Mediators of Inflammation

Mediators of Inflammation / 2020 / Article

Research Article | Open Access

Volume 2020 |Article ID 3640450 | https://doi.org/10.1155/2020/3640450

Giulia Collodel, Elena Moretti, Daria Noto, Francesca Iacoponi, Cinzia Signorini, "Fatty Acid Profile and Metabolism Are Related to Human Sperm Parameters and Are Relevant in Idiopathic Infertility and Varicocele", Mediators of Inflammation, vol. 2020, Article ID 3640450, 13 pages, 2020. https://doi.org/10.1155/2020/3640450

Fatty Acid Profile and Metabolism Are Related to Human Sperm Parameters and Are Relevant in Idiopathic Infertility and Varicocele

Academic Editor: Daniela Caccamo
Received27 Apr 2020
Revised18 Aug 2020
Accepted20 Aug 2020
Published31 Aug 2020

Abstract

Objectives. Fatty acids (FA) modulate oxidative stress, reactive oxygen species (ROS) production, and inflammatory processes in spermatogenesis. Methods. The amount of 17 different FAs and the level of F2-isoprostanes (F2-IsoPs) and cytoplasmic phospholipase A2 (cPLA2) were compared and correlated to sperm characteristics; these last ones were evaluated by light and electronic microscopy in varicocele and idiopathic infertile patients. Results. Total n-3 polyunsaturated acids (PUFAs) and docosahexaenoic acid (DHA), one of the n-3 PUFAs, were significantly reduced in idiopathic infertile men compared to controls (). In the whole studied population, oleic acid and total monounsaturated acids (MUFAs) correlated negatively with sperm concentration, progressive motility, normal morphology, vitality, and fertility index and positively with sperm necrosis. Eicosapentaenoic acid (EPA) amount was positively correlated with the percentage of sperm necrosis and cPLA2 level and negatively with sperm concentration. Sperm vitality was negatively correlated with the saturated fatty acids (SFAs). In infertile groups, cPLA2 was negatively correlated with DHA and n-3 PUFAs (both ) and positively with EPA (). In the varicocele group, sperm vitality was negatively correlated with palmitoleic acid and total n-6 PUFAs (); sperm apoptosis was positively correlated with the total SFA percentage (). Conclusions. FA composition in sperm membrane and the metabolism of sperm FAs are interrelated parameters, both relevant in sperm maturation processes and fertility.

1. Introduction

Phospholipids, glycerophospholipids, and cholesterol define the lipid composition of the plasma membranes [1]; in spermatozoa, it is directly related to sperm physiology regulating cellular osmotic balance [2], sperm motility [3], and acrosome reaction and sperm-oocyte fusion [4]. In animals, the lipid sperm composition is described, and it seems to influence external or internal fertilization in fishes [5], seasonal changes in stallions [6], and the ability to survive the cryopreservation process in different species [7].

Fatty acids (FAs) esterified to phospholipids are part of the cell membrane and contribute to the cell membrane structure. As we all know, FAs are classified, according to the presence of double bonds in their chain structure, as saturated FAs (SFAs), in the absence of double bonds, as monounsaturated FAs (MUFAs), when a single double bond is present, and as polyunsaturated FAs (PUFAs) when having two or more double bonds. In a sperm membrane, FA composition, as a different ratio in PUFAs, MUFAs, and SFAs, seems to be a relevant factor in explaining sperm quality. Cell flow and elasticity of plasma membranes are linked to the presence of PUFAs which are particularly abundant in mammalian spermatozoa. In human sperm, docosahexaenoic acid (DHA, C22:6n-3) and palmitic acid (C16:0) are the predominant PUFAs and SFAs [8], respectively. In addition, it has been reported that sperm motility, morphology, and concentration are positively associated with DHA levels [9, 10], whereas low concentrations of DHA have been observed in asthenozoospermic and oligozoospermic men [3, 11]. A negative correlation between increased SFAs [12] or trans FA levels [13] and normal sperm parameters was also described.

As a not secondary issue, the enzymatic, or free radical-induced, PUFA metabolism which influences inflammatory process and susceptibility to lipid peroxidation events in mammalian sperm membrane should be mentioned [4, 14, 15]. In particular, cyclooxygenase (COX) activities on arachidonic acid (ARA) lead to the synthesis of prostaglandins, prostacyclin, and thromboxane A2, whereas ARA metabolism by lipoxygenase (LOX) activities generates leukotrienes [15]. On the other hand, eicosapentaenoic acid (EPA) and DHA are converted, by the same enzymes, COX and LOX, to different series of prostanoids and leukotrienes, respectively. Moreover, specialized proresolving mediators (SPM) with potent anti-inflammatory and immunoregulatory actions are derived from enzymatic oxidation of DHA and EPA [16]. With reference to the free radical-induced PUFA metabolism, isoprostanes (IsoPs) and neuroprostanes originate from nonenzymatic PUFA (i.e., ARA, DHA, and EPA) metabolism and they are known as the secondary products of lipid peroxidation and markers of oxidative damage [17, 18]. Since IsoPs are originally formed in situ, on FA precursors esterified in phospholipids, and subsequently released from membrane phospholipid as free acids by phospholipase action [17], esterified IsoPs quantified from PUFA cell membrane are the specific index of membrane PUFA oxidation.

On PUFA metabolism, phospholipase activity and isoprostane formation are currently described as key factors in influencing male fertility and sperm membrane stability, capacitation, and immaturity [15, 1921].

The purpose of this manuscript was to investigate the membrane FA profile in the sperm of patients with different reproductive pathologies. As a relevant novelty in human male fertility, a close relationship between the FA pattern in a sperm membrane and the metabolism of sperm FA has been shown. We carried out comparisons and correlations between the amounts of 17 different FAs and the levels of F2-isoprostanes (F2-IsoPs) and phospholipase A2 (PLA2) with sperm characteristics. We reported these last ones evaluated by light and electronic microscopy.

2. Materials and Methods

2.1. Patients

From September 2018 to April 2019, we selected 24 infertile patients (aged 30 to 40 years) attending our centre for semen analysis. The duration of infertility was in the range 3–6 years of unprotected sexual intercourses without conception.

The inclusion criteria were as follows: nonazoospermic men with a normal karyotype evaluated by conventional cytogenetic analysis,  kg/m2 and no history of diabetes, radiotherapy, chemotherapy, chronic illness or medication, neither of the use of drugs, and alcohol and dietary supplements. All the individuals showed normal concentrations of follicle stimulating hormone (FSH), luteinizing hormone (LH), and testosterone (T). The patients declared that they were not professionally exposed to pesticides or heavy metal. Subjects with leukocytospermia, genitourinary infection, and heavy smoking habits (>10 cigarettes/day) were excluded. The presence of varicocele was detected by physical examination and scrotal eco-color Doppler analysis performed in all selected patients. The possible presence of clinically asymptomatic genitourinary infections was investigated by a bacteriological analysis. Patients with positive bacteriological cultures were considered infected and excluded consequently.

The patients enrolled for this study were then divided into two groups according to clinical diagnosis: (i)Group I: idiopathic infertility ()(ii)Group II: varicocele ()—where left sided varicocele was found in 10 cases (grade III in three patients, grade II in five patients, and grade I in two patients) and right sided varicocele and bilateral varicocele were diagnosed in one (grade II) and one (grade I-II), respectively

A group of six fertile men (aged 32-39 years) was used as controls. Men fathered at least one child in the last 3 years. They were not affected by infections, hormonal and anatomical problems.

All subjects were informed with respect to this study, and they provided an informed written consent before the inclusion on this research. The study was approved by the Ethics Committee of Azienda Ospedaliera Universitaria Senese, CEAOUS.

2.2. Light Microscopy

Subjects enrolled in this study were asked to abstain from intercourse and masturbation for a period of 3–4 days before the semen collection.

According to the WHO guidelines [22], samples were collected in a bathroom closed to the laboratory, in order to limit the exposure of the semen to fluctuations in temperature and to control the time between collection and analysis. Samples were analyzed after liquefaction for 30 min at 37°C; a normal liquefied semen sample had a homogeneous, grey opalescent appearance. Volume was formerly measured by pipetting the semen into calibrated containers, pH with pH indicator papers in the range 6.0–10.0.

The motility of sperm in a sample volume of 10 μl was evaluated microscopically at 400x magnification. Sperm were classified as progressively motile, locally motile, or immotile. Sperm morphology was assessed by the Papanicolaou (PAP) test modified for spermatozoa. The morphology of 200 spermatozoa was determined under a light microscope at 1,000x magnification using an oil-immersion objective. Sperm vitality was detected using 0.5% eosin Y (CI 45380) in a 0.9% aqueous sodium chloride solution; one hundred stained (dead) cells and unstained (living) cells were scored.

The possible presence of leukocytes was explored by peroxidase stain; a cell/ml was considered out of range and identified as leukocytospermia. A sample from each patient was analyzed. After semen evaluation, all the samples were divided in three aliquots to be used for the different analyses.

2.3. Transmission Electron Microscopy (TEM)

An aliquot was processed for transmission electron microscopy (TEM). Sperm samples were fixed in a cold Karnovsky fixative and maintained at 4°C for 2 h. Then, the semen was washed in 0.1 mol/l cacodylate buffer (pH 7.2) for 12 h, postfixed in 1% buffered osmium tetroxide for 1 h at 4°C, and washed again in 0.1 mol/l cacodylate buffer; the samples were dehydrated in a graded ethanol series and embedded in Epon Araldite. Ultrathin sections were cut with a Supernova ultramicrotome (Reichert Jung, Vienna, Austria), mounted on copper grids, stained with uranyl acetate and lead citrate, and then observed and photographed with a Philips CM12 transmission electron microscope (TEM; Philips Scientifics, Eindhoven, the Netherlands, Centro di Microscopie Elettroniche “Laura Bonzi,” ICCOM, Consiglio Nazionale delle Ricerche (CNR), Via Madonna del Piano 10, Firenze, Italy).

Three hundred sperm sections were analyzed from each sample. TEM data was elaborated using a mathematical formula [23] which provides numerical scores such as fertility index (number of sperm free of structural defects in a semen sample) and percentage of sperm pathologies such as immaturity, apoptosis, and necrosis [24], defined by distinctive ultrastructural characteristics. Immaturity includes the presence of cytoplasmic droplets, altered acrosomes, and roundish or elliptical nuclei with uncondensed chromatin. Marginated chromatin, translucent vacuoles included in cytoplasmic residues, and swollen and badly assembled mitochondria are the ultrastructural indicators of apoptosis, whereas sperm with reacted or absent acrosome, misshaped nuclei with disrupted chromatin, broken plasma membrane, and poor axonemal and periaxonemal cytoskeletal structures are affected by necrosis.

2.4. Analyses of Sperm Membrane Fatty Acid (FA) Composition

An aliquot of each semen sample was centrifuged at 400g for 15 min; then, the supernatant was discarded. Sperm membrane phospholipids were extracted and subsequently converted into fatty acid methyl esters (FAMEs) by a transesterification procedure performed in the presence of a methanol solution of 0.5 M KOH [25]. FAMEs were analyzed by a gas chromatography instrumentation (GC) (Agilent 6850, Milan) equipped with a capillary column (DB23, Agilent, USA) and a flame ionization detector. Hydrogen as carrier gas and FAMEs were identified by comparison with the retention times of authentic molecules [25]. All the determinations were performed by the Lipidomics Laboratory at Lipinutragen Srl, CNR Area della Ricerca di Bologna, Italy.

2.5. Phospholipase A2 (PLA2) Determination

An aliquot of each semen sample was centrifuged at 400 g for 15 min; then, the pellet was discarded. The supernatant was used for cytosolic PLA2 (cPLA2) and F2-isoprostane (F2-IsoP) evaluations.

Cytosolic PLA2 (cPLA2) amounts in seminal plasma samples were determined by an enzyme-linked immunosorbent assay (ELISA, AMSBIO, distributed by D.B.A. Italia). In particular, the determinations were carried out as a competitive immunoassay, where standard samples (known cPLA2 amounts) or seminal samples were added (50 μl) to wells precoated with a monoclonal antibody to cPLA2. cPLA2 is present in the standards, or human samples competed with the biotin-conjugated antigen to bind to the capture antibody. The final revelation was performed using an avidin horseradish peroxidase. The tetrametilbenzidina (TMB) substrate was then added to develop a color changing into yellow after the addition of an acidic stop solution. Spectrometric detection of color intensity at 450 nm allowed the determination of cPLA2 amounts by comparing the optical density of the seminal samples to the standard curve (ranging from 16 ng/ml to 0.5 ng/ml cPLA2 amounts).

2.6. F2-Isoprostane (F2-IsoP) Evaluation

A gas chromatography/negative-ion chemical ionization tandem mass spectrometry (GC/NICI-MS/MS) was applied to detect the amount of total (free plus esterified) F2-IsoPs in seminal plasma. After sample collection, butylated hydroxytoluene (BHT) was added (final concentration 90 μM) and a storage at −80°C was carried out until the assay time. As the first step in F2-IsoP detection, a basic hydrolysis was performed by incubation (45°C, 45 min) with 1 N KOH (1 : 0.5, v:v). At the end of the incubation, HCl 1 N was added (1 : 0.5, v : v). Progressively, in each sample, an internal standard (tetradeuterated derivative of prostaglandin F2α, PGF2α-d4, 500 pg) was added, and two different solid phase extractions (octadecylsilane, C18 cartridge and aminopropyl, NH2 cartridge) were consecutively carried out. The final eluate was derivatized to convert the carboxyl group of F2-IsoPs into pentafluorobenzyl ester and the hydroxyl group into trimethylsilyl ethers [26]. In GC/NICI-MS/MS, 299 and 303 ions, derived from the [M-181] precursor ions of derivatized 15-F2t-IsoP (i.e., 8-iso-PGF2α, the most represented isomer for F2-IsoP measurement) and PGF2α-d4, were, respectively, detected.

2.7. Statistical Analysis

Data was reported as the median and interquartile range (IQR: 75°-25° centile), and all analyses were performed by SPSS v. 26 (IBM SPSS Statistics). The Kruskal-Wallis test was used to compare the varicocele or idiopathic infertility group and fertile men, followed by, when significant, by Dunn’s post hoc test. In order to measure the correlations between the investigated variables, we used Spearman’s Rank Correlation Coefficient. A value < 0.05 (two-tailed) was considered statistically significant.

3. Results

3.1. Semen Characteristics

Semen characteristics and sperm lipid composition of the infertile groups (varicocele or idiopathic infertility) and controls (fertile men) are reported in Table 1. By considering that each sperm sample showed a different number of cells and that in the investigated patients’ groups, different amounts of membrane FAs could be present; the absolute values of each investigated parameters were normalized, i.e., expressed as a ratio, to FA contents and sperm parameters (as sperm/ml, sperm apoptosis, necrosis, and immaturity) in order to make them comparable in the investigated groups (Table 2). Fertile men showed a better semen quality compared to that of infertile groups; idiopathic infertile men had a significant decrease in sperm concentration (), progressive motility (), normal morphology (), vitality (), and fertility index () and an increase in sperm apoptosis () and sperm necrosis () compared to those observed in the control group.


Semen characteristics, sperm fatty acid abundance, and parameters of fatty acid metabolismFertility ()
Group 1
Idiopathic infertility ()
Group 2
Infertile Varicocele ()
Group 3
Kruskal-Wallis
value
Pairwise comparisons
Group 1 vs. group 2Group 1 vs. group 3Group 2 vs. group 3

Volume (ml)4.75 (1.00)3.75 (2.13)3.90 (2.00)0.07
118.50 (116.75)12.10 (51.95)53.35 (100.75)
Total sperm number454.50 (538.95)49.77 (184.65)182.00 (220.50)0.06
Motility (%)34.50 (28.75)17.50 (11.75)27.50 (21.00)
Morphology (%)17.00 (11.25)7.00 (4.75)8.50 (7.50)
Vitality (%)89.50 (14.25)50.00 (20.75)72.50 (20.50)
Apoptosis (A) (%)4.20 (3.18)8.81 (5.20)5.82 (2.07)
Necrosis (N) (%)22.40 (11.87)40.65 (16.11)37.18 (16.65)
Immaturity (I) (%)44.61 (4.53)53.55 (14.02)67.71 (12.95)
Fertility index (FI)2907921.50 (16036247.25)266546.00 (577090.50)837293.00 (1316138.25)
Seminal F2-IsoPs (ng/ml)8.16 (6.48)31.12 (22.33)62.40 (23.08)
Seminal cPLA2 (ng/ml)0.76 (0.30)0.86 (2.79)0.85 (2.23)0.52
Myristic acid (%)0.70 (0.73)1.00 (0.20)1.15 (0.45)0.28
Palmitic acid (%)29.85 (12.08)31.10 (7.95)28.40 (9.45)0.48
Margaritic acid (%)0.35 (0.18)0.40 (0.55)0.35 (0.28)0.27
Stearic acid (%)16.05 (16.70)15.50 (18.13)19.55 (15.05)0.89
Arachidic acid (%)0.15 (0.23)0.50 (0.40)0.20 (0.55)0.14
Total SFAs (%)52.50 (13.68)55.05 (14.20)52.60 (21.95)0.86
Palmitoleic acid (%)0.50 (0.55)0.90 (1.08)0.70 (0.85)0.48
Oleic acid (%)5.90 (9.13)10.45 (7.58)7.20 (3.95)0.07
Vaccenic acid (%)2.45 (1.90)3.20 (2.28)2.70 (1.05)0.49
Total MUFAs (%)8.75 (12.03)15.65 (8.60)10.95 (7.48)0.14
Linoleic acid (%)4.10 (2.75)5.00 (4.43)5.20 (3.83)0.76
Eicosadienoic acid (%)0.85 (0.20)0.80 (0.43)0.75 (0.45)0.75
Eicosatrienoic acid (%)2.85 (3.35)2.75 (2.83)4.70 (3.08)0.11
Arachidonic acid (%)3.00 (2.20)2.65 (2.18)2.85 (1.98)0.80
Total n-6 PUFAs (%)11.80 (6.00)10.35 (6.55)14.65 (9.20)0.21
Eicosapentaenoic acid (%)0.25 (0.23)0.70 (1.95)0.10 (1.23)0.17
Docosapentaenoic acid (%)1.05 (0.70)1.05 (0.58)0.90 (0.28)0.60
Docosahexaenoic acid (%)24.40 (9.90)14.55 (10.78)15.85 (10.88)
Total n-3 PUFAs (%)25.40 (10.38)17.45 (8.60)17.75 (8.85)
Trans 18 : 1 acid (%)0.00 (0.05)0.00 (0.10)0.00 (0.00)0.30
Trans 20 : 1 acid (%)0.00 (0.03)0.00 (0.08)0.00 (0.00)0.86

Volume: volume (ml) of the ejaculate; : sperm concentration in one ml; total sperm number: total number of sperm in the ejaculate; motility: percentage of progressive sperm motility; morphology: percentage of normal sperm morphology assessed with Papanicolaou staining; vitality: percentage of viable sperm by eosin Y; apoptosis: percentage of sperm apoptosis assessed by transmission electron microscopy; necrosis: percentage of sperm necrosis assessed by transmission electron microscopy; immaturity: percentage of sperm immaturity assessed by transmission electron microscopy; fertility index: the number of sperm probably devoid of defects assessed by transmission electron microscopy; F2-IsoPs: F2-isoprostanes; cPLA2: cytoplasmic phospholipase A2; total SFAs: sum of saturated fatty acids; total MUFAs: sum of monounsaturated acids; total n-6 PUFAs: sum of omega 6 polyunsaturated fatty acids; total n-3 PUFAs: sum of omega 3 polyunsaturated fatty acids. All FA amounts were reported as relative percentage concentration (%). Statistics are reported (, , and ).

Normalized seminal parametersFertility ()
Group 1
Idiopathic infertility ()
Group 2
Infertile Varicocele ()
Group 3
Kruskal-Wallis
value
Pairwise comparisons
Group 1 vs. group 2Group 1 vs. group 3Group 2 vs. group 3

EPA/ARA ratio0.05 (0.15)0.30 (0.68)0.10 (0.30)0.11
SFA/MUFA ratio5.55 (6.95)3.25 (3.15)4.80 (3.08)0.39
n-6/n-3 PUFA ratio0.45 (0.23)0.85 (0.48)0.75 (0.73)
F2-IsoP/ARA ratio2.97 (4.36)10.84 (21.38)27.07 (21.41)
F2-IsoP/sperm/ml ratio0.06 (0.10)1.01 (3.31)1.05 (2.47)
N/ARA ratio7.82 (6.97)13.90 (8.10)12.16 (14.71)
A/ARA ratio1.66 (1.50)3.18 (7.01)2.53 (2.56)0.13
N/n-6 PUFA ratio1.87 (2.44)3.94 (1.60)2.27 (1.45)nsnsns
A/n-6 PUFA ratio0.40 (0.39)0.77 (0.69)0.37 (0.35)
I/n-6 PUFA ratio3.85 (2.90)5.17 (3.44)4.07 (2.51)0.82
N/n-3 PUFA ratio0.86 (0.88)2.83 (1.64)1.58 (2.01)
A/n-3 PUFA ratio0.20 (0.17)0.55 (0.29)0.32 (0.23)
I/n-3 PUFA ratio1.71 (0.99)3.06 (4.40)3.72 (0.80)

EPA: eicosapentaenoic; ARA: arachidonic acid; SFAs: saturated fatty acids; MUFAs: monounsaturated fatty acids; PUFAs: polyunsaturated fatty acids; F2-IsoPs: F2-isoprostanes; N: sperm necrosis; A: sperm apoptosis; I: sperm immaturity. Statistics are reported (, , and ).

Fertility index was reduced also in the varicocele group () compared to control. Moreover, in the varicocele group, sperm immaturity percentage was significantly increased compared to that measured in the idiopathic infertility () and fertility () groups.

3.2. Seminal and Sperm Lipid Composition

The medians and interquartile ranges of seminal F2-IsoP and cPLA2 levels and investigated FAs are shown in Table 1.

In the varicocele group, higher levels of F2-IsoPs were detected with respect to those of the idiopathic infertility () and fertility () groups. Seminal cPLA2 level did not show differences among the investigated groups.

Regarding FA values for SFAs, MUFAs, and n-6 PUFAs, no statistical difference was observed between the control and the patient groups (Table 1).

Total n-3 PUFAs and DHA, one of the n-3 PUFAs, were significantly reduced in idiopathic infertile men compared to controls ().

Among the evaluated ratios (Table 2), a significant increase was detected in n-6/n-3 PUFA (), F2-IsoP/sperm/ml (), and immaturity (I)/n-3 PUFA () ratios in both infertile groups as compared to controls. Necrosis (N)/ARA () and N/n-3 PUFAs () increased in idiopathic the infertile group vs. fertile controls. The idiopathic infertile group also showed both a significant increase for the sperm apoptosis (A)/n-3 PUFA ratio, with respect to fertility () and varicocele groups (), and an increase for A/n-6 PUFA ratio as compared to the varicocele group ().

3.3. Relationships between Semen Characteristics and Sperm Fatty Acid Profile or Metabolism

The relationships among the investigated variables in the whole population are shown in Table 3. Oleic acid and total MUFAs correlated negatively with sperm concentration, total sperm number, sperm progressive motility, normal morphology, vitality, and fertility index and positively with sperm necrosis (Figure 1); oleic acid showed also positive correlation with cPLA2 levels (). Sperm concentration was negatively correlated with margaritic acid () and EPA () content. EPA amount was positively correlated with the percentage of sperm necrosis () and cPLA2 level (). Sperm vitality was negatively correlated with the SFAs palmitic acid (), whereas arachidic acid positively with sperm immaturity ().


Sperm fatty acidsVariables positively correlated to the fatty acidCorrelation coefficient (Rho) and statistical significance ()Variables negatively correlated to the fatty acidCorrelation coefficient (Rho) and statistical significance ()
Rho valueRho value

Palmitic acidSperm vitality- 0.362<0.05
Margaritic acidSperm concentration- 0.422<0.05
DHA- 0.505<0.01
Total n-3- 0.434<0.05
Arachidic acidSperm immaturity0.428<0.05
Oleic acidSperm necrosis0.655<0.01Sperm concentration- 0.561<0.01
cPLA20.414<0.05Total sperm number- 0.377<0.05
Total MUFAs0.968<0.01Sperm motility- 0.599<0.01
Sperm morphology- 0.401<0.05
Sperm vitality- 0.493<0.01
Fertility index- 0.477<0.01
DHA- 0.578 - 0.515<0.01
Total n-3<0.01
Total MUFAsSperm necrosis0.597<0.01Sperm concentration- 0.501<0.01
Oleic acid0.968<0.01Sperm motility- 0.573<0.01
Sperm morphology- 0.372<0.01
Sperm vitality- 0.462<0.05
Fertility index- 0.415<0.05
DHA- 0.483<0.01
Total n-3- 0.414<0.05
EPASperm necrosis0.446<0.05Sperm concentration- 0.479<0.01
Arachidonic acid0.471<0.05Total sperm number- 0.398<0.05
Oleic acid0.618<0.01DHA- 0.647<0.01
cPLA20.523<0.01Total n-3- 0.506<0.01
Total MUFAs0.559<0.01
Trans 18 : 10.603<0.01
DHASperm concentration0.405<0.05Sperm necrosis- 0.576<0.01
Total sperm number0.502<0.01cPLA2- 0.518<0.01
Sperm morphology0.580<0.01Margaritic acid- 0.505<0.01
Sperm vitality0.517<0.01Oleic acid- 0.578<0.01
Fertility index0.612<0.01Total MUFAs- 0.483<0.01
Total n-30.949<0.01EPA- 0.647<0.01
Trans 18 : 1- 0.437<0.05
Total n-3Sperm concentration0.475<0.01Sperm necrosis- 0.478<0.01
Total sperm number0.391<0.05cPLA2- 0.418<0.05
Sperm morphology0.364<0.05Margaritic acid- 0.434<0.05
Sperm vitality0.470<0.01Oleic acid- 0.515<0.01
Fertility index0.579<0.01Total MUFAs- 0.414<0.05
DHA0.949<0.01EPA- 0.506<0.01
Trans 18 : 1 acidcPLA20.371<0.05DHA- 0.437<0.05
EPA0.603<0.01Total sperm number- 0.551<0.01

Sperm concentration: number of ; total sperm number: total number of sperm in the ejaculate; sperm motility: percentage of progressive sperm motility; sperm morphology: percentage of normal sperm morphology assessed with Papanicolaou staining; sperm vitality: percentage of viable sperm by eosin Y; sperm necrosis: percentage of sperm necrosis assessed by transmission electron microscopy; sperm immaturity: percentage of sperm immaturity assessed by transmission electron microscopy; fertility index: the number of sperm probably devoid of defects assessed by transmission electron microscopy; cPLA2: cytoplasmic phospholipase seminal levels (ng/ml); total MUFAs: sum of monounsaturated acids; EPA: eicosapentaenoic acid; DHA: docosahexaenoic acid; total n-3: sum of omega 3 fatty acids. All FA amounts were reported as relative percentage concentration (%).

DHA and total n-3 FAs were positively correlated with sperm concentration, total sperm number, sperm normal morphology, vitality, and fertility index and negatively with sperm necrosis and cPLA2.

The amounts of cPLA2 were also positively correlated to trans 18 : 1 acid level (). These results were supported by the relationships between lipids; miristic acid, oleic acid, arachidic acid, EPA, and total MUFAs were positively correlated with each other and negatively correlated with DHA and total n-3 PUFAs. Levels of stearic, palmitoleic, vaccenic, linoleic, eicosadienoic, eicosatrienoic, arachidonic, docosapentaenoic, and trans 20 : 1 acids were not correlated to seminal parameters.

In Table 4, the significant correlations between the calculated ratios and sperm characteristics are reported. The EPA/ARA, n-6/n-3 PUFAs, F2-IsoPs/ARA, F2-IsoPs/sperm/ml, N/ARA, A/ARA, N/n-6, N/n-3, A/n-3, and I/n-3 ratios, except for SFA/MUFA ratio, were negatively correlated with sperm parameters. In addition, EPA/ARA, n-6/n-3 PUFA, and N/n-3 PUFA ratios had positive correlations with cPLA2.


Normalized parametersVariables positively correlated to normalized parametersCorrelation coefficient (Rho) and statistical significance ()Variables negatively correlated to normalized parametersCorrelation coefficient (Rho) and statistical significance ()
Rho valueRho value

EPA/ARASperm necrosis0.482<0.01Sperm concentration- 0.425<0.05
cPLA20.512<0.01Total sperm number- 0.410<0.05
N/ARA0.526<0.01Sperm vitality- 0.458<0.05
N/n-60.410<0.05SFA/MUFA- 0.405<0.05
A/n-30.384<0.05
N/n-30.491<0.01
SFA/MUFASperm concentration0.375<0.05Sperm necrosis- 0.504<0.01
Sperm motility0.516<0.05EPA/ARA- 0.405<0.05
n-6/n-3- 0.511<0.01
n-6/n-3Sperm necrosis0.599<0.01Sperm concentration- 0.602<0.01
cPLA20.519<0.01Total sperm number- 0.464<0.05
F2-IsoPs/sperm concentration0.616<0.05Sperm motility- 0.551<0.01
I/n-30.676<0.01Sperm morphology- 0.540<0.01
N/n-30.741<0.01Fertility index- 0.675<0.01
SFA/MUFA- 0.511<0.01
F2-IsoPs/sperm concentrationSperm necrosis0.412<0.01Sperm concentration- 0.833<0.01
Sperm immaturity0.368<0.05Sperm morphology- 0.617<0.05
F2-IsoPs0.370<0.05Sperm vitality- 0.370<0.05
n-6/n-30.616<0.05Fertility index- 0.643<0.01
I/n-30.558<0.01
N/n-30.523<0.01
F2-IsoPs/ARASperm immaturity0.522<0.01Sperm apoptosis- 0.499<0.01
F2-IsoPs0.854<0.01
N/ARA0.519<0.01
A/ARASperm apoptosis0.675<0.01Sperm vitality- 0.397<0.05
F2-IsoPs0.499<0.01
N/ARA0.677<0.01
N/n-60.392<0.05
N/ARASperm necrosis0.487<0.01Sperm vitality- 0.625<0.01
EPA/ARA0.526<0.01
F2-IsoPs/ARA0.519<0.01
A/ARA0.677<0.01
N/n-60.578<0.01
N/n-30.621<0.01
N/n-6Sperm necrosis0.557<0.01Sperm vitality- 0.638<0.01
EPA/ARA0.410<0.05
A/ARA0.392<0.05
N/ARA0.578<0.01
N/n-30.481<0.01
I/n-3Sperm necrosis0.411<0.05Sperm concentration- 0.447<0.05
Sperm immaturity0.530<0.01Total sperm number- 0.390<0.05
n-6/n-30.676<0.01Sperm morphology- 0.443<0.05
F2-IsoPs/sperm concentration0.558<0.01Sperm vitality- 0.389<0.05
N/n-30.745<0.01Fertility index- 0.507<0.01
A/n-3Sperm necrosis0.413<0.05Sperm concentration- 0.415<0.05
Sperm apoptosis0.666<0.05Sperm morphology- 0.444<0.01
EPA/ARA0.384<0.05Sperm vitality- 0.611<0.05
n-6/n-30.408<0.05Fertility index- 0.611<0.01
F2-IsoPs/sperm concentration0.376<0.05
A/ARA0.475<0.05
N/ARA0.362<0.01
N/n-60.523<0.01
I/n-30.570<0.01
N/n-3Sperm necrosis0.829<0.01Sperm concentration- 0.594<0.01
cPLA20.449<0.05Total sperm number- 0.458<0.01
EPA/ARA0.491<0.01Sperm motility- 0.468<0.01
n-6/n-30.741<0.01Sperm morphology- 0.467<0.01
F2-IsoPs/sperm concentration0.523<0.01Sperm vitality- 0.643<0.01
N/ARA0.621<0.01Fertility index- 0.692<0.01
N/n-60.481<0.01
I/n-30.745<0.01
A/n-30.653<0.01

Sperm concentration: number of ; total sperm number: total number of sperm in the ejaculate; sperm motility: percentage of progressive sperm motility; sperm morphology: percentage of normal sperm morphology assessed with Papanicolaou staining; sperm vitality: percentage of viable sperm; sperm apoptosis: percentage of sperm apoptosis assessed by transmission electron microscopy; sperm necrosis: percentage of sperm necrosis assessed by transmission electron microscopy; sperm immaturity: percentage of sperm immaturity assessed by transmission electron microscopy; fertility index: the number of sperm probably devoid of defects assessed by transmission electron microscopy; F2-IsoPs: F2-isoprostane seminal levels (ng/ml); cPLA2: phospholipase seminal levels (ng/ml); EPA: eicosapentaenoic acid; A: sperm apoptosis; ARA: arachidonic acid; N: sperm necrosis; SFAs: saturated fatty acids; MUFAs: monounsaturated fatty acids; n-3: sum of omega 3 FAs; n-6: sum of omega 6 FAs. All FA amounts were reported as relative percentage concentration (%).

Finally, the correlations were performed also in the three analyzed groups (data not shown). In fertile men, palmitoleic acid resulted to be negatively correlated with sperm concentration () and oleic and vaccenic acids with sperm normal morphology and fertility index (both ). Levels of cPLA2 were positively correlated with stearic and margaritic acids (). The idiopathic infertility group showed negative correlations between oleic and trans 18 : 1 acids with sperm concentration () and progressive motility () and positive correlations between ARA and EPA with sperm necrosis (both ). Moreover, cPLA2 was negatively correlated with DHA and n-3 PUFAs () and positively with EPA (). In varicocele, cPLA2 was negatively correlated with DHA and n-3 PUFAs () and positively with EPA (), trans 20: 1 acids and n-6/n-3 PUFA (), and EPA/ARA ratios (). In the same group, sperm vitality was negatively correlated with palmitoleic acid and total n-6 (); sperm apoptosis was positively correlated with the total SFA percentage ().

4. Discussion

Sperm quality and fertility capability have been related to sperm membrane FAs [10, 15]. In our study, this topic was explored by assessing specific semen features related to the sperm pathologies and fertility index [24] and FA oxidation and metabolism. In particular, sperm parameters were evaluated by light microscopy and sperm ultrastructure was analyzed by TEM. The relationships of sperm characteristics with sperm FA profile, seminal F2-IsoP, and cPLA2 levels in two groups of infertile patients (idiopathic infertility and varicocele) compared to fertile men were carried out.

As single molecules or as components of molecules, FAs show multiple biological functions involved from the cell membrane composition to energy suppliers and signaling molecules.

The seminal FA lipidome and its connections with sperm characteristics have been investigated in fertile and infertile men [10], and it was suggested that FA profiling may indicate markers of semen quality. Previously, sperm FA content was explored in sperm from patients with abnormal seminal conditions [3, 9, 27] showing that different percentage of FAs was correlated with seminal parameters.

The study of FA profile may individuate candidate markers of semen quality and suggest therapeutic treatments using appropriate FA supplementation [8, 10, 28]. Our study indicated that in the whole fertile and infertile population, oleic acid, total MUFAs, palmitic acid, and cPLA2 were linked to elevated levels of sperm necrosis and a reduction of fertility index in addition to a decrease in sperm concentration, progressive motility, normal morphology, and vitality. On the contrary, DHA and total n-3 content appeared strongly positively correlated to a good sperm quality in the total population and in the idiopathic infertile group.

Many studies reported a high concentration of DHA in the spermatozoa of normozoospermic subjects [3, 10, 27, 29]. Recently, it was observed that total n-3 PUFA of normozoospermic semen samples was significantly higher than those from oligozoospermic, asthenozoospermic, and oligoasthenozoospermic individuals [30].

In this study, a relevant characteristic is represented by the fertility index. The fertility index is the number of sperm devoid of defects obtained by TEM evaluation mathematically elaborated [23, 24]. The correlation of the fertility index with the FA content indicates that an abnormal FA metabolism causes spermatogenic dysfunction and consequently may influence male fertility.

The mathematical elaboration of TEM data calculates also the percentage of sperm pathologies such as sperm necrosis, apoptosis, and immaturity in a semen sample [23, 24]. In particular, necrotic sperm are characterized by broken plasma membranes, disrupted chromatin, altered mitochondria, and their percentage increases in the presence of an inflammatory status. It is reported that the membrane destabilization during necrosis is mediated by factors, such as acid-sphingomyelinase, PLA2, and calpains. In our research, cPLA2 level and sperm necrosis are negatively correlated with DHA and n-3 concentrations suggesting that high levels of cPLA2 are linked to an altered sperm and seminal condition. On this point, the membrane destabilization during necrosis is mediated by PLA2 [31].

We detected that sperm necrosis and cPLA2 were positively linked to palmitoleic and margaritic acids, respectively, and positively correlated to sperm vitality and concentration. This data agrees with that of other authors who detected elevated levels of palmitic, stearic, oleic, linoleic, and arachidonic acids in spermatozoa from patients with altered sperm parameters compared to normozoospermic subjects [32]. The amounts of cPLA2 were also negatively linked to normal sperm morphology, and they were increased in infertile varicocele patients [20, 21]. Accordingly, it is well known that cPLA2 is involved in inflammatory pathologies and its relationship with oxidative stress and phospholipid impairment has been previously described [33]. The role of cPLA2 was also investigated in a drug-induced reproductive impairment model in mice [34], considering that cPLA2 influences reproduction, as well as other physiological and pathological processes, by regulating production of prostaglandins, and other lipid mediators from PUFAs [35].

Related to the link between cPLA2 and PUFAs, our results showed some significant correlations. The opposite relationships of EPA and DHA with cPLA2 are in line with the results showed in depressive disorders where EPA, but not DHA, significantly increased cPLA2 gene expression [36]. Since cPLA2, besides COX 2, is a key enzyme of the PUFA metabolism, different biological pathways of EPA and DHA could be hypothesized in male infertility, as already reported in other human diseases [36].

Another parameter considered in our study was the level of F2-IsoPs, lipid mediators produced from ARA and released in biological fluid by PLA2 activity. In varicocele patients, the amount of F2-IsoPs was already related to sperm immaturity suggesting an influence of ARA in sperm maturation [19]. This data was confirmed, and the relevance of F2-IsoPs in infertility was also reinforced by considering that similar significant data are showed for both F2-IsoP absolute value and F2-IsoP levels normalized to the ARA content. Moreover, in the infertile varicocele group, palmitoleic acid and total n-6 PUFAs influenced sperm vitality and an increase of SFAs was linked to sperm apoptosis, another sperm pathology evaluated by TEM. Previously, the measurement of SFAs in seminal plasma showed that sperm concentration and motility were negatively correlated with stearic acid and elaidic acid [8, 10, 37]. Recently, in idiopathic infertility patients, a significant positive relationship between F4-Neuroprostane level and the percentage of sperm necrosis was observed [38].

An increase of the ratio n-6/n-3 PUFAs was detected in both infertile groups compared to fertile men; a previous study reported a lower seminal n-6/n-3 PUFA ratio in fertile men compared to the infertile ones, probably due to a significantly high amount of total n-3 PUFAs [39]. Safarinejad et al. [40] found higher levels of n-6 PUFAs but lower levels of n-3 PUFAs in the spermatozoa of infertile compared to fertile men.

Interestingly, the ratio EPA/ARA seems to be an index of poor sperm quality by confirming the relevance of an adequate balancing between n-3 and n-6 PUFAs in male fertility. On this issue, a reduced EPA amount with respect to ARA content is influencing inflammatory response [41], which is an event related to male infertility [42]. Considering the SFA/MUFA ratio, it appears to be significantly related to a good quality of sperm, assessed as progressive motility and concentration. This result appears to be linked to the greater presence of FAs without double bonds in their structure (i.e., SFAs), which are less vulnerable to oxidative damage than the FAs containing one double bond (C=C) in their aliphatic chain (i.e., MUFAs) [43]. Accordingly, it is known that the formation of FA radicals increases with increasing unsaturation [44].

The deep knowledge of sperm FA profiles could suggest on personalized nutraceutical treatments to improve male reproduction. Many studies described a semen improvement after n-3 PUFA supplementation in humans [28]; recently, it has been reported that supplemental dietary n-3 PUFAs resulted in an enrichment of membrane FA in rabbit sperm and testes [45].

Although the population investigated in the study can be considered not particularly wide, it is still enough for statistical processing. Moreover, it is important to underline that the strict selection of patients and the multiple investigations carried out on the same seminal sample are not easy to obtain and we believe they can make our study reliable.

5. Conclusions

In conclusion, FA composition in sperm membrane and the metabolism of sperm FAs are interrelated parameters, both relevant in sperm maturation processes and fertility.

Our future focus will be to investigate other pathological conditions affecting male fertility and to apply this same protocol in subjects on a controlled diet to explore the effect of dietary FA content in the sperm maturation process.

Data Availability

The data used to support the findings of this study is available from the corresponding author upon request.

Conflicts of Interest

The authors declare that they have no conflicts of interest.

Authors’ Contributions

All the authors gave substantial contributions to research design or drafting the paper or revising it critically and approval of the submitted version. Giulia Collodel, in particular, is responsible for the paper writing, research design, patient selection, TEM analysis, and data interpretation. Elena Moretti, in particular, is assigned to cPLA2 determination and TEM analysis, research design, and data interpretation. Francesca Iacoponi, in particular, contributed to the statistical analysis. Daria Noto, in particular, is responsible for the spermiograms, cPLA2 determination and TEM analysis, and data interpretation. Cinzia Signorini, in particular, did the research design, paper writing, lipid determinations, F2-isoprostane evaluation, and data interpretation.

Acknowledgments

This work was supported by the Plan of Research Support 2019 (PSR 2019) of the Department of Molecular Medicine and Development, University of Siena.

References

  1. D. E. Vance and J. E. Vance, Biochemistry of Lipids, Lipoproteins and Membranes, Elsevier, New York, 2008.
  2. F. M. Flesch and B. M. Gadella, “Dynamics of the mammalian sperm plasma membrane in the process of fertilization,” Biochimica et Biophysica Acta, vol. 1469, no. 3, pp. 197–235, 2000. View at: Publisher Site | Google Scholar
  3. Y. Aksoy, H. Aksoy, K. Altinkaynak, H. R. Aydin, and A. Ozkan, “Sperm fatty acid composition in subfertile men,” Prostaglandins Leukotrienes and Essential Fatty Acids, vol. 75, no. 2, pp. 75–79, 2006. View at: Publisher Site | Google Scholar
  4. J. A. Tapia, B. Macias-Garcia, A. Miro-Moran et al., “The membrane of the mammalian spermatozoa: much more than an inert envelope,” Reproduction Domestic Animals, vol. 47, no. 3, pp. 65–75, 2012. View at: Publisher Site | Google Scholar
  5. K. M. Engel, V. Dzyuba, A. Ninhaus-Silveira et al., “Sperm lipid composition in early diverged fish species: internal vs. external mode of fertilization,” Biomolecules, vol. 10, no. 2, p. 172, 2020. View at: Publisher Site | Google Scholar
  6. C. Aurich, C. Ortega Ferrusola, F. J. Peña Vega, N. Schrammel, D. Morcuende, and J. Aurich, “Seasonal changes in the sperm fatty acid composition of Shetland pony stallions,” Theriogenology, vol. 107, pp. 149–153, 2018. View at: Publisher Site | Google Scholar
  7. B. M. García, L. G. Fernández, C. O. Ferrusola et al., “Membrane lipids of the stallion spermatozoon in relation to sperm quality and susceptibility to lipid peroxidation,” Reproduction Domestic Animals, vol. 46, no. 1, pp. 141–148, 2011. View at: Publisher Site | Google Scholar
  8. V. Esmaeili, A. H. Shahverdi, M. H. Moghadasian, and A. R. Alizadeh, “Dietary fatty acids affect semen quality: a review,” Andrology, vol. 3, no. 3, pp. 450–461, 2015. View at: Publisher Site | Google Scholar
  9. N. M. Gulaya, V. M. Margitich, N. M. Govseeva, V. M. Klimashevsky, I. I. Gorpynchenko, and M. I. Boyko, “Phospholipid composition of human sperm and seminal plasma in relation to sperm fertility,” Archives of Andrology, vol. 46, no. 3, pp. 169–175, 2001. View at: Publisher Site | Google Scholar
  10. C. Zerbinati, L. Caponecchia, R. Rago et al., “Fatty acids profiling reveals potential candidate markers of semen quality,” Andrology, vol. 4, no. 6, pp. 1094–1101, 2016. View at: Publisher Site | Google Scholar
  11. J. A. Conquer, J. B. Martin, I. Tummon, L. Watson, and F. Tekpetey, “Fatty acid analysis of blood serum, seminal plasma, and spermatozoa of normozoospermic vs. asthernozoospermic males,” Lipids, vol. 34, no. 8, pp. 793–799, 1999. View at: Publisher Site | Google Scholar
  12. S. R. De Vriese, M. Dhont, and A. B. Christophe, “FA composition of cholesteryl esters and phospholipids in maternal plasma during pregnancy and at delivery and in cord plasma at birth,” Lipids, vol. 38, no. 1, pp. 1–7, 2003. View at: Publisher Site | Google Scholar
  13. J. E. Chavarro, L. Mínguez-Alarcón, J. Mendiola, A. Cutillas-Tolín, J. J. López-Espín, and A. M. Torres-Cantero, “Trans fatty acid intake is inversely related to total sperm count in young healthy men,” Human Reproduction, vol. 29, no. 3, pp. 429–440, 2014. View at: Publisher Site | Google Scholar
  14. J. K. Innes and P. C. Calder, “Omega-6 fatty acids and inflammation,” Prostaglandins Leukotrienes and Essential Fatty Acids, vol. 132, pp. 41–48, 2018. View at: Publisher Site | Google Scholar
  15. G. Collodel, C. Castellini, J. C.-Y. Lee, and C. Signorini, “Relevance of fatty acids to sperm maturation and quality,” Oxidative Medicine and Cellular Longevity, vol. 2020, Article ID 7038124, 14 pages, 2020. View at: Publisher Site | Google Scholar
  16. C. N. Serhan, C. B. Clish, J. Brannon, S. P. Colgan, N. Chiang, and K. Gronert, “Novel functional sets of lipid-derived mediators with antiinflammatory actions generated from omega-3 fatty acids via cyclooxygenase 2–nonsteroidal antiinflammatory drugs and transcellular processing,” The Journal of Experimental Medicine, vol. 192, no. 8, pp. 1197–1204, 2000. View at: Publisher Site | Google Scholar
  17. G. L. Milne, Q. Dai, and L. J. Roberts II, “The isoprostanes—25 years later,” Biochimica et Biophysica Acta, vol. 1851, no. 4, pp. 433–445, 2015. View at: Publisher Site | Google Scholar
  18. J. M. Galano, Y. Y. Lee, C. Oger et al., “Isoprostanes, neuroprostanes and phytoprostanes: an overview of 25years of research in chemistry and biology,” Progress in Lipid Research, vol. 68, pp. 83–108, 2017. View at: Publisher Site | Google Scholar
  19. G. Collodel, E. Moretti, M. Longini, N. A. Pascarelli, and C. Signorini, “Increased F2-isoprostane levels in semen and Immunolocalization of the 8-Iso prostaglandin F2αin spermatozoa from infertile patients with varicocele,” Oxidative Medicine and Cellular Longevity, vol. 2018, Article ID 7508014, 9 pages, 2018. View at: Publisher Site | Google Scholar
  20. E. Moretti, G. Collodel, M. C. Salvatici, G. Belmonte, and C. Signorini, “New insights into sperm with total globozoospermia: increased fatty acid oxidation and centrin1 alteration,” Systems Biology in Reproductive Medicine, vol. 65, no. 5, pp. 390–399, 2019. View at: Publisher Site | Google Scholar
  21. G. Collodel, C. Castellini, F. Iacoponi, D. Noto, and C. Signorini, “Cytosolic phospholipase A2 and F2 isoprostanes are involved in semen quality and human infertility-a study on leucocytospermia, varicocele and idiopathic infertility,” Andrologia, vol. 52, no. 2, article e13465, 2020. View at: Publisher Site | Google Scholar
  22. World Health Organization, WHO Laboratory Manual for the Examination and Processing of Human Semen, WHO Press, Geneva (Switzerland), 5th edition, 2010.
  23. B. Baccetti, G. Bernieri, A. G. Burrini et al., “Notulae seminologicae. 5. Mathematical evaluation of interdependent submicroscopic sperm alterations,” Journal of Andrology, vol. 16, no. 4, pp. 356–371, 1995. View at: Google Scholar
  24. G. Collodel and E. Moretti, “Morphology and meiotic segregation in spermatozoa from men of proven fertility,” Journal of Andrology, vol. 29, no. 1, pp. 106–114, 2007. View at: Publisher Site | Google Scholar
  25. C. Signorini, C. De Felice, S. Leoncini et al., “Altered erythrocyte membrane fatty acid profile in typical Rett syndrome: effects of omega-3 polyunsaturated fatty acid supplementation,” Prostaglandins Leukotrienes and Essential Fatty Acids, vol. 91, no. 5, pp. 183–193, 2014. View at: Publisher Site | Google Scholar
  26. C. Signorini, C. De Felice, T. Durand et al., “Relevance of 4-F4t-neuroprostane and 10-F4t-neuroprostane to neurological diseases,” Free Radical Biology and Medicine, vol. 115, pp. 278–287, 2018. View at: Publisher Site | Google Scholar
  27. T. H. Tavilani, M. Doosti, and H. Saeidi, “Malondialdehyde levels in sperm and seminal plasma of asthenozoospermic and its relationship with semen parameters,” Clinica Chimica Acta, vol. 356, no. 1-2, pp. 199–203, 2005. View at: Publisher Site | Google Scholar
  28. A. L. Falsig, C. S. Gleerup, and U. B. Knudsen, “The influence of omega-3 fatty acids on semen quality markers: a systematic PRISMA review,” Andrology, vol. 7, no. 6, pp. 794–803, 2019. View at: Publisher Site | Google Scholar
  29. L. X. Tang, D. J. Yuan, Q. L. Wang et al., “Association of decreased spermatozoa omega-3 fatty acid levels and increased oxidative DNA damage with varicocele in infertile men: a case control study,” Reproduction, Fertility and Development, vol. 28, no. 5, pp. 648–654, 2016. View at: Publisher Site | Google Scholar
  30. A. Lass and A. Belluzzi, “Omega-3 polyunsaturated fatty acids and IVF treatment,” Reproductive Biomedicine Online, vol. 38, no. 1, pp. 95–99, 2019. View at: Publisher Site | Google Scholar
  31. N. Vanlangenakker, T. Vanden Berghe, D. V. Krysko, N. Festjens, and P. Vandenabeele, “Molecular mechanisms and pathophysiology of necrotic cell death,” Current Molecular Medicine, vol. 8, no. 3, pp. 207–220, 2008. View at: Publisher Site | Google Scholar
  32. A. Khosrowbeygi and N. Zarghami, “Fatty acid composition of human spermatozoa and seminal plasma levels of oxidative stress biomarkers in subfertile males,” Prostaglandins, Leukotrienes and Essential Fatty Acids, vol. 77, no. 2, pp. 117–121, 2007. View at: Publisher Site | Google Scholar
  33. H. Qasem, L. Al-Ayadhi, H. Al Dera, and A. El-Ansary, “Increase of cytosolic phospholipase A2 as hydrolytic enzyme of phospholipids and autism cognitive, social and sensory dysfunction severity,” Lipids in Health and Disease, vol. 16, no. 1, p. 117, 2017. View at: Publisher Site | Google Scholar
  34. N. Fujisawa, W. Yoshioka, H. Yanagisawa, and C. Tohyama, “Roles of cytosolic phospholipase A2α in reproductive and systemic toxicities in 2,3,7,8-tetrachlorodibenzo-p-dioxin-exposed mice,” Archives of Toxicology, vol. 92, no. 2, pp. 789–801, 2018. View at: Publisher Site | Google Scholar
  35. C. C. Leslie, “Cytosolic phospholipase A2: physiological function and role in disease,” Journal of Lipid Research, vol. 56, no. 8, pp. 1386–1402, 2015. View at: Publisher Site | Google Scholar
  36. K.-P. Su, H.-T. Yang, J. P.-C. Chang et al., “Eicosapentaenoic and docosahexaenoic acids have different effects on peripheral phospholipase A2 gene expressions in acute depressed patients,” Progress in Neuropsychopharmacology and Biological Psychiatry, vol. 80, Part C, pp. 227–233, 2018. View at: Publisher Site | Google Scholar
  37. J. A. Attaman, T. L. Toth, J. Furtado, H. Campos, R. Hauser, and J. E. Chavarro, “Dietary fat and semen quality among men attending a fertility clinic,” Human Reproduction, vol. 27, no. 5, pp. 1466–1474, 2012. View at: Publisher Site | Google Scholar
  38. M. Longini, E. Moretti, C. Signorini, D. Noto, F. Iacoponi, and G. Collodel, “Relevance of seminal F2-dihomo-IsoPs, F2-IsoPs and F4-NeuroPs in idiopathic infertility and varicocele,” Prostaglandins and Other Lipid Mediators, vol. 149, p. 106448, 2020. View at: Publisher Site | Google Scholar
  39. J. C. Martínez-Soto, J. Landeras, and J. Gadea, “Spermatozoa and seminal plasma fatty acids as predictors of cryopreservation success,” Andrology, vol. 1, no. 3, pp. 365–375, 2013. View at: Publisher Site | Google Scholar
  40. M. R. Safarinejad, S. Y. Hosseini, F. Dadkhah, and M. A. Asgari, “Relationship of omega-3 and omega-6 fatty acids with semen characteristics, and anti-oxidant status of seminal plasma: a comparison between fertile and infertile men,” Clinical Nutrition, vol. 29, no. 1, pp. 100–105, 2010. View at: Publisher Site | Google Scholar
  41. J. D. Ramakers, R. P. Mensink, G. Schaart, and J. Plat, “Arachidonic acid but not eicosapentaenoic acid (EPA) and oleic acid activates NF-κB and elevates ICAM-1 expression in Caco-2 cells,” Lipids, vol. 42, no. 8, pp. 687–698, 2007. View at: Publisher Site | Google Scholar
  42. A. Agarwal, M. Rana, E. Qiu, H. AlBunni, A. D. Bui, and R. Henkel, “Role of oxidative stress, infection and inflammation in male infertility,” Andrologia, vol. 50, no. 11, article e13126, 2018. View at: Publisher Site | Google Scholar
  43. L. T. Rael, G. W. Thomas, M. L. Craun, C. G. Curtis, R. Bar-Or, and D. Bar-Or, “Lipid peroxidation and the thiobarbituric acid assay: standardization of the assay when using saturated and unsaturated fatty acids,” Journal of Biochemistry and Molecular Biology, vol. 37, no. 6, pp. 749–752, 2004. View at: Publisher Site | Google Scholar
  44. J. M. Yun and J. Surh, “Fatty acid composition as a predictor for the oxidation stability of Korean vegetable oils with or without induced oxidative stress,” Preventive Nutrition and Food Science, vol. 17, no. 2, pp. 158–165, 2012. View at: Publisher Site | Google Scholar
  45. C. Castellini, S. Mattioli, C. Signorini et al., “Effect of Dietaryn‐3Source on rabbit male reproduction,” Oxidative Medicine and Cellular Longevity, vol. 2019, Article ID 3279670, 13 pages, 2019. View at: Publisher Site | Google Scholar

Copyright © 2020 Giulia Collodel et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


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