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1.
Mol Biol Rep ; 51(1): 727, 2024 Jun 11.
Article in English | MEDLINE | ID: mdl-38861014

ABSTRACT

BACKGROUND: The presence of granulocyte-macrophage colony-stimulating factor (GM-CSF) and its receptor in various testicular cells and spermatozoa suggests a potential role in enhancing spermatogonial and postmeiotic cell development. Moreover, GM-CSF activates the pivotal pathways implicated in sperm motility regulation and glucose metabolism. However, the impact of GM-CSF on testicular biopsies from patients with obstructive azoospermia (OA) remains unexplored. Therefore, this study aimed to investigate the in vitro effects of GM-CSF on the expression of genes related to glucose transporters and signaling pathways, sperm motility, and viability in testicular biopsies. METHODS AND RESULTS: Following testicular sperm extraction from 20 patients diagnosed with OA, each sample was divided into two parts: the experimental samples were incubated with medium containing 2 ng/ml GM-CSF at 37 °C for 60 min, and the control samples were incubated with medium without GM-CSF. Subsequently, the oocytes retrieved from the partner were injected with sperm from the treatment and control groups. The sperm parameters (motility and viability), the expression levels of sperm motility-related genes (PIK3R1, PIK3CA, and AKT1), and the expression levels of sperm energy metabolism-related genes (GLUT1, GLUT3, and GLUT14) were assessed. Furthermore, the fertilization and day 3 embryo development rate and embryo quality were evaluated. Compared with those in the nontreated group, the motility parameters and the mRNA expression levels of PIK3R1, AKT1, and GLUT3 in testicular sperm supplemented with GM-CSF were significantly greater (p < 0.05). However, no significant differences in the mRNA expression of PIK3CA, GLUT1, or GLUT14 were detected. According to the ICSI results, compared with the control group, the GM-CSF treatment group exhibited significantly greater fertilization rates (p = 0.027), Day 3 embryo development rate (p = 0.001), and proportions of good-quality embryos (p = 0.002). CONCLUSIONS: GM-CSF increased the expression of genes related to motility and the energy metabolism pathway and effectively promoted the motility of testis-extracted spermatozoa, consequently yielding positive clinical outcomes.


Subject(s)
Azoospermia , Energy Metabolism , Granulocyte-Macrophage Colony-Stimulating Factor , Sperm Injections, Intracytoplasmic , Sperm Motility , Spermatozoa , Humans , Male , Sperm Motility/drug effects , Sperm Motility/genetics , Azoospermia/genetics , Azoospermia/drug therapy , Granulocyte-Macrophage Colony-Stimulating Factor/genetics , Granulocyte-Macrophage Colony-Stimulating Factor/metabolism , Granulocyte-Macrophage Colony-Stimulating Factor/pharmacology , Sperm Injections, Intracytoplasmic/methods , Energy Metabolism/drug effects , Energy Metabolism/genetics , Spermatozoa/metabolism , Spermatozoa/drug effects , Adult , Testis/metabolism , Testis/drug effects , Glucose Transporter Type 3/genetics , Glucose Transporter Type 3/metabolism , Signal Transduction/drug effects , Signal Transduction/genetics , Female , Gene Expression Regulation/drug effects
2.
Rep Biochem Mol Biol ; 12(2): 294-305, 2023 Jul.
Article in English | MEDLINE | ID: mdl-38317811

ABSTRACT

Background: Seminal plasma exosomes are now recognized to play a complex role in the regulation of the female reproductive system infertility. The objective of this study was to assess the effect of exosomes derived from the sperm of men with oligoasthenoteratozoospermia on endometrial implantation-related genes. Methods: To isolate the exosomes, we employed an ultracentrifugation method on samples derived from 10 fertile men with normal sperm parameters and 10 men with oligoasthenoteratozoospermia. The size distribution and ultrastructure of the exosomes were then characterized using transmission electron microscopy and dynamic light scattering. We detected an exosome marker using western blot analysis and confirmed the cytoplasmic localization of the exosomes by incubating them with DiI dye and visualizing them using fluorescence microscopy. After 6 hours of in vitro treatment of endometrial epithelial cells with 100 µg/ml seminal exosome, the endometrial receptivity genes were examined using qRT-PCR. To perform data analysis and quantification, we utilized Image J and Prism software. P< 0.05 were considered statistically significant. Results: After 6 hours of treatment, the mRNA levels of MUC1, LIF, G-CSF, CX3CL1, and VEGF were significantly downregulated in the endometrial epithelial cells treated with oligoasthenoteratozoospermia exosomes compared to the normal group. Although changes were observed in the mean mRNA levels of IL8 and TGF-ß genes in the oligoasthenoteratozoospermia group compared to the normal group, these differences did not reach statistical significance (p > 0.05). Conclusions: Oligoasthenoteratozoospermia exosomes have a distinct effect on endometrial receptivity compared to normal exosomes, leading to reduced expression of implantation-related genes.

3.
Andrologia ; 54(7): 1618-1630, 2022 Aug.
Article in English | MEDLINE | ID: mdl-35545511

ABSTRACT

Poor sperm quality in oligoasthenoteratospermia patients negatively affects assisted reproductive technology outcomes. Therefore, the development of sperm media is necessary to improve sperm parameters. This study investigated the effect of GM-CSF via PI3K/AKT pathway on sperm quality in OAT patients. Semen samples were collected from 20 OAT patients, and each sample was divided into two groups: Experiment and Control. In the experimental group, the samples were incubated with medium containing GM-CSF, and control samples were incubated without GM-CSF. Sperm parameters, mitochondrial membrane potential, acrosome reaction and DFI were studied; in addition, gene expression of PI3KR1, PI3KCA, GLUT1, GLUT3 and AKT1 was analysed, evaluation of PAKT/TAKT, and expression of GLUT 1, 3 was examined; subsequent fertilization rate and embryo quality were assessed. Our data showed that GM-CSF supplementation could significantly increase motility, mitochondrial activity, gene expression of PI3KCA, AKT1, the protein level of PAKT/TAKT and expression of GLUT 1, 3 while it decreases DNA fragmentation. The fertilization rate and embryo quality significantly improved in the treatment group. LY294002 had adverse effects on sperm motility and the PAKT/TAKT ratio. GM-CSF can improve in vitro sperm quality and could be a suitable supplement to sperm media for OAT patients.


Subject(s)
Asthenozoospermia , Fertilization in Vitro , Granulocyte-Macrophage Colony-Stimulating Factor , Asthenozoospermia/drug therapy , Granulocyte-Macrophage Colony-Stimulating Factor/therapeutic use , Humans , Male , Phosphatidylinositol 3-Kinases , Proto-Oncogene Proteins c-akt , Semen , Sperm Motility , Spermatozoa
4.
Comput Biol Chem ; 94: 107561, 2021 Oct.
Article in English | MEDLINE | ID: mdl-34461466

ABSTRACT

OBJECTIVE: The aim of our study was to detect a biomarker for selection of competent oocytes with acceptable fertilization potential. Calcium ion fluctuation play the most critical role of modulating intercellular signaling pathways in oocyte maturation, egg activation and the egg-to-embryo transition. Since, the stimulatory action of calcium ion is mediated by binding to certain proteins, the calcium/calmodulin-binding genes (CBGs), as the main calcium binding group, was analyzed in detail. METHODS: In this work, bioinformatics analysis was conducted on the CBGs of human cumulus cells (CCs) to elucidate a reliable biomarker for fertile oocyte selection. Calcineurin (CaN) or protein phosphatase 3 (PPP3) was selected which consists of a catalytic subunit A with PPP3CA (Aα), PPP3CB (Aß), and PPP3CC (Aγ) isoforms and a regulatory subunit B. Whereas CaN A regulates calcium ion function, our study gives insights to probable role of related isoforms within human oogenesis process. The presence of CaN A in CCs surrounding growing and mature oocytes was confirmed by western blotting and the expression patterns of related isoforms were examined by reverse transcription-quantitative PCR (RT-qPCR). RESULTS: Our results indicated the increased expression of the catalytic subunit of CaN protein in the CCs of metaphase (M) II oocytes. The expression level of PPP3CB was significantly elevated in CCs of fertile MII compared with those in the germinal vesicle (GV), MI and unfertilized MII oocytes (P ≤ 0.05). CONCLUSION: Elevated level of PPP3CB isoform in the CCs of fertile MII oocyte could be a reliable indication of oocyte fertilization potential. However, further researches are required to introduce CaN Aß as an appropriate biomarker for oocyte selection in assisted reproduction technique (ART) programs.


Subject(s)
Calcineurin/analysis , Fertilization , Oocytes/metabolism , Sequence Analysis, Protein , Biomarkers/analysis , Biomarkers/metabolism , Calcineurin/metabolism , Humans , Oocytes/cytology
5.
Caspian J Intern Med ; 12(1): 70-76, 2021.
Article in English | MEDLINE | ID: mdl-33680401

ABSTRACT

BACKGROUND: Intrauterine insemination (IUI) is a widely utilized method for treating the infertile couples. The aim of the present study was to determine the pregnancy and abortion rates after IUI and to examine the relationship of sperm parameters with these rates. METHODS: This retrospective study was performed on 911 infertile couples undergoing IUI treatment in Shahid Akbarabadi IVF Centre from May 2017 to May 2019. To evaluate the correlation of sperm parameters with the clinical pregnancy and abortion rates, odds ratio (OR) with 95% confidence intervals (CI) was calculated. RESULTS: In this study, the pregnancy rate following IUI was 15.7% (143/911), and among women who achieved pregnancy, the abortion rate was 42.0% (60/143). According to the multiple logistic regression analysis, none of the sperm parameters was associated with the pregnancy rate. Couples with either male or female factor infertility etiologies were more likely to get pregnant than those with unexplained infertility. Regarding the abortion rate, multiple logistic regression analysis revealed that normal sperm count was related to a lower abortion rate (adjusted OR=0.25, 95% CI=0.07-0.91). CONCLUSION: The present study did not reveal a significant relationship between none of the sperm parameters and pregnancy rate after IUI treatment. However, among women who got pregnant, continuation of the pregnancy was associated with the normal sperm count. Furthermore, analysis of all semen parameters together in comparison to one parameter alone might be more accurate to predict pregnancy or abortion. Further prospective cohort studies with a large number of couples are required.

6.
Int J Reprod Biomed ; 18(10): 815-824, 2020 Oct.
Article in English | MEDLINE | ID: mdl-33134794

ABSTRACT

BACKGROUND: Many researchers consider implantation and endometrial receptivity as pertinent issues in reproductive science. Although, several experiments have been performed and their results evaluated, yet there is no confirmed evidence about the related factors and the role of sperm in endometrial receptivity. OBJECTIVE: To investigate the effect of the sperm-endometrium interaction in regulating genes involved in the endometrial receptivity pathway. MATERIALS AND METHODS: In this experimental study, 10 male and 30 female NMRI mice were included, and half of the male cases were vasectomized. The subjects were divided into two groups as follows; group 1 (case) comprised of 15 females mated with 5 non-vasectomized male mice, while group 2 (control) consisted of 15 females mated with 5 vasectomized males. Cases were sacrificed and assessed after 36 hr and the endometrial tissue was extracted and kept at -80°C until the next use. The expression of the endometrial receptivity pathway genes, including VEGF, HBEGF, FGF2, EGF, LIF, LIFR, HOXA10, MUC1, PGR, and CSF, was examined in both groups. For statistical analysis, an independent samples test (Mean ± SD) was used. RESULTS: The mRNA levels of LIF (p = 0.045), LIFR (p = 0.040), MUC1 (p = 0.032), VEGF (p = 0.022), EFG (p = 0.035), and FGF2 (p = 0.040) were significantly upregulated in the case group compared with the control group. CONCLUSION: Finally, seminal plasma was observed to be effective in expressing the involved genes in the successful implantation pathway, including LIF, LIFR, MUC1, VEGF, EGF, and FGF2.

7.
J Reprod Infertil ; 21(4): 259-268, 2020.
Article in English | MEDLINE | ID: mdl-33209742

ABSTRACT

BACKGROUND: It is demonstrated that optimal preincubation time improves oocyte quality, fertilization potential and developmental rate. This study aimed to evaluate the effect of preincubation time in the simple and myo-inositol supplemented medium on the oocyte quality regarding oxidative stress and mitochondrial alteration. METHODS: Cumulus oocyte complexes (COCs) retrieved from superovulated NMRI mice were divided in groups of 0, 4 and 8 hr preincubation time in the simple and 20 mmol/L myo-inositol supplemented media. Intracellular reactive oxygen species (H2O2), glutathione (GSH), mitochondrial membrane potential (MMP), ATP content, and mitochondrial amount were measured and analyzed in experimental groups. One-way ANOVA and Kruskal-Wallis were respectively used for parametric and nonparametric variables. Statistical significance was defined as p<0.05. RESULTS: In comparison to control group, variables including ROS, GSH, mitochondrial amount, fertilization and developmental rates were significantly changed after 4 hr of preincubation in the simple medium, while MMP decreased following 8 hr of preincubation in the simple medium (p˂0.001). Preincubation of oocytes up to 8 hr in the simple medium could not decrease ATP content. For both 4 and 8 hr preincubation times, myo-inositole could decrease H2O2 and increase GSH and MMP levels and consequently could improve fertilization rate compared to oocytes preincubated in the simple culture. CONCLUSION: It seems that 4 hr or more preincubation time can decrease the oocyte quality and lead to reduced oocyte fertilization and developmental potential. Howevere, myo-inositol may prevent oocyte quality reduction and improve fertilization potential in comparision to the equivalent simple groups.

8.
Turk J Biol ; 44(6): 371-380, 2020.
Article in English | MEDLINE | ID: mdl-33402864

ABSTRACT

Odorant or olfactory receptors are mainly localized in the olfactory epithelium for the perception of different odors. Interestingly, many ectopic olfactory receptors with low expression levels have recently been found in nonolfactory tissues to involve in local functions. Therefore, we investigated the probable role of the olfactory signaling pathway in the surrounding microenvironment of oocyte. This study included 22 women in intracytoplasmic sperm injection cycle. The expression of olfactory target molecules in cumulus cells surrounding the growing and mature oocytes was evaluated by Western blotting and real-time polymerase chain reaction. Additionally, integrated bioinformatics analyses were carried out and 6 ectopic olfactory receptors were selected for further evaluation. The initiation of olfactory transduction cascade in cumulus cells of competent oocytes was confirmed by analyzing the expression of adenylyl cyclase type 3 and olfactory market protein. Moreover, the expression pattern of the selected olfactory receptors was evaluated and OR10H2 was selected due to a high level of expression in mature fertile oocytes. We suggested that OR10H2 could be considered as a reliable biomarker for oocyte selection in assisted reproduction technique programs. However, further studies are required to elucidate the role of olfactory transduction cascade in embryo quality and implantation.

9.
Eur J Obstet Gynecol Reprod Biol ; 229: 127-131, 2018 Oct.
Article in English | MEDLINE | ID: mdl-30173088

ABSTRACT

The tight junction between epithelial cells helps making connections in the fallopian tube and contributes to successful fertilization. Breaking the tight junction complex induces various diseases such as the EP. Previous studies have shown that glucocorticoids are effective in repairing and maintaining intercellular tight junctions in epithelial cells of the fallopian tube, although their mechanism is still unknown. This research is a genomic study of hydrocortisone's effect on epithelial cells of the fallopian tube. Using the human fallopian tube, epithelial cell line (OE-E6/E7) was cultured in four concentrations of hydrocortisone (0 nM, 50 nM, 100 nM and 200 nM) for three durations (24 h, 48 h and 72 h). Glucocorticoids are effective on the expression of Zona occluding-1(ZO-1), Claudin 4, Claudin3, Desmoglein and E-cadherin genes involved in the tight junctions of the fallopian tube. The expression of all genes was up-regulated in the concentrations of 100 nM after 48 h treatment, as compared with the control (0 nM). However, their expression was down-regulated significantly after 72 h treatment (P < 0.05). The present study showed that treatment of epithelial cells of the fallopian tube with glucocorticoid increased the expression of genes involved in tight junctions, including claudin-3, claudin-4, E-cadherin, zona occludin-1 and Desmoglein-1. The obtained data suggests that a new mechanism is developed for glucocorticoid induction of tight junctions by increasing the expression of claudin-3, claudin-4, E-cadherin, zona occludin-1 and Desmoglein-1 genes.


Subject(s)
Anti-Inflammatory Agents/pharmacology , Fallopian Tubes/drug effects , Hydrocortisone/pharmacology , Tight Junction Proteins/metabolism , Tight Junctions/drug effects , Cell Line , Epithelial Cells/drug effects , Female , Humans , Polymerase Chain Reaction , Tight Junctions/genetics , Tight Junctions/metabolism
10.
Int J Reprod Biomed ; 15(7): 413-422, 2017 Jul.
Article in English | MEDLINE | ID: mdl-29177242

ABSTRACT

BACKGROUND: Prenatal drug exposure, as a common public health concern, is associated with an increased risk of adverse effects on early embryo development. OBJECTIVE: To investigate the in vitro development of - embryo from experimentally Kerack-addicted mice. MATERIALS AND METHODS: Twenty-five female mice were studied in five groups: control, vehicle, and three experimental groups of Kerack-dependent mice (I, II, and III) which received different doses of Kerack for 14 days. After the establishment of addiction model (7 days), experimental groups I, II, and III were given Kerack intraperitoneally at the doses of 5, 35, and 70 mg/kg, twice a day for a period of 7 days, respectively. The vehicle group received normal saline and lemon juice whilst the control group just received water and food. Morulae were obtained through oviduct flashing. The survived embryos were cultured in T6+ 5mg/ml bovine serum albumin. The developmental rates up to hatched stage daily and embryo quality (differential staining and Tunnel staining) were also assessed. RESULTS: The developmental potential of embryos obtained from the addicted mother was significantly decreased in comparison with control group. There was a significant reduction in the rate of blastocyst formation in the high dose Kerack dependent group. However, in addicted mice there was reduction in the total cell number (40.92% vs. 65.08% in control) and, inner cell mass percentage (17.17% vs. 26.15% in control) while apoptotic cells numbers were increased (7.17 vs. 1.46 in control) (p<0.05). CONCLUSION: The Kerack addiction during pregnancy retards preimplantation development and induces apoptosis.

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