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Obesity and type 2 diabetes are becoming a global sociobiomedical burden. Beige adipocytes are emerging as key inducible actors and putative relevant therapeutic targets for improving metabolic health. However, in vitro models of human beige adipose tissue are currently lacking and hinder research into this cell type and biotherapy development. Unlike traditional bottom-up engineering approaches that aim to generate building blocks, here a scalable system is proposed to generate pre-vascularized and functional human beige adipose tissue organoids using the human stromal vascular fraction of white adipose tissue as a source of adipose and endothelial progenitors. This engineered method uses a defined biomechanical and chemical environment using tumor growth factor β (TGFβ) pathway inhibition and specific gelatin methacryloyl (GelMA) embedding parameters to promote the self-organization of spheroids in GelMA hydrogel, facilitating beige adipogenesis and vascularization. The resulting vascularized organoids display key features of native beige adipose tissue including inducible Uncoupling Protein-1 (UCP1) expression, increased uncoupled mitochondrial respiration, and batokines secretion. The controlled assembly of spheroids allows to translate organoid morphogenesis to a macroscopic scale, generating vascularized centimeter-scale beige adipose micro-tissues. This approach represents a significant advancement in developing in vitro human beige adipose tissue models and facilitates broad applications ranging from basic research to biotherapies., (© 2023 The Authors. Advanced Science published by Wiley-VCH GmbH.)

Background: Native bone marrow (BM) mesenchymal stem/stromal cells (BM-MSCs) participate in generating and shaping the skeleton and BM throughout the lifespan. Moreover, BM-MSCs regulate hematopoiesis by contributing to the hematopoietic stem cell niche in providing critical cytokines, chemokines and extracellular matrix components. However, BM-MSCs contain a heterogeneous cell population that remains ill-defined. Although studies on the taxonomy of native BM-MSCs in mice have just started to emerge, the taxonomy of native human BM-MSCs remains unelucidated., Methods: By using single-cell RNA sequencing (scRNA-seq), we aimed to define a proper taxonomy for native human BM non-hematopoietic subsets including endothelial cells (ECs) and mural cells (MCs) but with a focal point on MSCs. To this end, transcriptomic scRNA-seq data were generated from 5 distinct BM donors and were analyzed together with other transcriptomic data and with computational biology analyses at different levels to identify, characterize and classify distinct native cell subsets with relevant biomarkers., Results: We could ascribe novel specific biomarkers to ECs, MCs and MSCs. Unlike ECs and MCs, MSCs exhibited an adipogenic transcriptomic pattern while co-expressing genes related to hematopoiesis support and multilineage commitment potential. Furthermore, by a comparative analysis of scRNA-seq of BM cells from humans and mice, we identified core genes conserved in both species. Notably, we identified MARCKS, CXCL12, PDGFRA, and LEPR together with adipogenic factors as archetypal biomarkers of native MSCs within BM. In addition, our data suggest some complex gene nodes regulating critical biological functions of native BM-MSCs together with a preferential commitment toward an adipocyte lineage., Conclusions: Overall, our taxonomy for native BM non-hematopoietic compartment provides an explicit depiction of gene expression in human ECs, MCs and MSCs at single-cell resolution. This analysis helps enhance our understanding of the phenotype and the complexity of biological functions of native human BM-MSCs., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Bone marrow (BM) mesenchymal stromal cells (MSCs) are abnormal in multiple myeloma (MM) and play a critical role by promoting growth, survival, and drug resistance of MM cells. We observed higher Toll-like receptor 4 (TLR4) gene expression in MM MSCs than in MSCs from healthy donors. At the clinical level, we highlighted that TLR4 expression in MM MSCs evolves in parallel with the disease stage. Thus, we reasoned that the TLR4 axis is pivotal in MM by increasing the protumor activity of MSCs. Challenging primary MSCs with TLR4 agonists increased the expression of CD54 and interleukin-6 (IL-6), 2 factors directly implicated in MM MSC-MM cell crosstalk. Then, we evaluated the therapeutic efficacy of a TLR4 antagonist combined or not with conventional treatment in vitro with MSC-MM cell coculture and in vivo with the Vk*MYC mouse model. Selective inhibition of TLR4 specifically reduced the MM MSC ability to support the growth of MM cells in an IL-6-dependent manner and delayed the development of MM in the Vk*MYC mouse model by altering the early disease phase in vivo. For the first time, we demonstrate that specific targeting of the pathological BM microenvironment via TLR4 signaling could be an innovative approach to alter MM pathology development., (© 2022 by The American Society of Hematology. Licensed under Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0), permitting only noncommercial, nonderivative use with attribution. All other rights reserved.)

Bone is the most frequent metastasis site for breast cancer. As well as dramatically increasing disease burden, bone metastases are also an indicator of poor prognosis. One of the main challenges in investigating bone metastasis in breast cancer is engineering in vitro models that replicate the features of in vivo bone environments. Such in vitro models ideally enable the biology of the metastatic cells to mimic their in vivo behavior as closely as possible. Here, taking benefit of cutting-edge technologies both in microfabrication and cancer cell biology, we have developed an in vitro breast cancer bone-metastasis model. To do so we first 3D printed a bone scaffold that reproduces the trabecular architecture and that can be conditioned with osteoblast-like cells, a collagen matrix, and mineralized calcium. We thus demonstrated that this device offers an adequate soil to seed primary breast cancer bone metastatic cells. In particular, patient-derived xenografts being considered as a better approach than cell lines to achieve clinically relevant results, we demonstrate the ability of this biomimetic bone niche model to host patient-derived xenografted metastatic breast cancer cells. These patient-derived xenograft cells show a long-term survival in the bone model and maintain their cycling propensity, and exhibit the same modulated drug response as in vivo. This experimental system enables access to the idiosyncratic features of the bone microenvironment and cancer bone metastasis, which has implications for drug testing., (Copyright © 2020. Published by Elsevier Ltd.)

Human adipose-derived stem/stromal cells (hASCs) can differentiate into specialized cell types and thereby contribute to tissue regeneration. As such, hASCs have drawn increasing attention in cell therapy and regenerative medicine, not to mention the ease to isolate them from donors. Culture conditions are critical for expanding hASCs while maintaining optimal therapeutic capabilities. Here, we identified a role for transforming growth factor β1 (TGFβ1) in culture medium in influencing the fate of hASCs during in vitro cell expansion. Human ASCs obtained after expansion in standard culture medium (Standard-hASCs) and in endothelial cell growth medium 2 (EGM2-hASCs) were characterized by high-throughput transcriptional studies, gene set enrichment analysis and functional properties. EGM2-hASCs exhibited enhanced multipotency capabilities and an immature phenotype compared with Standard-hASCs. Moreover, the adipogenic potential of EGM2-hASCs was enhanced, including toward beige adipogenesis, compared with Standard-hASCs. In these conditions, TGFβ1 acts as a critical factor affecting the immaturity and multipotency of Standard-hASCs, as suggested by small mother of decapentaplegic homolog 3 (SMAD3) nuclear localization and phosphorylation in Standard-hASCs vs EGM2-hASCs. Finally, the typical priming of Standard-hASCs into osteoblast, chondroblast, and vascular smooth muscle cell (VSMC) lineages was counteracted by pharmacological inhibition of the TGFβ1 receptor, which allowed retention of SMAD3 into the cytoplasm and a decrease in expression of osteoblast and VSMC lineage markers. Overall, the TGFβ1 pathway appears critical in influencing the commitment of hASCs toward osteoblast, chondroblast, and VSMC lineages, thus reducing their adipogenic potential. These effects can be counteracted by using EGM2 culture medium or chemical inhibition of the TGFβ1 pathway., (©AlphaMed Press 2020.)

Multiple myeloma (MM) is an incurable B cell neoplasia characterized by the accumulation of tumor plasma cells within the bone marrow (BM). As a consequence, bone osteolytic lesions develop in 80% of patients and remain even after complete disease remission. We and others had demonstrated that BM-derived mesenchymal stromal cells (MSCs) are abnormal in MM and thus cannot be used for autologous treatment to repair bone damage. Adipose stromal cells (ASCs) represent an interesting alternative to MSCs for cellular therapy. Thus, in this study, we wondered whether they could be a good candidate in repairing MM bone lesions. For the first time, we present a transcriptomic, phenotypic, and functional comparison of ASCs from MM patients and healthy donors (HDs) relying on their autologous MSC counterparts. In contrast to MM MSCs, MM ASCs did not exhibit major abnormalities. However, the changes observed in MM ASCs and the supportive property of ASCs on MM cells question their putative and safety uses at an autologous or allogenic level., Competing Interests: The authors declare that there is no conflict of interest regarding the publication of this article., (Copyright © 2020 Nicolas Espagnolle et al.)

Mesenchymal stem cells (MSCs) are isolated from multiple biological tissues-adult bone marrow and adipose tissues and neonatal tissues such as umbilical cord and placenta. In vitro, MSCs show biological features of extensive proliferation ability and multipotency. Moreover, MSCs have trophic, homing/migration and immunosuppression functions that have been demonstrated both in vitro and in vivo. A number of clinical trials are using MSCs for therapeutic interventions in severe degenerative and/or inflammatory diseases, including Crohn's disease and graft-versus-host disease, alone or in combination with other drugs. MSCs are promising for therapeutic applications given the ease in obtaining them, their genetic stability, their poor immunogenicity and their curative properties for tissue repair and immunomodulation. The success of MSC therapy in degenerative and/or inflammatory diseases might depend on the robustness of the biological functions of MSCs, which should be linked to their therapeutic potency. Here, we outline the fundamental and advanced concepts of MSC biological features and underline the biological functions of MSCs in their basic and translational aspects in therapy for degenerative and/or inflammatory diseases.

Native human subcutaneous adipose tissue (AT) is well organized into unilocular adipocytes interspersed within dense vascularization. This structure is completely lost under standard culture conditions and may impair the comparison with native tissue. Here, we developed a 3-D model of human white AT reminiscent of the cellular architecture found in vivo. Starting with adipose progenitors derived from the stromal-vascular fraction of human subcutaneous white AT, we generated spheroids in which endogenous endothelial cells self-assembled to form highly organized endothelial networks among stromal cells. Using an optimized adipogenic differentiation medium to preserve endothelial cells, we obtained densely vascularized spheroids containing mature adipocytes with unilocular lipid vacuoles. In vivo study showed that when differentiated spheroids were transplanted in immune-deficient mice, endothelial cells within the spheroids connected to the recipient circulatory system, forming chimeric vessels. In addition, adipocytes of human origin were still observed in transplanted mice. We therefore have developed an in vitro model of vascularized human AT-like organoids that constitute an excellent tool and model for any study of human AT.

Mesenchymal stem/stromal cells (MSCs) delivered as cell therapy to individuals with degenerative and/or inflammatory disorders can help improve organ features and resolve inflammation, as demonstrated in preclinical studies and to some extent in clinical studies. MSCs have trophic, homing/migration, and immunosuppression functions, with many benefits in therapeutics. MSC functions are thought to depend on the paracrine action of soluble factors and/or the expression of membrane-bound molecules, mostly belonging to the molecular class of adhesion molecules, chemokines, enzymes, growth factors, and interleukins. Cutting-edge studies underline bioactive exchanges, including that of ions, nucleic acids, proteins, and organelles transferred from MSCs to stressed cells, thereby improving the cells' survival and function. From this aspect, MSC death modulation function appears as a decisive biological function that could carry a significant part of the therapeutic effects of MSCs. Identifying the function and modes of actions of MSCs in modulating cell death may be exploited to enhance consistency and efficiency of cell therapy that is based on MSCs as medical treatment for degenerative and/or inflammatory diseases. Here, we review the essentials of MSC functions in modulating cell death in unfit cells, and its modes of actions based on current advances and outline the clinical implications.

Osteosarcoma (OS) is suspected to originate from dysfunctional mesenchymal stromal/stem cells (MSC). We sought to identify OS-derived cells (OSDC) with potential cancer stem cell (CSC) properties by comparing OSDC to MSC derived from bone marrow of patients. This study included in vitro characterization with sphere forming assays, differentiation assays, cytogenetic analysis, and in vivo investigations of their tumorigenicity and tumor supportive capacities. Primary cell lines were isolated from nine high-grade OS samples. All primary cell lines demonstrated stromal cell characteristics. Compared to MSC, OSDC presented a higher ability to form sphere clones, indicating a potential CSC phenotype, and were more efficient at differentiation towards osteoblasts. None of the OSDC displayed the complex chromosome rearrangements typical of high grade OS and none of them induced tumors in immunodeficient mice. However, two OSDC demonstrated focused genomic abnormalities. Three out of seven, and six out of seven OSDC showed a supportive role on local tumor development, and on metastatic progression to the lungs, respectively, when co-injected with OS cells in nude mice. The observation of OS-associated stromal cells with rare genetic abnormalities and with the capacity to sustain tumor progression may have implications for future tumor treatments., Competing Interests: The authors declare no conflict of interest. The founding sponsors had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, and in the decision to publish the results.

The possibility of using adipose tissue-derived stromal cells (ATSC) as alternatives to bone marrow-derived stromal cells (BMSC) for bone repair has garnered interest due to the accessibility, high cell yield, and rapid in vitro expansion of ATSC. For clinical relevance, their bone forming potential in comparison to BMSC must be proven. Distinct differences between ATSC and BMSC have been observed in vitro and comparison of osteogenic potential in vivo is not clear to date. The aim of the current study was to compare the osteogenesis of human xenofree-expanded ATSC and BMSC in vitro and in an ectopic nude mouse model of bone formation. Human MSC were implanted with biphasic calcium phosphate biomaterials in subcutis pockets for 8 weeks. Implant groups were: BMSC, ATSC, BMSC and ATSC mixed together in different ratios, as well as MSC primed with either osteogenic supplements (250 μM ascorbic acid, 10 mM β-glycerolphosphate, and 10 nM dexamethasone) or 50 ng/ml recombinant bone morphogenetic protein 4 prior to implantation. In vitro results show osteogenic gene expression and differentiation potentials of ATSC. Despite this, ATSC failed to form ectopic bone in vivo, in stark contrast to BMSC, although osteogenic priming did impart minor osteogenesis to ATSC. Neovascularization was enhanced by ATSC compared with BMSC; however, less ATSC engrafted into the implant compared with BMSC. Therefore, in the content of bone regeneration, the advantages of ATSC over BMSC including enhanced angiogenesis, may be negated by their lack of osteogenesis and prerequisite for osteogenic differentiation prior to transplantation. Stem Cells Translational Medicine 2017;6:2160-2172., (© 2017 The Authors Stem Cells Translational Medicine published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.)

Mesenchymal stem (stromal) cells (MSCs) are being investigated for treating degenerative and inflammatory disorders because of their reparative and immunomodulatory properties. Intricate mechanisms relate cell death processes with immune responses, which have implications for degenerative and inflammatory conditions. We review the therapeutic value of MSCs in terms of preventing regulated cell death (RCD). When cells identify an insult, specific intracellular pathways are elicited for execution of RCD processes, such as apoptosis, necroptosis, and pyroptosis. To some extent, exacerbated RCD can provoke an intense inflammatory response and vice versa. Emerging studies are focusing on the molecular mechanisms deployed by MSCs to ameliorate the survival, bioenergetics, and functions of unfit immune or nonimmune cells. Given these aspects, and in light of MSC actions in modulating cell death processes, we suggest the use of novel functional in vitro assays to ensure the potency of MSCs for preventing RCD. Such analyses should be associated with existing functional assays measuring the anti-inflammatory capabilities of MSCs in vitro. MSCs selected on the basis of two in vitro functional criteria (i.e., prevention of inflammation and RCD) could possess optimal therapeutic efficacy in vivo. In addition, we underline the implications of these perspectives in clinical studies of MSC therapy, with particular focus on acute respiratory distress syndrome. Stem Cells Translational Medicine 2017;6:713-719., (© 2016 The Authors Stem Cells Translational Medicine published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.)

The biological effects of indium-tin-oxide (ITO) are of considerable importance because workers exposed to indium compounds have been diagnosed with interstitial lung disease or pulmonary alveolar proteinosis; however, the pathophysiology of these diseases is undefined. Here, mice intraperitoneally inoculated with ITO-nanoparticles (ITO-NPs) resulted in peritonitis dependent in NLRP3 inflammasome, with neutrophils recruitment and interleukin-1β (IL-1β) production. Withal peritoneal macrophages exposed ex vivo to ITO-NPs caused IL-1β secretion and cytolysis. Further, alveolar macrophages exposed to ITO-NPs in vitro showed ITO-NP endocytosis and production of tumor necrosis factor-α (TNF-α) and IL-1β, ensued cell death by cytolysis. This cell death was RIPK1-independent but caspase1-dependent, and thus identified as pyroptosis. Endocytosis of ITO-NPs by activated THP-1 cells induced pyroptosis with IL-1β/TNF-α production and cytolysis, but not in activated THP-1 cells with knockdown of NLRP3, ASC, or caspase1. However, exposing activated THP-1 cells with NLRP3 or ASC knockdown to ITO-NPs resulted in cell death but without cytolysis, with deficiency in IL-1β/TNF-α, and revealing features of apoptosis. While, mesenchymal stem cells (MSCs) co-cultured with macrophages impaired both inflammation and cell death induced by ITO-NPs. Together, our findings provide crucial insights to the pathophysiology of respiratory diseases caused by ITO particles, and identify MSCs as a potent therapeutic.

Similar to other adult tissue stem/progenitor cells, bone marrow mesenchymal stem/stromal cells (BM MSCs) exhibit heterogeneity at the phenotypic level and in terms of proliferation and differentiation potential. In this study such a heterogeneity was reflected by the CD200 protein. We thus characterized CD200(pos) cells sorted from whole BM MSC cultures and we investigated the molecular mechanisms regulating CD200 expression. After sorting, measurement of lineage markers showed that the osteoblastic genes RUNX2 and DLX5 were up-regulated in CD200(pos) cells compared to CD200(neg) fraction. At the functional level, CD200(pos) cells were prone to mineralize the extra-cellular matrix in vitro after sole addition of phosphates. In addition, osteogenic cues generated by bone morphogenetic protein 4 (BMP4) or BMP7 strongly induced CD200 expression. These data suggest that CD200 expression is related to commitment/differentiation towards the osteoblastic lineage. Immunohistochemistry of trephine bone marrow biopsies further corroborates the osteoblastic fate of CD200(pos) cells. However, when dexamethasone was used to direct osteogenic differentiation in vitro, CD200 was consistently down-regulated. As dexamethasone has anti-inflammatory properties, we assessed the effects of different immunological stimuli on CD200 expression. The pro-inflammatory cytokines interleukin-1β and tumour necrosis factor-α increased CD200 membrane expression but down-regulated osteoblastic gene expression suggesting an additional regulatory pathway of CD200 expression. Surprisingly, whatever the context, i.e. pro-inflammatory or pro-osteogenic, CD200 expression was down-regulated when nuclear-factor (NF)-κB was inhibited by chemical or adenoviral agents. In conclusion, CD200 expression by cultured BM MSCs can be induced by both osteogenic and pro-inflammatory cytokines through the same pathway: NF-κB., (© 2016 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

RNA delivery is an attractive strategy to achieve transient gene expression in research projects and in cell- or gene-based therapies. Despite significant efforts investigating vector-directed RNA transfer, there is still a requirement for better efficiency of delivery to primary cells and in vivo. Retroviral platforms drive RNA delivery, yet retrovirus RNA-packaging constraints limit gene transfer to two genome-molecules per viral particle. To improve retroviral transfer, we designed a dimerization-independent MS2-driven RNA packaging system using MS2-Coat-retrovirus chimeras. The engineered chimeric particles promoted effective packaging of several types of RNAs and enabled efficient transfer of biologically active RNAs in various cell types, including human CD34(+) and iPS cells. Systemic injection of high-titer particles led to gene expression in mouse liver and transferring Cre-recombinase mRNA in muscle permitted widespread editing at the ROSA26 locus. We could further show that the VLPs were able to activate an osteoblast differentiation pathway by delivering RUNX2- or DLX5-mRNA into primary human bone-marrow mesenchymal-stem cells. Thus, the novel chimeric MS2-lentiviral particles are a versatile tool for a wide range of applications including cellular-programming or genome-editing.

The aim of this study was to assess the immune modulatory properties of human mesenchymal stromal cells obtained from bone marrow (BM-MSCs), fat (ASCs), and cord blood (CB-MSCs) in the presence of a hydroxyapatite and tricalcium-phosphate (HA/TCP) biomaterial as a scaffold for MSC delivery. In resting conditions, a short-term culture with HA/TCP did not modulate the anti-apoptotic and suppressive features of the various MSC types toward T, B, and NK cells; in addition, when primed with inflammatory cytokines, MSCs similarly increased their suppressive capacities in the presence or absence of HA/TCP. The long-term culture of BM-MSCs with HA/TCP induced an osteoblast-like phenotype with upregulation of OSTERIX and OSTEOCALCIN, similar to what was obtained with dexamethasone and, to a higher extent, with bone morphogenetic protein 4 (BMP-4) treatment. MSC-derived osteoblasts did not trigger immune cell activation, but were less efficient than undifferentiated MSCs in inhibiting stimulated T and NK cells. Interestingly, their suppressive machinery included not only the activation of indoleamine-2,3 dioxygenase (IDO), which plays a central role in T-cell inhibition, but also cyclooxygenase-2 (COX-2) that was not significantly involved in the immune modulatory effect of human undifferentiated MSCs. Since COX-2 is significantly involved in bone healing, its induction by HA/TCP could also contribute to the therapeutic activity of MSCs for bone tissue engineering.

Bone marrow mesenchymal stem cells (MSCs) are plastic adherent cells that can differentiate into various tissue lineages, including osteoblasts, adipocytes and chondrocytes. However, this progenitor property is not shared by all cells within the MSC population. In addition, MSCs vary in their proliferation capacity and expression of markers. Because of heterogeneity of CD146 expression in the MSC population, we compared CD146(-/Low) and CD146(High) cells under clonal conditions and after sorting of the non-clonal cell population to determine whether this expression is associated with specific functions. CD146(-/Low) and CD146(High) bone marrow MSCs did not differ in colony-forming unit-fibroblast number, osteogenic, adipogenic and chondrogenic differentiation or in vitro haematopoietic-supportive activity. However, CD146(-/Low) clones proliferated slightly but significantly faster than did CD146(High) clones. In addition, a strong expression of CD146 molecule was associated with a commitment to a vascular smooth muscle cell (VSMC) lineage characterized by a strong up-regulation of calponin-1 and SM22α expression and an ability to contract collagen matrix. Thus, within a bone marrow MSC population, certain subpopulations characterized by high expression of CD146, are committed towards a VSMC lineage., (© 2013 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

Mesenchymal stem cells (MSCs) are multipotent cells that can be obtained from several sources such as bone marrow and adipose tissue. Depending on the culture conditions, they can differentiate into osteoblasts, chondroblasts, adipocytes, or neurons. In this regard, they constitute promising candidates for cell-based therapy aimed at repairing damaged tissues. In addition, MSCs display immunomodulatory properties through the expression of soluble factors including HLA-G. We here analyse both immunogenicity and immunosuppressive capacity of MSCs derived from bone marrow and adipose tissue before and after osteodifferentiation. Results show that HLA-G expression is maintained after osteodifferentiation and can be boosted in inflammatory conditions mimicked by the addition of IFN-γ and TNF-α. Both MSCs and osteodifferentiated MSCs are hypoimmunogenic and exert immunomodulatory properties in HLA-mismatched settings as they suppress T cell alloproliferation in mixed lymphocyte reactions. Finally, addition of biomaterials that stimulate bone tissue formation did not modify MSC immune properties. As MSCs combine both abilities of osteoregeneration and immunomodulation, they may be considered as allogenic sources for the treatment of bone defects.

Both human leukocyte antigen G (HLA-G) and multipotential mesenchymal stem/stromal cells (MSCs) exhibit immunomodulatory functions. In allogeneic tranplantation, the risks of acute and chronic rejection are still high despite improvement in immunosuppressive treatments, and the induction of a state of tolerance to alloantigens is not achieved. Immunomodulatory properties of MSCs and HLA-G in human allogeneic tranplantation to induce tolerance appears attractive and promising. Interestingly, we and others have demonstrated that MSCs can express HLA-G. In this review, we focus on the expression of HLA-G by MSCs and discuss how to ensure and improve the immunomodulatory properties of MSCs by selectively targeting MSCs expressing HLA-G (MSCs(HLA-G+)). We also discuss the possible uses of MSCs(HLA-G+) for therapeutic purposes, notably, to overcome acute and chronic immune rejection in solid-organ allogeneic transplantation in humans. Since MSCs are phenotypically and functionally heterogeneous, it is of primary interest to have specific markers ensuring that they have strong immunosuppressive potential and HLA-G may be a valuable candidate., (© AlphaMed Press.)

Bone homeostasis is maintained by the balance between bone-forming osteoblasts and bone-degrading osteoclasts. Osteoblasts have a mesenchymal origin whereas osteoclasts belong to the myeloid lineage. Osteoclast and osteoblast communication occurs through soluble factors secretion, cell-bone interaction and cell-cell contact, which modulate their activities. CD200 is an immunoglobulin superfamilly member expressed on various types of cells including mesenchymal stem cells (MSCs). CD200 receptor (CD200R) is expressed on myeloid cells such as monocytes/macrophages. We assume that CD200 could be a new molecule involved in the control of osteoclastogenesis and could play a role in MSC-osteoclast communication in humans. In this study, we demonstrated that soluble CD200 inhibited the differentiation of osteoclast precursors as well as their maturation in bone-resorbing cells in vitro. Soluble CD200 did not modify the monocyte phenotype but inhibited the receptor activator of nuclear factor kappa-B ligand (RANKL) signaling pathway as well as the gene expression of osteoclast markers such as osteoclast-associated receptor (OSCAR) and nuclear factor of activated T cells cytoplasmic 1 (NFATc1). Moreover, MSCs inhibited osteoclast formation, which depended on cell-cell contact and was associated with CD200 expression on the MSC surface. Our results clearly demonstrate that MSCs, through the expression of CD200, play a major role in the regulation of bone resorption and bone physiology and that the CD200-CD200R couple could be a new target to control bone diseases.

Bone-marrow mesenchymal stem cells (MSCs) are the origin of bone-forming cells with immunomodulation potential. HLA-G5 is among the generated immunosuppressive molecules. HLA-G proteins play a crucial role in promoting the acceptance of allografts. However, the mechanisms regulating the expression of HLA-G5 in human MSCs are unknown. We induced differentiation of MSCs and found that HLA-G5 was greatly up-regulated only in osteoblastic cells (+63% for mRNA). Growth plates and bone callus postfracture in adults showed that only bone-lining cells and mesenchymal progenitors were positive for HLA-G5. Use of gene silencing and dominant-negative factors revealed that HLA-G5 depends on the expression and function of the skeletogenesis master genes RUNX2 and DLX5. In addition, HLA-G5 could directly inhibit osteoclastogenesis by acting on monocytes through SHP1. However, in mature osteoblasts, the expression of HLA-G5 protein was greatly suppressed whereas the proosteoclastogenic factor, RANKL, was concomitantly increased. Down-regulation of HLA-G5 expression during the maturation of osteoblasts was due to binding of the repressor GLI3, a signal transducer of the Hedgehog pathway, to the GLI binding element within the HLA-G promoter. Our findings show that mesenchymal progenitors and osteoblastic cells specifically express HLA-G5 during osteogenesis, with a key role in bone homeostasis.

Objective: To determine the relationship between the number of CD14(+) cells, myocardial infarct (MI) size and left ventricular (LV) volumes in ST segment elevation MI (STEMI) and non-ST segment elevation MI (NSTEMI) patients., Methods: A total of 62 patients with STEMI (n=34) or NSTEMI (n=28) were enrolled. The number of CD14(+) cells was assessed at admission. Infarct size, left ventricular ejection fraction (LVEF) and LV volumes were measured using magnetic resonance imaging five days after MI and six months after MI., Results: In STEMI patients, the number of CD14(+) cells was positively and significantly correlated with infarct size at day 5 (r=0.40; P=0.016) and after six months (r=0.34; P=0.047), negatively correlated with LVEF at day 5 (r=-0.50; P=0.002) and after six months (r=-0.46; P=0.005) and positively correlated with end-diastolic (r=0.38; P=0.02) and end-systolic (r=0.49; P=0.002) volumes after six months. In NSTEMI patients, no significant correlation was found between the number of CD14(+) cells and infarct size, LVEF or LV volumes at day 5 or after six months., Conclusions: The number of CD14(+) cells at admission was associated with infarct size and LV remodelling in STEMI patients with large infarct size, whereas in NSTEMI patients, no relationship was observed between numbers of CD14(+) cells and LV remodelling.

Mesenchymal stem cells (MSCs) and pericyte progenitors (PPs) are both perivascular cells with similar multipotential properties regardless of tissue of origin. We compared the phenotype and function of the 2 cell types derived from the same bone-marrow samples but expanded in their respective media - pericyte conditions (endothelial cell growth medium 2 [EGM-2]) for PPs and standard medium (mesenchymal stem cell medium [MSM]) for MSCs. After 3 weeks of culture, whatever the expansion medium, all cells showed similar characteristics (MSC markers and adipo-osteo-chondroblastic differentiation potential), although neuronal potential was greater in EGM-2- than MSM-cultured cells. As compared with MSM-cultured MSCs, EGM-2-cultured PPs showed higher expression of the pericyte-specific antigen 3G5 than α-smooth muscle actin. In addition, EGM-2-cultured PPs showed an immature phenotype, with upregulation of stemness OCT4 and SOX2 proteins and downregulation of markers of osteoblastic, chondroblastic, adipocytic and vascular smooth muscle lineages. Despite having less effective in vitro immunosuppression capacities than standard MSCs, EGM-2-cultured PPs had higher engraftment potentials when combined with biomaterials heterotopically-transplanted in Nude mice. Furthermore, these engrafted cells generated more collagen matrix and were preferentially perivascular or lined trabeculae as compared with MSM-cultured MSCs. In conclusion, EGM-2-cultured PPs are highly immature cells with increased plasticity and engraftment potential.

Endothelial colony-forming cells (ECFCs) are known to increase after acute myocardial infarction (AMI). We examined whether the presence of ECFCs is associated with preserved microvascular integrity in the myocardium at risk by reducing microvascular obstruction (MVO). We enrolled 88 patients with a first ST elevation AMI. ECFC colonies and circulating progenitor cells were characterized at admission. MVO was evaluated at 5 days and infarct size at 5 days and at 6-month follow-up by magnetic resonance imaging. ECFC colonies were detected in 40 patients (ECFC(pos) patients). At 5 days, MVO was of greater magnitude in ECFC(neg) versus ECFC(pos) patients (7.7 ± 5.3 vs. 3.2 ± 5%, p = 0.0002). At 6 months, in ECFC(pos) patients, there was a greater reduction in infarct size (-32.4 ± 33 vs. -12.8 ± 24%; p = 0.003) and a significant improvement in left ventricular (LV) volumes and ejection fraction. Level of circulating CD34+/VEGF-R2+ cells was correlated with the number of ECFC colonies (r = 0.54, p < 0.001) and relative change in infarct size (r = 0.71, p < 0.0001). The results showed that the presence of ECFC colonies is associated with reduced MVO after AMI, leading to reduced infarct size and less LV remodelling and can be considered a marker of preserved microvascular integrity in AMI patients.

Mesenchymal stem cells (MSCs) are multipotent adult stem cells that are present in practically all tissues as a specialized population of mural cells/pericytes that lie on the abluminal side of blood vessels. Originally identified within the bone marrow (BM) stroma, not only do they provide microenvironmental support for hematopoietic stem cells (HSCs), but can also differentiate into various mesodermal lineages. MSCs can easily be isolated from the BM and subsequently expand in vitro and in addition they exhibit intriguing immunomodulatory properties, thereby emerging as attractive candidates for various therapeutic applications. This review addresses the concept of BM MSCs via a hematologist's point of view. In this context it discusses the stem cell properties that have been attributed to BM MSCs, as compared to those of the prototypic hematopoietic stem cell model and then gives a brief overview of the in vitro and vivo features of the former, emphasizing on their immunoregulatory properties and their hematopoiesis-supporting role. In addition, the qualitative and quantitative characteristics of BM MSCs within the context of a defective microenvironment, such as the one characterizing Myelodysplastic Syndromes are described and the potential involvement of these cells in the pathophysiology of the disease is discussed. Finally, emerging clinical applications of BM MSCs in the field of hematopoietic stem cell transplantation are reviewed and potential hazards from MSC use are outlined.

Objective: Circulating endothelial progenitor cells (EPCs) are a specialized subset of stem/progenitor cells found in bone marrow. They participate in neo-vascularization of injured vessels and predict cardiovascular outcome in patient at risk. Several factors influence their migration and proliferation, among which is the widely studied stromal-derived factor-1α (SDF-1α). In cardiovascular disease, regarding thoracic aortic aneurysms (TAAs), few studies have investigated the levels of EPC and SDF-1α. As rupture, acute dissection and hematoma are acute complications of idiopathic ascending thoracic aortic aneurysm (iATAA) that increase with the size of aneurysm, we aimed to evaluate a potential relationship between circulating EPC and SDF-1α and iATAA size., Methods: The aneurysm size of 27 consecutive patients suffering from iATAA and scheduled for surgery was assessed by computed tomography scan. In all patients, we measured levels of circulating EPCs by flow cytometer, and plasma levels of SDF-1α the day before surgery., Results: The median aneurysm size was 54 mm (interquartile range (IQR): 50.0-58.8]. The EPC levels of CD34+/CD144+/CD14- and CD34+/VEGF-R2+/CD14- were inversely correlated to aneurysm diameter (p = 0.038, r = -0.424 and p = 0.0046, r = -0.65, respectively) before surgery. Conversely, plasma levels of SDF-1α were positively correlated to aneurysm size (p = 0.042; r = 0.47)., Conclusions: Our findings indicate that EPC levels may be useful for monitoring ascending aorta aneurysms and that SDF-1α could be a biomarker of iATAA expansion., (Copyright © 2011 European Association for Cardio-Thoracic Surgery. Published by Elsevier B.V. All rights reserved.)

Mesenchymal stem cells/multipotent marrow stromal cells (MSC) have the ability to participate in there construction of tissues both directly by providing repair cells (essentially those originating from mesoderm)and indirectly by modulating inflammatory and immune responses. This wide range of properties makes these cells very appealing to treat various pathological conditions. They have been first used in 1995 as supportive cells to facilitate hematopoietic stem cells engraftment, and then to minimize the deleterious consequences of graft versus host disease by their immunosuppressive function. Their robust osteogenic differentiation capacity has also been evaluated in numerous preclinical settings of healing/repair but more rarely in human clinical trials. During the past 10-15 years, the potential benefit of their paracrine actions has been tested in various situations such as to facilitate repair after cutaneous defects after burns or lower consequences of ischemic strokes. The purpose of this series of short texts is not to give an exhaustive panorama, but to discuss some well-identified indications in four different fields : auto-immune diseases,bone repair, vascular regeneration and eye lesions such as corneal and retinal defects.

Providing fully mature and functional osteoblasts is challenging for bone tissue engineering and regenerative medicine. Such cells could be obtained from multipotent bone marrow mesenchymal stem cells (MSCs) after induction by different osteogenic factors. However, there are some discrepancies in results, notably due to the use of sera and to the type of osteogenic factor. In this study, we compared the osteogenic differentiation of bone marrow MSCs induced by dexamethasone (Dex) or bone morphogenetic proteins (BMPs) by assessing phenotypes in vitro and functional osteoblasts in vivo. Reducing the content of fetal calf serum from 10% to 2% significantly increased the mineral deposition and expression of osteoblastic markers during osteogenesis. In comparison to Dex condition, the addition of BMP4 greatly improved the differentiation of MSCs into fully mature osteoblasts as seen by high expression of Osterix. These results were confirmed in different supportive matrixes, plastic flasks, or biphasic calcium phosphate biomaterials. In contrast to Dex-derived osteoblasts, BMP4-derived osteoblasts from MSCs were significantly able to produce new bone in subcutis of nude mice in accordance with in vitro results. In conclusion, we describe a convenient ex vivo method to produce consistently mature functional osteoblasts from human MSCs with use of BMP4 and low serum.

HLA-G plays a particular role during pregnancy in which its expression at the feto-maternal barrier participates into the tolerance of the allogenic foetus. HLA-G has also been demonstrated to be expressed in some transplanted patients, suggesting that it regulates the allogenic response. In vitro data indicate that HLA-G modulates NK cells, T cells, and DC maturation through its interactions with various inhibitory receptors. In this paper, we will review the data reporting the HLA-G involvement of HLA-G in human organ transplantation, then factors that can modulate HLA-G, and finally the use of HLA-G as a therapeutic tool in organ transplantation.

Background: Osteosarcoma is the most common malignant primary bone tumour in young adult treated by neo adjuvant chemotherapy, surgical tumor removal and adjuvant multidrug chemotherapy. For correction of soft tissue defect consecutive to surgery and/or tumor treatment, autologous fat graft has been proposed in plastic and reconstructive surgery., Principal Findings: We report here a case of a late local recurrence of osteosarcoma which occurred 13 years after the initial pathology and 18 months after a lipofilling procedure. Because such recurrence was highly unexpected, we investigated the possible relationship of tumor growth with fat injections and with mesenchymal stem/stromal cell like cells which are largely found in fatty tissue. Results obtained in osteosarcoma pre-clinical models show that fat grafts or progenitor cells promoted tumor growth., Significance: These observations and results raise the question of whether autologous fat grafting is a safe reconstructive procedure in a known post neoplasic context.

After bone injuries, several molecular mechanisms establish bone repair from stem/progenitor cells. Inflammation factors attract regenerative cells which expand and differentiate in order to build up a bone highly similar to that before injury. Bone marrow (BM) mesenchymal stem cells (MSCs) as skeletal stem cells and endothelial progenitors (EPCs) are at the origin of such reparation mechanisms. However, discrepancies exist about their identities. Although cultured MSCs are extensively described, their in vivo native forms are poorly known. In addition, recent experiments show that several types of EPC exist. We therefore review up-to-date data on the characterization of such stem/progenitor cells and propose a new point of view of their function in bone regeneration.