血管内皮生长因子对人急性白血病细胞凋亡影响及相关基因表达的临床与实验研究

目的：探讨血管内皮生长因子(VEGF)对人急性白血病细胞株HL-60凋亡及相关基因Bcl-2和Mcl-1表达的影响,并从临床水平验证VEGF对人急性白血病细胞凋亡的影响。方法：采用单细胞凝胶电泳、流式细胞术检测VEGF处理后的HL-60细胞凋亡率;采用RT-PCR方法和免疫细胞化学法分别检测VEGF处理的HL-60细胞的Bcl-2和Mcl-1的表达水平。采用免疫细胞化学方法分别检测8例初发和复发、14例完全缓解的急性白血病患儿及5例正常儿童骨髓细胞的VEGF和Mcl-1蛋白表达。结果：VEGF可以明显抑制人急性白血病细胞株HL-60的凋亡,并且可以对抗VP16诱导的细胞凋亡效应。VEGF孵育HL-60细胞株18h后,其细胞内的Bcl-2和Mcl-1的mRNA和蛋白表达水平明显增加。初发和复发组白血病患儿骨髓细胞的VEGF和Mcl-1蛋白表达水平高于完全缓解组。结论：VEGF可能通过同时提高Bcl-2和Mcl-1的mRNA和蛋白表达,抑制急性白血病细胞株HL-60的凋亡,可能是白血病的发病机制之一。VEGF,Bcl-2和Mcl-1表达的增加可能对人急性白血病细胞的发生发展及预后产生影响,有可能成为临床白血病预后的参考指标之一。

Effect of vascular endothelial growth factor on apoptosis and expression of Bcl-2 and Mcl-1 in acute leukemia cells

OBJECTIVE: To investigate the effect of vascular endothelial growth factor (VEGF) on the apoptosis of human acute leukemia HL-60 cell line and to analyze the role of the related apoptosis genes, such as Bcl-2 and Mcl-1, in the process of apoptosis of human acute leukemia cells. METHODS: HL-60 cells were treated with different concentrations of VEGF (2 microg/L, 20 microg/L or 100 microg/L ) or 20 mg/L of etoposide (VP16, an apoptosis inducter) alone or VEGF plus VP16. After 18 hrs of treatment, the apoptosis rate of HL-60 cells was detected by single-cell gel electrophoresis and flow cytometry. The expressions of Bcl-2 and Mcl-1 of HL-60 cells were detected by RT-PCR. The Control group did not receive any treatment. Immunocytochemistry was used to detect the VEGF and Mcl-1 protein in bone marrow cells from 8 patients with newly diagnosed or relapsed leukemia, 14 leukemia patients in complete remission, and from 5 normal children. RESULTS: Different concentrations of VEGF markedly inhibited the apoptosis of HL-60 cells and decreased the apoptosis induced by VP16 exposure. The Bcl-2 and Mcl-1 mRNA and protein in HL-60 cells treated with VEGF were significantly higher than those in the Control group. The expressions of VEGF and Mcl-1 protein in bone marrow cells of the newly diagnosed and relapsed patients were significantly higher than in patients in complete remission. CONCLUSION: VEGF can inhibit the apoptosis of HL-60 cells possibly through increasing the expressions of Bcl-2 and Mcl-1 mRNA and protein, which may represent one of the mechanisms responsible for human acute leukemia. The expressions of VEGF, Bcl-2 and Mcl-1 might be used as the markers for the prognostic evaluation of leukemia.