Wharton's Jelly Mesenchymal Stem Cells Research Articles

Cryptomphalus aspersa Egg Extract Protects against Human Stem Cell Stress-Induced Premature Senescence.

Cellular senescence is a tightly regulated pathophysiologic process and is caused by replicative exhaustion or external stressors. Since naturally derived bioactive compounds with anti-ageing properties have recently captured scientific interest, we analysed the anti-ageing and antioxidant efficacy of Cryptomphalus aspersa egg extract (CAEE). Its effects on stemness, wound-healing properties, antioxidant defense mechanisms, and DNA damage repair ability of Human Wharton's jelly mesenchymal stem cells (WJ-MSCs) were analysed. Our results revealed that CAEE fortifies WJ-MSCs stemness, which possibly ameliorates their wound-healing ability. Additionally, we show that CAEE possesses a strong antioxidant capacity as demonstrated by the elevation of the levels of the basic antioxidant molecule, GSH, and the induction of the NRF2, a major antioxidant regulator. In addition, CAEE alleviated cells' oxidative stress and therefore prevented stress-induced premature senescence (SIPS). Furthermore, we demonstrated that the prevention of SIPS could be mediated via the extract's ability to induce autophagy, as indicated by the elevation of the protein levels of all basic autophagic molecules and the increase in formation of autophagolysosomes in CAEE-treated WJ-MSCs. Moreover, CAEE-treated cells exhibited decreased Caveolin-1 levels. We propose that Cryptomphalus aspersa egg extract comprises bioactive compounds that can demonstrate strong antioxidant/anti-ageing effects by regulating the Caveolin-1-autophagy-senescence molecular axis.

Engineered extracellular vesicles carrying let-7a-5p for alleviating inflammation in acute lung injury

BackgroundAcute lung injury (ALI) is a life-threatening respiratory condition characterized by severe inflammation and lung tissue damage, frequently causing rapid respiratory failure and long-term complications. The microRNA let-7a-5p is involved in the progression of lung injury, inflammation, and fibrosis by regulating immune cell activation and cytokine production. This study aims to use an innovative cellular electroporation platform to generate extracellular vesicles (EVs) carring let-7a-5p (EV-let-7a-5p) derived from transfected Wharton’s jelly-mesenchymal stem cells (WJ-MSCs) as a potential gene therapy for ALI.MethodsA cellular nanoporation (CNP) method was used to induce the production and release of EV-let-7a-5p from WJ-MSCs transfected with the relevant plasmid DNA. EV-let-7a-5p in the conditioned medium were isolated using a tangential flow filtration (TFF) system. EV characterization followed the minimal consensus guidelines outlined by the International Society for Extracellular Vesicles. We conducted a thorough set of therapeutic assessments, including the antifibrotic effects using a transforming growth factor beta (TGF-β)-induced cell model, the modulation effects on macrophage polarization, and the influence of EV-let-7a-5p in a rat model of hyperoxia-induced ALI.ResultsThe CNP platform significantly increased EV secretion from transfected WJ-MSCs, and the encapsulated let-7a-5p in engineered EVs was markedly higher than that in untreated WJ-MSCs. These EV-let-7a-5p did not influence cell proliferation and effectively mitigated the TGF-β-induced fibrotic phenotype by downregulating SMAD2/3 phosphorylation in LL29 cells. Furthermore, EV-let-7a-5p regulated M2-like macrophage activation in an inflammatory microenvironment and significantly induced interleukin (IL)-10 secretion, demonstrating their modulatory effect on inflammation. Administering EVs from untreated WJ-MSCs slightly improved lung function and increased let-7a-5p expression in plasma in the hyperoxia-induced ALI rat model. In comparison, EV-let-7a-5p significantly reduced macrophage infiltration and collagen deposition while increasing IL-10 expression, causing a substantial improvement in lung function.ConclusionThis study reveals that the use of the CNP platform to stimulate and transfect WJ-MSCs could generate an abundance of let-7a-5p-enriched EVs, which underscores the therapeutic potential in countering inflammatory responses, fibrotic activation, and hyperoxia-induced lung injury. These results provide potential avenues for developing innovative therapeutic approaches for more effective interventions in ALI.

Dual production of human mesenchymal stromal cells and derived extracellular vesicles in a dissolvable microcarrier-based stirred culture system

Background & aimsCell therapies based on mesenchymal stromal cells (MSCs) have gained an increasing therapeutic interest in the context of multiple disorders. Nonetheless, this field still faces important challenges, particularly concerning suitable manufacturing platforms. Here, we aimed at establishing a scalable culture system to expand umbilical cord-derived Wharton's jelly MSC (MSC(WJ)) and their derived extracellular vesicles (EVs) by using dissolvable microcarriers combined with xeno(geneic)-free culture medium. MethodsMSC(WJ) isolated from three donors were cultured at a starting density of 1 × 106 cells per spinner flask, i.e., 2.8 × 103 cells per cm2 of dissolvable microcarrier surface area. After a 6-day expansion period of MSC(WJ), extracellular vesicles (EVs) were produced for 24 h. ResultsTaking advantage of an intermittent agitation regimen, we observed high adhesion rates to the microcarriers (over 90% at 24 h) and achieved 15.8 ± 0.7-fold expansion after 6 days of culture. Notably, dissolution of the microcarriers was achieved through a pectinase-based solution to recover the cell product, reducing the hurdles of downstream processing. MSC identity was validated by detecting the characteristic MSC immunophenotype and by multilineage differentiation assays. Considering the growing interest in MSC-derived EVs, which are known to be mediators of the therapeutic features of MSC, this platform also was evaluated for EV production. Upon a 24-h period of conditioning, secreted EVs were isolated by ultrafiltration followed by anion-exchange chromatography and exhibited the typical cup-shaped morphology, small size distribution (162.6 ± 30.2 nm) and expressed EV markers (CD63, CD9 and syntenin-1). ConclusionsTaken together, we established a time-effective and robust scalable platform that complies with clinical-grade standards for the dual production of MSC(WJ) and their derived EV.

Miro1 improves the exogenous engraftment efficiency and therapeutic potential of mitochondria transfer using Wharton's jelly mesenchymal stem cells

Mitochondria are important for maintaining cellular energy metabolism and regulating cellular senescence. Mitochondrial DNA (mtDNA) encodes subunits of the OXPHOS complexes which are essential for cellular respiration and energy production. Meanwhile, mtDNA variants have been associated with the pathogenesis of neurodegenerative diseases, including MELAS, for which no effective treatment has been developed. To alleviate the pathological conditions involved in mitochondrial disorders, mitochondria transfer therapy has shown promise. Wharton's jelly mesenchymal stem cells (WJMSCs) have been identified as suitable mitochondria donors for mitochondria-defective cells, wherein mitochondrial functions can be rescued. Miro1 participates in mitochondria trafficking by anchoring mitochondria to microtubules. In this study, we identified Miro1 over-expression as a factor that could help to enhance the efficiency of mitochondrial delivery. More specifically, we reveal that Miro1 over-expressed WJMSCs significantly improved intercellular communications, cell proliferation rates, and mitochondrial membrane potential, while restoring mitochondrial bioenergetics in mitochondria-defective fibroblasts. Furthermore, Miro1 over-expressed WJMSCs decreased rates of induced apoptosis and ROS production in MELAS fibroblasts; although, Miro1 over-expression did not rescue mtDNA mutation ratios nor mitochondrial biogenesis. This study presents a potentially novel therapeutic strategy for treating mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS), and other diseases associated with dysfunctional mitochondria, while the pathophysiological relevance of our results should be further verified by animal models and clinical studies.

Unveiling the topographic cue rendered by micropatterns for steering cell differentiation by using extrinsic photobiomodulation

It is known that cell culture micropatterns have the ability to facilitate stem cell differentiation induced by specialized chemical factors and different differentiation directions have different optimal micropattern shapes. In this study, by utilizing extrinsic photobiomodulation (EPM) with verteporfin as photosensitizer and light irradiation of 690 nm wavelength as a universal, unbiased, and synchronizing way of inducing differentiation of human umbilical cord Wharton's Jelly mesenchymal stem cells (WJ-MSCs), the topographic cue for cell type specification conveyed in microislands is investigated. It is found that the topographic cues are encoded in the symmetry and aspect ratio of microislands and conformation of cells to microislands is necessary for acquiring the cue. F-actin vertical columns form and cell thickness increases for cells on microislands, and the two effects are enhanced by EPM and correlate with cell differentiation. EPM treatment poises cells in a stationary state to initiate differentiation and the process of making commitment takes two days. Our findings reveal the way to fully exploit topographic cues for promoting and controlling cell differentiation.

Umbilical cord mesenchymal stem cells from gestational diabetes show impaired ability to up-regulate paracellular permeability from sub-endothelial niche.

In vitro studies have shown that Wharton's jelly mesenchymal stem cells (WJ-MSCs) can cross umbilical and uterine endothelial barriers and up-regulate endothelial junctional integrity from sub-endothelial niches. This pericytic behaviour may be lost in pregnancies complicated by gestational diabetes (GDM), where increased vascular permeability and junctional disruption are reported. The aim of the present study was to investigate whether WJ-MSCs isolated from GDM pregnancies displayed any changes in morphology, proliferation, VEGF-A secretion, and their ability to influence paracellular junctional composition and permeability. WJ-MSCs were isolated from human umbilical cords from normal pregnancies (nWJ-MSCs, n=13) and those complicated by GDM (gWJ-MSCs), either diet-controlled (d-GDM, n=13) or metformin-treated (m-GDM, n=9). We recorded that 4-fold more WJ-MSCs migrated from m-GDM, and 2.5-fold from d-GDM cord samples compared with the normal pregnancy. gWJ-MSCs showed a less predominance of spindle-shaped morphology and secreted 3.8-fold more VEGF-A compared with nWJ-MSCs. The number of cells expressing CD105 (Endoglin) was higher in gWJ-MSCs compared with nWJ-MSCs (17%) at P-2. The tracer leakage after 24 h across the HUVEC + gWJ-MSCs bilayer was 22.13% and 11.2% higher in the m-GDM and d-GDM, respectively, HUVEC + nWJ-MSCs. Transfection studies with siRNAs that target Endoglin were performed in n-WJ-MSCs; transfected cells were co-cultured with HUVEC followed by permeability studies and VE-cadherin analyses. Loss of Endoglin also led to increased VEGF-A secretion, increased permeability and affected endothelial stabilization. These results reinforce the pericytic role of nWJ-MSCs to promote vascular repair and the deficient ability of gWJ-MSCs to maintain endothelial barrier integrity.

ADAMTS13 regulates angiogenic markers via Ephrin/Eph signaling in human mesenchymal stem cells under serum-deprivation stress

Mesenchymal stem cells (MSCs) are known to facilitate angiogenesis and promote neo-vascularization via secretion of trophic factors. Here, we explored the molecular mechanism adopted by ADAMTS13 in modulating the expression of some key angiogenic markers in human umbilical cord-derived MSCs under serum-deprivation stress. Wharton’s jelly MSCs (WJ-MSCs) were isolated from the perivascular region of human umbilical cords by explant culture. ADAMTS13 was upregulated at both mRNA and protein levels in WJ-MSCs under serum-deprivation stress. Correspondingly, some key angiogenic markers were also seen to be upregulated. By screening signaling pathways, p38 and JNK pathways were identified as negative and positive regulators for expression of ADAMTS13, and the angiogenic markers, respectively. Our results also indicated the Notch pathway and p53 as other probable partners modulating the expression of ADAMTS13 and the angiogenic markers. Knockdown of ADAMTS13 using siRNA led to reversal in the expression of these angiogenic markers. Further, ADAMTS13 was shown to act via the EphrinB2/EphB4 axis followed by ERK signaling to control expression of the angiogenic markers. Interestingly, stronger expression levels were noted for ADAMTS13, VEGF and PDGF under a more stringent nutrient stress condition. Thus, we highlight a novel role of ADAMTS13 in WJ-MSCs under nutrient stress condition.

BIOFUNCTIONALIZED 3D-BIOPRINTED VASCULAR-LIKE STRUCTURES: AN ADVANCED APPROACH TO PROMOTE ENDOTHELIAL TISSUE REGENERATION

Biofabrication is a popular technique to produce personalized constructs for tissue engineering. In this study we combined laponite (Lap), gellan gum (GG) with platelet-rich plasma (PRP) aiming to enhance the endothelial regeneration through the synergistic effects of their individual properties. Laponite has the ability to form porous three-dimensional networks mimicking the extracellular matrix structure, and PRP delivery of growth factors stimulates the endothelial cell proliferation and migration, offering a composite bioink for cell growth and support. The sustained release of these growth factors from the GG-laponite-PRP composite material over time provides a continuous source of stimulation for the cells, leading to more effective tissue engineering strategies for endothelial tissue regeneration. Four blend compositions comprising 1% w/v GG and 0.5 or 1% w/v Lap and 25% v/v PRP were combined with Wharton jelly mesenchymal stem cells (WJ-MSCs) and bioprinted into vessel-like structures with an inner diameter of 3 mm and a wall thickness of 1 mm. Stress/strain analysis revealed the elastomeric properties of the hydrogels with Young modulus values of 10 MPa. Increasing the Lap concentration led to a non-significant decrease of swelling ratio from 93 to 91%. Live/dead assay revealed cell viability of at least 76%, with the 0.5%Lap-GG viability exceeding 99% on day 21. Gradual increase of glycosaminoglycans accumulation and collagen production indicate promotion of ECM formation. The expression and membranous localization of PECAM-1 from day 7 and the granular intracellular localization of vWF after 2 weeks demonstrate in vitro endothelial functionality. In vivo subcutaneous implantation indicated the absence of any adverse immunological reactions. The results reveal the expression of both vWF and PECAM-1 by WJ-MSCs entrapped in all four construct compositions with significantly higher expression of vWF in the presence of PRP.

Early Detection of Functional Changes in an Intraocular Hypertension Rabbit Model Treated with Human Wharton Jelly Mesenchymal Stem Cells (hWJ-MSCs) Using Chromatic Pupillometry

Early Detection of Functional Changes in an Intraocular Hypertension Rabbit Model Treated with Human Wharton Jelly Mesenchymal Stem Cells (hWJ-MSCs) Using Chromatic Pupillometry

Bifunctional tissue-engineered composite construct for bone regeneration: The role of copper and fibrin.

Bifunctional tissue engineering constructs promoting osteogenesis and angiogenesis are essential for bone regeneration. Metal ion-incorporated scaffolds and fibrin encapsulation attract much attention due to low cost, nontoxicity, and tunable control over ion and growth factor release. Herein, we investigated the effect of Cu.nHA/Cs/Gel scaffold and fibrin encapsulation on osteogenic and angiogenic differentiation of Wharton's jelly mesenchymal stem cells (WJMSCs) in vitro and in vivo. Cu-laden scaffolds were synthesized using salt leaching/freeze drying and were characterized using standard techniques. WJMSCs were isolated from the human umbilical cord and characterized. WJMSCs with or without encapsulating in fibrin were seeded onto scaffolds, followed by differentiating into the osteogenic lineage for 7 and 21 days. Osteogenic and angiogenic differentiation were evaluated using real-time polymerase chain reaction, western blot, and Alizarin red staining. Then, scaffolds were implanted into critical-sized calvarial bone defects in rats and histological assessments were performed using hematoxylin/eosin, Masson's trichrome, and CD31 immunohistochemical staining at 4 and 12 weeks. The scaffolds had good physicochemical and biological characteristics suitable for cell attachment and growth. Cu and fibrin increased the expression of ALP, RUNX2, OCN, COLI, VEGF, and HIF1α in differentiated WJMSCs. Implanted scaffolds were also biocompatible and were integrated well with the host tissue. Increased collagen condensation, mineralization, and blood vessel formation were observed in Cu-laden scaffolds. The fibrin-encapsulated groups showed the highest collagen and cell densities, immune cell infiltration, and bone trabeculae. CD31-positive cell population increased with fibrin encapsulation and seeding onto Cu-laden scaffolds. Adding Cu to scaffolds and encapsulating cells in fibrin are promising methods that guide osteogenesis and angiogenesis cellular signaling, leading to better bone regeneration.

Chitosan microspheres loaded with WJMSCs facilitate the restoration of ovarian function in CTX‐induced premature ovarian failure mice

AbstractPremature ovarian failure (POF) is a complex disease affecting an increasing number of young women. Traditional treatments are limited in low efficacy and side effects. Wharton's jelly mesenchymal stem cells (WJMSCs) therapy holds promise in the treatment of POF. However, the clinical application of WJMSCs is hindered by challenges such as low cell viability, significant loss, and poor survival rates. In this study, we prepared three‐dimensional chitosan microspheres (CSM). CSM loading substantial improved the proliferative ability and cell viability of WJMSCs. Furthermore, transplantation of CSM loaded with WJMSCs into ovaries of POF mice increased serum levels of Estradiol (E2) and Anti‐Müllerian hormone (AMH), alongside decreased levels of follicle‐stimulating hormone (FSH). CSM+WJMSCs transplantation enhanced the number of growth follicles while reduced atresia follicles, and effective alleviated the ovarian fibrosis and apoptosis within the ovarian cortex. CSM also prolonged the residence time of WJMSCs in the ovarian capsule. Moreover, superovulation experiments illustrated that CSM+WJMSCs transplantation enhanced the total number of oocytes and reduced oocyte abnormality rates. Overall, CSM+WJMSCs transplantation promotes follicle development and regeneration, restoring ovarian function by inhibiting ovarian cell apoptosis and reducing levels of fibrosis. This study presents a novel therapeutic strategy that combines biomaterial and mesenchymal stem cells for the clinical treatment of premature ovarian failure.

The Comparative Effects of Schwann Cells and Wharton's Jelly Mesenchymal Stem Cells on the AIM2 Inflammasome Activity in an Experimental Model of Spinal Cord Injury

Inflammasome activation and the consequent release of pro-inflammatory cytokines play a crucial role in the development of sensory/motor deficits following spinal cord injury (SCI). Immunomodulatory activities are exhibited by Schwann cells (SCs) and Wharton's jelly mesenchymal stem cells (WJ-MSCs). In this study, we aimed to compare the effectiveness of these two cell sources in modulating the absent in melanoma 2 (AIM2) inflammasome complex in rats with SCI. The Basso, Beattie, Bresnahan (BBB) test, Nissl staining, and Luxol fast blue (LFB) staining were performed to evaluate locomotor function, neuronal survival, and myelination, respectively. Real-time polymerase chain reaction (RT-PCR), Western blotting, and enzyme-linked immunosorbent assay (ELISA) were employed to analyze the gene and protein expressions of inflammasome components, including AIM2, ASC, caspase-1, interleukin-1β (IL-1β), and IL-18. Both gene and protein expressions of all evaluated factors were decreased after SC or WJ-MSC treatment, with a more pronounced effect observed in the SCs group (P < 0.05). Additionally, SCs promoted neuronal survival and myelination. Moreover, the administration of 3 × 105 cells resulted in motor recovery improvement in both treatment groups (P < 0.05). Although not statistically significant, these effects were more prominent in the SC-treated animals. In conclusion, SC therapy demonstrated greater efficacy in targeting AIM2 inflammasome activation and the associated inflammatory pathway in SCI experiments compared to WJ-MSCs.

Effects of royal jelly on the antisenescence, mitochondrial viability and osteogenic differentiation capacity of umbilical cord-derived mesenchymal stem cells.

Mesenchymal stem cells (MSCs) are multipotent cells that have the ability to self-renew and regulate paracrine signalling and immune system processes. MSCs have extensive clinical applications in regeneration, functional reconstruction and cellular therapies. However, studies are needed to discover ways to improve the properties of MSCs, such as differentiation, and prevent senescence in culture, which are both very important for cell therapies. Royal jelly (RJ) is a nutritional substance produced by worker bees that contains a substantial amounts of proteins that are beneficial for cell growth and proliferation. RJ is widely used in traditional medicine today, and due to the specific components in its content, it has been reported to have antioxidant, antiproliferative, antimicrobial, neuroprotective, anti-inflammatory, immunomodulatory and anti-ageing properties. In our study, human Wharton's jelly mesenchymal stem cells (WJ-MSCs) derived from umbilical cord matrix were grown in culture medium supplemented with RJ. The control group comprised minimum essential medium (MEM) and 10% foetal bovine serum (FBS); RJ groups were formed using MEM, 10% FBS and 0.075mg/ml or 0.150mg/ml RJ. In our study, we evaluated the effect of RJ on WJ-MSC growth by MTT assay, proliferating cell nuclear antigen ELISA, β-galactosidase activity assay, MitoTracker Green staining and differentiation tests in adipogenic, osteogenic and chondrogenic cell lines. It was observed that the number of mitochondria increased, senescence decreased and osteogenic differentiation increased after differentiation induction after the addition of RJ to MSC culture. In general, the results of this study indicate that WJ-MSCs enhance mitochondrial numbers and important cellular activities, such as antisenescence and osteogenic differentiation, and with increasing evidence from further studies, RJ supplementation may be found beneficial for the use of MSCs in bone engineering regenerative medicine or cell therapy.

Wharton's Jelly mesenchymal stromal cell-derived extracellular vesicles promote nucleus pulposus cell anabolism in an in vitro 3D alginate-bead culture model.

Intradiscal transplantation of mesenchymal stromal cells (MSCs) has emerged as a promising therapy for intervertebral disc degeneration (IDD). However, the hostile microenvironment of the intervertebral disc (IVD) may compromise the survival of implanted cells. Interestingly, studies reported that paracrine factors, such as extracellular vesicles (EVs) released by MSCs, may regenerate the IVD. The aim of this study was to investigate the therapeutic effects of Wharton's Jelly MSC (WJ-MSC)-derived EVs on human nucleus pulposus cells (hNPCs) using an in vitro 3D alginate-bead culture model. After EV isolation and characterization, hNPCs isolated from surgical specimens were encapsulated in alginate beads and treated with 10, 50, and 100 μg/mL WJ-MSC-EVs. Cell proliferation and viability were assessed by flow cytometry and live/dead staining. Nitrite and glycosaminoglycan (GAG) content was evaluated through Griess and 1,9-dimethylmethylene blue assays. hNPCs in alginate beads were paraffin-embedded and stained for histological analysis (hematoxylin-eosin and Alcian blue) to assess extracellular matrix (ECM) composition. Gene expression levels of catabolic (MMP1, MMP13, ADAMTS5, IL6, NOS2), anabolic (ACAN), and hNPC marker (SOX9, KRT19) genes were analyzed through qPCR. Collagen type I and type II content was assessed with Western blot analysis. Treatment with WJ-MSC-EVs resulted in an increase in cell content and a decrease in cell death in degenerated hNPCs. Nitrite production was drastically reduced by EV treatment compared to the control. Furthermore, proteoglycan content was enhanced and confirmed by Alcian blue histological staining. EV stimulation attenuated ECM degradation and inflammation by suppressing catabolic and inflammatory gene expression levels. Additionally, NPC phenotypic marker genes were also maintained by the EV treatment. WJ-MSC-derived EVs ameliorated hNPC growth and viability, and attenuated ECM degradation and oxidative stress, offering new opportunities for IVD regeneration as an attractive alternative strategy to cell therapy, which may be jeopardized by the harsh microenvironment of the IVD.

Efficacy of Wharton Jelly Mesenchymal Stromal Cells infusions in moderate to severe SARS-Cov-2 related acute respiratory distress syndrome: a phase 2a double-blind randomized controlled trial.

The COVID-19 pandemic caused a wave of acute respiratory distress syndrome (ARDS) with a high in-hospital mortality, especially in patients requiring invasive mechanical ventilation. Wharton Jelly-derived Mesenchymal Stromal Cells (WJ-MSCs) may counteract the pulmonary damage induced by the SARS-CoV-2 infection through pro-angiogenic effects, lung epithelial cell protection, and immunomodulation. In this randomized, double-blind, placebo-controlled phase 2a trial, adult patients receiving invasive mechanical ventilation for SARS-CoV-2 induced moderate or severe ARDS were assigned to receive 1 intravenous infusion of 1 × 106 WJ-MSCs/kg or placebo within 48 h of invasive ventilation followed by 2 infusions of 0.5 × 106 WJ-MSCs/kg or placebo over 5 days. The primary endpoint was the percentage of patients with a PaO2/FiO2 > 200 on day 10. Thirty patients were included from November 2020 to May 2021, 15 in the WJ-MSC group and 15 in the placebo group. We did not find any significant difference in the PaO2/FiO2 ratio at day 10, with 18 and 15% of WJ-MSCs and placebo-treated patients reaching a ratio >200, respectively. Survival did not differ in the 2 groups with a 20% mortality rate at day 90. While we observed a higher number of ventilation-free days at 28 days in the WJ-MSC arm, this difference was not statistically significant (median of 11 (0-22) vs. 0 (0-18), p = 0.2). The infusions were well tolerated, with a low incidence of anti-HLA alloimmunization after 90 days. While treatment with WJ-MSCs appeared safe and feasible in patients with SARS-CoV2 moderate or severe ARDS in this phase 2a trial, the treatment was not associated with an increased percentage of patients with P/F > 200 at 10d, nor did 90 day mortality improve in the treated group. https://beta.clinicaltrials.gov/study/NCT04625738, identifier NCT04625738.

Human Wharton's jelly-derived mesenchymal stem cells prevent pregnancy loss in a rat by JAK/STAT-mediated immunomodulation.

Spontaneous abortion (SA) is a multiple-original syndrome with immune imbalance as one of its major risk factors. As Wharton's jelly-mesenchymal stem cells (WJ-MSCs) are considered to be able to prevent abortion, this study aims to explore the currently poorly understood underlining molecular signaling pathways and regulatory mechanisms of WJ-MSCs in pregnancy maintenance. Abortion mode is established by subcutaneous injection of bromocriptine in rat on day 9 and abortion prevention is achieved by WJ-MSCs injection via tail vein. WJ-MSCs were cultured with/without the inhibitors of JAK/STAT or NF-κB. The uterus was collected on the 14th day of gestation and the rate of embryo absorption was calculated. The expression of Th1/Th2/Th3 cytokines in decidual, placental tissue, and peripheral blood was analyzed. WJ-MSCs treatment significantly reduced the abortion rate in bromocriptine-treated pregnancy such that it was not significantly different from a normal pregnancy. JAK/STAT inhibition abolished pregnancy preserving effects of WJ-MSCs but NF-κB inhibition did not. The levels of Th1-related cytokines and mRNA levels in the bromocriptine abortion model were significantly higher than the normal pregnancy group and ethanol control group, while levels of the Th2-related cytokines and mRNA levels significantly decreased. WJ-MSCs transfusion into the abortion model restored cytokine profiles such that they were not significantly different from the normal pregnancy group and ethanol control group. JAK/STAT inhibition of WJ-MSCs prevented their effect on cytokine and mRNA levels, but NF-κB inhibition did not. WJ-MSCs significantly lower the rate of embryo resorption of spontaneous abortion by reducing Th1-related cytokines while increasing Th2 and Th3-related cytokines in JAK/STAT-dependent manner.

Wharton's jelly mesenchymal stem cells: a concise review of their secretome and prospective clinical applications.

Accumulating evidence indicates that most primary Wharton's jelly mesenchymal stem cells (WJ-MSCs) therapeutic potential is due to their paracrine activity, i.e., their ability to modulate their microenvironment by releasing bioactive molecules and factors collectively known as secretome. These bioactive molecules and factors can either be released directly into the surrounding microenvironment or can be embedded within the membrane-bound extracellular bioactive nano-sized (usually 30-150nm) messenger particles or vesicles of endosomal origin with specific route of biogenesis, known as exosomes or carried by relatively larger particles (100nm-1μm) formed by outward blebbing of plasma membrane called microvesicles (MVs); exosomes and MVs are collectively known as extracellular vesicles (EVs). The bioactive molecules and factors found in secretome are of various types, including cytokines, chemokines, cytoskeletal proteins, integrins, growth factors, angiogenic mediators, hormones, metabolites, and regulatory nucleic acid molecules. As expected, the secretome performs different biological functions, such as immunomodulation, tissue replenishment, cellular homeostasis, besides possessing anti-inflammatory and anti-fibrotic effects. This review highlights the current advances in research on the WJ-MSCs' secretome and its prospective clinical applications.

PSEN1 E280A Cholinergic-like Neurons and Cerebral Spheroids Derived from Mesenchymal Stromal Cells and from Induced Pluripotent Stem Cells Are Neuropathologically Equivalent.

Alzheimer's disease (AD) is a chronic neurological condition characterized by the severe loss of cholinergic neurons. Currently, the incomplete understanding of the loss of neurons has prevented curative treatments for familial AD (FAD). Therefore, modeling FAD in vitro is essential for studying cholinergic vulnerability. Moreover, to expedite the discovery of disease-modifying therapies that delay the onset and slow the progression of AD, we depend on trustworthy disease models. Although highly informative, induced pluripotent stem cell (iPSCs)-derived cholinergic neurons (ChNs) are time-consuming, not cost-effective, and labor-intensive. Other sources for AD modeling are urgently needed. Wild-type and presenilin (PSEN)1 p.E280A fibroblast-derived iPSCs, menstrual blood-derived menstrual stromal cells (MenSCs), and umbilical cord-derived Wharton Jelly's mesenchymal stromal cells (WJ-MSCs) were cultured in Cholinergic-N-Run and Fast-N-Spheres V2 medium to obtain WT and PSEN 1 E280A cholinergic-like neurons (ChLNs, 2D) and cerebroid spheroids (CSs, 3D), respectively, and to evaluate whether ChLNs/CSs can reproduce FAD pathology. We found that irrespective of tissue source, ChLNs/CSs successfully recapitulated the AD phenotype. PSEN 1 E280A ChLNs/CSs show accumulation of iAPPβ fragments, produce eAβ42, present TAU phosphorylation, display OS markers (e.g., oxDJ-1, p-JUN), show loss of ΔΨm, exhibit cell death markers (e.g., TP53, PUMA, CASP3), and demonstrate dysfunctional Ca2+ influx response to ACh stimuli. However, PSEN 1 E280A 2D and 3D cells derived from MenSCs and WJ-MSCs can reproduce FAD neuropathology more efficiently and faster (11 days) than ChLNs derived from mutant iPSCs (35 days). Mechanistically, MenSCs and WJ-MSCs are equivalent cell types to iPSCs for reproducing FAD in vitro.

Vitronectin Mediates Survival of Human WJ-MSCs Under Inflammatory Temperature Stress Via Cell Cycle Arrest

BACKGROUND: Due to their anti-inflammatory and immunomodulatory capabilities, mesenchymal stem cells (MSCs) are being widely used in cell-based therapies for the treatment of a wide spectrum of inflammatory disorders. Despite their promises, substantial cell loss post transplantation leads to compromised therapeutic benefits in clinical trials, which remains a challenge to overcome. Inflammatory microenvironment comprises the presence of pro-inflammatory cytokines, elevated temperature, etc., which could hamper MSC viability following transplantation. Thus, identifying the underlying molecular factors controlling survival mechanism under such stress conditions becomes important. Also, since MSCs from different origins have significantly varied biology, choosing the appropriate MSC source could be critical in determining the fate of transplanted MSCs in a stressful milieu. OBJECTIVE: As extracellular matrix (ECM) components can mediate cell survival signals, in the present study, we have evaluated the role of ECM matricellular protein, vitronectin (VTN), in the survival of human umbilical cord-derived Wharton’s Jelly MSCs (WJ-MSCs) under the condition of inflammatory temperature stress. METHODS AND RESULTS: On exposure to 40°C, WJ-MSCs underwent cell cycle arrest with no significant change in viability status, along with an induction in VTN expression both at mRNA and protein levels. Interestingly, inhibition of pro-survival signalling pathways, ERK or PI3K, at 40°C led to further upregulation in VTN expression without any significant impact on viability or cell cycle arrest status. Interestingly, esiRNA mediated knockdown of VTN in WJ-MSCs at 40°C, with or without pro-survival pathway inhibitors, resulted in reduced cellular viability accompanied by a reversal in G0/G1 cell cycle arrest. CONCLUSIONS: These results established VTN as a protective factor that maintained the viability of WJ-MSCs at 40°C temperature stress. Finally, autophagy was indicated as a mechanism adopted by VTN in executing its pro-survival role.

WHARTON'S JELLY MSCS-DERIVED EXOSOMES AS CELL-FREE THERAPEUTIC STRATEGY FOR INTERVERTEBRAL DISC REGENERATION

The aim of this study was to investigate the regenerative effects of Wharton's Jelly Mesenchymal Stem Cells (WJ-MSCs) derived exosomes (WJ-Exos) on human nucleus pulposus cells (hNPCs) in an in vitro 3D model.WJ-Exos were isolated by tangent flow filtration of WJ-MSCs conditioned media and characterized by TEM, WB for markers expression and quantified with NTA. WJ-Exos PKH26-labeled uptake in hNPCs was detected by confocal microscopy. hNPCs, isolated from surgical specimens (n=4), culture expanded in vitro and encapsulated in alginate beads, were pre-treated with IL1β (10 ng/ml) for 24 hours, then with WJ-Exos at 10, 50 and 100 µg/ml. Cells with growth medium were used as control. We examined: i) cell proliferation and viability (flow cytometry), ii) nitrite production (Griess) iii) glycosaminoglycan (GAG) amount (DMBB), iv) histological staining for extracellular matrix (ECM) analysis and v) gene expression levels of catabolic and anabolic genes (qPCR). The investigations were performed in triplicate for each donor. One-way ANOVA analysis was used to compare the groups under exam and data were expressed as mean ± S.D.A dose dependent increase in hNPCs proliferation was noticed at all exos concentrations under study. Cell death decreased significantly in WJ-Exos 50 µg/ml samples (p ≤ 0,05) compared to IL1β treated hNPCs. Nitrite production was significantly attenuated at 10µg/ml of WJ-Exos (p ≤ 0,01). GAG content and histological analysis showed a difference in ECM synthesis between treated and untreated hNPCs (p ≤ 0,05). Catabolic and inflammatory markers were modulated by WJ-Exos at 100 µg/ml concentration (p ≤ 0,05) whereas 10 µg/ml group increased anabolic gene expression levels (p ≤ 0,05).These findings offer new opportunities for the potential use of exosomes as an attractive alternative cell-free strategy of IDD. WJ-MSC exosomes ameliorate hNPCs growth and viability, attenuate ECM degradation and oxidative stress-related IDD progression after IL1β stimulation.Financial support was received from the “iPSpine” and “RESPINE” Horizon 2020 projects.