Search

Uterine leiomyomas are smooth-muscle tumors originating in the myometrium and are the most common pelvic tumors in women of reproductive age. Symptomatic tumors may result in abnormal uterine bleeding, bladder dysfunction, pelvic discomfort, and reproductive issues, such as infertility and miscarriage. There are currently few non-invasive treatments for leiomyoma, but there are no practical early intervention or preventive methods. In this study, human uterine leiomyoma and myometrial tissues were used to detect the protein and mRNA expression levels of UCHL1. To explore the effects of UCHL1 knockdown and inhibition in leiomyoma and myometrial cells, we determined the mRNA expressions of COL1A1 and COL3A1. Collagen gel contraction and wound-healing assays were performed on myometrial and leiomyoma cells. We found that UCHL1 expression was considerably higher in uterine leiomyomas than in the myometrium. COL1A1 and COL3A1 expression levels were downregulated after inhibition of UCHL1 in human leiomyoma cells. Furthermore, the elimination of UCHL1 significantly decreased the migration and contractility of leiomyoma cells. In conclusion, these results indicate that UCHL1 is involved in the growth of leiomyoma in humans. For the treatment of uterine leiomyoma, targeting UCHL1 activity may be a unique and possible therapeutic strategy.

Objective: To clarify whether a mediator complex subunit 12 (MED12) gain-of-function mutation induces leiomyoma cell properties in human uterine smooth muscle cells (USMCs)., Design: Experimental study., Setting: Academic research laboratory., Patient(s): Women undergoing hysterectomy for leiomyoma., Intervention(s): CRISPR/Cas9-mediated genome editing to introduce an MED12 gain-of-function mutation (G44D) into human USMCs., Main Outcome Measure(s): Cell proliferation, collagen production, and in vivo tumorigenicity of USMCs with vs. without the MED12 mutation., Result(s): Uterine smooth muscle cells isolated from the uterine myometrium of a 44-year-old patient were subjected to lentiviral vector-mediated gene transduction of the fluorescent protein Venus, followed by long-term passage. Uterine smooth muscle cells with a normal female karyotype, high cell proliferative activity, and Venus expression, but without stem/progenitor cell populations, were obtained and designated as USMC44. Using CRISPR/Cas9-mediated genome editing, mtUSMC44 (MED12, 131G>A, p.G44D) and mock USMC44 without MED12 mutation (wtUSMC44) were established from USMC44. wtUSMC44 and mtUSMC44 showed similar cell proliferation activity, even in the presence of estradiol and progesterone (EP) together with transforming growth factor-beta 3 (TGFB3). In addition, wtUSMC44 and mtUSMC44 generated similar tiny smooth muscle-like tissue constructs when xenotransplanted beneath the kidney capsule in immunodeficient mice treated with EP alone or TGFB3. In contrast, mtUSMC44 produced more collagen type I than wtUSMC in vitro, and this production was likely enhanced by EP and TGFB3., Conclusion(s): The results suggest that the MED12 gain-of-function mutation is involved in collagen production. Although approximately 70% of leiomyomas have MED12 mutations, additional factors and/or events other than MED12 and/or myometrial stem/progenitor cells may be required for fully inducing leiomyoma cell properties, including transformation, in USMCs., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Context: Uterine leiomyomas (fibroids) are the most common benign tumors in women. Recently, three populations of leiomyoma cells were discovered on the basis of CD34 and CD49b expression, but molecular differences between these populations remain unknown., Objective: To define differential gene expression and signaling pathways in leiomyoma cell populations., Design: Cells from human leiomyoma tissue were sorted by flow cytometry into three populations: CD34+/CD49b+, CD34+/CD49b-, and CD34-/CD49b-. Microarray gene expression profiling and pathway analysis were performed. To investigate the insulinlike growth factor (IGF) pathway, real-time quantitative polymerase chain reaction, immunoblotting, and 5-ethynyl-2'-deoxyuridine incorporation studies were performed in cells isolated from fresh leiomyoma., Setting: Research laboratory., Patients: Eight African American women., Interventions: None., Main Outcomes Measures: Gene expression patterns, cell proliferation, and differentiation., Results: A total of 1164 genes were differentially expressed in the three leiomyoma cell populations, suggesting a hierarchical differentiation order whereby CD34+/CD49b+ stem cells differentiate to CD34+/CD49b- intermediary cells, which then terminally differentiate to CD34-/CD49b- cells. Pathway analysis revealed differential expression of several IGF signaling pathway genes. IGF2 was overexpressed in CD34+/CD49b- vs CD34-/CD49b- cells (83-fold; P < 0.05). Insulin receptor A (IR-A) expression was higher and IGF1 receptor lower in CD34+/CD49b+ vs CD34-/CD49b- cells (15-fold and 0.35-fold, respectively; P < 0.05). IGF2 significantly increased cell number (1.4-fold; P < 0.001), proliferation indices, and extracellular signal-regulated kinase (ERK) phosphorylation. ERK inhibition decreased IGF2-stimulated cell proliferation., Conclusions: IGF2 and IR-A are important for leiomyoma stem cell proliferation and may represent paracrine signaling between leiomyoma cell types. Therapies targeting the IGF pathway should be investigated for both treatment and prevention of leiomyomas., (Copyright © 2017 by the Endocrine Society)

Uterine leiomyomas (fibroids) represent the most common class of benign tumors in women. Multiple leiomyomas usually arise from the uterus of a symptomatic woman. These tumors cause a variety of symptoms, including abnormal uterine bleeding, pelvic pain, bladder or bowel dysfunction, and recurrent pregnancy loss, and are responsible for more than 200,000 hysterectomies in the United States annually. Each leiomyoma seems to arise from the clonal expansion of a single myometrial smooth muscle cell transformed by a mutation. Tumor expansion is sustained by cell proliferation together with the production of large amounts of extracellular matrix. Estrogen and progesterone stimulate the growth of leiomyomas. Estrogen, together with its receptor ERα, enables progesterone action via induction of progesterone receptor (PR) expression. Progesterone induces the growth of leiomyoma by regulation of a set of key genes that control proliferation and apoptosis. A distinct cell population with stem-progenitor properties is indispensable for progesterone-dependent growth of leiomyomas. This stem-progenitor cell population is deficient in ERα and PR and dependent on the much higher levels of these steroid receptors in surrounding mature leiomyoma or myometrial cells. Progesterone sends paracrine signals from these mature cells to stem cells. The WNT/β-catenin pathway comprises a key component of this paracrine signaling system. The majority of medical treatments currently available for leiomyoma works by inhibiting estrogen or progesterone production or action, but tumors tend to regrow once treatment is stopped. Targeting stem cells and their paracrine interactions with more differentiated cell populations within leiomyoma may lead to the development of more effective therapeutics., (Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.)

Context: Uterine leiomyoma is the most common benign tumor in reproductive-age women. Using a dye-exclusion technique, we previously identified a side population of leiomyoma cells exhibiting stem cell characteristics. However, unless mixed with mature myometrial cells, these leiomyoma side population cells did not survive or grow well in vitro or in vivo., Objective: The objective of this study was to identify cell surface markers to isolate leiomyoma stem/progenitor cells., Design: Real-time PCR screening was used to identify cell surface markers preferentially expressed in leiomyoma side population cells. In vitro colony-formation assay and in vivo tumor-regeneration assay were used to demonstrate functions of leiomyoma stem/progenitor cells., Results: We found significantly elevated CD49b and CD34 gene expression in side population cells compared with main population cells. Leiomyoma cells were sorted into three populations based on the expression of CD34 and CD49b: CD34(+)/CD49b(+), CD34(+)/CD49b(-), and CD34(-)/CD49b(-) cells, with the majority of the side population cells residing in the CD34(+)/CD49b(+) fraction. Of these populations, CD34(+)/CD49b(+) cells expressed the lowest levels of estrogen receptor-α, progesterone receptor, and α-smooth muscle actin, but the highest levels of KLF4, NANOG, SOX2, and OCT4, confirming their more undifferentiated status. The stemness of CD34(+)/CD49b(+) cells was also demonstrated by their strongest in vitro colony-formation capacity and in vivo tumor-regeneration ability., Conclusions: CD34 and CD49b are cell surface markers that can be used to enrich a subpopulation of leiomyoma cells possessing stem/progenitor cell properties; this technique will accelerate efforts to develop new therapies for uterine leiomyoma.

Background: Uterine leiomyoma is the most common benign tumor in women and is thought to arise from the clonal expansion of a single myometrial smooth muscle cell transformed by a cellular insult. Leiomyomas cause a variety of symptoms, including abnormal uterine bleeding, pelvic pain, bladder or bowel dysfunction, and recurrent pregnancy loss, and are the most common indication for hysterectomy in the USA. A slow rate of cell proliferation, combined with the production of copious amounts of extracellular matrix, accounts for tumor expansion. A common salient feature of leiomyomas is their responsiveness to steroid hormones, thus providing an opportunity for intervention., Methods: A comprehensive search of PUBMED was conducted to identify peer-reviewed literature published since 1980 pertinent to the roles of steroid hormones and somatic stem cells in leiomyoma, including literature on therapeutics that target steroid hormone action in leiomyoma. Reviewed articles were restricted to English language only. Studies in both animals and humans were reviewed for the manuscript., Results: Estrogen stimulates the growth of leiomyomas, which are exposed to this hormone not only through ovarian steroidogenesis, but also through local conversion of androgens by aromatase within the tumors themselves. The primary action of estrogen, together with its receptor estrogen receptor α (ERα), is likely mediated via induction of progesterone receptor (PR) expression, thereby allowing leiomyoma responsiveness to progesterone. Progesterone has been shown to stimulate the growth of leiomyoma through a set of key genes that regulate both apoptosis and proliferation. Given these findings, aromatase inhibitors and antiprogestins have been developed for the treatment of leiomyoma, but neither treatment results in complete regression of leiomyoma, and tumors recur after treatment is stopped. Recently, distinct cell populations were discovered in leiomyomas; a small population showed stem-progenitor cell properties, and was found to be essential for ovarian steroid-dependent growth of leiomyomas. Interestingly, these stem-progenitor cells were deficient in ERα and PR and instead relied on the strikingly higher levels of these receptors in surrounding differentiated cells to mediate estrogen and progesterone action via paracrine signaling., Conclusions: It has been well established that estrogen and progesterone are involved in the proliferation and maintenance of uterine leiomyoma, and the majority of medical treatments currently available for leiomyoma work by inhibiting steroid hormone production or action. A pitfall of these therapeutics is that they decrease leiomyoma size, but do not completely eradicate them, and tumors tend to regrow once treatment is stopped. The recent discovery of stem cells and their paracrine interactions with more differentiated cell populations within leiomyoma has the potential to provide the missing link between developing therapeutics that temper leiomyoma growth and those that eradicate them., (© The Author 2014. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Tissue-specific (or somatic) stem cells constitute a subset of cells residing in normal adult tissues. By undergoing asymmetric division, they retain their ability to self-renew while producing daughter cells that go on to differentiate and play a role in tissue regeneration and repair. The human uterus consists primarily of endometrium and myometrium (the smooth muscle layer) that rapidly enlarges through its tremendous regenerative and remodeling capacity to accommodate the developing fetus. Such uterine enlargement and remodeling can take place repeatedly and cyclically over the course of a woman's reproductive life. These unique properties of the uterus suggest the existence of endometrial and myometrial stem cell systems. In addition, like somatic cells, tumor stem cells or tumor-initiating cells, a subset of cells within a tumor, retain the ability to reconstitute tumors. Uterine smooth muscle cells are thought to be the origin of leiomyomas that are the most common type of gynecologic tumor. Recent work has identified, isolated, and characterized putative stem/progenitor cells in the myometrium and in leiomyomas. Here, we review current studies of myometrial and leiomyoma stem/progenitor cells and provide a new paradigm for understanding myometrial physiology and pathology and how these cells might contribute to uterine remodeling during pregnancy and the formation of leiomyomas. The role of the WNT/CTNNB1 pathway in the pathogenesis of leiomyoma is also discussed., (© 2014 by the Society for the Study of Reproduction, Inc.)

Context: Uterine leiomyoma, or fibroids, represent the most common benign tumors of the female reproductive tract. A newly discovered epigenetic modification, 5-hydroxymethylation (5-hmC), and its regulators, the TET (Ten Eleven Translocation) enzymes, were implicated in the pathology of malignant tumors; however, their roles in benign tumors, including uterine fibroids, remain unknown., Objective: To determine the role of 5-hmC and TET proteins in the pathogenesis of leiomyoma using human uterine leiomyoma and normal matched myometrial tissues and primary cells., Design: 5-hmC levels were determined by ELISA and immunofluorescent staining in matched myometrial and leiomyoma tissues. TET expression was analyzed by quantitative RT-PCR and immunoblotting. TET1 or TET3 were silenced or inhibited by small interfering RNA or 2-hydroxyglutarate to study their effects on 5-hmC content and cell proliferation., Results: We demonstrated significantly higher 5-hmC levels in the genomic DNA of leiomyoma tissue compared to normal myometrial tissue. The increase in 5-hmC levels was associated with the up-regulation of TET1 or TET3 mRNA and protein expression in leiomyoma tissue. TET1 or TET3 knockdown significantly reduced 5-hmC levels in leiomyoma cells and decreased cell proliferation. Treatment with 2-hydroxyglutarate, a competitive TET enzyme inhibitor, significantly decreased both 5-hmC content and cell proliferation of leiomyoma cells., Conclusion: An epigenetic imbalance in the 5-hmC content of leiomyoma tissue, caused by up-regulation of the TET1 and TET3 enzymes, might lead to discovery of new therapeutic targets in leiomyoma.

Objective: To assess the effect of three WNT/β-catenin pathway inhibitors-inhibitor of β-catenin and TCF4 (ICAT), niclosamide, and XAV939-on the proliferation of primary cultures of human uterine leiomyoma cells., Design: Prospective study of human leiomyoma cells obtained from myomectomy or hysterectomy., Setting: University research laboratory., Patient(s): Women (n = 38) aged 27-53 years undergoing surgery., Intervention(s): Adenoviral ICAT overexpression or treatment with varying concentrations of niclosamide or XAV939., Main Outcome Measure(s): Cell proliferation, cell death, WNT/-catenin target gene expression or reporter gene regulation, β-catenin levels, and cellular localization., Result(s): Inhibitor of β-catenin and TCF4, niclosamide, or XAV939 inhibit WNT/β-catenin pathway activation and exert antiproliferative effects in primary cultures of human leiomyoma cells., Conclusion(s): Three WNT/-catenin pathway inhibitors specifically block human leiomyoma growth and proliferation, suggesting that the canonical WNT pathway may be a potential therapeutic target for the treatment of uterine leiomyoma. Our findings provide rationale for further preclinical and clinical evaluation of ICAT, niclosamide, and XAV939 as candidate antitumor agents for uterine leiomyoma., (Copyright © 2014 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

Uterine leiomyomas are extremely common estrogen and progesterone-dependent tumors of the myometrium and cause irregular uterine bleeding, severe anemia, and recurrent pregnancy loss in 15-30% of reproductive-age women. Each leiomyoma is thought to arise from a single mutated myometrial smooth muscle stem cell. Leiomyoma side-population (LMSP) cells comprising 1% of all tumor cells and displaying tumor-initiating stem cell characteristics are essential for estrogen- and progesterone-dependent in vivo growth of tumors, although they have remarkably lower estrogen/progesterone receptor levels than mature myometrial or leiomyoma cells. However, how estrogen/progesterone regulates the growth of LMSP cells via mature neighboring cells is unknown. Here, we demonstrate a critical paracrine role of the wingless-type (WNT)/β-catenin pathway in estrogen/progesterone-dependent tumorigenesis, involving LMSP and differentiated myometrial or leiomyoma cells. Estrogen/progesterone treatment of mature myometrial cells induced expression of WNT11 and WNT16, which remained constitutively elevated in leiomyoma tissues. In LMSP cells cocultured with mature myometrial cells, estrogen-progesterone selectively induced nuclear translocation of β-catenin and induced transcriptional activity of its heterodimeric partner T-cell factor and their target gene AXIN2, leading to the proliferation of LMSP cells. This effect could be blocked by a WNT antagonist. Ectopic expression of inhibitor of β-catenin and T-cell factor 4 in LMSP cells, but not in mature leiomyoma cells, blocked the estrogen/progesterone-dependent growth of human tumors in vivo. We uncovered a paracrine role of the WNT/β-catenin pathway that enables mature myometrial or leiomyoma cells to send mitogenic signals to neighboring tissue stem cells in response to estrogen and progesterone, leading to the growth of uterine leiomyomas.

The human uterus mainly consists of two layers: an inner endometrium and an outer layer, the myometrium, made of smooth muscle. The uterus is characterized by its unique capacity for regeneration. This capacity permits cyclical regeneration and remodeling of the tissue over the course of a woman's reproductive life. During each menstrual cycle, the endometrium regenerates, and the uterus enlarges to make room for fetal growth. This cyclic physiologic pattern suggests that myometrial stem/progenitor cells are present in the tissue and play a role in myometrial functions. Our group (and others) recently characterized and isolated putative stem/progenitor cells in the myometrium. These findings are permitting a better understanding of myometrial physiology and pathology. We review current studies of myometrial stem/progenitor cells and suggestions that, in combination with hypoxia, these cells may contribute to uterine remodeling during pregnancy and the formation of myomas., (Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.)

Background: Uterine leiomyoma is the most common benign tumor in reproductive-age women. Each leiomyoma is thought to be a benign monoclonal tumor arising from a single transformed myometrial smooth muscle cell; however, it is not known what leiomyoma cell type is responsible for tumor growth. Thus, we tested the hypothesis that a distinct stem/reservoir cell-enriched population, designated as the leiomyoma-derived side population (LMSP), is responsible for cell proliferation and tumor growth., Principal Findings: LMSP comprised approximately 1% of all leiomyoma and 2% of all myometrium-derived cells. All LMSP and leiomyoma-derived main population (LMMP) but none of the side or main population cells isolated from adjacent myometrium carried a mediator complex subunit 12 mutation, a genetic marker of neoplastic transformation. Messenger RNA levels for estrogen receptor-α, progesterone receptor and smooth muscle cell markers were barely detectable and significantly lower in the LMSP compared with the LMMP. LMSP alone did not attach or survive in monolayer culture in the presence or absence of estradiol and progestin, whereas LMMP readily grew under these conditions. LMSP did attach and survive when directly mixed with unsorted myometrial cells in monolayer culture. After resorting and reculturing, LMSP gained full potential of proliferation. Intriguingly, xenografts comprised of LMSP and unsorted myometrial smooth muscle cells grew into relatively large tumors (3.67 ± 1.07 mm(3)), whereas xenografts comprised of LMMP and unsorted myometrial smooth muscle cells produced smaller tumors (0.54 ± 0.20 mm(3), p<0.05, n = 10 paired patient samples). LMSP xenografts displayed significantly higher proliferative activity compared with LMMP xenografts (p<0.05)., Conclusions: Our data suggest that LMSP, which have stem/reservoir cell characteristics, are necessary for in vivo growth of leiomyoma xenograft tumors. Lower estrogen and progesterone receptor levels in LMSP suggests an indirect paracrine effect of steroid hormones on stem cells via the mature neighboring cells.