Endometrial-derived Stem Cells Research Articles

FGF2 promotes the proliferation of injured granulosa cells in premature ovarian failure via Hippo-YAP signaling pathway

Young women undergoing anticancer treatment are at risk of premature ovarian failure (POF). Endometrial-derived stem cells (EnSCs) have demonstrated significant therapeutic potential for treating ovarian insufficiency, although the underlying mechanisms remain to be fully understood. This study aims to further investigate the therapeutic effects of EnSCs, particularly through the paracrine action of fibroblast growth factor 2 (FGF2), on POF. The findings show that exogenous FGF2 enhances the survival of ovarian granulosa cells damaged by cisplatin. FGF2 stimulates the proliferation of these damaged cells by suppressing the Hippo signaling pathway and activating YAP expression. In vivo experiments also revealed that FGF2 treatment significantly improves ovarian reserve and endocrine function in mice with POF. These results suggest that FGF2 can boost the proliferative capacity of damaged ovarian granulosa cells through the Hippo-YAP signaling pathway, providing a theoretical foundation for using EnSCs and FGF2 in clinical treatments for POF.

Endometriotic mesenchymal stem cells promote the fibrosis process of endometriosis through paracrine TGF-β1 mediated RASAL1 inhibition.

Endometrial-derived stem cells are key players in endometriosis (EMs) pathogenesis, while the mechanism involved is still unclear. Herein, the role and regulatory mechanism of endometriotic mesenchymal stem cells (ecto-MSCs) in regulating fibrosis during EMs progression were investigated. The mRNA and protein expressions were assessed using qRT-PCR, western blot, and immunofluorescence. Flow cytometry was adopted to analyze the markers of MSCs. Transwell assay was adopted to examine endometriotic stromal cells (ESCs) migration and invasion. The interactions between DNMT3A and RASAL1 were analyzed by ChIP assay. In addition, MSP was employed to detect RASAL1 promoter methylation level. Ecto-MSCs promoted ESCs migration, invasion, and fibrosis process by TGF-β1 paracrine. It was subsequently revealed that TGF-β1 upregulated DNMT3A in ESCs in a SMAD3-dependent manner. As expected, DNMT3A knockdown abolished ecto-MSCs' facilitation on ESCs migration, invasion, and fibrosis process. DNMT3A, as a methyltransferase, reduced RASAL1 expression in TGF-β1-treated ESCs by increasing RASAL1 promoter methylation level. RASAL1, as an antifibrotic protein, was lowly expressed in TGF-β1-treated ESCs, and its overexpression ameliorated TGF-β1-induced increase in ESCs migration, invasion, and fibrosis process. TGF-β1 secreted by ecto-MSCs facilitated fibrogenesis in EMs through SMAD3/DNMT3A-mediated RASAL1 inhibition.

Artemisin and human endometrial-derived stem cells improve cognitive function and synaptic plasticity in a rat model of Alzheimer disease and diabetes.

Alzheimer disease (AD) is a common form of dementia associated with loss of memory and disruption of synaptic plasticity. There is a strong correlation between the pathophysiological features of AD and diabetes, including induction of oxidative stress, inflammation, and abnormality in blood vessels. Considering the brain's limited capacity to repair damage and the potential of stem cell-derived neural cells in the repair of neurodegenerative disease, we investigated the effects of artemisinin and TSP‑1‑human endometrial-derived-derived stem cells (TSP‑1‑hEDSCs) on the cognitive function and synaptic plasticity in AD-diabetes rats. The authors previously showed that artemisinin and TSP‑1‑hEDSCs suppressed oxidative stress and inflammation in AD-diabetes rats. Thrombospondins-1 (TSPs-1) is a glycoprotein that inhibits angiogenesis. AD and diabetes were induced using streptozotocin. Synaptic plasticity and learning and memory function were studied using the Morris water maze and electrophysiological test, respectively. Streptozotocin increased traveled swimming distance and escape latency in the morris water maze test, decreased the percent time spent in the target quadrant, inhibited the long-term potentiation (LTP), and increased the blood glucose levels. Simultaneous or separate administration of artemisinin and TSP‑1‑hEDSCs decreased the blood levels of glucose and improved cognitive tasks and synaptic plasticity by considerably reducing traveled swimming distance and escape latency, increasing the percent time spent in the target quadrant, and retrieval of the LTP; therefore, they could be utilized as an adjunct treatment for AD treatment. These results may be due to a decrease in oxidative stress and inflammation.

Stem Cell Therapy in Limb Ischemia: State-of-Art, Perspective, and Possible Impacts of Endometrial-Derived Stem Cells.

As an evidence-based performance, the rising incidence of various ischemic disorders has been observed across many nations. As a result, there is a growing need for the development of more effective regenerative approaches that could serve as main therapeutic strategies for the treatment of these diseases. From a cellular perspective, promoted complex inflammatory mechanisms, after inhibition of organ blood flow, can lead to cell death in all tissue types. In this case, using the stem cell technology provides a safe and regenerative approach for ischemic tissue revascularization and functional cell formation. Limb ischemia (LI) is one of the most frequent ischemic disease types and has been shown to have a promising regenerative response through stem cell therapy based on several clinical trials. Bone marrow-derived mononuclear cells (BM-MNCs), peripheral blood CD34-positive mononuclear cells (CD34+ PB-MNCs), mesenchymal stem cells (MSCs), and endothelial stem/progenitor cells (ESPCs) are the main, well-examined stem cell types in these studies. Additionally, our investigations reveal that endometrial tissue can be considered a suitable candidate for isolating new safe, effective, and feasible multipotent stem cells for limb regeneration. In addition to other teams’ results, our in-depth studies on endometrial-derived stem cells (EnSCs) have shown that these cells have translational potential for limb ischemia treatment. The EnSCs are able to generate diverse types of cells which are essential for limb reconstruction, including endothelial cells, smooth muscle cells, muscle cells, and even peripheral nervous system populations. Hence, the main object of this review is to present stem cell technology and evaluate its method of regeneration in ischemic limb tissue.

Effects of artemisinin and TSP-1-human endometrial-derived stem cells on a streptozocin-induced model of Alzheimer’s disease and diabetes in Wistar rats

Alzheimer's disease (AD) is an age‑associated dementia disorder characterized by Aβ plaques and neurofibrillary tangles. There is a strong link between cerebrovascular angiopathy, oxidative stress, inflammation, and glucose metabolism abnormalities with the development of AD. In this study, we investigated the therapeutic influences of artemisinin and TSP‑1‑human endometrial‑derived stem cells (TSP‑1‑hEDSCs) on the streptozocin‑induced model of AD and diabetes in rats. Hippocampal and intraperitoneal injections of streptozocin were used to induce AD and diabetes in male Wistar rats, followed by intranasal administration of a single dose of TSP‑1‑hEDSCs and intraperitoneal administration of artemisinin for 4 weeks. Hematoxylin together with eosin staining was performed for demonstrating Aβ plaque formation and for analyzing the influence of treatments on the pyramidal cells in the hippocampus. Biochemical analysis was used to assay the serum levels of glucose, MDA, ROS, and TAC. The expression of TNF‑α was measured using real‑time PCR. Streptozocin induced AD and diabetes via Aβ plaque formation and increasing blood glucose levels. It also increased the levels of ROS, MDA, and TNF‑α and decreased the levels of TAC. Simultaneous or separate administration of artemisinin and TSP‑1‑hEDSCs ameliorated this influence by considerably reducing Aβ plaque formation in the hippocampus, reducing glucose, MDA, ROS, and TNF‑α levels, and increasing TAC levels. It appears that artemisinin and TSP‑1‑hEDSCs improve the adverse features of AD in a rat model of AD and diabetes. Therefore, artemisinin and TSP‑1‑hEDSCs could be utilized as an adjunct treatment, as well as a protective agent, in AD patients.

MiRNA‑34a‑5p downregulation of VEGFA in endometrial stem cells contributes to the pathogenesis of endometriosis.

Endometrial-derived stem cells (EnSCs) serve an important role in the development of endometriosis via retrograde menstruation. Abnormal expression of miRNAs in EnSCs is involved in the etiology of endometriosis, however, the mechanisms remain unclear. The aim of the present study was to investigate the expression of miR‑34a‑5p and VEGFA in endometrial samples from patients with or without endometriosis, and then examine the underlying mechanism of microRNA‑34a‑5p regulation of VEGFA in EnSCs. Endometrial samples from patients with or without endometriosis were collected, and miR‑34a‑5p expression in the two groups was measured using RT‑PCR. Human endometrial‑derived stem cells (hEnSCs) were isolated from these endometrial samples, and hEnSCs were transfected with the miR‑34a‑5p mimics or control miRNAs. qPCR and western blotting were performed to assess the effects of miR‑34a‑5p on the expression of VEGFA in hEnSCs, and cell growth was assessed by an MTT assay. miR‑34a‑5p was significantly downregulated in patients with endometriosis when compared with that of those without endometriosis. VEGFA expression levels in hEnSCs with an overexpression of miR‑34a‑5p were significantly reduced when compared with those in the negative control (P<0.01). In addition, the upregulation of miR‑34a‑5p suppressed EnSCs proliferation by targeting the 3' untranslated region of VEGFA. miR‑34a‑5p provides a novel avenue for the understanding of the development of endometriosis, and may facilitate the development of potential therapeutics against endometriosis.

Enhanced expression of the stemness-related factors OCT4, SOX15 and TWIST1 in ectopic endometrium of endometriosis patients.

BackgroundCurrent evidence suggests that endometrial-derived stem cells, spilled in the peritoneal cavity via retrograde menstruation, are key players in the establishment of endometriotic lesions. The aim of this study was to determine the presence and distribution of the stemness-related factors OCT4, SOX15, TWIST1 and DCAMLK1 in women with and without endometriosis.MethodsImmunohistochemical analysis was used to determine stromal and epithelial expression of OCT4, SOX15, TWIST1 and DCAMLK1 in endometriosis patient (EP) endometrium (n = 69) and endometriotic tissue (n = 90) and in control endometrium (n = 50). Quantitative Real-Time PCR of OCT4, SOX15 TWIST1 and DCAMLK1 was performed in paired samples of EP endometrium and endometriotic tissue. Co-immunofluorescence staining was performed for OCT4 and SOX15. For statistical analyses we used unpaired t-test, Fisher combination test and Spearman test. For paired analyses, paired t-test and McNemar test were used.ResultsWe detected a significant correlation between the expression of the established stem cell marker OCT4 and the stemness-related markers SOX15 (p < 0.001) and TWIST1 (p = 0.002) but not DCAMLK1. We showed a colocalization of SOX15 and OCT4 in epithelial and stromal cells of endometriotic tissue by coimmunofluorescence. A concordant expression of OCT4 and SOX15 in the same sample was observed in epithelial cells of the endometriotic tissue (71.7%). The expression of stemness-related factors was not associated with proliferative or secretory phase of the menstrual cycle in endometriosis patients but was found to be differentially expressed during the menstrual cycle in the control group. Increased expression of epithelial OCT4, SOX15 and TWIST1 was detected in endometriotic tissue compared to EP endometrium in paired (p = 0.021, p < 0.001 and p < 0.001) and unpaired analysis (p = 0.040, p < 0.001 and p = 0.001).ConclusionOur findings support the hypothesis that upregulation of stem cell-related factors contribute to the establishment of endometriotic lesions.Trial registrationThe study was approved by the institutional review board (545/2010 on 6th of May 2014) of the Medical University of Vienna (http://ethikkommission.meduniwien.ac.at/fileadmin/ethik/media/dokumente/register/alle_2010.pdf).Electronic supplementary materialThe online version of this article (doi:10.1186/s12958-016-0215-4) contains supplementary material, which is available to authorized users.

Oleic acid promotes the expression of neural markers in differentiated human endometrial stem cells

Variety of neurodegenerative diseases in humans are caused by loss of cells along with loss of function and disability. Cell replacement therapy is a potential strategy to cure neurodegenerative diseases. Mesenchymal stem cells are pluripotent non-hematopoietic cells that can be isolated from numerous tissues. Human endometrial-derived stem cell (hEnSC) are the abundant and easy available source with no immunological response, for cell replacement therapy. In the nervous system, where fatty acids are found in huge amounts, they participate in its development and maintenance throughout life. Oleic acid is a kind of the saturated fatty acids which plays crucial role in brain development. Oleic acid released by astrocytes is used by neurons for the synthesis of phospholipids and is specifically incorporated into growth cones. Human endometrial-derived stem cells in the third passage were divided into 3 groups including: control, sham (cultured in full differentiation medium without oleic acid) and experimental group (cultured in full differentiation medium with oleic acid) to differentiate over a 18-day period. Data from Real-Time PCR showed that mRNA levels of NF and β-TUBULIN were increased significantly (p<0.05) in oleic acid treated cells in comparison to control and sham groups. Immunocytochemistry analysis of Chat and NF expression also showed the same results. The present study clearly demonstrates that oleic acid promotes neural differentiation of hEnSC through regulation of gene expression.

Local calcium signalling is mediated by mechanosensitive ion channels in mesenchymal stem cells

Mechanical forces are implicated in key physiological processes in stem cells, including proliferation, differentiation and lineage switching. To date, there is an evident lack of understanding of how external mechanical cues are coupled with calcium signalling in stem cells. Mechanical reactions are of particular interest in adult mesenchymal stem cells because of their promising potential for use in tissue remodelling and clinical therapy. Here, single channel patch-clamp technique was employed to search for cation channels involved in mechanosensitivity in mesenchymal endometrial-derived stem cells (hMESCs). Functional expression of native mechanosensitive stretch-activated channels (SACs) and calcium-sensitive potassium channels of different conductances in hMESCs was shown. Single current analysis of stretch-induced channel activity revealed functional coupling of SACs and BK channels in plasma membrane. The combination of cell-attached and inside-out experiments have indicated that highly localized Ca2+ entry via SACs triggers BK channel activity. At the same time, SK channels are not coupled with SACs despite of high calcium sensitivity as compared to BK. Our data demonstrate novel mechanism controlling BK channel activity in native cells. We conclude that SACs and BK channels are clusterized in functional mechanosensitive domains in the plasma membrane of hMESCs. Co-clustering of ion channels may significantly contribute to mechano-dependent calcium signalling in stem cells.

Human Endometrial Stem Cells May Differentiate into Schwann Cells in Fibrin Gel as 3D Culture

Damage in central nervous system plays an important role in biological life and causes severe paralysis of limbs and some organs. There are solutions to problems that can be a great revolution in the transplanted spinal cord and nerve injuries. Schwann cells (SCs) have important roles in development, myelination and regeneration in the peripheral nervous system. The applications of SCs in regenerative medicine are limited because of slow growth rate and difficulties in harvesting. Critical to the hypothesis is the experimental fact that human endometrial-derived stem cells (hEnSCs) as multipotent accessible source of cells are known as useful cell candidates in the field of nerve tissue engineering. We decided to use the three-dimensional culture of Schwann cells differentiated from endometrial stem cell in fibrin gel. In this study, we investigate the expression of differentiated Schwann cell markers by exposing of endometrial stem cells with induction media including FGF2/FSK/HRG/RA. Using immunocytochemistry, we show that differentiated cells express S100 and P75 markers. These results show that for the first time, human endometrial stem cells can be differentiated into Schwann cells in 2D and 3D culture. These novel differentiated cells in fibrin gel might open new opportunities for the management of cell survival and neurotrophic potential in tissue engineering approaches for nerve repair.

Menstrual blood-derived stromal stem cells from women with and without endometriosis reveal different phenotypic and functional characteristics.

Retrograde flow of menstrual blood cells during menstruation is considered as the dominant theory for the development of endometriosis. Moreover, current evidence suggests that endometrial-derived stem cells are key players in the pathogenesis of endometriosis. In particular, endometrial stromal stem cells have been suggested to be involved in the pathogenesis of this disease. Here, we aimed to use menstrual blood, as a novel source of endometrial stem cells, to investigate whether stromal stem cells from endometriosis (E-MenSCs) and non-endometriosis (NE-MenSCs) women differed regarding their morphology, CD marker expression pattern, proliferation, invasion and adhesion capacities and their ability to express certain immunomodulatory molecules. E-MenSCs were morphologically different from NE-MenSCs and showed higher expression of CD9, CD10 and CD29. Furthermore, E-MenSCs had higher proliferation and invasion potentials compared with NE-MenSCs. The amount of indoleamine 2,3-dioxygenase-1 (IDO1) and cyclooxygenase-2 (COX-2) in E-MenSCs co-cultured with allogenic peripheral blood mononuclear cells (PBMCs) was shown to be higher both at the gene and protein levels, and higher IDO1 activity was detected in the endometriosis group. However, NE-MenSCs revealed increased concentrations of forkhead transcription factor-3 (FOXP3) when compared with E-MenSCs. Nonetheless, interferon (IFN)-γ, Interleukin (IL)-10 and monocyte chemoattractant protein-1 (MCP-1) levels were higher in the supernatant of E-MenSCs-PBMC co-cultures. Here, we showed that there are inherent differences between E-MenSCs and NE-MenSCs. These findings propose the key role MenSCs could play in the pathogenesis of endometriosis and further support the retrograde and stem cell theories of endometriosis. Hence, considering its renewable and easily available nature, menstrual blood could be viewed as a reliable and inexpensive material for studies addressing the cellular and molecular aspects of endometriosis.

BMP-2 can promote the osteogenic differentiation of human endometrial stem cells

Abstract Background: Human endometrial-derived stem cells (hEnSCs) as multipotent accessible source of cells are known as useful cell candidates in the field of bone tissue engineering. However, the effect of bone morphogenic protein-2 (BMP-2) as an osteoinductive growth factor has not been clearly ascertained. Objective: To evaluate the effect of the remarkable osteoinductive growth factor BMP-2, on promotion of osteogenic differentiation in hEnSCs. Methods: Endometrial biopsies were obtained from healthy women referred to the hospital for infertility treatment. After tissue digestion in collagenase, the isolated endometrial cells were expanded in Dulbecco’s modified Eagle medium (DMEM) supplemented with 10% FBS. The propagated cells were characterized based on the expression of endometrial (CD90, CD105), endothelial (CD31), and hematopoietic (CD34, CD133) stem cell markers. Cells were differentiated in osteogenic medium containing DMEM supplemented with 10% FBS, 10 nM dexamethasone, 50 μg/ml Ascorbic acid, and 10 mM β-glycerophosphate in the presence or absence of BMP-2 for 21 days. Alizarin red staining was performed to verify the matrix mineralization. Immunocytochemical staining was conducted to detect the expression of OCT-4, CD133, and osteopontin as well as osteocalcin. The expression of osteoblast transcripts, including osteopontin, osteonectin, and alkaline phosphatase (ALP) were analyzed by semi quantitative PCR. Results: The expanded EnSCs were spindle shaped. They were positive for the expression of Oct-4, CD90, and CD105, while they were negative for endothelial and hematopoietic markers. The matrix mineralization was confirmed by Alizarin red in both groups at day 21. Although the expression of osteopontin and osteocalcin was detected in both groups by immunological staining, the expression of osteocalcin was more intense in the presence of BMP-2. ALP, Osteonectin and osteopontin transcripts were expressed in all groups; however, the expression of ALP and osteopontin was upregulated in the presence of BMP-2. Conclusion: BMP-2 as an osteoinductive growth factor, could promote the osteogenic differentiation of EnSCs in vitro.

Repair of critical size rat calvarial defects using endometrial-derived stem cells embedded within gelatin/apatite nanocomposite scaffold

Human stromal stem cells derived from endometrium (hESCs) are a type of multipotent stromal cells of the proven ability to differentiate into osteogenic lineage. Thus, it was suggested that these cells may be used to repair skeletal defects. In this study, Human ESCs were extracted from female endometrium and harvested. Biomimetic gelatin/apatite (Gel/Ap) scaffolds with and without harvested cells were implanted in a Critical size calvarial defects in the cranial bone of adult male rat. To CT-Scan and Histological studies were performed to investigate the level of bone formation after 8 weeks of surgery. Results confirmed that the treated defects with the bare and hESCs grafted Gel/Ap scaffold showed significant bone formation and maturation in comparison with the control group.

Induction of Insulin-Producing Cells Derived from Endometrial Mesenchymal Stem-like Cells

Studies have demonstrated that mesenchymal stem-like cells can be isolated from endometrium. However, the potential of endometrial-derived stem cells to differentiate into insulin-positive cells and functionally secrete insulin remains undetermined. We isolated endometrial mesenchymal stem-like cells (EMSCs) from human endometrial tissue from six donors. The insulin-secreting function of EMSCs was further analyzed in vitro and in transplanted grafts in vivo. We successfully isolated EMSCs from human endometrium, and our results showed that EMSCs expressed high levels of stemness genes (Nanog, Oct-4, Nestin). Under specific induction conditions for 2 weeks, EMSCs formed three-dimensional spheroid bodies (SBs) and secreted C-peptide. The high insulin content of SB-EMSCs was confirmed by enzyme-linked immunosorbent assay, and glucose responsiveness was demonstrated by measuring glucose-dependent insulin secretion. Using cDNA microarrays, we found that the expression profiles of SB-EMSCs are related to those of islet tissues. Insulin and C-peptide production in response to glucose was significantly higher in SB-EMSCs than in undifferentiated EMSC controls. Furthermore, upon differentiation, SB-EMSCs displayed increased mRNA expression levels of NKx2.2, Glut2, insulin, glucagon, and somatostatin. Our results also showed that SB-EMSCs were more resistant to oxidative damage and oxidative damage-induced apoptosis than fibroblasts from the same patient. It is noteworthy that SB-EMSCs xenotransplanted into immunocompromised mice with streptozotocin-induced diabetes restored blood insulin levels to control values and greatly prolonged the survival of graft cells. These data suggest that EMSCs not only play a novel role in the differentiation of pancreatic progenitors, but also can functionally enhance insulin production to restore the regulation of blood glucose levels in an in vivo transplantation model.

Is telomerase activity in cancer due to selection of stem cells and differentiation arrest?

Recent findings in stem cell biology have presented new perspectives and opportunities for the treatment of reproductive disease. In a departure from the long held dogma of embryologically fixed numbers of oocytes, current literature suggests that human ovaries contain stem cells which form new oocytes even in adulthood and that these stem cells can be cultured in vitro to develop into mature oocytes. These findings have provided new hope and broader options for fertility preservation. Evidence of endometrial regeneration by bone marrow stem cells in endometrial tissue of women who received bone marrow transplant highlight potential for the novel treatments of uterine disorders and supports new theories for the etiology of endometriosis – ectopic transdifferentiation of stem cells. Further, endometrial derived stem cells have been demonstrated to be useful in the treatment of several chronic and often debilitating diseases, including Parkinson's Disease and Diabetes. Other cells that may present future therapeutic benefits for a myriad of disease states include placental and fetal cells which enter maternal circulation during pregnancy and can later promote parenchymal regeneration in maternal tissue. These findings highlight novel functions of the uterus and ovaries. They demonstrate that the uterus is a dynamic organ permeable to fetal stem cells capable of transdifferentiation as well as a renewable source of multipotent stem cells. While we still have much to understand about stem cells, their potential applications in reproductive biology and medicine are countless.