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Updated: Jun 6, 2026
A Modified Co-Culture System for Understanding Granulosa-Theca Cell Interactions in the Bovine Ovary
Published on: September 19, 2025
AMPK/ULK1 enhances mitophagy and steroidogenesis in duck Granulosa cells under heat exposure
Xiaozhi Zhan1, Pei Luo1, Youtian Yang2
1College of Animal Science & Technology, Zhong Kai University of Agriculture and Engineering, Guangzhou, 510225, China; Guangdong Provincial Waterfowl Health Breeding Engineering Center, Guangzhou, 510225, China; Guangdong Engineering Technology Research Center of Biosafety and Intelligent Control for Aquatic Animals Diseases Guangzhou, 510225, China.
Abstract:
Mitochondria play crucial roles in energy metabolism and steroidogenesis. Heat stress suppresses steroidogenesis in ovarian granulosa cells (GCs) and impairs poultry reproduction. Our previous study revealed that mitophagy protects duck GCs from acute heat exposure. Notably, numerous studies have demonstrated that the AMP-activated protein kinase/Unc-51-like kinase 1 (AMPK/ULK1) pathway is a key regulatory pathway for mitophagy. However, how AMPK coordinates mitophagy and steroidogenesis under heat stress remains unclear. This study aimed to elucidate the role of the AMPK/ULK1 axis in heat-induced mitophagy and steroidogenesis. Using a duck heat-treatment model and in vitro GC culture models, we combined Western blotting, targeted metabolomics, siRNA-mediated gene knockdown, and mitochondrial function assays to assess the impacts of the AMPK/ULK1 axis on mitophagy and steroidogenesis. We showed that heat treatment induced mitochondrial dysfunction, activated the AMPK/ULK1 pathway, triggered mitophagy, and suppressed steroidogenesis in duck ovarian tissue. In vitro, GCs exhibited similar mitochondrial impairment and AMPK/ULK1-dependent mitophagic activation under heat treatment. The LC-MS/MS analysis also confirmed elevated intracellular AMP levels in heat treatment GCs. Furthermore, activation of AMPK by 5'-aminoimidazole-4´-carboxamide ribonucleotide (AICAR) triggered mitophagy and enhanced steroidogenesis in GCs under heat treatment by upregulating the expression of steroidogenic acute regulatory protein (StAR), cytochrome P450 11A1 (CYP11A1), and cytochrome P450 19A1 (CYP19A1), whereas inhibition of AMPK by compound C exerted the opposite effect. Additionally, genetic knockdown of AMPK or ULK1 via siRNA reduced mitophagy and steroidogenesis under heat treatment. Taken together, the AMPK/ULK1 axis mediates heat-induced mitophagy and enhances GC steroidogenesis, positioning AMPK as a therapeutic target to improve heat-stressed poultry reproduction.
