人脐血MSCs移植修复大鼠脊髓损伤的效果及机制的初步探讨

中国临床解剖学杂志 ›› 2011, Vol. 29 ›› Issue (2): 202-207.

人脐血MSCs移植修复大鼠脊髓损伤的效果及机制的初步探讨

孙　丽¹，　于　丽¹，　张华芳²，　王　力¹，　魏志新¹

1.潍坊医学院组织学与胚胎学教研室，山东潍坊 261053；　2 潍坊市人民医院保健三科，山东潍坊 261041

收稿日期:2010-10-03 出版日期:2011-03-25 发布日期:2011-03-28
通讯作者: 于　丽，博士，硕士生导师，E-mail:yulidoctor@163.com
作者简介:孙　丽（1985-），女，在读硕士，研究方向：脐血及脐带间充质干细胞分化研究
基金资助:
山东省自然科学基金项目（Y2008C26），山东省教育厅项目(J07YD01)

Effects and mechanism of the transplantation of human umbilical cord blood msenchymal stem cells to treat spinal cord injury

SUN Li1, YU Li1, ZHANG Hua-fang2, WANG Li1,WEI Zhi-xin1

1.Department of Histology and embryology ,Weifang medical college , Weifang 261053, China； 2. Weifang People Hospital, Weifang 261041

Received:2010-10-03 Online:2011-03-25 Published:2011-03-28

摘要/Abstract

摘要：

目的　探讨人脐血间充质干细胞（MSCs）移植修复大鼠脊髓损伤的作用及机制。方法　分离纯化人脐血MSCs；制备大鼠脊髓半横断损伤模型，随机分为三组，分别在术后3 d经尾静脉注射生理盐水、培养液和BrdU标记的MSCs。移植后7、14、21、28 d，采用BBB评分法评估各组大鼠脊髓功能恢复情况；免疫荧光双标法检测MSCs在脊髓内的迁移、存活和分化，免疫组织化学法检测炎症因子高迁移率族蛋白B1（HMGB1）和核因子（NF-κB）在脊髓损伤部位的表达规律。结果　移植后28 d，MSCs移植组大鼠肢体功能恢复明显，与生理盐水组和培养液组比较差别有统计学意义(P＜0.05)。移植后7、14、21d，脊髓损伤区及周边均可见大量Brdu+细胞，其中BrdU+GFAP+细胞约占53.3%，BrdU+NSE+细胞约占　22.15%。相同时间点MSCs移植组HMGB1和NF-κB的阳性表达率远低于生理盐水组和培养液组，差别有统计学意义(P＜0.05)。结论　人脐血MSCs移植后可替代损伤的神经细胞，并可减轻脊髓损伤后的炎症反应，从而促进大鼠脊髓功能的恢复。

关键词: 脐血, 间充质干细胞, 脊髓损伤, 修复

Abstract:

Objective　To explore the therapeutic effect and mechanism of mesenchymal stem cells (MSCs) derived from human umbilical cord blood on rat spinal cord injury.　Methods　MSCs were separated and purified from human umbilical cord blood. The spinal cord semi-transection was done on adult rats and then the animals were randomly divided into three groups which were treated with BrdU labeled MSCs, culture medium or saline respectively. Functional outcome was measured using the Basso-Beattie-Bresnehan (BBB) score method periodically up to 28 days post-surgery. MSCs survival and migration were observed by immunofluorescence double labeling techniques. The expression pattern of HMGB1 and NF-κB were detected by immunohistochemical techniques.　Results　BBB score showed that locomotor improvement in MSCs transplantation groups was much better than that of normal saline group and culture medium group at 28 days post-transplantation (P<0.05). BrdU-labeled MSCs could be detected in the lesion area at 7, 14 and 21 days post-transplantation. 53.3% of transplanted BrdU-positive cells were GFAP-positive, and 22.15% of them were NSE-positive. The positive rate of HMGB1 and NF-κB expression in MSCs transplantation group were lower than that in saline group and culture medium group, and there was a significant differences among three groups (P<0.05).　Conclusions　MSCs could promote functional recovery of the injuried spinal cord by replacing damaged nerve cells and alleviating inflammatory reaction.

Key words: Human umbilical cord blood, Mesenchymal stem cells, Spinal cord injury, Regeneration

中图分类号:

R749.1

孙　丽, 于　丽, 张华芳, 王　力, 魏志新. 人脐血MSCs移植修复大鼠脊髓损伤的效果及机制的初步探讨[J]. 中国临床解剖学杂志, 2011, 29(2): 202-207.

SUN 　Li, YU 　Li, ZHANG Hua-Fang, WANG 　Li, WEI Zhi-Xin. Effects and mechanism of the transplantation of human umbilical cord blood msenchymal stem cells to treat spinal cord injury[J]. Chinese Journal Of Clinical Anatomy, 2011, 29(2): 202-207.

参考文献

[1] 李德华,单伟,韩静,等
[G2]. 人脐静脉间充质干细胞向胰岛β样细胞诱导分化的实验研究
[J].中国临床解剖学杂志，2007，25（4）：424-426.

[2] 张炳强,于丽,张雪莉,等.丹参素联合生长因子体外诱导人脐血间充质干细胞分化为神经细胞的研究
[J].神经解剖学杂志,2009,25（1）:63-67.

[3] Cho SR, Kim YR, Kang HS, et al.Functional recovery after the transplantation of neurally differentiated mesenchymal stem cells derived from bone barrow in a rat model of spinal cord injury
[J]. Cell Transplant,2009,18(12):1359-1368.

[4] Lee JK, Jin HK, Endo S, et al. Intracerebral transplantation of bone marrow-derived mesenchymal stem cells reduces amyloid-beta deposition and rescues memory deficits in Alzheimer's disease mice by modulation of immune responses
[J]. Stem Cells, 2010, 28(2):329-343.

[5] Lee HJ, Lee JK, Lee H, et al. The therapeutic potential of human umbilical cord blood-derived mesenchymal stem cells in Alzheimer's disease
[J]. Neurosci Lett, 2010, 481(1):30-35.

[6] Lee HJ, Lee JK, Lee H, et al. Human umbilical cord blood-derived mesenchymal stem cells improve neuropathology and cognitive impairment in an Alzheimer's disease mouse model through modulation of neuroinflammation
[J]. Neurobiol Aging, 2010,3(24)
[Epub ahead of print]

[7] Cizkova D, Rosocha J, Vanicky I, et al. Transplants of human mesenchymal stem cells improve functional recovery after spinal cord injury in the rat
[J]. Cell Mol Neurobiol,2006, 26(7-8): 1165-1178.

[8] Bae JS, Carter JE, Jin HK. Adipose tissue-derived stem cells rescue Purkinje neurons and alleviate inflammatory responses in Niemann-Pick disease type C mice
[J]. Cell Tissue Res,2010,340(2):357-369.

[9] Erlandsson H, Andersson U. The nuclear protein HMGB1 as a proinflammatory mediator
[J].Eur J Immunol,2004,34(6):1503-1512.

[10]Palmubo R，Binaehi ME. High mobility group boxl，a cue for stem cell recruitment
[J].Biochem Pharmacol, 2004, 68(6):1165-1170.

[11]Degryse B, Virgilio MD. The nuclear Protein HMGBI，a new kind of chemokine
[J] . FEBS Lett, 2003, 553(l-2):11-17.

[12]Palumbo R, Sampaolesi M, Marchis FD. Extracellular HMGB1,a signal of tissue damage, induces mesoangioblast migration and proliferation
[J].Cell Biol, 2004,164(3):441-449.

[13]Compagno M, Lim WK, Grunn A. Mutations of multiple genes cause deregulation of NF-kappaB in diffuse large B-cell lymphoma
[J]. Nature, 2009, 459(7247):717-721.

[14]Huttunen HJ, Fages C, Rauvala H. Receptor for advanced glycation end products(RAGE)-mediated neurite outgrowth and activation of NF-kappaB require the cytoplasmic domain of the receptor but different downstream signaling pathways
[J].J Biol Chem,1999,274(28):19919-19924.

[15]Bachis A, Colangelo AM, Vicini S, et al. Interleukin-10 prevents glutamate-mediated cerebellar granule cell death by blocking caspase-3-like activity
[J].J Neurosci ,2001,21(9):3104-3012.