miR-223在CXCR4^+ Lewis肺癌细胞中的显著低表达及其靶基因预测  

Lowly expression of miR-223 in CXCR4 positive cells from Lewis lung carcinoma cell line and its target gene prediction

作　　者：辇伟奇[1] 陈芳琳[1] 敖绪军[2] 陈正堂[1] 

机构地区：[1]第三军医大学新桥医院全军肿瘤研究所,重庆400037 [2]中国人民解放军532医院普通外科,安徽黄山245041

出　　处：《第三军医大学学报》2009年第22期2202-2205,共4页Journal of Third Military Medical University

基　　金：国家高技术研究发展计划("863"计划)(2007AA02Z129);国家自然科学基金(30901790);重庆市自然科学基金(CSTC2008C166)~~

摘　　要：目的分析miR-146a、miR-206、miR-223、let-7c-1等细胞分化相关miRNAs,在小鼠Lewis肺癌细胞株(theLewis lung cancer cell lines,LLC)CXCR4+与CXCR4-亚群间的差异表达。方法免疫磁珠分选CXCR4+和CXCR4-LLC细胞,Trizol法抽提细胞总RNA,实时荧光定量PCR(TaqMan探针)检测miRNAs表达,对表达差异最为显著的miRNAs进行潜在靶基因预测,应用免疫组化方法初步分析具有研究价值的关键分子在两个不同亚群小鼠种植瘤组织中的表达情况,并通过BLAST对其3′非翻译端(untranslated region,UTR)进行直向同源基因序列比对分析。结果CXCR4+与CXCR4-LLC比较,各检测mirna表达均较低,其中以miR-223差异最为显著(Fold Change=8.26),通过软件分析预测,miR-223与IGF1R、IGFBP5、Pik3cb、ELK-1和E2F1等基因均具有可能靶位点,免疫组化检测发现CXCR4+亚群种植瘤组织IGF1R表达显著高于CXCR4-亚群,IGF1R3′UTR的238～244nt和688～695nt两个结合位点具有高度的进化保守性。结论miR-223在CXCR4+LLC中显著低表达,可能与肺腺癌细胞IGF1R信号通路调控有关。Objective To analyze the differential expressions of miR-146a,miR-206,miR-223 and let-7c-1,such as cell differentiation-related miRNAs,in CXCR4-positive and CXCR4-negative subsets of the Lewis lung cancer cell lines（LLC）.Methods CXCR4-positive and CXCR4-negative subsets were isolated from LLC by immunomagnetic beads sorting,and then their total cellular RNA were extracted by Trizol,expression of 4 miRNAs were detected by real-time fluorescence quantitative PCR（TaqMan probe）,and potential target genes of miRNA whose differential expression was the most significant were predicted. Immunohistochemistry was carried out to confirm differential expression of the key molecule of certain research value within CXCR4-positive and CXCR4-negative subsets growing tumor tissue, and a BLAST search was performed to identify homologies of its 3＇UTR. Results Compared to CXCR4-negative subsets, the expression of 4 miRNAs were lower in CXCR4-positive subsets, and expression of miR-223 had the most significant difference （Fold change=8.26）. By softwares forecasting, miR-223 had potential target sites of IGFIR, IGFBP5, Pik3cb, ELK-1 and E2F1 mRNA, such as key molecular of IGF1R signaling pathway. The expression of IGF1R of CX CR4-positive subsets growing tumor tissue was significantly higher than that of CXCR4-negative subsets. Conclusion miR-223 is lowly expressed in CXCR4 positive cells from Lewis lung carcinoma cell lines. Position 238 -244 nt and 688 -695 nt in target sequences of 3＇UTR of IGF1R mRNA was highly homologous by screening. Close correlation is found between miR-223 and IGF1R signaling pathway. The mechanisms underlying this biologically important finding need to be further explored.

关 键 词：miR-223 CXCR4 LEWIS肺癌细胞 

分 类 号：R394.3[医药卫生—医学遗传学] R730.23[医药卫生—基础医学]