目的 探讨影响脊髓损伤轴突生长的NogoA、NgR时相表达及电针治疗脊髓损伤的时间窗效应.方法 144只Sprague-Dawley雌性大鼠随机分成假手术组(A组,n=48)和造模组(n=96).造模组大鼠采用改良Al-len法制作脊髓损伤大鼠模型,随机分为模型组(B组,n=48)和电针组(C组,n=48).C组电针大椎、腰阳关及双侧次髎、足三里,选择疏密波,持续20 min,每天1次.分别在造模后1 d、7 d、14 d介入电针治疗,每个介入治疗点大鼠随机再分成2个亚组,分别持续治疗7 d和14 d.A组于术后1 d、3 d、7 d、14 d、21 d、28 d,B组和C组于造模后在相应时间点处死大鼠提取脊髓组织,所有大鼠处死前进行BBB评分.各组每个时间点随机选取4份样本,应用逆转录实时定量聚合酶链反应(RT-qPCR)和Western blotting检测损伤脊髓不同时间段NogoA、NgR mRNA和蛋白的表达.结果 B组术后14 d BBB评分开始有所提高,直至28 d,评分较术后1 d、3 d、7 d提高(P<0.05).C组术后1 d介入治疗7 d,术后7 d、14 d介入治疗7 d和14 d,大鼠BBB评分较相应时间点B组均提高(P<0.05);术后7 d和14 d介入治疗的评分要高于术后1 d介入的评分(P<0.05);不同时间点介入电针治疗7 d与治疗14 d评分比较无显著性差异(P>0.05).B组NogoA、NgR mRNA和蛋白在造模后呈逐渐升高趋势,21 d达到高峰;不同时间点,B组NogoA、NgR mRNA和蛋白表达均明显高于A组(P<0.01).C组电针介入治疗后,除术后1 d介入治疗7 d的NogoA mRNA (P>0.05)表达外,其他时间点NogoA mRNA和蛋白的表达较相应时间点的模型组均下降(P<0.05);术后14 d介入治疗NogoA、NgR mRNA和蛋白的表达大多要低于术后1 d介入治疗(P<0.05);术后14 d介入治疗与术后7 d介入治疗以及术后7 d介入治疗与术后1 d介入治疗之间比较,NogoA、NgR mRNA和蛋白的表达都无显著性差异(P>0.05);术后不同时间点介入电针治疗7 d与14 d NogoA、NgR mRNA和蛋白表达之间无显著性差异(P>0.05).结论 电针能改善脊髓损伤大鼠后肢运动功能,可能与电针抑制脊髓损伤后NogoA、NgR mRNA和蛋白的表达有关;且电针早期介入治疗有效,恢复期介入治疗效果更佳.%Objective To investigate the phased expression of gene and protein of NogoA and its receptor (NgR) that affects axon growth of spinal cord injury (SCI), and to explore the time window effect of electroacupuncture on SCI rats. Methods A total of 144 female Sprague-Dawley rats were randomly assigned to sham operation group (group A, n=48) and model group (n=96). In the model group, Allen's method was used to establish SCI rats model, and they were further subdivided into model control group (group B, n=48) and electroacupuncture group (group C, n=48). Group C received electroacupuncture on Dazhui (GV14), Yaoyangguan (GV3), bilateral Ciliao (BL32) and Zu-sanli (ST36) with loose-tight wave, for 20 minutes every day, one day, seven days and 14 days after modeling. The rats at every interventional therapy time were randomly subdivided into two subgroups, which accepted sev-en or 14 days of treatment. Groups A and B were killed and the injured spinal cord tissue was extracted one day, three days, seven days, 14 days and 28 days after modeling, group C at the corresponding time. The hind limb motor function was assessed with BBB score before all of rats were killed. Four samples at every time in each group were randomly selected to detect the expression of mRNA and protein of NogoA and NgR at different stage of SCI using reverse transcription real-time quantitative polymerase chain reaction (RT-qPCR) and Western blotting. Results The BBB score began to increase 14 days after modeling, and significantly increased until 28 days after model-ing (P<0.05), compared with one day, three days and seven days after modeling in group B. The BBB score in-creased in group C than in group B at all the time points (P<0.05), except 14 days after electroacupuncture one day after modeling. The BBB score was higher as electroacupuncture intervening seven days and 14 days after modeling than that at one day after modeling in group C, and no significant difference was found between seven days and 14 days of treatment at either electroacupuncture time point (P>0.05). The expression of gene and pro-tein of NogoA and NgR in group B was in the increasing tendency after SCI, and was at the peak until 21 days af-ter modeling, and was higher in group B than in group A at each time point (P<0.01). The expression of gene and protein of NogoA decreased at all the time points in group C than in group B (P<0.05), except seven days of elec-troacupuncture intervening one day after modeling in the expression of NogoA mRNA (P>0.05). The expression of gene and protein of NogoA and NgR was lower as electroacupuncture intervening 14 days after modeling than one day after modeling in group C (P<0.05). There was no significant difference in the expression of gene and protein of NogoA and NgR between electroacupuncture intervening 14 days and seven days after modeling, and seven days and one day after modeling (P>0.05); as well as between seven days and 14 days of treatment at each time point (P>0.05). Conclusion Elerctroacupuncture could improve the hind limb motor function, which may associate with the inhibition of the expression of gene and protein of NogoA and NgR in injured spinal cord of rats after SCI. Elerctroacu-puncture is effective in the treatment of SCI at the early time, however, it is much better in the recovery stage.
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