Types Of Mesenchymal Stromal Cells Research Articles

RNA-sequencing reveals positional memory of multipotent mesenchymal stromal cells from oral and maxillofacial tissue transcriptomes

BackgroundMultipotent mesenchymal stromal cells (MSCs) can be isolated from numerous tissues and are attractive candidates for therapeutic clinical applications due to their immunomodulatory and pro-regenerative capacity. Although the minimum criteria for defining MSCs have been defined, their characteristics are known to vary depending on their tissue of origin.ResultsWe isolated and characterized human MSCs from three different bones (ilium (I-MSCs), maxilla (Mx-MSCs) and mandible (Md-MSCs)) and proceeded with next generation RNA-sequencing. Furthermore, to investigate the gene expression profiles among other cell types, we obtained RNA-seq data of human embryonic stem cells (ESCs) and several types of MSCs (periodontal ligament-derived MSCs, bone marrow-derived MSCs, and ESCs-derived MSCs) from the Sequence Reads Archive and analyzed the transcriptome profile. We found that MSCs derived from tissues of the maxillofacial region, such as the jaw bone and periodontal ligament, were HOX-negative, while those derived from other tissues were HOX-positive. We also identified that MSX1, LHX8, and BARX1, an essential regulator of craniofacial development, were strongly expressed in maxillofacial tissue-derived MSCs. Although MSCs may be divided into two distinct groups, the cells originated from over the neck or not, on the basis of differences in gene expression profile, the expression patterns of all CD antigen genes were similar among different type of MSCs, except for ESCs.ConclusionsOur findings suggest that MSCs from different anatomical locations, despite meeting general characterization criteria, have remarkable differences in gene expression and positional memory. Although stromal cells from different anatomical sources are generally categorized as MSCs, their differentiation potential and biological functions vary. We suggested that MSCs may retain an original tissue memory about the developmental process, including gene expression profiles. This could have an important impact when choosing an appropriate cell source for regenerative therapy using MSCs.

Cytokine physiognomies of MSCs from varied sources confirm the regenerative commitment post-coculture with activated neutrophils.

The interaction of mesenchymal stromal cells (MSCs) with paracrine signals and immunological cells, and their responses and regenerative commitment thereafter, is understudied. In the current investigation, we compared MSCs from the umbilical cord blood (UCB), dental pulp (DP), and liposuction material (LS) on their ability to respond to activated neutrophils. Cytokine profiling (interleukin-1α [IL-1α], IL-2, IL-4, IL-6, IL-8, tumor necrosis factor-α [TNF-α], interferon-γ [IFN-γ], transforming growth factor-β [TGF-β]), cellular proliferation and osteogenic differentiation patterns were assessed. The results showed largely comparable cytokine profiles with higher TNF-α and IFN-γ levels in LSMSCs owing to their mature cellular phenotype. The viability and proliferation between LS/DP/UCB MSCs were comparable in the coculture group, while direct activation of MSCs with lipopolysaccharide (LPS) showed comparable proliferation with significant cell death in UCB MSCs and slightly higher cell death in the other two types of MSC. Furthermore, when MSCs post-neutrophil exposure were induced for osteogenic differentiation, though all the MSCs devoid of the sources differentiated, we observed rapid and significant turnover of DPMSCs positive of osteogenic markers rather than LS and UCB MSCs. We further observed a significant turnover of IL-1α and TGF-β at mRNA and cytokine levels, indicating the commitment of MSCs to differentiate through interacting with immunological cells or bacterial products like neutrophils or LPS, respectively. Taken together, these results suggest that MSCs have more or less similar cytokine responses devoid of their anatomical niche. They readily switch over from the cytokine responsive cell phenotype at the immunological microenvironment to differentiate and regenerate tissue in response to cellular signals.

Mesenchymal stromal cells from dermal and adipose tissues induce macrophage polarization to a pro-repair phenotype and improve skin wound healing

The process of wound healing restores skin homeostasis but not full functionality; thus, novel therapeutic strategies are needed to accelerate wound closure and improve the quality of healing. In this context, tissue engineering and cellular therapies are promising approaches. Although sharing essential characteristics, mesenchymal stromal cells (MSCs) isolated from different tissues might have distinct properties. Therefore, the aim of this study was to comparatively investigate, by a mouse model in vivo assay, the potential use of dermal-derived MSCs (DSCs) and adipose tissue–derived MSCs (ASCs) in improving skin wound healing. Human DSCs and ASCs were delivered to full-thickness mouse wounds by a collagen-based scaffold (Integra Matrix). We found that the association of both DSCs and ASCs with the Integra accelerated wound closure in mice compared with the biomaterial only (control). Both types of MSCs stimulated angiogenesis and extracellular matrix remodeling, leading to better quality scars. However, the DSCs showed smaller scar size,superior extracellular matrix deposition, and greater number of cutaneous appendages. Besides, DSCs and ASCs reduced inflammation by induction of macrophage polarization from a pro-inflammatory (M1) to a pro-repair (M2) phenotype. In conclusion, both DSCs and ASCs were able to accelerate the healing of mice skin wounds and promote repair with scars of better quality and more similar to healthy skin than the empty scaffold. DSCs associated with Integra induced superior overall results than the Integra alone, whereas scaffolds with ASCs showed an intermediate effect, often not significantly better than the empty biomaterial.

Stem cell-based interventions for the prevention and treatment of germinal matrix-intraventricular haemorrhage in preterm infants.

Germinal matrix-intraventricular haemorrhage (GMH-IVH) remains a substantial issue in neonatal intensive care units worldwide. Current therapies to prevent or treat GMH-IVH are limited. Stem cell-based therapies offer a potential therapeutic approach to repair, restore, and/or regenerate injured brain tissue. These preclinical findings have now culminated in ongoing human neonatal studies. To determine the benefits and harms of stem cell-based interventionsfor prevention or treatment of germinal matrix-intraventricular haemorrhage (GM-IVH) in preterm infants. We used the standard search strategy of Cochrane Neonatal to search the Cochrane Central Register of Controlled Trials (CENTRAL; 2019, Issue 1), in the Cochrane Library; MEDLINE via PubMed (1966 to 7 January 2019); Embase (1980 to 7 January 2019); and the Cumulative Index to Nursing and Allied Health Literature (CINAHL) (1982 to 7 January 2019). We also searched clinical trials databases, conference proceedings, and reference lists of retrieved articles for randomised controlled trials and quasi-randomised trials. We attempted to identify randomised controlled trials, quasi-randomised controlled trials, and cluster trials comparing (1) stem cell-based interventions versus control; (2) mesenchymal stromal cells (MSCs)of type or source versus MSCs of other type or source; (3) stem cell-based interventions other than MSCs of type or source versus stem cell-based interventions other than MSCs of other type or source; or (4) MSCs versus stem cell-based interventions other than MSCs. For prevention studies, we included extremely preterm infants (less than 28 weeks' gestation), 24 hours of age or less, without ultrasound diagnosis of GM-IVH; for treatment studies, we included preterm infants (less than 37 weeks' gestation), of any postnatal age, with ultrasound diagnosis of GM-IVH. For each of the included trials, two review authors independently planned to extract data (e.g. number of participants, birth weight, gestational age, type and source of MSCs, other stem cell-based interventions) and assess the risk of bias (e.g. adequacy of randomisation, blinding, completeness of follow-up). Primary outcomes considered in this review are all-cause neonatal mortality, major neurodevelopmental disability, GM-IVH, and extension of pre-existing non-severe GM-IVH. We planned to use the GRADE approach to assess the quality of evidence. Our search strategy yielded 769 references. We did not find any completed studies for inclusion. One randomised controlled trial is currently registered and ongoing. Five phase 1 trials are described in the excluded studies. Currently no evidence is available to show the benefits or harms of stem cell-based interventions for treatment or prevention of GM-IVH in preterm infants.

Characterization of the Effects of Mesenchymal Stromal Cells on Mouse and Human Islet Function.

Islet transplantation has the potential to cure type 1 diabetes, but current transplantation protocols are not optimal and there is extensive loss of islet β‐cell insulin secretory function during the immediate post‐transplantation period. Studies using experimental models of diabetes have shown that the coculture of islets with mesenchymal stromal cells (MSCs) prior to transplantation improves graft function, but several variables differed among research groups (e.g., type of MSCs used and the treatment conditions). We have therefore assessed the effects of MSCs on mouse and human islets by investigating the importance of tissue source for MSCs, the coculture protocol configuration and length, the effect of activated MSCs, and different β‐cell secretory stimuli. MSCs derived from adipose tissue (aMSCs) were the most effective at supporting β‐cell insulin secretion in both mouse and human islets, in a direct contact coculture configuration. Preculture with aMSCs enhanced both phases of glucose‐induced insulin secretion and further enhanced secretory responses to the non‐nutrients carbachol and arginine. These effects required a coculture period of 48–72 hours and were not dependent on activation of the MSCs. Thus, direct contact coculture with autologous, adipose‐derived MSCs for a minimum of 48 hours before implantation is likely to be an effective addition to human islet transplantation protocols. stem cells translational medicine 2019;8:935&944

Towards a consensus potency assay for mesenchymal stromal cells: a matrix analysis of cell source, donor variability and inflammatory stimuli to refine surrogate markers of immunomodulation

Background & Aim Reliable metrics to assess the therapeutic potential of mesenchymal stromal cell (MSC) batches are needed for successful translation of MSC-based products. Here, the veracity of proposed biomarkers was evaluated by matrix analysis of 3 donor populations each of umbilical cord-derived (UC) and bone marrow-derived (BM) MSCs primed by 3 different proinflammatory cytokines. Methods, Results & Conclusion High quality UC MSCs (Tissue Regeneration Therapeutics Inc., CAN) and BM MSCs (RoosterBio Inc. (RBI), USA) were cultured in xeno-free media (RBI). MSCs were primed by 24 hour exposure to TNF-α (50 ng/ml), IFN-γ (50 ng/ml) or IL-1β (80 pg/ml) in protein-free media (RBI) (n=3 replicates). Cells from replicate flasks were pooled for RNA extraction and transcriptome analysis by Affymetrix GeneChip™ U133A 2.0 microarray. Secreted proteins were analyzed by multiplex ELISA (BioPlex 27-plex Human Inflammation Panel II). The data were interrogated against the ISCT's proposed panel of 47 reference genes and proteins that report MSC priming to IFN-γ. The only gene markers of activation shared across MSC types and stimulants were ICAM1 (p 0.05), which increased >2-fold in primed versus resting cells. Eighteen genes responded significantly (p Eight secreted biomarkers proposed by the ISCT panel were represented in the multiplex data; none were induced by IFN-γ. TNF-α elicited significantly more IL-8 (CXCL8) and IL-1ra (IL1RN) from both MSC types, and specifically increased RANTES (CCL5) and IL-6 output from BM MSCs and MCP-1 (CCL2) from UC MSCs. In BM MSCs only, priming with TNF-α or IL-1β elevated IP-10 (CXCL10) levels, as well as IL-8 and IL-1ra. These outputs correlated with the array data, except for IL-1ra and IP-10 whose mRNA levels were unchanged. These data suggest that only a subset of proposed biomarkers may have broad utility for developing standardized potency assays for a range of MSC products. Ongoing work seeks to identify additional surrogate markers with applicability across MSC types, variable donors and inflammatory stimuli.

Isolation and multipotential differentiation of mesenchymal stromal cell‑like progenitor cells from human bladder.

Various types of mesenchymal stromal cells (MSCs) have been used in urological tissue engineering but to date the existence of MSCs has not been reported in the human bladder. The present study provided evidence that a small number of MSC-like cells exist in the human bladder and designated this class of cells ‘human bladder-derived MSC-like cells’ (hBSCs). It was demonstrated that hBSCs can be cultured to yield a large population. These hBSCs expressed the surface markers of MSCs and exhibited the capacity for osteogenic, adipogenic and chondrogenic differentiation. On induction with appropriate media in vitro, hBSCs could differentiate into bladder-associated cell types, including urothelial, endothelial and smooth muscle cell-like lineages. In addition, the average telomerase activity of adult hBSCs was higher compared with adult human bone marrow-derived MSCs, but lower than that of human umbilical cord Wharton's jelly-derived MSCs. These findings may inspire future studies on the role of hBSCs in urological tissue engineering applications and in other fields.

Comprehensive characterization of chorionic villi-derived mesenchymal stromal cells from human placenta

BackgroundStudies in which mesenchymal stromal cells (MSC) from the placenta are compared with multiple MSC types from other sources are rare. The chorionic plate of the human placenta is mainly composed of fetal blood vessels embedded in fetal stroma tissue, lined by trophoblastic cells and organized into chorionic villi (CV) structures.MethodsWe comprehensively characterized human MSC collected from postnatal human chorionic villi of placenta (CV-MSC) by analyzing their growth and proliferation potential, differentiation, immunophenotype, extracellular matrix production, telomere length, aging phenotype, and plasticity.ResultsImmunophenotypic characterization of CV-MSC confirmed the typical MSC marker expression as defined by the International Society for Cellular Therapy. The surface marker profile was consistent with increased potential for proliferation, vascular localization, and early myogenic marker expression. CV-MSC retained multilineage differentiation potential and extracellular matrix remodeling properties. They have undergone reduced telomere loss and delayed onset of cellular senescence as they aged in vitro compared to three other MSC sources. We present evidence that increased human telomerase reverse transcriptase gene expression could not explain the exceptional telomere maintenance and senescence onset delay in cultured CV-MSC. Our in-vitro tumorigenesis detection assay suggests that CV-MSC are not prone to undergo malignant transformation during long-term in-vitro culture. Besides SOX2 expression, no other pluripotency features were observed in early and late passages of CV-MSC.ConclusionsOur work brings forward two remarkable characteristics of CV-MSC, the first being their extended life span as a result of delayed replicative senescence and the second being a delayed aged phenotype characterized by improved telomere length maintenance. MSC from human placenta are very attractive candidates for stem cell-based therapy applications.

Mesenchymal Stromal Cell Therapy for Pancreatitis: A Systematic Review.

Background Based on animal studies, adult mesenchymal stromal cells (MSCs) are promising for the treatment of pancreatitis. However, the best type of this form of cell therapy and its mechanism of action remain unclear. Methods We searched the PubMed, Web of Science, Scopus, Google Scholar, and Clinical Trials.gov websites for studies using MSCs as a therapy for both acute and chronic pancreatitis published until September 2017. Results We identified 276 publications; of these publications, 18 met our inclusion criteria. In animal studies, stem cell therapy was applied more frequently for acute pancreatitis than for chronic pancreatitis. No clinical trials were identified. MSC therapy ameliorated pancreatic inflammation in acute pancreatitis and pancreatic fibrosis in chronic pancreatitis. Bone marrow and umbilical cord MSCs were the most frequently administered cell types. Due to the substantial heterogeneity among the studies regarding the type, source, and dose of MSCs used, conducting a meta-analysis was not feasible to determine the best type of MSCs. Conclusion The available data were insufficient for determining the best type of MSCs for the treatment of acute or chronic pancreatitis; therefore, clinical trials investigating the use of MSCs as therapy for pancreatitis are not warranted.

Mesenchymal Stromal Cell Therapy: After the Gold Rush.

Few medical advancements have captivated public interest as much as stem cell therapies. In fact, one of the original stem cell therapies was described in this very journal nearly 50 years ago1,2: the mesenchymal stem cell. Now known as mesenchymal stromal cells (MSCs),3 a more appropriate name due to their limited capacity to differentiate into somatic cell types, clinical evidence for their potential benefits is steadily mounting. The current nomenclature change reflects 2 important clinical advantages for MSC therapy in comparison to bona fide stem cells: no risk of teratoma formation and fewer ethical concerns related to isolation and expansion methods. Moreover, the ease in isolation and expansion makes MSC therapy economically attractive. These advantages coupled with credible immunosuppressive effects experimentally have led to much interest in their use in solid organ,4 cellular,5 and vascularized composite transplantation6,7 as well as other immune-mediated pathologies,8 and a rush toward clinical trials. Although there is some evidence that these cells have therapeutic effects through their regenerative properties,9-11 it is principally their broad immunosuppressive activity, and perhaps induction of regulatory T cells,12 that is of interest. The first clinical studies of MSC therapy were reported over a decade ago and hundreds of trials continue to be registered in European and North American databases. However, very little is understood regarding the clinical efficacy of these cells in transplantation. The paucity of evidence is striking given the widespread commercialization and availability of MSC therapy in many countries. The summary of the current state of the art of MSC therapy in renal transplantation from Reinders and colleagues13 is therefore timely and welcome. When investigating the field, the lack of consistency in cell description and the variety of isolation techniques mean that comparisons between studies are impossible. However, the International Society for Cellular Therapy’s position statement14 has provided much needed clarity. Their definition requires 3 components for defining MSCs: adherence to plastic, specific surface antigen expression, and the ability to differentiate into osteoblasts, adipocytes, and chondrocytes in vitro. Naturally, these are minimum requirements, and additional clinical release criteria will be required in different countries. Helpfully, the description provided in the review in this issue of Transplantation provides useful information regarding the types of MSCs used in ongoing trials in renal transplantation. MSCs may be derived from several sources, with each source exhibiting specific immunological properties.15 The most commonly used sources in reported clinical trials are bone marrow and adipose tissue. Reinders and colleagues16 summarized 7 published trials in renal transplantation with a total of 141 patients treated with MSCs derived from bone marrow. In these limited studies, there are hints of improved outcomes with reduced requirements for drug immunosuppression as well as enhanced renal function. However, the difficulty in defining useful short-term endpoints in renal transplantation clinical trials limits the ability to understand the true efficacy of MSC therapy. More concerning is the evidence of an increase in the incidence of opportunistic infections which was reported in a study from the same group in Leiden. This is not unexpected given that MSCs exert immunosuppressive effects that are nonspecific and not regulated or directed in the same manner as a leukocyte. Ostensibly, this broad activity raises some doubts regarding the overall benefits of MSC therapy versus traditional immunosuppression.17 There remain many questions regarding the specific techniques of clinical application as well as the practicalities of Good Manufacturing Practice manufacture, an issue which is examined by Reinders et al. A discussion of these issues is crucial given the lack of consistency and anecdotal nature of many clinical trials.

Ontogenic Identification and Analysis of Mesenchymal Stromal Cell Populations during Mouse Limb and Long Bone Development.

SummaryBone-derived mesenchymal stromal cells (MSCs) differentiate into multiple lineages including chondro- and osteogenic fates and function in establishing the hematopoietic compartment of the bone marrow. Here, we analyze the emergence of different MSC types during mouse limb and long bone development. In particular, PDGFRαposSCA-1pos (PαS) cells and mouse skeletal stem cells (mSSCs) are detected within the PDGFRαposCD51pos (PαCD51) mesenchymal progenitors, which are the most abundant progenitors in early limb buds and developing long bones until birth. Long-bone-derived PαS cells and mSSCs are most prevalent in newborn mice, and molecular analysis shows that they constitute distinct progenitor populations from the earliest stages onward. Differential expression of CD90 and CD73 identifies four PαS subpopulations that display distinct chondro- and osteogenic differentiation potentials. Finally, we show that cartilage constructs generated from CD90pos PαS cells are remodeled into bone organoids encompassing functional endothelial and hematopoietic compartments, which makes these cells suited for bone tissue engineering.

Correlation of CDC42 Activity with Cell Proliferation and Palmitate-Mediated Cell Death in Human Umbilical Cord Wharton's Jelly Derived Mesenchymal Stromal Cells.

RHO GTPases regulate cell migration, cell-cycle progression, and cell survival in response to extracellular stimuli. However, the regulatory effects of RHO GTPases in mesenchymal stromal cells (MSCs) are unclear. Herein, we show that CDC42 acts as an essential factor in regulating cell proliferation and also takes part in lipotoxic effects of palmitate in human umbilical cord Wharton's jelly derived MSCs (hWJ-MSCs). Cultured human bone marrow, adipose tissue, and hWJ-MSC derived cells had varying pro-inflammatory cytokine secretion levels and cell death rates when treated by palmitate. Strikingly, the proliferation rate of these types of MSCs correlated with their sensitivity to palmitate. A glutathione-S-transferase pull-down assay demonstrated that hWJ-MSCs had the highest activation of CDC42, which was increased by palmitate treatment in a time-dependent manner. We demonstrated that palmitate-induced synthesis of pro-inflammatory cytokines and cell death was attenuated by shRNA against CDC42. In CDC42 depleted hWJ-MSCs, population-doubling levels were notably decreased, and phosphorylation of ERK1/2 and p38 MAPK was reduced. Our data therefore suggest a mechanistic role for CDC42 activity in hWJ-MSC proliferation and identified CDC42 activity as a promising pharmacological target for ameliorating lipotoxic cell dysfunction and death.

Comparison and immunobiological characterization of retinoic acid inducible gene-I-like receptor expression in mesenchymal stromal cells

Due to their immunomodulatory and regenerative properties, Mesenchymal stromal cells (MSC) have generated major interests in several clinical settings including transplantation and inflammatory diseases. MSC functions can be influenced by their tissue origin. Their microenvironment strongly affects their biology notably through TLR sensing. In this study, we show that MSC isolated from four different sources express another type of cytosolic pathogen recognition receptors known as retinoic acid inducible gene-I (RIG-I)-like receptors (RLR). RLR activation in MSC induces the production of Type I IFN (IFN-β) and Type III IFN (IFN-λ1). The highest producers are adipose tissue(AT)-MSC. We further show that Interferon production is induced through TBK1/IKK-ε signaling and IRF7 phosphorylation. Depending on MSC source, the knockdown of TLR3 and/or RIG-I decreases the MSC response to RLR ligand poly(I:C)/Lyovec. Among the different MSC types, AT-MSCs display the highest sensitivity to viral stimuli as shown by the alteration of their viability after prolonged stimulation. Our work indicates that this could be linked to an increase of pro-apoptotic Noxa expression. Finally, the expression of IDO1 and LIF upon RLR activation indicate the increase of MSC immunomodulatory potential, especially in AT-MSCs. Altogether, these data should be considered when designing MSC-based therapy in clinical settings where inflammation or infection are present.

Expression of Surface Molecules in Human Mesenchymal Stromal Cells Co-Cultured with Nucleated Umbilical Cord Blood Cells.

We studied the expression of different classes of surface molecules (CD13, CD29, CD40, CD44, CD54, CD71, CD73, CD80, CD86, CD90, CD105, CD106, CD146, HLA-I, and HLA-DR) in mesenchymal stromal cells from human umbilical cord and bone marrow during co-culturing with nucleated umbilical cord blood cells. Expression of the majority of surface markers in both types of mesenchymal stromal cells was stable and did not depend on the presence of the blood cells. Significant differences were found only for cell adhesion molecules CD54 (ICAM-1) and CD106 (VCAM-1) responsible for direct cell-cell contacts with leukocytes and only for bone marrow derived cells.

Comparison of the expression profile of surface molecular markers on mesenchymal stromal cell cultures isolated from human endometrium and umbilical cord

Endometrial mesenchymal stromal cells (eMSCs), along with mesenchymal stromal cells (MSCs) isolated from other tissues, are promising for use in regenerative medicine. The benefits of eMSCs include their presence in adults, simplicity of isolation, high proliferative and differentiation capacity. In this study, we have employed the flow cytometry technique to assess expression of 28 molecular markers on the surface of two eMSCs cultures. The culture of MSCs isolated from Wharton's jelly of the umbilical cord (uMSCs) was used as a reference, because uMSCs were studied in details earlier and demonstrated their effectiveness in vivo. Both types of MSCs demonstrated similar expression profiles. They included stem cells surface molecules, cell adhesion molecules and their ligands, some receptor molecules responsible for cell metabolism and proliferation, as well as immunological response molecules.

Different Balance of Wnt Signaling in Adult and Fetal Bone Marrow-Derived Mesenchymal Stromal Cells.

Mesenchymal stromal cells (MSCs) are applied as novel therapeutics for their regenerative and immune-suppressive capacities. Clinical applications, however, require extensive expansion of MSCs. Fetal bone marrow-derived MSCs (FBMSCs) proliferate faster than adult bone marrow-derived MSC (ABMSCs). To optimize expansion and function of MSC in general, we explored the differences between ABMSC and FBMSC. Gene expression profiling implicated differential expression of genes encoding proteins in the Wnt signaling pathway, including excreted inhibitors of Wnt signaling, particularly by ABMSC. Both MSC types had a similar basal level of canonical Wnt signaling. Abrogation of autocrine Wnt production by inhibitor of Wnt production-2 (IWP2) reduced canonical Wnt signaling and cell proliferation of FBMSCs, but hardly affected ABMSC. Addition of exogenous Wnt3a, however, induced expression of the target genes lymphocyte enhancer-binding factor (LEF) and T-cell factor (TCF) faster and at lower Wnt3a levels in ABMSC compared to FBMSC. Medium replacement experiments indicated that ABMSC produce an inhibitor of Wnt signaling that is effective on ABMSC itself but not on FBMSC, whereas FBMSC excrete (Wnt) factors that stimulate proliferation of ABMSC. In contrast, FBMSC were not able to support hematopoiesis, whereas ABMSC displayed hematopoietic support sensitive to IWP2, the inhibitor of Wnt factor excretion. In conclusion, ABMSC and FBMSC differ in their Wnt signature. While FBMSC produced factors, including Wnt signals, that enhanced MSC proliferation, ABMSC produced Wnt factors in a setting that enhanced hematopoietic support. Thus, further unraveling the molecular basis of this phenomenon may lead to improvement of clinical expansion protocols of ABMSCs.

Effects of pro-inflammatory cytokines on chondrogenesis of equine mesenchymal stromal cells derived from bone marrow or synovial fluid

Mesenchymal stromal cells (MSCs) have the capacity to differentiate into cells of mesenchymal lineage, such as chondrocytes, and have potential for use in regeneration of equine articular cartilage. MSCs instilled intra-articularly would be exposed to the inflamed environment associated with equine osteoarthritis (OA), which may compromise their function and ability to heal a cartilaginous defect. The aim of this study was to assess the ability of equine adult MSCs to differentiate into chondrocytes when stimulated with pro-inflammatory cytokines. MSCs derived from equine bone marrow (BM) and from synovial fluid (SF) were cultured in chondrogenic induction medium containing transforming growth factor (TGF)-β1. BM-derived MSCs (BMMSCs) and SF-derived MSCs (SFMSCs) were stimulated with 100 ng/mL interferon (IFN)-γ and 10 ng/mL tumor necrosis factor (TNF)-α. Chondrogenic differentiation was measured quantitatively with the glycosaminoglycan (GAG) assay and qualitatively by immunofluorescence (IF) for SOX-9, TGF-β1, aggrecan and collagen II. The viability of equine MSCs was maintained in the presence of IFN-γ and TNF-α, but production of GAGs from both types of MSCs was decreased in stimulated medium. Exposure of BMMSCs to pro-inflammatory cytokines reduced the levels of SOX-9, TGF-β1, aggrecan and collagen II, whereas exposure of SFMSCs to these cytokines reduced the levels of aggrecan only. These data suggest that pro-inflammatory cytokines do not affect proliferation of MSCs, but could inhibit chondrogenesis of MSCs.

Different wound healing properties of dermis, adipose, and gingiva mesenchymal stromal cells.

Oral wounds heal faster and with better scar quality than skin wounds. Deep skin wounds where adipose tissue is exposed, have a greater risk of forming hypertrophic scars. Differences in wound healing and final scar quality might be related to differences in mesenchymal stromal cells (MSC) and their ability to respond to intrinsic (autocrine) and extrinsic signals, such as human salivary histatin, epidermal growth factor, and transforming growth factor beta1. Dermis-, adipose-, and gingiva-derived MSC were compared for their regenerative potential with regards to proliferation, migration, and matrix contraction. Proliferation was assessed by cell counting and migration using a scratch wound assay. Matrix contraction and alpha smooth muscle actin was assessed in MSC populated collagen gels, and also in skin and gingival full thickness tissue engineered equivalents (reconstructed epithelium on MSC populated matrix). Compared to skin-derived MSC, gingiva MSC showed greater proliferation and migration capacity, and less matrix contraction in full thickness tissue equivalents, which may partly explain the superior oral wound healing. Epidermal keratinocytes were required for enhanced adipose MSC matrix contraction and alpha smooth muscle actin expression, and may therefore contribute to adverse scarring in deep cutaneous wounds. Histatin enhanced migration without influencing proliferation or matrix contraction in all three MSC, indicating that salivary peptides may have a beneficial effect on wound closure in general. Transforming growth factor beta1 enhanced contraction and alpha smooth muscle actin expression in all three MSC types when incorporated into collagen gels. Understanding the mechanisms responsible for the superior oral wound healing will aid us to develop advanced strategies for optimal skin regeneration, wound healing and scar formation.

Horse adipose-derived mesenchymal stromal cells constitutively produce membrane vesicles: a morphological study.

Mesenchymal stromal cells (MSCs) are multipotent somatic cells that can differentiate into a variety of mature cell types. Over recent years, their biological in vitro and in vivo properties have elicited great expectations in the field of regenerative medicine, immunotherapy and tumour treatment. An increasing number of experimental observations suggest that their biological effects are probably related to a paracrine mechanism via the release of trophic factors and cytokines as well as through the production of membrane vesicles (MVs). These are nanometric membrane-bound structures, comprising shedding vesicles (SV) and exosomes (Ex), that enclose and transfer signalling molecules to target cells. We hypothesized that MVs may be implicated in the biological effects of MSCs from horse adipose tissue (E-AdMSCs), a type of MSC that has been extensively studied in recent years for its remarkable efficacy in tissue regeneration. By means of electron microscopy, we ascertained, for the first time, that equine adipose-derived MSCs constitutively produce MVs (E-Ad-MSCs). The analysis of MVs separated by ultracentrifugation allowed us to describe their general morphological features. Through the examination of cell monolayers by TEM, additionally, we distinguished the different pathways of SV and Ex formation, demonstrating that both fractions are produced by E-AdMSC. The accurate description of MV heterogeneous morphological characteristics led us to emphasize the possible implications of the relationship between different morphologies versus different functions. The data presented in this paper has an additional value, as they can be noteworthy for horses as well as for other mammalian species, including humans.

In vitro characterization of patches of human mesenchymal stromal cells.

Stem cells may represent an excellent strategy to improve the healing of skin ulcers. Today the administration mode of stem cells to skin defects remains unsatisfactory. Delivering stem cells with topical treatments represents a new strategy and answering the patients' need. Mesenchymal stromal cells (MSC) have been shown to improve wound healing of cutaneous lesions and amniotic membrane (AM) is known to represent a natural scaffold for cells. The aim of this study is to develop a tissue-engineered product combining MSC and AM for clinical use. In this work we investigated whether the stromal matrix of intact human AM could constitute a scaffold for human MSC derived from either bone marrow (BM) or adipose tissue (AT). For this purpose, clinical-grade AM, MSC, and culture medium were used. We performed experiments of short-term adherence and proliferation for 15 days after the seeding of the cells. Morphological aspects and secretion profiles of MSC onto AM were studied, respectively, by scanning electron microscopy and Luminex analysis. Results demonstrated that the stromal matrix allow the adherence in much greater amount of MSC from BM or AT compared to 2D material. Experiments of proliferation showed that both kinds of MSC could proliferate on the stromal matrix and remain viable 15 days after the seeding of the cells. The 3D analysis of MSC culture demonstrated that both types of MSC invaded the stromal matrix and grew in multiple layers while retaining their fibroblastic morphology. By studying the secretion profile of MSC onto the stromal matrix, we found that both kinds of MSC secrete important cytokines and growth factors for wound healing of cutaneous lesions, such as vascular endothelial growth factor, hepatocyte growth factor, and basic fibroblast growth factor. In conclusion, these results suggest that the stromal matrix of AM seeded with MSC represents a bioactive scaffold that should be evaluated in patients with a nonhealing cutaneous wound.