Adenomyosis and Abnormal Uterine Bleeding: Review of the Evidence.

BACKGROUND: Thomas Cullen described bleeding abnormalities and dysmenorrhea as the "expected" presentations of adenomyomas. Adenomyosis is included within the FIGO classification of structural causes of abnormal uterine bleeding (AUB). Nevertheless, this long-standing association has been questioned by some authors who reported a high incidence of adenomyosis in uteri removed for indications other than AUB or dysmenorrhea. Here, we examine evidence for the link between adenomyosis and AUB. METHODS: A comprehensive Medline literature review of all publications to October 2023. RESULTS: Sixty-three articles were identified and included in the review. Despite a large body of studies, the available literature does not provide conclusive evidence of a link between adenomyosis and AUB. This is because of unsuitable study design, or poor characterization of the study population or of the inclusion or exclusion criteria. Additional challenges arise because of the lack of agreed criteria for diagnosing adenomyosis and the often absence of detailed assessment of menstrual blood loss. Adenomyosis often coexists with other conditions that have also been linked to similar symptoms, and many cases of adenomyosis are asymptomatic. CONCLUSION: Most of the existing literature and studies that addressed treatment outcome of adenomyosis started from the premise that a link between the condition and AUB had been proven. Yet, published information shows that aspects such a relationship is still uncertain. Further research is needed to address the relation between AUB and adenomyosis burden (or subtypes), distribution, and concomitant pathology.

Is Neonatal Uterine Bleeding Involved in Early-Onset Endometriosis?

BACKGROUND: There has been considerable progress in our understanding of endometriosis, but its pathophysiology remains uncertain. Uncovering the underlying mechanism of the rare instances of endometriosis reported in early postmenarcheal years and in girls before menarche can have wide implications. METHODS: We conducted a literature review of all relevant articles on Medline. RESULTS: In the review, we explore the pathogenetic theories of premenarcheal endometriosis, the role of retrograde menstruation in the adult and its potential role in early-onset disease, as well as the factors that argue against the existence of a link between early-onset endometriosis (EOE) and neonatal uterine bleeding (NUB). CONCLUSIONS: As with endometriosis in adult women, the pathogenesis of early-onset disease remains unclear. A link between NUB and EOE is plausible, but there are considerable challenges to collating supporting evidence. The state of our understanding of early uterine development and of the pathophysiology of NUB leaves many unknowns that need exploration. These include proof of the existence of viable endometrial cells or endometrial mesenchymal stem cells in NUB, their passage to the pelvic cavity, their possible response to steroids, and whether they can reside within the pelvic cavity and remain dormant till menarche.

Hypermethylation of Klotho and Peroxisome Proliferator-Activated Receptor γ Concomitant with Overexpression of DNA Methyltransferase 1 in Adenomyosis.

Cellular senescence is known to be involved in tissue repair, but its role in adenomyosis remains unclear. This study was tasked to evaluate the expression of Klotho, a well-known aging-suppressing protein, as well as PPARγ and DNMT1 in adenomyotic lesions (AD) in comparison with that of control endometrium (CT). We performed immunohistochemistry analysis of markers of cellular senescence p16 and p21, along with Klotho, PPARγ and DNMT1 in CT and AD samples, followed by the quantification of gene expression of Klotho, PPARγ and DNMT1 in epithelial organoids derived from AD and CT samples and methylation-specific PCR to evaluate promoter methylation status. The effect of forced expression and knockdown of DNMT1 on Klotho and PPARγ expression in ectopic endometrial epithelial cells was evaluated. We found that both p16 and p21 immunoreactivity in AD was significantly higher while that of Klotho and PPARγ was significantly lower than CT samples, which was concomitant with elevated immunoexpression of DNMT1. The results were confirmed by transcriptional analysis using epithelial organoids derived from AD and CT samples. In addition, the promoter regions of both Klotho and PPARγ genes were hypermethylated in AD as compared with CT, and treatment with HDAC and DNMT inhibitors reactivated the expression of both Klotho and PPARγ. Forced expression of DNMT1 resulted in downregulation of both Klotho and PPARγ but its knockdown increased their expression. Thus, overexpression of DNMT1 seems to facilitate the promoter hypermethylation of both Klotho and PPARγ in AD, resulting in their reduced expression that is suggestive of the role of senescence in adenomyosis.

Meclizine improves endometrial repair and reduces simulated menstrual bleeding in mice with induced adenomyosis.

BACKGROUND: Adenomyosis is one of the structural causes of abnormal uterine bleeding, which often presents as heavy menstrual bleeding. Mostly because of the poor understanding of its pathophysiology, medical management of adenomyosis-induced heavy menstrual bleeding is still a challenge. We have previously reported that glycolysis is crucial to endometrial repair following menstruation and that suppressed glycolysis can cause heavy menstrual bleeding. OBJECTIVE: This study aimed to test the hypothesis that meclizine, a drug with an excellent safety profile, alleviates heavy menstrual bleeding in mice with induced adenomyosis using a simulated menstruation model. STUDY DESIGN: Adenomyosis was induced in 36 female C57BL/6 mice using endometrial-myometrial interface disruption. Three months after induction, the mice were randomly divided into the following 3 groups: low-dose meclizine, high-dose meclizine, and controls. Treatment with meclizine or vehicle started shortly before the simulated menstruation procedure and ended before progesterone withdrawal. The amount of blood loss was quantified and uterine tissue was harvested for histologic evaluation of the grade of endometrial repair. We performed immunohistochemistry analysis of 4 proteins critically involved in glycolysis: Glut1 (glucose transporter 1), Hk2 (hexokinase 2), Pfkfb3 (6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3), and Pkm2 (pyruvate kinase M2). The extent of tissue fibrosis in both ectopic and eutopic endometria was evaluated using Masson trichrome staining. RESULTS: In mice with induced adenomyosis, meclizine accelerated endometrial repair in a dose-dependent manner and reduced the amount of menstrual bleeding. Meclizine administration raised endometrial immunoexpression of Hk2 and Pfkfb3 but not of Glut1 or Pkm2. The extent of endometrial fibrosis was reduced following the meclizine administration. Remarkably, these favorable changes were accompanied by the suppression of lesional progression, as evidenced by the dose-dependent reduction in the extent of fibrosis (a surrogate for lesional progression). CONCLUSION: These encouraging results, taken together, suggest that glycolysis may be a promising therapeutic target and that meclizine may hold therapeutic potential as a nonhormonal treatment for adenomyosis-induced heavy menstrual bleeding without exacerbating the disease.

Systematic review and meta-analysis of reproductive outcomes after high-intensity focused ultrasound (HIFU) treatment of adenomyosis.

High-intensity focused ultrasound (HIFU) has emerged as a promising uterus-sparing and possibly fertility-sparing treatment modality for women with adenomyosis, especially those who desire to conceive. We conducted this systematic review and performed a meta-analysis on clinical studies aimed to improve reproduction in women with adenomyosis. After extensive search of PubMed and CNKI, we identified 10 studies published in English and Chinese involving a total of 557 patients with adenomyosis who desired to conceive after HIFU treatment. We found a pooled estimate of pregnancy rate of 53.4% and of the live birth rate of 35.2%, and there was a substantial heterogeneity among these studies. While there is a potential for HIFU treatment to improve fertility for patients with adenomyosis who desired to conceive, such evidence is very weak as of now. Comparative studies with much higher methodological rigor, preferably randomized clinical trials, are badly needed to further illuminate this issue.

Shorter Anogenital Distance in Women with Ovarian Endometriomas and Adenomyosis, but Not Uterine Leiomyomas.

We investigated whether anogenital distance (AGD) is associated with adenomyosis, endometriosis and uterine leiomyomas (UL, also called uterine fibroids). We recruited 81 women with UL, 105 with ovarian endometrioma (OE), 116 with adenomyosis, 28 with both adenomyosis and UL, and 100 control subjects with other acquired gynecological conditions but not endometriosis, adenomyosis, UL, or polycystic ovarian syndrome. Measurements from the anterior clitoral surface to the center of the anus (AGDAC), from the tip of the clitoris to the center of the anus (AGDACt), and from the posterior fourchette to the center of the anus (AGDAF) were made in all subjects. Multiple regression was performed to estimate the association between AGDs and presence of OE, adenomyosis, and UL while controlling for possible confounding factors. We found that, compared with controls, women with OE and adenomyosis, but not UL, had significantly shorter AGDAF, but not AGDAC. However, the amount of variance that could be explained by the disease status is rather moderate, suggesting that factors other than disease status, bodyweight and height were also responsible for AGD. Thus, prenatal exposure to reduced levels of androgen may increase the risk of developing endometriosis and adenomyosis. However, other factors may also contribute to the pathogenesis of endometriosis and adenomyosis.

Events Leading to the Establishment of Pregnancy and Placental Formation: The Need to Fine-Tune the Nomenclature on Pregnancy and Gestation.

Today, there is strong and diversified evidence that in humans at least 50% of early embryos do not proceed beyond the pre-implantation period. This evidence comes from clinical investigations, demography, epidemiology, embryology, immunology, and molecular biology. The purpose of this article is to highlight the steps leading to the establishment of pregnancy and placenta formation. These early events document the existence of a clear distinction between embryonic losses during the first two weeks after conception and those occurring during the subsequent months. This review attempts to highlight the nature of the maternal-embryonic dialogue and the major mechanisms active during the pre-implantation period aimed at "selecting" embryos with the ability to proceed to the formation of the placenta and therefore to the completion of pregnancy. This intense molecular cross-talk between the early embryo and the endometrium starts even before the blastocyst reaches the uterine cavity, substantially initiating and conditioning the process of implantation and the formation of the placenta. Today, several factors involved in this dialogue have been identified, although the best-known and overall, the most important, still remains Chorionic Gonadotrophin, indispensable during the first 8 to 10 weeks after fertilization. In addition, there are other substances acting during the first days following fertilization, the Early Pregnancy Factor, believed to be involved in the suppression of the maternal response, thereby allowing the continued viability of the early embryo. The Pre-Implantation Factor, secreted between 2 and 4 days after fertilization. This linear peptide molecule exhibits a self-protective and antitoxic action, is present in maternal blood as early as 7 days after conception, and is absent in the presence of non-viable embryos. The Embryo-Derived Platelet-activating Factor, produced and released by embryos of all mammalian species studied seems to have a role in the ligand-mediated trophic support of the early embryo. The implantation process is also guided by signals from cells in the decidualized endometrium. Various types of cells are involved, among them epithelial, stromal, and trophoblastic, producing a number of cellular molecules, such as cytokines, chemokines, growth factors, and adhesion molecules. Immune cells are also involved, mainly uterine natural killer cells, macrophages, and T cells. In conclusion, events taking place during the first two weeks after fertilization determine whether pregnancy can proceed and therefore whether placenta's formation can proceed. These events represent the scientific basis for a clear distinction between the first two weeks following fertilization and the rest of gestation. For this reason, we propose that a new nomenclature be adopted specifically separating the two periods. In other words, the period from fertilization and birth should be named "gestation", whereas that from the completion of the process of implantation leading to the formation of the placenta, and birth should be named "pregnancy".

Reduced endometrial expression of histone deacetylase 3 in women with adenomyosis who complained of heavy menstrual bleeding.

RESEARCH QUESTION: What role, if any, does histone deacetylase 3 (HDAC3) play in adenomyosis-associated heavy menstrual bleeding (HMB)? DESIGN: Seventy-two women with adenomyosis-associated HMB were recruited. Of these, 37 women reported moderate/heavy bleeding (MHB) and the remaining 35 women reported excessive bleeding (EXB). The stiffness of adenomyotic lesions and neighbouring endometrial-myometrial interface (EMI) was measured by transvaginal elastosonography, and full-thickness uterine tissue columns were processed for Masson trichrome staining and immunohistochemistry analyses. The protein expression levels of HDAC3 in endometrial cells cultured on substrates of different stiffnesses, and the protein concentrations of nuclear factor-κB (NF-κB) p65 subunit with HDAC3 suppression were evaluated. Mouse experiments were performed to assess the effect of adenomyosis on Hdac3 expression, endometrial repair and bleeding, and to evaluate the effect of HDAC3 inhibition on endometrial repair. RESULTS: Compared with controls, the endometrial staining of HDAC3 was significantly lower in women with adenomyosis-associated HMB, concomitant with a greater extent of fibrosis. The stiffness of lesions and neighbouring EMI was significantly higher in the EXB group compared with the MHB group, as was the extent of fibrosis in lesions, their neighboring EMI and endometrium. Expression of HDAC3 was reduced significantly when endometrial epithelial cells were cultured in stiff substrates. Suppression of HDAC3 abrogated the activation and signalling of NF-κB. Mice with induced adenomyosis exhibited reduced Hdac3 staining and elevated fibrosis in endometrium, concomitant with disrupted endometrial repair and more bleeding. Hdac3 inhibition resulted in botched inflammation and increased bleeding. CONCLUSIONS: Lesional fibrosis results in reduced endometrial HDAC3 expression and subsequent disruption in NF-κB signalling and inflammation, leading to adenomyosis-associated HMB.

The Asian Society of Endometriosis and Adenomyosis guidelines for managing adenomyosis.

This is the first guidelines for adenomyosis from the Asian Society of Endometriosis and Adenomyosis.

Is neonatal uterine bleeding responsible for early-onset endometriosis?

BACKGROUND: It has been hypothesized that the origin of early-onset endometriosis could be from endometrial mesenchymal stem cells (eMSCs) in neonatal uterine blood (NUB). There is no information on the possible mechanistic basis linking an association between NUB/neonatal endometrium and development of early-onset endometriosis. In this study we performed a series of experiments to clarify the mechanistic link between NUB and/or neonatal endometrium and development of early-onset endometriosis. METHODS: We retrospectively collected postmortem neonatal endometria (n = 15) and prospectively collected NUB (n = 18) of female babies for the analysis of different biological markers including eMSCs. Immunohistochemical analysis of neonatal endometria was performed to examine the expression patterns of ovarian steroid receptors (ER/PGR), decidualization (prolactin, IGFBP1), pre-decidualization (Glycodelin A, α-SMA), proliferation (Ki-67 index), vascularity (CD31 + cells), immunocompetent CD68+, CD45+, CD56 + cells and some putative markers of eMSCs. Cell transfer method and immunocytochemistry were used to investigate the eMSCs and/or endometrial cells in NUB. RESULTS: Immunohistochemical analysis of postmortem neonatal endometria revealed variable staining response to ER/PGR, decidual markers, and substantial proliferative and angiogenic activity. A moderate to strong immunoexpression of Glycodelin-A was found in both neonatal and adult endometria. The tissue infiltration of CD56+, CD45 + and CD68 + immunocompetent cells was significantly low in neonatal endometria than that in adult endometria (p = 0.0003, p < 0.0001, p = 0.034, respectively). No eMSCs or even endometrial cells were detected in NUB. However, a variable expression of some phenotypes of eMSCs (CD90/CD105) was found in neonatal endometria. CONCLUSIONS: Based on our serial experiments we did not find any supporting evidence for the role of NUB in early-onset endometriosis. Neonatal endometria showed variable expression of ovarian steroid receptors, decidualization, and a substantial amount of proliferative and angiogenic activity. As an alternative mechanism, a significantly less tissue accumulation of immunocompetent cells in neonatal endometria may explain the survival of ER + and PGR + cells should they make entry into the pelvis and consequent development of early endometriosis with the onset of ovarian function. Future study with large sample size and application of modified technological tools is warranted to test the NUB hypothesis and to clarify their biological or clinical significance. TRIAL REGISTRATION: not applicable.

An Appraisal of the Tissue Injury and Repair (TIAR) Theory on the Pathogenesis of Endometriosis and Adenomyosis.

As understanding their pathogenesis remains elusive, both endometriosis and adenomyosis are often referred to as "enigmatic diseases". The uncertainty and heightened interest are reflected in the range of expressed views and opinions. There is a sense of urgency because of the entailed patient suffering. The plethora of opinions calls for a critical analysis of proposed theories, both old and new. A series of papers published since 2009 proposed that both endometriosis and adenomyosis originate from the same aberrations occurring within the uterus. This came to be recognized as the tissue injury and repair theory, and the newly coined term "archimetrosis" posits that the two diseases share the same origin. While the theory opens an interesting channel for exploration, its claim as a unifying theory necessitates a critical appraisal. We, thus, undertook this review of the theory and analyzed its underpinnings based on a comprehensive review of the literature. Our appraisal indicates that the theory is open to a range of criticisms. Chief among these is the need for confirmatory evidence of features of abnormal uterine contractility and the lack of data addressing the question of causality. In addition, the theory has, as yet, no supporting epidemiological evidence, which is a major weakness. The theory suffers as it is not open to the test of falsifiability, and it lacks the ability to make useful predictions. It has not addressed the questions, such as why only a small percentage of women develop adenomyosis or endometriosis, given the ubiquity of uterine peristalsis. On the other hand, the triggers and prevention of hyper- or dys-peristalsis become critical to a theory of causation. We conclude that additional supportive evidence is required for the theory to be accepted.

Association between tissue stress reaction and ACE2/TMPRSS2 expression in endometria of reproductive aged women before and during Covid-19 pandemic.

BACKGROUND: A potential concern has been raised regarding fertility and reproductive outcome during the Covid-19 pandemic with growing stress and anxiety. However, information on the association between tissue stress reaction and expression profiles of SARS-CoV-2 viral entry proteins, ACE2 and TMPRSS2, in endometria collected from women before (pre-pandemic) and during the Covid-19 pandemic (in-pandemic) is unknown. We aim to investigate the relationship between the expression of stress-reactive proteins and of ACE2 and TMPRSS2 in endometria collected from women during these two different time frames. METHODS: We retrospectively retrieved tissue blocks of endometrial samples from 25 women in 2019 (pre-pandemic) and 25 women in 2020 (in-pandemic) who underwent hysterectomy for different gynecological indications. Immunohistochemical analysis was performed with endometrial tissue samples that were collected before and during the pandemic, using respective antibodies targeting ACE2/TMPRSS2, ADRB2 and NK1R (stress and anxiety receptor markers, respectively). The quantification of immunoreactive cells for each marker was calculated by the immunoreactive score (IRS) analysis. This retrospective cohort study was limited to small sample size. RESULTS: No significant differences in the IRS of ACE2 and TMPRSS2 were found between the endometria that were collected before and during the pandemic with a lack of correlation between ACE2 and TMPRSS2 expression in respective endometria (r = 0.11, pre-pandemic; r = 0.04, in-pandemic). The immunostaining levels of stress marker, ADRB2 were significantly higher in the endometria of in-pandemic group (p = 0.015) comparing to that of pre-pandemic group. Pearson's correlation coefficient analysis showed a significant correlation in the expression between ADRB2 and TMPRSS2 (r = 0.41, p = 0.042) in the endometria of in-pandemic group but not in the pre-pandemic group. CONCLUSION: The rise in stress and anxiety among women during current pandemic may elicit substantial amount of tissue stress reaction with consequent increase in the expression of SARS-CoV-2 viral entry proteins in their endometria. A lack of correlation between ACE2 and TMPRSS2 expression in endometria may reassure women in their reproductive age that they are not more susceptible to infection by SARS-CoV-2 and suggest that stressful women during this pandemic can safely decide to conceive naturally or by artificial reproductive technology.

In the thicket of fears, doubts, and murky facts: some reflections on treatment modalities for endometriosis-associated pain.

Endometriosis-associated pain can be managed by either surgery or hormonal therapy. The final decision as to which treatment modality to take is based on efficacy and possible complications of different treatment modalities, risk of recurrence, and the patient's wishes and preferences. But in the thicket of fears, doubts, and murky facts, the choice may ultimately be the trade-off between irrational fears and ignorance versus scientific evidence. We elaborate some pros and cons of the two treatment modalities and highlight some notable downsides of hormonal therapy, in particular the possible yet unquantified risk of long-term hormonal therapy for malignant transformation, perhaps with the only exception of combined oral contraceptives. Thus, when discussing with patients, we advocate the approach of discussing the advantages and disadvantages of all treatment options in detail, accounting for the known pros and cons with a full understanding of the predictive irrationality of human beings. For endometriosis-associated pain, surgery is definitely not a failure of medicine but, rather, a viable option, especially given the recently surfaced undercurrent of wariness and dissatisfaction with the current hormonal drugs among patients with endometriosis. Above all, there is a pressing need to fill the knowledge gap of perioperative interventions intended to reduce the risk of recurrence and to fulfill the demand for the development of safe and efficacious non-hormonal therapeutics.

Various types of adenomyosis and endometriosis: in search of optimal management.

Currently, hormonal drugs are the mainstay for treating endometriosis or adenomyosis. Aside from staying within the "therapeutic window" and cutting off the hormonal support to the ectopic endometrium, none of these treatments were developed with a previously identified target. The current lack of innovation is humbling, especially considering that among >34,000 PubMed-indexed publications on endometriosis or adenomyosis, approximately 65% of them were published in the last 2 decades, many of them presumably with all the benefits of omics and molecular biology. One major reason for so many doomed research and development (R&D) projects on nonhormonal drugs is the failure to understand the natural history of the ectopic endometrium. Other reasons include, but are not limited to, the number of side effects that these nonhormonal drug candidates presented, which the developers were either unaware of or simply had ignored when pursuing a too narrowly minded development-a common cognitive trap known as invisible gorillas. Unless we have a clear global picture with a good grasp of the natural history of ectopic endometrium, "potholes" and "blind alleys" in drug R&D are difficult to avoid. This review shall explore some new concepts in the progression of ectopic endometrium and explain how these new perspectives offer insights for devising better therapeutic options, explain why some drug R&D projects for endometriosis or adenomyosis have failed, provide a critical analysis of some issues encountered in everyday practice, expose knowledge gaps in need of further research, and, finally, offer possible ways to circumvent hurdles and potholes and avoid cul-de-sacs in nonhormonal drug R&D.

From Retrograde Menstruation to Endometrial Determinism and a Brave New World of "Root Treatment" of Endometriosis: Destiny or a Fanciful Utopia?

Practically unknown outside of China, the "endometrial determinism" theory was proposed to account for the apparent gap between the relatively low prevalence of endometriosis and nearly universal retrograde menstruation. Attracting uncritical advocacy, the theory culminates in a recent consensus by elite Chinese gynecologists in favor of "root treatment", intended to nip endometriosis in the bud. Correcting endometrial "defects" can gain further momentum by the presence of cancer-driver mutations such as KRAS mutations in the endometrium of women with endometriosis and the recent introduction of therapeutics aiming to rectify the effect of these mutations for cancer treatment. We provide a critical appraisal of evidence for endometrial aberrations in endometriosis and relevant experimental evidence. All available evidence of endometrial "defect" is invariably post hoc and may well be secondary to induced endometriosis. We propose that the theory of "endometrial determinism" needs to demonstrate a clear causal and a phylogenetic relationship between endometrial aberrations and endometriosis. We argue that while it is highly likely that endometriosis is a consequence of retrograde menstruation, the case that molecular aberrations as a sole or a necessary determinant remains to be proven. "Root treatment" is a worthy ambition but as of now it is close to a fanciful Utopia.

The Role of Platelets in the Pathogenesis and Pathophysiology of Adenomyosis.

Widely viewed as an enigmatic disease, adenomyosis is a common gynecological disease with bewildering pathogenesis and pathophysiology. One defining hallmark of adenomyotic lesions is cyclic bleeding as in eutopic endometrium, yet bleeding is a quintessential trademark of tissue injury, which is invariably followed by tissue repair. Consequently, adenomyotic lesions resemble wounds. Following each bleeding episode, adenomyotic lesions undergo tissue repair, and, as such, platelets are the first responder that heralds the subsequent tissue repair. This repeated tissue injury and repair (ReTIAR) would elicit several key molecular events crucial for lesional progression, eventually leading to lesional fibrosis. Platelets interact with adenomyotic cells and actively participate in these events, promoting the lesional progression and fibrogenesis. Lesional fibrosis may also be propagated into their neighboring endometrial-myometrial interface and then to eutopic endometrium, impairing endometrial repair and causing heavy menstrual bleeding. Moreover, lesional progression may result in hyperinnervation and an enlarged uterus. In this review, the role of platelets in the pathogenesis, progression, and pathophysiology is reviewed, along with the therapeutic implication. In addition, I shall demonstrate how the notion of ReTIAR provides a much needed framework to tether to and piece together many seemingly unrelated findings and how it helps to make useful predictions.

Decreased Glycolysis at Menstruation is Associated with Increased Menstrual Blood Loss.

Heavy menstrual bleeding (HMB) is common and severely affects the quality of life of the afflicted women. While HMB is known to be caused by impaired endometrial repair after menstruation, its more proximate cause remains unknown. To investigate whether glycolysis plays any role in endometrial repair and thus HMB, we conducted two mouse experiments using a mouse model of simulated menstruation. We performed immunohistochemistry analyses of proteins involved in glycolysis as well as pro- and anti-inflammatory cytokines in endometrium from decidualized and non-decidualized uterine horns. We also assessed the extent of endometrial repair by staging endometrial morphology from decidualization to full repair using histological scoring of uterine sections and quantitated the amount of menstrual blood loss (MBL). In addition, we employed the scratch assay and the CCK-8 assay to evaluate the effect of glycolysis suppression on cellular migration and proliferation, respectively. Finally, we performed an immunohistochemistry analysis of HK2 in endometrium from women with adenomyosis who experienced either moderate/heavy or excessive MBL. We found that endometrial repair coincided with increased glycolysis in endometrium and glycolysis suppression delayed endometrial repair, resulting in increased MBL. Additionally, glycolysis suppression significantly inhibited the proliferative and migratory capability of endometrial cells, and disrupted normal endometrial repair even when hypoxia was maintained. Women with adenomyosis who experienced excessive MBL had significantly lower HK2 staining than those who experienced moderate/heavy MBL. Thus, our study highlights the importance of glycolysis as well as inflammation in optimal endometrial repair, and provides clues for the cause of HMB in women with adenomyosis.

Are Adenomyosis and Endometriosis Phenotypes of the Same Disease Process?

In recent literature reviews, we concluded that the possibility that endometrial molecular aberrations are the sole or a necessary determinant of endometriosis and the Tissue Injury and Repair (TIAR) theory are yet to be convincingly proven. Here, we critically examine the theory that adenomyosis and endometriosis represent different phenotypes of a single disease. A common etiopathology for adenomyosis and endometriosis has been suggested because both conditions entail the presence of endometrial tissue at locations other than the lining of the uterus. There are wide differences in reported disease incidence and prevalence and, consequently, in estimates of the coexistence of both conditions. There are some similarities but also differences in their clinical features and predisposing factors. Each condition has a range of subtypes. These differences alone pose the question of whether subtypes of endometriosis and adenomyosis have different etiopathologies, and, in turn, this raises the question of whether they all share a common etiology. It is debatable whether the recognized differences between the eutopic endometrium in adenomyosis and endometriosis compared to those in unaffected women are the cause or the effect of the disease. The finding of common mutations, particularly of KRAS, lend support to the notion of shared predisposing factors, but this alone is insufficient evidence of causation.

Cracking the enigma of adenomyosis: an update on its pathogenesis and pathophysiology.

In brief: Traditionally viewed as enigmatic and elusive, adenomyosis is a fairly common gynecological disease but is under-recognized and under-researched. This review summarizes the latest development on the pathogenesis and pathophysiology of adenomyosis, which have important implications for imaging diagnosis of the disease and for the development of non-hormonal therapeutics. Abstract: Traditionally considered as an enigmatic disease, adenomyosis is a uterine disease that affects many women of reproductive age and is a contributing factor for pelvic pain, heavy menstrual bleeding (HMB), and subfertility. In this review, the new development in the pathogenesis and pathophysiology of adenomyosis has been summarized, along with their clinical implications. After reviewing the progress in our understanding of the pathogenesis and describing the prevailing theories, in conjunction with their deficiencies, a new hypothesis, called endometrial-myometrial interface disruption (EMID), which is backed by extensive epidemiologic data and demonstrated by a mouse model, is reviewed, along with recent data implicating the role of Schwann cells in the EMI area in the genesis of adenomyosis. Additionally, the natural history of adenomyotic lesions is elaborated and underscores that, in essence, adenomyotic lesions are fundamentally wounds undergoing repeated tissue injury and repair (ReTIAR), which progress to fibrosis through epithelial-mesenchymal transition, fibroblast-to-myofibroblast transdifferentiation, and smooth muscle metaplasia. Increasing lesional fibrosis propagates into the neighboring EMI and endometrium. The increased endometrial fibrosis, with ensuing greater tissue stiffness, results in attenuated prostaglandin E2, hypoxia signaling and glycolysis, impairing endometrial repair and causing HMB. Compared with adenomyosis-associated HMB, the mechanisms underlying adenomyosis-associated pain are less understood but presumably involve increased uterine contractility, hyperinnervation, increased lesional production of pain mediators, and central sensitization. Viewed through the prism of ReTIAR, a new imaging technique can be used to diagnose adenomyosis more accurately and informatively and possibly help to choose the best treatment modality.

ENDOCELL-Seud: a Delphi protocol to harmonise methods in endometrial cell culturing.

In vitro: culturing of endometrial cells obtained from the uterine mucosa or ectopic sites is used to study molecular and cellular signalling relevant to physiologic and pathologic reproductive conditions. However, the lack of consensus on standard operating procedures for deriving, characterising and maintaining primary cells in two- or three-dimensional cultures from eutopic or ectopic endometrium may be hindering progress in this area of research. Guidance for unbiased in vitro research methodologies in the field of reproductive science remains essential to increase confidence in the reliability of in vitro models. We present herein the protocol for a Delphi process to develop a consensus on in vitro methodologies using endometrial cells (ENDOCELL-Seud Project). A steering committee composed of leading scientists will select critical methodologies, topics and items that need to be harmonised and that will be included in a survey. An enlarged panel of experts (ENDOCELL-Seud Working Group) will be invited to participate in the survey and provide their ratings to the items to be harmonised. According to Delphi, an iterative investigation method will be adopted. Recommended measures will be finalised by the steering committee. The study received full ethical approval from the Ethical Committee of the Maastricht University (ref. FHML-REC/2021/103). The study findings will be available in both peer-reviewed articles and will also be disseminated to appropriate audiences at relevant conferences. Lay summary: Patient-derived cells cultured in the lab are simple and cost-effective methods used to study biological and dysfunctional or disease processes. These tools are frequently used in the field of reproductive medicine. However, the lack of clear recommendations and standardised methodology to guide the laboratory work of researchers can produce results that are not always reproducible and sometimes are incorrect. To remedy this situation, we define here a method to ascertain if researchers who routinely culture cells in the lab agree or disagree on the optimal laboratory techniques. This method will be used to make recommendations for future researchers working in the field of reproductive biology to reproducibly culture endometrial cells in the laboratory.