The Involvement of Lumican in Human Ovulatory Processes.

Based on a previous global transcriptome sequencing project, we hypothesized that Lumican (LUM) might play a role in ovulatory processes. We sought to determine LUM gene expression under various conditions in human preovulatory follicles. The in vitro expression of LUM mRNA in mural (MGCs) and cumulus (CGCs) granulosa cells was characterized using quantitative real-time polymerase chain reaction (qRT-PCR). Immunohistochemical staining was used to identify human LUM expression in follicles at different developmental stages. Cell signaling studies were performed by treating human MGCs with human chorionic gonadotropin (hCG) and both, different stimulators and inhibitors to determine their effect on LUM expression by using qRT-PCR. Cell confluence studies were carried out to study the correlation between LUM expression and follicle cell proliferation. Follicular MGCs and CGCs of women undergoing in vitro fertilization (IVF) procedures due to endometriosis were analyzed for differences in LUM expression patterns by qRT-PCR. LUM mRNA expression was significantly higher in MGCs as compared to CGCs. In CGCs, LUM mRNA was higher in mature metaphase II (MII) oocytes than in germinal vesicle (GV) and metaphase I (MI) oocytes. LUM expression was significantly upregulated in response to hCG in cultured MGCs. Immunohistochemistry of human ovaries revealed LUM was mostly present in MGCs of large preovulatory and postovulatory follicles and absent from primordial follicles. Using pharmacological activators and inhibitors, we demonstrated that LUM induction by luteinizing hormone (LH)/hCG is carried through the mitogen-activated protein kinase (MEK) pathway. LUM expression was induced in high-density cell cultures in a confluence-dependent manner. MGCs from follicles of subjects with endometriosis exhibited reduced mRNA transcription levels compared to control subjects. Our study confirms that LUM is a newly discovered ovulatory gene. LUM might play an important role during the preovulatory period up until ovulation as well as in endometriosis infertility. A better understanding of LUM's role might provide potential new treatment paradigms for some types of female infertility.

Day 5 vitrified blastocyst transfer versus day 6 vitrified blastocyst transfer in oocyte donation program.

The superiority of day 5 blastocysts compared to day 6 blastocysts in fresh cycle transfers was previously demonstrated and attributed mainly to endometrial asynchrony. Data from frozen blastocysts transfers showed conflicting results, possibly due to heterogeneous patient population and embryo quality. The aim of this study was to compare clinical pregnancy rate (CPR) and live birth rate (LBR) between transfers of vitrified day 5 blastocysts and day 6 blastocysts in oocyte donation, blastocyst-only cycles. In a retrospective, multi-center study, with a single oocyte donation program, a total of 1840 frozen embryo transfers (FET's) were analyzed, including 1180 day 5 blastocysts and 660 day 6 blastocysts transfers. Day 5 blastocyst transfers had better embryonic development and significantly higher CPRs (34.24% vs. 20.15%, P < 0.0001), higher LBRs (26.89% vs. 14.77%, P < 0.0001), less cycles to LBR (1.83 ± 0.08 vs. 2.39 ± 0.18, P = 0.003) and shorter time to LBRs (76.32 ± 8.7 vs. 123.24 ± 19.1 days, P = 0.01), compared to day 6 transfers, respectively. A multivariate stepwise logistic regression indicated, that day 5 transfer was an independent factor for CPRs (OR 1.91; 95% CI 1.43-2.54, P < 0.001) and LBRs (OR 2.26; 95% CI 1.19-4.28, P = 0.01), regardless of embryo quality, compared to day 6. In conclusion, day 5 blastocysts in oocyte donation program have significantly higher CPRs and LBRs, and present shorter time to delivery, compared to day 6 blastocysts, regardless of embryo quality.

Elucidating Decorin's role in the preovulatory follicle.

BACKGROUND: DCN (decorin) is a proteoglycan known to be involved in regulating cell proliferation, collagen fibril organization and migration. In our global transcriptome RNA-sequencing approach to systematically identify new ovulation-associated genes, DCN was identified as one of the highly regulated genes. We therefore hypothesize that DCN may have a role in ovulatory processes such as cell migration and proliferation. AIM: To characterize the expression, regulation and function of the proteoglycan DCN in the human ovarian follicles during the preovulatory period. METHODS: The in-vivo expression of DCN mRNA in mural (MGCs) and cumulus (CGCs) granulosa cells was characterized using quantitative RT-PCR and western blot. A signaling study was performed by treating human MGCs cultures with gonadotropins and different stimulators and inhibitors to determine their effect on DCN expression by qRT- PCR and elucidate the pathways regulating these proteins. In a functional study, KGN granulosa cell line was used to study cell migration with a scratch assay. RESULTS: DCN mRNA expression was significantly higher in MGCs compared to CGCs. DCN mRNA was significantly higher in CGCs surrounding mature metaphase II (MII) oocytes compared to CGCs of germinal vesicle (GV) and metaphase I (MI) oocytes. hCG significantly increased DCN mRNA and protein expression levels in cultured MGCs. Using signal transduction activators and inhibitors, we demonstrated that DCN induction by LH/hCG is carried out via PKA, PKC, ERK/MEK, and PI3K pathways. We showed that DCN expression is also induced in high-density cell cultures, in a dose-dependent pattern. In addition, progesterone induced a significant increase in DCN secretion to the media. MGCs from follicles of endometriosis patients exhibited reduced (about 20% of) mRNA transcriptions levels compared to MGCs follicles of control patients. More significantly, we found that DCN has an inhibiting effect on KGN cell migration. CONCLUSIONS: Our study indicates that DCN is a unique ovulatory gene. Our findings support the hypothesis that DCN plays an important new role during the preovulatory period and ovulation, and stress its involvement in endometriosis infertility. A better understanding of DCN role in ovulation and endometriosis may provide treatment for some types of infertility.

How Artificial Intelligence Can Improve Our Understanding of the Genes Associated with Endometriosis: Natural Language Processing of the PubMed Database.

Endometriosis is a disease characterized by the development of endometrial tissue outside the uterus, but its cause remains largely unknown. Numerous genes have been studied and proposed to help explain its pathogenesis. However, the large number of these candidate genes has made functional validation through experimental methodologies nearly impossible. Computational methods could provide a useful alternative for prioritizing those most likely to be susceptibility genes. Using artificial intelligence applied to text mining, this study analyzed the genes involved in the pathogenesis, development, and progression of endometriosis. The data extraction by text mining of the endometriosis-related genes in the PubMed database was based on natural language processing, and the data were filtered to remove false positives. Using data from the text mining and gene network information as input for the web-based tool, 15,207 endometriosis-related genes were ranked according to their score in the database. Characterization of the filtered gene set through gene ontology, pathway, and network analysis provided information about the numerous mechanisms hypothesized to be responsible for the establishment of ectopic endometrial tissue, as well as the migration, implantation, survival, and proliferation of ectopic endometrial cells. Finally, the human genome was scanned through various databases using filtered genes as a seed to determine novel genes that might also be involved in the pathogenesis of endometriosis but which have not yet been characterized. These genes could be promising candidates to serve as useful diagnostic biomarkers and therapeutic targets in the management of endometriosis.

Characterization of the human cumulus cell transcriptome during final follicular maturation and ovulation.

Cumulus expansion and oocyte maturation are central processes in ovulation. Knowledge gained from rodent and other mammalian models has revealed some of the molecular pathways associated with these processes. However, the equivalent pathways in humans have not been thoroughly studied and remain unidentified. Compact cumulus cells (CCs) from germinal vesicle cumulus oocyte complexes (COCs) were obtained from patients undergoing in vitro maturation (IVM) procedures. Expanded CCs from metaphase 2 COC were obtained from patients undergoing IVF/ICSI. Global transcriptome profiles of the samples were obtained using state-of-the-art RNA sequencing techniques. We identified 1746 differentially expressed (DE) genes between compact and expanded CCs. Most of these genes were involved in cellular growth and proliferation, cellular movement, cell cycle, cell-to-cell signaling and interaction, extracellular matrix and steroidogenesis. Out of the DE genes, we found 89 long noncoding RNAs, of which 12 are encoded within introns of genes known to be involved in granulosa cell processes. This suggests that unique noncoding RNA transcripts may contribute to the regulation of cumulus expansion and oocyte maturation. Using global transcriptome sequencing, we were able to generate a library of genes regulated during cumulus expansion and oocyte maturation processes. Analysis of these genes allowed us to identify important new genes and noncoding RNAs potentially involved in COC maturation and cumulus expansion. These results may increase our understanding of the process of oocyte maturation and could ultimately improve the efficacy of IVM treatment.

Head progression distance in prolonged second stage of labor: relationship with mode of delivery and fetal head station.

OBJECTIVE: To evaluate the clinical significance of fetal head progression distance (HPD), measured by transperineal ultrasound, during prolonged second stage of labor. METHODS: In this prospective study, a single operator, who was blinded to the results of the digital examination, assessed using transperineal ultrasound women at ≥ 37 weeks of gestation with failure to progress in the second stage of labor. Patients had an empty urinary bladder and the examination was performed during maternal pushing. HPD was defined as the length of the line perpendicular to the infrapubic line that would connect it to the lowest part of the fetal bony skull. We analyzed associations between HPD and digital examination of fetal head station, fetomaternal characteristics, mode of delivery and perinatal outcome. RESULTS: Sixty-five patients in prolonged second stage of labor participated in the study. The overall mean HPD was 6.50 (± 1.35; 95% CI, 6.16-6.83) cm. No correlation was found between HPD and head position or mode of delivery, but HPD was positively correlated with fetal head station and neonatal head circumference measured after delivery. Logistic regression and receiver-operating characteristics curve analysis demonstrated no significant predictive value of HPD with respect to mode of delivery. CONCLUSION: Although HPD in prolonged second stage of labor could not predict mode of delivery, it may have a role as an ancillary tool for fetal head station assessment.

Pubic arch angle in prolonged second stage of labor: clinical significance.

OBJECTIVE: To evaluate the clinical significance of the pubic arch angle (PAA) measured by transperineal ultrasound during prolonged second stage of labor. METHODS: We evaluated prospectively 62 women ≥ 37 weeks of gestation with failure to progress in the second stage of labor. Transperineal ultrasound (transverse plane) was used to measure the pubic arch angle. Correlations with fetomaternal characteristics, mode of delivery and perinatal outcome were evaluated. RESULTS: The mean PAA was 101.1° (± 13.1°; range, 80°-135°). We found a negative correlation with maternal age. Patients with an occipitotransverse fetal position had a significantly smaller angle compared with those with occipitoanterior positions (94.3° ± 5.5° vs. 103.2° ± 14.8°, P < 0.05), as did those with operative deliveries compared with those with spontaneous vaginal delivery (97.1° ± 11.5° vs. 110.1° ± 14.0°, P < 0.05). The prediction of operative delivery in prolonged second stage of labor by receiver-operating characteristics curve using PAA alone yielded an area under the curve of 0.75. The predicted probability for operative delivery increased as PAA decreased, with an odds ratio of 0.933 for each decrease in angle of 1°. CONCLUSION: Our study suggests a correlation between the PAA and mode of delivery in prolonged second stage of labor. This may be used as an adjunctive parameter when considering delivery mode.