Dnase

To prepare the host mice, all of the BALB/c mice were given 8 Gy of total body irradiation by linear accelerator before transplantation. To obtain the BMC and SMC suspensions, the C57BL/6 mice were killed by cervical dislocation and subsequently put into a 75% alcohol solution for 15 min. The bilateral femur, tibia, and spleen were harvested from the mice in an aseptic environment. The epiphyseal sides of the bones were cut off and the bone marrow was flushed out with RPMI 1640 medium (Gibco, Shanghai, China). RBC pyrolysis liquid (CW-Biotech, Beijing, China) was used to remove the erythrocytes from the cell suspension. For the most important step – removing the T lymphocyte cells – anti-Thy1.2 (Abcam, Shanghai, China) was used for 30 min at 4°C. The cell suspension was washed once with PBS and then treated with 10% immature rabbit complement (AbD Serotec, Beijing, China) and 2% DNase (CW-Biotech, Beijing, China) for 45 min at 37°C. Finally, the cell suspension was diluted to 2.0×10⁷ cells/ml with 1×PBS and stored at 4°C. The spleens were carefully milled through a 200-mesh cell sieve, and after lymphocyte cells separation medium (MPBio, Beijing, China) was added, the mononuclear cells were separated by density-gradient centrifugation. Finally, the cell suspension was diluted to 2.0×10⁶ cells/ml with 1×PBS and stored at 4°C.