干扰白纹伊蚊羽化激素对C6/36细胞糖代谢的影响

Effects of interfering eclosion hormone on carbohydrate metabolism in C6/36 cells of Aedes albopictus

目的观察干扰羽化激素对白纹伊蚊C6/36细胞糖代谢的影响。方法根据白纹伊蚊羽化激素基因设计RNAi干扰序列(EH-siRNA),转染白纹伊蚊C6/36细胞48 h后,用qRT-PCR检测细胞EH基因及糖代谢相关基因的转录水平,并检测细胞内糖原含量。结果 EH-siRNA干扰后,白纹伊蚊C6/36细胞EH基因的转录水平下降26.7%(P<0.05);糖酵解途径中葡萄糖-6-磷酸异构酶、磷酸甘油变位酶和烯醇酶基因的转录水平与阴性对照组相比,分别下降58.6%、55.4%和46.2%(均P<0.05),柠檬酸循环中的柠檬酸合成酶和异柠檬酸脱氢酶的转录水平与阴性对照组比较差异均无统计学意义(均P>0.05)。干预后白纹伊蚊C6/36细胞糖原含量为12.27 mg/10^(4)cell,与阴性对照组(21.38 mg/10^(4)cell)和空白对照组(27.66 mg/10^(4)cell)比较差异均有统计学意义(均P<0.05)。结论白纹伊蚊羽化激素对C6/36细胞的糖代谢起调节作用,羽化激素干扰可降低C6/36细胞糖酵解关键酶的表达以及糖原的含量。

Objective To investigate the effects of siRNA interference in expression of eclosion hormone(EH) on carbohydrate metabolism in C6/36 cells of Aedes albopictus. Methods An RNAi sequence(EH-siRNA) was designed in accordance with the eh gene of Aedes albopictus. EH-siRNA was transfected into C6/36 cells of Aedes albopictus. The level of transcription of eh and carbohydrate metabolism-related genes was determined using quantitative real-time PCR(qRT-PCR) and the intracellular glycogen content was determined with EH-siRNA 48 h post-transfection. Results After interference with EH-siRNA, the level of transcription of the eh gene in C6/36 cells decreased by 26.7%(t=5.998,P<0.05). The level of transcription of the gene coding for glucose-6-phosphate isomerase in the glycolytic pathway decreased by 58.6%, that of the gene coding for phosphoglycerate mutase decreased by 55.4%, and that of the gene coding for enolase decreased by 46.2%(t=4.351, 6.040, 4.884, P<0.05) compared to levels in the negative control group. In contrast, the level of transcription of the gene coding for citrate synthase and the gene coding for isocitrate dehydrogenase in the citrate cycle did not change post-interference(t=0.3814, 2.679, P>0.05). The glycogen level was 12.27 mg/104 cells in the interference group, which was significantly lower than that in the negative control group(21.38 mg/104 cells)(t=3.528, P<0.05) and the blank control group(27.66 mg/104 cells)(t=5.395, P<0.05). Conclusion The eclosion hormone may regulate carbohydrate metabolism in C6/36 cells of Aedes albopictus. Interference with EH-siRNA can decrease the expression of key glycolysis enzymes and glycogen content in C6/36 cells.