Long-Term Effects of Mouse Intracytoplasmic Sperm Injection with DNA-Fragmented Sperm on Health and Behavior of Adult Offspring1

作者：Raúl Fernández‐González, Pedro Nuno Moreira, Míriam Pérez-Crespo, Manuel Adolfo Sánchez-Martin, Miguel Ángel Ramírez, Eva Pericuesta, Ainhoa Bilbao, Pablo Bermejo‐Álvarez, Juan de Dios Hourcade, Fernando Rodrı́guez de Fonseca, Alfonso Gutiérrez‐Adán · 发表于：Biology of Reproduction · 年份：2008 · DOI：10.1095/biolreprod.107.065623 · 被引用次数：372 · 研究领域：Reproductive Biology and Fertility、Prenatal Screening and Diagnostics、Assisted Reproductive Technology and Twin Pregnancy

Genetic and environmental factors produce different levels of DNA damage in spermatozoa. Usually, DNA-fragmented spermatozoa (DFS) are used with intracytoplasmic sperm injection (ICSI) treatments in human reproduction, and use of DFS is still a matter of concern. The purpose of the present study was to investigate the long-term consequences on development and behavior of mice generated by ICSI with DFS. Using CD1 and B6D2F1 mouse strains, oocytes were injected with fresh spermatozoa or with frozen-thawed spermatozoa without cryoprotector. This treatment increased the percentage of TUNEL-positive spermatozoa, tail length as measured by comet assay, and loss of telomeres as measured by quantitative PCR. The ICSI-generated embryos were cultured for 24 h in KSOM, and 2-cell embryos were transferred into CD1 females. The DFS reduced both the rate of preimplantation embryo development and number of offspring. Immunofluorescence staining with an antibody against 5-methylcytosine showed a delay of 2 h on the active demethylation of male pronucleus in the embryos produced by ICSI. Moreover, ICSI affected gene transcription and methylation of some epigenetically regulated genes like imprinting, X-linked genes, and retrotransposon genes. At 3 and 12 mo of age, ICSI with DFS-produced animals and in vivo-fertilized controls were submitted to behavioral tests: locomotor activity (open field), exploratory/anxiety behavior (elevated plus maze, open field), and spatial memory (free-choice exp...