Gfi1.2基因在斑马鱼内耳的表达及顺铂的耳毒性研究

发布时间：2018-03-08 20:18

本文选题：斑马鱼　切入点：Gfi1.2　出处：《北京协和医学院》2009年博士论文　论文类型：学位论文

【摘要】：目的：建立以斑马鱼为模式动物的内耳损伤模型,研究Gfi1在斑马鱼内耳发育不同时间段的表达谱,以及耳毒性药物对内耳毛细胞损伤的发生机制。材料和方法：(1).选用发育至4.3hpf、12hpf、24hpf、48hpf的斑马鱼胚胎,运用原位杂交技术观察Gfi1.2基因在各时期胚胎中的表达情况。(2)选用48hpf的斑马鱼胚胎,分别用0.25mM.0.50mM、0.75mM、1.00mM、1.50mM的顺铂溶液培养8小时,用原位杂交技术观察Gfil.2在经过各种处理后胚胎中的表达情况。(3)选用48hpf的斑马鱼胚胎,用1.00mM的顺铂溶液培养4小时、8小时、12小时,用原位杂交技术观察Gfi1.2在经过各种处理后胚胎中的表达情况。结果：(1)Gfi1.2基因在发育至4.3hpf、12hpf、24hpf、48hpf的斑马鱼胚胎中均有明确表达,在48hpf的胚胎的内耳中有明显的表达。(2)用不同浓度顺铂处理的斑马鱼胚胎经过原位杂交,发现用越高浓度处理的胚胎,Gfil.2的表达越弱。(3)用相同浓度的顺铂处理的斑马鱼胚胎,处理时间越长,Gfil.2的表达越弱。结论：(1)斑马鱼是一种新兴的研究脊椎动物发育的模式生物,由于它具有发育周期短、胚胎透明内耳易于观察、价格相对低廉等优点,因而成为一种研究耳聋基因及内耳发育的优秀模型。本次实验证明,用斑马鱼作为研究药物对内耳毛细胞损害的模型也是可行的。(2)Gfi1是一种转录抑制因子,它在小鼠内耳毛细胞发育的过程中扮演着重要的角色,实验证明它的同源基因Gfil.2在斑马鱼内耳中也有持续表达,而且可以作为毛细胞损害的标记物。(3)顺铂对内耳毛细胞的损害具有浓度依赖性和时间依赖性。
[Abstract]:Aim: to establish an animal model of inner ear injury in zebrafish, and to study the expression profile of Gfi1 at different stages of development of zebrafish inner ear and the mechanism of ototoxic drugs on the damage of inner ear hair cells. Materials and methods zebrafish embryos developed to 4.3hpfn12hpfhfhfhfan24hpfan48hpf were used to observe the expression of Gfi1.2 gene in different stages of embryos by in situ hybridization. The zebrafish embryos of 48hpf were cultured in 0.25mM.0.50mM0.75mM 0.75mMU 1.50mm cisplatin solution for 8 hours, respectively. In situ hybridization technique was used to observe the expression of Gfil.2 in the embryos treated with various treatments. The zebrafish embryos with 48hpf were cultured in 1.00mm cisplatin solution for 4 hours, 8 hours and 12 hours, respectively. The expression of Gfi1.2 in embryos treated with in situ hybridization was observed. Results the Gfi1.2 gene was expressed in all zebrafish embryos which developed to 4.3hpfhpfl12hpfan24hpfan48hpf, and obviously expressed in the inner ear of 48hpf embryos) zebrafish embryos treated with different concentrations of cisplatin were hybridized in situ, and the zebrafish embryos with different concentrations of cisplatin were hybridized in situ, and the expression of Gfi1.2 gene was observed in the inner ear of 48hpf embryos. It was found that the weaker the expression of Gfil.2 was in the embryos treated with higher concentration of Cisplatin, the weaker the expression of Gfil.2 in the zebrafish embryos treated with the same concentration of cisplatin was. Conclusion zebrafish is a new model organism for studying the development of vertebrates, because it has the advantages of short developmental cycle, easy observation of transparent inner ear of embryo, and relatively low price. Therefore, it has become an excellent model to study the deafness gene and the development of the inner ear. This experiment proved that it is feasible to use zebrafish as a drug to study the damage of hair cells in the inner ear. It plays an important role in the development of mouse inner ear hair cells, and it has been proved that its homologous gene Gfil.2 is also expressed continuously in zebrafish inner ear. Cisplatin can be used as a marker of hair cell damage in a concentration-and time-dependent manner.
【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2009
【分类号】：R764.3;R-332

【共引文献】