转TrkC基因神经干细胞移植联合NT-3治疗脊髓缺血再灌注损伤中MAP-2表达的变化被引量：1

Expression changes of MAP-2 after spinal cord ischemia-reperfusion with the treatment of trans-TrkC gene neural stem cells combined with neurotrophin-3

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摘要目的研究转酪氨酸激酶C（tyrosine kinase C,Trk C）基因神经干细胞（neural stem cells,NSCs）移植联合神经营养素（neurotrophin,NT）-3局部应用对大鼠脊髓缺血再灌注损伤的治疗作用及微管相关蛋白-2（microtubule associated protein-2,MAP-2）表达的变化。方法 90只Wistar大鼠,雌雄不拘,清洁级,300~350 g,随机分为6组（每组15只）,分别为正常对照组（A组）、缺血再灌注损伤模型组（B组）、NSCs移植组（C组）、NSCs移植＋NT-3局部使用组（D组）、转Trk C基因NSCs移植组（E组）和转Trk C基因NSCs移植＋NT-3局部使用组（F组）。于左锁骨下动脉末梢处予降主动脉缠胶带法建立脊髓缺血损伤模型,50 min后,阻断的降主动脉被解除。术后第1天进行细胞移植。各组大鼠分别于细胞移植后6 h、24 h及48 h进行脊髓运动功能评分。术后24 h、1周,分别处死7和8只大鼠,迅速取出脊髓标本,苏木素-伊红、NISSLE染色观察神经元形态学变化,免疫组织化学法观察MAP-2的表达变化,实时定量聚合酶链反应法检测MAP-2 m RNA的表达变化。结果与缺血模型组（B、C、D、E组）和其他移植处理组相比,F组大鼠脊髓运动功能恢复情况最佳,脊髓神经元核溶解程度较轻,神经元结构相对完整,数量也较多,MAP-2表达增加;与对照组（A组）和缺血模型组（B组）相比,各移植处理组MAP-2 m RNA表达均有不同程度的增加,并且,F组MAP-2 m RNA表达增加最为明显,与其他各组相比,差异具有统计学意义（P〈0.01）。结论在局部给予的NT-3作用下,转Trk C基因NSCs对脊髓缺血再灌注损伤具有较好的治疗作用,其可能的机制是通过上调MAP-2的表达实现的。 Objectives To investigate the therapeutic effect of trans-tyrosine kinase （TrkC） gene neural stem cells （NSCs） combined with the regional application of neurotrophin （NT）-3 on the recovery of neural function after spinal cord ischemia-reperfusion injury through the observation of the change of microtubule associated protein-2 （MAP-2）. Methods Ninety Wistar rats of either sex, weighted 300-350 g, were randomly divided into six groups （n=15）： normal control group （A）, ischemia-reperfusion group （B）, NSCs transplantation group （C）, NSCs transplantation with the regional application of NT-3 group （D）, trans-TrkC gene NSCs transplantation group （E） and trans-TrkC gene NSCs transplantation with the regional application of NT-3 group （F）. Models of spinal cord ischemia-reperfusion injury were established by descending aorta at left subclavian artery occlusion. The descending aorta was unblocked after 50 mins. Cell transplantation into the injured spinal cord was performed at the postoperative day 1. After the cell transplantation, the motor function scores were recorded at 6 h, 24 h, 48 h. Seven and 8 rats were killed 24 h and 1 week after cell transplantation respectively. Then, the spinal cord specimens were taken out as quickly as possible. Hematoxylin and eosin staining method and NISSLE staining method were adopted to observe the morphological changes of spinal neurons, as well as immunohistochemical method were applied to observing the changes of MAP-2 and realtime quantitative polymerase chain reaction were adopted to detect the changes of MAP-2 mRNA. Results Compared with B, C, D, E group, the Tarlov score of F group restored the best. There was a slight damage and a higher expression changes of MAP-2 in F group. Compared with A and B group, the expression of MAP-2 mRNA in C, D, E and F group had an increase to various degrees. The expression of MAP-2 mRNA in F group was markedly increased compared with other groups （C,D,E group）, and the differences were statistically significant （P〈0.01）. Conclusions With the regional application of NT-3, trans-TrkC gene NSCs transplanted into injured spinal cord has a significant protective effect on spinal cord ischemia reperfusion injury and the potential mechanism may be related to the up-regulation of MAP-2.

作者丁皓朱平陈寄梅黄焕雷卢聪陈志衡蔡骞赵明一

机构地区广东省心血管病研究所心外科广东省华南结构性心脏病重点实验室广东省人民医院(广东省医学科学院) 广东医学院中南大学湘雅三医院儿科

出处《岭南心血管病杂志》 2016年第5期593-599,共7页 South China Journal of Cardiovascular Diseases

基金国家自然科学基金项目(项目编号:81370230 81500231) 中国科技部国际合作项目(项目编号:2010DFA32660 2008DFA31140) 湖南省自然科学基金(项目编号:13JJ6018 15JJ6118) 中南大学湘雅三医院"新湘雅人才工程"(项目编号:JY201524)

关键词酪氨酸激酶C 神经干细胞细胞移植脊髓缺血再灌注损伤神经营养素-3 微管相关蛋白-2 tyrosine C neural stell cell cell transplantation spinal cord ischemia-reperfusion injury neurotrophin-3 microtubule associated protein-2

分类号 R744 [医药卫生—神经病学与精神病学]

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参考文献15

1NAZLI Y, COLAK N, ALPAY M F, et al. Neuroprotective effect of atorvastatin in spinal cord ischemia-reperfusion injury [ J ]. Clinics (Sao Panlo), 2015, 70 ( 1 ) : 52-60.
2ACHER C W, WYNN M. A modem theory of paraplegia in the treatment of aneurysms of the thoracoabdominal aorta: an analysis of technique specific observed/expected ratios for paralysis[J]. J Vasc Surg, 2009, 49(5) : 1117-1124.
3REYNOLDS B A, TETZLAFF W, WEISS S. A multipotent EGF-responsive striatal embryonic progenitor cell produces neurons and astrocytes[J]. J Neurosci, 1992, 12(11): 4565- 4574.
4WONG M L, MEDRANO J F. Real-time PCR for mRNA quantitation[J]. Biotechniques, 2005, 39( 1 ) : 116-121.
5WAKAI T, SAKATA H, NARASIMHAN P, et al. Transplantation of neural stem cells that overexpress SOD1 enhances amelioration of intracerebral hemorrhage in mice [ J ]. J Cereb Blood Flow Metab, 2014, 34(3): 441-449.
6CAO Q L, ZHANG Y P, HOWARD R M, et al. Pluripotent stem cells engrafted into the normal or lesioned adult spinal cord are restricted to a glial lineage [J]. Exp Neurol, 2001, 167(1) : 48-58.
7粱日生.转TrkC基因神经干细胞移植治疗脊髓损伤的实验研究[D].上海:复旦大学,2005:82-86.
8CASTELLANOS D A, TSOULFAS P, FRYDEL B R, et al. TrkC overexpression enhances survival and migration of neural stem cell transplants in the rat spinal cord [ J ]. Cell Transplant, 2002, 11(3): 297-307.
9BOTHWELL M. NGF, BDNF, NT3, and NT4[J]. Handb Exp Pharmacol, 2014, 220: 3-15.
10JEFFERY N D, FITZGERALD M. Effects of red nucleus ablation and exogenous neurotrophin-3 on corticospinal axon terminal distribution in the adult rat [J]. Neuroscience, 2001, 104(2) :513-521.

二级参考文献18

1曾园山,吴良芳.哺乳动物脊髓损伤后的可塑性研究[J].解剖学报,1995,26(4):439-442. 被引量：3
2王俊梅,曾园山,刘然义,吴立志,薛刚,黄文林,熊轶,王延华.人神经营养素-3基因重组腺病毒载体的构建、表达和生物活性检测[J].解剖学报,2005,36(5):460-464. 被引量：4
3Zhou L, Baumgartner BJ, Hill-Felberg SJ, et al. Neurotrophin-3 expressed in situ induces axonal plasticity in the adult injured spinal cord. J Neurosci, 2003,23 (4): 1424-1431.
4Sehnell L, Schneider R, Kolbeck R, et al. Neurotropin-3 enhances sprouting of eorticospinal tract during development and after adult spinal cord lesion. Nature, 1994,367 (6549) : 170-173.
5Senut MC, Tuszynski MH, Raymon HK, et al. Regional differences in responsiveness of adult CNS axons to grafts of ceils expressing human neurotrophin-3. Exp Neurol, 1995,135 ( 1 ) : 36-55.
6Houweling DA, Lankhorst AJ, Gispen WH, et al. Collagen containing neurotrophin-3 (NT-3) attracts regrowing injured corticospinal axons in the adult rat spinal cord and promotes partial functional recovery. Exp Neurol, 1998, 153(1) :49-59.
7Jeffery ND, Fitzgerald M. Effects of red nucleus ablation and exogenous neurotrophin-3 on corticospinal axon terminal distribution in the adult rat. Neurosei, 2001,104 (2) : 513-521.
8Wang JM, Zeng YS, Wu JL, etal. Cograft of neural stem cells and Schwann cells overexpressing TrkC and neurotrophin-3 respectively after rat spinal cord transaction. Biomaterials, 2011, Jul 21. [Epub ahead of print].
9Zeng YS, Ding Y, Wu LZ, et al. Co-transplantation of schwann ceils promotes the survival and differentiation of neural stem cells transplanted into the injured spinal cord. Dev Neurosci, 2005,27 ( 1 ) : 20-26.
10Wang JM, Zeng YS, Liu RY, etal. Recombinant adenovirus vector-mediated functional expression of neu- rotropin-3 receptor (TrkC) in neural stem cells. Exp Neurol, 2007,203 ( 1 ) : 123-127.