Department of Reproductive Biology-CECOS, Rouen University Hospital, EA 4308 "Gametogenesis and Gamete Quality", UNIROUEN, Normandie Univ, 22 Boulevard Gambetta, 76000, Rouen, France.
Ann Biomed Eng. 2021 Jan;49(1):203-218. doi: 10.1007/s10439-020-02535-8. Epub 2020 May 21.
Testicular tissue freezing before gonadotoxic treatments allows the preservation of fertility for children suffering from cancer. Recently, the testis organ culture method was presented as a relevant method to restore the fertility of these patients. However, the yield of spermatozoa production is low in the mouse model and no gamete has been obtained in vitro in the rat model. Here, we assess different cryopreservation protocols and culture conditions to improve the efficiency of in vitro maturation of rat prepubertal testes. Testes from male rats aged 5 or 8 days post-partum were cultured onto agarose gels of different percentages. After determining the best culture conditions, different cryopreservation protocols were assessed. Finally, testicular tissues were cultured with media of various compositions and analyzed at different time points. Our results show that the cryopreservation protocols allow the preservation of tissue architecture, cell proliferation, with no or moderate increase of cell death. In vitro spermatogenesis did not proceed beyond the pachytene spermatocyte stage. Only 2 of the 6 tested media allowed the survival of differentiated germ cells over the 45-day culture period. In conclusion, this study highlights the necessity to further improve the organ culture method before applying it into the clinics.
在性腺毒性治疗前对睾丸组织进行冷冻保存，可使癌症患儿保留生育能力。最近，睾丸组织器官培养方法被提出作为恢复这些患者生育能力的相关方法。然而，在小鼠模型中精子产生的产量较低，在大鼠模型中体外也没有获得配子。在这里，我们评估了不同的冷冻保存方案和培养条件，以提高未成年大鼠睾丸体外成熟的效率。来自产后 5 或 8 天的雄性大鼠的睾丸在不同百分比的琼脂糖凝胶上培养。确定最佳培养条件后，评估了不同的冷冻保存方案。最后，用不同组成的培养基培养睾丸组织，并在不同时间点进行分析。我们的结果表明，冷冻保存方案允许组织结构、细胞增殖的保存，细胞死亡没有或适度增加。体外精子发生没有超过粗线期精母细胞阶段进行。在 45 天的培养期间，只有 6 种测试培养基中的 2 种允许分化的生殖细胞存活。总之，本研究强调在将该器官培养方法应用于临床之前，有必要进一步改进该方法。
