中国生物工程杂志

To investigate the effect of Hemangiopoietin (HAPO) on the hematopoiesis reconstitution in lethally irradiated Balb/c mice. Methods: In the live animal experiment, Balb/c mice underwent total body irradiation at 700 cGy 137Cs γ radiation and were treated after irradiation with HAPO or recombinant human granulocyte colony stimulating factor (rhG-CSF). Survival rate of mice after radiation, the number of bone marrow cells, blood routine, spleen node forming and colony forming units were detected. In vitro experiment, cells from bone marrow of Balb/c mice were washed and resuspended, then cultured in 96- well plates. HAPO or rhG-CSF were added in the culture system before or after irradiated for 24 hours or 72 hours. MTT assay was used to assess the viability of cells. Cells from bone marrow of Balb/c mice cultured in 24-well plates were irradiated, then cultured for another 3 weeks with HAPO in culture medium to assess ability of in vitro hematopoiesis reconstitution. Result: rhG-CSF and HAPO treated mice both showed increased survival rate and increased colony forming units. The changes of the peripheral WBC level after radiation were remarkable. The recovery of peripheral WBC number in HAPO group was quickest compared with that in rhG-CSF group and PBS control group. The number of bone marrow cells at day 14 of rhG-CSF group was higher than that in HAPO group, but the number of bone marrow cells at day 32 of rhG-CSF was lower than that in HAPO group. The number of bone marrow cells at day 42 of rhG-CSF was below normal. The number of bone marrow cells at day 42 of HAPO group was nearly normal. The number of CFU-GEMM in HAPO group was most compared with that in rhG-CSF group and PBS control group at day 7, 14 and 21 after radiation. The survival rate of cells after radiation in HAPO group was markedly higher than that in PBS control group, but the survival rate of cells after radiation in rhG-CSF group was no notable difference compared with that in PBS control group. In MTT assay, both HAPO and rhG-CSF incubation stimulated proliferation of bone marrow cell at 72 hours after radiation. Bone marrow cells formed Hematopoietic islands in HAPO group after radiation and were positive for sca-1 and CD31. CD31 positive endothelial cells increased around the Hematopoietic islands. There was no Hematopoietic islands formation, few CD31 positive endothelial cells and no sca-1 positive cells in PBS control group. Conclusion: we confirmed HAPO promoted hematopoiesis reconstitution in lethally irradiated Balb/c mice in vitro and in vivo study, increased survival rate of mice and stimulated hematopoietic stem cells proliferation.