骨髓间充质干细胞分泌基质细胞衍生因子1对异体T淋巴细胞和白血病细胞HL-60增殖的影响

背景：骨髓间充质干细胞持续分泌的基质细胞衍生因子1与其受体CXCR4的相互作用，不仅在细胞的炎症反应、造血干细胞的迁移与归巢等生物学过程中发挥作用，还在肿瘤的发生、转移、复发、免疫耐受及血管新生过程中扮演重要角色。 目的：探讨骨髓间充质干细胞分泌的基质细胞衍生因子1对T淋巴细胞及白血病细胞HL-60增殖的影响。 设计：细胞学体外对照观察。 材料：骨髓标本来自本院血液科异基因骨髓移植供者及骨髓象正常的非白血病患者，T淋巴细胞来源于健康志愿者，急性髓性白血病细胞株HL-60为本实验室液氮冷冻保存，CXCR4单克隆抗体12G5为eBioscience公司产品。 方法：应用Ficoll密度梯度离心法分离骨髓间充质干细胞，传至第3代后收集培养5 d的细胞上清，离心去沉淀后备用。应用尼龙棉柱法分离异体外周血T淋巴细胞，调整细胞密度为2×109 L-1备用。T淋巴细胞与骨髓间充质干细胞的共培养实验设立3组：T淋巴细胞组、T淋巴细胞+细胞上清组、T淋巴细胞+细胞上清+单抗组。HL-60细胞与骨髓间充质干细胞的共培养实验设立3组：HL-60细胞组、HL-60细胞+细胞上清组、HL-60细胞+细胞上清+单抗组。 主要观察指标：MTT法检测T淋巴细胞及HL-60细胞在应用单克隆抗体12G5前后的增殖情况。 结果：与T淋巴细胞组比较，T淋巴细胞+细胞上清组、T淋巴细胞+细胞上清+单抗组的吸光度值均明显降低（P < 0.05），后两组间比较差异无显著性意义（P > 0.05）。与HL-60细胞组比较，HL-60细胞+细胞上清组吸光度值明显升高（P < 0.05）；与HL-60细胞+细胞上清组比较，HL-60细胞+细胞上清+单抗组的吸光度值明显降低（P < 0.05）。 结论：加入单克隆抗体12G5阻断基质细胞衍生因子1的生物学效应后，不影响骨髓间充质干细胞上清液抑制T淋巴细胞增殖的效果，但可以抑制白血病细胞HL-60的增殖。

Effects of stromal cell derived factor 1 secreted by bone marrow mesenchymal stem cells on proliferation of allogeneic T lymphocytes and acute myelocytic leukemia cell line HL-60

BACKGROUND: Interaction of bone marrow mesenchymal stem cells-secreted stromal cell-derived factor-1 (SDF-1) and its receptor CXCR4 plays an important role in inflammatory reaction, migration and homing of hemopoietic stem cells, as well in occurrence, transfer, relapse, immune tolerance of tumor and angiogenesis. OBJECTIVE: To investigate the effects of bone marrow mesenchymal stem cells-secreted SDF-1 on proliferation of T lymphocytes and acute myelocytic leukemia cell line HL-60. DESIGN: A cytology in vitro control study. MATERIALS: Bone marrow samples were collected from donors of allogeneic bone marrow transplantation and non-leukemia patients with normal bone marrow image, who were enrolled at the Department of Hematology of this hospital. T lymphocytes were harvested from healthy volunteers. Acute myelocytic leukemia cell line HL-60 was kept in this laboratory via liquid nitrogen frozen. CXCR4 monoclonal antibody 12G5 was purchased from eBioscience. METHODS: Bone marrow mesenchymal stem cells were isolated by Ficoll-Hypaque density gradient centrifugation. At the third passage, cell supernatant cultured for 5 days was obtained, and centrifuged for usage. T lymphocytes were harvested by nylon column, and cells were 2×109/L. T lymphocytes and bone marrow mesenchymal stem cells were co-cultured. There were 3 groups, T lymphocyte group, T lymphocyte + cell supernatant group, T lymphocyte + cell supernatant + monoclonal antibody group. HL-60 cells were co-cultured with bone marrow mesenchymal stem cells. There were 3 groups, HL-60 cell group, HL-60 cell + cell supernatant group, and HL-60 cell + cell supernatant + monoclonal antibody group. MAIN OUTCOME MEASURES: Proliferation of T lymphocytes and HL-60 cells was examined by MTT prior to and following treatment of monoclonal antibody 12G5. RESULTS: Compared with the T lymphocyte group, absorbance in the T lymphocyte + cell supernatant group and T lymphocyte + cell supernatant + monoclonal antibody group was significantly reduced (P < 0.05). No significant differences were detected between the T lymphocyte + cell supernatant group and T lymphocyte + cell supernatant + monoclonal antibody group (P > 0.05). Compared with the HL-60 cell group, absorbance was significantly increased in the HL-60 cell + cell supernatant group (P < 0.05). Compared with the HL-60 cell + cell supernatant group, absorbance was significantly decreased in the HL-60 cell + cell supernatant + monoclonal antibody group (P < 0.05). CONCLUSION: Blocking SDF-1 activity can inhibit proliferation of leukemia cells HL-60, but at the same time proliferation of allogeneic T lymphocytes inhibited by bone marrow mesenchymal stem cell supernatant was not obviously effected.