亚铁螯合酶抑制诱导细胞内原卟啉IX积聚的肿瘤荧光成像

Tumor fluorescence detection technique researches on accumulation of protoporphyrin-IX in cancer cells induced by FECH-siRNA

目的:探讨靶向FECH的siRNA在肿瘤荧光成像中的作用。方法:设计并合成靶向FECH的siRNA,用脂质体转染剂Lipofectamine2000携带后转染H08910PM细胞,用RT—PCR半定量检测细胞中FECHmRNA的表达用荧光显微镜和激光共聚焦显微镜在体外检测H08910PM细胞的荧光强度。结果:siRNA转染后H08910PM细胞的FECHmRNA的表达水平低于未转染组,与转染剂组比敲减效率为70.1%。荧光显微镜成像图显示:随氨基酮戊酸(ALA)浓度的增高,红色荧光逐渐增强;同一ALA浓度下siRNA干扰后,其红色荧光增强。结论:化学合成的靶向FECH的siRNA能够下调H08910PM细胞中FECH基因的表达,增加肿瘤细胞内的PplX积聚,siRNA与小浓度ALA联用具有协同效应,可减少ALA系统毒性,并提高肿瘤细胞的检出率。

Aim：To explore the utilization of inhibitor FECH-siRNA in cancer cells.Methods： A siRNA targeting FECH was designed and synthesized chemically,and then loaded with dye Lipofectamine 2000,transferred into HO8910PM cells.Semi-quantitative RT-PCR was used to examine the FECH mRNA expression.Fluorescence microscopy and confocal laser icroscopy were used to detect the fluorescence intensity in HO891OPM cells in vitro.Results： The expression of FECH mRNA in siRNA-transfected cells was significantly lower than that in non-transfected cells,with a knocking-down efficiency of 70.1%（vs the transfer agent group）.The intensity of red fluorescence from high level ALA groups is significantly higher than that from low level ALA groups under fluorescent microscope.After interference with siRNA in the same concentration of ALA,the intensity of red fluorescence in siRNA-cells is significantly higher.Conclusion：The chemically synthesized siRNA targeting FECH can effectively downregulate the FECH expression in HO891 OPM cells which further cause the decline of FECH,increase of PplX accumulation,and plays a synergic and additive effect on PpIX accumulation to reduce the of systemic toxicity of ALA and improve the detection of tumor cells.