Development of evaluation method for spermatogenesis disorder using GFP mouse and analysis of mechanisms of reproduction disorder

为了明确己烯雌酚(DBS)的毒性作用靶点是生殖细胞还是支持生殖细胞的体细胞，在新生儿期DES暴露所致的生精障碍中，将GFP小鼠的生殖细胞移植到去自体生殖细胞的C57BL/6小鼠的生精小管内。首先，已证实新生小鼠暴露于0.5gGFP/只/只/天的DES 5天可引起μ小鼠和C57BL/6小鼠的生精功能障碍。当将暴露于DES的生殖细胞移植到未暴露于DES的无生殖细胞的小鼠时，精子发生正常发生。另一方面，当将正常生殖细胞移植到新生暴露于DES的无生殖细胞小鼠身上时，精子发生不正常，精子很少观察到。结果表明，DES的毒性作用靶点不是生殖细胞，而是体细胞。用ICR小鼠研究了新生小鼠DES(5μg/只/天)连续染毒5天对附睾腺的影响。DES可使雄激素受体蛋白表达延迟；雌激素受体α蛋白的表达范围扩大，沿附睾管的分布变窄；雌激素反应基因乳铁蛋白的表达区域近端变宽；附睾管间质和管腔增大，附睾管增厚。这些影响的机制将利用内部的小鼠附睾基因芯片进行分析。

In order to clarify which are the target of toxicity of diethylstilbestrol (DBS), germ cells or somatic cells supporting the germ cells, in the disorder of spermatogenesis caused by neonatal exposure to DES, germ cells of GFP mice were transplanted to seminiferous tubules of C57BL/6 mice which were free of own germ cells by Busulfan treatment. First, it was confirmed that neonatal exposure to 0.5 μg/mouse/day of DES for 5 days causes spermatogenesis disorder in GFP mice as well as C57BL/6 mice. When DES-exposed germ cells were transplanted to the germ cell-free mice that were not exposed to DES, spermatogenesis occurred normally. On the other hand, when normal germ cells were transplanted to the germ cell-free mice that were neonatally exposed to DES, spermatogenesis was not normal and spermatozoa were rarely observed. From these results, it was concluded that the target of DES toxicity is not germ cells but somatic cells.The effect of neonatal exposure to DES (5 μg/mouse/day) for 5 days on the epididymis was studied using ICR mice. By exposure to DES, (1) expression of androgen receptor protein was delayed ; (2) expression term of estrogen receptor α protein was widened and its distribution along the epididymal ductule was braodened ; (3) area of mRNA expression of lactoferrin, one of the estrogen-responsive genes, was proximally broadened ; and (4) various morphological changes, such as enlargement of interstitial space and lumen of epididymal tubule and thickening of the epithelium, occurred. Mechanisms of these effects will be analyzed using in-house mouse epididymis cDNA microarray.