Discovery of eleven novel factors essential for mouse zygote development

Research Overview
A research team led by Associate Professor Hirofumi Nishizono and graduate student Masaki Kato from the Research Support Center at the Medical Research Institute of Kanazawa Medical University has identified eleven novel factors essential for the development of fertilized eggs. This achievement was made possible through the integration of one-cell embryo cryopreservation technology, an inhibitor library screening, RNA-seq analysis, and CRISPR-Cas9-mediated gene editing. The findings have been published in Frontiers in Cell and Developmental Biology on October 17, 2025.

Background
Infertility affects approximately one in six individuals worldwide, posing a major healthcare challenge in both developed and developing countries (Cox et al., Hum. Reprod. Open, 2022). To advance assisted reproductive technologies (ART), such as in vitro fertilization, it is crucial to identify and understand the molecular regulators governing fertilization, early embryonic development, and implantation (Sang et al., Science, 2023). While many factors involved in these processes have been discovered, the molecular network underlying early embryogenesis is still only partially understood.

Key Findings
The team prepared a large number of genetically uniform mouse zygotes and stably cryopreserved them using a single-cell embryo freezing technique. By performing a comprehensive screening of an inhibitor library targeting various signaling pathways and integrating the results with RNA-seq data, the researchers identified multiple candidate factors involved in embryonic development. Two molecules, CXC chemokine receptor 2 (CXCR2) and cathepsin D (CTSD), were selected for further functional validation.

CRISPR-Cas9 knockout experiments revealed that deleting either gene resulted in arrested embryo development, demonstrating their essential roles during the preimplantation stage. These findings suggest the existence of previously unrecognized regulatory factors in early development and indicate the potential for modulating embryonic development through precise molecular control. These advances could contribute to next-generation ART.

Future Perspectives
While these results are promising, several limitations remain. These include the potential effects of inhibitor concentration and the stress caused by embryo cryopreservation.

Future studies will aim to systematically address these factors and expand screening efforts using broader chemical libraries. Ultimately, this research aims to develop novel ARTs that enable molecular-level control of embryo development. This would open new avenues for infertility treatment and developmental biology research.

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Reference
DOI: 10.3389/fcell.2025.1643551

Funding
This work was supported by the Takeda Science Foundation (Grant No. 2022048848), JST CREST (Grant No. JPMJCR23B6), JSPS KAKENHI (Grant No. 25H00436), the Lotte Foundation, and the Grant for Promoted Research from Kanazawa Medical University (S2023-1).

About Kanazawa Medical University (KMU), Japan
Kanazawa Medical University (KMU) is a reputed medical school located in Ishikawa, Japan. Established in 1972 with Albert Schweitzer's idea of “Ehrfurcht vor dem Leben (respect for life) and five “S” keywords: Spirit, Science, Skill, Speed, and Safety, KMU is pledged to the development of high-quality medical professionals.

KMU continues to grow by engaging over 4,500 graduates from the School of Medicine in medical care globally. Furthermore, KMU sends over 2,500 nurses, public health nurses, and midwives to gain practical exposure in the medical field.

By following the motto and five “S” elements along with sustainable development goals, KMU strives to contribute to the creation of innovative medical science technologies and the supply of healthcare workforce, promoting holistic societal development.

Website:
https://www.kanazawa-med.ac.jp/English/public_html/