scholarly journals The Immunomodulatory Effect of Triptolide on Mesenchymal Stromal Cells

2021 ◽  
Vol 12 ◽  
Author(s):  
Haiping He ◽  
Atsuko Takahashi ◽  
Takeo Mukai ◽  
Akiko Hori ◽  
Miwako Narita ◽  
...  
Keyword(s):  
T Cells ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Chinese Herb ◽  
Activated T Cells ◽  
Hematopoietic Stem ◽  
Proliferative Effect ◽  
Immunosuppressive Cell ◽  
Ifn Γ

Mesenchymal stromal cells (MSCs) are known to have immunosuppressive ability and have been used in clinical treatment of acute graft-versus-host disease, one of severe complications of the hematopoietic stem cell transplantation. However, MSCs are activated to suppress the immune system only after encountering an inflammatory stimulation. Thus, it will be ideal if MSCs are primed to be activated and ready to suppress the immune reaction before being administered. Triptolide (TPL) is a diterpene triepoxide purified from a Chinese herb-Tripterygium wilfordii Hook.f. It has been shown to possess anti-inflammatory and immunosuppressive properties in vitro. In this study, we aimed to use TPL to prime umbilical cord-derived MSCs (TPL-primed UC-MSCs) to enter a stronger immunosuppressive status. UC-MSCs were primed with TPL, which was washed out thoroughly, and the TPL-primed UC-MSCs were resuspended in fresh medium. Although TPL inhibited the proliferation of UC-MSCs, 0.01 μM TPL for 24 h was tolerable. The surface markers of TPL-primed UC-MSCs were identical to those of non-primed UC-MSCs. TPL-primed UC-MSCs exhibited stronger anti-proliferative effect for activated CD4+ and CD8+ T cells in the allogeneic mixed lymphocyte reaction assay than the non-primed UC-MSCs. TPL-primed UC-MSCs promoted the expression of IDO-1 in the presence of IFN-γ, but TPL alone was not sufficient. Furthermore, TPL-primed UC-MSCs showed increased expression of PD-L1. Conclusively, upregulation of IDO-1 in the presence of IFN-γ and induction of PD-L1 enhances the immunosuppressive potency of TPL-primed UC-MSCs on the proliferation of activated T cells. Thus, TPL- primed MSCs may provide a novel immunosuppressive cell therapy.

OP9-DL1 Cell Line Supports the Development of Phenotypically and Functionally Mature Tcrαβ And Tcrγδ T Cells, through Both Conventional and Aberrant Developmental Pathways

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2902-2902
Author(s):  
Tessa C. C. Kerre ◽  
Greet Verstichel ◽  
Stefanie Van Coppernolle ◽  
Imke Velghe ◽  
Frank Timmermans ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Line ◽  
Selection Process ◽  
Receptor Activation ◽  
Notch Receptor ◽  
Activated T Cells ◽  
Hematopoietic Stem ◽  
Ifn Γ

Abstract In vitro generation of mature T cells from human hematopoietic stem and progenitor cells (HSPC) could fulfill two existing needs. First, it could enhance and quicken T cell immune reconstitution after stem cell transplantation, which is very slow and generates a skewed TCR repertoire. Second, by generation of tumour antigen specific T cells it could provide an efficient therapy for numerous malignancies and could enhance GVT effect in the context of allogeneic SCT, without aggravating GVHD. T cells can be generated from human HSPC by culturing them on the murine stromal cell line OP9-transduced with the Notch ligand Delta-like-1 (OP9-DL1). Notch receptor activation is essential for T cell development. However, it is unclear whether Notch activation is sufficient for end maturation into functionally and phenotypically mature TCR positive cells. It was shown that human CD34+ cells cultured on OP9-DL1 differentiate to T cells which can proliferate and produce interferon-g upon polyclonal stimulation. The nature of the mature cells generated in these cultures, however, has not been well studied. CD34+ HSPC from postnatal thymus (PNT) or cord blood were cocultured with OP9-DL1, in the presence of the cytokines Flt-3L (5 ng/ml), SCF (2.5 ng/ml) and IL-7 (5 ng/ml). Every 3–5 days cells were harvested and transferred to fresh OP9-DL1 cells. At repetitive timepoints, an aliquot of the cells was analysed phenotypically. In some experiments, IL-15 was added to the culture. For some experiments, cells harvested from OP9-DL1 at the timepoint mature T cells were observed (usually about d 40 of culture), were transferred to feeder cells, consisting of JY cell line (5.104 cells/ml irradiated with 50 Gy and PBMC (5.105/ml irradiated with 40 Gy), in the presence of PHA (1 mg/ml). After 7 days, IL-2 (50 IU/ml) was added to the culture. Every 14 days, cells were restimulated with new feeders (irradiated JY and PBMC) and new addition of PHA. After 3 weeks of stimulation cells were stimulated overnight with 15 ng/ml PMA and 1500 ng/ml ionomycin, and 18 hours later cells were checked for intracellular presence of cytokines. We investigated whether the T cell population generated in these cultures contains mature cells with the characteristics of TCRγδ cells and of positively selected CD8 or CD4 single positive (SP) TCRαβ cells as observed in the human thymus. We found that under the described conditions, HSPC mature into CD1-CD27+ phenotypically mature T cells, with the TCRγδ fraction maturing faster and more efficiently compared to the TCRαβ fraction. Consistent with a mature phenotype, TCRγδ cells were mostly CD8αα or double negative (DN). No mature CD4 SP TCRαβ cells were observed and the mature CD8 SP cells co-expressed variable ratios of CD8αβ and CD8αα dimers, suggesting that these cells are not conventional positively selected TCRαβ cells. In support of this hypothesis, both mature CD1- TCRαβ and TCRγδ cells expressed the IL2Rβ receptor consitutively and both populations proliferated on IL-15 without prior antigen stimulation, CD8αα (TCRαβ and TCRγδ) cells being the most IL-15 responsive. Mature activated T cells secreted IFN-γ and TNFα, little or no IL-2 and IL-4, with no difference observed between TCRαβ and TCRγδ cells. These data suggest that CD8 TCRαβ cells generated in these cultures are unconventional CD8 cells possibly maturated through agonist selection. However, when cells harvested after 40 days of culture on OP9-DL1 were stimulated with PHA and IL-2 for 3 weeks, conventional appearing CD8αβ cells emerged, with a cytokine production profile similar to that of thymic CD8αβ TCRαβ T cells, with the majority of cells secreting IFN-γ and IL-2. We can conclude from these data that OP9-DL1 supports the development of both unconventional and conventional CD8+ TCRαβ cells, of which the generation and selection process are currently being investigated. Also the in vitro anti-tumor capacities of both populations need to be addressed.

Hematopoietic Multipotent Progenitor Cells Mobilized by G-CSF Can Inhibit Gvhd

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2969-2969
Author(s):  
Maud D'Aveni ◽  
Julien Rossignol ◽  
Ruddy Montandon ◽  
Marie Bouillie ◽  
Flora Zavala ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Progenitor Cells ◽  
Mechanisms Of Action ◽  
Activated T Cells ◽  
Hematopoietic Stem ◽  
Allogeneic Hsct ◽  
Ifn Γ

Abstract Abstract 2969 Backgound. Acute graft-versus-host-disease (aGVHD) is a frequent life threatening complication of allogeneic hematopoietic stem cell transplantation (HSCT). Despite the infusion of higher doses of T cells with the use of G-CSF-mobilized HSC grafts, the incidence of aGVHD is not increased. The mechanisms by which G-CSF-mobilized HSC can control GVHD are imperfectly elucidated. We previously described the mobilization of murin hematopoïetic progenitor cells (HPCs) by G-CSF and FLT3 ligand capable of inducing tolerance against autoimmune diabetes in the nude mice (Kared, Immunity 2006). We now show that G-CSF can mobilize murin HPCs with immunoregulatory functions in the allogeneic immune response and describe their mechanisms of action. Methods. Mobilization of HPCs is performed by subcutaneous administration of human recombinant G-CSF at 200μg/kg per day, for 4 consecutive days in the C57BL6 (H-2b) mouse. HPCs are collected in the spleen by FACS sorting according to their phenotype: Lin- Sca1high cKithigh FLT3low CD34+ CD106+ CD127−. In vitro, functions and mechanisms of action were analyzed by co-cultures with i) T cells (from C57BL6) activated by anti-CD28 and -CD3 mAbs or activated by BALB/c (H-2d) allogeneic splenic LPS matured dendritic cells, ii) C57BL6 splenic selected CD4+CD25high T regulatory T cells activated by anti-CD28 and -CD3 mAbs iii) activated antitumor specific CD8 T cells (C57BL6 ovalbumin specific TCR transgenic T cells). These different cultures were performed in the presence or absence of inhibitors of selective cytokines or other regulatory molecules. In vivo, we assessed the effect of donor HPCs on GVHD development by injecting C57BL6 derived HPCs (0.5×106/mouse), splenic T cells (1×106/mouse) and T depleted bone marrow cells (5×106/mouse) into lethally irradiated (8 Gy) Balb/c recipients. Results. In vitro, as compared to controls without HPCs, after 3 days of culture, HPCs: 1) promote the proliferation of natural T regs activated by anti-CD3 and anti-CD28 (>80% at 3 days of culture compared to control <50%), 2) inhibit the proliferation of activated T cells (>80% T cells blocked before 4 divisions as compared to control-T cells alone >80% after 4 divisions- p<0, 001) and 3) induce the apoptosis of activated T cells (30% increased, p=0, 01). The proliferation of T regs was cell contact dependant and required the presence of TGF-b. The inhibition of T cell activation required IFN γ produced by activated T-cells and some contact-dependent stimuli. In such pro-inflammatory conditions, HPCs differentiate after 4 days in myeloid derived suppressor cells (MDSC). These cells could then produce NO in response to IFN γ and suppress the proliferation of activated T cell. However, T cell suppression was not dependant on L-arginine depletion. Induction of apoptosis of T cells was Fas/Fas-L dependant. Although in the presence of HPCs the proliferation of CD8+ T TCR transgenic against the dominant ovalbumin epitope SIINFEKL was reduced, the cytotoxic response against the SIINFEKL-pulsed EL4 cell line was enhanced (cytotoxicity >90% with HPCs versus <90% w/o HPCs, p<0, 001). In addition, HPCs express CCR7 and CD62L, which should allow their migration to the sites of allopriming. In vivo, none of the mice that had received allogeneic HSCT with HPCs developed clinical or histological GVHD signs as compared to 50% of the control allografted mice without HPCs. Conclusion. Hematopoietic progenitor cells acquire an immunosuppressive potential after G-CSF mobilization. These cells can be isolated from mobilized peripheral stem cells and suppress GVHD while possibly preserving the GVL effect. Work is underway in humans to identify and amplify this population ex vivo for potential therapeutic application in allogeneic HSCT. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Extracellular Vesicles isolated from Mesenchymal Stromal Cells Modulate CD4+ T Lymphocytes Toward a Regulatory Profile

Cells ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 1059 ◽  
Author(s):  
Flavia Franco da Cunha ◽  
Vinicius Andrade-Oliveira ◽  
Danilo Candido de Almeida ◽  
Tamiris Borges da Silva ◽  
Cristiane Naffah de Souza Breda ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Confocal Microscopy ◽  
Extracellular Vesicles ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Target Genes ◽  
T Cell Response ◽  
Activated T Cells

Mesenchymal stromal cells (MSCs) can generate immunological tolerance due to their regulatory activity in many immune cells. Extracellular vesicles (EVs) release is a pivotal mechanism by which MSCs exert their actions. In this study, we evaluate whether mesenchymal stromal cell extracellular vesicles (MSC-EVs) can modulate T cell response. MSCs were expanded and EVs were obtained by differential ultracentrifugation of the supernatant. The incorporation of MSC-EVs by T cells was detected by confocal microscopy. Expression of surface markers was detected by flow cytometry or CytoFLEX and cytokines were detected by RT-PCR, FACS and confocal microscopy and a miRNA PCR array was performed. We demonstrated that MSC-EVs were incorporated by lymphocytes in vitro and decreased T cell proliferation and Th1 differentiation. Interestingly, in Th1 polarization, MSC-EVs increased Foxp3 expression and generated a subpopulation of IFN-γ+/Foxp3+T cells with suppressive capacity. A differential expression profile of miRNAs in MSC-EVs-treated Th1 cells was seen, and also a modulation of one of their target genes, TGFbR2. MSC-EVs altered the metabolism of Th1-differentiated T cells, suggesting the involvement of the TGF-β pathway in this metabolic modulation. The addition of MSC-EVs in vivo, in an OVA immunization model, generated cells Foxp3+. Thus, our findings suggest that MSC-EVs are able to specifically modulate activated T cells at an alternative regulatory profile by miRNAs and metabolism shifting.

scholarly journals Melatonin Synergizes With Mesenchymal Stromal Cells Attenuates Chronic Allograft Vasculopathy

2021 ◽  
Vol 12 ◽  
Author(s):  
Ya-fei Qin ◽  
De-jun Kong ◽  
Hong Qin ◽  
Yang-lin Zhu ◽  
Guang-ming Li ◽  
...  
Keyword(s):  
T Cells ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Th17 Cells ◽  
Memory T Cells ◽  
Transplant Recipients ◽  
Pathological Changes ◽  
Allograft Vasculopathy

BackgroundChronic rejection characterized by chronic allograft vasculopathy (CAV) remains a major obstacle to long-term graft survival. Due to multiple complicated mechanisms involved, a novel therapy for CAV remains exploration. Although mesenchymal stromal cells (MSCs) have been ubiquitously applied to various refractory immune-related diseases, rare research makes a thorough inquiry in CAV. Meanwhile, melatonin (MT), a wide spectrum of immunomodulator, plays a non-negligible role in transplantation immunity. Here, we have investigated the synergistic effects of MT in combination with MSCs in attenuation of CAV.MethodsC57BL/6 (B6) mouse recipients receiving BALB/c mouse donor aorta transplantation have been treated with MT and/or adipose-derived MSCs. Graft pathological changes, intragraft immunocyte infiltration, splenic immune cell populations, circulating donor-specific antibodies levels, cytokine profiles were detected on post-operative day 40. The proliferation capacity of CD4+ and CD8+ T cells, populations of Th1, Th17, and Tregs were also assessed in vitro.ResultsGrafts in untreated recipients developed a typical pathological feature of CAV characterized by intimal thickening 40 days after transplantation. Compared to untreated and monotherapy groups, MT in combination with MSCs effectively ameliorated pathological changes of aorta grafts indicated by markedly decreased levels of intimal hyperplasia and the infiltration of CD4+ cells, CD8+ cells, and macrophages, but elevated infiltration of Foxp3+ cells. MT either alone or in combination with MSCs effectively inhibited the proliferation of T cells, decreased populations of Th1 and Th17 cells, but increased the proportion of Tregs in vitro. MT synergized with MSCs displayed much fewer splenic populations of CD4+ and CD8+ T cells, Th1 cells, Th17 cells, CD4+ central memory T cells (Tcm), as well as effector memory T cells (Tem) in aorta transplant recipients. In addition, the percentage of splenic Tregs was substantially increased in the combination therapy group. Furthermore, MT combined with MSCs markedly reduced serum levels of circulating allospecific IgG and IgM, as well as decreased the levels of pro-inflammatory IFN-γ, TNF-α, IL-1β, IL-6, IL-17A, and MCP-1, but increased the level of IL-10 in the recipients.ConclusionsThese data suggest that MT has synergy with MSCs to markedly attenuate CAV and provide a novel therapeutic strategy to improve the long-term allograft acceptance in transplant recipients.

CD11b− Donor Dendritic Cells Enhance Donor T-Cells’ Th1 Polarization and Graft Versus Leukemia Activity in Allogeneic Hematopoietic Stem Cell Transplantation

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1252-1252 ◽  
Author(s):  
Jian-Ming Li ◽  
Kataryna A. Darlak ◽  
Ying Lu ◽  
Cynthia Giver ◽  
Wayne Harris ◽  
...  
Keyword(s):  
T Cells ◽  
Serum Levels ◽  
Hematopoietic Stem ◽  
Treatment Groups ◽  
Post Transplant ◽  
Tumor Bearing ◽  
Donor T Cells ◽  
Ifn Γ ◽  
Th1 Cytokines

Abstract Background: Based on a clinical association of donor plasmacytoid dendritic cell (DC) content with leukemia relapses after allogeneic BMT (Waller, Blood 2001), we have previously reported that CD11b− donor DC added to a graft containing FACS-purified hematopoietic stem cells (HSC) and T-cells enhanced interferon-γ (IFN-γ) production and GvL activity in MHC-mismatched allogeneic transplant mouse models (Li, Blood 2007). Objective: In this study, we studied the mechanisms whereby donor DC in the graft modulate donor T-cell activity and the graft-versus-leukemia (GvL) effect in MiHA (C3H.SW → C57BL/6J)- and MHC (C57BL/6J → B10.BR)- mismatched models of allogeneic hematopoietic stem cell transplantation (HSCT). Methods: Mice irradiated to 11 Gy received 5 × 104 log-phase viable MMB3.19 myeloid lymphoma cells via intraperitoneal injection or intravenous injection of 1 x 105 LBRM T-cell lymphoma cells one day before transplant. Allografts consisted of 5 × 104 FACS-purified donor BM CD11b− DC or CD11b+ DC plus 3 × 103 FACS-purified c-kit+ sca-1+ lineage− hematopoietic stem cells (HSC) in combination with either 3 × 105 T-cells, 3 × 105 CD8+ T-cells or no additional T-cells transplanted via tail vein. Graft-versus-host disease (GvHD) clinical scores (based on body weight loss, posture, skin, fur texture, activity) were recorded twice weekly in non-tumor bearing recipients. In vitro proliferation and cytotoxic activity of donor-derived T-cells against tumor targets was assessed by CFSE staining and a caspase flow cytometry assay (CyToxiLux PLUS) using donor T-cells harvested from recipients on day 34 and day 82 post transplant. Serum and intracellular Th1 cytokines (IFN-γ, IL-2, and TNF-α) and Th2 cytokines (IL-4, IL-5, and IL-10) from recipients’ peripheral blood and spleens day 3 and day 10 post-transplant was measured by ELISA and flow cytometry. IFN-γ direct killing of leukemia cells was tested by in vitro IFN-γ exposure. Results: In non-tumor bearing mice, recipients of all combinations of donor DC subsets, with and without donor T-cells had equivalent survival (75% – 85%) at 3 months post transplant without significant clinical signs of GvHD. Transplantation of tumor cells to recipients of HSC alone, HSC plus donor T-cells, or HSC plus T-cells and CD11b+ DC in the MiHA- and the MHC-mismatched transplant models led to 0% or 5% 3 month survival, respectively. Strikingly, tumor-bearing mice transplanted with CD11b− DC had significantly enhanced 3 month survival (35% in the MiHA-mismatched model and 45% in the MHCmismatched model) without increased GvHD (p&lt;0.001). There was no significant difference in survival between mice that received HSC plus CD11b− DC and a mixture of CD4+ and CD8+ donor T-cells versus mice that received HSC plus CD11b− DC and only CD8+ donor T-cells. Donor T-cells harvested from recipients of CD11b− DC 34 days after transplant in the MiHA-mismatched model as well as 82 days after transplant in the MHC-mismatched model displayed increased cell proliferation following co-culture with irradiated hosttype splenocytes as a source of alloantigen compared with donor T-cells harvested from recipients of CD11b+ DC or recipients of HSC plus T-cells without donor DC. Leukemia cell killing was greater following incubation of purified donor T-cells recovered from recipients of CD11b− DC with tumor targets compared to T-cells recovered from other treatment groups. Recipients of CD11b− DC had higher serum levels of Th1 cytokines IFN-γ and IL-2 and higher number of Th1 positive donor T-cells compared with recipients of other treatment groups. In contrast, recipients of CD11b+ DC had higher serum levels of Th2 cytokines IL-4, IL-5, and IL-10 and higher number of Th2 positive donor T-cells. IFN-γ added to in vitro cultures with MMB3.19, and LBRM, had no direct cell killing effect. Conclusion: CD11b− donor DC enhanced Th1 polarization of donor T-cells and GvL without increasing GvHD. Donor CD8+ T-cells mediated tumor killing effect. CD11b+ donor DC enhanced Th2 polarization of donor CD4+ T-cells and led to limited GvHD and GvL.

Interferon-Gamma Impairs Expansion and Hematopoietic Support of Bone Marrow Mesenchymal Stromal Cells

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 3884-3884
Author(s):  
Marieke Goedhart ◽  
Anne Cornelissen ◽  
Carlijn Kuijk ◽  
Sulima Geerman ◽  
Fernanda Pascutti ◽  
...  
Keyword(s):  
Stromal Cells ◽  
Stromal Compartment ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Ifn Γ ◽  
Immunomodulatory Properties ◽  
Pre Treatment ◽  
The Impact

Abstract Maintenance of hematopoietic stem cells (HSCs) and regulation of their quiescence and self-renewal is critical for maintaining a lifelong supply of blood cells. The ability of HSCs to stay quiescent is thought to depend on their specific niche in the bone marrow (BM). Mesenchymal stromal cells (MSC) in the BM are multipotent stem cells that form part of the vascular HSC niche and provide micro-environmental support to HSCs both in vivo and upon expansion ex vivo. Culture-expanded MSCs also exhibit immunomodulatory properties that can be enhanced by pre-treatment with interferon-gamma (IFN-γ). BM MSC are thus attractive candidates for cellular therapy after hematopoietic stem cell transplantation, for promoting rapid hematopoietic recovery and reducing the incidence or severity of graft versus host disease. Although IFN-γ pre-treatment can improve the immunomodulatory properties of MSCs, elevated IFN-γ levels have also been associated with anemia and BM failure in multiple chronic inflammatory diseases. While the impact of IFN-γ on HSC has been elucidated in recent years, it remains largely unknown whether IFN-γ can also influence hematopoietic support by BM stromal cells. In this study, we aim to elucidate the impact of IFN-γ on hematopoietic support of BM MSC. We show that in vitro expansion of primary BM MSC cultures from healthy donors was significantly reduced in the presence of IFN-γ, and this effect could be reproduced in the BM stromal cell line MS-5. Concurrently, IFN-γ diminished the clonal capacity of BM MSC, as measured by CFU-F assays. In addition, BM MSC that were pre-stimulated with IFN-γ produced significantly lower levels of CXCL12, suggesting a loss of hematopoietic support potential. Indeed, support of CD34+ hematopoietic stem and progenitor cells (HSPC) in a co-culture assay was greatly reduced in when MSC were pre-treated with IFN-γ. To determine the impact of IFN-γ on BM MSC in vivo, we investigated the BM stromal compartment of IFN-γ AU-rich element deleted (ARE-Del) mice, which constitutively express IFN-γ in steady state conditions. FACS analysis revealed a remodeling of the BM stromal compartment in ARE-Del mice compared to littermate controls, with significantly fewer MSCs, identified as CD45-Ter119-CD31-CD51+PDGFRa+ cells. Numbers of other stromal cell subsets, such as osteoblasts and fibroblasts, were not altered. The reduction of BM MSC in ARE-Del mice coincided with a loss of quiescence in HSCs; only 35% of long term HSC (LT-HSC) in ARE-Del mice were quiescent, compared to 70% in WT mice, as determined by Ki-67 staining. Loss of quiescence in LT-HSC did not lead to increased self-renewal, but rather induced increased differentiation towards short-term HSC and multi-potent progenitors. We then sorted LT-HSC from WT and ARE-Del mice and performed in vitro HSC culture assays in the absence of IFN-γ. Absolute numbers of LT-HSC were rapidly decreased in ARE-Del compared to WT cultures after 3 and 7 days of HSC culture, while numbers of more differentiated progenitors were increased. These data indicate that an IFN-γ-mediated loss of BM MSC in ARE-Del mice leads to loss of quiescent LT-HSCs and induces a tendency towards HSC differentiation over self-renewal. In conclusion, we have shown that IFN-γ has a negative impact on expansion and hematopoietic support of BM MSC in vitro and in vivo across species. Although IFN-γ treatment enhances the immunomodulatory function of MSCs in a clinical setting, it is obvious from our data that IFN-γ impairs their HSC supporting function. These data also provide more insight in the underlying mechanism by which IFN-γ contributes to the pathogenesis of anemia and BM failure. Disclosures No relevant conflicts of interest to declare.

Extracellular Vesicles Isolated from Mesenchymal Stromal Cells Primed with Hypoxia: Novel strategy in Regenerative Medicine

2020 ◽  
Vol 15 ◽  
Author(s):  
Shalmali Pendse ◽  
Vaijayanti Kale ◽  
Anuradha Vaidya
Keyword(s):  
Extracellular Vesicles ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Therapeutic Potential ◽  
Ex Vivo ◽  
Cell Types ◽  
Molecular Profile ◽  
Cell Contact ◽  
Hematopoietic Stem

: Mesenchymal stromal cells (MSCs) regulate other cell types through a strong paracrine component called the secretome, comprising of several bioactive entities. The composition of the MSCs’ secretome is dependent upon the microenvironment in which they thrive, and hence, it could be altered by pre-conditioning the MSCs during in vitro culture. The primary aim of this review is to discuss various strategies that are being used for pre-conditioning of MSCs, also known as “priming of MSCs”, in the context of improving their therapeutic potential. Several studies have underscored the importance of extracellular vesicles (EVs) derived from primed MSCs in improving their efficacy in the treatment of various diseases. We have previously shown that co-culturing hematopoietic stem cells (HSCs) with hypoxiaprimed MSCs improves their engraftment potential. Now the question we pose is would priming of MSCs with hypoxiafavorably alter theirsecretome and would this altered secretome work as effectively as the cell to cell contact did? Here we review the current strategies of using the secretome, specifically the EVs (microvesicles and exosomes), collected from the primed MSCs with the intention of expanding HSCs ex vivo. We speculate that an effective priming of MSCs in vitrocould modulate the molecular profile of their secretome, which could eventually be used as a cell-free biologic in clinical settings.

Mesenchymal Stromal Cells Suppress T-and B-Cells via Galectin-9 in a Donor Dependent Manner

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1248-1248
Author(s):  
Christopher Ungerer ◽  
Patricia Quade-Lyssy ◽  
Reinhard Henschler ◽  
Erhard Seifried ◽  
Heinfried Radeke ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
Immune Suppression ◽  
Immune Cells ◽  
Stromal Cells ◽  
Coagulation Factor ◽  
T And B Cells ◽  
Ifn Γ

Abstract Abstract 1248 Therapeutic approaches using multipotent mesenchymal stromal cells (MSCs) are advancing in regenerative medicine, transplantation and autoimmune diseases. Until now the way of action for MSC-mediated immune suppression is still controversial and relies most probably on a multifactorial mechanism. MSCs have been demonstrated to produce the suppressive molecules hepatocyte growth factor (HGF), tumor growth factor-β (TGF-β), prostaglandin E2 (PGE2) and indoleamine 2,3-dioxygenase (IDO). Furthermore, it has been described that immunosuppression by MSCs is enhanced via stimulation with interferon-γ (IFN-γ). Recently, galectin-1, a β-galactoside binding lectin with immune modulatory properties, has been added to the group of immune modulatory molecules that are responsible for MSC-mediated immune suppression. Here, we identified galectin-9 (Gal-9) as a new molecule involved in MSC-mediated immune modulation. First, we isolated MSCs from bone marrow of randomly selected donors and performed several in vitro experiments regarding their immune modulatory potential (e.g proliferation and IgG production). Interestingly, Gal-9 was the only investigated protein, which was strongly upregulated in MSCs upon activation with IFN-γ. We moreover demonstrate that Gal-9 is a major mediator of the anti-proliferative effect of MSCs on T-cells. Although a B-cell suppressive function of Gal-9 has previously not been reported, we were surprised to detect the same inhibitory effect on isolated B-cells. Proliferation of immune cells was triggered upon either stimulation with either PHA and LPS, or CD40L and PHA. Activation of MSCs with IFN-γ resulted in a major decrease of proliferation of both T-cells and B-cells. In addition, Gal-9 and activated MSCs contribute to the suppression of VZV triggered immunoglobulin release as well. Again activation of MSCs with IFN-γ decreased the IgG release, whereas blocking Gal-9 with lactose, a well characterized inhibitor of Gal-9 function, reversed the effect almost completely. Further, we determined that Gal-9 expression levels (mRNA and protein) distinguish between MSC cultures from different donors after activation. Among donors, we could differentiate between individuals with high Gal-9 levels and higher immune modulatory potential and such with low Gal-9 expression and lower immune modulatory potential. Compared to untreated MSCs we demonstrated a three- to fifty-fold rise in Gal-9 levels after prior activation with IFN-γ. In addition, we demonstrated the upregulation of Gal-9 in MSCs by cell-cell contacts with either T-or B-cells. The upregulation was additionally at least two fold increased by previeously activating MSCs with IFN-γ. Because our group is interested in the therapy of hemophilia A and because of the unxpected suppressive effect of Gal-9 on B-cells and B-cell function, we next tested the effect of MSCs and Gal-9 on the induction of inhibitory antibodies to coagulation factor VIII (FVIII). Mice were immunized with human coagulation factor VIII (FVIII) in the presence or absence of either human MSCs, anti-murine Gal-9 or human Gal-9. As predicted, MSCs suppressed and anti-Gal-9 antibodies anhanced antibody formation. However in contrary to the expected, human Gal-9 co-treatment enhanced the anti-FVIII antibody response. A set of additional experiments revealed, that human Gal-9 suppresses murine regulatory T-cells in vivo. Further, in contrast to human immune cells, murine-derived T- and B-cells did not respond to human recombinant Gal-9 in vitro, but human IFN-γ activated MSCs were able to suppress proliferation of murine immune cells. Because of only 60% homology of murine and human Gal-9 we assume that the murine model cannot predict the function of human Gal-9 and that MSC-mediated immune modulatory functions are exerted via alternative pathways in this setting. Experiments with murine Gal-9 to demonstrate the in vivo function of Gal-9 are ongoing. In conclusion, Gal-9 is novel mediator of MSC immunomodulatory functions and affectsmultiple immune cell types including B-cells. Gal-9 is differentially expressed in MSCs from different donors and may therefore serve as a predictive indicator for clinical MSC functionality. Disclosures: No relevant conflicts of interest to declare.

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