| Objective:To compare the effect of different concentrations of human umbilical cord mesenchymal stem cells (hUC-MSCs) transplanted into the injury site directly in rats after spinal cord injury.Methods:The rat SCI model was made by NYU Impactor model-Ⅱtype weight drop apparatus (10g,25mm) at T10. Eighty female Wistar rats were randomly divided into four groups:DMEM control group(group A, n=20), hUC-MSCs grafted 1×104/μl post injury(group B, n=20), hUC-MSCs grafted 1×105/μl post injury(group C, n=20) and hUC-MSCs grafted 1×106/μl post injury(group D, n=20). HUC-MSCs or DMEM was transplanted into the injury site directly in each groups. A week post injury, ten microliter DMEM were transplanted into the injury site directly in group A. The same volume of hUC-MSCs, isolated and cultured in vitro, the final concentration was prepared as metioned above, were transplanted into the injury site directly in group B. C and D, respectively. The hindlimb functional recovery of rats after spinal cord injury in each group was evaluated by Basso Beattie Bresnahan (BBB) score, HE, immunohistochemistry staining and Biotinylated Dextran Amine (BDA) antergrade tracing. The area of cavity in injured spinal cord was evaluated by HE staining in each group. Also, the imigration, differentiation and survive of hUC-MSCs post transplantation in the injury site were observed by immunohistochemistry. The area of glial scar in the injury site in each group was calculated by integrated optical density of immunostaining against GFAP. The regeneration of coticospinal tract (CST) was observed by using the biotinylated dextran amine (BDA) antergrade tracing.Result:Compared with the control group, grafted hUC-MSCs showed larger effect on functional recovery, especially for group D. As BBB score showed, group B,C and D are higher than group A 4 weeks post injury and the difference exists until 8 weeks post injury. BBB score in group D(13.273±0.810) is higher than group A(8.091±0.639),group B(11.364±0.834),group C(12.909±0.868). Statistic difference existed between the four groups (P<0.05). HE staining indicates that the area of cavity in injured spinal cord in group B. C and D is smaller than group A. Group D is the smallest. The result of immunohistochemical staining shows hUC-MSCs labeled by Brdu could imigration, and survive 8 weeks post transplantation. Double immunohistochemical staining showed hUC-MSCs kept undifferentiated post injury in vivo. The area of glial scar in the injury site in in group B. C and D is smaller than group A. The integrated optical density against GFAP of each group was calculated and statistic difference existed between the four groups (P<0.05). Group D is the lowest. The result of BDA antergrade tracing indicates that the axonal regeneration extension and the regeneration of coticospinal tract is more obvious in group B,C and D. Part of the regenerative fibers could get through the injury site.Conclusion:hUC-MSCs after its transplantation could promote axonal regeneration and functional recovery in rats after SCI. The internal mechanism includes improving the microenviroment near the injury site and protecting the remained neurons. hUC-MSCs could survive for a long time in vivo and inhibit the formation of glial scar post its transplantation. Ten to the power of six is the better concentration for transplanted hUC-MSCs than other groups.
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