AUTHOR=Qiu Lin , Xu Qi , Lin Yi , Wang Songxue , Zhang Huina , Liu Yi , Zhao Junzhao , Zhao Yingzheng , Xi Haitao 

TITLE=Optimizing in vitro maturation of oocytes with nitric oxide-loaded microbubbles

JOURNAL=Frontiers in Cell and Developmental Biology

VOLUME=Volume 13 - 2025

YEAR=2025

URL=https://www.frontiersin.org/journals/cell-and-developmental-biology/articles/10.3389/fcell.2025.1518226

DOI=10.3389/fcell.2025.1518226

ISSN=2296-634X

ABSTRACT=IntroductionNitric oxide (NO) plays a pivotal role in female reproductive processes, yet its clinical translation is limited by its short half-life and rapid systemic clearance. This study aimed to develop a stabilized NO delivery system to enhance oocyte maturation and quality for potential applications in assisted reproductive technologies (ART).MethodsLipid-based microbubbles (NO-MBs) were constructed via lyophilization to encapsulate and stabilize therapeutic NO gas. The effects of NO-MBs on in vitro oocyte maturation were evaluated using immature mouse oocytes, with conventional NO donors (e.g., SNP) as controls. Oocyte quality was assessed through Ca2+ levels, mitochondrial membrane potential (MMP), and intracellular reactive oxygen species (ROS). In vivo studies further examined oocyte retrieval, fertilization rates, and blastocyst cell apoptosis. Mechanistic investigations focused on the ERK signaling pathway.ResultsThe key parameters of the optimized lipid-based microbubbles for NO gas delivery were as follows: particle concentration (2.23 × 10⁷ MBs/mL) and diameter (0.919 ± 0.807 μm). NO-MBs demonstrated superior biosafety and efficacy compared to conventional NO donors. In vitro, NO-MBs significantly increased the maturation rate of immature mouse oocytes (72.00% vs. 57.77% in controls) while improving oocyte quality, as evidenced by elevated Ca2+ levels, enhanced MMP, and reduced ROS. In vivo, NO-MBs enhanced oocyte retrieval and fertilization rates and reduced blastocyst cell apoptosis. Mechanistically, NO-MBs uniquely activated phosphorylated ERK, suggesting ERK pathway involvement in oocyte maturation.DiscussionThese findings highlight NO-MBs as a novel, clinically relevant strategy for targeted NO delivery in reproductive medicine. By optimizing NO release and bioavailability, NO-MBs offer a promising approach to improve oocyte quality and developmental outcomes in ART. Further studies are warranted to explore their broader applications in reproductive health.