RNA干扰S100A4基因表达对肺癌SPC-A-1细胞侵袭能力的影响
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摘要
目的:研究小干扰RNA抑制高迁移族蛋白s10基因表达对肺癌细胞SPC-A-1增值和侵袭能力的影响。材料与方法:设计合成靶向S100A4基因的siRNA,采用阳离子脂质体试剂瞬时转染肺癌细胞株SPC-A-1,利用实时荧光定量PCR和Western blot检测RNA干扰后S100A4基因的沉默效果,评价siRNA设计的合理性及RNA干扰抑制S100A4表达的有效性;用细胞活力计数仪测定3组细胞活力;分别在RNA干扰后24、48、72和96 h应用MTT实验检测细胞生存状态,计算生长抑制率并绘制生长曲线;应用boyden chamber法检测侵袭能力的差异。结果:靶向S100A4的siRNA成功抑制肺癌细胞株SPC-A-1中S100A4基因及蛋白表达,细胞活力检测仪测定RNA干扰48 h后,细胞活力显著下降;MTT测定肺癌细胞生长受到明显抑制;boyden chamber小室细胞侵袭实验提示肺癌穿膜细胞数明显减少。结论:应用siRNA技术能有效地抑制S100A4基因的表达,同时有效抑制SPC-A-1细胞的体外增值和侵袭,为肺癌的生物学治疗提供了新思路。
Purpose: To study the effects and mechanism of S100A4 small interfering RNA( siRNA) on invasion and proliferation of human lung cancer cell. Materials and Methods: After lung cancer cell line SPC-A-1 were transfected by S100A4 siRNA,the mRNA and protein of PRL21 were determined by real time PCR and western blot assay,respectively. The anchorage-independent growth was exmined by clonformation in soft agar,and invasion ability was evaluated by boyden chamber model. Results: The siRNA could down-regulate the level of mRNA and protein of S100A4. Suppression of of S100A4 expression could inhibit invasion and proliferation ability in human lung cancer cell SPC-A-1. Conclusion: S100A4 gene might play an important role in development of human lung cancer,and down-regulation by S100A4 siRNA could inhibit invasion and proliferation of human lung cancer cell.
Purpose: To study the effects and mechanism of S100A4 small interfering RNA( siRNA) on invasion and proliferation of human lung cancer cell. Materials and Methods: After lung cancer cell line SPC-A-1 were transfected by S100A4 siRNA,the mRNA and protein of PRL21 were determined by real time PCR and western blot assay,respectively. The anchorage-independent growth was exmined by clonformation in soft agar,and invasion ability was evaluated by boyden chamber model. Results: The siRNA could down-regulate the level of mRNA and protein of S100A4. Suppression of of S100A4 expression could inhibit invasion and proliferation ability in human lung cancer cell SPC-A-1. Conclusion: S100A4 gene might play an important role in development of human lung cancer,and down-regulation by S100A4 siRNA could inhibit invasion and proliferation of human lung cancer cell.
引文
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15 齐瑞雪,徐向英.非小细胞肺癌中S1 00A4与上皮钙粘蛋白表达的关系及其临床意义.中华肿瘤杂志.2007;29(9):681-684。
16 Bartling B,Rehbein G,Schmitt WD,et al.S100A2-S100P expression profile and diagnosis of non-small cell lung carcinoma:impairment by advanced tumour stages and neoadjuvant chemotherapy.Eur J Cancer.2007;43(13):1935-1943.
2 Radke I,Gtte M,Kersting C,et al.Expression and prognostic impact of the Cancer,2006,95(3):347-354.
3 Liu J,Guo Y,Fu S,et al.Hypomethylation-induced expression of S100A4 increases the invasiveness of laryngeal squamous cell carcinoma.Oncol Rep.2010;23(4):1101-1107.
4 Intini G.Future approaches in periodontal regeneration:gene therapy,stem cells,and RNA interference.Dent Clin North Am.2010;54(1):141-55.
5 Chen M,Sinha M,Luxon BA,et al.Integrin alpha6beta4controls the expression of genes associated with cell motility,invasion,and metastasis,including S100A4/metastasin.J Biol Chem.2009;284(3):1484-1494.
6 Wang HY,Zhang JY,Cui JT,et al.Expression status of S100A14 and S100A4 correlates with metastatic potential and clinical outcome in colorectal cancer after surgery.Oncol Rep.2010;23(1):45-52.
7 Yang Y,Zhao S,Fan Y,et al.Detection and identification of potential biomarkers of non-small cell lung cancer.Technol Cancer Res Treat.2009;8(6):455-466.
8 Tabata T,Tsukamoto N,Fooladi AA,et al.RNA interference targeting against S100A4 suppresses cell growth and motility and induces apoptosis in human pancreatic cancer cells.Biochem Biophys Res Commun.2009;390(3):475-480.
9 Ikenaga N,Ohuchida K,Mizumoto K,et al.S100A4 mRNA is a diagnostic and prognostic marker in pancreatic carcinoma.J Gastrointest Surg.2009;13(10):1852-1858.
10 Petersson S,Shubbar E,Enerbck L,et al.Expression patterns of S100 proteins in melanocytes and melanocytic lesions.Melanoma Res.2009;19(4):215-225.
11 Matsumoto K,Irie A,Satoh T,et al.Expression of S100A2and S100A4 predicts for disease progression and patient survival in bladder cancer.Urology.2007;70(3):602-607.
12 Csorba T,Pantaleo V,Burgyán J.RNA silencing:an antiviral mechanism.Adv Virus Res.2009;75(1):35-71.
13 Grum-Schwensen B,Klingelhfer J,et al.Lung metastasis fails in MMTV-Py MT oncomice lacking S100A4 due to a Tcell deficiency in primary tumors.Cancer Res.2010;70(3):936-947.
14 Tsuna M,Kageyama S,Fukuoka J,et al.Significance of S100A4 as a prognostic marker of lung squamous cell carcinoma.Anticancer Res.2009;29(7):2547-54.
15 齐瑞雪,徐向英.非小细胞肺癌中S1 00A4与上皮钙粘蛋白表达的关系及其临床意义.中华肿瘤杂志.2007;29(9):681-684。
16 Bartling B,Rehbein G,Schmitt WD,et al.S100A2-S100P expression profile and diagnosis of non-small cell lung carcinoma:impairment by advanced tumour stages and neoadjuvant chemotherapy.Eur J Cancer.2007;43(13):1935-1943.