Endometrial Mesenchymal Stem Cells Research Articles

Is Neonatal Uterine Bleeding Involved in Early-Onset Endometriosis?

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. We conducted a literature review of all relevant articles on Medline. 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). 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.

The pathogenesis of endometriosis and adenomyosis: insights from single-cell RNA sequencing†.

Endometriosis and adenomyosis are two similar gynecological diseases that are characterized by ectopic implantation and the growth of the endometrial tissue. Previous studies have reported that they share a common pathophysiology in some respects, such as a similar cellular composition and resistance to the progestogen of lesions, but their underlying mechanisms remain elusive. Emerging single-cell ribonucleic acid sequencing (scRNA-seq) technologies allow for the dissection of single-cell transcriptome mapping to reveal the etiology of diseases at the level of the individual cell. In this review, we summarized the published findings in research on scRNA-seq regarding the cellular components and molecular profiles of diverse lesions. They show that epithelial cell clusters may be the vital progenitors of endometriosis and adenomyosis. Subclusters of stromal cells, such as endometrial mesenchymal stem cells and fibroblasts, are also involved in the occurrence of endometriosis and adenomyosis, respectively. Moreover, CD8+ T cells, natural killer cells, and macrophages exhibit a deficiency in clearing the ectopic endometrial cells in the immune microenvironment of endometriosis. It seems that the immune responses are activated in adenomyosis. Understanding the immune characteristics of adenomyosis still needs further exploration. Finally, we discuss the application of findings from scRNA-seq for clinical diagnosis and treatment. This review provides fresh insights into the pathogenesis of endometriosis and adenomyosis as well as the therapeutic targets at the cellular level.

A Novel Conductive Polypyrrole-Chitosan Hydrogel Containing Human Endometrial Mesenchymal Stem Cell-Derived Exosomes Facilitated Sustained Release for Cardiac Repair.

Myocardial infarction (MI) results in cardiomyocyte necrosis and conductive system damage, leading to sudden cardiac death and heart failure. Studies have shown that conductive biomaterials can restore cardiac conduction, but cannot facilitate tissue regeneration. This study aims to add regenerative capabilities to the conductive biomaterial by incorporating human endometrial mesenchymal stem cell (hEMSC)-derived exosomes (hEMSC-Exo) into poly-pyrrole-chitosan (PPY-CHI), to yield an injectable hydrogel that can effectively treat MI. In vitro, PPY-CHI/hEMSC-Exo, compared to untreated controls, PPY-CHI, or hEMSC-Exo alone, alleviates H2O2-induced apoptosis and promotes tubule formation, while in vivo, PPY-CHI/hEMSC-Exo improves post-MI cardiac functioning, along with counteracting against ventricular remodeling and fibrosis. All these activities are facilitated via increased epidermal growth factor (EGF)/phosphoinositide 3-kinase (PI3K)/AKT signaling. Furthermore, the conductive properties of PPY-CHI/hEMSC-Exo are able to resynchronize cardiac electrical transmission to alleviate arrythmia. Overall, PPY-CHI/hEMSC-Exo synergistically combines the cardiac regenerative capabilities of hEMSC-Exo with the conductive properties of PPY-CHI to improve cardiac functioning, via promoting angiogenesis and inhibiting apoptosis, as well as resynchronizing electrical conduction, to ultimately enable more effective MI treatment. Therefore, incorporating exosomes into a conductive hydrogel provides dual benefits in terms of maintaining conductivity, along with facilitating long-term exosome release and sustained application of their beneficial effects.

Isolation and characterization of feline endometrial mesenchymal stem cells.

Recently, there has been a growing interest in stem cells for human medicine. Limited feline endometrial mesenchymal stem cell (fEM-MSC) research in veterinary medicine necessitates reporting for future feline disease research and therapy. This study aimed to isolate fEM-MSCs from feline endometrial tissues and evaluate their morphology, proliferative ability, differentiation ability, and immunophenotype. Feline endometrial tissues were obtained from the ovariohysterectomies of healthy cats and isolated using an enzymatic method. The morphology and proliferative ability of the isolated cells were assessed using a doubling time (DT) assay from passages 3 to 6 (P3 - P6). We measured pluripotency gene expressions of cells in P2 using quantitative real-time polymerase chain reaction (qRT-PCR). To investigate MSC characteristics, a trilineage differentiation assay was conducted in P4, and cells in P4 were immunophenotyped using flow cytometry. fEM-MSCs showed a typical spindle-shaped morphology under a microscope, and the DT was maintained from P3 to P6. fEM-MSCs could differentiate into adipocytes, osteoblasts, and chondrocytes, and expressed three pluripotency markers (OCT4, SOX2, and NANOG) by qRT-PCR. Immunophenotypic analysis showed that the fEM-MSCs were CD14 -, CD34 -, CD45 -, CD9+, and CD44+. In this study, the feline endometrium was a novel source of MSCs, and to the best of our knowledge, this is the first report on the isolation method and characteristics of fEM-MSCs.

The Response of the Cell Genome of Endometrial Mesenchymal Stem Cells to the Procedure of Long-Term Cyropreservation

The Response of the Cell Genome of Endometrial Mesenchymal Stem Cells to the Procedure of Long-Term Cyropreservation

The Biological Characteristics of Eutopic and Ectopic Endometrial Progenitor Cells in Endometriosis.

The aim of this study was to identify the biological characteristics and potential roles of endometrial progenitor cells in the pathogenesis of endometriosis. It is generally believed that progenitor cells in human endometrium are responsible for rapid endometrial regeneration. However, the biological characteristics and potential roles of the paired eutopic and ectopic endometrial progenitor cells in endometriosis remain unclear. This study intends to isolate the epithelial progenitor (EP) cells and endometrial mesenchymal stem cells (eMSCs) from the eutopic and ectopic endometria from endometriosis patients, further to reveal their features and functions respectively. The distributions of EP cells and eMSCs and the expression of steroid hormone receptors in the endometrium and endometriotic tissues were assessed by immunohistochemistry. EP cells and eMSCs were sorted from paired eutopic and ectopic endometria with epithelial cell adhesion molecule (EpCAM) magnetic beads. The clonogenicity, cell viability after being treated with estradiol and progesterone, and cell markers expression were evaluated with colony forming on Matrigel, CCK-8 and immunofluorescence staining, respectively. The differentially expressed genes (DEGs) were further identified with RNA sequencing. SSEA-1- and PDGFRβ-positive cells were distributed in the epithelial and stromal layers. The ERβ staining was much more intense in endometriotic tissues, but PR expression was almost absent. The ectopic EP cells exhibit strong clonogenicity and ERβ expression but weak PR expression, leading to progesterone resistance. There are 12604 and 13242 DEGs revealed by RNA sequencing between eutopic and ectopic EP cells or eMSCs. GO and KEGG analyses revealed that the functions and pathways of DEGs enriched in cellular energy metabolism and regulation of the immune response, respectively. Additionally, ERβ targets were mainly enriched in ectopic EP cells. Both EP cells and eMSCs may engage in ectopic lesion formation in endometriosis by modifying the metabolic mode and immune tolerance. These data not only help to understand the molecular mechanism of endometriosis but also could potentially contribute to the discovery of therapeutic targets for endometriosis.

Retraction Note: intranasal administration of endometrial mesenchymal stem cells as a suitable approach for Parkinson's disease therapy.

Retraction Note: intranasal administration of endometrial mesenchymal stem cells as a suitable approach for Parkinson's disease therapy.

Response of the Endometrial Mesenchymal Stem Cell Genome to the Procedure of Long-Term Cryopreservation

Response of the Endometrial Mesenchymal Stem Cell Genome to the Procedure of Long-Term Cryopreservation

P-791 Supplementation of mitochondria from endometrial mesenchymal stem cells (EnMSCs) improves oocyte quality in aged mice

Abstract Study question To improve the quality of aging oocytes by supplementing the mitochondria from endometrial mesenchymal stem cells (EnMSCs) via microinjection. Summary answer Supplementation with EnMSC-derived mitochondria can improve the quality of oocytes and promote embryo development in aging mice. What is known already Maternal aging is one of the major causes of reduced ovarian reserve and low oocyte quality in elderly women. Decreased oocyte quality is the main cause of age-related infertility. Mitochondria are multifunctional energy stations that determine the oocyte quality. The mitochondria in aging oocytes display functional impairments with mtDNA damage, which leads to reduced competence and developmental potential of oocytes. To improve oocyte quality, mitochondrial supplementation is carried out as a potential therapeutic approach. However, the selection of suitable cells as the source of mitochondria remains controversial. Study design, size, duration Study design: Eight-month-old ICR mice were purchased and then were further raised to ten months of age for use in the experiments. EnMSCs were stably cultivated and oocytes were extracted from ten-month-old ICR mice. Study size: About 300 GV oocytes and 134 MII oocytes were counted . 80 fertilized oocytes were transplanted into the fallopian tubes of surrogate mothers. Study duraton: About one half a year. Participants/materials, setting, methods Participants:ten-month-old ICR mice. Setting and methods: We cultivated EnMSCs from aged mice and extracted mitochondria from EnMSCs. And then GV oocytes were supplemented with mitochondria via microinjection. To further assess the effects of mitochondrial supplementation on embryo development, MII oocytes from aging mice were fertilized by ICSI combining EnMSCs mitochondria microinjection.To test whether mitochondrial supplementation could improve in vivo embryo development in aging mice, embryo transplantation was performed after mitochondrial microinjection combined with ICSI. Main results and the role of chance In this study, we found that the mitochondria derived from EnMSCs could significantly improve the quality of aging oocytes.Supplementation with EnMSC mitochondria significantly increased the blastocyst ratio of MII oocytes from aging mice after intracytoplasmic sperm injection (ICSI). We also found that the birth rate of mitochondria-injected aging oocytes was significantly increased after embryo transplantation. Limitations, reasons for caution Microinjection into oocytes was invasive and some oocytes died after microinjection. The ability of this method to improve oocyte quality was limited because the MII rate of oocytes after mitochondria supplementation was 35.65±1.75% while the rate of oocytes from 6–8 weeks of mice in a previous study was nearly 80%. Wider implications of the findings For clinical treatment, the EnMSCs can be obtained from the patient's own menstrual blood as the sources of mitochondrial supplementation. A noninvasive procedure for cell extraction ensures safety, and the findings of this work might provide a valuable strategy for patients with declining oocyte quality due to aging. Trial registration number Not applicable

O-189 Extracellular vesicles derived from human myometrial cells support endometrial mesenchymal stromal/stem cells

Abstract Study question Whether other forms of intercellular communication such as extracellular vesicles (EVs) participate in stem cell maintenance in the human endometrium. Summary answer The present study revealed the involvement of EVs as a novel intercellular communication method between endometrial mesenchymal stem cells (eMSC) and their surrounding niche. What is known already In human endometrium, many studies have demonstrated the importance of EVs in mediating various physiological as well as pathological processes. Our group has demonstrated that the myometrial cells are candidate niche cells of eMSC modulating their biological function. We also demonstrate the Notch signaling pathway is involved in endometrial stem cell regulation. Although classical Notch signaling relies on direct cell contact dependent interactions, this pathway can also be activated at a distance by EVs containing Notch ligands. Therefore, we hypothesized that certain Notch ligand(s) are packaged into the myometrial EVs to mediate stem cell functions. Study design, size, duration Sequential beading with magnetic beads coated with anti-CD140b and anti-CD146 antibodies was used to isolate eMSC (CD140b+CD146+ cells) from endometrial tissues from proliferative (n = 15) and secretory (n = 10) phase. Myometrial derived EVs (n = 17) were isolated by ultracentrifugation. The EVs were characterized by transmission electron microscopy and western blotting. Participants/materials, setting, methods Endometrial samples obtained from women aged 41 – 52 years undergoing total abdominal hysterectomy. EMSC were cocultured with myometrial EVs and the percentage of eMSC was analysed by flow cytometry. Blockage the secretion of EVs was performed by transfection of Rab27a siRNA. Western blot analysis and gene silencing approach validated the role of Notch signaling in eMSC. The therapeutic features of transplanted eMSC/myometrial EVs was determined using the mouse injured endometrial model. Main results and the role of chance EVs released from myometrial cells can be internalized by eMSC, leading to a significant effect on stemness of eMSC. Pharmacological inhibition of Notch signaling with DAPT or silencing of Notch 1 nullified these stimulatory effects. Myometrial EVs contains a high amount of the notch ligand – JAG1, thus inducing a strong Notch activity in eMSC. When JAG1 was silenced in the myometrial EVs, the self-renewal and clonogenic activity was reduced. Combined transplantation of eMSC with myometrial EVs improves the therapeutic effect of eMSC in endometrial regeneration in vivo. The observed therapeutic feature was potentially achieved by elevating the cell proliferation and suppressing apoptosis in the injured mouse endometrium. Limitations, reasons for caution Whether other signaling pathway are involved in the stimulatory effect of myometrial EVs in eMSC activities remains unknown. Also, EVs derived from other endometrial cell types i.e. epithelial cells may also regulate eMSC function. Wider implications of the findings Our findings revealed that myometrial EVs stimulated the biological function of eMSC via the Notch signaling. In the near future, it would be important to explore whether Notch signaling pathway plays a role in the injured mouse endometrium transplanted with eMSC/myometrial EVs. Trial registration number Not applicable

P-796 Single-cell transcriptomics unveils a role for leptin receptor in human endometrial mesenchymal stromal/stem cells

Abstract Study question What are the characteristics of leptin receptor (LepR) expressing cells within the endometrial mesenchymal stromal/stem cells (eMSCs) population? Summary answer LepR+ eMSCs display better stem-cell capacities as well as distinctively quiescent properties compared to eMSCs and LepR- eMSCs. What is known already Single-cell RNA sequencing (scRNA-seq) reveals that cultured eMSCs are heterogeneous. LepR is highly expressed in bone marrow MSCs, and LepR+ cells contribute towards the formation of fibroblast colony-forming units (CFU-F). Mouse LepR+ MSCs are quiescent and activate upon injuries to form osteoblasts and adipocytes in the bone marrow. In this study, we hypothesized that LepR+ cells play a critical role in the stem cell population in human endometrium. Study design, size, duration We performed scRNA-seq on magnetic bead selected eMSCs (CD140b+CD146+ cells). Functional stem cell assays such as clonogenicity and serial subcloning were performed on the different stromal subsets (eMSC, LepR- eMSC, and LepR+ eMSC) isolated by fluorescence-activated cell sorting (FACS) cultured endometrial stromal cells. The cell cycle analysis was evaluated by multicolor flow cytometry on freshly isolated endometrial stromal cells. Participants/materials, setting, methods The endometrial samples were collected from women aged 37–53 years with regular menstrual cycles undergoing hysterectomy. The expression and localization of LepR in human endometrium were determined by flow cytometry and immunohistochemistry, respectively. The in vitro colony-forming and self-renewal abilities were assessed on FACS sorted endometrial stromal populations. The cell cycle status of the stromal subsets was determined by flow cytometry and the quiescent property was determined by qPCR (RB1, RBL2, CDKN1A, CDKN1B, and E2F4). Main results and the role of chance Clustering analysis of the scRNA-seq data revealed 5 subpopulations (SP0-4) of cultured eMSCs. A high proportion (>90%) of the cells in SP3 and SP4 were at the G1 phase. The majority of the SP3 cells (92%) were located at the root of the cell trajectory and pseudo time inference analysis. The expression of LepR was highly specific in SP3 suggesting LepR eMSCs are potentially the origin progenitors.The expression of LepR in endometrial stromal cells was 2.61 ± 1.43%, and in eMSCs was 25.59 ± 19.04%. In vitro experiments demonstrated that LepR+ eMSCs exhibit better stem-cell properties, including clonogenic and self-renewal ability, than its negative counterparts. QPCR validation of cell-quiescence associated genes revealed higher expression of RB1, RBL2, and CDKN1B in the LepR+ eMSCs than eMSCs and LepR- eMSCs. In addition, multicolor flow analysis on freshly isolated endometrial cells revealed significantly higher proportion of LepR+ eMSCs are in the G0 phase of the cell cycle when compared to eMSCs and LepR- eMSCs. The above results indicate in vivo LepR+ eMSCs are relatively more quiescent. Limitations, reasons for caution The mechanism involved in maintaining or activating quiescent LepR+ eMSCs remains unknown. The role of LepR+ is likely to be different in human compared to mouse. We also lack an in vivo model for studying LepR+ eMSCs during endometrial repair. Wider implications of the findings Our findings extend the understanding of heterogeneous human endometrial mesenchymal stem cells and unveil a new role for LepR in endometrial regeneration. Trial registration number not applicatble

Endometrial and placental stem cells in successful and pathological pregnancies.

The endometrium is a dynamic tissue that undergoes extensive remodeling during the menstrual cycle and further gets modified during pregnancy. Different kinds of stem cells are reported in the endometrium. These include epithelial stem cells, endometrial mesenchymal stem cells, side population stem cells, and very small embryonic-like stem cells. Stem cells are also reported in the placenta which includes trophoblast stem cells, side population trophoblast stem cells, and placental mesenchymal stem cells. The endometrial and placental stem cells play a pivotal role in endometrial remodeling and placental vasculogenesis during pregnancy. The dysregulation of stem cell function is reported in various pregnancy complications like preeclampsia, fetal growth restriction, and preterm birth. However, the mechanisms by which it does so are yet elusive. Herein, we review the current knowledge of the different type of stem cells involved in pregnancy initiation and also highlight how their improper functionality leads to pathological pregnancy.

Is neonatal uterine bleeding responsible for early-onset endometriosis?

BackgroundIt 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.MethodsWe 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.ResultsImmunohistochemical 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.ConclusionsBased 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 registrationnot applicable.

A Preliminary Study on the Correlation Between Age and Endometrial Receptivity.

It is well established that female fertility declines with age, primarily because of loss of ovarian function. However, few studies have clarified the relationship between increasing age and endometrial receptivity. Here, we aimed to study the impact of age on endometrial receptivity, meanwhile, we examined the expression of endometrial mesenchymal stem cell (eMSC) surface markers (CD146 and PDGF-Rβ), essential for endometrial development and re-growth, in different age groups. Participants were enrolled in this study between October 2020 and July 2021. All 31 patients were divided into three age groups; early (30-39 years old, n=10), intermediate (40-49 years old, n=12) and advanced (≥50 years old, n=9). We assessed localization and expression of CD146 and PDGF-Rβ by immunofluorescence and further analyzed selected endometrial receptivity markers (Homeobox A10 HOXA10, leukemia inhibitory factor LIF and osteopontin) and steroid hormone receptors by immunohistochemistry. There were no significant differences in expression of HOXA10 and OPN (p>0.05) among the three groups. However, we found a significant difference in LIF expression between the early and advanced age groups, with higher expression noted in the latter group (p=0.02). Similarly, estrogen receptor (ER) and progesterone receptor (PR) expression were significantly increased (p=0.01 and p=0.01, respectively) in the advanced age group compared with the early age group. There were no significant difference in CD146 and PDGF-Rβ expression among the three groups (p>0.05). These results suggest that the age of the patient does not influence their endometrial receptivity. So, this study serves to increase our understanding of the impact of age and eMSCs on endometrial receptivity and expands the etiology of age-related infertility.

Subtle changes in perivascular endometrial mesenchymal stem cells after local endometrial injury in recurrent implantation failure

Improvements in reproductive techniques have resulted in the live birth rates from IVF procedures increasing from 5% to approximately 30% in recent decades but has plateaued since. Emerging preclinical and clinical data implicates endometrial receptivity deficiencies in patients with recurrent implantation failure (RIF) as the predominant factor hindering successful implantation. Mechanisms on how local endometrial injury (LEI) improves implantation rates in patients with RIF are currently unknown. We hypothesized that LEI may influence perivascular endometrial mesenchymal stem/progenitor cells (eMSCs) which are thought to regenerate the stromal vascular component of the functional layer every month. Here, we assessed the effect of LEI on the proportion and function of eMSCs present in consecutive LEI biopsies. Consecutive paired mid-luteal phase endometrial biopsies obtained from patients with RIF were digested to single cells and the proportion of SUSD2-expressing cells determined. Growth kinetics and decidualization were compared between the consecutive LEI samples. A mid-luteal LEI altered the decidualization capacity of SUSD2+ eMSCs in women with RIF, but not their proportion or clonogenicity. With the potential of LEI to improve IVF outcomes in women with RIF, additional investigations are needed to understand the impact of the altered decidualization response in eMSCs.

Multi-Lineage Human Endometrial Organoids on Acellular Amniotic Membrane for Endometrium Regeneration.

Asherman's syndrome is an endometrial regeneration disorder resulting from injury to the endometrial basal layer, causing the formation of scar tissue in the uterus and cervix. This usually leads to uterine infertility, menstrual disorders, and placental abnormalities. While stem cell therapy has shown extensive progress in repairing the damaged endometrium and preventing intrauterine adhesion, issues of low engraftment rates, rapid senescence, and the risk of tumorigenesis remain to be resolved for efficient and effective application of this technology in endometrial repair. This study addressed these challenges by developing a co-culture system to generate multi-lineage endometrial organoids (MLEOs) comprising endometrial epithelium organoids (EEOs) and endometrial mesenchymal stem cells (eMSCs). The efficacy of these MLEOs was investigated by seeding them on a biocompatible scaffold, the human acellular amniotic membrane (HAAM), to create a biological graft patch, which was subsequently transplanted into an injury model of the endometrium in rats. The results indicated that the MLEOs on the HAAM patch facilitated endometrial angiogenesis, regeneration, and improved pregnancy outcomes. The MLEOs on the HAAM patch could serve as a promising strategy for treating endometrial injury and preventing Asherman's syndrome.

Supplementation of mitochondria from endometrial mesenchymal stem cells improves oocyte quality in aged mice.

Maternal ageing is one of the major causes of reduced ovarian reserve and low oocyte quality in elderly women. Decreased oocyte quality is the main cause of age-related infertility. Mitochondria are multifunctional energy stations that determine the oocyte quality. The mitochondria in aged oocytes display functional impairments with mtDNA damage, which leads to reduced competence and developmental potential of oocytes. To improve oocyte quality, mitochondrial supplementation is carried out as a potential therapeutic approach. However, the selection of suitable cells as the source of mitochondria remains controversial. We cultivated endometrial mesenchymal stem cells (EnMSCs) from aged mice and extracted mitochondria from EnMSCs. To improve the quality of oocytes, GV oocytes were supplemented with mitochondria via microinjection. And MII oocytes from aged mice were fertilized by intracytoplasmic sperm injection (ICSI), combining EnMSCs' mitochondrial microinjection. In this study, we found that the mitochondria derived from EnMSCs could significantly improve the quality of aged oocytes. Supplementation with EnMSC mitochondria significantly increased the blastocyst ratio of MII oocytes from aged mice after ICSI. We also found that the birth rate of mitochondria-injected ageing oocytes was significantly increased after embryo transplantation. Our study demonstrates that supplementation with EnMSC-derived mitochondria can improve the quality of oocytes and promote embryo development in ageing mice, which might provide a prospective strategy for clinical treatment.

Differentiation of human endometrial stem cells encapsulated in alginate hydrogel into oocyte-like cells.

Human endometrial mesenchymal stem cells (hEnMSCs) are a rich source of mesenchymal stem cells (MSCs) with multi-lineage differentiation potential, making them an intriguing tool in regenerative medicine, particularly for the treatment of reproductive and infertility issues. The specific process of germline cell-derived stem cell differentiation remains unknown, the aim is to study novel ways to achieve an effective differentiation method that produces adequate and functioning human gamete cells. We adjusted the optimum retinoic acid (RA) concentration for enhancement of germ cell-derived hEnSCs generation in 2D cell culture after 7 days in this study. Subsequently, we developed a suitable oocyte-like cell induction media including RA and bone morphogenetic protein 4 (BMP4), and studied their effects on oocyte-like cell differentiation in 2D and 3D cell culture media utilizing cells encapsulated in alginate hydrogel. Our results from microscopy analysis, real-time PCR, and immunofluorescence tests revealed that 10 µM RA concentration was the optimal dose for inducing germ-like cells after 7 days. We examined the alginate hydrogel structural characteristics and integrity by rheology analysis and SEM microscope. We also demonstrated encapsulated cell viability and adhesion in the manufactured hydrogel. We propose that in 3D cell cultures in alginate hydrogel, an induction medium containing 10 µM RA and 50 ng/mL BMP4 can enhance hEnSC differentiation into oocyte-like cells. The production of oocyte-like cells using 3D alginate hydrogel may be viable in vitro approach for replacing gonad tissues and cells.

Endometrial Mesenchymal Stem Cells and Their Role in the Origin and Treatment of Endometriosis

Background: To identify endometrial mesenchymal stem cells (eMSCs) in retrograde menstruation, in various endometriosis lesions, in normal control tissues, and to investigate the association between eMSCs and endometriosis. We also plan to evaluate the effect of gonadotropin-releasing hormone agonists (GnRH-a) on eMSCs. Methods: Patients diagnosed with endometriosis were included if they had experienced surgery during the time frame 1 January 2015 to 31 December 2019 in West China Second Hospital, Sichuan University. Immunofluorescence was performed to identify eMSCs in those tissues with cell surface markers PDGFR-β/CD146. The percents of eMSCs in various tissues were calculated, and compared using analysis of variance. A two-sided pvalue less than 0.05 showed significant difference. Results: This study included 508 patients. eMSCs were identified in retrograde menstruation and numerous pathologic specimen but were not detected in normal control tissues. There was no significant difference in the percent of eMSCs between the GnRH-a treatment group and the control group (p &gt; 0.05). Conclusions: Our study demonstrated that eMSCs played a critical role in the development and recurrence of endometriosis and that GnRH-a did not affect eMSCs. Gynecologists should regard endometriosis as a chronic disease requiring lifetime management, especially for patients with chronic pelvic pain.

Metformin as a potential agent for modulating the faulty endometriotic mesenchymal stem cells: A case-control study.

According to stem cell theory, it seems that the proliferation/differentiation imbalance in endometrial mesenchymal stem cells (enMSCs) is the leading cause of endometriosis, so targeting them to modulate stemness-relevant factors seems to be a wise choice for endometriosis treatment. We aimed to investigate the effects of metformin on stemness properties of enMSCs by evaluating the expression profile of stemness-related genes and microRNAs (miRNAs). In this case-control study, MSCs were isolated from the eutopic endometrium of 3 endometriotic and 3 healthy women. After their characterization and culture, they were treated with 0.1, 1, and 10 mM metformin for 72 hr. Finally, the expression of octamer-binding transcription factor (OCT) 4A, OCT4B, OCT4B1, sex determining region Y-Box transcription factor 2, nanog homeobox, microRNA-200b, microRNA-145, and lethal-7b were analyzed by quantitative reverse transcription-polymerase chain reaction. Metformin modulated the expression of stemness-related genes and miRNAs, OCT4A, OCT4B, OCT4B1, sex determining region Y-Box transcription factor 2, nanog homeobox, microRNA-200b, microRNA-145, and lethal-7b in enMSCs, especially at 1 and 10 mM concentration. Notably, metformin had a paradoxical effect on normal enMSCs. We showed that metformin could modulate the expression of deregulated genes and miRNAs in faulty enMSCs, and restore their skewed self-renewal/differentiation balance, so it might be a promising drug for endometriosis treatment. The paradoxical effect of metformin on enMSCs and normal enMSCs might be because of their different metabolic patterns, so it requires further investigation to illustrate.