白血病细胞TGF-β1含量减少及外源性TGF-β1基因对HL-60细胞的作用

目的：研究白血病细胞是否存在转化生长因子β1（TGF-β1）量的异常以及这种异常在白血病发病机制中的可能作用。 方法： 应用ELISA法检测白血病细胞株和原代白血病细胞培养上清TGF-β1水平，进一步应用脂质体介导基因转移法将TGF-β1基因转染HL-60细胞，采用有限稀释法及G418筛选得到抗性细胞，应用RT-PCR、白血病细胞集落培养、裸鼠移植瘤成瘤试验、片段化DNA分析、流式细胞术检测HL-60细胞内TGF-β1、bcl-2、端粒酶反转录酶（hTERT）mRNA的表达变化等方法了解外源性TGF-β1基因对HL-60细胞体内外增殖、凋亡的影响。 结果： 白血病细胞株和原代白血病细胞培养上清TGF-β1水平明显低于正常对照组(P<0.01)，转染TGF-β1基因后，HL-60细胞内TGF-β1 mRNA表达上调，bcl-2、hTERT mRNA表达下调，细胞体外增殖受抑制，集落形成抑制率达78.3%，裸鼠移植瘤生长受抑制，荷瘤鼠生存期延长，细胞呈现凋亡特征性的梯形条带，凋亡比例达73.4%。 结论： 内源性TGF-β1水平降低是白血病的发病机制之一,外源性TGF-β1基因能够抑制HL-60细胞增殖，诱导细胞凋亡,该基因通过下调bcl-2、hTERT mRNA的表达而发挥作用。

Deficiency of TGF - β1 in leukemic cells and the effects of exogenous TGF - β1gene on HL- 60 cells

AIM: To explore the existence of deficiency of TGF-β1 in leukemia cells and its possible mechanism in the pathogenesis of leukemia. METHODS: The levels of TGF-β1 were detected by ELISA in the cultured supernatant of leukemia cell lines and primary cells from patients with acute leukemia. TGF-β1 gene was transduced into HL-60 cells by lipofectin-mediated DNA transfection. In the presence of G418, the HL-60 clone expressing TGF-β1 was selected. The effects of exogenous TGF-β1 gene on the proliferation and apoptosis of HL-60 cells were studied by leukemic colony assay, tumorigenicity in athymic nude mice, DNA fragmentation and cell cycle analysis. The expression of intrinsic TGF-β1, bcl-2 oncogene, hTERT mRNA on the apoptosis of HL-60 cells induced by exogenous TGF-β1 gene were detected by RT-PCR. RESULTS: The levels of TGF-β1 were obviously lower in the supernatant of leukemia cell lines and primary cells from patients with acute leukemia, as compared with normal controls (P<0.01). The expression of intrinsic TGF-β1 mRNA was up-regulated, while the expression of bcl-2 and hTERT were down-regulated in HL-60 cells after transfected with exogenous TGF-β1 gene. Clongenicity was reduced 78.3% in vitro and apoptosis cells were 73.4% and tumorigenicity was weakened in vivo in athymic nude mice. CONCLUSIONS: The lower TGF-β1 expression may contribute to leukemogenesis. Exogenous TGF-β1 gene inhibits HL-60 cell proliferation and induces cells apoptosis via up-regulating the expression of intrinsic TGF-β1 and down-regulating the expression of bcl-2 and hTERT.