环丙沙星对小鼠原代肝细胞自噬的研究

发布时间：2018-05-30 02:45

  本文选题：自噬 + 环丙沙星　； 参考：《成都学院》2017年硕士论文

【摘要】：背景:研究发现自噬在治疗多种疾病中起着重要的作用,环丙沙星是亲溶酶体抗生素,溶酶体是自噬过程中的关键细胞器,环丙沙星可能通过干扰细胞自噬活性而影响肝细胞的生理或病理功能。目的:检测环丙沙星自噬活性,筛选出环丙沙星给药小鼠原代肝细胞的差异表达基因,富集环丙沙星小鼠功能基因以及信号通路,为进一步揭示环丙沙星给药后的体内代谢及分子机制研究提供理论依据。方法:17.0±2.0g昆明小鼠,饲养于SPF系统中,按照随机原则,分为实验组和对照组,实验组小鼠采用灌胃方式给环丙沙星,对照组予生理盐水,持续7天,于第7天通过胶原酶灌注法分离小鼠原代肝细胞,贴壁后换液,自噬溶酶体荧光探针试剂盒按照比例加入,拍摄荧光图片,Agilent小鼠全基因组表达谱芯片检测基因表达。结果:实验组及对照组都可见自噬荧光亮点,但实验组亮点较小且数量更少。按照Foldchange≥1.5或≤0.75,组间进行t-test分析,检验标准P0.05,筛选出实验组与对照组之间差异表达的基因458个,其中差异表达上调基因共有133个,差异表达下调基因有325个,并在GENECARDS输入关键词autophagy获得1971个与自噬相关的基因,将筛选的差异基因与1971个自噬相关的基因进行比对,本实验差异表达基因中自噬相关基因表达上调9个,而自噬相关基因表达下调27个,KEGG分析结果显示16个显著通路Pathway,包括:错配修复基因(Mismatch repair),粘多糖的降解(Glycosaminoglycan degradation),DNA复制(DNA replication),粘蛋白型O-聚糖的生物合成(Mucin type O-glycan biosynthesis),半乳糖代谢(Galactose metabolism),核苷酸切除修复(Nucleotide excision repair),同源重组(Homologous recombination),神经胶质瘤(Glioma),p53信号通路(p53 signaling pathway),鞘脂代谢(Sphingolipid metabolism),磷酸肌醇代谢(Inositol phosphate metabolism),肥厚性心肌病(HCM),蛋白质消化吸收(Protein digestion and absorption)。结论:小鼠原代肝细胞自噬活性被环丙沙星抑制,通过生物信息学分析发现环丙沙星可以显著影响小鼠肝脏基因表达,差异基因富集主要集中在信号传导、糖代谢等。根据实验获得的信号通路,我们将在后期实验,进一步验证环丙沙星影响小鼠原代肝细胞自噬活性的主要信号通路以及证实差异基因之间的相互作用。
[Abstract]:Background: autophagy plays an important role in the treatment of many diseases. Ciprofloxacin is a lysophile antibiotic and lysosome is the key organelle in autophagy. Ciprofloxacin may affect the physiological or pathological function of hepatocytes by interfering with autophagy. Objective: to detect the autophagy activity of ciprofloxacin and to screen the differentially expressed genes in primary liver cells of ciprofloxacin treated mice, and to enrich the functional genes and signal pathways of ciprofloxacin mice. It provides theoretical basis for further study on metabolism and molecular mechanism of ciprofloxacin after administration of ciprofloxacin. Methods the mice were divided into experimental group and control group according to the random principle. The mice in the experimental group were given ciprofloxacin by gavage, and the control group was given normal saline for 7 days. On the 7th day, primary mouse hepatocytes were isolated by collagenase perfusion method, and then injected into the lysosomal fluorescence probe kit. The whole genome expression profile of Agilent mice was detected by fluorescence microarray. Results: both the experimental group and the control group showed fluorescent bright spots of autophagy, but the light spots of the experimental group were smaller and fewer. According to Foldchange 鈮，

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