Simon Blanchard

Interferon-γ reverses the immunosuppressive and protumoral properties and prevents the generation of human tumor-associated macrophages

Tumor‐associated macrophages (TAM) are M2d‐polarized cells (IL‐10high, IL‐12low, ILT3high, CD86low) that accumulate in tumor microenvironment. TAM inhibit antitumor T lymphocyte generation and function, contribute to tumor tolerance and are trophic for tumors. In this study, we investigated whether some immunological factors may reverse TAM immunosuppressive properties. Among 32 cytokines, we have identified IFNγ on its ability to switch immunosuppressive TAM into immunostimulatory cells. Upon IFNγ exposure, TAM purified from ovarian cancer ascites recover a M1 phenotype (IL‐10low, IL‐12high), express high levels of CD86 and low levels of ILT3, enhance the proliferation of CD4+ T lymphocytes and potentiate the cytotoxic properties of a MelanA‐specific CD8+ T cell clone. IFNγ‐treated TAM also secreted reduced levels of mediators promoting suppressive T cell accumulation (CCL18) and trophic for tumors (VEGF and MMP9). As TAM derive from the local differentiation of peripheral blood monocytes, we investigated whether IFNγ may also affect TAM generation. In the presence of ovarian ascites, IFNγ skewed monocyte differentiation from TAM‐like cells to M1‐polarized immunostimulatory macrophages. Together, these data show that IFNγ overcomes TAM‐induced immunosuppression by preventing TAM generation and functions. These data highlight that IFNγ used locally at the tumor site could potentiate the efficacy of antitumor immunotherapies based on the generation of effector T cells.

IL-34 Induces the Differentiation of Human Monocytes into Immunosuppressive Macrophages. Antagonistic Effects of GM-CSF and IFNγ

IL-34 is a recently identified cytokine that signals via the M-CSF receptor and promotes monocyte survival. Depending on the environment, monocytes can differentiate into macrophages (Mφ) or dendritic cells (DC). A wide spectrum of Mφ and DC subsets, with distinct phenotypes and functions, has been described. To date, the phenotype of monocytes exposed to IL-34 remains unexplored. We report here that IL-34 induces the differentiation of monocytes into CD14high CD163high CD1a− Mφ (IL-34-Mφ). Upon LPS stimulation, IL-34-Mφ exhibit an IL-10high IL-12low M2 profile and express low levels of the costimulatory molecules CD80 and CD86. IL-34-Mφ exhibit poor T cell costimulatory properties, and have potent immunosuppressive properties (decrease of TCR-stimulated T cell proliferation). For all the parameters analyzed, IL-34-Mφ are phenotypically and functionally similar to M-CSF-Mφ. IL-34 appears as efficient as M-CSF in inducing the generation of immunosuppressive Mφ. Moreover, the generation of IL-34-Mφ is mediated through the M-CSF receptor, is independent of endogenous M-CSF consumption and is potentiated by IL-6. In an attempt to identify strategies to prevent a deleterious M2 cell accumulation in some pathological situations, we observed that IFNγ and GM-CSF prevent the generation of immunosuppressive Mφ induced by IL-34. IFNγ also switches established IL-34-Mφ into immunostimulatory Mφ. In conclusion, we demonstrate that IL-34 drives the differentiation of monocytes into immunosuppressive M2, in a manner similar to M-CSF, and that IFNγ and GM-CSF prevent this effect.

The Tachykinins Substance P and Hemokinin-1 Favor the Generation of Human Memory Th17 Cells by Inducing IL-1β, IL-23, and TNF-Like 1A Expression by Monocytes

The nervous system influences immune responses through the release of neural factors such as neuropeptides. Among them, the tachykinin substance P (SP) signals via the neurokinin 1 receptor (NK-1R), which is expressed by various immune cells. We thereby analyzed in this paper whether tachykinins may participate in human CD4+ Th cell polarization. We report that SP and hemokinin-1 (HK-1) upregulate IL-17A and IFN-γ production by human memory CD4+ T cells without affecting IL-4 and IL-10 production. SP and HK-1 switch non–Th17-committed CD4+ memory T cells into bona fide Th17 cells and Th1/Th17 cells. In contrast, SP and HK-1 do not modulate the polarization of naive CD4+ T cells. SP- and HK-1–induced Th17 cell generation is mediated through NK-1R and requires the presence of monocytes. SP and HK-1 trigger IL-1β, IL-6, and TNF-α production, upregulate IL-23 production, and enhance TNF-like 1A expression on monocyte surface. Neutralization experiments demonstrated that IL-1β, IL-23, and TNF-like 1A are involved in the SP- and HK-1–induced Th17 cell. The other members of the tachykinin family, neurokinins A and B, have no effect on the differentiation of naive and memory T cells. These results thereby show that SP and HK-1 are novel Th17 cell-inducing factors that may act locally on memory T cells to amplify inflammatory responses.

IL-34 and macrophage colony-stimulating factor are overexpressed in hepatitis C virus fibrosis and induce profibrotic macrophages that promote collagen synthesis by hepatic stellate cells

Chronic hepatitis C virus (HCV) infection is characterized by progressive hepatic fibrosis, a process dependent on monocyte recruitment and accumulation into the liver. The mediators expressed in chronically injured liver that control the differentiation of human monocytes into profibrotic macrophages (Mφ) remain poorly defined. We report that chronically HCV‐infected patients with high fibrosis stages have higher serum levels of macrophage colony‐stimulating factor (M‐CSF) and interleukin (IL)−34 than HCV‐infected patients with lower fibrosis stages and healthy subjects. Immunohistochemistry reveals an intense expression of IL‐34 and M‐CSF by hepatocytes around liver lesions. In addition, HCV infection and inflammatory cytokines enhance the in vitro production of IL‐34 and M‐CSF by hepatocytes. We next analyzed the acquisition of profibrotic properties by Mφ generated with M‐CSF (M‐CSF‐Mφ) or IL‐34 (IL‐34‐Mφ). M‐CSF and IL‐34 up‐regulate the expression, by differentiating monocytes, of chemokine (C‐C motif) ligand (CCL)2, CCL4, C‐C chemokine receptor (CCR)1, and CCR5, which are involved in monocyte recruitment/Mφ accumulation in liver lesions. M‐CSF‐Mφ and IL‐34‐Mφ also express the hepatic stellate cell (HSC) activators, platelet‐derived growth factor, transforming growth factor beta, and galectin‐3. IL‐34‐Mφ and M‐CSF‐Mφ induce type I collagen synthesis by HSCs, the main collagen‐producing cells in liver fibrosis. IL‐13, whose expression correlates with the fibrosis stage in HCV‐infected patients, decreases the expression of the collagenase, matrix metalloproteinase 1, by IL‐34‐Mφ and M‐CSF‐Mφ, thereby enhancing collagen synthesis. By inhibiting the production of interferon‐gamma (IFN‐γ) by activated natural killer cells, IL‐34‐Mφ and M‐CSF‐Mφ prevent the IFN‐γ‐induced killing of HSCs. Conclusion: These results identify M‐CSF and IL‐34 as potent profibrotic factors in HCV liver fibrosis. (Hepatology 2014;60:1878–1889)

Scavenger Receptors in Human Airway Epithelial Cells: Role in Response to Double-Stranded RNA

Scavenger receptors and Toll-like receptors (TLRs) cooperate in response to danger signals to adjust the host immune response. The TLR3 agonist double stranded (ds)RNA is an efficient activator of innate signalling in bronchial epithelial cells. In this study, we aimed at defining the role played by scavenger receptors expressed by bronchial epithelial cells in the control of the innate response to dsRNA both in vitro and in vivo. Expression of several scavenger receptor involved in pathogen recognition was first evaluated in human bronchial epithelial cells in steady-state and inflammatory conditions. Their implication in the uptake of dsRNA and the subsequent cell activation was evaluated in vitro by competition with ligand of scavenger receptors including maleylated ovalbumin and by RNA silencing. The capacity of maleylated ovalbumin to modulate lung inflammation induced by dsRNA was also investigated in mice. Exposure to tumor necrosis factor-α increased expression of the scavenger receptors LOX-1 and CXCL16 and the capacity to internalize maleylated ovalbumin, whereas activation by TLR ligands did not. In contrast, the expression of SR-B1 was not modulated in these conditions. Interestingly, supplementation with maleylated ovalbumin limited dsRNA uptake and inhibited subsequent activation of bronchial epithelial cells. RNA silencing of LOX-1 and SR-B1 strongly blocked the dsRNA-induced cytokine production. Finally, administration of maleylated ovalbumin in mice inhibited the dsRNA-induced infiltration and activation of inflammatory cells in bronchoalveolar spaces and lung draining lymph nodes. Together, our data characterize the function of SR-B1 and LOX-1 in bronchial epithelial cells and their implication in dsRNA-induced responses, a finding that might be relevant during respiratory viral infections.

PolyI:C plus IL‐2 or IL‐12 induce IFN‐γ production by human NK cells via autocrine IFN‐β

NK lymphocytes and type I IFN (IFN‐α/β) are major actors of the innate anti‐viral response that also influence adaptive immune responses. We evaluated type I IFN production by human NK cells in response to polyI:C, a potent type I IFN‐inducing TLR3 agonist. PolyI:C plus IL‐2/IL‐12 induced IFN‐β (but not IFN‐α) mRNA expression and protein production by highly pure human NK cells and by the human NK cell line NK92. Neutralizing anti‐IFNAR1 or anti‐IFN‐β Ab prevented the production of IFN‐γ induced by polyI:C plus IL‐2/IL‐12. Similarly, IFN‐γ production induced by polyI:C plus IL‐12 was reduced in NK cells isolated from IFNAR1−/− compared with WT mice. The ability of polyI:C plus IL‐12 to induce IFN‐γ production was related to an increase of TLR3, Mda5 and IFNAR expression and by an increase of STAT1 and STAT4 phosphorylation. Collectively, these data demonstrate that NK cells, in response to polyI:C plus IL‐2/IL‐12, produce IFN‐β that induce, in an autocrine manner, the production of IFN‐γ and thereby highlight that NK cells may control the outcome of protective or injurious immune responses through type I IFN secretion.

The scavenger receptors SRA-1 and SREC-I cooperate with TLR2 in the recognition of the hepatitis C virus non-structural protein 3 by dendritic cells

BACKGROUNDS & AIMS The hepatitis C virus NS3 protein is taken up by myeloid cells in a TLR2-independent manner and activates myeloid cells via TLR2. This study aimed to identify the endocytic receptor(s) involved in the uptake of NS3 by myeloid cells and its relation with TLR2. METHODS Inhibitors and transfected cells were used to identify the nature of the NS3-binding receptors expressed by myeloid cells. The cooperation between scavenger receptors (SRs) and TLR2 in the NS3-mediated activation of myeloid cells was evaluated using inhibitors, cells from TLR2(-/-) mice, and confocal microscopy. The involvement of SRs in NS3 cross-presentation was evaluated in vitro using an NS3-specific human T-cell clone. RESULTS We observed that SRs are the main binding structures for NS3 on myeloid cells and identified the SRs SRA-1 and SREC-I as endocytic receptors for NS3. Moreover, both SRs and TLR2 cooperate in NS3-induced myeloid cell activation. CONCLUSION This study highlights a central role for SRs in NS3 uptake and cross-presentation, and demonstrates a tightly orchestrated cooperation between signalling and endocytic innate receptors in NS3 recognition.

Detection of anti-PTX3 autoantibodies in systemic lupus erythematosus

levels amongst those with doctor-diagnosed arthritis. However, in the majority of people with arthritis, fish oil was not taken at analgesic/anti-inflammatory doses. This is especially important in patients with RA as symptomatic benefits are seen with doses between 2.6 g and 7.1 g per day, with no effect seen at 1 g per day [9], the most common dose seen in this study. However, the effectiveness of fish oil in OA has not yet been the subject of a randomized controlled trial. The pattern of usage we observed suggested that GPs or other therapists were recommending usage of fish oil in participants with doctor-diagnosed arthritis. Fish oil has been demonstrated to reduce coronary artery disease events and reduce triglyceride levels [10]. However, in this populationbased study, those at highest risk of CVD events (those with existing CVD, uncontrolled hypertension, current smokers and NSAID users) were less likely to be using fish oil. The symptomatic benefits of fish oil in arthritis have been well known for some time; however, most users are using suboptimal doses. In addition, less than one in five participants with RA were using fish oil, a proven intervention that is safe and inexpensive with favourable collateral benefits on cardiovascular risk and can reduce reliance on NSAIDs.

IL-26 is overexpressed in chronically HCV-infected patients and enhances TRAIL-mediated cytotoxicity and interferon production by human NK cells

Objective Interleukin-26 (IL-26) is a member of the IL-10 cytokine family, first discovered based on its peculiar expression by virus-transformed T cells. IL-26 is overexpressed in chronic inflammation (rheumatoid arthritis and Crohn’s disease) and induces proinflammatory cytokines by myeloid cells and some epithelial cells. We thus investigated the expression and potential role of IL-26 in chronic HCV infection, a pathology associated with chronic inflammation. Design IL-26 was quantified in a cohort of chronically HCV-infected patients, naive of treatment and its expression in the liver biopsies investigated by immunohistochemistry. We also analysed the ability of IL-26 to modulate the activity of natural killer (NK) cells, which control HCV infection. Results The serum levels of IL-26 are enhanced in chronically HCV-infected patients, mainly in those with severe liver inflammation. Immunohistochemistry reveals an intense IL-26 staining in liver lesions, mainly in infiltrating CD3+ cells. We also show that NK cells from healthy subjects and from HCV-infected patients are sensitive to IL-26. IL-26 upregulates membrane tumour necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) expression on CD16− CD56bright NK cells, enabling them to kill HCV-infected hepatoma cells, with the same efficacy as interferon (IFN)-α-treated NK cells. IL-26 also induces the expression of the antiviral cytokines IFN-β and IFN-γ, and of the proinflammatory cytokines IL-1β and TNF-α by NK cells. Conclusions This study highlights IL-26 as a new player in the inflammatory and antiviral immune responses associated with chronic HCV infection.

IL-34- and M-CSF-induced macrophages switch memory T cells into Th17 cells via membrane IL-1α.

Macrophages orchestrate the immune response via the polarization of CD4+ T helper (Th) cells. Different subsets of macrophages with distinct phenotypes, and sometimes opposite functions, have been described. M‐CSF and IL‐34 induce the differentiation of monocytes into IL‐10high IL‐12low immunoregulatory macrophages, which are similar to tumor‐associated macrophages (TAMs) in ovarian cancer. In this study, we evaluated the capacity of human macrophages induced in the presence of M‐CSF (M‐CSF macrophages) or IL‐34 (IL‐34 macrophages) and ovarian cancer TAMs to modulate the phenotype of human CD4+ T cells. Taken together, our results show that M‐CSF‐, IL‐34 macrophages, and TAMs switch non‐Th17 committed memory CD4+ T cells into conventional CCR4+ CCR6+ CD161+ Th17 cells, expressing or not IFN‐gamma. Contrary, the pro‐inflammatory GM‐CSF macrophages promote Th1 cells. The polarization of memory T cells into Th17 cells is mediated via membrane IL‐1α (mIL‐1α), which is constitutively expressed by M‐CSF‐, IL‐34 macrophages, and TAMs. This study elucidates a new mechanism that allows macrophages to maintain locally restrained and smoldering inflammation, which is required in angiogenesis and metastasis.