Wharton's Jelly-derived Mesenchymal Stem Cells Research Articles

Shockwave Therapy Combined with Autologous Adipose-Derived Mesenchymal Stem Cells Is Better than with Human Umbilical Cord Wharton’s Jelly-Derived Mesenchymal Stem Cells on Knee Osteoarthritis

Extracorporeal shockwave therapy (ESWT) and mesenchymal stem cells (MSCs) have been reported to have chondroprotective effects in knee osteoarthritis (OA). Here, we examined whether autologous adipose-derived mesenchymal stem cells (ADMSCs) and human umbilical cord Wharton’s jelly-derived mesenchymal stem cells (WJMSCs) increased the efficacy of ESWT in knee OA, and compared the efficacy of the two. The treatment groups exhibited significant improvement of knee OA according to pathological analysis, micro-computed tomography (CT), and immunohistochemistry (IHC) staining. The ADMSCs and ESWT+ADMSCs groups exhibited increased trabecular thickness and bone volume as compared with the ESWT, WJMSCs, and ESWT+WJMSCs groups individually. According to the results of IHC staining, Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) activity and caspase-3 were significantly reduced in the ADMSCs and ESWT+ADMSCs groups as compared with the WJMSCs and ESWT+WJMSC groups. In mechanistic factor analysis, the synergistic effect of ESWT+ADMSCs was observed as being greater than the efficacies of other treatments in terms of expressions of transforming growth factor (TGF)-β, runt-related transcription factor (RUNX)-2 and sex determining region Y-box (SOX)-9. The type II collagen was expressed at a higher level in the WJMSCs group than in the others. Furthermore, ESWT+ADMSCs reduced the expression of platelet-derived growth factor (PDGF)-BB and increased the expression of bone morphogenetic protein (BMP)-4. Therefore, we demonstrated that ESWT+ADMSCs had a synergistic effect greater than that of ESWT+WJMSCs for the treatment of early knee OA.

The effect of human wharton's jelly-derived mesenchymal stem cells on MC4R, NPY, and LEPR gene expression levels in rats with streptozotocin-induced diabetes.

Objective(s):Type 1 diabetes (T1D) is an autoimmune disease resulting from inflammatory destruction of islets β-cells. Nowadays, progress in cell therapy, especially mesenchymal stem cells (MSCs) proposes numerous potential remedies for T1D. We aimed to investigate the combination therapeutic effect of these cells with insulin and metformin on neuropeptide Y, melanocortin-4 receptor, and leptin receptor genes expression in TID. Materials and Methods:One hundreds male rats were randomly divided into seven groups: the control, diabetes, insulin (Ins.), insulin+metformin (Ins.Met.), Wharton’s Jelly-derived MSCs (WJ-MSCs), insulin+metformin+WJ-MSCs (Ins.Met.MSCs), and insulin+WJ-MSCs (Ins.MSCs). Treatment was performed from the first day after diagnosis as diabetes. Groups of the recipient WJ-MSCs were intraportally injected with 2× 10⁶ MSCs/kg at the 7th and 28th days of study. Fasting blood sugar was monitored and tissues and genes analysis were performed.Results:The blood glucose levels were slightly decreased in all treatment groups within 20th and 45th days compared to the diabetic group. The C-peptide level enhanced in these groups compared to the diabetic group, but this increment in Ins.MSCs group on the 45th days was higher than other groups. The expression level of melanocortin-4 receptor and leptin receptor genes meaningfully up-regulated in the treatment groups, while the expression of neuropeptide Y significantly down-regulated in the treatment group on both times of study. Conclusion:Our data exhibit that infusion of MSCs and its combination therapy with insulin might ameliorate diabetes signs by changing the amount of leptin and subsequent changes in the expression of neuropeptide Y and melanocortin-4 receptor.

Overview of methods of isolation, cultivation and genetic profiling on human umbilical cord stem cells

Abstract Stem cells possess unique properties, such as self-renewal ability or differentiation capacity into more specialized cells, which makes them particularly relevant for regenerative medicine and cellular therapies. Although embryonic stem cells (ESCs) are capable of differentiation into all cell lineages, their utilization is associated with ethical concerns since they are obtained from embryos. Furthermore, ESCs may form teratomas or cause immune rejection in the clinical setting. Therefore, an effort has been made to utilize stem cells derived from adult tissues, especially mesenchymal stem cells (MSCs). A particularly attractive source of MSCs is the human umbilical cord, which is typically discarded after birth and considered a medical waste, therefore the acquisition of the cells is not associated with any health risk for a patient. Moreover, umbilical cord-derived MSCs do not express MHCII, thus they exhibit reduced immunogenicity. MSCs have been isolated from all compartments of umbilical cord, however the Wharton’s jelly-derived MSCs (WJ-M-SCs) are the most clinically utilizable. There are two techniques of UC-MSCs isolation: the enzymatic and explant procedures. The explant method involves cell outgrowth of tissue pieces placed into plastic culture vessel after mechanical splitting, whereas the enzymatic technique involves minced tissue digestion in an enzymatic solution. In vitro culture conditions of the isolated cells are highly variable among the researchers, however the most commonly performed molecular assays are homogenous and include: RT-qPCR, flow cytometry and immunocytochemistry. Running title: Human umbilical cord stem cells isolation, cultivation and genetic profiling

Evaluation of reference genes for quantitative real-time PCR in Wharton's Jelly-derived mesenchymal stem cells after lentiviral transduction and differentiation.

Quantitative real time reverse transcription PCR, qRT-PCR, is one of the most important techniques for assessing the level of gene expression. Selecting the correct reference gene to normalize the results is a key step in this method. Inaccurate data can be generated if the correct reference gene is not selected. The level of the expression of reference genes is tissue-variable, and in the case of mesenchymal stem cells (MSC), it can be different depending on the source of their origin. The aim of this study was to select the reference gene for Wharton's Jelly-derived MSC (WJ- MSC) that were undergoing transduction and differentiation. In this work, the expression of 32 genes was analyzed, of which two (RPS17 and 18SrRNA), which had the most stable expression level, were selected. A comparative analysis of the expression stability of the selected genes was then performed with the genes that are most commonly used in the literature, i.e. β-actin and GAPDH. Next, it was determined that a false picture of the expression level of the studied genes can be obtained when a reference gene with variable expression level is used for normalization. RPS17 and 18SrRNA proved to be the most stable reference genes for the WJ-MSC that had been subjected to the lentiviral transfection procedure followed by differentiation. The expression of β-actin and GAPDH was highly unstable and therefore these genes are not suitable for use as reference genes in studies involving WJ- MSC.

Wharton's Jelly Mesenchymal Stromal Cells from Human Umbilical Cord: a Close-up on Immunomodulatory Molecules Featured In Situ and In Vitro.

Therapeutic options for end-stage organ failure are often limited to whole organ transplantation. The tolerance or rejection of the transplanted organ is driven by both early non-specific innate and specific adaptive responses. The use of mesenchymal stromal cells (MSCs) is considered a promising tool in regenerative medicine. Human umbilical cord (HUC) is an easily available source of MSCs, without relevant ethical issues. Moreover, Wharton's jelly-derived MSCs (WJ-MSCs), showed consistent immunomodulatory features that may be useful to promote immune tolerance in the host after transplantation. Few data are available on the phenotype of WJ-MSCs in situ. We investigated the expression of immune-related molecules, such as HLAs, IDO, CD276/B7-H3, and others, both in situ (HUC) and in in vitro-cultured WJ-MSCs. Morphological and biochemical techniques were used to define the expression of such molecules. In addition, we focused on the possible role of CD276/B7-H3 on T cells proliferation inhibition. We assessed CD276/B7-H3 expression by WJ-MSCs both in situ and alongside cell culture. WJ-MSCs were able to suppress T cell proliferation in mixed lymphocyte reaction (MLR). Moreover, we describe for the first time a specific role for CD276/B7-H3, since the immunomodulatory ability of WJ-MSCs was abolished upon anti-CD276/B7-H3 antibody addition to the MLR. These results further detail the immune regulation properties and tolerance induction exerted by human WJ-MSCs, in particular pointing to CD276/B7-H3 as one of the main involved factors. These data further suggest WJ-MSCs as potent tools to modulate local immune response in "support-type" regenerative medicine approaches.

Effect of Human Platelet Lysate in Differentiation of Wharton's Jelly Derived Mesenchymal Stem Cells.

Mesenchymal stem cells (MSCs) are highly preferred in clinical therapy for repair and regeneration of diseased tissues for their multipotent properties. Conventionally, MSCs have been cultured in media supplemented with animal derived serum, however, it is ideal to expand MSCs in media containing supplements of human origin for clinical therapy. Currently, a number of human derived products are being studied as an alternative to animal sources. Amongst these, platelet lysate (PL) has gained interest in the culture of MSCs without affecting their phenotypic property. In this study, we used various concentration of PL (2.5, 5, 7.5 & 10%) in the growth medium of MSCs to identify the least concentration of PL that could be an effective alternative to animal products. MSCs were isolated from Wharton's Jelly by using explant method and expanded in various concentration of PL supplemented medium against the standard FBS containing medium. WJ-MSCs were characterised as per the minimal criteria proposed by International Society for Cell therapy (ISCT), Proliferation study by BrdU assay, gene expression study by qRT-PCR, sterility test for bacteria, Mycoplasma by PCR and endotoxin detection by LAL assay. Whartons jelly derived MSCs (WJ-MSCs) cultured using standard medium supplemented with various concentration of PL exhibited enhanced proliferation and differentiation potential, unaltered immunophenotypic property and genetic stability when compared with the commercial medium containing 10% FBS. The least concentration of PL for an ideal expansion of MSCs was found to be 2.5% and was comparable to FBS.

Overexpression of anti-fibrotic factors ameliorates anti-fibrotic properties of Wharton's jelly derived mesenchymal stem cells under oxidative damage.

Transplantation with Wharton's jelly derived mesenchymal stem cells (WJ-MSCs) showed great benefits for restoring myocardial function. However, the outcome of WJ-MSCs transplantation was unsuccessful due to multiple factors including oxidative damage. The presence of oxidative stress due to myocardium injury influences fibrous tissue formation, which causes disability of cardiac muscle. Hepatocyte growth factor (HGF), insulin-like growth factor (IGF1), and sonic hedgehog (SHH) are well-known master regulators in anti-fibrosis when secreted by WJ-MSCs. They showed a beneficial role in the recovery of cardiac fibrosis after WJ-MSCs transplantation. This study hypothesizes whether the reduction of the anti-fibrosis property in WJ-MSCs from oxidative damage can be recovered by overexpression of the HGF, IGF1, or SHH gene. Overexpression was attained by transfection of WJ-MSCs with pCMV3-HGF, pCMV3-IGF1, or pCMV3-SHH followed by H2O2 exposure and co-culturing with cardiac fibroblasts. Myofibroblast specific markers comprised of alpha-smooth muscle actin (α-SMA) and collagen type 1 (COL1) were evaluated. The WJ-MSCs treated with H2O2 influenced the expression of myofibroblastic markers, whereas the overexpression of HGF, IGF1 or SHH reduced myofibroblastic formation. These results indicate that the oxidative stress impaired anti-fibrotic property of WJ-MSCs, leads to an increase of myofibroblasts. Overexpression of anti-fibrotic genes restored the endogenous HGF, IGF1, and SHH alleviating improvement of cardiac function.

Wharton’s jelly-derived mesenchymal stem cells and polycaprolactone/hydroxyapatite (PCL/HA) scaffold for bone tissue engineering

Background: Polycaprolactone (PCL) is a suitable polymer in the field of drug delivery and tissue engineering scaffolds.Aim of Work: This study aimed to design a Polycaprolactone /Nano hydroxyapatite (PCL/nHA10%) hybrid nanofiber scaffold and evaluate differentiation of Wharton’s jelly-derived mesenchymal stem cells (WJ-MSCs) to osteogenic lineage on scaffolds for bone tissue engineering.Materials and Methods: In this study, PCL/nHA(10%) composite polymeric nano scaffolds were prepared by electrospinning. Mesenchymal stem cells were isolated from the Warton’s Jelly and cultured in the PCL/nHA 10% scaffold. Biocompatibility of scaffolds was confirmed by MTT assay. Characterization of umbilical cord mesencymal stem cells were performed using flow cytometry. The morphological and cell adhesion characteristics of mesenchymal stem cells (MSCs) on the scaffolds were performed using scanning electron microscopy (SEM) imaging. Finally, the cells were treated with osteogenic differentiation medium for 21 days to investigate their differentiation potential on the scaffolds and then differentiated cells were stained with alizarin red and von Kossa stains.Results: The largest pore size of PCL/nHA (10%) nanofibers was about 22 μm. The culture of WJ-MSCs on the scaffolds showed that the addition of nHA 10% to PCL scaffold caused further attachment and proliferation of the cells. The stained mineral deposit scaffold with alizarin red and von Kossa were also compared. In both types of staining, nano-composite scaffold showed higher calcium deposits.Conclusion: The results showed that the PCL/nHA 10% scaffolds for bone tissue engineering is a good choice, and can be used for bone tissue repair.

Wharton's jelly-derived mesenchymal stem cells attenuate sepsis-induced organ injury partially via cholinergic anti-inflammatory pathway activation.

Sepsis induces organ dysfunction due to overexpression of the inflammatory host response, resulting in cardiopulmonary and autonomic dysfunction, thus increasing the associated morbidity and mortality. Wharton's jelly-derived mesenchymal stem cells (WJ-MSCs) express genes and secrete factors with anti-inflammatory properties, neurological and immunological protection, as well as improve survival in experimental sepsis. The cholinergic anti-inflammatory pathway (CAP) is mediated by α7-nicotinic acetylcholine receptors (α7nAChRs), which play an important role in the control of systemic inflammation. We hypothesized that WJ-MSCs attenuate sepsis-induced organ injury in the presence of an activated CAP pathway. To confirm our hypothesis, we evaluated the effects of WJ-MSCs as a treatment for cardiopulmonary injury and on neuroimmunomodulation. Male Wistar rats were randomly divided into four groups: control (sham-operated); cecal ligation and puncture (CLP) alone; CLP+WJ-MSCs (1 × 106 cells, at 6 h post-CLP); and CLP+methyllycaconitine (MLA)+WJ-MSCs (5 mg/kg body wt, at 5.5 h post-CLP, and 1 × 106 cells, at 6 h post-CLP, respectively). All experiments, including the assessment of echocardiographic parameters and heart rate variability, were performed 24 h after CLP. WJ-MSC treatment attenuated diastolic dysfunction and restored baroreflex sensitivity. WJ-MSCs also increased cardiac sympathetic and cardiovagal activity. WJ-MSCs reduced leukocyte infiltration and proinflammatory cytokines, effects that were abolished by administration of a selective α7nAChR antagonist (MLA). In addition, WJ-MSC treatment also diminished apoptosis in the lungs and spleen. In cardiac and splenic tissue, WJ-MSCs downregulated α7nAChR expression, as well as reduced the phospho-STAT3-to-total STAT3 ratio in the spleen. WJ-MSCs appear to protect against sepsis-induced organ injury by reducing systemic inflammation, at least in part, via a mechanism that is dependent on an activated CAP.

The Effect of Wharton Jelly-Derived Mesenchymal Stromal Cells and Their Conditioned Media in the Treatment of a Rat Spinal Cord Injury.

The transplantation of Wharton’s jelly derived mesenchymal stromal cells (WJ-MSCs) possesses therapeutic potential for the treatment of a spinal cord injury (SCI). Generally, the main effect of MSCs is mediated by their paracrine potential. Therefore, application of WJ-MSC derived conditioned media (CM) is an acknowledged approach for how to bypass the limited survival of transplanted cells. In this study, we compared the effect of human WJ-MSCs and their CM in the treatment of SCI in rats. WJ-MSCs and their CM were intrathecally transplanted in the three consecutive weeks following the induction of a balloon compression lesion. Behavioral analyses were carried out up to 9 weeks after the SCI and revealed significant improvement after the treatment with WJ-MSCs and CM, compared to the saline control. Both WJ-MSCs and CM treatment resulted in a higher amount of spared gray and white matter and enhanced expression of genes related to axonal growth. However, only the CM treatment further improved axonal sprouting and reduced the number of reactive astrocytes in the lesion area. On the other hand, WJ-MSCs enhanced the expression of inflammatory and chemotactic markers in plasma, which indicates a systemic immunological response to xenogeneic cell transplantation. Our results confirmed that WJ-MSC derived CM offer an alternative to direct stem cell transplantation for the treatment of SCI.

Effects of Wharton's jelly-derived mesenchymal stem cells on chronic obstructive pulmonary disease.

ObjectivesChronic obstructive pulmonary disease (COPD) is a fatal disease that shortens one's life expectancy and reduces the quality of life of patients. The current known treatments for COPD can only act to alleviate the symptoms. Recently, stem cells have demonstrated efficacy in various medical areas. The aim of this study was to investigate the possibility of using human Wharton's jelly-derived mesenchymal stem cells (MSC)s for lung recovery in a COPD mouse model.MethodsHuman Wharton's jelly was obtained during natural delivery or caesarean section from healthy women. Wharton's jelly-derived MSC was confirmed with expression of CD14, CD34, CD45, CD73, CD90, and CD105 using flow cytometry. Mice model (C57BL/6) of COPD were induced by injecting 10 μL elastase into the trachea and they were divided into three treatment groups (sham, vehicle, stem cell). The sham group was not induced COPD, nor provided any treatment; the vehicle group comprised of COPD-induced mice treated with normal saline; the stem cell group comprised of COPD-induced mice treated with Wharton's jelly-derived MSCs. The vehicle and mesenchymal stem cells (5 × 104 cells) were injected in tail vein 7 days following COPD induction. Mice were euthanized 7 days after vehicle and stem cell injection, and pathologic findings were confirmed. Mean Linear Intercept (MLI) was measured after emphysema-induced alveoli were identified.ResultsCell surface markers were positive for CD105, CD90, and CD73 and negative for CD45, CD34, and CD14. Pathological tests showed that COPD-induced mice had significantly increased emphysema volume as compared with that in the sham group. The degree of emphysema in the stem cell group was reduced based on pathologic findings. The mean MLI of the sham group was measured as 38.85 ± 6.45. The mean MLI of the vehicle and stem cell groups were 163.05 ± 48.94 and 123.59 ± 30.53, respectively, and there was a statistically significant difference between the two groups (p = 0.008).ConclusionsThough the number of mice in the experiment was not large, human Wharton's jelly-derived MSCs showed pulmonary regenerative effects in the COPD mouse model. Although we cannot confirm the effects of Wharton's jelly-derived MSCs in COPD through this experiment, it can be used as a basis for a larger clinical experiment.

The synergistic effects of graphene-contained 3D-printed calcium silicate/poly-ε-caprolactone scaffolds promote FGFR-induced osteogenic/angiogenic differentiation of mesenchymal stem cells.

Graphene-contained calcium silicate (CS)/polycaprolactone (PCL) scaffold (GCP) provides an alternative solution that can bring several bone formation properties, such as osteoinductive. This study finds out the optimal percentage of graphene additive to calcium silicate and polycaprolactone mixture for excellent in vitro and in vivo bone-regeneration ability, in addition, this scaffold could fabricate by 3D printing technology and demonstrates distinct mechanical, degradation, and biological behavior. With controlled structure and porosity by 3D printing, osteogenesis and proliferation capabilities of Wharton's Jelly derived mesenchymal stem cells (WJMSCs) were significantly enhanced when cultured on 3D printed GCP scaffolds. In this study, it was also discovered that fibroblast growth factor receptor (FGFR) plays an active role in modulating differentiation behavior of WJMSCs cultured on GCP scaffolds. The validation has been proved by analyzed the decreased cell proliferation, osteogenic-related protein (ALP and OC), and angiogenic-related protein (VEGF and vWF) with FGFR knockdown on all experimental groups. Moreover, this study infers that the GCP scaffold could induce the effects of proliferation, differentiation and related protein expression on WJMSCs through FGFR pathway. In summary, this research indicated the 3D-printed GCP scaffolds own the dual bioactivities to reach the osteogenesis and vascularization for bone regeneration.

Modulation of Cellular Colonization of Porous Polyurethane Scaffolds via the Control of Pore Interconnection Size and Nanoscale Surface Modifications.

Full-scale cell penetration within porous scaffolds is required to obtain functional connective tissue components in tissue engineering applications. For this aim, we produced porous polyurethane structures with well-controlled pore and interconnection sizes. Although the influence of the pore size on cellular behavior is widely studied, we focused on the impact of the size of the interconnections on the colonization by NIH 3T3 fibroblasts and Wharton's jelly-derived mesenchymal stem cells (WJMSCs). To render the material hydrophilic and allow good material wettability, we treated the material either by plasma or by polydopamine (PDA) coating. We show that cells weakly adhere on these surfaces. Keeping the average pore diameter constant at 133 μm, we compare two structures, one with LARGE (52 μm) and one with SMALL (27 μm) interconnection diameters. DNA quantification and extracellular matrix (ECM) production reveal that larger interconnections is more suitable for cells to move across the scaffold and form a three-dimensional cellular network. We argue that LARGE interconnections favor cell communication between different pores, which then favors the production of the ECM. Moreover, PDA treatment shows a truly beneficial effect on fibroblast viability and on matrix production, whereas plasma treatment shows the same effect for WJMSCs. We, therefore, claim that both pore interconnection size and surface treatment play a significant role to improve the quality of integration of tissue engineering scaffolds.

Doping of carbon nanodots for saving cells from silver nanotoxicity: A study on recovering osteogenic differentiation potential

Silver nanoparticles are explored for many advanced biological applications including the development of antimicrobial surfaces on implants, SERS imaging, nanotherapeutics, biosensing and much more. However, recent research findings suggest silver nanoparticles provide blockade of differentiation of mesenchymal stem cells (MSCs), especially into osteogenic developmental pathway via generation of reactive oxygen species. These studies suggest that the application of silver nanoparticles in medical implants should be prohibited. In the current study, carbon nanodots (CND) supported silver clusters (AgC) is explored as a remedy to this problem. The nanostructure was synthesized in microwave irradiation induced rapid method and characterization was conducted via UV–Vis spectroscopy, fluorescence spectroscopy, HRTEM, XRD, FTIR, Raman spectroscopy, DLS, AFM, and XPS. Fluorescence spectrum showed a quantum yield of 0.25 while Raman spectroscopy showed rapid amplification of CND specific peaks implicating significant SERS property. Further in vitro biocompatibility (MTT) and bio-imaging capability was assessed culturing Wharton's Jelly-derived MSCs. In this study, its efficacy as in-situ cellular oxidative stress scavenger is also studied using NBT and DCFH-DA assay. Via ALP assay, alizarin red staining, cell membrane nanoindentation studies, PCR analysis and immunocytochemistry for osteoblast-like gene expression it was confirmed that AgCs can control silver nanoparticle-induced inhibition of osteogenic differentiation in vitro. Thus, AgCs (Carbon nanodots supported silver clusters) are not only considered to be a dual-mode bio-imaging nanoprobe but also a remedy to the silver-induced ROS generation and osteogenic differentiation blockade of MSCs.

Cycloxygenase-2 inhibition potentiates trans-differentiation of Wharton's jelly–mesenchymal stromal cells into endothelial cells: Transplantation enhances neovascularization-mediated wound repair

BackgroundNeo-vascularization, an indispensible phenomenon for tissue regeneration, facilitates repair and remodeling of wound tissues. This process is impaired in chronic wounds due to reduced number and recruitment of endothelial cells (ECs), thereby necessitating development of newer strategies to enhance the EC repertoire as a therapeutic approach. MethodsWe explored the ‘plasticity’ of Wharton's jelly derived–mesenchymal stromal cells (WJ-MSCs) using an anti-inflammatory drug-mediated enhanced trans-differentiation into ECs, based on our observation of temporal decrease in COX-2 expression during trans-differentiation of MSCs into ECs at day 7 and 14 along with mature ECs. ResultsAt a physiological level, an increased DiI-labeled acetylated-low density lipoprotein (DiI-Ac-LDL) uptake, proliferation, migration and chick chorio allantoic membrane (CAM)-vasculogenesis occurred while at a molecular level significant up-regulation in messenger RNA (mRNA) and protein expression of endothelial-specific markers, Vegfr2, Pecam, eNOS, VE-Cadh and Tie-2, along with an activated p-VEGFR2 and its downstream mediators were observed in celecoxib-preconditioned ECs as compared with WJ-MSCs. Green fluorescent protein (GFP)-expressing stable WJ-MSCs and trans-differentiated EC-D14 in the absence/presence of celecoxib were generated using antibiotic selection for intradermal transplantation at the wound site on a murine ‘excisional splinting wound’ model. Engraftment of transplanted human cells in immunosuppressant-treated mice was confirmed by a significant increase in the expression levels of human gene-specific endothelial markers at the regenerated wound sites. Morphometrically, increased vascularity and percent wound closure were observed in regenerated wounds of mice transplanted with celecoxib-preconditioned-EC-D14. ConclusionCox-2 inhibition led to an enhanced trans-differentiation of WJ-MSCs into ECs that, when transplanted, accelerated the skin regeneration by engraftment and neo-vascularization at the wound bed, suggesting a plausible new therapeutic role of celecoxib.

Extracellular Vesicles from Wharton's Jelly Mesenchymal Stem Cells Suppress CD4 Expressing T Cells Through Transforming Growth Factor Beta and Adenosine Signaling in a Canine Model.

Mesenchymal stem cells (MSCs) are widely investigated as potential therapeutic agents due to their potent immunomodulatory capacity. Although specific mechanisms by which MSC acts on immune cells are emerging, many questions remain, including the potential of extracellular vesicles (EVs) to mediate biological activities. Canine MSCs are of interest for both veterinary and comparative models of disease and have been shown to suppress CD4pos T cell proliferation. The aim of this study was to determine whether EV isolated from canine Wharton's jelly-derived MSC (WJ-MSC EV) suppresses CD4pos T cell proliferation using biochemical mechanisms previously ascribed to soluble mediators [transforming growth factor beta (TGF-β) and adenosine]. WJ-MSC EV exhibited mode of 125 nm diameter, low buoyant density (1.1 g/mL), and expression of EV proteins Alix and TSG101. Functionally, EVs inhibited CD4pos T cell proliferation in a dose-dependent manner, which was absent in EV-depleted samples and EVs from non-MSC fibroblasts. EV suppression of CD4pos T cell proliferation was inhibited by a TGF-βRI antagonist, neutralizing antibodies to TGF-β, or A2A adenosine receptor blockade. TGF-β was present on EVs as latent complexes most likely tethered to EV membrane by betaglycan. These data demonstrate that canine WJ-MSC EV utilizes TGF-β and adenosine signaling to suppress proliferation of CD4pos T cell and will enable further investigation into mechanisms of immune cell modulation, as well as refinement of WJ-MSC and their EVs for therapeutic application.

Differentiation of human mesenchymal stem cells (MSC) to dopaminergic neurons: A comparison between Wharton’s Jelly and olfactory mucosa as sources of MSCs

The generation of dopaminergic neurons from stem cells is a potential therapeutic approach to treat neurodegenerative disorders, such as Parkinson’s disease. The current study aims to investigate the potential of two different types of mesenchymal stem cells derived from human Wharton’s jelly and nasal cavity for differentiation into dopaminergic neurons. The differentiation capacities of both cell types were evaluated using real-time PCR, immunocytochemistry, flow cytometry and HPLC. Wharton’s jelly-derived mesenchymal stem cells (WJ-MSCs) are noted for their capability to differentiate into mesodermal and non-mesodermal cells, including neurons. However, it was demonstrated that having the same neuroectodermal origin as the nervous system, the olfactory ectomesenchymal stem cells (OE-MSCs) expressed the neural marker MAP2 as well as dopaminergic markers such as tyrosine hydroxylase (TH), dopamine transporter (DAT) and PITX3 to a greater extent than the WJ-MSCs both at the level of mRNA and protein. Furthermore, quantitative flow cytometric evaluation of these markers at 12 days post-induction supported the above-mentioned results. Finally, the assessment of the functionality of differentiated cells and their ability to synthesize dopamine measured by HPLC revealed that the OE-MSC-derived dopaminergic cells released almost the same amount of dopamine as that secreted by WJ-MSC-derived cells. Thus it showed the difference in their functionality to be negligible. Overall, it may be concluded that higher proliferation and differentiation capacity of OE-MSCs, along with their easier harvestability and autologous transplantability compared with WJ-MSCs, makes them a better cell source for stem cell therapy of neurodegenerative disorders which are caused by degeneration of dopaminergic neurons.

Dose-dependent In Vitro Cytotoxic Effects of Phosphorus Doped Graphene Oxide Titanium Oxide Nanocomposite (P-GO-TiO2 NC) on Caprine Wharton’s Jelly Derived Mesenchymal Stem Cells (WJ-MSCs) and Erythrocytes

Dose-dependent In Vitro Cytotoxic Effects of Phosphorus Doped Graphene Oxide Titanium Oxide Nanocomposite (P-GO-TiO2 NC) on Caprine Wharton’s Jelly Derived Mesenchymal Stem Cells (WJ-MSCs) and Erythrocytes

Isolation of a novel embryonic stem cell cord blood–derived population with in vitro hematopoietic capacity in the presence of Wharton's jelly–derived mesenchymal stromal cells

BackgroundRecent studies highlight the existence of a population of cord blood (CB)–derived stem cells that bare embryonic features (very small embryonic-like stem cells [VSELs]) as the most primitive CB-stem cell population. In the present study, we present for the first time a novel and high purity isolation method of VSELs with in vitro hematopoietic capacity in the presence of Wharton's jelly–derived mesenchymal stromal cells (WJ-MSCs). MethodsThe experimental procedure includes isolation upon gradually increased centrifugation spins and chemotaxis to Stromal cell-derived factor 1a (SDF-1a). Τhis cell population is characterized with flow cytometry, alkaline phosphatase (ALP) staining and qRT-PCR. The functional role of the isolated VSELs is assayed following co-culture with WJ-MSCs or bone marrow–derived mesenchymal stromal cells (BM-MSCs), whereas the stimulation of the quiescent VSEL population is verified via cell cycle analysis. The in vitro hematopoietic capacity is evaluated in methylcellulose cultures and also through induction of erythroid differentiation. ResultsThe final isolated subpopulation is characterized as a small-sized CD45/Lineage-/CXCR4+/CD133+/SSEA-4+cell population, positive in ALP staining and overexpressing the Oct3/4, Nanog and Sox-2 transcription factors. Upon the co-culture with MSCs, a stimulation of the quiescent VSEL population is observed. An impressive increase in the co-expression of the CD34+/CD45+ markers is observed following the co-culture with the WJ-MSCs, which is confirmed by the intense clonogenic ability suggesting in vitro differentiation toward all of the hematopoietic cell lineages and successful differentiation toward erythrocytes. DiscussionConclusively, we propose a novel, rapid and rather simplified isolation method of CB-VSELs, capable of in vitro hematopoiesis.

Overexpression of apelin in Wharton\u2019 jelly mesenchymal stem cell reverses insulin resistance and promotes pancreatic \u03b2 cell proliferation in type 2 diabetic rats

BackgroundApelin plays a key beneficial role in energy metabolism by increasing glucose uptake and insulin sensitivity; however, apelin has a short half-life because it is rapidly cleared from the circulation limiting its therapeutic benefit. The aim of this study is to create a new approach to treat type 2 diabetes by inducing prolonged expression of apelin in Wharton’s jelly-derived mesenchymal stem cells (WJ-MSCs).MethodsA type 2 diabetic rat model was given a high-fat diet combined with low-dose streptozotocin (STZ) injection. The human WJ-MSCs were isolated and subsequently transduced with apelin-expressing lentiviral particles (WJMSCs-apelin), and expression was verified by flow cytometry, Western blot, ELISA, and RT-PCR analysis. Type 2 diabetic rats were infused with either WJMSCs-apelin (2 × 106 cells) or an equivalent dose of saline through the tail vein injection 7 days after STZ injection. The therapeutic effects of each infusion group were evaluated by monitoring plasma glucose levels and performing glucose tolerance tests (OGTTs), insulin tolerance tests (IPITTs), confocal microscopy, and immunocytochemical analysis for quantitating islet beta cells. Plasma inflammatory cytokines IL-6 and TNF-α and anti-inflammatory factors adiponectin were measured as well.ResultsType 2 diabetic rats infused with WJ-MSCs-apelin significantly decreased levels of blood glucose (from 26.03 ± 2.83 to 15.85 ± 2.13 mmol/L on 7 days P < 0.001, and to 9.41 ± 2.05 on 14 days, P < 0.001). Infusion of WJMSCs-apelin not only improved significantly insulin sensitivity and glucose disposal, but also promoted endogenous pancreatic ß cell proliferation (9.6-fold increase compared to the control group). Furthermore, infusion of the WJMSCs-apelin consistently increased insulin and C-peptide levels in the plasma, and the above effects persisted up to 42 days. The inflammatory cytokines IL-6 and TNF-α were significantly decreased, whereas anti-inflammatory factor adiponectin was significantly increased after WJ-MSC-apelin infusion.ConclusionIn this study, we report a novel approach to treat type 2 diabetic rats that combines apelin gene therapy with WJ-MSC cell therapy, which could provide a promising therapeutic option for management of type 2 diabetes clinically.