Overexpression of mitofusin 2 ameliorates inflammation and oxidative stress in lipopolysaccharide-induced mastitis model by regulating phosphofurin acidic cluster sorting protein 2

Abstract

Background: Mastitis seriously affects the mammary health of humans and animals. Studies have found that inflammation and oxidative stress play key roles in the occurrence and development of mastitis. Therefore, in-depth research on related molecular mechanisms is of great significance.

Methods: Postpartum mice were anesthetized with pentobarbital and administered lipopolysaccharide to develop the mouse mastitis model. Proteomic analysis was performed to compare protein expression in mitochondria-associated endoplasmic reticulum membranes (MAM) from two mouse mammary gland groups. Western blot was used to detect the expression of MAM-related proteins in mitochondria. AlphaFold3 was used to predict the molecular structures of phosphofurin acidic cluster sorting protein 2 (PACS2) and mitofusin 2 (MFN2) and their interaction levels. The MFN2–PACS2 interaction was investigated using co-immunoprecipitation and small interfering RNA.

Results: The results showed that the inflammation level in the mammary gland tissue of mice with mastitis significantly increased, the total antioxidant capacity decreased, and the expression of MAM-related proteins MFN2 and PACS2 was significantly downregulated. In cell experiments, overexpression of MFN2 can inhibit inflammation and oxidative stress responses, and promote the interaction between MFN2 and PACS2 to affect the formation of MAMs.

Conclusion: In summary, this study suggests that mastitis can alter the expression of MAM–related proteins in mouse breast tissue. The interaction between MFN2 and PACS2 regulates the formation of MAMs. Overexpression of MFN2 can promote the formation of MAMs and inhibit inflammation and oxidative stress response in mammary epithelial cells. Our results provided a new theoretical basis and potential therapeutic targets for the prevention and treatment of mastitis.

Keywords

mastitis / mitochondria-associated endoplasmic reticulum membranes (MAM) / mitofusin 2 (MFN2) / phosphofurin acidic cluster sorting protein 2 (PACS2)

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2025 The Author(s). Animal Models and Experimental Medicine published by John Wiley & Sons Australia, Ltd on behalf of The Chinese Association for Laboratory Animal Sciences.