A Case Study of Using Assisted Reproductive Technology to Rescue Genetically Modified Mice with Reproductive Disorder Phenotypes

doi:10.12300/j.issn.1674-5817.2024.107

Abstract

Abstract:

Objective The utilization of assisted reproductive technology to rescue genetically modified mouse strains with reproductive disorders provides a reference for improving techniques to preserve valuable experimental mouse strains. Methods In vitro fertilization-embryo transfer (IVF-ET) technology was performed on 28 strains of infertile male mice aged 9-18 months. Several indicators such as sperm density and sperm motility in infertile male mice were assessed to select the most viable sperm for IVF-ET experiments. Fertility rate, abnormal egg rate, and birth rate were recorded after the birth of the pups. An optimized ovarian transplantation procedure was applied to 12 strains of infertile female mice aged 8-18 months. 6-week-old female mice with the same genetic background were selected as recipients. One intact ovary was removed from each recipient mouse, and the contralateral oviduct was ligated. An ovary from a donor mouse was isolated and transplanted orthotopically into the side where the ovary had been removed in the recipient mouse. Twenty-one days post-surgery, recipient mice were co-housed with 8-week-old wild type male mice of the same genetic background for breeding. Data such as the pregnancy rate and live birth rate of the recipients were recorded after the birth of the pups. Results IVF-ET successfully rescued 28 mouse strains, with the oldest male mice being 18 months old. The success rate of the first round of IVF-ET experiments was 89.29% (25/28). The average fertility rate of IVF in infertile male mice was (51.01±14.97)%, the abnormal egg rate was (9.03±5.28)%, and the birth rate of offspring mice was (18.60±7.03)%. 39 out of 40 ovarian transplant recipient mice survived, with a pregnancy rate of 33.33% (13/39) for ovarian transplant recipients, and a live birth rate of 17.95% (7/39). Four mouse strains were successfully rescued using optimized ovarian transplantation technology, with the oldest female mice being 18 months old. 8 strains were not rescued as they failed to produce offspring that survived to sexual maturity. Conclusion IVF-ET is an effective approach for rescuing mice with reproductive disorders caused by different reasons, especially for those beyond the optimal breeding age. Ovarian transplantation technology can also be used as an alternative for aged female mice. But its success rate is relatively lower than that of IVF-ET, and carries a higher experimental risk.

Key words: Genetically modified mouse, Middle-aged and aged, Reproductive disorders, In vitro fertilization-embryo transfer, Ovarian transplantation

CLC Number:

Q95-33

WANG Qianqian,TAO Sijue,WEI Zhen,et al. A Case Study of Using Assisted Reproductive Technology to Rescue Genetically Modified Mice with Reproductive Disorder Phenotypes[J]. Laboratory Animal and Comparative Medicine, 2025, 45(1): 79-86. DOI: 10.12300/j.issn.1674-5817.2024.107.

Figures/Tables 5

Figure 1 Sperm masses collected from male mice and sperm status after 1 h of capacitationNote：A， The unilateral cauda epididymidis and a segment of the vas deferens from a male mouse； B， Sperm masses collected from the cauda epididymidis placed into droplets of sperm capacitation medium； C， The sperm status of two different strains of male mice after 1 h of capacitation： in the left culture droplet， the sperm were sparse， while in the right culture droplet， they were evenly dispersed.

Figure 2 Ovarian transplantation surgery in miceNote： A， Ligation was performed at two sites on the oviduct of one side of the recipient mouse； B， An intact ovary from one side of the donor mouse was harvested； C， The ovary and oviduct on the other side of the recipient were stabilized； D， The intact ovary from the donor was transplanted into the ovarian capsule of the recipient at the site where the ovary had been removed， followed by suturing of the capsule.

Table 1 Abnormal egg rate, fertility rate and birth rate of infertile genetically modified mice rescued by in vitro fertilization-embryo transfer

不育雄性小鼠月龄/月 Age of Infertile male mice/month	动物数量 n	异形卵率/% Abnormal egg rate/%	受精率/% Fertility rate/%	仔鼠出生率/% Birth rate/%
9	3	4.35±2.21	48.99±28.09	19.87±13.93
10	4	11.55±8.29	62.34±11.32	16.84±5.43
11	4	9.38±2.35	60.85±10.46	17.41±10.46
12	4	7.14±1.12	48.87±13.72	19.04±5.31
13	4	9.50±6.44	42.21±5.86	19.68±1.92
14	4	13.15±7.72	53.07±12.79	16.51±4.60
15	1	6.13	36.79	12.82
16	1	5.04	43.17	15.00
17	1	9.09	47.20	20.00
18	2	8.31±3.53	42.45±27.12	27.66±10.50

Figure 3 Comparison between the young group and the middle-aged and aged group in their fertility rates， abnormal egg rates， and birth ratesNote： A， Comparison of in vitro fertilization （IVF） fertilization rates between the young group and the middle-aged and aged group； B， Comparison of IVF abnormal egg rates between the young group and the middle-aged and aged group； C， Comparison of IVF-embryo transfer （ET） birth rate between the young group and the middle-aged and aged group. *P<0.05， ***P<0.001； ns， differences were not statistically significant； D， Correlation between male mice age and fertility rate.

Table 2 Age of donor mice, pregnancy rate and live birth rate of infertile genetically modified mice successfully rescued through ovarian transplantation

供体雌性小鼠月龄/月 Age of donor mice/month	左侧卵巢移植的受体小鼠数/只 The number of recipients for donor’s left ovarian transplantation	右侧卵巢移植的受体小鼠数/只 The number of recipients for donor’s right ovarian transplantation	妊娠率/% Pregnancy rate/%	活产率/% Live birth rate/%
10	1	1	100.00 （2/2）	100.00 （2/2）
15	1	1	50.00 （1/2）	100.00 （1/1）
15	1	2	66.67 （2/3）	100.00 （2/2）
18	1	1	100.00 （2/2）	100.00 （2/2）

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