bluuebunny/crossref_metadata_2025 huggingface dataset:
CD47-SIRPα axis restricts the therapeutic antitumor immunity through controlling tumor mitochondrial DNA-mediated innate sensing selectively in dendritic cells
Authors: Xu Meng , Fu Yang-Xin
Published: January 2023

STING (Stimulator of interferon genes)-dependent cytosolic DNA sensing is critical for the activation of antitumor immunity. However, the molecular determinants driving the differential DNA sensing of tumor cells in diverse professional phagocytes remain largely unexplored. Here we show that preventing the engagement of SIRPα (signal regulatory protein α) by CD47 promoted DNA sensing selectively in DCs, but not in macrophage, although both macrophages and DCs phagocytosed tumor cells efficiently. DCs were specialized to preserve mitochondrial DNA (mtDNA) from the engulfed tumor cells that served to initiate type I interferon-dependent antitumor immunity. Mechanically, interruption of CD47-SIRPα interactions prevented the translocation of SIRPα to NOX2 on DC phagosomes, which inhibited phago-lysosome fusion and mtDNA degradation. MtDNA is thus recognized by cyclic-GMP-AMP synthase (cGAS) in DC cytosol, which contributes to type I IFN production and antitumor adaptive immunity. Our findings identify CD47-SIRPα as a major innate checkpoint that dictates the unique capacity of DCs in immune recognition and therapeutic intervention of CD47-SIRPα axis could unleash DC-driven T-cell-mediated cancer immunity.
Inflammatory disease severity is ameliorated by inhibition of neutrophil-derived MPO that supports endothelial/epithelial integrity
Authors: Strzepa Anna , Dittel Bonnie N
Published: January 2023

Neutrophils are the most abundant leukocytes and provide a first line of defense against pathogens. Neutrophil granules store enzymes with anti-bacterial activities that can also damage tissues when released in an uncontrolled manner. Neutrophil infiltrates are prevalent in damaged tissues from mice with experimental autoimmune encephalomyelitis (EAE), dextran sodium sulfate (DSS)-colitis and contact hypersensitivity (CHS). The most cytotoxic and abundant enzyme expressed by neutrophils is myeloperoxidase (MPO). MPO utilizes hydrogen peroxide to generate highly reactive molecular moieties, that modify proteins, lipids and DNA, resulting in tissue damage. A pathogenic role for MPO was observed in EAE, DSS-colitis and CHS. MPO-deficiency, is associated with improved functioning of the vasculature, indicating that vasculature is a important target of MPO-activity. However, the precise molecular targets, associated with neutrophil-derived MPO are not known. To establish the importance of MPO as a candidate for therapeutic inhibition, we used animal models of colitis and CHS. Our strategy to modulate MPO activity was the use of MPO-deficient animals and the MPO-specific inhibitor KYC (N-acetyl lysyltyrosylcysteine amide). Our data indicated that MPO-deficient animals, exhibited significantly reduced disease severity in both DSS-colitis and CHS, which correlated with reduced intestinal and vascular permeability, respectively. In addition, MPO inhibition with KYC was also an effective strategy to reduce colitis and CHS severity. KYC mediated MPO inhibition enhanced endothelial integrity in vitro. Our data indicate that MPO could be potential therapeutic target in the treatment of inflammatory diseases.
Treatment with CD122 antibody durably reverses autoimmune skin disease in a mouse model of vitiligo
Authors: Richmond Jillian M , Zapata Lucio , Garg Madhuri , Strassner James , Rashighi Mehdi , Riding Rebecca , Ahmed Maggi , Essien Kingsley , Pell Lila , Agarwal Priti , Tsurushita Naoya , Tso J Yun , Harris John
Published: January 2023

Vitiligo is a disfiguring autoimmune skin disease characterized by patchy depigmentation. Vitiligo affects approximately 1% of the population worldwide, and it is mediated by CD8+ T cells that kill melanocytes, the pigment producing cells in the skin. Like many autoimmune diseases, treatment options for vitiligo are limited, and depigmentation rapidly recurs after therapy is stopped. We hypothesized that tissue resident memory T cells (Trm) persisted in the skin to reactivate disease. We first confirmed the presence of melanocyte-specific cells with Trm phenotypes in biopsies from human patients and in a mouse model of vitiligo. Functional analysis of Trm cells in mice with vitiligo revealed that they can sense antigen in the skin as determined by Nur77-GFP reporting, as well as secrete cytokines and chemokines as determined by IFNγ, CXCL9 and CXCL10 reporting. Treatment with the S1P1 inhibitor FTY720 resulted in rapid repigmentation, yet preserved Trm. Treatment with a CD122 blocking antibody effectively depleted autoreactive Trm. Importantly, CD122 antibody was also able to durably reverse disease mice. We propose that depleting Trm by targeting the IL-15 axis may prove to be a highly effective and durable treatment for vitiligo and other autoimmune diseases.
Implementation of just-in-time teaching strategy as an active learning instructional pedagogy for PharmD integrated microbiology and virology course
Authors: Madiraju Charitha , Tellez-Corrales Eglis , Hua Henry , Brown Deborah M
Published: January 2023

Just-In-Time Teaching (JiTT) is an active learning pedagogy that has been successfully adopted by various disciplines. However, it is unclear if JiTT presents value to a pharmacy curriculum. The current study implemented JiTT as an active-learning tool for an Integrated Microbiology and Virology course offered during the first year of a PharmD program. At the end of the course, students were provided with a survey that included items about JiTT. Several items were aggregated into a single rating about the degree that students agreed with various statements purporting JiTT as helpful, with score choices ranging from 1 (I strongly disagree) to 4 (I strongly agree). Overall, students strongly endorsed JiTT as helpful (M = 3.61, SD = 0.50), and even the lowest score, 2.50, was neutral in tone. Students were asked to endorse rationales for why JiTT questions were used in this course; 93.02% felt that JiTT helped them keep track with the material, and only 34.88% felt that JiTT gave them “extra work.” These results—and other findings stated in the poster —suggest that students in a pharmacy program find value in JiTT.
Tyrosine kinase Syk interacts with Hrs after TLR3 stimulation in murine microglial cells.
Authors: Mielcarska Matylda Barbara , Bossowska-Nowicka Magdalena , Gregorczyk Karolina Paulina , Wyzewski Zbigniew , Toka Felix Ngosa
Published: January 2023

TLR3 (Toll-like receptor 3) belongs to the family of receptors involved in innate immune response. Present in endosomal compartment in cells, TLR3 recognizes the dsRNA (double-stranded RNA), viral nucleic acid or intermediate during viral replication and initiates signal transduction leading to inflammatory response. However, the first steps of signaling cascade occurring in cells immediately after TLR3 stimulation are still not well understood and require careful attention. In this paper we demonstrate that after poly(I:C) (dsRNA mimetic) stimulation of murine microglial cells (C8D1A cell line) tyrosine kinase Syk interacts with Hrs (hepatocyte growth factor regulated tyrosine kinase substrate), one of the ESCRT-0 (endosomal sorting complex required for transportation-0) components. This protein complex was recently implicated in trafficking of other endosomal TLRs, TLR7 and TLR9, and plays a possible role in the TLR3 signaling. Phosphorylation of Hrs, which is a very likely result of Hrs-Syk interaction, also takes place after poly(I:C) stimulation, strongly suggesting that an association between activated TLR3 and ESCRT-0 exists. Therefore, Hrs may influence TLR3 signaling pathways, but this requires further investigation.
Utilization of protein-peptide conjugates for NK cell activation as a strategy for cancer immunotherapy.
Authors: Bitsaktsis Constantine , Phillips Mariana , Sabatino David
Published: January 2023

Cancer-based immunotherapy aims to develop biologics that can stimulate immune responses towards tumor eradication, as is the case of monoclonal antibodies (mAb) and their related analogs. Although the latter have been the most prevalently used biologics in cancer immunotherapy strategies, novel therapeutics may overcome their production, administration, and pharmacological limitations. The synthesis of intermediate size molecules, particularly chemically synthesized protein-peptide bioconjugates (PPB), possessing the targeting and effector functions of antibodies, represent a novel class of immunotherapeutics that may circumvent these limitations. In this study we proposed and executed the rational design of a novel class of cancer-targeting immunostimulatory protein-peptide conjugates (CTIPPC) that may potentially work as a synthetic antibody mimic. In this proof-of-concept, the CTIPPC aimed to target the tumor expressed GRP78, while recruiting and activating NK cells for tumor targeted immunotherapy. The sustained capabilities of the CTIPP conjugates to elicit NK-dependent immunostimmulatory activities were assessed via measuring cytokine secretion, NK cell migration, and the overall anti-tumor cytotoxic activity of NK cells. These studies shed important insights into the effective design, preparation and biological evaluation of a new class of semi-synthetic protein-peptide conjugates.
Eradication of tumors containing MHC class I deficient cells by inducing adaptive and innate immune responses
Authors: Seo Hyungseok , Jeon Insu , Bae Eun-Ah , Song Boyeong , Shin Kwang-Soo , Koh Choong-Hyun , Kang Chang-Yuil
Published: January 2023

During cancer immunoediting, tumor cells with spontaneous loss of MHC class I expression tend to gradually increase in tumor microenvironment by preferentially escaping from immune surveillance by cytotoxic T cells, although the MHC class I-deficient tumors have been historically considered to be susceptible to NK cell-dependent cytotoxicity. In line with this idea, recent studies demonstrated that most NK cells found in tumor microenvironment of advanced cancers are defected, releasing the malignant MHC class I-deficient tumors from NK cell-dependent immune control. Here we described that NKT cell-ligand loaded tumor-antigen expressing APC-based vaccine effectively eradicated these advanced tumors which have not been curable by an immunotherapy. In this process, we found that co-expression of Tim-3 and PD-1 marks functionally exhausted NK cells in advanced tumors, and that MHC class I down-regulation in tumors was closely associated with the induction of NK cell exhaustion in tumor-bearing mice as well as in cancer patients. Furthermore, recovery of NK cell function by NKT cell driven cytokines was critical for the anti-tumor effect of the vaccine against these advanced tumors. These results uncover the mechanisms involved in the induction of NK cell dysfunction in advanced cancers and provide a guidance to develop new strategies for cancer immunotherapy.
The lymphomagenic molecule LMP1 sequesters TRAF3 to induce enhanced B cell survival.
Authors: Stunz Laura L , Bangalore-Prakash Pradeep , Mambetsariev Nurbek , Bishop Gail A.
Published: January 2023

The human TRAF3 gene is frequently mutated in multiple myeloma and lymphoma. TRAF3 restrains homeostatic B cell survival. B cell TRAF3 deficiency promotes enhanced viability and over-expression of the anti-apoptotic molecules NIK, Mcl1 and Pim2, enhanced nuclear CREB, and increased pSTAT3 in response to IL6 signaling. We assessed TRAF3 protein expression in human lymphoma samples. The number of TRAF3low DLBCL was higher than predicted based on genetic loss, so we speculated that cellular TRAF3 can be depleted through other means. The oncogenic Epstein Barr virus protein LMP1 binds TRAF3 with much higher avidity than do CD40 or BAFF receptors, so we hypothesized that LMP1-sequesters TRAF3, causing functional depletion. We found evidence of induction of survival pathways in TRAF3 sufficient B cells expressing LMP1 consistent with those in B cells genetically lacking TRAF3.

B lymphoma cell lines stably transfected with inducible LMP1 were stimulated via CD40 or BAFFR. TRAF3 binding was studied via CD40/BAFFR immunoprecipitation and Western blotting. We also examined levels of cellular NIK, and used confocal fluorescence microscopy to assess nuclear CREB expression. We found LMP1 expression sequesters cellular and nuclear TRAF3, decreasing its availability to CD40 and BAFFR, and increases survival-promoting NIK, CREB and pSTAT3. LMP1 mutated at the major TRAF binding site only marginally induced TRAF3 sequestration and NIK. Thus, B cell LMP1 expression can enhance survival by functionally reducing TRAF3 availability.
Antigen-presenting tumor B cells impact the phenotype of CD4 tumor infiltrating T cells in lung cancer patients
Authors: Bruno Tullia Carmela , Ebner Peggy , Moore Brandon , Squalls Olivia , Waugh Katherine , Eruslanov Evgeniy B , Singhal Sunil , Mitchell John , Franklin Wilbur , Merrick Daniel , McCarter Martin , Palmer Brent E , Kern Jeffrey , Vignali Dario , Slansky Jill E
Published: January 2023

The focus of immunotherapy has been on CD8 and CD4 tumor infiltrating lymphocytes (TILs), however, tumor infiltrating B cells (TIL-Bs) are understudied with no focus on their role as antigen presenting cells. We hypothesize that TIL-Bs help generate potent, long-term immune responses against cancer by presenting tumor antigens to CD4 TILs.

Using un-manipulated, primary human B cells from fresh tumor, we generated a specific in vitro antigen presentation assay and observed three types of CD4 TIL responses when TIL-Bs presented autologous tumor antigens. There were activated responder CD4 TILs that proliferated when combined with TIL-Bs alone, which indicates stimulation with endogenous tumor antigens. There were antigen-associated responders that required autologous tumor lysate to elicit a CD4 TIL response, and there were patient CD4 TILs that did not respond to antigen presentation. Exhausted B cells have been described in chronic infection i.e. HIV, and thus, to parallel our studies to a model of chronic infection, we analyzed activated and exhausted TIL-Bs with our antigen presentation assay. If TIL-Bs were activated (HLADR+CD69+CD27+CD21+), the resultant CD4 TILs were T helper (anti-tumor) CD4 T cells and if the TIL-Bs were exhausted (HLADR+CD69+CD27−CD21−), the resultant CD4 TILs were T regulatory cells (pro-tumor). These data suggest that TIL-Bs influence CD4 TILs in NSCLC patient tumors.

In conclusion, determining if TIL-Bs are activated or exhausted in NSCLC patients will determine the extent of their anti-tumor function in human cancer. Ultimately, results from this study will dictate how to target TIL-Bs in future immunotherapies.
Characterization of bovine γδ T cells phenotype during post-natal development and following Mycobacterium bovis vaccination or virulent infection
Authors: Guerra-Maupome Mariana , Larsen Michelle H. , Palmer Mitchell V. , Waters Ray , McGill Jodi L
Published: January 2023

Bovine tuberculosis caused by Mycobacterium bovis is a globally significant veterinary health problem. γδ T cells participate in the immune control of mycobacterial infections. Data in human and non-human primates suggest that mycobacterial infection regulates memory/effector phenotype and adaptive immune functions of mycobacterium-responsive γδ T cells. To date, the impact of age or M. bovis infection on bovine γδ T cells memory/effector phenotype remains unknown. In this study, we addressed the age-dependent changes of circulating γδ T cells, analyzed functional and phenotypic differences in memory and activation marker expression, and evaluated functional responses of M. bovis-specific γδ T cells following M. bovis Bacille Calmette-Guerin (BCG) vaccination or virulent M. bovis infection. As expected, aerosol vaccination and virulent infection induced robust mucosal and systemic M. bovis-specific γδ T cell proliferative responses. Analysis of peripheral blood γδ T cells indicated phenotypic differences based on the expression of CD27 and CD45R that could suggest distinct functional properties. Recall responses after in vitro stimulation with mycobacterial antigens showed that expression of CD27 is critical for the expansion of peripheral IFN-γ-producing γδ T-cells and suggest that the CD27+ subset may compose the mycobacterium-responsive γδ T cell compartment in calves responding to M. bovis vaccination or infection. The ability of γδ T cells to mount a robust response during mycobacterial infections supports the notion that vaccine-elicited γδ T cell immunity might prove beneficial. Therefore, defining correlates of protection based on this population can accelerate the development of novel vaccines against TB.
Protective role of conventional dendritic cells in experimental model of sepsis
Authors: Gargaro Marco , Briseno Carlos , Vivek Durai , Scalisi Giulia , Theisen Derek J , Bagadia Prachi , Turco Antonella , Anderson David A , Murphy Theresa L , Murphy Kenneth M , Fallarino Francesca
Published: January 2023

Sepsis is a serious medical condition caused by an overwhelming immune response to infection and is a major challenge in the intensive care unit, where it’s one of the leading causes of death. Dendritic cells (DCs), acting as sentinels, constantly sense and respond to environmental stimuli. During the progression of sepsis, DCs have been reported to take part in the aberrant immune response and be necessary for survival. Recently, we demonstrated that activation of Aryl Hydrocarbon receptor (AhR), by tryptophan metabolites in DCs, represents a protective mechanism in an experimental animal model of Lipopolysaccharide (LPS)-induced septic shock. Based on these results we examined the role of DCs subsets in sepsis. To determine whether DCs are required for protection against LPS sterile septic shock we used Zbtb46DTR mouse model. We found that Zbtb46DTR chimeras treated with diphtheria toxin are unable to survive after sub-lethal LPS challenge compared to wild-type mice reconstituted with wild-type bone marrow. In addition, naive mice intravenously transferred with LPS-treated DCs survived to a lethal LPS challenge. Notably, we found that mice lacking of CD24+ DCs are extremely susceptible to LPS. Moreover, LPS selectively induces indoleamine-2,3-dioxygenase 1 (IDO1) in bone marrow-derived CD24+. Our data demonstrate for the first time that DCs and specifically CD24+ DCs, play a critical role in sepsis protection.
Dnase1L3 trafficking in inflammasome activation
Authors: Abbott Kennady N , Shi Guilan , Keyel Peter A
Published: January 2023

Pediatric-onset systemic lupus erythematosus (SLE) and the related autoimmune disease hypocomplementemic urticarial vasculitis syndrome (HUVS) are linked to deficiencies in the endonuclease Dnase1L3. Dnase1L3 prevents SLE by digesting apoptotic chromatin extracellularly, but the mechanism by which Dnase1L3 prevents HUVS is unknown. Two autoimmune murine strains, MRL and NZB/W F1, bear the activity-reducing T89I mutation in Dnase1L3. Previous data suggest that this mutation alters Dnase1L3 trafficking. Altered trafficking could allow Dnase1L3 to function intracellularly to regulate inflammation. We previously showed that Dnase1L3 is required for inflammasome activation by facilitating ASC nuclear export, though how Dnase1L3 reaches the nucleus is unknown. Here, we tested the hypothesis that Dnase1L3 trafficking regulates inflammasome activation. We showed that IL-1β secretion was reduced in the presence of T89I. We tested if other activity-reducing mutations alter Dnase1L3 trafficking. We transfected HeLa cells with Dnase1L3 containing point mutations in the signal sequence, C-terminus, and catalytic residues and determined the subcellular localization of Dnase1L3. We also determined their endonuclease activity relative to wild type Dnase1L3. Trafficking studies could establish a physiological intracellular function for Dnase1L3. The relationship between the mutations and localization could explain the mechanism of Dnase1L3 involvement in inflammasome activation. Mutational analysis could serve as a tool to evaluate the structure of Dnase1L3 necessary for inflammasome activation and IL-1β secretion, revealing the function of Dnase1L3 in the pathogenesis of autoimmune diseases like SLE and HUVS.
Hydroxychloroquine Improves Atherosclerosis in SLE
Authors: Niu Haitao , Liu Tianxing , Zhang Shuyang
Published: January 2023

Systemic lupus erythematosus (SLE) patients suffer from earlier premature and severer presentation of coronary artery disease (CAD). By using lupus combining atherosclerosis murine model, we proved that onset of SLE aggravates atherosclerosis. Lupus atheroma were characterized by inflammatory cell infiltration, such as dendritic cells and macrophages. Moreover, excessive immunoglobulin G deposition was also observed in lupus atheroma. Hydroxychloroquine (HCQ) prevented atherosclerosis progression mainly by reversing immune status abnormality caused by SLE. HCQ administration reduced aortic macrophage number and dendritic cell number in lupus mice. HCQ treatment also significantly suppressed IgG deposition. These findings strongly suggest that anti-inflammatory therapies and HCQ provides a new possible strategy for treating SLE patients suffered from atherosclerosis.
Novel functions CD11c+ IgM+ memory B cells suggested from transcriptome analysis
Authors: Levack Russell C , Papillion Amber M , Dishaw Lisa , Winslow Gary
Published: January 2023

Our laboratory has identified a distinct subset of CD11c+ T-bet+ IgM memory B cells during ehrlichial infection in mice. These cells are likely closely related to phenotypically similar B cells that have been described in a range of other conditions, including hepatitis, AIDs, malaria, SLE, and age-related autoimmunity. In order to identify novel functions of CD11c+ T-bet+ IgM memory B cells, we performed RNAseq analysis. The cells were purified from infected mice on day 30 post-infection, and were compared to CD11c-negative CD19+ B cells isolated from the same mice. The CD11c+ B cells exhibited a distinct transcriptomic profile, compared to the CD11c-negative B cells, as over 900 genes were differentially regulated. Differentially-expressed genes included those encoding for integrins, Fc receptors, transcription factors, complement and receptors involved in innate immunity. Differentially-expressed complement-related genes included those encoding for the C1q complex, properdin, and complement factor B, which were all upregulated. Other genes that were upregulated included those encoding the adenosine receptor Adora2a, Bcl-6, which encodes a transcription factor required for GC B cells and the B cell inhibitory FcR, Fcgr2b. These data open new avenues of exploration of CD11c+ T-bet+ B cell function, and will lead to a better understanding of the role of these novel B cells in human health and disease.
Comprehensive analysis of serum and innate immune cells revealed age-related alternations
Authors: Metcalf Talibah U , Wilkinson Peter , Wertheimer Anne , Nikolich-Zugich Janko , Haddad Elias
Published: January 2023

Age-related alterations in immunity have been linked to increased incidence of infections and decreased responses to vaccines in the aging population. These immune dysfunctions, termed immunosenescence, have been linked to the intricate interactions between the innate and adaptive immune responses. The impact of aging on innate immunity remains an open question. We performed a comprehensive analysis of serum and peripheral blood mononuclear cells (PMBCs) isolated from healthy non-frail adults (≤ 40 years) and old individuals (≥ 65 years). The basal level of circulating inflammatory cytokines and chemokines were measured, as well as transcriptome data was analyzed for both ex vivo and agonist stimulated DCs and monocyte subsets. We observed that sorted DC (BDCA-1, -2, and -3) and monocyte (CD14+CD16−,CD14dimCD16+, and CD14+CD16+) subsets were primarily distinct and have unique transcriptional profiles. Under ex vivo conditions, innate subsets did not reveal significant age-related alternations; however, agonist stimulated-monocytes from adults and old subjects did show differences at the transcriptional and functional levels. After stimulation of TLR4 (LPS), TLR7/8 (CLO97), and RIGI (5′-pppRNA), we observed higher production of interferons, CCL8, CCL20, and IL1B in monocytes isolated from adults compared to old individuals. Pathway analysis revealed that innate agonists induced functional pathways in monocytes isolated from both adults and old subjects. Overall, our data reveals a wide range of potential age-associated alternations in pattern recognition receptors (PRRs) signaling and have implications for strategies to enhance the immune response in the context of infection or immunization.
The IL-33/ST2 axis is crucial in type 2 airway responses induced by the Staphylococcus aureus protease SplD
Authors: Krysko Olga , Teufelberger Andrea , Nordengrün Maria Maria , Maes Tania , Bröker Barbara , Bachert Claus
Published: January 2023

Background

Chronic airway inflammatory diseases such as chronic rhinosinusitis with nasal polyps and asthma showed increased nasal Staphylococcus aureus (S. aureus) colonization. Serine protease like protein D (SplD) and other closely related proteases secreted by S. aureus have recently been identified as inducers of allergic asthma in humans and mice but their mechanism of action is largely unknown

Objective

We investigated the role of recombinant SplD in driving Th2-biased responses and IgE formation in a murine model of allergic asthma.

Methods

Allergic asthma was induced in C57BL/6 J wild type mice, TLR4−/− mice and Rag2 Rag2−/− mice by repeated intratracheal applications of SplD. Inflammatory parameters in the airways were assessed by flow cytometry, ELISA, Luminex and immunofluorescence. Serum SplD-specific IgE levels were analysed by ELISA.

Results

We observed that repeated intratracheal exposure to SplD led to IL-33 and eotaxin production, eosinophilia, bronchial hyperreactivity and goblet cell hyperplasia in the airways. Blocking the IL-33 activity using a sST2 receptor significantly decreased the numbers of eosinophils, CD4+ T cells, and IL-5 and IL-13 production by lymph node cells but had no effect on IgE production. SplD-induced airway inflammation and IgE production were largely dependent on the presence of the functional adaptive immune system and independent of TLR4 signalling.

Conclusion

The S. aureus-derived protein SplD is a potent allergen of S. aureus and induces a Th2-biased inflammatory response in the airways in an IL-33-dependent, but TRL4-independent manner. sST2 could be an efficient strategy to interfere with SplD-induced Th2 inflammation, but does not prevent the allergic sensitization.
Pathogenic conversion of RORγt+/Foxp3+ double positive regulatory T cells involves β-catenin up-regulation
Authors: Quandt Jasmin , Haghi Leila , Emmanuel Akinola Olumide , Gounari Fotini
Published: January 2023

The nature of regulatory T cells (Tregs) in cancer is ambivalent. In Colon Cancer (CC) patients and animal models of CC, we previously described a distinct subset of Tregs that suppresses anti-tumor effector T cell functions but simultaneously maintains inflammation. These Tregs co-express the Th17 lineage transcription factor RORγt along with Foxp3. Moreover, we causatively linked the pro-inflammatory skewing of T cells to Wnt/β-catenin-signaling. To elucidate how Wnt/β-catenin signaling orchestrates the induction of pathologic properties in Tregs we analyzed inflammatory bowel disease (IBD) patients. As IBD can progress to CC, this disease presents a unique platform to study the mechanisms underlying the emergence of RORγt+/Foxp3+Tregs. Indeed, in a sub-population of IBD patients we were able to detect RORγt+/Foxp3+Tregs, expressing high levels of β-catenin in the patients’ blood and tissue. To further establish that activated Wnt/β-catenin signaling is responsible for the pro-inflammatory skewing of Foxp3+ Tregs we over-expressed β-catenin Treg-specifically in a conditional Foxp3-Cre driven mouse model. Mice expressing constitutively active β-catenin in Tregs show striking X-linked immune pathology. Males die 4 weeks after birth of a severe scurfy-like phenotype. Although their Tregs are highly activated, they fail to suppress the proliferation of effector T cells. In heterozygous Foxp3-Cre mice, representing natural chimeras harboring wildtype and β-catenin over-expressing Tregs, the relative fitness of genetically altered Tregs is decreased. By further elucidating how Wnt/β-catenin signaling mediates the pathogenic conversion of Tregs, we hope to reveal novel targets for cancer immuno-therapy.
Transcriptomic analysis reveals the potential of Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus to modulate immune system activation related to host-pathogen and damage associated signaling in infected porcine monocytes.
Authors: Miller Laura C , Fleming Damarius , Li Xiangdong , Bayles Darrell , Blecha Frank , Sang Yongming
Published: January 2023

One of the largest risks to the continued stability of the swine industry is by pathogens like porcine reproductive and respiratory syndrome virus (PRRSV) that can decimate production as it spreads among individuals. These infections can be low or highly pathogenic, and because it infects monocytic cells, PRRSV can undermine the innate immune response. This is especially true of the interferon stimulated inflammatory response. To shed light on the effect of PRRSV pathogenicity on activation status of the immune response, we polarized infected and control monocyte-derived cells (mDCs), applied transcriptomic analysis, and predicted protein-protein interactions as networks to elucidate the interconnected biological processes related to low and highly pathogenic PRRSV infection. Differentially expressed genes (DEGs) from each polarized/unpolarized and PRRSV infected/ uninfected comparison was used to predict networks and subsequent molecular functions affected by the viruses and mediators. The analysis uncovered previously uncharacterized evidence of the ability of PRRSV to effect M1 activation by altering expression of genes related to viral defense, the extra-cellular matrix (ECM), toll-like receptor (TLR) signaling, and damage induced inflammatory signaling. Genes showing variability in expression, as well as the most common predicted biological network from the comparisons, were related to cellular structure and inflammatory immune responses. These results supply additional insight into the interplay of PRRSV pathogenicity and immune system evasion by affecting multiple routes of host inflammatory signaling, as well as mDC polarization.
CD11c+ cells are required for lymphocyte trafficking into previously infiltrated pancreatic islets during type 1 diabetes.
Authors: Sandor Adam , Lindsay Robin S , Gebert Matthew J , Bradley Brenda J , Haskins Kathryn , Jacobelli Jordan , Friedman Rachel S
Published: January 2023

Type 1 diabetes (T1D) is a largely T cell mediated autoimmune disease that destroys the beta cells of the pancreatic islets. Inhibition of cell trafficking to an active disease site has been an effective therapy for multiple autoimmune diseases, but the requirements of lymphocyte trafficking to the islets are not fully understood. We show that lymphocyte entry into previously infiltrated islets is dependent on the presence of CD11c+ cells in the islets. T cells and B cells transferred prior to CD11c+ cell depletion were able to enter the islets; whereas, short-term CD11c+ cell depletion rapidly prevented further entry. CD11c+ cells include cells that are highly efficient antigen presenters; however, we show entry of both activated or naïve T cells into previously infiltrated islets was not reliant on antigen. An alternative role for CD11c+ cells in lymphocyte entry into the islets is the production of chemokines and cytokines. These molecules can then either directly recruit lymphocytes through chemoattraction or increased adhesion to the vasculature. Initial experiments show that there are no changes in adhesion to the vasculature after CD11c depletion, suggesting that CD11c+ cells act through recruitment by chemokines following lymphocyte arrest on the vascular endothelium. CD11c+ cells in the islets express high levels of CXCL9, a chemoattractant for T and B lymphocytes. CXCR3, the receptor for CXCL9, is present on both T and B cells in the islets. We hypothesize that CD11c+ cells facilitate the process of extravasation into the islets through chemokine production. Targeting CD11c+ produced chemokine production may be useful therapeutically for the treatment of T1D by preventing T cell entry into remaining or transplanted islets.
The flavonoid galangin induces release of neutrophil extracellular traps and enhances resolution of E. coli pneumonia
Authors: dos Santos Everton OL , Mansouri Soukaina , Sekheri Meriem , El Kebir Driss , Filep Janos G
Published: January 2023

Killing of invading bacteria is essential for efficient control of infection and return to homeostasis. Phagocytosis of E. coli by neutrophils induces neutrophil apoptosis and subsequent efferocytosis, critical control points of the outcome of inflammation. Recently, we found that E. coli DNA impairs phagocytosis of E. coli by human neutrophils, delays neutrophil apoptosis and prolongs E. coli-evoked lung injury in mice. Here, we investigated whether the flavonoid galangin, which was shown to exert anti-inflammatory actions in several inflammation models, could also affect neutrophil function and the outcome of bacterial infection. In human neutrophils, galangin (100μM) did not affect viability or apoptosis; it efficiently countered the potent apoptosis-delaying cue from E. coli DNA by decreasing activation of ERK 1/2 and Akt, leading to Mcl-1 degradation and to collapse of mitochondrial trans-membrane potential. Galangin alone did not affect phagocytosis of live E. coli by neutrophils, and did not restore diminished phagocytosis in the presence of E. coli DNA. Consequently, phagocytosis-mediated bacterial killing was unaffected. By contrast, galangin triggered release of neutrophil extracellular traps (NETs). This was associated with enhanced bacterial killing, which was prevented in the presence of DNAse-I. In mice, galangin administered at the peak of inflammation, accelerated clearance of bacteria and the resolution of pulmonary injury evoked by intratracheal instillation of live E. coli. Collectively, these results identify novel mechanism, overriding survival cues from E. coli DNA and inducing release of NETs, by which galangin could facilitate bacterial clearance and the resolution of inflammation.
The kinetics of CCL27 expression in the skin
Authors: Davila Micha L , Xiong Na
Published: January 2023

The skin is exposed to a wide variety of microorganisms immediately after birth. The newly exposed skin establishes an immune ecosystem within its dermis and epidermis to promote homeostasis between the tissue and the colonizing commensal microflora. Elucidation of the mechanism by which immune cells in the skin are regulated is fundamentally important to understanding how skin homeostasis is established. We have found that the chemokine receptor CCR10 is important for the migration and function of various skin-homing lymphocytes, likely through interaction with its skin-specific ligand CCL27. However the mechanisms that regulate CCL27 expression are poorly understood. Skin sections of newborn, infant and adult mice were stained for CCL27 using a new antibody staining method to determine the kinetics of its expression. We found that CCL27 is markedly upregulated after birth, corresponding to the timeframe of increased effector and regulatory T cell migration to the skin. Further, we compared CCL27 expression in the skin of germ free and specific pathogen free mice to determine the effects of bacterial colonization. These results yield insight into the development of the skin immune system and the mechanisms by which these cells are continuously regulated.
Long term Immune Residence and Cellular Remodeling Follow Viral Infection of the Olfactory Neuroepithelial Barrier
Authors: Moseman E. Ashley , Ciesinski Alexa F. , McGavern Dorian B
Published: January 2023

Barrier immunity, particularly at mucosal surfaces, provides a first line of defense against invading pathogens. Unlike the more classical barrier surfaces in the gut, respiratory tract, genital tract, and skin, the olfactory epithelium within the nose is a specialized barrier structure containing layers of olfactory sensory neurons (OSNs) dedicated to our sense of smell. Because OSNs must contact the nasal airways, they are vulnerable to infection. Upper respiratory viral infection is known to damage the olfactory neuroepithelium and is associated with loss of olfaction. Neurotropic viruses, such as vesicular stomatitis virus (VSV), readily infect OSNs, yet the olfactory epithelial response to infection is unknown. Using a newly generated VSV expressing Cre recombinase, we tracked the fate of virus infected cells after clearance. Numerous cell types, including neural precursors and olfactory ensheathing cells, survived infection. In addition, several months after infection, we detected clonal patches of regenerated mature OSNs derived from previously infected neuroepithelial precursors. VSV infection drove a massive influx of CD45+ hematopoietic cells into the olfactory epithelium that established residence after viral clearance. This included antigen presenting cells and tissue resident CD4+ and CD8+ T cells that localized in and around the regenerating neuroepithelium. We postulate that these immune cells provide a barrier defense against viral rechallenge and might play a role in supporting neuronal regeneration. Collectively, these studies offer insights into how the nasal barrier remodels its neural-immune architecture to restore function and provide enhanced protection following a viral challenge.
Immune regulation in the female reproductive tract by the levonorgestrel-releasing intrauterine device during a baboon model of Chlamydia trachomatis infection
Authors: Eastman Alison J , Bergin Ingrid , Chai Daniel , Carter Kayla , Bell Jason
Published: January 2023

The levonorgestrel (LNG)-releasing intrauterine device (IUD) is increasing in popularity throughout the world, and is an attractive long-acting reversible contraceptive, particularly in areas of high maternal mortality. Its effects on immune responses in the female genital tract are unclear but, of note, LNG is associated with alterations in the CCL3/CCR5 axis. Chlamydia trachomatis (Ct) is the most common bacterial STI worldwide, and untreated infection can lead to infertility. Protective immune responses to Ct likely involve T cell recruitment. We hypothesized that LNG-IUS would result in decreased control of Ct infection in a baboon model. For this study, 20 baboons were randomized into the following groups: LNG-/Ct-, LNG+/Ct-, LNG-/Ct+, and LNG+/Ct+. LNG-IUS was inserted transcervically as in women, and baboons had 3 months to acclimate to the device. Baboons were inoculated with 107 IFU of serovar E Ct weekly for 5 weeks with weekly sampling of vaginal cytokines (assessed by ELISA) and Ct burden. LNG+/Ct+ baboons had delayed clearance in comparison to LNG-/Ct+ baboons (mean infection duration 10 weeks vs 3 weeks, respectively) and maintained higher bacterial burden as determined by assessed by quantitative nucleic acid amplification tests. LNG+/Ct+ baboons had lower expression of CCL3 than LNG-/Ct+ baboons during the first two weeks of infection, suggesting that the LNG-IUS altered T cell recruitment. CCL3 can be induced by IL-1β; in vitro experiments using cervical epithelium show that LNG induces less IL-1β than progesterone. We propose that LNG-IUS decreases IL-1β production in response to Ct infection, thus decreasing T cell recruitment to the infected area via the CCL3/CCR5 axis.
Identification of MAIT Cells in Human Term Decidua
Authors: Chavarria Melina , Vazquez Jessica , Stanic-Kostic Aleksandar
Published: January 2023

Mucosal-associated invariant T (MAIT) cells are a CD161+Vα7+ MR1-restricted T-cell subset, that has a complex relationship in regulating the microbiome and defense from invasive bacteria. Recent discovery of a placental microbiome, and genital mucosa MAIT cells, raises the possibility that MAIT are present at the maternal fetal interface and perform regulatory functions in this setting. As a first step, we analyzed human term decidua to detect MAIT cells and analyze their transcriptional programming.

Decidua was dissected from human term placentas and mononuclear cells (MCs) were isolated by mechanical (GentleMACS) and enzymatic (Collagenase V, DNAse I) disruption. MCs labeled by flurochrome-conjugated antibodies against CD3, 4, 8, 19, 27, 45RO, 161, and TCR Vα7.2 and intracellular Eomes, RORγt, and T-bet. Data was acquired on a 5 laser (355, 405, 488, 562, 633nm), 18-color BD Fortessa cytometer and analyzed with FlowJo 10.1r7.

Data revealed MAIT (CD3+CD161+Vα7.2+) cells in human term decidua, particularly of a memory CD8+ phenotype (CD27+CD45R0+), while fewer of CD4-CD8- and scant CD4+ cells were seen. Transcription factor expression analysis demonstrated uniform expression of PLZF, RORγt, and EOMES in phenotypic MAIT cells, confirming MAIT programming.

MAIT cells are present in human term decidua and add a new dimension to the immunological complex regulating the maternal-fetal interface. Mucosal immune dysregulation is implicated in pregnancy pathology, and our findings open a novel area to mechanistic investigation.
Microbiome-dependent modulation of mucosal immunity at the ocular surface.
Authors: St. Leger Anthony J , Desai Jigar V , Drummond Rebecca , Kugadas Abirami , Almaghrabi Fatimah , Silver Phyllis , Gadjeva Mihaela , Iwakura Yoichiro , Lionakis Michail S , Caspi Rachel R
Published: January 2023

Mucosal sites such as the intestine, oral cavity, nasopharynx, and female reproductive tract all have their associated commensal flora. The surface of the eye (conjunctiva) is also a mucosal site, but existence of a resident microbiome on the ocular surface is highly controversial. We used a mouse model of ocular surface disease to study whether commensal microbes are present in ocular mucosa and modulate immunity. We found that IL-17 is constitutively produced within the conjunctiva-associated lymphoid tissue (CALT) and recruits neutrophils to the ocular surface in the steady state and after a bacterial challenge. IL-17 sources in CALT include γδ T cells, αβ T cells and innate lymphoid cells (ILCs), in that order. Notably, a strain of Corynebacterium isolated from ocular tissue of mice, and known to occur on the ocular surface of humans, induced the conjunctival γδ T cells to secrete IL-17, which in turn modified the local inflammatory signature and barrier function. Elimination of this putative commensal by antibiotic treatment, or its introduction into non-colonized mice, correlated inversely with severity of an experimental Candida albicans infection. Our results thus indicate that a relationship exists between commensals and immune cells at the ocular surface, which is critical for maintenance of homeostasis and host defense within the ocular mucosa.
MAIT cells in the human oral mucosa are activated and perforinlow
Authors: Sobkowiak Michal , Davanian Haleh , Tjernlund Annelie , Gibbs Anna , Nayak Sushrusha , Leeansyah Edwin , Dias Joana , Heymann Robert , Krüger-Weiner Carina , Aleman Soo , Moll Markus , Sällberg-Chen Margaret , Sandberg Johan K
Published: January 2023

Mucosa-associated invariant T (MAIT) cells are a class of non-classical T lymphocytes defined by their invariant Vα7.2-containing MR1-restricted T cell receptor, as well as high expression of CD161. At present, the characteristics of MAIT cells in the oral mucosa are poorly defined. In this study, we have analyzed buccal biopsies and matched blood samples from a cohort of healthy volunteers. MAIT cells were present in the buccal mucosa, with a tendency to cluster around the basal layer, a region also found to contain a higher frequency of MR1-expressing antigen-presenting cells. The frequency of MAIT cells was similar in the buccal mucosa compared to matched peripheral blood, in contrast to other major T cell subsets. However, the buccal mucosal MAIT cells displayed a tissue resident activated profile, with high CD69, HLA-DR and PD-1 expression, as well as a skewed subset distribution with higher representation of CD4/CD8 double-negative cells and of CD8αα+ cells within the CD8+ subset. The Jα chain usage in the MAIT cell TCR was more variable in oral mucosal MAIT cells than in peripheral blood MAIT cells. Functionally, the oral mucosal MAIT cells had lower perforin levels, indicative of a lower cytolytic potential. Interestingly, aspects of the oral MAIT cell population were associated with life-style factors such as consumption of bacterially fermented milk products and alcohol intake. Together, these data show that MAIT cells form a part of the oral mucosal T cell compartment, where they exhibit a tissue-resident activated profile. Thus, MAIT cells are positioned in the oral cavity and equipped to handle resident microbiota as well as invading pathogens.
Modulation of CD4+ T cell polarization using non-canonical co-receptors
Authors: Meeks Christian Matthew , Horton Christopher G
Published: January 2023

CD4+ T helper cells are a diverse group of cells promoting target cell death and production of antibodies. These cells acquire one of several fates following signals through the T cell receptor, co-receptors, and cytokines. The cytokine requirements inducing the polarization of T cell fate has been heavily evaluated and reasonably well defined. However, the influence provided by co-receptors has not been completely elucidated. Traditionally, in vitro T cell polarization assays utilize CD28 stimulation as the primary co-receptor signal required for differentiation. Others have observed the importance of supplementary co-receptors in activation of T helper cells, though little is known about their function in early stage polarization. We hypothesized that the addition of signaling through supplemental co-receptors would alter T helper cell polarization. To conduct these studies, we isolated naïve T cells and polarized them towards Th1 or Tfh fates in the presence or absence of recombinant B7-DC Fc chimera or recombinant B7-H4 Fc chimera. We observed a significant and selective alteration in T helper cell polarization in the presence of these non-canonical co-receptors. These data suggest a complex interplay between cytokine and co-receptor signals that cooperate for robust Th cell polarization. Furthermore, these findings illustrate the potential for novel mechanisms of T cell manipulation for vaccine design and treatment of T cell-mediated diseases.