Protective features of resveratrol on human spermatozoa cryopreservation may be mediated through 5’ AMP‐activated protein kinase activation

Abstract

Biochemical and physical modifications during the freeze-thaw process adversely influence the restoration of energy-dependent sperm functions required for fertilization. Resveratrol, a phytoalexin, has been introduced to activate 5' AMP-activated protein kinase which is a cell energy sensor and a cell metabolism regulator. The cryoprotection of resveratrol on sperm cryoinjury via activation of AMP-activated protein kinase also remains to be elucidated. Our aim, thus, was to investigate: (i) the presence and intracellular localization of AMP-activated protein kinase protein; (ii) whether resveratrol may exert a protective effect on certain functional properties of fresh and post-thaw human spermatozoa through modulation of AMP-activated protein kinase. Spermatozoa from normozoospermic men were incubated with or without different concentrations of Compound C as an AMP-activated protein kinase inhibitor or resveratrol as an AMP-activated protein kinase activator for different lengths of time and were then cryopreserved. AMP-activated protein kinase is expressed essentially in the entire flagellum and the post-equatorial region. Viability of fresh spermatozoa was not significantly affected by the presence of Compound C or resveratrol. However, although Compound C caused a potent inhibition of spermatozoa motility parameters, resveratrol did not induce negative effect, except a significant reduction in motility at 25 μm for 1 h. Furthermore, resveratrol significantly increased AMP-activated protein kinase phosphorylation and mitochondrial membrane potential and decreased reactive oxygen species and apoptosis-like changes in frozen-thawed spermatozoa. Nevertheless, it was not able to compensate decreased sperm viability and motility parameters following cryopreservation. In contrast, Compound C showed opposite effects to resveratrol on AMP-activated protein kinase phosphorylation, reactive oxygen species, apoptosis-like changes, mitochondrial membrane potential, and motility parameters. These findings, although preliminary, suggest that resveratrol-induced improvement of cryopreserved sperm functions may be mediated through activation of AMP-activated protein kinase, indicating the importance of AMP-activated protein kinase activity for human spermatozoa functions. Further investigations are required to elucidate the mechanism by which resveratrol ameliorates oxidative stress-mediated damages in an AMP-activated protein kinase-dependent mechanism.

5’-AMP-Activated Protein Kinase Regulates Goat Sperm Functions via Energy Metabolism In Vitro

“…AMPK was expressed in the acrosome, midpiece and the entire flagellum of goat sperm, which was in consistence with that in boar [30], human [16,17] and chicken [15] sperm. However, this observation was different to the report by Swegen et al [31] who demonstrated that AMPK was predominantly present in the midpiece and the entire flagellum of stallion sperm.…”

“…On the other hand, AMPK inhibition led to a significant decrease in motility, mitochondria membrane potential and acrosome integrity during long-term storage of boar semen [21]. However, Shabani Nashtaei et al demonstrated that AMPK inhibition did not affect membrane integrity in human sperm [17]. Moreover, Cordova et al [14] reported that addition of neither AMPK activators nor AMPK inhibitors to the freezing extenders affected motility, membrane integrity and acrosome integrity in stallion sperm.…”

“…Subsequently, evidences have shown that AMPK was also present in stallion [14], chicken [15] and human [16,17] sperm. On one hand, AMPK activation enhanced sperm motility [13,[15][16][17][18][19], acrosome reaction [15,19,20]. On the other hand, AMPK inhibition led to a significant decrease in motility, mitochondria membrane potential and acrosome integrity during long-term storage of boar semen [21].…”

Impact of 5'-amp-activated Protein Kinase on Male Gonad and Spermatozoa Functions

“…Relationships between metabolism and fertility are well established, particularly in the female. Recent discoveries show that AMPK is also present in male gonad and spermatozoa in different species [Caenorhabditis elegans (Lee et al, 2008), oyster (Guevelou et al, 2013), chicken (Nguyen et al, 2014), boar (Hurtado de Llera et al, 2012, 2013), mouse (Tanwar et al, 2012; Tartarin et al, 2012), stallion (Cordova et al, 2014), and human Calle-Guisado et al, 2016; Shabani Nashtaei et al, 2016]. It is these roles in male gonad and spermatozoa that I will now describe.…”

“…Furthermore, the presence of AMPK, especially in its active form, phospho-AMPK-Thr172, was highlighted in spermatozoa of boar, stallion, mouse, chicken and human (Hurtado de Llera et al, 2012, 2013; Tartarin et al, 2012; Cordova et al, 2014; Nguyen et al, 2014; Calle-Guisado et al, 2016; Shabani Nashtaei et al, 2016). In boar, the phospho-AMPK form is mainly localized in the acrosome, the sub-equatorial segment of the head and in the intermediate part (Hurtado de Llera et al, 2013).…”