SEL1L-HRD1 endoplasmic reticulum-associated degradation controls STING-mediated innate immunity by limiting the size of the activable STING pool.

Stimulator of interferon genes (STING) orchestrates the production of proinflammatory cytokines in response to cytosolic double-stranded DNA; however, the pathophysiological significance and molecular mechanism underlying the folding and maturation of nascent STING protein at the endoplasmic reticulum (ER) remain unknown. Here we report that the SEL1L-HRD1 protein complex-the most conserved branch of ER-associated degradation (ERAD)-is a negative regulator of the STING innate immunity by ubiquitinating and targeting nascent STING protein for proteasomal degradation in the basal state. SEL1L or HRD1 deficiency in macrophages specifically amplifies STING signalling and immunity against viral infection and tumour growth. Mechanistically, nascent STING protein is a bona fide substrate of SEL1L-HRD1 in the basal state, uncoupled from ER stress or its sensor inositol-requiring enzyme 1α. Hence, our study not only establishes a key role of SEL1L-HRD1 ERAD in innate immunity by limiting the size of the activable STING pool, but identifies a regulatory mechanism and therapeutic approach to targeting STING.

Interleukin-37 promotes DMBA/TPA skin cancer through SIGIRR-mediated inhibition of glycolysis in CD103+DC cells.

Interleukin 37 (IL-37), a member of the IL-1 family, is considered a suppressor of innate and adaptive immunity and, hence is a regulator of tumor immunity. However, the specific molecular mechanism and role of IL-37 in skin cancer remain unclear. Here, we report that IL-37b-transgenic mice (IL-37tg) treated with the carcinogenic 7,12-dimethylbenzoanthracene (DMBA)/12-o-tetradecylphorbol-13-acetate (TPA) exhibited enhanced skin cancer and increased tumor burden in the skin by inhibiting the function of CD103+ dendritic cells (DCs). Notably, IL-37 induced rapid phosphorylation of adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK), and via single immunoglobulin IL-1-related receptor (SIGIRR), inhibited the long-term Akt activation. Specifically, by affecting the SIGIRR-AMPK-Akt signaling axis, which is related to the regulation of glycolysis in CD103+DCs, IL-37 inhibited their anti-tumor function. Our results show that a marked correlation between the CD103+DC signature (IRF8, FMS-like tyrosine kinase 3 ligand, CLEC9A, CLNK, XCR1, BATF3, and ZBTB46) and chemokines C-X-C motif chemokine ligand 9, CXCL10, and CD8A in a mouse model with DMBA/TPA-induced skin cancer. In a word, our results highlight that IL-37 as an inhibitor of tumor immune surveillance through modulating CD103+DCs and establishing an important link between metabolism and immunity as a therapeutic target for skin cancer.

Hepatic SEL1L-HRD1 ER-associated degradation regulates systemic iron homeostasis via ceruloplasmin.

Iron homeostasis is critical for cellular and organismal function and is tightly regulated to prevent toxicity or anemia due to iron excess or deficiency, respectively. However, subcellular regulatory mechanisms of iron remain largely unexplored. Here, we report that SEL1L-HRD1 protein complex of endoplasmic reticulum (ER)-associated degradation (ERAD) in hepatocytes controls systemic iron homeostasis in a ceruloplasmin (CP)-dependent, and ER stress-independent, manner. Mice with hepatocyte-specific Sel1L deficiency exhibit altered basal iron homeostasis and are sensitized to iron deficiency while resistant to iron overload. Proteomics screening for a factor linking ERAD deficiency to altered iron homeostasis identifies CP, a key ferroxidase involved in systemic iron distribution by catalyzing iron oxidation and efflux from tissues. Indeed, CP is highly unstable and a bona fide substrate of SEL1L-HRD1 ERAD. In the absence of ERAD, CP protein accumulates in the ER and is shunted to refolding, leading to elevated secretion. Providing clinical relevance of these findings, SEL1L-HRD1 ERAD is responsible for the degradation of a subset of disease-causing CP mutants, thereby attenuating their pathogenicity. Together, this study uncovers the role of SEL1L-HRD1 ERAD in systemic iron homeostasis and provides insights into protein misfolding-associated proteotoxicity.

Interleukin-38 promotes skin tumorigenesis in an IL-1Rrp2-dependent manner.

Interleukin-38 (IL-38) is strongly associated with chronic inflammatory diseases; however, its role in tumorigenesis is poorly understood. We demonstrated that expression of IL-38, which exhibits high expression in the skin, is downregulated in human cutaneous squamous cell carcinoma and 7,12-dimethylbenzanthracene/12-O-tetradecanoyl phorbol-13-acetate-induced mouse skin tumorigenesis. IL-38 keratinocyte-specific knockout mice displayed suppressed skin tumor formation and malignant progression. Keratinocyte-specific deletion of IL-38 was associated with reduced expression of inflammatory cytokines, leading to reduced myeloid cell infiltration into the local tumor microenvironment. IL-38 is dispensable for epidermal mutagenesis, but IL-38 keratinocyte-specific deletion reduces proliferative gene expression along with epidermal cell proliferation and hyperplasia. Mechanistically, we first demonstrated that IL-38 activates the c-Jun N-terminal kinase (JNK)/activator protein 1 signal transduction pathway to promote the expression of cancer-related inflammatory cytokines and proliferation and migration of tumor cells in an IL-1 receptor-related protein 2 (IL-1Rrp2)-dependent manner. Our findings highlight the role of IL-38 in the regulation of epidermal cell hyperplasia and pro-tumorigenic microenvironment through IL-1Rrp2/JNK and suggest IL-38/IL-1Rrp2 as a preventive and potential therapeutic target in skin cancer.

Interleukin-37 promotes colitis-associated carcinogenesis via SIGIRR-mediated cytotoxic T cells dysfunction.

Interleukin-37b (hereafter called IL-37) was identified as fundamental inhibitor of natural and acquired immunity. The molecular mechanism and function of IL-37 in colorectal cancer (CRC) has been elusive. Here, we found that IL-37 transgenic (IL-37tg) mice were highly susceptible to colitis-associated colorectal cancer (CAC) and suffered from dramatically increased tumor burdens in colon. Nevertheless, IL-37 is dispensable for intestinal mutagenesis, and CRC cell proliferation, apoptosis, and migration. Notably, IL-37 dampened protective cytotoxic T cell-mediated immunity in CAC and B16-OVA models. CD8+ T cell dysfunction is defined by reduced retention and activation as well as failure to proliferate and produce cytotoxic cytokines in IL-37tg mice, enabling tumor evasion of immune surveillance. The dysfunction led by IL-37 antagonizes IL-18-induced proliferation and effector function of CD8+ T cells, which was dependent on SIGIRR (single immunoglobulin interleukin-1 receptor-related protein). Finally, we observed that IL-37 levels were significantly increased in CRC patients, and positively correlated with serum CRC biomarker CEA levels, but negatively correlated with the CD8+ T cell infiltration in CRC patients. Our findings highlight the role of IL-37 in harnessing antitumor immunity by inactivation of cytotoxic T cells and establish a new defined inhibitory factor IL-37/SIGIRR in cancer-immunity cycle as therapeutic targets in CRC.

EDIL3 deficiency ameliorates adverse cardiac remodelling by neutrophil extracellular traps (NET)-mediated macrophage polarization.

AIMS: After myocardial infarction (MI), injured cardiomyocytes recruit neutrophils and monocytes/macrophages to myocardium, which in turn initiates inflammatory and reparative cascades, respectively. Either insufficient or excessive inflammation impairs cardiac healing. As an endogenous inhibitor of neutrophil adhesion, EDIL3 plays a crucial role in inflammatory regulation. However, the role of EDIL3 in MI remains obscure. We aimed to define the role of EDIL3 in cardiac remodelling after MI. METHODS AND RESULTS: Serum EDIL3 levels in MI patients were negatively associated with MI biomarkers. Consistently, WT mice after MI showed low levels of cardiac EDIL3. Compared with WT mice, Edil3-/- mice showed improvement of post-MI adverse remodelling, as they exhibited lower mortality, better cardiac function, shorter scar length, and smaller LV cavity. Accordingly, infarcted hearts of Edil3-/- mice contained fewer cellular debris and lower amounts of fibrosis content, with decreased collagen I/III expression and the percentage of α-smooth muscle actin myofibroblasts. Mechanistically, EDIL3 deficiency did not affect the recruitment of monocytes or T cells, but enhanced neutrophil recruitment and following expansion of pro-inflammatory Mertk-MHC-IIlo-int (myeloid-epithelial-reproductive tyrosine kinase/major histocompatibility complex II) macrophages. The injection of neutrophil-specific C-X-C motif chemokine receptor 2 antagonist eliminated the differences in macrophage polarization and cardiac function between WT and Edil3-/- mice after MI. Neutrophil extracellular traps (NETs), which were more abundant in the hearts of Edil3-/- mice, contributed to Mertk-MHC-IIlo-int polarization via Toll-like receptor 9 pathway. The inhibition of NET formation by treatment of neutrophil elastase inhibitor or DNase I impaired macrophage polarization, increased cellular debris and aggravated cardiac adverse remodelling, thus removed the differences of cardiac function between WT and Edil3-/- mice. Totally, EDIL3 plays an important role in NET-primed macrophage polarization and cardiac remodelling during MI. CONCLUSION: We not only reveal that EDIL3 deficiency ameliorates adverse cardiac healing via NET-mediated pro-inflammatory macrophage polarization but also discover a new crosstalk between neutrophil and macrophage after MI.

IL-30 ameliorates imiquimod and K14-VEGF induced psoriasis-like disease by inhibiting both innate and adaptive immunity disorders.

Psoriasis is a severe skin disease with significant physical and psychological health consequences. As a typical type of immune disease, both innate and adaptive immunity disorders play key roles in the development of psoriasis. Interleukin (IL)-30 was thought as a natural antagonist of gp130-mediated signaling that affects T helper type 1 and 17 cell polarization by inhibiting IL-6 and IL-27 signaling pathways. Here, we found that, in vitro, IL-30 reduced cytokine levels of HaCaT keratinocytes and dendritic cells (DCs), weakened the maturationS of DCs, inhibited DC-mediated T cell proliferation, and blocked the activation of nuclear factor-κB. In vivo, IL-30 inhibited the development of skin disease in two animal models: Krt14-Vegfa and imiquimod (IMQ)-induced psoriasis-like skin disease. Thus, IL-30 may be useful as a therapeutic agent for controlling psoriasis.

Autophagy-based unconventional secretion of HMGB1 by keratinocytes plays a pivotal role in psoriatic skin inflammation.

The precise mechanism through which macroautophagy/autophagy affects psoriasis is poorly understood. Here, we found that keratinocyte (KC) autophagy, which was positively correlated with psoriatic severity in patients and mouse models and could be inhibited by mitogen-activated protein kinase (MAPK) family inactivation. The impairment of autophagic flux alleviated psoriasisform inflammation. We also found that an autophagy-based unconventional secretory pathway (autosecretion) dependent on ATG5 (autophagy related 5) and GORASP2 (golgi reassembly stacking protein 2) promoted psoriasiform KC inflammation. Moreover, the alarmin HMGB1 (high mobility group box 1) was more effective than other autosecretory proteins in regulating psoriasiform cutaneous inflammation. HMGB1 neutralization in autophagy-efficient KCs eliminated the differences in psoriasiform inflammation between Krt14+/+-Atg5f/f KCs and Krt14Cre/+-atg5f/f KCs, and conversely, recombinant HMGB1 almost completely restored psoriasiform inflammation in Krt14Cre/+-atg5f/f KCs in vivo. These results suggest that HMGB1-associated autosecretion plays a pivotal role in cutaneous inflammation. Finally, we demonstrated that Krt14Cre/+-hmgb1f/f mice displayed attenuated psoriatic inflammation due to the essential crosstalk between KC-specific HMGB1-associated autosecretion and γδT cells. Thus, this study uncovered a novel autophagy mechanism in psoriasis pathogenesis, and the findings imply the clinical significance of investigating and treating psoriasis.Abbreviations: 3-MA: 3-methyladenine; ACTB: actin beta; AGER: advanced glycosylation end-product specific receptor; Anti-HMGB1: anti-HMGB1 neutralizing antibody; Anti-IL18: anti-IL18 neutralizing antibody; Anti-IL1B: anti-IL1B neutralizing antibody; ATG5: autophagy related 5; BAF: bafilomycin A1; BECN1: beclin 1; CASP1: caspase 1; CCL: C-C motif chemokine ligand; CsA: cyclosporine A; ctrl shRNA: lentivirus harboring shRNA against control; CXCL: C-X-C motif chemokine ligand; DCs: dendritic cells; DMEM: dulbecco's modified Eagle's medium; ELISA: enzyme-linked immunosorbent assay; EM: electron microscopy; FBS: fetal bovine serum; GORASP2 shRNA: lentivirus harboring shRNA against GORASP2; GORASP2/GRASP55: golgi reassembly stacking protein 2; GR1: a composite epitope between LY6 (lymphocyte antigen 6 complex) locus C1 and LY6 locus G6D antigens; H&E: hematoxylin and eosin; HMGB1: high mobility group box 1; HMGB1 shRNA: lentivirus harboring shRNA against HMGB1; IFNG/IFN-γ: interferon gamma; IL17A: interleukin 17A; IL18: interleukin 18; IL1A/IL-1α: interleukin 1 alpha; IL1B/IL-1ß: interleukin 1 beta; IL22/IL-22: interleukin 22; IL23A: interleukin 23 subunit alpha; IL23R: interleukin 23 receptor; IMQ: imiquimod; ITGAM/CD11B: integrin subunit alpha M; ITGAX/CD11C: integrin subunit alpha X; IVL: involucrin; KC: keratinocyte; KD: knockdown; KO: knockout; Krt14+/+-Atg5f/f mice: mice bearing an Atg5 flox allele, in which exon 3 of the Atg5 gene is flanked by two loxP sites; Krt14+/+-Hmgb1f/f: mice bearing an Hmgb1 flox allele, in which exon 2 to 4 of the Hmgb1 gene is flanked by two loxP sites; Krt14Cre/+-atg5f/f mice: keratinocyte-specific atg5 knockout mice generated by mating Atg5-floxed mice with mice expressing Cre recombinase under the control of the promoter of Krt4; Krt14Cre/+-hmgb1f/f mice: keratinocyte-specific hmgb1 knockout mice generated by mating Hmgb1-floxed mice with mice expressing Cre recombinase under the control of the promoter of Krt14; Krt14-Vegfa mice: mice expressing 164-amino acid Vegfa splice variant recombinase under the control of promoter of Krt14; LAMP1: lysosomal associated membrane protein 1; LDH: lactate dehydrogenase; LORICRIN: loricrin cornified envelope precursor protein; M5: TNF, IL1A, IL17A, IL22 and OSM in combination; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAPK: mitogen-activated protein kinase; MKI67: marker of proliferation Ki-67; MTT: thiazolyl blue tetrazolium bromide; NFKB/NF-κB: nuclear factor kappa B; NHEKs: primary normal human epidermal keratinocytes; NS: not significant; OSM: oncostatin M; PASI: psoriasis area and severity index; PtdIns3K: class III phosphatidylinositol 3-kinase; qRT-PCR: quantitative RT-PCR; RELA/p65: RELA proto-oncogene, NF-kB subunit; rHMGB1: recombinant HMGB1; rIL18: recombinant interleukin 18; rIL1B: recombinant interleukin 1 beta; S100A: S100 calcium binding protein A; SQSTM1/p62: sequestosome 1; T17: IL17A-producing T; TCR: T-cell receptor; tcrd KO mice: tcrd (T cell receptor delta chain) knockout mice, which show deficient receptor expression in all adult lymphoid and epithelial organs; TLR: toll-like receptor; TNF/TNF-α: tumor necrosis factor; WOR: wortmannin; WT: wild-type; γδT17 cells: IL17A-producing γδ T cells.

IL-37 ameliorates the inflammatory process in psoriasis by suppressing proinflammatory cytokine production.

IL-37 is a potent inhibitor of innate immunity by shifting the cytokine equilibrium away from excessive inflammation. Psoriasis is thought to be initiated by abnormal interactions between the cutaneous keratinocytes and systemic immune cells, triggering keratinocyte hyperproliferation. In the current study, we assessed IL-37 in two well-known psoriasis models: a human keratinocyte cell line (HaCaT) and the keratin 14 VEGF-A-transgenic mouse model. First, we used the HaCaT cell line, which was transiently transfected with an overexpressing IL-37 vector, and tested the effect of IL-37 on these cells using a mixture of five proinflammatory cytokines. IL-37 was effective in suppressing the production of CXCL8, IL-6, and S100A7, which were highly upregulated by the mixture of five proinflammatory cytokines. Keratin 14 VEGF-A-transgenic mice were treated with plasmid coding human IL-37 sequence-formulated cationic liposomes, and we observed potent immunosuppressive effects over the 18-d period. In this model, we observed reduced systemic IL-10 levels, local IFN-γ gene transcripts, as well as mild mast cell infiltration into the psoriatic lesions of the mice. Immunohistochemical analysis indicated that IL-37 was expressed by effector memory T cells, as well as macrophages, in human psoriatic plaques. In conclusion, our studies strongly indicate that IL-37 plays a potent immunosuppressive role in the pathogenesis of both experimental psoriasis models in vitro and in vivo by downregulating proinflammatory cytokines. Importantly, our findings highlight new therapeutic strategies that can be designed to use this immunosuppressive anti-inflammatory cytokine in psoriasis and other inflammatory cutaneous diseases.

[A case of Leigh syndrome associated with respiratory chain complex I deficiency due to mitochondrial gene 13513G>A mutation].

Leigh syndrome is a genetically heterogeneous disease caused by defects in enzymes involved in aerobic energy metabolism and the Krebs', cycle. Mitonchondrial complex I deficiency is a main cause of Leigh syndrome. In this study, a Chinese child with Leigh syndrome caused by 13513G>A mutation was reported. The proband was the first child of his parents. The previously healthy boy presented with generalized epilepsy at 12 years of age. When he visited Peking University First Hospital at 13 years of age, he had subacute loss of vision in two eyes and temporal defect of visual field in the left eye. He walked with a spastic gait. His blood lactate and pyruvate levels were elevated. Muscle biopsy showed mild lipid accumulation in muscle fiber. An electrocardiogram showed incomplete right bundle branch block. Brain magnetic resonance imaging showed bilateral, symmetrical lesions in the basal ganglia, supporting the diagnosis of Leigh syndrome. 13513G>A mutation was identified by gene analysis in the patient, which led to mitochondrial respiratory chain complex I deficiency. Multivitamins and L-carnitine were administered. At present, the patient is 16 years old and has progressive deterioration with significant muscle weakness and body weight loss. He is absent from school. He has no obvious retardation in intelligence. 13513G>A mutation was first identified by gene analysis in Chinese population with Leigh syndrome. This may be helpful in genetic counseling.