人胎儿卵巢组织异种移植后卵母细胞的发育及成熟能力  

Capability of Oocyte Maturation in Human Cryopreserved Ovarian Tissue Following Xenografting

作　　者：晁岚[1] 姜爱芳[2] 邓晓惠[1] 于红玲[1] 甄军晖[3] 

机构地区：[1]山东大学齐鲁医院不孕不育诊疗中心,济南250012 [2]潍坊医学院妇产科教研室,山东潍坊261041 [3]山东大学齐鲁医院病理科,济南250012

出　　处：《中国医学科学院学报》2008年第5期583-588,I0004,共7页Acta Academiae Medicinae Sinicae

摘　　要：目的探讨人胎儿卵巢组织冷冻复苏、体外培养和异种移植后卵母细胞的发育和成熟情况。方法取5例胎龄20周引产的胎儿卵巢,采用丙二醇慢速冷冻保存,复苏后在组织培养条件下进行体外培养,6d后移植到免疫缺陷小鼠的肾被膜下,应用卵泡刺激激素作用23周后取出卵巢组织,机械法取出窦卵泡内的卵母细胞,结合体外成熟培养,观察卵子成熟情况,并通过移植前后组织学和增殖细胞核抗原(PCNA)表达变化观察卵泡发育情况。结果冷冻复苏组织和新鲜卵巢组织中各期卵泡的构成比之间差异无显著性(P>0.05);培养组织与新鲜组织和冻融组织相比,增殖期卵泡的比例显著增加(P<0.05);移植组织与新鲜组织、冻融组织和培养组织相比,增殖期卵泡比例亦显著增加(P<0.05)。各组织中原始卵泡均未见PCNA阳性表达,其表达最早出现于初级卵泡,新鲜组织和冻融组织中偶可见PCNA阳性表达,而培养组织和移植组织中可见明显的PCNA阳性表达。从移植卵巢的窦卵泡中获得未成熟卵子42个,经体外成熟培养有21.43%的卵子可发育到MII期。结论人胎儿卵巢组织能耐受冻融、组织培养和移植过程,其卵泡能够重新生长发育,恢复内分泌功能,且不影响卵母细胞的发育及成熟能力。Objective To investigate the development and maturation competence of oocytes retrieved from cryopreserved and transplanted human fetal ovarian tissue by techniques of tissue culture, inducing ovary, oocyte retrieval, and in vitro maturation （IVM）. Methods Fetal ovaries of 20 weeks were frozen-thawed and cultured for 6 days in vitro, then xenografted into kidney capsules of immunodeficient mice All mice were stimulated with follicle stimulating hormone every second day for 23 weeks, starting 1 week after grafting. Then oocytes were retrieved from antral follicles 13 hours after human chorionic gonadotrophin injection. IVM was performed to evaluate the maturation competence of the oocytes from ovarian grafts. Human fetal ovarian tissues were examined with histological and proliferating cell nuclear antigen （PCNA） evaluation. Results There was no difference between fresh ovarian tissues and frozen-thawed ovarian tissues in the percentage of follicles at dif-ferent growth stages （ P 〉 0.05 ）. The proportion of the primary follicles and preantral follicles in the cultured ovarian tissues was significantly larger than that of fresh ovarian tissues and frozen-thawed ovarian tissues （ P 〈 0.05 ）. The proportion of the primary follicles, preantral follicles, and antral follicles in the transplanted ovarian tissues was significantly higher than that of cultured ovarian tissues, fresh ovarian tissues, and frozen-thawed ovarian tissues （ P 〈 0.05 ）. No significant signals of PCNA in the primordial follicles in all ovarian tissues were observed. PCNA immunoreactivity first appeared in primary follicles. However, the obviously positive signals of PCNA were seen in the oocytes and/or the granular cells of cultured ovarian tissues and transplanted ovarian tissues. Oocytes from antral follicles were collected and matured in vitro, and 21.43 % of the oocytes reached to MII within 48 hours IVM. Conclusions Human ovarian follicles can survive and develop well after cryopreservation, tissue culture, and xen

关 键 词：胎儿卵巢组织 冷冻保存 组织培养 异种移植 增殖细胞核抗原 卵泡发育 卵母细胞 体外成熟培养 

分 类 号：R617[医药卫生—外科学]