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Although constitutive nuclear factor(NF)-kB activation has been reported in many humantumors,the role of the NF-kB pathway in esophageal squamous cell carcinoma(ESCC)has not been known.In this study,NF-kB pathway in two ESCC cell lines was investigated using immunocytochemistry,Westernblot and reverse transciiption-polymerase chain reaction.The activation of NF-kB DNA binding was determinedby electrophoretic mobility-shift assay.RNA interference was used to specifically inhibit the expression ofp65.Growth of cells Was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay.The results showed that p50,p65,IkBα,p-IkBα and IkB kinase β were expressed and mainly localized in thecytoplasm.Reverse transcription-polymerase chain reaction results showed the constitutive expressions ofp50,p65 and IkBα mRNA in the two ESCC cell lines.Furthermore,the nuclear extracts revealed that p50and p65 translocated to the nucleus had DNA-binding activity.Finally,small interfering RNA of p65 decreasedthe expression of p65,and the viability of ceils transfected with p65 small interfering RNA was significantlysuppressed at the same concentration of 5-fluorouracil(P<0.05)compared to untransfected cells.The resultsof this study showed that there was the constitutively activated NF-kB signaling pathway in the ESCC celllines.RNA interference targeting at p65 increased the sensitivity of the ESCC cell lines to 5-fluorouracil,suggesting that NF-kB might be a good target for cancer treatment.
The constitutive nuclear factor (NF) -kB activation has been reported in many humantumors, the role of the NF-kB pathway in esophageal squamous cell carcinoma (ESCC) has not been known.In this study, NF-kB pathway in two ESCC cells lines was investigated using immunocytochemistry, Western blot and reverse transciiption-polymerase chain reaction. The activation of NF-kB DNA binding was determined by electrophoretic mobility-shift assay. RNA interference was used to specifically inhibit the expression of p65. Growth of cells by evaluated 3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide assay. These results showed that p50, p65, IkBα, p-IkBα and IkB kinase β were expressed and mainly localized in the cytoplasm. Reverse transcription-polymerase chain reaction results showed the constitutive expressions of p50, p65 and IkBα mRNA in the two ESCC cell lines. Frthermore, the nuclear extracts revealed that p50 and p65 translocated to the nucleus had DNA-binding activity .Finally, small interfering RNA of p65 decreased the expression of p65, and the viability of ceils transfected with p65 small interfering RNA was significantlysuppressed at the same concentration of 5-fluorouracil (P <0.05) compared to untransfected cells. The resultsof this study showed that there was the constitutively activated NF-kB signaling pathway in the ESCC celllines. RNA interference targeting at p65 increased the sensitivity of the ESCC cell lines to 5-fluorouracil, suggesting that NF-kB might be a good target for cancer treatment.