Solenne Vigne

IL-36R ligands are potent regulators of dendritic and T cells

IL-36α (IL-1F6), IL-36β (IL-1F8), and IL-36γ (IL-1F9) are members of the IL-1 family of cytokines. These cytokines bind to IL-36R (IL-1Rrp2) and IL-1RAcP, activating similar intracellular signals as IL-1, whereas IL-36Ra (IL-1F5) acts as an IL-36R antagonist (IL-36Ra). In this study, we show that both murine bone marrow-derived dendritic cells (BMDCs) and CD4(+) T lymphocytes constitutively express IL-36R and respond to IL-36α, IL-36β, and IL-36γ. IL-36 induced the production of proinflammatory cytokines, including IL-12, IL-1β, IL-6, TNF-α, and IL-23 by BMDCs with a more potent stimulatory effect than that of other IL-1 cytokines. In addition, IL-36β enhanced the expression of CD80, CD86, and MHC class II by BMDCs. IL-36 also induced the production of IFN-γ, IL-4, and IL-17 by CD4(+) T cells and cultured splenocytes. These stimulatory effects were antagonized by IL-36Ra when used in 100- to 1000-fold molar excess. The immunization of mice with IL-36β significantly and specifically promoted Th1 responses. Our data thus indicate a critical role of IL-36R ligands in the interface between innate and adaptive immunity, leading to the stimulation of T helper responses.

IL-36 signaling amplifies Th1 responses by enhancing proliferation and Th1 polarization of naive CD4+ T cells

The interleukin-1 (IL-1) superfamily of cytokines comprises a set of pivotal mediators of inflammation. Among them, the action of IL-36 cytokines in immune responses has remained elusive. In a recent study, we demonstrated a direct effect of IL-36 on immune cells. Here we show that, among T cells, the IL-36 receptor is predominantly expressed on naive CD4(+) T cells and that IL-36 cytokines act directly on naive T cells by enhancing both cell proliferation and IL-2 secretion. IL-36β acts in synergy with IL-12 to promote Th1 polarization and IL-36 signaling is also involved in mediating Th1 immune responses to Bacillus Calmette-Guerin infection in vivo. Our findings point toward a critical function of IL-36 in the priming of Th1 cell responses in vitro, and in adaptive immunity in a model of mycobacterial infection in vivo.

Distinct expression of interleukin (IL)-36α, β and γ, their antagonist IL-36Ra and IL-38 in psoriasis, rheumatoid arthritis and Crohn's disease.

Interleukin (IL)‐36α, IL‐36β and IL‐36γ are expressed highly in skin and are involved in the pathogenesis of psoriasis, while the antagonists IL‐36Ra or IL‐38, another potential IL‐36 inhibitor, limit uncontrolled inflammation. The expression and role of IL‐36 cytokines in rheumatoid arthritis (RA) and Crohns disease (CD) is currently debated. Here, we observed that during imiquimod‐induced mouse skin inflammation and in human psoriasis, expression of IL‐36α, γ and IL‐36Ra, but not IL‐36β and IL‐38 mRNA, was induced and correlated with IL‐1β and T helper type 17 (Th17) cytokines (IL‐17A, IL‐22, IL‐23, CCL20). In mice with collagen‐induced arthritis and in the synovium of patients with RA, IL‐36α, β, γ, IL‐36Ra and IL‐38 were all elevated and correlated with IL‐1β, CCL3, CCL4 and macrophage colony‐stimulating factor (M‐CSF), but not with Th17 cytokines. In the colon of mice with dextran sulphate sodium‐induced colitis and in patients with CD, only IL‐36α, γ and IL‐38 were induced at relatively low levels and correlated with IL‐1β and IL‐17A. We suggest that only a minor subgroup of patients with RA (17–29%) or CD (25%) had an elevated IL‐36 agonists/antagonists ratio, versus 93% of patients with psoriasis. By immunohistochemistry, IL‐36 cytokines were produced by various cell types in skin, synovium and colonic mucosa such as keratinocytes, CD68+ macrophages, dendritic/Langerhans cells and CD79α+ plasma cells. In primary cultures of monocytes or inflammatory macrophages (M1), IL‐36β and IL‐36Ra were produced constitutively, but IL‐36α, γ and IL‐38 were produced after lipopolysaccharide stimulation. These distinct expression profiles may help to explain why only subgroups of RA and CD patients have a potentially elevated IL‐36 agonists/antagonists ratio.

The mouse interleukin (Il)33 gene is expressed in a cell type‐ and stimulus‐dependent manner from two alternative promoters

GenBank entries for mouse Il33 reveal the existence of two transcripts, Il33a and Il33b, with different 5′UTRs but coding for the same protein. We investigated expression of these transcripts in different mouse organs and cell types in basal and inflammatory conditions. Il33a and Il33b mRNAs start with different noncoding first exons, transcribed from different promoter regions, which both contain a consensus TATA‐like sequence. Constitutive Il33a mRNA expression was detected in mouse stomach, lung, spleen, and brain, whereas basal Il33b mRNA expression was observed only in the stomach. Expression of both transcripts increased after systemic LPS administration. In vitro, we observed high constitutive expression of Il33 transcripts in MEFs. Constitutive Il33a mRNA expression was observed also in BMDCs, where it was preferentially increased in response to poly(I:C), whereas LPS increased levels of Il33a and Il33b mRNA. In contrast, BMMs and Raw 264.7 cells did not express Il33 mRNA constitutively, and LPS stimulation selectively induced expression of Il33b mRNA in these cells. Our data indicate that the Il33 gene is expressed from two alternative promoters in the mouse and that the relative expression of Il33a and Il33b transcripts is cell type‐ and stimulus‐dependent.

Distinct Expression of IL‐36α, β, γ, their antagonist IL‐36Ra and IL‐38 in psoriasis, rheumatoid arthritis and Crohn's disease

Interleukin (IL)‐36α, IL‐36β and IL‐36γ are expressed highly in skin and are involved in the pathogenesis of psoriasis, while the antagonists IL‐36Ra or IL‐38, another potential IL‐36 inhibitor, limit uncontrolled inflammation. The expression and role of IL‐36 cytokines in rheumatoid arthritis (RA) and Crohns disease (CD) is currently debated. Here, we observed that during imiquimod‐induced mouse skin inflammation and in human psoriasis, expression of IL‐36α, γ and IL‐36Ra, but not IL‐36β and IL‐38 mRNA, was induced and correlated with IL‐1β and T helper type 17 (Th17) cytokines (IL‐17A, IL‐22, IL‐23, CCL20). In mice with collagen‐induced arthritis and in the synovium of patients with RA, IL‐36α, β, γ, IL‐36Ra and IL‐38 were all elevated and correlated with IL‐1β, CCL3, CCL4 and macrophage colony‐stimulating factor (M‐CSF), but not with Th17 cytokines. In the colon of mice with dextran sulphate sodium‐induced colitis and in patients with CD, only IL‐36α, γ and IL‐38 were induced at relatively low levels and correlated with IL‐1β and IL‐17A. We suggest that only a minor subgroup of patients with RA (17–29%) or CD (25%) had an elevated IL‐36 agonists/antagonists ratio, versus 93% of patients with psoriasis. By immunohistochemistry, IL‐36 cytokines were produced by various cell types in skin, synovium and colonic mucosa such as keratinocytes, CD68+ macrophages, dendritic/Langerhans cells and CD79α+ plasma cells. In primary cultures of monocytes or inflammatory macrophages (M1), IL‐36β and IL‐36Ra were produced constitutively, but IL‐36α, γ and IL‐38 were produced after lipopolysaccharide stimulation. These distinct expression profiles may help to explain why only subgroups of RA and CD patients have a potentially elevated IL‐36 agonists/antagonists ratio.

The severity of experimental arthritis is independent of IL-36 receptor signaling

IntroductionInterleukin (IL)-36 refers to three related IL-1 family cytokines, IL-36α, IL-36β, and IL-36γ, that bind to the IL-36 receptor (IL-36R). IL-36 exerts proinflammatory effects in skin and lung and stimulates T cell responses. In the present study, we examined the expression and function of IL-36R and its ligands in experimental arthritis.MethodsCollagen-induced arthritis (CIA), antigen-induced arthritis (AIA), and K/BxN serum transfer-induced arthritis were induced according to standard protocols. Messenger RNA levels for IL-36R and its ligands in the joints of mice with CIA were determined by RT-qPCR. Mice with CIA were injected with a blocking monoclonal anti-IL-36R, a blocking anti-IL-1RI, or their isotype-matched control antibodies at the time of arthritis onset. Anti-IL-36R or control antibodies were also injected at the time of AIA induction. Finally, IL-36R-deficient mice were examined in AIA and serum transfer-induced arthritis. The development and severity of arthritis were assessed by clinical and histological scoring.ResultsIL-36R, IL-36Ra and IL-36γ mRNA were detected in the joints of mice with CIA, but their levels did not correlate with arthritis severity. As opposed to anti-IL-1RI antibody treatment, the injection of an anti-IL-36R antibody was devoid of effect on the development and severity of CIA. The severity of joint inflammation and structural damage in AIA was also unaltered by anti-IL-36R antibody treatment. Finally, the severity of AIA and K/BxN serum transfer-induced arthritis was similar in IL-36R-deficient and wild-type mice.ConclusionsThe development and severity of experimental arthritis are independent of IL-36R signaling.

Mouse neutrophils express the decoy type 2 interleukin-1 receptor (IL-1R2) constitutively and in acute inflammatory conditions

The proinflammatory activities of IL‐1 are tightly controlled at different levels. IL‐1R2 acts as a decoy receptor and has been shown to regulate the biological effects of IL‐1 in vitro and in vivo. However, little is known about its natural expression in the mouse in physiologic and pathologic conditions. In this study, we examined IL‐1R2 mRNA and protein expression in isolated cells and tissues in response to different stimulatory conditions. Data obtained using ex vivo CD11b+Ly6G+ peripheral blood cells and in vitro‐differentiated CD11b+Ly6G+ BMG indicated that neutrophils are the major source of constitutively expressed IL‐1R2 in the mouse. The expression of IL‐1R2 on BMG and ex vivo Ly6G+ peripheral blood cells was highly up‐regulated by HC. IL‐1R2 pull‐down experiments showed that mouse rIL‐1β binds to BMG IL‐1R2, whereas binding of IL‐1Ra could not be detected. Furthermore, LPS treatment induced shedding of IL‐1R2 from the neutrophil membrane in vitro and in vivo, executed mainly by ADAM17. Finally, in in vivo models of inflammation, including thioglycolate‐induced acute peritonitis and acute lung injury, infiltrating Ly6G+ neutrophils, expressed IL‐1R2. Our data show that in the mouse, neutrophils mainly express the decoy receptor IL‐1R2 under naïve and inflammatory conditions. These data suggest that neutrophils may contribute to the resolution of acute inflammation.

Disease severity in K/BxN serum transfer-induced arthritis is not affected by IL-33 deficiency

IntroductionInterleukin (IL)-33 is a cytokine of the IL-1 family, which signals through the ST2 receptor. Previous work suggested implication of the IL-33/ST2 axis in the pathogenesis of human and mouse arthritis. Here, we directly investigated the role of endogenous IL-33 in K/BxN serum transfer-induced arthritis by using IL-33 knockout (KO) mice.MethodsArthritis was induced by injection of complete K/BxN serum or purified IgG. Disease severity was monitored by clinical and histological scoring.ResultsK/BxN serum transfer induced pronounced arthritis with similar incidence and severity in IL-33 KO and wild-type (WT) mice. In contrast, disease development was significantly reduced in ST2 KO mice. IL-33 expression in synovial tissue was comparable in arthritic WT and ST2 KO mice, and absent in IL-33 KO mice. Transfer of purified arthritogenic IgG instead of complete K/BxN serum also resulted in similar arthritis severity in IL-33 KO and WT mice, excluding a contribution of IL-33 contained in the serum of donor mice to explain this result. We investigated additional potential confounding factors, including purity of genetic background, but the mechanisms underlying reduced arthritis in ST2 KO mice remained unclear.ConclusionsThe data obtained with IL-33 KO mice indicate that endogenous IL-33 is not required for the development of joint inflammation in K/BxN serum transfer-induced arthritis. On the contrary, arthritis severity was reduced in ST2 KO mice. This observation might relate to IL-33 independent effects of ST2, and/or reveal the existence of confounding variables affecting the severity of joint inflammation in these KO strains.

Limited Contribution of IL-36 versus IL-1 and TNF Pathways in Host Response to Mycobacterial Infection

IL-36 cytokines are members of the IL-1 family of cytokines that stimulate dendritic cells and T cells leading to enhanced T helper 1 responses in vitro and in vivo; however, their role in host defense has not been fully addressed thus far. The objective of this study was to examine the role of IL-36R signaling in the control of mycobacterial infection, using models of systemic attenuated M. bovis BCG infection and virulent aerogenic M. tuberculosis infection. IL-36γ expression was increased in the lung of M. bovis BCG infected mice. However, IL-36R deficient mice infected with M. bovis BCG showed similar survival and control of the infection as compared to wild-type mice, although their lung pathology and CXCL1 response were transiently different. While highly susceptible TNF-α deficient mice succumbed with overwhelming M. tuberculosis infection, and IL-1RI deficient mice showed intermediate susceptibility, IL-36R-deficient mice controlled the infection, with bacterial burden, lung inflammation and pathology, similar to wild-type controls. Therefore, IL-36R signaling has only limited influence in the control of mycobacterial infection.

Severe Neutrophil-Dominated Inflammation and Enhanced Myelopoiesis in IL-33–Overexpressing CMV/IL33 Mice

IL-33 is a cytokine of the IL-1 family, which signals through the ST2 receptor. Previous studies emphasized a role for IL-33 in shaping innate and adaptive immune responses. IL-33 was also reported to modulate myelopoiesis and myeloid cell activity. In this article, we describe IL-33–overexpressing CMV/IL33 and LysM/IL33 mice, which display an inflammatory phenotype associated with growth retardation and paw swelling. The phenotype of CMV/IL33 mice is dependent on activation of the ST2 receptor and is characterized by extensive neutrophil infiltration into different organs, including the paws. Local or systemic levels of proinflammatory mediators such as IL-1β, Cxcl-1, G-CSF, and IL-6 are increased. CMV/IL-33 mice also suffer from anemia, thrombocytosis, and a marked dysregulation of myelopoiesis, leading to an important increase in myeloid cell production or accumulation in bone marrow (BM), spleen, and peripheral blood. Consistently, recombinant IL-33 induced proliferation of myeloid lineage cells in BM-derived granulocyte cultures, whereas IL-33 knockout mice exhibited minor deficiencies in spleen and BM myeloid cell populations. Our observations reveal a neutrophil-dominated inflammatory phenotype in IL-33–overexpressing CMV/IL33 and LysM/IL33 mice, and highlight important regulatory effects of IL-33 on myelopoiesis in vitro and in vivo, where excessive IL-33 signaling can translate into the occurrence of a myeloproliferative disorder.