Article

Mimicking the Events of Menstruation in the Murine Uterus

Department of Physiology and Pharmacology, The University of Western Ontario, London, Ontario, Canada
Biology of Reproduction (Impact Factor: 3.32). 11/2003; 69(4):1273-80. DOI: 10.1095/biolreprod.103.016550
Source: PubMed

ABSTRACT

Menstruation and endometrial regeneration occur during every normal reproductive cycle in women and some Old World primates. Many of the cellular and molecular events of menstruation have been identified by correlative or in vitro studies, but the lack of a convenient model for menstruation in a laboratory animal has restricted functional studies. In this study, a mouse model for menstruation first described by Finn in the 1980s has been modified for use in a commonly used inbred strain of mouse. A decidual stimulus was applied into the uterine lumen of appropriately primed mice and leukocyte numbers and apoptosis were examined over time following progesterone withdrawal. Endometrial tissue breakdown was initiated after 12-16 h, and by 24 h, the entire decidual zone had been shed. Re-epithelialization was nearly complete by 36 h and the endometrium was fully restored by 48 h. Leukocyte numbers increased significantly in the basal zone by 12 h after progesterone withdrawal, preceding stromal destruction. Stromal apoptosis was detected by TUNEL staining at 0 and 12 h but decreased by 16 h after progesterone withdrawal. This mouse model thus mimics many of the events of human menstruation and has the potential to assist in elucidation of the functional roles of a variety of factors thought to be important in both menstruation and endometrial repair.

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    • "This approach was used to label all endometrial cells with adenovirus encoding red fluorescent protein (Wang et al., 2014) or recombinant lentivirus conferring yellow fluorescent protein or click beetle red-emitting luciferase (Masuda et al., 2007b) for non-invasive monitoring of cell xenografts over several menstrual-like cycles. These animal models need further adapting to monitor labelled endometrial stem/progenitor cells, transplanted into the peritoneal cavity and subjected to a similar hormonal regime as the mouse model of menstruation (Brasted et al., 2003) for several cycles to examine the role of endometrial stem/progenitor cells in the initiation and progression of endometriosis. "
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    ABSTRACT: Background: The existence of stem/progenitor cells in the endometrium was postulated many years ago, but the first functional evidence was only published in 2004. The identification of rare epithelial and stromal populations of clonogenic cells in human endometrium has opened an active area of research on endometrial stem/progenitor cells in the subsequent 10 years. Methods: The published literature was searched using the PubMed database with the search terms 'endometrial stem cells and menstrual blood stem cells' until December 2014. Results: Endometrial epithelial stem/progenitor cells have been identified as clonogenic cells in human and as label-retaining or CD44(+) cells in mouse endometrium, but their characterization has been modest. In contrast, endometrial mesenchymal stem/stromal cells (MSCs) have been well characterized and show similar properties to bone marrow MSCs. Specific markers for their enrichment have been identified, CD146(+)PDGFRβ(+) (platelet-derived growth factor receptor beta) and SUSD2(+) (sushi domain containing-2), which detected their perivascular location and likely pericyte identity in endometrial basalis and functionalis vessels. Transcriptomics and secretomics of SUSD2(+) cells confirm their perivascular phenotype. Stromal fibroblasts cultured from endometrial tissue or menstrual blood also have some MSC characteristics and demonstrate broad multilineage differentiation potential for mesodermal, endodermal and ectodermal lineages, indicating their plasticity. Side population (SP) cells are a mixed population, although predominantly vascular cells, which exhibit adult stem cell properties, including tissue reconstitution. There is some evidence that bone marrow cells contribute a small population of endometrial epithelial and stromal cells. The discovery of specific markers for endometrial stem/progenitor cells has enabled the examination of their role in endometrial proliferative disorders, including endometriosis, adenomyosis and Asherman's syndrome. Endometrial MSCs (eMSCs) and menstrual blood stromal fibroblasts are an attractive source of MSCs for regenerative medicine because of their relative ease of acquisition with minimal morbidity. Their homologous and non-homologous use as autologous and allogeneic cells for therapeutic purposes is currently being assessed in preclinical animal models of pelvic organ prolapse and phase I/II clinical trials for cardiac failure. eMSCs and stromal fibroblasts also exhibit non-stem cell-associated immunomodulatory and anti-inflammatory properties, further emphasizing their desirable properties for cell-based therapies. Conclusions: Much has been learnt about endometrial stem/progenitor cells in the 10 years since their discovery, although several unresolved issues remain. These include rationalizing the terminology and diagnostic characteristics used for distinguishing perivascular stem/progenitor cells from stromal fibroblasts, which also have considerable differentiation potential. The hierarchical relationship between clonogenic epithelial progenitor cells, endometrial and decidual SP cells, CD146(+)PDGFR-β(+) and SUSD2(+) cells and menstrual blood stromal fibroblasts still needs to be resolved. Developing more genetic animal models for investigating the role of endometrial stem/progenitor cells in endometrial disorders is required, as well as elucidating which bone marrow cells contribute to endometrial tissue. Deep sequencing and epigenetic profiling of enriched populations of endometrial stem/progenitor cells and their differentiated progeny at the population and single-cell level will shed new light on the regulation and function of endometrial stem/progenitor cells.
    Preview · Article · Nov 2015 · Human Reproduction Update
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    • "In mammals such as humans and rodents, the estrous cycle prepares the female for reproduction. The estrous cycle is under control of sex steroids hormones (Brasted et al., 2003) but also under cytokines instruction (López and Stanley, 2005; Blitek et al., 2012). Cytokines are essential components of the complex communication network in the female reproductive tract during estrous cycle (Schaefer et al., 2005). "
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    ABSTRACT: The dynamic regulation of NF-κB activity in the uterus maintains a favorable environment of cytokines necessary to prepare for pregnancy throughout the estrous cycle. Recently, the mechanisms that directly regulate the NF-κB transcriptional activity in different tissues are of growing interest. IκBNS and BCL-3 are negative nuclear regulators of NF-κB activity that regulate IL-6 and TNF-α transcription, respectively. Both cytokines have been described as important factors in the remodeling of uterus for blastocyst implantation. In this work we analyzed in ICR mice the mRNA expression and protein production profile of IL-6, TNF-α, and their correspondent negative transcription regulators IκBNS or BCL-3 using real-time PCR, western blot and immunochemistry. We found that the expression of TNF-α and IL-6 was oscillatory along the estrous cycle, and its low expression coincided with the presence of BCL-3 and IκBNS, and vice versa, when the presence of the regulators was subtle, the expression of TNF-α and IL-6 was exacerbated. When we compared the production of TNF-α and IL-6 in the different estrous stages relating with diestrus we found that at estrus there is an important increase of the cytokines (p<0.05) decreasing at metestrus to reach the basal expression at diestrus. In the immunochemistry analysis we found that at diestrus BCL-3 is distributed all over the tissue with a barely detected TNF-α, but on the contrary, at estrus the expression of BCL-3 is not detected with TNF-α clearly observable along the tissue; the same phenomenon occur in the analysis of IκBNS and IL-6. With that evidence we suggest that the expression of TNF-α and IL-6 might be regulated through NF-κB nuclear regulators BCL-3 and IκBNS in the uterus of mice as has been demonstrated in other systems.
    Full-text · Article · Oct 2015 · Molecular Immunology
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    • "All animal procedures were carried out in accordance with UK legal requirements and in under licensed approval from the UK Home Office. In the current study a mouse model of menstruation described by Brasted et al [15] was modified to include non-surgical induction of decidualisation and a longer decidualisation period. Uterine tissues were also collected during a period of active shedding and repair, time-points that have not been previously described. "
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    ABSTRACT: In women dynamic changes in uterine tissue architecture occur during each menstrual cycle. Menses, characterised by the shedding of the upper functional layer of the endometrium, is the culmination of a cascade of irreversible changes in tissue function including stromal decidualisation, inflammation and production of degradative enzymes. The molecular mechanisms that contribute to the rapid restoration of tissue homeostasis at time of menses are poorly understood. A modified mouse model of menses was developed to focus on the events occurring within the uterine lining during endometrial shedding/repair. Decidualisation, vaginal bleeding, tissue architecture and cell proliferation were evaluated at 4, 8, 12, and 24 hours after progesterone (P4) withdrawal; mice received a single injection of bromodeoxyuridine (BrdU) 90 mins before culling. Expression of genes implicated in the regulation of mesenchymal to epithelial transition (MET) was determined using a RT2 PCR profiler array, qRTPCR and bioinformatic analysis. Mice exhibited vaginal bleeding between 4 and 12 hours after P4 withdrawal, concomitant with detachment of the decidualised cell mass from the basal portion of the endometrial lining. Immunostaining for BrdU and pan cytokeratin revealed evidence of epithelial cell proliferation and migration. Cells that appeared to be in transition from a mesenchymal to an epithelial cell identity were identified within the stromal compartment. Analysis of mRNAs encoding genes expressed exclusively in the epithelial or stromal compartments, or implicated in MET, revealed dynamic changes in expression, consistent with a role for reprogramming of mesenchymal cells so that they could contribute to re-epithelialisation. These studies have provided novel insights into the cellular processes that contribute to re-epithelialisation post-menses implicating both epithelial cell migration and mesenchymal cell differentiation in restoration of an intact epithelial cell layer. These insights may inform development of new therapies to induce rapid healing in the endometrium and other tissues and offer hope to women who suffer from heavy menstrual bleeding.
    Full-text · Article · Jan 2014 · PLoS ONE
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