INDUCTING DIFFERENTIATION EFFECT OF SCIATIC NERVE EXTRACTS ON RABBIT ADIPOSE-DERIVED STEM CELLS IN VITRO_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

ZHAOBin ,  MAXinlong , SUNXiaolei , LIXiulan , ZHANGYang , GUOYue , YANGQiang

Cell Engineering Laboratory of Orthopaedic Institute, Tianjin Hospital, Tianjin, 300050, P. R. China;

Corresponding?author：

MAXinlong, Email: agzhb@sina.com

Keywords：

Adipose-derived stem cells; Schwann cells; Neuron; Sciatic nerve tissue extracts; Induction and differentiation

DOI：

10.7507/1002-1892.20150103

Video：

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Objective To study the inducting differentiation effect of the sciatic nerve extracts on rabbit adipose-derived stem cells (ADSCs) in vitro. Methods The ADSCs were isolated from 2 healthy 4-month-old New Zealand rabbits (weighing, 2.0-2.5 kg) and cultured to passage 3, which were pretreated with 10 ng/mL basic fibroblast growth factor (bFGF) for 24 hours before induction. Then the induction media containing the extracts of normal sciatic nerve (group B) and injured sciatic nerve at 3, 7, and 14 days (group C, group D, and group E) were used, and D-Hank was used in group A as blank control group. The morphological changes of the cells were observed. At 7 days of induction, the gene expressions of neuron-specific enolase (NSE), nestin (NES), and S-100 were detected by real-time fluorescent quantitative PCR. The S-100 protein expression was tested by immunocytochemical staining. Results At 4 days after induction, some ADSCs of groups C, D, and E showed the morphology of Schwann-like cells or neuron-like cells, the change of group D was more obvious; and the ADSCs of group A and B had no obvious change, which were still spindle. The S-100 immunocytochemical staining showed positive expression in groups C, D, and E (more obvious in group D) and negative expression in groups A and B. The gene expression of S-100 displayed time-dependent increases in groups C and D, which was significantly higher than that of groups A, B, and E (P<0.05), but no significant difference was found between groups C and D (P>0.05). The gene expression of NSE showed the same tendency to S-100, which reached the peak in group D; the gene expression of NSE in groups D and E was significantly higher than that of groups A, B, and C (P<0.05), and groups D and E showed significant difference (P<0.05). However, the gene expression of Nestin showed no significant difference among different groups (P>0.05). Conclusion The ADSCs can be induced to differentiate into Schwann-like cells or neuron-like cells with sciatic nerve extracts; and the early stage (3-7 days) after injury is the best time for stem cell transplantation.

Citation： ZHAOBin, MAXinlong, SUNXiaolei, LIXiulan, ZHANGYang, GUOYue, YANGQiang. INDUCTING DIFFERENTIATION EFFECT OF SCIATIC NERVE EXTRACTS ON RABBIT ADIPOSE-DERIVED STEM CELLS IN VITRO. Chinese Journal of Reparative and Reconstructive Surgery, 2015, 29(4): 477-482. doi: 10.7507/1002-1892.20150103 Copy

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13.	Luk JM, Wang PP, Lee CK, et al. Hepatic potential of bone marrow stromal cells:development of in vitro co-culture and intra-portal transplantation models. J Immunol Methods, 2005, 305(1):39-47.
14.	Choi KS, Shin JS, Lee JJ, et al. In vitro trans-differentiation of rat mesenchymal cells into insulin-producing cells by rat pancreatic extracts. Biochem Biophys Res Commun, 2005, 330(4):1299-1305.
15.	陳增海, 馬萬里, 高春正, 等. 人臍血干細胞培養及其向神經元樣細胞定向誘導分化的研究. 山東大學學報:醫學版, 2006, 44(11):1085-1089.
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20.	Hofstetter CP, Schwarz EJ, Hess D, et al. Marrow stromal cells form guiding strands in the injured spinal cord and promate recovery. Proc Natl Acad Sci U S A, 2002, 99(4):2199-2204.

1. Utsunomiya T, Shimada M, Imura S, et al. Human adipose-derived stem cells:potential clinical applications in surgery. Surg Today, 2011, 41(1):18-23.
2. Kingham PJ, Kalbermatten DF, Mahay D, et al. Adipose-derived stem cells differentiate into s Schwann cell phenotype and promote neurite outgrowth in vitro. Exp Neurol, 2007, 207(2):267-274.
3. Razavi S, Ahmadi N, Kazemi M, et al. Efficient transdifferentiation of human adipose-derived stem cells into Schwann-like cells:A promise for treatment of demyelinating diseases. Adv Biomed Res, 2012, 1:12.
4. 任志午, 趙喆, 盧世璧, 等. 大鼠脂肪干細胞向雪旺細胞誘導分化能力的研究. 中國應用生理學雜志, 2011, 27(4):385-388.
5. Marconi S, Castiglione G, Turano E, et al. Human adipose-derived mesenchymal stem cells systemically injected promote peripheral nerve regeneration in the mouse model of sciatic crush. Tissue Eng Part A, 2012, 18(11-12):1264-1272.
6. Shen CC, Yang YC, Liu BS. Peripheral nerve repair of transplanted undifferentiated adipose tissue-derived stem cells in a biodegradable reinforced nerve conduit. J Biomed Mater Res A, 2012, 100(1):48-63.
7. Liu GB, Cheng YX, Feng YK, et al. Adipose-derived stem cells promote peripheral nerve repair. Arch Med Sci, 2011, 7(4):592-596.
8. Brohlin M, Mahay D, Novikov LN, et al. Characterisation of human mesenchymal stem cells following differentiation into Schwann cell-like cell. Neurosci Res, 2009, 64(1):41-49.
9. 陳國玲, 劉執玉. 人結膜組織勻漿上清液體外誘導BMSCs定向分化的研究. 中華創傷雜志, 2012, 28(9):858-861.
10. Spradling A, Drummond-Barbosa D, Kai T. Stem cells find their niche. Nature, 2001, 414(6859):98-104.
11. Fuchs E, Tumbar T, Guasch G. Socializing with the neighbors:stem cells and their niche. Cell, 2004, 116(6):769-778.
12. 王曉萃, 吳金生, 付文玉, 等. 誘導大鼠間充質干細胞形成的神經元樣細胞的形態特征. 解剖學報, 2007, 38(1):107-110.
13. Luk JM, Wang PP, Lee CK, et al. Hepatic potential of bone marrow stromal cells:development of in vitro co-culture and intra-portal transplantation models. J Immunol Methods, 2005, 305(1):39-47.
14. Choi KS, Shin JS, Lee JJ, et al. In vitro trans-differentiation of rat mesenchymal cells into insulin-producing cells by rat pancreatic extracts. Biochem Biophys Res Commun, 2005, 330(4):1299-1305.
15. 陳增海, 馬萬里, 高春正, 等. 人臍血干細胞培養及其向神經元樣細胞定向誘導分化的研究. 山東大學學報:醫學版, 2006, 44(11):1085-1089.
16. Ye Y, Zeng YM, Wan MR, et al. Induction of human bone marrow mesenchymal stem cells differentiaion into neural-Like cells using cerebrospinal fluid. Cell Biochem Biophys, 2011, 59(3):179-184.
17. 張儒有, 鄭永日, 郭薇, 等. 不同損傷時期人腦挫裂傷對神經干細胞分化影響的體外研究. 航空航天醫學雜志, 2011, 22(6):641-643.
18. Ankeny DP, McTigue DM, Jakeman LB, et al. Bone marrow transplants provide tissue protection and directional guidance for axons after contusive spinal cord injury in rat. Exp Neurol, 2004, 190(1):17-31.
19. 孫曉雷,趙斌, 馬信龍, 等. 兔坐骨神經損傷后內源性生長因子含量的變化及意義. 中華創傷雜志, 2014, 30(5):471-474.
20. Hofstetter CP, Schwarz EJ, Hess D, et al. Marrow stromal cells form guiding strands in the injured spinal cord and promate recovery. Proc Natl Acad Sci U S A, 2002, 99(4):2199-2204.

Chinese Journal of Reparative and Reconstructive Surgery

INDUCTING DIFFERENTIATION EFFECT OF SCIATIC NERVE EXTRACTS ON RABBIT ADIPOSE-DERIVED STEM CELLS IN VITRO

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