• Lin Zhong, Hui Zhang, Weilu Gao, and Zongsheng Yin.
• Department of Orthopedics, the First Affiliated Hospital of Anhui Medical University, Hefei Anhui, 230022, PR China.
• Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2012 Jul 1; 26 (7): 855-60.

ObjectiveTo compare single cell suspension of neural stem cells (NSCs) with neurospheres transplantation for spinal cord injury (SCI) so as to explore the therapeutic effectiveness of two NSCs transplantation methods for SCI.MethodsThe NSCs were isolated from the spinal cord of adult Sprague Dawley (SD) rats, purified and cultured. At passage 3, the cells were identified by Hoechst33342, Nestin staining, and glial fibrillary acidic protein staining for differentiated cells. Sixty adult SD rats (weighing 230-250 g) were made the SCI models at T10 level with modified Allen method and randomly divided into 3 groups (20 rats in each). The injury sites were treated by injecting 5 microL saline (group A), 5 microL single cells suspensions of NSCs at passage 3 (group B), and 5 microL neurospheres cell suspensions at passage 3 (group C). At preoperation and 3, 7, 14, 21, and 28 days after operation, the locomotor functions of each group were assessed using the Basso, Beattie, and Bresnahan (BBB) rating scale. HE staining was applied to observe the morphology of spinal cord. Subsequently immunofluorescence staining was used to observe microtubule-associated protein 2 (MAP-2).ResultsThe cells cultured were NSCs by morphological observation and immunofluorescence staining. After 3 days of modeling surgery, BBB score significantly decreased when compared with preoperative score, and there was no significant difference among 3 groups at 3 and 7 days (P > 0.05). BBB score increased in different degrees with time; at 14, 21, and 28 days, BBB score of groups B and C was better than that of group A, and group C was better than group B, showing significant differences (P < 0.05). HE staining showed that spinal cord structure of group C was more clear than that of groups A and B, and had less scar. There was no significant difference in the number of MAP-2 positive cells among 3 groups at 3 and 7 days (P > 0.05). At 14, 21, and 28 days, the number of MAP-2 positive cells of groups B and C was significantly more than that of group A, and group C was more than group B, showing significant differences (P < 0.05).ConclusionTransplantation of neurospheres suspension compared with single cell can significantly promote NSCs to differentiate into neurons and is conducive to recover the lower extremity function after SCI.

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