DNA甲基转移酶1抑制对涎腺腺样囊性癌细胞生长、侵袭及转移的影响

目的 研究DNA甲基转移酶1(DNA methyltmnderases 1,DNMT-1)表达抑制对涎腺腺样囊性癌(salivary adenoid cystic carcinoma,SACC)细胞体内、体外的生物学影响,探讨DNMT-1在SACC发生、侵袭及转移中的作用.方法 对前期实验建立的DNMT-1表达稳定抑制的ACC-M细胞(实验干扰组)、空载对照ACC-M细胞系(空载对照组)及ACC-M细胞系(空白对照组),在体外分别应用甲基噻唑基四唑(MTT)法生长曲线分析、流式细胞周期分析、平板克隆实验、体外侵袭能力分析等方法,体内对裸鼠皮下、尾静脉注射肿瘤细胞,综合分析评估DNMT-1表达抑制对ACC-M细胞体内增殖、侵袭及转移的影响.结果 实验干扰组细胞倍增时间延长[(34.7±2.1)h]、细胞周期S期比例[(17.4±117)%]、克隆形成率[(43.0±1.3)%]降低,与空白对照组[分别为(26.2±3.1)h、(31.5±2.0)%、(71.0±4.7)%]及空载对照组[分别为(28.4±3.9)h、(39.0±2.0)%、(66.0±5.2)%]差异有统计学意义(P＜0.05);各组体外侵袭能力的差异无统计学意义(P＞0.05);体内DNMT-1表达稳定抑制的ACC-M细胞,其皮下成瘤率(6/10)、瘤体体积[(2.18±0.83)mm~3]、重量[(0.0156±0.0046)g]均显著低于空白对照组[分别为10/10,(155.44±1.67)mm~3、(0.0724±0.0157)g]及空载对照组[分别为10/10、(147.46±1.73)mm~3、(0.0729±0.0177)g],差异均有统计学意义(P＜0.05);各组的肺转移率差异无统计学意义(P＞0.05);DNMT-1表达稳定抑制的ACC-M细胞,其肺转移瘤的个数[(2.0±0.5)个]、直径[(70.0±20.3)μm]均显著低于空白[(28.0±5.5)个、(195.0±25.4)μm]及空载(27.0±4.5)个、(190.0±19.9)μm]对照组,P＜0.05.结论 抑制DNMT-1的表达能有效降低ACC细胞的生长、增殖及转移能力.

Effect of DNA methyltransferases 1 inhibition on proliferation, invasion, and metastasis in ACC-M line

Objective To investigate the effect of DNA methyltransferases 1(DNMT-1) inhibition on the ACC-M cells in vitro and in vivo and discuss the role of DNMT-1 in the development,invasion and metastasis of salivary adenoid cystic carcinoma (SACC).Methods ACC-M cells of stable DNMT-1 inhibition were established in a previous research.In vitro,the growth and invasion of ACC-M cells which stably inhibited DNMT-1 were detected and analyzed by methyl thiazolyl tetrazolium (MTF) growth curve,flow cytometry,plating efficiency and invasion assay.In vivo,the growth and metastasis of ACC-M cells which persistently inhibited DNMT-1 were observed and analyzed by subcutaneous injection and tail vein injection into the nude mice.Results In vitro,the doubling time [(34.7±2.1) h],S phase fraction [(17.4±1.7)%],plating efficiency [(43.0±1.3)%]of ACC-M cells was significantly different from those of blank [(26.2±3.1) h,(31.5±2.0)%,(71.0±4.7)%],empty load control [(28.4±3.9) h,(39.0±2.0)%,(66.0±5.2)%],P＜0.05,and the invasion ability was not significantly different among these groups(P＞0.05).In vivo,the subcutaneous tumor forming rate(6/10),volume[(2.18±0.83) mm~3],weight[(0.0156±0.0046) g]of ACC-M cells was also significantly lower than that of blank 10/10,(155.44±1.67) mm~3,(0.0724±0.0157) g],empty load control[10/10,(147.46±1.73) mm3,(0.0729±0.0177) g],P＜0.05,but the rate of lung metastasis was not significantly different among these groups(P＞0.05),and the masses(2.0±0.5),diameter(70.0±20.3) μm of ACC-M cells was significantly lower than that of blank [(28.0±5.5),(195±25.4) μm],empty load control[(27.0±4.5),(190.0±19.9) μm],P＜0.05.Conclusions Inhibition of DNMT-1 is able to inhibit the proliferation and metastasis of ACC-M cells in vitro and in vivo.