Pairs of ovarian tissue were cultured for 3 h, and proteins were extracted using RIPA buffer containing a protease inhibitor cocktail (APExBIO, United States) and phosphatase inhibitors (Roche, Switzerland). Protein concentrations were determined by BCA protein assay (Beyotime Institute of Biotechnology, China). Equal amounts of protein lysates were separated on 12% PAGE Bis-Tris gels (Epizyme) and transferred onto nitrocellulose membranes. Proteins were visualized, and images were obtained using an Odyssey infrared imager.

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