外源性Wnt-3a促进急性T淋巴细胞白血病细胞增殖与细胞周期.docVIP

外源性Wnt-3a促进急性T淋巴细胞白血病细胞增殖与细胞周期进展的研究 胡艳妮 徐酉华 郑改焕 罗庆 沈亚莉 (400014 重庆 , 重庆医科大学附属儿童医院血液科，) [摘要] 目的? 应用重组腺病毒介导的Wnt-3a激活急性T白血病细胞株Jurkat中的经典Wnt/β-catenin信号途径，观察该途径激活后对Jurkat细胞增殖及细胞周期的影响，初步探讨其在急性T淋巴细胞白血病分子发病机制中的作用。 方法 将携带Wnt-3a基因的重组腺病毒（Ad5-Wnt-3a）感染Jurkat细胞，实验分为三组：实验组（Jurkat/Ad5-Wnt-3a组）、空载体组（Jurkat/Ad5-GFP组）和空白对照组（Jurkat组）。转染后，采用RT-PCR方法检测Jurkat细胞内Wnt-3a mRNA的表达；Western Blot检测细胞内β-catenin蛋白的表达；MTT法检测细胞增殖，FCM法检测细胞周期，RT-PCR检测Wnt信号通路下游靶基因C-myc和Cyclin D1 mRNA表达。 结果 重组腺病毒Ad5-Wnt-3a转染Jurkat细胞后48h，实验组有效表达Wnt-3a的基因。Western Blot结果显示，与对照组比较，实验组细胞内β-catenin蛋白表达明显增加，与对照组相比差异有统计学意义（P＜0.05）。MTT结果显示，Wnt-3a转染后48h和72h可明显促进Jurkat细胞的增殖，与对照组相比差异有统计学意义（P＜0.05）。FCM检测发现，Wnt-3a转染后可引起细胞G0/G1期下降，S期升高（P＜0.05）。RT-PCR检测结果显示细胞内C-myc和Cyclin D1 mRNA表达升高，与对照组相比差异有统计学意义（P＜0.05）。结论 腺病毒介导的Wnt-3a可以激活Jurkat细胞中的经典Wnt/β-catenin信号途径，使Jurkat细胞增殖，促进细胞周期进展，其可能的机制是通过调控其下游靶基因C-myc和Cyclin D1的表达实现。 [关键词] 经典Wnt/β-catenin信号途径；急性T淋巴细胞白血病；细胞增殖；细胞周期 [中图法分类号] R730. 5; R733. 7 [文献标志码] A Exogenous Wnt-3a mediates T-lineage acute lymphoblastic leukemia cell proliferation and cell cycle progression HU Yan-ni,XU You-hua,ZHENG Gai-huan,LUO Qing，SHEN Ya-li(Department of Hematology , Affiliated Children’s Hospital,Chongqing Medical University,Chongqing 400014, China) [Abstract] Objective To study the effect of canonical Wnt/β-catenin signaling activated by recombinant adenovirus vector-mediated Wnt-3a on proliferation and cell cycle in human leukemia cell line Jurkat ,and explore its molecular pathogenesis in T-lineage acute lymphocytic leukemia. Methods Jurkat cells were infected with recombinant adenovirus expressing Wnt-3a as experimental group (Jurkat/Ad5-Wnt-3a),and empty vector group (Jurkat/Ad5-GFP) and blank control group (Jurkat) as controls. Wnt-3a mRNA expression was detected by RT-PCR.The protein levels of β-catenin was measured by Western Blot. The proliferation, cell cycle and C-myc and Cyclin D1 mRNA expressions were determined by MTT assay , flow cytometry and RT-PCR, respectivel