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| SOCS1、SHP1在JAK2V617F突变阳性骨髓增殖性肿瘤中的表达及鲁索替尼的调控作用 |
| 谢旭磊1,2,杨圣俊1,郝洪岭1,王素云1*,王红杰3,齐峰2,成志勇2,刘贵敏2 |
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(1. 河北省人民医院血液内科, 石家庄 050051;
2. 保定市第一医院血液内科, 保定 071000;
3. 河北大学附属医院麻醉科, 保定 071000
*通信作者) |
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| 摘要: |
| 目的 研究JAK2V617F突变阳性骨髓增殖性肿瘤（MPN）组织中JAK2V617F突变量与磷酸化Janus激酶2（p-JAK2）、细胞因子信号转导抑制蛋白1（SOCS1）、含SH2结构域蛋白酪氨酸磷酸酶1（SHP1）表达的关系，并探讨JAK2抑制剂鲁索替尼（ruxolitinib）对JAK2V617F突变阳性人红白血病细胞系HEL细胞的增殖和HEL细胞中SOCS1、SHP1表达的影响。方法 纳入2012年7月至2016年8月于河北省人民医院和保定市第一医院就诊的48例JAK2V617F突变阳性的MPN患者为MPN组，同期24例贫血患者为对照组。采用免疫组织化学法检测骨髓组织标本中p-JAK2、SOCS1和SHP1的蛋白表达水平。SOCS1、SHP1蛋白表达量与JAK2V617F突变量的相关分析采用Spearman等级相关分析。用不同浓度（50、100、250、500、1 000 nmol/L）的鲁索替尼处理HEL细胞后，采用CCK-8法检测HEL细胞的活力，qPCR法检测MPN组织和HEL细胞中JAK2V617F突变量以及HEL细胞中JAK2、SOCS1、SHP1 mRNA表达水平，蛋白质印迹法检测HEL细胞中JAK2、p-JAK2、SOCS1、SHP1的蛋白表达水平。结果 （1）MPN组织中JAK2V617F/JAK2比值为（57.33±20.82）%，对照组为0%。MPN患者骨髓细胞质中p-JAK2、SOCS1、SHP1蛋白质表达量与对照组相比差异均有统计学意义（P均<0.01）。（2）MPN组织中SOCS1、SHP1蛋白表达量均与JAK2V617F突变量呈负相关（r=-0.648、-0.692，P均<0.05）。JAK2V617F/JAK2比值<50% MPN患者的SOCS1、SHP1蛋白表达水平高于JAK2V617F/JAK2比值≥ 50%的MPN患者（P均<0.01），p-JAK2的蛋白表达水平低于JAK2V617F/JAK2比值≥ 50%者（P<0.01）。（3）250 nmol/L鲁索替尼处理24、48、72 h后，HEL细胞活力分别为（60.06±3.87）%、（52.05±2.88）%、（36.43±2.01）%。随着鲁索替尼浓度的增加，HEL细胞中JAK2的mRNA和蛋白表达与p-JAK2的蛋白表达水平逐渐降低（P<0.01，P<0.05），SOCS1和SHP1的mRNA和蛋白表达逐渐增加（P均<0.01）。结论 鲁索替尼能够抑制HEL细胞中JAK2的mRNA和蛋白表达及其磷酸化水平，增加SOCS1、SHP1 mRNA和蛋白表达，降低HEL细胞的活力。 |
| 关键词: 骨髓增殖性肿瘤 JAK2V617F突变 JAK激酶 细胞因子信号转导抑制蛋白 蛋白酪氨酸磷酸酶 SH2域 |
| DOI：10.16781/j.0258-879x.2018.01.0074 |
| 投稿时间：2017-06-27修订日期：2017-09-22 |
| 基金项目: |
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| Expression of SOCS1 and SHP1 in JAK2V617F mutation positive myeloproliferative neoplasms and regulation effect of ruxolitinib |
| XIE Xu-lei1,2,YANG Sheng-jun1,HAO Hong-ling1,WANG Su-yun1*,WANG Hong-jie3,QI Feng2,CHENG Zhi-yong2,LIU Gui-min2 |
(1. Department of Hematology, Hebei General Hospital, Shijiazhuang 050051, Hebei, China;
2. Department of Hematology, The First People's Hospital of Baoding, Baoding 071000, Hebei, China;
3. Department of Anesthesiology, Affiliated Hospital of Hebei University, Baoding 071000, Hebei, China
*Corresponding author) |
| Abstract: |
| Objective To analyze the relationship between the JAK2V617F mutation and the expressions of phosphorylated Janus kinase 2 (p-JAK2), suppressor of cytokine signaling 1 (SOCS1), and SH2-containing protein tyrosine phosphatase 1 (SHP1) in JAK2V617F mutation positive myeloproliferative neoplasm (MPN) tissues, and to investigate the effects of JAK2 inhibitor ruxolitinib on regulating the proliferation of JAK2V617F mutation positive human erythroleukemia cell lines HEL and the expressions of SOCS1 and SHP1 in HEL cells. Methods A total of 48 patients with JAK2V617F mutation positive MPN (MPN group) and 24 patients with anemia (control group) in Hebei General Hospital and The First People's Hospital of Baoding from Jul. 2012 to Aug. 2016 were enrolled in this study. The protein expressions of p-JAK2, SOCS1 and SHP1 in bone marrow biopsies (BMBs) were detected by immunohistochemistry. The correlations between JAK2V617F mutation level and the protein expressions of SOCS1 and SHP1 were analyzed by Spearman rank correlation analysis. HEL cells were treated with ruxolitinib at different concentrations (50, 100, 250, 500 and 1 000 nmol/L), and the viability of cells was determined by CCK-8 assay. The JAK2V617F mutation levels in MPN tissues and HEL cells and the mRNA expressions of JAK2, SOCS1 and SHP1 in HEL cells were detected by qPCR. The protein expressions of JAK2, SOCS1 and SHP1 in HEL cells were detected by Western blotting analysis. Results The ratio of JAK2V617F/JAK2 was (57.33±20.82)% in the MPN group and was zero in the control group. The protein expressions of p-JAK2, SOCS1 and SHP1 in BMBs of MPN patients were significantly different from those in the control group (all P<0.01). The protein expressions of SOCS1 and SHP1 were negatively correlated with the mutation level of JAK2V617F (r=-0.648, -0.692; P<0.05). The expressions of SOCS1 and SHP1 in MPN patients with JAK2V617F/JAK2<50% were significantly higher than those in MPN patients with JAK2V617F/JAK2 ≥ 50% (P<0.01), while the expression of p-JAK2 was significantly lower than that in MPN patients with JAK2V617F/JAK2 ≥ 50% (P<0.01). After treatment with 250 nmol/L ruxolitinib for 24 h, 48 h, and 72 h, the viabilities of HEL cells were (60.06±3.87)%, (52.05±2.88)%, and (36.43±2.01)%, respectively. With the increase of ruxolitinib concentrations, the mRNA and protein expressions of JAK2 and the protein expression of p-JAK2 were gradually decreased (P<0.01, P<0.05), while the mRNA and protein expressions of SOCS1 and SHP1 were gradually increased (all P<0.01). Conclusion Ruxolitinib can inhibit the expressions of JAK and the phosphorylation of JAK in HEL cells, enhance the expressions of SOCS1 and SHP1, and reduce the viability of HEL cells. |
| Key words: myeloproliferative neoplasm JAK2V617F mutation Janus kinase suppressor of cytokine signaling protein tyrosine phosphatase Src homology 2 domain |
