Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study

Michal Kups; Kamil Gill; Aleksandra Rosiak-Gill; Patryk Harasny; Tomasz Machalowski; Marta Grabowska; Rafal Kurzawa; Olimpia Sipak; Malgorzata Piasecka

doi:10.12688/f1000research.121622.1

Home Browse Evaluation of selected semen parameters and biomarkers of male infertility...

ALL Metrics

-

Views

-

Downloads

Get PDF

Get XML

Export

▬

✚

Research Article

Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study

[version 1; peer review: 1 approved, 2 approved with reservations]

Michal Kups^1,2, Kamil Gill³, Aleksandra Rosiak-Gill^2,3, [...] Patryk Harasny^3,4, Tomasz Machalowski^3,5, Marta Grabowska³, Rafal Kurzawa^2,6, Olimpia Sipak⁷, Malgorzata Piasecka ³

Michal Kups^1,2, Kamil Gill³, [...] Aleksandra Rosiak-Gill^2,3, Patryk Harasny^3,4, Tomasz Machalowski^3,5, Marta Grabowska³, Rafal Kurzawa^2,6, Olimpia Sipak⁷, Malgorzata Piasecka ³

PUBLISHED 30 May 2022

Author details Author details

¹ Department of Urology and Oncological Urology, Regional Specialist Hospital in Szczecin, Szczecin, West Pomeranian Voivodeship, 71-455, Poland
² The Fertility Partnership Vitrolive in Szczecin, Szczecin, West Pomeranian Voivodeship, 70-483, Poland
³ Department of Histology and Developmental Biology, Faculty of Health Sciences, Pomeranian Medical University in Szczecin, Szczecin, West Pomeranian Voivodeship, 71-210, Poland
⁴ Department of Urology and Urological Oncology, Faculty of Medicine and Dentistry, Pomeranian Medical University in Szczecin, Szczecin, West Pomeranian Voivodeship, 70-111, Poland
⁵ Department of Perinatology, Obstetrics and Gynecology, Faculty of Medicine and Dentistry, Pomeranian Medical University in Szczecin, Police, West Pomeranian Voivodeship, 72-010, Poland
⁶ Department of Gynecology and Reproductive Health, Faculty of Health Sciences, Pomeranian Medical University in Szczecin, Szczecin, West Pomeranian Voivodeship, 71-210, Poland
⁷ Department of Obstetrics and Pathology of Pregnancy, Faculty of Health Sciences, Pomeranian Medical University in Szczecin, Szczecin, Szczecin, West Pomeranian Voivodeship, 71-210, Poland

Michal Kups
Roles: Investigation, Writing – Original Draft Preparation, Writing – Review & Editing

Kamil Gill
Roles: Conceptualization, Formal Analysis, Methodology, Visualization, Writing – Original Draft Preparation, Writing – Review & Editing

Aleksandra Rosiak-Gill
Roles: Formal Analysis

Patryk Harasny
Roles: Investigation

Tomasz Machalowski
Roles: Investigation

Marta Grabowska
Roles: Formal Analysis

Rafal Kurzawa
Roles: Investigation

Olimpia Sipak
Roles: Investigation

Malgorzata Piasecka
Roles: Conceptualization, Formal Analysis, Funding Acquisition, Methodology, Project Administration, Supervision, Visualization, Writing – Original Draft Preparation, Writing – Review & Editing

OPEN PEER REVIEW

REVIEWER STATUS

Abstract

Background: Because the etiopathogenesis of male infertility is multifactorial our study was designed to clarify the relationship between standard semen parameters, testicular volume, levels of reproductive hormones and the fragmentation of sperm nuclear DNA (SDF).
Methods: Patients (n = 130) were clustered as subjects: 1) with an abnormal volume (utrasonography) of at least one testis (<12 mL) or with a normal volume of testes and 2) with abnormal levels of at least one of the reproductive hormones (FSH, LH, PRL, TSH, total T – electrochemiluminescence method) or with normal hormonal profiles and 3) with high level of SDF (>30%), moderate (>15–30%) or low (≤15%) (sperm chromatin dispersion test).
Results: In subjects with a decreased testicular volume and in subjects with abnormal levels of reproductive hormones, decreased basic semen parameters were found. Participants with abnormal testicular volume had a higher percentage of SDF and a higher level of FSH (Mann–Whitney U test). In turn, men with a high level of SDF had lower testicular volume and conventional sperm parameters than men with a low level of SDF (Kruskal–Wallis test).
Conclusions: We showed that spermatogenesis disorders coexisted with decreased testicular volume and increased FSH levels. The disorders of spermatogenesis were manifested by reduced basic sperm characteristics and a high level of sperm nuclear DNA damage.

Keywords

male infertility, semen characteristics, testicular volume, reproductive hormones, sperm nuclear DNA integrity

Corresponding author: Malgorzata Piasecka

Competing interests: No competing interests were disclosed.

Grant information: This research was supported by Pomeranian Medical University in Szczecin, Poland, grant number WNoZ-322-01/S/19/2021 to M.P.
The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Copyright: © 2022 Kups M et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

How to cite: Kups M, Gill K, Rosiak-Gill A et al. Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study [version 1; peer review: 1 approved, 2 approved with reservations]. F1000Research 2022, 11:591 (https://doi.org/10.12688/f1000research.121622.1) First published: 30 May 2022, 11:591 (https://doi.org/10.12688/f1000research.121622.1) Latest published: 30 May 2022, 11:591 (https://doi.org/10.12688/f1000research.121622.1)

Introduction

The etiopathogenesis of male infertility is a multifactorial medical problem and is correlated with many congenital and acquired defects of the urogenital tract, cancers, urogenital infections, heat stress in the scrotum, hormonal disorders, genetic abnormalities and immunological factors. It is estimated that approximately 30–50% of male infertility cases are recognized as idiopathic, very often associated with low-quality of spermatozoa.¹^–⁴ On the other hand, unexplained infertility (couples where male patients have normal basic semen parameters and female patients have normal ovulation and fallopian tube potency) is diagnosed in 15–30% of cases.¹^–⁴ Therefore, the comprehensive evaluation of male fertility status should be developed using scrotal ultrasonography (USG) and assessment of the key reproductive hormone as well as advanced seminological tests.³^,⁵^–¹²

Available data has suggested that it is possible that infertile men could have normal standard semen characteristics.¹^,¹³^,¹⁴ Therefore, it is important to look beyond conventional semen analysis. Many authors report that among the advanced sperm tests, the assays that verify sperm nuclear DNA fragmentation (SDF) are the most clinically useful. Furthermore, evaluation of the percentage of SDF could significantly help in determining the most beneficial treatment algorithm for couples trying to have offspring.⁴^,⁵^,⁷^,¹⁵^–²⁰ An SDF ≤15% is considered a normal value (low level of nuclear DNA damage) and correlates with high male fertility potential. In these cases, the chance of becoming pregnant naturally or by intrauterine insemination (IUI) is high. In turn, SDF >15–30% (moderate level of DNA damage) can be associated with a reduced chance of becoming pregnant through natural conception and IUI or even in vitro fertilization (IVF) treatment. This range of SDF values and history of previous unsuccessful attempts to achieve pregnancy might indicate the need to introduce intracytoplasmic sperm injection (ICSI). Finally, a high level of SDF (>30%) is strongly associated with a significantly increased risk of reproductive failure, including ICSI treatment. It should be highlighted that even if pregnancy due to assisted reproductive technology (ART) is achievable, the percentage of sperm cells with a fragmented genome >30%, especially >40%, may significantly increase the risk of pregnancy loss.⁴^,¹⁶^,¹⁹^,²¹^–³⁰ These three ranges of sperm DNA damage (≤15%, >15–30% and >30) were primarily recommended for interpretation of the SCSA test results,¹⁶^,²²^,²⁵^,²⁶^,²⁹ however similar ranges also have been successfully adapted to sperm chromatin dispersion test (SCD).¹⁹^,²³^,³¹ Hence, an in-depth assessment of male fertility status, including testicular ultrasound, the levels of reproductive hormones and basic and advanced semen analysis, is clinically justified. Therefore, our study was designed to 1) determine the relationship between testicular volume, levels of reproductive hormones (follicle-stimulating hormone – FSH, luteinizing hormone – LH, prolactin – PRL, total testosterone – total T, thyroid-stimulating hormone – TSH), standard semen analysis and sperm genomic integrity and 2) compare standard semen parameters and investigated biomarkers of male infertility between groups of participants with low, moderate and high levels of nuclear DNA fragmentation.

Methods

Ethical considerations

In accordance with the Declaration of Helsinki, all participants in the study indicated their written conscious and voluntary consent to participate in the scientific project. The study was conducted according to the guidelines of the Declaration of Helsinki and approved by the Ethics Committee of Pomeranian Medical University, Szczecin, Poland (KB-0012/21/18, date of approval: 5 February 2018).

Study population

The study population consisted of 130 male infertile participants (median age: 33.00 years; range: 23–51 years) who were treated in 2018-2021 in the Individual Specialist Medical Practice (Szczecin, Poland) and The Fertility Partnership Vitrolive in Szczecin (Poland) – Gynaecology and Fertility Clinic and who gave their consent to participate in the study. All patients were partners of women (n = 130; median age: 30.00 years; range: 22–46 years) who did not become pregnant during one year (median: 2 years; range: 1.00–14.00 years) of regular intercourse without contraception (Figure 1). All initially qualified participants during a medical interview reported to the Laboratory of Andrology in the Department of Histology and Developmental Biology (Pomeranian Medical University, Szczecin, Poland) for seminological analysis. Based on the performed basic semen analysis, men with azoospermia and cryptozoospermia were excluded from the study group.

Figure 1. The strategy developed in study analyses.

FSH – follicle-stimulating hormone, LH – luteinizing hormone, SDF – sperm DNA fragmentation, PRL – prolactin, TSH – thyroid-stimulating hormone, total T – total testosterone.

The infertile status of subjects was verified based on an in-depth medical interview conducted by a specialist in urology (M. K.). The interview included information about factors that may affect fertility potential (genital injuries, cryptorchidism, varicocele, urogenital infections, chronic diseases, pharmacotherapy, use of anabolic steroids, operations and treatments, exposure to harmful factors, lifestyle, stimulants and others) (the interview form can be found as Extended data.¹¹¹ Moreover, the physical medical examination included body assessment (body and hair proportions), and palpation (penis, gonads, epididymides, seminal cords, inguinal canal, prostate and mammary glands) was carried out.

Conventional semen analysis

Standard semen analysis was carried out according to World Health Organization (WHO) recommendations.³² Semen samples were collected in a sterile urine container by masturbation after a two- to seven-day sexual abstinence. After complete liquefaction of semen (at 37 °C), standard semen analysis was carried out at room temperature – 22°C. The macroscopic evaluation of the semen included color, viscosity, volume and pH. In turn, the microscopic assessment (light/phase-contrast microscope DM500, Lecia, Heerbrugg, Switzerland) included the initial verification of the samples (presence of mucus bands, erythrocytes, epithelial cells, spermine crystals, residual bodies, aggregation and agglutination of sperm) as well as the assessment of the sperm concentration and the total sperm count, motility (progressive and nonprogressive motility), morphology, vitality and the concentration of inflammatory cells. Sperm concentration (analyzed in an improved Neubauer hemocytometer – Heinz Hernez Medizinalbedarf GmbH, Hamburg, Germany, ref no 1080339), sperm motility and vitality (eosin-positive cells and hypoosmotic-reactive cells: HOS test-positive cells) were assessed under a light/phase-contrast microscope using a 40× objective. To evaluate sperm cell morphology (including the teratozoospermia index reflecting multiple morphological defects per spermatozoon – TZI), native sperm smears were fixed and stained according to the Papanicolaou method (Aqua-Med, Lodz, Poland) and were analyzed under a bright light microscope using a 100× objective oil immersion lens. The concentration of leukocytes (peroxidase-positive cells) was calculated using the Endtz test (LeucoScreen kit, FertiPro N.V., Beernem, Belgium) and assessed in an improved Neubauer hemocytometer.

Sperm Chromatin Dispersion (SCD) test

To verify sperm nuclear DNA fragmentation, a commercial chromatin dispersion test – a Halosperm G2^® kit (Halotech DNA, Madrid, Spain) – was applied. The procedure was performed strictly according to the manufacturer’s guidelines and was described in detail in our previous publications.³³^–³⁶

To calculate the percentage of sperm cells with fragmented DNA, a minimum of 300 sperm cells per sample was counted under the 100x objective of a bright light microscope. According to the manufacturer’s guidelines, the following evaluation criteria were used: (1) sperm cells without nuclear DNA fragmentation (spermatozoa with a large halo – equal to or higher than the diameter of the core of spermatozoa and spermatozoa with a medium-sized halo – >1/3 of the diameter of the core of spermatozoa) and (2) sperm cells with nuclear DNA fragmentation (spermatozoa with a small halo – ≤1/3 of the diameter of the core of spermatozoa and spermatozoa without a halo but with a strongly stained core or without a halo and degraded chromatin – sperm cells showing no halo and simultaneously presenting an irregularly or weakly stained core) (Figure 2). The results of the SCD test (SDF) are presented as the sum of spermatozoa with nuclear DNA fragmentation divided by the total number of assessed sperm cells and multiplied by 100%.

Figure 2. Visualization of sperm chromatin dispersion test (SCD).

Micrographs obtained by light microscopy, 100×. Scale bar = 5 μm. Raw micrographs were edited in Corel Photo-Paint 2019 (Corel Corporation, Ottawa, Canada, RRID:SCR_014235). Editing included only: cropping (to center the presented sperm cells), rotation, brightening, contrast enhancing and enlargement.

Hormone profile of infertile subjects

To assess the panel of basic hormones influencing male fertility, potential vein blood was collected from participants in the morning (7.30–09.00), and the following hormones presented in Table 1 were measured. The hormone levels were determined by the electrochemiluminescence method (ECLIA) using the Cobas e801 analytical unit (Roche Diagnostics GmbH, Mannheim, Germany). The ECLIA method is based on the binding of biotinylated monoclonal-specific antibodies directed against the measured hormones and specific monoclonal antibodies labeled with a complex containing ruthenium metal (sandwich complex). In the next step, streptavidin-bound microparticles were used to bind to biotinylated antibodies directed against the measured hormones. Unbound substances were removed. The bound microparticles were magnetically trapped on the electrode surface. The electrode voltage induced chemiluminescence emission, which was measured by a photomultiplier. The result was determined based on a two-step calibration. All hormonal analyses were assessed strictly in accordance with the manufacturer's instructions.

Table 1. Reproductive hormones evaluated in study population.

Hormone	Short description of biological function	Reference value
FSH	FSH levels are considered a marker of impaired spermatogenesis⁸^,¹⁰^,¹¹^,¹⁰⁵^,¹⁰⁶	1.5–12.4 μIU/mL
LH	LH levels are helpful for differentiating primary (testicular) hypogonadism from secondary (nontesticular) hypogonadism¹⁰^,¹¹^,⁵⁴^,¹⁰⁶^,¹⁰⁷	1.7–8.6 uIU/mL
total T	Total T levels influences spermatogenesis through Sertoli cells¹⁰^,¹¹^,¹⁰⁸	8.0–41.7 nmol/L
PRL	PRL levels may result in decreased libido and erectile function¹⁰^,⁵⁵	4.04–15.02 ng/mL
TSH	Both hyperthyroidism and hypothyroidism can negatively affect spermatogenesis¹⁰^,⁵⁴^,⁵⁶	0.27–4.2 μIU/mL

Ultrasonography of the scrotum

To verify testicular volume, USG of the scrotum (ultrasound system Z-5 with a 75L38EA linear head; frequency range of 5–10 MHz, Mindray, Shenzhen, China) was performed by a senior urologist. The measurements were calculated using the following formula: length × width × height × 0.71. Furthermore, they were expressed in mL. According to the most commonly accepted criterion in clinical practice, the hypotrophic gonad was considered when the volume of the testis was less than 12 mL.³⁷ Additionally, the homogeneity and echogenicity of the gonadal parenchyma as well as the presence of possible focal lesions and microcalcifications were assessed.

Statistical analysis

Because the Shapiro–Wilk test showed that the data were not normally distributed, a nonparametric Mann–Whitney U test and Kruskal–Wallis test were applied to compare quantitative variables between two or more studied groups, respectively. Quantitative variables are expressed as the median with the range and mean ± standard deviation (SD). Additionally, to verify the relationships between study parameters, the Spearman’s rank (r_s) correlation coefficient was calculated (Figure 1). To interpret the strength of dependence between parameters, the following levels of correlation were presumed: <0.2 – lack of linear dependence (regardless of the p value), 0.2–0.4 – weak dependence, >0.4–0.7 – moderate dependence, >0.7–0.9 – strong dependence and >0.9 very strong dependence. For all statistical analyses, a p value < 0.05 was considered significant. Data analysis was performed using Statistica version 13.3 (StatSoft, Krakow, Poland, RRID:SCR_014213) and MedCalc version 18.2.1 (MedCalc Software, Ostend, Belgium, RRID:SCR_015044). Open source statistical software which can be used in the study – GNU PSPP.

Results

Seminological characteristics of study population

Of 130 obtained semen samples, 26 were classified as normozoospermia (total sperm count ≥39 × 10⁶ cells, sperm progressive motility ≥32%, normal sperm morphology ≥4%), 36 as teratozoospermia (abnormal sperm morphology), 36 as oligoasthenoteratozoospermia (simultaneously abnormal total sperm count, progressive motility and morphology), 19 as oligoteratozoospermia (simultaneously abnormal total sperm count and morphology), nine as asthenoteratozoospermia (simultaneously abnormal progressive sperm motility and morphology), three as oligozoospermia (abnormal sperm total count) and one as asthenozoospermia (abnormal sperm progressive motility). Moreover, 51 men had abnormal levels of at least one of the assessed reproductive hormones (FSH, LH, PRL, total T, TSH), and 37 men had an abnormal volume of at least one testis (<12 mL). The descriptive statistics of the investigated parameters are provided in Table 2.

Table 2. Descriptive statistics of study parameters in the infertile group of men (n=130).

Study parameters	median (range) mean ± SD
Age (y)	33.00 (23.00–51.00) 33.77 ± 5.28
LTV (mL)	n = 128 15.00 (2.40–25.00) 14.99 ± 4.81
RTV (mL)	n = 128 15.00 (4.30–25.00) 15.50 ± 4.95
FSH (mIU/mL)	n = 128 6.33 (1.17–21.35) 6.84 ± 3.90
LH (mIU/mL)	5.51 (1.21–17.10) 5.66 ± 2.63
total T (nmol/L)	16.60 (3.37–39.48) 17.53 ± 6.22
PRL (ng/mL)	13.22 (5.55–46.56) 14.25 ± 5.90
TSH (μU/mL)	n = 129 1.78 (0.63–14.40) 2.17 ± 1.50
Semen volume (mL)	3.00 (0.50–8.50) 3.34 ± 1.63
Sperm concentration (×10⁶/mL)	13.82 (0.40–251.00) 21.60 ± 28.54
Total number of spermatozoa (×10⁶)	42.70 (0.84–426.70) 67.76 ± 79.57
Morphologically normal spermatozoa (%)	1.00 (0.00–11.00) 1.80 ± 2.49
TZI	1.75 (1.34–2.50) 1.80 ± 0.29
Sperm progressive motility (%)	44.00 (0.00–87.00) 43.22 ± 22.25
Sperm nonprogressive motility (%)	5.00 (0.00–29.00) 5.56 ± 4.40
Total sperm motility (%)	50.00 (1.00–98.00) 48.79 ± 22.32
Eosin-negative spermatozoa – live cells (%)	76.50 (30.00–96.00) 74.33 ± 13.29
HOS test-positive spermatozoa – live cells (%)	n = 107 78.00 (26.00–92.00) 75.60 ± 10.51
Peroxidase-positive cells (%)	0.25 (0.00–10.25) 0.66 ± 1.32
SDF (%)	20.00 (3.00–58.00) 22.06 ± 12.04

Comparison of study parameters between infertile subjects with an abnormal volume of at least one testis (<12 mL) and subjects with a normal volume of testes (each ≥12 mL)

Compared groups did not differ in age. The subjects from the group with abnormal testicular volume (n = 37) had significantly higher levels of FSH than the reference group (n = 91) (median: 8.05 mIU/mL vs. 5.29 mIU/mL), whereas the levels of other study hormones (LH, PRL, total T, TSH) did not differ significantly (Table 3). On the other hand, in case of seminological parameters, patients with decreased testicular volume had a significantly reduced sperm concentration (medians: 6.25 × 10⁶ cells/mL vs. 19.00 ×10⁶ cells/mL), total sperm count (medians: 15.35 × 10⁶ cells vs. 60.12 × 10⁶ cells), sperm morphology (medians: 0.00% vs. 1.00%), progressive motility (medians: 37.00% vs. 51.00%), total motility (medians: 43.50% vs. 57.00%) and vitality – eosin-negative sperm cells (medians: 73.00% vs. 79.00%) and hypoosmotic (HOS) test-positive sperm cells (medians: 73.50% vs. 78.00%). It should be highlighted that in the group of men with abnormal testicular volume, a significantly higher percentage of SDF was found (medians: 27.00% vs. 17.00%) (Table 4).

Table 3. Descriptive statistics and comparison of age, testicular volume and levels of reproductive hormones between infertile subjects with an abnormal volume of at least one testis (<12 mL) and subjects with a normal volume of testes (each ≥12 mL).

Study parameters	Subjects with abnormal volume of at least one testis (<12 mL) n = 37	Subjects with normal volume of testes (each ≥12 mL) n = 91	p
median (range) mean ± SD
Age (y)	33.00 (23.00–49.00) 33.34 ± 4.89	33.00 (22.00–48.00) 33.60 ± 5.06	NS
LTV (mL)	10.00 (2.40–18.00) 9.81 ± 3.27	16.00 (12.00–25.00) 17.10 ± 3.58	<0.000000
RTV (mL)	10.00 (4.30–24.00) 10.82 ± 4.10	16.70 (9.00–25.00) 17.40 ± 3.90	<0.000000
FSH (mIU/mL)	8.05 (2.10–21.35) 8.66 ± 4.23	n = 90 5.29 (1.17–17.00) 6.05 ± 3.30	0.002677
LH (mIU/mL)	5.86 (2.30–17.10) 6.37 ± 3.32	5.49 (1.21–11.20) 5.41 ± 2.22	NS
total T (nmol/L)	16.12 (4.16–27.07) 4.76 ± 1.62	17.75 (3.37–39.48) 5.18 ± 1.87	NS
PRL (ng/mL)	13.12 (6.80–25.52) 13.84 ± 5.28	13.20 (5.55–46.56) 14.39 ± 6.18	NS
TSH (μU/mL)	1.61 (0.63–14.40) 2.13 ± 2.21	n = 90 2.01 (0.71–6.70) 2.19 ± 1.09	NS

Table 4. Descriptive statistics and comparison of semen parameters between infertile subjects with an abnormal volume of at least one testis (<12 mL) and subjects with a normal volume of testes (each ≥12 mL).

Study parameters	Subjects with abnormal volume of at least one testis (<12 mL) n = 37	Subjects with normal volume of testes (each ≥12 mL) n = 91	p
median (range) mean ± SD
Semen volume (mL)	3.00 (0.75–6.50) 3.04 ± 1.25	3.00 (0.50–8.50) 3.48 ± 1.76	NS
Sperm concentration (×10⁶/mL)	6.25 (0.40–57.25) 11.26 ± 12.63	19.00 (0.45–251.00) 26.05 ± 32.17	0.000111
Total number of spermatozoa (×10⁶)	15.35 (0.88–169.00) 33.28 ± 39.25	60.12 (0.84–426.70) 82.64 ± 87.72	0.000046
Morphologically normal spermatozoa (%)	0.00 (0.00–10.00) 1.05 ± 1.95	1.00 (0.00–11.00) 2.14 ± 2.63	0.028614
TZI	1.71 (1.36–2.50) 1.79 ± 0.29	1.75 (1.34–2.48) 1.81 ± 0.28	NS
Sperm progressive motility (%)	37.00 (0.00–87.00) 37.36 ± 20.47	51.00 (2.00–86.00) 45.98 ± 22.49	0.032661
Sperm nonprogressive motility (%)	5.00 (0.00–18.00) 5.50 ± 3.98	5.00 (0.00–29.00) 5.63 ± 4.59	NS
Total sperm motility (%)	43.50 (1.00–90.00) 42.86 ± 20.70	57.00 (4.00–88.00) 51.62 ± 22.42	0.026757
Eosin-negative spermatozoa –live cells (%)	73.00 (41.00–95.00) 70.07 ± 13.08	79.00 (30.00–96.00) 76.16 ± 13.10	0.003811
HOS test-positive spermatozoa – live cells (%)	n = 26 73.50 (54.00–88.00) 72.38 ± 9.60	n = 80 78.00 (26.00–92.00) 76.73 ± 10.66	0.033568
Peroxidase-positive cells (%)	0.25 (0.00–10.25) 0.65 ± 1.66	0.25 (0.00–9.50) 0.66 ± 1.17	NS
SDF (%)	27.00 (3.00–58.00) 29.00 ± 14.30	17.00 (4.00–46.00) 19.16 ± 9.75	0.000127

Comparison of study parameters between infertile subjects with abnormal levels of at least one of the assessed hormones and subjects with normal hormonal profiles

There were no significant differences in age between the compared groups, whereas higher levels of FSH, LH, PRL and TSH were noted in infertile men with abnormal hormonal profiles (n = 51) than in infertile men with normal hormonal profiles (n = 79). Unexpectedly, the study groups did not differ in the level of total T (Table 5). Regarding the semen parameters, infertile men with hormonal disorders had significantly lower total sperm count (medians: 30.25 × 10⁶ cells vs. 54.00 × 10⁶ cells), sperm morphology (medians: 0.00% vs. 1.00%), progressive motility (medians: 35.00% vs. 50.00%) and total motility (medians: 41.00% vs. 57.00%). Furthermore, the percentage of SDF was increased in the group with hormonal abnormalities, but the difference was not statistically significant (medians: 22.00% vs. 18.00%). Additionally, the compared groups did not differ in testicular volume (Table 6).

Table 5. Descriptive statistics and comparison of age, testicular volume and levels of reproductive hormones between infertile subjects with abnormal hormone profiles (abnormal levels of at least one evaluated hormone) and subjects with normal hormone profiles.

Study parameters	Subjects with abnormal hormone profile n = 51	Subjects with normal hormone profile n = 79	p
median (range) mean ± SD
Age (y)	33.00 (25.00–47.00) 33.43 ± 4.45	33.00 (22.00–49.00) 33.48 ± 5.44	NS
LTV (mL)	n = 49 14.00 (2.40–23.00) 13.98 ± 5.02	15.40 (6.00–25.00) 15.62 ± 4.59	NS
RTV (mL)	n = 50 14.00 (4.30–24.00) 14.61 ± 4.98	n = 78 15.00 (6.80–25.00) 16.06 ± 4.78	NS
FSH (mIU/mL)	7.70 (1.17–21.35) 7.90 ± 4.23	n = 78 5.40 (1.80–17.87) 6.16 ± 3.53	0.011483
LH (mIU/mL)	6.82 (1.45–17.10) 6.94 ± 3.07	4.49 (1.21–8.55) 4.83 ± 1.91	0.000024
total T (nmol/L)	15.60 (3.37–28.77) 16.08 ± 6.37	17.95 (8.32–39.48) 5.31 ± 1.74	NS
PRL (ng/mL)	19.10 (6.42–46.56) 18.85 ± 6.35	11.79 (5.55–17.90) 11.29 ± 3.00	< 0.000000
TSH (μU/mL)	n = 50 2.17 (0.63–14.40) 2.49 ± 1.98	1.65 (0.70–6.70) 1.97 ± 1.05	0.028703

Table 6. Descriptive statistics and comparison of semen parameters between infertile subjects with abnormal hormone profiles (abnormal levels of at least one evaluated hormone) and subjects with normal hormone profiles.

Study parameters	Subjects with abnormal hormone profile n = 51	Subjects with normal hormone profile n = 79	p
median (range) mean ± SD
Semen volume (mL)	3.00 (0.75–7.90) 3.07 ± 1.49	3.50 (0.50–8.50) 3.52 ± 1.70	NS
Sperm concentration (×10⁶/mL)	12.87 (0.40–118.50) 17.42 ± 20.70	15.62 (0.45–251.00) 24.31 ± 32.46	NS
Total number of spermatozoa (×10⁶)	30.25 (0.88–412.77) 52.53 ± 69.77	54.00 (0.84–426.70) 77.60 ± 84.27	0.018847
Morphologically normal spermatozoa (%)	0.00 (0.00–8.00) 1.17 ± 1.95	1.00 (0.00–11.00) 2.21 ± 2.72	0.013886
TZI	1.75 (1.35–2.50) 1.82 ± 0.32	1.75 (1.34–2.45) 1.79 ± 0.27	NS
Sperm progressive motility (%)	35.00 (0.00–86.00) 37.19 ± 21.44	50.00 (1.00–87.00) 47.11 ± 22.03	0.011304
Sperm nonprogressive motility (%)	5.00 (0.00–18.00) 4.88 ± 3.19	5.00 (0.00–29.00) 6.01 ± 4.99	NS
Total sperm motility (%)	41.00 (1.00–88.00) 42.07 ± 21.56	57.00 (2.00–90.00) 53.12 ± 21.84	0.005652
Eosin-negative spermatozoa – live cells (%)	74.00 (44.00–96.00) 73.47 ± 11.54	79.00 (30.00–95.00) 74.88 ± 14.35	NS
HOS test-positive spermatozoa – live cells (%)	n = 41 76.00 (54.00–91.00) 75.12 ± 8.66	n = 66 78.50 (26.00–92.00) 75.90 ± 11.56	NS
Peroxidase-positive cells (%)	0.25 (0.00–10.25) 0.94 ± 2.00	0.25 (0.00–2.15) 0.48 ± 0.51	NS
SDF (%)	22.00 (7.00–58.00) 23.54 ± 12.05	18.00 (3.00–54.00) 21.10 ± 12.02	NS

Comparison of study parameters between infertile subjects with SDF >30%, >15–30% and ≤15%

Based on the publications of other authors,¹⁶^,¹⁹^,²²^,²³^,²⁵^,²⁶^,²⁹^,³¹ the study group was divided into three subgroups: 1) with a high level of sperm nuclear DNA damage (SDF >30%, low fertility potential), 2) with a moderate level of sperm nuclear DNA damage (SDF >15–30%, moderate fertility potential) and 3) with a low level of sperm nuclear DNA damage (SDF ≤15%, high fertility potential) (Tables 7, 8).

Table 7. Descriptive statistics and comparison of age, testicular volume and levels of reproductive hormones between infertile subjects with SDF >30%, >15–30% and ≤15%.

Study parameters	Subjects with SDF >30 (1) n = 28	Subjects with SDF >15–30% (2) n = 59	Subjects with SDF ≤15% (3) n = 43	p 1 vs. 2	p 1 vs. 3	p 2 vs. 3
median (range) mean ± SD
Age (y)	31.00 (27.00–51.00) 33.00 ± 5.63	34.00 (25.00–49.00) 34.35 ± 5.90	34.00 (23.00–43.00) 33.48 ± 4.04	NS	NS	NS
LTV (mL)	13.00 (4.00–23.00) 13.48 ± 4.76	n = 57 14.00 (2.40–25.00) 14.55 ± 5.02	16.00 (9.00–25.00) 16.58 ± 4.15	NS	0.024251	NS
RTV (mL)	n = 27 12.00 (8.00–23.00) 13.78 ± 4.86	n = 58 15.00 (4.30–25.00) 15.38 ± 5.12	16.00 (8.00–24.00) 16.73 ± 4.52	NS	0.032806	NS
FSH (mIU/mL)	6.32 (1.80–14.50) 6.60 ± 3.55	6.70 (1.93–21.35) 7.51 ± 4.49	n = 42 5.44 (1.17–13.40) 6.06 ± 3.06	NS	NS	NS
LH (mIU/mL)	5.07 (2.10–9.30) 5.45 ± 2.47	5.77 (1.45–17.10) 5.93 ± 3.09	5.20 (1.21–11.20) 5.43 ± 2.01	NS	NS	NS
total T (nmol/L)	18.16 (4.16–28.08) 5.10 ± 1.51	16.19 (3.37–37.20) 17.05 ± 6.76	16.12 (8.66–39.48) 18.09 ± 6.10	NS	NS	NS
PRL (ng/mL)	11.89 (6.42–25.52) 13.89 ± 5.48	14.16 (6.42–28.30) 14.68 ± 4.95	12.14 (5.55–46.56) 13.91 ± 7.30	NS	NS	NS
TSH (μU/mL)	2.01 (0.63–14.40) 2.48 ± 2.52	n = 58 1.73 (0.84–6.70) 2.07 ± 1.06	1.68 (0.71–5.48) 2.11 ± 1.07	NS	NS	NS

Table 8. Descriptive statistics and comparison of semen parameters between infertile subjects with SDF >30%, >15–30% and ≤15%.

Study parameters	Subjects with SDF >30 (1) n = 28	Subjects with SDF >15–30% (2) n = 59	Subjects with SDF ≤15% (3) n = 43	p 1 vs. 2	p 1 vs. 3	p 2 vs. 3
median (range) mean ± SD
Semen volume (mL)	3.50 (1.50–8.50) 3.66 ± 1.78	3.00 (0.50–8.00) 3.06 ± 1.47	3.00 (1.50–8.00) 3.52 ± 1.72	NS	NS	NS
Sperm concentration (×10⁶/mL)	5.65 (0.40–60.25) 10.90 ± 13.50	13.04 (0.80–104.50) 20.40 ± 21.46	21.75 (2.50–251.00) 30.22 ± 39.93	NS	0.000175	NS
Total number of spermatozoa (×10⁶)	20.02 (0.88–221.00) 42.42 ± 59.41	41.25 (0.84–365.75) 63.75 ± 78.07	70.76 (8.10–426.70) 89.77 ± 88.40	NS	0.000841	0.025102
Morphologically normal spermatozoa (%)	0.00 (0.00–3.00) 0.46 ± 0.83	0.00 (0.00–8.00) 1.20 ± 1.74	3.00 (0.00–11.00) 3.51 ± 3.11	NS	0.000011	0.000497
TZI	1.79 (1.36–2.48) 1.82 ± 0.26	1.69 (1.35–2.50) 1.82 ± 0.33	1.75 (1.34–2.30) 1.77 ± 0.24	NS	NS	NS
Sperm progressive motility (%)	26.00 (6.00–67.00) 28.82 ± 16.00	40.00 (0.00–70.00) 38.54 ± 19.72	63.00 (15.00–87.00) 59.02 ± 19.86	NS	<0.000001	0.000016
Sperm nonprogressive motility (%)	4.50 (0.00–10.00) 4.39 ± 2.97	6.00 (0.00–29.00) 6.74 ± 5.50	4.00 (0.00–13.00) 4.72 ± 2.89	NS	NS	NS
Total sperm motility (%)	31.00 (6.00–71.00) 33.21 ± 17.18	48.00 (1.00–77.00) 45.28 ± 20.01	68.00 (15.00–90.00) 63.74 ± 19.47	NS	<0.000001	0.000103
Eosin-negative spermatozoa – live cells (%)	67.00 (30.00–86.00) 62.71 ± 15.82	74.00 (40.00–86.00) 73.13 ± 9.49	86.00 (47.00–96.00) 83.53 ± 8.87	NS	<0.000001	<0.000001
HOS test-positive spermatozoa – live cells (%)	n = 17 67.00 (26.00–84.00) 65.82 ± 13.67	n = 49 72.00 (53.00–85.00) 72.97 ± 7.44	n = 41 83.00 (67.00–92.00) 82.80 ± 7.01	NS	<0.000001	0.000001
Peroxidase-positive cells (%)	0.25 (0.00–10.25) 0.81 ± 1.90	0.25 (0.00–9.50) 0.76 ± 1.41	0.25 (0.00–2.00) 0.41 ± 0.51	NS	NS	NS

Statistical analysis revealed some significant differences between men with SDF >30% (n = 28) and men with SDF ≤15% (n = 43). The first group had significantly lower left testis volume (medians: 13.00 mL vs. 16.00 mL) and right testis volume (medians: 12.00 mL vs. 16.00 mL), sperm concentration (medians: 5.65 ×10⁶ cells/mL vs. 21.75 ×10⁶ cells/mL), total sperm count (medians: 20.02 ×10⁶ cells vs. 70.76 ×10⁶ cells), sperm morphology (medians: 0.00% vs. 3.00%), progressive motility (medians: 26.00% vs. 63.00%), total motility (medians: 33.00% vs. 68.00%) and vitality – eosin-negative sperm cells (medians: 67.00% vs. 86.00%) and HOS test-positive sperm cells (medians: 67.00% vs. 83.00%) (Tables 7, 8).

In addition, in contrast to men with SDF ≤15%, infertile men with SDF >15–30% (n = 59) had a significantly lower total sperm count (medians: 41.25 × 10⁶ cells vs. 70.76 × 10⁶ cells), sperm morphology (medians: 0.00% vs. 3.00%), progressive motility (medians: 40.00% vs. 63.00%), total motility (medians: 48.00% vs. 68.00%) and sperm vitality – eosin-negative sperm cells (medians: 74.00% vs. 86.00%) and HOS test-positive sperm cells (medians: 72.00% vs. 83.00%) (Table 7).

On the other hand, we did not observe any significant differences between men with SDF >30% and men with SDF >15–30% in any study parameters. Additionally, in the case of age, hormone levels (FSH, LH, PRL, total T, TSH), TZI index, sperm nonprogressive motility and concentration of peroxidase-positive cells in semen, no significant differences between the compared three groups were recorded (Tables 7, 8).

Spearman’s rank correlations between the study parameters

Correlations between SDF, male age, basic semen parameters, testicular volume and hormone levels

Analysis of the Spearman’s rank correlation coefficient showed a linear relationship between SDF and selected parameters. SDF was negatively correlated with sperm concentration (r_s = –0.3461; weak dependence), total sperm count (r_s = –0.3343; weak dependence), sperm morphology (r_s = –0.4482; moderate dependence), progressive motility (r_s = –0.5476; moderate dependence), total motility (r_s = –0.5374; moderate dependence) and vitality – eosin-negative sperm cells (r_s = –0.6389; moderate dependence) and HOS test-positive sperm cells (r_s = –0.5811; moderate dependence). In turn, there were no significant correlations between SDF and age, ejaculate volume, nonprogressive sperm motility or peroxidase-positive cell concentration. Moreover, a negative correlation between SDF and the volume of the left testis was found (r_s = –0.2055; weak dependence), whereas there were no significant correlations between SDF and the volume of the right testis or study hormone levels (Tables 9, 10).

Table 9. Spearman's rank correlation coefficient (r_s) between age, testicular volume and levels of reproductive hormones.

Study parameters	LTV (mL)	RTV (mL)	FSH (mIU/mL)	LH (mIU/mL)	total T (nmol/L)	PRL (ng/mL)	TSH (μU/mL)	SDF (%)
Age (y)	n = 128 r_s = -0.0320 NS	n = 128 r_s = -0.0767 NS	n = 129 r_s = 0.1698 NS	r_s = 0.1203 NS	r_s = 0.0657 NS	r_s = -0.0599 NS	n = 129 r_s = -0.0398 NS	r_s = -0.0783 NS
LTV (mL)	-	n = 127 r_s = 0.8259 p < 0.000001	n = 127 r_s = -0.2491 p = 0.004745	n = 128 r_s = -0.1529 NS	n = 128 r_s = 0.1225 NS	n = 128 r_s = -0.0195 NS	n = 127 r_s = 0.0832 NS	n = 128 r_s = -0.2055 p = 0.019947
RTV (mL)	n = 127 r_s = 0.8259 p < 0.000001	-	n = 127 r_s = -0.2402 p = 0.006525	n = 128 r_s = -0.1596 NS	n = 128 r_s = 0.1418 NS	n = 128 r_s = -0.0759 NS	n = 127 r_s = 0.0564 NS	n = 128 r_s = -0.1666 NS
FSH (mIU/mL)	n = 127 r_s = -0.2491 p = 0.004745	n = 127 r_s = -0.2402 p = 0.006525	-	n = 129 r_s = 0.5018 p < 0.000001	n = 129 r_s = 0.0524 NS	n = 129 r_s = 0.0720 NS	n = 128 r_s = 0.0533 NS	n = 129 r_s = 0.0459 NS
LH (mIU/mL)	n = 128 r_s = -0.1529 NS	n = 128 r_s = -0.1596 NS	n = 129 r_s = 0.5018 p < 0.000001	-	r_s = 0.0568 NS	r_s = 0.1673 NS	n = 129 r_s = 0.0874 NS	r_s = 0.0029 NS
total T (nmol/L)	n = 128 r_s = 0.1225 NS	n = 128 r_s = 0.1418 NS	n = 129 r_s = 0.0524 NS	r_s = = 0.0568 NS	-	r_s = -0.0703 NS	n = 129 r_s = --0.0892 NS	r_s = 0.0102 NS
PRL (ng/mL)	n = 128 r_s = -0.0195 NS	n = 128 r_s = -0.0759 NS	n = 129 r_s = 0.0720 NS	r_s = 0.1673 NS	r_s = -0.0703 NS	-	n = 128 r_s = 0.1514 NS	r_s = 0.0802 NS
TSH (μU/mL)	n = 127 r_s = 0.0832 NS	n = 127 r_s = 0.0564 NS	n = 128 r_s = 0.0533 NS	n = 129 r_s = 0.0874 NS	n = 129 r_s = -0.0892 NS	n = 129 r_s = 0.1514 NS	-	n = 129 r_s = 0.0321 NS

Table 10. Spearman's rank correlation coefficient (r_s) between testicular volume, levels of reproductive hormones and semen parameters.

Study parameters	LTV (mL)	RTV (mL)	FSH (mIU/mL)	LH (mIU/mL)	total T (nmol/L)	PRL (ng/mL)	TSH (μU/mL)	SDF (%)
Semen volume (mL)	n = 128 r_s = 0.0376 NS	n = 128 r_s = -0.0524 NS	n = 129 r_s = -0.0102 NS	r_s = -0.1285 NS	r_s = -0.0994 NS	r_s = -0.1039 NS	n = 129 r_s = -0.0278 NS	r_s = -0.0265 NS
Sperm concentration (×10⁶/mL)	n = 128 r_s = 0.4345 p < 0.000001	n = 128 r_s = 0.4019 p = 0.000003	n = 129 r_s = -0.1720 NS	r_s = -0.2205 p = 0.011703	r_s = -0.0564 NS	r_s = -0.0760 NS	n = 129 r_s = -0.0152 NS	r_s = -0.3461 p = 0.000055
Total number of spermatozoa (×10⁶)	n = 128 r_s = 0.4191 p < 0.000001	n = 128 r_s = 0.3452 p = 0.000066	n = 129 r_s = -0.1850 p = 0.035873	r_s = -0.2350 p = 0.007108	r_s = -0.1067 NS	r_s = -0.1011 NS	n = 129 r_s = -0.0220 NS	r_s = -0.3343 p = 0.000101
Morphologically normal spermatozoa (%)	n = 128 r_s = 0.1634 NS	n = 128 r_s = 0.1579 NS	n = 129 r_s = 0.0063 NS	r_s = -0.0681 NS	r_s = -0.0269 NS	r_s = -0.1368 NS	n = 129 r_s = -0.0006 NS	r_s = -0.4482 p < 0.000001
TZI	n = 128 r_s = 0.0062 NS	n = 128 r_s = 0.0549 NS	n = 129 r_s = 0.0152 NS	r_s = 0.0235 NS	r_s = 0.1821 p = 0.037884	r_s = 0.0066 NS	n = 129 r_s = 0.1334 NS	r_s = 0.0525 NS
Sperm progressive motility (%)	n = 128 r_s = 0.2048 p = 0.020393	n = 128 r_s = 0.1313 NS	n = 129 r_s = 0.0378 NS	r_s = -0.0940 NS	r_s = -0.0095 NS	r_s = -0.1121 NS	n = 129 r_s = -0.1199 NS	r_s = -0.5476 p < 0.000001
Sperm nonprogressive motility (%)	n = 128 r_s = 0.0562 NS	n = 128 r_s = -0.0665 NS	n = 129 r_s = 0.0030 NS	r_s = 0.0059 NS	r_s = -0.0287 NS	r_s = 0.0462 NS	n = 129 r_s = -0.0200 NS	r_s = 0.0202 NS
Total sperm motility (%)	n = 128 r_s = 0.2115 p = 0.016524	n = 128 r_s = 0.1236 NS	n = 129 r_s = 0.0207 NS	r_s = -0.1126 NS	r_s = -0.0096 NS	r_s = -0.1008 NS	n = 129 r_s = -0.1180 NS	r_s = -0.5374 p < 0.000001
Eosin-negative spermatozoa – live cells (%)	n = 128 r_s = 0.1956 p = 0.026909	n = 128 r_s = 0.0945 NS	n = 129 r_s = -0.0102 NS	r_s = -0.0470 NS	r_s = 0.1652 NS	r_s = -0.0577 NS	n = 129 r_s = -0.0300 NS	r_s = -0.6389 p < 0.000001
HOS test-positive spermatozoa – live cells (%)	n = 105 r_s = 0.1904 NS	n = 105 r_s = 0.0934 NS	n = 106 r_s = -0.0359 NS	n = 107 r_s = -0.0147 NS	n = 107 r_s = 0.1867 NS	n = 107 r_s = -0.0708 NS	n = 106 r_s = 0.0016 NS	n = 107 r_s = -0.5811 p < 0.000001
Peroxidase-positive cells (%)	n = 128 r_s = 0.1248 NS	n = 128 r_s = 0.1629 NS	n = 129 r_s = -0.0553 NS	r_s = 0.0387 NS	r_s = 0.1138 NS	r_s = -0.0247 NS	n = 129 r_s = 0.1441 NS	r_s = 0.1906 p = 0.026503

Correlations between testicular volume, hormone levels and basic semen parameters

In the examined group, the left and right testis volumes were negatively correlated with the level of FSH (r_s = –0.2491 and r_s = –0.2402, respectively; weak dependences) but was not correlated with other hormones (LH, PRL, total T, TSH). Moreover, the volumes of the left and right testes were positively correlated with sperm concentration (r_s = 0.4345 and r_s = 0.4019, respectively; moderate dependences) and total sperm number (r_s = 0.4191 and r_s = 0.3452, respectively; moderate and weak dependences). Additionally, positive correlations between left testis volume and sperm progressive motility (r_s = 0.2048) as well as total motility (r_s = 0.2115; weak dependence) were found. Furthermore, the LH level was negatively correlated with sperm concentration (r_s = –0.2205; weak dependence) and total sperm count (r_s = –0.2350; weak dependence), but there were no other significant correlations between the levels of assessed hormones and conventional semen parameters (Tables 9, 10).

The raw data can be found as Underlying data.¹⁰⁹^,¹¹⁰

Discussion

Reduced basic semen parameters can result from impaired spermatogenesis

Based on the obtained data, it can be suggested that the failure to become a biological father could be due to disorders of spermatogenesis manifested by reduced standard seminological parameters. It is worth noting that in our study, the median morphologically normal sperm was only 1%, and as many as 100 out of 130 infertile men had teratozoospermia (isolated or coexisting with other semen disorders). Additionally, studies conducted by other authors confirm the relationship between standard sperm parameters and male fertility.³⁸^–⁴⁰ Slama et al.⁴⁰ proved a significantly shorter time to pregnancy (TTP) in women whose partners had a higher percentage of sperm with normal morphology. Moreover, it was found that the percentage of morphologically normal sperm was decreased in men from couples with a history of recurrent miscarriage.⁴¹^–⁴⁴ Additionally, morphologically normal sperm cells play an important role not only in the case of natural conception but also in medically assisted conception (IUI, fertilization in vitro),⁴⁵ and it has been shown that sperm morphology may also influence embryo development.⁴³ On the other hand, reproductive success might be achieved even when morphologically normal sperm cells are not observed in the semen. Shabtaie et al.⁴⁶ emphasize that in the case of only abnormal sperm morphology (assuming no female infertility factor), first-line therapy should not assist ART without undertaking a sufficiently long attempt at natural conception. Therefore, opinions about the predictive value of sperm morphological assessment for both natural conception and medically assisted conception are controversial.⁴⁶^–⁴⁹

Additionally, in our study, 46 cases of asthenozoospermia (isolated or coexisting) and 58 cases of oligozoospermia (isolated or coexisting) were observed. Many authors confirm that progressive motility is one of the most important parameters influencing reproductive success both in terms of natural conception and medically assisted conception.³⁸^,⁵⁰^,⁵¹ Furthermore, Lotti et al.³⁹ revealed a negative correlation between sperm vitality and TTP. Also, analyzing a large group of infertile men and men from the control group (candidates to be sperm donors), Li et al.⁴³ showed that in the first group, there were significantly more men with azoospermia, asthenozoospermia and oligoasthenozoospermia, whereas surprisingly isolated oligoozospermia were detected more often in men from the control group. Some authors⁵² even suggest greater clinical implications of the total sperm count in relation to the sperm concentration.

However, it should be emphasized that low standard seminological parameters are not always synonymous with infertility status. Not all authors⁵³ recognize the arbitrary division of men into fertile and infertile groups based only on the basic semen characteristics according to the WHO.³² Therefore, except for azoospermia, necrozoospermia, and globo- and macrozoospermia, it is difficult to determine male fertility potential considering only standard seminological parameters. This thesis is also confirmed by our previous studies, in which 19 cases of reduced basic semen parameters were found in a group of men with proven fertility (n = 64).³³ Therefore, in this research, the standard seminological assessment was not only one criterion for qualifying a man as infertile. The patient's infertility was established by an interview indicating unsuccessful attempts for offspring during one year of regular intercourse without the use of contraception.

Relationships between testicular volume, hormone levels, basic semen parameters and sperm genome integrity

The results of our research suggest the coexistence of spermatogenesis disorders with a reduced testicle volume and a higher FSH level. It is known that the process of spermatogenesis, reflecting testicular function, depends on a hypothalamic–pituitary–gonadal axis function, in which gonadotropins LH and FSH play a key role in maintaining testosterone biosynthesis and the function of the seminiferous epithelium, respectively. In addition, it has been proven that the function of the male gonad is influenced by thyroid hormones and prolactin.¹⁰^,⁵⁴^–⁵⁷ Importantly, a significant decrease in testicular volume can be associated with both reduced hormonal activity (lower levels of androgens) and reproductive activity manifested by seminiferous tubule atrophy.⁵⁸^–⁶⁰ Therefore, our study included an evaluation of not only standard seminological parameters but also testicular volume and reproductive hormone levels (FSH, LH, PRL, total T, TSH). It should be noted that the selection of the assessed hormones was based on data from the literature.¹¹^,⁶¹ Unfortunately, to date, there have been no strict guidelines regarding the hormonal test profile that should be determined in the routine diagnosis of male infertility. On the other hand, the European Academy of Andrology (EAA) in guidelines from 2018 postulates evaluation of total T, FSH and LH in every case of an infertile man with oligoasthenoteratozoospermia (OAT).⁶¹ These recommendations are in line with the guidelines of the European Society Urology (EAU) from 2021.¹¹ However, it is believed that the remaining hormonal tests should be performed based on an individual assessment of the patient. The levels of commonly recognized markers of spermatogenesis and Sertoli cell function (FSH, inhibin B) have been most frequently studied in the available literature.⁶²^–⁶⁵ In addition, the authors of the study also paid attention to the analysis of the levels of SHBG,⁶⁶ prolactin,⁶⁷ estradiol,⁶⁶ TSH,⁶⁸^–⁷⁰ cortisol,⁷¹^,⁷² growth hormone (GH) and insulin-like growth factor 1 (IGF-1).⁷³

Based on testicular volume measurement in our study, two groups of infertile participants were distinguished: men with a volume of at least one testis below the norm (<12 mL) and men with a normal volume of both testes.⁷⁴ Our results showed that infertile men with a reduced volume of at least one testis had a significantly higher FSH level and a lower sperm count, sperm morphology, motility and vitality. Moreover, it should be especially highlighted that we found a significantly increased fragmentation of sperm nuclear DNA in the first group. These results were confirmed by correlation analysis. The testicular volume was negatively correlated with the level of FSH and the SDF value and positively correlated with the number and motility of sperm. Surprisingly, we did not find an association between testicular volume and total T level.

The obtained findings were partially consistent with the data published by other authors. Numerous researchers have reported relationships between testicular volume, conventional semen parameters, gonadotropin and testosterone levels as well as the results of advanced sperm tests (chromatin status, mitochondrial potential, apoptosis).⁵⁷^,⁶⁰^,⁷⁵^–⁷⁸ The coexistence of reduced standard semen parameters, decreased testosterone levels and testicular volume presented by Bahk et al.⁷⁵ and Condorelli et al.⁵⁷ suggest that the reduction of testicular volume may be associated not only with impaired spermatogenesis but also with decreased hormonal function of male gonads. Condorelli et al.⁵⁷ recommend periodic assessment of testosterone levels for patients with hypotrophic gonads. On the other hand, the obtained results presented by other authors are not always unambiguous. For example, in contrast to our results, Condorelli et al.⁵⁷ revealed a relationship between testicular volume and testosterone levels, but they did not find a correlation between testicular volume and gonadotropin levels.

As mentioned above, we discovered that a group of men with at least one testis volume <12 mL had significantly reduced integrity of the sperm genome. The data could suggest that spermatogenesis disorders coexist with decreased testicular volume and are manifested not only by reduced conventional sperm parameters but also by molecular disorders of sperm chromatin. The relationship between testicular volume and sperm DNA strand brakes was also confirmed by our other findings. The participants with a high level of SDF (>30%) had significantly smaller testes than men with a low level of SDF (≤15%). Moreover, we noted a negative correlation between testicular volume and sperm chromatin fragmentation. Similar results were obtained by other authors who observed a negative correlation between the fragmentation of nuclear sperm DNA (verification using the TUNEL method), its denaturation (verification using acridine orange), sperm chromatin density (verified using propidium iodide) and the volume of testes.⁵⁷^,⁷⁶^,⁷⁸

In the next step of our research, we compared two groups of subjects: men with abnormal levels of at least one of the assessed hormones and men with normal hormonal profiles. The obtained findings provided nonobvious data. We noted that in the first group, sperm count, morphology and motility were reduced, but testicular volume did not differ significantly between the two groups. Moreover, the LH level was negatively correlated with the total sperm count. Additionally, other authors have confirmed the statistical relationship between the level of selected hormones and standard seminological parameters.⁶³^,⁷⁹^,⁸⁰ Wei et al.⁸⁰ showed that in patients with OAT, total T was positively correlated with sperm morphology, whereas PRL was correlated with sperm concentration and motility. Moreover, Lu et al.⁷⁹ and Uhler et al.⁶³ revealed a negative correlation between FSH level and semen volume, sperm concentration, morphology and motility as well as between LH level and sperm concentration in infertile men or healthy volunteers.

It should be pointed out that we did not find significant differences in the percentage of sperm cells with fragmented DNA between participants with abnormal levels of at least evaluated hormones and those with normal hormonal profiles. This comparative analysis was consistent with Spearman's rank correlation test, which did not show a significant correlation between the SDF value and the level of the assessed hormones. However, other authors’ data indicated statistical relationships between sperm chromatin quality and the hormonal profile⁷⁹^,⁸¹^,⁸² The coexistence of sperm DNA fragmentation with abnormally high or low levels of gonadotropins was shown in research published by Wdowiak et al.⁸² These results were partially consistent with the studies of Lu et al.⁸³ and Smit et al.,⁸¹ who showed a negative correlation between sperm DNA fragmentation and elevated levels of FSH and LH. In turn, the association between sperm nuclear DNA damage and testosterone level is not always unequivocal. Some researchers Wdowiak et al.⁸² have found a negative correlation between these parameters, whereas others did not confirm these findings.⁸¹^,⁸³

The open question is why there was no statistically significant difference in our research in the percentage of SDF between the groups of men differing in the level of at least one of the assessed hormones. There is no doubt that the obtained results could have been influenced by the limited number of infertile men (n = 130) enrolled in our study and the hormonal heterogeneity of the group of men with abnormal levels of at least one of the verified hormones. Disturbances in the level of hormones can be both a factor influencing infertility and a consequence of such a state. In other words, an abnormal hormonal profile can be responsible for reduced semen quality or may be only a secondary effect of pathological processes in testes. In addition, it should be emphasized that there are many potential factors (e.g., obesity, occupational exposure, comorbidities, age, pharmacotherapy, stress) that may affect the interrelationship between spermatogenesis and hormone levels.⁸⁴^–⁹⁰

Sperm genome integrity is a key point for male fertility

It can be assumed that in our investigated group of infertile patients, one of the major factors that limited the ability of male gametes to fertilize was probably an increased level of sperm nuclear DNA fragmentation. It was found that the group of men with SDF >30% had a significantly reduced sperm count, morphology, motility and vitality in comparison to infertile men with a normal SDF rate of ≤15%. Similarly, Erenpreiss et al.⁹¹ showed that if males had diagnosed astheno- and teratozoospermia, the odds ratio (OR) for >20% DFI or for >30% DFI was 1.9–4.0-fold higher or 2.8–6.2-fold higher, respectively, than in subjects with normal sperm motility and morphology. Additionally, Vinnakota et al.²⁹ observed a decrease in sperm motility in participants with SDF >30%. Moreover, we showed that the level of SDF was negatively correlated (Spearman’s rank correlation test) with sperm count, morphology, motility and vitality, and our findings have been confirmed by the research of other authors.³⁰^,⁸³^,⁹²^,⁹³ However, some researchers have not always found a correlation between SDF and basic sperm parameters.⁹⁴^–⁹⁶

Importantly, it should be highlighted that in our study, the median SDF was 20%. In fact, according to the manufacturer of the Halosperm G2^® kit, these results are in the normal range (SDF below 30%). It seems that the threshold of 30% SDF is too high (risk of a false-negative result). In our previous publications, we demonstrated that the median SDF in the group of men with confirmed fertility and/or with high reproductive potential (healthy volunteers with normozoospermia) ranged from 12% to 14%.³³^–³⁶ In addition, these studies also showed a significantly satisfactory predictive value of the sperm chromatin dispersion (SCD) test to discriminate males with normal reproductive potential from those with reduced reproductive potential (based on receiver operating characteristic [ROC] analysis). The cut-off value was 18% and 20% SDF.³⁴^–³⁶ Moreover, we obtained a threshold of 18% SDF to distinguish infertile men from fertile men (unpublished data).

These observations were in agreement with other authors who also clearly showed that the level of sperm nuclear DNA fragmentation was correlated with male infertility and that the acceptable threshold for sperm genome fragmentation was not below 30% but rather below 20%.⁴^,¹⁶^,²¹^,²⁷^–²⁹^,⁹¹^,⁹⁷^–¹⁰² For example, Bungum et al.⁹⁷ showed that in the case of subjects with sperm nuclear DNA fragmentation in the range of 0–20%, the chance of spontaneous pregnancy is constant, but an increase in sperm DNA fragmentation >20% is associated with a reduced ability to achieve pregnancy. Moreover, Majzoub et al.¹⁰¹ estimated that the mean value of SDF for fertile subjects was 15.68%, whereas in the infertile group, it was 27.60%. In turn, comparing the groups of fertile and infertile men, Wiweko and Utami¹⁰² found not only significant differences in the SDF value between the study groups (medians: 19.90% vs. 29.95%, respectively) but also reported that SDF at the cutoff point of 26.1% had a higher diagnostic value. Similar results were presented by Javed et al.¹⁰⁰ (the SDF at the cutoff point was 24.47%). Moreover, Evenson¹⁶ emphasized that the percentage of sperm with damaged chromatin >15–25 could increase the risk of male infertility and that >20–35% spermatozoa with damaged DNA could significantly reduce the chances of becoming pregnant using in vitro fertilization. Therefore, based on own research and analysis of the results of other researchers, Evenson¹⁶ concluded that when the percentage of sperm with abnormal chromatin status was >20, male fertility was decreased, and in vitro fertilization as first-line therapy should be considered. These conclusions were also confirmed by Giwercman et al.,¹⁰³ who performed a comparison of ORs for the occurrence of infertility depending on the percentage of DFI. The authors showed that in the group of men with DFI 10–20%, the risk of reproductive failure was higher (OR = 2.5) than that in men with DFI <10%. In addition, Giwercman et al.¹⁰³ observed a significant increase in the risk of infertility (OR = 8.4) in men with DFI> 20% compared to men with DFI <10%. Finally, in two most recent publications both Esteves et al.⁴ and Agarwal et al.²¹ reported that cut-off point of 20% sperm cells with fragmented DNA (verified both by SCSA, TUNEL and SCD assay) is the best criterion to discriminate fertile men from infertile.

Additionally, the influence of DFI on the fertilization process has been confirmed. Simon et al.⁵¹ revealed a higher risk (OR = 9.5) of a low percentage of fertilized oocytes (<40% fertilized oocytes) when men had DFI >40% compared to men with DFI ≤40%. Therefore, we can assume that sperm chromatin abnormalities may be accompanied by lowered standard sperm parameters synergistically affecting male fertility.

Conclusions

Our comprehensive assessment of male infertility factors allowed us to conclude that in the study clinical cases, spermatogenesis disorders coexisted with decreased testicular volume and increased FSH levels. Moreover, they were manifested not only by reduced basic sperm characteristics but also, very importantly, by a high level of sperm nuclear DNA damage, which has great clinical utility both in terms of natural conception and in terms of ART (Figure 1). Furthermore, our current and previous findings suggest that the cut-off value of 30% SDF given by manufacturer of the Halosperm G2^® kit seems too high and should be revised/downgraded to 20%, for better prognosis of male fertility. What’s more, clarification of the relationship between standard semen parameters, testicular volume, levels of reproductive hormones, SDF and clinical features might help to develop new personalized strategies for therapeutic interventions. In the case of infertile men, a complete andrological examination including in-depth medical interview, physical examination, standard semen analysis, scrotal ultrasound, assessment of reproductive hormones and integrity of sperm genome is justified. This medical approach is necessary not only due for verification of the causes of infertility but also due to the need to detect serious health disorders that may be life-threatening. For example, it has been proven that infertile men have an increased risk of testicular cancer, which determines the recommendation of periodic ultrasound examinations of the scrotum and gonadal self-examination.¹¹^,¹⁰⁴ Therefore, the introduction of a complex diagnosis of male infertility factors is justified and needed.

Limitations of the study

Some limitations of our study should be addressed. One of the most important factors influencing our results is the limited number of participants. In total, 130 men from couples with confirmed infertility were included to this project. It is known that the most reliable data are obtained from well-designed studies on large cohorts of patients. Due to the limited number of participants in our research, the presented results should be approached critically, and it should be borne in mind that studies conducted on a larger group could provide different results and conclusions. Moreover, the number of compared men in particular groups was not equal, which may affect the obtained statistical differences between groups. In the assessed hormonal profile, we did not include the determinations of some markers which could also be important for assessing the status of male fertility (i.a. inhibin B, SHBG, GH, estradiol, cortisol). Finally, sperm chromatin dispersion (SCD) test was performed to assess SDF. This test is a standardized diagnostic method but often not considered the gold standard for sperm DNA assessment because it does not directly evaluate breaks of DNA.

Data availability

Underlying data

Zenodo: Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study. https://doi.org/10.5281/zenodo.6536196.¹⁰⁹

This project contains the following underlying data:

- Kups et al. for database.xlsx (raw data)

Zenodo: Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study. https://doi.org/10.5281/zenodo.6538474.¹¹⁰

This project contains the following underlying data:

- Raw microphotographs

Extended data

Zenodo: Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study. https://doi.org/10.5281/zenodo.6542238.¹¹¹

This project contains the following extended data:

- Urological and andrological medical interview Michal Kups.pdf (Patient card used during the medical interview)

Data are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).

References

1. Minhas S, Bettocchi C, Boeri L, et al.: European Association of Urology Guidelines on Male Sexual and Reproductive Health: 2021 Update on Male Infertility. Eur. Urol. 2021; 80: 603–620. PubMed Abstract | Publisher Full Text
2. Quaas A, Dokras A: Diagnosis and Treatment of Unexplained Infertility. Rev. Obstet. Gynecol. 2008; 1: 69–76. PubMed Abstract
3. Agarwal A, Parekh N, Panner Selvam MK, et al.: Male Oxidative Stress Infertility (MOSI): Proposed Terminology and Clinical Practice Guidelines for Management of Idiopathic Male Infertility. World J. Mens Health. 2019; 37: 296–312. PubMed Abstract | Publisher Full Text
4. Esteves SC, Zini A, Coward RM, et al.: Sperm DNA Fragmentation Testing: Summary Evidence and Clinical Practice Recommendations. Andrologia. 2021; 53: e13874. PubMed Abstract | Publisher Full Text
5. Agarwal A, Baskaran S, Parekh N, et al.: Male Infertility. Lancet. 2021; 397: 319–333. Publisher Full Text
6. Arafa M, Henkel R, Agarwal A, et al.: Correlation of Oxidation-Reduction Potential with Hormones, Semen Parameters and Testicular Volume. Andrologia. 2019; 51: e13258. PubMed Abstract | Publisher Full Text
7. Barbăroșie C, Agarwal A, Henkel R: Diagnostic Value of Advanced Semen Analysis in Evaluation of Male Infertility. Andrologia. 2021; 53: e13625. PubMed Abstract | Publisher Full Text
8. Casarini L, Crépieux P, Reiter E, et al.: FSH for the Treatment of Male Infertility. Int. J. Mol. Sci. 2020; 21: E2270. PubMed Abstract | Publisher Full Text
9. Krausz C, Riera-Escamilla A: Genetics of Male Infertility. Nat. Rev. Urol. 2018; 15: 369–384. Publisher Full Text
10. Lateef OM, Akintubosun MO: Sleep and Reproductive Health. J. Circadian Rhythms. 2020; 18: 1. Publisher Full Text
11. Salonia A, Bettocchi C, Carvalho J, et al.: Sexual and Reproductive Health. Uroweb. 2021.
12. Thurston L, Abbara A, Dhillo WS: Investigation and Management of Subfertility. J. Clin. Pathol. 2019; 72: 579–587. Publisher Full Text
13. Faduola P, Kolade CO: Sperm Chromatin Structure Assay Results in Nigerian Men with Unexplained Infertility. Clin. Exp. Reprod. Med. 2015; 42: 101–105. PubMed Abstract | Publisher Full Text
14. Mayorga-Torres BJM, Camargo M, Cadavid ÁP, et al.: Are Oxidative Stress Markers Associated with Unexplained Male Infertility?. Andrologia. 2017; 49: e12659. PubMed Abstract | Publisher Full Text
15. Dutta S, Henkel R, Agarwal A: Comparative Analysis of Tests Used to Assess Sperm Chromatin Integrity and DNA Fragmentation. Andrologia. 2021; 53: e13718. PubMed Abstract | Publisher Full Text
16. Evenson DP: Evaluation of Sperm Chromatin Structure and DNA Strand Breaks Is an Important Part of Clinical Male Fertility Assessment. Transl. Androl. Urol. 2017; 6: S495–S500. PubMed Abstract | Publisher Full Text
17. Rashki Ghaleno L, Alizadeh A, Drevet JR, et al.: Oxidation of Sperm DNA and Male Infertility. Antioxidants (Basel). 2021; 10: 97. Publisher Full Text
18. Martinez M, Majzoub A: Best Laboratory Practices and Therapeutic Interventions to Reduce Sperm DNA Damage. Andrologia. 2021; 53: e13736. PubMed Abstract | Publisher Full Text
19. Nguyen TT, Trieu TS, Tran TO, et al.: Evaluation of Sperm DNA Fragmentation Index, Zinc Concentration and Seminal Parameters from Infertile Men with Varicocele. Andrologia. 2019; 51: e13184. Publisher Full Text
20. Ribas-Maynou J, Yeste M, Becerra-Tomás N, et al.: Clinical Implications of Sperm DNA Damage in IVF and ICSI: Updated Systematic Review and Meta-Analysis. Biol. Rev. Camb. Philos. Soc. 2021; 96: 1284–1300. PubMed Abstract | Publisher Full Text
21. Agarwal A, Farkouh A, Parekh N, et al.: Sperm DNA Fragmentation: A Critical Assessment of Clinical Practice Guidelines. World J. Mens Health. 2022; 40: 30–37. PubMed Abstract | Publisher Full Text
22. Abdelbaki SA, Sabry JH, Al-Adl AM, et al.: The Impact of Coexisting Sperm DNA Fragmentation and Seminal Oxidative Stress on the Outcome of Varicocelectomy in Infertile Patients: A Prospective Controlled Study. Arab. J. Urol. 2017; 15: 131–139. PubMed Abstract | Publisher Full Text
23. Al Omrani B, Al Eisa N, Javed M, et al.: Associations of Sperm DNA Fragmentation with Lifestyle Factors and Semen Parameters of Saudi Men and Its Impact on ICSI Outcome. Reprod. Biol. Endocrinol. 2018; 16: 49. PubMed Abstract | Publisher Full Text
24. Cissen M, van Wely M , Scholten I, et al.: Measuring Sperm DNA Fragmentation and Clinical Outcomes of Medically Assisted Reproduction: A Systematic Review and Meta-Analysis. PLoS One. 2016; 11: e0165125. PubMed Abstract | Publisher Full Text
25. Cheng H, Han M, Ding J, et al.: Importance of a Semen Analysis Report for Determining the Relationship between SCSA Sperm DNA Fragmentation Index and Assisted Reproductive Technology Pregnancy Rate. Reprod. Biol. 2020; 20: 460–464. Publisher Full Text
26. Leach M, Aitken RJ, Sacks G: Sperm DNA Fragmentation Abnormalities in Men from Couples with a History of Recurrent Miscarriage. Aust. N. Z. J. Obstet. Gynaecol. 2015; 55: 379–383. Publisher Full Text
27. Oleszczuk K, Giwercman A, Bungum M: Sperm Chromatin Structure Assay in Prediction of in vitro Fertilization Outcome. Andrology. 2016; 4: 290–296. Publisher Full Text
28. Simon L, Emery BR, Carrell DT: Review: Diagnosis and Impact of Sperm DNA Alterations in Assisted Reproduction. Best Pract. Res. Clin. Obstet. Gynaecol. 2017; 44: 38–56. PubMed Abstract | Publisher Full Text
29. Vinnakota C, Cree L, Peek J, et al.: Incidence of High Sperm DNA Fragmentation in a Targeted Population of Subfertile Men. Syst. Biol. Reprod. Med. 2019; 65: 451–457. PubMed Abstract | Publisher Full Text
30. Yang H, Li G, Jin H, et al.: The Effect of Sperm DNA Fragmentation Index on Assisted Reproductive Technology Outcomes and Its Relationship with Semen Parameters and Lifestyle. Transl. Androl. Urol. 2019; 8: 356–365. PubMed Abstract | Publisher Full Text
31. Khalafalla K, Majzoub A, Elbardisi H, et al.: The Effect of Sperm DNA Fragmentation on Intracytoplasmic Sperm Injection Outcome. Andrologia. 2021; 53: e14180. PubMed Abstract | Publisher Full Text
32. World Health Organization: WHO Laboratory Manual for the Examination and Processing of Human Semen. 5th ed.Geneva, Switzerland:World Health Organization Press;2010.
33. Gill K, Kups M, Harasny P, et al.: The Negative Impact of Varicocele on Basic Semen Parameters, Sperm Nuclear DNA Dispersion and Oxidation-Reduction Potential in Semen. Int. J. Environ. Res. Public Health. 2021; 18: 5977. PubMed Abstract | Publisher Full Text
34. Gill K, Rosiak-Gill A, Jakubik J, et al.: The Higher Risk for Sperm DNA Damage in Infertile Men. Ginekol. Pol. 2019; 90: 684–691. PubMed Abstract | Publisher Full Text
35. Gill K, Jakubik J, Rosiak-Gill A, et al.: Utility and Predictive Value of Human Standard Semen Parameters and Sperm DNA Dispersion for Fertility Potential. Int. J. Environ. Res. Public Health. 2019; 16: E2004. PubMed Abstract | Publisher Full Text
36. Jakubik-Uljasz J, Gill K, Rosiak-Gill A, et al.: Relationship between Sperm Morphology and Sperm DNA Dispersion. Transl. Androl. Urol. 2020; 9: 405–415. PubMed Abstract | Publisher Full Text
37. Pedersen MR, Rafaelsen SR, Møller H, et al.: Testicular Microlithiasis and Testicular Cancer: Review of the Literature. Int. Urol. Nephrol. 2016; 48: 1079–1086. PubMed Abstract | Publisher Full Text
38. Elbashir S, Magdi Y, Rashed A, et al.: Relationship between Sperm Progressive Motility and DNA Integrity in Fertile and Infertile Men.2018. Publisher Full Text
39. Lotti F, Frizza F, Balercia G, et al.: The European Academy of Andrology (EAA) Ultrasound Study on Healthy, Fertile Men: Clinical, Seminal and Biochemical Characteristics. Andrology. 2020; 8: 1005–1020. PubMed Abstract | Publisher Full Text
40. Slama R, Eustache F, Ducot B, et al.: Time to Pregnancy and Semen Parameters: A Cross-Sectional Study among Fertile Couples from Four European Cities. Hum. Reprod. 2002; 17: 503–515. PubMed Abstract | Publisher Full Text
41. Cao X, Cui Y, Zhang X, et al.: The Correlation of Sperm Morphology with Unexplained Recurrent Spontaneous Abortion: A Systematic Review and Meta-Analysis. Oncotarget. 2017; 8: 55646–55656. PubMed Abstract | Publisher Full Text
42. Jayasena CN, Radia UK, Figueiredo M, et al.: Reduced Testicular Steroidogenesis and Increased Semen Oxidative Stress in Male Partners as Novel Markers of Recurrent Miscarriage. Clin. Chem. 2019; 65: 161–169. PubMed Abstract | Publisher Full Text
43. Li B, Ma Y, Huang J, et al.: Probing the Effect of Human Normal Sperm Morphology Rate on Cycle Outcomes and Assisted Reproductive Methods Selection. PLoS One. 2014; 9: e113392. PubMed Abstract | Publisher Full Text
44. McQueen DB, Zhang J, Robins JC: Sperm DNA Fragmentation and Recurrent Pregnancy Loss: A Systematic Review and Meta-Analysis. Fertil. Steril. 2019; 112: 54–60.e3. Publisher Full Text
45. Gatimel N, Moreau J, Parinaud J, et al.: Sperm Morphology: Assessment, Pathophysiology, Clinical Relevance, and State of the Art in 2017. Andrology. 2017; 5: 845–862. PubMed Abstract | Publisher Full Text
46. Shabtaie SA, Gerkowicz SA, Kohn TP, et al.: Role of Abnormal Sperm Morphology in Predicting Pregnancy Outcomes. Curr. Urol. Rep. 2016; 17: 67. PubMed Abstract | Publisher Full Text
47. Danis RB, Samplaski MK: Sperm Morphology: History, Challenges, and Impact on Natural and Assisted Fertility. Curr. Urol. Rep. 2019; 20: 43. Publisher Full Text
48. Kohn TP, Kohn JR, Lamb DJ: Role of Sperm Morphology in Deciding Between Various Assisted Reproduction Technologies. Eur. Urol. Focus. 2018; 4: 311–313. PubMed Abstract | Publisher Full Text
49. Kovac JR, Smith RP, Cajipe M, et al.: Men with a Complete Absence of Normal Sperm Morphology Exhibit High Rates of Success without Assisted Reproduction. Asian J. Androl. 2017; 19: 39–42. PubMed Abstract | Publisher Full Text
50. Björndahl L: The Usefulness and Significance of Assessing Rapidly Progressive Spermatozoa. Asian J. Androl. 2010; 12: 33–35. Publisher Full Text
51. Simon L, Lewis SEM: Sperm DNA Damage or Progressive Motility: Which One Is the Better Predictor of Fertilization in Vitro?. Syst. Biol. Reprod. Med. 2011; 57: 133–138. Publisher Full Text
52. Kuriya A, Agbo C, Dahan MH: Do Pregnancy Rates Differ with Intra-Uterine Insemination When Different Combinations of Semen Analysis Parameters Are Abnormal?. J. Turk. Ger. Gynecol. Assoc. 2018; 19: 57–64. PubMed Abstract | Publisher Full Text
53. Patel AS, Leong JY, Ramasamy R: Prediction of Male Infertility by the World Health Organization Laboratory Manual for Assessment of Semen Analysis: A Systematic Review. Arab. J. Urol. 2018; 16: 96–102. PubMed Abstract | Publisher Full Text
54. Zivlak-Radulovic N, Banjac V, Miskovic M: Hyperprolactinemia as a Side Effect of Long-Acting Injectable Risperidone Therapy or a Symptom of Klinefelter Syndrome - a Diagnostic and Therapeutic Dilemma. Psychiatr. Danub. 2018; 30: 99–103. PubMed Abstract | Publisher Full Text
55. Dabbous Z, Atkin SL: Hyperprolactinaemia in Male Infertility: Clinical Case Scenarios. Arab. J. Urol. 2018; 16: 44–52. PubMed Abstract | Publisher Full Text
56. La Vignera S, Vita R: Thyroid Dysfunction and Semen Quality. Int. J. Immunopathol. Pharmacol. 2018; 32: 2058738418775241. PubMed Abstract | Publisher Full Text
57. Condorelli R, Calogero AE, La Vignera S: Relationship between Testicular Volume and Conventional or Nonconventional Sperm Parameters. Int. J. Endocrinol. 2013; 2013: 145792–145796. PubMed Abstract | Publisher Full Text
58. Huang I-S, Huang WJ, Lin AT: Distinguishing Non-Obstructive Azoospermia from Obstructive Azoospermia in Taiwanese Patients by Hormone Profile and Testis Size. J. Chin. Med. Assoc. 2018; 81: 531–535. PubMed Abstract | Publisher Full Text
59. Nieschlag E, Vorona E: MECHANISMS IN ENDOCRINOLOGY: Medical Consequences of Doping with Anabolic Androgenic Steroids: Effects on Reproductive Functions. Eur. J. Endocrinol. 2015; 173: R47–R58. PubMed Abstract | Publisher Full Text
60. Sakamoto H, Yajima T, Nagata M, et al.: Relationship between Testicular Size by Ultrasonography and Testicular Function: Measurement of Testicular Length, Width, and Depth in Patients with Infertility. Int. J. Urol. 2008; 15: 529–533. PubMed Abstract | Publisher Full Text
61. Colpi GM, Francavilla S, Haidl G, et al.: European Academy of Andrology Guideline Management of Oligo-Astheno-Teratozoospermia. Andrology. 2018; 6: 513–524. PubMed Abstract | Publisher Full Text
62. Corinne TM, Anatole PC, Jeanne NY: Comparison of Serum Inhibin B and Follicle-Stimulating Hormone (FSH) Level between Normal and Infertile Men in Yaoundé. Int. J. Reprod. Med. 2020; 2020: 4765809–4765809. PubMed Abstract | Publisher Full Text
63. Uhler ML, Zinaman MJ, Brown CC, et al.: Relationship between Sperm Characteristics and Hormonal Parameters in Normal Couples. Fertil. Steril. 2003; 79 Suppl 3: 1535–1542. PubMed Abstract | Publisher Full Text
64. Mabeck LM, Jensen MS, Toft G, et al.: Fecundability According to Male Serum Inhibin B--a Prospective Study among First Pregnancy Planners. Hum. Reprod. 2005; 20: 2909–2915. PubMed Abstract | Publisher Full Text
65. Meeker JD, Godfrey-Bailey L, Hauser R: Relationships between Serum Hormone Levels and Semen Quality among Men from an Infertility Clinic. J. Androl. 2007; 28: 397–406. Publisher Full Text
66. Ring JD, Lwin AA, Köhler TS: Current Medical Management of Endocrine-Related Male Infertility. Asian J. Androl. 2016; 18: 357–363. PubMed Abstract | Publisher Full Text
67. Vilar L, Abucham J, Albuquerque JL, et al.: Controversial Issues in the Management of Hyperprolactinemia and Prolactinomas - An Overview by the Neuroendocrinology Department of the Brazilian Society of Endocrinology and Metabolism. Arch. Endocrinol. Metab. 2018; 62: 236–263. Publisher Full Text
68. Krajewska-Kulak E, Sengupta P: Thyroid Function in Male Infertility. Front. Endocrinol. 2013; 4. PubMed Abstract | Publisher Full Text
69. Krassas GE, Papadopoulou F, Tziomalos K, et al.: Hypothyroidism Has an Adverse Effect on Human Spermatogenesis: A Prospective, Controlled Study. Thyroid. 2008; 18: 1255–1259. PubMed Abstract | Publisher Full Text
70. Singh R, Hamada J, Agarwal A: Thyroid Hormones in Male Reproduction and Fertility. Open Reprod. Sci. J. 2011; 3: 98–104. Publisher Full Text
71. Fozooni R, Jafarzadeh Shirazi MR, Saedi S, et al.: Male Subfertility Effects of Sub-Chronic Ethanol Exposure during Stress in a Rat Model. Alcohol. 2021; 96: 63–71. PubMed Abstract | Publisher Full Text
72. Perumal P, Chang S, De AK, et al.: Slow Release Exogenous Melatonin Modulates Scrotal Circumference and Testicular Parameters, Libido, Endocrinological Profiles and Antioxidant and Oxidative Stress Profiles in Mithun. Theriogenology. 2020; 154: 1–10. Publisher Full Text
73. Simopoulou M, Philippou A, Maziotis E, et al.: Association between Male Infertility and Seminal Plasma Levels of Growth Hormone and Insulin-like Growth Factor-1. Andrologia. 2018; 50: e13048. PubMed Abstract | Publisher Full Text
74. Pedersen MR, Osther PJS, Rafaelsen SR: Ultrasound Evaluation of Testicular Volume in Patients with Testicular Microlithiasis. Ultrasound Int. Open. 2018; 04: E99–E103. Publisher Full Text
75. Bahk JY, Jung JH, Jin LM, et al.: Cut-off Value of Testes Volume in Young Adults and Correlation among Testes Volume, Body Mass Index, Hormonal Level, and Seminal Profiles. Urology. 2010; 75: 1318–1323. PubMed Abstract | Publisher Full Text
76. Lotti F, Tamburrino L, Marchiani S, et al.: DNA Fragmentation in Two Cytometric Sperm Populations: Relationship with Clinical and Ultrasound Characteristics of the Male Genital Tract. Asian J. Androl. 2017; 19: 272–279. PubMed Abstract | Publisher Full Text
77. Schurich M, Aigner F, Frauscher F, et al.: The Role of Ultrasound in Assessment of Male Fertility. Eur. J. Obstet. Gynecol. Reprod. Biol. 2009; 144(Suppl 1): S192–S198. Publisher Full Text
78. Zorn B, Golob B, Ihan A, et al.: Apoptotic Sperm Biomarkers and Their Correlation with Conventional Sperm Parameters and Male Fertility Potential. J. Assist. Reprod. Genet. 2012; 29: 357–364. PubMed Abstract | Publisher Full Text
79. Lu J-C, Jing J, Dai J-Y, et al.: Body Mass Index, Waist-to-Hip Ratio, Waist Circumference and Waist-to-Height Ratio Cannot Predict Male Semen Quality: A Report of 1231 Subfertile Chinese Men. Andrologia. 2015; 47: 1047–1054. Publisher Full Text
80. Wei T-C, Huang WJ, Lin ATL, et al.: The Role of Hormones on Semen Parameters in Patients with Idiopathic or Varicocele-Related Oligoasthenoteratozoospermia (OAT) Syndrome. J. Chin. Med. Assoc. 2013; 76: 624–628. PubMed Abstract | Publisher Full Text
81. Smit M, Romijn JC, Wildhagen MF, et al.: Sperm Chromatin Structure Is Associated with the Quality of Spermatogenesis in Infertile Patients. Fertil. Steril. 2010; 94: 1748–1752. PubMed Abstract | Publisher Full Text
82. Wdowiak A, Raczkiewicz D, Stasiak M, et al.: Levels of FSH, LH and Testosterone, and Sperm DNA Fragmentation. Neuro Endocrinol. Lett. 2014; 35: 73–79. PubMed Abstract
83. Lu J-C, Jing J, Chen L, et al.: Analysis of Human Sperm DNA Fragmentation Index (DFI) Related Factors: A Report of 1010 Subfertile Men in China. Reprod. Biol. Endocrinol. 2018; 16: 23. PubMed Abstract | Publisher Full Text
84. Amjad S, Baig M, Zahid N, et al.: Association between Leptin, Obesity, Hormonal Interplay and Male Infertility. Andrologia. 2019; 51: e13147. Publisher Full Text
85. Bundhun PK, Janoo G, Bhurtu A, et al.: Tobacco Smoking and Semen Quality in Infertile Males: A Systematic Review and Meta-Analysis. BMC Public Health. 2019; 19: 36. PubMed Abstract | Publisher Full Text
86. Ilacqua A, Izzo G, Emerenziani GP, et al.: Lifestyle and Fertility: The Influence of Stress and Quality of Life on Male Fertility. Reprod. Biol. Endocrinol. 2018; 16: 115. PubMed Abstract | Publisher Full Text
87. Kesari KK, Agarwal A, Henkel R: Radiations and Male Fertility. Reprod. Biol. Endocrinol. 2018; 16: 118. PubMed Abstract | Publisher Full Text
88. Miranda-Contreras L, Gómez-Pérez R, Rojas G, et al.: Occupational Exposure to Organophosphate and Carbamate Pesticides Affects Sperm Chromatin Integrity and Reproductive Hormone Levels among Venezuelan Farm Workers. J. Occup. Health. 2013; 55: 195–203. PubMed Abstract | Publisher Full Text
89. Palmer NO, Bakos HW, Fullston T, et al.: Impact of Obesity on Male Fertility, Sperm Function and Molecular Composition. Spermatogenesis. 2012; 2: 253–263. PubMed Abstract | Publisher Full Text
90. Semet M, Paci M, Saïas-Magnan J, et al.: The Impact of Drugs on Male Fertility: A Review. Andrology. 2017; 5: 640–663. Publisher Full Text
91. Erenpreiss J, Elzanaty S, Giwercman A: Sperm DNA Damage in Men from Infertile Couples. Asian J. Androl. 2008; 10: 786–790. Publisher Full Text
92. Belloc S, Benkhalifa M, Cohen-Bacrie M, et al.: Sperm Deoxyribonucleic Acid Damage in Normozoospermic Men Is Related to Age and Sperm Progressive Motility. Fertil. Steril. 2014; 101: 1588–1593. PubMed Abstract | Publisher Full Text
93. Giwercman A, Richthoff J, Hjøllund H, et al.: Correlation between Sperm Motility and Sperm Chromatin Structure Assay Parameters. Fertil. Steril. 2003; 80: 1404–1412. Publisher Full Text
94. Boushaba S, Belaaloui G: Sperm DNA Fragmentation and Standard Semen Parameters in Algerian Infertile Male Partners. World J. Mens Health. 2015; 33: 1–7. PubMed Abstract | Publisher Full Text
95. Bounartzi T, Dafopoulos K, Anifandis G, et al.: Pregnancy Prediction by Free Sperm DNA and Sperm DNA Fragmentation in Semen Specimens of IVF/ICSI-ET Patients. Hum. Fertil. (Camb.). 2016; 19: 56–62. PubMed Abstract | Publisher Full Text
96. Cohen-Bacrie P, Belloc S, Ménézo YJR, et al.: Correlation between DNA Damage and Sperm Parameters: A Prospective Study of 1,633 Patients. Fertil. Steril. 2009; 91: 1801–1805. PubMed Abstract | Publisher Full Text
97. Bungum M, Bungum L, Giwercman A: Sperm Chromatin Structure Assay (SCSA): A Tool in Diagnosis and Treatment of Infertility. Asian J. Androl. 2011; 13: 69–75. PubMed Abstract | Publisher Full Text
98. Bungum M: Sperm DNA Integrity Assessment: A New Tool in Diagnosis and Treatment of Fertility. Obstet. Gynecol. Int. 2012; 2012: 531042. Publisher Full Text
99. Evenson DP: The Sperm Chromatin Structure Assay (SCSA(^®)) and Other Sperm DNA Fragmentation Tests for Evaluation of Sperm Nuclear DNA Integrity as Related to Fertility. Anim. Reprod. Sci. 2016; 169: 56–75. PubMed Abstract | Publisher Full Text
100. Javed A, Talkad MS, Ramaiah MK: Evaluation of Sperm DNA Fragmentation Using Multiple Methods: A Comparison of Their Predictive Power for Male Infertility. Clin. Exp. Reprod. Med. 2019; 46: 14–21. PubMed Abstract | Publisher Full Text
101. Majzoub A, Arafa M, Mahdi M, et al.: Oxidation-Reduction Potential and Sperm DNA Fragmentation, and Their Associations with Sperm Morphological Anomalies amongst Fertile and Infertile Men. Arab. J. Urol. 2018; 16: 87–95. PubMed Abstract | Publisher Full Text
102. Wiweko B, Utami P: Predictive Value of Sperm Deoxyribonucleic Acid (DNA) Fragmentation Index in Male Infertility. Basic Clin. Androl. 2017; 27: 1. PubMed Abstract | Publisher Full Text
103. Giwercman A, Lindstedt L, Larsson M, et al.: Sperm Chromatin Structure Assay as an Independent Predictor of Fertility in Vivo: A Case-Control Study. Int. J. Androl. 2010; 33: e221–e227. PubMed Abstract | Publisher Full Text
104. Baird DC, Meyers GJ, Hu JS: Testicular Cancer: Diagnosis and Treatment. Am. Fam. Physician. 2018; 97: 261–268. PubMed Abstract
105. Wang F, Zhao S, Xie Y, et al.: Novo SOX10 Nonsense Mutation in a Patient with Kallmann Syndrome, Deafness, Iris Hypopigmentation, and Hyperthyroidism. Ann. Clin. Lab. Sci. 2018; 48: 248–252. PubMed Abstract
106. Santi D, Crépieux P, Reiter E, et al.: Follicle-Stimulating Hormone (FSH) Action on Spermatogenesis: A Focus on Physiological and Therapeutic Roles. J. Clin. Med. 2020; 9: E1014. PubMed Abstract | Publisher Full Text
107. Holota H, Thirouard L, Monrose M, et al.: FXRα Modulates Leydig Cell Endocrine Function in Mouse. Mol. Cell. Endocrinol. 2020; 518: 110995. PubMed Abstract | Publisher Full Text
108. Sertkaya Z, Tokuç E, Özkaya F, et al.: Acute Effect of Microdissection Testicular Sperm Extraction on Blood Total Testosterone and Luteinising Hormone Levels. Andrologia. 2020; 52: e13655. PubMed Abstract | Publisher Full Text
109. Kups M, Gill K, Rosiak-Gill A, et al.: Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study (Final) [Data set]. Zenodo. 2022; Publisher Full Text
110. Kups M, Gill K, Rosiak-Gill A, et al.: Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study. [Data set]. Zenodo. 2022. Publisher Full Text
111. Kups M, Gill K, Rosiak-Gill A, et al.: Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study. [Data set]. Zenodo. 2022. Publisher Full Text

Comments on this article Comments (0)

Version 1

VERSION 1 PUBLISHED 30 May 2022

Author details Author details

¹ Department of Urology and Oncological Urology, Regional Specialist Hospital in Szczecin, Szczecin, West Pomeranian Voivodeship, 71-455, Poland
² The Fertility Partnership Vitrolive in Szczecin, Szczecin, West Pomeranian Voivodeship, 70-483, Poland
³ Department of Histology and Developmental Biology, Faculty of Health Sciences, Pomeranian Medical University in Szczecin, Szczecin, West Pomeranian Voivodeship, 71-210, Poland
⁴ Department of Urology and Urological Oncology, Faculty of Medicine and Dentistry, Pomeranian Medical University in Szczecin, Szczecin, West Pomeranian Voivodeship, 70-111, Poland
⁵ Department of Perinatology, Obstetrics and Gynecology, Faculty of Medicine and Dentistry, Pomeranian Medical University in Szczecin, Police, West Pomeranian Voivodeship, 72-010, Poland
⁶ Department of Gynecology and Reproductive Health, Faculty of Health Sciences, Pomeranian Medical University in Szczecin, Szczecin, West Pomeranian Voivodeship, 71-210, Poland
⁷ Department of Obstetrics and Pathology of Pregnancy, Faculty of Health Sciences, Pomeranian Medical University in Szczecin, Szczecin, Szczecin, West Pomeranian Voivodeship, 71-210, Poland

Michal Kups
Roles: Investigation, Writing – Original Draft Preparation, Writing – Review & Editing

Kamil Gill
Roles: Conceptualization, Formal Analysis, Methodology, Visualization, Writing – Original Draft Preparation, Writing – Review & Editing

Aleksandra Rosiak-Gill
Roles: Formal Analysis

Patryk Harasny
Roles: Investigation

Tomasz Machalowski
Roles: Investigation

Marta Grabowska
Roles: Formal Analysis

Rafal Kurzawa
Roles: Investigation

Olimpia Sipak
Roles: Investigation

Malgorzata Piasecka
Roles: Conceptualization, Formal Analysis, Funding Acquisition, Methodology, Project Administration, Supervision, Visualization, Writing – Original Draft Preparation, Writing – Review & Editing

Competing interests

No competing interests were disclosed.

Grant information

This research was supported by Pomeranian Medical University in Szczecin, Poland, grant number WNoZ-322-01/S/19/2021 to M.P.
The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Article Versions (1)

version 1

Published: 30 May 2022, 11:591

https://doi.org/10.12688/f1000research.121622.1

Copyright

© 2022 Kups M et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Download

Export To

metrics

	Views	Downloads
F1000Research	-	-
PubMed Central Data from PMC are received and updated monthly.	-	-

Citations

0

SEE MORE DETAILS

CITE

how to cite this article

Kups M, Gill K, Rosiak-Gill A et al. Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study [version 1; peer review: 1 approved, 2 approved with reservations]. F1000Research 2022, 11:591 (https://doi.org/10.12688/f1000research.121622.1)

NOTE: If applicable, it is important to ensure the information in square brackets after the title is included in all citations of this article.

track

receive updates on this article

Track an article to receive email alerts on any updates to this article.

Open Peer Review

Current Reviewer Status: ?

Key to Reviewer Statuses VIEW HIDE

ApprovedThe paper is scientifically sound in its current form and only minor, if any, improvements are suggested

Approved with reservations A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit.

Not approvedFundamental flaws in the paper seriously undermine the findings and conclusions

Version 1

VERSION 1

PUBLISHED 30 May 2022

Views

8

Reviewer Report 20 Feb 2024

Eva Tvrdá, Slovak University of Agriculture, Nitra, Slovakia

Approved with Reservations

https://doi.org/10.5256/f1000research.133505.r238146

This is a nicely written and appropriately designed study evaluating and interconnecting standard semen parameters in infertile men.
While I appreciate papers that turn "back to the basics" and re-evaluate/re-verify the importance of traditional parameters of sperm quality, I ... Continue reading

This is a nicely written and appropriately designed study evaluating and interconnecting standard semen parameters in infertile men.
While I appreciate papers that turn "back to the basics" and re-evaluate/re-verify the importance of traditional parameters of sperm quality, I am missing the originality of the study. The authors should place more emphasis on the originality and scientific rigour of their experiments as well as provide a solid justification as to why such types of papers on generally well-accepted knowledge are necessary.
Finally, the title of the article hints evaluation of biomarkers of male infertility. if so, what are the biomarkers and why should these be more of a center focus for the andrologists?

Is the work clearly and accurately presented and does it cite the current literature?

Yes
Is the study design appropriate and is the work technically sound?

Yes
Are sufficient details of methods and analysis provided to allow replication by others?

Yes
If applicable, is the statistical analysis and its interpretation appropriate?

Yes
Are all the source data underlying the results available to ensure full reproducibility?

Yes
Are the conclusions drawn adequately supported by the results?

Yes

Competing Interests: No competing interests were disclosed.

Reviewer Expertise: Sperm quality, oxidative stress, antioxidants, molecular andrology.

I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above.

CITE

Report a concern

Respond or Comment

Views

6

Reviewer Report 20 Feb 2024

Nafisa Balasinor, ICMR-National Institute for Research in Reproductive and Child Health, Parel, India

Approved with Reservations

https://doi.org/10.5256/f1000research.133505.r238141

In the present manuscript the authors have studied the association between standard semen parameters, testicular volume, reproductive hormones profile and the sperm nuclear DNA fragmentation.
Overall the study is well done and has clinical utility. However, there are few ... Continue reading

In the present manuscript the authors have studied the association between standard semen parameters, testicular volume, reproductive hormones profile and the sperm nuclear DNA fragmentation.
Overall the study is well done and has clinical utility. However, there are few points which needs to be addressed:
1. How was the sample size of 130 calculated?
2. Was infertility due to known female factor ruled out?
3. The authors should mention how they categorized abnormal hormonal levels.
4. Table 3: volume in the right testis of normal volume group ranged between 9 to 25ml. However, testicular volume of > or equal to12 was taken as normal testicular volume. Please check the data.
5. Table 5: Men with abnormal hormonal profile group had increase in all hormones except Testosterone. However, the values do not indicate so. Hence need to check if correct statistical tool was used.
6. The authors need to check testosterone levels in this infertile group as compared to fertile group or normal range.
7. SDF in men with normal semen and sperm parameters should be analyzed. If SDF is found in this group too than the importance to including SDF in male infertility work out will be evident.
8. It would be interesting to see the sperm parameters in individuals in which all 3 parameters, namely, Volume, hormone profile and SDF.

Is the work clearly and accurately presented and does it cite the current literature?

Yes
Is the study design appropriate and is the work technically sound?

Partly
Are sufficient details of methods and analysis provided to allow replication by others?

Partly
If applicable, is the statistical analysis and its interpretation appropriate?

I cannot comment. A qualified statistician is required.
Are all the source data underlying the results available to ensure full reproducibility?

Yes
Are the conclusions drawn adequately supported by the results?

Partly

Competing Interests: No competing interests were disclosed.

Reviewer Expertise: Reproductive endocrinology, male fertility and epigenetics

I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above.

CITE

Report a concern

Respond or Comment

Views

9

Reviewer Report 01 Feb 2024

Abdul S Ansari, Department of Zoology, University of Rajasthan, Jaipur, Rajsthan, India

Approved

https://doi.org/10.5256/f1000research.133505.r228728

In the present manuscript the author(s) evaluated etiopathogenesis of male infertility by investigation of the relationship between standard semen parameters, testicular volume, levels of reproductive hormones and the fragmentation of sperm nuclear DNA (SDF). A total of 130 subjects, divided ... Continue reading

In the present manuscript the author(s) evaluated etiopathogenesis of male infertility by investigation of the relationship between standard semen parameters, testicular volume, levels of reproductive hormones and the fragmentation of sperm nuclear DNA (SDF). A total of 130 subjects, divided into three groups, with at least one abnormal testis volume, minimum low level among any reproductive hormones and with SDF. Results obtained from the study revealed that the subjects with decreased testicular volume and abnormal levels of hormones were observed with decreased basic semen parameters. Abnormal testicular volume also had, respectively, higher percentage of SDF and enhanced levels of FSH. Subjects with high SDF level had low testicular volume and seminal parameters. Authors concluded that spermatogenesis disorders coexisted with decreased testicular volume and increased FSH levels which manifested by reduced seminal characteristics and high sperm nuclear damage. However, the research article contains several lacunae in terms of writing the manuscript, language, presentation of data, etc. Moreover, the title of the article does not match the contents of the article. It correlated several parameters.

Specific comments:
Abstract:

Page 1: Methods: The participants of the study were not patients, instead they are subjects. Modify the sentence as “Participant subjects (n=130) were divided into”.

Introduction:

Page 3, Introduction: Write full form of SCSA test followed by its abbreviation SCSA in parentheses.
Page 3, Methods (Study population): The study population , i.e., 130 male infertile subjects is very small. If the number would have been more, it is easy to draw a conclusion.
Page 3, Methods (Study population): Replace “All patients …” with All subjects …”.
Page 3, Methods (Study population): How many subjects with azoospermia and cryptozoospermia were excluded? Mention their number.
Page 3, Methods (Study population): Replace the phrase “seminal cords” with “spermatic cords”.
Page 4, Methods (Conventional semen analysis): Room temperature is not -20 °C. Correct the sentence.
Page 4, Methods (Conventional semen analysis): Add “liquefaction time” of semen as a macroscopic semen analysis parameter.
Page 4, Methods (Conventional semen analysis): The terminology total sperm count has changed by WHO as “sperm density” (million/ejaculate). Replace the same with new terminology.
Page 4, Methods (Conventional semen analysis): The sperm vitality or viability is carried out the mixing of a well mixed semen sample with eosin-nigrosin stain and and spreaded on a slide and dried. The unstained spermatozoa are regarded as alive/vital, while pink coloured sperms are considered dead. The HOS test is done with the mixing of a semen sample with a hypoosmotic solution and incubated. After incubation spermatozoa with coiled tails are counted, and calculated in percent (%), termed as HOS positive sperms. How vitality indicating both eosin-positive cells and hypoosmotic-reactive cells (HOS test-positive cells can be assessed?
Page 4, Methods (Sperm Chromatin Dispersion (SCD) test): The chromatin dispersion test carried out in the study with Halosperm G2 kit is sort of In vitro Nuclear Chromatin Decondensation (NCD) Test of WHO routinely carried out for infertility diagnosis for fertilizing ability. In the test Sperm heads decondensed which results into swelling of heads, appears as, halos. Whereas, Non-fertilizable sperm heads remain condensed due to defective nuclear chromatin. Decondensed sperms heads are counted versus condensed sperm heads and represented as percent. I am surprised that why so expensive SCD test was carried out when other several simple tests are available. A detailed comment is required from author(s) on this point.
Page 4, Methods (Hormone profile of infertile subjects): Add Inter-assay and Intra-asssay coefficient of variation of each hone assayed.

Results:

Page 6, Seminological Characteristics of study population: Replace the word “Seminological” with “Seminal”.
Page 6, Comparison … (each ≥ 12 mL): Write P (significant) value of FSH.

Discussion:

Page 14, Para 1, Line 5: This is not a thesis, instead a study or investigation. Change the phrase “This thesis” with an appropriate phrase.
Page 14, Para 3, Line 2: Complete the word “norm” as “normal”.
The write-up of discussion is too lengthy, reduced the same to half length.
Conclusions:
A brief conclusion of the study carried out similar with Abstract is required.
Data availability:
Replace the word “project” with “research article”.
References:
Reference Nos. 27, 51: Italicize the words “in Vitro”.
Reference 59: The title of the reference in upper (capital) case. Change the same as per Instruction to Authors.
References 109 to 111 are related to the present research article, thus should be delated from the list of References.
Tables:
Table 1: This table contains all assayed hormones in the study and well described in the text, therefore, should be deleted.
Tables 2, 4, 6, 8, 10: In all these tables data on Sperm nonprogressive motility (%) have been depicted. When data on Sperm progressive motility (%) are presented, there is no use of depicting this. The rest of Sperm progressive motility (%) will obviously related to the Sperm nonprogressive motility (%).
The present study contains a total 10 tables. Presentation of data in the present research article merely only in tables is not so impressive and very hard to understand by readers. Prepare histograms or bar diagrams for depiction of data.
Table 10: The data of this table should be depicted with Scatter diagrams.

Is the work clearly and accurately presented and does it cite the current literature?

Yes
Is the study design appropriate and is the work technically sound?

Yes
Are sufficient details of methods and analysis provided to allow replication by others?

Yes
If applicable, is the statistical analysis and its interpretation appropriate?

I cannot comment. A qualified statistician is required.
Are all the source data underlying the results available to ensure full reproducibility?

Yes
Are the conclusions drawn adequately supported by the results?

Partly

References

1. Kups M, Gill K, Rosiak-Gill A, Harasny P, et al.: Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study. F1000Research. 2022; 11. Publisher Full Text

Competing Interests: No competing interests were disclosed.

Reviewer Expertise: Reproductive physiology, Male contraception, infertility diagnosis, cancer diagnosis, endocrinology

I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.

CITE

Report a concern

Respond or Comment

Comments on this article Comments (0)

Version 1

VERSION 1 PUBLISHED 30 May 2022

Open Peer Review

Reviewer Status

Reviewer Reports

	Invited Reviewers
	1	2	3
Version 1 30 May 22	read	read	read

Abdul S Ansari, University of Rajasthan, Jaipur, India
Nafisa Balasinor, ICMR-National Institute for Research in Reproductive and Child Health, Parel, India
Eva Tvrdá, Slovak University of Agriculture, Nitra, Slovakia

Comments on this article

All Comments(0)

Add a comment

Sign up for content alerts

Browse by related subjects

Back to all reports

Reviewer Report

8 Views

20 Feb 2024 | for Version 1

Eva Tvrdá, Slovak University of Agriculture, Nitra, Slovakia

8 Views Cite this report Responses(0)

Approved With Reservations

This is a nicely written and appropriately designed study evaluating and interconnecting standard semen parameters in infertile men.
While I appreciate papers that turn "back to the basics" and re-evaluate/re-verify the importance of traditional parameters of sperm quality, I am missing the originality of the study. The authors should place more emphasis on the originality and scientific rigour of their experiments as well as provide a solid justification as to why such types of papers on generally well-accepted knowledge are necessary.
Finally, the title of the article hints evaluation of biomarkers of male infertility. if so, what are the biomarkers and why should these be more of a center focus for the andrologists?

Is the work clearly and accurately presented and does it cite the current literature?

Yes
Is the study design appropriate and is the work technically sound?

Yes
Are sufficient details of methods and analysis provided to allow replication by others?

Yes
If applicable, is the statistical analysis and its interpretation appropriate?

Yes
Are all the source data underlying the results available to ensure full reproducibility?

Yes
Are the conclusions drawn adequately supported by the results?

Yes

Competing Interests

No competing interests were disclosed.

Reviewer Expertise

Sperm quality, oxidative stress, antioxidants, molecular andrology.

I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above.

Respond to this report

Responses (0)

Back to all reports

Reviewer Report

6 Views

20 Feb 2024 | for Version 1

Nafisa Balasinor, ICMR-National Institute for Research in Reproductive and Child Health, Parel, India

6 Views Cite this report Responses(0)

Approved With Reservations

In the present manuscript the authors have studied the association between standard semen parameters, testicular volume, reproductive hormones profile and the sperm nuclear DNA fragmentation.
Overall the study is well done and has clinical utility. However, there are few points which needs to be addressed:
1. How was the sample size of 130 calculated?
2. Was infertility due to known female factor ruled out?
3. The authors should mention how they categorized abnormal hormonal levels.
4. Table 3: volume in the right testis of normal volume group ranged between 9 to 25ml. However, testicular volume of > or equal to12 was taken as normal testicular volume. Please check the data.
5. Table 5: Men with abnormal hormonal profile group had increase in all hormones except Testosterone. However, the values do not indicate so. Hence need to check if correct statistical tool was used.
6. The authors need to check testosterone levels in this infertile group as compared to fertile group or normal range.
7. SDF in men with normal semen and sperm parameters should be analyzed. If SDF is found in this group too than the importance to including SDF in male infertility work out will be evident.
8. It would be interesting to see the sperm parameters in individuals in which all 3 parameters, namely, Volume, hormone profile and SDF.

Is the work clearly and accurately presented and does it cite the current literature?

Yes
Is the study design appropriate and is the work technically sound?

Partly
Are sufficient details of methods and analysis provided to allow replication by others?

Partly
If applicable, is the statistical analysis and its interpretation appropriate?

I cannot comment. A qualified statistician is required.
Are all the source data underlying the results available to ensure full reproducibility?

Yes
Are the conclusions drawn adequately supported by the results?

Partly

Competing Interests

No competing interests were disclosed.

Reviewer Expertise

Reproductive endocrinology, male fertility and epigenetics

I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above.

Respond to this report

Responses (0)

Back to all reports

Reviewer Report

9 Views

01 Feb 2024 | for Version 1

Abdul S Ansari, Department of Zoology, University of Rajasthan, Jaipur, Rajsthan, India

9 Views Cite this report Responses(0)

Approved

In the present manuscript the author(s) evaluated etiopathogenesis of male infertility by investigation of the relationship between standard semen parameters, testicular volume, levels of reproductive hormones and the fragmentation of sperm nuclear DNA (SDF). A total of 130 subjects, divided into three groups, with at least one abnormal testis volume, minimum low level among any reproductive hormones and with SDF. Results obtained from the study revealed that the subjects with decreased testicular volume and abnormal levels of hormones were observed with decreased basic semen parameters. Abnormal testicular volume also had, respectively, higher percentage of SDF and enhanced levels of FSH. Subjects with high SDF level had low testicular volume and seminal parameters. Authors concluded that spermatogenesis disorders coexisted with decreased testicular volume and increased FSH levels which manifested by reduced seminal characteristics and high sperm nuclear damage. However, the research article contains several lacunae in terms of writing the manuscript, language, presentation of data, etc. Moreover, the title of the article does not match the contents of the article. It correlated several parameters.

Specific comments:
Abstract:

Page 1: Methods: The participants of the study were not patients, instead they are subjects. Modify the sentence as “Participant subjects (n=130) were divided into”.

Introduction:

Page 3, Introduction: Write full form of SCSA test followed by its abbreviation SCSA in parentheses.
Page 3, Methods (Study population): The study population , i.e., 130 male infertile subjects is very small. If the number would have been more, it is easy to draw a conclusion.
Page 3, Methods (Study population): Replace “All patients …” with All subjects …”.
Page 3, Methods (Study population): How many subjects with azoospermia and cryptozoospermia were excluded? Mention their number.
Page 3, Methods (Study population): Replace the phrase “seminal cords” with “spermatic cords”.
Page 4, Methods (Conventional semen analysis): Room temperature is not -20 °C. Correct the sentence.
Page 4, Methods (Conventional semen analysis): Add “liquefaction time” of semen as a macroscopic semen analysis parameter.
Page 4, Methods (Conventional semen analysis): The terminology total sperm count has changed by WHO as “sperm density” (million/ejaculate). Replace the same with new terminology.
Page 4, Methods (Conventional semen analysis): The sperm vitality or viability is carried out the mixing of a well mixed semen sample with eosin-nigrosin stain and and spreaded on a slide and dried. The unstained spermatozoa are regarded as alive/vital, while pink coloured sperms are considered dead. The HOS test is done with the mixing of a semen sample with a hypoosmotic solution and incubated. After incubation spermatozoa with coiled tails are counted, and calculated in percent (%), termed as HOS positive sperms. How vitality indicating both eosin-positive cells and hypoosmotic-reactive cells (HOS test-positive cells can be assessed?
Page 4, Methods (Sperm Chromatin Dispersion (SCD) test): The chromatin dispersion test carried out in the study with Halosperm G2 kit is sort of In vitro Nuclear Chromatin Decondensation (NCD) Test of WHO routinely carried out for infertility diagnosis for fertilizing ability. In the test Sperm heads decondensed which results into swelling of heads, appears as, halos. Whereas, Non-fertilizable sperm heads remain condensed due to defective nuclear chromatin. Decondensed sperms heads are counted versus condensed sperm heads and represented as percent. I am surprised that why so expensive SCD test was carried out when other several simple tests are available. A detailed comment is required from author(s) on this point.
Page 4, Methods (Hormone profile of infertile subjects): Add Inter-assay and Intra-asssay coefficient of variation of each hone assayed.

Results:

Page 6, Seminological Characteristics of study population: Replace the word “Seminological” with “Seminal”.
Page 6, Comparison … (each ≥ 12 mL): Write P (significant) value of FSH.

Discussion:

Page 14, Para 1, Line 5: This is not a thesis, instead a study or investigation. Change the phrase “This thesis” with an appropriate phrase.
Page 14, Para 3, Line 2: Complete the word “norm” as “normal”.
The write-up of discussion is too lengthy, reduced the same to half length.
Conclusions:
A brief conclusion of the study carried out similar with Abstract is required.
Data availability:
Replace the word “project” with “research article”.
References:
Reference Nos. 27, 51: Italicize the words “in Vitro”.
Reference 59: The title of the reference in upper (capital) case. Change the same as per Instruction to Authors.
References 109 to 111 are related to the present research article, thus should be delated from the list of References.
Tables:
Table 1: This table contains all assayed hormones in the study and well described in the text, therefore, should be deleted.
Tables 2, 4, 6, 8, 10: In all these tables data on Sperm nonprogressive motility (%) have been depicted. When data on Sperm progressive motility (%) are presented, there is no use of depicting this. The rest of Sperm progressive motility (%) will obviously related to the Sperm nonprogressive motility (%).
The present study contains a total 10 tables. Presentation of data in the present research article merely only in tables is not so impressive and very hard to understand by readers. Prepare histograms or bar diagrams for depiction of data.
Table 10: The data of this table should be depicted with Scatter diagrams.

Is the work clearly and accurately presented and does it cite the current literature?

Yes
Is the study design appropriate and is the work technically sound?

Yes
Are sufficient details of methods and analysis provided to allow replication by others?

Yes
If applicable, is the statistical analysis and its interpretation appropriate?

I cannot comment. A qualified statistician is required.
Are all the source data underlying the results available to ensure full reproducibility?

Yes
Are the conclusions drawn adequately supported by the results?

Partly

References

1. Kups M, Gill K, Rosiak-Gill A, Harasny P, et al.: Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study. F1000Research. 2022; 11. Publisher Full Text

Competing Interests

No competing interests were disclosed.

Reviewer Expertise

Reproductive physiology, Male contraception, infertility diagnosis, cancer diagnosis, endocrinology

I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.

Respond to this report

Responses (0)

[1] 1. Minhas S, Bettocchi C, Boeri L, et al.: European Association of Urology Guidelines on Male Sexual and Reproductive Health: 2021 Update on Male Infertility. Eur. Urol. 2021; 80: 603–620. PubMed Abstract | Publisher Full Text

[2] 2. Quaas A, Dokras A: Diagnosis and Treatment of Unexplained Infertility. Rev. Obstet. Gynecol. 2008; 1: 69–76. PubMed Abstract

[3] 3. Agarwal A, Parekh N, Panner Selvam MK, et al.: Male Oxidative Stress Infertility (MOSI): Proposed Terminology and Clinical Practice Guidelines for Management of Idiopathic Male Infertility. World J. Mens Health. 2019; 37: 296–312. PubMed Abstract | Publisher Full Text

[4] 4. Esteves SC, Zini A, Coward RM, et al.: Sperm DNA Fragmentation Testing: Summary Evidence and Clinical Practice Recommendations. Andrologia. 2021; 53: e13874. PubMed Abstract | Publisher Full Text

[5] 5. Agarwal A, Baskaran S, Parekh N, et al.: Male Infertility. Lancet. 2021; 397: 319–333. Publisher Full Text

[6] 6. Arafa M, Henkel R, Agarwal A, et al.: Correlation of Oxidation-Reduction Potential with Hormones, Semen Parameters and Testicular Volume. Andrologia. 2019; 51: e13258. PubMed Abstract | Publisher Full Text

[7] 7. Barbăroșie C, Agarwal A, Henkel R: Diagnostic Value of Advanced Semen Analysis in Evaluation of Male Infertility. Andrologia. 2021; 53: e13625. PubMed Abstract | Publisher Full Text

[8] 8. Casarini L, Crépieux P, Reiter E, et al.: FSH for the Treatment of Male Infertility. Int. J. Mol. Sci. 2020; 21: E2270. PubMed Abstract | Publisher Full Text

[9] 9. Krausz C, Riera-Escamilla A: Genetics of Male Infertility. Nat. Rev. Urol. 2018; 15: 369–384. Publisher Full Text

[10] 10. Lateef OM, Akintubosun MO: Sleep and Reproductive Health. J. Circadian Rhythms. 2020; 18: 1. Publisher Full Text

[11] 11. Salonia A, Bettocchi C, Carvalho J, et al.: Sexual and Reproductive Health. Uroweb. 2021.

[12] 12. Thurston L, Abbara A, Dhillo WS: Investigation and Management of Subfertility. J. Clin. Pathol. 2019; 72: 579–587. Publisher Full Text

[13] 13. Faduola P, Kolade CO: Sperm Chromatin Structure Assay Results in Nigerian Men with Unexplained Infertility. Clin. Exp. Reprod. Med. 2015; 42: 101–105. PubMed Abstract | Publisher Full Text

[14] 14. Mayorga-Torres BJM, Camargo M, Cadavid ÁP, et al.: Are Oxidative Stress Markers Associated with Unexplained Male Infertility?. Andrologia. 2017; 49: e12659. PubMed Abstract | Publisher Full Text

[15] 15. Dutta S, Henkel R, Agarwal A: Comparative Analysis of Tests Used to Assess Sperm Chromatin Integrity and DNA Fragmentation. Andrologia. 2021; 53: e13718. PubMed Abstract | Publisher Full Text

[16] 16. Evenson DP: Evaluation of Sperm Chromatin Structure and DNA Strand Breaks Is an Important Part of Clinical Male Fertility Assessment. Transl. Androl. Urol. 2017; 6: S495–S500. PubMed Abstract | Publisher Full Text

[17] 17. Rashki Ghaleno L, Alizadeh A, Drevet JR, et al.: Oxidation of Sperm DNA and Male Infertility. Antioxidants (Basel). 2021; 10: 97. Publisher Full Text

[18] 18. Martinez M, Majzoub A: Best Laboratory Practices and Therapeutic Interventions to Reduce Sperm DNA Damage. Andrologia. 2021; 53: e13736. PubMed Abstract | Publisher Full Text

[19] 19. Nguyen TT, Trieu TS, Tran TO, et al.: Evaluation of Sperm DNA Fragmentation Index, Zinc Concentration and Seminal Parameters from Infertile Men with Varicocele. Andrologia. 2019; 51: e13184. Publisher Full Text

[20] 20. Ribas-Maynou J, Yeste M, Becerra-Tomás N, et al.: Clinical Implications of Sperm DNA Damage in IVF and ICSI: Updated Systematic Review and Meta-Analysis. Biol. Rev. Camb. Philos. Soc. 2021; 96: 1284–1300. PubMed Abstract | Publisher Full Text

[21] 21. Agarwal A, Farkouh A, Parekh N, et al.: Sperm DNA Fragmentation: A Critical Assessment of Clinical Practice Guidelines. World J. Mens Health. 2022; 40: 30–37. PubMed Abstract | Publisher Full Text

[22] 22. Abdelbaki SA, Sabry JH, Al-Adl AM, et al.: The Impact of Coexisting Sperm DNA Fragmentation and Seminal Oxidative Stress on the Outcome of Varicocelectomy in Infertile Patients: A Prospective Controlled Study. Arab. J. Urol. 2017; 15: 131–139. PubMed Abstract | Publisher Full Text

[23] 23. Al Omrani B, Al Eisa N, Javed M, et al.: Associations of Sperm DNA Fragmentation with Lifestyle Factors and Semen Parameters of Saudi Men and Its Impact on ICSI Outcome. Reprod. Biol. Endocrinol. 2018; 16: 49. PubMed Abstract | Publisher Full Text

[24] 24. Cissen M, van Wely M , Scholten I, et al.: Measuring Sperm DNA Fragmentation and Clinical Outcomes of Medically Assisted Reproduction: A Systematic Review and Meta-Analysis. PLoS One. 2016; 11: e0165125. PubMed Abstract | Publisher Full Text

[25] 25. Cheng H, Han M, Ding J, et al.: Importance of a Semen Analysis Report for Determining the Relationship between SCSA Sperm DNA Fragmentation Index and Assisted Reproductive Technology Pregnancy Rate. Reprod. Biol. 2020; 20: 460–464. Publisher Full Text

[26] 26. Leach M, Aitken RJ, Sacks G: Sperm DNA Fragmentation Abnormalities in Men from Couples with a History of Recurrent Miscarriage. Aust. N. Z. J. Obstet. Gynaecol. 2015; 55: 379–383. Publisher Full Text

[27] 27. Oleszczuk K, Giwercman A, Bungum M: Sperm Chromatin Structure Assay in Prediction of in vitro Fertilization Outcome. Andrology. 2016; 4: 290–296. Publisher Full Text

[28] 28. Simon L, Emery BR, Carrell DT: Review: Diagnosis and Impact of Sperm DNA Alterations in Assisted Reproduction. Best Pract. Res. Clin. Obstet. Gynaecol. 2017; 44: 38–56. PubMed Abstract | Publisher Full Text

[29] 29. Vinnakota C, Cree L, Peek J, et al.: Incidence of High Sperm DNA Fragmentation in a Targeted Population of Subfertile Men. Syst. Biol. Reprod. Med. 2019; 65: 451–457. PubMed Abstract | Publisher Full Text

[30] 30. Yang H, Li G, Jin H, et al.: The Effect of Sperm DNA Fragmentation Index on Assisted Reproductive Technology Outcomes and Its Relationship with Semen Parameters and Lifestyle. Transl. Androl. Urol. 2019; 8: 356–365. PubMed Abstract | Publisher Full Text

[31] 31. Khalafalla K, Majzoub A, Elbardisi H, et al.: The Effect of Sperm DNA Fragmentation on Intracytoplasmic Sperm Injection Outcome. Andrologia. 2021; 53: e14180. PubMed Abstract | Publisher Full Text

[32] 32. World Health Organization: WHO Laboratory Manual for the Examination and Processing of Human Semen. 5th ed.Geneva, Switzerland:World Health Organization Press;2010.

[33] 33. Gill K, Kups M, Harasny P, et al.: The Negative Impact of Varicocele on Basic Semen Parameters, Sperm Nuclear DNA Dispersion and Oxidation-Reduction Potential in Semen. Int. J. Environ. Res. Public Health. 2021; 18: 5977. PubMed Abstract | Publisher Full Text

[34] 34. Gill K, Rosiak-Gill A, Jakubik J, et al.: The Higher Risk for Sperm DNA Damage in Infertile Men. Ginekol. Pol. 2019; 90: 684–691. PubMed Abstract | Publisher Full Text

[35] 35. Gill K, Jakubik J, Rosiak-Gill A, et al.: Utility and Predictive Value of Human Standard Semen Parameters and Sperm DNA Dispersion for Fertility Potential. Int. J. Environ. Res. Public Health. 2019; 16: E2004. PubMed Abstract | Publisher Full Text

[36] 36. Jakubik-Uljasz J, Gill K, Rosiak-Gill A, et al.: Relationship between Sperm Morphology and Sperm DNA Dispersion. Transl. Androl. Urol. 2020; 9: 405–415. PubMed Abstract | Publisher Full Text

[37] 37. Pedersen MR, Rafaelsen SR, Møller H, et al.: Testicular Microlithiasis and Testicular Cancer: Review of the Literature. Int. Urol. Nephrol. 2016; 48: 1079–1086. PubMed Abstract | Publisher Full Text

[38] 38. Elbashir S, Magdi Y, Rashed A, et al.: Relationship between Sperm Progressive Motility and DNA Integrity in Fertile and Infertile Men.2018. Publisher Full Text

[39] 39. Lotti F, Frizza F, Balercia G, et al.: The European Academy of Andrology (EAA) Ultrasound Study on Healthy, Fertile Men: Clinical, Seminal and Biochemical Characteristics. Andrology. 2020; 8: 1005–1020. PubMed Abstract | Publisher Full Text

[40] 40. Slama R, Eustache F, Ducot B, et al.: Time to Pregnancy and Semen Parameters: A Cross-Sectional Study among Fertile Couples from Four European Cities. Hum. Reprod. 2002; 17: 503–515. PubMed Abstract | Publisher Full Text

[41] 41. Cao X, Cui Y, Zhang X, et al.: The Correlation of Sperm Morphology with Unexplained Recurrent Spontaneous Abortion: A Systematic Review and Meta-Analysis. Oncotarget. 2017; 8: 55646–55656. PubMed Abstract | Publisher Full Text

[42] 42. Jayasena CN, Radia UK, Figueiredo M, et al.: Reduced Testicular Steroidogenesis and Increased Semen Oxidative Stress in Male Partners as Novel Markers of Recurrent Miscarriage. Clin. Chem. 2019; 65: 161–169. PubMed Abstract | Publisher Full Text

[43] 43. Li B, Ma Y, Huang J, et al.: Probing the Effect of Human Normal Sperm Morphology Rate on Cycle Outcomes and Assisted Reproductive Methods Selection. PLoS One. 2014; 9: e113392. PubMed Abstract | Publisher Full Text

[44] 44. McQueen DB, Zhang J, Robins JC: Sperm DNA Fragmentation and Recurrent Pregnancy Loss: A Systematic Review and Meta-Analysis. Fertil. Steril. 2019; 112: 54–60.e3. Publisher Full Text

[45] 45. Gatimel N, Moreau J, Parinaud J, et al.: Sperm Morphology: Assessment, Pathophysiology, Clinical Relevance, and State of the Art in 2017. Andrology. 2017; 5: 845–862. PubMed Abstract | Publisher Full Text

[46] 46. Shabtaie SA, Gerkowicz SA, Kohn TP, et al.: Role of Abnormal Sperm Morphology in Predicting Pregnancy Outcomes. Curr. Urol. Rep. 2016; 17: 67. PubMed Abstract | Publisher Full Text

[47] 47. Danis RB, Samplaski MK: Sperm Morphology: History, Challenges, and Impact on Natural and Assisted Fertility. Curr. Urol. Rep. 2019; 20: 43. Publisher Full Text

[48] 48. Kohn TP, Kohn JR, Lamb DJ: Role of Sperm Morphology in Deciding Between Various Assisted Reproduction Technologies. Eur. Urol. Focus. 2018; 4: 311–313. PubMed Abstract | Publisher Full Text

[49] 49. Kovac JR, Smith RP, Cajipe M, et al.: Men with a Complete Absence of Normal Sperm Morphology Exhibit High Rates of Success without Assisted Reproduction. Asian J. Androl. 2017; 19: 39–42. PubMed Abstract | Publisher Full Text

[50] 50. Björndahl L: The Usefulness and Significance of Assessing Rapidly Progressive Spermatozoa. Asian J. Androl. 2010; 12: 33–35. Publisher Full Text

[51] 51. Simon L, Lewis SEM: Sperm DNA Damage or Progressive Motility: Which One Is the Better Predictor of Fertilization in Vitro?. Syst. Biol. Reprod. Med. 2011; 57: 133–138. Publisher Full Text

[52] 52. Kuriya A, Agbo C, Dahan MH: Do Pregnancy Rates Differ with Intra-Uterine Insemination When Different Combinations of Semen Analysis Parameters Are Abnormal?. J. Turk. Ger. Gynecol. Assoc. 2018; 19: 57–64. PubMed Abstract | Publisher Full Text

[53] 53. Patel AS, Leong JY, Ramasamy R: Prediction of Male Infertility by the World Health Organization Laboratory Manual for Assessment of Semen Analysis: A Systematic Review. Arab. J. Urol. 2018; 16: 96–102. PubMed Abstract | Publisher Full Text

[54] 54. Zivlak-Radulovic N, Banjac V, Miskovic M: Hyperprolactinemia as a Side Effect of Long-Acting Injectable Risperidone Therapy or a Symptom of Klinefelter Syndrome - a Diagnostic and Therapeutic Dilemma. Psychiatr. Danub. 2018; 30: 99–103. PubMed Abstract | Publisher Full Text

[55] 55. Dabbous Z, Atkin SL: Hyperprolactinaemia in Male Infertility: Clinical Case Scenarios. Arab. J. Urol. 2018; 16: 44–52. PubMed Abstract | Publisher Full Text

[56] 56. La Vignera S, Vita R: Thyroid Dysfunction and Semen Quality. Int. J. Immunopathol. Pharmacol. 2018; 32: 2058738418775241. PubMed Abstract | Publisher Full Text

[57] 57. Condorelli R, Calogero AE, La Vignera S: Relationship between Testicular Volume and Conventional or Nonconventional Sperm Parameters. Int. J. Endocrinol. 2013; 2013: 145792–145796. PubMed Abstract | Publisher Full Text

[58] 58. Huang I-S, Huang WJ, Lin AT: Distinguishing Non-Obstructive Azoospermia from Obstructive Azoospermia in Taiwanese Patients by Hormone Profile and Testis Size. J. Chin. Med. Assoc. 2018; 81: 531–535. PubMed Abstract | Publisher Full Text

[59] 59. Nieschlag E, Vorona E: MECHANISMS IN ENDOCRINOLOGY: Medical Consequences of Doping with Anabolic Androgenic Steroids: Effects on Reproductive Functions. Eur. J. Endocrinol. 2015; 173: R47–R58. PubMed Abstract | Publisher Full Text

[60] 60. Sakamoto H, Yajima T, Nagata M, et al.: Relationship between Testicular Size by Ultrasonography and Testicular Function: Measurement of Testicular Length, Width, and Depth in Patients with Infertility. Int. J. Urol. 2008; 15: 529–533. PubMed Abstract | Publisher Full Text

[61] 61. Colpi GM, Francavilla S, Haidl G, et al.: European Academy of Andrology Guideline Management of Oligo-Astheno-Teratozoospermia. Andrology. 2018; 6: 513–524. PubMed Abstract | Publisher Full Text

[62] 62. Corinne TM, Anatole PC, Jeanne NY: Comparison of Serum Inhibin B and Follicle-Stimulating Hormone (FSH) Level between Normal and Infertile Men in Yaoundé. Int. J. Reprod. Med. 2020; 2020: 4765809–4765809. PubMed Abstract | Publisher Full Text

[63] 63. Uhler ML, Zinaman MJ, Brown CC, et al.: Relationship between Sperm Characteristics and Hormonal Parameters in Normal Couples. Fertil. Steril. 2003; 79 Suppl 3: 1535–1542. PubMed Abstract | Publisher Full Text

[64] 64. Mabeck LM, Jensen MS, Toft G, et al.: Fecundability According to Male Serum Inhibin B--a Prospective Study among First Pregnancy Planners. Hum. Reprod. 2005; 20: 2909–2915. PubMed Abstract | Publisher Full Text

[65] 65. Meeker JD, Godfrey-Bailey L, Hauser R: Relationships between Serum Hormone Levels and Semen Quality among Men from an Infertility Clinic. J. Androl. 2007; 28: 397–406. Publisher Full Text

[66] 66. Ring JD, Lwin AA, Köhler TS: Current Medical Management of Endocrine-Related Male Infertility. Asian J. Androl. 2016; 18: 357–363. PubMed Abstract | Publisher Full Text

[67] 67. Vilar L, Abucham J, Albuquerque JL, et al.: Controversial Issues in the Management of Hyperprolactinemia and Prolactinomas - An Overview by the Neuroendocrinology Department of the Brazilian Society of Endocrinology and Metabolism. Arch. Endocrinol. Metab. 2018; 62: 236–263. Publisher Full Text

[68] 68. Krajewska-Kulak E, Sengupta P: Thyroid Function in Male Infertility. Front. Endocrinol. 2013; 4. PubMed Abstract | Publisher Full Text

[69] 69. Krassas GE, Papadopoulou F, Tziomalos K, et al.: Hypothyroidism Has an Adverse Effect on Human Spermatogenesis: A Prospective, Controlled Study. Thyroid. 2008; 18: 1255–1259. PubMed Abstract | Publisher Full Text

[70] 70. Singh R, Hamada J, Agarwal A: Thyroid Hormones in Male Reproduction and Fertility. Open Reprod. Sci. J. 2011; 3: 98–104. Publisher Full Text

[71] 71. Fozooni R, Jafarzadeh Shirazi MR, Saedi S, et al.: Male Subfertility Effects of Sub-Chronic Ethanol Exposure during Stress in a Rat Model. Alcohol. 2021; 96: 63–71. PubMed Abstract | Publisher Full Text

[72] 72. Perumal P, Chang S, De AK, et al.: Slow Release Exogenous Melatonin Modulates Scrotal Circumference and Testicular Parameters, Libido, Endocrinological Profiles and Antioxidant and Oxidative Stress Profiles in Mithun. Theriogenology. 2020; 154: 1–10. Publisher Full Text

[73] 73. Simopoulou M, Philippou A, Maziotis E, et al.: Association between Male Infertility and Seminal Plasma Levels of Growth Hormone and Insulin-like Growth Factor-1. Andrologia. 2018; 50: e13048. PubMed Abstract | Publisher Full Text

[74] 74. Pedersen MR, Osther PJS, Rafaelsen SR: Ultrasound Evaluation of Testicular Volume in Patients with Testicular Microlithiasis. Ultrasound Int. Open. 2018; 04: E99–E103. Publisher Full Text

[75] 75. Bahk JY, Jung JH, Jin LM, et al.: Cut-off Value of Testes Volume in Young Adults and Correlation among Testes Volume, Body Mass Index, Hormonal Level, and Seminal Profiles. Urology. 2010; 75: 1318–1323. PubMed Abstract | Publisher Full Text

[76] 76. Lotti F, Tamburrino L, Marchiani S, et al.: DNA Fragmentation in Two Cytometric Sperm Populations: Relationship with Clinical and Ultrasound Characteristics of the Male Genital Tract. Asian J. Androl. 2017; 19: 272–279. PubMed Abstract | Publisher Full Text

[77] 77. Schurich M, Aigner F, Frauscher F, et al.: The Role of Ultrasound in Assessment of Male Fertility. Eur. J. Obstet. Gynecol. Reprod. Biol. 2009; 144(Suppl 1): S192–S198. Publisher Full Text

[78] 78. Zorn B, Golob B, Ihan A, et al.: Apoptotic Sperm Biomarkers and Their Correlation with Conventional Sperm Parameters and Male Fertility Potential. J. Assist. Reprod. Genet. 2012; 29: 357–364. PubMed Abstract | Publisher Full Text

[79] 79. Lu J-C, Jing J, Dai J-Y, et al.: Body Mass Index, Waist-to-Hip Ratio, Waist Circumference and Waist-to-Height Ratio Cannot Predict Male Semen Quality: A Report of 1231 Subfertile Chinese Men. Andrologia. 2015; 47: 1047–1054. Publisher Full Text

[80] 80. Wei T-C, Huang WJ, Lin ATL, et al.: The Role of Hormones on Semen Parameters in Patients with Idiopathic or Varicocele-Related Oligoasthenoteratozoospermia (OAT) Syndrome. J. Chin. Med. Assoc. 2013; 76: 624–628. PubMed Abstract | Publisher Full Text

[81] 81. Smit M, Romijn JC, Wildhagen MF, et al.: Sperm Chromatin Structure Is Associated with the Quality of Spermatogenesis in Infertile Patients. Fertil. Steril. 2010; 94: 1748–1752. PubMed Abstract | Publisher Full Text

[82] 82. Wdowiak A, Raczkiewicz D, Stasiak M, et al.: Levels of FSH, LH and Testosterone, and Sperm DNA Fragmentation. Neuro Endocrinol. Lett. 2014; 35: 73–79. PubMed Abstract

[83] 83. Lu J-C, Jing J, Chen L, et al.: Analysis of Human Sperm DNA Fragmentation Index (DFI) Related Factors: A Report of 1010 Subfertile Men in China. Reprod. Biol. Endocrinol. 2018; 16: 23. PubMed Abstract | Publisher Full Text

[84] 84. Amjad S, Baig M, Zahid N, et al.: Association between Leptin, Obesity, Hormonal Interplay and Male Infertility. Andrologia. 2019; 51: e13147. Publisher Full Text

[85] 85. Bundhun PK, Janoo G, Bhurtu A, et al.: Tobacco Smoking and Semen Quality in Infertile Males: A Systematic Review and Meta-Analysis. BMC Public Health. 2019; 19: 36. PubMed Abstract | Publisher Full Text

[86] 86. Ilacqua A, Izzo G, Emerenziani GP, et al.: Lifestyle and Fertility: The Influence of Stress and Quality of Life on Male Fertility. Reprod. Biol. Endocrinol. 2018; 16: 115. PubMed Abstract | Publisher Full Text

[87] 87. Kesari KK, Agarwal A, Henkel R: Radiations and Male Fertility. Reprod. Biol. Endocrinol. 2018; 16: 118. PubMed Abstract | Publisher Full Text

[88] 88. Miranda-Contreras L, Gómez-Pérez R, Rojas G, et al.: Occupational Exposure to Organophosphate and Carbamate Pesticides Affects Sperm Chromatin Integrity and Reproductive Hormone Levels among Venezuelan Farm Workers. J. Occup. Health. 2013; 55: 195–203. PubMed Abstract | Publisher Full Text

[89] 89. Palmer NO, Bakos HW, Fullston T, et al.: Impact of Obesity on Male Fertility, Sperm Function and Molecular Composition. Spermatogenesis. 2012; 2: 253–263. PubMed Abstract | Publisher Full Text

[90] 90. Semet M, Paci M, Saïas-Magnan J, et al.: The Impact of Drugs on Male Fertility: A Review. Andrology. 2017; 5: 640–663. Publisher Full Text

[91] 91. Erenpreiss J, Elzanaty S, Giwercman A: Sperm DNA Damage in Men from Infertile Couples. Asian J. Androl. 2008; 10: 786–790. Publisher Full Text

[92] 92. Belloc S, Benkhalifa M, Cohen-Bacrie M, et al.: Sperm Deoxyribonucleic Acid Damage in Normozoospermic Men Is Related to Age and Sperm Progressive Motility. Fertil. Steril. 2014; 101: 1588–1593. PubMed Abstract | Publisher Full Text

[93] 93. Giwercman A, Richthoff J, Hjøllund H, et al.: Correlation between Sperm Motility and Sperm Chromatin Structure Assay Parameters. Fertil. Steril. 2003; 80: 1404–1412. Publisher Full Text

[94] 94. Boushaba S, Belaaloui G: Sperm DNA Fragmentation and Standard Semen Parameters in Algerian Infertile Male Partners. World J. Mens Health. 2015; 33: 1–7. PubMed Abstract | Publisher Full Text

[95] 95. Bounartzi T, Dafopoulos K, Anifandis G, et al.: Pregnancy Prediction by Free Sperm DNA and Sperm DNA Fragmentation in Semen Specimens of IVF/ICSI-ET Patients. Hum. Fertil. (Camb.). 2016; 19: 56–62. PubMed Abstract | Publisher Full Text

[96] 96. Cohen-Bacrie P, Belloc S, Ménézo YJR, et al.: Correlation between DNA Damage and Sperm Parameters: A Prospective Study of 1,633 Patients. Fertil. Steril. 2009; 91: 1801–1805. PubMed Abstract | Publisher Full Text

[97] 97. Bungum M, Bungum L, Giwercman A: Sperm Chromatin Structure Assay (SCSA): A Tool in Diagnosis and Treatment of Infertility. Asian J. Androl. 2011; 13: 69–75. PubMed Abstract | Publisher Full Text

[98] 98. Bungum M: Sperm DNA Integrity Assessment: A New Tool in Diagnosis and Treatment of Fertility. Obstet. Gynecol. Int. 2012; 2012: 531042. Publisher Full Text

[99] 99. Evenson DP: The Sperm Chromatin Structure Assay (SCSA(^®)) and Other Sperm DNA Fragmentation Tests for Evaluation of Sperm Nuclear DNA Integrity as Related to Fertility. Anim. Reprod. Sci. 2016; 169: 56–75. PubMed Abstract | Publisher Full Text

[100] 100. Javed A, Talkad MS, Ramaiah MK: Evaluation of Sperm DNA Fragmentation Using Multiple Methods: A Comparison of Their Predictive Power for Male Infertility. Clin. Exp. Reprod. Med. 2019; 46: 14–21. PubMed Abstract | Publisher Full Text

[101] 101. Majzoub A, Arafa M, Mahdi M, et al.: Oxidation-Reduction Potential and Sperm DNA Fragmentation, and Their Associations with Sperm Morphological Anomalies amongst Fertile and Infertile Men. Arab. J. Urol. 2018; 16: 87–95. PubMed Abstract | Publisher Full Text

[102] 102. Wiweko B, Utami P: Predictive Value of Sperm Deoxyribonucleic Acid (DNA) Fragmentation Index in Male Infertility. Basic Clin. Androl. 2017; 27: 1. PubMed Abstract | Publisher Full Text

[103] 103. Giwercman A, Lindstedt L, Larsson M, et al.: Sperm Chromatin Structure Assay as an Independent Predictor of Fertility in Vivo: A Case-Control Study. Int. J. Androl. 2010; 33: e221–e227. PubMed Abstract | Publisher Full Text

[104] 104. Baird DC, Meyers GJ, Hu JS: Testicular Cancer: Diagnosis and Treatment. Am. Fam. Physician. 2018; 97: 261–268. PubMed Abstract

[105] 105. Wang F, Zhao S, Xie Y, et al.: Novo SOX10 Nonsense Mutation in a Patient with Kallmann Syndrome, Deafness, Iris Hypopigmentation, and Hyperthyroidism. Ann. Clin. Lab. Sci. 2018; 48: 248–252. PubMed Abstract

[106] 106. Santi D, Crépieux P, Reiter E, et al.: Follicle-Stimulating Hormone (FSH) Action on Spermatogenesis: A Focus on Physiological and Therapeutic Roles. J. Clin. Med. 2020; 9: E1014. PubMed Abstract | Publisher Full Text

[107] 107. Holota H, Thirouard L, Monrose M, et al.: FXRα Modulates Leydig Cell Endocrine Function in Mouse. Mol. Cell. Endocrinol. 2020; 518: 110995. PubMed Abstract | Publisher Full Text

[108] 108. Sertkaya Z, Tokuç E, Özkaya F, et al.: Acute Effect of Microdissection Testicular Sperm Extraction on Blood Total Testosterone and Luteinising Hormone Levels. Andrologia. 2020; 52: e13655. PubMed Abstract | Publisher Full Text

[109] 109. Kups M, Gill K, Rosiak-Gill A, et al.: Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study (Final) [Data set]. Zenodo. 2022; Publisher Full Text

[110] 110. Kups M, Gill K, Rosiak-Gill A, et al.: Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study. [Data set]. Zenodo. 2022. Publisher Full Text

[111] 111. Kups M, Gill K, Rosiak-Gill A, et al.: Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study. [Data set]. Zenodo. 2022. Publisher Full Text

Evaluation of selected semen parameters and biomarkers of male infertility – preliminary study

Abstract

Keywords

Introduction

Methods

Ethical considerations

Study population

Figure 1. The strategy developed in study analyses.

Conventional semen analysis

Sperm Chromatin Dispersion (SCD) test

Figure 2. Visualization of sperm chromatin dispersion test (SCD).

Hormone profile of infertile subjects

Table 1. Reproductive hormones evaluated in study population.

Ultrasonography of the scrotum

Statistical analysis

Results

Seminological characteristics of study population

Table 2. Descriptive statistics of study parameters in the infertile group of men (n=130).

Comparison of study parameters between infertile subjects with an abnormal volume of at least one testis (<12 mL) and subjects with a normal volume of testes (each ≥12 mL)

Table 3. Descriptive statistics and comparison of age, testicular volume and levels of reproductive hormones between infertile subjects with an abnormal volume of at least one testis (<12 mL) and subjects with a normal volume of testes (each ≥12 mL).

Table 4. Descriptive statistics and comparison of semen parameters between infertile subjects with an abnormal volume of at least one testis (<12 mL) and subjects with a normal volume of testes (each ≥12 mL).

Comparison of study parameters between infertile subjects with abnormal levels of at least one of the assessed hormones and subjects with normal hormonal profiles

Table 5. Descriptive statistics and comparison of age, testicular volume and levels of reproductive hormones between infertile subjects with abnormal hormone profiles (abnormal levels of at least one evaluated hormone) and subjects with normal hormone profiles.

Table 6. Descriptive statistics and comparison of semen parameters between infertile subjects with abnormal hormone profiles (abnormal levels of at least one evaluated hormone) and subjects with normal hormone profiles.

Comparison of study parameters between infertile subjects with SDF >30%, >15–30% and ≤15%

Table 7. Descriptive statistics and comparison of age, testicular volume and levels of reproductive hormones between infertile subjects with SDF >30%, >15–30% and ≤15%.

Table 8. Descriptive statistics and comparison of semen parameters between infertile subjects with SDF >30%, >15–30% and ≤15%.

Spearman’s rank correlations between the study parameters

Table 9. Spearman's rank correlation coefficient (rs) between age, testicular volume and levels of reproductive hormones.

Table 10. Spearman's rank correlation coefficient (rs) between testicular volume, levels of reproductive hormones and semen parameters.

Discussion

Reduced basic semen parameters can result from impaired spermatogenesis

Relationships between testicular volume, hormone levels, basic semen parameters and sperm genome integrity

Sperm genome integrity is a key point for male fertility

Conclusions

Limitations of the study

Data availability

Underlying data

Extended data

References

Comments on this article Comments (0)

Open Peer Review

Comments on this article Comments (0)

Open Peer Review

Reviewer Status

Reviewer Reports

Comments on this article

Browse by related subjects

Competing Interests Policy

Stay Updated

Table 9. Spearman's rank correlation coefficient (r_s) between age, testicular volume and levels of reproductive hormones.

Table 10. Spearman's rank correlation coefficient (r_s) between testicular volume, levels of reproductive hormones and semen parameters.