CAJ | 학술논문

目的探讨CD133阳性胃癌起始细胞对常规化疗药物氟尿嘧啶(5-FU)的敏感性及其耐药机制.方法免疫磁珠法分选KATO-Ⅲ、SGC7901和MKN-45胃癌细胞株并分为未分选组、CD133+组及CD133-组.Western blot法和RT-PCR法分别检测分选后胃癌细胞CD133、P-gp、Bax和Bcl-2蛋白及mRNA表达水平.免疫荧光法检测各组细胞P-gp及Bcl-2蛋白的表达.CCK-8法和Hoechest染色检测各组细胞对5-FU的敏感性.siRNA干扰MKN45细胞CD133表达后,检测CD133、P-gp、Bcl-2、Akt及p-Akt蛋白及mRNA表达.结果 CD133+组胃癌细胞中CD133、P-gp及Bcl-2蛋白和mRNA表达水平均显著高于未分选组及CD133-组(均P＜0.05),而促凋亡因子Bax的表达水平则明显降低(P＜0.05).在相同药物浓度下,5-FU对CD133+组的抑制率显著低于CD133-组(P＜0.05).5-FU处理胃癌细胞后,CD133+组胃癌细胞中凋亡细胞比率明显低于CD133-组及未分选组(P＜0.05).CD133特异siRNA干扰胃癌细胞后,干扰组P-gp、Bcl-2和p-Akt蛋白及mRNA表达显著降低(均P＜0.05),而Bax表达水平则显著增加(P＜0.05).结论 CD133可能通过调节P-gp和Bcl-2的表达来促进胃癌的化疗耐药；在胃癌化疗耐药产生中,CD133+细胞可通过PI3K/Akt通路起作用.
목적 탐토CD133양성위암기시세포대상규화료약물불뇨밀정(5-FU)적민감성급기내약궤제.방법 면역자주법분선KATO-Ⅲ、SGC7901화MKN-45위암세포주병분위미분선조、CD133+조급CD133-조.Western blot법화RT-PCR법분별검측분선후위암세포CD133、P-gp、Bax화Bcl-2단백급mRNA표체수평.면역형광법검측각조세포P-gp급Bcl-2단백적표체.CCK-8법화Hoechest염색검측각조세포대5-FU적민감성.siRNA간우MKN45세포CD133표체후,검측CD133、P-gp、Bcl-2、Akt급p-Akt단백급mRNA표체.결과 CD133+조위암세포중CD133、P-gp급Bcl-2단백화mRNA표체수평균현저고우미분선조급CD133-조(균P＜0.05),이촉조망인자Bax적표체수평칙명현강저(P＜0.05).재상동약물농도하,5-FU대CD133+조적억제솔현저저우CD133-조(P＜0.05).5-FU처리위암세포후,CD133+조위암세포중조망세포비솔명현저우CD133-조급미분선조(P＜0.05).CD133특이siRNA간우위암세포후,간우조P-gp、Bcl-2화p-Akt단백급mRNA표체현저강저(균P＜0.05),이Bax표체수평칙현저증가(P＜0.05).결론 CD133가능통과조절P-gp화Bcl-2적표체래촉진위암적화료내약；재위암화료내약산생중,CD133+세포가통과PI3K/Akt통로기작용.
Objective To explore the relationship between CD133 + subsets cells in human gastric cancer (GC) and molecules of drug resistance and their sensitivity to 5-FU.Methods Three gastriccancer cell lines therein KATO-Ⅲ,SGC7901 and MKN45 were sorted by immunomagnetic beads cell sorting method.Then above cell lines were further divided into un-sorted GC cells,CD133+ subgroup and CD133-subgroup.The expressions of CD133,P-gp,Bax and Bcl-2 were determined by RT-PCR,Western blot and immunoflurescence.Meanwhile,the sensitivity to 5-FU of three subgroups was detected by CCK-8 Kit.The apoptosis induced by 5-FU in three subgroups was determined by Hoechst 33258.Results Expressions of CD133 in three CD133+ subgroups were significantly higher than those in un-sorted GC cells and CD133-subgroup (all P＜0.05).Expressions of P-gp and Bcl-2 in the three GC cell lines were different (all P＜0.05).There were significant differences of expressions of P-gp,Bcl-2 and Bax among CD133+ cells,un-sorted GC cells and CD133-cells (all P＜0.05).CCK-8 detection showed that CD133-subgroup of MKN45 GC cell line was more sensitive than CD133+ cells to 5-FU (P＜0.05).Hoechst 33258 staining showed that there were more apoptotic cells in CD133-subgroup as compared to other two subgroups,and the least apoptotic cells were observed in CD133+ subgroup of MKN45 GC cell line (P＜0.05).CD133 siRNA was transfected into MKN45 GC cell line and could down-regulate the expressions of CD133,P-gp,Bcl-2 and p-Akt,while the expression of Bax increased (all P＜0.05).Conclusions CD133 may contribute to the resistance of GC cells to chemotherapy drug through P-gp,Bcl-2 and Bax.PI3K/Akt signal pathway may be involved in this process.