中华胰腺病杂志
中華胰腺病雜誌
중화이선병잡지
CHINESE JOURNAL OF PANCREATOLOGY
2012年
4期
242-245
,共4页
邵叶波%靳大勇%戎叶飞%许雪峰
邵葉波%靳大勇%戎葉飛%許雪峰
소협파%근대용%융협비%허설봉
RNA,小分子干扰%基因沉默%成纤维细胞活化蛋白%胰腺肿瘤%细胞增殖%细胞凋亡
RNA,小分子榦擾%基因沉默%成纖維細胞活化蛋白%胰腺腫瘤%細胞增殖%細胞凋亡
RNA,소분자간우%기인침묵%성섬유세포활화단백%이선종류%세포증식%세포조망
RNA,small interfering%Gene silence%Fibroblast activation protein%Pancreatic neoplasms%Cell proliferation%Apoptosis
目的 应用RNA干扰技术沉默小鼠胰腺癌相关成纤维细胞的成纤维活化蛋白(FAP)表达,观察其对小鼠原代胰腺癌细胞增殖及凋亡的影响.方法 构建靶向小鼠FAP基因的重组表达质粒siFAP及对照质粒siMOCK,分别转染小鼠原代胰腺癌相关成纤维细胞mPCa-FCs-1212,采用定量RT-PCR法及蛋白质印迹法检测转染细胞的FAP mRNA及蛋白表达.将该细胞与小鼠原代胰腺癌细胞mPCa-1212按1∶1的比例共培养,应用MTT比色法检测mPCa-1212细胞的增殖抑制率,应用Annexin V-FTTC/PI染色及流式细胞仪检测细胞凋亡.结果 转染重组质粒siFAP的mPCa-FCs-1212细胞的FAP mRNA及蛋白表达较转染对照质粒siMOCK的细胞的表达明显下调[0.584±0.029比1.052±0.281,P=0.0213;(27.18±3.23)％比(61.58±4.72)％,P=0.0317].转染siFAP和转染siMOCK的mPCa-FCs-1212分别与mPCa-1212细胞共培养3d后,mPCa-1212的增殖抑制率分别为(23.02±3.32)％和(1.11±0.23)％；细胞凋亡率分别为(42.31±5.34)％和(7.38±2.09)％,两组细胞的差异具有统计学意义(P=0.000).结论 沉默FAP基因的mPCa-FCs-1212在体外可有效抑制mPCa-1212细胞的增殖,诱导细胞凋亡,可能是一种潜在的基因治疗新方法.
目的 應用RNA榦擾技術沉默小鼠胰腺癌相關成纖維細胞的成纖維活化蛋白(FAP)錶達,觀察其對小鼠原代胰腺癌細胞增殖及凋亡的影響.方法 構建靶嚮小鼠FAP基因的重組錶達質粒siFAP及對照質粒siMOCK,分彆轉染小鼠原代胰腺癌相關成纖維細胞mPCa-FCs-1212,採用定量RT-PCR法及蛋白質印跡法檢測轉染細胞的FAP mRNA及蛋白錶達.將該細胞與小鼠原代胰腺癌細胞mPCa-1212按1∶1的比例共培養,應用MTT比色法檢測mPCa-1212細胞的增殖抑製率,應用Annexin V-FTTC/PI染色及流式細胞儀檢測細胞凋亡.結果 轉染重組質粒siFAP的mPCa-FCs-1212細胞的FAP mRNA及蛋白錶達較轉染對照質粒siMOCK的細胞的錶達明顯下調[0.584±0.029比1.052±0.281,P=0.0213;(27.18±3.23)％比(61.58±4.72)％,P=0.0317].轉染siFAP和轉染siMOCK的mPCa-FCs-1212分彆與mPCa-1212細胞共培養3d後,mPCa-1212的增殖抑製率分彆為(23.02±3.32)％和(1.11±0.23)％；細胞凋亡率分彆為(42.31±5.34)％和(7.38±2.09)％,兩組細胞的差異具有統計學意義(P=0.000).結論 沉默FAP基因的mPCa-FCs-1212在體外可有效抑製mPCa-1212細胞的增殖,誘導細胞凋亡,可能是一種潛在的基因治療新方法.
목적 응용RNA간우기술침묵소서이선암상관성섬유세포적성섬유활화단백(FAP)표체,관찰기대소서원대이선암세포증식급조망적영향.방법 구건파향소서FAP기인적중조표체질립siFAP급대조질립siMOCK,분별전염소서원대이선암상관성섬유세포mPCa-FCs-1212,채용정량RT-PCR법급단백질인적법검측전염세포적FAP mRNA급단백표체.장해세포여소서원대이선암세포mPCa-1212안1∶1적비례공배양,응용MTT비색법검측mPCa-1212세포적증식억제솔,응용Annexin V-FTTC/PI염색급류식세포의검측세포조망.결과 전염중조질립siFAP적mPCa-FCs-1212세포적FAP mRNA급단백표체교전염대조질립siMOCK적세포적표체명현하조[0.584±0.029비1.052±0.281,P=0.0213;(27.18±3.23)％비(61.58±4.72)％,P=0.0317].전염siFAP화전염siMOCK적mPCa-FCs-1212분별여mPCa-1212세포공배양3d후,mPCa-1212적증식억제솔분별위(23.02±3.32)％화(1.11±0.23)％；세포조망솔분별위(42.31±5.34)％화(7.38±2.09)％,량조세포적차이구유통계학의의(P=0.000).결론 침묵FAP기인적mPCa-FCs-1212재체외가유효억제mPCa-1212세포적증식,유도세포조망,가능시일충잠재적기인치료신방법.
Objective Small interfering RNA (siRNA) was used to silence the fibroblast activation protein4 (FAP) expression of mouse pancreatic cancer related fibroblast cells (mPCa-FCs-1212),and to observe the effects of mPCa-FCs-1212 silencing FAP gene on mouse pancreatic cancer cells (mPCa-1212) proliferation and apoptosis.Methods The small interfering RNA targeting FAP gene was designed; the recombinant siRNA plasmid siFAP and control plasmid siMOCK was constructed,which were transfected into mPCa-FCs-1212,respectively.The FAP mRNA and protein expression in transfected cells were examined by real-time PCR and Western blotting.The mPCa-1212 and transfected mPCa-FCs-1212 were co-cultured with a 1:1 ratio in vitro.The growth inhibitory rates and apoptosis rates of mPCa-1212 were detected by MTT assay and Annexin V-FTTC/PI staining and FCM assay.Results The mRNA and protein expressions of FAP in siFAP transfected mPCa-FCs-1212 were significantly down-regulated when compared with that in siMOCK transfected mPCa-FCs-1212[0.584 ±0.029vs.1.052±0.281,P=0.0213; (27.18±3.23)％ vs.(61.58±4.72)％,P=0.0317].The mPCa-1212 was co-cultured with the mPCa-FCs-1212 transfected with siFAP or siMOCK for 3 d,and the inhibitory rates of mPCa-1212 were (23.02 ±3.32)％ and (1.11 ±0.23)％,and the apoptosis rates were (42.31 ±5.34)％ and (7.38 ± 2.09)％,the difference between the two groups was statistically significant (P =0.000).Conclusions mPCa-FCs-1212 silencing FAP gene can inhibit the proliferation of mPCa-1212 in vitro and induce cell apoptosis,and may be a potential new approach to gene therapy.
