Metabolomic profiling of embryo culture media in patients with repeated implantation failure during assisted reproductive technology cycles.

Objective: This study investigated the metabolic status of the spent culture media from embryos of patients with repeated implantation failure (RIF) undergoing in vitro fertilization-intracytoplasmic sperm injection cycles in comparison with the embryos from healthy fertile women. Methods: Metabolite levels in spent culture media were assessed and compared between embryos from RIF patients (n=35) and oocyte donors as controls (n=15). Protein levels of insulin-like growth factor 1 (IGF-1) were determined using Western blotting. Concentrations of glucose, pyruvate, and lactate were measured using spectrophotometry. Ionic colorimetric assay kits were utilized to analyze the concentrations of sodium, chloride, calcium, and magnesium ions. High-performance liquid chromatography was employed to measure the concentrations of glutamic acid, aspartic acid, methionine, phenylalanine, and histidine. Results: Glucose consumption and lactate secretion were higher in the control group than in the RIF group. The magnesium concentration was significantly higher in the control group than in the RIF group, but glutamic acid and aspartic acid concentrations were lower in the control group than in the RIF patients (p<0.05). The levels of IGF-1, sodium, calcium, chloride, methionine, histidine, and phenylalanine did not show statistically significant differences between the two groups. Conclusion: The metabolic profile of the culture medium of the embryos in the RIF group differed from that of the control group. These findings suggest potential factors that may affect implantation capacity in RIF patients and provide a new perspective on embryo selection.

CGRP neuropeptide levels in patients with endometriosis-related pain treated with dienogest: a comparative study.

BACKGROUND AND OBJECTIVE: Endometriosis (EM) involves the peripheral nervous system and causes chronic pain. Sensory nerves innervating endometriotic lesions contribute to chronic pain and influence the growth phenotype by releasing neurotrophic factors and interacting with nearby immune cells. Calcitonin gene-related peptide (CGRP), a pain-signaling neurotransmitter, has a significant role. This study examines the effect of Dienogest (DNG), a hormone therapy used for managing EM -related pain, on serum CGRP levels in EM patients. MATERIALS AND METHODS: The Visual Analog Scale (VAS) assessed pain in diagnosed EM. INDIVIDUALS: Serum samples were obtained to measure CGRP concentration. Participants received a 2 mg/day oral dose of DNG for six months as prescribed treatment. Additional serum samples were collected after this period to measure CGRP levels. RESULTS: In the EM group, 6.7%, 33.3%, and 20% had ovarian EM, ovarian plus uterosacral, and ovarian plus bladder, respectively. The EM group showed higher CGRP serum levels than the control group (80.53 ± 16.13 vs. 58.55 ± 6.93, P < 0.0001). Still, after drug administration, CGRP serum levels significantly decreased compared to pre-treatment levels (69.66 ± 11.53 vs. 80.53 ± 16.13, P < 0.05). The EM group showed higher pain compared to the control group (7.93 ± 1.58 vs. 0.13 ± 0.35, P < 0.0001), but after drug administration, pain significantly decreased compared to pre-treatment levels (1.00 ± 2.00 vs. 7.93 ± 1.58, P < 0.05). CONCLUSION: DNG administration reduces pain and serum CGRP levels in EM patients, offering the potential for innovative treatments and tailored options. Understanding neurotransmitter roles and drug effects can aid in discovering more effective modulators for these pathways.

Toxicological Impact of Bisphenol A on Females' Reproductive System: Review Based on Experimental and Epidemiological Studies.

The study encompassing research papers documented in the last two decades pertaining to the possible influence of bisphenol A (BPA) on the fertility of females are appraised with emphasis on the influence of BPA in reproductive organs (uterus and ovaries) and pregnancy outcomes including discussion on the reproductive process (implantation, estrous cycle, hormone secretion); outcomes reveal a connection amongst BPA and female infertility. Ovary, uterus, and its shape as well as function can alter a person's ability to become pregnant by influencing the hypothalamus-pituitary axis in the ovarian model. Additionally, implantation and the estrous cycle may be affected by BPA. However, more research is warranted to comprehend the underlying action mechanisms and to promptly identify any imminent reproductive harm.

NLRP3 concentration, oxidants, and antioxidants in plasma of endometriosis patients undergoing treatment with dienogest.

OBJECTIVE: NOD-like receptor pyrin domain-containing 3 (NLRP3) is a cytosolic multi-protein complex that induces inflammation and is negatively regulated by progesterone. Previous researches have reported abnormal induction of reactive oxygen species (ROS) and progesterone resistance in endometriosis (EM). Since progesterone regulates ROS level and, consequently, inflammation, our objective is to investigate whether dienogest (DNG) regulates NLRP3 and whether the regulation of NLRP3 inflammasome by DNG in the blood plasma of patients with EM can affect oxidant and antioxidant markers. METHODS: Plasma samples were obtained from control and EM patients experiencing pain symptoms to measure the level of NLRP3, oxidants, and antioxidants. Subsequently, these patients were given oral DNG 2 mg/day for six months for drug treatment. After six months, plasma samples were collected from the patients for re-examination. RESULTS: The findings indicate that DNG reduced NLRP3 concentration and oxidant production while increasing antioxidant production in blood plasma. By reducing NLRP3, DNG was able to alleviate inflammation and pain caused by inflammation in EM patients. CONCLUSION: In conclusion, the use of DNG in EM patients resulted in a decrease in NLRP3 concentration in the patient's plasma. Furthermore, this effect was enhanced by balancing oxidant/antioxidant levels, which may contribute to reducing inflammation associated with EM.

The effect of temperature and storage time on DNA integrity after freeze-drying sperm from individuals with normozoospermia.

OBJECTIVE: This study evaluated the effects of temperature and storage time on the quality and DNA integrity of freeze-dried sperm from individuals with normozoospermia. METHODS: Normal sperm samples from 15 men aged 24 to 40 years were studied. Each sample was divided into six groups: fresh, freezing (frozen in liquid nitrogen), freeze-dried then preserved at room temperature for 1 month (FD-1m-RT), freeze-dried then preserved at room temperature for 2 months (FD-2m-RT), freeze-dried then preserved at 4 °C for 1 month (FD-1m-4 °C), and freeze-dried then preserved at 4 °C for 2 months (FD-2m-4 °C). The morphology, progressive motility, vitality, and DNA integrity of the sperm were evaluated in all groups. RESULTS: In all freeze-dried groups, sperm cells were immotile after rehydration. The freeze-dried groups also showed significantly less sperm vitality than the fresh and frozen groups. Significantly more morphological sperm abnormalities were found in the freeze-dried groups, but freeze-drying did not lead to a significantly higher DNA fragmentation index (DFI). The DFI was significantly higher in the FD-2m-RT group than in the other freeze-dried groups. CONCLUSION: The freeze-drying method preserved the integrity of sperm DNA. The temperature and duration of storage were also identified as factors that influenced the DFI. Accordingly, more research is needed on ways to improve sperm quality in the freeze-drying process.

Does transcutaneous electrical nerve stimulation reduce the laparoscopic related shoulder pain?

OBJECTIVE: Although laparoscopic surgery is a good substitute for laparotomy in reducing postsurgical pain, many patients complain of shoulder pain after laparoscopic surgery and require pain-relief. Post-operative pain management leads to increased patient satisfaction. Transcutaneous Electrical Nerve Stimulation (TENS) is a non-pharmacological, noninvasive modality that reduces pain by activating the descending inhibitory systems in the central nervous system. Given the importance of decreasing shoulder pain after gynecological laparoscopy, the current study aimed to investigate the management of shoulder pain in these patients using TENS. METHODS: This was a retrospective case-control study. A total of 112 women aged 18-45 years who experienced shoulder pain due to gynecologic laparoscopic surgery were included in the study. Patients were divided into TENS and control groups. In the TENS group, TENS was used twice for 20 minutes each, but in the control group, the patients received regular treatment. Patients were evaluated at intervals of 2, 4, 8, 24, 48, and 72 hours after laparoscopy for shoulder pain score. RESULTS: The results showed a significant decrease in visual analog scale scores at 2, 4, and 8-hour in the TENS group compared with the control group. At 24 hours evaluation, although the pain was reduced, the difference was not significant. At 48- and 72-hour assessment, all patients in each group reported zero score for severity of pain. CONCLUSION: The findings suggest that TENS significantly reduces postoperative shoulder pain.

The Effect of Vitamin D3 Supplemented with PhytoSolve on Genes Involved in Implantation in an NMRI Mice Model of Polycystic Ovary Syndrome: An Experimental Study.

OBJECTIVE: Vitamin D3 has been shown to be effective in the treatment of PCOS. However, due to its poor solvability and bioavailability, effective time is delayed and dosage requirements are increased. In our previous study, we demonstrated that PhytoSolve containing VD3 is more effective than vitamin D3 alone in the treatment of PCOS. In this study, we aimed to investigate the effect of this vitamin D3 formulation on gene expression involved in implantation in patients with PCOS. METHODS: To create PhytoSolve, Lipid S75, glycerol, and MCT oil were combined using a sonicator probe. Six groups, each consisting of 36 female Naval Medical Research Institute (NMRI) mice, were included in the following groups: control; sham; PCOS; PhytoSolve; PhytoSolve containing VD3; and vitamin D3. The mice were given DHEA injections to induce PCOS. After administering PhytoSolve containing VD3 and vitamin D3 by gavage for one week from the 13th day of model creation, the female mice were mated and endometrial tissue was collected for analysis of LIF, ß-integrin, and HOXA10 proteins and genes. RESULTS: Compared to the group receiving vitamin D3 alone, the group receiving PhytoSolve containing vitamin D3 showed a significant increase in the expression of LIF, ß-integrin, and HOXA10 genes (p<0.05). Although there was an increase in the expression of ß-integrin and HOXA10 proteins in the group given PhytoSolve containing vitamin D3 compared to the group given vitamin D3, this increase was not significant. However, the increase in LIF protein expression in the group given PhytoSolve containing vitamin D3 was significant when compared to the group given vitamin D3 (p<0.05). CONCLUSIONS: The use of PhytoSolve containing vitamin D3 was more effective than vitamin D3 alone. The PhytoSolve formulation might be a useful solution for medications with limited solubility and bioavailability.

TLR-1, TLR-2, and TLR-6 MYD88-dependent signaling pathway: A potential factor in the interaction of high-DNA fragmentation human sperm with fallopian tube epithelial cells.

OBJECTIVE: The DNA integrity of spermatozoa that attach to fallopian tube (FT) cells is higher than spermatozoa that do not attach. FT epithelial cells can distinguish normal and abnormal sperm chromatin. This study investigated the effects of sperm with a high-DNA fragmentation index (DFI) from men with unexplained repeated implantation failure (RIF) on the Toll-like receptor (TLR) signaling pathway in human FT cells in vitro. METHODS: Ten men with a RIF history and high-DFI and 10 healthy donors with low-DFI comprised the high-DFI (>30%) and control (<30%) groups, respectively. After fresh semen preparation, sperm were co-cultured with a human FT epithelial cell line (OE-E6/E7) for 24 hours. RNA was extracted from the cell line and the human innate and adaptive immune responses were tested using an RT2 profiler polymerase chain reaction (PCR) array. RESULTS: The PCR array data showed significantly higher TLR-1, TLR-2, TLR-3, TLR-6, interleukin 1α (IL-1α), IL-1ß, IL-6, IL-12, interferon α (IFN-α), IFN-ß, tumor necrosis factor α (TNF-α), CXCL8, GM-CSF, G-CSF, CD14, ELK1, IRAK1, IRAK2, IRAK4, IRF1, IRF3, LY96, MAP2K3, MAP2K4, MAP3K7, MAP4K4, MAPK8, MAPK8IP3, MYD88, NFKB1, NFKB2, REL, TIRAP, and TRAF6 expression in the high-DFI group than in the control group. These factors are all involved in the TLR-MyD88 signaling pathway. CONCLUSION: The MyD88-dependent pathway through TLR-1, TLR-2, and TLR-6 activation may be one of the main inflammatory pathways activated by high-DFI sperm from men with RIF. Following activation of this pathway, epithelial cells produce inflammatory cytokines, resulting in neutrophil infiltration, activation, phagocytosis, neutrophil extracellular trap formation, and apoptosis.

Evaluation of expression of biomarkers of PLAGL1 (ZAC1), microRNA, and their non-coding RNAs in patients with endometriosis.

Since the PLAGL1 (ZAC1) gene is expressed in the human endometrium. It may be involved in the etiology of endometrial disorders by its abnormal regulation and expression. This study aimed to investigate the Zac1 gene and related microRNA and LncRNA and its alterations in patients with endometriosis. Blood plasma, ectopic (EC) and eutopic (EU) endometrial samples were gathered from 30 patients with endometriosis and 30 healthy fertile women, and the Q-PCR technique was used to determine the expression level of Zac1 mRNA and microRNAs (miR-1271-5p, hsa-miR-490-3pin) and LncRNAs (TONSL-AS1 TONSL, KCNQ1OT1 KCNQ1). According to the results, the Zac1 gene and KCNQ1OT1 KCNQ1, TONSL-AS1 TONSL LncRNA expression were significantly decreased in the endometriosis group versus the control group (P < 0.05). MiR-1271-5p and hsa-miR-490-3pin microRNA expression were significantly raised in the endometriosis group as opposed to the control group (P < 0.05). In summary, this research for the first time revealed that identifying Zac1 expression provides us with new indicators for evaluating endometriosis.

Cutting-edge techniques provide insights regarding repeated implantation failure patients.

RESEARCH QUESTION: Can time-lapse parameters and the transcriptional profile of cumulus cells be used to achieve a more stringent and non-invasive method of embryo assessment and to identify possible factors affecting the embryo's ability to implant in repeated implantation failure (RIF) patients? DESIGN: A total of 190 embryos from 18 oocyte donors and 145 embryos from 15 RIF patients were evaluated based on time-lapse parameters. Three morphokinetic parameters including T5 (time to reach five cells), T3 (time to reach three cells) and CC2 (time to two to three cells) were recorded for all embryos. Embryos that had all three parameters in the normal range were graded as high quality and comparison between these parameters were compared in high-quality embryos between two groups. The transcriptional profile of cumulus cells related to high-quality embryos of both groups were analysed by RNA sequencing and compared. Finally, the possible relationship between differentially expressed genes and time-lapse parameters was examined. RESULTS: T5 was significantly lower in the RIF group than the donor group (P = 0.011). The cumulus cell transcriptome analysis showed 193 genes were down-regulated and 222 genes up-regulated. The mammalian target of rapamycin and the transforming growth factor beta pathways were significantly increased in the RIF group compared to the donor group (P = 0.007 and 0.01, respectively). Vitamin B12 and fatty acid beta-oxidation pathways were also significantly reduced in the RIF group compared to the donor group (P = 0.006 and 0.01, respectively). CONCLUSIONS: Differences in the transcriptomic profiles of cumulus cells and some morphokinetic parameters may be one of the main factors contributing to unexplained RIF.

Effects of Au@Ag core-shell nanostructure with alginate coating on male reproductive system in mice.

Despite the widespread use of silver nanoparticles (NPs), these NPs can accumulate and have toxic effects on various organs. However, the effects of silver nanostructures (Ag-NS) with alginate coating on the male reproductive system have not been studied. Therefore, this study aimed to investigate the impacts of this NS on sperm function and testicular structure. After the synthesis and characterization of Ag-NS, the animals were divided into five groups (n = 8), including one control group, two sham groups (received 1.5 mg/kg/day alginate solution for 14 and 35 days), and two treatment groups (received Ag-NS at the same dose and time). Following injections, sperm parameters, apoptosis, and autophagy were analyzed by the TUNEL assay and measurement of the mRNA expression of Bax, Bcl-2, caspase-3, LC3, and Beclin-1. Fertilization rate was assessed by in vitro fertilization (IVF), and testicular structure was analyzed using the TUNEL assay and hematoxylin and eosin (H&E) staining. The results showed that the NS was rod-shaped, had a size of about 60 nm, and could reduce sperm function and fertility. Gene expression results demonstrated an increase in the apoptotic markers and a decrease in autophagy markers, indicating apoptotic cell death. Moreover, Ag-NS invaded testicular tissues, especially in the chronic phase (35 days), resulting in tissue alteration and epithelium disintegration. The results suggest that sperm parameters and fertility were affected. In addition, NS has negative influences on testicular tissues, causing infertility in men exposed to these NS.

Does Gold-Silver Core-Shell Nanostructure with Alginate Coating Induce Apoptosis in Human Lymphoblastic Tumoral (Jurkat) Cell Line?

Background: T-cell acute lymphoblastic leukemia (T-ALL) is known as an aggressive malignant disease resulting from the neoplastic alteration of T precursor cells. Although treatment with stringent chemotherapy regimens has achieved an 80% cure rate in children, it has been associated with lower success rates in adult treatment. Silver nanoparticles (Ag-NPs) have a toxic effect on human breast cancer cells, human glioblastoma U251 cells, and chronic myeloid leukemia cells in vitro. This study aimed to investigate the effect of Ag nanostructures (Ag-NSs) on Jurkat cells' viability and apoptosis. Methods: The Jurkat cell line was acquired. Following the synthesis Ag-NSs and their characterization, they were incubated with Jurkat cells at different doses for 24, 48, and 72 hours to determine the optimal time and dose. Two groups were examined: a control group with Jurkat cells without nanostructure maintained in the same medium as the cells in the treatment group without changing the medium, and a treatment group with cells treated with the Ag nanostructure solution at a dose of 75 µg/ml for 48 hours according to the MTT results. After 48 hours, the cells from the two groups were used for the q RT-PCR of the apoptotic genes (BAX, BCL-2, and CASPASE-3). Results: According to our results, the rod-shaped silver nanostructures had a size of about 50 nm, increased apoptotic markers, including BAX and CASPASE-3, and induced cell death. Conclusions: Ag-NSs have anticancer properties and can induce apoptosis of cells; therefore, they may be a potential candidate for the treatment of T-cell acute lymphoblastic leukemia.

Treatment of Ovarian Hyperstimulation Syndrome in a Mouse Model by Cannabidiol, an Angiogenesis Pathway Inhibitor.

Studies suggest that ovarian hyperstimulation syndrome (OHSS) can be treated by reducing the level of vascular endothelial growth factor (VEGF). However, due to the side effects of commercially available VEGF-reducing drugs, they can be ruled out as a suitable treatment for OHSS; therefore, researchers are looking for new medications to treat OHSS. This study is aimed at investigating the effects of cannabidiol (CBD) in an OHSS model and to evaluate its efficacy in modulating the angiogenesis pathway and VEGF gene expression. For this purpose, 32 female mice were randomly divided into four groups (eight mice per group): control group, group 2 with OHSS induction, group 3 receiving 32 nmol of dimethyl sulfoxide after OHSS induction, and group 4 receiving 30 mg/kg of CBD after OHSS induction. The animals' body weight, ovarian weight, vascular permeability (VP), and ovarian follicle count were measured, and the levels of VEGF gene and protein expression in the peritoneal fluid were assessed. Based on the results, CBD decreased the body and ovarian weights, VP, and corpus luteum number compared to the OHSS group (p < 0.05). The peritoneal VEGF gene and protein expression levels reduced in the CBD group compared to the OHSS group (p < 0.05). Also, CBD caused OHSS alleviation by suppressing VEGF expression and VP. Overall, CBD downregulated VEGF gene expression and improved VP in OHSS.

Accompaniment of Time-Lapse Parameters and Cumulus Cell RNA-Sequencing in Embryo Evaluation.

The aim of this study was to investigate the use of time-lapse morphokinetic parameters and cumulus cells transcriptomic profile to achieve a more accurate and non-invasive method in embryo evaluation. Two hundred embryos from 20 couples were evaluated based on morphokinetic characteristics using time-lapse. Embryos were divided into the high-quality, moderate-quality, and bad-quality groups. Non-fertilized oocytes were considered as the fourth group. T5 (time to five cells), S2 (time from three to four cells), and CC2 (time from two to three cells) were recorded. Also, the cumulus cells of the respective oocytes were divided into high-quality, moderate-quality, bad-quality, and non-fertilized groups based on the grading of the embryos. Then their transcriptomic profiles were analyzed by RNA-sequencing. Finally, the correlation between differentially expressed genes and embryo time-lapse parameters was investigated. T5 was the only timing that showed a statistically significant difference between high-quality group and other groups. RNA-sequencing results showed that 37 genes were downregulated and 106 genes were upregulated in moderate, bad-quality, and non-fertilized groups compared to high-quality group (q value < 0.05). These genes were involved in the main biological processes such as cell cycle, DNA repair, cell signaling and communication, transcription, and cell metabolism. Embryos graded in different groups showed different transcriptomic profiles in the related cumulus cells. Therefore, it seems that embryo selection using the combination of cytokinetics and cumulus cells gene expression can improve the accuracy of the embryo selection and pregnancy rate in ART clinics.