硫酸軟骨素酶ABC聯合BMSCs修復大鼠脊髓損傷的實驗研究_《中國修復重建外科雜志》

作者：

張純 , 賀西京 , 李浩鵬

西安交通大學醫學院第二附屬醫院骨二科（西安，710004）;

關鍵詞：

BMSCs 硫酸軟骨素酶ABC 脊髓損傷細胞治療大鼠

DOI：

10.7507/1002-1892.20130121

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目的探討硫酸軟骨素酶ABC（chondroitinase ABC，ChABC）聯合BMSCs對大鼠脊髓損傷的修復作用。方法取新生SD大鼠股骨、脛骨骨髓分離培養原代BMSCs。取成年雄性SD大鼠24只，體重200～230 g，制備脊髓擠壓傷模型后，隨機分為脊髓損傷對照組（A組）、BMSCs移植組（B組）、ChABC注射組（C組）和ChABC聯合BMSCs移植組（D組），每組6只。傷后7、14 d采用BBB評分法評價大鼠后肢運動功能；傷后14 d取材行HE染色計算損傷區面積；采用免疫熒光染色法觀察膠質纖維酸性蛋白（glial fibrillary acidic protein，GFAP）/硫酸軟骨素蛋白多糖（chondroitin sulfate proteoglycan，CSPG）和GFAP/ 生長相關蛋白43（growth associated protein 43，GAP43）雙標表達情況。結果傷后7 d各組大鼠均恢復了后肢3個關節的運動，各組BBB評分比較差異無統計學意義（P gt; 0.05）；傷后14 d A、B、C組大鼠后肢3個關節無負重拖動，D組大鼠足底位于負重位或偶爾足背負重步行，D組BBB評分高于其余3組（P lt; 0.05）。傷后14 d HE染色示，各組損傷區形成空洞，空腔內及其周邊存在大量巨噬細胞，空洞周圍被瘢痕組織包裹；D組損傷區面積顯著小于其余3組（P lt; 0.05）。傷后14 d，GFAP/CSPG雙重免疫染色示D組星形膠質細胞瘢痕損傷區明顯減少，未見空洞形成；相對于A組的熒光強度值結果顯示，C、D組顯著低于B組（P lt; 0.05）。GFAP/GAP43雙重免疫染色示D組GAP43免疫組織化學染色陽性纖維穿過損傷區；相對于A組的熒光強度值結果顯示，D組顯著高于B、C組（P lt; 0.05）。結論采用ChABC早期抑制星形膠質細胞分泌CSPG，并聯合BMSCs移植可促進神經纖維再生，對大鼠脊髓損傷有一定修復作用。

引用本文： 張純,賀西京,李浩鵬. 硫酸軟骨素酶ABC聯合BMSCs修復大鼠脊髓損傷的實驗研究. 中國修復重建外科雜志, 2013, 27(5): 541-546. doi: 10.7507/1002-1892.20130121 復制

1.	沈學峰. 大鼠脊髓損傷后二次繼發性損傷的發現及其治療. 西安: 第四軍醫大學: 2006.
2.	焦西英, 程希平, 錢新宏, 等. 脊髓壞死面積的測量方法介紹. 神經解剖學雜志, 2006, 22(3): 356-358.
3.	Fawcett JW, Asher RA. The glial scar and central nervous system repair. Brain Res Bull, 1999, 49(6): 377-391.
4.	Hofstetter CP, Schwarz EJ, Hess D, et al. Marrow stromal cells from guiding strands in the injuryed spinal cord and promote recovery. Proc Natl Acad Sci U S A, 2002, 99(4): 2199-2204.
5.	Woodlbury D, Schwarz EJ, Proclop DJ, et al. Adult rat and hunan bone marrow stromal cells ditterentiate into neurons. J Neurosci Res, 2000, 61(4): 364-370.
6.	Novikova LN, Brohlin M, Kingham PJ, et al. Neuroprotective and growth-promoting effects of bone marrow stromal cells after cervical spinal cord injury in adult rats. Cytotherapy, 2011, 13(7): 873-887.
7.	Zurita M, Otero L, Aguayo C, et al. Cell therapy for spinal cord repair: optimization of biologic scaffolds for survival and neural differentiation of human bone marrow stromal cells. Cytotherapy, 2010, 12(4): 522-537.
8.	Park KW, Eglitis MA, Mouradian MM. Protection of migral neurous by GDNF-engineered marrow cell trancplantation. Neurosci Res, 2001, 40(4): 315-323.
9.	Yazdani SD, Pedram M, Hafizi M, et al. A comparison between neurally induced bone marrow derived mesenchymal stem cells and olfactory ensheathing glial cells to repair spinal cord injuries in rats. Tissue Cell, 2012, 44(4): 205-213.
10.	Pal R, Gopinath C, Rao NM, et al. Functional recovery after transplantation of bone marrow-derived human mesenchymal stromal cells in a rat model of spinal cord injury. Cytotherapy, 2010, 12(6): 792-806.
11.	Zhang JF, Zhao FS, Wu G, et al. Therapeutic effect of co-transplantation of neuregulin 1-transfected-Schwann cells and bone marrow stromal cells on spinal cord hemisection syndrome. Neuroscience, 2011, 497(2): 128-133.
12.	祖波, 尹宗生. 機械性脊髓損傷模型的建立及其評價. 中國臨床康復, 2006, 10(12): 151-153.
13.	Coumans JV, Lin TT, Dai HN, et al. Axonal regeneration and functional recovery after complete spinal cord transection in rats by delayed treatment with transplants and neurotrophins. J Neurosci, 2001, 21(23): 9334-9344.
14.	Benowitz LI, Popovich PG. Inflammation and axon regeneration. Curr Opin Neurol, 2011, 24(6): 577-583.
15.	游思維, 蘇國輝. 促進中樞神經損傷再生的探索之路. 中國處方藥, 2004, 3(2): 30-33.
16.	Rhodes KE, Moon LD, Fawcett JW. Inhibiting cell proliferation during formation of the glial scar effect on axon regeneration in the CNS. Neuroscience, 2003, 120(1): 41-56.
17.	Tang X, Davies JE, Davies SJ. Changes in distribution, cell associations and protein expression levels of NG2, neurocan, phosphacan, brevican, versican V2 and tenascin-C during acute to chronic maturation of spinal cord scar. J Neurosci Res, 2003, 71(3): 427-444.
18.	Minor K, Tang X, Kahrilas G, et al. Decorin promotes robust axon growth on inhibitory CSPGs and myelin via a direct effect on neurons. Neuroboil Dis, 2008, 32(1): 88-95.
19.	Willerth SM, Sakiyama-Elbert SE. Cell therapyfor spinal cord regeneration. Adv Drug Deliv Rev, 2008, 60(2): 263-276.
20.	Evans GR. Peripheral nerve injury: a review and approach to tissue engineered constructs. Anay Rec, 2001, 263(4): 396-404.
21.	Ruff CA, Wilcox JT, Fehlings MG, et al. Cell-based transplantation strategies to promote plasticity following spinal cord injury. Exp Neurol, 2012, 235(1): 78-90.
22.	Wu B, Sun L, Li P, et al. Transplantation of oligodendrocyte precursor cells improves myelination and promotes functional recovery after spinal cord injury. Injury, 2012, 43(6): 794-801.

1. 沈學峰. 大鼠脊髓損傷后二次繼發性損傷的發現及其治療. 西安: 第四軍醫大學: 2006.
2. 焦西英, 程希平, 錢新宏, 等. 脊髓壞死面積的測量方法介紹. 神經解剖學雜志, 2006, 22(3): 356-358.
3. Fawcett JW, Asher RA. The glial scar and central nervous system repair. Brain Res Bull, 1999, 49(6): 377-391.
4. Hofstetter CP, Schwarz EJ, Hess D, et al. Marrow stromal cells from guiding strands in the injuryed spinal cord and promote recovery. Proc Natl Acad Sci U S A, 2002, 99(4): 2199-2204.
5. Woodlbury D, Schwarz EJ, Proclop DJ, et al. Adult rat and hunan bone marrow stromal cells ditterentiate into neurons. J Neurosci Res, 2000, 61(4): 364-370.
6. Novikova LN, Brohlin M, Kingham PJ, et al. Neuroprotective and growth-promoting effects of bone marrow stromal cells after cervical spinal cord injury in adult rats. Cytotherapy, 2011, 13(7): 873-887.
7. Zurita M, Otero L, Aguayo C, et al. Cell therapy for spinal cord repair: optimization of biologic scaffolds for survival and neural differentiation of human bone marrow stromal cells. Cytotherapy, 2010, 12(4): 522-537.
8. Park KW, Eglitis MA, Mouradian MM. Protection of migral neurous by GDNF-engineered marrow cell trancplantation. Neurosci Res, 2001, 40(4): 315-323.
9. Yazdani SD, Pedram M, Hafizi M, et al. A comparison between neurally induced bone marrow derived mesenchymal stem cells and olfactory ensheathing glial cells to repair spinal cord injuries in rats. Tissue Cell, 2012, 44(4): 205-213.
10. Pal R, Gopinath C, Rao NM, et al. Functional recovery after transplantation of bone marrow-derived human mesenchymal stromal cells in a rat model of spinal cord injury. Cytotherapy, 2010, 12(6): 792-806.
11. Zhang JF, Zhao FS, Wu G, et al. Therapeutic effect of co-transplantation of neuregulin 1-transfected-Schwann cells and bone marrow stromal cells on spinal cord hemisection syndrome. Neuroscience, 2011, 497(2): 128-133.
12. 祖波, 尹宗生. 機械性脊髓損傷模型的建立及其評價. 中國臨床康復, 2006, 10(12): 151-153.
13. Coumans JV, Lin TT, Dai HN, et al. Axonal regeneration and functional recovery after complete spinal cord transection in rats by delayed treatment with transplants and neurotrophins. J Neurosci, 2001, 21(23): 9334-9344.
14. Benowitz LI, Popovich PG. Inflammation and axon regeneration. Curr Opin Neurol, 2011, 24(6): 577-583.
15. 游思維, 蘇國輝. 促進中樞神經損傷再生的探索之路. 中國處方藥, 2004, 3(2): 30-33.
16. Rhodes KE, Moon LD, Fawcett JW. Inhibiting cell proliferation during formation of the glial scar effect on axon regeneration in the CNS. Neuroscience, 2003, 120(1): 41-56.
17. Tang X, Davies JE, Davies SJ. Changes in distribution, cell associations and protein expression levels of NG2, neurocan, phosphacan, brevican, versican V2 and tenascin-C during acute to chronic maturation of spinal cord scar. J Neurosci Res, 2003, 71(3): 427-444.
18. Minor K, Tang X, Kahrilas G, et al. Decorin promotes robust axon growth on inhibitory CSPGs and myelin via a direct effect on neurons. Neuroboil Dis, 2008, 32(1): 88-95.
19. Willerth SM, Sakiyama-Elbert SE. Cell therapyfor spinal cord regeneration. Adv Drug Deliv Rev, 2008, 60(2): 263-276.
20. Evans GR. Peripheral nerve injury: a review and approach to tissue engineered constructs. Anay Rec, 2001, 263(4): 396-404.
21. Ruff CA, Wilcox JT, Fehlings MG, et al. Cell-based transplantation strategies to promote plasticity following spinal cord injury. Exp Neurol, 2012, 235(1): 78-90.
22. Wu B, Sun L, Li P, et al. Transplantation of oligodendrocyte precursor cells improves myelination and promotes functional recovery after spinal cord injury. Injury, 2012, 43(6): 794-801.

《中國修復重建外科雜志》

硫酸軟骨素酶ABC聯合BMSCs修復大鼠脊髓損傷的實驗研究

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《中國修復重建外科雜志》

硫酸軟骨素酶ABC聯合BMSCs修復大鼠脊髓損傷的實驗研究

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