bluuebunny/crossref_metadata_2025 huggingface dataset:
Egr2 deficiency significantly impairs the development of pathogenic TH17 cells
Authors: Gao Yuanyuan , Wang Yan , Guan Yukun , Bouladoux Nicolas Jean , Break Timothy , Lionakis Michail S , Belkaid Yasmine , Lazarevic Vanja
Published: January 2023

TH17 cells have critical functions in host defense at mucosal barriers, but they can also be the key drivers of immunopathology. Since their discovery, multiple transcription factor networks have been described that direct Th17 lineage commitment; however, molecular switches that control the development of “pathogenic” versus “non-pathogenic” TH17 cells remain largely unknown. In this study, we have identified the transcription factor Egr2 as a critical regulator of the TH17 pathogenic program. When ectopically expressed under TH17-polarizing conditions, Egr2 significantly enhanced the expression of TH17 signature genes in a RORγt-dependent manner. Although Egr2-deficient CD4+ T cells could be polarized in vitro into TH17 cells, these cells lacked the pathogenic potential in a TH17-dependent disease model, experimental autoimmune encephalomyelitis (EAE). In both active EAE and passive EAE, Egr2-deficeint TH17 cells failed to induce neuroinflammatory response in the CNS. Analysis of CNS infiltrates revealed reduced accumulation of IL-17A-producing CD4+ T cells and impaired recruitment of monocytes and myeloid dendritic cells in mice with T cell-specific loss of Egr2. Most importantly, transcriptional analysis of ex vivo isolated Egr2-deficient CD4+ T cells revealed that Egr2 controlled the expression of pathogenicity-associated genes in TH17 cells in the CNS. We further show that Egr2 deficiency did not affect the effector function of protective TH17 cells at mucosal barriers in Citrobacter rodentium and Candida albicans infections, suggesting that Egr2 is an attractive candidate for the therapeutic targeting of pathogenic TH17 cells while preserving tissue-protective functions of TH17 cells at barrier sites.
Persistent antigen can maintain lung resident memory CD8 T cells long-term
Authors: Uddback Ida Elin Maria , Cartwright Emily , Schøller Amalie Skak , Hayward Sarah L , Lobby Jenny , Takamura Shiki , Thomsen Allan R , Kohlmeier Jacob E , Christsensen Jan Pravsgaard
Published: January 2023

A sub-population of memory T cells, tissue resident memory T cells (TRM), have in recent years gained attention for their indispensable role in heterologous protection against respiratory viruses. For this reason, they are of great interest for vaccine research. However, the number of lung TRM decline within a few months after infection and researchers have been unable to determine how the TRM population, and protection, can be made to last long-term in the lung. We have previously shown that an Adenovirus expressing nucleoprotein (AdNP) from influenza A virus induce CD8 T cell mediated heterosubtypic immunity lasting up to at least 8 months post vaccination when administered locally and systemically, but the underlying reason was not investigated.

Here, we show that the number of CD8 T cell in the lung induced by AdNP is significantly higher compared to cells induced by an influenza infection. Using AdNP, we show that CD8 TRM in the lung can be maintained for at least one year post-vaccination. Our results revealed that lung TRM continued to proliferate in-situ 8 months after AdNP vaccination. Importantly, this required pulmonary vaccination and antigen persistence in the lung, as non-respiratory vaccination routes failed to support lung TRM maintenance. Additionally, parabiosis experiments show that in AdNP vaccinated mice the lung TRM pool is sustained by continual replenishment from circulating memory CD8 T cells that differentiate into lung TRM, a phenomenon not observed in influenza infected parabiont partners. Concluding, these results demonstrates requirements for long-lived cellular immunity to influenza virus, knowledge that could be utilized in future vaccine design.
Aire deficiency during early pregnancy leads to maternal T cell imbalance and embryo loss
Authors: Gillis-Buck Eva Mae , Gardner James M. , Bautista Jhoanne L. , Anderson Mark S. , Erlebacher Adrian , MacKenzie Tippi C.
Published: January 2023

Healthy pregnancy relies on maternal tolerance to a unique set of pregnancy associated self-antigens (PAAs), which are encoded in the maternal genome, but have not been produced since the mother herself was a fetus with a placenta. We hypothesize that Aire-mediated expression of PAAs promotes deletion of PAA-reactive T cells and induction of PAA-specific Tregs, and that Aire-deficiency may play a role in pregnancy complications. We used an Aire-diphtheria toxin receptor (DTR) transgenic mouse model to ablate maternal Aire-expressing cells only during the first nine days of pregnancy. AireDTR+DT plugged females were 6.5 times as likely to show complete embryo resorption by E9.5, compared to WT+DT dams (RR 6.5; 95% CI 1.61–26.1; p<0.01), and twice as likely to show no signs of implantation at E9.5 compared to WT+DT dams (RR 2.0; 95% CI 1.05–3.94; p<0.05). AireDTR+DT dams had significantly fewer FoxP3+ Tregs (p<0.0001) and more CD4+ T cells (p<0.0001) in the maternal thymus, and increased frequency of CD3+ CD4− CD8− T cells in the uterus (p<0.05). AireKO mice are known to develop ovarian insufficiency and subsequent progesterone (P4) deficiency. However, we found no significant differences in serum P4 levels of AireDTR+DT compared to WT+DT dams. P4 supplementation failed to prevent embryo resorption in AireDTR+DT dams or to change the alteration in thymic Tconv:Treg. Thus, Aire deficiency during early pregnancy leads to maternal T cell imbalance and embryo loss without inducing ovarian insufficiency, suggesting a novel mechanism for autoimmune mediated infertility.
Absence of conventional dendritic cells type 1 (cDC1) impairs intestinal homeostasis and permeability and leads to the development of obesity
Authors: Hamade Hussein , Stamps Jamsine T , Thomas Lisa S , Stamps Dalton , Shimodaira Yosuke , Michelsen Kathrin S
Published: January 2023

The maintenance of intestinal homeostasis depends on a complex interaction between microbiota, intestinal epithelial barrier and immune system. Alteration in of one of these components could lead to the development of chronic inflammatory diseases such as inflammatory bowel diseases (IBD). Recent studies have associated obesity with more severe IBD. In obesity, pro-inflammatory macrophages accumulate in adipose tissue and their numbers correlate with inflammation and insulin resistance. BATF3 is a transcription factor implicated in the development of conventional dendritic cells type 1 (cDC1). Here, we show that aged Batf3−/− mice developed obesity as characterized by increased body weight, fasting glucose levels (Batf3−/−: 213, WT: 115 mg/dl, p < 0.005), size of abdominal white adipocyte, and development of hepatosteatosis. We also observed increased intestinal permeability in Batf3−/− mice (FITC-dextran gavage; p < 0.01) suggesting impaired epithelial barrier function. Feeding high-fat diet for 2 months further enhanced the metabolic phenotype in Batf3−/− mice, leading to increased fasting glucose levels (300 vs. 246 mg/dl, p < 0.05), and increased adipocyte size. Moreover, during high-fat diet Batf3−/− mice were more susceptible to dextran sulfate sodium (DSS) colitis with more severe cecal inflammation, increased gut and mesenteric fat M1 macrophages (p < 0.05). Our data suggest that deficiency of the transcription factor BATF3 and lack of cDC1 alters the intestinal epithelial barrier, promotes the development of obesity, and increases susceptibility to DSS. In conclusion, BATF3-dependent cDC1 play an important role in maintaining the intestinal epithelial barrier and are protective for the development of obesity.
Isolation of mouse CD45 positive leukocytes from tissues
Authors: Antignano Frann , Poon Grace , Facca Valerie , Separovic Lea , Valdez Yanet , Kokaji Andy I , Woodside Steven M. , Eaves Allen C. , Thomas Terry E.
Published: January 2023

Immune cell function is often tissue-specific, therefore isolating cells from their tissue microenvironment is necessary to better understand their role in health and disease. This can be challenging in the presence of non-immune, tissue-derived cells and cellular debris from tissue dissociation. These factors reduce the leukocyte start frequency, which can prolong the isolation process and limit the identification of small but critical leukocyte subsets. To overcome these obstacles, we developed a simple and rapid selection method to enrich for CD45+ leukocytes from mouse tissues. Starting with a single cell suspension, cells are labelled with an antibody complex that links CD45+ cells to magnetic particles and following magnetic separation, the isolated CD45+ cells are ready for use. Using healthy mouse lung tissue as an example, CD45+ leukocytes were enriched from 63.5 ± 9.4% to 97.1 ± 1.2% purity, and the recovery of viable CD45+ cells was 37.7 ± 14.5% (mean ± SD, n=37). Preliminary testing on tumors from a mouse 4T1 breast tumor model resulted in efficient enrichment of tumor-infiltrating leukocytes. Importantly, the composition of immune subsets from both healthy lung tissues and tumor samples was maintained following CD45 selection. To demonstrate functionality, T cells within the CD45 selected population from spleen were able to upregulate CD25 and CD69, and proliferate upon stimulation. In as little as 20 minutes, the EasySep™ Mouse CD45 Isolation Kit allows researchers to easily enrich for functional leukocytes from tissue samples. Examining immune cells directly from healthy and tumor tissues will enhance our understanding of how these cells function and aid in developing new approaches to combat disease progression.
Identifying predictive cytokine biomarkers in asthma and chronic obstructive pulmonary disease using the Invitrogen ProQuantum high-sensitivity immunoassay platform
Authors: Bourdon David M , Ekoff Helena , Sjölander Anders , Fujisawa Takao , Nagao Mizuho , Molin Magnus
Published: January 2023

Asthma and chronic obstructive pulmonary disease (COPD) affect hundreds of millions of people worldwide and are a significant cause of morbidity and mortality. These heterogenous and complex inflammatory diseases have few good biomarkers for disease etiology, diagnosis, and monitoring of treatment efficacy. Characterization has previously been performed at genomic and proteomic levels, as well as airway histology, lung function, and quality of life metrics. Since inflammation typically involves one or more cytokine cascades, or pathways responsible for triggering and maintaining such inflammation, cytokine profiles are critical to advancing our understanding of the disease. However, a powerful immunoassay platform with a simplified workflow that is capable of early cytokine detection and minimum sample consumption is lacking in the asthma/COPD research space.

The goal of the following study was to determine whether the Invitrogen ProQuantum high-sensitivity immunoassay platform (www.thermofisher.com/proquantum) could prove useful in uncovering biomarker profiles in asthmatic populations. Additional benefits included running the proteomic assay on ubiquitous qPCR instrumentation and the small 2 μL sample requirement. In this study, 168 human serum samples were analyzed (IL-5, IL-6, IL-8, IL-13, IFN-γ and eotaxin-1) across wheezing and asthmatic phenotypes, as well as healthy age-matched controls among both cohorts. Data presented include group-wise analyses that indicate that the ProQuantum high-sensitivity immunoassay has helped uncover predictive biomarkers in samples from wheezing and asthmatic children, that were not previously distinguishable using more traditional immunoassay testing platforms.
Immunoregulatory effects of Mesenchymal Stem Cells-Derived Exsomes in a Rabbit Model of Autoimmune Dacryoadenitis
Authors: Nian Hong , Li Na , Wei Yankai , Zhao Lu , Chen Sisi , Wei Ruihua
Published: January 2023

We previously demonstrated that mesenchymal stem cells (MSCs) ameliorated autoimmune dacryoadenitis in rabbits. Recently, MSC-derived exsomes (MSC-Exo) were considered to carry functions of MSCs. In this study, we investigated the effect of local administration of human MSC-Exo on rabbit autoimmune dacryoadenitis. Diseased rabbits were treated with MSC-Exo or PBS via subconjunctival injection on days 1, 3, 5, 7 and 9 after infusions of activated autologous lymphocytes, and clinical scores were determined by assessing tear production, break-up time and fluorescein. The in vitro effects of MSC-Exo on activated peripheral blood mononuclear cells (PBMCs) were determined by BrdU proliferation assay and Q-PCR assessing the mRNA expression levels of T cell or macrophage related cytokine genes and subpopulation-related marker genes. We found that subconjunctivally administered MSC-Exo greatly reduced the severity of autoimmune dacryoadenitis as their parent cells by reducing the infiltration of T cell subsets and other inflammatory-related factors in lacrimal glands. In vitro, the proliferation of autoreactive T cells was inhibited by MSC-Exo in a dose-dependent manner. MSC-Exo treatment significantly down-regulated the expression of M1 macrophage-related gene, while promoted the expression of anti-inflammatory M2 macrophage related genes. These results suggests that MSC-Exo effectively ameliorated autoimmune dacryoadenitis partially by inducing the generation of M2 macrophage, indicating a potential novel therapy of MSC-Exo for autoimmune dacryoadenitis.
N-acetylcysteine reduces reactive oxygen species and pro-inflammatory cytokine production in HLA-B27 positive macrophages and shapes them towards a quiescent metabolic state
Authors: Navid Fatemeh , Nguyen Breanna , Cougnoux Antony , Violet Pierre-Christian , Levine Mark , Colbert Robert A
Published: January 2023

HLA-B27, which predisposes to spondyloarthritis (SpA), has a tendency to misfold, generate endoplasmic reticulum (ER) stress, and activate the unfolded protein response (UPR). The UPR may contribute to SpA pathogenesis by promoting the production of pro-inflammatory cytokines like IL-23 and TNF. During ER stress, cells produce more reactive oxygen species (ROS), which can further worsen the UPR and promote inflammation. Here, we examined whether reducing oxidative stress in HLA-B27+ macrophages affects pro-inflammatory cytokine production and UPR gene expression. Bone marrow-derived macrophages from HLA-B27-transgenic (Tg) treated with IFNg and LPS (stimulated) showed increased ROS production compared to HLA-B7-Tg and wild type (WT) control rats. This was accompanied by robust activation of the UPR only in HLA-B27-expressing cells. Treatment with N-acetylcysteine (NAC), a ROS scavenger, significantly reduced ROS levels in HLA-B27+ macrophages and strongly decreased the transcription of many pro-inflammatory cytokines including Il-23, Il-12, Tnf, Il-6 and Il-1b. The effect of NAC was further verified by ELISA assays available for rat IL-1b, IL-6, and TNFa. NAC treatment also partially reduced UPR markers including spliced Xbp1, and BiP, Chop, Manf and Derlin-3 in HLA-B27+ macrophages. Interestingly, NAC reduced metabolic activity in stimulated macrophages to levels similar to untreated control cells as measured by the Seahorse Assay. Our data demonstrate that reducing ROS levels with NAC ameliorates ER stress-related pro-inflammatory effects of HLA-B27 misfolding. Alteration of the oxidative stress pathway should be further explored for therapeutic intervention in SpA.
Interleukin-17 Drives Interstitial Entrapment of Tissue Lipoproteins in Experimental Psoriasis
Authors: Huang Paul , Zinselmeyer Bernd , Saunders Brian T , Swartz Melody A , Kim Brian S , Randolph Gwendalyn J
Published: January 2023

Lipoproteins trapped in arteries drive atherosclerosis. Extravascular low-density lipoprotein undergoes receptor uptake, whereas high-density lipoprotein (HDL) interacts with cells to acquire cholesterol and then recirculates to plasma. We developed photoactivatable apoA-I to understand how HDL passage through tissue is regulated. We focused on skin and arteries of healthy mice versus those with psoriasis, which carries cardiovascular risk in man. Our findings suggest that psoriasis-affected skin lesions program interleukin-17-producing T cells in draining lymph nodes to home to distal skin and later to arteries. There, these cells mediate thickening of the collagenous matrix, such that larger molecules including lipoproteins become entrapped. HDL transit was rescued by depleting CD4 + T cells, neutralizing interleukin-17, or inhibiting lysyl oxidase that crosslinks collagen. Experimental psoriasis also increased vascular stiffness and atherosclerosis via this common pathway. Thus, interleukin-17 can reduce lipoprotein trafficking and increase vascular stiffness by, at least in part, remodeling collagen.
The cross-talk between infiltrating macrophages and inflamed or malignant colonic epithelium promotes overexpression of ST6GALNAC1 and epithelial MUC1 tumor form MUC1-sTn
Authors: Cascio Sandra , Kvorjak Michael , Ahmed Yamine , Miller Michelle , Al Hashash Jana , Hartman Douglas , Miskov-Zivanov Natasa , Telmer Cheryl , Finn Olivera J
Published: January 2023

Patients with inflammatory bowel disease (IBD) have an increased risk of developing colitis-associated colon cancer (CACC). Despite the strong relationship between inflammation and cancer, the mechanistic events that contribute to the transition from IBD to CACC remain undefined. Changes in the glycosylation profile of the known oncoprotein MUC1 commonly occur in chronic inflammation and may facilitate progression to cancer. We used a transwell coculture model system to examine the effect of polarized macrophages, such as those contributing to inflammation, on the expression of glycosylation-related enzymes in colon cancer cells. We found that M2-like macrophages induce the expression of ST6GALNAC1 glycosyltransferase which, by adding sialic acid to O-linked GalNAc residues, promotes the formation of the tumor-associated MUC1-sTn glycoform. Cytokine antibody arrays and blocking antibody experiments revealed that high levels of IL-13 and CCL17, present in the conditioned medium of colon cells cocultured with M2 macrophages, activate ST6GALNAC1 expression in colon cancer cells. In silico and in vivo murine and human models of colitis and CACC showed that IL-13 induces the phosphorylation of STAT-6 that directly binds ST6GALNAC1 promoter resulting in its transcription activation and protein over-expression. On the other hand, CCL17 activates ST6GALNAC1 expression via NF-kB pathway signaling. Our findings revealed a novel cross-talk between M2-like macrophages and inflamed and malignant colon cells that contributes to the pathogenesis of colitis and progression to CACC.
Antigen processing and autophagy function in adjuvant treated dendritic cell
Authors: Hashimoto Kahoko , Ootomo Takuya , Hasegawa Akihiro , Kotera Ryoma , Kobayashi Daichi , Nakayama Toshinori
Published: January 2023

Dendritic cell (DC) and macrophage are the powerful antigen presenting cells. These show remarkable antigen processing function. It has been reported that antigen processing is enhanced by adjuvant. In DC and macrophage, LPS or IFNγ induces cell proliferation, cytokine production, antigen presentation to naive T cell. Among DC subset, CD8 positive DC enhances autophagy function especially in cross presentation. It is still nuclear how DC processes antigen by ubiquitin proteasome pathway or by autophagy pathway. To induce more effective immune response on vaccination or immune therapy, we study the effect of adjuvant treatment for autophagy function in DC and macrophage. Three types of different culture condition were used to establish DC subsets and compared to those function of macrophage. Then analyzed the antigen processing capacity as well as the autophagy function.

We induced DC in vitro culture with GM-CSF, GM-CSF with IL-4, GM-CSF with IL-15, then stimulated DCs or macrophage by using LPS or IFNγ for up to 24 hours before antigen uptake. On antigen uptake and processing, GM-CSF induced or GM-CSF with IL-15 induced DC showed antigen processing, however GM-CSF with IL-4 induced DC showed active antigen processing within 60 min. Contrary to active antigen processing, GM-CSF with IL-14 –DC did not show any inducible effect after adjuvant treatment, but GM-CSF or GM-CSF with IL-15-DC increased the autophagy function. On the other hand, macrophage could not obtain autophagy effect thorough adjuvant treatment. DCs were studied the gene expression of cytokines, cell surface molecules such as co-stimulation molecules, mannose receptor, to evaluate antigen uptake capacity, processing pattern and autophagy.
Batf3-dependent Dendritic Cells are Required to Present Cell-associated Antigen to CD4 T Cells
Authors: Ferris Stephen T , Durai Vivek , Wu Renee , Murphy Theresa L , Murphy Kenneth M
Published: January 2023

Dendritic cells (DCs) are required to prime T cells during an immune response. The dogma in the field is that Batf3-dependent DCs (DC1s) prime CD8 T cells and IRF4-dependent DCs (DC2s) prime CD4 T cells. However, recent studies have shown that DC1s are required for induction of Th1 immune responses and to prime autroreactive CD4 T cells in the NOD mouse model of diabetes. Therefore, this segregation of priming theory is flawed. To further study the contribution of DC1s to CD4 T cell priming, we generated an XCR1-cre mouse and crossed it to the MHC class II floxed mouse to generate MHC class II deficient DC1s. We found that the form of antigen directs the priming of T cells. DC2s are superior at presenting soluble antigen as evidenced by normal CD4 and CD8 OVA specific T cell responses in mice lacking DC1s. However, DC1s are superior at presenting cell-associated antigen as evidenced by a lack of OVA specific CD4 and CD8 T cell priming in mice lacking DC1s. Furthermore, CD4 priming during tumor immune responses is absent when DC1s lack MHC class II. Our findings show that the DC subsets differ in the form of antigen that they present. DC1s present cell-associated material; whereas DC2s present soluble antigen.
Loss of gut barrier integrity triggers autoimmune diabetes through microbiota-induced activation of islet-reactive T cells
Authors: Falcone Marika , Sorini Chiara , Cosorich Ilaria , Conte Marta Lo , Facciotti Federica , Sanvito Francesca , Ferrarese Roberto , Canducci Filippo
Published: January 2023

Recent evidence suggests that the intestinal environment and, specifically, modifications of the microbiome profile, regulate the pathogenesis of extra-intestinal autoimmune diseases such as Type 1 Diabetes (T1D) by inducing intestinal inflammation and increasing gut permeability. Although low-grade intestinal inflammation and alterations of gut barrier integrity are found in humans and animal models of T1D, a direct causal link between enteropathy and triggering of beta cell autoimmunity is yet to be established. Here we show that breakage of the gut barrier integrity (by low-dose DSS administration) in BDC2.5XNOD mice carrying a transgenic TCR specific for a beta cell-autoantigen leads to activation of the islet-reactive T cell clone (BDC2.5) within the gut mucosa and onset of T1D. The intestinal activation of islet-reactive T cells requires the presence of gut microbiota and is abolished when mice are depleted of endogenous commensal microbiota. Importantly, we found that gut microbial antigens are directly capable to activate the diabetogenic BDC2.5 T cells with a TCR-mediated mechanism. Our results indicate that loss of the intestinal barrier continuity is directly responsible for activation of islet-specific T cells by commensal gut microbiota and provide a strong rationale to design innovative therapeutic interventions in “at risk” individuals aimed at restoring gut barrier integrity to prevent T1D.
Impact of genetic polymorphisms on human immune cell gene expression
Authors: Schmiedel Benjamin J , Singh Divya , Madrigal Ariel , Valdovino-Gonzalez Alan G , White Brandie M , Zapardiel-Gonzalo Jose , Ha Brendan , Altay Gokmen , Greenbaum Jason A , McVicker Graham , Seumois Grégory , Rao Anjana , Kronenberg Mitchell , Peters Bjoern , Vijayanand Pandurangan
Published: January 2023

While many genetic variants have been associated with risk for human diseases, how these variants affect gene expression in various cell types remains largely unknown. To address this gap, the DICE (database of immune cell expression, expression quantitative trait loci (eQTLs), and epigenomics) project was established. Considering all human immune cell types and conditions studied, we identified cis-eQTLs for a total of 12,254 unique genes, which represent 61% of all protein-coding genes expressed in these cell types. Strikingly, a large fraction (41%) of these genes showed a strong cis-association with genotype only in a single cell type. We also found that biological sex is associated with major differences in immune cell gene expression in a highly cell-specific manner. These datasets will help reveal the effects of disease risk-associated genetic polymorphisms on specific immune cell types, providing mechanistic insights into how they might influence pathogenesis (https://dice-database.org).
Idiopathic inflammatory myopathy patients with different autoantibody exhibit unique peripheral blood T cell subsets
Authors: Li Ju-Pi , Chang Shih-Hsin , Tsay Gregory J , Chen Der-Yuan , Lan Joung-Liang
Published: January 2023

Idiopathic inflammatory myopathy (IIM), a heterogeneous group of systemic autoimmune diseases, is characterized clinically by weakness of skeletal muscle. Patients with IIM often show a myositis-specific autoantibody (MSA) such as anti-melanoma differentiation-associated gene-5 (MDA5 group) or anti-aminoacyl-tRNA synthetase (ARS group, including Jo-1, EJ, PL-12, PL-7, OJ, and KS), indicating that the adaptive immune system may alter in IIM patients. T cells (CD3+) play important roles in regulation of the adaptive immune system and usually subdivide into naïve cells (TN) and memory T cells (TM). Based on the surface expression of different cell markers, TM cells can be divided into central memory T (TCM, CD45RA−CD62L+) and effector memory T (TEM, CD45RA−CD62L−) cells. After antigenic stimulation, a subset of TEM cells re-expresses CD45RA, called TTEMRA. Here we examined peripheral blood T cell subsets in MDA5 group (n=22), ARS group (n=34) and healthy control individuals (HC group, n=30) using flow cytometry analysis. We found that TTEMRA population was increased in MDA5 group. TN population was reduced and TTEMRA population was enhanced in ARS group. Th17 and Th1 cells were more abundant in MDA5 and in ARS group, respectively. Treg cells was increased in both of MDA5 and ARS group. In addition, in MDA5 group, death patients (n=5) had fewer Treg cells compared with alive patients (n=17), suggesting reduced Treg cells may be associated with mortality. Therefore, peripheral blood T cell subsets may be correlated with specific clinical manifestations for IIM patients with distinct MSA.
Novel roles of lipid mediators in psoriatic skin inflammation
Authors: Sumida Hayakazu , Kita Yoshihiro , Shimizu Takao , Sato Shinichi
Published: January 2023

Psoriasis is a common chronic inflammatory cutaneous disease thought to arise as a result of infiltration of inflammatory cells and activation of keratinocytes. Psoriasis has been considered as a classical Th1 disease; however, Th17 cells are attracting much interest. Previous knowledge on the pathology of psoriasis has been obtained mostly through the study of human data and samples. In addition, recently developed animal models for psoriatic skin inflammation recapitulates the hallmarks of human psoriasis and provide us new insights on psoriasis. Lipid mediators and their receptors regulate a variety of physiological processes and involved in multiple pathologies including immune-related diseases. Here, we explored novel lipid mediators involved in psoriatic skin inflammation using clinical samples and imiquimod-induced psoriasis model. In particular, mass spectrometry analysis for multiplex quantitation of eicosanoids using imiquimod-treated murine skin samples revealed unique kinetics of various lipid mediators in the time-series from early induction phase to chronic resolution phase. These data motivated us to focus on some lipid mediators such as leukotriene B4 (LTB4) and prostaglandin E2 (PGE2). As an example, BLT1 (known as LTB4 receptor) and CXCR2 have been revealed to coordinately promote neutrophil infiltration into the skin during the early phase of imiquimod-induced inflammation. Further analysis on PGE2 is in progress and some data indicates the novel functions of PGE2 and mPGES-1, an upstream enzyme for PGE2 production, in the pathogenesis in psoriasis. These insights may aid in the rational design of novel treatments for broad spectrum of inflammatory and immune disorders beyond psoriasis.
Role of CD137L in lumbar spinal cord microglial cytokine responses following sciatic nerve crush
Authors: Cao Ling
Published: January 2023

Previously, we showed that CD137 ligand (CD137L; AKA 4-1BBL) contributes to the development of behavioral hypersensitivity and functional recovery in mice following peripheral nerve injury-induced neuropathic pain. Mice that lack CD137L showed reduced sensory sensitivity and faster functional recovery. Here, we sought to further investigate the potential involvement of lumbar spinal cord microglial responses in CD137L-mediated behavioral changes following sciatic nerve crush. Lumbar spinal cord microglia were isolated from both wild type B6 and CD137L knockout (KO)_B6 mice via positive selection based on CD11b expression using magnetic beads at days 0 (naïve), 3, 7, 14, 28 and 56 following sciatic nerve crush or sham surgery. Following RNA extraction, qRT-PCR were performed to examine cytokine/chemokine expression in isolated microglia. Our preliminary results suggest that male mice had more measurable cytokine/chemokine responses than female mice. Despite that many cytokines are below the detection levels, we were able to observe increased expression of several chemokines, CCL2, CCL3, CCL4 and CCL5 in microglia from CD137L KO mice, which appeared to peak early following injury (Days 3–14) than that in microglia isolated from WT mice. Future study will determine the involvement of microglial chemokine responses in CD137L mediated sensory hypersensitivity and delayed functional recovery following peripheral nerve injury.
The effects of altered TGFβ signaling on human T follicular helper cell development and function
Authors: Schmiechen Zoe C , Weissler Katherine , Laky Karen , Guerrerio Pamela
Published: January 2023

T follicular helper (Tfh) cells are a subset of CD4+ T cells that play a crucial role in the development of memory B cells and plasma cells. T follicular regulatory (Tfr) cells oppose Tfh cells and suppress humoral immune responses. Patients with Loeys-Dietz Syndrome (LDS) type 1 and 2 have an autosomal dominant mutation in the genes encoding TGFβ Receptor 1 or 2, respectively, and have a strong predisposition to develop allergic diseases. We hypothesized that altered development and function of Tfh cells could contribute to increased IgE-mediated allergic phenotypes in LDS. Comparing a cohort of pediatric LDS patients with age-matched healthy volunteers, we found that LDS patients have significantly higher levels of serum IgE and IgG and lower levels of IgM. Congruently, LDS patients have an increased frequency of memory T cells, Th2 cells, and Tfh cells and a significantly lower frequency of Tfr cells. The increase in Tfh cells in LDS was restricted to the CCR4+ CCR6− subset, which has been associated with Th2 immune responses. In vitro, Tfh cells from LDS patients co-cultured with autologous naïve B cells demonstrated a greater ability to induce plasma cell differentiation compared to age-matched healthy volunteers. Additionally, naïve CD4+ T cells from LDS patients cultured in the presence of recombinant TGFβ1 and IL-12 or IL-4 were more likely to differentiate into Tfh cells compared to healthy age-matched controls, suggesting a cell-intrinsic propensity for naïve CD4+ T cells in LDS patients to acquire a Tfh fate. Collectively, our data suggest that LDS mutations alter the development and function of Tfh cells in a manner that promotes IgE-mediated disease.
FcγRIIB on CD11c+ cells alters liver and adipose tissue inflammation affecting glucose tolerance and atherosclerosis in female mice
Authors: Marvin Jennifer , Balsamo Joseph A , Rhoads Jillian P , Major Amy S
Published: January 2023

Circulating levels of oxidized lipoprotein (oxLDL) correlate with myocardial infarction (MI) risk. Previous work demonstrates that oxLDL immune complexes can signal through FcγRs on bone marrow-derived dendritic cells (BMDCs), enhancing their activation and inflammatory cytokine secretion. While global FcγR−/− studies show that activating FcγRs are proatherogenic, the role of inhibitory FcγRIIb is unclear. We sought to determine the role of DC-specific FcγRIIb on atherosclerosis and glucose tolerance, as both are important MI risk factors. Bone marrow chimeras were generated by rescuing lethally irradiated female Ldlr−/− mice with hematopoietic cells from littermate CD11c-Cre+or CD11c-Cre− Fcgr2bfl/fl donors. Four weeks after transplant, recipients were placed on a Western diet for eight weeks. Quantitation of atherosclerosis in the proximal aorta demonstrated a 58% increase CD11c− Cre+ Fcgr2bfl/fl recipients. We discovered that hepatic cholesterol, scavenger receptor CD36, and MHC Class II were increased on CD11c+CD11b+ cells in CD11c-Cre+ Fcgr2bfl/fl recipients. Furthermore, study of aortic draining lymph nodes revealed a 50% reduction in TREGS. To study glucose tolerance, six-week-old CD11c-Cre+ and CD11c-Cre− Fcgr2bfl/fl mice were placed on a 60 kcal% fat diet. Despite similar weight gain, CD11c-Cre+ Fcgr2bfl/fl mice were significantly more glucose intolerant after six weeks. Analysis of white adipose tissue (WAT) demonstrated a two-fold increase in pro-inflammatory AT DCs with increased MHC Class II and CD86. Collectively, these results demonstrate that the absence of FcγRIIb on CD11c+ cells results in increased liver and AT inflammation, influencing atherogenesis and development of glucose intolerance.
BeO-exposure induces B cell-mediated Noninfectious Granuloma Formation in the Lung
Authors: Atif Shaikh M , Mack Douglas , McKee Amy S , Rangel-Moreno Javier , Martin Allison K , Getahun Andrew , Maier Lisa A , Cambier John C , Tuder Rubin M , Fontenot Andrew P
Published: January 2023

Susceptibility to chronic beryllium disease (CBD)is linked to HLA-DP molecules possessing a glutamic acid at the 69thposition of the β-chain (βGlu69), with the most prevalent βGlu69-containing molecule being HLA-DP2. We have previously shown that exposure of HLA-DP2 transgenic mice to beryllium oxide (BeO) results in the development of mononuclear infiltrates in a peribronchovascular distribution and a beryllium-specific, HLA-DP2-restricted CD4+T cell response. In addition to T cells, B cells constituted a major portion of infiltrated leukocytes in the lung of BeO-exposed HLA-DP2 Tg mice and were localized within ectopic lymphoid aggregates and granulomas. B cell depletion had no effect on the development of Be-specific CD4+T cells. However, B cell depletion was associated with a loss of lymphoid aggregates and granulomas as well as a significant increase in lung injury in BeO-exposed mice compared to the isotype-treated group. Furthermore, B cell recruitment to the lung was independent of antigen and dependent on MyD88 signaling through the secretion of CXCL13. Adoptively-transferred WT B cells into BeO-exposed μMT mice significantly reduced lung injury. B cells also surrounded granulomas in transbronchial biopsies from CBD patients, confirming a role for B cells in CBD. Overall, our data suggest a novel modulatory role for B cells in the formation of granulomas and lymphoid aggregates, resulting in the protection of the lung against sterile particulate exposure.
Group A Streptococcal Infection, Acute Rheumatic Fever and Rheumatic Heart Disease in the Solomon Islands
Authors: Sika-Paotonu Dianne , Liligeto Urijah
Published: January 2023

Group A Streptococcal (GAS) infection of the upper respiratory tract (and possibly skin) can trigger Acute Rheumatic Fever (ARF), an autoimmune condition if left untreated. Severe or recurrent attacks of ARF that also remain untreated, can lead to a type of permanent cardiac damage known as Rheumatic Heart Disease (RHD). RHD remains a significant cause of morbidity and mortality globally and is rare in developed countries. In Australia, New Zealand (NZ) and the Pacific Region, the disease burden of ARF and RHD amongst Indigenous and Pacific communities remain one of the highest in the world, usually affecting children and young adults. Since a GAS vaccine remains unavailable, the most effective recommended treatment and management strategy to prevent further GAS infections that can lead to ARF and RHD constitute of painful monthly intramuscular injections of Benzathine Penicillin G (BPG). These BPG injections are administered for at least 10 years or more and are referred to as secondary prophylaxis (SP). Due to the pain, formulation and other issues associated with regular BPG injections, adherence to SP remains problematic globally. A new Penicillin that supports better adherence to SP is urgently needed. This work sought to further understand the local context in the Solomon Islands concerning GAS, ARF and RHD. Preliminary findings indicate very high ARF and RHD rates present in the local setting, in addition to issues with access to ARF/RHD prevention measures with BPG. An electronic database to monitor ARF and RHD as a national disease register will be of clinical relevance to the local setting in the Solomon Islands. Research efforts in the Pacific Region contributing towards the generation of a new Penicillin for BPG SP will be of benefit.
Prevention of erythrocyte fragility and anaemia induced by extracellular histones
Authors: Kordbacheh Farzaneh , O’Meara Connor H , Coupland Lucy A , Lelliott Patrick M , Parish Christopher R
Published: January 2023

Introduction

Neutrophil extracellular traps (NETs) are released from neutrophils following activation by pathogens and/or tissue injury and exhibit anti-microbial activity. Uncontrolled formation of NETs can, however, become pathogenic, particularly via their associated histones, and induce endothelial cell death, systemic vascular obstruction, multiple organ failure and initiate coagulation by both activating platelets and damaging erythrocytes such that they become pro-thrombotic.

Study Objectives

Recently we discovered an additional effect of histones on erythrocyte, which is to enhance their fragility when they are subjected to shear stress associated with blood flow through the vascular system and spleen in particular. Our laboratory has synthesised a family of small polyanions (SPAs) which have been screened for their ability to neutralise the pathogenic effects of histones and NETs.

Methods

We have used a novel mechanically-induced shear stress method as well as an in vitro spleen filtration model to investigate the effects of mammalian histones ± SPAs, on RBC fragility. The in vitro findings were mirrored in vivo with the injection of histones and SPAs in mice.

Results

Our results revealed that free histones bind to erythrocytes with high affinity and render erythrocytes susceptible to lysis by shear stress in a dose-dependent manner and some SPAs could totally inhibit or reverse this phenomenon.

Conclusions

Based on these data we propose that erythrocytes can neutralise the damaging effects of histones in the circulation and also transport them to the spleen for disposal. Overload of this disposal system could cause the unexplained anaemia associated with sepsis, cancer and other pathological conditions.
Inhibitory effects of Klotho hormone on antigen-specific clonal expansion of CD4+ T cells
Authors: Hwang Haun , Bamezai Anil K
Published: January 2023

Early signaling events through the antigen receptor in CD4+ T cells occur in and around the lipid microenvironment of the plasma membrane. The plasma membrane of all eukaryotic cells have phospholipids, cholesterol and transmembrane/lipid anchored proteins organized in small and dynamic ordered (Io) and disordered (Id) –like phases. Lipid rafts are nanometer-sized, cholesterol and saturated phospholipid-rich domains that contribute to the Io phase. Interaction between CD4+ T cells with antigen presenting cells during antigen presentation stabilize and/or promote coalescing of Io nanometer size domains that are normally scattered on the plasma membrane. To assess the role of membrane order in CD4+ T cell responses we used recombinant klotho protein that is known to bind glycolipids in membrane lipid rafts and destabilize these nanometer-sized domains. Klotho is a naturally occurring protein in mice and humans - the soluble form of this protein circulates in the body, and acts as a hormone to promote longevity in mice as well as enhance cognitive function in humans. However, its possible immunologic effects are unknown. We have examined the effects of recombinant klotho protein on CD4+ T cell responses in-vitro. We found that klotho hormone inhibits clonal expansion of CD4+ T cell in an antigen-specific manner, promotes apoptosis in CD4+ T cells from younger mice (< 8 weeks) although these inhibitory effects are minimal or not observed in CD4+ T cells from older mice (>8 weeks). -In-vivo experiments to further investigate the role of soluble klotho protein are underway, and will be reported.
Dysregulation of Lipid Metabolism during Gram-Negative Sepsis in Mkp-1 Deficient Mice
Authors: Batty Abel J , Li Jinhui , Wang Xiantao , Ackerman William E. , Kirk Sean G , White William M. , Wang Xianxi , Anastasakis Dimitrios , Samavati Lobelia , Buhimschi Irina , Nelin Leif D. , Hafner Markus , Liu Yusen
Published: January 2023

Mitogen-activated protein kinase phosphatase (Mkp)-1 exerts its anti-inflammatory activities during Gramnegative sepsis by deactivating p38 and JNK. We have previously shown that Mkp-1+/+ mice, but not Mkp-1−/− mice, exhibit hypertriglyceridemia and hepatic glycogen depletion during severe sepsis. However, the regulation of hepatic lipid stores and the underlying mechanism of lipid dysregulation during sepsis remains an enigma. To understand the molecular mechanism underlying the sepsis-associated metabolic changes and the role of Mkp-1 in the process, we infected Mkp-1+/+ and Mkp-1−/− mice with Escherichia coli i.v., and assessed the effects of Mkp-1 deficiency on tissue lipid contents. We also examined global gene expression profile by RNA-seq in the livers. We found that in the absence of E. coli infection, Mkp-1 deficiency decreased liver triglyceride levels. Upon E. coli infection, Mkp-1+/+ mice, but not Mkp-1−/− mice, developed hepatocyte ballooning and increased lipid deposition in the livers. E. coli infection caused profound changes in gene expression profile of a large number of proteins that regulate lipid metabolism in wildtype mice, while these changes were substantially disrupted in Mkp-1−/− mice. Interestingly, in Mkp-1+/+ mice E. coli infection resulted in down-regulation of genes that facilitate fatty acid synthesis but upregulation of CD36 and Dgat2, whose protein products mediate fatty acid uptake and triglyceride synthesis, respectively. Taken together, our studies indicate that sepsis leads to a profound change in metabolic gene expression programs and Mkp-1 plays an important role in this process.
The ssDNA exonucleases PLD3 and PLD4 are required to prevent lethal primary HLH-like disease in mice
Authors: Gavin Amanda L , Huang Deli , Nemazee David
Published: January 2023

Leukocyte sensing of microbial genetic material often elicits beneficial proinflammatory cytokines, but dysregulated responses can cause severe pathogenesis. Genome-wide association studies have linked Phospholipase D3 (PLD3) to Alzheimer’s disease and PLD4 to rheumatoid arthritis and systemic sclerosis. PLD3 and PLD4 are endolysosomal proteins with no detectable phospholipase activity. PLD4 deficient mice have a mild inflammatory disease, marked by elevated interferon-γ (IFN γ) and splenomegaly. These phenotypes were traced to an altered responsiveness of PLD4-deficient dendritic cells to ligands of the single-stranded DNA (ssDNA) sensor Toll-like receptor 9 (TLR9). Thioglycolate elicited peritoneal macrophages from PLD3-deficient mice also had exaggerated TLR9 responses to particular ligands. Although PLD4 and PLD3 were presumed to be phospholipases, we found that they are 5′-ssDNA exonucleases that break down TLR9 ligands.

Mice deficient in both PLD3 and PLD4 died pre-weaning from an inflammatory disease with many similarities to primary hemophagocytic lymphohistiocytosis (HLH). The levels of IFN γ, IL-6, TNF α, IL-10 and ferritin were markedly elevated in the serum of Pld3−/−Pld4−/− mice. Low blood platelets, severe anemia, liver steatosis, and hemophagocytosis were evident in Pld3−/−Pld4−/− mice. The lethal inflammation in PLD3 and PLD4 deficient animals was only marginally improved by IFN γ deficiency, however inactivation of UNC93B1 rescues their survival, revealing excessive endosomal TLR signaling is responsible for driving the disease.
Microbiota-derived butyrate promotes cellular metabolism and memory potential of antigen-activated CD8+ T cells
Authors: Bedoui Sammy , Binger Katrina , Kallies Axel , Turner Stephen John , McConville Malcolm , Gebhardt Thomas , Bachem Annabell
Published: January 2023

Microbiota modulate the immune system and recent studies suggest a functional relationship between microbiota, its metabolites and CD8+ T cells, but whether this link is also relevant for CD8+ T cell memory is unclear. We show that antigen-activated CD8+ T cells transferred into germ-free mice had transcriptional impairments in oxidative metabolism and failed to transition into memory cells with enhanced recall capacity. Conversely, the microbiotaderived metabolite butyrate promoted memory potential and secondary recall responses of activated CD8+ T cells. This was accompanied by metabolic rewiring of the T cells and uncoupling of the tricarboxylic acid cycle from glycolytic input, enabling preferential fuelling of enhanced oxidative phosphorylation through glutamine utilization and fatty acid catabolism. As such energetic adaptions promote memory T cell differentiation, our findings uncover an important role for microbiota-derived butyrate in rewiring the metabolism of activated CD8+ T cells in support of their long-term survival as memory cells.