The Essential Role of PGF2α/PTGFR in Molding Endometrial Breakdown and Vascular Dynamics, Regulated by HIF-1α in a Mouse Menstrual-like Model.

In women of childbearing age, extensive decidualization, shedding and remodeling of the endometrium during the menstrual cycle are fundamental for successful pregnancy. The role of prostaglandins (PGs) in menstruation has long been proposed in humans, and the rate-limiting enzyme cyclooxygenase was shown to play a key role in endometrial breakdown and shedding in a mouse menstrual-like model in our previous study. However, the specific types of PGs involved and their respective roles remain unclear. Therefore, our objective was to investigate the mechanism through which PGs regulate endometrial disintegration. In this study, the microscopy was observed by HE; the protein levels of prostaglandins E1 (PGE1), prostaglandins E2 (PGE2), prostaglandin F2α (PGF2α) and Prostaglandin I2 (PGI2) were detected by ELISA; the mRNA level of Pfgfr2, Vascular Endothelial Growth Factor(Vegf), Angiostatin and Hypoxia inducible factor-1α (Hif1α) were examined by real-time PCR; PTGFR Receptor (PTGFR), VEGF, Angiostatin and HIF-1α protein levels were investigated by western blotting; the locations of protein were observed by Immunohistochemistry; HIF-1α binding PTGFR promoter was detected by Chromatin Immunoprecipitation (ChIP) and real-time PCR. We found that the concentrations of PGE1, PGE2, and PGF2α all increased significantly during this process. Furthermore, Ptgfr mRNA increased soon after Progesterone (P4) withdrawal, and PTGFR protein levels increased significantly during abundant endometrial breakdown and shedding processes. PTGFR inhibitors AL8810 significantly suppressed endometrial breakdown and shedding, promoted Angiostatin expression, and reduced VEGF-A expressions and vascular permeability. And HIF-1α and PTGFR were mainly located in the luminal/gland epithelium, vascular endothelium, and pre-decidual zone. Interestingly, HIF-1α directly bound to Ptgfr promoter. Moreover, a HIF-1α inhibitor 2-methoxyestradiol (2ME) significantly reduced PTGFR expression and suppressed endometrial breakdown which was in accord with PTGFR inhibitor's effect. Similar changes occurred in human stromal cells relevant to menstruation in vitro. Our study provides evidence that PGF2α/PTGFR plays a vital role in endometrial breakdown via vascular changes that are regulated by HIF-1α during menstruation.

HGF Secreted by Menstrual Blood-Derived Endometrial Stem Cells Ameliorates Non-Alcoholic Fatty Liver Disease Through Downregulation of Hepatic Rnf186.

Mesenchymal stem cells (MSCs) have been demonstrated to protect against fatty liver diseases, but the mechanism is still not clear. Menstrual blood-derived endometrial stem cells (MenSCs) are a substantial population of MSCs that can be obtained in a noninvasive manner. In the present study, we investigated the therapeutic effects and underlying mechanisms of MenSC transplantation in mouse models of diet-induced nonalcoholic fatty liver disease (NAFLD). The results revealed that MenSCs markedly promoted hepatic glycogen storage and attenuated lipid accumulation after transplantation. We further identified Rnf186 as a novel regulator involved in MenSC-based therapy for NAFLD mice. Rnf186 deficiency substantially inhibited high-fat diet-induced insulin resistance and abnormal hepatic glucose and lipid metabolism in mice. Mechanistically, Rnf186 regulated glucose and lipid metabolism through the AMPK-mTOR pathway. More importantly, hepatocyte growth factor (HGF) is identified as the key functional cytokine secreted by MenSCs and decreases the expression of hepatic Rnf186. HGF deficient MenSCs cannot attenuate glucose and lipid accumulation after transplantation in NAFLD mice. Collectively, our results provide preliminary evidence for the protective roles of HGF secreted by MenSCs in fatty liver diseases through downregulation of hepatic Rnf186 and suggest that MenSCs or Rnf186 may be an alternative therapeutic approach/target for the treatment of NAFLD.

Single-cell RNA sequencing and lineage tracing confirm mesenchyme to epithelial transformation (MET) contributes to repair of the endometrium at menstruation.

The human endometrium experiences repetitive cycles of tissue wounding characterised by piecemeal shedding of the surface epithelium and rapid restoration of tissue homeostasis. In this study, we used a mouse model of endometrial repair and three transgenic lines of mice to investigate whether epithelial cells that become incorporated into the newly formed luminal epithelium have their origins in one or more of the mesenchymal cell types present in the stromal compartment of the endometrium. Using scRNAseq, we identified a novel population of PDGFRb + mesenchymal stromal cells that developed a unique transcriptomic signature in response to endometrial breakdown/repair. These cells expressed genes usually considered specific to epithelial cells and in silico trajectory analysis suggested they were stromal fibroblasts in transition to becoming epithelial cells. To confirm our hypothesis we used a lineage tracing strategy to compare the fate of stromal fibroblasts (PDGFRa+) and stromal perivascular cells (NG2/CSPG4+). We demonstrated that stromal fibroblasts can undergo a mesenchyme to epithelial transformation and become incorporated into the re-epithelialised luminal surface of the repaired tissue. This study is the first to discover a novel population of wound-responsive, plastic endometrial stromal fibroblasts that contribute to the rapid restoration of an intact luminal epithelium during endometrial repair. These findings form a platform for comparisons both to endometrial pathologies which involve a fibrotic response (Asherman's syndrome, endometriosis) as well as other mucosal tissues which have a variable response to wounding.

The human uterus is a formidable organ. From puberty to menopause, it completely sheds off its internal lining every 28 days or so, creating what is in effect a large open wound. Unlike the skin or other parts of the body, however, this tissue can quickly repair itself without scarring. This fascinating process remains poorly understood, partly because human samples and animal models that mimic human menstruation are still lacking. This makes it difficult to grasp how various types of uterine cells get mobilised for healing. To fill this gap, Kirkwood et al. focused on fibroblasts, a heterogenous cell population which helps to support the epithelial cells lining the inside of the uterus. How these cells responded to the advent of menstruation was examined in female mice genetically manipulated to have human-like periods. A method known as single-cell RNAseq was used to track which genes were active in each of these cells before, one day and two days after period onset. This revealed the existence of a subpopulation of cells which only appeared when wound healing was most needed. These 'repair-specific' fibroblasts expressed a mixture of genes; those typical of fibroblasts but also some known to be active in the epithelial cells lining the uterus. This suggests that the cells were in the process of changing their identity so they could remake the uterine layer lost during a period. And indeed, labelling these fibroblasts with a fluorescent tag showed that, during healing, they had migrated from within the uterine tissue to become part of its newly restored internal surface. These results represent the first evidence that fibroblasts play a direct role in repairing the uterus during menstruation. From endometriosis to infertility, the lives of millions of people around the world are impacted by disorders which affect the uterine lining. A better understanding of how the uterus can fix itself month after month could help to find new treatments for these conditions. This knowledge could also be useful for to address abnormal wound healing in the skin and other tissues, as this process often involves fibroblasts.

Endometrial macrophages in health and disease.

Macrophages are present in the endometrium throughout the menstrual cycle and are most abundant during menstruation. Endometrial macrophages contribute to tissue remodeling during establishment of pregnancy and are thought to play key roles in mediating tissue breakdown and repair during menstruation. Despite these important roles, the phenotype and function of endometrial macrophages remains poorly understood. In this review, we summarize approaches used to characterize endometrial macrophage phenotype, current understanding of the functional role of macrophages in normal endometrial physiology as well as the putative contribution of macrophage dysfunction to women's reproductive health disorders.

A Comparative Study of Gene Expression in Menstrual Blood-Derived Stromal Cells between Endometriosis and Healthy Women.

BACKGROUND: Research into the pathogenesis of endometriosis would substantially promote its effective treatment and early diagnosis. Currently, accumulating evidence has shed light on the importance of endometrial stem cells within the menstrual blood which are involved in the establishment and progression of endometriotic lesions in a retrograde manner. OBJECTIVES: We aimed to identify the differences in some genes' expression between menstrual blood-derived mesenchymal stem cells (MenSCs) isolated from endometriosis patients (E-MenSCs) and MenSCs from healthy women (NE-MenSCs). METHODS: Menstrual blood samples (2-3 mL) from healthy and endometriosis women in the age range of 22-35 years were collected. Isolated MenSCs by the Ficoll-Paque density-gradient centrifugation method were characterized by flow cytometry. MenSCs were evaluated for key related endometriosis genes by real-time-PCR. RESULTS: E-MenSCs were morphologically different from NE-MenSCs and showed, respectively, higher and lower expression of CD10 and CD9. Furthermore, E-MenSCs had higher expression of Cyclin D1 (a cell cycle-related gene) and MMP-2 and MMP-9 (migration- and invasion-related genes) genes compared with NE-MenSCs. Despite higher cell proliferation in E-MenSCs, the BAX/BCL-2 ratio was significantly lower in E-MenSCs compared to NE-MenSCs. Also, the level of inflammatory genes such as IL1ß, IL6, IL8, and NF-κB and stemness genes including SOX2 and SALL4 was increased in E-MenSCs compared with NE-MenSCs. Further, VEGF, as a potent angiogenic factor, showed a significant increase in E-MenSCs rather than NE-MenSCs. However, NE-MenSCs showed increased ER-α and ß-catenin when compared with E-MenSCs. CONCLUSION: Here, we showed that there are gene expression differences between E-MenSCs and NE-MenSCs. These findings propose that MenSCs could play key role in the pathogenesis of endometriosis and further support the menstrual blood retrograde theory of endometriosis formation. This could be of great importance in exploiting promising therapeutic targets and new biomarkers for endometriosis treatment and prognosis.

Effect of estrogen/progesterone ratio on the differentiation and the barrier function of epidermal keratinocyte and three-dimensional cultured human epidermis.

AIMS: Estrogen (E) and progesterone (P) are the major female hormones and are secreted with changing concentration ratios throughout the menstrual cycle. These hormones have been studied individually regarding their physiological function in the skin, but their concentration ratio (E/P) and its effect on the skin has not yet been investigated. The purpose of this study was to clarify the effect of the E/P ratio on skin barrier function. The menstrual cycle was divided into the menstrual, follicular, ovulation, and luteal phases. MATERIALS AND METHODS: The E/P concentration ratios corresponding with each phase were added to a three-dimensional epidermal model or normal human epidermal keratinocytes for 5 days. Gene and protein expression levels of several markers of cell differentiation, including loricrin (LOR) and transglutaminase (TGase), were quantified by real-time PCR and western blotting, respectively. Transepidermal water loss (TEWL) of the three-dimensional epidermal model was measured, and ceramide content was quantified by thin-layer chromatography. KEY FINDINGS: Gene expression of the epidermal differentiation markers, LOR and TGase, increased when applying the concentration ratio of E/P associated with the menstrual and luteal phases. The LOR protein level decreased from menstrual to luteal phases, and the TGase protein level increased from menstrual to luteal phases. During the same phases, ceramide NS increased and TEWL decreased. SIGNIFICANCE: Skin barrier function was improved by culturing cells at specific E/P concentration ratios, which would, therefore, be considered beneficial for female skin. This suggests that dysregulated E/P concentration ratios may be the cause of certain skin problems.

Prevalence, impact, and management perception of dysmenorrhea among university students: A cross-sectional study

Abstract Dysmenorrhea is a common condition among females that is characterized by painful cramps before or during menstruation. It is considered as a common gynecological complaint that affects the quality of women's life. The study evaluated prevalence of dysmenorrhea, its impact, associated risk factors, and the management strategies adopted by female university students in Taif city, Saudi Arabia. A cross-sectional study was conducted among 562 female students aged 18-30 years at the university level. The results showed a high prevalence rate of dysmenorrhea (79.4%) among the students. The most common risk factors were family history (87.4%) and length of menstruation (79%). Half (50.2%) of the respondents were absent at the university at least 1 day every month. The most widely used medications by the respondents were ibuprofen (42%) and paracetamol (40%), whereas only 3% used mefenamic acid, despite that they experienced complete pain relief with mefenamic acid. High prevalence rate of dysmenorrhea associated with risk factors such as family history and length of menstruation, was found among university students. However, pain and associated symptoms affect the quality of life.

Reproductive aging and menopause-like transition in the menstruating spiny mouse (Acomys cahirinus).

STUDY QUESTION: Does the naturally menstruating spiny mouse go through menopause? SUMMARY ANSWER: Our study is the first to show a natural and gradual menopausal transition in a rodent. WHAT IS KNOWN ALREADY: Age-related depletion of the human ovarian reserve (OvR) leads to menopause, the permanent cessation of menstruation and reproduction. Current rodent models of menopause are inappropriate for inferences of the human condition, as reproductive senescence is abrupt or induced through ovariectomy. The spiny mouse is the only confirmed rodent with a naturally occurring menstrual cycle. STUDY DESIGN, SIZE, DURATION: Histological assessment of virgin spiny mice occurred in females aged 6 months (n = 14), 1 year (n = 7), 2 years (n = 13), 3 years (n = 9) and 4 years (n = 9). Endocrinology was assessed in a further 9 females per age group. Five animals per group were used for ovarian stereology with additional ovaries collected at prenatal Day 35 (n = 3), day of birth (n = 5), postnatal Days 35 (n = 5) and 100 (n = 5) and 15 months (n = 5). PARTICIPANTS/MATERIALS, SETTING, METHODS: Morphological changes in the reproductive system were examined using hematoxylin and eosin stains. Proliferating cell nuclear antigen immunohistochemistry assessed endometrial proliferation and sex steroids estradiol and testosterone were assayed using commercial ELISA kits. MAIN RESULTS AND THE ROLE OF CHANCE: The proportion of females actively cycling was 86% at 6 months, 71% at 1 year, 69% at 2 years, 56% at 3 years and 44% at 4 years. Uterine and ovarian weights declined steadily from 1 year in all groups and corresponded with loss of uterine proliferation (P < 0.01). Estradiol was significantly decreased at 1 and 2 years compared to 6-month-old females, before becoming erratic at 3 and 4 years, with no changes in testosterone across any age. Fully formed primordial follicles were observed in prenatal ovaries. Aging impacted on both OvR and growing follicle numbers (P < 0.001-0.0001). After the age of 3 years, the follicle decline rate increased more than 5-fold. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: This is a descriptive study in a novel research rodent whereby reagents validated for use in the spiny mouse were limited. WIDER IMPLICATIONS OF THE FINDINGS: The gradual, rather than sudden, menopausal transition suggests that the spiny mouse is a more appropriate perimenopausal model than the current rodent models in which to examine the neuroendocrine pathways that encompass all hormonal interactions in the hypothalamic-pituitary-gonadal axis. The logistic, ethical and economic advantages of such a model may reduce our reliance on primates in menopause research and enable more thorough and invasive investigation than is possible in humans. STUDY FUNDING/COMPETING INTEREST(S): Hudson Institute is supported by the Victorian State Government Operational Infrastructure Scheme. The authors declare no competing interests.

Mouse model of menstruation: An indispensable tool to investigate the mechanisms of menstruation and gynaecological diseases (Review).

Abnormal menstruation may result in several pathological alterations and gynaecological diseases, including endometriosis, menstrual pain and miscarriage. However, the pathogenesis of menstruation remains unclear due to the limited number of animal models available to study the menstrual cycle. In recent years, an effective, reproducible, and highly adaptive mouse model to study menstruation has been developed. In this model, progesterone and oestrogen were administered in cycles following the removal of ovaries. Subsequently, endometrial decidualisation was induced using sesame oil, followed by withdrawal of progesterone administration. Vaginal bleeding in mice is similar to that in humans. Therefore, the use of mice as a model organism to study the mechanism of menstruation and gynaecological diseases may prove to be an important breakthrough. The present review is focussed ond the development and applications of a mouse model of menstruation. Furthermore, various studies have been described to improve this model and the research findings that may aid in the treatment of menstrual disorders in women are presented.

Detection of five specified menstrual fluid mRNA markers over the uterine cycle.

Matrix metallopeptidases (MMPs) 7, 10, and 11 are currently the most commonly employed messenger RNAs (mRNAs) for the identification of menstrual fluid (MF) in forensic analysis. However, no comprehensive study has been carried out to date to explore their time-dependent detection in vaginal samples. This research investigated the detection of MMPs 7, 10, and 11, as well as MMP3 and stanniocalcin 1 (STC1) over the uterine cycle. The aim was to associate relative transcript levels with cycle stages and thus determine which of these transcripts is most suitable for MF identification in a forensic context. Additionally, the effect of hormonal contraceptives (HCs) on their abundance was explored. A total of 300 vaginal swab samples were collected from eight female donors, including a pregnant woman, naturally cycling women, and contraceptive users. Differences among individuals were observed, but these were not consistent within the groups. Only MMP10 and STC1 mRNA abundance appeared to be unaffected by the use of HCs. MMP3, MMP7, and MMP11 transcripts were less abundant in MF samples of some HC users. Overall, MMP3 was most specific to MF, although this transcript was still detected in one of four vaginal material (VM) samples. STC1 was less specific than MMP3 (detected in 39.6 % of VM samples). However, over the days of menstruation, STC1 was more consistently detectable than the MMPs. MMP10 was least specific, with a 78.3 % detection rate in VM samples, but the presence/absence in VM was individual-specific and consistent. MMP10 may therefore be more useful as a VM marker with elevated abundance during menstruation in some individuals. MMP7 and MMP11 were the least reliable mRNAs for MF identification, despite an increased specificity compared to MMP10. Detection rates in MF were lower than those of MMP3 and STC1, whereas detection rates in VM were higher. MMP7 abundance additionally increased approximately 2-5 days after the end of menstruation in all donors except one naturally cycling individual. In view of these results, MMP3 and STC1 were identified as the most useful MF markers for forensic use. Nevertheless, mRNA typing results need to be interpreted with utmost caution.

Effect of Tahiti lime (Citrus latifolia) juice on the Production of the PGF2α/PGE2 and Pro-Inflammatory Cytokines involved in Menstruation.

Tahiti lemon juice (Citrus latifolia) (TLJ), as a natural source of flavonoids, has been used as an alternative to anti-inflammatory drugs for the treatment of dysmenorrhea and menstrual excessive bleeding, often associated with an imbalance of the prostaglandins (PG) levels. However, despite the positive effects, the mechanisms that rule menstruation control are still unknown. Therefore, the objectives were to characterize the TLJ and analyze its effect on the production of PGF2α, PGE2 and pro-inflammatory cytokines involved inmenstruation. Flavonoids from TLJ were discriminated by UPLC-DAD-MS/MS (Qq-TOF) and the effects of TLJ were studied in vitro by quantification of the contraction of myoblasts in culture and PGF2α and PGE2 productions. Further, the systemic and menstrual fluid levels of PGF2α, PGE2, IL-1ß, TNF-α, IL-6, AK1B1 and AK1C3 enzymes produced by women during the menstrual period were compared after exposition or not to TLJ or meloxicam. The results showed that TLJ induces an increase in the contraction of myoblasts and the PGF2α supernatant level. Regarding in vivo analysis, a higher concentration of PGF2α and an unaltered PGE2 level was also found in the menstrual blood of women treated with TLJ, in contrast with a lower level of PGE2 and PGF2α observed in the meloxicam group. Concerning cytokines, only menstrual TNF-α levels decrease after treatment with TLJ or meloxicam. In conclusion, TLJ may favor the control of menstruation events via a PGF2α mediated muscle contractile response.

The Effect of Twin Sex on Menstrual Characteristics.

Background and objectives: The purpose of this project is to evaluate the association between twin sex discordance and menstrual characteristics. We hypothesize that sharing the uterus with a male twin can change ovulation programming, hence changing the menstrual cycle characteristics during adulthood. This project could be novel in discovering new physiological mechanisms of hormone exposure and menstrual cycles. Materials and methods: This is a cross-sectional study. We asked females from sex-concordant (n = 1290) and sex-discordant (n = 168) twin pairs in the Washington State Twin Registry about characteristics of menstrual cycles. Generalized Estimating Equation (GEE) analysis was used to compare groups. The main outcome measures included the amount of bleeding, duration of menstruation, the timing of menstruation, length of menstruation, and a number of periods per year. Results: We found a statistically significant association between the amount of menstrual period bleeding and twin sex discordance (0.42 (95% CI 0.18-0.94)). However, twin sex discordance was not associated with period duration, length of menstrual cycle, cycle regularity, or a number of periods per year. Conclusions: Twin sex discordance is not a predictor of clinical characteristics of menstruation during adulthood except for the amount of bleeding. Future studies should focus on the impact of male hormones on the amount of bleeding during menstruation.

Hormonal Profiles of Menstrual Bleeding Patterns During the Luteal-Follicular Transition.

CONTEXT: Menstrual cycle function is determined by a complex endocrine axis that controls the ovaries and endometrium. While the late luteal phase is characterized by declining progesterone and estrogen, how these hormonal profiles relate to menstrual bleeding patterns is not well understood. OBJECTIVE: Characterize associations between luteal phase hormonal profiles and subsequent menstrual bleeding patterns, specifically spotting before bleeding. DESIGN, SETTING, AND PARTICIPANTS: We examined creatinine-adjusted urinary estrone 3-glucuronide (E13G) and pregnanediol 3-glucuronide (Pd3G) levels in relation to spotting in 116 premenopausal women (ages 20-47) who kept daily menstrual diaries and collected first morning urine samples for ≥ 2 consecutive cycles or 1 luteal-follicular transition (n = 283 transitions). We used linear mixed models to estimate associations between luteal phase hormone levels and spotting before bleeding. MAIN OUTCOME MEASURE(S) AND RESULTS: Transitions with ≥ 1 days of spotting before menstrual bleeding (n = 118) had greater luteal phase Pd3G levels vs nonspotting transitions (n = 165). Differences in Pd3G between spotting and nonspotting transitions were largest at menses onset (34.8%, 95% confidence interval, 18.9%, 52.7%). Pd3G levels for spotting transitions dropped to similar levels as nonspotting transitions an average of 1 day later, which aligned with the first day of bleeding for transitions with contiguous spotting. Spotting transitions were preceded by slower rates of Pd3G decline than nonspotting transitions, whereas E13G declines were similar. CONCLUSIONS: Self-reported bleeding patterns may provide insight into luteal phase Pd3G levels. First bleed appears to be the best choice for defining the end of the luteal phase and achieving hormonal consistency across transitions.

Distinct spectrum of microRNA expression in forensically relevant body fluids and probabilistic discriminant approach.

MicroRNA is attracting worldwide attention as a new marker for the identification of forensically relevant body fluids. A probabilistic discriminant model was constructed to identify venous blood, saliva, semen, and vaginal secretion, based on microRNA expression assessed via RT-qPCR. We quantified 15 candidate microRNAs in four types of body fluids by RT-qPCR and found that miR-144-3p, miR-451a-5p, miR-888-5p, miR-891a-5p, miR-203a-3p, miR-223-3p and miR-1260b were helpful to discriminate body fluids. Using the relative expression of seven candidate microRNAs in each body fluid, we implemented a partial least squares-discriminant analysis (PLS-DA) as a probabilistic discriminant model and distinguished four types of body fluids. Of 14 testing samples, 13 samples were correctly identified with >90% posterior probability. We also investigated the effects of microRNA expression in skin, semen infertility, and vaginal secretion during different menstrual phases. Semen infertility and menstrual phases did not affect our body fluid identification system. Therefore, the selected microRNAs were effective in identifying the four types of body fluids, indicating that probabilistic evaluation may be practical in forensic casework.

Physiologic Uterine Uptake of Radioiodine During Menstruation Demonstrated by SPECT/CT.

A 35-year-old woman with papillary thyroid cancer underwent I therapy after thyroidectomy. Post-therapy whole body scan revealed increased activity in the pelvis, in addition to the activity in the neck. On SPECT/CT images, the radioactivity in the pelvis was localized in the rectum and cervix. Further inquiry discovered that the patient was menstruating. We concluded that abnormal radioiodine uptake in menstrual uterus might be an exceptional finding mimicking a metastasis.

Activation of the Hippo/TAZ pathway is required for menstrual stem cells to suppress myofibroblast and inhibit transforming growth factor ß signaling in human endometrial stromal cells.

STUDY QUESTION: Can menstrual stem cells (MenSCs) inhibit myofibroblast differentiation and reverse transforming growth factor ß (TGFß)-mediated activation of myofibroblast phenotypes in human endometrial stromal cells (ESCs)? SUMMARY ANSWER: MenSCs suppressed endometrial myofibroblast differentiation and reversed TGFß-mediated activation of myofibroblast phenotypes, which might be associated with activation of the Hippo/TAZ pathway. WHAT IS KNOWN ALREADY: The potential effect of MenSCs as a cell therapy include attenuation of intrauterine adhesions, but the underlying mechanisms by which MenSCs exerts these effects are not entirely understood. STUDY DESIGN, SIZE, DURATION: We evaluated the antagonistic effects of MenSCs on myofibroblast differentiation as well as the broader effect of the Hippo/TAZ signaling pathway on TGFß-mediated induction of myofibroblast gene expression. The study design was based on a cohort of clinical proliferative phase endometrial samples obtained from three healthy premenopausal females with regular menstrual cycles. PARTICIPANTS/MATERIALS, SETTING, METHODS: ESCs were cocultured with MenSCs or in MenSC-conditioned medium. Fibrotic markers (αSMA, collagen I, CTGF and fibronectin) as well as proliferation and wound-healing abilities were evaluated. Components of the Hippo/TAZ pathway (TAZ, p-TAZ, MOB1, p-MOB1, LATS1 and p-LATS1) were also investigated. Cell Counting Kit 8, wound healing assay, real-time PCR, western blotting, immunofluorescence and shRNA knockdown approaches were used to validate the findings. MAIN RESULTS AND THE ROLE OF CHANCE: MenSCs inhibited myofibroblast activation, resulting in more rapid proliferation of ESCs. MenSCs downregulated the expression of myofibroblast markers αSMA and collagen I and promoted endometrial wound healing. Coculture with MenSCs also attenuated the TGFß-mediated increase in expression of fibrotic marker genes αSMA, collagen I, CTGF and fibronectin, and restored the wound-healing ability inhibited by TGFß. MenSCs induced Hippo/TAZ pathway activation, resulting in nuclear export and cytoplasmic retention of TAZ. TAZ inhibition was demonstrated to have similar effects even in the absence of MenSCs, and inhibition of TAZ was sufficient to attenuate TGFß-mediated myofibroblast activation. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: This study included only in vitro experiments. Thus, additional data from in vivo experiments are needed in a future study. WIDER IMPLICATIONS OF THE FINDINGS: The Hippo/TAZ pathway may be an important therapeutic target for endometrial fibrosis. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by the National Natural Science Foundation of China (No. 81601236) and Zhejiang Provincial Natural Science Foundation of China (LY19H040009). None of the authors has any competing interests to declare.

GABAA dysregulation as an explanatory model for late-onset postpartum depression associated with weaning and resumption of menstruation.

It is well established that a subgroup of women are particularly vulnerable to affective dysregulation during times of hormonal fluctuation. One underrecognized reproductive transition may be late-onset postpartum depression (PPD) in the context of weaning from breastfeeding and the resumption of menstruation. The goal of this review is to propose a biologically plausible mechanism for affective dysregulation during these transitions. The relationship between affective symptoms and neurohormonal changes associated with weaning will be investigated through a hypothesis-driven review of relevant literature. Neurosteroids, like allopregnanolone (ALLO), are widely recognized for augmenting GABAergic inhibition and having a powerful anxiolytic effect (Belelli D and Lambert JL, Nature Reviews Neuroscience 6:565-575, 2005). However, when ALLO is administered after prolonged withdrawal, there may be a paradoxical anxiogenic effect (Smith et al., Psychopharmacology 186:323-333, 2006; Shen et al., Nat Neurosci 10:469-477, 2007). Weaning from breastfeeding is a physiologic example of fluctuating levels of ALLO after prolonged withdrawal. We propose that the complex hormonal milieu during weaning and resumption of menstruation may modify GABAA receptors such that ALLO may contribute to rather than ameliorate depressive symptoms in vulnerable individuals. The proposed model provides an initial step for understanding the mechanisms by which the changing hormonal environment during weaning and resumption of menstruation may contribute to an increased risk of depression in a subgroup of women who are hormonally sensitive. Future research investigating this model would be valuable both to identify women at increased risk for developing mood symptoms late in postpartum and to inform treatment for this and related reproductive depressive disorders.

Menstrual fluid factors facilitate tissue repair: identification and functional action in endometrial and skin repair.

Repair after damage is essential for tissue homeostasis. Postmenstrual endometrial repair is a cyclical manifestation of rapid, scar-free, tissue repair taking ∼3-5 d. Skin repair after wounding is slower (∼2 wk). In the case of chronic wounds, it takes months to years to restore integrity. Herein, the unique "rapid-repair" endometrial environment is translated to the "slower repair" skin environment. Menstrual fluid (MF), the milieu of postmenstrual endometrial repair, facilitates healing of endometrial and keratinocyte "wounds" in vitro, promoting cellular adhesion and migration, stimulates keratinocyte migration in an ex vivo human skin reconstruct model, and promotes re-epithelialization in an in vivo porcine wound model. Proteomic analysis of MF identified a large number of proteins: migration inhibitory factor, neutrophil gelatinase-associated lipocalin, follistatin like-1, chemokine ligand-20, and secretory leukocyte protease inhibitor were selected for further investigation. Functionally, they promote repair of endometrial and keratinocyte wounds by promoting migration. Translation of these and other MF factors into a migration-inducing treatment paradigm could provide novel treatments for tissue repair.-Evans, J., Infusini, G., McGovern, J., Cuttle, L., Webb, A., Nebl, T., Milla, L., Kimble, R., Kempf, M., Andrews, C. J., Leavesley, D., Salamonsen, L. A. Menstrual fluid factors facilitate tissue repair: identification and functional action in endometrial and skin repair.

Characterization of human-like menstruation in the spiny mouse: comparative studies with the human and induced mouse model.

STUDY QUESTION: Is the newly discovered menstruating rodent, the spiny mouse, a valid model for studying endometrial morphology and menstruation? SUMMARY ANSWER: Our study is the first to demonstrate a primate-like pattern of natural menstruation in a rodent, with decidualization, spiral arteriole remodeling and piece-meal endometrial shedding. WHAT IS KNOWN ALREADY: The spiny mouse has a naturally occurring menstrual cycle. This unique feature has the potential to reduce the heavy reliance on primates and provide a more appropriate small animal model for menstrual physiology research. STUDY DESIGN, SIZE, DURATION: This study compares morphological changes in the endometrium during early, mid and late menstruation of the spiny mouse (n = 39), human (n = 9) and the induced mouse model of menstruation (n = 17). PARTICIPANTS/MATERIALS, SETTING, METHODS: We assessed tissue morphology with hematoxylin and eosin and erythrocyte patterns with Mallory's trichrome. We conducted staining for neutrophil gelatinase associated lipocalin (NGAL), cytokeratin and interleukin-11 (IL-11) in all species. We used double immunofluorescence staining for vascular endothelial growth factor and alpha-smooth muscle actin to detect vasculature remodeling and western immunoblot to detect interleukin-8 (IL-8) and macrophage migration inhibitory factor (MIF) in the menstrual fluid of spiny mice. MAIN RESULTS AND THE ROLE OF CHANCE: Menstruation occurs in the spiny mouse over a 72-h period, with heaviest menstrual breakdown occurring 24 h after initial observation of red blood cells in the vaginal cytology. During menstruation, the endometrium of the spiny mouse appeared to resemble human menstrual shedding with focal epithelial breakdown observed in conjunction with lysis of underlying stroma and detection of IL-8 and MIF in menstrual fluid. The mouse exhibits extensive decidualization prior to induced menses, with transformation of the entire uterine horn and cytokeratin expression absent until initiation of repair. Decidualization occurred spontaneously and was less marked in the spiny mouse, where epithelial integrity remained intact. In all species, the decidua was positive for IL-11 secretion and neutrophil recruitment was similar in each. Spiral arteriole formation was confirmed in the spiny mouse. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: This is a descriptive study comparing primarily morphological traits between the spiny mouse, the mouse and the human. Reagents specific to the spiny mouse were limited and resources for global use of this novel species are few. WIDER IMPLICATIONS OF THE FINDINGS: Our work supports the spiny mouse as a viable model, sharing many attributes of physiological menstruation with humans. The strength of a natural as opposed to an artificial model is validated through the striking similarities observed between the spiny mouse and human in uterine breakdown. The spiny mouse may be highly useful in large-scale investigations of menstruation and menstrual disorders. STUDY FUNDING/COMPETING INTEREST(S): N.B. and S.R. are each recipients of a Research Training Program scholarship supported by Monash University. This work was supported by the Victorian Government Operational Infrastructure and laboratory funds to H.D. The authors declare no competing interests.

Bone metabolic responses to low energy availability achieved by diet or exercise in active eumenorrheic women.

PURPOSE: We aimed to explore the effects of low energy availability (EA)[15 kcal·kg lean body mass (LBM)-1·d-1] achieved by diet or exercise on bone turnover markers in active, eumenorrheic women. METHODS: By using a crossover design, ten eumenorrheic women (VO2 peak: 48.1 ±â€¯3.3 ml·kg-1·min-1) completed all three, 3-day conditions in a randomised order: controlled EA (CON; 45 kcal·kgLBM-1·d-1), low EA through dietary energy restriction (D-RES; 15 kcal·kgLBM-1·d-1) and low EA through increasing exercise energy expenditure (E-RES; 15 kcal·kgLBM-1·d-1), during the follicular phase of three menstrual cycles. In CON, D-RES and E-RES, participants consumed diets providing 45, 15 and 45 kcal·kgLBM-1·d-1. In E-RES only, participants completed supervised running sessions (129 ±â€¯10 min·d-1) at 70% of their VO2 peak that resulted in an exercise energy expenditure of 30 kcal·kg LBM-1·d-1. Blood samples were collected at baseline (BASE) and at the end of the 3-day period (D6) and analysed for bone turnover markers (ß-CTX and P1NP), markers of calcium metabolism (PTH, albumin-adjusted Ca, Mg and PO4) and hormones (IGF-1, T3, insulin, leptin and 17ß-oestradiol). RESULTS: In D-RES, P1NP concentrations at D6 decreased by 17% (BASE: 54.8 ±â€¯12.7 µg·L-1, D6: 45.2 ±â€¯9.3 µg·L-1, P < 0.001, d = 0.91) and were lower than D6 concentrations in CON (D6: 52.5 ±â€¯11.9 µg·L-1, P = 0.001). P1NP did not change significantly in E-RES (BASE: 55.3 ±â€¯14.4 µg·L-1, D6: 50.9 ±â€¯15.8 µg·L-1, P = 0.14). ß-CTX concentrations did not change following D-RES (BASE: 0.48 ±â€¯0.18 µg·L-1, D6: 0.55 ±â€¯0.17 µg·L-1) or E-RES (BASE: 0.47 ±â€¯0.24 µg·L-1, D6: 0.49 ±â€¯0.18 µg·L-1) (condition × time interaction effect, P = 0.17). There were no significant differences in P1NP (P = 0.25) or ß-CTX (P = 0.13) responses between D-RES and E-RES. Both conditions resulted in reductions in IGF-1 (-13% and - 23% from BASE in D-RES and E-RES, both P < 0.01) and leptin (-59% and - 61% from BASE in D-RES and E-RES, both P < 0.001); T3 decreased in D-RES only (-15% from BASE, P = 0.002) and PO4 concentrations decreased in E-RES only (-9%, P = 0.03). CONCLUSIONS: Low EA achieved through dietary energy restriction resulted in a significant decrease in bone formation but no change in bone resorption, whereas low EA achieved through exercise energy expenditure did not significantly influence bone metabolism. Both low EA conditions elicited significant and similar changes in hormone concentrations.