DNA methylation patterns of spermatozoa and two generations of offspring obtained after murine spermatogonial stem cell transplantation.

摘要

BACKGROUND: Apart from its use in research, spermatogonial stem cell transplantation (SSCT) may have important clinical applications. This controlled study aimed at evaluating the safety of SSCT by analyzing the DNA methylation pattern of Igf2, Peg1 and alpha-Actin both in spermatozoa and live born offspring obtained after SSCT in mice. METHODS: Testicular cell suspensions were transplanted to the testes of genetically sterile WW recipients. Transplanted males were mated with fertile females and their first and second generation offspring were examined and compared with controls with respect to weight, length and DNA methylation patterns. Sodium-bisulfite treated genomic DNA extracted from post-transplantation spermatozoa, liver, kidney and placenta of first and second generation offspring was PCR-amplified to obtain Igf2, Peg1 and alpha-Actin gene fragments. Pyrosequencing was used to individually quantify the resulting artificial C/T sequence variation at CpG sites. RESULTS: First and second generation offspring developed normally with their length and weight not being different from controls. Also the DNA methylation patterns of Igf2, Peg1 and alpha-Actin were not different among controls and first and second generation offspring after SSCT. CONCLUSIONS: SSCT between syngenic individuals was not associated with changes in fetal development nor with differences in the DNA methylation patterns of Igf2, Peg1 and alpha-Actin in spermatozoa or other tissues from two subsequent generations of offspring obtained after SSCT.