# Effects of Insulin-like Growth Factor I and Follicular Fluid on In Vitro Growth of Cultured Oocytes

## TL;DR

This study shows that 50 ng/mL IGF-I improves the in vitro growth of porcine oocytes by reducing cell death, while follicular fluid had no beneficial effects.

## Contribution

The study identifies 50 ng/mL IGF-I as an effective concentration for promoting oocyte growth and reducing granulosa cell apoptosis in vitro.

## Key findings

- 50 ng/mL IGF-I significantly increased antrum-like structure formation and oocyte growth rate.
- 50 ng/mL IGF-I reduced granulosa cell apoptosis by modulating BAX and BCL-2 gene expression.
- PFF at tested concentrations failed to promote oocyte development and caused abnormal morphology.

## Abstract

In vitro culture of oocyte granulosa cell complexes (OGCs) is one approach to obtain large quantities of usable oocytes, supporting assisted reproductive technologies, yet the current technical system remains incomplete. This study aimed to explore whether insulin-like growth factor I (IGF-I) and porcine follicular fluid (PFF) could improve porcine OGCs in vitro culture efficiency. Results showed that 50 ng/mL IGF-I significantly increased OGCs’ antrum-like structure formation rate and oocyte growth rate; it also reduced granulosa cell apoptosis by decreasing pro-apoptotic gene BAX expression and increasing anti-apoptotic factor BCL-2 expression. In contrast, the different concentrations of PFF used in this study (2.5%, 5% and 10%, v/v) caused abnormal morphology of OGCs and failed to promote oocyte in vitro development. We conclude that 50 ng/mL IGF-I effectively promotes in vitro oocyte growth. This finding provides data support for optimizing reproductive technologies, obtaining large quantities of oocytes, and conserving germplasm resources of livestock or endangered animals.

Oocyte-granulosa cell complexes (OGCs) cultivation is crucial for advancing reproductive biotechnology but remains incomplete and needs further optimization. Insulin-like growth factor-I (IGF-I) regulates granulosa cell proliferation and apoptosis, and numerous studies have confirmed its role in promoting ovarian follicle development. Porcine follicular fluid (PFF) contains factors beneficial for oocyte growth, which may enhance oocyte development. To investigate whether IGF-I and PFF improve the in vitro culture efficiency of porcine OGCs, we cultured OGCs with IGF-I (0, 10, 50, 100 ng/mL) and PFF (from 3 to 6 mm follicles) at concentrations of 0, 2.5%, 5%, 10%, respectively. The results revealed that 50 and 100 ng/mL IGF-I significantly increased the antrum formation rate of OGCs (from 61.11 ± 7.35% to 88.89 ± 7.35%) and diameter growth of oocytes (from 108.77 ± 0.27 µm to 114.94 ± 0.58 and 113.29 ± 0.50 µm, respectively). However, only the 50 ng/mL group, but not the 100 ng/mL group, significantly improved the maturation rate (38.13 ± 3.77% vs. 25.00 ± 3.27%, p < 0.05) of oocytes. Additionally, 50 ng/mL IGF-I downregulated BAX (a pro-apoptotic gene) and upregulated BCL-2 (an anti-apoptotic factor) in granulosa cells, ultimately reducing apoptosis. In contrast, none of the PFF doses used in this study induced the formation of enclosed antrum-like structures in OGCs, nor did they significantly enhance their in vitro development. Our findings demonstrate that 50 ng/mL IGF-I effectively promotes the in vitro growth of porcine early antral follicle-derived OGCs by reducing apoptosis, whereas tested PFF concentrations had no beneficial effects and induced abnormal granulosa cell growth. How PFF modulates the adherent and spreading growth of granulosa cells has not been fully elucidated and requires further clarification.