Autophagy and Apoptosis of Porcine Ovarian Granulosa Cells During Follicular Development

Yuxin Zheng,Bin Zhang,Xiaorong Tang,Zhongliang Jiang,Ning Liu,Shun Guo,Lizhu Ma

doi:10.3390/ani9121111

Abstract

Simple SummaryGranulosa cells (GCs) provide nutrients and information for oocytes in porcine follicles. Follicular atresia is closely related to both apoptosis and autophagy of granulosa cells in ovarian follicles; however, the follicular stages of granulosa cell apoptosis or autophagy during follicular development or atresia are poorly understood. We found that autophagy and apoptosis of GCs occurred in GCs from different size follicles during follicular development, and autophagy was mainly found in GCs of medium follicles, while apoptosis was mainly found in GCs of large follicles. These data provided some useful information to understand follicular atresia which is related to the fertility of sows.Follicular atresia is closely related to both apoptosis and autophagy of granulosa cells (GCs) in ovarian follicles. In the present study, GCs were isolated from pig ovaries in small, medium and large antral follicles, and the current results showed that the proliferation of GCs was higher in medium follicles, and lower in large follicles compared to small follicles. The Bax and Caspase 3 mRNA levels were significantly higher, but the ratio of Bcl-2/Bax was lower in GCs of large follicles. The marker genes of autophagy, Atg3, Atg7 and LC3 mRNA levels were higher in GCs from medium follicles. Apoptosis- and autophagy-related proteins had a similar expression pattern to the mRNA level. Our results showed that phosphorylated ERK (p-ERK) was activated in GCs of large follicles, while phosphorylated AKT (p-AKT) and phosphorylated mTOR (p-mTOR) were inhibited in GCs of medium follicles. Labeling of autophagic vesicles with 4’,6-diamidino-2-phenylindole (DAPI) and monodansylcadaverine (MDC) confirmed the results of gene transcription and protein expression in GCs of different size follicles. We conclude that autophagy and apoptosis of GCs occurred in different size follicles during follicular development, and autophagy was mainly found in GCs of medium follicles, while apoptosis was mainly found in GCs of large follicles.

Highlights

Many factors regulate the process of ovarian follicle development, which includes endocrine, paracrine and autocrine factors aimed to promote ovulation; only 1% or less of follicles in the ovary reach the preovulatory stage and about 99% of follicles undergo the degenerative processAnimals 2019, 9, 1111; doi:10.3390/ani9121111 www.mdpi.com/journal/animalsAnimals 2019, 9, 1111 called atresia [1]
Granulosa cell (GC) apoptosis is confirmed to be the main reason for follicular atresia [2], increasing evidence suggests that autophagy is induced in ovarian granulosa cells (GCs) during follicular atresia to promote apoptotic cell death by excessive self-digestion and degradation of essential cellular constituents [3,4]
A significantly higher level of phosphorylated Extracellular signal-regulated kinase (ERK) (p-ERK) was found in large follicles than those of small and medium follicles (Figure 7C). These results showed that the phosphorylation of mTOR and AKT might be involved in the activation of autophagy in GCs of medium follicles

Summary

Introduction

Many factors regulate the process of ovarian follicle development, which includes endocrine, paracrine and autocrine factors aimed to promote ovulation; only 1% or less of follicles in the ovary reach the preovulatory stage and about 99% of follicles undergo the degenerative processAnimals 2019, 9, 1111; doi:10.3390/ani9121111 www.mdpi.com/journal/animalsAnimals 2019, 9, 1111 called atresia [1]. Granulosa cell (GC) apoptosis is confirmed to be the main reason for follicular atresia [2], increasing evidence suggests that autophagy is induced in ovarian GCs during follicular atresia to promote apoptotic cell death by excessive self-digestion and degradation of essential cellular constituents [3,4]. Autophagy is a process of degradation and recycling of cellular constituents resulting in constitutive, dynamic, evolutionarily conserved catabolism to maintain cell survival under various conditions of starvation, hypoxia, and interruption of growth signaling. The results of many studies suggest that autophagy promotes cell death and can be triggered by various stimuli that induce apoptosis [7,8]. Autophagy may be involved in folliculogenesis, as granulosa cells are the primary site of apoptosis during follicle atresia [3]

Methods

Results

Discussion

Conclusion