DNA methylation patterns of spermatozoa and two generations of offspring obtained after murine spermatogonial stem cell transplantation
DNA methylation patterns of spermatozoa and two generations of offspring obtained after murine spermatogonial stem cell transplantation
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DOI：
10.1093/humrep/dep213
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发表时间：
2009-09-01
影响因子：
6.1
通讯作者：
Tournaye, H.
中科院分区：
文献类型：
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作者：
Goossens, E.;De Rycke, M.;Tournaye, H.
除了在研究中的应用外，精原干细胞移植（SSCT）可能具有重要的临床应用。本研究采用小鼠睾丸细胞悬液移植于遗传性不育的WW受体小鼠睾丸，通过检测小鼠精子和活产后代中Igf 2、Peg 1和α-Actin的DNA甲基化水平，评价SSCT的安全性。将移植的雄性与可育雌性交配，检查它们的第一代和第二代后代，并与对照组在体重、长度和DNA甲基化模式方面进行比较。从移植后第一代和第二代后代的精子、肝脏、肾脏和胎盘中提取经亚硫酸氢钠处理的基因组DNA，进行PCR扩增以获得Igf 2、Peg 1和α-肌动蛋白基因片段。焦磷酸测序用于单独定量在CpG位点产生的人工C/T序列变异。第一代和第二代后代发育正常，其长度和重量与对照没有差异。同基因个体之间的SSCT与胎儿发育的变化无关，也与SSCT后两代后代精子或其他组织中Igf 2、Peg 1和α-肌动蛋白DNA甲基化模式的差异无关。
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.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.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.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.