Tumor Fluorescence Detection Technique Researches On Accumulation Of Protoporphyrin-Ⅸ In Ovarian Cancer Cell Induced By Fech-sirna

AIMS:Fluorescence intensity of protoporphyrin-IX in human ovarian cancer HO8910PM cell were observed under fluorescent microscopy and confocal microscopy, studied the characteristics of accumulation of protoporphyrin-IX in HO8910 cell induced by FECH-siRNA and small dose 5-ALA, to explore the utilization potentiality of inhibitor-FECH-siRNA-based on early carcinoma diagnosis.METHODS:1. Ovarian cancer HO8910PM cell culture.2. The siRNA targeting to FECH was designed and synthesized chemically, and then with Lipofectamine 2000, transferred into HO8910PM cells.3. RT-PCR and quantitative PCR were used to examine the FECH mRNA expression.4. The experiment was divided 8 groups:①blank control group,②siRNA groups,③0.1mmol/L ALA group,④0.1 mmol/L ALA+siRNA groups,⑤0.5 mmol/L ALA groups,⑥0.5 mmol/L ALA+siRNA group,⑦1mmol/L ALA group,⑧1 mmol/L ALA+siRNA group.The intensity of red fluorescence in HO8910PM cell was observed under fluorescent microscopy and confocal microscopy.RESULTS:1. The expression of FECH mRNA in siRNA-cells significantly was lower than in no-siRNA-cells.2. The intensity of red fluorescence from high level ALA groups is significantly higher than that from low level ALA groups under fluorescent microscopy. In the same concentration of ALA, The intensity of red fluorescence in siRNA-cells is significantly higher.3.④0.1 mmol/L ALA+siRNA group is minimal group,which can efficiently observe red fluorescence.4.②group (135.0307±7.84902),④group (201.6248±7.36810),⑥group (361.8257±55.79851),⑧group (655.9286±72.31014) and①group (115.3773±4.53428),③(130.6456±1.54809),⑤group (259.9907±31.33794),⑦group (388.2075±64.03168),the difference of fluorescence content is significant(all P<0.01),Group①<Group③<Group②<Group④<Group⑤<Group⑥<Group⑦<Group⑧.CONCLUSIONS:1. The synthesized chemically specific siRNA targeting to FECH can effectively downregulate the FECH expression in HO8910PM cell and further lead to the decline of cell FECH, thus increase PpIX accumulation,and possesses the synergy and additive effect on PpIX accumulation.2. The minimum concentration of use of associating ALA with siRNA which red fluorescence was observed is 0.1mmol/L.