重组人源白细胞介素-24联合重组人源核心蛋白聚糖对人类肝细胞癌HepG2细胞增殖抑制和凋亡诱导协同效应的研究

目的 观察外源基因重组人源白细胞介素（IL）-24（rhIL-24）联合重组人源核心蛋白聚糖（rhDCN）对人类肝细胞癌HepG2细胞的增殖、凋亡和细胞周期的影响.方法 采用脂质体瞬时转染法将pcDNA3.1（＋）-IL-24和pcDNA3.1（＋）-DCN质粒转染至HepG2细胞,48 h后倒置显微镜下观察细胞生长状况、形态学改变.分别在转染后24h、48 h和72 h,采用四甲基偶氮唑蓝（MTT）法观察IL-24和DCN单独及联合转染对HepG2细胞的增殖抑制效应.转染后48 h流式细胞术检测HepG2细胞凋亡情况,并进行细胞周期阻滞分析.结果 与对照组相比,转染目的基因48 h后显微镜下可见细胞生长不同程度地被抑制,并可见典型的凋亡细胞形态改变,以联合转染组改变更为明显.MTT结果显示转染后48 h和72 h,双基因联合转染组的抑制率分别是（31.88±6.57）％和（36.83±3.76）％,与对照组和单独转染组比较,差异均有统计学意义（P＜0.01）.流式细胞术结果显示,单独转染IL-24和DCN基因可诱导HepG2细胞出现不同程度凋亡,而双基因联合转染组细胞凋亡率可达（32.56±0.90）％,与空质粒转染组的（2.98±0.72）％、空白细胞对照组的（3.50±0.92）％、IL-24组的（20.01±1.08）％和DCN组的（22.20±0.91）％比较,差异均有统计学意义（P＜0.01）.细胞周期分析结果表明,与对照组相比,IL-24基因转染组的G2/M期和DCN基因转染组的G0/G1期细胞比例明显增高分别为（11.24±0.35）％、（77.93±0.67）％,而双基因联合转染组G0/G1期和G2/M期细胞比例同时增高,分别是（71.36±0.60）％和（10.39±0.67）％,差异有统计学意义（P＜0.01）.结论 联合转染IL-24和DCN基因可以协同发挥较单独应用更强的抑制HepG2细胞增殖和诱导HepG2细胞凋亡的作用.IL-24可使HepG2细胞发生G2/M期阻滞,DCN可使HepG2细胞发生G0/G1期阻滞,双基因联合使HepG2细胞同时发生G2/M期和G0/G1期阻滞,是IL-24和DCN对HepG2细

Study on the synergistic proliferation-inhibiting and apoptosis-inducing effects of recombinant human-derived interleukin-24 and recombinant human-derived decorin on human hepatocellular carcinoma cells HepG2

Objective To investigate the synergistic proliferation-inhibiting and apoptosis-inducing effects of recombinant human-derived interleukin-24 （rhIL-24） and recombinant human-derived decorin （rhDCN） on human hepatocellular carcinoma cells HepG2.Methods Cellular growth and morphological changes of HepG2 cells were observed under the inverted microscope at 48 h after being transiently transfected with pcDNA3.1 （＋）-IL-24 and pcDNA3.1 （＋）-DCN by Lipofectamine.The proliferation-inhibiting effects of IL-24 and DCN on HepG2 cells,respectively and jointly,were observed with MTT assay at 24 h,48 h and 72 h post-transfection.Apoptosis and cell-cycle of HepG2 cells were analyzed by flow cytometry at 48 h post-transfection.Results Compared to control groups,the cells of target gene groups presented typically changes of proliferation inhibition and apoptotic morphology,which occurred obviously in co-transfection group.The results of MTT assay showed that at 48 h and 72 h post-transfection,the profiferation-inhibiting rates in the group of cells co-transfected with IL-24 and DCN were （31.88±6.57） ％ and （36.83±3.76） ％,respectively,displaying significant difference with those of other groups （P ＜ 0.01）.The results of flow cytometry showed that IL-24 and DCN can induce HepG2 cells apoptosis to some extent.Compared to the early apoptosis rate of cells of control groups,plasmid （2.98±0.72） ％,blank cell （3.50±0.92） ％,IL-24 （20.01±1.08） ％ and DCN （22.20±0.91） ％,a statistically remarkable apoptosis rate,（32.56±0.90） ％,can be seen in the group of cells treated with IL-24 and DCN jointly （P ＜ 0.01）.The result of cell cycle analysis revealed that,compared to control groups,the proportion of cells was higher in the phase of G2/M in the IL-24 group （11.24±0.35） ％ and in the phase of G0/G1 in the DCN group （77.93±0.67） ％.The proportions of cells in the phases of G2/M increased to （71.36±0.60） ％ and that of G0/G1 statistically increased to （