新生小鼠睾丸组织异体异位移植物中精子形态及功能的研究

目的采用卵胞浆内单精子显微注射(ICSI)技术探讨新生小鼠睾丸组织异体异位移植到裸鼠体内后所生成精子的受精能力。方法将含有原始生精细胞的新生1-2 d小鼠睾丸组织异体异位移植到BALB/c-nu/nu免疫缺陷小鼠背部皮下。8周后取出移植物,通过常规染色和电镜技术对生精小管中生精细胞组成和形态进行观察;以薄层明胶水解法检测精子顶体蛋白酶活性;采用ISCI技术检测移植物中具有成熟形态精子的受精能力。结果移植物的HE染色发现,新生小鼠睾丸组织在裸鼠体内已启动并进行精子发生过程,生精小管中含有包括精子在内的所有类型生精细胞;移植物组织经处理后,可计数到具有镰刀头状的长尾精子约1×103个/mg移植物;透射电镜标本中观察到的精原细胞和精母细胞均具有正常超微结构;薄层明胶水解法检测可观察到顶体蛋白酶呈阳性反应的精子;ICSI结果显示,单纯ICSI处理组与ICSI+电激活组均有受精和卵裂现象发生。结论仅含有原始生精细胞的新生小鼠睾丸组织在异体异位移植到裸鼠体内后,可以产生形态正常并具有受精能力的精子。

Functional study of spermatozoa derived from neonatal mouse testes grafted into immunodeficient mice

Objective To investigate the fertilization ability of spermatozoa derived from neonatal mouse testes grafted into immunodeficient mice by intracytoplasmic sperm injection(ICSI).Methods Neonatal Kun-ming mouse testes containing primitive spermatogenic cells as the only germ cells were grafted under the back skin of castrated BALB/c-nu/nu mice.Grafts were taken out after 8 weeks.The initiation and restoration of spermatogenesis in grafts was documented by histological analyses.The ultrastructure of spermatogenic cells in grafts was observed using transmission electron microscopy.The activity of acrosomal enzyme of spermatozoa was analyzed with gelatin substrate test.The fertilization ability of spermatozoa was determined by using intracytoplasmic sperm injection.Results Histological analysis showed complete restoration of spermatogenesis in the grafts after 8 weeks.1×10~(3) spermatozoa with normal morphological appearance in 1mg suspended graft tissue were counted.Spermatogonia and spermatocytes with normal ultrastructure were observed.Gelatin substrate test positive spermatozoa were found implicating the activity of acrosomal enzyme of the spermatozoa.Cleavages were observed both in the ICSI group and the ICSI plus electric stimulation group.Conclusion Spermatozoa with normal morphology and fertilization ability can be developed by allografting neonatal mouse testes under the back skin of immunodeficient mice.