Cryoprotective effect of resveratrol on DNA damage and crucial human sperm messenger RNAs, possibly through 5′ AMP-activated protein kinase activation

Abstract

This work aimed at investigating the effect of resveratrol on (1) DNA integrity and (2) fertilizing capacity of sperm by quantifying the presence of key paternal transcripts considered as markers for male fertility (protamine 1 [PRM1] and protamine 2 [PRM2]) and pregnancy success (adducin 1 alpha [ADD1]) in cryopreserved human spermatozoa through modulation of AMP-activated protein kinase (AMPK). The study populations was drawn from 22 normozoospermic healthy volunteers which were incubated with or without AMPK activator (resveratrol [RSV], 15 µM) or inhibitor (Compound C [CC], 30 µM) for 1 h and were then cryopreserved. Untreated frozen-thawed spermatozoa served as controls. The RSV-induced AMPK activation decreased the level of DNA fragmentation in comparison with the control (21.18 ± 0.92 vs. 22.50 ± 0.40; p < 0.01). The relative mRNA expression levels of protamines (1 and 2) and ADD1 in RSV pretreated frozen-thawed human spermatozoa were also improved significantly compared to the control (p < 0.05). Conversely, the inhibitory effect of CC on AMPK activity deteriorated the deleterious effects of cryopreservation on these parameters (p < 0.01). In conclusion, these results demonstrated the cryoprotective effect of the RSV-induced increase in AMPK activity on DNA integrity and key paternal transcripts of cryopreserved human spermatozoa. These findings are of great importance for improving the available cryopreservation protocols in terms of the number of lesions that produced over key genes and the dramatic effects on sperm DNA fragmentation.

Inhibitory effect of Lycium�barbarum polysaccharide on sperm damage during cryopreservation

“…Sperm cryopreservation is not only a routine part of the daily work performed at sperm banks but also an indispensable part of assisted reproductive technology. Ultralow temperatures may lead to impaired sperm motility and the formation of ice crystals during freezing and thawing, as well as excessive oxidative stress during this process that may damage the structure and function of sperm (1,2). According to the World Health Organization (WHO), ~50% of sperm cells are damaged or killed during thawing (3).…”

AMPK Function in Mammalian Spermatozoa

“…Many studies have tried to improve male gamete cryopreservation by adding antioxidants to the semen extender to reduce oxidative stress [ 116 ]. In this regard, the natural compound resveratrol that activates AMPK in some somatic cell types, when added to cryopreservation media, had positive effects on human spermatozoa [ 64 , 117 ]. Thus, AMPK was activated and ROS levels were reduced in human spermatozoa cryopreserved in the presence of resveratrol [ 64 ].…”

“…Thus, AMPK was activated and ROS levels were reduced in human spermatozoa cryopreserved in the presence of resveratrol [ 64 ]. Moreover, DNA damage on human cryopreserved spermatozoa was ameliorated when resveratrol was added before cryopreservation, likely mediated by AMPK [ 117 ]. However, when resveratrol was added to boar seminal doses preserved at 17 °C, no improvement was detected in any quality sperm parameter analyzed, and negative effects were reported in mitochondria membrane potential, intracellular ATP content and motility [ 118 ].…”

“…In general, the effects of the manipulation (up or down) of AMPK activity during ART protocols seem contradictory and are basically dependent on the pharmacological agent, species, and/or the ART protocol studied. As an example, whereas resveratrol did not have any positive effect during boar spermatozoa preservation at 17 °C [ 118 ], positive effects were described in cryopreserved human spermatozoa [ 64 , 117 ]. While metformin did not have any positive effect during stallion spermatozoa cryopreservation [ 59 ], very positive outcomes were described for cryopreserved mouse spermatozoa [ 74 ].…”

“…Nevertheless, although differences were observed between species and depending on the ART used, in general, beneficial effects were found when AMPK was shown to be over-activated by metformin in cryopreserved mouse spermatozoa [ 74 ], rosiglitazone in stallion spermatozoa preserved at room temperature [ 60 ], AICAR in cryopreserved chicken spermatozoa [ 119 ] and resveratrol in cryopreserved human spermatozoa [ 64 , 117 ]. The positive effects observed were: (i) improvement of sperm survival, motility, and velocity; (ii) improvement of sperm antioxidant defense and decrease of lipid peroxidation (LPO); and (iii) improvement of embryo quality ( Table 2 ).…”