RESUMO
Mesenchymal stromal cells (MSCs) are multipotent adult cells with relevant biological properties making them interesting tools for cell-based therapy. These cells have the ability to home to sites of injury and secrete bioactive factors as part of their therapeutic functions. However, depending on the local environment, diverse functions of MSCs can be modulated and thus can influence their therapeutic value. The specific cytokine milieu within the site of inflammation is vital in determining the fate and cell behaviors of MSCs. Indeed, inflammatory signals (called as inflammatory priming), may induce critical changes on the phenotype, multilineage potential, hematopoietic support and immunomodulatory capacity of MSCs. Thus, for appropriate clinical application of MSCs, it is important to well know and understand these effects. In summary, investigating MSC interactions with the inflammatory environment is necessary to empower the therapeutic value of MSCs.
Assuntos
Inflamação/imunologia , Células-Tronco Mesenquimais/imunologia , Animais , Humanos , Imunomodulação , FenótipoRESUMO
AIM: Uncertainty about the safety of cell therapy continues to be a major challenge to the medical community. Inflammation and the associated immune response represent a major safety concern hampering the development of long-term clinical therapy. In vivo interactions between the cell graft and the host immune system are mediated by functional environmental sensors and stressors that play significant roles in the immunobiology of the graft. Within this context, human liver stellate cells (HSC) demonstrated marked immunological plasticity that has main importance for future liver cell therapy application. METHODS: By using qPCR technique, we established the cytokine gene expression profile of HSCs and investigated the effect of an inflammatory environment on the immunobiology of HSCs. RESULTS AND DISCUSSION: HSCs present a specific immunological profile as demonstrated by the expression and modulation of major immunological cytokines. Under constitutive conditions, the cytokine pattern expressed by HSCs was characterized by the high expression of IL-6. Inflammation critically modulated the expression of major immunological cytokines. As evidenced by the induction of the expression of several inflammatory genes, HSCs acquire a pro-inflammatory profile that ultimately might have critical implications for their immunological shape. CONCLUSION: These new observations have to be taken into account in any future liver cell therapy application based on the use of HSCs.
Assuntos
Células Estreladas do Fígado/imunologia , Hepatite/imunologia , Interleucina-6/imunologia , Células Cultivadas , Células Estreladas do Fígado/patologia , Hepatite/patologia , Humanos , Inflamação/imunologia , Inflamação/patologiaRESUMO
Because of their well-recognized immunomodulatory properties, mesenchymal stromal cells (MSCs) represent an attractive cell population for therapeutic purposes. In particular, there is growing interest in the use of MSCs as cellular immunotherapeutics for tolerance induction in allogeneic transplantations and the treatment of autoimmune diseases. However, multiple mechanisms have been identified to mediate the immunomodulatory effects of MSCs, sometimes with several ambiguities and inconsistencies. Although published studies have mainly reported the role of soluble factors, we believe that a sizeable cellular component plays a critical role in MSC immunomodulation. We refer to these cells as regulatory immune cells, which are generated from both the innate and adaptive responses after co-culture with MSCs. In this review, we discuss the nature and role of these immune regulatory cells as well as the role of different mediators, and, in particular, regulatory immune cell induction by MSCs through interleukin-10. Once induced, immune regulatory cells accumulate and converge their regulatory pathways to create a tolerogenic environment conducive for immunomodulation. Thus, a better understanding of these regulatory immune cells, in terms of how they can be optimally manipulated and induced, would be suitable for improving MSC-based immunomodulatory therapeutic strategies.
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Linfócitos B Reguladores/imunologia , Terapia Baseada em Transplante de Células e Tecidos/métodos , Células Dendríticas/imunologia , Imunomodulação/imunologia , Células-Tronco Mesenquimais/imunologia , Linfócitos T Reguladores/imunologia , Doenças Autoimunes , Técnicas de Cocultura , Humanos , Fatores Imunológicos , Interleucina-10/imunologia , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/citologiaRESUMO
BACKGROUND AIMS: Because of their self-renewal capacity, multilineage potential and immunomodulatory properties, MSCs are an attractive tool for cell-based immunotherapy strategies. Foreskin, considered as a biological waste material, has been shown to be a reservoir of therapeutic cells. METHODS: MSCs were isolated from different foreskin samples, maintained under in vitro culture and defined according to the International Society for Cellular Therapy (ISCT) criteria. We subsequently determined their main cell characteristics as well as their immunobiological properties. The following parameters were determined: (i) morphology and phenotype, (ii) proliferative and clonogenic potentials, (iii) tri-lineage differentiation ability, (iv) immunological profile, (v) immunomodulatory properties and (vi) protein and messenger RNA expression/secretion profile of immunoregulatory cytokines/factors as well as the pattern of toll-like receptors (TLRs). By using a pro-inflammatory cytokine cocktail, we also evaluated the influence of an inflammatory environment on their biology. RESULTS: With a typical fibroblast-like morphology and an ISCT-compliant phenotype, foreskin-MSCs (FSK-MSCs) were highly proliferative and had a great clonogenic potential. They displayed multilineage capacities and interesting immunomodulatory properties. Of importance, FSK-MSCs were not immunogenetic and were further able to inhibit T-cell proliferation. We showed that several immunoregulatory cytokines and factors might be potentially involved in FSK-MSC immunomodulation with particular attention to hepatocyte growth factor and interleukin-11. Moreover, FSK-MSCs expressed several TLRs and were sensitive to the inflammatory environment by properly adjusting their profile and fate. CONCLUSIONS: Foreskin represents a new alternative source for MSCs that is compliant with ISCT criteria. Their unique immunobiological properties allow consideration of FSK-MSCs as a valuable tolerogenic product for cell-based immunotherapy.
Assuntos
Separação Celular/métodos , Terapia Baseada em Transplante de Células e Tecidos , Prepúcio do Pênis/citologia , Imunomodulação/fisiologia , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/fisiologia , Técnicas de Cultura de Células/métodos , Diferenciação Celular , Proliferação de Células/fisiologia , Terapia Baseada em Transplante de Células e Tecidos/métodos , Células Cultivadas , Fator de Crescimento de Hepatócito/metabolismo , Humanos , Recém-Nascido , Interleucina-11/metabolismo , MasculinoRESUMO
Human skin-derived precursors (hSKPs) are multipotent somatic stem cells that persist within the dermis throughout adulthood and harbor potential clinical applicability. In this study, we investigated their immunogenicity and immunosuppressive features, both in vitro and in vivo. As such, this study provides a solid basis for developing their future clinical applications. We found that hSKPs express HLA-ABC molecules, but not HLA-DR, rendering them poorly immunogenic. Using a coculture set-up, we could further demonstrate that hSKPs inhibit the proliferation of allogeneic activated T cells and alter their cytokine secretion profile, in a dose-dependent manner. Cotransplantation of hSKP and human peripheral blood leukocytes (PBL) into severe combined immune-deficient mice also showed a significant impairment of the graft-versus-host response 1 week post-transplantation and a drastic increase in survival time of 60%. From a mechanistic point of view, we found that hSKPs require cell contact as well as secretion of soluble inhibitory factors in order to modulate the immune response. The expression/secretion levels of these factors further increases upon inflammation or in the presence of activated T cells. As such, we believe that these features could be beneficial in a later allogeneic clinical setting, because rejection of engrafted allogeneic hSKP might be delayed or even avoided due to their own promotion of a tolerogenic microenvironment.
Assuntos
Aloenxertos/imunologia , Células-Tronco Multipotentes/imunologia , Pele/citologia , Pele/imunologia , Animais , Técnicas de Cocultura , Dinoprostona/biossíntese , Citometria de Fluxo , Antígenos HLA/biossíntese , Fator de Crescimento de Hepatócito/biossíntese , Humanos , Imuno-Histoquímica , Fator Inibidor de Leucemia/biossíntese , Ativação Linfocitária/imunologia , Teste de Cultura Mista de Linfócitos , Camundongos , Camundongos SCID , Células-Tronco Multipotentes/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
BACKGROUND AIMS: Stem cell therapy for liver diseases has recently emerged as a promising alternative to liver transplantation. Eligible cells should have an appropriate immunophenotype. The aim of the present study was to define the immunological profile of two human liver-derived mesenchymal stromal cell populations, namely, stem cells (ADHLSC) and hepatic stellate cells (HSC). METHODS: The study was conducted under normal and inflammatory conditions with the use of human bone marrow mesenchymal stromal cells (BM-MSC) as reference. RESULTS: Like BM-MSC and ADHLSC, HSC were negative for hematopoietic (CD45) and endothelial (CD34) markers but positive for stromal markers. All cell types were constitutively positive for HLA class I and negative for human leukocyte antigen (HLA) class II and co-stimulatory molecules (CD80, CD86, CD134 and CD252). Inflammation induced the expression of CD40 in all cell types, but the highest values were observed on HSCs; high CD252 expression was only observed on HSC as compared with ADHLSC and BM-MSC. The expression of various adhesion molecules (CD54, CD58, CD106 and CD166) was dissimilar in these three cell types and was differentially influenced by inflammation as well. ADHLSC and HSC constitutively expressed the immunosuppressive molecule HLA-G, whereas CD274 expression was induced by inflammation, as in the case of BM-MSC. Moreover, all cell types expressed the two major natural killer ligands CD112 and CD115. CONCLUSIONS: Toll-like receptors (TLR) 1, 3, 4 and 6 messenger RNA was expressed by both cell types, whereas TLR 2, 5, 7, 9 and 10 were only expressed by ADHLSC. Inflammation increased the expression of TLR 2 and 3 by ADHLSC and HSC. Finally, both liver-derived cell types were immunosuppressive because they inhibited the proliferation of mitogen-activated T cells.
Assuntos
Células Estreladas do Fígado/imunologia , Imunomodulação/imunologia , Inflamação/imunologia , Ativação Linfocitária/imunologia , Células-Tronco Mesenquimais/imunologia , Antígenos CD34/metabolismo , Antígeno B7-H1/metabolismo , Biomarcadores/metabolismo , Células da Medula Óssea/citologia , Terapia Baseada em Transplante de Células e Tecidos , Antígenos HLA-G/metabolismo , Células-Tronco Hematopoéticas/imunologia , Células Estreladas do Fígado/citologia , Humanos , Imunofenotipagem , Subunidade beta de Receptor de Interleucina-2/metabolismo , Antígenos Comuns de Leucócito/metabolismo , Fígado/citologia , Hepatopatias/terapia , Células-Tronco Mesenquimais/citologia , Receptor de Fator Estimulador de Colônias de Macrófagos/metabolismo , Receptores Toll-Like/biossínteseRESUMO
Mesenchymal stromal cells (MSC) can be expanded from different sources. We compared the influence of inflammation and TLR ligation on the phenotype and function of MSC derived from bone marrow (BM), adipose tissue (AT), and Wharton's jelly (WJ). WJ-MSC were featured by a lack of TLR4 expression. While inflammation upregulated TLR3 in all three MSC types, TLR4 upregulation was observed only on BM-MSC. TLR ligation increased the production of inflammatory cytokines in BM- and AT-MSC but not in WJ-MSC and augmented anti-inflammatory cytokines in AT-MSC. Although inflammation increased in all MSC types the secretion of inflammatory cytokines, additional TLR triggering did not have further effect on WJ-MSC. The immunosuppressive potential of WJ-MSC on MLR was affected neither by inflammation nor by TLR triggering. This resistance was related to an overproduction of HGF. These data indicate that MSC source could be of importance while designing immunomodulating cell therapy in transplantation.
Assuntos
Células-Tronco Mesenquimais/imunologia , Células Estromais/imunologia , Receptores Toll-Like/metabolismo , Tecido Adiposo/citologia , Tecido Adiposo/imunologia , Sequência de Bases , Células da Medula Óssea/citologia , Células da Medula Óssea/imunologia , Separação Celular , Citocinas/genética , Citocinas/metabolismo , Dinoprostona/biossíntese , Feminino , Fator de Crescimento de Hepatócito/biossíntese , Humanos , Técnicas In Vitro , Recém-Nascido , Inflamação/genética , Inflamação/metabolismo , Teste de Cultura Mista de Linfócitos , Células-Tronco Mesenquimais/citologia , Gravidez , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transdução de Sinais , Células Estromais/citologia , Receptor 3 Toll-Like/genética , Receptor 3 Toll-Like/metabolismo , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/metabolismo , Receptores Toll-Like/genética , Cordão Umbilical/citologia , Cordão Umbilical/imunologiaRESUMO
Due to their immunomodulatory properties, adipose tissue (AT) and Wharton's Jelly (WJ) constitute valuable alternatives to BM as sources of MSCs for managing graft-versus-host disease. To ensure the efficiency of AT- and WJ-MSCs implies the characterization of their immunomodulatory functions in comparison to those of BM. In this study, we investigated the capacity of AT- and WJ-MSCs to modulate lymphocyte reactions in response to different stimuli as well as the specificity of this immunomodulation. AT- and WJ-MSC displayed potent immunosuppressive effects on lymphocyte responses in a dose-dependent manner. These effects included the prevention of lymphocyte activation as well as the suppression of T-cell proliferation regardless of the stimuli used to activate lymphocytes. These effects were mediated through the expression of COX1/COX2 enzymes and by the production of PGE2. CD4(+) and CD8(+) T-lymphocytes were equally targeted by MSCs demonstrating that the immunomodulation was not restricted to a specific T-cell subpopulation.
Assuntos
Tecido Adiposo/patologia , Medula Óssea/patologia , Dinoprostona/biossíntese , Células-Tronco Mesenquimais/metabolismo , Cordão Umbilical/patologia , Adolescente , Adulto , Antígenos CD/biossíntese , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Técnicas de Cocultura , Dinoprostona/antagonistas & inibidores , Dinoprostona/genética , Dinoprostona/imunologia , Feminino , Humanos , Imunomodulação , Indometacina/farmacologia , Recém-Nascido , Ativação Linfocitária/efeitos dos fármacos , Subpopulações de Linfócitos/imunologia , Subpopulações de Linfócitos/metabolismo , Subpopulações de Linfócitos/patologia , Células-Tronco Mesenquimais/imunologia , Células-Tronco Mesenquimais/patologia , Gravidez , Linfócitos T/imunologia , Linfócitos T/metabolismo , Linfócitos T/patologiaRESUMO
BACKGROUND AIMS: Mesenchymal stromal cells (MSC) have been shown to possess immunomodulatory functions and proposed as a tool for managing or preventing graft-versus-host disease (GvHD) as well as promoting clinical transplantation tolerance. We investigated the capacity of human bone marrow (BM) MSC to modulate the proliferation of T cells obtained from peripheral blood (PB) and umbilical cord blood (CB). We addressed the importance of the MSC:T-cell ratio, requirement for cell contact and impact of soluble factors on the MSC-mediated effects. We also analyzed whether regulatory T cells could be modulated by MSC in co-cultures. METHODS: The effect of different MSC concentrations on T-cell proliferation induced by allogeneic, mitogenic or CD3/CD28 stimulation was analyzed using bromodeoxyuridine (BrdU) incorporation and carboxyfluorescein diacetate-succinimidyl ester (CFDA-SE) labeling. The level of regulatory T cells was assessed using quantitative real-time polymerase chain reaction (PCR) and flow cytometry analysis. RESULTS: MSC induced a dose- and contact-dependent inhibition of T-cell proliferation but lymphocytes from CB and PB were differentially affected. At low concentrations, MSC supported both CB and PB T-cell proliferation, rather than inhibiting their proliferation. This supportive effect was contact independent and soluble factors such interleukin-6 (IL-6) appeared to be involved. Interestingly, among the expanded T-cell population in both CB and PB, regulatory T cells were increased and were a part of the new cells promoted by MSC at low doses. CONCLUSIONS: MSC represent an attractive tool for reducing the lymphocyte response by inhibiting T-cell activation and proliferation as well as promoting tolerance by maintaining and promoting the expansion of regulatory cells. Nevertheless, the dual ability of MSC to either sustain or suppress T-cell proliferation according to conditions should be considered in the context of clinical applications.
Assuntos
Sangue Fetal/citologia , Interleucina-6/metabolismo , Células-Tronco Mesenquimais/citologia , Células Estromais/citologia , Linfócitos T/citologia , ADP-Ribosil Ciclase 1/metabolismo , Adulto , Antígenos CD4/metabolismo , Contagem de Células , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Técnicas de Cocultura , Ensaio de Unidades Formadoras de Colônias , Meios de Cultivo Condicionados/farmacologia , Humanos , Tolerância Imunológica , Subunidade alfa de Receptor de Interleucina-2/metabolismo , Subunidade alfa de Receptor de Interleucina-7/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Mitógenos/farmacologia , Células Estromais/efeitos dos fármacos , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Adulto JovemRESUMO
BACKGROUND: Being non-immunogenic and capable of achieving major metabolic liver functions, adult-derived human liver stem/progenitor cells (ADHLSCs) are of special interest in the field of liver cell therapy. The cytokine repertoire of engrafted cells may have critical impacts on the immune response balance, particularly during cell transplantation. METHODS: In this work, we analyzed the cytokinome of ADHLSCs during hepatogenic differentiation (HD) following stimulation with a mixture of inflammatory cytokines (I) in vitro and compared it to that of mature hepatocytes. RESULTS: Independent of their hepatic state, ADHLSCs showed no constitutive expression of pro-inflammatory cytokines, which were significantly induced by inflammation (IL-1ß, IL-6, IL-8, TNFα, CCL5, IL-12a, IL-12b, IL-23p19, IL-27p28 and EBI-3). IL1-RA and IDO-1, as immunoregulatory cytokines, were highly induced in undifferentiated ADHLSCs, whereas TGF-ß was downregulated by both hepatic and inflammatory events. Interestingly, TDO-1 was exclusively expressed in ADHLSCs after hepatic differentiation and enhanced by inflammatory cytokines. Compared to mature hepatocytes, hepatic-differentiated ADHLSCs showed significantly different cytokine expression patterns. CONCLUSIONS: By establishing the cytokinome of ADHLSCs and highlighting their immunological and inflammatory features, we can enhance our knowledge about the safety and efficiency of the transplantation strategy.
RESUMO
OBJECTIVES: We aimed at characterizing the transcription profiles of immunological receptors associated with the biology of mesenchymal stromal cells (MSCs). MATERIALS AND METHODS: In this experimental study, quantitative real time-polymerase chain reaction (qRTPCR) was performed to establish the transcription profiles of advanced glycation end-products (RAGE) receptor, C-type lectin receptors (CLRs, including DECTIN-1, DECTIN-2 and MINCLE), leukotriene B4 (LTB4) receptors (BLT1 and BLT2) and cysteinyl leukotrienes (CysLTs) receptors (CYSLTR1 and CYSLTR2) in distinct populations of MSCs grown under basic or inflammatory conditions. RESULTS: MSCs derived from adipose tissue (AT), foreskin (FSK), Wharton's jelly (WJ) and bone marrow (BM) exhibited significantly different transcription levels for these genes. Interestingly, these transcription profiles substantially changed following exposure of MSCs to inflammatory signals. CONCLUSIONS: Collectively, for the first time, our data highlights that MSCs depending on their tissue-source, present several relevant receptors potentially involved in the regulation of inflammatory and immunological responses. Understanding the roles of these receptors within MSCs immunobiology will incontestably improve the efficiency of utilization of MSCs during cell-based therapies.
RESUMO
Human hepatic stellate cells (HSCs) demonstrated great immunological plasticity with important consequences for liver cell therapy. Activated HSCs (aHSCs) are in vitro reverted (rHSCs) to a quiescent-like phenotype with potential benefit to reduce liver fibrosis. The goal of this study is to establish and compare the immunological profile of activated and in vitro reverted HSCs and to investigate the impact of inflammatory priming on the immunobiology of both HSCs populations. The distribution of inflammatory primed activated and reverted HSCs across the different phases of the cell cycle is assessed by flow cytometry. In addition, Flow analysis was done to assess the expression level of neuronal, endothelial and stromal markers, cell adhesion molecules, human leucocyte antigens, co-stimulatory molecules, immunoregulatory molecules and natural killer ligands. Our results showed that the cell cycle distribution of both HSCs populations is significantly modulated by inflammation. Accordingly, activated HSC that were in G1 phase switch to S- and G2 phases when exposed to inflammation, while reverted HSCs mostly redistribute into sub-G0 phase. In a HSC state dependent manner, inflammatory priming modulated the expression of the stromal marker CD90, biological receptors (CD95 and CD200R), cell adhesion molecules (CD29, CD54, CD58, CD106 and CD166), human leucocyte antigen HLA-G, co-stimulatory molecules (CD40 and CD252), as well as the immunoregulatory molecules (CD200 and CD274). In conclusion, the immunologic profile of HSCs is significantly modulated by their activation state and inflammation and is important for the development of novel HSC liver cell-based therapy.
Assuntos
Células Estreladas do Fígado/imunologia , Fígado/imunologia , Biomarcadores/metabolismo , Moléculas de Adesão Celular/metabolismo , Ciclo Celular/imunologia , Linhagem Celular , Antígenos HLA-G/imunologia , Células Estreladas do Fígado/metabolismo , Humanos , Fatores Imunológicos/metabolismo , Inflamação/imunologia , Inflamação/metabolismo , Fígado/metabolismo , Cirrose Hepática/imunologia , Cirrose Hepática/metabolismoRESUMO
BACKGROUND Mesenchymal stromal cells (MSCs) possess potent immunomodulatory properties that increase their value as a cell-based therapeutic tool for managing various immune-based disorders. Over the past years, accumulated results from trials using MSCs-based therapy have shown substantial contradictions. Although the reasons underlying these discrepancies are still not completely understood, it is well known that the immunomodulatory activities mediated by distinct MSCs differ in a manner dependent on their tissue origin and adequate response to inflammation priming. Thus, characterization of new molecular pathway(s) through which distinct MSC populations can exert their immunomodulatory effects, particularly during inflammation, will undoubtedly enhance their therapeutic potential. MATERIAL AND METHODS After confirming their compliance with ISCT criteria, quantitative real time-PCR (qRT-PCR) was used to screen new immunoregulatory genes in MSCs, derived from adipose tissue, foreskin, Wharton's jelly or the bone-marrow, after being cultivated under normal and inflammatory conditions. RESULTS FGL2, GAL, SEMA4D, SEMA7A, and IDO1 genes appeared to be differentially transcribed in the different MSC populations. Moreover, these genes were not similarly modulated following MSCs-exposure to inflammatory signals. CONCLUSIONS Our observations suggest that these identified immunoregulatory genes may be considered as potential candidates to be targeted in order to enhance the immunomodulatory properties of MSCs towards more efficient clinical use.
Assuntos
Inflamação/genética , Células-Tronco Mesenquimais/fisiologia , Tecido Adiposo/citologia , Tecido Adiposo/imunologia , Tecido Adiposo/fisiologia , Antígenos CD/genética , Antígenos CD/metabolismo , Células da Medula Óssea/citologia , Células da Medula Óssea/imunologia , Células da Medula Óssea/fisiologia , Proliferação de Células/fisiologia , Células Cultivadas , Fibrinogênio/genética , Fibrinogênio/metabolismo , Prepúcio do Pênis/citologia , Prepúcio do Pênis/imunologia , Prepúcio do Pênis/fisiologia , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/metabolismo , Humanos , Indolamina-Pirrol 2,3,-Dioxigenase/genética , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Inflamação/imunologia , Inflamação/patologia , Masculino , Células-Tronco Mesenquimais/imunologia , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/patologia , Semaforinas/genética , Semaforinas/metabolismo , Geleia de Wharton/citologia , Geleia de Wharton/imunologia , Geleia de Wharton/fisiologiaRESUMO
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-ß) 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.
Assuntos
Proteína DEAD-box 58/genética , Regulação da Expressão Gênica , Células-Tronco Mesenquimais/metabolismo , Sobrevivência Celular/genética , Citocinas/metabolismo , Proteína DEAD-box 58/metabolismo , Humanos , Quinase I-kappa B/metabolismo , Imunofenotipagem , Fator Regulador 7 de Interferon/metabolismo , Helicase IFIH1 Induzida por Interferon/genética , Helicase IFIH1 Induzida por Interferon/metabolismo , Fator Inibidor de Leucemia/genética , Fator Inibidor de Leucemia/metabolismo , Especificidade de Órgãos , Poli I-C/farmacologia , RNA Mensageiro/genética , Receptores Imunológicos , Receptores do Ácido Retinoico/genética , Receptores do Ácido Retinoico/metabolismo , Ligante Indutor de Apoptose Relacionado a TNF/genética , Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Receptor 3 Toll-Like/metabolismoRESUMO
Mesenchymal stromal cells (MSCs) have recently been the subject of great interest in the fields of regenerative medicine and immunotherapy due to their unique biological properties. In particular, MSCs possess immunoregulatory properties that can modulate immune as well as inflammatory responses. Although there are many studies about MSC immunomodulation, several complex and conflicting mechanisms have been reported. Herein, we aim to review these mechanisms and identify a link between these pathways. We focus on human studies in which bone marrow-derived MSCs and T cells were investigated. We propose that MSC-induced immunomodulation exists as a network where converging regulatory pathways compete to establish a tolerogenic state. As interleukin-10 seems to play a central role in this network, we also discuss the relationship between this cytokine and other regulatory factors in the context of immunomodulation.
Assuntos
Imunoterapia , Células-Tronco Mesenquimais/imunologia , Medicina Regenerativa , Células Estromais/imunologia , Linfócitos T/imunologia , Medula Óssea/imunologia , Humanos , Tolerância Imunológica , Imunomodulação , Interleucina-10/metabolismoRESUMO
BACKGROUND: Bone marrow mesenchymal stromal cells (BM-MSCs) display immunomodulatory features, representing a promising tool for cell-based therapies. However, the mechanisms used by MSCs to regulate T cell fate remain unclear. AIMS: We investigated the potential of BM-MSCs to modulate T cell activation, proliferation, cytokine secretion and immunophenotype. MATERIALS AND METHODS: T cells were co-cultured with BM-MSCs to assess their immunomodulatory impact. T cell characterization was performed using cell tracing, ELISA, intracellular and surface staining, flow cytometry analysis and qPCR. RESULTS: The activation and proliferation of T cells were downregulated during coculture with BM-MSCs. We also observed that BM-MSCs upregulated IL-10 secretion as well as the expression of its receptor CD210 on T cells, thus creating a loop favoring the expansion of IL-10-producing T cells. IL-10 neutralization restored T cell proliferation, demonstrating that IL-10 is functionally relevant during immunomodulation. Moreover, BM-MSCs differently modulated CD4 and CD8 T-cell immunophenotype by inducing broad changes in their molecular pattern. CONCLUSIONS: We provide a comprehensive functional and molecular characterization of T cells that are immunomodulated by BM-MSCs. Indeed, a better understanding of the immunological interplay between T cells and MSCs will facilitate the development of new efficient approaches to improve cell-based immune therapies.
Assuntos
Diferenciação Celular/genética , Subunidade alfa de Receptor de Interleucina-10/biossíntese , Interleucina-10/biossíntese , Células-Tronco Mesenquimais/citologia , Tecido Adiposo/citologia , Tecido Adiposo/imunologia , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Proliferação de Células/genética , Técnicas de Cocultura , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Interleucina-10/metabolismo , Ativação Linfocitária/imunologia , Células-Tronco Mesenquimais/imunologia , Células-Tronco Mesenquimais/metabolismo , Linfócitos T/citologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismoRESUMO
Preparations of mesenchymal stromal cells (MSCs) are generally obtained from unfractionated tissue cells, resulting in heterogeneous cell mixtures. Several markers were proposed to enrich these cells, but the majority of these markers are defined for bone marrow (BM). Moreover, the surface markers of freshly isolated MSCs also differ from those of cultured MSCs in addition to a phenotypic variation depending on the MSC source. For tissue engineering applications, it is crucial to start with a well-defined cell population. In this study, we performed immunomagnetic selections with five single surface markers to isolate MSC subpopulations from BM and adipose tissue (AT): CD271, SUSD2, MSCA-1, CD44, and CD34. We determined the phenotype, the clonogenicity, the proliferation, the differentiation capacity, and the immunoregulatory profile of the subpopulations obtained in comparison with unselected cells. We showed that native BM-MSCs can be enriched from the positive fractions of MSCA-1, SUSD2, and CD271 selections. In contrast, we observed that SUSD2 and MSCA-1 were unable to identify MSCs from AT, meaning they are not expressed in situ. Only the CD34(+) selection successfully isolated MSCs from AT. Interestingly, we observed that CD271 selection can define AT cell subsets with particular abilities, but only in lipoaspiration samples and not in abdominoplasty samples. Importantly, we found a population of clear CD34(+) fresh BM-MSCs displaying different properties. A single marker-based selection for MSC enrichment should be more advantageous for cell therapy and would enable the standardization of efficient and safe therapeutic intervention through the use of a well-identified and homogeneous cell population.
Assuntos
Adipócitos/citologia , Tecido Adiposo/citologia , Células da Medula Óssea/citologia , Citometria de Fluxo/métodos , Células-Tronco Mesenquimais/citologia , Adolescente , Antígenos de Superfície/metabolismo , Biomarcadores/metabolismo , Diferenciação Celular/fisiologia , Proliferação de Células , Feminino , Humanos , Masculino , Glicoproteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Fenótipo , Receptores de Fator de Crescimento Neural/metabolismoRESUMO
Stem cell therapy is a potential method for the treatment of numerous diseases. The most frequent cellular source is bone-marrow-derived mesenchymal stromal cells (BM-MSCs). Human adipose-derived stromal cells (ADSCs) share similar properties with BM-MSCs as they support hematopoiesis, modulate ongoing immune responses, and differentiate into cells of mesodermal origin. On the other hand, ADSCs have higher frequency in situ, higher availability, and very few ethical issues compared with BM-MSCs, giving them an advantage over BM-MSCs for clinical use. Most of the methods used to isolate ADSCs contain a collagenase digestion step, but the type of collagenase and time of sample digestion vary among studies and these differences could have an impact on the cell properties and thus in result comparison. To overcome this obstacle, we propose a new method to isolate ADSCs from lipoaspirate without collagenase digestion step. We compared ADSCs obtained with our method versus classical protocol using collagenase digestion. Cells obtained with our method are equivalent but they have a better long-term hematopoietic support than those obtained with classical method. Moreover, our method has an advantage over the classical one as it is easier, safer, faster, less expensive, and more consistent with good manufacturing practices to obtain large number of ADSCs ex vivo.
Assuntos
Adipócitos/citologia , Células da Medula Óssea/citologia , Diferenciação Celular/fisiologia , Proliferação de Células , Hematopoese/fisiologia , Células-Tronco Mesenquimais/citologia , Tecido Adiposo/citologia , Proliferação de Células/fisiologia , Separação Celular , Células Cultivadas , Feminino , HumanosRESUMO
Mesenchymal stromal cells (MSCs) isolated from different tissue sources may present distinct immunomodulatory profiles. As lymphocyte responses are a combination of several distinct steps, we evaluated and compared the impact of MSCs from different sources on the activation, proliferation and migration of T-cells. We demonstrated that tissue-derived MSCs have important immunomodulatory effects. AT-MSCs induced potent anti-proliferative and anti-inflammatory (IFN-γ downregulation) effects and differentially modulated several T-cell activation markers (CD23, CD26, CD45, and CD69). Among all the MSC types tested, only AT-MSCs induced significant downregulation of CD26 and CD45 expression. Of importance, AT-MSCs maintained a sustained expression of CD69. AT-MSCs, particularly following exposure to an inflammatory environment, promoted the migration of lymphocytes into their surrounding environment. The AT-MSCs may increase recruitment of T lymphocytes by upregulation of IL-8 and CCL5 secretion. Following their migration, T-cells interact with MSCs, which can impair lymphocyte proliferation and activation depending on their origin. Inflammatory T-cells appeared to be progressively suppressed, which may lead to a population of lymphocytes with a regulatory phenotype. These findings are relevant, as they increase our understanding of the different immunomodulatory effect of MSCs as well as their behavior in an inflammatory environment.
Assuntos
Movimento Celular/imunologia , Proliferação de Células , Imunomodulação , Ativação Linfocitária/imunologia , Células-Tronco Mesenquimais/imunologia , Linfócitos T/imunologia , Células Cultivadas , Técnicas de Cocultura , Citocinas/imunologia , Citocinas/metabolismo , Citometria de Fluxo , Humanos , Especificidade de ÓrgãosRESUMO
Mesenchymal stromal cells (MSCs) possess a specific immunological profile that makes them potentially useful for immune-based therapies. Adipose tissue (AT) and Wharton's jelly (WJ) are considered to be valuable alternatives to bone marrow (BM) as sources of MSCs. These MSCs exhibit strong immunomodulatory properties that affect lymphocyte responses. The CD200/CD200R axis has been reported to be important in regulating the immune responses. Engagement of CD200R by CD200 initiates an inhibitory pathway that displays immunosuppressive effects. Because the CD200/CD200R axis is involved in immunoregulation, we investigated the expression and role of this ligand/receptor pair in MSCs and T-lymphocytes during co-culture. CD200 is differently expressed and modulated on MSCs depending on the tissue of origin and the culture conditions. Among the different MSC sources, WJ-MSCs express CD200 in the greatest proportion. This high constitutive CD200 expression may represent a distinctive marker for WJ-MSCs. A pro-inflammatory environment and IFN-γ in particular induce an increase in CD200 expression by BM-MSCs. In T-lymphocytes, CD200R and CD200 are differently distributed between the CD4(+) and CD8(+) T-cell subsets. During co-culture, blocking CD200-CD200R interactions does not prevent MSC-mediated inhibition of lymphocyte proliferation. However, depending on their origin, MSCs are able to modulate the expression of both CD200 and CD200R on some T-cells. Further study is required to understand the function of CD200 expression by nonmyeloid cells such MSCs and the significance of CD200 and C200R expression by T-cells. The findings presented here support bidirectional communication between MSCs and T-lymphocytes. Understanding the role of this ligand-receptor pair during co-culture will improve and increase the clinical use of MSCs.