In Vitro Growth of Human Follicles: Current and Future Perspectives

Abstract

Ovarian tissue cryopreservation is gaining importance as a successful method to restore fertility to girls and young women at high risk of sterility. However, there are concerns regarding the safety of transplantation after ovarian tissue cryopreservation due to the high risk of reintroducing cancer cells and causing disease recurrence. In these cases, the development of culture systems that support oocyte development from the primordial follicle stage is required. Notable achievements have been reached in human follicle in vitro growth in the past decade. Currently, systems for the in vitro culture of ovarian tissue are based on two-dimensional substrates that do not support the survival of follicles or recapitulate the mechanical heterogenicity in the mammalian ovary. Recognition of the importance of special arrangements between cells has spurred research in three-dimensional culture systems, and the provision of a precise culture system that maximizes the diffusion of nutrients and gases through the follicles has raised interest in advanced biomimetic models. The current review critically examines various culture systems employed for the in vitro development of follicles, with a particular focus on solutions utilizing Organ-on-a-Chip (OOC) technology. The emphasis on OOC technology underscores its role as a promising avenue in ensuring the successful cultivation and maintenance of follicular structures during the culture period.

Keywords: cancer; cryopreservation; fertility preservation; in vitro follicular growth; organ-on-chip; ovarian tissue; prepubertal; three-dimensional culture systems; two-dimensional culture systems.

Figure 1

Fertility preservation protocol in young women suffering from cancer. Created with BioRender.com.

Figure 2

In vivo folliculogenesis. Folliculogenesis can be separated into early folliculogenesis and late folliculogenesis. Early folliculogenesis is under paracrine control and is gonadotrophin-independent, whereas late folliculogenesis is under endocrine control and is gonadotrophin-dependent. Follicle and oocyte promedium sizes for human and mouse are mentioned, as well as time of appearance [13]. Created with BioRender.com.

Figure 3

Diagrammatic representation of a multi-step culture system to support IVG of oocytes from human primordial follicles through to maturation as described by McLaughlin et al. [28]. Created with BioRender.com.

Figure 4

Diagram illustrating the encapsulation of follicles and cells within degradable PEG hydrogels containing DexVS fibers and functionalized with ECM-sequestering BMB peptides. In the presence of BMB-functionalized Dextran-VS fibers, extracellular matrix (ECM) secreted by cells, including laminin and collagen, is sequestered along DexVS fibers. This facilitates cell adhesion and the organization of cells into tissue-like aggregates, which subsequently undergo restoration. Adapted from Nason-Tomaszewski et al. [52].

Figure 5

Ovary-on-a-chip. (1) A PDMS microfluidic device designed for ovarian follicle culture by Aziz et al. [86]. (a) Schematic representation of the device featuring 5 inlets and 1 outlet, containing a single chamber for follicle cultivation. (b) Real microfluidic device. (2) A microfluidic platform for culturing individual follicles and ovarian explants developed by Zubizarreta and Xiao [81]. (a) Each microfluidic platform comprises 4 replicates of a fluidic circuit, with each replicate containing three interconnected modules: the inlet media donor module (DO), tissue culture module (T), and outlet media acceptor module (AC). (b) The ovary-on-a-chip supported the extended culture of mouse ovarian explants, facilitating follicle development, oocyte maturation, ovulation, and luteinization. CL: corpus luteum. Scale bar: 300 μm. (c) The ovary-on-a-chip generated a 28-day menstrual cycle-like hormone secretion profile, encompassing both the follicular and luteal phases.