临床与病理杂志
臨床與病理雜誌
림상여병리잡지
International Journal of Pathology and Clinical Medicine
2015年
6期
1080-1086
,共7页
钱静%陈不尤%刘贤称%顾红梅%姚宁华
錢靜%陳不尤%劉賢稱%顧紅梅%姚寧華
전정%진불우%류현칭%고홍매%요저화
非小细胞肺癌%人表皮生长因子受体%胰岛素样生长因子受体1%苦鬼臼毒%厄洛替尼
非小細胞肺癌%人錶皮生長因子受體%胰島素樣生長因子受體1%苦鬼臼毒%阨洛替尼
비소세포폐암%인표피생장인자수체%이도소양생장인자수체1%고귀구독%액락체니
non-small cell lung cancer%epithelial growth factor receptor (EGFR)%insulin-like growth factor receptor 1 (IGF1R)%picropodophyllotoxin%erlotinib
目的:构建厄洛替尼耐药人肺腺癌细胞模型PC-9/ER,观察单用人表皮生长因子受体(epithelial growth factor receptor,EGFR)酪氨酸激酶抑制剂厄洛替尼或联合胰岛素样生长因子受体1(insulin-like growth factor receptor 1,IGF1R)酪氨酸激酶抑制剂苦鬼臼毒(picropodophyllotoxin,PPP)作用于该细胞后,该细胞对厄洛替尼耐药性的影响,并探讨耐药相关机制。方法:选择人肺腺癌细胞株PC-9,采用逐步递增厄洛替尼浓度的方法体外诱导构建耐药株PC-9/ER。CCK-8法检测耐药指数;细胞计数法绘制PC-9和PC-9/ER的生长曲线,并计算出两细胞系的倍增时间;流式细胞术检测两细胞系的细胞周期;Western Blot法检测p-EGFR及p-IGF1R的表达水平,并进一步检测厄洛替尼及PPP单独或联合作用于PC-9/ER后,各组Akt,ERK,p-Akt及p-ERK的表达水平。结果:PC-9/ER细胞株的耐药指数是72.3。细胞生长曲线显示,PC-9/ER细胞生长较亲代细胞慢,PC-9与PC-9/ER细胞的倍增时间分别为32.9及36.9 h (P=0.003)。与PC-9相比,PC-9/ER细胞的G1期细胞增多(P=0.001),而S期细胞则明显下降(P=0.015)。Western Blot结果表明,PC-9/ER细胞中p-IGF1R表达比亲代细胞明显增多(P=0.016),而p-EGFR无明显变化(P=0.152)。在亲代及耐药细胞系,厄洛替尼联合PPP组均较其他组更能显著的抑制细胞增殖(P<0.05);且Western Blot法表明联合用药组EGFR下游磷酸化的Akt、ERK的表达水平明显减少。结论:成功构建了人肺腺癌厄洛替尼耐药细胞株PC-9/ER,IGF1R通路可能与肺腺癌EGFR-TKI获得性耐药有关。
目的:構建阨洛替尼耐藥人肺腺癌細胞模型PC-9/ER,觀察單用人錶皮生長因子受體(epithelial growth factor receptor,EGFR)酪氨痠激酶抑製劑阨洛替尼或聯閤胰島素樣生長因子受體1(insulin-like growth factor receptor 1,IGF1R)酪氨痠激酶抑製劑苦鬼臼毒(picropodophyllotoxin,PPP)作用于該細胞後,該細胞對阨洛替尼耐藥性的影響,併探討耐藥相關機製。方法:選擇人肺腺癌細胞株PC-9,採用逐步遞增阨洛替尼濃度的方法體外誘導構建耐藥株PC-9/ER。CCK-8法檢測耐藥指數;細胞計數法繪製PC-9和PC-9/ER的生長麯線,併計算齣兩細胞繫的倍增時間;流式細胞術檢測兩細胞繫的細胞週期;Western Blot法檢測p-EGFR及p-IGF1R的錶達水平,併進一步檢測阨洛替尼及PPP單獨或聯閤作用于PC-9/ER後,各組Akt,ERK,p-Akt及p-ERK的錶達水平。結果:PC-9/ER細胞株的耐藥指數是72.3。細胞生長麯線顯示,PC-9/ER細胞生長較親代細胞慢,PC-9與PC-9/ER細胞的倍增時間分彆為32.9及36.9 h (P=0.003)。與PC-9相比,PC-9/ER細胞的G1期細胞增多(P=0.001),而S期細胞則明顯下降(P=0.015)。Western Blot結果錶明,PC-9/ER細胞中p-IGF1R錶達比親代細胞明顯增多(P=0.016),而p-EGFR無明顯變化(P=0.152)。在親代及耐藥細胞繫,阨洛替尼聯閤PPP組均較其他組更能顯著的抑製細胞增殖(P<0.05);且Western Blot法錶明聯閤用藥組EGFR下遊燐痠化的Akt、ERK的錶達水平明顯減少。結論:成功構建瞭人肺腺癌阨洛替尼耐藥細胞株PC-9/ER,IGF1R通路可能與肺腺癌EGFR-TKI穫得性耐藥有關。
목적:구건액락체니내약인폐선암세포모형PC-9/ER,관찰단용인표피생장인자수체(epithelial growth factor receptor,EGFR)락안산격매억제제액락체니혹연합이도소양생장인자수체1(insulin-like growth factor receptor 1,IGF1R)락안산격매억제제고귀구독(picropodophyllotoxin,PPP)작용우해세포후,해세포대액락체니내약성적영향,병탐토내약상관궤제。방법:선택인폐선암세포주PC-9,채용축보체증액락체니농도적방법체외유도구건내약주PC-9/ER。CCK-8법검측내약지수;세포계수법회제PC-9화PC-9/ER적생장곡선,병계산출량세포계적배증시간;류식세포술검측량세포계적세포주기;Western Blot법검측p-EGFR급p-IGF1R적표체수평,병진일보검측액락체니급PPP단독혹연합작용우PC-9/ER후,각조Akt,ERK,p-Akt급p-ERK적표체수평。결과:PC-9/ER세포주적내약지수시72.3。세포생장곡선현시,PC-9/ER세포생장교친대세포만,PC-9여PC-9/ER세포적배증시간분별위32.9급36.9 h (P=0.003)。여PC-9상비,PC-9/ER세포적G1기세포증다(P=0.001),이S기세포칙명현하강(P=0.015)。Western Blot결과표명,PC-9/ER세포중p-IGF1R표체비친대세포명현증다(P=0.016),이p-EGFR무명현변화(P=0.152)。재친대급내약세포계,액락체니연합PPP조균교기타조경능현저적억제세포증식(P<0.05);차Western Blot법표명연합용약조EGFR하유린산화적Akt、ERK적표체수평명현감소。결론:성공구건료인폐선암액락체니내약세포주PC-9/ER,IGF1R통로가능여폐선암EGFR-TKI획득성내약유관。
Objective:To establish a human lung adenocarcinoma cell line PC-9/ER that is resistant to erlotinib; to observe the effect of erlotinib alone or in combination with PPP (a tyrosine kinase inhibitor of insulin-like growth factor receptor 1) on PC-9/ER and to discuss its possible mechanism.Methods: A erlotinib resistant humanlung adenocarcinoma cell line PC-9/ER was induced by continuously exposing human lung adenocarcinoma cell line PC-9 to gradually increased doses of erlotinib. The drug resistant ability to erlotinib was measured by CCK-8 assay. The growth curve and cell cycle of PC-9 and PC-9/ER were compared. The expression of Akt, ERK, p-EGFR, p-IGF1R, p-Akt and p-ERK was measured by Western Blot in different groups.Results: The drug resistant index of PC-9/ER to erlotinib was 72.3. Double time of PC-9 and PC-9/ER were 32.9 and 36.9 h (P=0.003), respectively, as evaluated by the growth curve. Figures in G1 phase was decreased in PC-9 than PC-9/ER (P=0.001). hTe expression of p-IGF1R signiifcantly increased in PC-9/ER (P=0.016) but not p-EGFR (P=0.152). More signiifcant inhibition of cell proliferation than other groups was observed in combination group that erlotinib combination with PPP (P<0.05). hTe expression of p-Akt and p-ERK was decreased in combination group.Conclusion: The resistant cell model PC-9/ER is established and IGF1R may associate with drug resistance to EGFR-TKI.