摘要
[目的]观察基质金属蛋白酶组织抑制因子-1(TIMP-1)基因表达水平对人卵巢癌细胞生长的影响。[方法]采用质脂体法将正、反义TIMP-1表达载体体外转染到卵巢癌A2780细胞中,用G418筛选稳定表达外源基因的克隆并用RT-PCR鉴定后扩大培养。以未转染组细胞作为对照,分别用直接计数法、四氮唑蓝(MTT)比色法、平皿克隆形成试验测定3组细胞的增殖活性,利用流式细胞术检测肿瘤细胞周期与凋亡的变化。[结果]与对照组相比,转染TIMP-1对卵巢癌细胞A2780呈现增殖抑制效应:生长曲线示增殖速度减慢(P<0.01);在平皿上的集落形成能力下降(38±3.05)%;MTT法结果显示增殖抑制率增加(P<0.01);细胞周期显示G0/G1期比例增加(58.41±0.94)%。转染反义TIMP-1对卵巢癌细胞A2780呈现促增殖效应:生长曲线示增殖速度加快(P<0.01);在平皿上的集落形成能力增强(59±2.08)%;MTT显示增殖抑制率减小(P<0.01);细胞周期显示G0/G1期比例减小(44.82±0.31)%。[结论]体外转染TIMP-1基因提高TIMP-1的表达可抑制卵巢癌细胞的增殖,TIMP-1可能成为卵巢癌基因治疗的一个新靶点。
[ Objective] To observe the effects on growth of A2780 cells by TIMP - 1 gene transfection. [ Methods] Transferred sense and antisense TIMP- 1 into A2780 gramulosa carcinoma cells by liposomal transfection reagent. Then we screened stable expression sublines by G418 solution and further confirmed by RT - PCRo Then the subtypes of A2780 were marked as 2780 - PTs,2780 - Ptas and A - 2780, respectively. We also evaluated the proliferation activities in these three subtypes by direct cells count, MTT assay, plate clone formation test, respectively;The change of cell cycle were detected by FCM. [ Results] Compared with the normal A2780, 2780- PTs showed sush characteristics as followed: Partial inhibition of proliferation activities reflected by growth curves( P 〈 0.01 ) ; Decreasing clone formation on plate(38 ± 3.05)%;the proliferation inhibition rate increase revealed by MTT method result( P 〈 0.01 ); cell cycle reflected G0/G1 increase(58.41 ± 0.94)%. On the contrary, the 2780- PTas subline showed promoted proliferation, which was indicated as followed: Promoted proliferation activities under cells culture( P 〈 0.01 ) ; Increasing clone formation on plate(59 ± 2.08) % ;decreaseing proliferation inhibition rate revealed by MTT method result (P 〈 0.01 );cell cycle reflected Go/Gl decrease (44.82 ± 0.31)%. [Conclusion] TIM P- 1 transfection can inhibit the growth of A2780 gramulosa carcinoma cell. ThusTIMP- 1 can be a novel target for gramulosa carcinoma gene therapy.
出处
《大连医科大学学报》
CAS
2006年第4期273-275,295,共4页
Journal of Dalian Medical University
基金
国家自然科学基金资助(30370620)
关键词
TIMP-1
卵巢癌
基因转染
TIMP - 1
gramulosa carcinoma
gene transfection