Duygu Kütük

Background: Diminished ovarian reserve is characterized by a decrease in oocyte count and estrogen levels, which leads to infertility. The genetic and epigenetic mechanisms in MI to MII transition or complete MII phase in the oocyte maturation process estrogen receptor alpha and piRNA relationship were evaluated. Methods: This study analyzed 100 cumulus oophorous complex samples from normoresponder and DOR patients undergoing IVF, subdivided into metaphase I and metaphase II stages. To elucidate the ER-α, PIWIL3, piR-651, and piR-823 genes qRT-PCR was used and qualitative ER-α protein expressions were determined by immunohistochemistry. Pearson’s correlation analysis was utilized to evaluate the interactions between genes within each experimental group. Results: The DOR samples exhibited significant downregulation of ER-α gene and protein expression compared to the NOR controls. PIWIL3 gene, piR-651, and piR-823 expressions reduced in DOR MI and MII. Strong positive correlations among ER-α, PIWIL3, piR-651, and piR-823 were observed in NOR, whereas DOR showed weaker correlations and immunohistochemistry verified lower ER-α protein levels in DOR. Conclusions: The disruption of ER-α and piRNA-related gene networks in DOR may underlie the suboptimal maturation of oocytes, and monitoring ER-α, PIWIL3, piR-651, and piR-823 expressions could facilitate early determination of maturation stages and improve assessment of ovarian reserve. The potential for transposition to MII in NOR and DOR oocytes was observed in relation to the association between ER-α protein/gene expression and PIWIL3, which regulates ER-α. Moreover, piR-651 and piR-823, whose expressions depend on estrogen level, indirectly regulate oocyte maturation from MI to MII in both NOR and DOR epigenetically. We suggest that the MI and MII stages of oocytes could be determined earlier in NOR and DOR cases by controlling ER-α, PIWIL3, piR-651 and piR-823 expressions. These molecular markers indicate promise for diagnostic applications in reproductive medicine, warranting further validation in larger cohorts.

Aims: Cytochrome C (Cyt C) is a small heme protein located in the mitochondria, playing a dual and crucial role in cellular physiology as cellular respiration and apoptosis. This study aimed to investigate Cyt C gene expression in spent embryo culture fluid (SECF) as a non-invasive biomarker for distinguishing between Normoresponder (NOR), Early-Diminished Ovarian Reserve (E-DOR), Advanced-Diminished Ovarian Reserve (A-DOR), and polycystic ovarian syndrome (PCOS) patients undergoing in vitro fertilization (IVF). Methods: SECF samples were collected from patients at Bahçeci Health Group Umut IVF Laboratory, İstanbul, with informed consent and ethical approval (2023/19-22). Forty-eight samples from each group (NOR, E-DOR, A-DOR, PCOS) were analyzed. Ovarian stimulation followed an antagonist protocol, and embryos were cultured individually. On day 3, embryos were morphologically graded. Total RNA was isolated from SECF, converted to cDNA, and Cyt C gene expression was quantified using RT-PCR, with GAPDH as an internal control. Statistical analyses included the Kolmogorov-Smirnov and Shapiro-Wilk tests for normality, Sidak for multiple comparisons, and the Pearson correlation test for group relationships. Results: Compared to NOR samples, Cyt C gene expression was significantly upregulated in A-DOR samples, while E-DOR samples showed downregulation (p0.05). Pearson correlation revealed a significant relationship only between NOR and A-DOR SECF samples (r<0.01). Conclusion: These findings suggest that Cyt C expression in SECF can differentiate between early and advanced stages of diminished ovarian reserve, with age-related differences in apoptosis mechanisms. PCOS cases, characterized by a high quantity but low quality of oocytes, showed no significant variation in Cyt C expression, likely due to less active apoptotic pathways. Monitoring Cyt C in SECF may provide a valuable, non-invasive tool for assessing embryo quality and optimizing IVF outcomes. Further research is needed to validate these findings and establish clinical protocols.

Background/Objectives: Oocyte maturation is a process involving both nuclear and cytoplasmic development regulated by epigenetic changes in gene expression. Cyclin-B3 (CCNB3) and cyclin-A2 (CCNA2) genes are thought to be involved in oocyte maturation; however, the expression profiles and key function in Metaphase-I (MI) and Metaphase-II (MII) phases have yet to be fully elucidated. Small non-coding RNA sequences are involved in epigenetic regulation of specific transcriptional targets, whereas microRNAs (miRNAs) participate in the post-transcriptional and translational repression of target genes. This study examined the expression levels of CCNB3, CCNA2, and their associated miRNAs (miR-17, miR-106b, miR-190a, miR-1275) in cumulus oophorous complex (COC) cells derived from MI and MII oocytes of NOR and DOR IVF cases, with particular emphasis on elucidating their functions during the transition from MI to MII stage. Methods: Follicular fluid containing cumulus–oocyte complex (COC) cells obtained from oocytes of 120 cases in each group NOR MI (n = 30), NOR MII (n = 30), DOR MI (n = 30), and DOR MII (n = 30) who were admitted to the Istanbul Bahçeci Health Group Assisted Reproductive Treatment Center. Following total RNA isolation from COC cells, the gene and protein expression levels of CCNB3 and CCNA2, along with the expression of miR-17, miR-106b, miR-190a, and miR-1275, were assessed using (qPCR-based assay) and immunohistochemistry (IHC). To investigate the functional roles of COC cell populations, morphological analysis was performed using H&E staining. Additionally, metadata of the cases, including age, number of oocytes, fertilization, and embryonic development rates, were evaluated. Results: The expressions of miR-17 and miR-1275 were significantly elevated in both NOR MI and DOR MI groups compared to their respective NOR MII and DOR MII groups (p < 0.05). Additionally, miR-106b levels were higher in the NOR MII group relative to NOR MI (p < 0.05), while an increase was also observed in DOR MI compared to DOR MII (p < 0.05). No difference was observed in miR-190a expression between the NOR and DOR (p > 0.05). Based on the results of H and E staining, the NOR MI, NOR MII, DOR MI, and DOR MII groups exhibited distinct variations in cellular morphology, nuclear characteristics, cytoplasmic volume, and cell density. Conclusions: CCNB3 is predicted to be a potential candidate for determining MI between the NOR and DOR cases. On the other hand, only for the NOR MII cases could CCNA2 provide evidence of oocyte maturation. Moreover, we determined the relationship between related genes and miRNAs which target CCNA2 and CCNB3. Genetic and protein expression analysis across diverse molecular pathways and miRNAs yielded comprehensive preliminary data regarding the developmental stages of oocytes at the MI and MII phases, and their fertilization potential following maturation shows potential and warrants prospective validation with clinical performance evaluation.

An embryo culture medium is a specialized set of ambient conditions, technological equipment, and nutrients that embryos require to grow properly. We aimed to investigate the Ki-67, hTERT, and HIF-1α gene expression differences between developing and non-developing embryos in spent embryo culture medium. Ki-67, hTERT, and HIF-1α gene expressions were determined from the spent embryo culture medium containing developing and non-developing embryos of 20 normoresponder patients admitted to the Bahçeci Umut IVF Center. An increase in hTERT gene expression (p < 0.05) and a decrease in HIF-1α gene expression (p < 0.001) were observed in mediums of developing compared to the non-developing embryos. No difference was observed in Ki-67 gene expression (p > 0.05). While there was a correlation between Ki-67 and HIF-1α genes in the non-growing group (r < 0.01); no correlation was observed in the developing group (r > 0.05). Both normoresponder groups will be similar in terms of proliferation rate. The low HIF-1α expression that observed high telomerase activity in embryo development maintains continuity and avoids mechanisms that result in cell death. A molecular study of the embryo development in patients with similar characteristics may help to understand the pathogenesis of the disease and establish a diagnosis and specific treatment.

Incorporating artificial intelligence (AI) into in vitro fertilization (IVF) laboratories signifies a significant advancement in reproductive medicine. AI technologies, such as neural networks, deep learning, and machine learning, promise to enhance quality control (QC) and quality assurance (QA) through increased accuracy, consistency, and operational efficiency. This comprehensive review examines the effects of AI on IVF laboratories, focusing on its role in automating processes such as embryo and sperm selection, optimizing clinical outcomes, and reducing human error. AI’s data analysis and pattern recognition capabilities offer valuable predictive insights, enhancing personalized treatment plans and increasing success rates in fertility treatments. However, integrating AI also brings ethical, regulatory, and societal challenges, including concerns about data security, algorithmic bias, and the human–machine interface in clinical decision-making. Through an in-depth examination of current case studies, advancements, and future directions, this manuscript highlights how AI can revolutionize IVF by standardizing processes, improving patient outcomes, and advancing the precision of reproductive medicine. It underscores the necessity of ongoing research and ethical oversight to ensure fair and transparent applications in this sensitive field, assuring the responsible use of AI in reproductive medicine.

Mitochondria is an important organelle for the oocyte-to-embryo transition in the early embryonic development period. The oocyte uses mitochondria functionally and its mitochondrial DNA (mtDNA) content as the main energy source in the embryo development at the preimplantation stage. The aim of this study is to compare mitophagic, apoptotic and humanin gene expressions from the culture medium fluid in which embryos are developed and monitored among normoresponder (NOR), polycystic ovary syndrome (PCOS), young and older patients with poor ovarian reserve (POR). The study groups consisted of infertile patients who applied to the Bahçeci Umut IVF Center as NOR (Control), PCOS, POR-Advanced (POR-A) and POR-Young (POR-Y). After the isolation of total RNA from the collected samples, MFN1, MFN2, PINK1, PARKIN, SMN1, SMN2, p53 and Humanin gene expressions were determined by Real Time-PCR. The average age of only the POR-A was determined to be higher than the NOR (p < 0.001). The MFN1, SMN2 (p < 0.05), Humanin and p53 gene expressions (p < 0.001) increased, while PINK1 gene expression decreased (p < 0.05), in the POR-Y compared to the NOR. A decrease in MFN2, PARKIN (p < 0.05) and PINK1 gene expressions was determined in the PCOS compared to the NOR (p < 0.001). Furthermore, a decrease was observed in MFN2, PINK1 (p < 0.001) and Humanin gene expressions compared to the NOR (p < 0.05). The current data are the first in the literature determining the apoptotic and mitophagic status of the oocyte. The current results prove that waste embryo culture fluid may provide a non-invasive profile for important cellular parameters such as mitochondrial dysfunction in female infertility. The evaluation of significant cellular parameters can be performed much earlier without any intervention into the embryo.