摘要
目的:体外研究刀豆蛋白A(Concanavalin A,Con A)联合淋巴细胞对人卵巢癌细胞株SKOV3细胞形态、活力、凋亡以及顺铂敏感性的影响。方法:应用密度梯度离心法分离健康成人新鲜血中的单个核细胞,培养3～5d待单核细胞完全贴壁后分离出悬浮的淋巴细胞,与SKOV3细胞混合培养,并设对照组(未处理的SKOV3)、实验组1(SKOV3培养体系中仅加入5μg/mL Con A)、实验组2(SKOV3和淋巴细胞混合培养)和实验组3(SKOV3和淋巴细胞混合培养体系中加入5μg/mL Con A)。培养7d后将SKOV3分离出继续培养,并进行以下实验,在光学显微镜下观察细胞形态变化;CCK8检测24和48h细胞活性;流式细胞术检测24和48h细胞凋亡情况;用浓度0和60μmol/L顺铂处理SKOV3细胞24和48h后,流式细胞术检测细胞凋亡率以判断细胞对顺铂的敏感性。结果:与对照组比较,实验组1和实验组2中细胞无明显改变,而实验组3中SKOV3与淋巴细胞混合培养后发生明显改变,如细胞突起减少,细胞变圆。与对照组比较,实验组1中24和48h细胞存活率分别为(105.68±7.12)%(z=-1.256,P=0.30)和(111.90±8.28)%(z=-1.766,P=0.13),实验组2分别为(101.05±5.92)%(z=-0.215,P=0.79)和(109.72±6.54)%(z=-1.766,P=0.13),差异均无统计学意义;而实验组3分别为(25.08±7.71)%(z=-2.087,P=0.004)和(14.6±4.41)%(z=-3.431,P=0.001),差异有统计学意义。与对照组比较,实验组1中24和48h细胞凋亡率分别为(4.43±1.49)%(z=-2.78,P=0.61)和(10.95±1.09)%(z=-1.982,P=0.08),实验组2分别为(4.79±0.83)%(z=-1.349,P=0.28)和(10.59±1.03)%(z=-1.349,P=0.28),差异均无统计学意义;实验组3分别为(13.88±1.00)%(z=-2.247,P=0.003)和(21.37±2.35)%(z=-2.137,P=0.006),差异有统计学意义。顺铂处理各组SKOV3细胞后,与对照组比较,实验组1中24和48h细胞凋亡率分别为(11.25±1.86)%(z=-0.361,P=0.59)和(33.66±4.96)%(z=-0.536,P=0.54),实验组2分别为(10.32±0.14)%(z=-0.225,P=0.65)和(31.56±6.21)%(z=-0.163,P=0.83),差异均无统计学意义;实验组3分别为(41.99±6.66)%(z=-2.14,P=0.03)和(66.5±2.55)%(z=-1.909,P=0.04),差异有统计学意义。结论:Con A联合淋巴细胞可显著改变SKOV3细胞形态,降低细胞活力,促进细胞凋亡,并增加细胞对顺铂的敏感性,而Con A或淋巴细胞单独处理并无此效应。
OBJECTIVE： To investigate the effects of Con A combined with lymphocytes on morphology,viability, apoptosis and sensitivity to cisplatin of ovarian cancer cell line SKOV3 in vitro. METHODS： Peripheral blood mononuclear cells （PBMCs） from normal healthy blood donors were prepared by Ficoll-Hypaque gradient centrifugation. Lymphocytes were isolated after monocytes adherent followed by coculture with SKOV3,groups were divided as follows： Control group （SKOV3 without treatment）,Experimental group 1 （SKOV3 were treated with 5 μg/mL Con A）,Experimental group 2 （coculture of SKOV3 and lymphocytes）, Experimental group 3 （coculture of SKOV3 and lymphocytes treated with 5 μg/mL Con A）. After 7 days,SKOV3 cells were isolated and continued to be cultured followed by several experiments： Cell morphology was observed by optical microscope; Cell viability was detected using CCK8 at 24 and 48 h; Apoptosis was assessed by flow cytometry at 24 and 48 h; 4） Apoptosis was assessed by flow cytometry after SKOV3 ceils were treated with 0 and 60 μmol/L cisplatin for 24 and 48 h. RESULTS：Compared with control group, there was no change in experimental group 1 and 2,but there were significant changes happened in SKOV3 cells： cellular protruding decreased and cells turned round; Compared with control group, cell viability at 24 and 48 h in experimental 1 were （105.68± 7.12）% （z=-1. 256,P=0.30） and （111.90±8.28）% （z=-1. 766,P=0.13）,and in experimental 2 were （101.05±5.92） % （z= - 0. 215, P = 0.79） and （ 109.72 ±6.54） % （z = - 1. 766, P = 0.13）, there was no significant difference ; but in experimental 3 the cell viability at 24 and 48 h were （25.08±7.71）% （z=-2. 087,P=0. 004） and （14. 6±4.41）% （z=-3. 431 ,P=0. 001）,the difference was significant. Compared with control group,cell apoptosis at 24 and 48 h were in experimental 1 were （4.43±1.49）% （z=-2.78,P=0.61） and （10. 95±1.09）% （z=-1. 982,P=0.08） ,and in ex- perimental 2 were （4.79±0.83）% （z=-1. 349,P=0.28） and （10.59±1.03）% （z=-1. 349,P=0.28） ,there was no significant difference;but in experimental 3 the cell viability at 24 and 48 h were （13.88 ±1.00）% （z =- 2. 247, P = 0. 003） and （21.37±2.35）% （z=-2. 137,P=0. 006）,the difference was significant. Compared with control group,cell apoptosis after treated with cisplatin for 24 and 48 h were in experimental 1 were （11.25±1.86）% （z= -0. 361,P= 0.59） and （33.66±4. 96）% （z=-0. 536,P=0.54）,and in experimental 2 were （10.32±0.14）% （z= -0. 225,P= 0.65） and （31.56±6.21）% （z=-0. 163 ,P=0.83）,there was no significant difference;but in experimental 3 the cell vi- ability at 24 and 48 hwere （41.99±6.66）% （z=-2.14,P=0.03） and （66.5±2.55）% （z=-1. 909 , P= 0. 04） , the difference was significant. CONCLUSION.. Con A combined with lymphocytes, but not Con A or lymphocytes, can change the morphology of SKOV3, decrease its viability, promote its apoptosis,and increase the sensitivity to cisplatin in vitro.
出处
《中华肿瘤防治杂志》
CAS
北大核心
2014年第14期1068-1072,共5页
Chinese Journal of Cancer Prevention and Treatment
