Effect of STAT3 siRNA-Induced Inhibition of STAT3 Gene Expression on the Growth and Apoptosis of Lewis Lung Cancer Cells

OBJECTIVE To determine the effect of short interference RNA （siRNA） against STAT3 induced inhibition of STAT3 gene expression and on the growth and apoptosis of Lewis lung cancer cells. METHODS pSilencer 2.1-U6 STAT3 siRNA against STAT3-mRNA was synthesized, Lewis lung cancer cells were divided into 3 groups： vehicle, plasmid, and STAT3 siRNA in which the ceils were treated with RPMI- 1640 culture media, or transfected with pSilencer empty vector, or pSilencer STAT3 siRNA, Semiquantitative RT-PCR and Western blot analysis of STAT3 gene expression in the cells was performed 72 h after transfection, MFr assay for cell proliferation, flow cytometry and DNA laddering electrophoresis were used for determination of cell proliferation and apoptosis, RESULTS STAT3 was markedly expressed at both the mRNA and protein levels in the cells treated with RPMI-1640 media or transfected with the plasmid vector, whereas STAT3 expression was significantly reduced in cells treated with STAT3 siRNA, These findings suggest that STAT3 siRNA effectively inhibited STAT3 expression. Transfection of the cells with STAT3 siRNA resulted in significant cellular growth inhibition and enhanced apoptosis, CONCLUSION Transfection of Lewis lung cancer cells with synthetic STAT3 siRNA resulted in effective inhibition of STAT3 gene expression at both protein and mRNA levels, leading to induced apoptosis and growth suppression.

In vivo Anti-tumor Effect of siRNA Against STAT3 on Transplanted Lewis Lung Cancer in Mice

This study aims at determining the therapeutic effect of knockdown of signal transducers and activators of transcription3（STAT3） gene expression by short interference RNA（siRNA） on transplanted Lewis lung cancer in mice in vivo. pSilencer 2.1-U6 STAT3 siRNA against STAT3（STAT3 siRNA） was synthesized. Lewis lung cancer cells were inoculated subcutaneously into C57BL/6 mice. Seven days after inoculation, the tumor-bearing mice were randomly divided into 3 groups and received intratumoral injection of （1） vehicle（PBS solution）, （2） vector（negative control）, and （3） STAT3 siRNA. Tumor volume and weight were calculated. Tumors and the lungs were excised for 21 days after inoculation. Expressions of STAT3, vascular endothelial growth factor（VEGF）, and matrix metalloproteinase-2（MMP2） were analyzed by RT-PCR, Western blot, and immunohistochemical staining. HE staining and TUNEL assay were used to confirm the apoptosis of tumors. The synthetic STAT3 siRNA effectively suppressed tumor growth, prevented tumor from pulmonary metastasis, and induced tumor apoptosis in vivo compared with vehicle and vectore in controls. It significantly inhibited STAT3 expression to contribute to downregulation of VEGF and MMP2 expression within tumors in vivo. This study demonstrates that STAT3 siRNA can effectively inhibit the expression of STAT3 gene within tumors, leading to suppression of tumor growth, prevention of cancer from pulmonary metastasis, and enhancing apoptosis of the transplanted Lewis lung cancer in mice in vivo.

Lewis肺癌小鼠血管内皮生长因子的时间节律性表达

目的:检测日间和夜间Lewis肺癌小鼠血清中内血管内皮生长因子(VEGF)水平和肺癌组织中VEGF蛋白表达的差别,探讨肿瘤血管生成的昼夜节律。方法:选择C57BL小鼠30只,制备Lewis肺癌小鼠模型后随机分为日间组(D组)和夜间组(N组),在光照-黑暗条件下饲养建立统一同步化日夜节律。随着成瘤过程,应用ElISA方法测定两组小鼠模型第0、3、5、7天血清中VEGF浓度水平;成瘤后第9天处死小鼠,应用Westeon-blot法检测瘤体中VEGF蛋白的表达,并分别进行相关分析。结果:随着成瘤过程,D组小鼠血清中VEGF浓度均显著高于N组(P<0.05);D组小鼠瘤体组织中VEGF蛋白表达灰度值(8.87±1.20)均明显高于N组(6.43±1.35),有统计学意义(P<0.05)。结论:Lewis小鼠休息期(白天)血清中VEGF浓度及瘤体中VEGF蛋白表达水平均明显高于活动期(夜间),存在着明显的日夜差异,说明肺癌组织的血管生成可能具有一定日夜节律。