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Evolutionary forward genomics reveals novel insights into the genes and pathways dysregulated in recurrent early pregnancy loss.

ABSTRACT: Are the genes that gained novel expression in the endometria of Eutherian (placental) mammals more likely to be dysregulated in patients with endometrial-associated recurrent early pregnancy loss (REPL)?There was a significant enrichment of genes dysregulated in REPL patients among the Eutherian-specific endometrial genes.Pregnancy loss is the most common complication of human pregnancy. REPL has multiple etiologies, including dysregulation of endometrial function, leading to 'suboptimal' implantation. Although the implantation process is tightly regulated in Eutherian (placental) mammals, the molecular factors contributing to dysregulated endometrial gene expression patterns in women with REPL are largely unknown.Endometrial biopsies were obtained from 32 REPL patients during the mid-luteal phase, and evaluated for glandular development arrest based on elevated nuclear cyclin E levels in gland cells, and for out-of-phase endometrial development based on histology. Gene expression levels were measured using Illumina Human HT-12v4 BeadChip arrays.Differentially expressed genes were identified between patients with (i) out-of-phase (n = 10) versus normal (n = 22) histological dating and (ii) abnormally elevated (n = 9) versus normal (n = 23) cyclin E levels in the nuclei of endometrial glands, using a likelihood ratio test. Enrichment of dysregulated genes in REPL endometria among Eutherian-specific genes was tested by permutation. Gene ontology and pathway enrichment analyses were carried out for the dysregulated genes.Fifty-eight and eighty-one genes were identified as differentially expressed at P < 0.001 in women with out-of-phase histological dating and abnormally elevated glandular cyclin E levels, respectively. Genes that were recruited into endometrial expression during the evolution of pregnancy in Eutherian mammals were significantly enriched for dysregulated genes (P = 0.002 for histology, P = 0.021 for cyclin E), as well as for genes involved in immune response and signaling pathways with essential roles in implantation and endometrial biology.Small sample size limits the statistical power to detect dysregulated genes, and the lack of non-REPL control women does not allow us to test for the contribution of these genes to overall risk of REPL.Enrichment of functional gene categories, as well as genes gained expression in the Eutherian endometria, help to identify molecular etiologies that contribute to normal functioning of the endometrium. These pathways are also strong candidates for successful pregnancy outcomes. Using the evolutionary history of mammalian gene expression in the endometrial tissue may be a promising approach to discover genes involved in female reproductive disorders.This work is supported by National Institutes of Health (NIH) grant R01 HD21244 to C.O. Authors declare no competing interests.

SUBMITTER: Kosova G

PROVIDER: S-EPMC4325674 | biostudies-literature | 2015 Mar

REPOSITORIES: biostudies-literature

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Evolutionary forward genomics reveals novel insights into the genes and pathways dysregulated in recurrent early pregnancy loss.

Kosova Gülüm G Stephenson Mary D MD Lynch Vincent J VJ Ober Carole C

Human reproduction (Oxford, England) 20150112 3

<h4>Study question</h4>Are the genes that gained novel expression in the endometria of Eutherian (placental) mammals more likely to be dysregulated in patients with endometrial-associated recurrent early pregnancy loss (REPL)?<h4>Summary answer</h4>There was a significant enrichment of genes dysregulated in REPL patients among the Eutherian-specific endometrial genes.<h4>What is known already</h4>Pregnancy loss is the most common complication of human pregnancy. REPL has multiple etiologies, inc ...[more]

PMID: 25586782

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Project description:Study hypothesisExome sequencing can identify genetic causes of idiopathic recurrent pregnancy loss (RPL).Study findingWe identified compound heterozygous deleterious mutations affecting DYNC2H1 and ALOX15 in two out of four families with RPL. Both genes have a role in early development. Bioinformatics analysis of all genes with rare and putatively pathogenic mutations in miscarriages and couples showed enrichment in pathways relevant to pregnancy loss, including the complement and coagulation cascades pathways.What is known alreadyNext generation sequencing (NGS) is increasingly being used to identify known and novel gene mutations in children with developmental delay and in fetuses with ultrasound-detected anomalies. In contrast, NGS is rarely used to study pregnancy loss. Chromosome microarray analysis detects putatively causative DNA copy number variants (CNVs) in ∼2% of miscarriages and CNVs of unknown significance (predominantly parental in origin) in up to 40% of miscarriages. Therefore, a large number of miscarriages still have an unknown cause.Study design, samples/materials, methodsWhole exome sequencing (WES) was performed using Illumina HiSeq 2000 platform on seven euploid miscarriages from four families with RPL. Golden Helix SVS v8.1.5 was used for data assessment and inheritance analysis for deleterious DNA variants predicted to severely disrupt protein-coding genes by introducing a frameshift, loss of the stop codon, gain of the stop codon, changes in splicing or the initial codon. Webgestalt (http://bioinfo.vanderbilt.edu/webgestalt/) was used for pathway and disease association enrichment analysis of a gene pool containing putatively pathogenic variants in miscarriages and couples in comparison to control gene pools.Main results and the role of chanceCompound heterozygous mutations in DYNC2H1 and ALOX15 were identified in miscarriages from two families with RPL. DYNC2H1 is involved in cilia biogenesis and has been associated with fetal lethality in humans. ALOX15 is expressed in placenta and its dysregulation has been associated with inflammation, placental, dysfunction, abnormal oxidative stress response and angiogenesis. The pool of putatively pathogenic single nucleotide variants (SNVs) and small insertions and deletions (indels) detected in the miscarriages showed enrichment in 'complement and coagulation cascades pathway', and 'ciliary motility disorders'. We conclude that CNVs, individual SNVs and pool of deleterious gene mutations identified by exome sequencing could contribute to RPL.Limitations, reasons for cautionThe size of our sample cohort is small. The functional effect of candidate mutations should be evaluated to determine whether the mutations are causative.Wider implications of the findingsThis is the first study to assess whether SNVs may contribute to the pathogenesis of miscarriage. Furthermore, our findings suggest that collective effect of mutations in relevant biological pathways could be implicated in RPL.Study funding and competing interestsThe study was funded by Canadian Institutes of Health Research (grant MOP 106467) and Michael Smith Foundation of Health Research Career Scholar salary award to ERS.

Project description:Pregnancy loss is the most common complication of human pregnancy. Recurrent early pregnancy loss (REPL) has multiple etiologies, including endometrial dysregulation leading to “suboptimal” implantation. Although the implantation process is tightly regulated in Eutherian (placental) mammals, the molecular factors contributing to dysregulated endometrial gene expression patterns in women with REPL are largely unknown. We hypothesized that genes that gained novel expression in the endometria of mammals that evolved in the Eutherian stemlineage, coincident with the evolution of pregnancy, are likely essential for establishment and maintenance of normal pregnancy and are, therefore, good candidates for genes whose expression may be dysregulated in disorders such as REPL. To test this hypothesis, we took an evolutionary forward genomics approach to characterize gene expression profiles of midsecretory endometria from women with REPL associated with abnormal endometria based either on histology or molecular expression of cyclin E. We identified 58 genes that were differentially expressed (P<0.001) between women with out-of-phase histological dating vs normal histology, and 81 genes that were differentially expressed (P<0.001) between women with abnormally elevated cyclin E levels vs normal cyclin E. Remarkably, genes that were recruited into endometrial expression during the evolution of pregnancy in Eutherian mammals were significantly enriched for dysregulated genes (P=0.002 for histology, P=0.021 for cyclin E), as well as for immune and signaling pathways with essential roles in endometrial biology. Thus, our novel evolutionary-based forward genomics approach identified genes whose dysregulation during the mid-secretory phase likely contributes to the molecular etiologies of recurrent early pregnancy loss. Total RNA obtained from mid luteal phase endometrium (two replicates per biopsy) from women with recurrent early pregnancy loss (REPL). Endometrial gene expression levels were compared 1) between women with out-of-phase (n=10) and normal histological dating (n=22), 2) between women with abnormally elevated (n=9) and normal (n=23) cyclin E levels. For 5 additional women with abnormally high cyclin E levels, biopsy samples were collected before and after progesterone treatment to investigate the gene expression profiles in response to progesterone.

Project description:Study questionWhat is the recommended management of women with recurrent pregnancy loss (RPL) based on the best available evidence in the literature?Summary answerThe guideline development group formulated 77 recommendations answering 18 key questions on investigations and treatments for RPL, and on how care should be organized.What is known alreadyA previous guideline for the investigation and medical treatment of recurrent miscarriage was published in 2006 and is in need of an update.Study design size durationThe guideline was developed according to the structured methodology for development of ESHRE guidelines. After formulation of key questions by a group of experts, literature searches and assessments were performed. Papers published up to 31 March 2017 and written in English were included. Cumulative live birth rate, live birth rate and pregnancy loss rate (or miscarriage rate) were considered the critical outcomes.Participants/materials setting methodsBased on the collected evidence, recommendations were formulated and discussed until consensus was reached within the guideline group. A stakeholder review was organized after finalization of the draft. The final version was approved by the guideline group and the ESHRE Executive Committee.Main results and the role of chanceThe guideline provides 38 recommendations on risk factors, prevention and investigations in couples with RPL, and 39 recommendations on treatments. These include 60 evidence-based recommendations - of which 31 were formulated as strong recommendations and 29 as conditional - and 17 good practice points. The evidence supporting investigations and treatment of couples with RPL is limited and of moderate quality. Of the evidence-based recommendations, only 10 (16.3%) were supported by moderate quality evidence. The remaining recommendations were supported by low (35 recommendations: 57.4%), or very low quality evidence (16 recommendations: 26.2%). There were no recommendations based on high quality evidence. Owing to the lack of evidence-based investigations and treatments in RPL care, the guideline also clearly mentions investigations and treatments that should not be used for couples with RPL.Limitations reasons for cautionSeveral investigations and treatments are offered to couples with RPL, but most of them are not well studied. For most of these investigations and treatments, a recommendation against the intervention or treatment was formulated based on insufficient evidence. Future studies may require these recommendations to be revised.Wider implications of the findingsThe guideline provides clinicians with clear advice on best practice in RPL, based on the best evidence available. In addition, a list of research recommendations is provided to stimulate further studies in RPL. One of the most important consequences of the limited evidence is the absence of evidence for a definition of RPL.Study funding/competing interestsThe guideline was developed and funded by ESHRE, covering expenses associated with the guideline meetings, with the literature searches and with the dissemination of the guideline. The guideline group members did not receive payment. J.E. reports position funding from CARE Fertility. S.L. reports position funding from SpermComet Ltd. S.M. reports research grants, consulting and speaker's fees from GSK, BMS/Pfizer, Sanquin, Aspen, Bayer and Daiichi Sankyo. S.Q. reports speaker's fees from Ferring. The other authors report no conflicts of interest.ESHRE Pages are not externally peer reviewed. This article has been approved by the Executive Committee of ESHRE.

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Evolutionary forward genomics reveals novel insights into the genes and pathways dysregulated in recurrent early pregnancy loss.

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Evolutionary forward genomics reveals novel insights into the genes and pathways dysregulated in recurrent early pregnancy loss.

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