Animal Bioscience

Asian Australasian Association of Animal Production Societies (아세아태평양축산학회)
권호 표지
DOI QR코드

DOI QR Code

Protocatechuic acid promotes lactate synthesis in Sertoli cells of Tibetan sheep through AMPK/mTOR-mediated autophagy

  • Xingcai Qi (College of Animal Science and Technology, Gansu Agricultural University) ;
  • Yi Wu (College of Animal Science and Technology, Gansu Agricultural University) ;
  • Qiao Li (College of Animal Science and Technology, Gansu Agricultural University) ;
  • Huihui Wang (College of Animal Science and Technology, Gansu Agricultural University) ;
  • Youji Ma (College of Animal Science and Technology, Gansu Agricultural University)
  • Received : 2025.10.15
  • Accepted : 2026.02.08
  • Published : 2026.06.01
Download PDF
⟨ Previous Next ⟩

Abstract

Objective: This study aimed to investigate the effects of protocatechuic acid (PCA) on Sertoli cells (SCs) in Tibetan sheep, focusing on its impact on cell proliferation, lactate synthesis, and the underlying molecular mechanisms. Methods: Primary SCs were isolated from the testes of Tibetan sheep and subsequently cultured. The effects of PCA on the functionality of SCs were evaluated through in vitro experiments, with an additional investigation into its molecular mechanisms. In vivo and in vitro mouse models were employed to validate the regulatory mechanisms of PCA on male reproductive function and SCs. Results: PCA significantly upregulated the expression of proliferation and lactate synthesis-related genes in Tibetan sheep SCs, and increased the intracellular levels of pyruvate and lactate. Mechanistically, PCA activated the AMP-activated protein kinase (AMPK) pathway and inhibited the mechanistic target of rapamycin (mTOR) pathway, leading to the induction of autophagy in SCs. The use of autophagy inhibitors and AMPK inhibitors effectively blocked PCA-induced promotion of SCs proliferation and lactate synthesis. In mouse model studies, PCA demonstrated enhancements in sperm count, pregnancy rates, and litter size, while reduction in the rate of sperm abnormality. It also elevated activin A levels and decreased inhibin B levels in testicular tissues, increased serum follicle-stimulating hormone, luteinizing hormone, and testosterone levels, enhanced testicular antioxidant enzyme activities, and reduced malondialdehyde content. Furthermore, PCA upregulated the expression of proteins associated with proliferation and lactate synthesis in mouse testicular tissues and primary SCs, modulated the AMPK/mTOR pathway, and increased autophagic activity. Conclusion: PCA promotes proliferation and lactate synthesis in SCs of Tibetan sheep through AMPK/mTOR-mediated autophagy.

Keywords

Acknowledgement

This research was funded by the National Natural Science Foundation of China (32260833), Discipline Team Project of Gansu Agricultural University (GAU-XKTD-2022-20).

References

  1. Sha Y, Guo X, He Y, et al. Synergistic responses of Tibetan sheep rumen microbiota, metabolites, and the host to the plateau environment. Int J Mol Sci 2023;24:14856. https://doi.org/10.3390/ijms241914856
  2. Li C, Chen B, Langda S, et al. Multi-omic analyses shed light on the genetic control of high-altitude adaptation in sheep. Genom Proteomics Bioinform 2024;22:qzae030. https://doi.org/10.1093/gpbjnl/qzae030
  3. Zhang Y, Xue X, Liu Y, et al. Genome-wide comparative analyses reveal selection signatures underlying adaptation and production in Tibetan and Poll Dorset sheep. Sci Rep 2021; 11:2466. https://doi.org/10.1038/s41598-021-81932-y
  4. Li X, Han B, Liu D, et al. Whole-genome resequencing to investigate the genetic diversity and mechanisms of plateau adaptation in Tibetan sheep. J Anim Sci Biotechnol 2024;15:164. https://doi.org/10.1186/s40104-024-01125-1
  5. Wen Y, Ma X, Wang X, et al. hnRNPU in Sertoli cells cooperates with WT1 and is essential for testicular development by modulating transcriptional factors Sox8/9. Theranostics 2021;11:10030-46. https://doi.org/10.7150/thno.66819
  6. Boussouar F, Benahmed M. Lactate and energy metabolism in male germ cells. Trends Endocrinol Metab 2004;15:345-50. https://doi.org/10.1016/j.tem.2004.07.003
  7. Rato L, Alves MG, Socorro S, Duarte AI, Cavaco JE, Oliveira PF. Metabolic regulation is important for spermatogenesis. Nat Rev Urol 2012;9:330-8. https://doi.org/10.1038/nrurol.2012.77
  8. Zhang XN, Tao HP, Li S, et al. Ldha-dependent metabolic programs in Sertoli cells regulate spermiogenesis in mouse testis. Biology 2022;11:1791. https://doi.org/10.3390/biology11121791
  9. Courtens JL, Plöen L. Improvement of spermatogenesis in adult cryptorchid rat testis by intratesticular infusion of lactate. Biol Reprod 1999;61:154-61. https://doi.org/10.1095/biolreprod61.1.154
  10. Paul C, Rhind SM, Kyle CE, Scott H, McKinnell C, Sharpe RM. Cellular and hormonal disruption of fetal testis development in sheep reared on pasture treated with sewage sludge. Environ Health Perspect 2005;113:1580-7. https://doi.org/10.1289/ehp.8028
  11. Jiang L, Yang F, Liao H, et al. Molybdenum and cadmium cause blood–testis barrier dysfunction through ROS-mediated NLRP3 inflammasome activation in sheep. Sci Total Environ 2024;906:167267. https://doi.org/10.1016/j.scitotenv.2023.167267
  12. Li WH, Weng XX, Yuan LF, Li F, Yue XP, Li FD. Effect of feeding linseed diet on testis development, antioxidant capacity, and epididymal cauda sperm concentration in Chinese Hu lamb. Theriogenology 2021;159:69-76. https://doi.org/10.1016/j.theriogenology.2020.10.014
  13. Li W, Yao R, Xie L, et al. Dietary supplementation of grape seed tannin extract stimulated testis development, changed fatty acid profiles and increased testis antioxidant capacity in pre-puberty hu lambs. Theriogenology 2021;172:160-8. https://doi.org/10.1016/j.theriogenology.2021.06.015
  14. Li LJ, Chao S, Zhao SX, et al. Protocatechuic acid delays postovulatory oocyte ageing in mouse. Mol Nutr Food Res 2023;67:2200363. https://doi.org/10.1002/mnfr.202200363
  15. Cadena-Iñiguez J, Santiago-Osorio E, Sánchez-Flores N, et al. The cancer-protective potential of protocatechuic acid: a narrative review. Molecules 2024;29:1439. https://doi.org/10.3390/molecules29071439
  16. Adedara IA, Omole O, Okpara ES, et al. Impact of prepubertal exposure to dietary protocatechuic acid on the hypothalamic-pituitary-testicular axis in rats. Chem Biol Interact 2018;290:99-109. https://doi.org/10.1016/j.cbi.2018.05.013
  17. Hu R, Yang X, Gong J, et al. Patterns of alteration in boar semen quality from 9 to 37 months old and improvement by protocatechuic acid. J Anim Sci Biotechnol 2024;15:78. https://doi.org/10.1186/s40104-024-01031-6
  18. An X, Li T, Chen N, et al. miR-1285-3p targets TPI1 to regulate the glycolysis metabolism signaling pathway of Tibetan sheep Sertoli cells. PLOS ONE 2022;17:e0270364. https://doi.org/10.1371/journal.pone.0270364
  19. Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods 2001;25:402-8. https://doi.org/10.1006/meth.2001.1262
  20. Zhang S, Gai Z, Gui T, et al. Antioxidant effects of protocatechuic acid and protocatechuic aldehyde: old wine in a new bottle. Evid Based Complement Alternat Med 2021;2021:6139308. https://doi.org/10.1155/2021/6139308
  21. Liu R, Liu B, Tian L, et al. Induction of reproductive injury by bisphenol A and the protective effects of cyanidin-3-Oglucoside and protocatechuic acid in rats. Sci Total Environ 2023;883:163615. https://doi.org/10.1016/j.scitotenv.2023.163615
  22. Tao R, Miao L, Yu X, et al. Cynomorium songaricum Rupr demonstrates phytoestrogenic or phytoandrogenic like activities that attenuates benign prostatic hyperplasia via regulating steroid 5-α-reductase. J Ethnopharmacol 2019;235:65-74. https://doi.org/10.1016/j.jep.2019.01.038
  23. Adedara IA, Okpara ES, Busari EO, Omole O, Owumi SE, Farombi EO. Dietary protocatechuic acid abrogates male reproductive dysfunction in streptozotocin-induced diabetic rats via suppression of oxidative damage, inflammation and caspase-3 activity. Eur J Pharmacol 2019;849:30-42. https://doi.org/10.1016/j.ejphar.2019.01.033
  24. Owumi SE, Ochaoga SE, Odunola OA, Farombi EO. Protocatechuic acid inhibits testicular and epididymal toxicity associated with methotrexate in rats. Andrologia 2019;51:e13350. https://doi.org/10.1111/and.13350
  25. Namwanje M, Brown CW. Activins and inhibins: roles in development, physiology, and disease. Cold Spring Harb Perspect Biol 2016;8:a021881. https://doi.org/10.1101/cshperspect.a021881
  26. Marchetti C, Hamdane M, Mitchell V, et al. Immunolocalization of inhibin and activin α and βB subunits and expression of corresponding messenger RNAs in the human adult testis. Biol Reprod 2003;68:230-5. https://doi.org/10.1095/biolreprod.102.004424
  27. Burger LL, Haisenleder DJ, Aylor KW, Dalkin AC, Prendergast KA, Marshall JC. Regulation of luteinizing hormone-β and follicle-stimulating hormone (FSH)-β gene transcription by androgens: testosterone directly stimulates FSH-β transcription independent from its role on follistatin gene expression. Endocrinology 2004;145:71-8. https://doi.org/10.1210/en.2003-1047
  28. Wiater E, Lewis KA, Donaldson C, Vaughan J, Bilezikjian L, Vale W. Endogenous betaglycan is essential for high-potency inhibin antagonism in gonadotropes. Mol Endocrinol 2009; 23:1033-42. https://doi.org/10.1210/me.2009-0021
  29. Tian X, Teng J, Chen J. New insights regarding SNARE proteins in autophagosome-lysosome fusion. Autophagy 2021;17:2680-8. https://doi.org/10.1080/15548627.2020.1823124
  30. Wang M, Zeng L, Su P, Ma L, Zhang M, Zhang YZ. Autophagy: a multifaceted player in the fate of sperm. Hum Reprod Update 2022;28:200-31. https://doi.org/10.1093/humupd/dmab043
  31. Liu C, Wang H, Shang Y, et al. Autophagy is required for ectoplasmic specialization assembly in sertoli cells. Autophagy 2016;12:814-32. https://doi.org/10.1080/15548627.2016.1159377
  32. Ma Q, You X, Zhu K, et al. Changes in the tight junctions of the testis during aging: role of the p38 MAPK/MMP9 pathway and autophagy in Sertoli cells. Exp Gerontol 2022;161:111729. https://doi.org/10.1016/j.exger.2022.111729
  33. Yu T, Ye X, Deng F, et al. Protective effect of polysaccharides from Ostrea rivularis against reproductive injury via regulating autophagy induced by oxidative stress. Int J Biol Macromol 2022;209:655-67. https://doi.org/10.1016/j.ijbiomac.2022.04.053
  34. Wang H, Wang L, Hu F, et al. Neuregulin-4 attenuates diabetic cardiomyopathy by regulating autophagy via the AMPK/mTOR signalling pathway. Cardiovasc Diabetol 2022;21:205. https://doi.org/10.1186/s12933-022-01643-0
  35. Chen M, Zhang C, Wu Z, et al. Bta-miR-365-3p-targeted FK506-binding protein 5 participates in the AMPK/mTOR signaling pathway in the regulation of preadipocyte differentiation in cattle. Anim Biosci 2024;37:1156-67. https://doi.org/10.5713/ab.23.0328
  36. Guo H, Ouyang Y, Yin H, et al. Induction of autophagy via the ROS-dependent AMPK-mTOR pathway protects copperinduced spermatogenesis disorder. Redox Biol 2022;49:102227. https://doi.org/10.1016/j.redox.2021.102227
  37. Duan H, Yang S, Xiao L, et al. Melatonin promotes progesterone secretion in sheep luteal cells by regulating autophagy via the AMPK/mTOR pathway. Theriogenology 2024;214:342-51. https://doi.org/10.1016/j.theriogenology.2023.11.010
  38. Zhou Y, Yan J, Qiao L, et al. Bone marrow mesenchymal stem cell-derived exosomes ameliorate aging-induced BTB impairment in porcine testes by activating autophagy and inhibiting ROS/NLRP3 inflammasomes via the AMPK/mTOR signaling pathway. Antioxidants 2024;13:183. https://doi.org/10.3390/antiox13020183
  39. Eid AH, Gad AM, Fikry EM, Arab HH. Venlafaxine and carvedilol ameliorate testicular impairment and disrupted spermatogenesis in rheumatoid arthritis by targeting AMPK/ERK and PI3K/AKT/mTOR pathways. Toxicol Appl Pharmacol 2019;364:83-96. https://doi.org/10.1016/j.taap.2018.12.014
  40. Arab HH, Gad AM, Reda E, Yahia R, Eid AH. Activation of autophagy by sitagliptin attenuates cadmium-induced testicular impairment in rats: targeting AMPK/mTOR and Nrf2/HO-1 pathways. Life Sci 2021;269:119031. https://doi.org/10.1016/j.lfs.2021.119031