文章摘要
傅 蔷,唐 娜,张 越,周 略,刘安生.CDA基因沉默对人慢性髓系白血病细胞增殖和凋亡的影响[J].,2020,(12):2239-2245
CDA基因沉默对人慢性髓系白血病细胞增殖和凋亡的影响
Effect of CDA Gene Silencing on Proliferation and Apoptosis of Human Chronic Myeloid Leukemia Cells
投稿时间:2019-12-01  修订日期:2019-12-25
DOI:10.13241/j.cnki.pmb.2020.12.008
中文关键词: 慢性髓系白血病  胞苷脱氨酶  KCL-22细胞  PI3K / Akt信号通路  基因沉默
英文关键词: Chronic myeloid leukemia  Cytidine deaminase  KCL-22 cells  PI3K / Akt signaling pathway  Gene silencing
基金项目:国家自然科学基金青年科学基金项目(81702359);深圳市科技计划项目(JCYJ20180301170035531)
作者单位
傅 蔷 西安市儿童医院血液肿瘤科 陕西 西安 710003 
唐 娜 暨南大学第二临床医学院 南方科技大学第一附属医院 深圳市人民医院病理科 广东 深圳 518020 
张 越 西安市儿童医院中心实验室 陕西 西安 710003 
周 略 深圳市人民医院肿瘤放疗科 广东 深圳 518020 
刘安生 西安市儿童医院血液肿瘤科 陕西 西安 710003 
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中文摘要:
      摘要 目的:探讨胞苷脱氨酶(CDA)基因沉默在治疗人慢性髓系白血病(CML)中的潜在价值。方法:通过RT-PCR和Western blot检测CML患者和造血干细胞移植供体的骨髓单个核细胞中的CDA表达。对CML KCL-22细胞系转染shRNA和过表达CDA的pBS / U6-Neo质粒来诱导CDA基因沉默或过表达。通过细胞计数试剂盒8(CCK-8)测定和细胞集落形成实验评价细胞增殖,通过流式细胞仪检测细胞凋亡。此外,将0.2 mL不同处理的细胞悬浮液(106个细胞/ mL)注射到裸鼠中建立裸鼠肿瘤异种移植模型。结果:与造血干细胞移植供体相比,CML患者的骨髓单个核细胞中的CDA mRNA和蛋白表达显著升高(P <0.05)。转染shRNA-CDA显著降低了KCL-22细胞的细胞活力和细胞集落数(P<0.05)。与对照组(4.32%)相比,shRNA-CDA组(13.45%)的细胞凋亡率显著升高(P<0.05)。与对照组相比,shRNA-CDA组的BCL-2蛋白表达水平显著降低,而cleaved caspase-3显著升高(P<0.05)。与对照组相比,shRNA-CDA组的PI3K蛋白表达水平和Akt磷酸化水平显著降低(P<0.05)。接种30 d后,与对照组相比,shRNA-CDA组裸鼠的肿瘤重量和肿瘤体积均显著降低(P<0.05)。结论:CDA在人慢性髓系白血病中高表达,CDA基因沉默可在体内和体外抑制肿瘤细胞的生长。其机制与抑制PI3K / Akt信号通路的激活有关。
英文摘要:
      ABSTRACT Objective: To explore the potential value of cytidine deaminase (CDA) gene silencing in the treatment of human chronic myeloid leukemia (CML). Methods: The expression of CDA in bone marrow mononuclear cells of CML patients and hematopoietic stem cell transplantation donors was detected by RT-PCR and Western blot. CML KCL-22 cell line was transfected with shRNA and overexpressing CDA pBS/U6-Neo plasmid to induce CDA gene silencing or overexpression. Cell proliferation was evaluated by the Cell Counting Kit 8(CCK-8) assay and cell colony formation experiments, and apoptosis was detected by flow cytometry. In addition, 0.2 mL of differently treated cell suspensions (106 cells / mL) were injected into nude mice to establish nude mice tumor xenograft models. Results: Compared with hematopoietic stem cell transplantation donors, the expression of CDA mRNA and protein in bone marrow mononuclear cells of CML patients was significantly increased (P<0.05). Transfection of shRNA-CDA significantly reduced the cell viability and cell colony number of KCL-22 cells (P<0.05). Compared with the control group (4.32%), the apoptosis rate of the shRNA-CDA group (13.45%) was significantly increased (P<0.05). Compared with the control group, the expression level of BCL-2 protein in the shRNA-CDA group was significantly reduced, while cleaved caspase-3 was significantly increased (P<0.05). Compared with the control group, the expression level of PI3K protein and Akt phosphorylation level in shRNA-CDA group were significantly reduced (P<0.05). After 30 days of inoculation, the tumor weight and tumor volume of the nude mice in the shRNA-CDA group were significantly reduced compared with the control group (P<0.05). Conclusion: CDA is highly expressed in human chronic myeloid leukemia, and CDA gene silencing can inhibit tumor cell growth in vivo and in vitro. Its mechanism is related to inhibiting the activation of PI3K / Akt signaling pathway.
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