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山东大学学报（医学版） ›› 2014, Vol. 52 ›› Issue (9): 1-5.doi: 10.6040/j.issn.1671-7554.0.2014.271

• 基础医学 •    下一篇

干扰CD147表达抑制白血病细胞SHI-1增殖及移行

涂燕1,2, 李振江1, 汤爱平1, 费妍1, 李慧慧1, 李剑1, 贺文凤1   

  1. 1. 南昌大学第二附属医院血液科 江西省血液重点实验室, 江西 南昌 330006;
    2. 浙江大学金华医院血液科, 浙江 金华 321000
  • 收稿日期:2014-04-14 修回日期:2014-09-05 出版日期:2014-09-10 发布日期:2014-09-10
  • 基金资助:
    国家青年自然科学基金(30800490)；江西省科技厅基金

RNAi-mediated silencing of CD147 inhibits the proliferation and invasion of leukemic cells SHI-1

TU Yan1,2, LI Zhenjiang1, TANG Aiping1, FEI Yan1, LI Huihui1, LI Jian1, HE Wenfeng1   

  1. 1. Department of Hematology, Second Hospital Affiliated to Nanchang University, Key Laboratory of Hematology in Jiangxi Province, Nanchang 330006, Jiangxi, China;
    2. Department of Hematology, Jinhua Hospital of Zhejiang University, Jinhua 321000, Zhejiang, China
  • Received:2014-04-14 Revised:2014-09-05 Online:2014-09-10 Published:2014-09-10
  • Contact: 李振江。E-mail:lzjdgh@163.com E-mail:lzjdgh@163.com

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摘要： 目的 构建针对细胞外基质金属蛋白酶诱导因子（CD147）的慢病毒干扰载体，探讨干扰CD147表达后对白血病细胞系SHI-1细胞体外增殖、侵袭、转移能力的影响。方法 设计针对CD147的干扰序列片段，构建于慢病毒载体pGCSIL-GFP上，重组慢病毒载体与包装载体共转染293T细胞，收集病毒上清，测定病毒滴度。病毒上清感染SHI-1细胞，RT-PCR法检测CD147、MMP-2及MMP-9 mRNA表达，Western blotting法检测CD147蛋白水平；MTT法检测细胞体外增殖能力，体外SHI-1细胞与骨髓基质细胞共培养跨Matrigel基质胶检测SHI-1细胞的体外侵袭能力。结果 成功构建慢病毒干扰载体，获得重组慢病毒，滴度达1×109 TU/mL。病毒感染SHI-1细胞后，SHI-1/CD147i细胞的CD147 mRNA较感染阴性对照病毒的SHI-1/NC细胞下调86.7%，CD147蛋白表达下降91%，SHI-1/CD147i细胞MMP-2及MMP-9 mRNA表达较SHI-1/NC细胞分别下降78.3%和70.6%。SHI-1/CD147i细胞体外增殖能力较SHI-1/NC及SHI-1细胞明显下降；与骨髓基质细胞共培养24h后，SHI-1、SHI-1/NC、SHI-1/CD147i细胞移行至Transwell下层的细胞占接种细胞数比例分别为（18.2±2.5）%、（16.5±2.7）%、（4.5±1.2）%，SHI-1/CD147细胞移行能力显著低于SHI-1和SHI-1/NC细胞。结论 干扰SHI-1细胞的CD147表达后，通过下调MMPs表达抑制SHI-1细胞体内外增殖及移行能力，CD147可能成为治疗急性白血病的靶点。

关键词: 细胞增殖, 细胞外基质金属蛋白酶诱导因子, 急性单核细胞白血病细胞株, RNA干扰, 基质金属蛋白酶, 细胞移行

Abstract: Objective To construct lentivirus vector for RNA interference (RNAi) targeting extracellular matrix metalloproteinase inducer (CD147) and to investigate the role of CD147 gene on the proliferation and infiltration of a human monocytic leukemic cell line SHI-1. Methods The complementary shRNA targeting CD147 was synthesized, and connected with pGCSIL-GFP vector. The 293T cells were transfected with recombined viral vector, packing plasmid pHelper 1.0 and pHelper 2.0 using Lipfectamine 2000. Then the recombinant lentiviruses were collected and infected the SHI-1 cells. The expressions of CD147, MMP-2 and MMP-9 in SHI-1 cells were detected by real time PCR. The protein of CD147 was detected by Western blotting. The capability of proliferation and infiltration of SHI-1 cell were examined by MTT and trans-matrigel invasion assay co-cultured with leukemia BMSCs in vitro. Results The lentivirus containing the CD147 shRNA was constructed successfully. The titer of recombined virus was 1×109 TU/mL. The mRNA and protein of CD147 in SHI-1/CD147i cells decreased by 86.7% and 91% respectively after the SHI-1 cells were infected by the lentivirus containing the CD147 siRNA. The proliferation capability of SHI-1/CD147i cells significantly decreased compared with that of SHI-1 and SHI-1/NC cells. The mRNA expressions of MMP-2 and MMP-9 in SHI-1/CD147i cells were significantly lower than that in SHI-1/NC and SHI-1 cells. The SHI-1/CD147i cells demonstrated significantly lower invasion rate than SHI-1 cells and SHI-1/NC cells when co-cultured with BMSCs. Conclusion CD147 plays important roles in the leukemia cell proliferation and infiltration. CD147 should be a potential target for the treatment of acute leukemia.

Key words: matrix metalloproteinase, Extracellular matrix metalloproteinase inducer, RNA interference, Cell invasion, Acute monocytic leukemia cell line, Cell proliferation

中图分类号: 

  • R733.7
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