Abstract: BACKGROUND: As the incidence of spinal cord injury increases with the years and axon regeneration after spinal cord injury was very difficult. How to promote the recovery from spinal cord injury and improve the transplantation efficiency of stem cells and other therapeutic cells after spinal cord injury has been the focus of clinical and scientific research.  
OBJECTIVE: To establish the efficient transplantation and replacement of mouse spinal cord microglia in the spinal cord injury model.
METHODS: CX3CR1 creER-/+::LSL-BDNF-/+-tdTomato mice, CX3CR1+/GFP mice, β-actin GFP mice and C57 BL/6J wild-type mice at 8-10 weeks of age were selected. According to the requirements of the experiment, they were randomly divided into six groups. (1) Sham operation group: eight C57 BL/6J wild-type mice were used when only the lamina was removed without injury. (2) Spinal cord contusion injury group: eight C57 BL/6J wild-type mice were used. (3) Spinal cord crush injury group: eight C57 BL/6J wild-type mice were used. (4) Conjoined symbiotic spinal cord strike injury group: β-actin GFP mice with green fluorescent blood were surgically stitched together with C57 BL/6J wild-type mice, using eight β-actin GFP mice and eight C57 BL/6J wild-type mice. (5) Mr BMT-X Ray group (using PLX5622 to eliminate the spinal microglia and bone marrow transplantation with X-ray radiation): Bone marrow cells from four CX3CR1 creER-/+::LSL-BDNF-/+-tdTomato mice were extracted and transplanted into eight C57 BL/6J wild-type mice for spinal cord injury modeling. (6) Mr BMT-Busulfan group (using PLX5622 to eliminate the spinal microglia and bone marrow transplantation with Busulfan): Bone marrow cells from four CX3CR1+/GFP mice were transplanted into eight C57 BL/6J wild-type mice. The percentage of cell transplantation replacement in this group was observed, and the spinal cord injury model was not established in this group. The sham operation group, spinal cord contusion injury group and spinal cord crush injury group were sampled by perfusion on day 14 after spinal cord injury. The conjoined symbiotic spinal cord strike injury group was sampled by perfusion on day 7 after spinal cord injury. Mr BMT-X Ray group was sampled by perfusion on day 28 after spinal cord injury. Mr BMT-Busulfan group was sampled by perfusion on day 28 after transplantation. The sampling site was a 1.2 cm long spinal cord with the T10 segment as the center. In the Mr BMT-X Ray group and Mr BMT-Busulfan group, additional mouse brain tissue was retained to see if it would lead to brain transplantation and replacement. The number and proportion of transplanted and replaced cells in the damaged area were measured using transgenic mice, symbiosis and immunofluorescence.  
RESULTS AND CONCLUSION: Compared with the traditional peripheral blood transplantation (9.8%) of mice in the conjoined symbiotic spinal cord strike injury group, the new transplantation methods, Mr BMT-X Ray and Mr BMT-Busulfan, could greatly improve the proportion of spinal microglia transplantation and replacement, which could reach 84.8% and 95.6%, respectively. The difference was significant (P < 0.05). The results showed that Mr BMT-X Ray and Mr BMT-Busulfan could achieve efficient replacement of spinal microglia cells, and could improve the problems of low cell transplantation efficiency, few survival numbers and unclear differentiation of the traditional cell transplantation methods. In addition, Mr BMT-X Ray can only replace the microglia in the spinal cord, while Mr BMT-Busulfan could avoid brain inflammation and injury caused by X-ray radiation transplantation.

Key words: spinal cord injury, cell transplantation and replacement, microglia, macrophage, Mr BMT-X Ray, Mr BMT-Busulfan