bluuebunny/crossref_metadata_2025 huggingface dataset:
Single cell analysis of transitional B cells reveals autocrine IFNβ sustains a dynamic type I IFN network in lupus
Authors: Hamilton Jennie A , Yang PingAr , Liu Shanrun , Li Jun , Wu Qi , Luo Bao , Walter Mark R , Sanz Ignacio , Fish Eleanor N. , Chatham Winn , Hsu Hui-Chen , Mountz John D
Published: January 2023

Increased selection of transitional B cells reactive with nucleic acid antigens is a prominent feature of systemic lupus erythematosus (SLE). We found that transitional stage 1 (T1) B cells from both human SLE patients and BXD2 lupus mice intrinsically express significantly higher levels of IFNα and IFNβ compared to normal individuals. IFNβ expression in T1 B cells was significantly correlated with the development of mature autoreactive 9G4+ B cells in SLE patients and La+ B cells in BXD2 mice. IFNβ was also specifically required for optimal B cell survival and TLR7 responses. In single T1 B cells isolated from Ifnb−/− vs. WT B6 bone marrow chimeras, upregulation of Ifnα7 was completely abrogated in single Ifnb−/− T1 B cells, while other genes (Ifna1, Ifna4, Cd86, Tlr7) were significantly but incompletely diminished implicating both autocrine and paracrine IFNβ activity. To resolve these signals in an autoimmune environment, single cell gene expression analysis was carried out on sorted BXD2 T1 B cells. Hierarchical clustering revealed three distinct gene expression patterns. Cluster 1 cells were “IFN producers” characterized by high expression of type I IFN genes, but low co-expression of IFN receptor genes, Ifnar1 and Ifnar2. Cluster 2 “IFN responders” exhibited the highest co-expression of both Ifnar1 and Ifnar2 as well as immunomodulatory IFN response genes (Irg1, IL6, Cd69, Cd86). Cluster 3 had low expression of type I IFN and IFNAR genes. Clusters 1 and 2 exhibited the highest Tlr7 expression, while Cluster 3 had the highest expression of Baffr and Tlr9. The heterogeneity of IFN signatures at the T1 stage suggests that varying interferonogenic phenomena can differentially prime T1 B cells for subsequent responses and cell fates.
Estrogen promotes SLE serum-induced skin inflammation via the estrogen membrane receptor GPER1
Authors: Deng Guo-Min , Cai Zhenming , Xie Changhao , Guo Xuanxuan , Liu Huicheng , Li Xiaoyan , Fang Xiang , Fei Xibin , Guo Yacong , Xu Guangqiong , Dou Hui
Published: January 2023

Skin injury is the second most common clinical manifestation in patients with systemic lupus erythematosus (SLE). Estrogen may affect the onset and development of SLE. In this study, we investigated the role of estrogen and its membrane receptor GPER1 in SLE-related skin injury in mice treated with SLE serum in vivo, and monocytes from mouse spleen in vitro. We found that skin injury induced by SLE serum was more severe in female mice and required monocytes. Estrogen promoted these effects through the membrane receptor GPER1 located in lipid rafts and that inhibition of lipid rafts and GPER1 suppressed SLE serum-induced skin inflammation and expression of inflammatory molecules. From these studies, we conclude that estrogen promotes the development of skin injury induced by SLE serum through the membrane receptor GPER1 and that lipid rafts play an important role in the regulatory effect of GPER1 in SLE skin inflammation.
Leptin promotes plasma cell generation and exacerbates collagen-induced arthritis
Authors: CHEN QIAN , LU LIWEI
Published: January 2023

Plasma cells (PCs) contribute to the relapse of many autoimmune diseases, especially in patients with rheumatoid arthritis (RA) after rituximab treatment. Increasing evidence indicates a crucial role of leptin in autoimmune pathogenesis via enhancing CD4+ T cell responses, but whether and how leptin regulates PCs remain largely unclear. In collagen-induced arthritis (CIA) mice, we detected increased levels of leptin and anti-collagen II (CII) antibodies in the synovial fluid (SF) and sera of mice at acute and chronic stages of CIA. Higher percentage and cell number of plasma cells (PCs) were observed in the SF of CIA mice. Leptin receptor-deficient (db/db) mice exhibited ameliorated CIA development with reduced PCs responses and CII-specific antibody production. Intra-articular injection of recombinant leptin enhanced PCs responses and CII-specific antibody production and joint damage. Importantly, intra-articular injection of a soluble leptin blocker (ObR:Fc) deceased PCs responses, CII-specific antibody production and joint damage. Mechanistic studies revealed that leptin promoted CD138+IFR4+ PC generation from GL-7+Fas+ germinal center B cells via activating Akt-mTOR-IRF4 axis. Moreover, rapamycin treatment attenuated leptin-induced IRF4 expression and PCs generation. In RA patients, leptin levels and autoantibody production were positively correlated with disease activity (DAS28 score) with increased CD38+CD27+ plasmablasts detected in the synovium of active RA patients. Together, these findings demonstrated that leptin enhanced PCs responses via ObR-Akt-mTOR-IRF4 axis and exacerbated CIA development, indicating that leptin blockade may serve as a potential therapeutic target for the treatment of RA.
A Methodology and Application for Statistical Polarization in the Immune Response
Authors: Qeadan Fares , Dwivedi Sunita , Page Kimberly , Gravitt Patti
Published: January 2023

Polarization in the CD4+ mediated T-helper cell immune response is a very well characterized phenomenon in immunology; towards either Th1, Th2, or Th17 pathways. However, to-date, this concept has neither been developed nor demonstrated in a robust statistical framework. In this study, we provide a novel statistical methodology to facilitate the idea of polarization within a multivariate setting, and apply it to luminal fluid cytokine data of 90 women from the Peruvian Amazon. The panel of cytokines and chemokines for this study were assayed using separate BioRad Multiplex Array kits. In our application, we show that soil transmitted helminth (STH) infection induces statistically significant Th2 polarization in the female genital tract.

To provide statistical evidence of Th2 polarization in STH-infected women at the cervix, we compare the clustering of cytokines among STH-infected versus STH-uninfected women while examining the shift in the strength of correlation for Th2 cytokines with IL-4, which is seen as a central Th2 cytokine. The proposed statistical test is one that tests if, significantly, k>1 for f(L1)=f(L2) where L1 and L2 are the loadings in the principal component analysis (PCA) framework for the STH-infected and STH-infected groups respectively.

The proposed method is a confirmatory one and could be supported by visual representation of the Th polarization through heat maps. It has the potential to become the reference or gold standard when studying the polarization phenomenon in immunology as it tailors the clinical characteristics and mechanisms associated with the entire panel of cytokines and chemokines in the context of a system.
IL-17c promotes peripheral nerve growth during human herpes simplex virus type 2 (HSV-2) infection
Authors: Zhu Jia , Peng Tao , Chanthaphavong Savahn , Sun Sijie , Trigilio James , Jin Lei , Vazquez Julio , O’Day Diana , Glass Ian , Knipe David , Wald Anna , Corey Larry
Published: January 2023

During human HSV-2 infection, peripheral nerve destruction and sensory anesthesia are rarely developed, despite frequent virus reactivation. The mechanisms underlying this clinical observation are unclear. Here, we describe a novel interaction between HSV infected keratinocytes and the peripheral nerve system via IL-17c that promotes neurite growth and neuron survival during reactivations. Skin biopsies obtained during asymptomatic human HSV-2 reactivation exhibit a higher nerve fiber density in relation to biopsies obtained during the virological and clinical quiescent phase. Keratinocytes, the peripheral targets of HSV reactivation, produce IL-17c both in vivo during HSV-2 reactivation in humans and in vitro in primary cultures. Further characterization indicates that IL17RE, the receptor specific for IL-17c signaling, is expressed on nerve endings in human skin and on nerve fibers, glia cells and sensory neurons in human dorsal root ganglia. The addition of exogenous human IL-17c promotes neurite growth and branching and provides directional guidance in a microfluidic device. Furthermore, pretreatment of IL-17c reduces apoptosis in HSV-2 infected primary neurons. Taken together, these results suggest that IL-17c might function as a neurotrophic factor to induce peripheral nerve ending growth and protect neurons from apoptosis during HSV reactivation. This interaction provides a potential mechanism for the lack of clinical nerve damage from a lifelong recurrent neurotropic viral infection. It also highlights the role of immune cytokines in regulating the functions of the peripheral nervous system.
An unadjuvanted model for oral immunization revealing complex regulatory control of gut IgA responses to soluble protein
Authors: Gribonika Inta , Schön Karin , Strömberg Anneli , Lycke Nils
Published: January 2023

We have identified a unique mouse model in which oral immunization with soluble protein, without a potent mucosal adjuvant, is highly effective at stimulating strong gut IgA responses. In fact, cholera toxin (CT), which is the gold standard for oral adjuvants, did not enhance the gut IgA response to ovalbumin (OVA) in this model. As we dissected the mechanism allowing for an unadjuvanted strong gut IgA response we found that adoptively transferred OVA-peptide-specific TCR Tg DO11.10 CD4 T cells into nude Balb/c mice were sufficient to promote the gut IgA response. However, only CD4 T cells with a rearranged TCR a-chain, reactive to unknown, but possibly microbial, antigens in the gut, were effective, because SCID DO11.10 CD4 T cells failed to support a gut IgA response. Co-transfer of wild-type CD4 T cells with DO11.10 CD4 T cells into nude Balb/c hosts completely obliterated the response, but not if the CD4 T cells were isolated from IL-10−/− mice. Hence, the gut IgA response was under strict regulatory CD4 T cell control. By contrast, if CT adjuvant was admixed with OVA we completely rescued the gut IgA response following oral immunization. Thus, oral CT adjuvant primarily exerted a modulating effect on the regulatory CD4 T cell control in Peyer’s patches (PP), impacting on the balance between follicular helper and regulatory T cells. While both Th17 and CD25+ Tregs, have been claimed sufficient for supporting gut IgA responses, our model clearly suggests that the regulatory environment in PP is more complex, as will be discussed.
The microbiota-derived metabolite indole is an endogenous regulator of autophagy
Authors: Hunt Carrie , Zhao Xiao , Crisp Zeni , Choi Kyungoh , de Figueiredo Paul , Jayaraman Arul , Karpac Jason , Alaniz Robert Christopher
Published: January 2023

Autophagy is an evolutionarily conserved cellular mechanism that is crucial for promoting cell survival, pathogen clearance, and homeostasis. Dysregulated autophagy destabilizes GI tract homeostasis, promotes Crohn’s Disease, and perturbs the microbiota, a prominent feature during gut inflammation. Classically, energy substrate starvation induces cellular autophagy, yet host dietary intake makes the GI tract nutritionally abundant. Thus, the discrete gut signals able to induce autophagy and overcome the cues from a fully energy replete environment remain largely unknown. Here we demonstrate that indole, a strictly microbiota-derived metabolite, induces autophagy at a level comparable with the canonical autophagy-inducer rapamycin.

Consistent with the induction of autophagy, indole-conditioned dendritic cells exhibited increased LC3-II expression and puncta formation, enhanced bacterial killing, and reduced inflammasome activity. Indole effects were largely similar to rapamycin yet also distinct, suggesting a complementary mechanism of action for these molecules. Both indole and rapamycin suppressed mTOR signaling and IL-1beta production, suggesting a functional link. Unlike rapamycin, we have reported indole is a ligand of the aryl hydrocarbon receptor (AhR), yet regulation of autophagy by indole was AhR-independent. To our knowledge, this is the first evidence of a tryptophan microbiota metabolite functioning as a regulator of autophagy. These findings reveal novel biology by which the microbiota regulates gut autophagy and has therapeutic implications in inflammatory and autophagy-mediated disorders.
The differentiation and maintenance of RORγt+ Tregs respectively require integrin αvβ8 expression by dendritic cells and commensal microbiota
Authors: Chen Zhengtao , Wang Meixiang , Zhang Zhongyun , Liu Weixiu , Song Wengang , Tang Hua
Published: January 2023

The generation of RORγt+ regulatory T cells (Tregs), which regulate type 2 immune respond, is dependent on dendritic cells and microbiota, but mechanisms are poorly understood. Here we show that the differentiation of RORγt+ Tregs was critically dependented on integrin αvβ8 expressed in DCs. In mice, lack of αvβ8 in DCs resulted in loss of RORγt+ Tregs in the small intestine and mesenteric lymph nodes. Moreover, we also report that the maintenance of RORγt+ Tregs needs the existence of commensal bacteria but not DCs in small intestine. The percentage and the proliferation of RORγt+ Tregs were decreased after treated with antibiotic while these were unaffected by the absence of DCs in a short time. These date demonstrate that αvβ8 expressed in small intestinal DCs play a critical role in the induction of RORγt+ Tregs and microbiota act as a key factor on maintenance of RORγt+ Tregs, respectively. Our results may help to explain the mechanisms of oral tolerance via αvβ8 in DCs and commensal microbiota regulate the generation of RORγt+ Tregs to suppress type 2 immune respond.
Suppression of glial inflammatory responses by IL-24
Authors: Burmeister Amanda Rae , Marriott Ian
Published: January 2023

Maintenance of an immunoquiescent environment and/or resolution of inflammation is of critical importance within the confines of the central nervous system (CNS). The cytokine IL-10 has previously been demonstrated to play a critical role in resolving inflammation within the CNS and in suppressing damaging host responses. However, little is known regarding the role played by other IL-10 family members in the control of neuroinflammation. We have previously reported that bacterially challenged glia produce IL-10 and the related cytokine IL-19 in a delayed manner, and can respond to these cytokines by upregulating the expression of suppressor of cytokine signaling 3 (SOCS3), a key anti-inflammatory response gene. More recently, we have determined that murine astrocytes can similarly express another IL-10 family member, IL-24, in a delayed manner following activation, and constitutively express both subunits of the receptor for this cytokine. Here, we have investigated the ability of glial cells to respond to IL-24 and assessed its effect on the inflammatory responses of these cells to bacterial ligands. We demonstrate that recombinant murine IL-24 (rmIL-24) elicits the rapid expression of mRNA encoding SOCS3 by primary murine astrocytes. Importantly, we have determined that pretreatment with rmIL-24 significantly limits the production of the key pro-inflammatory cytokine IL-6 by bacterially challenged astrocytes. These findings are consistent with previous reports in peripheral tissues indicating that IL-24 acts to suppress host responses and provides another potential mechanism whereby resident CNS cells can limit detrimental neuroinflammation.
Palmitate induces interleukin-8 expression in human monocytic cells via TLR4/MyD88 dependent pathway
Authors: Ahmad Rasheed , AlRoub Areej Abu , Sindhu Sardar
Published: January 2023

Infiltrated immune cells in adipose tissue in obesity produce various cytokines and chemokines which are involved in metabolic inflammation and insulin resistance. Obese individuals are known to have increased IL-8 plasma level and it plays an important role in metabolic disorders. Since in obesity, the levels of circulatory saturated free fatty acid palmitate are increased and modulate the expression of other cytokines and chemokines, the role of palmitate in the regulation of IL-8 in monocytic cells is not well defined. Here, we provide the evidence that palmitate induces IL-8 expression at both mRNA and protein levels in human monocytic cells (P<0.05). Palmitate-mediated IL-8 secretion was suppressed either by neutralizing anti-TLR4 antibody or by genetic silencing of TLR4. Furthermore, MyD88−/− cells did not express IL-8 in response to palmitate. However, IRF3 deficiency did not affect IL-8 production. NF-κB/AP-1 activity was also elevated in palmitate-treated cells. Altogether, these results show that palmitate induces TLR4/MyD88 dependent activation of IL-8 gene expression leading to activation of NF-kB/AP1 transcription factors.
NLRP3 inflammasomes are required for MUC5AC expression induced by S100A8, S100A9 and S100A12
Authors: Chung Il Yup , Kim Karam , Kang Jin Hyun
Published: January 2023

S100A8, S100A9, and S100A12, damage-associated molecular pattern (DAMP) molecules, initiate and modulate local inflammation and innate immune responses. NLRP3 is a component of the innate immune system that detects danger signals. Activation of NLRP3 is known to be associated with the pathogenesis of pulmonary obstructive diseases such as asthma and COPD. We have previously demonstrated that these three S100 proteins are able to induce MUC5AC production in airway epithelial cells through activation of NF-kB and ERK pathways. In this study, we examine the involvement of NLRP3 inflammasomes in the S100 proteins-induced MUC5AC production. Treatment of NCI-H292 cells with the S100 proteins results in synthesis of NLRP3 mRNA and protein along with increased procaspase-1 and IL-1b secretion. Introduction of NLRP3 siRNA leads to a decrease in MUC5AC expression, which is also accompanied by diminished IL-1b mRNA. In addition, NLRP3 siRNA inhibits S100A12-induced activation of NF-kB. In line, MCC950, an inhibitor of NLRP3 inflammasome oligomerization, abolishes MUC5AC production, with no effect on synthesis of NLRP3 molecule. Collectively, these data suggest that NLRP3 inflammasome plays an active role in S100 protein-induced mucin production in airway epithelial cells.
Interleukin 7 promotes motility of naive T cells in the lymph node
Authors: Byrum Janie Rae , Fricke Matthew , Tafoya Justyna , Moses Melanie , Cannon Judy L
Published: January 2023

Naïve T cells interact with dendritic cells (DCs) in lymph nodes to become activated and effect a specific immune response. The chemokine CCL21 and lysophosphatidic acid (LPA) have both been demonstrated to be important for T cell movement within the T cell zone of the lymph node. However, the residual motility in the absence of both CCL21 signaling via CCR7 and LPA suggest that additional molecules can play a role in driving T cell movement within lymph nodes. Using two photon microscopy, we show interleukin-7 (IL-7) promotes T cell motility in vitro and in vivo, in addition to its known function as a survival factor. Blocking IL-7 signaling in the lymph node elicited a reduction in T cell speed. IL-7 treatment of primary T cells in vitro resulted in increased migration across a permeable barrier and small GTPase RhoA activation. We further demonstrate that IL-7 combines with CCL21 and LPA to promote naïve T cell motility. Our results indicate that IL-7 is a novel regulator of T cell motion in lymph nodes, in addition to its effect on T cell survival.
Inflammasomes promote the initiation of Brucella induced arthritis in an IL-18 dependent manner and mediate Brucella clearance independently of IFN-γ
Authors: Lacey Carolyn , Skyberg Jerod A
Published: January 2023

Brucellosis, caused by the intracellular bacterial pathogen Brucella, is a zoonotic disease for which infection of the joints is the most common focal complication in humans. The purpose of this study was to identify if Brucella activation of inflammasomes and in turn caspase1/11 activation is responsible for inducing arthritis. Here we report caspase-1/11−/− (casp1/11−/−) mice had reduced swelling, inflammation and inflammatory cytokine levels early in infection relative to wild-type “WT” mice. Later in infection, joint inflammation was similar between WT and casp1/11−/− mice, but bacterial loads and swelling were greater in casp1/11−/− joints. While IL-1Rdeficiency reduced joint swelling, this effect occurred later and was not of the same magnitude as casp1/11 deficiency, indicating an IL-1 independent role of casp1/11. Casp1/11 induced inflammation was partially dependent on IL-18, and IFN-γ levels were reduced in both casp1/11−/− and IL-18-deficient mice relative to WT mice early in infection. However, while IFN-γ and casp1/11 both played a role in controlling joint Brucella burdens, an additive effect of these deficiencies was observed indicating that the protective effects of IFN-γ and casp1/11 on bacterial clearance are not interdependent. Enhanced arthritis and musculoskeletal inflammation was observed in IFN-γ-deficient, relative to WT mice. However, neutralization of IFN-γ in casp1/11−/− mice did not result in enhanced inflammation. Collectively these data demonstrate casp1/11 induces early inflammation in an IL-18 dependent manner. Moreover, there is an IFN-γ independent role of casp1/11 in joint Brucella clearance, and casp1/11 is responsible for enhanced inflammation observed in IFN-γ-deficient mice.
The prostaglandin D2 receptor CRTH2 suppresses epithelial cell responses during intestinal helminth infection
Authors: Oyesola Oyebola Oluwakemi , Webb Lauren M , Solouki Sabrina , Pham Duc , Campioli Pamela , Cubitt Rebecca , Wojno Elia D Tait
Published: January 2023

Type 2 inflammation is required for expulsion of intestinal helminth parasites and is characterized by immune cell activation, type 2 cytokine production, and increased mucin production by epithelial goblet cells. Previous studies show that the prostaglandin D2 (PGD2) receptor chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2) promotes type 2 inflammation in the lung via effects on immune cells, but how CRTH2 influences helminth-induced type 2 inflammation in the intestine was unclear. Here we show that CRTH2-deficient mice cleared infection with the mouse-adapted helminth parasite Nippostrongylus brasiliensis more efficiently and had increased numbers of mucin-producing goblet cells in the small intestine post-infection compared to wild type mice, despite similar levels of worm establishment. These data suggest that CRTH2 restrains goblet cell responses during intestinal helminth infection, in contrast to its pro-inflammatory role in the lung. Consistent with this idea, bone marrow-chimeric mice in which only non-hematopoietic cells lacked CRTH2 also cleared parasites more efficiently than chimeric wild type mice and had increased numbers of small intestinal goblet cells. In addition, murine small intestinal organoids that were stimulated with type 2 cytokines downregulated expression of goblet cell-associated genes following culture with PGD2. Taken together, these data suggest that the PGD2-CRTH2 pathway suppresses epithelial goblet cell responses following helminth-induced type 2 inflammation in the intestine. These findings may inform the development and use of drugs that inhibit this pathway during intestinal type 2 inflammatory disease, such as food allergy.
Broccoli-derived nanoparticle inhibits mouse colitis by activating dendritic cell AMP-activated protein kinase
Authors: Deng Zhongbin , Rong Yuan , Teng Yun , Mu Jingyao , Zhuang Xiaoying , Samykutty Abhilash , Zhang Lifeng , Yan Jun , Miller Donald , Suttles Jill , zhang huang-ge
Published: January 2023

The intestinal immune system is continuously exposed to massive amounts of nanoparticles derived from food. Whether nanoparticles from plants we eat daily have a role in maintaining intestinal immune homeostasis is poorly defined. Here, we present evidence supporting our hypothesis that edible nanoparticles regulate intestinal immune homeostasis by targeting DC cell. With three mouse colitis models, our data show that orally given nanoparticles isolated from broccoli extracts protect mice against colitis. Broccoli-derived nanoparticles (BDN) mediated activation of adenosine monophosphate-activated protein kinase (AMPK) in dendritic cell (DC) plays a role in not only prevention of DC activation but also induction of tolerant DCs. Adoptively transferring DCs pre-pulsed with total BDN lipids but not sulforaphane depleted BDN lipids prevented DSS induced colitis in B6 mice, supporting the role of BDN sulforaphane in the induction of DC tolerance. Adoptively transferring AMPK+/+ but not AMPK−/− DCs pre-pulsed with sulforaphane prevented DSS induced colitis in B6 mice, further supporting the DC AMPK role in sulforaphane mediated prevention of DSS induced colitis. This finding could open new preventive/therapeutic avenues to address intestinal related inflammatory diseases via activating AMPK.
IL-15 regulates IL-23 production and murine IBD-like disease
Authors: Eken Ahmet , Singh Akhilesh K , Fry Mallory , Bhise Gauri , Bettelli Estelle , Oukka Mohamed
Published: January 2023

Group 3 innate lymphoid cells (ILC3) have been recently implicated in various autoimmune conditions including inflammatory bowel diseases (IBD). The mechanisms by which ILC3s contibute to IBD pathogenesis remain elusive. In this study, using genetic deleletions that impact ILC3s to varying degrees we sought to determine their contribution to murine IBD development. We show that IL-23-dependent T cell-induced colitis develops independently of ILC3s. Host IL-2 receptor common γ chain (IL-2Rγc) is needed for both innate and adaptive cell-driven colitis, since its deficiency impairs colitogenic T cell proliferation, expansion and recruitment to intestines. We also provide evidence that resistance to colitis in IL-2Rγc Rag2−/− mice may occur as a result of defective IL-15R signaling, and that IL–15−/− Rag1−/− mice recapitulates the murine IBD phenotype of IL-2Rγc Rag2−/− mice. Data we presented herein demonstrates that IL-15 plays an important role in IBD pathogenesis and may contribute to gut inflammation by regulating IL-23 production of antigen presenting cells.
Down-modulation of colitis-associated colorectal cancer development by treatment with helminth-derived molecules
Authors: Terrazas Luis I , Callejas Blanca , Reyes Sandy , Ledesma-Soto Yadira , Olguín Efraín J , León-Cabrera Sonia , Rodríguez-Sosa Miriam
Published: January 2023

A role for chronic inflammation in tumor development has been well established and supported in both, animal and epidemiological studies. A classic association between inflammation and cancer is colitis-associated colorectal cancer (CAC). Helminth infections and helminth-derived molecules have been recognized as inducers of immunoregulation. Taenia crassiceps modulates inflammatory diseases such as type 1 diabetes, colitis, and autoimmune encephalomyelitis. Here we analyzed the pathophysiologic implication of T. crassiceps excreted/secreted (TcES) products on CAC development. CAC was induced in BALB/c mice with azoxymethane followed by 3 cycles of 2% dextran sodium sulfate exposure to elicit intestinal inflammation. After cycle 1, mice received i.p. TcES (200 μg/mouse 3-times/week) until the end of the experiment. On days 20, 40 and 68 after CAC induction mice were killed to evaluate colonic inflammation, proliferation, and tumorigenesis. Untreated mice developed tumors as early as 20 days after CAC induction and by the end of the experiment, 100% of these mice displayed tumors. In contrast, only 30% of mice receiving TcES developed tumors. Tumor numbers per mouse were significantly reduced in TcES-treated mice (4.5 tumors) versus untreated mice (14.5 tumors). Both inflammation and proliferation scores in the colon from CAC-TcES-treated mice were significantly lower than CAC-untreated mice. High levels of IL-1β, TNF-α, and IL-17 in CAC-untreated mice were down-modulated by TcES-treatment. Lower expression of COX2, β-catenin and, STAT3 activation were detected in colonic tissue from mice receiving TcES. Our data suggest a main role for TcES in generating an anti-inflammatory environment that suppresses CAC progression
Phosphorylation of SNAP-23 regulates its dynamic subcellular localization and regulated exocytosis in mast cells
Authors: Naskar Pieu , Puri Niti
Published: January 2023

Mast cells (MCs) play an important role in inflammatory response to pathogens and in inflammatory disorders like allergies. This role depends on their ability to release prestored mediators from secretory granules in response to some external trigger like crosslinking of FcɛRI. The membrane fusion events involved therein are mediated by SNAREs. The target-SNARE SNAP-23 has been shown to be very important for MC exocytosis, and our previous studies revealed presence of one basal (Thr102) and two induced (Ser95 and Ser120) phosphorylation sites in its linker region, whose significance was not very clear. To study the role of SNAP-23 phosphorylation in the regulation of exocytosis, GFP-tagged wildtype SNAP-23 (GFP-SNAP-23) and its phosphorylation mutants were transfected into RBL MCs, that revealed some dynamic changes in SNAP-23 membrane association immediately after activation. SNAP-23 was associated with plasma membrane in resting mast cells, however on activation, it translocated to internal LAMP-3 containing lysosomal membranes, many of which contained secretory granule cargo. This dynamic change in the membrane association of SNAP-23 in MCs may be important for mediating internal granule-granule fusions in compound exocytosis. Further studies with mutants revealed, for the first time, that the basal phosphorylation site Thr102 plays an important role in initial SNAP-23 membrane association. During exocytosis, the induced phosphorylation sites Ser95 and Ser120 regulate SNAP-23 internal membrane association and subsequently mast cell exocytosis. Hence this study reveals a novel mechanism in mast cell inflammatory response that can be a therapeutic target.
Mast cells’ interaction with Leishmania leads to effector responses in reducing initial parasite load
Authors: SELVAPANDIYAN ANGAMUTHU , Naqvi Nilofer , Ahuja Kavita , Dey Ranadhir , Nakhasi Hira L , Puri Niti
Published: January 2023

Mast cells (MCs) are involved in clearing invading pathogens at the skin surface and play an important role in innate and adaptive defenses. Hence they may have an important role in the control of Leishmaniasis due to the protozoa Leishmania that are inoculated by infected sandfly vector on the skin surface. It is still unclear if their contributions occur directly through active pathogen killing or antigen uptake. Our objectives were to use Leishmania donovoni the causative agent of fatal visceral Leishmaniasis and L. tropica the causative agent of cutaneous leishmaniasis for studying the interactions between the parasite and MCs to clarify if MCs are directly involved in active antigen uptake, and its clearance. We found that co-culture of Leishmania with MCs led to significant killing of L. tropica and to a lesser extent of L. donovoni. Also, while there was significant uptake of L. tropica by MCs, L. donovoni was not phagocytosed by MCs. Uptake increased with time of exposure in case of L. tropica. We also found significant generation of reactive oxygen species by MCs on co-culture with these species of Leishmania, which may contribute to their clearance. However interactions of MCs with Leishmania led to generation of MC extracellular traps comprising of DNA, histones and tryptase probably to ensnare these pathogens. These results clearly establish that MCs may contribute to host defences to Leishmania in a differential manner by actively taking up these pathogens and also by mounting effector responses for their direct clearance by extracellular and non-phagocytic means. This gives us an understanding that MCs can also be used as a target for therapeutic approaches to leishmaniasis.
Antibody-dependent NK cell control of Plasmodium falciparum infection
Authors: Hart Geoffrey T , Theorell Jakob , Tran Tuan , Arora Gunjan , Crompton Peter D , Bryceson Yenan , Long Eric O
Published: January 2023

Plasmodium falciparum (P.f) infection is a major cause of morbidity and mortality world-wide where symptoms and death occur as a result of the blood stage of the P.f. life cycle. To date, there is no effective blood stage malarial vaccine. Natural Killer (NK) cells are key players in the control of hematopoietic cancers and viral infections, however their role in blood stage malaria is unknown. We undertook a comprehensive analysis of NK cell phenotype and function in a cohort of subjects from a malaria clinical study in Mali, Africa. Using an unbiased analysis of different NK cell subsets, we found that ‘adaptive’ NK cells (lacking FcR gamma chain) increased with age and correlated with reduced malaria risk. Adaptive NK cells had enhanced antibody dependent cellular cytotoxicity (ADCC) function relative to other NK cell subsets. Antibodies are known to be protective in malaria, however the mechanism of this protection is still being elucidated. We hypothesized the protective mechanism of adaptive NK cells may be adaptive NK cells pairing with malaria antibodies to clear infected RBCs via ADCC. In a parallel study, we have shown that NK cells from healthy US donors can inhibit the growth of Plasmodium falciparum 3D7 strain in vitro in the presence of IgG from malaria-exposed adults. Adaptive NK cells from Mali also have enhanced ADCC to infected RBCs. This work indicates that protective antibodies may pair with the strong ADCC activity of adaptive NK cells to protect human subjects from malaria symptoms. These findings thus open a new avenue for vaccine development exploring antibody-dependent cell mediated killing of malaria parasites.
Wild mouse gut microbiome protects laboratory mice against lethal influenza virus infection and colorectal cancer
Authors: Rosshart Stephan Patrick , Vassallo Brian G. , Angeletti Davide , Hutchinson Diane S. , Morgan Andrew P. , Takeda Kazuyo , Hickman Heather D. , Ajami Nadim J. , de Villena Fernando Pardo-Manuel , Yewdell Jonathan W. , Rehermann Barbara
Published: January 2023

Mouse models are paramount for understanding basic immunological mechanisms, but can be limited in recapitulating human diseases. The microbiome has been identified as a main factor influencing host physiology as illustrated by many studies that disrupt or modify host-microbe interactions in laboratory mice. Differences in the gut microbiome contribute to the variability of research results obtained with genetically identical animals from different vendors. In an effort to identify an external reference that better recapitulates physiologically important interactions found in a natural habitat, we asked how far removed the gut microbiome of laboratory mice is from that of their outbred, wild-living relative. Through genetic analysis we identify the closest wild relatives to classical laboratory strains among 21 distinct populations from Europe, Asia and the Americas. We establish that their gut microbiome differs significantly from that of C57BL/6 mice from the leading breeders worldwide, and that it can be transferred and maintained over several generations under vivarium conditions. Offspring of pregnant germ-free C57BL/6 mice reconstituted with the gut microbiome of wild mice exhibit a significantly reduced inflammatory response and increased survival following influenza A virus infection. This restoration of the natural ‘microbial identity’ of laboratory mice also improved their resistance against mutagen- and inflammation-induced colorectal cancer. Collectively, these data show the beneficial effects of the wild mouse microbiome in two diseases of global relevance. They also illustrate a novel approach towards developing animal models of greater biological relevance and translational research value.
Minced muscle grafts reduce the proinflammatory immune response and partially restore musculoskeletal healing
Authors: Wager Chrissy M Leopold , Goldman Stephen M , Pollot Beth E , Greising Sarah M , Wenke Joseph C , Corona Benjamin T
Published: January 2023

The immune response plays a key role in healing of bone and muscle injuries. An unrestrained proinflammatory response generally results in impaired healing. Extremity fracture is often complicated by traumatic volumetric muscle loss (VML), which can lead to delayed or non-union of the fracture (i.e., impaired healing). In a rat model, we recently showed that the VML injury of the tibialis anterior muscle prevented healing of the adjacent tibia fracture and induced a robust proinflammatory response in the musculature early post-injury that was not observed with the fracture alone. In the current studies, we show that a fracture treated with rhBMP-2, a highly osteoinductive protein, was impaired by the VML injury and corresponded with a heightened proinflammatory response. We utilized an autologous minced muscle graft (MMG) to repair the VML which, interestingly, reduced the proinflammatory response. Gene transcript analysis of the muscle injury indicates that MMG repair results in decreased gene expression of proinflammatory mediators including ITGAM, IL18, and CCL12, compared to non-repaired muscle. In the bone defect, MMG repair also induced a modest proinflammatory response with lower expression of IFNG and TNF than the non-repaired group. The restrained proinflammatory response corresponded with restored healing of the bone fracture and, remarkably, partial restoration of muscle function which does not normally occur clinically. Altogether, data suggest that MMG repair controls the proinflammatory response in a manner conducive to improved healing responses in both muscle and bone. These data reveal that immunomodulation to dampen proinflammatory responses could provide an effective therapy for musculoskeletal trauma.
Deletion of CCL2 in mesenchymal stem cells suppresses experimental aortic aneurysms
Authors: Xu Baohui , Deng Hongping , Li Gang , Fujimura Naoki , Furusho Yuko , Dalman Ronald L
Published: January 2023

Abdominal aortic aneurysm (AAA) is a macrophage-mediated chronic inflammatory disease. Aortic mural macrophages arise from bone marrow-derived circulating monocytes, recruited to the aneurysmal aorta by CCL2/CCR2 signaling. Given the importance of mesenchymal stem cell (MSC)-derived CCL2 in bone marrow monocyte mobilization, we hypothesized that MSC-derived CCL2 contributes to AAA pathogenesis. AAAs were created in 10–12 weeks old, male MSC-specific CCL2 knockout (Nestin+/CCL2 Flox+/+) and wild type (CCL2 Flox+/+) mice via intra-aortic elastase infusion. AAA incidence and progression were assessed via serial in vivo ultrasound aortic diameter measurements and histopathological analysis at sacrifice. MSC cell-specific CCL2 ablation significantly reduced circulating Ly-6Chigh inflammatory monocytes at baseline as well as 3 hours following lipopolysaccharide injection. Following elastase infusion, wild type mice demonstrated progressive, time-dependent aortic enlargement beginning on day 3. In MSC-specific CCL2 knockout mice, elastase-induced aortic enlargement was substantially suppressed, and medial elastin lamellae and smooth muscle cell cellularity were preserved compared to that demonstrated in wild type AAA mice. Mural macrophages, T and B cell infiltration were all also significantly attenuated in knockout, as compared to wild type, mice. Additionally, aortic neoangiogenesis, as defined by mural CD31+ capillary vessel density, was significantly reduced in knockout mice. Taken together, this evidence suggests that MSC-specific CCL2 deletion suppresses progression of experimental AAAs, in conjunction with attenuation of aortic mural inflammation and angiogenesis.
Correction of mitochondrial and peroxisomal defects by lovastatin and AICAR in EAE.
Authors: Singh Inderjit , Saxena Nishant , Singh Avtar K , Won Jeseong
Published: January 2023

In multiple sclerosis, the CNS disease progression continues despite treatment with FDA approved immunomodulatory drugs. Recent studies report that loss of mitochondria and peroxisomes contributes to the CNS disease of MS. In EAE, the loss of myelin proteins paralleled the loss of peroxisomes (catalase and PMP70) and loss of mitochondria (complex I and citrate synthase) as well as levels of ATP. Using EAE animal model, we investigated the efficacy of immune modulatory drugs (lovastatin or AMPK activator AICAR) for protections of mitochondria, peroxisomes, and myelin in relation to the clnical disease. While lovastatin or AICAR attenuated the EAE disease, but individually these drugs provided only partial protection of mitochondria and peroxisomes. However, combination therapy of lovastatin and AICAR was able to protect mitochondria, peroxisomes, and myelin as well as greater efficacy against clinical disease of EAE as compared to individual drug treatments. Lovastatin-mediated inhibition of RhoA induced the expression of transcription factors, PPARα, PPARγ, and catalase and thus biogenesis of peroxisomes and AICAR-mediated activation of AMPK induced expression of transcription factors, PGC-1α, SIRT1, and MnSOD and thus biogenesis of mitochondria in EAE spinal cords. In summary, these studies document that oral medication of combination of immunomodulatory drugs (lovastatin and AICAR) protect against the loss of mitochondria and peroxisomes and energy homeostasis in EAE resulting in greater efficacy in EAE supporting a potential therapeutic approach for progressive MS. Supported in-part by VA:BX002829 and NIH:NS037766.
Breadth of function and homing capacity of peripheral CD4+ Th responses to intradermal (ID) immunization with enterotoxigenic E. coli fimbrial tip adhesin CfaE plus adjuvant LTR192G
Authors: Trop Stefanie , Laird Renee M , Gutierrez Ramiro L. , Riddle Mark S. , Porter Chad K , Maciel Milton
Published: January 2023

ETEC are a leading cause of traveler’s diarrhea and a vaccine development priority for the US military. A subunit vaccine comprised of ETEC fimbrial tip adhesin, CfaE, adjuvanted with single-mutant heat-labile toxin, LTR192G, was given ID 3 times, 21 days apart, and evaluated for immunogenicity and efficacy against moderate to severe diarrhea (MSD) following CFA/I+ ETEC oral challenge. The homing phenotype, activation status, and cytokine production of circulating CD4+ T cells following vaccination were assessed by flow cytometry to understand the response generated by mucosal adjuvant, LT, introduced via the skin, and the potential mechanisms involved in protection from an enteric pathogen.

Following in vitro stimulation with CfaE or LT 15mers, PBMCs of most vaccinated subjects expressed IFNγ or IL-17, within multiple homing subsets. Specifically, activated CfaE-specific α4β7+IFNγ+ cells were detected in 69% of vaccinated subjects, suggesting potential for gut homing, while 58% subjects generated CD62L+IFNγ+ responses. LT-specific IFNγ was observed in CD62L+ and CLA+ cells (52% and 50% subjects, respectively). Both CfaE- and LT-specific IL-17 were identified in CCR6+ (65%, 66% subjects, respectively) and CD62L+ cells (65%, 58%).

Subjects without MSD following experimental challenge had significantly higher CfaE-specific activated (CD154+) α4β7+ cells (p=0.043, t-test), as well as IFNγ (p = 0.036) and IL-17 (p = 0.008) responses to vaccination. LT-specific activation of CLA+ cells (p = 0.035) and IL-17 (p = 0.032) levels were also higher in subjects without MSD. Overall, ID vaccination with CfaE and LTR192G elicited broadly homing and functional Th cell responses that may play a role in protection at the mucosal level.
Immune response and early transcriptional changes after primary and booster vaccination against Hepatitis B in young and old adults
Authors: Weinberger Birgit , Haks Marielle C , Katzgraber Franz , Ottenhoff Tom HM , Grubeck-Loebenstein Beatrix
Published: January 2023

Many currently used vaccines are less immunogenic and effective in the elderly compared to younger adults due to age-related changes of the immune system. Most vaccines utilized in the elderly contain antigens, with which the target population had previous contact. Therefore, most studies investigating vaccine-induced immune responses in the elderly do not analyze responses to neo-antigen but rather booster responses. It can be hypothesized that age-related differences in the immune response are distinct for primary and recall responses. We therefore aimed to investigate primary and recall responses using the same antigen in young and older adults and chose Hepatitis B vaccine as a model antigen. Young (20–40 years) and elderly (>60 years) healthy volunteers received either a primary series (no prior vaccination) or a single booster shot (documented primary vaccination more than 10 years ago) of the registered vaccine Twinrix. Expression of immunity-related genes was measured before and 1 day after vaccination and antibody titers were determined at several time points with a long-term follow-up 6 months after the last vaccination. Antibody responses were lower and seemed to be delayed in the elderly compared to young adults. Non-responders after the 3-dose primary series were more frequent in the elderly group. Antibody titers after booster vaccination increased in all participants. Associations between early expression-profiles on day 1 and antibody responses were found.
Antigenic competition affects the magnitude and breadth of CD8 T cell immunity following immunization with a nanoparticle neoantigen cancer vaccine
Authors: Blobel Nicolas J , Ramirez-Valdez Andrei , Ishizuka Andrew S , Lynn Geoffrey M , Seder Robert A
Published: January 2023

Induction of T cell immunity against neoantigens of cancers is a promising personalized vaccine approach against tumors. We developed a novel vaccine platform to enhance the breadth and magnitude of neoantigen-specific CD8 T cell responses by coupling neoantigen peptides to a polymer containing a Toll-Like Receptor-7/8 agonist that self-assembles into nanoparticles (termed SPP-7/8a). As the SPP-7/8 vaccine platform can be formulated to immunize against multiple neoantigen peptides, a critical issue is how the delivery influences the magnitude and breadth of CD8 T cell responses. Thus, C57BL/6 mice were injected subcutaneously against a single neoantigen at either a single site, or divided across four sites. CD8 T cell responses were ~3% in the mice immunized at multiple sites compared to ~1% at a single injection site. To assess breadth, mice were immunized with four antigens, with either a single antigen at each of four injection sites or a combination of all four antigens across four injection sites. In mice that received all four antigens in each site, CD8 T cell responses against two of the antigens were undetectable. By contrast, in animals that received 1 antigen per site, responses were detected against all four antigens, and the total magnitude was higher than animals receiving 4 antigens per site. Collectively, these data suggest that despite increasing the CD8 T cell response of a single antigen via multi-site injection, vaccination even with as few as 4 antigens can interfere with the induction of neoantigen-specific CD8 T cells. Current work is focused on altering the delivery of the SPP-7/8a immunization approach to optimize both magnitude and breadth of CD8 T cell immunity that will be critical for tumor treatment and protection.