酪氨酸蛋白磷酸酶2高表达对p210bcr/abl诱导的慢性粒细胞白血病细胞增殖与凋亡抵抗的影响  

Overexpression of Shp-2 is associated with the unlimited growth and apoptosis resistance of p210 bcr-abl-mediated chronic myeloid leukemia

作　　者：朱旭贞 虞瑛姿[1] 方永明[2] 梁赟[1] 吕庆华[2] 徐荣臻[1] 

机构地区：[1]浙江大学医学院附属第二医院血液科 [2]浙江大学肿瘤研究所

出　　处：《中华医学杂志》2005年第27期1903-1906,共4页National Medical Journal of China

基　　金：国家自然科学基金资助项目(30270572);浙江省自然科学基金资助项目(Y204113)

摘　　要：目的研究Src癌基因同源的酪氨酸蛋白磷酸酶2(Shp2)在慢性粒细胞白血病(CML)细胞中的表达,及其在p210bcr/abl诱导的白血病细胞恶性增殖和凋亡抵抗中的作用。方法收集25例p210bcr/abl阳性CML患者白血病细胞样本,8例非肿瘤病人骨髓和10例正常人外周血细胞样本作阴性对照,K562和KU812白血病细胞系作为p210bcr/abl阳性对照,KG1白血病细胞作为Shp2阳性对照。用Western印迹技术定量分析与比较Shp2在白血病细胞和正常骨髓造血细胞中的表达情况,通过特异性抑制剂分别下调Shp2和白血病融合基因p210bcr/abl后,观察Shp2表达对p210bcr/abl阳性白血病细胞增殖与凋亡的作用。细胞增殖与凋亡检测应用流式细胞仪。结果(1)磷酸化Shp2蛋白在92%患者CML白血病细胞样本中呈高表达状态,而在8例非肿瘤患者骨髓和10例正常人外周血细胞中低表达或不表达。磷酸化Shp2/β肌动蛋白比值分别为0.91±0.62、0.16±0.09和0.03±0.05(P均<0.01)。(2)下调Shp2蛋白表达后,白血病细胞凋亡率从4.89%上升到38.69%(P<0.01),而S期细胞从33.6%下降到10.8%(P<0.01)。(3)特异性抑制p210bcr/abl融合基因表达蛋白后,白血病细胞中磷酸化Shp2蛋白明显下降,同时出现明显细胞凋亡与生长抑制现象。结论(1)磷酸化Shp2蛋白在慢性粒细胞白血病细胞患者中呈过度表达状态,并且与白血病细胞恶性增殖与凋亡抵抗密切相关。(2)bcr/abl融合基因阳性患者的白血病细胞中Shp2的过度表达可能由p210bcr/abl蛋白激活引起。Objective To investigate the expression level of Shp-2 tyrosine phosphatase in chronic myeloid leukemia (CML) and its relationship with the unlimited growth and apoptosis resistance of p210 bcr/abl-induced malignant cells. Methods In this study, p210 bcr/abl positive leukemia cell specimens were obtained from 25 CML cases, meanwhile, bone marrow and peripheral blood cell samples from 8 non-tumor individuals and 10 normal individuals were used as p210 bcr/abl negative controls. K562 and KU812 leukemia cells were used as p210 bcr/abl positive controls, and KG-1 leukemia cell line was used as Shp-2 positive control. Specimens of peripheral blood and bone marrow of 25 adult patients of chronic myelocytic leukemia, 15 males and 10 females, aged 28~64, were collected. Specimens of bone marrow of 8 basically healthy adult volunteers and specimens of peripheral blood of 10 healthy adult volunteers were used as controls. The total cell protein was collected and the expression of Shp-2 was examined by Western blotting. Human leukemia cells of the line K562 were cultured. Shp-2 specific sense and antisense oligonucleotides were added into the culture fluid respectively. The cell apoptosis was detected by flow cytometry (FCM). STI571, specific inhibitor of p210 bcr/abl was added into the cultured fluid of K562 cells, then Western blotting and FCM were used to detect the protein expression of Shp-2 and p210 bcr/abl, and cell apotosis. Results Phospharylated Shp-2 (pShp)-2 protein was overexpressed in 92% (23/25) of the CML cells, but lowly expressed or not expressed in the normal hematopoietic cells. The mean pShp-2 protein/β-actin ratio of the primary CML leukemia cells was 0.91±0.62, significantly higher than those of the normal bone marrow cells and peripheral blood hematopoietic cells (0.16±0.09 and 0.03±0.05 respectively, both P<0.01). The apoptotic rates of the CML cells treated by Shp-2 specific antisense oligonucleotide of the concentrations of 1 μmol/L and 4 μmol/L respectively for 72h was 7.98% and 20.29% resp

关 键 词：慢性粒细胞白血病 蛋白磷酸酶 细胞增殖 高表达 酪氨酸 抵抗 bcr/abl融合基因 Shp-2蛋白 细胞恶性增殖 人外周血细胞 Western Src癌基因 白血病细胞系 骨髓造血细胞 特异性抑制剂 Β-肌动蛋白 基因表达蛋白 阳性对照 过度表达 

分 类 号：R733.72[医药卫生—肿瘤] R749.16[医药卫生—临床医学]