Antinori S, Versaci C, Dani G, Antinori M, Selman H A
RAPRUI, Day Hospital, Rome, Italy.
Hum Reprod. 1997 Mar;12(3):554-6. doi: 10.1093/humrep/12.3.554.
The effect of cryopreservation on the integrity and fertilizing capacity of round spermatids was studied in two azoospermic patients. In December 1995 the patients, both with maturation arrest of spermatogenesis, were submitted to testicular sperm extraction (TESE) after an extensive examination of their ejaculate. Only round spermatids were found after testicular biopsy. Some of the spermatids were isolated and used for a first injection, while the remainder of the preparation was cryopreserved for successive cycles. Because of the failure of the first attempt, 3 months later, the same two patients were submitted to a second one. The frozen preparation was thawed and examined to evaluate the integrity and the viability of surviving round spermatids. More than 70% of the thawed spermatids were viable for injection. Fifteen oocytes at metaphase II, retrieved from the patients' wives, were microinjected with thawed round spermatids. Eighteen hours after the injection, seven out of 15 oocytes showed normal fertilization, with the presence of two pronuclei. The zygotes were cultured to observe embryonic development. After 48 h, six cleaving embryos had developed to at least the two-cell stage, while one had arrested at the pronuclear stage. At 72 h, the cleaving embryos showed further development to the four- to six-cell stage. They were then transferred into the uterus. After 3 weeks a clinical pregnancy was established in one patient [beta-human chorionic gonadotrophin (beta HCG) concentration was 2100 UI]. At 16 weeks of gestation, chromosomic analysis was performed, which confirmed the presence of a fetus with normal karyotype. The pregnancy is ongoing.
在两名无精子症患者中研究了冷冻保存对圆形精子细胞完整性和受精能力的影响。1995年12月，两名精子发生成熟停滞的患者在对其精液进行全面检查后接受了睾丸精子提取（TESE）。睾丸活检后仅发现圆形精子细胞。分离出一些精子细胞用于首次注射，而制剂的其余部分则冷冻保存以备后续周期使用。由于首次尝试失败，3个月后，这两名患者又接受了第二次尝试。解冻冷冻制剂并检查以评估存活圆形精子细胞的完整性和活力。超过70%的解冻精子细胞可用于注射。从患者妻子体内获取15个处于中期II的卵母细胞，用解冻的圆形精子细胞进行显微注射。注射18小时后，15个卵母细胞中有7个显示正常受精，有两个原核。对受精卵进行培养以观察胚胎发育。48小时后，6个分裂胚胎至少发育到二细胞阶段，而1个在原核阶段停滞。72小时时，分裂胚胎进一步发育到四至六细胞阶段。然后将它们转移到子宫。3周后，一名患者确诊临床妊娠[β-人绒毛膜促性腺激素（β-HCG）浓度为2100 UI]。妊娠16周时进行染色体分析，证实存在一名核型正常的胎儿。妊娠仍在继续。
