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. 2022 Jan 27;13(1):87. doi: 10.1038/s41419-022-04541-1

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© The Author(s) 2022

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 4 — a The timeline of the experiment was shown. Ovaries at 2 dpp were cultured in standard (control) or pyruvate-free (pyr-free) medium for 6 days, or cultured in pyruvate-free medium for 2 days and then in standard medium for 4 days (recovery). b, c Morphological comparison of the ovaries (b) and the number of primordial, growing (red arrows, c) and atretic follicles (yellow arrows, c) in the control, pyruvate-free and recovery groups. Nuclei were stained by hematoxylin. Scale bars: 50 μm. d DDX4 protein levels in the control, pyruvate-free, and recovery groups. e The mRNA levels of Pcna, Ki-67, Bax/Bcl-2, and Caspase-3 in the control, pyruvate-free and recovery groups. f The protein levels of PCNA, BAX, BCL-2, and Cleaved Caspase-3 in the control, pyruvate-free and recovery groups. g Immunofluorescence stain of PCNA, Ki-67, TUNEL and Cleaved Caspase-3 (green) in the control, pyruvate-free and recovery groups. DAPI, blue. Arrows show the cells with positive signals. Scale bars: 50 μm. h The percentage of granulosa cells with PCNA- and Ki-67-positive signals, and the number of cells with TUNEL- and Cleaved Caspase-3-positive signals in each section in the control, pyruvate-free and recovery groups. i The protein levels of GLUT4, HK1, PFKL, and PKM2 in the control, pyruvate-free and recovery groups. All the experiments were repeated three times, and the representative images are shown. In western blot results, β-actin was used as an internal control. Bars indicate the mean ± SD. *P < 0.05, **P < 0.01, and ***P < 0.001 vs. control.