Evaluation of an ovary‐on‐a‐chip in large mammalian models: Species specificity and influence of follicle isolation status

Abstract

The ability to grow oocytes from immature ovarian follicles in vitro has significant potential for fertility preservation; yet, it has proved challenging in large mammalian species due to the complex metabolic needs and long-term culture requirements. Currently, follicular incubations are based on a "static" system with manual exchange of medium. Despite the numerous advantages of conventional culturing approaches, recapitulating the native microenvironment and supporting the survival of ovarian follicles from large mammalian species still represent challenges. In this study, we utilized an innovative, dynamic microfluidic system to support the in vitro survival of domestic cat and dog follicles enclosed within the ovarian cortex or isolated from ovarian cortex. Results indicate both species-specific and tissue type-specific differences in response to microfluidic culture. Domestic cat but not dog ovarian cortical tissues maintained viability under flow similar to conventional agarose gel controls. Preantral stage isolated follicles from both species that grew most favourably in conventional alginate bead culture, but overall, there was no influence of culture system on expression of follicle development or oocyte health markers. This system represents an important exploration toward the development of an improved ovarian in vitro culture system of large mammalian species (e.g., cats and dogs), which has potential applications for fertility preservation, reproductive toxicology, and endangered mammal conservation efforts.

3D printed mold leachates in PDMS microfluidic devices

“…In all 'static' conditions, culture medium was refreshed every second day. Mammalian ovaries contain hundreds of thousands of immature or "primordial" follicles, comprising granulosa cells and the enclosed oocytes 46 . The majority of the oocytes from these primordial follicles will never be ovulated and the ability to rescue and grow these follicles in vitro is of extreme importance to rescue female fertility 46 .…”

“…Mammalian ovaries contain hundreds of thousands of immature or "primordial" follicles, comprising granulosa cells and the enclosed oocytes 46 . The majority of the oocytes from these primordial follicles will never be ovulated and the ability to rescue and grow these follicles in vitro is of extreme importance to rescue female fertility 46 . Ovarian tissues are sensitive to culture; specifically, nutrient availability is a problem for their survival and a significant reduction in follicle viability has been reported/observed when these tissues are statically cultured 45 .…”

“…Tissues were cultured for four days, after which the device was disassembled by removing the sealing tape, and ovarian tissues were fixed in 4% paraformaldehyde for evaluation of follicle density and morphology compared with fresh tissues that were fixed at the time of initial tissue collection from the same individual. Morphologically normal follicle counts were determined in 10 sections of a 300-µm thick tissue, as previously described 46 . The percentage of live follicles (number of morphologically normal primordial, transitional, primary and secondary follicles, divided by the total number of live and atretic follicles, standardized per mm 2 tissue), was compared among treatment groups.…”

Organs-On-Chip Models of the Female Reproductive System

“…Microfluidics allowed to overcome limitations of conventional ovarian tissue culture enabling the monitoring of follicle development over time and the continuous nutrient replenishment. However, no significant differences in terms of follicle morphology and growth were observed between the microfluidic and the control cultures and also among the different tested flow rates [32].…”

Bioengineering models of female reproduction

“…In addition to mouse ovarian tissues, the microfluidic culture has also been applied to large mammalian species. Nagashima et al used the microfluidic system to culture both individual follicles and ovarian cortex tissues isolated from domestic cats and dogs [54]. The dynamic fluidic flow significantly promoted the growth of both primordial follicles and preantral growing follicles, indicating a promising exploration toward the development and maturation of ovarian tissues or follicles of large mammalian species.…”