scholarly journals In VitroExpansion and Characterization of Mesenchymal Stromal Cells from Peritoneal Dialysis Effluent in a Human Protein Medium

2018 ◽  
Vol 2018 ◽  
pp. 1-10 ◽  
Author(s):  
Bo La Han ◽  
Lan Zhou ◽  
Qiunong Guan ◽  
Gerald da Roza ◽  
Hao Wang ◽  
...  
Keyword(s):  
Peritoneal Dialysis ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Therapeutic Potential ◽  
Human Protein ◽  
Positive Staining ◽  
Alizarin Red ◽  
Hla Dr ◽  
Marker Expression ◽  
Peritoneal Dialysis Effluent

The therapeutic potential of mesenchymal stromal cells (MSCs) from various tissue origins have extensively been explored in both experimental and clinical studies, and peritoneal dialysis effluent-derived MSC (pMSC) may be an easily obtainable MSC source for clinical applications. In this study, we expanded and characterized the pMSCs after expansion in a human protein culture medium. The pMSCs were expanded in plastic dishes with the human protein medium. MSC marker expression was examined by flow cytometry. Spherical formation was tested by hanging drop method, and osteogenic, adipogenic, and chondrogenic differentiation capacities were confirmed by positive staining with Alizarin red, Oil red O, and Alcian blue, respectively. Here, we showed that after four passages of culturing in plastic dishes, pMSCs in the human protein medium displayed a homogeneous pattern of classical MSC markers (positive: CD29, CD44, CD73, CD90, and CD166; negative: CD14, CD34, CD45, CD79a, CD105, CD146, CD271, HLA-DR, SSEA-4, and Stro-1), while in the standard medium, pMSCs from some donors were CD45 or HLA-DR positive. For nonclassical MSC markers, pMSCs were CD200 positive from all the donors, negative for CD163, CD271, CD36, and CD248, and either positive or negative for CD274 and CD140b. Further, pMSCs from the human protein medium had the spherical formation capacity and multipotent differentiation capacityin vitro. In conclusion, upon expansion in a human protein medium, pMSCs showed a differential MSC marker expression profile from those of bone marrow or adipose tissue-derived MSCs and could maintain the multipotency. The therapeutic potential of the pMSCs requires further investigation.

Crosslinked layered surfaces of heparin and poly(L-lysine) enhance mesenchymal stromal cells behavior in the presence of soluble interferon gamma

2021 ◽  
Author(s):  
Mahsa Haseli ◽  
Luis Pinzon-Herrera ◽  
Jorge Almodovar
Keyword(s):  
Interferon Gamma ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Photoelectron Spectroscopy ◽  
Therapeutic Potential ◽  
Layer By Layer ◽  
Alizarin Red ◽  
Final Layer ◽  
Extracellular Matrix Components ◽  
Ifn Γ

Human mesenchymal stromal cells (hMSCs) are multipotent cells that have been proposed for the treatment of immune-mediated diseases. Culturing hMSCs on tissue culture plastic reduces their therapeutic potential in part due to the lack of extracellular matrix components. The aim of this study is to evaluate multilayers of heparin and poly(L-lysine) (HEP/PLL) as a bioactive surface for hMSCs stimulated with soluble interferon gamma (IFN‐γ). Multilayers were formed, via layer-by-layer assembly, with HEP as the final layer and supplemented with IFN-γ in the culture medium. Multilayer construction and chemistry were confirmed using Azure A staining, quartz crystal microbalance (QCM), and X-ray photoelectron spectroscopy. hMSCs adhesion, viability, and differentiation, were assessed. Results showed that (HEP/PLL) multilayer coatings were poorly adhesive for hMSCs. However, performing chemical crosslinking using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide (EDC/NHS) significantly enhanced hMSCs adhesion and viability. The immunosuppressive properties of hMSCs cultured on crosslinked (HEP/PLL) multilayers were confirmed by measuring the level of indoleamine 2,3-dioxygenase (IDO) secretion. Lastly, hMSCs cultured on crosslinked (HEP/PLL) multilayers in the presence of soluble IFN- γ successfully differentiated towards the osteogenic and adipogenic lineages as confirmed by Alizarin red, and oil-red O staining, as well as alkaline phosphatase activity. This study suggests that crosslinked (HEP/PLL) films can modulate hMSCs response to soluble factors, which may improve hMSCs-based therapies aimed at treating several immune diseases.

scholarly journals Protection of the Peritoneal Membrane by Peritoneal Dialysis Effluent-Derived Mesenchymal Stromal Cells in a Rat Model of Chronic Peritoneal Dialysis

2019 ◽  
Vol 2019 ◽  
pp. 1-11 ◽  
Author(s):  
Lan Zhou ◽  
Ming Zong ◽  
Qiunong Guan ◽  
Gerald da Roza ◽  
Hao Wang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Peritoneal Dialysis ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Therapeutic Potential ◽  
Ex Vivo ◽  
Flow Cytometric Analysis ◽  
Peritoneal Membrane ◽  
Peritoneal Mesothelial Cells

Peritoneal dialysis (PD) is a renal replacement option for patients with end-stage renal disease. However, a long-term exposure to hypertonic PD solutions leads to peritoneal membrane (PM) injury, resulting in ultrafiltration (UF) failure. This study was designed to primarily evaluate efficacy of PD effluent-derived mesenchymal stromal cells (pMSCs) in the prevention of PM injury in rats. The pMSCs were isolated from PD effluent. Male Wistar rats received daily intraperitoneal (IP) injection of 10 mL of Dianeal (4.25% dextrose) and were treated with pMSCs (1.2‐1.5×106/rat/wk, IP). UF was determined by IP injection of 30 mL of Dianeal (4.25% dextrose) with dwell time of 1.5 h, and PM injury was examined by histology. Apoptosis was quantitated by using flow cytometric analysis, and gene expression by using the PCR array and Western blot. Here, we showed that as compared to naive control, daily IP injection of the Dianeal PD solution for 6 weeks without pMSC treatment significantly reduced UF, which was associated with an increase in both PM thickness and blood vessel, while pMSC treatment prevented the UF loss and reduced PM injury and blood vessels. In vitro incubation with pMSC-conditioned medium prevented cell death in cultured human peritoneal mesothelial cells (HPMCs) and downregulated proinflammatory (i.e., CXCL6, NOS2, IL1RN, CCL5, and NR3C1) while upregulated anti-inflammatory (i.e., CCR1, CCR4, IL9, and IL-10) gene expression in activated THP1 cells. In conclusion, pMSCs prevent bioincompatible PD solution-induced PM injury and UF decline, suggesting that infusing back ex vivo-expanded pMSCs intraperitoneally may have therapeutic potential for reduction of UF failure in PD patients.

Evaluation of Potential Application of Wharton’s Jelly-Derived Human Mesenchymal Stromal Cells and its Conditioned Media for Dermal Regeneration using Rat Wound Healing Model

2021 ◽  
pp. 1-14
Author(s):  
Caroline Mathen ◽  
Mrunal Ghag Sawant ◽  
Raghubansh Gupta ◽  
Wilfrid Dsouza ◽  
Shilpa G. Krishna
Keyword(s):  
Wound Healing ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Therapeutic Potential ◽  
Wound Care ◽  
Free Cell ◽  
Conditioned Media ◽  
Human Umbilical Cord ◽  
Human Mscs ◽  
Wound Model

Mesenchymal stromal cells and the derived conditioned media represent an area of tremendous medical interest and, among other clinical applications, are currently being extensively explored for wound healing. The aim of this study was to comparatively evaluate the wound healing potential of xeno-free human umbilical cord-derived mesenchymal stromal cells (MSCs) and the conditioned media (CM) in a full-thickness excision wound model in rats. The evaluation parameters included rate of wound healing, serum cytokine analyses, collagen content, histopathology, and hyperspectral imaging as an independent qualitative and quantitative tool. Both the cell-based and cell-free approaches scored better in lower inflammation, as evidenced in lower IL-10 and stable IL-6 levels, and improved rate of wound healing (<i>p</i> &#x3c; 0.0001). More importantly, no adverse reaction or rejection was observed although human MSCs and CM were used in a xenogeneic model. The presence of hFGF, hHGF, hGCSF, hIL-1Ra, hVEGF, and hIL-6 in the secretome may elucidate the regenerative potential of the xeno-free cell-based and cell-free approaches which have translational value for advanced wound care. The results revealed the therapeutic potential of both the cell-based and cell-free approaches for wound healing.

scholarly journals Characterization and Comparison of Human and Ovine Mesenchymal Stromal Cells from Three Corresponding Sources

2020 ◽  
Vol 21 (7) ◽  
pp. 2310 ◽  
Author(s):  
El-Mustapha Haddouti ◽  
Thomas M. Randau ◽  
Cäcilia Hilgers ◽  
Werner Masson ◽  
Klaus J. Walgenbach ◽  
...  
Keyword(s):  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Therapeutic Option ◽  
Large Animal Model ◽  
Osteogenic Potential ◽  
Large Animal ◽  
Human Mscs ◽  
Mineralization Process ◽  
Hla Dr ◽  
Different Sources

Currently, there is an increasing focus on mesenchymal stromal cells (MSC) as therapeutic option in bone pathologies as well as in general regenerative medicine. Although human MSCs have been extensively characterized and standardized, ovine MSCs are poorly understood. This limitation hampers clinical progress, as sheep are an excellent large animal model for orthopedic studies. Our report describes a direct comparison of human and ovine MSCs from three corresponding sources under the same conditions. All MSCs presented solid growth behavior and potent immunomodulatory capacities. Additionally, we were able to identify common positive (CD29, CD44, CD73, CD90, CD105, CD166) and negative (CD14, CD34, CD45, HLA-DR) surface markers. Although both human and ovine MSCs showed strong osteogenic potential, direct comparison revealed a slower mineralization process in ovine MSCs. Regarding gene expression level, both human and ovine MSCs presented a comparable up-regulation of Runx2 and a trend toward down-regulation of Col1A during osteogenic differentiation. In summary, this side by side comparison defined phenotypic similarities and differences of human and ovine MSCs from three different sources, thereby contributing to a better characterization and standardization of ovine MSCs. The key findings shown in this report demonstrate the utility of ovine MSCs in preclinical studies for MSC-based therapies.

scholarly journals Vadadustat, a HIF Prolyl Hydroxylase Inhibitor, Improves Immunomodulatory Properties of Human Mesenchymal Stromal Cells

Cells ◽  
2020 ◽  
Vol 9 (11) ◽  
pp. 2396
Author(s):  
Katarzyna Zielniok ◽  
Anna Burdzinska ◽  
Beata Kaleta ◽  
Radoslaw Zagozdzon ◽  
Leszek Paczek
Keyword(s):  
Real Time ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Real Time Pcr ◽  
Mononuclear Cells ◽  
Therapeutic Potential ◽  
Human Peripheral Blood ◽  
Immunomodulatory Activity ◽  
Peripheral Blood Mononuclear ◽  
Immunomodulatory Properties

The therapeutic potential of mesenchymal stromal cells (MSCs) is largely attributed to their immunomodulatory properties, which can be further improved by hypoxia priming. In this study, we investigated the immunomodulatory properties of MSCs preconditioned with hypoxia-mimetic Vadadustat (AKB-6548, Akebia). Gene expression analysis of immunomodulatory factors was performed by real-time polymerase chain reaction (real-time PCR) on RNA isolated from six human bone-marrow derived MSCs populations preconditioned for 6 h with 40 μM Vadadustat compared to control MSCs. The effect of Vadadustat preconditioning on MSCs secretome was determined using Proteome Profiler and Luminex, while their immunomodulatory activity was assessed by mixed lymphocyte reaction (MLR) and Culturex transwell migration assays. Real-time PCR revealed that Vadadustat downregulated genes related to immune system: IL24, IL1B, CXCL8, PDCD1LG1, PDCD1LG2, HIF1A, CCL2 and IL6, and upregulated IL17RD, CCL28 and LEP. Vadadustat caused a marked decrease in the secretion of IL6 (by 51%), HGF (by 47%), CCL7 (MCP3) (by 42%) and CXCL8 (by 40%). Vadadustat potentiated the inhibitory effect of MSCs on the proliferation of alloactivated human peripheral blood mononuclear cells (PBMCs), and reduced monocytes-enriched PBMCs chemotaxis towards the MSCs secretome. Preconditioning with Vadadustat may constitute a valuable approach to improve the therapeutic properties of MSCs.

Therapeutic potential of allogeneic mesenchymal stromal cells transplantation for lupus nephritis

Lupus ◽  
2018 ◽  
Vol 27 (13) ◽  
pp. 2161-2165 ◽  
Author(s):  
J Barbado ◽  
S Tabera ◽  
A Sánchez ◽  
J García-Sancho
Keyword(s):  
Lupus Nephritis ◽  
Mesenchymal Stromal Cells ◽  
Lupus Erythematosus ◽  
Stromal Cells ◽  
Therapeutic Potential ◽  
Activity Index ◽  
Disease Activity Index ◽  
Controlled Clinical Trial ◽  
Allogeneic Mscs

Animal and human studies have suggested the potential of mesenchymal stromal cells (MSCs) to treat systemic lupus erythematosus (SLE). Here, we present the results of compassionate MSC treatments for three SLE patients to provide the proof of concept for a randomized and controlled clinical trial. Three patients of different ethnicities who suffer from chronic SLE, and who presented with class IV active proliferative nephritis confirmed by biopsy, were treated with allogeneic MSCs from healthy donors. Ninety million cells were infused intravenously into each patient during high and very high activity disease flare-ups and follow-up was continued for 9 months. Multi-organic affectation was quantified by the SLE disease activity index (SLEDAI), and indicators of lupus nephritis activity, such as proteinuria, as well as lymphocyte and monocyte antigens and anti-HLA antibodies were measured at 1, 3, 6, and 9 months after treatment. Proteinuria levels improved dramatically during the 1st month after treatment and the ameliorations were sustained throughout the follow-up period. SLEDAI scores revealed early, durable, and substantial remissions that were complete for two patients and partial for the third patient and that permitted medication doses to be reduced 50–90%. These favourable outcomes support completion of the randomized and controlled MSC trial for SLE.

In Vitro Characterization of Multipotent Mesenchymal Stromal Cells Isolated from Palatal Subepithelial Tissue Grafts

2013 ◽  
Vol 19 (2) ◽  
pp. 370-380 ◽  
Author(s):  
Alexandra Roman ◽  
Andrada Şoancă ◽  
Adrian Florea ◽  
Emőke Páll
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Embryoid Body ◽  
Basic Research ◽  
Multipotent Mesenchymal Stromal Cells ◽  
Positive Staining ◽  
Basic Characteristics ◽  
Tissue Grafts

AbstractThe aim of this study was to analyze whether the mesenchymal stromal cells (MSCs) isolated from palatal tissue grafts harvested in order to cover gingival recessions have the basic characteristics of stem cells. The palatal tissue cells were processed using a special culture medium that stimulated the development of only undifferentiated cellular lines. Cells at passage 4 were evaluated by flow cytometry to examine the expression of specific surface markers and were tested for multilineage differentiation capacity. These cells collected at passage 4 were also investigated for the capacity to cluster into embryoid body aggregates. Palatal MSCs displayed positive staining for the mesenchymal markers CD29, CD73, CD105, CD 49e, and CD44, but did not express hematopoietic markers CD34/45. The palatal MSCs successfully differentiated into osteogenic, adipogenic, and chondrogenic lineages. When seeded in special conditions, palatal MSCs propagated into unattached spheres resembling embryoid body aggregates consisting both of differentiated and undifferentiated cells as revealed at the ultrastructural evaluation. It is concluded that the isolated palatal MSCs fulfilled the basic criteria defining the stem cells. This new source of stem cells characterized here for the first time opens new perspectives on possible applications in basic research and in regenerative medicine.

Role Of Long Pentraxin 3 (PTX3) In Wound Closure Induced By Bone Marrow-Derived Mesenchymal Stromal Cells

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 1220-1220
Author(s):  
Claudia Cappuzzello ◽  
Andrea Doni ◽  
Erica Dander ◽  
Fabio Pasqualini ◽  
Manuela Nebuloni ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Wound Closure ◽  
Conflicts Of Interest ◽  
Histological Evaluation ◽  
Pentraxin 3 ◽  
Alizarin Red ◽  
Matrix Deposition

Abstract Although several studies have shown the capacity of mesenchymal stromal cells (MSCs) to repair and regenerate different tissues, the mechanisms underlying these processes are not understood. Long Pentraxin 3 (PTX3) is a multifunctional protein produced by MSCs and other cell subsets upon activation with inflammatory cytokines. PTX3 is involved in innate immunity, inflammation and extracellular matrix deposition. In the present study we analyzed the potential role of PTX3 in wound repair process induced by MSCs. PTX3 knockout MSCs (PTX3-/-MSCs) were collected from bone marrow of PTX3-/- mice. After 3-5 culture passages the expression of surface markers was analyzed by flow cytometry and their osteogenic and adipogenic differentiation capacity was detected by alizarin red O and oil red S staining, respectively. The ability of PTX3-/-MSCs to abrogate T cell proliferation was evaluated by co-culturing MSCs and PBMCs previously activated with Phytohaemagglutinin. Finally, equal number of both PTX3-/-MSCs and wild type (WT) MSCs were implanted into excisional wounds created by a biopsy punch on the back of allogenic WT and PTX3-/- mice. Wound area was measured up to 14 day and calculated using an image analysis program. The wound specimens were collected at 2, 7 and 14 days and processed for histological analysis. We demonstrated that PTX3-/-MSCs, similarly to WT MSCs, displayed typical fibroblastoid morphology, they expressed common MSC markers and were able to differentiate into adipocytes and osteoblasts. In addition, they drastically decreased the mitogen-induced proliferation of lymphocyte. Importantly, in a mouse model of wound healing, PTX3-/- MSCs showed a highly significant defect in wound closure compared to WT MSCs at each time point. Histological evaluation of skin samples treated with PTX3-/- MSCs showed a reduction of the granulation tissue and a significant increase of neutrophils (GR-1+) in the wound bed. Moreover, wounds treated with PTX3-/- MSCs were characterized by an excessive accumulation of fibrin at the 2nd day after injury. Accordingly, PTX3-/- MSCs showed a defective ability to degrade the fibrin matrix in vitro. Finally, PTX3-/- MSCs failed to close the ulcers in PTX3-/- mice. In conclusion, we demonstrated that PTX3 deficiency does not alter the phenotype and the capacity of MSCs to differentiate into mesengetic lineages; however, the production of PTX3 represents an essential requirement for MSC ability of enhancing tissue repair. Disclosures: No relevant conflicts of interest to declare.

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