目的 探讨“三通针法”治疗对大鼠脊髓损伤后p75神经营养素受体(p75NTR) mRNA及蛋白表达的影响。方法 72只Sprague-Dawley雌性大鼠随机分成假手术组(A,n=8)和造模组(n=64)。造模组大鼠采用改良Allen重物打击法制作脊髓损伤大鼠模型,造模成功后存活48只大鼠,随机分为模型组(B,n=12)、电针组(C,n=12)、阻断剂NEP1-40组(D,n=12)、电针+阻断剂NEP1-40组(E,n=12)。电针治疗取大椎、腰阳关及双侧次髎、足三里,选择疏密波,持续时间20 min,每天1次。分别于治疗后7 d、14 d提取损伤处脊髓组织,采用实时荧光定量PCR、原位杂交、Western blotting方法分别检测p75NTR mRNA及蛋白的表达,采用BBB评分评估大鼠后肢活动功能。结果 BBB评分显示,治疗后各组评分均较B组提高,且E组BBB评分分别高于C组和D组(P<0.05),各组14 d评分均高于7 d (t>2.623, P<0.05)。脊髓损伤后,各治疗组与B组比较,脊髓组织中p75NTR mRNA及蛋白的表达均降低(P<0.05);C组、D组和E组之间无显著性差异(P>0.05)。结论 “三通针法”电针能改善脊髓损伤大鼠后肢运动功能,抑制脊髓损伤后p75NTR的表达活动,这可能是电针治疗脊髓损伤的机制之一。
Objective To investigate the effect of Santong electroacupuncture (EA) on mRNA and protein expression of p75 neurotrophin receptor (p75NTR) in rats with spinal cord injury (SCI). Methods A total of 72 female Sprague-Dawley rats were randomly assigned to sham operation group (group A, n=8) and model group (n=64). In the model group, Allen's method was used to make SCI rats model, in which 48 survived model rats were further subdivided into model control group (group B, n=12), EA group (group C, n=12), inhibitor Nogo extra cellular peptide residues 1-40 (NEP1-40) group (group D, n=12) and EA+inhibitor NEP1-40 group (group E, n=12) according to design proposal. The treatment groups were electroacupunctured on Dazhui (GV14) and Yaoyangguan (GV3), bilateral Ciliao (BL32) and Zusanli (ST36) with loose-tight wave, for 20 minutes every day. After 7 and 14 days of treatment, injured spinal cord tissue was extracted for detecting. The mRNA and protein expression of p75NTR was detected by real-time fluorescent quantitative PCR, in situ hybridization and Western blotting respectively. The hind limb motor function was assessed with Basso-Beattie-Bresnahan (BBB) score. Results The BBB score increased in the treatment groups compared with group B, and was higher in group E than in groups C and D (P<0.05), as well as on the 14th day than on the 7th day in all the treatment groups (t>2.623, P<0.05). The mRNA and protein expression of p75NTR in spinal cord tissues decreased in the treatment groups compared with group B (P<0.05), and no significant difference was found among the treatment groups (P>0.05). Conclusion Santong elerctroacupuncture treatment could improve the hind limb motor function, which may associate with inhibition of the mRNA and protein expression of p75NTR in rats after SCI.
[1] 高洁,程立红,闵友江. 针灸治疗脊髓损伤恢复期的临床研究概况[J]. 按摩与康复医学, 2015, 6(3): 18-21.
[2] 王智琴,闵友江. 热敏灸治疗不完全性脊髓损伤性尿潴留的疗效观察[J]. 成都中医药大学学报, 2013, 36(3): 60-63.
[3] 王文春,卢家春,王倩,等. “截瘫三联针法”对外伤性脊髓损伤患者日常生活活动能力的影响[J]. 中国针灸, 2012, 32(10): 877-881.
[4] 闵友江,程立红,高洁,等. 三通针法治疗脊髓损伤恢复期截瘫患者临床观察[J]. 上海针灸杂志, 2013, 32(10): 1010-1013.
[5] 闵志云,程立红,闵友江. Nogo/NgR通路在脊髓神经再生中的研究进展[J]. 江西中医药大学学报, 2016, 28(3): 103-105.
[6] 杨俊锋,张亚峰,马勇,等. 大鼠脊髓损伤后勿动蛋白受体的动态表达[J]. 中国康复理论与实践, 2014, 20(10): 919-923.
[7] Wang KC, Kim JA, Sivasankaran R, et al. p75 interacts with the Nogo receptor as a coreceptor for Nogo, MAG and OMgp [J]. Nature, 2002, 420(691): 74-78.
[8] GrandPré T, Li S, Strittmatter SM. Nogo-66 receptor antagonist peptide promotes axonal regeneration [J]. Nature, 2002, 417(6888): 547-551.
[9] 尹明锡,时素华,宋萌,等. 电针对大鼠脊髓损伤后Nogo-A 表达的实验研究[J]. 陕西中医, 2010, 31(3): 371-373.
[10] 伍芳,龚标,李学智,等. 电针对局灶性脑梗死大鼠Nogo-A及其受体NgR和运动诱发电位的影响[J]. 第三军医大学学报, 2013, 35(3): 228-232.
[11] 高洁,张云杰,闵友江,等. 针灸治疗截瘫的腧穴研究及应用[J]. 江西中医药, 2013, 44(2): 54-56.
[12] 张露芬.实验针灸学[M]. 北京:化学工业出版社, 2010: 219-220.
[13] 郭爱丽,朱薇薇,王丽,等. Nogo-A受体拮抗剂NEP1-40对新生大鼠HIBD中神经元轴突再生的影响[J]. 中国儿童保健杂志, 2008, 16(6): 679-681.
[14] Basso DM, Beattie MS, Bresnahan JC. A sensitive and reliable locomotor rating scale for open field testing in rats [J]. J Neurotrauma, 1995, 12: 1-21.
[15] 闵志云,程立红,闵友江.电针对脊髓损伤大鼠Nogo/NgR信号通路相关因子基因表达的影响[J]. 北京中医药大学学报, 2016, 39(11): 926-932.
[16] Zhang L, Ni YQ, Li YF. Ligustrazine facilitates hair cell regeneration in the cochlea following gentamicin ototoxicity [J]. Neural Regen Res, 2010, 5(10): 735-740.
[17] Yang TW, Song XS, Wang P, et al. Effect of extracellular signal-regulated kinase and nitric oxide on compressive neuralgia formation and maintenance [J]. Neural Regen Res, 2010, 5(10):757-763.
[18] 闵友江,程立红,张举玲,等. 基于临床路径模式的脊髓损伤恢复期针灸、康复、护理三位一体治疗方案制订的初探[J]. 医学信息, 2012, 12(25): 273-274.
[19] 于金娜,马晓晶,刘志顺,等. 电针“次髎”穴对逼尿肌反射亢进大鼠骶髓排尿中枢c-fos表达的影响[J]. 针刺研究, 2010, 35(3): 204-208.
[20] 邓娜,高敏. 电针治疗脊髓损伤的基础理论研究[J]. 甘肃中医, 2009, 22(3): 11-12.
[21] 李志刚,刘如春,耿直,等. 电针对急性脊髓损伤大鼠差异表达基因及钙离子作用的实验研究[J]. 北京中医药大学学报, 2008, 31(7): 486-489.
[22] 杜旭,王瑞辉. 电针对脊髓损伤大鼠尼氏体和神经功能的影响[J]. 时珍国医国药, 2011, 22(4): 995-996.
[23] Beattie MS, Harrington AW, Lee R, et al. ProNGF induces p75-mediated death of oligodendro-cytes following spinal cord injury [J]. Neuron, 2002, 36(3): 375-386.
[24] Harrington AW, Leiner B, Blechschmitt C, et al. Secreted proNGF is a pathophysiological death-inducing ligand after adult CNS injury [J]. Proc Natl Acad Sci USA, 2004, 101(16): 6226-6230.
[25] 吕威,莫雨平,李冰,等. 督脉电针对不同时间段脊髓损伤大鼠运动功能及p75神经营养素受体表达的影响[J]. 中国康复理论与实践, 2016, 22(8): 876-883.
