| Background and objectiveSpinal cord injury (SCI) is a kind of serious injury occurred in central nervous system. It becomes nodus for medical research because of its high morbidity and high fatality rate. As the development of neurobiology and regenerative medicine, the treatment of SCI with stem cell transplantation has become a hotspot. Many kinds of stem cells such as bone marrow mesenchymal stem cells and cord blood stem cells have proved its therapeutical effect throw animal experiment and clinical treatment effect, but the further research and clinical application are limited because of some disadvantages such as its source and low quantity cells extracted. While adipose-derived stem cells throw off the disadvantages upon, and it has the advantages of wide source, easy large amount of in vitro expansion and multiplex differentiation potential potentiality. At present, domestic and overseas scholars research proves that the characteristics of ADSCs in vitro can be induced neuron-sample cells is superior than the other organizations sources of stem cells. The study of the the treatment of SCI with ADSCs transplantation is rarely reported. To explore and discuss the motor function and influence of various factors expression to spinal cord injury tissues throw ADSCs transplantation via vein in this experiment, and observe the therapeutical effect of SCI with ADSCs at the acute and chronic phase.MethodsCells cultured and identification:Adipose tissue were cleaned, sheared ground, digested, filtering, and made single-celled levitation liquid transferred to join DMEM/F12 medium (contain 10% FBS and 20g/L bFGF) train culture at sterile conditions. Take the train to the fourth generation ADSCs cells using flow cytometric analysis detection cell surface markers:CD29, CD34, CD44, CD45, HLA-ABC, HLA-DR.Animal model and grouping:All the experimental rats were divide into acute period group (group A) and chronic period group (group B). Group A divide again for A1 group (acute period control group), A2 group (acute period false transplantation group) A3 group (acute period transplantation group). Group B controls into B1 group (chronic period control group), B2 group (chronic period false transplantation group), B3 group (chronic period transplantation group). Spinal cord injury rat model production with self-made blow regulator, with abdominal vertical strike method, cause rats closed spinal cord injuries. Each rat was made mode for scoring after injury, with behavioral grade no more than 7 in one week into rating criteria.Cellular marker and transplantation and related observation index:take fourth-generation adipose-derived stem cells, with contain 10umol/L Brdu markers and collect cells. Cell transplants were taken in respectively for A3 and B3 rats with intravenous way for the first week and forth week postoperation, A2 and B2 group criterion with equal saline alternative, Al and B1 group do not anything others. BBB score were taken for each rat respectively on cell transplantation to partial executed and collected model rat damaged spinal cord segmental at 3d, 1w,2 w,3w,4w, And observat group damaged spinal cord segmental spinal cord tissues pathological changes, cell migration and differentiation by immunohistochemical (SP) dyeing method, and observe GFAP, NSE, Nestin, SYN expression neurons markers.ResultsBasso, Beattie, Bresnahan locomotor scales (BBB) motor scores of group A3 and group B3 at 4w after transplantation were significantly higher than control group and false transplantation group. The BBB scores of group A3 and group B3 were 15.33±1.21 and 13.66±1.03, the BBB scores of group A1 and group A2 were 9.60±1.34. and 10.00±1.87, the BBB scores of group B1 and group B2 were 10.60±1.82 and 10.80±2.05. there was significant difference between transplantation group and control group or false transplantation group (P<0.05), while there was not significant difference between the false transplantation group and control group (P> 0.05).Immunohistochemistry showed that, the ADSCs labeled by Brdu can be found in the injury spinal cord of group A3 and group B3 at 3d after ADSCs transplantation. A3 group and the B3 group The migrated cells per high magnification of A3 group A3 and the B3 group B3 were 15.03±3.67 and 10.27±3.13, while the difference was significant (P=0.024). The GFAP-positive rate began to decrease at 1w after ADSCs transplantation of group A3, the positive expression area ratio was 4.33%±1.58%, while the positive expression area ratio of group Al and group A2 were 8.68%±2.09% and 8.88%±2.33%, The ratio of group A3 was lower than that of group A1 and group A2 (P=0.002); The GFAP-positive expression area ratios of group B1, B2 and B3 were 4.92%±1.38%,4.52%±1.41% and 4.68%±1.35%, there were no difference among the three groups (P=0.253). The NSE, Nestin, SYN positive area ratio of group A3 and group B3 at 2w after ADSCs transplantation were 2.33%±1.56%,1.83%±1.44%,0.97%±1.13% and 1.53%±1.28%,1.03%±1.22%,0.56%±0.65%. The NSE positive area ratio of group A3 and group B3 was lower than that of the control group and flase transplant group (P<0.05), while the Nestin, SYN positive area ratios were significantly higher than that of the control group and flase transplant group (P<0.05). The GFAP, NSE positive area ratios of chronic groups were lower than that of the acute phase groups (P<0.05), while the Nestin, SYN positive area ratios of group A3 were higher than that group B3 (P< 0.05).Conclusions1. Via the tail vein graft ADSCs, can be seen in the damaged spinal cord tissue were labeled local ADSCs, that ADSCs can be moved to the damaged spinal cord tissue.2. The expression of neuron-related factors changed after ADSCs transplantation, the GFAP was lower, while the NSE, Nestin,SYN were higher, and BBB score was higher. These shown that ADSCs transplanted via vein can promote recovery of injuried spinal cord tissues of SCI rats.3. The SCI motor function recovery was better taken at acute phase than at chronic phase after ADSCs transplantation, shown that it is effective to improve the function of the damaged spinal cord tissues at the early phase of ADSCs transplanted. |
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