沉默LIMK1促进DADS抑制SW480细胞迁移与侵袭

摘要

目的 探讨LIMK1沉默对DADS抑制SW480细胞迁移与侵袭的影响.方法 RNA干扰技术建立稳定LIMK1-miRNA/SW480细胞株;免疫组化和Western blot检测DADS对沉默LIMK1结肠癌SW480细胞LIMK1和磷酸化LIMK1蛋白表达的影响.划痕实验和侵袭实验检测LIMK1 RNA沉默与DADS对SW480细胞迁移与侵袭的影响.结果 RT-PCR与Western blot显示,LIMK1-miR/SW480细胞LIMK1mNRA与蛋白表达明显下调,表明成功构建稳定沉默LIMK1基因的SW480细胞株.免疫组化和Western blot显示,45 mg· L-1 DADS处理和沉默LIMK1组较未转染组与空载体组SW480细胞LIMK1蛋白和磷酸化LIMK1明显下调 (P<0.05).划痕实验和侵袭实验发现,沉默LIMK1或DADS处理SW480细胞的迁移与侵袭能力较未转染组与空载体组明显抑制 (P<0.05).而DADS处理沉默组抑制SW480细胞迁移和侵袭能力更为明显 (P<0.05).结论 沉默LIMK1基因可抑制SW480细胞迁移与侵袭,增强DADS抑制SW480细胞迁移与侵袭作用.%Aim To investigate the effects of silence LIMK1 on diallyl disulfide ( DADS ) inhibiting the migration and invasion of human colon cancer SW480 cells. Methods The stable low expression of LIMK1 cells ( LIMK1-miR/SW480 ) was established by mi-croRNA interference in SW480 cell line. The effects of DADS on the expression of LIMK1 and p-LIMKl in LIMK1 -miR/SW480 cells were detected by immunocy-tochemistry and Western blot. Migration and invasion potentials were examined by scratch test and transwell membrane assays. Results RT-PCR and Western blot showed that the expression of LIMK1 mRNA and protein was significantly downregulated in LIMK1-miR/ SW480, which indicated the stable low expression of LIMK1 cells was successfully established by microRNA interference. The expression of LIMK1 and p-LIMK1 in the silence LIMK1 group and treatment group with 45 mg · L-1 DADS compared to the no-transfection and vector groups was significantly downregulated, which was observed by Western blot and immunocyto-chemistry. Scratch test and transwell membrane assay showed that the ability of migration and invasion was obviously reduced by silence LIMK1 or treatment with DADS ( P <0. 05 ). Especially, the migration and invasion of SW480 cells were inhibited markedly in silence LIMK1 and simultaneously treatment with DADS ( P <0. 05 ). Conclusion Silence LIMK1 gene could inhibit the migration and invasion of SW480 cells and enhance the inhibition of DADS on the migration and invasion of SW480 cells.