骨髓巨噬细胞M1/M2亚群失衡对免疫介导的新型再生障碍性贫血模型小鼠发病的影响

OBJECTIVE: To investigate the role of macrophages (Mø) in the pathogenesis of modified immune-mediated aplastic anemia (AA) mice model. METHODS: Before the establishment of the F1 AA mice model by total-body irradiation combined with allogeneic lymphocyte infusion, the mice of the CLO+AA group were treated with clodronate (CLO) liposomes to remove macrophages, and those of the PBS+AA group were treated with phosphate-buffered saline (PBS) liposomes and used as control. The severity of AA was observed by bone marrow (BM) pathological examination and peripheral blood cell count. Flow cytometry (FCM) was used to detect the CD4(+)/CD8(+) T lymphocyte subsets in the BM and Mø subsets in the BM and spleen of each group. The levels of IFN-γ, TNF-α, G-CSF, GM-CSF, EPO, and TPO in the peripheral blood were detected using enzyme-linked immunosorbent assay. Finally, the relationships between inflammatory factors and Mø subsets were analyzed. RESULTS: The BM fatty conversion of mice in the CLO+AA group was significantly alleviated compared with the PBS+AA group. Hemoglobin counts were (91.50±31.63) and (110.65±24.15) g/L, respectively, and the platelet counts were (90.85±121.90)× 10(6)/L and (461.13±483.45)×10(6)/L, respectively. The differences were all statistically significant (all P<0.05). After removing macrophages, the proportions of CD4(+) and CD8(+) T lymphocytes in BM of mice in the CLO+AA group decreased, but the reduction of CD8(+) T cells was more significant. The proportions of CD4(+) T cells and CD8(+) T cells in BM of the PBS+AA group were (18.5±10.17)％ and (36.23±6.40)％, respectively, and in the CLO+AA group were (7.58±8.00)％ and (6.67±5.78)％, respectively. Similarly, the percentage of macrophages in the spleen and BM in the CLO+AA group was significantly reduced compared with the PBS+AA group, most of which were M1 macrophages (P<0.05). The levels of IFN-γ in peripheral blood of the PBS+AA and CLO+AA groups were (602.37±104.62) ng/L and (303.01± 87.22) ng/L, respectively, the levels of TNF-α were (34.46±1.42) ng/L and (23.25±4.21) ng/L, respectively, the levels of GM-CSF were (9.32 ± 2.00) ng/L and (64.85±12.25) ng/L, respectively, the levels of G-CSF were (5 891.78±2 632.39) ng/L and (17 784.16±488.36) ng/L, respectively, the levels of EPO were (9 667.31±4 501.95) ng/L and (2 078.02±897.56) ng/L, respectively, and the levels of TPO were (6.36±2.09) ng/L and (11.67±2.86) ng/L, respectively (all P<0.05). CONCLUSION: This study confirmed that macrophages were involved in the pathogenesis of AA, and the degree of BM damage in AA mice was improved by removing macrophages in advance. The imbalance of M1/M2 macrophages and the changes of IFN-γ and TNF-α may be important mechanisms that eventually lead to AA.