One-step automated bioprinting-based method for cumulus-oocyte complex microencapsulation for 3D in vitro maturation

Area of Science:

• Reproductive Biology
• Biotechnology
• Oocyte Maturation

Background:

• Traditional two-dimensional in vitro maturation (2D IVM) faces challenges with cumulus-oocyte complex (COC) flattening, potentially compromising oocyte quality.
• Existing methods for three-dimensional in vitro maturation (3D IVM) often lack reproducibility and extensive validation.
• Preserving the structural and functional integrity of COCs is crucial for efficient IVM.

Purpose of the Study:

• To optimize and validate a bioprinting-based process for generating COC-microbeads for 3D IVM.
• To compare the efficiency of 3D IVM using COC-microbeads against standard 2D IVM in a sheep model.
• To assess the impact of 3D IVM on oocyte nuclear maturation, bioenergetic status, and gene expression.

Main Methods:

• A bioprinting technique was developed to encapsulate COCs within alginate microbeads (COC-microbeads).
• Microbeads were characterized for size, shape, and stability under culture conditions.
• COC-microbeads and standard COCs were subjected to IVM, followed by analysis of nuclear chromatin, mitochondrial activity, reactive oxygen species (ROS), and specific gene expression (TFAM, ATP6, ATP8, KHDC3, NLRP5, OOEP, TLE6).

Main Results:

• The bioprinting method demonstrated high efficiency and reproducibility in producing COC-microbeads, preserving cumulus integrity.
• 3D IVM significantly improved oocyte nuclear maturation compared to 2D IVM (P<0.05).
• Ooplasmic mitochondrial activity and ROS generation increased in 3D IVM (P<0.05), with altered expression of key maturation and mitochondrial genes.

Conclusions:

• The developed bioprinting method for COC-microbeads is reproducible and efficient for 3D IVM.
• 3D IVM enhances oocyte nuclear and cytoplasmic maturation parameters, including mitochondrial function.
• This 3D IVM approach holds potential for clinical applications and toxicological studies in assisted reproduction.