METTL3-mediated m6A methylation regulates granulosa cell autophagy during follicular atresia in porcine ovaries (2023)

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Follicle isolation and GC collection

All experimental procedures were performed according to the guidelines for the care and use of laboratory animals and approved by the Animal Ethics Committee of Guangxi University. The pig ovaries were collected from a local slaughterhouse and returned to the laboratory in normal saline with high pressure treatment within 1 hour. The ovarian tissue was washed with 75% ethanol and cut with ophthalmic scissors. The single follicle with a diameter of 3-5 mm was separated and

Detection of autophagy in GCs from porcine follicles with different degrees of atresia

To examine the difference in GC autophagy of porcine follicles with different degrees of atresia, the GCs of the three groups were pooled. MDC staining, WB and TEM were implemented to detect autophagy. With increasing extent of atresia, the number of MDC-positive cells increased significantly and the LC3-II/LC3-I ratio showed an increasing trend. TEM showed that the GCs in the EAF and PAF groups had more autophagic vesicles than those in the HF group (Fig. 2).

Follicular atresia induced by GC autophagy

To check if GC can induce autophagy


Although the hypothalamic-pituitary-ovarian axis plays an important role in follicular atresia through endocrine regulation, more cellular processes such as autophagy are also thought to be involved in this regulation [2]. Autophagy is an adaptive response to maintain cell homeostasis and normal physiological activity by removing intracellular waste through a lysosomal mechanism [22]. However, excessive activation of autophagy can lead to programmed cell death [23]. ovules in follicles


The present study found that the degree of follicular atresia was positively correlated with the autophagy level of GCs and negatively with the m6A modification level of GCs. In addition, autophagy can induce follicular atresia. The m6A modification may regulate GC autophagy by altering theULK1transcription levels. Our study reveals a possible mechanism by which the m6A modification regulates follicular atresia, providing new insight into the mechanism of follicular atresia


This work was jointly supported by theNatural Science Foundation of China(grant number32160788);Natural Science Foundation of Guangxi Province(grant number2021GXNSFDA075001,2020GXNSFBA297090);Project to improve basic skills for junior and junior high school teachers in Guangxi Province(2020KY07004);Self-funded scientific research project of the Health Commission of the Guangxi Zhuang Autonomous Region in China(Z-A20220028);Young Scientist Research Award Fund of Guangxi People's Hospital

CRediT Authorship Contribution Statement

Zhengda Li:Conceptualization, Resources, Formal Analysis, Writing - Original Draft.Ziyun Ruan:Resources, Research, Methodology, Writing - Original Draft.Yun Feng:resources, methodology.Yanxin Wang:Resources.Jun Zhang:Investigation.Canqiang Lu:Formal Analysis.Deshun Shi:Conceptualization, resources, fundraising.Fenghua Lu:Conception, acquisition of funding, writing – review & editing.

declarations of competing interests

The authors declare no conflict of interest.

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