Intrauterine botulinum toxin A administration promotes endometrial regeneration mediated by IGFBP3-dependent OPN proteolytic cleavage in thin endometrium.

Adequate endometrial growth is a critical factor for successful embryo implantation and pregnancy maintenance. We previously reported the efficacy of intrauterine administration of botulinum toxin A (BoTA) in improving the endometrial angiogenesis and the rates of embryo implantation. Here, we further evaluated its potent therapeutic effects on the uterine structural and functional repair and elucidated underlying molecular regulatory mechanisms. This study demonstrated that a murine model of thin endometrium was successfully established by displaying dramatically decreased endometrial thickness and the rates of embryo implantation compared to normal endometrium. Interestingly, the expressions of insulin-like growth factor binding protein-3 (IGFBP3) and an active 35 kDa-form of osteopontin (OPN) were significantly reduced in thin endometrium, which were almost fully restored by intrauterine BoTA administration. Neutralization of BoTA-induced IGFBP3 subsequently suppressed proteolytic cleavage of OPN, exhibiting un-recovered endometrial thickness even in the presence of BoTA administration, suggesting that BoTA-induced endometrial regeneration might be mediated by IGFBP3-dependent OPN proteolytic cleavage. Our findings suggest that intrauterine BoTA administration improves the endometrial environment in our murine model with thin endometrium by increasing endometrial receptivity and angiogenesis in a manner dependent on the regulatory effect of IGFBP3 on OPN proteolytic cleavage, proposing BoTA as an efficient therapeutic strategy for the patients with thin endometrium.

Endometrial mesenchymal stromal/stem cells improve regeneration of injured endometrium in mice.

BACKGROUND: The monthly regeneration of human endometrial tissue is maintained by the presence of human endometrial mesenchymal stromal/stem cells (eMSC), a cell population co-expressing the perivascular markers CD140b and CD146. Endometrial regeneration is impaired in the presence of intrauterine adhesions, leading to infertility, recurrent pregnancy loss and placental abnormalities. Several types of somatic stem cells have been used to repair the damaged endometrium in animal models, reporting successful pregnancy. However, the ability of endometrial stem cells to repair the damaged endometrium remains unknown. METHODS: Electrocoagulation was applied to the left uterine horn of NOD/SCID mice causing endometrial injury. Human eMSC or PBS was then injected into the left injured horn while the right normal horn served as controls. Mice were sacrificed at different timepoints (Day 3, 7 and 14) and the endometrial morphological changes as well as the degree of endometrial injury and repair were observed by histological staining. Gene expression of various inflammatory markers was assessed using qPCR. The functionality of the repaired endometrium was evaluated by fertility test. RESULTS: Human eMSC successfully incorporated into the injured uterine horn, which displayed significant morphological restoration. Also, endometrium in the eMSC group showed better cell proliferation and glands formation than the PBS group. Although the number of blood vessels were similar between the two groups, gene expression of VEGF-α significantly increased in the eMSC group. Moreover, eMSC had a positive impact on the regeneration of both stromal and epithelial components of the mouse endometrium, indicated by significantly higher vimentin and CK19 protein expression. Reduced endometrial fibrosis and down-regulation of fibrosis markers were also observed in the eMSC group. The eMSC group had a significantly higher gene expression of anti-inflammatory factor Il-10 and lower mRNA level of pro-inflammatory factors Ifng and Il-2, indicating the role of eMSC in regulation of inflammatory reactions. The eMSC group showed higher implantation sites than the PBS group, suggesting better endometrial receptivity with the presence of newly emerged endometrial lining. CONCLUSIONS: Our findings suggest eMSC improves regeneration of injured endometrium in mice.

Bioactive Injectable and Self-Healing Hydrogel Via Cell-Free Fat Extract for Endometrial Regeneration.

The damaged endometrium and the formation of fibrosis are key barriers to pregnancy and further lead to infertility. However, how to promote endometrium repair is always a challenge. Here, a bioactive injectable and self-healing hydrogel is developed by physically combination of thiolated polyethylene (PEG), Cu2+ and cell-free fat extract (CEFFE, CF) for endometrial regeneration and fertility. By inheriting the advantages of various active proteins contained in CEFFE, it could induce the overall repair of endometrial microenvironment for intrauterine adhesion (IUA). In vitro, CF@Cu-PEG reduces endometrial cell apoptosis by more than 50%, and increases angiogenesis by 92.8%. In the IUA mouse, injection of CF@Cu-PEG significantly reduces the rate of uterine hydrometra and prevents the formation of endometrial fibrosis. Remarkably, CF@Cu-PEG contributes to the repair of endometrial microstructure, especially increases the number of endometrial pinopodes, significantly improves endometrial receptivity, and increases the pregnancy rate of IUA mice from 7.14% to 66.67%. In summary, through the physically combination of CEFFE and Cu-PEG, the construction of loaded bioactive injectable hydrogel not only inhibits the IUA, but also induces the self-repair of endometrial cells in situ and improves fertility, providing a new strategy for IUA repair in clinical application.

Melatonin-pretreated human umbilical cord mesenchymal stem cells improved endometrium regeneration and fertility recovery through macrophage immunomodulation in rats with intrauterine adhesions†.

Intrauterine adhesions (IUA) are a common gynecological problem. Stem cell therapy has been widely used in the treatment of IUA. However, due to the complex and harsh microenvironment of the uterine cavity, the effectiveness of such therapy is greatly inhibited. This study aimed to investigate whether melatonin pretreatment enhances the efficacy of human umbilical cord mesenchymal stem cells (HucMSCs) in IUA treatment in rats. First, we explored the effect of melatonin on the biological activity of HucMSCs in vitro through a macrophage co-culture system, Cell Counting Kit 8 (CCK-8), 5-Ethynyl-2'-deoxyuridine (EdU), flow cytometry, immunofluorescence staining, and qRT-PCR. Subsequently, we established the IUA rat model and tracked the distribution of HucMSCs in this model. In addition, we observed the number of M1 and M2 macrophages through immunofluorescence staining and detected the levels of inflammatory cytokines. Four weeks after cell transplantation, HE, Masson, and immunohistochemical staining were performed. In vitro experiments showed that melatonin pretreatment of HucMSCs promoted proliferation, reduced apoptosis, up-regulated the stemness gene, and regulated macrophage polarization. In vivo, melatonin pretreatment caused more HucMSCs to remain in the uterine cavity. Melatonin-pretreated HucMSCs recruited more macrophages, regulated macrophage polarization, and reduced inflammation. Melatonin-pretreated HucMSCs relieved fibrosis, increased endometrium thickness, and up-regulated CD34, vimentin, proliferating cell nuclear antigen (PCNA), and alpha small muscle antigen (α-SMA) expression. Fertility tests showed that melatonin-pretreated HucMSCs increased the number of embryos. In summary, pretreatment with melatonin was beneficial for HucMSC treatment because it enhanced the cell's ability to recruit macrophages and regulate macrophage polarization, which led to the regeneration of the endometrium and improved pregnancy outcomes.

A re-appraisal of mesenchymal-epithelial transition (MET) in endometrial epithelial remodeling.

Mesenchymal-epithelial transition (MET) is a mechanism of endometrial epithelial regeneration. It is also implicated in adenocarcinoma and endometriosis. Little is known about this process in normal uterine physiology. Previously, using pregnancy and menses-like mouse models, MET occurred only as an epithelial damage/repair mechanism. Here, we hypothesized that MET also occurs in other physiological endometrial remodeling events, outside of damage/repair, such as during the estrous cycle and adenogenesis (gland development). To investigate this, Amhr2-Cre-YFP/GFP mesenchyme-specific reporter mice were used to track the fate of mesenchymal-derived (MD) cells. Using EpCAM (epithelial marker), EpCAM+YFP+ MD-epithelial cells were identified in all stages of the estrous cycle except diestrus, in both postpartum and virgin mice. EpCAM+YFP+ MD-epithelial cells comprised up to 80% of the epithelia during estrogen-dominant proestrus and significantly declined to indistinguishable from control uteri in diestrus, suggesting MET is hormonally regulated. MD-epithelial cells were also identified during postnatal epithelial remodeling. MET occurred immediately after birth at postnatal day (P) 0.5 with EpCAM+GFP+ cells ranging from negligible (0.21%) to 82% of the epithelia. EpCAM+GFP+ MD-epithelial cells declined during initiation of adenogenesis (P8, avg. 1.75%) and then increased during gland morphogenesis (P14, avg. 10%). MD-epithelial cells expressed markers in common with non-MD-epithelial cells (e.g., EpCAM, FOXA2, ESR1, PGR). However, MD-epithelial cells were differentially regulated postnatally and in adults, suggesting a functional distinction in the two populations. We conclude that MET occurs not only as an epithelial damage/repair mechanism but also during other epithelial remodeling events, which to our knowledge has not been demonstrated in other tissues.

Exploration of eMSCs with HA-GEL system in repairing damaged endometrium after endometrial cancer with fertility-sparing treatment.

Despite the high complete response rate of fertility-sparing treatment in early-stage endometrial cancer (EC), the low pregnancy rate is a clinical challenge. Whether endometrium-derived mesenchymal stem cells (eMSCs) can repair damaged endometrium after EC reversal remains unclear. This study explored the potential therapeutic effects of eMSCs with suitable scaffold materials on endometrial damage caused by EC. Here, appropriate engineering scaffold materials were compared to identify the most suitable materials to carry eMSCs. Then, safety and efficacy evaluations of eMSCs with a suitable hyaluronic acid hydrogel (eMSCs/HA-GEL) were investigated in in vivo experiments with subcutaneous xenotransplantation in Balb/C nude mice and a model of endometrial mechanical injury in rats. HA-GEL has minimal cytotoxicity to eMSCs compared to other materials. Then, in vitro experiments demonstrate that eMSCs/HA-GEL enhance the inhibitory effects of progestins on EC cell biological behaviors. eMSCs/HA-GEL significantly inhibit EC cell growth and have no potential safety hazards of spontaneous tumorigenesis in Balb/C nude mouse subcutaneous xenotransplantation assays. eMSCs/HA-GEL intrauterine transplantation effectively increases endometrial thickness and glandular number, improves endometrial blood supply, reduces fibrotic areas, and improves pregnancy rates in a rat endometrial mechanical injury model. GFP-eMSCs/HA-GEL intrauterine transplantation in rats shows more GFP-eMSCs in the endometrium than GFP-eMSCs transplantation alone, and no tumor formation or suspicious cell nodules are found in the liver, kidney, or lung tissues. Our results reveal the safety and efficacy of eMSCs/HA-GEL in animal models and provide preliminary evidence for the use of eMSCs/HA-GEL as a treatment for EC-related endometrial damage.

Effect of Human Peripheral Blood Mononuclear Cells on Mouse Endometrial Cell Proliferation: A Potential Therapeutics for Endometrial Regeneration.

OBJECTIVES: The persistently thin endometrium is a major cause of repeated implantation failure; however, there is no definite treatment for it yet. This study aimed to confirm the potential of human peripheral blood mononuclear cells (hPBMCs) as a therapeutic agent for endometrial regeneration. DESIGN: An experimental study was carried out. PARTICIPANTS/MATERIALS, SETTING, METHODS: To assess the in vitro effect of hPBMC, the human primary endometrial epithelial cell lines SNU-685 and SNU-1077 were co-cultured with or without 1 × 105 hPBMCs for 24 h. To evaluate the in vivo effect, either 1 × 105 hPBMCs in PBS or PBS alone were injected into the left uterine horn of nonobese diabetic-severe combined immune-deficient mice, and the right untreated uterine horn was used as control. RESULTS: Co-culture with hPBMCs stimulated significant proliferation in both SNU-685 and SNU-1077 cell lines (p = 0.002 and 0.044, respectively). Moreover, treatment with hPBMCs significantly increased the thickness in all parts of the endometrium compared with that in the untreated control uterine horn (proximal: 1.69 ± 0.19 vs. 1.00 ± 0.10, p = 0.009; middle: 1.51 ± 0.14 vs. 1.00 ± 0.12, p = 0.010; distal: 1.72 ± 0.22 vs. 1.00 ± 0.12, p = 0.003, respectively). Compared with the PBS injection group, the hPBMC injection group had significantly thickened endometrium in the middle (p = 0.036) and distal segments (p = 0.002) of the uterine horn. Immunohistochemical analysis revealed the presence of exogenously injected hPBMCs in the uterus of recipient mice. hPBMC-recipient mice had cyclic uterus with normal histology in the endometrium. LIMITATIONS: hPBMCs were not applied directly to a mouse model with thin endometrium, so further study is needed. CONCLUSION: The beneficial effect of hPBMCs on endometrium may suggest their clinical feasibility for the safe treatment of infertile patients with persistently thin endometrium.

Strategies for managing Asherman's syndrome and endometrial atrophy: Since the classical experimental models to the new bioengineering approach.

Endometrial function is essential for embryo implantation and pregnancy, but managing endometrial thickness that is too thin to support pregnancy or an endometrium of compromised functionality due to intrauterine adhesions is an ongoing challenge in reproductive medicine. Here, we review current and emerging therapeutic and experimental options for endometrial regeneration with a focus on animal models used to study solutions for Asherman's syndrome and endometrial atrophy, which both involve a damaged endometrium. A review of existing literature was performed that confirmed the lack of consensus on endometrial therapeutic options, though promising new alternatives have emerged in recent years (platelet-rich plasma, exosomes derived from stem cells, bioengineering-based techniques, endometrial organoids, among others). In the future, basic research using established experimental models of endometrial pathologies (combined with new high-tech solutions) and human clinical trials with large population sizes are needed to evaluate these emerging and new endometrial therapies.

Adult stem cells in endometrial regeneration: Molecular insights and clinical applications.

Endometrial damage is an important cause of female reproductive problems, manifested as menstrual abnormalities, infertility, recurrent pregnancy loss, and other complications. These conditions are collectively termed "Asherman syndrome" (AS) and are typically associated with recurrent induced pregnancy terminations, repeated diagnostic curettage and intrauterine infections. Cancer treatment also has unexpected detrimental side effects on endometrial function in survivors independently of ovarian effects. Endometrial stem cells act in the regeneration of the endometrium and in repair through direct differentiation or paracrine effects. Nonendometrial adult stem cells, such as bone marrow-derived mesenchymal stem cells and umbilical cord-derived mesenchymal stem cells, with autologous and allogenic applications, can also repair injured endometrial tissue in animal models of AS and in human studies. However, there remains a lack of research on the repair of the damaged endometrium after the reversal of tumors, especially endometrial cancers. Here, we review the biological mechanisms of endometrial regeneration, and research progress and challenges for adult stem cell therapy for damaged endometrium, and discuss the potential applications of their use for endometrial repair after cancer remission, especially in endometrial cancers. Successful application of such cells will improve reproductive parameters in patients with AS or cancer. Significance: The endometrium is the fertile ground for embryos, but damage to the endometrium will greatly impair female fertility. Adult stem cells combined with tissue engineering scaffold materials or not have made great progress in repairing the injured endometrium due to benign lesions. However, due to the lack of research on the repair of the damaged endometrium caused by malignant tumors or tumor therapies, the safety and effectiveness of such stem cell-based therapies need to be further explored. This review focuses on the molecular insights and clinical application potential of adult stem cells in endometrial regeneration and discusses the possible challenges or difficulties that need to be overcome in stem cell-based therapies for tumor survivors. The development of adult stem cell-related new programs will help repair damaged endometrium safely and effectively and meet fertility needs in tumor survivors.

Stem cell paracrine actions in tissue regeneration and potential therapeutic effect in human endometrium: a retrospective study.

OBJECTIVE: Determining genetic and paracrine mechanisms behind endometrial regeneration in Asherman's syndrome and endometrial atrophy (AS/EA) patients after autologous CD133+ bone marrow-derived stem cell (CD133+ BMDSC) transplantation. DESIGN: Retrospective study using human endometrial biopsies and mouse models. SETTING: Fundación-IVI, IIS-La Fe, Valencia, Spain. SAMPLES: Endometrial biopsies collected before and after CD133+ BMDSC therapy, from eight women with AS/EA (NCT02144987) from the uterus of five mice with only left horns receiving CD133+ BMDSC therapy. METHODS: In human samples, haematoxylin and eosin (H&E) staining, RNA arrays, PCR validation, and neutrophil elastase (NE) immunohistochemistry (IHQ). In mouse samples, PCR validation and protein immunoarrays. MAIN OUTCOME MEASURES: H&E microscopic evaluation, RNA expression levels, PCR, and growth/angiogenic factors quantification, NE IHQ signal. RESULTS: Treatment improved endometrial morphology and thickness for all patients. In human samples, Jun, Serpine1, and Il4 were up-regulated whereas Ccnd1 and Cxcl8 were down-regulated after treatment. The significant decrease of NE signal corroborated Cxcl8 expression. Animal model analysis confirmed human results and revealed a higher expression of pro-angiogenic cytokines (IL18, HGF, MCP-1, MIP2) in treated uterine horns. CONCLUSIONS: CD133+ BMDSC seems to activate several factors through a paracrine mechanism to help tissue regeneration, modifying endometrial behaviour through an immunomodulatory milieu that precedes proliferation and angiogenic processes. Insight into these processes could bring us one step closer to a non-invasive treatment for AS/EA patients. TWEETABLE ABSTRACT: CD133+ BMDSC therapy regenerates endometrium, modifying the immunological milieu that precedes proliferation and angiogenesis.

An update on stem cell therapy for Asherman syndrome.

The current treatment for Asherman syndrome is limited and not very effective. The aim of this review is to summarize the most recent evidence for stem cells in the treatment of Asherman syndrome. The advent of stem cell therapy has propagated experimentation on mice and humans as a novel treatment. The consensus is that the regenerative capacity of stem cells has demonstrated improved outcomes in terms of fertility and fibrosis in both mice and humans with Asherman syndrome. Stem cells have effects on tissue repair by homing to the injured site, recruiting other cells by secreting chemokines, modulating the immune system, differentiating into other types of cells, proliferating into daughter cells, and potentially having antimicrobial activity. The studies reviewed examine different origins and administration modalities of stem cells. In preclinical models, therapeutic systemic injection of stem cells is more effective than direct intrauterine injection in regenerating the endometrium. In conjunction, bone marrow-derived stem cells have a stronger effect on uterine regeneration than uterine-derived stem cells, likely due to their broader differentiation potency. Clinical trials have demonstrated the initial safety and effectiveness profiles of menstrual, bone marrow, umbilical cord, and adipose tissue-derived stem cells in resumption of menstruation, fertility outcomes, and endometrial regeneration.

The role of mesenchymal-epithelial transition in endometrial function and receptivity.

OBJECTIVE: During reproductive age of a woman, endometrium undergoes frequent stuctural and functional changes. Abilities of regeneration, remodelation and differentiation are precondition of endometrial receptivity and implantation and development of an embryo. These processes are conditioned by mutual transformation between mesenchymal and epithelial fenotype of endometrial cells: epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET). The aim of this study is to present contemporary knowledge of transformation between epithelial and mesenchymal endometrial cells and its influence on human fertility. DESIGN: Review article. SETTING: Department of Obstetrics and Gynecology, Faculty of Medicine, Masaryk university and University Hospital Brno; Department of Obstetrics and Gynecology, University Hospital Faculty of Medicine, Palacky University, Olomouc. METHODS: PubMed was searched for articles in English indexed until February 2019 with terms of „endometrial receptivity“, „embryo implantation“, „endometrial regeneration“, „mesenchymal-epithelial transition/transformation“. RESULTS: It has been proved, that mesenchymal stromal cells participate on regeneration of not only the endometrial stroma, but also of the epithelium. During endometrial decidualisation under influence of ovarian steroids, the MET is under way. Stromal fibroblasts gain the morfological and functional properties of epithelial cells. During implantaion of an embryo, the trofoblast interacts with decidualised endometrium. Epithelial cells transform into mesenchymal (EMT), which mediate the growth of trofoblast. CONCLUSION: Mutual transformation between stromal and epithelial cells in essential for normal function of endometrium and implantation and development of an embryo.

Endometrial Stem Cell Markers: Current Concepts and Unresolved Questions.

The human endometrium is a highly regenerative organ undergoing over 400 cycles of shedding and regeneration over a woman's lifetime. Menstrual shedding and the subsequent repair of the functional layer of the endometrium is a process unique to humans and higher-order primates. This massive regenerative capacity is thought to have a stem cell basis, with human endometrial stromal stem cells having already been extensively studied. Studies on endometrial epithelial stem cells are sparse, and the current belief is that the endometrial epithelial stem cells reside in the terminal ends of the basalis glands at the endometrial/myometrial interface. Since almost all endometrial pathologies are thought to originate from aberrations in stem cells that regularly regenerate the functionalis layer, expansion of our current understanding of stem cells is necessary in order for curative treatment strategies to be developed. This review critically appraises the postulated markers in order to identify endometrial stem cells. It also examines the current evidence supporting the existence of epithelial stem cells in the human endometrium that are likely to be involved both in glandular regeneration and in the pathogenesis of endometrial proliferative diseases such as endometriosis and endometrial cancer.

Endometrial stem/progenitor cells.

Human endometrium regenerates and regresses with each menstrual cycle under hormonal control throughout a woman's reproductive life. The cyclical regeneration and remodeling potentials allude to the existence of stem/progenitor cells in the endometrium. There is increasing evidence that human endometrium contains small numbers of stem-like cells capable of self-renewal, multiple differentiation and tissue reconstitution. Although the precise identity of endometrial stem/progenitor cells remains elusive, these cells are thought to play pivotal role(s) in the physiological remodeling and regeneration of the human endometrium and also in the pathogenesis of endometrium-associated diseases, such as endometriosis.

The Endometrial Stem/Progenitor Cells and Their Niches.

Endometrial stem/progenitor cells are a type of stem cells with the ability to self-renew and differentiate into multiple cell types. They exist in the endometrium and form niches with their neighbor cells and extracellular matrix. The interaction between endometrial stem/progenitor cells and niches plays an important role in maintaining, repairing, and regenerating the endometrial structure and function. This review will discuss the characteristics and functions of endometrial stem/progenitor cells and their niches, the mechanisms of their interaction, and their roles in endometrial regeneration and diseases. Finally, the prospects for their applications will also be explored.

Restoration of Pregnancy Function Using a GT/PCL Biofilm in a Rabbit Model of Uterine Injury.

Biomaterial scaffolds have been used successfully to promote the regenerative repair of small endometrial lesions in small rodents, providing partial restoration of gestational function. The use of rabbits in this study allowed us to investigate a larger endometrial tissue defect and myometrial injury model. A gelatin/polycaprolactone (GT/PCL) gradient-layer biofilm was sutured at the defect to guide the reconstruction of the original tissue structure. Twenty-eight days postimplantation, the uterine cavity had been restored to its original morphology, endometrial growth was accompanied by the formation of glands and blood vessels, and the fragmented myofibers of the uterine smooth muscle had begun to resemble the normal structure of the lagomorph uterine cavity, arranging in a circular luminal pattern and a longitudinal serosal pattern. In addition, the repair site supported both embryonic implantation into the placenta and normal embryonic development. Four-dimensional label-free proteomic analysis identified the cell adhesion molecules, phagosome, ferroptosis, rap1 signaling pathways, hematopoietic cell lineage, complement and coagulation cascades, tricarboxylic acid cycle, carbon metabolism, and hypoxia inducible factor (HIF)-1 signaling pathways as important in the endogenous repair process of uterine tissue injury, and acetylation of protein modification sites upregulated these signaling pathways.

Nano-biomaterial Fibrinogen/P(LLA-CL) for prevention of intrauterine adhesion and restoration of fertility.

Endometrial damage resulting from surgical procedures is a significant cause of intrauterine adhesion, thin endometrium, and subsequent miscarriage and infertility. Unfortunately, there is currently no effective clinical solution to promote endometrial regeneration after severe injury. In this study, we combined fibrinogen (Fg) and P(LLA-CL) by electrostatic spinning to form a stable nano-biomaterial Fg/P(LLA-CL), which can promote endometrial regeneration. After inducing physical injury to rat endometrium, we found that Fg/P(LLA-CL) membranes placed in the uterine cavities increased endometrial thickness and the number of glands after injury, while reducing the area of endometrial fibrosis. In addition, Fg/P(LLA-CL) increased neovascularization and decreased COL1A1 deposition. The expression of TGF-ß1, a cytokine that promotes fibrosis, was down-regulated in the early stage of injury. Finally, fertility assays confirmed that Fg/P(LLA-CL) improved the pregnancy rate in rats with endometrial injury, and its safety was verified by blood tests and pathological examination of heart, liver, spleen, lung, and kidney. Therefore, Fg/P(LLA-CL) shows great potential as a safe and nontoxic biomaterial for endometrial regeneration, ultimately improving pregnancy outcomes in patients with intrauterine adhesion.

Effect of autologous bone marrow stem cells-scaffold transplantation on the ongoing pregnancy rate in intrauterine adhesion women: a randomized, controlled trial.

Intrauterine adhesion is a major cause of female reproductive disorders. Although we and others uncontrolled pilot studies showed that treatment with autologous bone marrow stem cells made a few patients with severe intrauterine adhesion obtain live birth, no large sample randomized controlled studies on this therapeutic strategy in such patients have been reported so far. To verify if the therapy of autologous bone marrow stem cells-scaffold is superior to traditional treatment in moderate to severe intrauterine adhesion patients in increasing their ongoing pregnancy rate, we conducted this randomized controlled clinical trial. Totally 195 participants with moderate to severe intrauterine adhesion were screened and 152 of them were randomly assigned in a 1:1 ratio to either group with autologous bone marrow stem cells-scaffold plus Foley balloon catheter or group with only Foley balloon catheter (control group) from February 2016 to January 2020. The per-protocol analysis included 140 participants: 72 in bone marrow stem cells-scaffold group and 68 in control group. The ongoing pregnancy occurred in 45/72 (62.5%) participants in the bone marrow stem cells-scaffold group which was significantly higher than that in the control group (28/68, 41.2%) (RR=1.52, 95%CI 1.08-2.12, P=0.012). The situation was similar in live birth rate (bone marrow stem cells-scaffold group 56.9% (41/72) vs. control group 38.2% (26/68), RR=1.49, 95%CI 1.04-2.14, P=0.027). Compared with control group, participants in bone marrow stem cells-scaffold group showed more menstrual blood volume in the 3rd and 6th cycles and maximal endometrial thickness in the 6th cycle after hysteroscopic adhesiolysis. The incidence of mild placenta accrete was increased in bone marrow stem cells-scaffold group and no severe adverse effects were observed. In conclusion, transplantation of bone marrow stem cells-scaffold into uterine cavities of the participants with moderate to severe intrauterine adhesion increased their ongoing pregnancy and live birth rates, and this therapy was relatively safe.

E2-Loaded Microcapsules and Bone Marrow-Derived Mesenchymal Stem Cells with Injectable Scaffolds for Endometrial Regeneration Application.

Bone marrow-derived mesenchymal stem cells (BMSCs) have been recognized as new candidates for the treatment of serious endometrial injuries. However, owing to the local microenvironment of damaged endometrium, transplantation of BMSCs yielded disappointing results. In this study, Pectin-Pluronic® F-127 hydrogel as scaffolds were fabricated to provide three-dimensional architecture for the attachment, growth, and migration of BMSCs. E2 was encapsulated into the W/O/W microspheres to construct pectin-based E2-loaded microcapsules (E2 MPs), which has the potential to serve as a long-term reliable source of E2 for endometrial regeneration. Then, the BMSCs/E2 MPs/scaffolds system was injected into the uterine cavity of mouse endometrial injury model for treatment. At 4 weeks after transplantation, the system increased proliferative abilities of uterine endometrial cells, facilitated microvasculature regeneration, and restored the ability of endometrium to receive an embryo, suggesting that the BMSCs/E2 MPs/scaffolds system is a promising treatment option for endometrial regeneration. Furthermore, the mechanism of E2 in promoting the repair of endometrial injury was also investigated. Exosomes are critical paracrine mediators that act as biochemical cues to direct stem cell differentiation. In this study, it was found that the expression of endometrial epithelial cell (EEC) markers was upregulated in BMSCs treated by exosomes secreted from endometrial stromal cells (ESCs-Exos). Exosomes derived from E2-stimulated ESCs further promoted the expression level of EECs markers in BMSCs, suggesting exosomes released from ESCs by E2 stimulation could enhance the differentiation efficiency of BMSCs. Therefore, exosomes derived from ESCs play paracrine roles in endometrial regeneration stimulated by E2 and provide optimal estrogenic response.

Minimally invasive delivery of human umbilical cord-derived mesenchymal stem cells by an injectable hydrogel via Diels-Alder click reaction for the treatment of intrauterine adhesions.

Intrauterine adhesions (IUA) are the most common cause of uterine infertility, and conventional treatments have not consistently achieved satisfactory pregnancy rates. Stem cell therapy shows promising potential for the clinical treatment of IUA. Although various advanced biomaterials have been designed for delivering stem cells to the uterine cavity, there remain significant challenges, particularly in devising therapeutic strategies for clinical application that minimize surgical incisions and conform to the intricate structure of uterine cavity. Herein, an injectable hydrogel loaded with human umbilical cord-derived mesenchymal stem cells (UCMSCs) was synthesized via the Diels-Alder click reaction for endometrial regeneration and fertility restoration, exhibiting suitable mechanical properties, good biocompatibility, and desirable degradation properties. Notably, this hydrogel permitted minimally invasive administration and integrated seamlessly with surrounding tissue. Our study revealed that the UCMSCs-laden injectable hydrogel enhanced cell proliferation, migration, angiogenesis, and exhibited anti-fibrotic effects in vitro. The implantation of this hydrogel significantly facilitated endometrium regeneration and restored fertility in a rat endometrial damage model. Mechanistically, in vivo results indicated that the UCMSCs-laden injectable hydrogel effectively promoted macrophage recruitment and facilitated M2 phenotype polarization. Collectively, this hydrogel demonstrated efficacy in regenerating damaged endometrium, leading to the restoration of fertility. Consequently, it holds promise as a potential therapeutic strategy for endometrial damage and fertility decline arising from intrauterine adhesions. STATEMENT OF SIGNIFICANCE: Severe endometrial traumas frequently lead to intrauterine adhesions and subsequent infertility. Stem cell therapy shows promising potential for the clinical treatment of IUA; however, challenges remain, including low delivery efficiency and compromised stem cell activity during the delivery process. In this study, we fabricated an injectable hydrogel loaded with UCMSCs via the Diels-Alder click reaction, which exhibited unique bioorthogonality. The in situ-gelling hydrogels could be introduced through a minimally invasive procedure and adapt to the intricate anatomy of the uterus. The UCMSCs-laden injectable hydrogel promoted endometrial regeneration and fertility restoration in a rat endometrial damage model, efficaciously augmenting macrophage recruitment and promoting their polarization to the M2 phenotype. The administration of UCMSCs-laden injectable hydrogel presents a promising therapeutic strategy for patients with severe intrauterine adhesion.