Journal of Reproduction & Infertility

Journal of Reproduction & Infertility

Leptin Improves Sperm Cryopreservation via Antioxidant Defense

Authors
Paula Fontoura 1
Mariana Mello 2
Paulo Gallo-Sá 2
Maria Cecília Erthal-Martins 2
Maria Cecília Cardoso 2
Cristiane Ramos 1
1 University Hospital Pedro Ernesto, State University of Rio de Janeiro, Rio de Janeiro, Brazil
2 Vida Centro de Fertilildade da Rede D’Or Clinic, Rio de Janeiro, Brazil
Abstract
Background: Leptin and its receptor are present in spermatozoa; however, the role of leptin in sperm function is still controversial. Our present study aimed at demonstrating the effect of cryopreservation on sperm DNA fragmentation (DNAf) and investigating the possible effects of sperm capacitation techniques and leptin in vitro incubation on frozen-thawed sperm DNAf and oxidative stress. Methods: Samples of 45 normospermic men attending for infertility investigation at Vida Centro de Fertilidade, Rio de Janeiro, Brazil, were frozen and thawed with or without capacitation and leptin incubation prior to freezing. Sperm DNA fragmentation was evaluated by Sperm Chromatin Dispersion Assay before and after cryopreservation and oxidative stress parameters were measured by spectrophotometry with and without leptin incubation. Statistical analysis was performed using paired t test to compare DNAf between groups before and after freeze-thaw cycle, to compare groups before and after capacitation and leptin incubation and oxidative measurements before and after leptin incubation. Statistical significance was considered when p≤0.05. Results: Our results revealed a significant post-thaw rise in sperm DNAf compared with fresh samples (p=0.0003). Sperm DNAf was significantly reduced when sperm capacitation was performed before freezing, when compared to those frozen with no previous capacitation (p=0.01). The addition of leptin to capacitated sperm before freezing reduced DNAf (p<0.0001) and enhanced superoxide dismutase (p=0.001) and glutathione peroxidase (p=0.02) antioxidant enzymes activity. Conclusion: The addition of leptin to capacitated sperm can improve sperm DNA quality following cryopreservation, possibly by inducing the activity of certain antioxidant enzymes.
Keywords
DNA Fragmentation
freeze
thaw cycle
leptin
oxidative stress
Sperm
  1. Petyim S, Neungton C, Thanaboonyawat I, Laokirkkiat P, Choavaratana R. Sperm preparation before freezing improves sperm motility and reduces apoptosis in post-freezing-thawing sperm compared with post-thawing sperm preparation. J Assist Reprod Genet. 2014;31(12):1673-80.
  2. Sieme H, Oldenhof H, Wolkers WF. Sperm membrane behaviour during cooling and cryopreservation. Reprod Domest Anim. 2015;50 Suppl 3:20-6.
  3. Yeste M, Estrada E, Rocha LG, Marin H, Rodriguez-Gil JE, Miro J. Cryotolerance of stallion spermatozoa is related to ROS production and mitochondrial membrane potential rather than to the integrity of sperm nucleus. Andrology. 2015;3(2):395-407.
  4. Di Santo M, Tarozzi N, Nadalini M, Borini A. Human Sperm Cryopreservation: Update on Techniques, Effect on DNA Integrity, and Implications for ART. Adv Urol. 2012;2012:854837.
  5. Aitken RJ, Baker MA, De Iuliis GN, Nixon B. New insights into sperm physiology and pathology. Handb Exp Pharmacol. 2010;(198):99-115.
  6. Helfenstein F, Losdat S, Moller AP, Blount JD, Richner H. Sperm of colourful males are better protected against oxidative stress. Ecol Lett. 2010;13(2):213-22.
  7. Cocuzza M, Sikka SC, Athayde KS, Agarwal A. Clinical relevance of oxidative stress and sperm chromatin damage in male infertility: an evidence based analysis. Int Braz J Urol. 2007;33(5):603-21.
  8. Sakkas D, Alvarez JG. Sperm DNA fragmentation: mechanisms of origin, impact on reproductive outcome, and analysis. Fertil Steril. 2010;93(4):1027-36.
  9. Zhao J, Zhang Q, Wang Y, Li Y. Whether sperm deoxyribonucleic acid fragmentation has an effect on pregnancy and miscarriage after in vitro fertilization/intracytoplasmic sperm injection: a systematic review and meta-analysis. Fertil Steril. 2014;102(4):998-1005.
  10. Zhang W, Li F, Cao H, Li C, Du C, Yao L, et al. Protective effects of l-carnitine on astheno- and normozoospermic human semen samples during cryopreservation. Zygote. 2016;24(2):293-300.
  11. Mata-Campuzano M, Alvarez-Rodriguez M, Alvarez M, Tamayo-Canul J, Anel L, de Paz P, et al. Post-thawing quality and incubation resilience of cryopreserved ram spermatozoa are affected by antioxidant supplementation and choice of extender. Theriogenology. 2015;83(4):520-8.
  12. Garcia-Galiano D, Allen SJ, Elias CF. Role of the adipocyte-derived hormone leptin in reproductive control. Horm Mol Biol Clin Investig. 2014;19(3):141-9.
  13. Li HW, Chiu PC, Cheung MP, Yeung WS, O WS. Effect of leptin on motility, capacitation and acrosome reaction of human spermatozoa. Int J Androl. 2009;32(6):687-94.
  14. Jope T, Lammert A, Kratzsch J, Paasch U, Glander HJ. Leptin and leptin receptor in human seminal plasma and in human spermatozoa. Int J Androl. 2003;26(6):335-41.
  15. Aquila S, Giordano F, Guido C, Rago V, Carpino A. Nitric oxide involvement in the acrosome reaction triggered by leptin in pig sperm. Reprod Biol Endocrinol. 2011;9:133.
  16. Lampiao F, du Plessis SS. Insulin and leptin enhance human sperm motility, acrosome reaction and nitric oxide production. Asian J Androl. 2008;10(5):799-807.
  17. Haron MN, D'Souza UJ, Jaafar H, Zakaria R, Singh HJ. Exogenous leptin administration decreases sperm count and increases the fraction of abnormal sperm in adult rats. Fertil Steril. 2010;93(1):322-4.
  18. Solinas G. Leptin signalling coordinates lipid oxidation with thermogenesis and defence against oxidative stress. Clin Exp Pharmacol Physiol. 2010;37(10):953-4.
  19. Zheng J, Fang J, Yin YJ, Wang XC, Ren AJ, Bai J, et al. Leptin protects cardiomyocytes from serum-deprivation-induced apoptosis by increasing anti-oxidant defence. Clin Exp Pharmacol Physiol. 2010;37(10):955-62.
  20. Yamagishi SI, Edelstein D, Du XL, Kaneda Y, Guzman M, Brownlee M. Leptin induces mitochondrial superoxide production and monocyte chemoattractant protein-1 expression in aortic endothelial cells by increasing fatty acid oxidation via protein kinase A. J Biol Chem. 2001;276(27):25096-100.
  21. Feijo CM, Esteves SC. Diagnostic accuracy of sperm chromatin dispersion test to evaluate sperm deoxyribonucleic acid damage in men with unexplained infertility. Fertil Steril. 2014;101(1):58-63.
  22. Draper HH, Hadley M. Malondialdehyde determination as index of lipid peroxidation. Methods Enzymol. 1990;186:421-31.
  23. Ellman GL. Tissue sulfhydryl groups. Arch Biochem Biophys. 1959;82(1):70-7.
  24. Bannister JV, Calabrese L. Assays for superoxide dismutase. Methods Biochem Anal. 1987;32:279-312.
  25. Aebi H. Catalase in vitro. Methods Enzymol. 1984;105:121-6.
  26. Flohe L, Gunzler WA. Assays of glutathione peroxidase. Methods Enzymol. 1984;105:114-21.
  27. Paoli D, Lombardo F, Lenzi A, Gandini L. Sperm cryopreservation: effects on chromatin structure. Adv Exp Med Biol. 2014;791:137-50.
  28. Sharma R, Kattoor AJ, Ghulmiyyah J, Agarwal A. Effect of sperm storage and selection techniques on sperm parameters. Syst Biol Reprod Med. 2015;61(1):1-12.
  29. Meamar M, Zribi N, Cambi M, Tamburrino L, Marchiani S, Filimberti E, et al. Sperm DNA fragmentation induced by cryopreservation: new insights and effect of a natural extract from Opuntia ficus-indica. Fertil Steril. 2012;98(2):326-33.
  30. Tataru DA, Markova EV, Osadchuk LV, Sheina EV, Svetlakov AV. [The optimal conditions of storage of spermatozoa for analysis of DNA fragmentation]. Klin Lab Diagn. 2015;60(4):52-6. Russian.
  31. Vutyavanich T, Piromlertamorn W, Nunta S. Rapid freezing versus slow programmable freezing of human spermatozoa. Fertil Steril. 2010;93(6):1921-8.
  32. Ozkavukcu S, Erdemli E, Isik A, Oztuna D, Karahuseyinoglu S. Effects of cryopreservation on sperm parameters and ultrastructural morphology of human spermatozoa. J Assist Reprod Genet. 2008;25(8):403-11.
  33. Basu S, Pant M, Rachana R. Protective effect of Salacia oblonga against tobacco smoke-induced DNA damage and cellular changes in pancreatic β-cells. Pharm Biol. 2016;54(3):458-64.
  34. Olaciregui M, Luno V, Gonzalez N, De Blas I, Gil L. Freeze-dried dog sperm: Dynamics of DNA integrity. Cryobiology. 2015;71(2):286-90.
  35. Khodayari Naeini Z, Hassani Bafrani H, Nikzad H. Evaluation of ebselen supplementation on cryopreservation medium in human semen. Iran J Reprod Med. 2014;12(4):249-56.
  36. Ghorbani M, Amiri I, Khodadadi I, Fattahi A, Atabakhsh M, Tavilani H. Influence of BHT inclusion on post-thaw attributes of human semen. Syst Biol Reprod Med. 2015;61(1):57-61.
  37. Kalthur G, Adiga SK, Upadhya D, Rao S, Kumar P. Effect of cryopreservation on sperm DNA integrity in patients with teratospermia. Fertil Steril. 2008;89(6):1723-7.
  38. Saritha KR, Bongso A. Comparative evaluation of fresh and washed human sperm cryopreserved in vapor and liquid phases of liquid nitrogen. J Androl. 2001;22(5):857-62.
  39. Martínez-Soto JC, Landeras J, Gadea J. Spermatozoa and seminal plasma fatty acids as predictors of cryopreservation success. Andrology. 2013;1(3):365-75.
  40. Thomson LK, Fleming SD, Aitken RJ, De Iuliis GN, Zieschang JA, Clark AM. Cryopreservation-induced human sperm DNA damage is predominantly mediated by oxidative stress rather than apoptosis. Hum Reprod. 2009;24(9):2061-70.
  41. Zribi N, Feki Chakroun N, El Euch H, Gargouri J, Bahloul A, Ammar Keskes L. Effects of cryopreservation on human sperm deoxyribonucleic acid integrity. Fertil Steril. 2010;93(1):159-66.
  42. Zorn B, Ihan A, Kopitar AN, Kolbezen M, Sesek-Briski A, Meden-Vrtovec H. Changes in sperm apoptotic markers as related to seminal leukocytes and elastase. Reprod Biomed Online. 2010;21(1):84-92.
  43. Brugnon F, Ouchchane L, Pons-Rejraji H, Artonne C, Farigoule M, Janny L. Density gradient centrifugation prior to cryopreservation and hypotaurine supplementation improve post-thaw quality of sperm from infertile men with oligoasthenoteratozoospermia. Hum Reprod. 2013;28(8):2045-57.
  44. Bateni Z, Azadi L, Tavalaee M, Kiani-Esfahani A, Fazilati M, Nasr-Esfahani MH. Addition of Tempol in semen cryopreservation medium improves the post-thaw sperm function. Syst Biol Reprod Med. 2014;60(4):245-50.
  45. Paravicini TM, Touyz RM. NADPH oxidases, reactive oxygen species, and hypertension: clinical implications and therapeutic possibilities. Diabetes Care. 2008;31 Suppl 2:S170-80.
Journal of Reproduction & Infertility
Volume 18, Issue 1
Winter 2017
Pages 172-178