Open access peer-reviewed chapter

Nitric Oxide: Key Features in Spermatozoa

Written By

Florentin-Daniel Staicu and Carmen Matas Parra

Submitted: 18 May 2016 Reviewed: 20 January 2017 Published: 17 May 2017

DOI: 10.5772/67551

From the Edited Volume

Nitric Oxide Synthase - Simple Enzyme-Complex Roles

Edited by Seyed Soheil Saeedi Saravi

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Abstract

Several in vitro studies have pointed to the importance of nitric oxide (NO) in the female and male reproductive system in mammals. Its functions vary from preventing oocyte aging, improving the integrity of the microtubular spindle apparatus in aged oocytes, modulating the contraction of the oviduct, to regulating sperm physiology by affecting the motility, inducing chemotaxis in spermatozoa, regulating tyrosine phosphorylation, enhancing the sperm-zona pellucida binding ability, and modulating the acrosomal reaction. In spermatozoa, NO exerts its functions in different ways, which involve key elements such as the soluble isoform of guanylate cyclase, cyclic guanosine monophosphate (cGMP), cyclic adenosine monophosphate (cAMP), protein kinase A (PKA), adenylate cyclase, and the extracellular signal-regulated kinase (ERK) pathway. Furthermore, NO leads to the S-nitrosylation of several sperm proteins, among them a substantial group associated with energy generation and cell movement, but also with signal transduction, suggesting a role for S-nitrosylation in sperm motility and in modulating the sperm function, respectively. In this chapter, an overview of how NO modulates the sperm physiology is presented, based on the knowledge acquired to this day.

Keywords

  • nitric oxide
  • nitric oxide synthase
  • S-nitrosylation
  • spermatozoa
  • fertilization

1. Introduction

NO is a small hydrophobic molecule which can easily diffuse through biological membranes [1]. In vivo, it is synthesized during the conversion of L-arginine to L-citrulline by nitric oxide synthase, with the help of co-factors such as the reduced form of nicotinamide adenine dinucleotide phosphate, flavin mononucleotide, flavin adenine dinucleotide, and tetrahydrobiopterin [2].

The nitric oxide synthase (NOS) may be found in three different isoforms. Two of them, the endothelial and the neuronal NOS (eNOS, nNOS), require calcium/calmodulin to be activated and are responsible for the continuous basal release of NO. The third isoform, known as inducible NOS (iNOS), is calcium independent [3, 4]. Since NOS activity depends on the availability of its substrate and its co-factors, all these elements jointly determine the cellular rates of NO synthesis [5].

Substantial evidence indicates that NO is a crucial biological messenger involved in a wide variety of physiological and pathological processes in different systems in mammals, including the vascular, nervous, and reproductive system [1, 6].

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2. NOS/NO duo in the reproductive system

The NOS/NO duo regulates key functions in both the female and male reproductive systems [6].

All three isoforms of NOS have been identified in the oviduct [7, 8], oocytes, and cumulus and corona cells [9, 10] of several species [7, 11, 12]. The expression of NOS isoforms differs during the estrous cycle in the follicles as well as in the oviduct [13]. Tao et al. [10] showed that the immunoreactivity of eNOS in early antral follicles was restricted to the oocyte and increased from small and medium to large follicle-enclosed oocytes. In contrast, no immunoreactivity for iNOS was found in primordial, early antral follicle, or the cumulus-oocyte complexes aspirated from small and medium follicles.

In the oviduct, the endogenous basal release of NO regulates its contraction and the ciliary beating of the ciliated epithelial cells and induces chemotaxis in human spermatozoa via activation of the nitric oxide/soluble isoform of guanylate cyclase/cyclic guanosine monophosphate pathway (NO/sGC/cGMP) [1416]. NO forms a vital component of the oocyte microenvironment and has been positively implicated in meiotic resumption [17], in preventing oocyte aging and improving the integrity of the microtubular spindle apparatus in aged oocytes [18]. It may also contribute as an anti-platelet agent during implantation [19].

As far as the male gamete is concerned, research was first concentrated on determining the effects of NO-releasing compounds on sperm motility and viability. Low concentrations of sodium nitroprusside (SNP), an NO-releasing compound, stimulated sperm hyperactivation in mouse, fish, and hamster [2022] and were beneficial to the maintenance of post-thaw human sperm motility [23]. On the other hand, high concentrations of NO-releasing compounds decreased sperm motility [20, 2426].

Numerous studies have also been conducted to determine the presence and localization of NOS in sperm from several species (Table 1). For example, Herrero et al. [27] located nNOS in the head of freshly ejaculated human spermatozoa, with a more concentrated fluorescent staining toward the equatorial region. O’Bryan et al. [28] described the pattern of eNOS expression in human spermatozoa, finding that morphologically normal spermatozoa exhibited post-acrosomal and equatorial eNOS immunostaining. Interestingly, though, abnormally shaped sperm cells exhibited aberrant staining, especially in the midpiece and/or head region, which correlated negatively with the percentage of motile sperm.

SpeciesAuthorsTechniquesIdentified isoforms
HumanHerrero et al. [27]ImmunofluorescencenNOS
O’Bryan et al. [28]ImmunocytochemistryeNOS
MouseHerrero et al. [29]Kinetic assays measuring the conversion of L-arginine to L-citrullinenNOS, iNOS, eNOS
Western blot
BullMeiser and Schulz [30]Modified griess reaction [92]nNOS, eNOS
Western blot
Immunofluorescence
BoarHou [31]NO assay kit with the Griess reagentnNOS, iNOS, eNOS
Aquila et al. [32]Western blot
StallionOrtega Ferrusola et al. [33]Flow cytometrynNOS, eNOS
Western blot
CatLiman and Alan [34]HistochemistrynNOS, iNOS, eNOS
Immunohistochemistry
Western blot

Table 1.

Summary of in vitro studies and the techniques used to identify NOS isoforms in different species.

NOSs were revealed in mature mouse spermatozoa by means of biochemical techniques and Western blot. Herrero et al. [29] showed that mouse spermatozoa can synthesize L-citrulline, depending on the concentration of L-arginine present in the incubation medium while different concentrations of N(G)-nitro-L-arginine methyl ester (L-NAME) inhibit the formation of the amino acid. Furthermore, when sperm protein extracts were incubated under denaturing and nonreducing conditions and then subjected to immunoblotting assay, a protein fraction of 140 kDa was recognized by the three anti-NOS antibodies.

Bull spermatozoa were examined for the presence of constitutive NOS [30]. NO generation seemed to be enhanced by L-arginine and abolished by the NOS-inhibitor, L-NAME. In addition, Meiser and Schulz [30] verified the presence of NOS in bull sperm cells by immunohistochemistry, which was confirmed by Western blot. Confocal laser microscopy localized nNOS-related immunofluorescence at the acrosome cap and the main part of the flagellum. The same technique also identified eNOS staining spread over the spermatozoan head. Moreover, when these findings were confirmed by Western blot, immunoreactive bands at 161 kDa (nNOS) and 133 kDa (eNOS) were identified.

Hou et al. [31] investigated whether boar sperm can generate NO, finding that porcine spermatozoa synthesized low levels of NO under noncapacitating conditions, but that the NO concentration almost doubled when sperms were capacitated. Furthermore, NO production was significantly inhibited when capacitated sperms were treated with L-NAME. In another study [32], Western blot analysis was performed to identify NOS enzymes in boar sperm samples. The immunoblots showed three distinct bands: ~160, ~130, and ~135 kDa, corresponding to nNOS, iNOS, and eNOS, respectively.

NO production was evaluated in stallion spermatozoa before and after freezing/thawing [33] by means of flow cytometry, after loading the sperm suspension with an NO detection probe. NO synthesis was positively correlated with sperm motility after thawing and, interestingly, the presence of egg yolk in the semen extender radically reduced the amount of NO produced. The authors further investigated in fresh and frozen/thawed stallion sperm the presence of NOS enzymes by Western blot, using anti-nNOS, anti-eNOS, and anti-universal NOS antibodies. Two bands of approximately 83 kDa and 96 kDa were labeled by the antibodies anti-nNOS and anti-eNOS, respectively. Moreover, the other antibody, which recognized an epitope present in all the NOS isoforms described so far, showed two similar bands of 84 and 92 kDa.

Recently, Liman and Alan [34] investigated the localization of NOS isoforms in spermatozoa within the intratesticular and excurrent duct systems of adult domestic cats. Overall, the spermatozoa head did not exhibit immunoreactivity. On the other hand, immunoreactivity for all three isoforms was observed in the flagellum, in the proximal cytoplasmic droplets of spermatozoa (located in the neck region) within the lumen of the intratesticular and efferent ducts, in the epididymal duct of the caput epididymis, and in the distal cytoplasmic droplets of spermatozoa (located at the mid-principal piece junction of the tail) within the lumen of corpus and cauda epididymis and the vas deferens.

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3. Role of NO on sperm functionality

Several in vitro studies were conducted in order to determine the effects that NO has on sperm physiology (Figure 1). It has been shown that NO affects sperm motility [28, 35, 36], acts as chemoattractant [16, 37], regulates the tyrosine phosphorylation of different sperm proteins [38, 39], enhances the sperm-zona pellucida binding ability [40], and modulates the acrosomal reaction [41, 42].

Figure 1.

Some aspects of the sperm physiology modulated by the NOS/NO system. At physiological levels, endogenous NO has a beneficial role in maintaining sperm motility, enhances tyrosine phosphorylation, which, in turn, promotes the capacitation process. NO also increases the sperm-zona pellucida binding ability and leads to a rise in the percentage of reacted spermatozoa, especially in the presence of follicular fluid or protein-enriched extracts of follicular fluid.

In detail, NO seems to play an important role in the maintenance of sperm motility at physiological levels. A study [36] showed that the basal release of NO by spermatozoa from normozoospermic samples tended to be greater than that from asthenozoospermic samples, suggesting a physiological and beneficial role for endogenous NO in the preservation of sperm motility. These observations agree with a previous report that normozoospermic spermatozoa express more NOS and generate more nitrite than asthenozoospermic spermatozoa [35]. On the other hand, as previously mentioned, it has been shown that spermatozoa with an abnormal morphology show aberrant staining for eNOS, which was negatively correlated with the motility [28]. A detrimental effect on motility has also been reported by Rosselli et al. [24] and Weinberg et al. [25] when millimolar concentrations of exogenous NO donors were added to sperm samples.

It has been suggested that, upon approaching and entering the cumulus oophorus, both NO and progesterone, which are synthesized by the cumulus cells [9, 10, 4345], provide a synergistic stimulus to mobilize stored calcium in the sperm neck/midpiece [46]. As a consequence, they can modulate flagellar activity and contribute to the hyperactivation that is vital for penetration of the oocyte vestments [47].

Interestingly, it has also been suggested that NO may exert a chemoattractant effect on spermatozoa. In fact, the percentage of mouse sperm migrating toward the medium containing an NO donor increased significantly [37]. Similar results were obtained when human spermatozoa were exposed to an NO donor [16]. In the latter case, the signal transduction pathway was also studied. It was proposed that NO exerts its chemoattractant effect through the activation of the NO/sGC/cGMP pathway, since the use of an NO scavenger and/or an sGC and cGMP-dependent protein kinase inhibitor reverted the NO donor-induced migration of sperm.

Since tyrosine phosphorylation in different sperm proteins is associated with the capacitation process [48], this aspect was investigated in order to further define the involvement of NO in capacitation. Herrero et al. [38] observed an increase in tyrosine phosphorylation when human sperm capacitation was accelerated by an NO-releasing compound. On the other hand, when sperm capacitation was inhibited by L-NAME, there was an attenuation in the tyrosine phosphorylation of sperm proteins. In addition, Thundathil et al. [39] reported that L-NAME prevented, and a NO donor promoted, the increase in threonine, glutamine, and tyrosine phosphorylation in human spermatozoa. Furthermore, the addition of L-arginine reversed the inhibitory effect of L-NAME on the capacitation and the associated increase in phosphorylation.

The correlation between NO and sperm-zona pellucida binding ability was investigated by Sengoku et al. [40], who reported that when treated with low concentrations of a NO donor, the number of spermatozoa which binds to the hemizona is higher than in sperm treated with a higher concentration. Additionally, a NO quencher lowered the enhancement of sperm binding by the NO donor.

NO also seems to modulate the acrosome reaction. The percentage of acrosome loss induced by human follicular fluid or by calcium ionophore was studied when human spermatozoa were capacitated in the presence/absence of NO-releasing compounds or NOS inhibitors [38]. NO donors induced sperm cells to respond faster to human follicular fluid, whereas NOS inhibitors decreased the percentage of acrosome reaction. Similar results were obtained by Revelli et al. [41], who showed that different NO-releasing compounds were able to increase the percentage of reacted spermatozoa in the presence of protein-enriched extracts of human follicular fluid. Also, hemoglobin, a NO scavenger, inhibited the follicular fluid–induced acrosomal reaction. In an in-depth analysis of the signaling pathway of the nitric oxide–induced acrosome reaction in human spermatozoa [42], the authors suggested that the acrosome reaction-inducing effect of exogenous NO on capacitated human spermatozoa is accomplished via the NO/sGC/cGMP pathway, which leads to the activation of cGMP-dependent protein kinase (PKG). In fact, both the intracellular cGMP levels and the percentage of reacted spermatozoa were significantly increased after incubation with SNP. Furthermore, the SNP-induced acrosome reaction was significantly reduced in the presence of sGC inhibitors, a reduction that was reversed by the addition of a cell-permeating cGMP analogue to the incubation medium. Finally, PKG inhibition reduced the SNP-induced acrosome reaction.

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4. NOS-activating molecules

As previously stated, NOS activity depends on the availability of its substrate and co-factors [5]. However, the scientific literature does not include many studies on the molecules present in the female reproductive tract which may activate, in one way or another, NOS enzymes in spermatozoa.

Starting from follicular fluid samples, Revelli et al. [41] obtained a protein-enriched follicular fluid solution (PFF), which was then used to study its effects on NOS activity, citrulline synthesis, and acrosome reaction in human sperm. Interestingly, this study showed for the first time that the endogenous NOS activity of human sperm may be increased by PFF. Moreover, the authors demonstrated that PFF-mediated induction of sperm NOS activity leads to acrosome reaction in the same cells, thereby, establishing a link between follicle-derived substances, the activation of NO synthesis in sperm and biological responses.

Furthermore, the increase in NO synthesis mediated by PFF was not associated with a rise in the expression of NOS catalytic units, which is not surprising since specialized cells possess very poor, if any, transcriptional activity [41]. The authors hypothesized that PFF first determines the transient enzyme activation of sperm NOS, which is subsequently strengthened by a more stable modification of the enzyme.

However, more studies should be performed in order to identify the NOS-activating molecule(s) in the follicular fluid.

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5. NO pathway in spermatozoa

In spermatozoa, NO acts via three main pathways (Figure 2) [13]. First, NO is able to activate sGC, leading to a rise in the intracellular levels of cGMP [49]. The latter activates the cyclic nucleotide-gated channels (CNG) localized in the flagellum of mammalian spermatozoa [50, 51]. These channels seem to play an important role in the sperm motility control, by allowing the entry of Ca2+ ions to the cytoplasm during the capacitation process of mammal sperm [50]. Their activation is one of the first events that occur during capacitation in the mouse spermatozoa [52]. cGMP also activates PKG [53, 54], which is involved in the serine/threonine phosphorylation of proteins that promote sperm capacitation and the acrosome reaction [55, 56]. Furthermore, since cGMP and cAMP compete for the catalytic sites of phosphodiesterases [57, 58], an increase in the intracytoplasmic cGMP concentration may inhibit cAMP degradation via cyclic nucleotide phosphodiesterase type 3 [59], thus increasing cAMP intracellular levels and activating protein kinase A (PKA). The latter leads to an increase in protein tyrosine phosphorylation [60].

Figure 2.

Representation of the main pathways through which NO acts in spermatozoa. NO leads to an increase in the intracellular levels of cyclic guanosine monophosphate (cGMP) by activating the soluble isoform of guanylate cyclase (sGC). The cGMP can activate the cyclic nucleotide-gated channels (CNG) localized in the flagellum of mammalian spermatozoa, which regulate the influx of Ca2+ ions to the cytoplasm during the capacitation process and also activates the cGMP-dependent protein kinase (PKG), leading to the serine/threonine phosphorylation of different proteins. It can also inhibit cyclic adenosine monophosphate (cAMP) degradation via cyclic nucleotide phosphodiesterase (PDE), which leads to the activation of cAMP-dependent protein kinase A (PKA) and tyrosine phosphorylation. Furthermore, NO is involved in the tyrosine phosphorylation process in a direct manner, by activating adenylate cyclase (AC) and the extracellular signal-regulated kinase (ERK) pathway. Finally, NO determines post-translational protein modification in spermatozoa via S-nitrosylation.

Second, NO is directly involved in tyrosine phosphorylation by modulating the cAMP/PKA and the extracellular signal-regulated kinase (ERK) pathways. The cAMP/PKA pathway can be influenced by NO via activation of sGC (as described above), but it can also be regulated directly. In fact, S-nitrosylation of adenylate cyclase (AC) has been suggested as a possible mechanism of action of NO [61]. Low levels of NO may activate AC, consequently increasing the cAMP concentration and activating PKA [62]. However, high levels of NO can inhibit AC [61]. As far as the ERK pathway is concerned, NO reacts with the cysteine residues of the RAS protein, inducing its activation [35]. In turn, RAS triggers the RAF, MEK, and ERK1/2 complex, necessary for tyrosine phosphorylation [63].

Third, NO regulates the post-translational protein modification in spermatozoa via S-nitrosy-lation [64], a process similar to phosphorylation and acetylation [65, 66]. S-nitrosylation consists of the covalent incorporation of NO into thiol groups (-SH) to form S-nitrosothiols (S-NO), a modification that is selective and reversible [13].

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6. Function of S-nitrosoproteins in spermatozoa

An extensive study by Lefièvre et al. [64] described a large number of proteins present in the sperm of normozoospermic men, which can be subjected to S-nitrosylation in the presence of NO donors. Although the function of some nitrosylated proteins remains to be discovered, a considerable group of them are known to be metabolic proteins and proteins associated with energy generation and cell movement, suggesting a role for S-nitrosylation in sperm motility. This agrees with a previous proteomic analysis [67], in which the most abundant group was also involved in energy production.

Other considerable groups of proteins were those involved in signal transduction, which agrees with a role for S-nitrosylation in modulating the sperm function [64]. Interestingly, since sperms are generally assumed to be transcriptionally inactive, a small percentage of the S-nitrosylated proteins identified by Lefièvre et al. [64] was related to transcription. Previous proteomic studies in sperm also observed the presence of proteins involved in transcription [67, 68]. However, when comparing the human sperm S-nitrosoproteome with proteins identified during a proteomic study of sperm-oocyte interaction, only three proteins were found in common, suggesting that S-nitrosylation is not a regulatory mechanism employed during fertilization [64, 69].

It is known that the mobilization of Ca2+ stored in the sperm neck/midpiece is necessary for the hyperactivation process [46]. The Ca2+ store in the neck of the sperm coincides with the region occupied by the redundant nuclear envelope (RNE) [70] and in order to mobilize Ca2+ from this site, ryanodine receptors (RyRs), which are intracellular Ca2+-release channels involved in regulation of cytosolic calcium levels [71], need to be activated. These proteins contain a large number of thiol groups and are thus prone to S-nitrosylation by NO [64, 72, 73]. S-nitrosylation can potentiate the opening of RyRs [7479], probably through the generation of the membrane permanent product S-nitrosocysteine [80]. It has been shown that an increase in Ca2+ induced by NO is accompanied by an increase in S-nitrosylation levels of endogenous RyRs [81, 82] while these Ca2+ channels may be inhibited under strongly nitrosylating conditions or at high doses of NO (Figure 3) [76, 79, 82]. Furthermore, progesterone acts synergistically with NO to mobilize Ca2+ in the sperm neck/midpiece by activation of RyRs [47], contributing to the hyperactivation process.

Figure 3.

S-nitrosylation process. NO acts on the thiol groups (-SH) of the cysteines in proteins to form S-nitrosothiols (S-NO). At the sperm neck/midpiece, the S-nitrosylation occurs in ryanodine receptors (RyRs) allowing the release of calcium from the redundant nuclear envelope (RNE), which is required for sperm hyperactivation. Adapted and modified from López-Úbeda and Matás [13].

Other examples of proteins which can undergo S-nitrosylation in sperm and have a known biological significance are the A-kinase anchoring proteins (AKAPs) [64]. Both AKAP3 and AKAP4 are present in the fibrous sheath of the sperm flagellum, control PKA activity and undergo phosphorylation during the capacitation process [8385]. AKAP complexes also modulate the motility of sperm. In fact, phosphodiesterase inhibitors were seen to significantly increase sperm motility [86], whereas PKA-anchoring inhibitor peptides arrested sperm motility [87]. Since the effects of NO on sperm motility are well established, the S-nitrosylation of AKAPs would be an interesting subject for additional studies.

A number of heat shock proteins (HSPs) may also be targets of S-nitrosylation in sperm [64], and some of them have been reported to act as important modulators of sperm capacitation. For instance, Asquith et al. [88] reported that heat shock protein 1 and endoplasmin undergo tyrosine phosphorylation during mouse sperm capacitation, whereas Nixon et al. [89] suggested that they form part of a zona pellucida complex, allowing successful sperm-egg interaction in the same species. Heat shock 70 kDa protein 8 and heat shock protein 90α also undergo tyrosine phosphorylation during human sperm capacitation [83], but whether they function in a zona receptor complex is still unknown [64]. Furthermore, HspA2 has been shown to be a marker of sperm maturity [90], its expression in infertile men with idiopathic oligoteratozoospermia being lower than in normozoospermic men [91].

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7. Concluding remarks

In recent years, our knowledge of the involvement of the NOS/NO system in mammalian fertilization has grown, and there is clear evidence that NO acts as a significant modulator of the male and female gamete. However, many aspects regarding the NOS/NO duo, such as the presence of NOS-activating molecule(s) in the fertilization site or how the biological function of the S-nitrosylated proteins changes, remain to be discovered. Shedding light on these mechanisms will increase our understanding of the etiopathology of subfertility/infertility problems and how such problems can be overcome.

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Acknowledgments

This work was supported by H2020 MSC-ITN-EJD 675526 REP-BIOTECH, the Spanish Ministry of Economy and Competitiveness (MINECO) and the European Regional Development Fund (FEDER), Grants AGL2015–66341-R.

References

  1. 1. Pacher P, Beckman JS, Liaudet L. Nitric oxide and peroxynitrite in health and disease. Physiol Rev. 2007;87(1):315–424. DOI: http://dx.doi.org/10.1152/physrev.00029.2006.
  2. 2. Halliwell B, Gutteridge JMC. Free Radicals in Biology and Medicine. 3rd ed. Clarendon Press, Oxford:1999.
  3. 3. Griffith OW, Stuehr DJ. Nitric oxide synthases: properties and catalytic mechanism. Annu Rev Physiol. 1995;57:707–736. DOI: http://dx.doi.org/10.1146/annurev.ph.57.030195.003423.
  4. 4. Snyder SH. Nitric oxide. No endothelial NO. Nature. 1995;377(6546):196–197. DOI: http://dx.doi.org/10.1038/377196a0.
  5. 5. Rosselli M, Keller PJ, Dubey RK. Role of nitric oxide in the biology, physiology and pathophysiology of reproduction. Hum Reprod Update. 1998;4(1):3–24. DOI: 10.1093/humupd/4.1.3.
  6. 6. O’Flaherty C, Rodriguez P, Srivastava S. L-arginine promotes capacitation and acrosome reaction in cryopreserved bovine spermatozoa. Biochim Biophys Acta. 2004;1674(2):215–221. DOI: http://dx.doi.org/10.1016/j.bbagen.2004.06.020.
  7. 7. Rosselli M, Dubey RK, Rosselli MA, Macas E, Fink D, Lauper U, Keller PJ, Imthurn B. Identification of nitric oxide synthase in human and bovine oviduct. Mol Hum Reprod. 1996;2(8):607–612. DOI: 10.1093/molehr/2.8.607.
  8. 8. Lapointe J, Roy M, St-Pierre I, Kimmins S, Gauvreau D, MacLaren LA, Bilodeau JF. Hormonal and spatial regulation of nitric oxide synthases (NOS) (neuronal NOS, inducible NOS, and endothelial NOS) in the oviducts. Endocrinology. 2006;147(12):5600–5610. DOI: http://dx.doi.org/10.1210/en.2005–1548.
  9. 9. Reyes R, Vázquez ML, Delgado NM. Detection and bioimaging of nitric oxide in bovine oocytes and sperm cells. Arch Androl. 2004;50(4):303–309. DOI: http://dx.doi.org/10.1080/01485010490448471.
  10. 10. Tao Y, Fu Z, Zhang M, Xia G, Yang J, Xie H. Immunohistochemical localization of inducible and endothelial nitric oxide synthase in porcine ovaries and effects of NO on antrum formation and oocyte meiotic maturation. Mol Cell Endocrinol. 2004;222(1–2):93–103. DOI: http://dx.doi.org/10.1016/j.mce.2004.04.014.
  11. 11. Ekerhovd E, Brännström M, Alexandersson M, Norström A. Evidence for nitric oxide mediation of contractile activity in isolated strips of the human fallopian tube. Hum Reprod. 1997;12(2):301–305. DOI: 10.1093/humrep/12.2.301.
  12. 12. Bryant CE, Tomlinson A, Mitchell JA, Thiemermann C, Willoughby DA. Nitric oxide synthase in the rat fallopian tube is regulated during the oestrous cycle. J Endocrinol. 1995;146(1):149–157. DOI: 10.1677/joe.0.1460149.
  13. 13. López-Úbeda R, Matás C. An approach to the factors related to sperm capacitation process. Andrology. 2015;4:1.
  14. 14. Rosselli M, Imthurn B, Macas E, Keller PJ, Dubey RK. Endogenous nitric oxide modulates endothelin-1 induced contraction of bovine oviduct. Biochem Biophys Res Commun. 1994;201(1):143–148. DOI: http://dx.doi.org/10.1006/bbrc.1994.1680.
  15. 15. Zhan X, Li D, Johns RA. Expression of endothelial nitric oxide synthase in ciliated epithelia of rats. J Histochem Cytochem. 2003;51(1):81–87. DOI: 10.1177/002215540305100110.
  16. 16. Miraglia E, Rullo ML, Bosia A, Massobrio M, Revelli A, Ghigo D. Stimulation of the nitric oxide/cyclic guanosine monophosphate signaling pathway elicits human sperm chemotaxis in vitro. Fertil Steril. 2007;87(5):1059–1063. DOI: http://dx.doi.org/10.1016/j.fertnstert.2006.07.1540.
  17. 17. Romero-Aguirregomezcorta J, Santa ÁP, García-Vázquez FA, Coy P, Matás C. Nitric oxide synthase (NOS) inhibition during porcine in vitro maturation modifies oocyte protein S-nitrosylation and in vitro fertilization. PLoS One. 2014;9(12):e115044. DOI: 10.1371/journal.pone.0115044.
  18. 18. Goud AP, Goud PT, Diamond MP, Abu-Soud HM. Nitric oxide delays oocyte aging. Biochemistry. 2005;44(34):11361–11368. DOI: http://dx.doi.org/10.1021/bi050711f.
  19. 19. Cameron IT, Campbell S. Nitric oxide in the endometrium. Hum Reprod Update. 1998;4(5):565–569. DOI: 10.1093/humupd/4.5.565.
  20. 20. Herrero MB, Cebral E, Boquet M, Viggiano JM, Vitullo A, Gimeno MA. Effect of nitric oxide on mouse sperm hyperactivation. Acta Physiol Pharmacol Ther Latinoam. 1994;44(3):65–69.
  21. 21. Creech MM, Arnold EV, Boyle B, Muzinich MC, Montville C, Bohle DS, Atherton RW. Sperm motility enhancement by nitric oxide produced by the oocytes of fathead minnows, Pimephelas promelas. J Androl. 1998;19(6):667–674. DOI: 10.1002/j.1939–4640.1998.tb02076.x.
  22. 22. Yeoman RR, Jones WD, Rizk BM. Evidence for nitric oxide regulation of hamster sperm hyperactivation. J Androl. 1998;19(1):58–64. DOI: 10.1002/j.1939–4640.1998.tb02470.x.
  23. 23. Hellstrom WJ, Bell M, Wang R, Sikka SC. Effect of sodium nitroprusside on sperm motility, viability, and lipid peroxidation. Fertil Steril. 1994;61(6):1117–1122.
  24. 24. Rosselli M, Dubey RK, Imthurn B, Macas E, Keller PJ. Effects of nitric oxide on human spermatozoa: evidence that nitric oxide decreases sperm motility and induces sperm toxicity. Hum Reprod. 1995;10(7):1786–1790.
  25. 25. Weinberg JB, Doty E, Bonaventura J, Haney AF. Nitric oxide inhibition of human sperm motility. Fertil Steril. 1995;64(2):408–413.
  26. 26. Tomlinson MJ, East SJ, Barratt CL, Bolton AE, Cooke ID. Preliminary communication: possible role of reactive nitrogen intermediates in leucocyte-mediated sperm dysfunction. Am J Reprod Immunol. 1992;27(1–2):89–92. DOI: 10.1111/j.1600–0897.1992.tb00730.x.
  27. 27. Herrero MB, Pérez Martínez S, Viggiano JM, Polak JM, de Gimeno MF. Localization by indirect immunofluorescence of nitric oxide synthase in mouse and human spermatozoa. Reprod Fertil Dev. 1996;8(5):931–934.
  28. 28. O’Bryan MK, Zini A, Cheng CY, Schlegel PN. Human sperm endothelial nitric oxide synthase expression: correlation with sperm motility. Fertil Steril. 1998;70(6):1143–1147. DOI: http://dx.doi.org/10.1016/S0015–0282(98)00382–3.
  29. 29. Herrero MB, Goin JC, Boquet M, Canteros MG, Franchi AM, Perez Martinez S, Polak JM, Viggiano JM, Gimeno MA. The nitric oxide synthase of mouse spermatozoa. FEBS Lett. 1997;411(1):39–42. DOI: 10.1016/S0014–5793(97)00570-X.
  30. 30. Meiser H, Schulz R. Detection and localization of two constitutive NOS isoforms in bull spermatozoa. Anat Histol Embryol. 2003;32(6):321–325. DOI: 10.1111/j.1439–0264.2003.00459.x.
  31. 31. Hou ML, Huang SY, Lai YK, Lee WC. Geldanamycin augments nitric oxide production and promotes capacitation in boar spermatozoa. Anim Reprod Sci. 2008;104(1):56–68. DOI: http://dx.doi.org/10.1016/j.anireprosci.2007.01.006.
  32. 32. 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. DOI: http://dx.doi.org/10.1186/1477–7827-9–133.
  33. 33. Ortega Ferrusola C, González Fernández L, Macías García B, Salazar-Sandoval C, Morillo Rodríguez A, Rodríguez Martinez H, Tapia JA, Peña FJ. Effect of cryopreservation on nitric oxide production by stallion spermatozoa. Biol Reprod. 2009;81(6):1106–1111. DOI: 10.1095/biolreprod.109.078220.
  34. 34. Liman N, Alan E. Region-specific localization of NOS isoforms and NADPH-diaphorase activity in the intratesticular and excurrent duct systems of adult domestic cats (Felis catus). Microsc Res Tech. 2016;79(3):192–208. DOI: 10.1002/jemt.22619.
  35. 35. Lewis SE, Donnelly ET, Sterling ES, Kennedy MS, Thompson W, Chakravarthy U. Nitric oxide synthase and nitrite production in human spermatozoa: evidence that endogenous nitric oxide is beneficial to sperm motility. Mol Hum Reprod. 1996;2(11):873–878. DOI: 10.1093/molehr/2.11.873.
  36. 36. Donnelly ET, Lewis SE, Thompson W, Chakravarthy U. Sperm nitric oxide and motility: the effects of nitric oxide synthase stimulation and inhibition. Mol Hum Reprod. 1997;3(9):755–762. DOI: 10.1093/molehr/3.9.755.
  37. 37. Sliwa L, Stochmal E. Effect of sodium nitroprusside on mouse sperm migration in vitro. Arch Androl. 2000;45(1):29–33.
  38. 38. Herrero MB, de Lamirande E, Gagnon C. Nitric oxide regulates human sperm capacitation and protein-tyrosine phosphorylation in vitro. Biol Reprod. 1999;61(3):575–581. DOI: 10.1095/biolreprod61.3.575.
  39. 39. Thundathil J, de Lamirande E, Gagnon C. Nitric oxide regulates the phosphorylation of the threonine-glutamine-tyrosine motif in proteins of human spermatozoa during capacitation. Biol Reprod. 2003;68(4):1291–1298. DOI: http://dx.doi.org/10.1095/biolreprod.102.008276.
  40. 40. Sengoku K, Tamate K, Yoshida T, Takaoka Y, Miyamoto T, Ishikawa M. Effects of low concentrations of nitric oxide on the zona pellucida binding ability of human spermatozoa. Fertil Steril. 1998;69(3):522–527. DOI: http://dx.doi.org/10.1016/S0015–0282(97)00537–2.
  41. 41. Revelli A, Soldati G, Costamagna C, Pellerey O, Aldieri E, Massobrio M, Bosia A, Ghigo D. Follicular fluid proteins stimulate nitric oxide (NO) synthesis in human sperm: a possible role for NO in acrosomal reaction. J Cell Physiol. 1999;178(1):85–92. DOI: http://dx.doi.org/10.1002/(SICI)1097–4652(199901)178:1%3C85::AID-JCP11%3E3.0.CO;2-Y.
  42. 42. Revelli A, Costamagna C, Moffa F, Aldieri E, Ochetti S, Bosia A, Massobrio M, Lindblom B, Ghigo D. Signaling pathway of nitric oxide-induced acrosome reaction in human spermatozoa. Biol Reprod. 2001;64(6):1708–1712. DOI: 10.1095/biolreprod64.6.1708.
  43. 43. Chian RC, Ao A, Clarke HJ, Tulandi T, Tan SL. Production of steroids from human cumulus cells treated with different concentrations of gonadotropins during culture in vitro. Fertil Steril. 1999;71(1):61–66. DOI: http://dx.doi.org/10.1016/S0015–0282(98)00416–6.
  44. 44. Mingoti GZ, Garcia JM, Rosa-e-Silva AA. Steroidogenesis in cumulus cells of bovine cumulus-oocyte-complexes matured in vitro with BSA and different concentrations of steroids. Anim Reprod Sci. 2002;69(3–4):175–186. DOI: http://dx.doi.org/10.1016/S0378–4320(01)00187–7.
  45. 45. Yamashita Y, Shimada M, Okazaki T, Maeda T, Terada T. Production of progesterone from de novo-synthesized cholesterol in cumulus cells and its physiological role during meiotic resumption of porcine oocytes. Biol Reprod. 2003;68(4):1193–1198. DOI: 10.1095/biolreprod.102.010934.
  46. 46. Bedu-Addo K, Costello S, Harper C, Machado-Oliveira G, Lefievre L, Ford C, Barratt C, Publicover S. Mobilisation of stored calcium in the neck region of human sperm—a mechanism for regulation of flagellar activity. Int J Dev Biol. 2008;52(5–6):615–626. DOI: 10.1387/ijdb.072535kb.
  47. 47. Machado-Oliveira G, Lefièvre L, Ford C, Herrero MB, Barratt C, Connolly TJ, Nash K, Morales-Garcia A, Kirkman-Brown J, Publicover S. Mobilisation of Ca2+ stores and flagellar regulation in human sperm by S-nitrosylation: a role for NO synthesised in the female reproductive tract. Development. 2008;135(22):3677–3686. DOI: 10.1242/dev.024521.
  48. 48. Emiliozzi C, Fenichel P. Protein tyrosine phosphorylation is associated with capacitation of human sperm in vitro but is not sufficient for its completion. Biol Reprod. 1997;56(3):674–679. DOI: 10.1095/biolreprod56.3.674.
  49. 49. Murad F. The nitric oxide-cyclic GMP signal transduction system for intracellular and intercellular communication. Recent Prog Horm Res. 1994;49:239–248.
  50. 50. Wiesner B, Weiner J, Middendorff R, Hagen V, Kaupp UB, Weyand I. Cyclic nucleotide-gated channels on the flagellum control Ca2+ entry into sperm. J Cell Biol. 1998;142(2):473–484. DOI: 10.1083/jcb.142.2.473.
  51. 51. Weyand I, Godde M, Frings S, Weiner J, Müller F, Altenhofen W, Hatt H, Kaupp UB. Cloning and functional expression of a cyclic-nucleotide-gated channel from mammalian sperm. Nature. 1994;368(6474):859–863. DOI: http://dx.doi.org/10.1038/368859a0.
  52. 52. Cisneros-Mejorado A, Hernández-Soberanis L, Islas-Carbajal MC, Sánchez D. Capacitation and Ca(2+) influx in spermatozoa: role of CNG channels and protein kinase G. Andrology. 2014;2(1):145–154. DOI: 10.1111/j.2047–2927.2013.00169.x.
  53. 53. Lohmann SM, Vaandrager AB, Smolenski A, Walter U, De Jonge HR. Distinct and specific functions of cGMP-dependent protein kinases. Trends Biochem Sci. 1997;22(8):307–312. DOI: http://dx.doi.org/10.1016/S0968–0004(97)01086–4.
  54. 54. Pfeifer A, Ruth P, Dostmann W, Sausbier M, Klatt P, Hofmann F. Structure and function of cGMP-dependent protein kinases. Rev Physiol Biochem Pharmacol. 1999;135:105–149.
  55. 55. Rahman MS, Kwon WS, Pang MG. Calcium influx and male fertility in the context of the sperm proteome: an update. Biomed Res Int. 2014;2014:841615. DOI: 10.1155/2014/841615.
  56. 56. Miraglia E, De Angelis F, Gazzano E, Hassanpour H, Bertagna A, Aldieri E, Revelli A, Ghigo D. Nitric oxide stimulates human sperm motility via activation of the cyclic GMP/protein kinase G signaling pathway. Reproduction. 2011;141(1):47–54. DOI: 10.1530/REP-10–0151.
  57. 57. Bender AT, Beavo JA. Cyclic nucleotide phosphodiesterases: molecular regulation to clinical use. Pharmacol Rev. 2006;58(3):488–520. DOI: http://dx.doi.org/10.1124/pr.58.3.5.
  58. 58. Conti M, Beavo J. Biochemistry and physiology of cyclic nucleotide phosphodiesterases: essential components in cyclic nucleotide signaling. Annu Rev Biochem. 2007;76:481–511. DOI: http://dx.doi.org/10.1146/annurev.biochem.76.060305.150444.
  59. 59. Kurtz A, Götz KH, Hamann M, Wagner C. Stimulation of renin secretion by nitric oxide is mediated by phosphodiesterase 3. Proc Natl Acad Sci U S A. 1998;95(8):4743–4747.
  60. 60. Beavo JA. Cyclic nucleotide phosphodiesterases: functional implications of multiple isoforms. Physiol Rev. 1995;75(4):725–748.
  61. 61. McVey M, Hill J, Howlett A, Klein C. Adenylyl cyclase, a coincidence detector for nitric oxide. J Biol Chem. 1999;274(27):18887–18892. DOI: 10.1074/jbc.274.27.18887.
  62. 62. Belén Herrero M, Chatterjee S, Lefièvre L, de Lamirande E, Gagnon C. Nitric oxide interacts with the cAMP pathway to modulate capacitation of human spermatozoa. Free Radic Biol Med. 2000;29(6):522–536. DOI: http://dx.doi.org/10.1016/S0891–5849(00)00339–7.
  63. 63. de Lamirande E, Gagnon C. The extracellular signal-regulated kinase (ERK) pathway is involved in human sperm function and modulated by the superoxide anion. Mol Hum Reprod. 2002;8(2):124–135. DOI: 10.1093/molehr/8.2.124.
  64. 64. Lefièvre L, Chen Y, Conner SJ, Scott JL, Publicover SJ, Ford WC, Barratt CL. Human spermatozoa contain multiple targets for protein S-nitrosylation: an alternative mechanism of the modulation of sperm function by nitric oxide?. Proteomics. 2007;7(17):3066–3084. DOI: http://dx.doi.org/10.1002/pmic.200700254.
  65. 65. Foster MW, Stamler JS. New insights into protein S-nitrosylation. Mitochondria as a model system. J Biol Chem. 2004;279(24):25891–25897. DOI: http://dx.doi.org/10.1074/jbc.M313853200.
  66. 66. Hess DT, Matsumoto A, Kim SO, Marshall HE, Stamler JS. Protein S-nitrosylation: purview and parameters. Nat Rev Mol Cell Biol. 2005;6(2):150–166. DOI: http://dx.doi.org/10.1038/nrm1569.
  67. 67. Martínez-Heredia J, Estanyol JM, Ballescà JL, Oliva R. Proteomic identification of human sperm proteins. Proteomics. 2006;6(15):4356–4369. DOI: http://dx.doi.org/10.1002/pmic.200600094.
  68. 68. Lalancette C, Faure RL, Leclerc P. Identification of the proteins present in the bull sperm cytosolic fraction enriched in tyrosine kinase activity: a proteomic approach. Proteomics. 2006;6(16):4523–4540. DOI: http://dx.doi.org/10.1002/pmic.200500578.
  69. 69. Stein KK, Go JC, Lane WS, Primakoff P, Myles DG. Proteomic analysis of sperm regions that mediate sperm-egg interactions. Proteomics. 2006;6(12):3533–3543. DOI: http://dx.doi.org/10.1002/pmic.200500845.
  70. 70. Costello S, Michelangeli F, Nash K, Lefievre L, Morris J, Machado-Oliveira G, Barratt C, Kirkman-Brown J, Publicover S. Ca2+-stores in sperm: their identities and functions. Reproduction. 2009;138(3):425–437. DOI: 10.1530/REP-09–0134.
  71. 71. Kakizawa S, Yamazawa T, Iino M. Nitric oxide-induced calcium release: activation of type 1 ryanodine receptor by endogenous nitric oxide. Channels (Austin). 2013;7(1):1–5. DOI: 10.4161/chan.22555.
  72. 72. Takeshima H, Nishimura S, Matsumoto T, Ishida H, Kangawa K, Minamino N, Matsuo H, Ueda M, Hanaoka M, Hirose T, et al. Primary structure and expression from complementary DNA of skeletal muscle ryanodine receptor. Nature. 1989;339(6224):439–445. DOI: http://dx.doi.org/10.1038/339439a0.
  73. 73. Otsu K, Willard HF, Khanna VK, Zorzato F, Green NM, MacLennan DH. Molecular cloning of cDNA encoding the Ca2+ release channel (ryanodine receptor) of rabbit cardiac muscle sarcoplasmic reticulum. J Biol Chem. 1990;265(23):13472–13483.
  74. 74. Stoyanovsky D, Murphy T, Anno PR, Kim YM, Salama G. Nitric oxide activates skeletal and cardiac ryanodine receptors. Cell Calcium. 1997;21(1):19–29. DOI: http://dx.doi.org/10.1016/S0143–4160(97)90093–2.
  75. 75. Xu L, Eu JP, Meissner G, Stamler JS. Activation of the cardiac calcium release channel (ryanodine receptor) by poly-S-nitrosylation. Science. 1998;279(5348):234–237. DOI: 10.1126/science.279.5348.234.
  76. 76. Hart JD, Dulhunty AF. Nitric oxide activates or inhibits skeletal muscle ryanodine receptors depending on its concentration, membrane potential and ligand binding. J Membr Biol. 2000;173(3):227–236.
  77. 77. Li N, Zou AP, Ge ZD, Campbell WB, Li PL. Effect of nitric oxide on calcium-induced calcium release in coronary arterial smooth muscle. Gen Pharmacol. 2000;35(1):37–45. DOI: http://dx.doi.org/10.1016/S0306–3623(01)00089–1.
  78. 78. Heunks LM, Machiels HA, Dekhuijzen PN, Prakash YS, Sieck GC. Nitric oxide affects sarcoplasmic calcium release in skeletal myotubes. J Appl Physiol. 1985;91(5):2117–2124.
  79. 79. Zima AV, Blatter LA. Redox regulation of cardiac calcium channels and transporters. Cardiovasc Res. 2006;71(2):310–321. DOI: http://dx.doi.org/10.1016/j.cardiores.2006.02.019.
  80. 80. Zhang Y, Hogg N. The mechanism of transmembrane S-nitrosothiol transport. Proc Natl Acad Sci U S A. 2004;101(21):7891–7896. DOI: http://dx.doi.org/10.1073/pnas.0401167101.
  81. 81. Kakizawa S. Nitric oxide-induced calcium release: activation of type 1 ryanodine receptor, a calcium release channel, through non-enzymatic post-translational modification by nitric oxide. Front Endocrinol (Lausanne). 2013;4:142. DOI: http://dx.doi.org/10.3389%2Ffendo.2013.00142.
  82. 82. Zahradníková A, Minarovic I, Venema RC, Mészáros LG. Inactivation of the cardiac ryanodine receptor calcium release channel by nitric oxide. Cell Calcium. 1997;22(6):447–454.
  83. 83. Ficarro S, Chertihin O, Westbrook VA, White F, Jayes F, Kalab P, Marto JA, Shabanowitz J, Herr JC, Hunt DF, Visconti PE. Phosphoproteome analysis of capacitated human sperm. Evidence of tyrosine phosphorylation of a kinase-anchoring protein 3 and valosin-containing protein/p97 during capacitation. J Biol Chem. 2003;278(13):11579–11589. DOI: http://dx.doi.org/10.1074/jbc.M202325200.
  84. 84. Visconti PE, Johnson LR, Oyaski M, Fornés M, Moss SB, Gerton GL, Kopf GS. Regulation, localization, and anchoring of protein kinase A subunits during mouse sperm capacitation. Dev Biol. 1997;192(2):351–363. DOI: http://dx.doi.org/10.1006/dbio.1997.8768.
  85. 85. Bajpai M, Fiedler SE, Huang Z, Vijayaraghavan S, Olson GE, Livera G, Conti M, Carr DW. AKAP3 selectively binds PDE4A isoforms in bovine spermatozoa. Biol Reprod. 2006;74(1):109–118. DOI: http://dx.doi.org/10.1095/biolreprod.105.043588.
  86. 86. Fisch JD, Behr B, Conti M. Enhancement of motility and acrosome reaction in human spermatozoa: differential activation by type-specific phosphodiesterase inhibitors. Hum Reprod. 1998;13(5):1248–1254. DOI: 10.1093/humrep/13.5.1248.
  87. 87. Vijayaraghavan S, Goueli SA, Davey MP, Carr DW. Protein kinase A-anchoring inhibitor peptides arrest mammalian sperm motility. J Biol Chem. 1997;272(8):4747–4752. DOI: 10.1074/jbc.272.8.4747.
  88. 88. Asquith KL, Baleato RM, McLaughlin EA, Nixon B, Aitken RJ. Tyrosine phosphorylation activates surface chaperones facilitating sperm-zona recognition. J Cell Sci. 2004;117(Pt 16):3645–3657. DOI: http://dx.doi.org/10.1242/jcs.01214.
  89. 89. Nixon B, Asquith KL, John Aitken R. The role of molecular chaperones in mouse sperm-egg interactions. Mol Cell Endocrinol. 2005;240(1–2):1–10. DOI: http://dx.doi.org/10.1016/j.mce.2005.06.004.
  90. 90. Ergur AR, Dokras A, Giraldo JL, Habana A, Kovanci E, Huszar G. Sperm maturity and treatment choice of in vitro fertilization (IVF) or intracytoplasmic sperm injection: diminished sperm HspA2 chaperone levels predict IVF failure. Fertil Steril. 2002;77(5):910–918. DOI: http://dx.doi.org/10.1016/S0015–0282(02)03073-X.
  91. 91. Cedenho AP, Lima SB, Cenedeze MA, Spaine DM, Ortiz V, Oehninger S. Oligozoospermia and heat-shock protein expression in ejaculated spermatozoa. Hum Reprod. 2006;21(7):1791–1794. DOI: http://dx.doi.org/10.1093/humrep/del055.
  92. 92. Guevara I, Iwanejko J, Dembińska-Kieć A, Pankiewicz J, Wanat A, Anna P, Gołabek I, Bartuś S, Malczewska-Malec M, Szczudlik A. Determination of nitrite/nitrate in human biological material by the simple Griess reaction. Clin Chim Acta. 1998;274(2):177–188. DOI: http://dx.doi.org/10.1016/S0009–8981(98)00060–6.

Written By

Florentin-Daniel Staicu and Carmen Matas Parra

Submitted: 18 May 2016 Reviewed: 20 January 2017 Published: 17 May 2017