Three-Dimensional Hydrogel-Based Culture to Study the Effects of Toxicants on Ovarian Follicles

: Various toxicants, such as drugs and their metabolites, can cause potential ovarian toxicity. As the functional units of the ovary, ovarian follicles are susceptible to this type of damage at all developmental stages. Studying the effects of toxicants on ovarian follicles is an important task. Three-dimensional (3D) hydrogels, such as fibrin alginate interpenetrating networks (FA-IPNs), can support ovarian follicle culture in vitro for extended periods of time and serve as a suitable tool for studying ovotoxicity. Growing follicles encapsulated in the FA-IPN can proteolytically degrade the fibrin component in the FA-IPN. The degradation of fibrin mirrors the follicle growth and serves as a surrogate reporter for follicle health. The speed of fibrin degradation can be further controlled by aprotinin, a small molecule that inhibits plasmin-driven proteolytic degradation, which further expands the application of the described system. In this chapter, we describe methods to (1) isolate and encapsulate mouse ovarian follicles in FA-IPN, (2) follow follicle growth and development in vitro, and (3) evaluate the effects of toxicants on folliculogenesis using fibrin degradation.