Abstract
Natural variability in menstrual cycle length, coupled with rapid changes in endometrial gene expression, makes it difficult to accurately define and compare different stages of the ...endometrial cycle. Here we develop and validate a method for precisely determining endometrial cycle stage based on global gene expression. Our ‘molecular staging model’ reveals significant and remarkably synchronised daily changes in expression for over 3400 endometrial genes throughout the cycle, with the most dramatic changes occurring during the secretory phase. Our study significantly extends existing data on the endometrial transcriptome, and for the first time enables identification of differentially expressed endometrial genes with increasing age and different ethnicities. It also allows reinterpretation of all endometrial RNA-seq and array data that has been published to date. Our molecular staging model will significantly advance understanding of endometrial-related disorders that affect nearly all women at some stage of their lives, such as heavy menstrual bleeding, endometriosis, adenomyosis, and recurrent implantation failure.
Abstract
Study question
Are uterine natural killer (uNK) cell numbers and their distribution relative to endometrial arterioles altered in women with recurrent implantation failure (RIF) compared to ...women with embryo implantation success (IS)?
Summary answer
uNK cell numbers and their distribution relative to endometrial arterioles are not significantly different in women with RIF compared to women in whom embryo implantation occurs successfully following IVF.
What is already known
uNK cells are regulators of decidual angiogenesis and spiral arteriole remodelling during early pregnancy. Although some studies have shown that uNK cell numbers may be altered in women with RIF, the methods used to measure uNK cell numbers have proven inconsistent, making reproduction of these results difficult. It is unclear, therefore, whether the results reported so far are reproducible. Moreover, it is not known how uNK cell numbers may impact IVF outcomes. Despite the lack of conclusive evidence, uNK cell numbers are often evaluated as a prognostic criterion in women undergoing assisted reproductive procedures.
Study design, size, duration
Endometrial pipelle biopsies were collected 6–8 days post-LH surge in natural cycles from women with RIF (n = 14), women with IS (n = 11) and women with potential RIF at the time of the study (PRIF; n = 9) from 2013 to 2015.
Participants/materials, setting, methods
uNK cells (i.e. CD56+ and/or CD16+ phenotypes) and their distribution relative to endometrial arterioles were investigated by standard immunohistochemistry protocols and quantified using Aperio ScanScopeXT images digitized by ImageJ and deconvoluted into binary images for single cell quantification using a Gaussian Blur and Yen algorithm.
Main results and the role of chance
There was no significant difference in the cell density of CD56+ or CD16+ uNK cells in women with RIF compared to women with IS or PRIF. There was a higher proportion of uNK cells in the distal regions compared to the regions closest to the arterioles in all patient groups. Further, we identified a significant reduction in uNK cell density in women who had a previous pregnancy compared to those who had not, regardless of their current implantation status.
Large scale data
Not applicable.
Limitations, reasons for caution
Spiral arterioles could not always be accurately identified by digital image analysis; therefore, all endometrial arterioles were selected and analysed. Patient numbers for the study were low. However, as the clinical phenotypes of each patient were well defined, and endometrial dating was accurately determined by three independent pathologists, differences between patient groups with respect to the uNK numbers and distribution should have been measurable if uNK cell counts were to be useful as a prognostic marker of RIF.
Wider implications of the findings
Our findings demonstrate that CD56+ and CD16+ uNK cell numbers are not significantly different in women with RIF in a typical cohort of women undergoing IVF. Further, prior pregnancy was associated with a significantly reduced number of uNK cells in both the RIF and IS patient groups, suggestive of a long-term pregnancy induced suppression of uNK cells. Combined, these findings do not support the clinical value of using uNK cell numbers as a prognostic indicator of implantation success with IVF treatment.
Study funding/competing interest(s)
Funding for this work was provided by Royal Women’s Hospital Foundation. P.P. was supported by an NHMRC Early Career Fellowship TF 11/14 and W.T.T. was supported by an NHMRC Postgraduate Scholarship 1055814. The authors do not have any competing interests with this study.
Do menstrual cycle-dependent changes occur in the histological appearance of superficial peritoneal endometriotic lesions, and are they equivalent to those observed in the eutopic endometrium?
Only a ...small subset of superficial peritoneal endometriotic lesions exhibits some histological features in phase with menstrual cycle-related changes observed in eutopic endometrium.
Endometriotic lesions are frequently described as implants that follow menstrual cycle-related changes in morphology, as per the eutopic endometrium. This concept has been widely accepted despite the lack of conclusive published evidence.
This was a retrospective cohort study of 42 patients, from across the menstrual cycle, with surgically and histologically confirmed endometriosis. Patients were a subset selected from a larger endometriosis study being conducted at the Royal Women's Hospital, Melbourne since 2012.
Histological features of epithelium, stroma and gland morphology were examined in haematoxylin and eosin stained sections of superficial peritoneal endometriotic lesions and matched eutopic endometrium (menstrual: n = 4, proliferative: n = 11, secretory: n = 17, hormone-treated: n = 10). At least two biopsies (average = 4, range = 2-8 biopsies) and a matched endometrial sample were analysed for each patient and results were presented per endometriotic gland profile (n = 1051). Data were analysed using mixed effects logistic regression to account for multiple patients and multiple endometriotic biopsies, each with multiple endometriotic gland profiles. This model also enabled analysis of endometriotic lesions versus eutopic endometrium.
There was considerable inter- and intra-patient variability in the morphology of superficial peritoneal endometriotic lesions. Menstrual cycle-associated changes were only observed for some features in a subset of endometriotic gland profiles. The proportion of endometriotic gland profiles with epithelial mitoses significantly increased in the proliferative phase (18% of gland profiles) relative to the menstrual phase (0% of endometriotic gland profiles) (odds ratios (OR) 9.30; 95% confidence intervals (CI) = 3.71-23.32; P < 0.001). Fewer blood-filled gland lumens were observed in the secretory phase (45% of endometriotic gland profiles) compared to the menstrual phase (67% of endometriotic gland profiles) (OR, 0.30; 95% CI = 0.11-0.79; P = 0.015). The features of the eutopic endometrium analysed in this study did not reflect the results in matched endometriotic lesions (P > 0.05).
Not applicable.
This study focused on features observed in sections of superficial peritoneal lesions and these may differ from features of deep infiltrating endometriosis or ovarian endometriomas. Cycle phases were limited to menstrual, proliferative and secretory phases to allow appropriate statistical modelling.
This study highlights heterogeneity in the histological characteristics of superficial peritoneal lesions. It challenges the assumption that lesion morphology consistently reflects menstrual cycle-associated changes.
Research reported in this publication was supported in part by National Health and Medical Research Council (NHMRC) project grants GNT1012245, GNT1105321 and GNT1026033 (P.A.W.R., J.E.G. and S.J.H.-C.). There are no competing interests.
The anatomy and biomechanics of the volar plate are reviewed. From a study of 60 fresh adult specimens, the authors present a consolidated anatomic concept of this structure, emphasizing the ...distinctiveness of the proximal attachments, the variable nature of the distal attachments, and the complex relationships to the flexor sheaths, the vincula, and the collateral ligaments. Experimental biomechanical data suggest that the nature and site of injury to this structure is dependent on the rate of application of force: rapid rates produce rupture at the distal attachment, slow rates attenuate the proximal check ligaments.
The discovery of somatic mutations in the isocitrate dehydrogenase (IDH) enzymes through a genome-wide mutational analysis in glioblastoma represents a milestone event in cancer biology. The nature ...of the heterozygous, point mutations mapping to arginine residues involved in the substrate binding inspired several research teams to investigate their impact on the biochemical activity of these enzymes. Soon, it became clear that the mutations identified impaired the ability of IDH1 and IDH2 to catalyze the conversion of isocitrate to α-ketoglutarate (αKG), whereas conferring a gain of a novel enzymatic activity leading to the reduction of αKG to the metabolite D2-hydroxyglutarate (D-2HG). Across glioma as well as several hematologic malignancies, mutations in IDH1 and IDH2 have shown prognostic value. Several hypotheses implicating the elevated levels of D-2HG and tumorigenesis, and the therapeutic potential of targeting mutant IDH enzymes will be discussed.
Chronic exposure to stress is associated with increased incidence of depression, generalized anxiety, and PTSD. However, stress induces vulnerability to such disorders only in a sub-population of ...individuals, as others remain resilient. Inflammation has emerged as a putative mechanism for promoting stress vulnerability. Using a rodent model of social defeat, we have previously shown that rats with short-defeat latencies (SL/vulnerable rats) show increased anxiety- and depression-like behaviors, and these behaviors are mediated by inflammation in the ventral hippocampus. The other half of socially defeated rats show long-latencies to defeat (LL/resilient) and are similar to controls. Because gut microbiota are important activators of inflammatory substances, we assessed the role of the gut microbiome in mediating vulnerability to repeated social defeat stress. We analyzed the fecal microbiome of control, SL/vulnerable, and LL/resilient rats using shotgun metagenome sequencing and observed increased expression of immune-modulating microbiota, such as Clostridia, in SL/vulnerable rats. We then tested the importance of gut microbiota to the SL/vulnerable phenotype. In otherwise naive rats treated with microbiota from SL/vulnerable rats, there was higher microglial density and IL-1β expression in the vHPC, and higher depression-like behaviors relative to rats that received microbiota from LL/resilient rats, non-stressed control rats, or vehicle-treated rats. However, anxiety-like behavior during social interaction was not altered by transplant of the microbiome of SL/vulnerable rats into non-stressed rats. Taken together, the results suggest the gut microbiome contributes to the depression-like behavior and inflammatory processes in the vHPC of stress vulnerable individuals.
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