Dctn1在小鼠睾丸和精子中的定位及在变形过程中的功能初探

Location of Dctn1 in the mouse testis and sperm and its role in spermiogenesis

目的:探讨动力蛋白激活蛋白1(Dctn1)在小鼠精子变形过程中的作用。方法:通过Western印迹及间接免疫荧光技术分析Dctn1在小鼠睾丸及精子中的表达和定位。分别运用体外GC2-spd细胞系以及在体两种策略筛选出具有最高干扰效率的Dctn1的小干扰RNA(siRNA),进而应用睾丸网微注射技术,将混有示踪剂(0.4%台盼蓝)的Dctn1 siRNA注射至3周ICR小鼠的生精小管内,对照侧的睾丸注射阴性对照siRNA,正常组为不加任何处理的3周ICR小鼠。在注射后第3周取附睾尾部精子进行形态学分析。结果:Dctn1主要定位于精子的尾部。干涉后Dctn1 siRNA组的精子尾部畸形率为(23.57±0.55)%,明显高于对照组[(12.35±2.29)%,P<0.01,n=3],而正常组精子尾部畸形率为(3.37±0.69)%。结论:Dctn1在精子变形中可能发挥重要作用,它主要影响精子尾部的形成。

Objective： To investigate the role of dynactin 1（Dctn1） in the process of mouse spermiogenesis.Methods： Western blot and indirect immunofluorescence were used to analyze the expression and location of Dctn1 in the mouse testis and spermatozoa.The highest efficiency of small interference RNA（siRNA） was verified by GC2-spd cell line in vitro and in vivo studies,respectively.Dctn1 siRNA mixed with the indicator（0.4% trypan blue） was injected into the seminiferous tubules of 3-week-old ICR mice through rete testis microinjection,and negative control siRNA injected into the control testes.The normal group included 3-week-old ICR mice that did not receive any treatment.Spermatozoa were collected from the cauda epididymis 3 weeks after siRNA injection for morphological analysis.Results： Dctn1 was mainly localized in the tail of spermatozoa.After interference,the sperm tail abnormality in the Dctn1 siRNA group was（23.57±0.55）%,significantly higher than（12.35±2.29）% in the control（P0.01,n=3）,and it was（3.37±0.69）% in the normal group.Conclusion： Dctn1 plays an important role in mouse spermiogenesis,and mainly affects the formation of the tail of spermatozoa.