CAJ | 학술논문

目的应用无血清培养基分离胰腺癌干细胞，检测其miR-590-3p的表达。方法运用无血清培养基克隆培养ASPC-1、PANC1细胞，检测其单克隆形成、分化及细胞周期、半数抑制浓度(IC50)和表面标记物CD24+、CD44+表达。实时定量PCR法检测细胞miR-590-3p的表达。结果经无血清培养基培养,(0.94±0.53)％的ASPC-1细胞和(0.57±0.12)％的PANC1细胞能存活，呈克隆球样悬浮生长，并可以在体外连续传代。加入血清后细胞球又重新贴壁生长。ASPC-1细胞球G0/G1期比例和CD24+、C44+、CD24+ CD44+的细胞比例及IC50分别为(75.3±5.4)％、0.96％～2.01％、27.52％～34.47％、0.35％～0.44％和(224.37±5.71) μg/ml，均显著高于亲本细胞的(43.7±3.8)％、0.38％～0.42％、17.65％～ 18.25％、0.05％～ 0.08％、(11.43±2.10) μg/ml(P值均＜0.05)。PANC1细胞球G0/G1期比例和CD24+、CD44+、CD24 +CD44+的细胞比例及IC50分别为(80.1±4.7)％、5.31％～9.84％、72.05％～93.06％、4.91％～5.21％、(296.58±4.27) μg/ml，均显著高于亲本细胞的(46.1±5.3)％、4.09％～4.97％、47.71％～55.66％、1.48％～2.63％、(26.17±3.81) μg/ml(P值均＜0.05)。ASPC-1、PANC1细胞球miR-590-3p表达分别是亲本细胞的4.67和4.52倍。结论应用无血清培养基可以从ASPC-1、PANC1细胞系中分离出具有干细胞特性的胰腺癌细胞球，其miR-590-3p表达上调，该基因可能是胰腺癌干细胞特性维持的关键基因。
목적 응용무혈청배양기분리이선암간세포，검측기miR-590-3p적표체。방법 운용무혈청배양기극륭배양ASPC-1、PANC1세포，검측기단극륭형성、분화급세포주기、반수억제농도(IC50)화표면표기물CD24+、CD44+표체。실시정량PCR법검측세포miR-590-3p적표체。결과 경무혈청배양기배양,(0.94±0.53)％적ASPC-1세포화(0.57±0.12)％적PANC1세포능존활，정극륭구양현부생장，병가이재체외련속전대。가입혈청후세포구우중신첩벽생장。ASPC-1세포구G0/G1기비례화CD24+、C44+、CD24+ CD44+적세포비례급IC50분별위(75.3±5.4)％、0.96％～2.01％、27.52％～34.47％、0.35％～0.44％화(224.37±5.71) μg/ml，균현저고우친본세포적(43.7±3.8)％、0.38％～0.42％、17.65％～ 18.25％、0.05％～ 0.08％、(11.43±2.10) μg/ml(P치균＜0.05)。PANC1세포구G0/G1기비례화CD24+、CD44+、CD24 +CD44+적세포비례급IC50분별위(80.1±4.7)％、5.31％～9.84％、72.05％～93.06％、4.91％～5.21％、(296.58±4.27) μg/ml，균현저고우친본세포적(46.1±5.3)％、4.09％～4.97％、47.71％～55.66％、1.48％～2.63％、(26.17±3.81) μg/ml(P치균＜0.05)。ASPC-1、PANC1세포구miR-590-3p표체분별시친본세포적4.67화4.52배。결론 응용무혈청배양기가이종ASPC-1、PANC1세포계중분리출구유간세포특성적이선암세포구，기miR-590-3p표체상조，해기인가능시이선암간세포특성유지적관건기인。
Objectives To isolate cancer stem cells (CSCs) in pancreatic cancer cell lines PANC1 and ASPC-1 with serum-free medium( SFM ), and to detect the expression of miR-590-3p in CSCs. Methods PANC1 and ASPC-1 cells was cultured in serum-free medium. The monoclonal formation, differentiation and cell cycle, half inhibitory concentration ( IC50 ), and the expression of the surface markers CD24 + , CD44 + were detected. qRT-PCR was used to detect the expression of miR-590-3p. Results After SFM culture, (0.94 ±0.53 ) ％ of ASPC-1 and (0.57 + 0. 12 ) ％ PANC1 survived, and they formed spheres, and could continuously passage in vitro. Cell spheres differentiation recurred when serum was supplemented in SFM. The G0/G1 stage proportion, CD24+ , CD44 + , CD24+ CD44+ cells proportion, IC50 in ASPC-1 cell were (75.3 ± 5.4)％,0.96％～ 2.01％, 27.52％～ 34.47％, 0.35％～ 0.44％ and (224.37 ± 5.71 ) μg/ml, which were significantly higher than that those in parent cell [ (43.7 ± 3.8 ) ％, 0. 38％～ 0.42％, 17.65％～ 18.25％,0.05％～0.08％, (11.43 ±2.10)μg/ml, P＜0.05]. The G0/G1 stage proportion, CD24+ ,CD44+ ,CD24 +CD44 + cells proportion, IC50 in PANC 1 cell were ( 80. 1 ± 4.7) ％, 5.31％～ 9.84％, 72.05％～ 93.06％,4.91％～5.21％, (296.58±4.27) μg/ml, which were significantly higher than that those in parent cell [ (46.1 ±5.3)％, 4.09％～4.97％, 47.71％～55.66％, 1.48％～2.63％, (26.17 ±3.81)μg/ml, P＜0.05]. The expression of miR-590-3p in ASPC-1, PANC1 spheres was 4.67 and 4.52 times higher than the expression in parent cell lines. Conclusions Pancreatic cancer cell spheres can be isolated from ASPC-1, PANC1 by culture with SFM. miR-590-3p is up-regulated and may play an important role in regulating biological characteristics of pancreatic cancer stem cells.