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目的:探讨mi R-223调控急性淋巴细胞白血病(acute lymphoblastic leukemia,ALL)E6-1细胞的增殖和体内成瘤能力及其作用机制。方法:应用实时荧光定量PCR检测mi R-223在不同ALL细胞株中的表达水平,免疫荧光染色法检测携带mi R-223mimic慢病毒感染E6-1细胞株的感染效率,双荧光素酶实验检测mi R-223和Lmo2的相互结合关系,MTT增殖实验和克隆形成实验检测过表达mi R-223对E6-1细胞株增殖和克隆形成能力的影响。裸鼠体内成瘤实验检测mi R-223过表达对E6-1细胞成瘤能力的影响。Western blotting检测mi R-223对MAPK信号通路相关蛋白表达的影响。结果:mi R-223在E6-1细胞株中表达水平相对较低(P<0.05),双荧光素酶实验证实mi R-223可以直接靶向结合Lmo2的3’UTR区序列。过表达mi R-223后:(1)抑制E6-1细胞的增殖能力(0.16±0.02 vs 1.15±0.21,P<0.05);(2)抑制E6-1细胞的裸鼠体内成瘤能力[(0.56±0.08)vs(1.69±0.22)g,P<0.05];(3)调控MAPK信号通路的活性。结论:mi R-223可靶向作用Lmo2通过MAPK信号通路调控ALL细胞的增殖、克隆形成和体内成瘤能力。
OBJECTIVE: To investigate the proliferation and in vivo tumorigenicity of mi R-223 in E6-1 cells and its mechanism of action in acute lymphoblastic leukemia (ALL). Methods: The expression of mi R-223 in different ALL cell lines was detected by real-time fluorescence quantitative PCR. The infection efficiency of E6-1 cells infected with mi R-223mimic lentivirus was detected by immunofluorescence staining. The results of dual luciferase assay mi R-223 and Lmo2, MTT proliferation assay and clonogenic assay were used to detect the effect of overexpression mi R-223 on the proliferation and clonality of E6-1 cells. The effect of mi R-223 overexpression on tumorigenicity of E6-1 cells in nude mice in vivo. Western blotting was used to detect the effect of mi R-223 on MAPK signal pathway related protein expression. Results: The expression of mi R-223 in E6-1 cells was relatively low (P <0.05). Dual luciferase assay confirmed that mi R-223 could directly bind to the 3’UTR region of Lmo2. Overexpression of mi R-223: (1) inhibited the proliferation of E6-1 cells (0.16 ± 0.02 vs. 1.15 ± 0.21, P <0.05); (2) inhibited the tumorigenicity of E6-1 cells in nude mice [ 0.56 ± 0.08) vs (1.69 ± 0.22) g, P <0.05]. (3) The activity of MAPK signal pathway was regulated. Conclusion: mi R-223 can target Lmo2 to regulate the proliferation, colony formation and in vivo tumorigenicity of ALL cells via MAPK signal pathway.