Takeuchi Takumi, Neri Queenie V, Katagiri Yukiko, Rosenwaks Zev, Palermo Gianpiero D
The Center for Reproductive Medicine and Infertility, Weill Medical College of Cornell University, New York, New York 10021, USA.
Biol Reprod. 2005 Mar;72(3):584-92. doi: 10.1095/biolreprod.104.032391. Epub 2004 Nov 3.
Germinal vesicle transplantation (GVT) has been proposed as a possible treatment to correct age-related oocyte aneuploidy caused by dysfunctional ooplasm. How healthy ooplasm regulates normal meiosis and subsequent development has yet to be elucidated, but impaired mitochondrial metabolism may be attributable to incomplete segregation of the oocyte chromosomes. In the present study, after ooplasmic mitochondrial damage by photoirradiating chloromethyl-X-rosamine, examination of the oocyte nuclei's ability to survive after transfer into healthy ooplasts was performed. To assess their fertilizability and potential for development, GVT oocytes were fertilized by intracytoplasmic sperm injection (ICSI) and transferred to foster mice. Condition of the offspring at birth was assessed, and epigenetic analysis was performed. Photosensitization consistently inhibited oocyte maturation. However, after GVT of photosensitized nuclei into healthy ooplasts, 67.2% were reconstituted, and 76.2% of these matured normally, with an overall rate of 51.2%, much higher than that (6.0%) in the mitochondrially injured oocytes. After ICSI, 65.8% (52/79) of GVT oocytes were fertilized normally, and 21.1% (11/52) eventually reached the blastocyst stage. The transfer of 132 two-cell GVT embryos into the oviducts of pseudopregnant females resulted in 17 apparently healthy live offspring. For some key developmental genes, a high level of expression was identified in the GVT and "rescue"-derived fetal adnexa. Thus, one can induce in oocyte mitochondria a photosensitization-based type of damage, which consistently inhibits GV breakdown, meiotic spindle formation, chromosomal segregation, and polar body extrusion. Germinal vesicle transplanted and rescued oocytes were able to undergo maturation, fertilization, and embryonic cleavage and, ultimately, to develop to term. This approach may provide a model with which to study the age-related ooplasmic dysfunction seen in human oocytes.
生发泡移植（GVT）已被提议作为一种可能的治疗方法，用于纠正由卵质功能障碍引起的与年龄相关的卵母细胞非整倍体。健康的卵质如何调节正常减数分裂及后续发育尚未阐明，但线粒体代谢受损可能归因于卵母细胞染色体的不完全分离。在本研究中，在用氯甲基-X-若丹明进行光照射造成卵质线粒体损伤后，对转移到健康卵质体中的卵母细胞核的存活能力进行了检测。为评估其受精能力和发育潜力，通过胞浆内精子注射（ICSI）使GVT卵母细胞受精，并将其转移到代孕小鼠体内。评估出生时后代的状况，并进行表观遗传分析。光致敏持续抑制卵母细胞成熟。然而，将光致敏的细胞核移植到健康卵质体中后，67.2%得到重构，其中76.2%正常成熟，总体率为51.2%，远高于线粒体受损卵母细胞的成熟率（6.0%）。ICSI后，65.8%（52/79）的GVT卵母细胞正常受精，21.1%（11/52）最终发育到囊胚阶段。将132个二细胞GVT胚胎转移到假孕雌性的输卵管中，产生了17只明显健康的活后代。对于一些关键的发育基因，在GVT和“挽救”衍生的胎儿附件中鉴定出高水平的表达。因此，可以在卵母细胞线粒体中诱导基于光致敏的损伤类型，这种损伤持续抑制GV破裂、减数分裂纺锤体形成、染色体分离和极体排出。生发泡移植和挽救的卵母细胞能够进行成熟、受精和胚胎分裂，并最终发育至足月。这种方法可能提供一个模型，用于研究人类卵母细胞中出现的与年龄相关的卵质功能障碍。
