IL-10 Deficient Mice Research Articles

Histochemical assessment of accelerated bone remodeling and reduced mineralization in Il-6 deficient mice

ObjectivesInterleukin-6 (IL-6) contributes to the regulation of functions in various tissues and organs. Even though IL-6 has been reported to modulate bone metabolism in previous studies, this finding is controversial. This study aims to evaluate the possible involvement of IL-6 in bone metabolism by examining the histological activity of osteoblasts and osteoclasts in the femora of Il-6 deficient (Il-6−/−) mice. MethodsEight-week-old male Il-6−/− mice and their wild-type littermates were fixed with a paraformaldehyde solution, and their femora were extracted for micro-CT analysis, immunohistochemistry, and real-time PCR analysis. ResultsIl-6−/− femora showed an increased bone volume/tissue volume (TV) but a reduced bone mineral density compared with the wild-type. Furthermore, the tissue-nonspecific alkaline phosphatase positive area/TV ratio, the expression of Runx2, Osterix, and Rankl, and the number of tartrate-resistant acid phosphatase-positive osteoclasts were all increased in the Il-6−/− mice. A considerable number of unmineralized areas within the bone matrix and abundant sclerostin-reactive osteocytes were observed in Il-6−/− femoral metaphyses but not in the wild-type. Interestingly, the gene expression of Cd206 was elevated in Il-6−/− femora, and many F4/80-positive macrophages/monocytes and CD206-immunoreactive macrophages in the primary trabeculae had migrated closer to the growth plate, where intense RANKL immunoreactivity was detected. These results suggest that, in an IL-6-deficient state, CD206-positive macrophages may differentiate into osteoclasts when in contact with RANKL-reactive osteoblastic cells. ConclusionIn a state of IL-6 deficiency, the population and cell activities of osteoblast, osteoclasts, and macrophages seemed to be facilitated, except for the reduced mineralization in bone.

Depletion of Foxp3+ regulatory T cells but not the absence of CD19+IL-10+ regulatory B cells hinders tumor growth in a para-orthotopic neuroblastoma mouse model.

Adaptive immune cells with regulatory function reportedly mediate immune escape in a variety of tumors. Little is known regarding the relevance of the most prominent regulatory cell populations, namely Foxp3+ T regulatory cells (Tregs) and CD19+IL-10+ B regulatory cells (Bregs), for neuroblastoma (NB) survival. After establishing a novel immunocompetent syngeneic NB mouse model where orthotopic tumors can be generated after intrarenal injection of NB975A cells, we studied the importance of Tregs and Bregs in Foxp3-DTR mice whose Tregs can be depleted by diphtheria toxin (DT) application as well as in CD19-specific IL-10 deficient mice that lack IL-10+ Bregs (CD19cre+/- × IL-10fl/fl mice). We observed Foxp3 Treg cells in tumors from wild type mice. On the contrary, Bregs or B cells were scarce. Specific depletion of Tregs in Foxp3-DTR mice resulted in an 85% reduction of tumor volume and weight compared to DT-treated wild type mice and untreated Foxp3-DTR mice. In contrast, NB tumor growth was not affected in CD19-specific IL-10 deficient mice. Similarly, mice lacking mature B cells (μMT mice) and CD19 deficient mice (CD19cre mice) showed no change in growth pattern of NB tumors. In Treg-depleted mice, reduced tumor growth was associated with an increased concentration of IFN-gamma, TNF-alpha, IL-4, IL-6, and IL-10 in isolated splenocytes. In summary, transient ablation of Tregs but not absence of Bregs hindered the growth of NB, strongly suggesting the therapeutic potential of targeting Tregs for this aggressive childhood tumor.

Blockade of IL-6 inhibits tumor immune evasion and improves anti–PD-1 immunotherapy

Long-standing inflammatory bowel disease predisposes to the development of colorectal cancer (CRC). Interleukin (IL) -6, a pivotal link between chronic inflammation and tumor progression, has recently been recognized as a potential therapeutic target. The effect of IL-6 on proliferation and metastasis of CRC by activating the STAT3 pathway has been widely demonstrated in recent years, but few on mediating tumor immune evasion. In this study, we found that IL-6 was remarkably overexpressed in CRC and its elevation was associated with a poor prognosis. We studied CRC tumorigenesis in vivo by inoculating MC38 tumors and induced-CRC model via AOM/DSS (azoxymethane/dextransulfate sodium) in IL-6 deficient (IL-6-/-) and wild-type (WT) mice and found that IL-6-/- mice were less susceptible to develop tumors, compared to WT mice. We detected CD8+ T cells via immunofluorescence and found they exhibit high expression in tumor of IL-6-/- mice. High level of IL-6 was found in colitis model, with down-regulation of MHC-I molecules. In in vitro experiments, we found that IL-6 may act as a negative regulator in IFNγ-STAT1-MHC-I signaling. In addition, vivo trials also confirmed that MHC-I mRNA level was negatively related to the existence of IL-6. Furthermore, the blockade of IL-6 also activated CD8+T-cell accumulation and led to the high PD-L1 expression in CRC, which can sensitize animals to anti-PD-1 therapy. Our study provides a research basis for the significant role of IL-6 in tumor evasion and highlights a novel target to improve the efficacy of immunotherapy.

Eosinophils protect pressure overload- and β-adrenoreceptor agonist-induced cardiac hypertrophy.

Blood eosinophil (EOS) counts and EOS cationic protein (ECP) levels associate positively with major cardiovascular disease (CVD) risk factors and prevalence. This study investigates the role of EOS in cardiac hypertrophy. A retrospective cross-section study of 644 consecutive inpatients with hypertension examined the association between blood EOS counts and cardiac hypertrophy. Pressure overload- and β-adrenoreceptor agonist isoproterenol-induced cardiac hypertrophy was produced in EOS-deficient ΔdblGATA mice. This study revealed positive correlations between blood EOS counts and left ventricular (LV) mass and mass index in humans. ΔdblGATA mice showed exacerbated cardiac hypertrophy and dysfunction, with increased LV wall thickness, reduced LV internal diameter, and increased myocardial cell size, death, and fibrosis. Repopulation of EOS from wild-type (WT) mice, but not those from IL4-deficient mice ameliorated cardiac hypertrophy and cardiac dysfunctions. In ΔdblGATA and WT mice, administration of ECP mEar1 improved cardiac hypertrophy and function. Mechanistic studies demonstrated that EOS expression of IL4, IL13, and mEar1 was essential to control mouse cardiomyocyte hypertrophy and death and cardiac fibroblast TGF-β signalling and fibrotic protein synthesis. The use of human cardiac cells yielded the same results. Human ECP, EOS-derived neurotoxin, human EOS, or murine recombinant mEar1 reduced human cardiomyocyte death and hypertrophy and human cardiac fibroblast TGF-β signalling. Although blood EOS counts correlated positively with LV mass or LV mass index in humans, this study established a cardioprotective role for EOS IL4 and cationic proteins in cardiac hypertrophy and tested a therapeutic possibility of ECPs in this human CVD.

Therapeutic Evaluation of Fecal Microbiota Transplantation in an Interleukin 10-deficient Mouse Model.

With the development of microecology in recent years, the relationship between intestinal bacteria and inflammatory bowel disease (IBD) has attracted considerable attention. Accumulating evidence suggests that dysbiotic microbiota plays an active role in triggering or worsening the inflammatory process in IBD and that fecal microbiota transplantation (FMT) is an attractive therapeutic strategy since transferring a healthy microbiota to IBD patient could restore the appropriate host-microbiota communication. However, the molecular mechanisms are unclear, and the efficacy of FMT has not been very well established. Thus, further studies in animal models of IBD are necessary. In this method, we applied FMT from wild-type C57BL/6J mice to IL-10 deficient mice, a widely used mouse model of colitis. The study elaborates on collecting fecal pellets from the donor mice, making the fecal solution/suspension, administering the fecal solution, and monitoring the disease. We found that FMT significantly mitigated the cardiac impairment in IL-10 knockout mice, underlining its therapeutic potential for IBD management.

Multi-omics analyses of the ulcerative colitis gut microbiome link Bacteroides vulgatus proteases with disease severity.

Ulcerative colitis (UC) is driven by disruptions in host-microbiota homeostasis, however current treatments exclusively target host inflammatory pathways. To understand how host-microbiota interactions become disrupted in UC, we collected and analyzed six fecal or serum based –omic datasets (metaproteomic, metabolomic, metagenomic, metapeptidomic, and amplicon sequencing profiles of fecal samples and proteomic profiles of serum samples) from 40 UC patients at a single inflammatory bowel disease centre, as well as various clinical, endoscopic, and histologic measures of disease activity. A validation cohort of 210 samples (73 UC, 117 Crohn’s disease (CD), 20 healthy controls) was collected and analyzed separately and independently. Data integration across both cohorts showed that a subset of the clinically active UC patients had an over-abundance of proteases that originated from the bacterium, Bacteroides vulgatus. To test whether B. vulgatus proteases contribute to UC disease activity, we first profiled B. vulgatus proteases found in patients and bacterial cultures. Use of a broad-spectrum protease inhibitor improved B. vulgatus-induced barrier dysfunction in vitro, and prevented colitis in B. vulgatus monocolonized, IL-10 deficient mice. Furthermore, transplantation of feces from UC patients with a high abundance of B. vulgatus proteases into germ-free mice induced colitis dependent on protease activity. These results, stemming from a multi-omics approach, improve understanding of functional microbiota alterations that drive UC and provides a resource for identifying other pathways that could be inhibited as a strategy to treat this disease.

Abstract PO-102: Fibroblast-derived interleukin-33 promotes pancreatic ductal adenocarcinoma as a result of tumor cell KRASG12D

Abstract Pancreatic ductal adenocarcinoma (PDA) is a deadly disease that is difficult to detect early and limited in its therapeutic options. As a result, the five-year survival rate of PDA is only 10%. Expression of the oncogene KRAS is present in over 94% of PDA cases, with the most common KRAS mutation being KRASG12D. Another hallmark of PDA is its reactive, fibroinflammatory tumor microenvironment, which is established in tandem with the earliest stages of tumorigenesis and is essential to the growth and maintenance of the tumor. In this work, we have endeavored to uncover the ways by which tumor cell KRASG12D expression influences extracellular signals that shape the tumor microenvironment to the benefit of the disease, with the overall goal of identifying new potential therapeutic vulnerabilities downstream of KRAS signaling. Importantly, using genetically engineered mouse models (GEMMs) and in vitro assays by which tumor cell expression of KrasG12D can be turned on and off, we have found that tumor cell KRASG12D activity stimulates the expression of Interleukin-33 (IL33) in pancreatic fibroblasts through a mechanism requiring both JAK/STAT3 and Focal Adhesion Kinase (FAK) signaling. Through immunohistochemical staining and single cell RNA sequencing of patient tumor samples, we have also confirmed the expression of IL33 in the cancer-associated fibroblast compartment of human PDA. IL33 is a dual function cytokine - it is sequestered in the nucleus where it can impact the cellular transcriptome, and it can also be released to signal to cells expressing its receptor, ST2L. Across the cancer biology field, IL33 expression has been reported to be regulated by different signaling factors, and its impact can be either tumor promoting or tumor restricting depending on the tissue context. In PDA, the role of IL33 is currently unclear, with recent publications alternatively describing it as anti- or pro-tumor. While previous studies have focused on epithelial IL33, we have used a Pdgfra-CreERT2/+;Il33f/f GEMM to knock out Il33 in fibroblasts prior to orthotopic implantation of syngeneic PDA cells. Tumor growth in Il33-deficient mice was reduced, and we observed fewer infiltrating immune cells, including macrophages. As our group and others have previously shown that macrophages are a significant driver of immunosuppression required for the maintenance of PDA, we hypothesized that fibroblast-derived Il33 promotes pancreatic tumorigenesis through either direct or indirect recruitment of immunosuppressive macrophages. As we continue to dissect the role of IL33 in the pancreatic tumor microenvironment, we aim to fully understand the biological role of IL33 during the onset of carcinogenesis and in advanced disease. Overall, this work will shed new light on the ways by which fibroblasts and macrophages are co-opted by tumor cells as a result of KRASG12D, further elucidating prospective therapeutic avenues that may be exploited in the future. Citation Format: Katelyn Donahue, Wenting Du, Carlos Espinoza, Eileen Carpenter, Kristee Brown, Nina Steele, Marina Pasca di Magliano. Fibroblast-derived interleukin-33 promotes pancreatic ductal adenocarcinoma as a result of tumor cell KRASG12D [abstract]. In: Proceedings of the AACR Virtual Special Conference on Pancreatic Cancer; 2021 Sep 29-30. Philadelphia (PA): AACR; Cancer Res 2021;81(22 Suppl):Abstract nr PO-102.

Interleukin-10 improves stroke outcome by controlling the detrimental Interleukin-17A response

BackgroundLymphocytes have dichotomous functions in ischemic stroke. Regulatory T cells are protective, while IL-17A from innate lymphocytes promotes the infarct growth. With recent advances of T cell-subtype specific transgenic mouse models it now has become possible to study the complex interplay of T cell subpopulations in ischemic stroke.MethodsIn a murine model of experimental stroke we analyzed the effects of IL-10 on the functional outcome for up to 14 days post-ischemia and defined the source of IL-10 in ischemic brains based on immunohistochemistry, flow cytometry, and bone-marrow chimeric mice. We used neutralizing IL-17A antibodies, intrathecal IL-10 injections, and transgenic mouse models which harbor a deletion of the IL-10R on distinct T cell subpopulations to further explore the interplay between IL-10 and IL-17A pathways in the ischemic brain.ResultsWe demonstrate that IL-10 deficient mice exhibit significantly increased infarct sizes on days 3 and 7 and enlarged brain atrophy and impaired neurological outcome on day 14 following tMCAO. In ischemic brains IL-10 producing immune cells included regulatory T cells, macrophages, and microglia. Neutralization of IL-17A following stroke reversed the worse outcome in IL-10 deficient mice and intracerebral treatment with recombinant IL-10 revealed that IL-10 controlled IL-17A positive lymphocytes in ischemic brains. Importantly, IL-10 acted differentially on αβ and γδ T cells. IL-17A producing CD4+ αβ T cells were directly controlled via their IL-10-receptor (IL-10R), whereas IL-10 by itself had no direct effect on the IL-17A production in γδ T cells. The control of the IL-17A production in γδ T cells depended on an intact IL10R signaling in regulatory T cells (Tregs).ConclusionsTaken together, our data indicate a key function of IL-10 in restricting the detrimental IL-17A-signaling in stroke and further supports that IL-17A is a therapeutic opportunity for stroke treatment.

Akkermansia muciniphila in inflammatory bowel disease and colorectal cancer.

Akkermansia muciniphila in inflammatory bowel disease and colorectal cancer.

Endogenous Interleukin-33 Acts as an Alarmin in Liver Ischemia-Reperfusion and Is Associated With Injury After Human Liver Transplantation.

Ischemia and reperfusion injury is an early inflammatory process during liver transplantation that impacts on graft function and clinical outcomes. Interleukin (IL)-33 is a danger-associated molecular pattern involved in kidney ischemia/reperfusion injury and several liver diseases. The aims were to assess whether IL-33 was released as an alarmin responsible for ischemia/reperfusion injury in a mouse model of warm hepatic ischemia, and whether this hypothesis could also apply in the setting of human liver transplantation. First, a model of warm hepatic ischemia/reperfusion was used in wild-type and IL-33–deficient mice. Severity of ischemia/reperfusion injury was assessed with ALT and histological analysis. Then, serum IL-33 was measured in a pilot cohort of 40 liver transplant patients. Hemodynamic postreperfusion syndrome, graft dysfunction (assessed by model for early allograft scoring >6), renal failure, and tissue lesions on time-zero biopsies were assessed. In the mouse model, IL-33 was constitutively expressed in the nucleus of endothelial cells, immediately released in response to hepatic pedicle clamping without neosynthesis, and participated in the recruitment of neutrophils and tissue injury on site. The kinetics of IL-33 in liver transplant patients strikingly matched the ones in the animal model, as attested by serum levels reaching a peak immediately after reperfusion, which correlated to clinical outcomes including postreperfusion syndrome, posttransplant renal failure, graft dysfunction, and histological lesions of ischemia/reperfusion injury. IL-33 was an independent factor of graft dysfunction with a cutoff of IL-33 at 73 pg/ml after reperfusion (73% sensitivity, area under the curve of 0.76). Taken together, these findings establish the immediate implication of IL-33 acting as an alarmin in liver I/R injury and provide evidence of its close association with cardinal features of early liver injury-associated disorders in LT patients.

Transcriptomics Underlying Pulmonary Ozone Pathogenesis Regulated by Inflammatory Mediators in Mice.

Ozone (O3) is the predominant oxidant air pollutant associated with airway inflammation, lung dysfunction, and the worsening of preexisting respiratory diseases. We previously demonstrated the injurious roles of pulmonary immune receptors, tumor necrosis factor receptor (TNFR), and toll-like receptor 4, as well as a transcription factor NF-κB, in response to O3 in mice. In the current study, we profiled time-dependent and TNFR- and NF-κB-regulated lung transcriptome changes by subacute O3 to illuminate the underlying molecular events and downstream targets. Mice lacking Tnfr1/Tnfr2 (Tnfr-/-) or Nfkb1 (Nfkb1-/-) were exposed to air or O3. Lung RNAs were prepared for cDNA microarray analyses, and downstream and upstream mechanisms were predicted by pathway analyses of the enriched genes. O3 significantly altered the genes involved in inflammation and redox (24 h), cholesterol biosynthesis and vaso-occlusion (48 h), and cell cycle and DNA repair (48–72 h). Transforming growth factor-β1 was a predicted upstream regulator. Lack of Tnfr suppressed the immune cell proliferation and lipid-related processes and heightened epithelial cell integrity, and Nfkb1 deficiency markedly suppressed lung cell cycle progress during O3 exposure. Common differentially regulated genes by TNFR and NF-κB1 (e.g., Casp8, Il6, and Edn1) were predicted to protect the lungs from cell death, connective tissue injury, and inflammation. Il6-deficient mice were susceptible to O3-induced protein hyperpermeability, indicating its defensive role, while Tnf-deficient mice were resistant to overall lung injury caused by O3. The results elucidated transcriptome dynamics and provided new insights into the molecular mechanisms regulated by TNFR and NF-κB1 in pulmonary subacute O3 pathogenesis.

IL-37 regulates allergic inflammation by counterbalancing pro-inflammatory IL-1 and IL-33.

Children with asthma have impaired production of interleukin (IL) 37; in mice, IL-37 reduces hallmarks of experimental allergic asthma (EAA). However, it remains unclear how IL-37 exerts its inhibitory properties in asthma. This study aimed to identify the mechanism(s) by which IL-37 controls allergic inflammation. IL-37 target cells were identified by single-cell RNA-seq of IL-1R5 and IL-1R8. Airway tissues were isolated by laser-capture microdissection and examined by microarray-based gene expression analysis. Mononuclear cells (MNC) and airway epithelial cells (AECs) were isolated and stimulated with allergen, IL-1β, or IL-33 together with recombinant human (rh) IL-37. Wild-type, IL-1R1- and IL-33-deficient mice with EAA were treated with rhIL-37. IL-1β, IL-33, and IL-37levels were determined in sputum and nasal secretions from adult asthma patients without glucocorticoid therapy. IL-37 target cells included AECs, T cells, and dendritic cells. In mice with EAA, rhIL-37led to differential expression of >90genes induced by IL-1β and IL-33. rhIL-37 reduced production of Th2 cytokines in allergen-activated MNCs from wild-type but not from IL-1R1-deficient mice and inhibited IL-33-induced Th2 cytokine release. Furthermore, rhIL-37 attenuated IL-1β- and IL-33-induced pro-inflammatory mediator expression in murine AEC cultures. In contrast to wild-type mice, hIL-37had no effect on EAA in IL-1R1- or IL-33-deficient mice. We also observed that expression/production ratios of both IL-1β and IL-33 to IL-37 were dramatically increased in asthma patients compared to healthy controls. IL-37 downregulates allergic airway inflammation by counterbalancing the disease-amplifying effects of IL-1β and IL-33.

Anti-IL5 mepolizumab minimally influences residual blood eosinophils in severe asthma.

Neutralising antibodies against the cytokine interleukin (IL)5 have become widely used for the control of severe eosinophilic asthma. Remarkably, patients receiving neutralising anti-IL5 biological therapies retain a very stable population of residual blood eosinophils. Whether these residual eosinophils are endowed with particular biological activity has not yet been studied, but is of importance in predicting potential long-term effects of IL5 neutralisation in patients. To tackle the effect of IL5 depletion on residual eosinophils, we used a comparative RNA-sequencing approach and compared the gene expression programme of eosinophils arising in IL5-depleted or IL5-replete human or murine hosts, at steady-state in vivo and following in vitro stimulation with the eosinophil-activating alarmin IL33. We compared blood eosinophils from patients with severe allergic eosinophilic asthma treated with anti-IL5 mepolizumab therapy to those of healthy controls and matched asthma patients receiving anti-IgE omalizumab therapy. We made similar comparisons on bone marrow eosinophils from mice genetically deficient or not for IL5. We report that restriction of IL5 availability did not elicit any detectable transcriptional response in steady-state residual eosinophils in mepolizumab-treated patients or IL5-deficient mice, and influenced only a handful of genes in their response to IL33. Together, these results support the notion that treatment with IL5 neutralising antibodies spares a pool of circulating residual eosinophils largely resembling those of healthy individuals.

Targeting sorting nexin 10 improves mouse colitis via inhibiting PIKfyve-mediated TBK1/c-Rel signaling activation

Sorting nexin 10 (SNX10) has been reported as a critical regulator in macrophage function, and germline SNX10 knockout effectively alleviated mouse colitis. Here, we investigated the precise role of SNX10 in inflammatory responses in macrophages in mouse colitis, and explored the druggability of SNX10 as a therapeutic target for inflammatory bowel disease (IBD). Our results revealed that myeloid-specific SNX10 deletion alleviated inflammation and pathological damage induced by dextran sulfate sodium (DSS). In vitro experiments showed that SNX10 deletion contributed to inflammation elimination by inhibiting PIKfyve-mediated TANK-binding kinase 1 (TBK1) /c-Rel signaling activation. Further study provided rational mechanism that SNX10 was required for the recruitment of PIKfyve to the TRIF-positive endosomes, through which PIKfyve activated TBK1/c-Rel for LPS-induced inflammation response. Based on the structure of SNX10, we discovered a new small-molecule inhibitor DC-SX029, which targeted SNX10 to block the SNX10-PIKfyve interaction, thereby decreased the TBK1/c-Rel signaling activation. Additionally, therapeutic efficiency of DC-SX029 was evaluated in both DSS-induced and IL10-deficient mouse colitis models. Our data demonstrate a new mechanism by which SNX10-PIKfyve interaction regulates LPS-induced inflammation response in macrophages via the TBK1/c-Rel signaling pathway. In vivo and in vitro pharmacological studies of SNX10 protein-protein interaction (PPI) inhibitor DC-SX029 demonstrate the feasibility of targeting SNX10 in IBD treatment.

Minimal role of interleukin 6 and toll-like receptor 2 and 4 in murine models of immune-mediated bone marrow failure.

Immune aplastic anemia (AA) results from T cell attack on hematopoietic cells, resulting in bone marrow hypocellularity and pancytopenia. Animal models have been successfully developed to study pathophysiological mechanisms in AA. While we have systemically defined the critical components of the adaptive immune response in the pathogenesis of immune marrow failure using this model, the role of innate immunity has not been fully investigated. Here, we demonstrate that lymph node (LN) cells from B6-based donor mice carrying IL-6, TLR2, or TLR4 gene deletions were fully functional in inducing severe pancytopenia and bone marrow failure (BMF) when infused into MHC-mismatched CByB6F1 recipients. Conversely, B6-based recipient mice with IL-6, TLR2, and TLR4 deletion backgrounds were all susceptible to immune-mediated BMF relative to wild-type B6 recipients following infusion of MHC-mismatched LN cells from FVB donors, but the disease appeared more severe in IL-6 deficient mice. We conclude that IL-6, TLR2, and TLR4, molecular elements important in maintenance of normal innate immunity, have limited roles in a murine model of immune-mediated BMF. Rather, adaptive immunity appears to be the major contributor to the animal disease.

Ustekinumab for Ulcerative Colitis

Ustekinumab for Ulcerative Colitis

TRANSFER RNA QUEUOSINE MODIFICATION ENZYME MANIPULATES TIGHT JUNCTION PROTEINS IN INFLAMMATORY BOWEL DISEASE

Abstract Background Transfer RNA (tRNA) queuosine (Q)-modification occur at the wobble anticodon position of four special cellular tRNAs. In eukaryotes, tRNA-Q modification relies on the intestinal microbial product queuine and eukaryotic tRNA-guanine transglycosylase complex contains of Q tRNA ribosyltransferase catalytic subunit 1 (QTRT1). Q-tRNA modification is critical for the fidelity and accuracy to translate RNA to protein. It is known that dysfunction of Q-tRNA associates with cancer proliferation and malignancy. However, mechanisms of how Q and Q-tRNA modifications influence intestinal epithelial biology and chronic inflammation are unknown. Methods We explored the expression of QTRT1 in patients with IBD by investigating human biopsy tissues and re-analyzing gene expression datasets (with both UC and CD patients). To further investigate the QTRT1 role in IBD, the dextran sodium sulfate (DSS) induced colitis mouse model and IL10-deficient mouse model were used to evaluate the molecular mechanism. We further investigated the impact of QTRT1 reduction on intestinal cells by generating QTRT1 knockdown HCT116 and Caco-2 BBE cells using a Double Nickase Plasmid. Results We found that the QTRT1 expression in IHC staining was significantly downregulated in the colon of both Ulcerative Colitis (UC) and Crohn’s Disease (CD) patients compared with normal controls. After extracting and analyzing the datasets, we found the gene expression of QTRT1 was significantly suppressed in colonic mucosa of UC and intestinal mucosa of CD, compared with controls. Moreover, the four tRNA-Q-related tRNA synthetase (asparaginyl-tRNA synthetase, aspartyl-tRNA synthetase, histidyl-tRNA synthetase and tyrosyl-tRNA synthetase) were also decreased in IBD patients. The decreased QTRT1-expression was further confirmed in both DSS-induced colitis mice and IL10-defecient mice. Meanwhile, the significantly downregulated beta-catenin and Claudin-5, and upregulated Caludin-2 were identified in these two colitis models. All these molecular alterations, which are important in cell adhesion and tight junction formation, were further verified in protein expression and mRNA level using QTRT1 reduction HCT116 and Caco-2 BBE cells in vitro. Conclusion Our results demonstrate that the QTRT1 plays a novel role in human IBD by altering intestinal cell adhesion and junctions. Microbiome-dependent Q supply can be altered during disease development or when a limited variety of food-types are ingested. Investigations on the gut microbiome-derived tRNA modification in IBD will uncover novel molecular mechanisms for potential prevention and therapy.

Requirement of brain interleukin33 for aquaporin4 expression in astrocytes and glymphatic drainage of abnormal tau

Defective aquaporin4 (AQP4)-mediated glymphatic drainage has been linked to tauopathy and amyloid plaque in Alzheimer’s disease. We now show that brain interleukin33 (IL33) is required for regulation of AQP4 expression in astrocytes, especially those at neuron-facing membrane domain (n-AQP4). First, IL33-deficient (Il33−/−) mice showed a loss of n-AQP4 after middle age, which coincided with a rapid accumulation of abnormal tau in neurons and a reduction in drainage of abnormal tau to peripheral tissues. Second, injection of recombinant IL33 induced robust expression of AQP4 at perivascular endfoot (p-AQP4) of astrocytes, but not n-AQP4, in Il33−/− brains. Although the increased p-AQP4 greatly accelerated drainage of intracerebroventricularly injected peptides, it did not substantially accelerate drainage of abnormal tau. These results suggest that p-AQP4 drives overall convective flow toward perivenous space, i.e., glymphatics, whereas n-AQP4 may generate an aqueous flow away from neurons to remove neuronal wastes, e.g., abnormal tau. We have previously shown the role of brain IL33 in DNA repair and autophagy in neurons with oxidative stress. Now, we show that IL33 deficiency also impairs glymphatic drainage. Defects in those mechanisms together may lead to chronic neurodegeneration and tauopathy at old age in IL33-deficient mice.

IL-23 Contributes to Campylobacter jejuni-Induced Intestinal Pathology via Promoting IL-17 and IFNγ Responses by Innate Lymphoid Cells.

Human pathogen Campylobacter jejuni is a significant risk factor for the development of long-term intestinal dysfunction although the cellular and molecular mechanisms remain scantily defined. IL-23 is an emerging therapeutic target for the treatment of inflammatory intestinal diseases, however its role in C. jejuni-driven intestinal pathology is not fully understood. IL-10 deficient mice represent a robust model to study the pathogenesis of C. jejuni infection because C. jejuni infection of mice lacking IL-10 results in symptoms and pathology that resemble human campylobacteriosis. To determine the role of IL-23 in C. jejuni-driven intestinal inflammation, we studied the disease pathogenesis in IL-23-/- mice with inhibited IL-10Rα signaling. These mice exhibited reduced intestinal pathology independent from bacterial clearance. Further, levels of IFNγ, IL-17, IL-22, TNF, and IL-6 were reduced and associated with reduced accumulation of neutrophils, monocytes and macrophages in the colon. Flow cytometry analysis revealed reduced production of IL-17 and IFNγ by group 1 and 3 innate lymphoid cells. Thus, our data suggest that IL-23 contributes to intestinal inflammation in C. jejuni infected mice by promoting IL-17 and IFNγ production by innate lymphoid cells.

Role of Interleukin33 in Rejuvenation of Aged Neurons and Age-Related Dementias.

Late-onset Alzheimer’s disease (LOAD) is the most common age-related dementia, and its etiology remains unclear. Recent studies have linked abnormal neuronal aging to LOAD. Neurons are non-proliferative, and thus, majority of aged neurons must be rejuvenated through repairing or eliminating damaged molecules to regain their healthy status and functionalities. We discovered a surge of oxidative stress in neurons at middle age in mice. A rapid upregulation of neuronal rejuvenation is vital, while astrocyte-expressed interleukin33 (IL33), an IL1-like cytokine, is critical for this process. Thus, IL33-deficiency cripples the neuronal rejuvenation mechanisms, such as repairing DNA double strand breaks, eliminating damaged molecules by autophagy or by glymphatic drainage. IL33-deficient mice develop tau deposition and age-related dementia following a path similar to LOAD. We hypothesize that any interferences on IL33-initiated rejuvenation process for aged neurons after middle life is a potential risk for LOAD development.