Brca2^{(p.T1942fs/+)} dissipates ovarian reserve in rats through oxidative stress in follicular granulosa cells

Abstract

Pathogenic variants of BRCA1/2 constitute hereditary breast and ovarian cancer (HBOC) syndrome, and BRCA1/2 mutant is a risk for various cancers. Whereas the clinical guideline for HBOC patients has been organized for the therapy and prevention of cancer, there is no recommendation on the female reproductive discipline. Indeed, the role of BRCA1/2 pathogenic variants in ovarian reserve has not been established due to the deficiency of appropriate animal models. Here, we used a rat model of Brca2^{(p.T1942fs/+)} mutant of Sprague-Dawley strain with CRISPR-Cas9 editing to evaluate ovarian reserve in females. Fertility and ovarian follicles were evaluated and anti-Müllerian hormone (AMH) was measured at 8–32 weeks of age with a comparison between the wild-type and the mutant rats (MUT). MUT revealed a significantly smaller number of deliveries with fewer total pups. Furthermore, MUT showed a significant decrease in primordial follicles at 20 weeks and a low AMH level at 28 weeks. RNA-sequencing of the ovary at 10 weeks detected acceleration of the DNA damage repair pathway, which was accompanied by oxidative stress-induced DNA double-strand breaks, a decrease in PTEN, and an increase in mTOR in follicular granulosa cells. In conclusion, Brca2^{(p.T1942fs/+)} dissipates primordial follicles via early activation of granulosa cells through oxidative stress, leading to earlier termination of fertility.

Keywords:

• BRCA2
• pathogenic variant
• ovarian reserve
• follicular granulosa cell
• fertility

Acknowledgements

BioRender.com was used for Figure 9. The authors thank Nobuaki Misawa (Department of Pathology and Biological Responses, Nagoya University Graduate School of Medicine), Bayasula (Bell Research Center for Reproductive Health and Cancer, Nagoya University Graduate School of Medicine), and the Division for Medical Research Engineering, Nagoya University Graduate School of Medicine for excellent technical assistance.

Authors’ contributions

HT, YMo and ST conceived the design of the experiments and HT, YMo and YMa collected and analyzed the data. HT, YMo and ST wrote and organized the manuscript, and prepared the figures and the tables. TM contributed to the generation of Brca2^{(p.T1942fs/+)} rats. RS, TN and HK contributed to the data acquisition, discussion and revision of the manuscript.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Additional information

Funding

This work was supported in part by JST CREST (JPMJCR19H4) and JSPS Kakenhi (JP19H05462, JP20H05502 and JP16H06276 [AdAMS (Aa210038)]) to ST.