with remifentanil on electrophysiology and hindlimb functionon in spinal cord injured rats.Methods The spinal cord injury model was established in 83 adult Wistar rats,the model rats were randomly divided into four groups.① UCMSC transplantation group,via the tail vein infusion of equal volumes UCMSC cell sap.② control group,refers to tail vein injection medium group.③ remifentanil group,remifentanil injection (2 mL · kg · h-1) via tail vein infusion of 4 h.④ combined group,UCMSC cells injected intravenously via the tail vein and combined with infusion of remifentanil injection solution (2 ml · kg · h-1) continued 4 h.The spinal cord injury (BBB) score was evaluated before and 1 week,2 weeks,4 weeks and 6 weeks after transplantation.The modified Tarlov score and the slant plate test were used to evaluate the motor function.The survival and pathology of PKH-26-labeled UCMSC were observed by fluorescence microscopy at 4 weeks after transplantation.HRP (Horse Reddish Peroxidase) was used to analyze the regeneration of nerve fibers in the fourth week.MEP motor evoked potential and SEP somatosensory evoked potential were used to analyze the nerve Physiological recovery.Results Compared with UCMSC transplantation group and remifentanil group,UCMSC transplantation group and remifentanil group were better than the control group.There was a small amount of axonal-like structure in the injury area of Rufentini group and UCMSC transplantation group.The syndromes of the syringes were relatively small,and the ganglion-like structure was seen in the combined group.No syringomyelia was found.HE staining,the control group can be seen spinal cord loss and syringomyelia formation,no nerve axon through.The number of HRP-positive nerve fibers and PKH-26 positive cells were the lowest in the control group and the most in the control group,UCMSC transplantation group and remifentanil group were significantly different between the two groups at 4 weeks after transplantation (P＜0.05).MEP incubation group＞ remifentanil group and UCMSC transplantation group＞ and the difference between the groups were statistically significant (P＜0.05);SEP incubation period,control group,remifentanil group and UCMSC group,and the difference was statistically significant (P＜0.05).Conclusion Remifentanil may play a role in neuroprotective effects on spinal cord injury.At the same time,UCMSC transplanted with remifentanil can promote the regeneration of neuronal synapses in rats with spinal cord injury and improve the electrophysiological function and limb motor function of rats.