CAJ | 학술논문

目的探讨缺氧条件下核因子(NF) κB对胰腺癌细胞上皮-间质转化的调控作用.方法在常氧和缺氧条件下培养人胰腺癌细胞株BxPC-3和Panc-1,分为常氧组和缺氧组.通过光镜观察细胞形态学变化,通过Western blot法检测上皮标志物和间质标志物蛋白表达,通过Western blot 法和电泳凝胶迁移实验检测NF-κB P65的蛋白表达和DNA结合活性,通过Transwell小室观察细胞迁移侵袭能力,通过cell counting kit-8法检测细胞生存率.分别采用药物及siRNA法抑制缺氧细胞NF-κB P65表达后,重复检测上述指标.结果胰腺癌细胞在缺氧条件下培养48 h后,细胞失去多边形,获得纺锤形态,细胞间黏附减少并出现伪足,迁移侵袭能力增强,对吉西他滨药物敏感性下降,上皮标志物蛋白表达下调,间质标志物蛋白表达上调,NF-κB P65蛋白表达和DNA结合活性升高；阻断缺氧细胞NF-κB P65表达后可逆转上述现象.结论缺氧可能通过上调胰腺癌细胞NF-κB P65活性,调控上皮-间质转化相关蛋白表达,增强其迁移侵袭能力,降低对吉西他滨的药物敏感性.
목적 탐토결양조건하핵인자(NF) κB대이선암세포상피-간질전화적조공작용.방법 재상양화결양조건하배양인이선암세포주BxPC-3화Panc-1,분위상양조화결양조.통과광경관찰세포형태학변화,통과Western blot법검측상피표지물화간질표지물단백표체,통과Western blot 법화전영응효천이실험검측NF-κB P65적단백표체화DNA결합활성,통과Transwell소실관찰세포천이침습능력,통과cell counting kit-8법검측세포생존솔.분별채용약물급siRNA법억제결양세포NF-κB P65표체후,중복검측상술지표.결과 이선암세포재결양조건하배양48 h후,세포실거다변형,획득방추형태,세포간점부감소병출현위족,천이침습능력증강,대길서타빈약물민감성하강,상피표지물단백표체하조,간질표지물단백표체상조,NF-κB P65단백표체화DNA결합활성승고；조단결양세포NF-κB P65표체후가역전상술현상.결론결양가능통과상조이선암세포NF-κB P65활성,조공상피-간질전화상관단백표체,증강기천이침습능력,강저대길서타빈적약물민감성.
Objective To investigate the function of nuclear factor ( NF)-κB in the epithelial to mesenchymal transition induced by hypoxia in pancreatic cancer cells.Methods For cultured pancreatic cancer cells( BxPC-3 and Panc-1 ) under hypoxic and normoxic conditions,the differences in the morphology were observed by optical microscope.The expression of markers of epithelial and mesenchymal phenotypes,E-cadherin,vimentin and N-cadherin,were determined by Western blot.NF-κB P65 activity was measured by electrophoretic mobility shift assay.Invasion and gemcitabine resistance of pancreatic cancer cells were evaluated in matrigel invasion assay and cell counting kit-8 assay.Both molecular and pharmacologic means of inhibiting NF-κB P65 were used in these hypoxic cells and then the above resulting phenotypes were compared with those of the control-treated cells.Results After cultured pancreatic cancer cells under hypoxic conditions for 48 h,normoxic cells exhibited a polygonal shape and formed tight clusters of cells,whereas hypoxic cells took on an elongated,fibroblastoid morphology associated with a more highly invasive character and resistance to gemcitabine ; hypoxic cells exhibited an suppression of E-cadherin and increase in vimentin and N-cadherin expression.NF-κB P65 activity was elevated in hypoxic cells.On the contrary,on molecular or pharmacologic inhibition of NF-κB P65,hypoxic cells regained expression of E-cadherin,lost expression of N-cadherin,and reversed their highly invasive and drug resistant phenotype.Conclusions Pancreatic cancer cells underwent epithelial to mesenchymal transition exposed to hypoxia,exhibited highly invasive and drug resistant phenotype.Inhibition of NF-κB P65 under hypoxic conditions,pancreatic cancer cells regained expression of E-cadherin,lost expression of N-cadherin,and reversed their highly invasive and drug resistant phenotype.