Impact of caffeic acid on the testicular damages in D-galactose induced aging model in mice

Document Type : Original Article


1 Department of Anatomical Sciences, School of Medicine, Ardabil University of Medical Sciences, Ardabil, Iran

2 School of Medicine, Ardabil University of Medical Sciences, Ardabil, Iran


Objective(s): Aging is a biological phenomenon that causes various disorders and diseases in body systems such as the reproductive system. One of the important factors in aging is oxidative stress, which facilitates the aging process through various mechanisms. The aim of this study is the investigation of  effects of caffeic acid on the testicular damages in D‑galactose induced aging model in mice. 
Materials and Methods: Forty male mice were randomly divided into 5 groups (n=8): 1) Control, 2) Sham, 3) Aging, 4) Aging + caffeic acid, and 5) Caffeic acid. Aging was induced through daily injection of D-Galactose (300 mg/kg, intraperitoneal) for 6 weeks. Caffeic acid (60 mg/kg, intraperitoneal) was injected daily for 6 weeks. One day after the last injection mice were killed and the testicle and epididymis were removed. Then, sperm parameters, factors of oxidative stress, and histopathological changes were evaluated. 
Results: The results showed that aging significantly decreased the count, motility, and viability of sperm, and increased abnormal sperm and sperm DNA fragmentation in contrast to the control group (P<0.05). In addition, MDA levels increased significantly in this group, and SOD, GPx, and TAC activity decreased (P<0.05). Histological studies also showed the destruction of seminiferous tubules, and Johnson’s score decreased (P<0.05). Caffeic acid  administration significantly improved the above disarrays (P<0.05). 
Conclusion: The results showed that caffeic acid reduces the adverse effects of aging on spermatogenesis in mice by reducing oxidative stress and increasing antioxidant defenses.


1. Hamza RZ, Al-Harbi MS, Al-Hazaa MA. Neurological alterations and testicular damages in aging induced by D-Galactose and neuro and testicular protective effects of combinations of chitosan nanoparticles, Resveratrol and Quercetin in male mice. Coatings 2021; 11: 435-453.  
2. Li L, Chen B, An T, Zhang H, Xia B, Li R, et al. Bazibushen alleviates altered testicular morphology and spermatogenesis and modulates sirt6/p53 and sirt6/nf-κb pathways in aging mice induced by d-galactose and nano2. J Ethnopharmacol 2021; 271:113810. 
3. Taba MY, Mohammadi S, Jalali M, Beheshti F, Attari SS. Effects of different doses of curcumin on testicular histopathology, apoptosis, and reproductive organs weight index in mice D-galactose-induced aging model. Comp Clin Path 2019; 28: 997-1002.
4. Liao C-H, Chen B-H, Chiang H-S, Chen C-W, Chen M-F, Ke C-C, et al. Optimizing a male reproductive aging mouse model by D-galactose injection. Int J Mol Sci 2016; 17: 98-107.
5. Johnson SL, Dunleavy J, Gemmell NJ, Nakagawa S. Consistent age-dependent declines in human semen quality: A systematic review and meta-analysis. Ageing Res Rev 2015; 19:22-33.
6. Brahem S, Mehdi M, Elghezal H, Saad A. The effects of male aging on semen quality, sperm DNA fragmentation and chromosomal abnormalities in an infertile population. J Assist Reprod Genet 2011; 28: 425-432.
7. Agarwal A, Virk G, Ong C, Du Plessis SS. Effect of oxidative stress on male reproduction. World J Mens Health 2014; 32: 1-17.
8. Ji M, Su X, Liu J, Zhao Y, Li Z, Xu X, et al. Comparison of naturally aging and D‑galactose induced aging model in beagle dogs. Exp Ther Med 2017; 14: 5881-5888. 
9. Al-Shahari EA, Alkhazendar AA, ElBealy ER, Alm-Eldeen AA. Quercetin modulates aging induced-testicular damage via increasing the antioxidant defense and the immunolocalization of the proliferating cell nuclear antigen in male rats. Pak J Zool 2021; 53: 1081-1083.
10. Mohammadi E, Mehri S, Bostan HB, Hosseinzadeh H. Protective effect of crocin against d-galactose-induced aging in mice. Avicenna J Phytomed 2018; 8: 14-23. 
11. Golmohammadi MG, Khoshdel F, Salimnejad R. Protective effect of resveratrol against bisphenol A-induced reproductive toxicity in male mice. Toxin Rev 2021.
12. Mazani M, Ojarudi M, Banaei S, Salimnejad R, Latifi M, Azizi H, et al. The protective effect of cinnamon and ginger hydro‐alcoholic extract on carbon tetrachloride‐induced testicular damage in rats. Andrologia 2020;52:e13651.
13. Gülçin İ. Antioxidant activity of caffeic acid (3, 4-dihydroxycinnamic acid). Toxicology 2006; 217: 213-220.
14. Mercantepe T, Unal D, Tümkaya L, Yazici ZA. Protective effects of amifostine, curcumin and caffeic acid phenethyl ester against cisplatin-induced testis tissue damage in rats. Exp Ther Med 2018; 15: 3404-3412.
15. El-Refaei MF, Abdallah EA. Protective effects of caffeic acid phenethyl ester on cadmium-induced testicular injury: A crucial role of antioxidant enzymes in male mice infertility. Heliyon 2021; 7:e06965.
16. Huyut Z, Alp HH, Yaman T, Keleş ÖF, Yener Z, Türkan F, et al. Comparison of the protective effects of curcumin and caffeic acid phenethyl ester against doxorubicin‐induced testicular toxicity. Andrologia 2021; 53: e13919.
17. Moghadam MD, Baghshani H, Azadi HG, Moosavi Z. Ameliorative Effects of Caffeic Acid Against Arsenic-Induced Testicular Injury in Mice. Biol Trace Elem Res 2021; 199:3772-3780.
18. Fernandes FDP, Menezes APF, de Sousa Neves JC, Fonteles AA, da Silva ATA, de Araújo Rodrigues P, et al. Caffeic acid protects mice from memory deficits induced by focal cerebral ischemia. Behav Pharmacol 2014; 25: 637-647.
19. Timar M, Banaei S, Mehraban Z, Salimnejad R, Golmohammadi MG. Protective effect of saponin on sperm DNA fragmentation of mice treated with cyclophosphamide. Andrologia 2021:e14336.
20. Salimnejad R, Soleimani Rad J, Mohammad Nejad D, Roshangar L. Effect of ghrelin on total antioxidant capacity, lipid peroxidation, sperm parameters and fertility in mice against oxidative damage caused by cyclophosphamide. Andrologia 2018; 50: e12883.
21. Ohkawa H, Ohishi N, Yagi K. Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction. Anal Biochem 1979; 95: 351-358.
22. Yousefi H, Ahmadiasl N, Salimnejad R, Bagheri E, Roshangar L, Alihemmati A. Effects of renal ischemia-reperfusion on biochemical factors and histopathological alterations in the liver of male rats. Physiology and Pharmacology 2019; 23: 44-50.
23. Ameli M, Moghimian M, Saeb F, Bashtani E, Shokoohi M, Salimnejad R, et al. The effect of clomiphene citrate and human chorionic gonadotropin on the expression of CatSper1, CatSper2, LHCGR, and SF1 genes, as well as the structural changes in testicular tissue of adult rats. Mol Reprod Dev 2019; 86: 738-748. 
24. Abdallah FB, Fetoui H, Zribi N, Fakhfakh F, Keskes L. Protective role of caffeic acid on lambda cyhalothrin-induced changes in sperm characteristics and testicular oxidative damage in rats. Toxicol Ind Health 2012; 28: 639-647.
25. Zhao H, Song L, Ma N, Liu C, Dun Y, Zhou Z, et al. The dynamic changes of Nrf2 mediated oxidative stress, DNA damage and base excision repair in testis of rats during aging. Exp Gerontol 2021; 152: 111460. 
26. Nakano T, Nakata H, Kadomoto S, Iwamoto H, Yaegashi H, Iijima M, et al. Three-dimensional morphological analysis of spermatogenesis in aged mouse testes. Sci Rep 2021; 11: 1-10.