RESUMO
Objectives: Cryopreservation has destructive effects on the function and structure of spermatozoa. It is known that leptin and prolactin play an active role in decreasing the rates of reactive oxygen species and DNA fragmentation, as well as enhancing sperm motility. Hence, this experiment aimed to investigate the effects of leptin and prolactin as pro-survival factors on the normozoospermic human semen samples during cryopreservation. Material and methods: Semen samples were collected from 15 healthy, fertile men ranging from 25 to 40 years. Cryopreservation of the samples was performed in liquid nitrogen over a period of two weeks, using five varying concentrations of leptin/prolactin, 0, 10, 100, 500, and 1000ng/ml respectively. Sperm motility, total caspase activity, and mitochondrial and cytosolic ROS were measured by flowcytometry, TUNEL, and other appropriate tests after thawing of the samples. Results: Both hormones were observed to have positive effects on the motility of the samples post-cryopreservation, the highest improvement being in the 100ng/ml concentration leptin and prolactin in comparison to the control group (P=0.01 and P=0.041, respectively). A significant reduction of mitochondrial ROS was also observed in 100 and 1000ng/ml of leptin (P=0.042), and there was a considerable decrease in the cytosolic ROS in the 100ng/ml of prolactin in comparison to the control group (P=0.048). Total caspase activity was also highly reduced in the 100, 500, and 1000ng/ml of leptin compared to the control group (P=0.039). Interestingly, both hormones also significantly decreased DNA fragmentation in 1000ng/ml compared to the control group (P=0.042). (AU)
Objetivos: La criopreservación tiene efectos destructivos sobre la función y estructura de los espermatozoides. Se sabe que la leptina y la prolactina desempeñan un papel activo en la disminución de las tasas de especies reactivas de oxígeno (ROS) y la fragmentación del ADN, así como en la mejora de la motilidad de los espermatozoides. Por lo tanto, este experimento tuvo como objetivo investigar los efectos de la leptina y la prolactina como factores de supervivencia en las muestras de semen humano normozoospérmico durante la criopreservación. Material y métodos: Se recolectaron muestras de semen de 15 hombres sanos y fértiles de entre 25 y 40 años. La crioconservación de las muestras se realizó en nitrógeno líquido durante un período de 2 semanas, utilizando 5 concentraciones variables de leptina/prolactina: 0, 10, 100, 500 y 1000ng/ml respectivamente. La motilidad de los espermatozoides, la actividad de caspasa total y las ROS mitocondriales y citosólicas se midieron mediante citometría de flujo, TUNEL y otras pruebas apropiadas después de descongelar las muestras. Resultados: Se observó que ambas hormonas tienen efectos positivos sobre la motilidad de las muestras después de la crioconservación, la mayor mejora se encuentra en la concentración de leptina y prolactina de 100ng/ml en comparación con el grupo de control (p=0,01 y p=0,041, respectivamente). También se observó una reducción significativa de las ROS mitocondriales en 100 y 1000ng/ml de leptina (p=0,042), y hubo una disminución considerable en las ROS citosólicas en los 100ng/ml de prolactina en comparación con el grupo de control (p=0,048). La actividad de la caspasa total también se redujo considerablemente en los 100, 500 y 1000ng/ml de leptina en comparación con el grupo de control (p=0,039). Curiosamente, ambas hormonas también redujeron significativamente la fragmentación del ADN en 1000ng/ml en comparación con el grupo de control (p=0,042). (AU)
Assuntos
Humanos , Masculino , Adulto , Sêmen , Prolactina , Caspases/farmacologia , Leptina/farmacologia , Espécies Reativas de Oxigênio , Criopreservação , Motilidade dos Espermatozoides , EspermatozoidesRESUMO
Introduction: Sperm motility is a crucial factor in male infertility and it depends on mitochondrial tail movements. Photobiomodulation light therapy allows the cells to produce their energy through activation of the mitochondria. The aim of the present study was to examine the impact of photobiomodulation on sperm motility in astenozoospermic individuals. Materials and methods: Following semen analyses of 20 astenozoospermic individuals, collected semen samples were centrifuged. Pellet was obtained and homogenized through mixing with culture media in 1:1 ratio. Each semen samples were divided into 3 groups. In the first group, control samples were not exposed to laser irradiation. The Group 2 and Group 3 were exposed to 650nm wavelength of photobiomodulation from 10cm distance in dark environment via a 36cm2 aperture sizer with 200mW output power for 30 and 60min duration, respectively. Sperm motilities were evaluated and chromatin condensation of sperms was determined. Results: Sperm motilities were significantly increased in photobiomodulation groups compared with the controls. Sperm motilities tended to be different between the 30 and 60min red light exposure groups; however, it was not statistically significant. When the motility grades were compared, no significant difference was observed in non-progressive motility sperms. While immotile sperms decreased significantly in the photobiomodulation groups compared to the control group, progressive sperms increased. (AU)
Introducción: La motilidad espermática es un factor crucial en la infertilidad masculina y depende de los movimientos de la cola mitocondrial. La fototerapia de fotobiomodulación permite que las células produzcan su energía a través de la activación de las mitocondrias. El objetivo del presente estudio fue examinar el impacto de la fotobiomodulación en la motilidad de los espermatozoides en individuos astenozoospérmicos. Materiales y métodos: Luego de los análisis de semen de 20 individuos astenozoospérmicos, se centrifugaron las muestras de semen recolectadas. Se obtuvo el sedimento y se homogeneizó mezclándolo con medios de cultivo en una proporción de 1:1. Cada muestra de semen se dividió en 3 grupos. En el primer grupo, las muestras de control no se expusieron a la irradiación láser. El grupo 2 y el grupo 3 fueron expuestos a una longitud de onda de 650nm de fotobiomodulación desde una distancia de 10cm en un ambiente oscuro a través de un medidor de apertura de 36cm2 con una potencia de salida de 200mW durante 30 y 60min de duración, respectivamente. Se evaluaron las motilidades de los espermatozoides y se determinó el tamaño de la cromatina de los espermatozoides. Resultados: La motilidad de los espermatozoides aumentó significativamente en los grupos de fotobiomodulación en comparación con los controles. La motilidad de los espermatozoides tendió a ser diferente entre los grupos de exposición a la luz roja de 30 y 60min; sin embargo, no fue estadísticamente significativo. Cuando se compararon los grados de motilidad, no se observaron diferencias significativas en los espermatozoides de motilidad no progresiva. Mientras que los espermatozoides inmóviles disminuyeron significativamente en los grupos de fotobiomodulación en comparación con el grupo de control, los espermatozoides progresivos aumentaron. (AU)
Assuntos
Humanos , Masculino , Adulto , Terapia com Luz de Baixa Intensidade/métodos , Sêmen , Análise do Sêmen , Espermatozoides/fisiologia , Motilidade dos EspermatozoidesRESUMO
Introduction: Human semen analysis must be performed after the liquefaction of the ejaculate. This takes place about 30min after ejaculation and samples must be maintained in the lab during this time. The temperatures for this incubation and the final analysis of motility are crucial but seldom taken into account. This study aims to examine the effect of these temperatures on various sperm parameters both manually (sperm count, motility, morphology, viability, chromatin condensation and maturation and DNA fragmentation) and CASA (kinematics and morphometrics, using an ISAS®v1 CASA-Mot and CASA-Morph systems, respectively) analyzed. Methods: Seminal samples from thirteen donors were incubated for 10min at 37°C followed by additional 20min at either room temperature (RT, 23°C) or 37°C and then examined following WHO 2010 criteria. Results: The data obtained show that there were no significant differences (P>0.05) in the subjective sperm quality parameters with incubation temperature. On the other hand, the head sperm morphometric parameters were significantly higher after room temperature incubation showing, in addition, lower ellipticity (P<0.05). Furthermore, kinematic parameters were evaluated both at RT and 37°C for the two incubation temperatures. In general, the four temperature combinations showed that kinematic parameters followed this order: RT-RT Conclusions: Our results showed that temperature control during both incubation and analysis is needed for accurate semen analysis, recommending the use of 37°C during the entire process. (AU)
Introducción: El análisis de semen humano debe realizarse después de la licuefacción del eyaculado. Esto ocurre aproximadamente a los 30minutos después de la eyaculación. Las temperaturas para esta incubación y el análisis final de la motilidad son cruciales, pero rara vez se tienen en cuenta. Este estudio tiene como objetivo examinar el efecto de estas temperaturas en varios parámetros de los espermatozoides tanto de forma manual (recuento de espermatozoides, motilidad, morfología, viabilidad, condensación y maduración de la cromatina y fragmentación del ADN) como CASA (cinemática y morfometría, utilizando un CASA-Mot ISAS®v1 y Sistemas CASA-Morph, respectivamente) analizados. Métodos: Las muestras seminales de 13 donantes se incubaron durante 10minutos a 37°C, seguidas de 20minutos adicionales a temperatura ambiente (TA, 23°C) o a 37°C y luego se examinaron siguiendo los criterios de la OMS 2010. Resultados: Los datos obtenidos muestran que no hubo diferencias significativas (p>0,05) en los parámetros subjetivos de calidad del esperma con la temperatura de incubación. Por otro lado, los parámetros morfométricos de la cabeza de los espermatozoides fueron significativamente más altos después de la incubación a temperatura ambiente, mostrando, además, una elipticidad más baja (p<0,05). Además, los parámetros cinemáticos se evaluaron tanto a temperatura ambiente como a 37°C para las dos temperaturas de incubación. En general, las cuatro combinaciones de temperatura mostraron que los parámetros cinemáticos siguieron este orden: RT-RT < RT-37 < 37-37 < 37-RT (temperaturas de incubación y análisis, respectivamente). Conclusiones: Nuestros resultados mostraron que el control de la temperatura durante la incubación y el análisis es necesario para un análisis de semen preciso, recomendando el uso de 37°C durante todo el proceso. (AU)
Assuntos
Humanos , Masculino , Adulto Jovem , Adulto , Pessoa de Meia-Idade , Sêmen , Motilidade dos Espermatozoides , Espermatozoides , Análise do Sêmen/métodos , Fenômenos BiomecânicosRESUMO
BACKGROUND: Semen cryopreservation is a critical tool for breed improvement and preservation of biodiversity. However, instability of sperm freezability affects its application. The Mediterranean buffalo is one of the river-type buffaloes with the capacity for high milk production. Until now, there is no specific cryopreservation system for Mediterranean buffalo, which influences the promotion of excellent cultivars. To improve the semen freezing extender used in cryopreservation of Mediterranean buffalo, different protein datasets relating to freezability sperm were analyzed by iTRAQ-based proteomics. This study will be beneficial for further understanding the sperm freezability mechanism and developing new cryopreservation strategy for buffalo semen. RESULTS: 2652 quantified proteins were identified, including 248 significantly differentially expressed proteins (DEP). Gene Ontology (GO) analysis indicated that many these were mitochondrial proteins, enriched in the molecular function of phospholipase A2 activity and enzyme binding, and biological processes of regulation of protein kinase A signaling and motile cilium assembly. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis identified 17 significant pathways, including oxidative phosphorylation (OXPHOS). Furthermore, 7 DEPs were verified using parallel reaction monitoring or western blot, which confirmed the accuracy of the iTRAQ data. Peroxiredoxin 6 (PRDX6), which expressed 1.72-fold higher in good freezability ejaculate (GFE) compared to poor freezability ejaculate (PFE) sperms, was selected to explore the function in sperm freezability by adding recombinant PRDX6 protein into the semen freezing extender. The results showed that the motility, mitochondrial function and in vitro fertilization capacity of frozen-thawed sperm were significantly increased, while the oxidation level was significantly decreased when 0.1 mg/L PRDX6 was added compared with blank control. CONCLUSIONS: Above results revealed the metabolic pattern of freezability of Mediterranean buffalo sperms was negatively associated with OXPHOS, and PRDX6 had protective effect on cryo-damage of frozen-thawed sperms.
Assuntos
Búfalos , Preservação do Sêmen , Animais , Masculino , Peroxirredoxina VI/genética , Peroxirredoxina VI/análise , Proteômica , Preservação do Sêmen/veterinária , Preservação do Sêmen/métodos , Espermatozoides/fisiologia , Criopreservação/veterinária , Criopreservação/métodos , Proteínas Recombinantes , Motilidade dos EspermatozoidesRESUMO
Toll-like receptor 2 (TLR2) signaling pathway is involved in the sperm-triggered uterine inflammatory response at insemination, but its precise mechanism at molecular-level remains unknown. According to the ligand specificity, TLR2 forms a heterodimer with TLR1 or TLR6 as an initial step to mediate intracellular signaling, leading to a specific type of immune response. Hence, the present study aimed to identify the active TLR2 heterodimer (TLR2/1 or TLR2/6) that is involved in sperm-uterine immune crosstalk in bovine using various models. First, in-vitro (bovine endometrial epithelial cells, BEECs) and ex-vivo (bovine uterine explant) models were employed to test different TLR2 dimerization pathways in endometrial epithelia after exposure to sperm or TLR2 agonists; PAM3 (TLR2/1 agonist), and PAM2 (TLR2/6 agonist). Additionally, in-silico approaches were performed to confirm the dimer stability using de novo protein structure prediction model for bovine TLRs. The in-vitro approach revealed that sperm triggered the mRNA and protein expression of TLR1 and TLR2 but not TLR6 in BEECs. Moreover, this model disclosed that activation of TLR2/6 heterodimer, triggers a much stronger inflammatory response than TLR2/1 and sperm in bovine uterine epithelia. In the ex-vivo model that mimics the intact uterine tissue at insemination, sperm also induced the protein expression of both TLR1 and TLR2, but not TLR6, in bovine endometrium, particularly in uterine glands. Importantly, PAM3 and sperm induced similar and low mRNA expression of pro-inflammatory cytokines and TNFA protein to a lesser extent than PAM2 in endometrial epithelia. This implied that sperm might trigger a weak inflammatory response via TLR2/TLR1 activation which is similar to that of PAM3. Additionally, the in-silico analyses showed that the existence of bridging ligands is essential for heterodimer stability in bovine TLR2 with either TLR1 or TLR6. Altogether, the present findings revealed that sperm utilize TLR2/1, but not TLR2/6, heterodimerization to trigger a weak physiological inflammatory response in the bovine uterus. This might be the way to remove excess dead sperm remaining in the uterine lumen without tissue damage for providing an ideal uterine environment for early embryo reception and implantation.
Assuntos
Receptor 1 Toll-Like , Receptor 2 Toll-Like , Feminino , Masculino , Animais , Bovinos , Receptor 2 Toll-Like/metabolismo , Receptor 1 Toll-Like/genética , Receptor 1 Toll-Like/metabolismo , Dimerização , Receptor 6 Toll-Like/metabolismo , Sêmen/metabolismo , Endométrio/metabolismo , Ligantes , Espermatozoides/metabolismo , RNA Mensageiro/metabolismoRESUMO
Proteasomes are highly sophisticated protease complexes that degrade non-lysosomal proteins, and their proper regulation ensures various biological functions such as spermatogenesis. The proteasome-associated proteins, PA200 and ECPAS, are predicted to function during spermatogenesis; however, male mice lacking each of these genes sustain fertility, raising the possibility that these proteins complement each other. To address this issue, we explored these possible roles during spermatogenesis by producing mice lacking these genes (double-knockout mice; dKO mice). Expression patterns and quantities were similar throughout spermatogenesis in the testes. In epididymal sperm, PA200 and ECPAS were expressed but were differentially localized to the midpiece and acrosome, respectively. Proteasome activity was considerably reduced in both the testes and epididymides of dKO male mice, resulting in infertility. Mass spectrometric analysis revealed LPIN1 as a target protein for PA200 and ECPAS, which was confirmed via immunoblotting and immunostaining. Furthermore, ultrastructural and microscopic analyses demonstrated that the dKO sperm displayed disorganization of the mitochondrial sheath. Our results indicate that PA200 and ECPAS work cooperatively during spermatogenesis and are essential for male fertility.
Assuntos
Complexo de Endopeptidases do Proteassoma , Sêmen , Masculino , Animais , Camundongos , Complexo de Endopeptidases do Proteassoma/metabolismo , Sêmen/metabolismo , Espermatogênese , Espermatozoides/metabolismo , Camundongos Knockout , Fosfatidato Fosfatase/metabolismo , Proteínas Nucleares/metabolismoRESUMO
The Caenorhabditis elegans spe-38 gene encodes a four-pass transmembrane molecule that is required in sperm for fertilization. In previous work, the localization of the SPE-38 protein was examined using polyclonal antibodies on spermatids and mature amoeboid spermatozoa. SPE-38 is localized to unfused membranous organelles (MOs) in nonmotile spermatids. Different fixation conditions revealed that SPE-38 either localized to fused MOs and the cell body plasma membrane or the pseudopod plasma membrane of mature sperm. To address this localization paradox in mature sperm, CRISPR/Cas9 genome editing was used to tag endogenous SPE-38 with fluorescent wrmScarlet-I. Homozygous male and hermaphrodite worms encoding SPE-38::wrmScarlet-I were fertile indicating the fluorescent tag does not interfere with SPE-38 function during sperm activation or fertilization. We found that SPE-38::wrmScarlet-I localized to MOs in spermatids consistent with previous antibody localization. In mature and motile spermatozoa we found SPE-38::wrmScarlet-I in fused MOs, the cell body plasma membrane, and the pseudopod plasma membrane. We conclude that the localization pattern observed with SPE-38::wrmScarlet-I represents the complete distribution of SPE-38 in mature spermatozoa and this localization pattern is consistent with a hypothesized role of SPE-38 directly in sperm-egg binding and/or fusion.
Assuntos
Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Animais , Masculino , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Sistemas CRISPR-Cas/genética , Proteínas de Membrana/metabolismo , Sêmen/metabolismo , Espermatozoides/metabolismoRESUMO
C. elegans spermiogenesis converts non-motile spermatids into motile, fertilization-competent spermatozoa. Two major events include the building of a pseudopod required for motility and fusion of membranous organelles (MOs)-intracellular secretory vesicles-with the spermatid plasma membrane required for the proper distribution of sperm molecules in mature spermatozoa. The mouse sperm acrosome reaction-a sperm activation event occurring during capacitation-is similar to MO fusion in terms of cytological features and biological significance. Moreover, C. elegans fer-1 and mouse Fer1l5, both encoding members of the ferlin family, are indispensable for MO fusion and acrosome reaction, respectively. Genetics-based studies have identified many C. elegans genes involved in spermiogenesis pathways; however, it is unclear whether mouse orthologs of these genes are involved in the acrosome reaction. One significant advantage of using C. elegans for studying sperm activation is the availability of in vitro spermiogenesis, which enables combining pharmacology and genetics for the assay. If certain drugs can activate both C. elegans and mouse spermatozoa, these drugs would be useful probes to explore the mechanism underlying sperm activation in these two species. By analyzing C. elegans mutants whose spermatids are insensitive to the drugs, genes functionally relevant to the drugs' effects can be identified.
Assuntos
Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Masculino , Animais , Camundongos , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Mutação , Sêmen/metabolismo , Espermatogênese/genética , Espermatozoides/metabolismoRESUMO
Quantitative and qualitative spermatogenic impairments are major causes of men's infertility. Although in vitro fertilization (IVF) is effective, some couples persistently fail to conceive. To identify causal variants in patients with severe male infertility factor and repeated IVF failures, we sequenced the exome of two consanguineous family members who underwent several failed IVF cycles and were diagnosed with low sperm count and motility. We identified a rare homozygous nonsense mutation in a previously uncharacterized gene, RNF212B, as the causative variant. Recurrence was identified in another unrelated, infertile patient who also faced repeated failed IVF treatments. scRNA-seq demonstrated meiosis-specific expression of RNF212B. Sequence analysis located a protein domain known to be associated with aneuploidy, which can explain multiple IVF failures. Accordingly, FISH analysis revealed a high aneuploidy rate in the patients' sperm cells and their IVF embryos. Finally, inactivation of the Drosophila orthologs significantly reduced male fertility. Given that members of the evolutionary conserved RNF212 gene family are involved in meiotic recombination and crossover maturation, our findings indicate a critical role of RNF212B in meiosis, genome stability, and in human fertility. Since recombination is completely absent in Drosophila males, our findings may indicate an additional unrelated role for the RNF212-like paralogs in spermatogenesis.
Assuntos
Infertilidade Masculina , Ligases , Sêmen , Humanos , Masculino , Aneuploidia , Fertilização In Vitro , Infertilidade Masculina/genética , Ligases/genética , Espermatozoides , Domínios RING FingerRESUMO
Studies of socially mediated phenotypic plasticity have demonstrated adaptive male responses to the 'competitive' environment. Despite this, whether variation in the paternal social environment also influences offspring reproductive potential in an intergenerational context has not yet been examined. Here, we studied the descendants of wild-caught house mice, a destructive pest species worldwide, to address this knowledge gap. We analysed traits that define a 'competitive' phenotype in the sons of males (sires) that had been exposed to either a high-male density (competitive) or high-female density (non-competitive) environment. We report disparate reproductive strategies among the sires: high-male density led to a phenotype geared for competition, while high-female density led to a phenotype that would facilitate elevated mating frequency. Moreover, we found that the competitive responses of sires persisted in the subsequent generation, with the sons of males reared under competition having elevated sperm quality. As all sons were reared under common-garden conditions, variation in their reproductive phenotypes could only have arisen via nongenetic inheritance. We discuss our results in relation to the adaptive advantage of preparing sons for sperm competition and suggest that intergenerational plasticity is a previously unconsidered aspect in invasive mammal fertility control.
Assuntos
Sêmen , Espermatozoides , Animais , Camundongos , Masculino , Feminino , Espermatozoides/fisiologia , Reprodução , Adaptação Fisiológica , Mamíferos , Comportamento Sexual AnimalRESUMO
While exogenous metal/metalloid (metal) exposure has been associated with reduced human semen quality, no study has assessed the associations of exogenous metals in human spermatozoa with semen quality. Here, we developed a strategy to explore the associations between exogenous metals in spermatozoa at single-cell resolution and human semen quality among 84 men screened as sperm donors, who provided 266 semen samples within 90 days. A cellular atlas of exogenous metals at the single-cell level was created with mass cytometry (CyTOF) technology, which concurrently displayed 18 metals in more than 50â¯000 single sperm. Exogenous metals in spermatozoa at single-cell resolution were extremely heterogeneous and diverse. Further analysis using multivariable linear regression and linear mixed-effects models revealed that the heterogeneity and prevalence of the exogenous metals at single-cell resolution were associated with semen quality. The heterogeneity of lead (Pb), tin (Sn), yttrium (Y), and zirconium (Zr) was negatively associated with sperm concentration and count, while their prevalence showed positive associations. These findings revealed that the heterogeneous properties of exogenous metals in spermatozoa were associated with human semen quality, highlighting the importance of assessing exogenous metals in spermatozoa at single-cell resolution to evaluate male reproductive health risk precisely.
Assuntos
Análise do Sêmen , Sêmen , Humanos , Masculino , Espermatozoides , Contagem de Espermatozoides , Metais , Motilidade dos EspermatozoidesRESUMO
Background: Autosomal dominant polycystic kidney disease (ADPKD) is a hereditary kidney disorder mostly caused by mutations in PKD1 or PKD2 genes. Here, we report thirteen ADPKD males with infertility and investigated the sperm morphological defects associated with PC1 disruption. Methods: Targeted next-generation sequencing was performed to detect PKD1 variants in patients. Sperm morphology was observed by immunostaining and transmission electron microscopy, and the sperm motility was assessed using the computer-assisted sperm analysis system. The Hippo signaling pathway was analyzed with by quantitative reverse transcription polymerase chain reaction (qPCR) and western blotting in vitro. Results: The ADPKD patients were infertile and their sperm tails showed morphological abnormalities, including coiled flagella, absent central microtubules, and irregular peripheral doublets. In addition, the length of sperm flagella was shorter in patients than in controls of in in. In vitro, ciliogenesis was impaired in Pkd1-depleted mouse kidney tubule cells. The absence of PC1 resulted in a reduction of MST1 and LATS1, leading to nuclear accumulation of YAP/TAZ and consequently increased transcription of Aurka. which might promote HDAC6-mediated ciliary disassembly. Conclusion: Our results suggest the dysregulated Hippo signaling significantly contributes to ciliary abnormalities in and may be associated with flagellar defects in spermatozoa from ADPKD patients.
Assuntos
Via de Sinalização Hippo , Rim Policístico Autossômico Dominante , Canais de Cátion TRPP , Animais , Humanos , Masculino , Camundongos , Rim Policístico Autossômico Dominante/complicações , Rim Policístico Autossômico Dominante/genética , Sêmen , Motilidade dos Espermatozoides , Espermatozoides/patologia , Canais de Cátion TRPP/genéticaRESUMO
BACKGROUND: The emergence and the spread of coronavirus disease (COVID-19) induced by the SARS-CoV-2 virus has multiple consequences in all countries around the world. Male germ cells of infertile patients which are shown to be vulnerable to many environmental conditions, could be particularly vulnerable to such an exceptional pandemic situation. We aimed through the current study to investigate the potential variations in sperm quality of infertile patients during the COVID-19 pandemic in Tunisia. METHODS: This was a cohort study including 90 infertile patients addressed to Laboratory of Cytogenetics and Reproductive Biology of Monastir Department of Maternity and Neonatology in Monastir, during the two first COVID-19 waves in Tunisia and who already have a spermogram before the pandemic period. RESULTS: We have pointed out a significant decrease in both total and progressive sperm motility during COVID-19 pandemic (p<0.0001 and p = 0.001 respectively). The percentage of morphologically abnormal spermatozoa increased from 90.99±7.38 to 93.67±4.55% during the pandemic (p< 0.001). The remaining sperm parameters were similar between the two compared timepoints. Interestingly, the univariate analysis didn't show any other associated factor to the observed impairment in sperm mobility and morphology. CONCLUSION: These data highlight the severe impact of the pandemic of the male reproductive health of hypofertile patients. Delaying infertility investigations and management after pandemic waves is recommended to hope a better gamete quality and hence to improve conception potential.
Assuntos
COVID-19 , Infertilidade Masculina , Humanos , Masculino , Feminino , Gravidez , Análise do Sêmen , Contagem de Espermatozoides , Infertilidade Masculina/epidemiologia , Pandemias , Motilidade dos Espermatozoides , Sêmen , Estudos de Coortes , COVID-19/epidemiologia , SARS-CoV-2 , EspermatozoidesRESUMO
INTRODUCTION: Spermatozoa are highly specialized cells with unique morphology. In addition, spermatozoa lose a considerable amount of cytoplasm during spermiogenesis, when they also compact their DNA, resulting in a transcriptionally quiescent cell. Throughout the male reproductive tract, sperm will acquire proteins that enable them to interact with the female reproductive tract. After ejaculation, proteins undergo post-translational modifications for sperm to capacitate, hyperactivate, and fertilize the oocyte. Many proteins have been identified as predictors of male infertility and also investigated in diseases that compromise reproductive potential. AREAS COVERED: In this review, we proposed to summarize the recent findings about the sperm proteome and how they affect sperm structure, function, and fertility. A literature search was performed using PubMed and Google Scholar databases within the past 5 years until August 2022. EXPERT OPINION: Sperm function depends on protein abundance, conformation, and PTMs; understanding the sperm proteome may help to identify pathways essential to fertility, even making it possible to unravel the mechanisms involved in idiopathic infertility. In addition, proteomics evaluation offers knowledge regarding alterations that compromise the male reproductive potential.
Assuntos
Infertilidade Masculina , Proteoma , Humanos , Masculino , Feminino , Proteoma/metabolismo , Sêmen/metabolismo , Espermatozoides/metabolismo , Fertilidade/genética , Infertilidade Masculina/genéticaRESUMO
A manual assessment of sperm motility requires microscopy observation, which is challenging due to the fast-moving spermatozoa in the field of view. To obtain correct results, manual evaluation requires extensive training. Therefore, computer-aided sperm analysis (CASA) has become increasingly used in clinics. Despite this, more data is needed to train supervised machine learning approaches in order to improve accuracy and reliability in the assessment of sperm motility and kinematics. In this regard, we provide a dataset called VISEM-Tracking with 20 video recordings of 30 seconds (comprising 29,196 frames) of wet semen preparations with manually annotated bounding-box coordinates and a set of sperm characteristics analyzed by experts in the domain. In addition to the annotated data, we provide unlabeled video clips for easy-to-use access and analysis of the data via methods such as self- or unsupervised learning. As part of this paper, we present baseline sperm detection performances using the YOLOv5 deep learning (DL) model trained on the VISEM-Tracking dataset. As a result, we show that the dataset can be used to train complex DL models to analyze spermatozoa.
Assuntos
Sêmen , Motilidade dos Espermatozoides , Espermatozoides , Humanos , Masculino , Reprodutibilidade dos Testes , Gravação em VídeoRESUMO
Sperm motility is a prerequisite for achieving pregnancy, and alterations in sperm motility, along with sperm count and morphology, are commonly observed in subfertile men. The aim of the study was to determine whether the expression level of genes annotated with the Gene Ontology (GO) term 'sperm motility' differed in sperm collected from healthy men and men diagnosed with oligoasthenozoospermia. Reverse transcription quantitative real-time PCR (RT-qPCR), quantitative mass spectrometry (LC-MS/MS), and enrichment analyses were used to validate a set of 132 genes in 198 men present at an infertility clinic. Out of the 132 studied sperm-motility-associated genes, 114 showed differentially expressed levels in oligoasthenozoospermic men compared to those of normozoospermic controls using an RT-qPCR analysis. Of these, 94 genes showed a significantly lower expression level, and 20 genes showed a significantly higher expression level. An MS analysis of sperm from an independent cohort of healthy and subfertile men identified 692 differentially expressed proteins, of which 512 were significantly lower and 180 were significantly higher in oligoasthenozoospermic men compared to those of the normozoospermic controls. Of the 58 gene products quantified with both techniques, 48 (82.75%) showed concordant regulation. Besides the sperm-motility-associated proteins, the unbiased proteomics approach uncovered several novel proteins whose expression levels were specifically altered in abnormal sperm samples. Among these deregulated proteins, there was a clear overrepresentation of annotation terms related to sperm integrity, the cytoskeleton, and energy-related metabolism, as well as human phenotypes related to spermatogenesis and sperm-related abnormalities. These findings suggest that many of these proteins may serve as diagnostic markers of male infertility. Our study reveals an extended number of sperm-motility-associated genes with altered expression levels in the sperm of men with oligoasthenozoospermia. These genes and/or proteins can be used in the future for better assessments of male factor infertility.
Assuntos
Infertilidade Masculina , Sêmen , Gravidez , Feminino , Humanos , Masculino , Sêmen/metabolismo , Cromatografia Líquida , Espectrometria de Massas em Tandem , Espermatozoides/metabolismo , Infertilidade Masculina/genética , Infertilidade Masculina/metabolismo , Fertilidade/genéticaRESUMO
In this study, the semen parameters, sperm chromatin integrity, antioxidant enzyme levels, and reproductive hormone levels of subfertile male subjects from Pakistan were assessed in relation to their age. Data on the demographic characteristics of the 750 study participants, including their general health, body mass index (BMI), and reproductive status, were collected from subfertile men from Pakistan. Semen and blood were collected to determine standard semen parameters, sperm chromatin dispersion (Halosperm-SCD), sperm chromatin integrity using toluidine blue (TB) staining, sperm chromatin maturity using chromomycin A3 (CMA3+) staining, and reproductive hormone (FSH, LH, prolactin and testosterone levels). The patients were divided into three groups according to their age: Group 1 included male subjects aged 30 years or less (n = 90), Group 2 included male subjects between the ages of 31 and 40 years (n = 330), and Group 3 included male subjects over 40 years of age (n = 330). Conventional semen parameters, reactive oxygen species (ROS), superoxide dismutase (SOD), guaiacol peroxidase (GPX), catalase (CAT), and lipid peroxidation (MDA) did not statistically (p > 0.05) differ with increasing male age or between different age groups. When compared to younger men (<30 years), sperm SCD (23.2 ± 0.88%) was significantly (p = 0.01) lower as compared to male patients aged >40 years (26.6 ± 0.6%). The concentration of LH, FSH, and testosterone levels were comparable between the groups (p > 0.05), while a significant (p = 0.04) increase in sperm chromatin immaturity CMA3+ (30 ± 0.71%) was observed in the old age group (>40 years) compared to the <30-year group (26.6 ± 1.03%). A positive association was observed between advanced male age and sperm chromatin dispersion (SCD) (r = 0.124, p = 0.001) and decondensation (CMA3+) (r = 0.1, p = 0.009). Despite potential limitations, this study has been carried out with extensive information on the potential risk of male age on sperm integrity. The present study demonstrated the impact of male age on male reproductive health, as these patients had a higher percentage of sperm chromatin damage (SCD) in their semen. Sperm DNA damage assessment will help in the evaluation and diagnosis of the underlying cause of poor fertility and can help clinicians in selecting the right treatment options. Male age is one of the factors that have an impact on the decline in male fertility. As a result, it is preferable for patients receiving assisted reproductive technology to be younger.
Assuntos
Infertilidade Masculina , Sêmen , Humanos , Masculino , Cromatina , Infertilidade Masculina/diagnóstico , Contagem de Espermatozoides , Motilidade dos Espermatozoides , Espermatozoides , Prolactina/genética , Hormônio Foliculoestimulante , Testosterona , BiomarcadoresRESUMO
Role of male factor in recurrent abortion and in vitro fertilization failure has not been fully defined yet and there is much controversy about evaluating male patients with normal semen analysis. One of the factors that might help establish the male role is DNA fragmentation index. However, strong correlation between this factor and quality of semen, has caused many clinicians to believe that it does not help in abortion and implantation failure. We aim to assess this factor in our patients. In a prospective observational study, we assessed age, duration of infertility, undesired fertility related events (assisted reproductive techniques attempts and abortions), semen parameters and DNA fragmentation index in patients with multiple abortions or in vitro fertilization failures and analysed the results by statistical software SPSS version 24. DNA fragmentation index was remarkably correlated with age, duration of infertility and semen parameters. Among all groups in our study, patients with abnormal semen analysis had statistically significant higher level of DNA fragmentation. Ten percent of patients with normal or slightly abnormal semen analysis had abnormally high SDFI (sperm DNA fragmentation index). Checking DNA fragmentation index is recommended in all couples with fertilization problems even in the presence of normal semen analysis. It might be more reasonable to assess it in aged men, long duration of infertility or candidates with remarkable semen abnormality.
Assuntos
Aborto Habitual , Infertilidade Masculina , Gravidez , Feminino , Masculino , Humanos , Idoso , Sêmen , Infertilidade Masculina/diagnóstico , Infertilidade Masculina/genética , Fragmentação do DNA , Fertilização In Vitro/métodos , Espermatozoides , Análise do SêmenRESUMO
Despite there have been many experiments conducted about antioxidants, the best sole or combination use of antioxidants to include as a standard ingredient to freezing extenders is yet to be found. This study was designed to investigate the different doses of methionine (2.5 and 5 mM), cysteine (1 and 2 mM), and butylated hydroxytoluene (BHT) (1 and 2 mM) for ram semen cryopreservation on post-thaw and post-incubation (6 h) time points over spermatological parameters. Semen samples were collected from Kivircik rams via electro-ejaculator in breeding season. After essential spermatological evaluations, appropriate samples were pooled then split into 7 equal aliquots to create study groups (antioxidant free control, 2.5 mM methionine, 5 mM methionine, 1 mM cysteine, 2 mM cysteine, 1 mM BHT, and 2 mM BHT). Semen samples were put into French straws (0.25 mL), and freezing procedure (two-step) was conducted via a programmable gamete freezer. At both time points, motility, HOST, PSA-FITC, and TUNEL assays were made to discover the impacts of cryopreservation and incubation process over sperm cells. Antioxidant supplemented groups yielded better results compared to the control groups in terms of various spermatological parameters not only at post-thaw time point but after incubation for 6 h of time. The study demonstrated that supplementing sperm freezing extenders with previous antioxidants may create new approaches to cryopreservation procedures, and through increasing success rate of freezing, fertility results may increase to better results in near future.
Assuntos
Preservação do Sêmen , Sêmen , Masculino , Ovinos , Animais , Hidroxitolueno Butilado/farmacologia , Cisteína , Metionina , Motilidade dos Espermatozoides , Preservação do Sêmen/veterinária , Preservação do Sêmen/métodos , Espermatozoides , Antioxidantes , Criopreservação/veterinária , Racemetionina , Carneiro Doméstico , Crioprotetores/farmacologiaRESUMO
Studies on the gene regulation of spermatogenesis are of unusual significance for maintaining male reproduction and treating male infertility. Here, we have demonstrated, for the first time, that a loss of ZBTB40 function leads to abnormalities in the morphological and phenotypic characteristics of mouse spermatocytes and spermatids as well as male infertility. We revealed that Zbtb40 was expressed in spermatocytes of mouse testes, and it was co-localized with γH2AX in mouse secondary spermatocytes. Interestingly, spermatocytes of Zbtb40 knockout mice had longer telomeres, compromised double-strand break (DSB) repair in the sex chromosome, and a higher apoptosis ratio compared to wild-type (WT) mice. The testis weight, testicular volume, and cauda epididymis body weight of the Zbtb40+/- male mice were significantly lower than in WT mice. Mating tests indicated that Zbtb40+/- male mice were able to mate normally, but they failed to produce any pups. Notably, sperm of Zbtb40+/- mice showed flagellum deformities and abnormal acrosome biogenesis. Furthermore, a ZBTB40 mutation was associated with non-obstructive azoospermia. Our results implicate that ZBTB40 deficiency leads to morphological and phenotypic abnormalities of spermatocytes and spermatids and causes male infertility. This study thus offers a new genetic mechanism regulating mammalian spermatogenesis and provides a novel target for gene therapy in male infertility.
