Key Laboratory of Animal Reproduction and Germplasm Enhancement in Universities of Shandong, Qingdao Agricultural University, Qingdao, China.
Stem Cells Dev. 2013 Feb 15;22(4):567-80. doi: 10.1089/scd.2012.0436. Epub 2012 Nov 2.
The efficiency of in vitro culture systems for a premeiotic female germ cell is still low, mostly because of our incomplete understanding of the mechanisms controlling oogenesis and the obvious difficulties in reproducing the complex in vivo environment of such a process under in vitro conditions. Here we explored the possibility of recovering the developmental potential of mouse oocytes generated in vitro from premeiotic germ cells by transplantation under a kidney capsule of adult animals. To this aim, mouse embryonic ovaries of 12.5 days postcoitum cultured in vitro in a serum-free medium for 7 or 14 days, were transplanted beneath the kidney capsule of immunodeficient mice and analyzed after 21 (7+21 group) or 14 days (14+14 group). Cultured ovaries before transplantation showed delayed oocyte meiotic progression and follicle development. Interestingly, grafted ovaries of both groups, especially those of the 7+21 group, seemed able to restore the reproductive cycle of recipients. While the almost complete absence of primordial follicles was observed in grafted ovaries, oocytes from these ovaries showed transcript levels of genes associated to oocyte maturation similar to control. Moreover, the developmental stage of follicles and oocytes of the 7+21 group ovaries were comparable to that of 21 days post partum in vivo ovaries, whereas significant developmental delay were found in the 14+14 group ovaries. Nevertheless, oocytes retrieved from transplanted ovaries of both groups matured (around 80%) and were fertilized in vitro (around 20%-45%). Two-cell embryos from the fertilized oocytes developed to hatching blastocysts (about 50%) or gave rise to healthy live offspring (from 6% to 10%) when transplanted in a host mother. In conclusion, our results indicate that premeiotic female germ cells cultured in vitro up to primordial/primary follicle stages preserve their capability to complete oogenesis and can be fertilized and generate live pups after transplantation into a suitable in vivo environment.
体外培养系统对原始生殖细胞的效率仍然较低，主要是因为我们对卵母细胞发生的机制了解不完整，并且在体外条件下明显难以复制该过程的复杂体内环境。在这里，我们探索了通过将体外培养的原始生殖细胞来源的小鼠卵母细胞移植到成年动物的肾包膜下，恢复其发育潜能的可能性。为此，我们将体外培养的 12.5 天龄合胞体小鼠胚胎卵巢在无血清培养基中培养 7 或 14 天，然后将其移植到免疫缺陷小鼠的肾包膜下，并在 21 天（7+21 组）或 14 天（14+14 组）后进行分析。移植前的培养卵巢显示出卵母细胞减数分裂进程和卵泡发育的延迟。有趣的是，两组移植的卵巢，尤其是 7+21 组的卵巢，似乎能够恢复受体的生殖周期。虽然在移植的卵巢中几乎完全没有原始卵泡，但这些卵巢的卵母细胞显示出与卵母细胞成熟相关的基因的转录水平与对照相似。此外，7+21 组卵巢的卵泡和卵母细胞的发育阶段与体内 21 天产后的卵巢相当，而 14+14 组卵巢则存在明显的发育延迟。尽管如此，从移植卵巢中取出的卵母细胞在体外成熟（约 80%）并受精（约 20%-45%）。来自受精卵母细胞的二细胞胚胎在移植到宿主母体内时发育为孵化囊胚（约 50%）或产生健康的活后代（6%-10%）。总之，我们的结果表明，体外培养至原始/初级卵泡阶段的原始生殖细胞仍然能够完成卵母细胞发生，并在移植到合适的体内环境后能够受精并产生活后代。
