AIM: To investigate the expression of tissue factor(TF) induced by oxidized high density lipoprotein(oxHDL) in human umbilical vein cell line,ECV304,and the related mechanisms.METHODS: Four main groups were designed: the negative,the positive(ECV304 with histamine),the HDL group and the oxHDL group.Quantitative real-time polymerase chain reaction(RQ-PCR) and Western blotting were used to detect the expression level of TF.The specific inhibitors of MAPKs,SP600125(c-jun terminal NH2 kinase,JNK),SB203580(p38 MAP kinase,p38 MAPK),PD98059(extracellular signal-regulated kinase,ERK1/2) were used to investigate the underlying mechanisms.RESULTS: The TF expression in normal ECV304 cell line was not detected.Histamine administration resulted in a significant expression of TF in ECV304 cell line,with strongest effect after 1 h co-incubation at concentration of 1?10-5 mol/L histamine(about 4.8-fold higher expression of TF compared with that of 1?10-9 mol/L histamine).Expression level of TF was detected after stimulated with oxHDL in dose-and time-dependent manners.The highest expression of TF mRNA was found at 20 mg/L oxHDL and 6 h co-incubation,with 1.8-fold and 5.3-fold increase in TF expression,respectively,compared with that at 10 mg/L oxHDL and 2 h co-incubation.20 mg/L oxHDL also caused an apparent augmentation of TF protein expression,about 1.5-fold higher compared with that stimulated by 40 mg/L oxHDL.HDL co-incubation did not cause a detectable expression of TF protein.The mRNA levels of TF in ECV304 cell line induced by oxHDL were decreased by 95.0%,81.0%,87.0%,respectively(all P

OBJECTIVE: To explore the effects of c-Met-siRNA on the proliferation, movement and invasion of laryngeal carcinoma Hep-2 cells in vitro. METHOD: Firstly, the pSilencer 2.0/c-Met-shRNA recombinant plasmid was transfected into laryngeal carcinoma Hep-2 cells with transfecting agent of cationic liposome Lipofectamine 2000. Secondly,the transfection efficacy was tested by RT-PCR and Western-Blot, then the most inhibitive c-Met-siRNA sequence was elected. Cell proliferation, movement and invasion were detected with MTT, cell migration assay and cell invasion assay, respectively. RESULT: After the transfection of pSilencer 2.0/c-Met-shRNA recombinant plasmid into laryngeal carcinoma Hep-2 cells, the expression of mRNA and protein of c-Met decreased significantly in Hep-2 cells, and ability of the proliferation, movement and invasion of laryngeal carcinoma Hep-2 cells were also inhibited. CONCLUSION The results indicated that c-Met-siRNA can down-regulated the expression of c-Met and markedly inhibited laryngeal carcinoma Hep-2 cell proliferation, movement and invasion. It may have the potential as a therapeutic modality to treat human laryngeal carcinoma.
Apoptosis ; genetics ; Carcinoma, Squamous Cell ; genetics ; pathology ; Cell Line, Tumor ; Cell Proliferation ; Humans ; Laryngeal Neoplasms ; genetics ; pathology ; Liposomes ; Proto-Oncogene Proteins c-met ; genetics ; RNA, Messenger ; genetics ; RNA, Small Interfering ; genetics ; Transfection