| 苏坚，潘志兵，史玲，向姝霖，夏红，董琳，廖爱军，苏琦.LIMK1在结肠癌中表达与沉默对人结肠癌细胞迁移与侵袭的影响.[J].中南医学科学杂志.,2014,42(2):109-115. |
| LIMK1在结肠癌中表达与沉默对人结肠癌细胞迁移与侵袭的影响 |
| Expression of LIMK1 and the Effect of Silence LIMK1 forMigration and Invasion in Human Colon Cancer |
| 投稿时间：2013-10-12 |
| DOI： |
| 中文关键词: 结肠癌 SW480细胞 LIMK1 RNA干扰 迁移与侵袭 |
| 英文关键词:human colon cancer SW480 cell LIMK1 RNA interference migration and invasion |
| 基金项目:国家自然科学基金（31000629，81102854）. |
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| 中文摘要: |
| 目的探讨LIMK1在结肠癌中的表达与临床病理参数的关系和沉默LIMK1对人结肠癌SW480细胞迁移与侵袭的影响。方法免疫组化检测LIMK1在结肠癌表达；RNA干扰建立LIMK1-miR/SW480细胞株；RT-PCR与Western blot分别检测LIMK1 mNRA与蛋白及磷酸化LIMK1表达；划痕实验和侵袭实验检测沉默LIMK1对SW480细胞迁移与侵袭的影响。结果免疫组化显示，LIMK1在结肠癌中表达明显高于结肠正常组织，高分化腺癌表达明显低于中分化与低分化腺癌，而低分化高于中分化腺癌；<5.0 cm的结肠癌表达明显低于≥5.0 cm；淋巴结转移显著高于无转移；Dukes分期A+B组表达明显低于C+D组（P<0.05）。RT-PCR与Western blot显示，LIMK1-miR/SW480细胞LIMK1 mNRA与蛋白表达明显下调，表明成功构建稳定沉默LIMK1基因的SW480细胞。免疫细胞化学证实，沉默组LIMK1表达较未转染组与空载体组明显降低。Western blot显示，沉默组LIMK1和磷酸化LIMK1较未转染组与空载体组明显下调(P<0.05)。划痕实验显示，沉默组癌细胞迁移率（22.53%）较对照组（76.50%）与空载体组（72.14%）明显降低（P<0.05）。侵袭实验发现，沉默组穿膜细胞（43.67±1.51个）较未处理组（143.33±1.52个）与空载体组（136.34±1.53个）明显减少（P<0.05）。结论LIMK1表达与结肠癌发生、大小、淋巴结转移及Dukes分期有关。沉默LIMK1基因可抑制人结肠癌SW480细胞迁移与侵袭。 |
| 英文摘要: |
| ObjectiveTo investigate the expression of LIMK1 and the effects of silence LIMK1 for migration and invasion in human colon cancer.MethodsThe stable low expression of LIMK1-miR/SW480 cells was established by RNA interference and validated by RT-PCR and Western blot in SW480 cells.The effects of expression of LIMK1 and p-LIMK1 in LIMK1-miR/SW480 cells was detected by Western blot and Immunocytochemistry.Migration and invasion potentials were examined by scratch test and transwell membrane assays.ResultsLIMK1 expression in colon cancer was significantly higher than in the normal tissue,lower in the well-differentiated than in the moderately differentiated and poorly differentiated adenocarcinoma,and lower in moderately differentiated than in poorly differentiated adenocarcinoma;The expression of LIMK1 in volume<5.0 cm was lower than in ≥5.0 cm;that lymph node metastasis was significantly higher than that of non-lymph node metastasis;Dukes stage A+B was significantly lower than in C+D (P<0.05).RT-PCR and Western blot showed that the expression of LIMK1 mRNA and protein was significantly downregulated in LIMK1-miR/SW480,indicating the stable low expression of LIMK1 cells were successfully established by microRNA interference.The expression of LIMK1 and p-LIMK1 in the silence LIMK1 group compared to the no-transfection and vector groups was significantly downregulated by Western blot,and the expression of LIMK1 in the silence group compared to the no-transfection and vector groups was significantly decreased by Immunocytochemistry.Scratch test revealed that the cell transport ratio of silence group was 22.53%,which was lower than 76.50% of no-transfection and 72.14% of vector groups (P<0.05).Transwell assay exhibited that the number of cancer cells which passed through the matrigel membrane was 43.67±1.51 in silence group which was decreased compared with 143.33±1.52 in no-transfection and 136.34±1.53 in vector groups (P<0.05).ConclusionThe expression of LIMK1 is closely related to carcinogenesis,tumor size,differentiation,lymph node metastasis and Dukes stage in colon cancer.Silence LIMK1 gene could inhibit the migration and invasion in SW480 cells. |
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