Amelioration of experimental autoimmune encephalomyelitis by in vivo reprogramming of macrophages using pro-resolving factors.

BACKGROUND: Reinstating inflammation resolution represents an innovative concept to regain inflammation control in diseases marked by chronic inflammation. While most therapeutics target inflammatory molecules and inflammatory effector cells and mediators, targeting macrophages to initiate inflammation resolution to control neuroinflammation has not yet been attempted. Resolution-phase macrophages are critical in the resolution process to regain tissue homeostasis, and are programmed through the presence and elimination of apoptotic leukocytes. Hence, inducing resolution-phase macrophages might represent an innovative therapeutic approach to control and terminate dysregulated neuroinflammation. METHODS: Here, we investigated if the factors released by in vitro induced resolution-phase macrophages (their secretome) are able to therapeutically reprogram macrophages to control neuroinflammation in the model of experimental autoimmune encephalomyelitis (EAE). RESULTS: We found that injection of the pro-resolutive secretome reduced demyelination and decreased inflammatory cell infiltration in the CNS, notably through the in vivo reprogramming of macrophages at the epigenetic level. Adoptive transfer experiments with in vivo or in vitro reprogrammed macrophages using such pro-resolutive secretome confirmed the stability and transferability of this acquired therapeutic activity. CONCLUSIONS: Overall, our data confirm the therapeutic activity of a pro-resolution secretome in the treatment of ongoing CNS inflammation, via the epigenetic reprogramming of macrophages and open with that a new therapeutic avenue for diseases marked by neuroinflammation.

A critical function for TGF-beta signaling in the development of natural CD4+CD25+Foxp3+ regulatory T cells.

The molecular mechanisms directing the development of 'natural' CD4+CD25+Foxp3+ regulatory T cells (T(reg) cells) in the thymus are not thoroughly understood. We show here that conditional deletion of transforming growth factor-beta receptor I (TbetaRI) in T cells blocked the appearance of CD4+CD25+Foxp3+ thymocytes at postnatal days 3-5. Paradoxically, however, beginning 1 week after birth, the same TbetaRI-mutant mice showed accelerated expansion of thymic CD4+CD25+Foxp3+ populations. This rapid recovery of Foxp3+ thymocytes was attributable mainly to overproduction of and heightened responsiveness to interleukin 2, as genetic ablation of interleukin 2 in TbetaRI-mutant mice resulted in a complete absence of CD4+CD25+Foxp3+ cells from the thymus and periphery. Thus, transforming growth factor-beta signaling is critical to the thymic development of natural CD4+CD25+Foxp3+ T(reg) cells.

Photopheresis efficacy in the treatment of rheumatoid arthritis: a pre-clinical proof of concept.

BACKGROUND: Despite major advances in rheumatoid arthritis outcome, not all patients achieve remission, and there is still an unmet need for new therapeutic approaches. This study aimed at evaluating in a pre-clinical murine model the efficacy of extracorporeal photopheresis (ECP) in the treatment of rheumatoid arthritis, and to provide a relevant study model for dissecting ECP mechanism of action in autoimmune diseases. METHODS: DBA/1 mice were immunized by subcutaneous injection of bovine collagen type II, in order to initiate the development of collagen-induced arthritis (CIA). Arthritic mice received 3 ECP treatments every other day, with psoralen + UVA-treated (PUVA) spleen cells obtained from arthritic mice. Arthritis score was measured, and immune cell subsets were monitored. RESULTS: ECP-treated mice recovered from arthritis as evidenced by a decreasing arthritic score over time. Significant decrease in the frequency of Th17 cells in the spleen of treated mice was observed. Interestingly, while PUVA-treated spleen cells from healthy mouse had no effect, PUVA-treated arthritic mouse derived-spleen cells were able to induce control of arthritis development. CONCLUSIONS: Our results demonstrate that ECP can control arthritis in CIA-mice, and clarifies ECP mechanisms of action, showing ECP efficacy and Th17 decrease only when arthritogenic T cells are contained within the treated sample. These data represent a pre-clinical proof of concept supporting the use of ECP in the treatment of RA in Human.

LXR agonist treatment of blastic plasmacytoid dendritic cell neoplasm restores cholesterol efflux and triggers apoptosis.

Blastic plasmacytoid dendritic cell (PDC) neoplasm (BPDCN) is an aggressive hematological malignancy with a poor prognosis that derives from PDCs. No consensus for optimal treatment modalities is available today and the full characterization of this leukemia is still emerging. We identified here a BPDCN-specific transcriptomic profile when compared with those of acute myeloid leukemia and T-acute lymphoblastic leukemia, as well as the transcriptomic signature of primary PDCs. This BPDCN gene signature identified a dysregulation of genes involved in cholesterol homeostasis, some of them being liver X receptor (LXR) target genes. LXR agonist treatment of primary BPDCN cells and BPDCN cell lines restored LXR target gene expression and increased cholesterol efflux via the upregulation of adenosine triphosphate-binding cassette (ABC) transporters, ABCA1 and ABCG1. LXR agonist treatment was responsible for limiting BPDCN cell proliferation and inducing intrinsic apoptotic cell death. LXR activation in BPDCN cells was shown to interfere with 3 signaling pathways associated with leukemic cell survival, namely: NF-κB activation, as well as Akt and STAT5 phosphorylation in response to the BPDCN growth/survival factor interleukin-3. These effects were increased by the stimulation of cholesterol efflux through a lipid acceptor, the apolipoprotein A1. In vivo experiments using a mouse model of BPDCN cell xenograft revealed a decrease of leukemic cell infiltration and BPDCN-induced cytopenia associated with increased survival after LXR agonist treatment. This demonstrates that cholesterol homeostasis is modified in BPDCN and can be normalized by treatment with LXR agonists which can be proposed as a new therapeutic approach.

Plasmacytoid Dendritic Cells Die by the CD8 T Cell-Dependent Perforin Pathway during Acute Nonviral Inflammation.

Regulation of the inflammatory response involves the control of dendritic cell survival. To our knowledge, nothing is known about the survival of plasmacytoid dendritic cells (pDC) in such situation. pDC are specialized in type I IFN (IFN-I) secretion to control viral infections, and IFN-I also negatively regulate pDC survival during the course of viral infections. In this study, we asked about pDC behavior in the setting of virus-free inflammation. We report that pDC survival was profoundly reduced during different nonviral inflammatory situations in the mouse, through a mechanism independent of IFN-I and TLR signaling. Indeed, we demonstrated that during inflammation, CD8(+) T cells induced pDC apoptosis through the perforin pathway. The data suggest, therefore, that pDC have to be turned down during ongoing acute inflammation to not initiate autoimmunity. Manipulating CD8(+) T cell response may therefore represent a new therapeutic opportunity for the treatment of pDC-associated autoimmune diseases, such as lupus or psoriasis.

Concise Review: Apoptotic Cell-Based Therapies-Rationale, Preclinical Results and Future Clinical Developments.

The objectives of this review are to summarize the experimental data obtained using apoptotic cell-based therapies, and then to discuss future clinical developments. Indeed, apoptotic cells exhibit immunomodulatory properties that are reviewed here by focusing on more recent mechanisms. These immunomodulatory mechanisms are in particular linked to the clearance of apoptotic cells (called also efferocytosis) by phagocytes, such as macrophages, and the induction of regulatory T cells. Thus, apoptotic cell-based therapies have been used to prevent or treat experimental inflammatory diseases. Based on these studies, we have identified critical steps to design future clinical trials. This includes: the administration route, the number and schedule of administration, the appropriate apoptotic cell type to be used, as well as the apoptotic signal. We also have analyzed the clinical relevancy of apoptotic-cell-based therapies in experimental models. Additional experimental data are required concerning the treatment of inflammatory diseases (excepted for sepsis) before considering future clinical trials. In contrast, apoptotic cells have been shown to favor engraftment and to reduce acute graft-versus-host disease (GvHD) in different relevant models of transplantation. This has led to the conduct of a phase 1/2a clinical trial to alleviate GvHD. The absence of toxic effects obtained in this trial may support the development of other clinical studies based on this new cell therapy. Stem Cells 2016;34:1464-1473.

Human monocyte-derived suppressor cells control graft-versus-host disease by inducing regulatory forkhead box protein 3-positive CD8+ T lymphocytes.

BACKGROUND: Adoptive transfer of immunosuppressive cells has emerged as a promising strategy for the treatment of immune-mediated disorders. However, only a limited number of such cells can be isolated from in vivo specimens. Therefore efficient ex vivo differentiation and expansion procedures are critically needed to produce a clinically relevant amount of these suppressive cells. OBJECTIVE: We sought to develop a novel, clinically relevant, and feasible approach to generate ex vivo a subpopulation of human suppressor cells of monocytic origin, referred to as human monocyte-derived suppressive cells (HuMoSCs), which can be used as an efficient therapeutic tool to treat inflammatory disorders. METHODS: HuMoSCs were generated from human monocytes cultured for 7 days with GM-CSF and IL-6. The immune-regulatory properties of HuMoSCs were investigated in vitro and in vivo. The therapeutic efficacy of HuMoSCs was evaluated by using a graft-versus-host disease (GvHD) model of humanized mice (NOD/SCID/IL-2Rγc(-/-) [NSG] mice). RESULTS: CD33+ HuMoSCs are highly potent at inhibiting the proliferation and activation of autologous and allogeneic effector T lymphocytes in vitro and in vivo. The suppressive activity of these cells depends on signal transducer and activator of transcription 3 activation. Of therapeutic relevance, HuMoSCs induce long-lasting memory forkhead box protein 3-positive CD8+ regulatory T lymphocytes and significantly reduce GvHD induced with human PBMCs in NSG mice. CONCLUSION: Ex vivo-generated HuMoSCs inhibit effector T lymphocytes, promote the expansion of immunosuppressive forkhead box protein 3-positive CD8+ regulatory T cells, and can be used as an efficient therapeutic tool to prevent GvHD.

Transforming growth factor-ß released by apoptotic white blood cells during red blood cell storage promotes transfusion-induced alloimmunomodulation.

BACKGROUND: Red blood cell (RBC) alloimmunization is a major immunologic risk of transfusion. However, RBC storage facilitates white blood cell (WBC) apoptosis and apoptotic cells have immunomodulatory properties. We investigated the behavior of WBCs, and apoptosis in particular, in RBC units during storage and then studied the impact of WBC apoptosis on the modulation of posttransfusion alloimmunization in RBC products stored short term. STUDY DESIGN AND METHODS: We used a mouse model of alloimmunization to transfused HEL-ovalbumin-Duffy (HOD) surface antigen expressed specifically on RBCs. The presence of circulating anti-HOD immunoglobulin G detected by flow cytometry confirmed immunization to HOD+ RBCs. WBC apoptosis and factors released by apoptotic WBCs during storage were determined and in particular the role of transforming growth factor (TGF)-ß was assessed on RBC alloimmunization. RESULTS: In blood stored 72 hours, 30% of WBCs were apoptotic, and transfusion of short-term-stored blood resulted in lesser immunization than did fresh blood or stored leukoreduced (LR) RBCs. WBCs undergoing apoptosis released during short-term storage factors modulating RBC alloimmunization. Indeed apoptotic cell-released factors modulate alloimmunization whereas exogenous apoptotic cells directly transfused with LR RBCs did not. While microparticles released during RBC storage had no immunomodulatory role, TGF-ß found in the supernatant of stored blood demonstrated the capacity to favor Treg polarization of naïve CD4+CD25- T cells in vitro and limited RBC alloimmunization in vivo. Indeed, addition of recombinant TGF-ß to stored LR RBC transfusion strongly limited posttransfusion RBC alloimmunization. CONCLUSION: Our findings show that short-term storage of non-LR blood facilitates WBC apoptosis therefore releasing TGF-ß that modulates posttransfusion RBC alloimmunization.

Macrophage phenotype is determinant for fibrosis development in keloid disease.

Keloid refers to a fibroproliferative disorder characterized by an accumulation of extracellular matrix (ECM) components at the dermis level, overgrowth beyond initial wound, and formation of tumor-like nodule areas. Treating keloid is still an unmet clinical need and the lack of an efficient therapy is clearly related to limited knowledge about keloid etiology, despite the growing interest of the scientific community in this pathology. In past decades, keloids were often studied in vitro through the sole prism of fibroblasts considered as the major effector of ECM deposition. Nevertheless, development of keloids results from cross-interactions of keloid fibroblasts (KFs) and their surrounding microenvironment, including immune cells such as macrophages. Our study aimed to evaluate the effect of M1 and M2 monocyte-derived macrophages on KFs in vitro. We focused on the effects of the macrophage secretome on fibrosis-related criteria in KFs, including proliferation, migration, differentiation, and ECM synthesis. First, we demonstrated that M2-like macrophages enhanced the fibrogenic profile of KFs in culture. Then, we surprisingly founded that M1-like macrophages can have an anti-fibrogenic effect on KFs, even in a pro-fibrotic environment. These results demonstrate, for the first time, that M1 and M2 macrophage subsets differentially impact the fibrotic fate of KFs in vitro, and suggest that restoring the M1/M2 balance to favor M1 in keloids could be an efficient therapeutic lever to prevent or treat keloid fibrosis.

Functions of TGF-ß-exposed plasmacytoid dendritic cells.

Plasmacytoid dendritic cells (pDCs) belong to the family of dendritic cells and possess specific features that distinguish them from conventional dendritic cells. For instance, pDC are the main interferon-alpha-secreting cells. Plasmacytoid dendritic cells exert both proinflammatory and regulatory functions. This is attested by the involvement of pDC through interferon-alpha secretion in several autoimmune diseases, and by the implication of pDC in tolerance. The same is true for TGF-ß that plays a dual role in inflammation. In this review, we discuss recent data on pDC and TGF-ß interactions. As with many cell types, pDCs are able to respond to TGF-ß using the classic Smad signaling pathway. In addition, pDCs are capable to secrete TGF-ß, in particular in response to TGF-ß exposure. Exposure of pDCs to TGF-ß prevents type I interferon secretion in response to TLR7/9 ligands. In contrast, the consequences of TGF-ß on the antigen-presenting cell capacities of pDC are less clear, since TGF-ß-exposed pDCs may lead to both regulatory T-cell and interleukin-17-secreting cell polarization. Here, we discuss the factors that may influence this polarization. We also discuss how pDCs exposed to TGF-ß may participate in tolerance induction and maintenance, or, on the contrary, in autoimmune diseases.

Plasmacytoid dendritic cells play a major role in apoptotic leukocyte-induced immune modulation.

Several APCs participate in apoptotic cell-induced immune modulation. Whether plasmacytoid dendritic cells (PDCs) are involved in this process has not yet been characterized. Using a mouse model of allogeneic bone marrow engraftment, we demonstrated that donor bone marrow PDCs are required for both donor apoptotic cell-induced engraftment and regulatory T cell (Treg) increase. We confirmed in naive mice receiving i.v. syngeneic apoptotic cell infusion that PDCs from the spleen induce ex vivo Treg commitment. We showed that PDCs did not interact directly with apoptotic cells. In contrast, in vivo macrophage depletion experiments using clodronate-loaded liposome infusion and coculture experiments with supernatant from macrophages incubated with apoptotic cells showed that PDCs required macrophage-derived soluble factors--including TGF-ß--to exert their immunomodulatory functions. Overall, PDCs may be considered as the major APC involved in Treg stimulation/generation in the setting of an immunosuppressive environment obtained by apoptotic cell infusion. These findings show that like other APCs, PDC functions are influenced, at least indirectly, by exposure to blood-borne apoptotic cells. This might correspond with an additional mechanism preventing unwanted immune responses against self-antigens clustered at the cell surface of apoptotic cells occurring during normal cell turnover.

TGF-beta-exposed plasmacytoid dendritic cells participate in Th17 commitment.

TGF-ß is required for both Foxp3(+) regulatory T cell (Treg) and Th17 commitment. Plasmacytoid DCs (pDC) have been shown to participate to both Treg and Th17 commitment as well. However, few studies have evaluated the direct effect of TGF-ß on pDC, and to our knowledge, no study has assessed the capacity of TGF-ß-exposed pDC to polarize naive CD4(+) T cells. In this paper, we show that TGF-ß-treated pDC favor Th17 but not Treg commitment. This process involves a TGF-ß/Smad signal, because TGF-ß treatment induced Smad2 phosphorylation in pDC and blockade of TGF-ß signaling with the SD208 TGF-ßRI kinase inhibitor abrogated Th17 commitment induced by TGF-ß-treated pDC. Moreover, TGF-ß mRNA synthesis and active TGF-ß release were induced in TGF-ß-treated pDC and anti-TGF-ß Ab blocked Th17 commitment. Unexpectedly, TGF-ß treatment also induced increased IL-6 production by pDC, which serves as the other arm for Th17 commitment driven by TGF-ß-exposed pDC, because elimination of IL-6-mediated signal with either IL-6- or IL-6Rα-specific Abs prevented Th17 commitment. The in vivo pathogenic role of TGF-ß-treated pDC was further confirmed in the Th17-dependent collagen-induced arthritis model in which TGF-ß-treated pDC injection significantly increased arthritis severity and pathogenic Th17 cell accumulation in the draining lymph nodes. Thus, our data reveal a previously unrecognized effect of TGF-ß-rich environment on pDC ability to trigger Th17 commitment. Such findings have implications in the pathogenesis of autoimmune diseases or immune responses against mucosal extracellular pathogens.

Resolution therapy: Harnessing efferocytic macrophages to trigger the resolution of inflammation.

Several chronic inflammatory diseases are associated with non-resolving inflammation. Conventional anti-inflammatory drugs fail to completely cure these diseases. Resolution pharmacology is a new therapeutic approach based on the use of pro-resolving mediators that accelerate the resolution phase of inflammation by targeting the productive phase of inflammation. Indeed, pro-resolving mediators prevent leukocyte recruitment and induce apoptosis of accumulated leukocytes. This approach is now called resolution therapy with the introduction of complex biological drugs and cell-based therapies. The main objective of resolution therapy is to specifically reduce the duration of the resolution phase to accelerate the return to homeostasis. Under physiological conditions, macrophages play a critical role in the resolution of inflammation. Indeed, after the removal of apoptotic cells (a process called efferocytosis), macrophages display anti-inflammatory reprogramming and subsequently secrete multiple pro-resolving factors. These factors can be used as resolution therapy. Here, we review the different mechanisms leading to anti-inflammatory reprogramming of macrophages after efferocytosis and the pro-resolving factors released by these efferocytic macrophages. We classify these mechanisms in three different categories: macrophage reprogramming induced by apoptotic cell-derived factors, by molecules expressed by apoptotic cells (i.e., "eat-me" signals), and induced by the digestion of apoptotic cell-derived materials. We also evoke that macrophage reprogramming may result from cooperative mechanisms, for instance, implicating the apoptotic cell-induced microenvironment (including cellular metabolites, specific cytokines or immune cells). Then, we describe a new drug candidate belonging to this resolution therapy. This candidate, called SuperMApo, corresponds to the secretome of efferocytic macrophages. We discuss its production, the pro-resolving factors present in this drug, as well as the results obtained in experimental models of chronic (e.g., arthritis, colitis) and acute (e.g., peritonitis or xenogeneic graft-versus-host disease) inflammatory diseases.

Lethal effect of CD3-specific antibody in mice deficient in TGF-beta1 by uncontrolled flu-like syndrome.

CD3-specific Ab therapy results in a transient, self-limiting, cytokine-associated, flu-like syndrome in experimental animals and in patients, but the underlying mechanism for this spontaneous resolution remains elusive. By using an in vivo model of CD3-specific Ab-induced flu-like syndrome, we show in this paper that a single injection of sublethal dose of the Ab killed all TGF-beta1(-/-) mice. The death of TGF-beta1(-/-) mice was associated with occurrence of this uncontrolled flu-like syndrome, as demonstrated by a sustained storm of systemic inflammatory TNF and IFN-gamma cytokines. We present evidence that deficiency of professional phagocytes to produce TGF-beta1 after apoptotic T cell clearance may be responsible, together with hypersensitivity of T cells to both activation and apoptosis, for the uncontrolled inflammation. These findings indicate a key role for TGF-beta1 and phagocytes in protecting the recipients from lethal inflammation and resolving the flu-like syndrome after CD3-specific Ab treatment. The study may also provide a novel molecular mechanism explaining the early death in TGF-beta1(-/-) mice.

Regulation of T-Cell Immune Responses by Pro-Resolving Lipid Mediators.

Both the initiation and the resolution of inflammatory responses are governed by the sequential activation, migration, and control/suppression of immune cells at the site of injury. Bioactive lipids play a major role in the fine-tuning of this dynamic process in a timely manner. During inflammation and its resolution, polymorphonuclear cells (PMNs) and macrophages switch from producing pro-inflammatory prostaglandins and leukotrienes to specialized pro-resolving lipid mediators (SPMs), namely, lipoxins, resolvins, protectins, and maresins, which are operative at the local level to limit further inflammation and tissue injury and restore homeostasis. Accumulating evidences expand now the role and actions of these lipid mediators from innate to adaptive immunity. In particular, SPMs have been shown to contribute to the control of chronic inflammation, and alterations in their production and/or function have been associated with the persistence of several pathological conditions, including autoimmunity, in human and experimental models. In this review, we focus on the impact of pro-resolving lipids on T cells through their ability to modulate T-cell responses. In particular, the effects of the different families of SPMs to restrain effector T-cell functions while promoting regulatory T cells will be reviewed, along with the underlying mechanisms. Furthermore, the emerging concept of SPMs as new biological markers for disease diagnostic and progression and as putative therapeutic tools to regulate the development and magnitude of inflammatory and autoimmune diseases is discussed.

Mini-Review: The Administration of Apoptotic Cells for Treating Rheumatoid Arthritis: Current Knowledge and Clinical Perspectives.

Rheumatoid arthritis (RA) is a chronic immune-mediated disease managed by conventional synthetic drugs, such as methotrexate (MTX), and targeted drugs including biological agents. Cell-based therapeutic approaches are currently developed in RA, mainly mesenchymal stroma cell-based approaches. Early-stage apoptotic cells possess direct and indirect anti-inflammatory properties. During the elimination of dying cells (a process called efferocytosis), specific mechanisms operate to control immune responses. There are compelling evidences in experimental models of arthritis indicating that apoptotic cell administration may benefit joint inflammation, and may even have therapeutic effects on arthritis. Additionally, it has been demonstrated that apoptotic cells could be administered with standard treatments of RA, such as MTX or TNF inhibitors (TNFi), given even a synergistic response with TNFi. Interestingly, apoptotic cell infusion has been successfully experienced to prevent acute graft-vs.-host disease after hematopoietic cell transplantation in patients with hematologic malignancies, with a good safety profile. In this mini-review, the apoptotic cell-based therapy development in arthritis is discussed, as well as its transfer in the short-term to an innovative treatment for patients with RA. The use of apoptotic cell-derived factors, including secretome or phosphatidylserine-containing liposomes, in RA are also discussed.

Pro-Resolving Factors Released by Macrophages After Efferocytosis Promote Mucosal Wound Healing in Inflammatory Bowel Disease.

Nonresolving inflammation is a critical driver of several chronic inflammatory diseases, including inflammatory bowel diseases (IBD). This unresolved inflammation may result from the persistence of an initiating stimulus or from the alteration of the resolution phase of inflammation. Elimination of apoptotic cells by macrophages (a process called efferocytosis) is a critical step in the resolution phase of inflammation. Efferocytosis participates in macrophage reprogramming and favors the release of numerous pro-resolving factors. These pro-resolving factors exert therapeutic effects in experimental autoimmune arthritis. Here, we propose to evaluate the efficacy of pro-resolving factors produced by macrophages after efferocytosis, a secretome called SuperMApo, in two IBD models, namely dextran sodium sulfate (DSS)-induced and T cell transfer-induced colitis. Reintroducing these pro-resolving factors was sufficient to decrease clinical, endoscopic and histological colitis scores in ongoing naive T cell-transfer-induced colitis and in DSS-induced colitis. Mouse primary fibroblasts isolated from the colon demonstrated enhanced healing properties in the presence of SuperMApo, as attested by their increased migratory, proliferative and contractive properties. This was confirmed by the use of human fibroblasts isolated from patients with IBD. Exposure of an intestinal epithelial cell (IEC) line to these pro-resolving factors increased their proliferative properties and IEC acquired the capacity to capture apoptotic cells. The improvement of wound healing properties induced by SuperMApo was confirmed in vivo in a biopsy forceps-wound colonic mucosa model. Further in vivo analysis in naive T cell transfer-induced colitis model demonstrated an improvement of intestinal barrier permeability after administration of SuperMApo, an intestinal cell proliferation and an increase of α-SMA expression by fibroblasts, as well as a reduction of the transcript coding for fibronectin (Fn1). Finally, we identified TGF-ß, IGF-I and VEGF among SuperMApo as necessary to favor mucosal healing and confirmed their role both in vitro (using neutralizing antibodies) and in vivo by depleting these factors from efferocytic macrophage secretome using antibody-coated microbeads. These growth factors only explained some of the beneficial effects induced by factors released by efferocytic macrophages. Overall, the administration of pro-resolving factors released by efferocytic macrophages limits intestinal inflammation and enhance tissue repair, which represents an innovative treatment of IBD.

Pro-Resolving Factor Administration Limits Cancer Progression by Enhancing Immune Response Against Cancer Cells.

Cancers are consequences of cellular dysfunction leading to an aberrant cellular multiplication and proliferation, subsequently yielding metastasis formation. Inflammatory reaction, with immune cell recruitment, is the main defense against precancerous lesions. However, an inflammatory environment also favors cancer cell progression, with cancer cell evasion from immune surveillance, leading to cancer development. Current therapeutic strategies enhance this natural immune response in order to restore immunosurveillance. The variety of these strategies is a predominant source of inflammatory mediators used by cancer cells to grow, differentiate, and migrate, therefore encouraging metastasis formation. For this reason, during cancer progression, limiting inflammation appears to be an innovative strategy to avoid the escape of cancer cells and potentially enhance the efficacy of antitumor therapies. Thus, this study aims to investigate the impact of administering pro-resolving factors (SuperMApo® drug candidate), which are inducers of inflammation resolution, in the framework of cancer treatment. We have observed that administering pro-resolving mediators issued from apoptotic cell efferocytosis by macrophages controlled peritoneal cancer progression by limiting cancer cell dissemination to the blood and mesenteric lymph nodes. This observation has been linked to an increase of macrophage mobilization in both peritoneal cavity and mesenteric lymph nodes. This control is associated to a restricted immunosuppressive myeloid cell circulation and to an IFN-γ-specific anti-tumor T-cell response. Altogether, these results suggest that administering proresolving factors could provide a new additional therapeutic alternative to control cancer progression.

Regulatory T-cell expansion and function do not account for the impaired alloreactivity of ex vivo-expanded T cells.

CD3- and CD28-activated T cells expanded for 12 days ex vivo to produce suicide gene-modified T cells are hyporesponsive to alloantigens. To investigate whether this impaired alloreactivity is a result of preferential expansion of regulatory T (Treg) cells, we compared peripheral blood mononuclear cells (PBMC) activated with CD3 and CD28 antibodies co-immobilized on beads and expanded for 12 days with interleukin (IL)-2 (Co(CD3/CD28) cells) to the respective unactivated PBMC in terms of proliferation, cytokine production, and expression of Treg markers [cytotoxic T-lymphocyte antigen 4 (CTLA4), glucocorticoid-induced tumour necrosis factor receptor (GITR) and forkhead box P3 (FoxP3)] after allostimulation. Alloreactive cells were identified by carboxyfluoresceine succinimidyl ester staining dilution. Alloreactive cells in Co(CD3/CD28) cells had a lower proliferative response and a lower potential for IL-2 and interferon-gamma secretion than did those in PBMC, demonstrating a functional impairment of alloreactive cells during ex vivo expansion. Expression of Treg markers transiently increased during ex vivo expansion and was unaffected by depletion of CD25(+) cells (containing Treg cells) before ex vivo PBMC expansion. Such prior CD25(+) depletion did not restore the alloreactivity of Co(CD3/CD28) cells. After allostimulation, expression of Treg markers was restricted to proliferative (alloreactive) cells among PBMC or Co(CD3/CD28) cells. Lastly, CD4(+) CD25(+) cells purified from Co(CD3/CD28) cells lacked suppressive activity when used as a third party, in contrast to CD4(+) CD25(+) cells purified from PBMC. In conclusion, the impaired alloreactivity of T cells expanded ex vivo is not a result of preferential Treg cell expansion and/or enhanced suppressive Treg activity.