Gene pathway analysis of the endometrium at the start of the window of implantation in women with unexplained infertility and unexplained recurrent pregnancy loss: is unexplained recurrent pregnancy loss a subset of unexplained infertility?

This study aims to understand differences/similarities in the genetic profile of the endometrium at the start of window of implantation (WOI) in women with unexplained infertility (UI) and unexplained recurrent pregnancy loss (uRPL). Differentially expressed genes (DEGs) from the endometrium were evaluated using gene expression array and pathway enrichment analysis was performed to analyse gene expression pathways involved in both conditions. We found 2,171 genes arranged in 117 pathways and 730 genes arranged in 33 pathways differentially expressed in endometrium of patients in UI and uRPL, respectively. Complement-coagulation cascades, morphine addiction pathway, and PI3K-Akt signalling pathway were predominantly differentially expressed in UI. Cancer pathways, NF-κB signalling pathway, and actin cytoskeleton regulation pathway showed significant changes in uRPL. Forty-eight percent of DEGs and 84% of differentially expressed pathways in uRPL were found in the endometrium of UI patients. Unexpected close association in gene expression pathways between UI and uRPL is observed supporting the hypothesis 'uRPL is a clinical subset of UI'. Yet 100% DEGs overlap wasn't found suggesting the endometrium has still some different gene expression patterns at start of WOI in UI and uRPL. Lastly, diagnostic tools may be developed for uRPL because more specific genes-pathways are involved compared with UI, which shows broader genetic expression profile.

Integrated bioinformatics analysis of validated and circulating miRNAs in ovarian cancer.

Recent studies have focused on the early detection of ovarian cancer (OC) using tumor materials by liquid biopsy. The mechanisms of microRNAs (miRNAs) to impact OC and signaling pathways are still unknown. This study aims to reliably perform functional analysis of previously validated circulating miRNAs' target genes by using pathfindR. Also, overall survival and pathological stage analyses were evaluated with miRNAs' target genes which are common in the The Cancer Genome Atlas and GTEx datasets. Our previous studies have validated three downregulated miRNAs (hsa-miR-885-5p, hsa-miR-1909-5p, and hsalet7d-3p) having a diagnostic value in OC patients' sera, with high-throughput techniques. The predicted target genes of these miRNAs were retrieved from the miRDB database (v6.0). Active-subnetwork-oriented Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis was conducted by pathfindR using the target genes. Enrichment of KEGG pathways assessed by the analysis of pathfindR indicated that 24 pathways were related to the target genes. Ubiquitin-mediated proteolysis, spliceosome and Notch signaling pathway were the top three pathways with the lowest p-values (p < 0.001). Ninety-three common genes were found to be differentially expressed (p < 0.05) in the datasets. No significant genes were found to be significant in the analysis of overall survival analyses, but 24 genes were found to be significant with pathological stages analysis (p < 0.05). The findings of our study provide in-silico evidence that validated circulating miRNAs' target genes and enriched pathways are related to OC and have potential roles in theranostics applications. Further experimental investigations are required to validate our results which will ultimately provide a new perspective for translational applications in OC management.

Regulatory effect of miR-195 in the placental dysfunction of preeclampsia.

Purpose: Preeclampsia (PE) is a pregnancy specific disease soon after 20 weeks of gestation where major symptoms are hypertension and proteinuria. The underlying pathology is believed to be abnormal placentation. Epigenetic and genetic factors have significant roles in abnormal placental development. MicroRNA's (miRNAs), being one of the most important epigenetic regulators, take part in abnormal placentation. Hsa-miR-195 is a molecule associated with abnormal placental growth mechanisms such as impaired cellular proliferation, inadequate trophoblastic invasion causing defective spiral artery remodeling, and apoptosis. We aimed to evaluate miRNA functions, namely miR-195 expression profile, in order to divulge PE pathogenesis.Methods: In this study, we extracted circulating miRNAs from maternal plasma and placenta from 20 PE patients and 20 normotensive pregnant women. miR-195 was quantified using quantitative real time reverse transcriptase PCR (qRT-PCR). The target genes of miR-195 were predicted by Diana Tools-mirPath, TargetScan, and miRDB databases.Results: We found that miR-195 levels were downregulated (3.83-fold decrease, p < .05) in preeclamptic placenta samples, however miR-195 were undetected in preeclamptic and normotensive plasma samples. The steep down-regulation of miR-195 points to its importance of PE pathogenesis.Conclusion: miR-195 is suggested to regulate PE via its target genes manipulating biological processes such as placental proliferation, apoptosis, and angiogenesis. We propose that detection of decreased miR-195 levels in preeclamptic placentas could be used to enlighten the pathophysiology of PE.

Regulation of HMGA2 and KRAS genes in epithelial ovarian cancer by miRNA hsa-let-7d-3p.

AIM OF THE STUDY: The purpose of this study was to identify specific circulating microRNAs (miRNAs) and investigate expression level of their target genes for evaluation of pathogenesis of epithelial ovarian cancer (EOC). MATERIALS AND METHODS: In this study, we have studied on EOC patients' serum and whole blood, healthy control (HC) serum, and whole blood samples. Sixteen serum samples were collected to compare miRNA expression analysis through microarray. According to microarray results, one of the dysregulated miRNA in serum, hsa-let-7d-3p, was validated by RT-qPCR for discriminate two groups. The hsa-let-7d-3p is one of the tumor suppressive let-7d family members. Let-7d is downregulated in numerous types of cancer, including ovarian cancer and directly targets various oncogenes. We analyzed the let-7d targets, which are High Mobility Group A2 (HMGA2) and (Kirsten Rat Sarcoma Viral Oncogene Homolog), as the oncogenes that are associated with EOC. The relation between target genes of hsa-let-7d-3p and EOC has been examined by Pathway Studio. Twenty serum and whole blood samples collected to analyze expression level of target genes were analyzed by real-time PCR. RESULTS: 31 significantly dysregulated miRNAs were identified by microarray in serum. Hsa-let-7d-3p has been selected for the validation, according to P-value and dysregulated level. RT-qPCR results showed that hsa-let-7d-3p could discriminate EOC patients from HC (P = 0.0484, AUC = 0.7). Furthermore, we identified hsa-let-7d-3p's target genes (HMGA2, KRAS) by bioinformatic analysis. The expression level of genes could discriminate patients with EOC from HC, with a power area under the ROC curves (AUC) of 62 and 64.2, respectively. CONCLUSION: HMGA2 and KRAS could be translationally downregulated by the hsa-let-7d-3p, and the loss of hsa-let-7d-3p expression led to the progression of EOC related to the tumorigenesis, invasion, and metastasis.

Potential Marker Pathways in the Endometrium That May Cause Recurrent Implantation Failure.

The aim of this prospective cohort study was to identify altered biologic processes in the endometrium that may be potential markers of receptive endometrium in patients with repeated implantation failure (RIF) as compared with fertile controls. The study was conducted in a university-affiliated in vitro fertilization (IVF) gynecology clinic and molecular biology and genetics laboratory. Healthy fertile controls (n = 24) and patients with RIF (n = 24) were recruited. Window of implantation gene profiling associated with RIF was performed. Six hundred forty-one differentially expressed genes were identified, and 44 pathways were found enriched. Upon clustering of the enriched pathways, 9 representative pathways were established. The important pathways that were identified included circadian rhythm, pathways in cancer, proteasome, complement and coagulation cascades, citrate cycle, adherens junction, immune system and inflammation, cell cycle, and renin-angiotensin system. The involvement of the circadian rhythm pathway and other related pathways may alter the endometrium's functioning to ultimately cause RIF. Furthermore, we found that the pathogenesis of RIF was multifaceted and that numerous processes were involved. We believe that a better understanding of the underlying mechanisms of RIF will ultimately give rise to better treatment opportunities and to better outcomes in IVF.

Potential biomarker of circulating hsa-miR-1273g-3p level for detection of recurrent epithelial ovarian cancer.

PURPOSE: Ovarian cancer (OC) is first gynaecologic cancer that causes women death and epithelial ovarian cancer (EOC) is the most lethal ovarian cancer type. While treatment is commonly successful, some cases (10-20%) show resistance to chemotherapy which is followed by recurrence. MicroRNA (miRNA) based diagnosis methods are slightly important for recurrent ovarian cancer diagnosis. We aimed to detect novel circulating miRNAs to be used as an early diagnosis and prediction tools for recurrent EOC. METHODS: In this study, recurrent EOC serum samples and healthy control serum samples were compared for miRNA expression analysis by microarray. Microarray results were analyzed by bioinformatics tools and differentially expressed hsa-miR-1273g-3p was obtained. After microarray analysis, differentially expressed hsa-miR-1273g-3p was validated by Real-Time PCR (RT-qPCR). The relation between target genes of hsa-miR-1273g-3p and ovarian cancer were examined by Pathway Studio® (v.11.4.0.8). RESULTS: The expression of hsa-miR-1273g-3p was found to be significantly down-regulated by t test Bonferroni FWER corrected p < 0.05 and fold change > 2, in recurrence EOC compare with healthy controls groups. The RT-qPCR results confirmed that relative expressions of the serum hsa-miR-1273g-3p were significantly down-regulated in patients with recurrent EOC (p = 0.0275). Serum hsa-miR-1273g-3p levels could discriminate patients with recurrent EOC from healthy controls, with a power area under the curve (AUC) of 0.7. CONCLUSION: This study suggested that hsa-miR-1273g-3p plays a significant role in regulation of related genes, which are TNF-alfa, COL1A1, MMP-2, MMP-9, with recurrent EOC outcome. hsa-miR-1273g-3p may be used as a prognostic marker for recurrent EOC after chemotherapy.

MicroRNA expression profiling in placenta and maternal plasma in early pregnancy loss.

Early pregnancy loss (EPL), also termed early miscarriage, is determined as the unintentional expulsion of an embryo or fetus prior to the 12th week of gestation. EPL frequency is ~15% in pregnancies. Fetal development and growth is associate with placental function and vessel development; therefore, the placental genome would represent a useful miscarriage model for (epi)genetic and genomic studies. An important factor of placental development and function is epigenetic regulation of gene expression. microRNAs (miRNAs) are the primary epigenetic regulators which have an important role in placental development and function. In the present study, maternal plasma and villous tissue were collected from 16 EPL cases in 6th­8th gestational weeks (GWs) and 8 abortions (control group) in 6th­8th GWs. Detection of the differences in miRNA expression was performed using microarrays and dysregulated miRNAs were validated by reverse transcription­quantitative polymerase chain reaction (RT­qPCR). miRNA microarray findings revealed that four miRNAs, including hsa­miRNA (miR)­125a­3p, hsa­miR­3663­3p, hsa­miR­423­5p and hsa­miR­575 were upregulated in tissue samples. In maternal plasma, two miRNAs (hsa­let­7c, hsa­miR­122) were upregulated and one miRNA (hsa­miR­135a) was downregulated. A total of 6 out of 7 dysregulated miRNAs were validated using RT­qPCR. The target genes of these dysregulated miRNAs were detected using the GeneSpring database. The aim of the present study was to detect dysregulated miRNAs in maternal plasma and villous cells and identify the target genes of dysregulated miRNAs and their associated pathways. The target gene analyses have revealed that the affected genes are primarily associated with cell migration, proliferation, implantation, adhesion, angiogenesis and differentiation and all are involved with EPL pathogenesis. Therefore, the present study may contribute to the understanding of the molecular mechanisms which lead to EPL.

Expression profiling of maternal plasma and placenta microRNAs in preeclamptic pregnancies by microarray technology.

Preeclampsia (PE) is one of the leading causes of maternal and fetal morbidity and mortality, occurring usually in the second half of pregnancy and affecting approximately 5-8% of pregnancies in the world. miRNAs play critical role in the regulation of placental development processes. We aimed to determine specific novel miRNAs for early diagnosis of preeclampsia which is one of the most dangerous pregnancy diseases. In this study 72 samples, maternal age 22 ≤ and ≤36, have been analyzed; maternal plasma and placental miRNAs were isolated from 18 severe preeclampsia (sPE) patients and 18 controls, respectively. Profiling of human miRNAs (1368 probe) was performed in samples with Agilent v16 microarrays for detection of the differences in miRNA expression between two groups. The results were validated by using TaqMan RT-qPCR method. The analysis indicated that 406 of these miRNAs in all placentas and 42 of these miRNAs in all maternal plasma were expressed. The relative expression analysis has shown that 12 miRNAs (p < 0.05 and >2-fold) in maternal plasma were differentially expressed in PE and control group. However, five miRNAs were validated by qRT-PCR. Once validated miRNAs have been searched in databases for their target genes and function, it has been shown that there are some preeclampsia related pathways as a target such as angiogenesis, cardiovascular, hypertension, placental abruption and preeclampsia disorders. Differentially expressed and validated plasma miRNAs might be used as notable biomarkers for non-invasive early diagnosis of preeclampsia and treatment of disease.

Effect of angiotensin I-converting enzyme and α-actinin-3 gene polymorphisms on sport performance.

Genetic polymorphism is considered to be associated with human physical performance. The angiotensin I-converting enzyme insertion/deletion (ACE I/D) and the α-actinin-3 gene (ACTN3) R577X polymorphisms have been widely investigated for such associations, and functional ACE I/D and ACTN3 R577X polymorphisms have been associated with sprinter performance. The aim of this study was to determine the effect of these polymorphisms on sport performance among 37 elite athletes and 37 healthy controls. The ACE II genotype was identified in 32.43% of the control group and 8.11% of elite athletes, the DD genotype in 37.84% of the control group and 51.35% of the elite athletes, and the ID genotype in 29.73% of the control group and 40.54% of the elite athletes. With regard to the ACTN3 gene, the XX genotype, which confers an advantage for endurance activities, was identified in 10.81% of the control group and 35.14% of the elite athletes. The XX genotype was observed more frequently than the RR genotype (advantageous for sprinting), which was identified in 2.70% of the control group and 10.81% of elite athletes. The RX genotype (observed in 86.48% of the control group and in 54.05% of the elite athletes) was the most common genotype of the individuals in the present study. The study showed that ACTN3 and ACE gene polymorphisms have an effect on muscle power; however, larger studies are required.

Current approaches on non-invasive prenatal diagnosis: Prenatal genomics, transcriptomics, personalized fetal diagnosis.

Recent developments in molecular genetics improved our knowledge on fetal genome and physiology. Novel scientific innovations in prenatal diagnosis have accelerated in the last decade changing our vision immensely. Data obtained from fetal genomic studies brought new insights to fetal medicine and by the advances in fetal DNA and RNA sequencing technology novel treatment strategies has evolved. Non-invasive prenatal diagnosis found ground in genetics and the results are widely studied in scientific arena. When Lo and colleges proved fetal genetic material can be extracted from maternal plasma and fetal DNA can be isolated from maternal serum, the gate to many exciting discoveries was open. Microarray technology and advances in sequencing helped fetal diagnosis as well as other areas of medicine. Today it is a very crucial prerequisite for physicians practicing prenatal diagnosis to have a profound knowledge in genetics. Prevailing practical use and application of fetal genomic tests in maternal and fetal medicine mandates obstetricians to update their knowledge in genetics. The purpose of this review is to assist physicians to understand and update their knowledge in fetal genetic testing from maternal blood, individualized prenatal counseling and advancements on the subject by sharing our experiences as Istanbul University Fetal Nucleic Acid Research Group.