Disrupted PGR-B and ESR1 signaling underlies defective decidualization linked to severe preeclampsia
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Curated by eLife
Evaluation Summary:
Garrido-Gomez et al generate a unique dataset profiling the transcriptomes late-stage endometrium of women with prior pregnancies leading to severe preeclampsia (sPE) compared to that of non-preeclamptic pregnancies. Although the question of molecular drivers preeclampsia has been explored at the molecular level directly in the placenta, this study provides a unique view in the endometrium of women years after the pregnancy in question, which is consistent with the fact that a previous preeclamptic pregnancy is one of the best predictors for a subsequent preeclamptic pregnancy.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. The reviewers remained anonymous to the authors.)
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Abstract
Decidualization of the uterine mucosa drives the maternal adaptation to invasion by the placenta. Appropriate depth of placental invasion is needed to support a healthy pregnancy; shallow invasion is associated with the development of severe preeclampsia (sPE). Maternal contribution to sPE through failed decidualization is an important determinant of placental phenotype. However, the molecular mechanism underlying the in vivo defect linking decidualization to sPE is unknown.
Methods:
Global RNA sequencing was applied to obtain the transcriptomic profile of endometrial biopsies collected from nonpregnant women who suffer sPE in a previous pregnancy and women who did not develop this condition. Samples were randomized in two cohorts, the training and the test set, to identify the fingerprinting encoding defective decidualization in sPE and its subsequent validation. Gene Ontology enrichment and an interaction network were performed to deepen in pathways impaired by genetic dysregulation in sPE. Finally, the main modulators of decidualization, estrogen receptor 1 ( ESR1 ) and progesterone receptor B ( PGR-B ), were assessed at the level of gene expression and protein abundance.
Results:
Here, we discover the footprint encoding this decidualization defect comprising 120 genes—using global gene expression profiling in decidua from women who developed sPE in a previous pregnancy. This signature allowed us to effectively segregate samples into sPE and control groups. ESR1 and PGR were highly interconnected with the dynamic network of the defective decidualization fingerprint. ESR1 and PGR-B gene expression and protein abundance were remarkably disrupted in sPE.
Conclusions:
Thus, the transcriptomic signature of impaired decidualization implicates dysregulated hormonal signaling in the decidual endometria in women who developed sPE. These findings reveal a potential footprint that could be leveraged for a preconception or early prenatal screening of sPE risk, thus improving prevention and early treatments.
Funding:
This work has been supported by the grant PI19/01659 (MCIU/AEI/FEDER, UE) from the Spanish Carlos III Institute awarded to TGG. NCM was supported by the PhD program FDGENT/2019/008 from the Spanish Generalitat Valenciana. IMB was supported by the PhD program PRE2019-090770 and funding was provided by the grant RTI2018-094946-B-100 (MCIU/AEI/FEDER, UE) from the Spanish Ministry of Science and Innovation with CS as principal investigator. This research was funded partially by Igenomix S.L.
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Author Response:
Reviewer #1 (Public Review):
The authors generate a unique dataset profiling the transcriptomes of late-stage secretory endometrium from women with past pregnancies confirmed to be (i) with sPE or (ii) without sPE, divided into pre-term and full-term pregnancies. They divide the dataset into training and testing samples in order to identify a signature of sPE. Intriguingly, they observe long lasting anomalies in the transcriptomes of endometria from women with prior sPE, which indicates that elements of PE pathogenesis survive the end of the pregnancy. This observation is consistent with the fact that a previous preeclamptic pregnancy is one of the best predictors for a subsequent preeclamptic pregnancies. The authors then perform functional enrichment analysis on the signature, which appears to be related to …
Author Response:
Reviewer #1 (Public Review):
The authors generate a unique dataset profiling the transcriptomes of late-stage secretory endometrium from women with past pregnancies confirmed to be (i) with sPE or (ii) without sPE, divided into pre-term and full-term pregnancies. They divide the dataset into training and testing samples in order to identify a signature of sPE. Intriguingly, they observe long lasting anomalies in the transcriptomes of endometria from women with prior sPE, which indicates that elements of PE pathogenesis survive the end of the pregnancy. This observation is consistent with the fact that a previous preeclamptic pregnancy is one of the best predictors for a subsequent preeclamptic pregnancies. The authors then perform functional enrichment analysis on the signature, which appears to be related to hormonal signaling (i.e. estrogen and progesterone signaling).
Although this study presents a seemingly well-controlled dataset (with stratified controls at pre-term and full-term) and on a rarely assessed tissue (post-pregnancy endometrium), there are a number of concerns on the processing of the data and the statistical methods employed (for instance the use of fold-change thresholding, which is known to lead to extremely poor FDR control in standard processing pipelines). There is also not enough information about how some analyses were performed for full transparency and reproducibility of studies in the long-term.
We would like to thank the reviewer for the feedback on our manuscript and the recommendations formulated to improve it. Regarding methodology, we supply more details to provide full transparency and reproducibility. Specifically, the statistical methods applied to obtain differentially expressed genes have been clarified to enable full understanding of the use of fold change thresholding and FDR cutoff.
Reviewer #2 (Public Review):
Garrido-Gomez et al. perform RNAseq on human endometrial biopsies from women with a history of sPE and women who never had sPE, to better define the signature of decidualization that may lead to sPE. The authors report on 166 differentially expressed genes in sPE compared to control, and many of which are connected to hormonal signaling through progesterone receptor-B and estrogen receptor 1 pathways. Therefore, changes in these genes could connect some of the dots between decidualization impairment and the development of sPE during pregnancy. The strengths of this manuscript include the use of human endometrial biopsies, a training and validation set of samples and the rigorous analyses to define the decidualization disorder genotype/phenotype of sPE. While this is a mostly descriptive study, the conclusions are mostly well supported by the data; however, some aspects of the patient population need to be further explained.
The clinical definition of sPE and whether it was early or late onset is not included anywhere in the manuscript. Please clearly define these variables and discuss how these pathways may impact the timing of disease onset.
Thank you for this observation that let us improve the clarity of the manuscript. Following this recommendation, we included the clinical definition of sPE in the “Study design” section of the Materials and Methods so that the definition is not just in the Introduction (Lines 35–37). Regarding preeclampsia classification, our work is focused on severe preeclampsia (sPE) due to the severity of its symptoms and health risks. Early onset preeclampsia (EOPE) tends to be more severe than late onset, but late onset preeclampsia could also be severe. Furthermore, all sPE cases included in the study were associated with prematurity (gestational week of <37) regardless of early or late onset. An increasing number of studies are focused on this classification, but severity is present in the clinical practice (PMID: 32443079). However, the discussion proposed is interesting, and we take it into consideration for future studies.
The conclusion (and title) that these gene signatures could be useful for preconception or early prenatal screening is overreaching since all these biopsies were collected from women who already experienced sPE and not from women who had yet to be diagnosed with PE. As risks of many diseases (i.e. cardiovascular and metabolic disease) increase in women with a history of PE, sPE itself might have a long-lasting impact on the endometrial environment, independent from decidualization.
There is increasing evidence to support that inappropriate endometrial maturation before pregnancy may contribute to reproductive disorders (PMID: 32521725). In addition, the role of defective decidualization in the origin of sPE was recently described (PMID: 33007270). It is worth highlighting recent work demonstrating a connection between decidual immaturity and shallow extravillous trophoblast invasion in preeclampsia (PMID: 25421975; PMID: 31356122). These studies identified DEGs in decidual tissue obtained in a chorionic villous sampling from women at ~11.5 gestational weeks who developed sPE symptoms 6 months later compared with normal pregnancies. Forty percent of these DEGs overlapped with DEGs associated with various stages of normal endometrial maturation before and after implantation, as identified by other data sets. These results reinforce the concept that a decidualization defect during the secretory phase and early pregnancy preceded the development of preeclampsia. Recently, it has been further demonstrated share molecular pathways of defective decidualization in preeclampsia and endometrial disorders, such as implantation failure, recurrent miscarriage, and endometriosis.
In this context, we demonstrated [Garrido-Gómez T. et al. (PMID: 28923940)] that human endometrial stromal cells isolated from women who experienced sPE in a previous pregnancy failed to decidualize in vitro. Additionally, decidua from tissue sections of the maternal–fetal interface at delivery in sPE had transcriptomic alterations, isolated decidual cells failed to redecidualize in culture, and conditioned medium from these cells failed to support cytotrophoblast (CTB) invasion.
Our findings reinforce a maternal cause for sPE through defective decidualization, and therefore is an important step toward the development of new strategies that will enable early assessment of women’s risk of experiencing sPE and would open the door to possible new therapeutic interventions to treat this enigmatic condition. Ideally, translational efforts could be targeted to noninvasive monitoring of maternal, placental, and fetal dynamics during pregnancy, which was recently proposed by Munchel et al. (PMID: 32611681). Following the reviewer’s recommendation, the conclusion and the title have been qualified regarding preconception and early pregnancy prenatal screening.
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Evaluation Summary:
Garrido-Gomez et al generate a unique dataset profiling the transcriptomes late-stage endometrium of women with prior pregnancies leading to severe preeclampsia (sPE) compared to that of non-preeclamptic pregnancies. Although the question of molecular drivers preeclampsia has been explored at the molecular level directly in the placenta, this study provides a unique view in the endometrium of women years after the pregnancy in question, which is consistent with the fact that a previous preeclamptic pregnancy is one of the best predictors for a subsequent preeclamptic pregnancy.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. The reviewers remained anonymous to the authors.)
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Reviewer #1 (Public Review):
The authors generate a unique dataset profiling the transcriptomes of late-stage secretory endometrium from women with past pregnancies confirmed to be (i) with sPE or (ii) without sPE, divided into pre-term and full-term pregnancies. They divide the dataset into training and testing samples in order to identify a signature of sPE. Intriguingly, they observe long lasting anomalies in the transcriptomes of endometria from women with prior sPE, which indicates that elements of PE pathogenesis survive the end of the pregnancy. This observation is consistent with the fact that a previous preeclamptic pregnancy is one of the best predictors for a subsequent preeclamptic pregnancies. The authors then perform functional enrichment analysis on the signature, which appears to be related to hormonal signaling (i.e. …
Reviewer #1 (Public Review):
The authors generate a unique dataset profiling the transcriptomes of late-stage secretory endometrium from women with past pregnancies confirmed to be (i) with sPE or (ii) without sPE, divided into pre-term and full-term pregnancies. They divide the dataset into training and testing samples in order to identify a signature of sPE. Intriguingly, they observe long lasting anomalies in the transcriptomes of endometria from women with prior sPE, which indicates that elements of PE pathogenesis survive the end of the pregnancy. This observation is consistent with the fact that a previous preeclamptic pregnancy is one of the best predictors for a subsequent preeclamptic pregnancies. The authors then perform functional enrichment analysis on the signature, which appears to be related to hormonal signaling (i.e. estrogen and progesterone signaling).
Although this study presents a seemingly well-controlled dataset (with stratified controls at pre-term and full-term) and on a rarely assessed tissue (post-pregnancy endometrium), there are a number of concerns on the processing of the data and the statistical methods employed (for instance the use of fold-change thresholding, which is known to lead to extremely poor FDR control in standard processing pipelines). There is also not enough information about how some analyses were performed for full transparency and reproducibility of studies in the long-term. -
Reviewer #2 (Public Review):
Garrido-Gomez et al. perform RNAseq on human endometrial biopsies from women with a history of sPE and women who never had sPE, to better define the signature of decidualization that may lead to sPE. The authors report on 166 differentially expressed genes in sPE compared to control, and many of which are connected to hormonal signaling through progesterone receptor-B and estrogen receptor 1 pathways. Therefore, changes in these genes could connect some of the dots between decidualization impairment and the development of sPE during pregnancy. The strengths of this manuscript include the use of human endometrial biopsies, a training and validation set of samples and the rigorous analyses to define the decidualization disorder genotype/phenotype of sPE. While this is a mostly descriptive study, the …
Reviewer #2 (Public Review):
Garrido-Gomez et al. perform RNAseq on human endometrial biopsies from women with a history of sPE and women who never had sPE, to better define the signature of decidualization that may lead to sPE. The authors report on 166 differentially expressed genes in sPE compared to control, and many of which are connected to hormonal signaling through progesterone receptor-B and estrogen receptor 1 pathways. Therefore, changes in these genes could connect some of the dots between decidualization impairment and the development of sPE during pregnancy. The strengths of this manuscript include the use of human endometrial biopsies, a training and validation set of samples and the rigorous analyses to define the decidualization disorder genotype/phenotype of sPE. While this is a mostly descriptive study, the conclusions are mostly well supported by the data; however, some aspects of the patient population need to be further explained.
The clinical definition of sPE and whether it was early or late onset is not included anywhere in the manuscript. Please clearly define these variables and discuss how these pathways may impact the timing of disease onset.
The conclusion (and title) that these gene signatures could be useful for preconception or early prenatal screening is overreaching since all these biopsies were collected from women who already experienced sPE and not from women who had yet to be diagnosed with PE. As risks of many diseases (i.e. cardiovascular and metabolic disease) increase in women with a history of PE, sPE itself might have a long-lasting impact on the endometrial environment, independent from decidualization.
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