The presence of GM-CSF and IL-4 interferes with effect of TGF-beta1 on antigen presenting cells in patients with multiple sclerosis and in rats with experimental autoimmune encephalomyelitis

The possibility to generate and expand tolerogenic dendritic cells (DC) with TGF-β1 in vitro opens new therapeutic perspectives for the treatment of autoimmune diseases. In the present study, GM-CSF+IL-4 induced the differentiation of DC from adherent peripheral blood mononuclear cells, which had a higher expression of HLA-DR, CD86 and CD1a and the capacity to stimulate T cells. TGF-β1 alone slightly promoted the generation of antigen presenting cells (APC) with higher expression of CD14, but did not differentiate them into E-cadherin + Langerhans cell (LC)-like DC. TGF-β1-driven APC exhibited the morphology, phenotypes and functions of tolerogenic immature DC, and had lower capacity to stimulate T cells. In vivo experiment demonstrates that TGF-β1-treated APC exhibited the therapeutic potential in Lewis rats with experimental autoimmune encephalomyelitis (EAE), followed by increase of IL-10 production in lymph nodes and decrease of inflammatory cells in spinal cords. Most importantly, GM-CSF/IL-4 used in DC preparation abolished the effect of TGF-β1 to induce tolerogenic APC in vitro and in vivo. The results reveal that the usage of GM-CSF for the generation of tolerogenic DC should not be copied from DC preparation for anti-tumor therapy.     

Induction of TGF-β and IL-10 production in dendritic cells using astilbin to inhibit dextran sulfate sodium-induced colitis

Astilbin, a major bioactive compound from Rhizoma smilacis glabrae, has been reported to possess anti-inflammatory properties. Our study first evaluated astilbin on dextran sulfate sodium (DSS)-induced acute colitis in mice. By intraperitoneal injection of astilbin, the severity of colitis was attenuated, and the serum levels of IL-10 and TGF-β were increased. Using flow cytometry, a higher number of IL-10⁺ dendritic cells (DCs) and TGF-β⁺ DCs and a lower number of CD86⁺ DCs, IL-12 p40⁺ DCs, and IL-1β⁺ DCs were detected in the spleen of mice with colitis after astilbin treatment. The administration of astilbin also resulted in the upregulation of CD103⁺ expression in colonic DCs. In a coculture system, murine bone marrow-derived DCs pretreated with astilbin resulted in an enhanced production of CD4⁺CD25⁺Foxp3⁺ T cells. The results of this study show that astilbin could be a candidate drug for inflammatory bowel disease by mediating the regulatory functions of DCs.     

Increased expression of mGITRL on D2SC/1 cells by particulate β-glucan impairs the suppressive effect of CD4+CD25+ regulatory T cells and enhances the effector T cell proliferation

β-Glucans have been shown to enhance immune responses for centuries, which contributes to their anti-tumor property. However, their mechanisms of action are still elusive. Dectin-1, the C-type lectin receptor for β-glucan, is expressed abundantly on dendritic cells (DCs). Activation of DCs via Dectin-1 can lead to the maturation of DC, inducing both innate and adaptive immune responses against tumor development and microbial infection. In this study, we found that particulate yeast-derived β-glucans could induce the maturation of murine dendritic cell line D2SC/1 cells and increase the expression of mGITRL on D2SC/1 cells via Dectin-1/Syk pathway in a dose dependent manner. Furthermore, we demonstrated that the increased mGITRL on D2SC/1 cells could impair the suppressive activity of CD4⁺CD25⁺ regulatory T cells (Tregs) and enhance the proliferation of CD4⁺CD25⁻ effector T cells (Teffs). These findings suggest that particulate β-glucan can be used as immunomodulator to stimulate potent T cell-mediated adaptive immunity while down-regulate immune suppressive activity, leading to a more efficient defense mechanism against tumor development or infectious diseases.     

Developmental Regulation by Cytokines of Bone Marrow-Derived Dendritic Cells and Epidermal Langerhans Cells

Dendritic cells (DC) are specialized antigen-presenting cells involved in T cell-mediated immune responses. Differentiation and functional maturation of the DC are now known to be regulated by various cytokines, including TGF-β1. The experiments of this study examined the effect of other cytokines, such as IL-4, IL-10 and IL-6, on the differentiation and maturation of bone marrow (BM)-derived DC (BM-DC) and epidermal Langerhans cells (LC). When IL-6 or IL-10 was added to cultures of BM cells in the presence of GM-CSF, both cytokines, as in the case of TGF-β1, suppressed the maturation of DC in terms of the expression of adhesion and costimulatory molecules and T cell-stimulating activity. In contrast, IL-4 was not suppressive but rather supportive for the differentiation of DC. However, these suppressive cytokines hardly counteracted the maturation-inducing activity of TNF-α when added to cultures of immature DC. In addition, they appeared to block the overmaturation of DC, which is characterized by a loss of MHC class II molecules. Regarding LC maturation in epidermal cell cultures, IL-6 and IL-10 were inhibitory for the expression of CD86 and CD80 in a dose-dependent fashion. Unlike BM-DC, LC maturation was slightly enhanced by TGF-β1. The protein antigen-presentation by LC to Th1 clone was not affected by IL-6, but slightly reduced by IL-10. These results suggest that each cytokine contributes to regulate the differentiation and maturation of DC at a different developmental stage.     

Regulatory CD8 T cells induced by exposure to all-trans retinoic acid and TGF-β suppress autoimmune diabetes

Antigen-specific regulatory CD4⁺ T cells have been described but there are few reports on regulatory CD8⁺ T cells. We generated islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP)-specific regulatory CD8⁺ T cells from 8.3-NOD transgenic mice. CD8⁺ T cells from 8.3-NOD splenocytes were cultured with IGRP, splenic dendritic cells (SpDCs), TGF-β, and all-trans retinoic acid (ATRA) for 5 days. CD8⁺ T cells cultured with either IGRP alone or IGRP and SpDCs in the absence of TGF-β and ATRA had low Foxp3⁺ expression (1.7 ± 0.9% and 3.2 ± 4.5%, respectively). In contrast, CD8⁺ T cells induced by exposure to IGRP, SpDCs, TGF-β, and ATRA showed the highest expression of Foxp3⁺ in IGRP-reactive CD8⁺ T cells (36.1 ± 10.6%), which was approximately 40-fold increase compared with that before induction culture. CD25 expression on CD8⁺ T cells cultured with IGRP, SpDCs, TGF-β, and ATRA was only 7.42%, whereas CD103 expression was greater than 90%. These CD8⁺ T cells suppressed the proliferation of diabetogenic CD8⁺ T cells from 8.3-NOD splenocytes in vitro and completely prevented diabetes onset in NOD-scid mice in cotransfer experiments with diabetogenic splenocytes from NOD mice in vivo. Here we show that exposure to ATRA and TGF-β induces CD8⁺Foxp3⁺ T cells ex vivo, which suppress diabetogenic T cells in vitro and in vivo.     

Role of TGF-β and IL-6 in dendritic cells,Treg and Th17 mediated immune response during experimental cerebral malaria

The role of cytokines in Plasmodium infection have been extensively investigated, but pro and anti inflammatory cytokines mediated imbalance during malaria immune-pathogenesis is still unrevealed. Malaria is associated with the circulating levels of Interleukin-6 (IL-6) and transforming growth factor β (TGF-β), but association between these two cytokines in immune response remains largely obscured. Using mouse model, we proposed that IL-6 and TGF-β are involved in immune regulation of dendritic cells (DC), regulatory T cells (Treg), T-helper cells (Th17) during P. berghei ANKA (PbA) infection. Association between the cytokines and the severity of malaria was established with anti-TGF-β treatment resulting in increased parasitemia and increased immunopathology, whereas; anti-IL-6 treatment delays immunopathology during PbA infection. Further, splenocytes revealed differential alteration of myeloid DC (mDC), plasmocytoid DC (pDC), Treg, Th17 cells following TGF-β and IL-6 neutralization. Interestingly anti-TGF-β reduces CD11c⁺CD8⁺ DC expression, whereas anti-IL-6 administration causes a profound increase during PbA infection in Swiss mice. We observed down regulation of TGF-β, IL-10, NFAT, Foxp3, STAT-5 SMAD-3 and upregulation of IL-6, IL-23, IL-17 and STAT-3 in splenocytes during PbA infection. The STAT activity probably plays differential role in induction of Th17 and Treg cells. Interestingly we found increase in STAT-3 and decrease in STAT-5 expression during PbA infection. This pattern of STAT indicates that possibly TGF-β and IL-6 play a crucial role in differentiation of DCs subsets and Treg/Th17 imbalance during experimental cerebral malaria (ECM).     

Exposure of CD34+ precursors to cytostatic anthraquinone-derivatives induces rapid dendritic cell differentiation: implications for cancer immunotherapy

Appropriate activation of dendritic cells (DC) is essential for successful active vaccination and induction of cell-mediated immunity. The scarcity of precursor cells, as well as long culture methods, have hampered wide-scale application of DC vaccines derived from CD34⁺ precursors, despite their suggested superior efficacy over the more commonly applied monocyte-derived DC (MoDC). Here, employing the CD34⁺/CD14⁺ AML-derived human DC progenitor cell line MUTZ3, we show that cytostatic anthraquinone-derivatives (i.e., the anthracenedione mitoxantrone and the related anthracyclin doxorubicin) induce rapid differentiation of CD34⁺ DC precursors into functional antigen-presenting cells (APC) in a three-day protocol. The drugs were found to act specifically on CD34⁺, and not on CD14⁺ DC precursors. Importantly, these observations were confirmed for primary CD34⁺ and CD14⁺ DC precursors from peripheral blood. Mitoxantrone-generated DC were fully differentiated within three days and after an additional 24 h of maturation, were as capable as standard 9-day differentiated and matured DC to migrate toward the lymph node-homing chemokines CCL19 and CCL21, to induce primary allogeneic T cell proliferation, and to prime functional MART1-specific CD8⁺ T lymphocytes. Our finding that anthraquinone-derivatives like mitoxantrone support rapid high-efficiency differentiation of DC precursors may have consequences for in vitro production of DC vaccines as well as for novel immunochemotherapy strategies.     

Neutrophil elastase treated dendritic cells promote the generation of CD4(+)FOXP3(+) regulatory T cells in vitro

We have previously shown that neutrophilic elastase converts human immature dendritic cells (DCs) into TGF-β secreting cells and reduces its allostimulatory ability. Since TGF-β has been involved in regulatory T cells (Tregs) induction we analyzed whether elastase or neutrophil-derived culture supernatant treated DCs induce CD4⁺FOXP3⁺ Tregs in a mixed lymphocyte reaction (MLR). We found that elastase or neutrophil-derived culture supernatant treated DCs increased TGF-β and decreased IL-6 production. Together with this pattern of cytokines, we observed a higher number of CD4⁺FOXP3⁺ cells in the MLR cultures induced by elastase or neutrophil-derived culture supernatant treated DCs but not with untreated DCs. The higher number of CD4⁺FOXP3⁺ T cell population was not observed when the enzymatic activity of elastase was inhibited with an elastase specific inhibitor and also when a TGF-β1 blocking antibody was added during the MLR culture. The increased number of CD4⁺ that express FOXP3 was also seen when CD4⁺CD25^- purified T cells were cocultured with the TGF-β producing DCs. Furthermore, these FOXP3⁺ T cells showed suppressive activity in vitro.These results identify a novel mechanism by which the tolerogenic DCs generated by elastase exposure contribute to the immune regulation and may be relevant in the pathogenesis of several lung diseases where the inflammatory infiltrate contains high numbers of neutrophils and high elastase concentrations.     

Endogenous cortisol and TGF-beta in human aqueous humor contribute to ocular immune privilege by regulating dendritic cell function

Aqueous humor (AqH) has been shown to have significant immunosuppressive effects on APCs in animal models. We wanted to establish whether, in humans, AqH can regulate dendritic cell (DC) function and to identify the dominant mechanism involved. Human AqH inhibited the capacity of human peripheral blood monocyte-derived DC to induce naive CD4(+) T cell proliferation and cytokine production in vitro, associated with a reduction in DC expression of the costimulatory molecule CD86. This was seen both for DC cultured under noninflammatory conditions (immature DC) and for DC stimulated by proinflammatory cytokines (mature DC). DC expression of MHC classes I/II and CD83 was reduced (mature DC only). Myeloid DC from peripheral blood were similarly sensitive to the effects of human AqH, but only under inflammatory conditions. The addition of α-melanocyte stimulating hormone and vasoactive intestinal peptide did not cause significant inhibition at physiological levels. However, the addition of exogenous cortisol at physiological levels recapitulated the AqH-induced reduction in CD86 and inhibition of DC-induced T cell proliferation, and blockade of cortisol in AqH partially reversed its suppressive effects. TGF-β2 had an additional effect with cortisol, and although simultaneous blockade of cortisol and TGF-β2 in AqH reduced its effectiveness, there was still a cortisol- and TGF-β-independent component. In humans, AqH regulates DC maturation and function by the combined actions of cortisol and TGF-β2, a pathway that is likely to contribute to the maintenance of immune privilege in the eye.     

CD40 ligation converts TGF-β-secreting tolerogenic CD48 dendritic cells into IL-12-secreting immunogenic ones

CD40L, the ligand for CD40 on dendritic cells (DCs), plays an important role in maturation and activation of DCs leading to induction of immune responses. Our previous studies showed that the mouse splenic CD4⁻8⁻ DCs are tolerogenic and capable of stimulating suppressive type 1 CD4⁺ regulatory T (Tr1) cell responses via TGF-β secretion. In this study, we investigated whether CD40 ligation is able to convert tolerogenic CD4⁻8⁻ DCs into immunogenic ones by in vitro treatment of DCs with anti-CD40 antibody. Our data showed that in vitro CD40 ligation with anti-CD40 antibody converted TGF-β-secreting tolerogenic CD4⁻8⁻ DCs into IL-12-secreting immunogenic ones capable of stimulating type 1 CD4⁺ helper T (Th1) and CD8⁺ cytotoxic T lymphocyte (CTL) responses leading to induction of antitumor immunity. In addition, in vivo CD40 ligation by intratumoral injection of adenoviral vector AdVCD40L expressing CD40 ligand also induced tumor growth inhibition and regression of established P815 tumors with infiltration of tolerogenic CD4⁻8⁻ DCs. Therefore, our data provide new information for and may thus have useful impacts in CD40 ligation-based immunotherapy of cancer.     

Immunological properties of Fc receptor on lymphocytes. 5. Suppressive regulation of humoral immune response by Fc receptor bearing B lymphocytes.

High anti-DNP PFC responses to DNP-DE or DNP-KLH were obtained by transferring normal or primed FcR^? B cell fractions into irradiated syngeneic recipients. On the other hand, the FcR⁺ B cell fraction showed a low precursor activity. Trypsinization of the FcR⁺ B cells, to eliminate remaining antigen-antibody complexes on the surface, failed to augment the response in comparison with that of trypsin-untreated FcR⁺ B cells. Therefore, the weak precursor activity of FcR⁺ B cells seemed to be inherent. No synergistic interaction between the FcR⁺ B and precursor FcR^? B cells, to give rise to the maximum PFC response, was observed. On the contrary, the FcR⁺ B cells significantly suppressed the PFC responses of FcR^? B cells. This kind of suppression could be mediated by a factor released from the FcR⁺ B cell, but not from the FcR^? B or original-unrosetted spleen cell fraction. The factor was not attributable to macrophages, because the FcR⁺ B cells isolated from normal spleen cells, of which macrophages were depleted by Sephadex G-10 columns, could produce the factor with the same activity. Stimulation by specific antigen is not necessary for the induction of the factor(s) as well as of the suppressing FcR⁺ B cells. It seems to be necessary to stimulate FcR by antigen-antibody complexes to produce or release this factor.     

Blocking retinoic acid receptor-α enhances the efficacy of a dendritic cell vaccine against tumours by suppressing the induction of regulatory T cells

The immune system has evolved regulatory mechanisms to control immune responses to self-antigens. Regulatory T (Treg) cells play a pivotal role in maintaining immune tolerance, but tumour growth is associated with local immunosuppression, which can subvert effector immune responses. Indeed, the induction and recruitment of Treg cells by tumours is a major barrier in the development of effective immunotherapeutics and vaccines against cancer. Retinoic acid (RA) has been shown to promote conversion of naïve T cells into Treg cells. This study addresses the hypothesis that blocking RA receptor alpha (RARα) may enhance the efficacy of a tumour vaccine by inhibiting the induction of Treg cells. We found that RA significantly enhanced TGF-β-induced expression of Foxp3 on naïve and committed T cells in vitro and that this was blocked by an antagonist of RARα (RARi). In addition, RARi significantly suppressed TGF-β and IL-10 and enhanced IL-12 production by dendritic cells (DC) in response to killed tumour cells or TLR agonists. Furthermore, RARi augmented the efficacy of an antigen-pulsed and TLR-activated DC vaccine, significantly attenuating growth of B16 tumours in vivo and enhancing survival of mice. This protective effect was associated with significant reduction in tumour-infiltrating FoxP3⁺ and IL-10⁺ Treg cells and a corresponding increase in tumour-infiltrating CD4⁺ and CD8⁺ T cells that secreted IFN-γ. Our findings demonstrate that RARα is an important target for the development of effective anti-tumour immunotherapeutics and for improving the efficacy of cancer vaccines.     

Mesenchymal Stem Cells Derived from Human Exfoliated Deciduous Teeth (SHEDs) Induce Immune Modulatory Profile in Monocyte-Derived Dendritic Cells

Background

Mesenchymal stem cells have prominent immune modulatory properties, which may have clinical applications; however their major source, bone marrow, is of limited availability. On the other hand, mesenchymal stem cells derived from human exfoliated deciduous teeth (SHEDs) are readily accessible, but their immune regulatory properties have not been completely investigated. This study was designed, therefore, to evaluate the SHEDs influence on DCs differentiation, maturation, ability to activate T cells and to expand CD4⁺Foxp3⁺ T cells.

Methodology/Principal Findings

The experiments were based in cellular co-culture during differentiation and maturation of monocyte derived-DCs (moDCs), with, or not, presence of SHEDs. After co-culture with SHEDs, (moDCs) presented lower expression of BDCA-1 and CD11c, in comparison to DC cultivated without SHEDs. CD40, CD80, CD83 and CD86 levels were also decreased in mature DCs (mDCs) after co-cultivation with SHEDs. To assess the ability of SHEDs-exposed moDCs to modulate T cell responses, the former were separated from SHEDs, and co-cultured with peripheral blood lymphocytes. After 5 days, the proliferation of CD4⁺ and CD8⁺ T cells was evaluated and found to be lower than that induced by moDCs cultivated without SHEDs. In addition, an increase in the proportion of CD4⁺Foxp3⁺IL-10⁺ T cells was observed among cells stimulated by mature moDCs that were previously cultivated with SHEDs. Soluble factors released during co-cultures also showed a reduction in the pro-inflammatory cytokines (IL-2, TNF-α and IFN-γ), and an increase in the anti-inflammatory molecule IL-10.

Conclusion/Significance

This study shows that SHEDs induce an immune regulatory phenotype in moDCs cells, evidenced by changes in maturation and differentiation rates, inhibition of lymphocyte stimulation and ability to expand CD4⁺Foxp3⁺ T cells. Further characterization and validation of this phenomenon could support the use of SHEDs, directly or indirectly for immune modulation in the clinical practice.     

Role of macrophage colony-stimulating factor in the differentiation and expansion of monocytes and dendritic cells from CD34+ progenitor cells

The present study focused on whether it is possible to expand monocytic cells from CD34⁺ progenitor cells by using macrophage colony-stimulating factor (M-CSF) in the absence and presence of mast cell growth factor (MGF) and IL-6. It was demonstrated that CD34⁺ cells differentiate without expansion to functional mature monocytic cells in the presence of M-CSF or combinations of M-CSF plus IL-6 and MGF. A different response pattern was observed for the number of clonogenic cells. The addition of IL-6 or both IL-6 and MGF to M-CSF containing cultures resulted in significant higher numbers of colony-forming unit-macrophage (CFU-M) as tested in clonogenic and³H-thymidine assays. Furthermore, M-CSF plus both IL-6 and MGF appeared to be the most potent combination to preserve the monocytic precursor in cell suspension culture assays. These results indicate that IL-6 and MGF in conjunction with M-CSF affect CD34⁺ cells especially at precursor level without distinct effect on the more mature stages. Secondly we studied whether M-CSF is only critical for the monocytic lineage or also affects dendritic cell (DC) development. Indeed, we were able to culture CD83⁺ DC from CD34⁺ progenitor cells in the presence of M-CSF in conjunction with TNF-α, IL-4, and MGF although their absolute number is almost threefold lower than the number of CD83⁺ cells yielded from GM-CSF plus TNF-α, IL-4, and MGF stimulated CD34⁺ cells.     

The Mycobacterium avium subsp. Paratuberculosis protein MAP1305 modulates dendritic cell-mediated T cell proliferation through Toll-like receptor-4

In this study, we show that Mycobacterium avium subsp. Paratuberculosis MAP1305 induces the maturation of bone marrow-derived dendritic cells (BMDCs), a representative antigen presenting cell (APC). MAP1305 protein induces DC maturation and the production of pro-inflammatory cytokines (Interleukin (IL)-6), tumor necrosis factor (TNF)-α, and IL-1β) through Toll like receptor-4 (TLR-4) signaling by directly binding with TLR4. MAP1305 activates the phosphorylation of MAPKs, such as ERK, p38MAPK, and JNK, which is essential for DC maturation. Furthermore, MAP1305-treated DCs transform naïve T cells to polarized CD4⁺ and CD8⁺ T cells, thus indicating a key role for this protein in the Th1 polarization of the resulting immune response. Taken together, M. avium subsp. Paratuberculosis MAP1305 is important for the regulation of innate immune response through DC-mediated proliferation of CD4⁺ and CD8⁺ T cells. [BMB Reports 2014; 47(2): 115-120]     

CD1d‐mediated lipid presentation by CD11c+ cells regulates intestinal homeostasis

Intestinal homeostasis relies on a continuous dialogue between the commensal bacteria and the immune system. Natural killer T (NKT) cells, which recognize CD1d‐restricted microbial lipids and self‐lipids, contribute to the regulation of mucosal immunity, yet the mechanisms underlying their functions remain poorly understood. Here, we demonstrate that NKT cells respond to intestinal lipids and CD11c⁺ cells (including dendritic cells (DCs) and macrophages) are essential to mediate lipid presentation within the gut ultimately controlling intestinal NKT cell homeostasis and activation. Conversely, CD1d and NKT cells participate in the control of the intestinal bacteria composition and compartmentalization, in the regulation of the IgA repertoire and in the induction of regulatory T cells within the gut. These changes in intestinal homeostasis require CD1d expression on DC/macrophage populations as mice with conditional deletion of CD1d on CD11c⁺ cells exhibit dysbiosis and altered immune homeostasis. These results unveil the importance of CD11c⁺ cells in controlling lipid‐dependent immunity in the intestinal compartment and reveal an NKT cell–DC crosstalk as a key mechanism for the regulation of gut homeostasis.     

Opisthorchis viverrini antigens up-regulates the expression of CD80 and MHC class II in JAWSII mouse dendritic cells and promotes IL-10 and TGF-β secretions

Dendritic cells (DCs) are antigen-presenting cells (APC) involved in the initiation of immune responses. Maturation of DCs is characterized by the high expression of major histocompatibility complex (MHC) class II and co-stimulatory clusters of differentiation (CD) 40, CD80, and CD86 molecules. Matured DCs are required for T cell differentiation and proliferation. However, the response of DCs to Opisthorchis viverrini antigens has not yet been understood. Therefore, this study sought to determine the expression of surface molecules of JAWSII mouse DCs stimulated by crude somatic (CS) and excretory-secretory (ES) antigens of O. viverrini. ES antigen significantly induced only mRNA expression of CD80 and MHC class II in JAWSII mouse DCs, while CS antigen promoted up-regulation of both mRNA and protein levels of CD80 and MHC class II, indicating relative maturation of JAWII mouse DCs. Moreover, the secreted cytokines from the co-cultures of O. viverrini antigens stimulated JAWSII DC with naïve CD4⁺ T cells was determined. Significantly increased levels of immunosuppressive cytokines interleukin (IL)-10 and transforming growth factor beta (TGF-β) were found. The up-regulation of these cytokines may indicate the response of regulatory T cells (Treg) to CS antigen-stimulated JAWSII DC. These findings may lead to a better understanding of the role that DCs play in O. viverrini infection.     

Application of immunotherapy based on dendritic cells stimulated by tumor cell-derived exosomes in a syngeneic breast tumor mouse model

We here evaluated the therapeutic effect of tumor cell-derived exosomes (TEXs)-stimulated dendritic cells (DCs) in a syngeneic orthotopic breast tumor model. The DC line DC2.4 and breast cancer cell line E0771 originally isolated from C57BL/6 mice were used. E0771 cells stably expressing the exosomal CD63-RFP or luciferase (Luc) and DC2.4 cells stably expressing GFP were produced using lentivirus. TEXs were purified from conditioned medium of E0771/CD63-RFP cells. Breast tumor model was established by injecting E0771/Luc cells into mammary gland fat pad of mice. TEXs contained immune modulatory molecules such as HSP70, HSP90, MHC I, MHC II, TGF-β, and PD-L1. TEXs were easily taken by DC2.4 cells, resulting in a significant increase in the in vitro proliferation and migration abilities of DC2.4 cells, accompanied by the upregulation of CD40. TEX-DC-treated group exhibited a decreased tumor growth compared with control group. CD8⁺ cells were more abundant in the tumors and lymph nodes of TEX-DC-treated group than in those of control group, whereas many CD4⁺ or FOXP3+ cells were localized in those of control group. Our results suggest a potential application of TEX-DC-based cancer immunotherapy.     

Immunological properties of Fc receptor on lymphocytes. 3. Stimulating activities of Fc receptor-bearing and -nonbearing cells in mixed-lymphocyte reaction.

By separating FcR⁺ and FcR^? cells from stimulator spleens using an EA rosetting procedure, it was found that EA-rosetting (FcR⁺) cells stimulate mixed-lymphocyte culture reaction (MLR) far more effectively than do non-EA-rosetting (FcR^?) cells. The difference in stimulatory activity is observed in MLR of both H-2 and M-locus different combinations and cannot be explained by the proportion of B cells and macrophages contained in each population. The finding that FcR⁺ cells can stimulate allogeneic responding T cells more effectively than FcR^? cells suggests a close association of FcR with Ia and Mls antigens on the cell surface.