摘要目的 观察KISS-1基因对食管鳞癌细胞EC-1侵袭及增殖能力的影响,探讨KISS-1基因与食管鳞癌生物学行为的关系.方法 检测KISS-1在EC-1、Eca109、EC9706和TE-1四种食管癌细胞株中的表达;利用脂质体介导在体外将KISS-1基因转染人食管鳞癌细胞EC-1,利用Western blot和RT-PCR方法检测转染之后KISS-1表达的改变,并采用Boyden小室体外侵袭实验及MTT、软琼脂克隆形成实验,观察KISS-1对食管鳞癌细胞侵袭及增殖能力的影响.结果 4种食管癌细胞株的Western blot和RT-PCR检测结果显示:EC-1中KISS-1蛋白(0.715±0.109)及mRNA(0.670±0.176)表达均最低;转染之后Western blot和RT-PCR结果显示:KISSS-1蛋白和mRNA在转基因组的表达分别为1.143±0.218和0.877±0.162,均显著高于转空质粒组(0.745±0.130,0.685±0.128;t=3.850,2.481,P均<0.05)和对照组EC-1细胞(0.855±0.184,0.677±0.138;t=2.275,2.306,P均<0.05);Boyden小室检测细胞体外侵袭力实验发现转基因组在培养24、48和72 h后的穿膜细胞数分别为91.8±11.7,117.8±11.1和139.2±11.8,均显著低于转空质粒组(118.1±14.7,141.7±13.2,162.2±22.7;t=3.153,4.215,3.569,P值均<0.01)和对照组EC-1细胞(112.2±15.6,138.1±13.0,162.3±14.0;t=4.154,3.797,2.702,P值均<0.05).MTT检测结果显示,转基因组细胞在培养48 h和72 h后的增殖能力分别为0.517±0.127和0.394±0.137,与转空质粒组(0.636±0.186,0.513±0.150;t=2.054,2.709,P值均<0.05)和对照组(0.646±0.135,0.511±0.153;t=2.276,2.205,P值均<0.05)相比,细胞生长受到明显抑制;克隆形成实验结果显示,转基因组细胞的克隆形成数为157.2±36.4,明显低于转空质粒组(236.3±78.1;t=3.441,P<0.01)和对照组(242.5±48.6;t=2.250,P<0.05).结论 KISS-1基因在食管癌细胞株EC-1中可抑制细胞的体外侵袭能力及细胞的增殖能力.

abstractsObjective To investigate the effect of KISS-1 expression on the potential of invasion and proliferation of esophageal squamous carcinoma cell EC-1. Methods Protein and mRNA expressions of KISS-1 were evaluated by Western blot and RT-PCR in four esophageal carcinoma cell lines (EC-1, Ecal09, EC9706 and TE-1). Using liposome-mediated transfection, an eukaryotic expression vector (pcDNA3.1-KISS-1) of KISS-1 gene was transfected into EC-1 cells. Boyden chamber model, MTT and clone formation assay were used to detect the potential of invasion and proliferation. Results Western blot and RT-PCR showed a baseline low level of expression of KISS-1 protein (0.715±0.109) and mRNA (0.670±0.176) in EC-1 cells, pcDNA3.1-KISS-1 expression vector was successfully transfected into EC-1 cells. Western blot and RT-PCR showed that the expression of KISS-1 protein (1.143±0.218) and mRNA (0.877±0.162) in EC-1 cells transfected with pcDNA3.1-KISS-1 were significantly higher than those transfected with the control vector pcDNA3. 1 (0.745±0.130,0.685±0.128;t =3.850,2.481, P < 0.05) and the control cells (0.855±0.184,0.677±0.138;t = 2.275,2.306,P <0.05). Boyden chamber analysis showed that the invasiveness of the coils transfected with KISS-1 at 24 h (91.8±11.7), 48 h (117.8±11.1) and 72 h (139.2±11.8) were significantly reduced than that of the cells transfected with the control vector pcDNA3.1 (118.1±14.7, 141.7±13.2, 162.2±22.7;t = 3.153, 4.215, 3.569, P<0.01) and the control cells (112.2±15.6,138.1±13.0,162.3±14.0;t =4. 154,3.797, 2.702,P <0.05). MTT showed that the proliferation potential of ceils after transfection with KISS-1 at 48 h (0.517±0.127) and 72 h (0.394±0.137) were significantly reduced than that of cells transfected with the control vector pcDNA3.1(0.636±0.186, 0.513±0.150;t =2.054, 2.709, P <0.05) and the control ceils (0.646±0.135, 0.511±0.153;t =2.276, 2.205, P <0.05). Clone formation assay suggested that cells transfected with KISS-1 (157.2±36.4) showed significantly decreased clone formation than cells transfected with the control vector pcDNA3.1 (236.3±78.1;t= 3.441, P < 0.01) and the control cells (242.5±48.6;t =2.250,P <0.05). Conclusion KISS-1 gene inhibits the potential of invasion and proliferation of EC-1 cells.