摘要
目的研究EJ细胞中CD44小RNA干扰后细胞增殖、迁移功能以及其与特异性单抗KMP1结合力的改变。方法小RNA干扰的方法抑制EJ细胞中CD44表达,Western blot和流式细胞仪检测EJ细胞与CD44特异性单抗KMP1的结合功能变化,细胞划痕实验和Boyden小室检测shCD44-EJ细胞的迁移功能,[3H]掺入法检测shCD44-EJ细胞的增殖能力。结果 shCD44-EJ细胞与KMP1的结合能力下降,CD44的表达量也降低,Wide type组和shCD44-EJ组的迁移到划痕中央的细胞数分别为257±32个和132±29个,shCD44-EJ组和Vector组细胞迁移到膜中和下室中的平均细胞数为356±13和672±17个,显示shCD44-EJ细胞的迁移能力较Wide type组和Vector组分别降低52%和53%,shCD44-EJ细胞的增殖活性是Vector组的73%。结论特异性沉默CD44可降低EJ细胞与其特异性单抗KMP1的结合功能,并抑制EJ细胞的迁移和增殖能力。
Objective To investigate the effect of CD44 targeted RNA interference（RNAi） on the growth and migration of human bladder carcinoma EJ cells,and research the change of EJ cells immuno-bounding to the monoclonal antibody KMP1.Methods We transfected bladder cancer cell line EJ with well-designed CD44 siRNA.Flow cytometry and Western blot analysis were used to assess the CD44 immuno-bounding to the monoclonal antibody KMP1.Wound healing assay and Boyden chamber were used to investigate the change of shCD44-EJ cell＇s migration.Proliferation of cells was measured using a -thymidine incorporation assay.Results CD44 knockdown significantly decreased KMP1 staining signal compared with the EJ cells transfected by pSuper-puro empty vector control,Flow cytometry analysis found that CD44 knockdown decreases EJ cells immuno-bounding to the KMP1.Western blot analysis revealed a remarkable decrease of CD44 expression in shCD44-EJ cells,but not in pSuper-puro vector transfected.CD44silencing also significantly inhibited cell migration through wound healingassay.The modified Boyden chamber assay showed similar results also.-thymidine incorporation assay revealed that CD44 silencing can extremely decrease the proliferation of EJ cells.Conclusions CD44 silencing can effectively decrease EJ cells immuno-bounding to the monoclonal antibody KMP1 and inhibit the migration and the proliferation of EJ cells in vitro.
出处
《基础医学与临床》
CSCD
北大核心
2012年第3期313-316,共4页
Basic and Clinical Medicine
基金
云南省科技厅高层次人才培引工程经费资助项目(200807015)
