干扰CXCR4抑制急性单核细胞白血病细胞株SHI-1黏附、侵袭及成瘤能力

RNAi-mediated Silencing of CXCR4 Inhibits the Adhesion,Invasion and Tumorigenicity of Acute Monocyice Leukemic Cell Line SHI-1

目的:研究干扰人趋化因子受体4(CXCR4)基因表达对白血病细胞株SHI-1细胞体外增殖、黏附、侵袭能力及裸鼠皮下成瘤能力的影响。方法:通过构建干扰CXCR4表达的慢病毒载体,重组病毒颗粒感染SHI-1细胞,干扰SHI-1细胞CXCR4表达,定量PCR检测CXCR4、MMP-2和MMP-9表达;流式细胞术检测SHI-1细胞膜表面CXCR4蛋白表达;MTT法检测细胞体外增殖能力;将SHI-1等与骨髓基质细胞共培养,检测SHI-1细胞黏附能力及跨Matrigel基质胶迁移能力;将白血病细胞SHI-1接种于裸鼠皮下并观察其在裸鼠皮下生长情况。结果:干扰CXCR4表达的慢病毒颗粒感染SHI-1细胞后,SHI-1/CXCR4i细胞的CXCR4 mRNA表达较感染阴性对照病毒的SHI-1/NC细胞下调76%,SHI-1/CXCR4i细胞膜表面CXCR4表达明显下调;SHI-1/CXCR4i细胞MMP-2、MMP-9表达分别也分别下降63%和62%;SHI-1/CXCR4i细胞体外增殖能力无显著改变,但其黏附能力及跨Matrigel基质胶迁移能力明显下降;SHI-1/CXCR4i细胞不能在裸鼠皮下形成肿瘤,而SHI-1及SHI-1/NC细胞均能在裸鼠皮下形成肿瘤,且肿瘤体积无显著差异。结论:干扰CXCR4表达可使SHI-1的黏附、移行能力显著下降,并能完全抑制SHI-1在裸鼠皮下形成肿瘤,CXCR4可作为白血病治疗的一个靶点。

Objective : To investigate the effect of CXCR4 gene on the proliferation,adhesion,invasion and tumorigenicity of a human monocytic leukemic cell line SHI-1. Methods: The lentivirus vector silencing the expression of CXCR4 was constructed for infection of SHI-1 cells silencing expression of CXCR4 in SHI-1 cells. The expression of CXCR4,MMP-2 and MMP-9 was detected by real time PCR. The expression of CXCR4 on membrane of SHI-1 cells was detected by flow cytometry. The SHI-1 cell proliferation ability was detected by MTT. The co-culture system of the leukemia cells and bone marrow stromal cells was used to detect the adhesion and migration ability of leukemia cells.SHI-1 cells were inoculated subcutaneously in nude mice to investigate the growth ability in vivo. Results: After SHI-1cells were infected by lentivirus silencing expression of CXCR4,the expression of CXCR4 mRNA in SHI-1 CXCR-4i cells decreased by 76% as compared with expression of SHI-1 /NC of negative control virus,the expression of CXCR4 on membrane of SHI-1 /CXCR4 i obviously downregulated; the expression of MMP-2 and MMP-9 in SHI-1 /CXCRi cells also declined by 63% and 62% respectively; the proliferation ability of SHI-1 /CXCR4 i in vitro did not obviously changed,but the adhesion and trans-matrigel invasion ability significantly decreased,the SHI-1 /CXCR4 i cells could not form neoplasm subcutaneously in mice,but the SHI-1 and SHI-1 /NC cells could form neoplasm subcutaneously in mice,and there was no siguificant difference in volum of neoplasm mass. Conclusion: The silencing expression of CXCR4 can decline the adhesion and migration ability of SHI-1 cells,and can completely suppress the formation of neoplasm subcutaneously,so the CXCR4 may serve as a target for treating leukemia.acute monocytic leukemia cell line; CXCR4; RNA interference; MMP; cell proliferation; cell adhesion; extramedullary infiltration; nude mice J Exp Hematol 2015;( 5) : 1286- 1291