CAJ | 학술논문

目的：研究雷酚萜甲醚( TME )在体外对人胃癌AGS细胞的增殖抑制和诱导凋亡作用。方法采用 MTT法观察TME对人胃癌AGS细胞、正常人胃黏膜上皮细胞GES-1的增殖抑制作用；克隆形成实验观察细胞克隆的形成；光镜下及AO / EB染色观察细胞形态；流式细胞术检测细胞凋亡和细胞周期；JC-1染色和DCFH-DA荧光探针分别检测TME对AGS细胞线粒体膜电位的改变和活性氧产生的影响；West-ern blot检测凋亡蛋白caspase-3、caspase-8和Bcl-2、Bax表达情况,以及 caspase 广谱抑制剂 z-VAD-fmk 对 caspase-3、caspase-8蛋白表达的影响。结果 TME可明显抑制人胃癌AGS细胞的增殖,并诱导其凋亡,作用48 h时IC50为23.85μmol·L-1,而对正常人胃黏膜上皮细胞GES-1的抑制作用明显低于AGS。 TME能够抑制AGS细胞克隆形成,并使细胞出现明显的凋亡形态改变。 Annexin V-FITC / PI双染实验表明,随TME剂量的增加,细胞凋亡百分数也增加。 JC-1和DCFH-DA结果显示, TME使细胞内线粒体膜电位降低、细胞内活性氧水平增加。 Western blot结果显示, TME增加了Bax / Bcl-2的比例,激活caspase-8和caspase-3,并且加入z-VAD-fmk后, caspase-3、caspase-8蛋白的表达降低。 TME可使AGS细胞周期阻滞于G0/G1期。结论 TME能抑制人胃癌AGS细胞增殖和诱导凋亡,抗肿瘤作用机制与激活凋亡通路、影响细胞周期及Bcl-2蛋白家族相关。
목적：연구뢰분첩갑미( TME )재체외대인위암AGS세포적증식억제화유도조망작용。방법채용 MTT법관찰TME대인위암AGS세포、정상인위점막상피세포GES-1적증식억제작용；극륭형성실험관찰세포극륭적형성；광경하급AO / EB염색관찰세포형태；류식세포술검측세포조망화세포주기；JC-1염색화DCFH-DA형광탐침분별검측TME대AGS세포선립체막전위적개변화활성양산생적영향；West-ern blot검측조망단백caspase-3、caspase-8화Bcl-2、Bax표체정황,이급 caspase 엄보억제제 z-VAD-fmk 대 caspase-3、caspase-8단백표체적영향。결과 TME가명현억제인위암AGS세포적증식,병유도기조망,작용48 h시IC50위23.85μmol·L-1,이대정상인위점막상피세포GES-1적억제작용명현저우AGS。 TME능구억제AGS세포극륭형성,병사세포출현명현적조망형태개변。 Annexin V-FITC / PI쌍염실험표명,수TME제량적증가,세포조망백분수야증가。 JC-1화DCFH-DA결과현시, TME사세포내선립체막전위강저、세포내활성양수평증가。 Western blot결과현시, TME증가료Bax / Bcl-2적비례,격활caspase-8화caspase-3,병차가입z-VAD-fmk후, caspase-3、caspase-8단백적표체강저。 TME가사AGS세포주기조체우G0/G1기。결론 TME능억제인위암AGS세포증식화유도조망,항종류작용궤제여격활조망통로、영향세포주기급Bcl-2단백가족상관。
Aim Toinvestigatetheeffectsoftriptonot-erpene methyl ether ( TME ) , a diterpene derived from the medicinal plant Triptergium wilfordii, on human gastric cancer AGS cell proliferation inhibition and ap-optosisinducedinvitro.Methods MTTassaywas used for screening tumor spectrum and detecting the vi-ability of AGS cells and normal human gastric epitheli-al cells GES-1 . Cell morphology was observed by light microscopy and AO / EB staining. Flow cytometry was used to detect cell apoptotic rate and cell cycle. JC-1 staining and fluorescence probe DCFH-DA were em-ployed to detect the changes of mitochondrial mem-brane potential and reactive oxygen species ( ROS ) . The effect of inhibiting AGS clonogenic survival was as-sayed by the method of plate clone formation. Western blot was used to analyse the expression of caspase-3 , caspase-8,Bcl-2andBax.Results MTTresults showed that TME exhibited significantly higher cytotox-icity to gastric cancer AGS cell line than to noncancer-ous cell line GES-1. IC50 for AGS of 48 h treatment was 23 . 85 μmol · L-1 . TME significantly inhibited colony formation and caused morphological changes in AGS cells. Annexin V-FITC / PI double staining showed the apoptotic rate increased. DCFH-DA stai-ning showed TME resulted in an increase in intracellu-lar ROS levels. Mitochondrial membrane potential de-creased after TME treatment. Western blot results showed that TME increased the proportion of Bax /Bcl-2 , with the activation of caspase-8 and caspase-3 . The broad-spectrum caspase inhibitor z-VAD-fmk pre-treatment reduced the expression of caspase-8 and caspase-3. TME enabled AGS cell cycle arrest in G0/G1phase.Conclusion TMEpossessespotenttumor selected toxicity and can induce apoptosis of AGS cells through cell cycle arrest, which is associated with Bcl-2 protein family.