Pre-Incubation with Kisspeptin Improves the Adverse Effects of Freeze-Thawed Human Ejaculated Sperms

Abstract Objectives: Sperm cryopreservation reduces sperm quality. Kisspeptin (KP), as an antioxidant, has beneficial effects on the sperm functions. Therefore, the present study was conducted to use kisspeptin in order to mitigate detrimental effects of the sperm freeze-thawing process and compare it with Glutathione (GSH), as positive control. Materials and methods: 30 normal semen samples, prepared by swim-up procedure, were divided into 3 aliquots: negative control with no treatment; positive control receiving 1mM GSH; and experimental aliquot treated with 10 µM KP for 30 min. All aliquots were cryopreserved, and then thawed after 48 hr. Sperm motility was assessed according to WHO guidelines. Acrosomal reaction and capacitation were evaluated by Fluorescein Isothiocyanate-Conjugated Peanut Agglutinin (FITC-PNA), Wheat Germ Agglutinin (WGA), and Concanavalin A (ConA); percentage of positive cells was evaluated by flow cytometry. Sperm DNA quality was evaluated by Acridine Orange (AO), Aniline Blue (AB), Chromomycin A3 (CMA3), and TUNEL (Terminal deoxynucleotidyl transferase dUTP Nick End Labeling) staining methods. Statistical analyses were performed using ANOVA and LSD.Results: Results of the study showed that, KP supplementation improved motility and led to an increase in the percentage of capacitated and acrosome-intact sperms compared to both controls. Freeze-thaw procedure damaged the DNA integrity severely, and KP pre-treatment significantly reduced frequency of apoptotic sperms as well as those with histone-protamine substitution impairment. Conclusion: Pre-exposure of the sperms to KP can protect the sperm quality including motility and DNA integrity against detrimental influence of freeze–thaw cycle. Therefore, it can be considered as a good pre-additive substance to control the sperm quality during freezing and thawing procedure.