The composition and signaling of the IL-35 receptor are unconventional.

Interleukin 35 (IL-35) belongs to the IL-12 family of heterodimeric cytokines but has a distinct functional profile. IL-35 suppresses T cell proliferation and converts naive T cells into IL-35-producing induced regulatory T cells (iTr35 cells). Here we found that IL-35 signaled through a unique heterodimer of receptor chains IL-12Rß2 and gp130 or homodimers of each chain. Conventional T cells were sensitive to IL-35-mediated suppression in the absence of one receptor chain but not both receptor chains, whereas signaling through both chains was required for IL-35 expression and conversion into iTr35 cells. Signaling through the IL-35 receptor required the transcription factors STAT1 and STAT4, which formed a unique heterodimer that bound to distinct sites in the promoters of the genes encoding the IL-12 subunits p35 and Ebi3. This unconventional mode of signaling, distinct from that of other members of the IL-12 family, may broaden the spectrum and specificity of IL-35-mediated suppression.

Mesothelin CAR-T cells expressing tumor-targeted immunocytokine IL-12 yield durable efficacy and fewer side effects.

Chimeric antigen receptor (CAR)-modified T cell therapy has achieved remarkable efficacy in treating hematological malignancies, but it confronts many challenges in treating solid tumors, such as the immunosuppressive microenvironment of the solid tumors. These factors reduce the antitumor activity of CAR-T cells in clinical trials. Therefore, we used the immunocytokine interleukin-12 (IL-12) to enhance the efficacy of CAR-T cell therapy. In this study, we engineered CAR-IL12R54 T cells that targeted mesothelin (MSLN) and secreted a single-chain IL-12 fused to a scFv fragment R54 that recognized a different epitope on mesothelin. The evaluation of the anti-tumor activity of the CAR-IL12R54 T cells alone or in combination with anti-PD-1 antibody in vitro and in vivo was followed by the exploration of the functional mechanism by which the immunocytokine IL-12 enhanced the antitumor activity. CAR-IL12R54 T cells had potency to lyse mesothelin positive tumor cells in vitro. In vivo studies demonstrated that CAR-IL12R54 T cells were effective in controlling the growth of established tumors in a xenograft mouse model with fewer side effects than CAR-T cells that secreted naked IL-12. Furthermore, combination of PD-1 blockade antibody with CAR-IL12R54 T cells elicited durable anti-tumor responses. Mechanistic studies showed that IL12R54 enhanced Interferon-γ (IFN-γ) production and dampened the activity of regulatory T cells (Tregs). IL12R54 also upregulated CXCR6 expression in the T cells through the NF-κB pathway, which facilitated T cell infiltration and persistence in the tumor tissues. In summary, the studies provide a good therapeutic option for the clinical treatment of solid tumors.

IL-12 p40 monomer is different from other IL-12 family members to selectively inhibit IL-12Rß1 internalization and suppress EAE.

Multiple sclerosis (MS) is the most common human demyelinating disease of the central nervous system. The IL-12 family of cytokines has four members, which are IL-12 (p40:p35), IL-23 (p40:p19), the p40 monomer (p40), and the p40 homodimer (p402). Since all four members contain p40 in different forms, it is important to use a specific monoclonal antibody (mAb) to characterize these molecules. Here, by using such mAbs, we describe selective loss of p40 in serum of MS patients as compared to healthy controls. Similarly, we also observed decrease in p40 and increase in IL-12, IL-23, and p402 in serum of mice with experimental autoimmune encephalomyelitis (EAE), an animal model of MS, as compared to control mice. Interestingly, weekly supplementation of mouse and human recombinant p40 ameliorated clinical symptoms and disease progression of EAE. On the other hand, IL-12, IL-23, and p402 did not exhibit such inhibitory effect. In addition to EAE, p40 also suppressed collagen-induced arthritis in mice. Using IL-12Rß1-/-, IL-12Rß2-/-, and IL-12Rß1+/-/IL-12Rß2-/- mice, we observed that p40 required IL-12Rß1, but not IL-12Rß2, to suppress EAE. Interestingly, p40 arrested IL-12-, IL-23-, or p402-mediated internalization of IL-12Rß1, but neither IL-12Rß2 nor IL-23R, protected regulatory T cells, and suppressed Th1 and Th17 biasness. These studies identify p40 as an anti-autoimmune cytokine with a biological role different from IL-12, IL-23, and p402 in which it attenuates autoimmune signaling via suppression of IL-12Rß1 internalization, which may be beneficial in patients with MS and other autoimmune disorders.

Production of IL-17A at Innate Immune Phase Leads to Decreased Th1 Immune Response and Attenuated Host Defense against Infection with Cryptococcus deneoformans.

IL-17A is a proinflammatory cytokine produced by many types of innate immune cells and Th17 cells and is involved in the elimination of extracellularly growing microorganisms, yet the role of this cytokine in the host defense against intracellularly growing microorganisms is not well known. Cryptococcus deneoformans is an opportunistic intracellular growth fungal pathogen that frequently causes fatal meningoencephalitis in patients with impaired immune responses. In the current study, we analyzed the role of IL-17A in the host defense against C. deneoformans infection. IL-17A was quickly produced by γδT cells at an innate immune phase in infected lungs. In IL-17A gene-disrupted mice, clearance of this fungal pathogen and the host immune response mediated by Th1 cells were significantly accelerated in infected lungs compared with wild-type mice. Similarly, killing of this fungus and production of inducible NO synthase and TNF-α were significantly enhanced in IL-17A gene-disrupted mice. In addition, elimination of this fungal pathogen, Th1 response, and expression of IL-12Rß2 and IFN-γ in NK and NKT cells were significantly suppressed by treatment with rIL-17A. The production of IL-12p40 and TNF-α from bone marrow-derived dendritic cells stimulated with C. deneoformans was significantly suppressed by rIL-17A. In addition, rIL-17A attenuated Th1 cell differentiation in splenocytes from transgenic mice highly expressing TCR for mannoprotein 98, a cryptococcal Ag, upon stimulation with recombinant mannoprotein 98. These data suggest that IL-17A may be involved in the negative regulation of the local host defense against C. deneoformans infection through suppression of the Th1 response.

Increased Interleukin-35 suppresses peripheral CD14+ monocytes function in patients with Kawasaki disease.

BACKGROUND: Interleukin-35 (IL-35) is a newly identified IL-12 cytokine family member, which regulates the activity of immune cells in infectious diseases and autoimmune disorders. However, the regulatory function of IL-35 in Kawasaki disease is not well elucidated. METHODS: Thirty-three patients with Kawasaki disease and seventeen healthy controls were studied. Peripheral IL-35 concentration was measured by enzyme linked immunosorbent assay. CD14+ monocytes were purified, and mRNA expression of IL-35 receptor (IL-12Rß2 and gp130) was semi-quantified by real-time polymerase chain reaction. CD14+ monocytes were stimulated with recombinant IL-35. The modulatory role of IL-35 treated CD14+ monocytes to naïve CD4+ T cell activation was investigated by flow cytometry. The influence of IL-35 to cytotoxicity of CD14+ monocytes was assessed by measuring target cell death, cytokine and granzyme secretion. RESULTS: Plasma IL-35 concentration was elevated in patients with Kawasaki disease. There was no significant differences of either IL-12Rß2 or gp130 mRNA expression in CD14+ monocytes between Kawasaki disease patients and controls. IL-35 suppressed CD14+ monocytes induced naïve CD4+ T cell activation in Kawasaki disease, and this process required direct cell-to-cell contact. IL-35 also inhibited tumor necrosis factor-α and granzyme B secretion by CD14+ monocytes from patients with Kawasaki disease, however, only granzyme B was responsible for the cytotoxicity of CD14+ monocytes. CONCLUSIONS: IL-35 played an important immunosuppressive role to CD14+ monocytes function in Kawasaki disease.

The monogenic basis of human tuberculosis.

The pathogenesis of tuberculosis (TB) remains poorly understood, as no more than 5-10% of individuals infected with Mycobacterium tuberculosis go on developing clinical disease. The contribution of human genetics to TB pathogenesis has been amply documented by means of classic genetics since the turn of the twentieth century. Over the last 20 years, following-up on the study of Mendelian susceptibility to mycobacterial disease (MSMD), monogenic disorders have been found to underlie TB in some patients. Rare inborn errors of immunity, such as autosomal recessive, complete IL-12Rß1 and TYK2 deficiencies, impairing the IL-12- and IL-23-dependent induction of IFN-γ, were initially identified in a few patients. More recently, homozygosity for a common variant of TYK2 (P1104A) that selectively disrupts cellular responses to IL-23 was found in two cohorts of TB patients. It shows high penetrance in areas endemic for TB and appears to be responsible for about 1% of TB cases in populations of European descent. Both rare and common genetic etiologies of TB affect IFN-γ immunity, providing a rationale for novel preventive and therapeutic approaches for TB control, including the use of recombinant IFN-γ.

JNK2 modulates the CD1d-dependent and -independent activation of iNKT cells.

Invariant natural killer T (iNKT) cells play critical roles in autoimmune, anti-tumor, and anti-microbial immune responses, and are activated by glycolipids presented by the MHC class I-like molecule, CD1d. How the activation of signaling pathways impacts antigen (Ag)-dependent iNKT cell activation is not well-known. In the current study, we found that the MAPK JNK2 not only negatively regulates CD1d-mediated Ag presentation in APCs, but also contributes to CD1d-independent iNKT cell activation. A deficiency in the JNK2 (but not JNK1) isoform enhanced Ag presentation by CD1d. Using a vaccinia virus (VV) infection model known to cause a loss in iNKT cells in a CD1d-independent, but IL-12-dependent manner, we found the virus-induced loss of iNKT cells in JNK2 KO mice was substantially lower than that observed in JNK1 KO or wild-type (WT) mice. Importantly, compared to WT mice, JNK2 KO mouse iNKT cells were found to express less surface IL-12 receptors. As with a VV infection, an IL-12 injection also resulted in a smaller decrease in JNK2 KO iNKT cells as compared to WT mice. Overall, our work strongly suggests JNK2 is a negative regulator of CD1d-mediated Ag presentation and contributes to IL-12-induced iNKT cell activation and loss during viral infections.

Behçet's disease: An immunogenetic perspective.

Behçet's disease (BD) is a chronic and rare multisystemic disorder defined by autoimmunity and inflammatory characteristics, manifested by ocular lesions, recurrent genital and oral ulcers, skin symptoms and arthritis as well as neurological, intestinal, and vascular involvement. Despite the unknown cause of BD, there is some strong documentation for immunological, genetic, environmental, and infectious factors playing a role in the pathogenesis of BD. While the nature of the genetic variants remains unidentified, many genetic risk factors are considered to contribute to BD susceptibility. Along with human leukocyte antigen gene encoding B*51 (HLA-B*51) and areas including the major histocompatibility complex class I, genome-wide association studies have recognized numerous other BD susceptibility genes including those encoding interleukin (IL)-10, IL-12 receptor ß 2 (IL-12RB2), IL-23 receptor (IL-23R), C-C chemokine receptor 1 gene, signal transducer and activator of transcription 4 (STAT4), endoplasmic reticulum aminopeptidase (ERAP1), and genes encoding killer cell lectin-like receptor family members (KLRC4-KLRK1). It is believed that BD could be considered as a disorder lying in between autoimmune and autoinflammatory syndromes. The positive responses to classical immunosuppressive agents like azathioprine and cyclosporine and involvement of autoantigens in the initiation of the disorder are the main BD features that reflect the autoimmune nature of the disorder. In this review, we address recent findings on the role of common cytokines, antibodies and immunogenetic factors in BD.

Interleukin-12 receptor ß2 from grass carp: Molecular characterization and its involvement in Aeromonas hydrophila-induced intestinal inflammation.

Interleukin-12 receptor ß2 (IL-12Rß2) is a signaling subunit of heterodimeric receptors for IL-12 and IL-35. It plays important regulatory functions in the development of Th1 cells and in the expression of inflammatory cytokines in mammals and other higher vertebrates. However, little is known about IL-12Rß2 in teleost fish. In this work, we have cloned and characterized IL-12Rß2 from grass carp (Ctenopharyngodon idella). The full-length cDNA of grass carp IL-12Rß2 is 2875 bp, which encodes a mature protein with 741 amino acids. This mature protein contains three fibronectin type III domains, a transmembrane helix, and CXW and WSXWS-like motifs that are characteristic of the type I cytokine receptor family. Phylogenetic analysis revealed that cyprinid fish IL-12Rß2 formed a single branch, clearly separated from those of other vertebrates. We expressed and purified a recombinant grass carp IL-12Rß2 protein containing major antigenic regions, which was used to raise a polyclonal antibody. The specificity of the antibody was assessed by Western blotting analysis of whole cell lysates from Escherichia coli cells expressing the recombinant IL-12Rß2, grass carp intestinal intraepithelial lymphocytes, and cultured C. idella kidney cells. To explore the potential regulatory role of IL-12Rß2 in inflammation, we generated an intestinal inflammation model by anal intubation of fish with Aeromonas hydrophila. Immunohistochemical staining of the inflamed intestines revealed that IL-12Rß2 expression is consistent with inflammatory cell recruitment during intestinal inflammation. Real-time quantitative PCR revealed that IL-12Rß2 is widely expressed in normal tissues and is up-regulated in most tissues after infecting with A. hydrophila. We found that IL-12Rß2, IL-12p35, and interferon-γ were expressed in similar patterns in the intestines during inflammation. Taken together, our results suggest that IL-12Rß2 is involved in the regulation of intestinal inflammation.

H3K27 Methylation Dynamics during CD4 T Cell Activation: Regulation of JAK/STAT and IL12RB2 Expression by JMJD3.

The changes to the epigenetic landscape in response to Ag during CD4 T cell activation have not been well characterized. Although CD4 T cell subsets have been mapped globally for numerous epigenetic marks, little has been done to study their dynamics early after activation. We have studied changes to promoter H3K27me3 during activation of human naive and memory CD4 T cells. Our results show that these changes occur relatively early (1 d) after activation of naive and memory cells and that demethylation is the predominant change to H3K27me3 at this time point, reinforcing high expression of target genes. Additionally, inhibition of the H3K27 demethylase JMJD3 in naive CD4 T cells demonstrates how critically important molecules required for T cell differentiation, such as JAK2 and IL12RB2, are regulated by H3K27me3. Our results show that H3K27me3 is a dynamic and important epigenetic modification during CD4 T cell activation and that JMJD3-driven H3K27 demethylation is critical for CD4 T cell function.

Susceptibility to mycobacterial disease due to mutations in IL-12Rß1 in three Iranian patients.

In the last decade, autosomal recessive interleukin-12 receptor ß1 (IL-12Rß1) deficiency, the most common cause of Mendelian susceptibility to mycobacterial disease (MSMD), has been diagnosed in a few children and adults with severe tuberculosis in Iran. Here, we report three cases referred to the Immunology, Asthma and Allergy ward at the National Research Institute of Tuberculosis and Lung Diseases (NRITLD) at Masih Daneshvari Hospital from 2012 to 2017 with Mycobacterium tuberculosis and non-tuberculous mycobacteria infections due to defects in IL-12Rß1 but with different clinical manifestations. All three were homozygous for either an IL-12Rß1 missense or nonsense mutation that caused the IL-12Rß1 protein not to be expressed on the cell membrane and completely abolished the cellular response to recombinant IL-12. Our findings suggest that the presence of IL-12Rß1 deficiency should be determined in children with mycobacterial infections at least in countries with a high prevalence of parental consanguinity and in areas endemic for TB like Iran.

Sequential polarization and imprinting of type 1 T helper lymphocytes by interferon-gamma and interleukin-12.

Differentiation of naive T lymphocytes into type I T helper (Th1) cells requires interferon-gamma and interleukin-12. It is puzzling that interferon-gamma induces the Th1 transcription factor T-bet, whereas interleukin-12 mediates Th1 cell lineage differentiation. We use mathematical modeling to analyze the expression kinetics of T-bet, interferon-gamma, and the IL-12 receptor beta2 chain (IL-12Rbeta2) during Th1 cell differentiation, in the presence or absence of interleukin-12 or interferon-gamma signaling. We show that interferon-gamma induced initial T-bet expression, whereas IL-12Rbeta2 was repressed by T cell receptor (TCR) signaling. The termination of TCR signaling permitted upregulation of IL-12Rbeta2 by T-bet and interleukin-12 signaling that maintained T-bet expression. This late expression of T-bet, accompanied by the upregulation of the transcription factors Runx3 and Hlx, was required to imprint the Th cell for interferon-gamma re-expression. Thus initial polarization and subsequent imprinting of Th1 cells are mediated by interlinked, sequentially acting positive feedback loops of TCR-interferon-gamma-Stat1-T-bet and interleukin-12-Stat4-T-bet signaling.

Gain-of-function STAT1 mutations are associated with intracranial aneurysms.

Chronic mucocutaneous candidiasis, characterized by persistent or recurrent fungal infections, represents the clinical hallmark in gain-of-function (GOF) signal transducer and activator of transcription 1 (STAT1) mutation carriers. Several cases of intracranial aneurysms have been reported in patients with GOF STAT1 mutation but the paucity of reported cases likely suggested this association still as serendipity. In order to endorse this association, we link the development of intracranial aneurysms with STAT1 GOF mutation by presenting the two different cases of a patient and her mother, and demonstrate upregulated phosphorylated STAT4 and IL-12 receptor ß1 upon stimulation in patient's blood cells. We also detected increased transforming growth factor (TGF)-ß type 2 receptor expression, particularly in CD14+ cells, and a slightly higher phosphorylation rate of SMAD3. In addition, the mother of the patient developed disseminated bacille Calmette-Guérin disease after vaccination, speculating that GOF STAT1 mutations may confer a predisposition to weakly virulent mycobacteria.

Advantage of tacrolimus/mycophenolate mofetil regimen for cytotoxic T cell-mediated defence and its inhibition by additive steroid administration in high-risk liver transplant recipients.

Our previous work revealed that the recipients with the highest pre-existing numbers of CD8(+) effector T cells (TE ) [hyperparathyroidism (HPT)E recipients] occupied approximately 30% of adult transplant recipients performed in our hospital. HPTE recipients demonstrated very poor clinical outcome compared with the remaining 70% of recipients with the lowest pre-existing TE (LPTE recipient). This study aimed to clarify the best combined immunosuppressive regimen related to function of cytotoxic T lymphocytes (CTLs) for HPTE recipients. Eighty-one HPTE recipients were classified into three types, according to the immunosuppressive regimens: type 1, tacrolimus (Tac)/glucocorticoid (GC); type 2, Tac/mycophenolate mofetil (MMF)/GC; and type 3, Tac/MMF. Frequencies of severe infection, rejection and hospital death were the highest in types 1 and 2, whereas the lowest occurred in type 3. The survival rate in type 3 was the highest (100%) during follow-up until post-operative day 2000. Regarding the immunological mechanism, in type 1 TE perforin and interferon (IFN)-γ were generated through the self-renewal of CD8(+) central memory T cells (TCM ), but decreased in the early post-transplant period due to marked down-regulation of interleukin (IL)-12 receptor beta-1 of TCM. In type 2, the self-renewal TCM did not develop, and the effector function could not be increased. In type 3, in contrast, the effectors and cytotoxicity were correlated inversely with IL-12Rß1(+) TCM levels, and increased at the highest level around the pre-transplant levels of IL-12Rß1(+) TCM . However, the immunological advantage of Tac/MMF therapy was inhibited strongly by additive steroid administration.

Toxoplasma gondii profilin promotes recruitment of Ly6Chi CCR2+ inflammatory monocytes that can confer resistance to bacterial infection.

Ly6C+ inflammatory monocytes are essential to host defense against Toxoplasma gondii, Listeria monocytogenes and other infections. During T. gondii infection impaired inflammatory monocyte emigration results in severe inflammation and failure to control parasite replication. However, the T. gondii factors that elicit these monocytes are unknown. Early studies from the Remington laboratory showed that mice with a chronic T. gondii infection survive lethal co-infections with unrelated pathogens, including L. monocytogenes, but a mechanistic analysis was not performed. Here we report that this enhanced survival against L. monocytogenes is due to early reduction of bacterial burdens and elicitation of Ly6C+ inflammatory monocytes. We demonstrate that a single TLR11/TLR12 ligand profilin (TgPRF) was sufficient to reduce bacterial burdens similar to T. gondii chronic infection. Stimulation with TgPRF was also sufficient to enhance animal survival when administered either pre- or post-Listeria infection. The ability of TgPRF to reduce L. monocytogenes burdens was dependent on TLR11 and required IFN-γ but was not dependent on IL-12 signaling. TgPRF induced rapid production of MCP-1 and resulted in trafficking of Ly6Chi CCR2+ inflammatory monocytes and Ly6G+ neutrophils into the blood and spleen. Stimulation with TgPRF reduced L. monocytogenes burdens in mice depleted with the Ly6G specific MAb 1A8, but not in Ly6C/Ly6G specific RB6-8C5 depleted or CCR2-/- mice, indicating that only inflammatory monocytes are required for TgPRF-induced reduction in bacterial burdens. These results demonstrate that stimulation of TLR11 by TgPRF is a mechanism to promote the emigration of Ly6Chi CCR2+ monocytes, and that TgPRF recruited inflammatory monocytes can provide an immunological benefit against an unrelated pathogen.

Early coupled up-regulation of interleukin-12 receptor beta-1 in CD8+ central memory and effector T cells for better clinical outcomes in liver transplant recipients.

This study aimed to investigate the role of initial priming of interleukin (IL)-12 receptor beta-1 in CD8(+) central memory T cells (initial IL-12RTCM priming) and CCR7-negative subsets (CNS) in effector cell expansion and clinical outcome after living donor liver transplantation (LDLT). One hundred and six patients who underwent LDLT were classified into the following three groups according to hierarchical clustering of CD8(+) CD45 isoforms before LDLT: I, naive-dominant; II, effector memory-dominant; and III, effector-dominant. The pre-existing CD8(+) effector cells (TE ) and activated immune status increased progressively from group I to group II to group III. Groups I, II and III received tacrolimus (Tac)/glucocorticoid (GC) regimens. Eighteen group III recipients received Tac/mycophenolate mofetil (MMF) and were defined as group IV. Initial IL-12RTCM priming was slightly, moderately and markedly decreased in droups I, II, and III, respectively. Initial priming of IL-12Rß1 in CNS was decreased markedly in the three groups with marked decreases of TE , perforin and interferon (IFN)-γ; all parameters were restored by up-regulation of IL-12Rß1(+) TCM through the self-renewal of TCM . The lag time required until coupled up-regulation of IL-12Rß1 of TCM and CNS to above baseline was 12, 20 and 32 days in groups I, II and III, respectively. Inferior clinical outcomes were associated with increasing lag time. In contrast, the initial priming of IL-12Rß1 in TCM and CNS remained above baseline in group IV due to MMF-mediated increase of IL-12Rß1. Early coupled up-regulation of TCM and CNS leads to efficient TE differentiation and optimal clinical outcomes.

Remission of systemic lupus erythematosus disease activity with regulatory cytokine interleukin (IL)-35 in Murphy Roths Large (MRL)/lpr mice.

The immunological mechanisms mediated by regulatory cytokine interleukin (IL)-35 are unclear in systemic lupus erythematosus (SLE). We investigated the frequency of CD4(+) CD25(+) forkhead box protein 3 (FoxP3)(+) regulatory T (Treg ) and IL-10(+) regulatory B (Breg ) cells and related immunoregulatory mechanisms in a female Murphy Roths Large (MRL)/lpr mouse model of spontaneous lupus-like disease, with or without IL-35 treatment. A remission of histopathology characteristics of lupus flare and nephritis was observed in the MRL/lpr mice upon IL-35 treatment. Accordingly, IL-35 and IL-35 receptor subunits (gp130 and IL-12Rß2) and cytokines of MRL/lpr and BALB/c mice (normal controls) were measured. The increased anti-inflammatory cytokines and decreased proinflammatory cytokines were possibly associated with the restoration of Treg and Breg frequency in MRL/lpr mice with IL-35 treatment, compared to phosphate-buffered saline (PBS) treatment. mRNA expressions of Treg -related FoxP3, IL-35 subunit (p35 and EBI3) and soluble IL-35 receptor subunit (gp130 and IL12Rß2) in splenic cells were up-regulated significantly in IL-35-treated mice. Compared with the PBS treatment group, IL-35-treated MRL/lpr mice showed an up-regulation of Treg -related genes and the activation of IL-35-related intracellular Janus kinase/signal transducer and activator of transcription signal pathways, thereby indicating the immunoregulatory role of IL-35 in SLE. These in vivo findings may provide a biochemical basis for further investigation of the regulatory mechanisms of IL-35 for the treatment of autoimmune-mediated inflammation.

Autoimmune risk loci of IL12RB2, IKZF1, XKR6, TMEM39A and CSK in Chinese patients with systemic lupus erythematosus.

Recent genome-wide or follow-up studies conducted in European or Caucasian populations have identified single nucleotide polymorphisms (SNPs) conferring increased risk to autoimmune diseases. It is unclear whether these observations can apply to systemic lupus erythematosus (SLE) in China. An association study was performed on 395 SLE patients and 378 healthy controls recruited from the Chinese population, in which the IL12RB2 rs3790567, IKZF1 rs2366293, XKR6 rs4240671, TMEM39A rs1132200 and CSK rs34933034 polymorphisms were examined by Matrix Assisted Laser Desorption Time of Flight Mass Spectrometry. The frequency of the A allele of IL12RB2 rs3790567 was lower in the cases compared with the controls (24.8% vs 30.2%, P = 0.018) and significant difference among the AA, AG and GG genotypes of rs3790567 was detected between the SLE patients and healthy controls (P = 0.020). We also found a statistically significant difference in the dominant model (GG+AG vs AA, P = 0.008). There was no correlation between the genotypes and specific sub-phenotypes in the current cohort. Associations with IKZF1 rs2366293, XKR6 rs4240671, TMEM39A rs1132200 and CSK rs34933034 were also lacking (P > 0.05). The results supported the theory that IL12RB2 is associated with SLE in the Chinese population.

Deregulation of Fas ligand expression as a novel cause of autoimmune lymphoproliferative syndrome-like disease.

Autoimmune lymphoproliferative syndrome is frequently caused by mutations in genes involved in the Fas death receptor pathway, but for 20-30% of patients the genetic defect is unknown. We observed that treatment of healthy T cells with interleukin-12 induces upregulation of Fas ligand and Fas ligand-dependent apoptosis. Consistently, interleukin-12 could not induce apoptosis in Fas ligand-deficient T cells from patients with autoimmune lymphoproliferative syndrome. We hypothesized that defects in the interleukin-12 signaling pathway may cause a similar phenotype as that caused by mutations of the Fas ligand gene. To test this, we analyzed 20 patients with autoimmune lymphoproliferative syndrome of unknown cause by whole-exome sequencing. We identified a homozygous nonsense mutation (c.698G>A, p.R212*) in the interleukin-12/interleukin-23 receptor-component IL12RB1 in one of these patients. The mutation led to IL12RB1 protein truncation and loss of cell surface expression. Interleukin-12 and -23 signaling was completely abrogated as demonstrated by deficient STAT4 phosphorylation and interferon γ production. Interleukin-12-mediated expression of membrane-bound and soluble Fas ligand was lacking and basal expression was much lower than in healthy controls. The patient presented with the classical symptoms of autoimmune lymphoproliferative syndrome: chronic non-malignant, non-infectious lymphadenopathy, splenomegaly, hepatomegaly, elevated numbers of double-negative T cells, autoimmune cytopenias, and increased levels of vitamin B12 and interleukin-10. Sanger sequencing and whole-exome sequencing excluded the presence of germline or somatic mutations in genes known to be associated with the autoimmune lymphoproliferative syndrome. Our data suggest that deficient regulation of Fas ligand expression by regulators such as the interleukin-12 signaling pathway may be an alternative cause of autoimmune lymphoproliferative syndrome-like disease.

Developmental expression of IL-12Rß2 on murine naive neonatal T cells counters the upregulation of IL-13Rα1 on primary Th1 cells and balances immunity in the newborn.

Upon exposure to Ag on the day of birth, neonatal mice mount balanced primary Th1 and Th2 responses, with the former displaying upregulated IL-13Rα1 expression. This chain associates with IL-4Rα to form a heteroreceptor (IL-4Rα/IL-13Rα1) that marks the Th1 cells for death by IL-4 produced by Th2 cells during rechallenge with Ag, hence the Th2 bias of murine neonatal immunity. The upregulation of IL-13Rα1 on neonatal Th1 cells was due to the paucity of IL-12 in the neonatal environment. In this study, we show that by day 8 after birth, naive splenic T cells are no longer susceptible to IL-13Rα1 upregulation even when exposed to Ag within the neonatal environment. Furthermore, during the 8-d lapse, the naive splenic T cells spontaneously and progressively upregulate the IL-12Rß2 chain, perhaps due to colonization by commensals, which induce production of IL-12 by cells of the innate immune system such as dendritic cells. In fact, mature T cells from the thymus, a sterile environment not accessible to microbes, did not upregulate IL-12Rß2 and were unable to counter IL-13Rα1 upregulation. Finally, the 8-d naive T cells were able to differentiate into Th1 cells even independently of IL-12 but required the cytokine to counter upregulation of IL-13Rα1. Thus, in neonatal mice, IL-12, which accumulates in the environment progressively, uses IL-12Rß2 to counter IL-13Rα1 expression in addition to promoting Th1 differentiation.