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Menstrual flow as a non-invasive source of endometrial organoids.

ABSTRACT: Assessment of the endometrium often necessitates a biopsy, which currently involves an invasive, transcervical procedure. Here, we present an alternative technique based on deriving organoids from menstrual flow. We demonstrate that organoids can be derived from gland fragments recovered from menstrual flow. To confirm they faithfully reflect the in vivo state we compared organoids derived from paired scratch biopsies and ensuing menstrual flow from patients undergoing in vitro fertilisation (IVF). We demonstrate that the two sets of organoids share the same transcriptome signature, derivation efficiency and proliferation rate. Furthermore, they respond similarly to sex steroids and early-pregnancy hormones, with changes in morphology, receptor expression, and production of 'uterine milk' proteins that mimic those during the late-secretory phase and early pregnancy. This technique has wide-ranging impact for non-invasive investigation and personalised approaches to treatment of common gynaecological conditions, such as endometriosis, and reproductive disorders, including failed implantation after IVF and recurrent miscarriage.

SUBMITTER: Cindrova-Davies T

PROVIDER: S-EPMC8211845 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Menstrual flow as a non-invasive source of endometrial organoids

Project description:Organoid cultures derived from menstrual flow faithfully replicate secretory phase endometrial glands and provide a novel, non-invasive technique for the clinical assessment of endometrial function. Paired organoid cultures derived from scratch biopsies and ensuing menstrual flow share the same transcriptome signature, responses to early pregnancy hormones, and pathological changes in cases of early-pregnancy loss. The technique opens new avenues for investigating endometrial function in cases of subfertility and gynaecological disorders.

2021-04-13 | E-MTAB-9284 | biostudies-arrayexpress

Menstrual flow as a non-invasive source of endometrial organoids

Project description:Menstrual flow as a non-invasive source of endometrial organoids

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The Extracellular Vesicles Proteome of Endometrial Cells Simulating the Receptive Menstrual Phase Differs from That of Endometrial Cells Simulating the Non-Receptive Menstrual Phase.

Project description:Successful embryo implantation into a receptive endometrium requires mutual endometrial-embryo communication. Recently, the function of extracellular vehicles (EVs) in cell-to-cell interaction in embryo-maternal interactions has been investigated. We explored isolated endometrial-derived EVs, using RL95-2 cells as a model of a receptive endometrium, influenced by the menstrual cycle hormones estrogen (E2; proliferative phase), progesterone (P4; secretory phase), and estrogen plus progesterone (E2P4; the receptive phase). EV sized particles were isolated by differential centrifugation and size exclusion chromatography. Nanoparticle tracking analysis was used to examine the different concentrations and sizes of particles and EV proteomic analysis was performed using shotgun label-free mass spectrometry. Our results showed that although endometrial derived EVs were secreted in numbers independent of hormonal stimulation, EV sizes were statistically modified by it. Proteomics analysis showed that hormone treatment changes affect the endometrial EV's proteome, with proteins enhanced within the EV E2P4 group shown to be involved in different processes, such as embryo implantation, endometrial receptivity, and embryo development, supporting the concept of a communication system between the embryo and the maternal endometrium via EVs.

| S-EPMC9953153 | biostudies-literature

The Extracellular Vesicles Proteome of Endometrial Cells Simulating the Receptive Menstrual Phase Differs from That of Endometrial Cells Simulating the Non-Receptive Menstrual Phase

Project description:Successful embryo implantation into a receptive endometrium requires mutual endometrial-embryo communication. Recently, the function of extracellular vehicles (EVs) in cell-to-cell interaction in embryo-maternal interactions has been investigated. We explored isolated endometrial derived EVs, using RL95-2 cells as a model of a receptive endometrium, influenced by menstrual cycle hormones estrogen (E2; proliferative phase) progesterone (P4; secretory phase) and estrogen plus progesterone (E2P4; the receptive phase). EV sized particles were isolated by differential centrifugation and size exclusion chromatography. Nanoparticle tracking analysis was used to examine the different concentration and size of particles and EV proteomic analysis per-formed using shotgun label-free mass spectrometry. Our results showed that although endome-trial derived EVs were secreted in numbers independent of hormonal stimulation, EVs sizes were statistically modified by it. Proteomics analysis showed that hormone treatment changes affect the endometrial EVs proteome, with proteins enhanced within the EV E2P4 group shown to be in-volved in different processes such as embryo implantation, endometrial receptivity, and embryo development, supporting the concept of a communication system between the embryo and the maternal endometrium via EVs.

2023-02-13 | PXD039764 | Pride

Potential utility of a non-invasive menstrual blood immunophenotype analysis in reproductive medicine.

Project description:Purpose Can a comprehensive flow cytometry panel be used to assess immunophenotype profiles in menstrual blood of patients experiencing reproductive failure and age matched controls of proven fertility? Methods 58 recurrent pregnancy loss and repeated implantation failure patients, along with 15 age matched controls of proven fertility, had menstrual blood samples obtained within the first 24 hours of the onset of menstruation to non-invasively assess the local immunophenotype. Using a comprehensive multi-parameter flow panel the lymphocyte sub-populations were described and compared. Results Relative to well established peripheral blood immunophenotyping values, distinct lymphocyte population differences were noted between the subgroups. The ratios of CD4+ and CD8+ T-cells were inverted relative to peripheral blood and uterine NK cells represented by CD56bright were distinctly visualised, emphasising the distinction of menstrual and peripheral blood. Relative to controls there were marked increases in CD3+ve T-cells (p=0.009), CD4:CD8 ratio (p=0.004), CD19 B-cells (p=0.026) and CD56dim NK's (p=0.002) in the reproductive failure cases. Conclusions Flow cytometric evaluation can provide a rapid and objective analysis of lymphocyte subpopulations in many forms of tissue and fluid. The findings show significant variations in cellular composition of immune cells indicating a distinct compartment, with differences between cases and controls. Immunological assessment of the menstrual blood immunophenotype, in clinically appropriate patients, may provide insight into the aetiology of adverse reproductive outcome, without the risks and inconveniences associated with a more invasive endometrial biopsy.

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Non-Invasive Blood Flow Speed Measurement Using Optics.

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Progesterone Signaling in Endometrial Epithelial Organoids.

Project description:For pregnancy to be established, uterine cells respond to the ovarian hormones, estrogen, and progesterone, via their nuclear receptors, the estrogen receptor (ESR1) and progesterone receptor (PGR). ESR1 and PGR regulate genes by binding chromatin at genes and at distal enhancer regions, which interact via dynamic 3-dimensional chromatin structures. Endometrial epithelial cells are the initial site of embryo attachment and invasion, and thus understanding the processes that yield their receptive state is important. Here, we cultured and treated organoids derived from human epithelial cells, isolated from endometrial biopsies, with estrogen and progesterone and evaluated their transcriptional profiles, their PGR cistrome, and their chromatin conformation. Progesterone attenuated estrogen-dependent gene responses but otherwise minimally impacted the organoid transcriptome. PGR ChIPseq peaks were co-localized with previously described organoid ESR1 peaks, and most PGR and ESR1 peaks were in B (inactive) compartment regions of chromatin. Significantly more ESR1 peaks were assigned to estrogen-regulated genes by considering chromatin loops identified using HiC than were identified using ESR1 peak location relative to closest genes. Overall, the organoids model allowed a definition of the chromatin regulatory components governing hormone responsiveness.

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Biomarker discovery in pre-menstrual endometrial biopsies

Project description:The endometrium undergoes profound progesterone-driven remodeling during the secretory phase of the menstrual cycle in a process called decidualization. In the absence of pregnancy, circulating progesterone levels fall and tissue-wide inflammation and influx of neutrophils precede tissue breakdown and menstrual shedding. These changes are accompanied by wide-scale transcriptomic changes that co-ordinate temporal changes throughout the secretory phase. Here, we sequenced whole endometrial biopsies to identify molecular biomarkers that mark specific stages of the secretory phase, including pre-menstrual tissues. 20 biopsies were timed to specific phases of the secretory cycle based on the donor’s reported LH, the physical morphology of tissues observed via IHC and serum progesterone levels.

2024-10-27 | GSE246529 | GEO

Murine Endometrial Organoids to Model Chlamydia Infection.

Project description:The obligate intracellular bacterium Chlamydia trachomatis is the leading cause of bacterial sexually transmitted infections. Once internalized in host cells, C. trachomatis undergoes a biphasic developmental cycle within a membrane-bound compartment, known as the inclusion. Successful establishment of the intracellular niche relies on bacterial Type III effector proteins, such as Inc proteins. In vitro and in vivo systems have contributed to elucidating the intracellular lifestyle of C. trachomatis, but additional models combining the archetypal environment of infection with the advantages of in vitro systems are needed. Organoids are three-dimensional structures that recapitulate the microanatomy of an organ's epithelial layer, bridging the gap between in vitro and in vivo systems. Organoids are emerging as relevant model systems to study interactions between bacterial pathogens and their hosts. Here, we took advantage of recently developed murine endometrial organoids (EMOs) and present a C. trachomatis-murine EMO infection model system. Confocal microscopy of EMOs infected with fluorescent protein-expressing bacteria revealed that inclusions are formed within the cytosol of epithelial cells. Moreover, infection with a C. trachomatis strain that allows for the tracking of RB to EB transition indicated that the bacteria undergo a full developmental cycle, which was confirmed by harvesting infectious bacteria from infected EMOs. Finally, the inducible gene expression and cellular localization of a Chlamydia Inc protein within infected EMOs further demonstrated that this model is compatible with the study of Type III secreted effectors. Altogether, we describe a novel and relevant system for the study of Chlamydia-host interactions.

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Development of Polarity-Reversed Endometrial Epithelial Organoids.

Project description:The uterine epithelium is composed of a single layer of hormone responsive polarized epithelial cells that line the lumen and form tubular glands. Endometrial epithelial organoids (EEO) can be generated from uterine epithelia and recapitulate cell composition and hormone responses in vitro. As such, the development of EEO represents a major advance for facilitating mechanistic studies in vitro. However, a major limitation for the use of EEO cultured in basement membrane extract and other hydrogels is the inner location of apical membrane, thereby hindering direct access to the apical surface of the epithelium to study interactions with the embryo or infectious agents such as viruses and bacteria. Here, a straightforward strategy was developed that successfully reverses the polarity of EEO. The result is an apical-out organoid that preserves a distinct apical-basolateral orientation and remains responsive to ovarian steroid hormones. Our investigations highlight the utility of polarity-reversed EEO to study interactions with E. coli and blastocysts. This method of generating apical-out EEO lays the foundation for developing new in vitro functional assays, particularly regarding epithelial interactions with embryos during pregnancy or other luminal constituents in a pathological or diseased state.

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