Abstract Although positive CD28 costimulation is needed for the generation of natural CD4+CD25+ regulatory T cells, we report that negative CTLA-4 costimulation is necessary for generating phenotypically and functionally similar adaptive CD4+CD25+ suppressor cells. TGF-β could not induce CD4+CD25− cells from CTLA-4−/− mice to express normal levels of FoxP3 or to develop suppressor activity. Moreover, blockade of CTLA-4 following activation of wild-type CD4+ cells abolished the ability of TGF-β to induce FoxP3-expressing mouse suppressor cells. TGF-β accelerated expression of CTLA-4, and time course studies suggested that CTLA-4 ligation of CD80 shortly after T cell activation enables TGF-β to induce CD4+CD25− cells to express FoxP3 and develop suppressor activity. TGF-β also enhanced CD4+ cell expression of CD80. Thus, CTLA-4 has an essential role in the generation of acquired CD4+CD25+ suppressor cells in addition to its other inhibitory effects. Although natural CD4+CD25+ cells develop normally in CTLA-4−/− mice, the lack of TGF-β-induced, peripheral CD4+CD25+ suppressor cells in these mice may contribute to their rapid demise.
Group 2 innate lymphoid cells (ILC2s) are recently reported to play a more critical role in allergic diseases. We previously identified that mesenchymal stromal cells (MSCs) elicited therapeutic effects on allergic airway inflammation. Small extracellular vesicles (sEV) derived from MSCs possess striking advantages including low immunogenicity and high biosafety, and is extremely promising cell-free therapeutic agents. However, the effects of MSC-sEV on ILC2s are still unclear. Additionally, scalable isolation protocols are required for the mass production of homogenous MSC-sEV especially in clinical application. We previously reported that induced pluripotent stem cells-derived MSCs were the ideal cellular source for the large preparation of MSC-sEV. Here we developed a standardized scalable protocol of anion-exchange chromatography for isolation of MSC-sEV, and investigated the effects of MSC-sEV on ILC2 function from patients with allergic rhinitis and in a mouse ILC2-dominant asthma model. The characterization of MSC-sEV was successfully demonstrated in terms of size, morphology and specific markers. Using flow cytometry and human Cytokine Antibody Array, MSC-sEV but not fibroblasts-sEV (Fb-sEV) were found to significantly inhibit the function of human ILC2s. Similarly, systemic administration of MSC-sEV but not Fb-sEV exhibited an inhibition of ILC2 levels, inflammatory cell infiltration and mucus production in the lung, a reduction in levels of T helper 2 cytokines, and alleviation of airway hyperresponsiveness in a mouse model of asthma. Using RNA sequencing, miR-146a-5p was selected as the candidate to mediate the above effects of MSC-sEV. We next revealed the uptake of ILC2s to MSC-sEV, and that transfer of miR-146a-5p in MSC-sEV to ILC2s in part contributed to the effects of MSC-sEV on ILC2s in vitro and in a mouse model. In conclusion, we demonstrated that MSC-sEV were able to prevent ILC2-dominant allergic airway inflammation at least partially through miR-146a-5p, suggesting that MSC-sEV could be a novel cell-free strategy for the treatment of allergic diseases.
Abstract TGF-β is crucial for induction of CD4+Foxp3+ Tregs and maintenance of immunologic tolerance. It is, however, unclear if TGF-β also induces the similar CD8+ Tregs. While CD8+ cells primed with TGF-β (CD8+ iTregs) displayed much lower Foxp3 expression compared with compartment CD4+ cells, their suppression activity in vitro and in vivo was equivalent or even better than CD4+ Tregs. These cells did not express Granzyme A, Granzyme B or Perforin and lacked cytotoxic activity on T response cells. Additionally, CD8+ iTregs generated from Granzyme B and Perforin KO mice still suppressed autoimmunity. Transwell experiments revealed that cell-contact is required for their suppressive activity. Adoptive transfer of the CD8+ iTregs markedly suppressed experimental colitis, EAE and cGVHD. We further found both Foxp3- and Foxp3+ subsets from TGF-primed CD8+ cells had suppressive activities. Among CD8+Foxp3- cells, we identified CD103 expression is crucial for their generation and function since CD8+ but not CD4+ iTreg production decreased on CD103 KO mice. Adoptive transfer of CD8+CD103+Foxp3- subset suppressed colitis, and IL-10 and TGF-β signal seem to be crucial for this therapeutic effect. TGF-β can induce CD8+Foxp3- and CD8+Foxp3+ iTreg subsets that displayed suppressive activity in cell contact-dependent, non-cytotoxic manner and have protective effects on autoimmune diseases. Generation of CD8+ iTregs may have considerable therapeutic potential on patients with autoimmune diseases.
Graft-versus-host disease (GVHD) is a severe adverse effect that results from bone marrow or peripheral blood cells transplantation and has a high rate of mortality. About 50% of the patients are accompanied with acute Graft-versus-Host Disease (aGVHD) after bone marrow cell transplantation and need systematic treatment. It has an important clinical significance to evaluate the prevention and treatment effects of GVHD. The stable and reliable approaches of humanized animal models are crucial for advancing on the study the biology of GVHD. Relative models transplanting the human immune cells into the mouse body can trigger immunoreaction similar to the humans. As it is a disease triggered by human immune cells, any intervention research prior to clinical treatment has more clinical interrelations compared with the general animal models. In this review, we update the current understanding on humanized animal disease models on studying Graft-versus-host disease and expect to provide more theoretical basis to further study on Graft-versus-host disease.
Atherosclerosis is a chronic inflammatory disease which results in thickening of the vessel wall and narrowing of the lumen. It is a leading cause of death worldwide. Preventive treatment is taken into prioritized consideration since currently no effective approaches to cure atherosclerosis are available. These treatments mainly focus on lowering blood cholesterol levels, especially LDL-C, by statins. Even so, lowering lipid levels is not sufficient to reduce the risk of cardiovascular events in all patients. Recently, atherosclerosis has increasingly been recognized as a chronic inflammatory disease involving the immune system, initiating new therapeutic approaches which could alleviate or prevent atherosclerosis by modulating inflammation. Mesenchymal stem cells (MSCs) have emerged as a promising option to relieve inflammation and balance immune responses in inflammatory diseases. Several studies including our group also reported that MSCs may be a new therapeutic option for atherosclerosis. This review summarizes the updated state of our knowledge in the administration of MSCs to alleviate atherosclerosis and discusses some of the key unresolved challenges that need to be solved in future studies.
Background: Rheumatoid arthritis (RA) is a progressive, chronic, even disabling systemic autoimmune disease. Imbalance between pathogenic immune cells and immunosuppressive cells is associated with the pathogenesis and development of RA and other autoimmune diseases. As Foxp3 is also expressed on activated CD4 cells in the presence of inflammation, the identification of Treg cells in patients with RA remains a challenge.
Interleukin 1 is a critical inflammatory mediator and involved in host defense to several pathogens. Oral T. gondii infection causes lethal ileitis in C57BL/6 (BL6) mice and serves to investigate the mechanisms of acute intestinal inflammation. Here we show that IL-1 is expressed upon oral T. gondii (76K strain) infection in the small intestine and mediates ileitis as IL-1R1 deficient mice have reduced neutrophil recruitment in the lamina propria, parasite invasion, inflammatory lesions and enhanced survival as compared to BL6 infected control mice. Protection in the absence of IL-1R1 signaling was associated with reduced IFN-γ expression and preserved Paneth cells, while these cells were eliminated in infected BL6 mice. Furthermore, blockade of IL-1 by IL-1β antibody attenuated inflammation in BL6 mice. In conclusion, IL-1 signaling contributes to the inflammatory response with increase IFN-γ expression and Paneth cell depletion upon oral T. gondii infection.
Retinal ischemia-reperfusion injury (IRI) is one of the main pathogenic mechanisms of glaucoma, which are largely unknown, including neuroinflammation and neuronal death in the pathological process. In our previous studies, mesenchymal stem cells (MSCs) have been reported to play anti-inflammatory and neuroprotective roles. Additionally, conditioned culture medium (CM) of MSCs stimulated by TNF-α have achieved better antiallergic effects in an experimental allergic conjunctivitis mouse model. However, there is an urgent need for cell-free therapy approaches, like exosomes, to reduce the side effects of autoimmunity. The present study aimed to elucidate the pathways involving TNF-α-stimulated gingival MSC (GMSC)-exosomes (TG-exos), in modulating inflammatory microglia and alleviating apoptosis. In this study, exosomes from the CM of GMSCs were isolated by ultracentrifugation and were injected into the vitreous of mice. The results showed that intraocular injection of TG-exos into mice with IRI notably reduced inflammation and cell loss than that with G-exos (GMSC-exosomes). Similar results were observed in vitro. Additionally, with the microRNA (miR) arrays, it was found that miR-21-5p acted as a crucial factor in TG-exos for neuroprotection and anti-inflammation. Following target prediction and dual-luciferase assay suggested that miR-21-5p played a role by combining with programmed cell death 4 (PDCD4), which was regulated by the long non-coding RNA (lncRNA) maternally expressed gene 3 (MEG3) as a competing endogenous RNA (ceRNA). This study demonstrates a new therapeutic pathway for neuroprotection against IRI by delivering miR-21-5p-enriched exosomes through MEG3/miR-21-5p/PDCD4 axis and paves the way for the establishment of a cell-free therapeutic approach for glaucoma.
Whereas TGF-beta is essential for the development of peripherally induced Foxp3(+) regulatory T cells (iTreg cells) and Th17 cells, the intracellular signaling mechanism by which TGF-beta regulates development of both cell subsets is less understood. In this study, we report that neither Smad2 nor Smad3 gene deficiency abrogates TGF-beta-dependent iTreg induction by a deacetylase inhibitor trichostatin A in vivo, although the loss of the Smad2 or Smad3 gene partially reduces iTreg induction in vitro. Similarly, SMAD2 and SMAD3 have a redundant role in development of Th17 in vitro and in experimental autoimmune encephalomyelitis. In addition, ERK and/or JNK pathways were shown to be involved in regulating iTreg cells, whereas the p38 pathway predominately modulated Th17 and experimental autoimmune encephalomyelitis induction. Therefore, selective targeting of these intracellular TGF-beta signaling pathways during iTreg and Th17 cell development might lead to the development of therapies in treating autoimmune and other chronic inflammatory diseases.
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