对培美曲塞耐药胰腺癌Patu8988细胞株克隆、侵袭能力及相关耐药基因的研究
杨军，石欣,严伟,林士波,顾海涛,钱程佳
【摘要】 目的：探讨对培美曲塞耐药的胰腺癌Patu8988细胞株相关于亲本株在细胞克隆、侵袭能力及多药耐药相关蛋白1(MDR1)、ABC多药转运蛋白G家族成员2(ABCG2)、ABC多药转运蛋白C家族成员3(ABCC3)耐药基因的转变。方式：采纳双层软琼脂克隆实验别离在无药及加入500 μmol·L-1培美曲塞的环境下测定胰腺癌Patu8988耐药株及亲本株克隆生成能力的改变；采纳Transwell侵袭小室检测两种条件下胰腺癌Patu8988侵袭能力的不同；采纳RT
PCR
法检测胰腺癌Patu8988细胞MDR一、ABCG二、ABCC3的mRNA表达水平,Wersten blot检测ABCG2和ABCC3的蛋白表达水平,免疫荧光及激光共聚焦检测ABCG2的表达及散布。结果：在无药及加入500 μmol·L-1培美曲塞的环境下,对培美曲塞耐药胰腺癌Patu8988细胞株的增殖克隆能力均与其亲本株有不同(P＜,而两种培育条件下胰腺癌Patu8988细胞株的侵袭能力转变不同无统计学意义(P＞。RT
PCR
检测显示,在耐药株中ABCG二、ABCC3 mRNA表达别离是其亲本株的倍和倍左右,而MDR1 mRNA表达仅提高到倍；Werstenblot结果显示蛋白表达为其亲本株的倍和倍。免疫荧光检测结果显示,耐药株中ABCG2的荧光强度较强,而其亲本株细胞较弱。激光共聚焦显示，耐药株中ABCG2表达要紧散布于细胞膜，细胞质内也有少量散布。结论：相关
于亲本株，对培美曲塞耐药的胰腺癌Patu8988细胞株克隆能力增强,而侵袭能力无明显改变。多药耐药基因表达均有不同程度的提高,以ABCG2基因表达提高尤其明显,而且其在耐药株中要紧散布在细胞膜,细胞质内也有散布。
【关键词】 胰腺肿瘤； 取得性耐药； 培美曲塞； ABC多药转运蛋白G家族成员2
［Abstract］ Objective:To investigate the cloning capability,invasion and associated multigenes:MDR1(P
drug resistance
gp),ABCG2,ABCC3 of Patu8988 human pancreatic
cancer cells resistance to the pemetrexed compared with it′s parental :The cloning capability of Patu8988 human pancreatic cancer and it′s parental strain were detected in condition free or adding 500 μmol·L-1 pemetrexed by soft agar cloning invasion ability betweent them were detected by transwell invasion mRNA expression of MDR1,ABCG2 and ABCC3 were determined by RT
PCR,ABCG2 and ABCC3 protein expression levels
were detected by Wersten blot,the expression and distribution of ABCG2 were detected by immunofluorescence and laser scanning confocal :The cloning capability between drug
free and adding
500 μmol·L-1 pemetrexed was significantly different(P
＜,whereas there was no significant changes of the invasion ability between them(P＞.RT
PCR and Wersten blot showed the
mRNA and protein levels of ABCG2,ABCC3 were almost 250% and 150% in the drugthe
resistant cells than their parental cells, while
of
MDR1
mRNA
increased
only
expression
20%.Immunofluorescence showed drugresistant cells of ABCG2
stronger fluorescence intensity than their parental confocal detection of drug
resistant cells showed ABCG2 expression was
mainly distributed in the cell membrane,also a small amount of the :The cloning capability is significantly different between Patu8988 human paucreatic cancer cells resistance to the pemetrexed and it′s parental strain, whereas invasion ability is no is particularly increase than other multi
drug
resistance genes and it is mainly distributed in the cell membrane and distribution．
［Key words］ pancreatic neoplasms； acquired resistance； pemetrexed； ABCG2
本实验室前期通过体外多次大剂量冲击诱导法成功诱导出对培美曲塞耐药的胰腺癌Patu8988细胞株［1］(IC50=倍),通过Affymetrix基因芯片比对分析发觉某些与耐药相关基因的转变。本研
对培美曲塞耐药胰腺癌Patu8988细胞株克隆侵袭能力及相关耐药基因的研究
