Embryonic Mesenchymal Stem Cells Research Articles

Stem cell therapy for interstitial cystitis/bladder pain syndrome.

Interstitial cystitis/bladder pain syndrome (IC/BPS) is a chronic disease with limited treatment options. Current multidisciplinary approach targeting bladder inflammation and urothelial dysfunction has limited durable effect that major surgery is ultimately required for both Hunner and non-Hunner type IC. Various investigational attempts are underway to avoid such operations and preserve the urinary bladder. Stem cell therapy is a fascinating option for treating chronic illnesses. Stem cells can self-renew, restore damaged tissue, and have paracrine effects. The therapeutic efficacy and safety of stem cell therapy have been demonstrated in numerous preclinical models, primarily chemically induced cystitis rat models. Only one clinical trial (phase 1 study) has investigated the safety of human embryonic stem cell-derived mesenchymal stem cells in three Hunner-type IC patients. Under general anesthesia, participants underwent cystoscopic submucosal stem cell injection (2.0 × 107 stem cells/5 mL). No safety issues were reported up to 12 months of follow-up and long-term follow-up (up to 3 years). Although there were variations in therapeutic response, all patients reported significant improvement in pain at 1 month postoperatively. One patient underwent fulguration of the Hunner lesion after the trial, but others reported an overall improvement in pain. The analysis on phase 1/2a trial which had several modifications in protocol is currently ongoing. Despite several limitations that need to be overcome, stem cell therapy could be a potential therapeutic option for treating IC/BPS. Clinical outcome on phase 1/2a trial is important and might provide more insight into the clinical application of stem cell therapy for IC/BPS.

THU-230 Human embryonic stem cell-derived mesenchymal stem cells improve mitochondrial oxidative dysfunction in metabolic dysfunction-associated liver disease via the AMPK pathway

THU-230 Human embryonic stem cell-derived mesenchymal stem cells improve mitochondrial oxidative dysfunction in metabolic dysfunction-associated liver disease via the AMPK pathway

CpG methylation changes in human mesenchymal and neural stem cells in response to in vitro niche modifications

Stem cell therapies hold promise in addressing the burden of neurodegenerative diseases with human embryonic neural stem cells (hNSC-H9s) and bone marrow-derived human mesenchymal stem cells (hMSCs) as viable candidates. The induction of hMSC neurospheres (hMSC-IN) generate a more lineage-restricted common neural progenitor-like cell population, potentially tunable by heparan sulfate proteoglycans (HSPGs). We examined CpG (5 mC) site methylation patterns using Illumina Infinium 850 K EPIC arrays in hNSC-H9, hMSCs and hMSC-IN cultures with HSPG agonist heparin at early and late phases of growth. We identified key regulatory CpG sites in syndecans (SDC2; SDC4) that potentially regulate gene expression in monolayers. Unique hMSC-IN hypomethylation in glypicans (GPC3; GPC4) underscore their significance in neural lineages with Sulfatase 1 and 2 (SULF1 &2) CpG methylation changes potentially driving the neurogenic shift. hMSC-INs methylation levels at SULF1 CpG sites and SULF2:cg25401628 were more closely aligned with hNSC-H9 cells than with hMSCs. We further suggest SOX2 regulation governed by lncSOX2-Overall Transcript (lncSOX2-OT) methylation changes with preferential activation of ENO2 over other neuronal markers within hMSC-INs. Our findings illuminate epigenetic dynamics governing neural lineage commitment of hMSC-INs offering insights for targeted mechanisms for regenerative medicine and therapeutic strategies.

The use of mesenchymal stem cells in tendon regeneration in rats.

Mesenchymal stem cells are of great interest due to their regenerative potential and capacity to transdifferentiate into various cell lineages. The present study executed a narrative review of the literature on the influence of mesenchymal stem cells on the regeneration of tendon injuries. To acomplish this, a search was carried out in the Pubmed and Scielo databases using descriptors such as: Mesenchymal stem cell regeneration and application tendon rat; Embryonic Stem Cell-Derived Mesenchymal Stem Cells for Tendon Tissue rat. Only research articles carried out on rats between 2011 and 2023 were included. 935 articles were found in the Pubmed database, and 43 articles were included based on the criteria of categorization, analysis of titles and abstracts. The articles showed benefits in relation to the use of stem cells in rat tendon injuries, but some studies showed that this repair was not 100% guaranteed to the state of the tendon originally, but associated with growth factors obtained a better induction of tenogenesis.

Cell Reprogramming Strategies for Treating Osteoarthritis and Intervertebral Disc Degeneration.

Osteoarthritis (OA) and intervertebral disc degeneration (IVDD) are the most common degenerative bone and joint diseases, posing a major threat to patients' physical and mental health due to the occurrence of chronic pain and disability. Within this context, the absence of efficacious therapies has led to a growing interest in regenerative medicine. In particular, as a method that can erase the memory of differentiation and re-endow cells with pluripotency, cell reprogramming technologies have ushered in a new era of personalized therapy, which not only show great potential for the treatment of degenerative osteoarthropathies but also promise to achieve tissue regenerative and repair. However, compared to other areas of research, reprogramming technologies to treat OA and IVDD are still in the preliminary stages and require further investigation. This paper briefly introduces the characteristics of cell reprogramming; summarizes the pathological mechanisms of reprogramming to improves energy metabolism, aging, inflammation, oxidative stress, and immune imbalance in OA and IVDD under the background of microenvironment and immunity; highlights the significant advantages of reprogramming-derived cells compared to embryonic stem cells and mesenchymal stem cells, based on these advances, providing important strategies for its development and clinical application in OA and IVDD.

Extracellular vesicles derived from human ESC–MSCs target macrophage and promote anti-inflammation process, angiogenesis, and functional recovery in ACS-induced severe skeletal muscle injury

BackgroundAcute compartment syndrome (ACS) is one of the most common complications of musculoskeletal injury, leading to the necrosis and demise of skeletal muscle cells. Our previous study showed that embryonic stem cells-derived mesenchymal stem cells (ESC–MSCs) are novel therapeutics in ACS treatment. As extracellular vesicles (EVs) are rapidly gaining attention as cell-free therapeutics that have advantages over parental stem cells, the therapeutic potential and mechanisms of EVs from ESC–MSCs on ACS need to be explored.MethodIn the present study, we examined the protective effects in the experimental ACS rat model and investigated the role of macrophages in mediating these effects. Next, we used transcriptome sequencing to explore the mechanisms by which ESC–MSC-EVs regulate macrophage polarization. Furthermore, miRNA sequencing was performed on ESC–MSC-EVs to identify miRNA candidates associated with macrophage polarization.ResultsWe found that intravenous administration of ESC–MSC-EVs, given at the time of fasciotomy, significantly promotes the anti-inflammation process, angiogenesis, and functional recovery of muscle in ACS. The beneficial effects were associated with ESC–MSC-EVs affecting macrophage polarization by delivering various miRNAs which regulate NF-κB, JAK/STAT, and PI3K/AKT pathways. Our data further illustrate that ESC–MSC-EVs mainly modulate macrophage polarization via the miR-21/PTEN, miR-320a/PTEN, miR-423/NLRP3, miR-100/mTOR, and miR-26a/TLR3 axes.ConclusionTogether, our results demonstrated the beneficial effects of ESC–MSC-EVs in ACS, wherein the miRNAs present in ESC–MSC-EVs regulate the polarization of macrophages.

Mesenchymal stem cell-derived exosomes attenuate DNA damage response induced by cisplatin and bleomycin.

Stem cell-derived exosomes (SC-Exos) have been shown to protect cells from chemical-induced deoxyribonucleic acid (DNA) damage. However, there has been no systematic comparison of the efficacy of exosomes against different types of DNA damage. Therefore, in this study, we assessed the protective effect of exosomes derived from human embryonic stem cell-induced mesenchymal stem cells (hESC-MSC-Exos) on two types of DNA damage, namely, intra-/inter-strand crosslinks and DNA double-strand breaks induced by cisplatin (Pt) and bleomycin (BLM), respectively, in HeLa cells. The alkaline comet assay demonstrated that hESC-MSC-Exos effectively inhibited Pt- and BLM-induced DNA damage in a dose-dependent manner. When the concentration of hESC-MSC-Exos reaches 2.0×106 and 4.0×106 particles/mL in Pt- and BLM-treated groups, respectively, there was a significant decrease in tail DNA percentage (Pt: 20.80±1.61 vs 9.40±1.14, p<0.01; BLM: 21.80±1.31 vs 6.70±0.60, p<0.01), tail moment (Pt: 10.00±1.21 vs 2.08±0.51, p<0.01; BLM: 12.00±0.81 vs 2.00±0.21, p<0.01), and olive tail moment (Pt: 6.01±0.55 vs 2.09±0.25, p<0.01; BLM: 6.03±0.37 vs 1.53±0.13, p<0.01). Phospho-histone H2AX (γH2AX) immunofluorescence and western blotting showed an over 50 % decrease in γH2AX expression when the cells were pretreated with hESC-MSC-Exos. As reactive oxygen species (ROS) are important mediators of Pt- and BLM-induced DNA damage, dichloro-dihydro-fluorescein diacetate staining indicated that hESC-MSC-Exos inhibited the increase in intracellular ROS in drug-treated cells. In conclusion, our findings suggest that hESC-MSC-Exos can protect cells from the two types of DNA-damaging drugs and that reduced intracellular ROS is involved in this effect.

A tissue engineered 3D printed calcium alkali phosphate bioceramic bone graft enables vascularization and regeneration of critical-size discontinuity bony defects in vivo.

Introduction: Recently, efforts towards the development of patient-specific 3D printed scaffolds for bone tissue engineering from bioactive ceramics have continuously intensified. For reconstruction of segmental defects after subtotal mandibulectomy a suitable tissue engineered bioceramic bone graft needs to be endowed with homogenously distributed osteoblasts in order to mimic the advantageous features of vascularized autologous fibula grafts, which represent the standard of care, contain osteogenic cells and are transplanted with the respective blood vessel. Consequently, inducing vascularization early on is pivotal for bone tissue engineering. The current study explored an advanced bone tissue engineering approach combining an advanced 3D printing technique for bioactive resorbable ceramic scaffolds with a perfusion cell culture technique for pre-colonization with mesenchymal stem cells, and with an intrinsic angiogenesis technique for regenerating critical size, segmental discontinuity defects in vivo applying a rat model. To this end, the effect of differing Si-CAOP (silica containing calcium alkali orthophosphate) scaffold microarchitecture arising from 3D powder bed printing (RP) or the Schwarzwalder Somers (SSM) replica fabrication technique on vascularization and bone regeneration was analyzed in vivo. In 80 rats 6-mm segmental discontinuity defects were created in the left femur. Methods: Embryonic mesenchymal stem cells were cultured on RP and SSM scaffolds for 7d under perfusion to create Si-CAOP grafts with terminally differentiated osteoblasts and mineralizing bone matrix. These scaffolds were implanted into the segmental defects in combination with an arteriovenous bundle (AVB). Native scaffolds without cells or AVB served as controls. After 3 and 6months, femurs were processed for angio-µCT or hard tissue histology, histomorphometric and immunohistochemical analysis of angiogenic and osteogenic marker expression. Results: At 3 and 6months, defects reconstructed with RP scaffolds, cells and AVB displayed a statistically significant higher bone area fraction, blood vessel volume%, blood vessel surface/volume, blood vessel thickness, density and linear density than defects treated with the other scaffold configurations. Discussion: Taken together, this study demonstrated that the AVB technique is well suited for inducing adequate vascularization of the tissue engineered scaffold graft in segmental defects after 3 and 6months, and that our tissue engineering approach employing 3D powder bed printed scaffolds facilitated segmental defect repair.

Human embryonic stem cell-derived mesenchymal stem cell secretome reverts silica-induced airway epithelial cell injury by regulating Bmi1 signaling.

Silicosis is an irreversible chronic pulmonary disease caused by long-term inhalation and deposition of silica particles, which is currently incurable. The exhaustion of airway epithelial stem cells plays a pathogenetic role in silicosis. In present study, we investigated therapeutic effects and potential mechanism of human embryonic stem cell (hESC)-derived MSC-likes immune and matrix regulatory cells (IMRCs) (hESC-MSC-IMRCs), a type of manufacturable MSCs for clinical application in silicosis mice. Our results showed that the transplantation of hESC-MSC-IMRCs led the alleviation of silica-induced silicosis in mice, accompanied by inhibiting epithelia-mesenchymal transition (EMT), activating B-cell-specific Moloney murine leukemia virus integration site 1 (Bmi1) signaling and airway epithelial cell regeneration. In consistence, the secretome of hESC-MSC-IMRC exhibited abilities to restore the potency and plasticity of primary human bronchial epithelial cells (HBECs) proliferation and differentiation following the SiO2 -induced HBECs injury. Mechanistically, the secretome resolved the SiO2 -induced HBECs injury through the activation of BMI1 signaling and restoration of airway basal cell proliferation and differentiation. Moreover, the activation of BMI1 significantly enhanced the capacity of HBEC proliferation and differentiation to multiple airway epithelial cell types in organoids. Cytokine array revealed that DKK1, VEGF, uPAR, IL-8, Serpin E1, MCP-1 and Tsp-1 were the main factors in the hESC-MSC-IMRC secretome. These results demonstrated a potential therapeutic effect of hESC-MSC-IMRCs and their secretome for silicosis, in part through a mechanism by activating Bmi1 signaling to revert the exhaustion of airway epithelial stem cells, subsequentially enhance the potency and plasticity of lung epithelial stem cells.

Isolation of a Population of Cells Co-Expressing Markers of Embryonic Stem Cells and Mesenchymal Stem Cells from the Rudimentary Uterine Horn of a Patient with Uterine Aplasia

More than 50% cells isolated from the endometrial cavity scraping and the myometrium of the rudimentary horn of an underdeveloped uterus removed from a patient with uterine aplasia and maintained under culturing conditions normal for mesenchymal stem cells (MSC) expressed embryonic transcription factors Oct4 and Nanog, embryonic cell membrane sialyl glycolipid SSEA4, and MSC markers. After 2-3 passages, the cells lost the expression of the early embryogenesis markers, but retained MSC markers. The presence of dormant stem cells in the underdeveloped endometrium and in the uterus indicates that this tissue has a regenerative potential that can be activated and used for completion of organ morphogenesis. This task requires the development of methods of early diagnosis of morphogenesis impairment and tools for safe reactivation of the ontogenesis.

ALK-5 Inhibitors for Efficient Derivation of Mesenchymal Stem Cells from Human Embryonic Stem Cells.

Objectives: Successful tissue regeneration requires a clinically viable source of mesenchymal stem cells (MSCs). We explored activin receptor-like kinase (ALK)-5 inhibitors to rapidly derive an MSC-like phenotype with high cartilage forming capacity from a xeno-free human embryonic cell line. Methods: Embryonic stem cell (ESC) lines (H9 and HADC100) were treated with the ALK-5 inhibitor SB431542; HADC100 cells were additionally treated with ALK-5 inhibitors SB525334 or GW788388. Cells were then seeded upon human fibronectin in the presence of fibroblast growth factor 2 (FGF2) in a serum-free medium. Flow cytometry was used to assess MSC markers (positive for CD73, CD90, and CD105; negative for CD34 and CD45). Differentiation status was assessed through quantitative polymerase chain reaction. Cartilage forming capacity was determined in high-density pellet cultures, in fibrin gels containing extracellular matrix (fibrin-ECM), and after implantation in ex vivo human osteoarthritic cartilage. Gene expression, histology, and immunostaining were used to assess cartilage phenotype, tissue regeneration, and integration. Results: Exposure to all three ALK-5 inhibitors lead to expression of mesodermal gene markers and differentiation into MSC-like cells (embryonic stem cell-derived mesenchymal stem cells [ES-MSCs]) based on surface marker expression. ES-MSC in pellet cultures or in fibrin-ECM gels expressed high levels of chondrogenic genes: COL2A1, ACAN, and COMP; and low levels of COL1A1 and RUNX2. Cell pellets or fibrin constructs implanted into ex vivo human osteoarthritic cartilage defects produced GAG-rich (safranin O positive) and collagen type II-positive neocartilage tissues that integrated well with native diseased tissue. Conclusions: We developed a protocol for rapid differentiation of xeno-free ESC into MSC-like cells with high cartilage forming capacity with potential for clinical applications. Impact statement Osteoarthritis (OA) is a common disease resulting in significant disability and no approved disease modifying treatment (other than total joint replacement). Embryonic stem cell-derived cell therapy has the potential to benefit patients with cartilage lesions leading to OA and may prevent or delay the need for total joint replacement.

Hypoxia enhances the hair growth-promoting effects of embryonic stem cell-derived mesenchymal stem cells via NADPH oxidase 4

Human embryonic stem cell (hES)-derived mesenchymal stem cells (-MSCs) are an unlimited source of MSCs. The hair growth-promoting effects of diverse MSCs have been reported, but not that of hES-MSCs. In the present study, we investigated the hair growth-promoting effects of hES-MSCs and their underlying mechanisms. hES-MSCs or conditioned medium of hES-MSCs exhibited hair-growth effects, which increased the length of mouse vibrissae and human hair follicles. hES-MSCs accelerated the telogen-to-anagen transition in C3H mice and were more effective than adipose-derived stem cells. We further examined whether hypoxia could enhance the hair-growth promoting effects of hES-MSCs. The injection of hES-MSCs or conditioned medium (Hyp-CM) cultured under hypoxia (2% O2) enhanced the telogen-to-anagen transition in C3H mice. Additionally, Hyp-CM increased the length of mouse vibrissae, human hair follicles, and the proliferation of human dermal papilla and outer root sheath cells. Moreover, fibroblast growth factor 7, interleukin 12B, and teratocarcinoma-derived growth factor 1 were upregulated under hypoxia, and the co-treatment with these three proteins increased the hair length and induced telogen-to-anagen transition. Hypoxia increased reactive oxygen species (ROS) production, and ROS scavenging attenuated the secretion of growth factors. NADPH oxidase 4 was primarily expressed in hES-MSCs and generated ROS under hypoxia. Collectively, our results suggest that hES-MSCs exhibit hair-growth effects, which is enhanced by hypoxia.

Distribution of Embryonic Stem Cell-Derived Mesenchymal Stem Cells after Intravenous Infusion in Hypoxic-Ischemic Encephalopathy.

Systemic administration of mesenchymal stem cells (MSCs) has been reported to improve neurological function in brain damage, including hypoxic-ischemic encephalopathy (HIE), though the action mechanisms have not been fully elucidated. In this study, the cells were tracked live using a Pearl Trilogy Small Animal fluorescence imaging system after human embryonic stem Cell-Derived MSCs (ES-MSCs) infusion for an HIE mouse model. ES-MSC-treated HIE mice showed neurobehavioral improvement. In vivo imaging showed similar sequential migration of ES-MSCs from lungs, liver, and spleen within 7 days in both HIE and normal mice with the exception of lungs, where there was higher entrapment in the HIE 1 h after infusion. In addition, ex vivo experiments confirmed time-dependent infiltration of ES-MSCs into the organs, with similar findings in vivo, although lungs and brain revealed small differences. ES-MSCs seemed to remain in the brain only in the case of HIE on day 14 after the cell infusion. The homing effect in the host brain was confirmed with immunofluorescence staining, which showed that grafted cells remained in the brain tissue at the lesion area with neurorestorative findings. Further research should be carried out to elucidate the role of each host organ's therapeutic effects when stem cells are systemically introduced.

Promising Therapeutic Effects of Embryonic Stem Cells-Origin Mesenchymal Stem Cells in Experimental Pulmonary Fibrosis Models: Immunomodulatory and Anti-Apoptotic Mechanisms.

Interstitial lung disease (ILD) involves persistent inflammation and fibrosis, leading to respiratory failure and even death. Adult tissue-derived mesenchymal stem cells (MSCs) show potential in ILD therapeutics but obtaining an adequate quantity of cells for drug application is difficult. Daewoong Pharmaceutical's MSCs (DW-MSCs) derived from embryonic stem cells sustain a high proliferative capacity following long-term culture and expansion. The aim of this study was to investigate the therapeutic potential of DW-MSCs in experimental mouse models of ILD. DW-MSCs were expanded up to 12 passages for in vivo application in bleomycin-induced pulmonary fibrosis and collagen-induced connective tissue disease-ILD mouse models. We assessed lung inflammation and fibrosis, lung tissue immune cells, fibrosis-related gene/protein expression, apoptosis and mitochondrial function of alveolar epithelial cells, and mitochondrial transfer ability. Intravenous administration of DW-MSCs consistently improved lung fibrosis and reduced inflammatory and fibrotic markers expression in both models across various disease stages. The therapeutic effect of DW-MSCs was comparable to that following daily oral administration of nintedanib or pirfenidone. Mechanistically, DW-MSCs exhibited immunomodulatory effects by reducing the number of B cells during the early phase and increasing the ratio of Tregs to Th17 cells during the late phase of bleomycin-induced pulmonary fibrosis. Furthermore, DW-MSCs exhibited anti-apoptotic effects, increased cell viability, and improved mitochondrial respiration in alveolar epithelial cells by transferring their mitochondria to alveolar epithelial cells. Our findings indicate the strong potential of DW-MSCs in the treatment of ILD owing to their high efficacy and immunomodulatory and anti-apoptotic effects.

Effective Combination Therapy with Human Amniotic Membrane Mesenchymal Stem Cells and Low-Level Laser Therapy in Wound Healing in Animal Models.

Introduction: Many studies have recognized the importance of new methods in wound healing. This study aims to investigate the healing effects of allograft extra embryonic mesenchymal stem cells (MSCs) with and without low-level diode laser irradiation when grafted into full-thickness skin defects in diabetic animal models. Methods: In this experimental study, in order to make the rats diabetic, we used an intra-peritoneal injection of streptozotocin. Human amniotic membrane derived mesenchymal stem cells (hAMSCs) were irradiated with a low-level diode laser. Two full-thickness excisions were made on the backs of the rats. Next, the rats were divided into the following groups: group 1: low-level laser (LLL) irradiated hAMSCs and group 2: hAMSCs alone transplanted into skin wound. Histopathologic, ultrasound and elasticity evaluations were performed 7, 14 and 21 days after grafting. Results: In the evaluated rats, epithelial formation was on day 7 and increased until day 14. On days 7, 14 and 21, the percentage of epithelial formation in the irradiated cell group was significantly higher than that in the cell group, so that, on day 21, the epithelium in this group completely covered the wound surface while in the control group the wound surface was still not completely covered. In terms of angiogenesis, on day 7, the irradiated cells were significantly lower than the cells. Also, the formation of collagen in the cellular hydrogel group could confirm the effectiveness of amniotic MSCs in collagen production and thus accelerate the wound healing process. In comparison with hAMSCs alone, irradiated hAMSCs increased the thickness and elasticity of the skin. Conclusion: Low-power laser along with MSCs can be effective in improving chronic wound condition in the animal model.

The Protective Effects of Human Embryonic Stem Cell-Derived Mesenchymal Stem Cells in Noise-Induced Hearing Loss of Rats.

A few prior animal studies have suggested the transplantation or protective effects of mesenchymal stem cells (MSCs) in noise-induced hearing loss. This study intended to evaluate the fates of administered MSCs in the inner ears and the otoprotective effects of MSCs in the noise-induced hearing loss of rats. Human embryonic stem cell-derived MSCs (ES-MSCs) were systematically administered via the tail vein in adult rats. Eight-week-old Sprague-Dawley rats were randomly allocated to the control (n = 8), ES-MSC (n = 4), noise (n = 8), and ES-MSC+noise (n = 10) groups. In ES-MSC and ES-MSC+noise rats, 5 × 105 ES-MSCs were injected via the tail vein. In noise and ES-MSC+noise rats, broadband noise with 115 dB SPL was exposed for 3 h daily for 5 days. The hearing levels were measured using auditory brainstem response (ABR) at 4, 8, 16, and 32 kHz. Cochlear histology was examined using H&E staining and cochlear whole mount immunofluorescence. The presence of human DNA was examined using Sry PCR, and the presence of human cytoplasmic protein was examined using STEM121 immunofluorescence staining. The protein expression levels of heat shock protein 70 (HSP70), apoptosis-inducing factor (AIF), poly (ADP-ribose) (PAR), PAR polymerase (PARP), caspase 3, and cleaved caspase 3 were estimated. The ES-MSC rats did not show changes in ABR thresholds following the administration of ES-MSCs. The ES-MSC+ noise rats demonstrated lower ABR thresholds at 4, 8, and 16 kHz than the noise rats. Cochlear spiral ganglial cells and outer hair cells were more preserved in the ES-MSC+ noise rats than in the noise rats. The Sry PCR bands were highly detected in lung tissue and less in cochlear tissue of ES-MSC+noise rats. Only a few STEM121-positivities were observed in the spiral ganglial cell area of ES-MSC and ES-MSC+noise rats. The protein levels of AIF, PAR, PARP, caspase 3, and cleaved caspase 3 were lower in the ES-MSC+noise rats than in the noise rats. The systemic injection of ES-MSCs preserved hearing levels and attenuated parthanatos and apoptosis in rats with noise-induced hearing loss. In addition, a tiny number of transplanted ES-MSCs were observed in the spiral ganglial areas.

Engineering elastic bioactive composite hydrogels for promoting osteogenic differentiation of embryonic mesenchymal stem cells.

The development of bioactive materials with good mechanical properties and promotion of stem cell osteogenic differentiation has important application prospects in bone tissue engineering. In this paper, we designed a novel organic‒inorganic composite hydrogel (FPIGP@BGN-Sr) utilizing diacrylated F127 (DA-PF127), β-glycerophosphate-modified polyitaconate (PIGP) and strontium-doped bioactive glass nanoparticles (BGN-Sr) through free radical polymerization and coordination interactions and then evaluated its promoting effect on the osteogenic differentiation of mouse embryonic mesenchymal stem cells in detail. The results showed that the FPIGP@BGN-Sr hydrogel exhibited a controlled storage modulus by changing the amount of BGN-Sr. Notably, the FPIGP@BGN-Sr hydrogel possessed excellent elastic ability with a compressive strain of up to 98.6% and negligible change in mechanical properties after 10 cycles of compression. In addition, the FPIGP@BGN-Sr hydrogel had good cytocompatibility, maintained the activity and proliferation of mouse embryonic mesenchymal stem cells (C3H10T1/2), and effectively enhanced the activity of alkaline phosphatase, osteogenic gene expression and biomineralization ability of the cells. In conclusion, the excellent mechanical properties and osteogenic biological activity of the FPIGP@BGN-Sr hydrogel make it a promising organic‒inorganic composite bioactive material for stem cell-based bone regeneration.

Uniparental parthenogenetic embryonic stem cell derivatives adaptable for bone and cartilage regeneration

Cells with the desired phenotype and number are critical for regenerative medicine and tissue engineering. Uniparental parthenogenetic embryonic stem cells (pESCs) share fundamental properties with embryonic stem cells. This study aims to determine the viability of pESC-based tissue engineering for bone and cartilage reconstruction. The mouse pESCs were cultured in suspension to form embryoid bodies. An adherent cultivation approach was employed to obtain parthenogenetic embryonic mesenchymal stem cells (pMSCs) from the embryoid bodies. Then, the pMSCs were cultured in conditional media to differentiate into osteogenic and chondrogenic lineages. The pESC-derived osteoblasts and chondroblasts were seeded into coral and sodium alginate scaffolds, respectively. The cell-seeded scaffolds were implanted into dorsal subcutaneous pockets of nude mice to evaluate ectopic reconstruction of bone and cartilage. We demonstrated that pESCs display the capacity to differentiate into all three germ layers. The generated pMSCs were able to differentiate into osteogenic and chondrogenic lineages, which survived well after seeding into coral and alginate acid scaffolds. Six weeks after cell-scaffold implantation, gross inspection and histological examination revealed that ectopic bone and cartilage tissues had successfully regenerated in the specimen. According to the findings of this study, pESC derivatives have a high potential for bone and cartilage regeneration.

Embryonic stem cells-derived mesenchymal stem cells do not differentiate into ovarian cells but improve ovarian function in POF mice

Premature ovarian failure (POF) is characterized by amenorrhea, hypoestrogenism, elevated gonadotropin levels, and infertility. Although some of previous studies reported that Mesenchymal stem cells (MSC) transplantation could rescue the ovary function of POF animal models through the paracrine pathways, these mechanisms require further investigation. Here, we aimed to investigate the possible mechanisms of therapeutic effects of human embryonic stem cells derived MSC (ES-MSC) in a mice model of chemotherapy-induced POF. For this purpose, Cyclophosphamide (Cy) was injected intraperitoneally into female mice to induce POF. 10 days after Cy injection, we evaluated follicle count, follicle-stimulating hormone (FSH) and estradiol (E2) hormone concentrations, and TUNEL assay. Then, ES-MSC was transplanted into mice and the expression of Anti-müllerian hormone (AMH) and apoptosis was evaluated in ovary. Results indicated that ES-MSC reduced apoptosis in the follicles and increased the expression of AMH protein in the ovary of POF mice. So, ES-MSC may inhibit the apoptosis of ovarian granulosa cells. Then, to investigate the potential mechanisms of therapeutic effects of ES-MSC and their fate in the ovary, MSC were labeled with green fluorescent protein (GFP) before transplantation. Immunofluorescence staining indicated that although GFP-labeled ES-MSC was located in the ovarian stroma, they did not express granulosa cell markers: AMH and Follicle-stimulating hormone receptor (FSHR), theca cell marker: luteinizing hormone receptor (LHR), and oocyte marker: Growth/differentiation factor 9 (GDF9). Therefore, ES-MSC may not differentiate into ovarian cells directly and they might restore ovarian function in chemotherapy-induced POF mice by paracrine mechanisms.

Abstract PR003: YAP1 is a key regulator of EWS-FLI1-dependent malignant transformation upon IGF1 mediated reprogramming of bone mesenchymal stem cell-like cells

Abstract Ewing sarcoma (EwS) is driven by the EWS-FLI1 (EF) oncoprotein and is assumed to originate from mesenchymal progenitors during bone development. Attempts to generate a EwS genetic mouse model by targeting EF expression to the mesenchymal lineage so far failed to result in tumorigenesis. Here, we investigated the influence of bone niche-derived factors on EF dependent transformation. We focused on IGF1 which is highly enriched in the bone during puberty when EwS incidence peaks, and which was previously proposed to play an essential role in EwS pathogenesis. We restricted EF expression to mesenchymal stem cells during endochondral bone formation by crossing an EF knock-in mouse with a mouse expressing Prx1-driven Cre recombinase. The resulting EFprx1 progeny died after birth as consequence of severe skeletal malformations resulting from differentiation arrest of early chondrocytes. In vitro, these embryonic EF transgenic chondrocyte-like mesenchymal stem cells (EFprx1 MSCLC) are immortal, but largely failed to form colonies in soft-agar as functional surrogate of full malignant transformation. In contrast, when treated with IGF1 for 4 weeks, a significant increase in soft-agar colony formation was observed, which was dependent on sustained EF expression. Strikingly, cells from these colonies maintained their transformed phenotype in absence of further IGF1 treatment and gave rise to aggressively growing tumors upon xenotransplantation in mice. Analysis of differentially expressed genes from these stably IGF1-reprogrammed, fully transformed EFprx1 MSCLC showed a significant enrichment for the EwS specific EF transcriptional signature with upregulation of Foxm1 and cell cycle-related pathways, and down-regulation of apoptotic gene sets. Cluster analysis of open chromatin regions in EF immortalized versus fully transformed and wildtype cells revealed five distinct chromatin modules. Of these, module 2 comprises genomic regions that open up exclusively in IGF1 reprogrammed EF expressing cells. Among associated genes, Yap1 was most highly induced in IGF1 reprogrammed EFprx1 MSCLC. Consistent with this finding, RNA-seq revealed enrichment of a Yap1 transcriptional signature in IGF1 reprogrammed versus parental EF expressing cells. In addition, chromatin module 4, which comprises regions open in parental and IGF1 reprogrammed EFprx1 MSCLC but not in wildtype MSC, is highly enriched in binding motifs for TEAD transcription factors, the functional effectors of YAP1 signaling. We therefore hypothesize that YAP1 induction by IGF1 reprogramming synergizes with EF in the activation of module 4 genes. Strikingly, modules 2 and 4 comprise hallmark genes of stemness and self-renewal which also characterize human EwS, including Bcl11b, Prkcb, Sox2, Ccnd1, and Klf5, as well as the top EwS dependency gene Igf2bp1. Our data suggest that transient IGF1 exposure in the bone microenvironment during puberty may be required for EF driven EwS tumor initiation, and we identify a central role of YAP1 in the early phases of EwS pathogenesis. Citation Format: Heinrich Kovar, Rahil Noorizadeh, Florian Halbritter, Utkarsh Kapoor, Wolfgang Mikulits, Richard Moriggl, Barbara Sax. YAP1 is a key regulator of EWS-FLI1-dependent malignant transformation upon IGF1 mediated reprogramming of bone mesenchymal stem cell-like cells [abstract]. In: Proceedings of the AACR Special Conference: Sarcomas; 2022 May 9-12; Montreal, QC, Canada. Philadelphia (PA): AACR; Clin Cancer Res 2022;28(18_Suppl):Abstract nr PR003.