神经毡蛋白-1在急性髓系白血病细胞中的表达和作用

Expression and Effect of Neuropilin-1 in Acute Myeloid Leukemic Cells

本研究旨在探索神经毡蛋白(neuropilin-1,NRP-1)和NRP-2在髓系白血病细胞中的表达及抑制NRP-1基因表达后对白血病细胞生长和迁移的影响。采用RT-PCR观察NRP-1和NRP-2mRNA在24例急性髓系白血病(AML)患者骨髓单个核细胞(MNC)及7种髓系白血病细胞系中的表达;以小干扰RNA(siRNA)干扰NRP-1基因在HEL细胞的表达,用MTT和细胞迁移试验观察细胞增殖、迁移的变化。结果显示:NRP-1在24例AML患者骨髓MNC中表达,其阳性率为100%,显著高于正常对照组67%(p=0.03)。79%AML患者表达NRP-2,67%正常人表达NRP-2,两者无明显差异(p=0.6)。NRP-1的表达与AML患者骨髓、外周血原始细胞比例呈正相关(r=0.5,r=0.4,p<0.05)。NRP-1和NRP-2mRNA分别在6/7和3/7种髓系白血病细胞系表达。用siRNA转染HEL细胞24小时后NRP-1mRNA和蛋白表达水平明显降低,当有VEGF165作用时,对照组细胞增殖明显增加,而干扰组无变化(MTT值:对照组vs干扰组:0.6±0.01vs0.49±0.02,p<0.001)。转染24小时后干扰组细胞迁移显著降低,其迁移细胞数:干扰组vs对照组为500±17vs123±7(p<0.001)。结论:NRP-1在AML患者骨髓MNC中表达增高,并在髓系白血病细胞的增殖和迁移中起重要作用。

This study was aimed to investigate the expression of neuropilin-1 （NRP-1） and NRP-2 mRNA in myeloid leukemia cells and the effect of NRP-1 on cell proliferation and migration. The expressions of NRP-1 and NRP- 2 mRNA in bone marrow mononnclear cells of 24 patients with acute myeloid leukemia and in 7 myeloid leukemic cell lines （HL-60, KGIa, NB4, U937, HEL MEG01 and K562 ） were detected by RT-PCR. The effects of NRP-1 interfered with siRNA on proliferation and migration in leukemic cell line HEL were examined by MTT and migration test. The results showed that the expression of NRP-1 mRNA was found in bone marrow mononuclear cells （BMMNCS） of 24 AML patients, the positive rate was 100% and significantly higher than that in control group （positive rate 67% ）. The expressions of NRP-2 mRNA were seen in 79% AML patients and in 67% health control, there was no significant difference between them. The increased NRP-1 expression was directly correlated with the blast percentage in both peripheral blood and bone marrow of AML patients （ r = 05, r = 0. 4, p 〈 0.05 ）. The expressions of NRP-1 and NRP-2 mRNA were observed in 6/7 and 3/7 myeloid leukemic cell lines respectively. After HEL cells were transfected with siRNA for 24 hours, the expression levels of NRP-1 mRNA and protein decreased obviously. Under VEGF action, the cell number in control group significantly increased, while the cell proliferation in interfered group had been not changed. After being transfected for 24 hous, the migration in interfered group decreased significantly. It is concluded that the higher level of NRP-1 mRNA is expressed in bone marrow mononucler cells of leukemia patients and plays a pivotal role in proliferation and migration of myeloid leukemic cells. Inhibition of NRP-1 functions may provide a new therapeutic strategy for AML.