bluuebunny/crossref_metadata_2025 huggingface dataset:
The Role of NK/DC cross-talk in HIV pathogenesis (94.5)
Authors: Valentin-Torres Alice , Bernstein Helene
Published: January 2023

NK and dendritic cells(DCs) interact with each other inducing reciprocal activation and maturation of both cell types. Interactions between NK cells and DCs are important in controlling viral and bacterial infections. Furthermore, NK/DC interactions potentially link the innate and adaptive immune response via NK cell maturation of DCs. Our laboratory previously identified CD4-expressing NK cells. Using an NK/DC co-culture system, we find that both autologous immature and mature DCs(mDCs) induce CD4 expression on NK cells. CD4 expression requires cell to cell contact and is dependent on the NK/DC ratio. Cellular activation is bidirectional, as CD4+ NK cells induce dendritic cell maturation as assessed by enhanced CD40, CD83, CD86, and HLA-DR expression. Moreover, CD4+ NK cells up-regulate co-stimulatory molecule and HLA-DR expression, better than CD4- NK cells, beyond levels seen in LPS matured DCs. These findings can impact HIV pathogenesis, as mDCs are capable of transferring HIV virus to T cells, a process called trans-infection. We also find that mDCs can trans-infect CD4+ NK cells with HIV, providing a potential mechanism for in vivo HIV-infection of NK cells. In summary, NK/DC crosstalk results in reciprocal activation, including induction of CD4 expression on NK cells and DC maturation. CD4+ NK cells are susceptible to HIV infection, and can be trans-infected by mDCs, supporting a critical role for NK/DC interactions in HIV pathogenesis.
Pigment epithelium-derived factor (PEDF) is an endogenous inflammatory factor (94.6)
Authors: Chavan Sangeeta , Hudson LaQueta , Li Jianhua , Ochani Mahendar , Tracey Kevin
Published: January 2023

Obesity is a major risk factor for conditions ranging from metabolic syndrome and type 2 diabetes mellitus to atherosclerosis. Low grade inflammation leading to insulin resistance is a central feature of the pathophysiology of most obesity-related disorders; however the factors linking these disorders are not well defined. Here a potential mediator of inflammation is identified and characterized in animal models. We identified pigment epithelium-derived factor (PEDF), a noninhibitory serine protease inhibitor, as a mediator released from cultured adipocytes. Adipocyte PEDF levels were elevated after treatment with TNF. Addition of adipocyte-conditioned media to macrophages resulted in increased proinflammatory cytokine production, and this activity was recapitulated by addition of recombinant PEDF to macrophages in culture. PEDF administration to mice mediated increased serum levels of TNF and IL6, and administration of neutralizing PEDF antibodies attenuated this cytokine release. Animals injected with recombinant PEDF developed insulin resistance. Considering these observations, in light of the recently reported elevated levels of circulating PEDF in patients with type 2 diabetes and metabolic syndrome, indicates that adipocyte secreted PEDF may contribute to the development of chronic inflammation and insulin resistance observed in obesity.
Cigarette smoke suppresses the anti-viral capacity of dendritic cells (94.7)
Authors: Castro Shawn , Guerrero-Plata Antonieta
Published: January 2023

The innate immune system is the first line of host defense and against viral pathogens via effector molecules, such as type-I interferon (IFN-α/β). Plasmacytoid dendritic cells (pDC) and monocyte-derived DC (moDC) are potent producers of IFN in response to viruses through the respective activation of recognition toll-like receptors (TLRs) and cytosolic RNA helicases MDA5 and RIG-I. Exposure to environmental tobacco smoke (ETS) is a risk factor associated with an increase in the frequency and severity of respiratory infections. However, the mechanisms by which ETS alters innate immune responses have not been fully elucidated. Therefore, we investigated the ability of pDC and moDC to respond to respiratory syncytial virus (RSV) and human metapneumovirus (hMPV) in the presence of cigarette smoke extract (CSE). In viral infected pDC and moDC, CSE led to a significant dose-dependent decrease in IFN-α production and in pDC alone cytokines IL-10 and IP-10. CSE treatment attenuated IFN induction by TLR agonists TLR7 and TLR9 in pDC and TLR3 in moDC. CSE also significantly decreased TLR7 and TLR9 gene expression in viral infected pDC. In hMPV infected moDC, the presence of CSE down regulated the gene and protein expression of both MDA5 and RIG-I and phosphorylation of PKR regulated eukaryotic initiation factor-2alpha (eIF2α). Overall, these findings provides novel evidence of the IFN signaling pathways abrogated by CSE in viral infected pDC and moDC.
Stress-induced increases in bacterial killing are p38 dependent (94.9)
Authors: Allen Rebecca , Stiner La'Tonia , Hufnagle Amy , Galley Jeffrey , Sheridan John , Bailey Michael
Published: January 2023

Social Disruption (SDR), a murine social stressor, results in an increased reactivity of splenic CD11b+ macrophages to inflammatory stimuli. For example, SDR leads to increased cytokine production and glucocorticoid insensitivity upon lipopolysaccharide stimulation. Moreover, SDR enhances the ability of these macrophages to kill Escherichia coli partly through TLR4-induced expression of inducible nitric oxide synthase (iNOS). The MAPK p38 has extensive known relationships to a number of these SDR characteristics and previous work has also shown that levels of phosphorylated p38 are increased in CD11b+ splenocytes following SDR. As such, p38 is a likely common link among these various stress-induced immunological changes in this model. These experiments were designed to test the hypothesis that inhibiting p38 would significantly reduce the stress-induced increase in killing of E. coli. SDR normally significantly (p < 0.05) increases the percentage of bacteria killed; however, when p38 is inhibited, we found that the percentage killed is reduced to baseline. Additionally, iNOS expression in stressed animals stimulated with E.coli is significantly enhanced in SDR-exposed splenocytes but, p38 inhibition eliminated this increase. Taken together, the negation of the enhanced killing effect and significant decrease in iNOS expression following p38 inhibition supports the hypothesis that the MAPK p38 has an important role in the stress-induced increase in bacterial killing.
ISG15 as a Novel Target for Tumor Immunotherapy (95.13)
Authors: Wood Laurence , Seavey Matthew , Pan Zhen-Kun , Paterson Yvonne
Published: January 2023

Passive immunotherapy is widely used in the treatment of cancer, however, active therapeutic vaccination is achieving increased effectiveness in clinic trials and in preclinical models of cancer. Our laboratory specializes in constructing therapeutic cancer vaccines that utilize Listeria monocytogenes (Lm) as a vector to induce a strong CTL response against tumor-associated antigens (TAAs) when fused to non-hemolytic Listeriolysin O (LLO). Recently, elevated expression of ISG15, an ubiquitin-like protein, has been found in human breast cancer cell lines and clinical breast cancer. After confirming elevated expression of this potential TAA in murine breast cancer models, we constructed an Lm vaccine targeting cells with elevated expression of ISG15. Our recombinant Lm vaccine, Lm-LLO-ISG15, produces and secretes a fusion protein of LLO and ISG15 proficiently. After tumor implantation and subsequent vaccinations, Lm-LLO-ISG15 significantly slowed tumor growth and reduced metastatic lung lesions in comparison to a non-specific Lm control vaccine. In addition, prior vaccination with Lm-LLO-ISG15 offered some protection against implantable tumor challenge and reduced time to tumor progression in a mouse model for sporadic breast cancer. Further work has identified two hypothetical ISG15 H2d-restricted CD8 T cell epitopes. Additional work to determine the mechanism of Lm-LLO-ISG15 anti-tumor efficacy is being pursued.
TGFβ1 Modulation of Inflammation in Premalignant Squamous Lesions is IL-17-dependent and Independent (95.5)
Authors: Mohammed Javed , Ryscavage Andrew , Perez-Lorenzo Rolando , Gunderson Andrew , Blazanin Nicholas , Glick Adam
Published: January 2023

The proinflammatory functions of TGFβ1 are dependent on the local cytokine milieu. Overexpression of TGFβ1 in epidermis provokes an inflammatory response in the skin while its role in skin tumors is still not clear. To evaluate the effects of TGFβ1 in modulating inflammatory cytokine production, we overexpressed TGFβ1 in premalignant primary keratinocytes derived from an inducible TGFβ1 transgenic mouse. TGFβ1 induced expression of TNFα, IL-1β and IL-23a in vitro and conditioned media from these cells polarized naïve T cells to a Th17 phenotype that was dependent on TGFβ1 signaling. Overexpression of TGFβ1 in premalignant squamous lesions resulted in an increase in IL-17 production by CD4+ and γδ+ T cells as well as rapid regression of the tumors. Microarray analysis of gene expression following induction of TGFβ1 in premalignant skin tumors indicated a significant upregulation of proinflammatory genes at day 2. This was followed by an increase in MPO+, F4/80+ and CD8+ cells in the tumors and increase in CD8+ effectors and IFNγ+ cells in the skin draining lymph nodes at day 5. Concomitantly, the percentage of tumor CD11b+Ly6G+ cells was reduced in TGFβ1 overexpressing tumors. Neutralization of IL-17 blocked TGFβ1 induced CD11b+Ly6g- macrophage infiltration in the tumor but did not alter the reduction of CD11b+Ly6g+ cells or tumor regression. Thus, TGFβ1 overexpression causes IL-17 dependent and independent changes in the premalignant tumor inflammatory microenvironment.
Effective prevention and therapy of ovarian cancer with sperm protein 17 vaccination (95.7)
Authors: Chiriva Maurizio , Yu Yuefei , Mirandola Leonardo , Jenkins Marjorie , Chapman Caroline , Cannon Martin , Cobos Everardo , Kast W. Martin
Published: January 2023

Sperm protein (Sp17) is an attractive target for cancer vaccines because of its over-expression in primary and metastatic ovarian cancer (OC). Our studies suggest that a Sp17-based vaccine can induce an enduring defense against OC development in C57BL/6 mice with ID8 OC cells, following prophylactic and therapeutic treatments. Our results show that a SP17 protein based vaccine combined with CpG induced an effective CD8 mediated immune response against Sp17 protein. This is the first time that a mouse counterpart of a cancer testis antigen (Sp17) was shown to be expressed in an OC mouse model. and that vaccination against this antigen significantly controlled tumor growth . The CpG-adjuvanted vaccine are activating Th17 cells while inhibiting immunosuppressive T-reg cells, suggesting a reprogramming of active cellular-mediated response against Sp17-expressing tumor cells. Taken together, these results indicate that prophylactic and therapeutic vaccinations can induce long-standing protection against OC and delay tumor growth, suggesting that this strategy may provide additional treatments of human OC and the prevention of disease onset in women with a family history of OC.
Cannabinoid receptors as novel targets in the treatment of T cell-mediated fibrosis (96.26)
Authors: Kanada Shunsuke , Nagarkatti Mitzi , Nagarkatti Prakash
Published: January 2023

Liver diseases are one of the main causes of mortality in the world, especially chronic hepatitis leading to the development of liver cirrhosis or liver cancer. Previous studies have shown that natural cannabinoids such as Δ9-tetrahydrocannabinol (THC) exhibit immunosuppressive and anti-inflammatory properties. In this study, we investigated the effect of THC on chronic hepatitis caused by repeated toxic insult. For the chronic hepatitis, ConA or CCl4 was administered to the C57BL/6 mice for several weeks. THC was injected after every toxic insult. Livers were collected for the histological assessment and hepatic cells were harvested for flow cytometric analysis to characterize the nature of hepatic inflammatory cells. Fibrosis related gene expression in injured liver was investigated by real time PCR. The results showed that THC treatment suppressed the mRNA expression of liver fibrosis markers and flow cytometric analysis suggested that THC treatment decreased the inflammatory cell accumulation in the liver in ConA-induced model. On the contrary, THC did not suppress the progression of fibrosis in CCl4-induced model. Together, these results demonstrated that THC can suppress chronic autoimmune hepatitis but not chemical-induced hepatitis and that cannabinoid receptors can be the new therapeutic targets preventing immune cell-mediated liver injury (Supported in part by NIH grants R01AI053703, R01ES09098, R01 AI058300, R01DA016545, R01HL058641, R01ES019313 and P01AT003961)
OR3: an anti-TCR autoantibody and immunomodulation (96.7)
Authors: Ranganathan Parvathi , Schluter Samuel
Published: January 2023

The presence of low levels of circulating self-reactive autoantibodies (AAbs) is a normal feature of a healthy immune system. One suggested function for autoantibodies is the control of autoreactivity and immune homeostasis. Healthy humans spontaneously express low levels of serum AAbs directed against defined epitopes of the T Cell Receptor (TCR) Vβ chains that can be markedly elevated in individuals with autoimmune diseases such as Rheumatoid Arthritis (RA). Here, we describe a monoclonal IgM AAb, OR3, derived from peripheral blood B cells of a patient with RA. OR3 was selected for its ability to bind to residues 23-38 of the complementarity determining region 1 (CDR1) segment of human Vβ8.1. It binds to Jurkat T cells expressing human Vβ8.1 and to ovalbumin-specific transgenic murine DO11.10 T cells. OR3 significantly reduces ova peptide stimulated activation of DO11.10 CD4+ T cells and inhibits IFN-γ, IL-2 and IL-10 cytokine secretion by antigen specific DO11.10 T cells. OR3 also blocks antigen induced proliferation of DO11.10 T cells, but does not initiate apoptosis. The interaction between the TCR and MHC:peptide is a critical point in the activation of the adaptive immune response, disruption of which, can lead to a global dampening of the immune response. We show evidence for the development of a unique anti-TCR autoantibody that binds to only a subset of T cells and possesses potent blocking and anti-inflammatory properties that can be harnessed for therapeutic use.
Induction of regulatory T cells by a novel immunoregulatory therapy suppresses the development of allergic airways disease (97.10)
Authors: Thorburn Alison , Foster Paul , Gibson Peter , Hansbro Philip
Published: January 2023

Evidence suggests that exposure to Streptococcus pneumoniae may be protective against asthma. We have identified key S. pneumoniae components that form an immunoregulatory therapy (IRT) and suppress the development of allergic airways disease (AAD) using mouse models. AAD was induced in adult BALB/c mice by intraperitoneal sensitization and intranasal challenge with ovalbumin. At the time of sensitization, the IRT was administered intratracheally. IRT treatment suppressed hallmark features of AAD including eosinophil influx, local and systemic Th2 cytokines, airways hyperresponsiveness and mucus hypersecretion. A time course analysis showed that the IRT induced CD4+CD25+Foxp3+ Tregs in the lymph nodes 4 days post treatment. This was associated with increased IL-2, TGF-β and Foxp3 expression in the lungs. IRT treatment also increased the proportion of CD4+CD25+/- cells expressing Foxp3 in the lungs and expression of markers associated with Treg function, during the induction of AAD. Anti-CD25 administration 3 days before IRT treatment abolished the suppressive effects on AAD. However, anti-CD25 administration 9 days after IRT treatment had no effect. This indicated a requirement for IL-2 signalling via CD25 (IL-2R) during the induction of Tregs by the IRT treatment. In conclusion, a S. pneumoniae-based IRT induces CD25-dependent Tregs, suppresses the development of AAD and may be useful as a novel therapy for asthma.
Role of IL-17 in a murine model of allergen immunotherapy for allergic asthma (97.5)
Authors: Boehm Livia , Soo-Becker Kinga , Übel Caroline , Schuster Petra , Karwot Roman , Lehr Hans-Anton , Schild Hansjörg , Finotto Susetta
Published: January 2023

Human studies have demonstrated that allergen specific immunotherapy (IT) is validated for allergic diseases. IT involves repeated subcutaneous administration of the allergen following allergen sensitization. We now established a mouse model in which subcutaneous applications with allergen suppressed the asthma-like manifestations. In this model we found an antigen dependent decrease of airway hyperresponsiveness, IgE and lung eosinophils mediated inflammation as compared to untreated mice. Consistently, Th2 as well as Th17 type cytokines were found to be reduced in the lung of antigen treated mice. Moreover, we found that this established subcutaneous immunotherapy decreased the number of lung gamma-delta (γδ)+IL17+ T cells. IL-17 production by αβ T helper cells (Th17) and the conditions of their development received much attention over the last few years, however the role of IL-17-producing γδ T cells was not fully investigated. In fact, it has been recently reported that γδ T cells represent a vast majority of IL-17-producing cells in murine lung. Revealing the role of IL-17 and lung γδ+ IL17+ T cells in asthma and in an immunotherapy model will probably lead to therapeutical improvement for this disease.
Serum but not secreted Ag-specific IgA inhibits oral Ag induced anaphylaxis. (97.6)
Authors: Strait Richard , Mahler Ashley , Hogan Simon , Khodoun Marat , Finkelman Fred
Published: January 2023

Epidemiologic evidence suggests that IgA, the main mucosal Ig isotype, inhibits allergy, but it is not known whether it suppresses anaphylaxis. We used mouse models to evaluate whether IgA can suppress systemic anaphylactic responses to ingested allergens by neutralizing them before or after their systemic absorption. Passive systemic anaphylaxis (PSA), measured as hypothermia, was induced by sensitizing mice with IgE anti-TNP mAb, then challenging them orally (og) with TNP-OVA. The ability of IgA to inhibit PSA by neutralizing Ag prior to or after systemic absorption was evaluated by pretreating mice with IgA anti-TNP mAb either og or iv. TNP-OVA was detectible in serum by 5 min after og administration. PSA developed within 5 min and was inhibited by iv, but not og IgA. Extent of inhibition was proportional to the IgA dose and Fcα/μR- and FcγRIIb-independent. Intestinal anaphylaxis (IA), defined as diarrhea <60 minutes after OVA challenge, was induced by priming mice ip with OVA/alum, then challenging them og 3x/wk with OVA. IA development was delayed in J-chain- and PIgR-deficient mice, which have markedly increased serum IgA and decreased intestinal IgA. Thus, Ag-specific IgA acts in the blood but not the gut to inhibit systemic and intestinal anaphylaxis. This indicates that ingested antigen must be absorbed systemically to induce systemic anaphylaxis and suggests that immunotherapy should aim at increasing blood, rather than gut levels of non-IgE Abs to food allergens.
Administration of dendritic cells signaled via the neurokinin 1 receptor favors type-1 cellular immunity by mechanisms involving exogenous and endogenous dendritic cell populations (98.12)
Authors: Janelsins Brian , Mathers Alicia , Tkacheva Olga , Erdos Geza , Shufesky William , Morelli Adrian , Larregina Adriana
Published: January 2023

Induction of type-1 immunity is the focus of current dendritic cell (DC)-vaccination approaches for tumors and intracellular infections. The pro-inflammatory neuropeptides, substance P and hemokinin-1, favor cell-mediated immunity by binding the neurokinin 1 receptor (NK1R). DCs express functional NK1R; however, the ability of NK1R agonists to modulate the T cell stimulatory and biasing functions of DCs remains unknown. In the present work, we analyzed the effects of ex vivo-generated DCs signaled via the NK1R to exert T cell stimulatory and type-1 biasing functions. We demonstrate that agonistic signaling via the NK1R promotes the maturation of murine bone marrow-derived DCs (BMDCs) and inhibits their secretion of IL-10. Adoptive transfer of NK1R-signaled BMDCs favors enhanced Ag specific type-1 CD4+ (Th1) and CD8+ (CTL/Tc1) responses in vivo. The individual roles of adoptively transferred BMDCs and endogenous DCs, under our experimental conditions, were further addressed by comparing the development of type-1 immunity in C57BL/6 wild type, IL-12p35-/- and CD11c+DTR mice (the latter depleted of endogenous DCs). Our results demonstrate that generation of type-1 immunity required inhibition of IL-10 by transferred BMDCs and secretion of IL-12p70 by endogenous DCs. Together, our data strongly suggest that adoptive transfer of NK1R-signaled BMDCs promotes type-1 immunity by mechanisms involving both the adoptively transferred and endogenous DC populations.
Prostaglandin E2 Receptor Signaling Induces IL-23p19 Expression through CREB and STAT-3 Transcriptional Regulation (98.2)
Authors: Kocieda Virginia , Adhikary Sabina , Emig Fran , Ganea Doina
Published: January 2023

Previously, we have shown that PGE2 inhibits IL-12p35 expression & promotes IL-23p19 expression in dendritic cells (DC) treated with LPS or Poly I:C. The effect is mediated through EP4 and to a lesser degree EP2 receptors and induction of cAMP. This study is focused on essential transcriptional factors and signaling pathways involved in PGE1-OH (EP4 agonist) induced p19 expression. Previous studies reported involvement of NFkB (c-Rel), ATF-2, SMAD-3 & AP-1. CREB is a target of cAMP-activated PKA and two CREB binding sites have been identified in the p19 promoter. Therefore, CREB is a potential transcription factor activated by PGE. Experiments with p19-luciferase constructs containing wt & CREB-mutated sites indicate the importance of CREB sites in p19 transcriptional activity. PGE2/PGE1-OH also induced STAT-3 tyrosine phosphorylation in DC and in the DC cell line. Blocking STAT-3 phosphorylation with the chemical inhibitor S3I-201 and the specific peptide inhibitor PpYLKTK-mts reduced p19 expression in DC. Although no STAT-3 binding site was found in the p19 promoter, STAT-3 was proposed to act as a co-activator for c-Rel to induce p19 expression. These data show for the first time that CREB and STAT-3 are important regulators of PGE2-induced p19 expression in DC. These observations show the versatility that prostaglandins have to differentially regulate expression patterns of cytokines that orchestrate an immune response.
The activation state of Ly-6Chi inflammatory monocytes determines their immune regulatory function (98.23)
Authors: Zhu Bing , Kennedy Jennifer , Wang Yue , Sandoval Garcia Carolina , Elyaman Wassim , Xiao Sheng , Khoury Samia
Published: January 2023

We reported that CD11b+Ly-6ChiLy-6G- inflammatory monocytes (IMCs) obtained after EAE immunization strongly induced apoptosis in pre-activated CD4 T cells through the production of nitric oxide. However, it was recently reported that transfer of wild type IMCs worsened EAE disease in GM-CSF-/- or CCL2-/- recipients, suggesting a pathogenic function of IMCs. We hypothesized that the immune function of IMCs depends on their activation state. We found that ex vivo purified IMCs induced antigen-specific proliferation of naïve TCR transgenic CD4 T cells, suggesting that these IMCs can function as antigen presenting cells. In contrast, when the same IMCs were first activated with IFN-γ, LPS and GM-CSF, they strongly suppressed T cell proliferation induced by professional APCs, which could be rescued by NOS2 inhibition. Similar dichotomy of T cell regulatory function was observed in inflammatory dendritic cells (IDCs) differentiated from IMCs with GM-CSF. In addition, IMCs isolated from the central nervous system (CNS) of MOG-immunized B6 mouse before EAE onset supported T cell proliferation, survival and Th1/Th17 differentiation, but CNS IMCs obtained at EAE peak suppressed T cell proliferation and differentiation. Furthermore, transfer of in vitro activated IMC and IDCs markedly suppressed EAE severity and reduced CNS inflammation. In summary, we demonstrate that the immune function of IMCs and IDCs is plastic and dependent on their activation state.
Requirement of voltage-gated calcium (Cav) channels in T cell receptor-mediated calcium response and effector functions of T lymphocytes (99.13)
Authors: Jha Mithilesh , Badou Abdallah , Flavell Richard
Published: January 2023

Calcium signals are required for the activation, differentiation and effector functions of T lymphocytes. However, the molecular machinery required for calcium influx upon TCR stimulation of T cells is not yet clear. Previously, we have shown that members of the Cav1 family of voltage-gated calcium channels are expressed in T lymphocytes and regulate calcium influx into these cells. Recently, we have demonstrated an essential role for the beta3 regulatory subunit of Cav channels in the maintenance of naive CD8 T cells since deficiency in beta3 resulted in a profound survival defect of naïve CD8 T cells. Here, we investigated the expression and function of Cav1 channels in differentiated CD4 T lymphocyte populations and discovered a novel role of Cav1 channels in effector helper T cell population. We found a selective expression of Cav1.3 in effector Th2 cells but not naïve CD4 T cells or effector Th1 cells. Both TCR-mediated calcium response and NFATs nuclear localization were severely impaired in Cav1.3-deficient Th2 cells. A profound and selective impairment of Th2 effector cytokine production was observed in vitro and in vivo after immunization of mice. Our data demonstrate a novel and essential role of Cav1 channels in different T cell functions. Therefore, Cav1 channels represent a specific and new therapeutic target for T cell-mediated inflammatory diseases.
Humoral responses require a multi-stage T:B lymphocyte interaction process (99.5)
Authors: Cannons Jennifer , Qi Hai , Lu Kristina , Ghai Mala , Gomez-Rodriguez Julio , Germain Ronald , Schwartzberg Pamela
Published: January 2023

X-linked lymphoproliferative (XLP) disease is an immune disorder caused by mutations encoding SLAM-associated protein (SAP, Sh2d1a), an adaptor that facilitates Fyn recruitment to SLAM family of receptors, including 2B4, Ly9, SLAM, CD84 and Ly108. SAP-/- CD4 T cells exhibit a selective impairment in adhesion to antigen presenting B cells but not dendritic cells (DC), resulting in defective germinal centre (GC) formation. However, the nature of this selective adhesion defect is unclear. We provide evidence that while T:DC interactions are primarily integrin-dependent, T:B cell interactions have an early integrin-dependent component and a sustained phase that also requires SAP. We further demonstrate that T cells adhere to the SLAM family members, CD84 and Ly108, in a SAP-dependent manner. Both CD84 and Ly108 are highly expressed on T follicular helper (TFH) cells, a specialized subset of CD4 T cells that reside within GCs and are critical for their formation. Cd84-/- mice exhibit a partial defect in GC responses to T-dependent antigens. This defect correlated with impaired cognate T:B cell conjugate pairing observed in vivo by dynamic imaging as well as reduced TFH cell differentiation and function. Strikingly, the combined loss of CD84 and Ly108 markedly reduced T:B conjugate pairing in vitro. Thus, our results identify SLAM family members as critical receptors involved in the dynamic regulation of sustained T:B cell interactions.
Functional effects of ERAAP polymorphisms linked to ankylosing spondylitis (100.18)
Authors: Reeves Emma , Elliott Tim , James Edd
Published: January 2023

Optimal peptide loading on the MHC I molecule is essential for antigen presentation to CD8+ cytotoxic T lymphocytes. Endoplasmic Reticulum Aminopeptidase associated with Antigen Processing (ERAAP) is responsible for N-terminal trimming of peptides to optimal length for stable loading and presentation through MHC I. In humans, ERAAP is also known to play a role in pro-inflammatory cytokine receptor cleavage as well as regulating blood pressure and angiogenesis. Recently, a single nucleotide polymorphism (SNP) linkage analysis study has identified ERAAP as being associated with increased risk of the autoimmune inflammatory disorder, ankylosing spondylitis (AS). Here, we show that SNP mutation of ERAAP alters its ability to trim peptides. Transfection of ERAAP-/- cells with ERAAP molecules incorporating each of the SNPs identified in the linkage analysis show a hierarchy of reduced function. More pronounced functional phenotypes are revealed with additional mutations suggesting a cumulative effect. Although the mechanism for the effect of SNP mutation on ERAAP function is unclear, it appears that they cause a dramatic effect on peptide trimming.
Self-antigen peptidome carried by the human lymph (100.3)
Authors: Santambrogio Laura , Clement Cristina , Stern Lawrence
Published: January 2023

The pre-nodal afferent lymph is the fluid which directly derives from the extracellular milieu of every parenchymal organ collecting catabolic and metabolic products. A comprehensive qualitative and quantitative investigation of the self-antigenic repertoire transported by the human lymph was the goal of our investigation. A major difference between lymph and plasma could be visualized by FPLC and 2D gel in the amount of low molecular weight products corresponding to peptide fragments. Naturally processed peptides in normal pre-nodal human lymph were fractionated by HPLC and characterized by MS/MS. Analysis of more then 300 sequences, identified self-peptides derived from both intracellular and extracellular proteins, transported by the human lymph. Quantitative analysis established that some of these peptides are present in nanomolar concentration. The peptidome, generated by physiological tissue catabolism and transported by the pre-nodal lymph, is in addition to the self-peptidome generated in endosomal compartment. Unlike self antigen processed by local or nodal APC, which produce epitopes constrained by the endosomal processing activity, self antigens present in the lymph could derived from a wider variety of processing pathways; including caspases, involved in cellular apoptosis, ADAM and MMPs involved in receptors/cytokines editing and matrix remodeling. Altogether, expanding the tissue-specific self-repertoire available for the maintenance of immunological tolerance.
Preterm labor leads to activation and proliferation of maternal T cells which recognize the paternal alloantigen via the indirect pathway (100.46)
Authors: Wegorzewska Marta , Nijagal Amar , Le Tom , Tucker Andrew , Tang Qizhi , MacKenzie Tippi
Published: January 2023

Purpose: Multiple tolerance mechanisms protect the semi-allogeneic fetus from attack by the maternal adaptive immune system. We hypothesize preterm labor (PTL) may represent a breakdown of tolerance to the allogeneic fetus. Methods: B6 females were bred to BALB/c males and CFSE-labeled TCR transgenic T cells (TCR75 CD4, indirect; 4C CD4, direct; 2C CD8, direct) which recognize BALB/c antigen were adoptively transferred on E12.5 to study the roles of indirect and direct antigen presentation. PTL was induced by LPS on E13.5 and mice were analyzed at E18.5 for expression of CD44, CD62L, and CFSE. Controls included syngeneic matings with PTL and allogeneic matings without PTL. Results: 4C and 2C T cells did not proliferate during PTL. TCR75 cells proliferated and upregulated CD44 in allogeneic pregnancies with PTL compared to those without PTL (p<0.05) or syngeneic pregnancies with PTL (p<0.05). Although TCR75 cells proliferated in all allogeneic pregnancies, they only accumulated in animals undergoing PTL, suggesting that apoptosis of alloreactive T cells maintains maternal-fetal tolerance in normal pregnancies. Conclusions: PTL leads to proliferation, activation, and accumulation of maternal T cells that recognize the paternal alloantigen via the indirect pathway, indicating these cells participate in the pathogenesis of the disease. Understanding their role may uncover novel pathways for therapeutic intervention in PTL.
Myosin 1c participates in B cell cytoskeleton rearrangements, is recruited to the immunological synapse and contributes to antigen presentation. (100.48)
Authors: Maravillas-Montero Jose , Gillespie Peter , Patiño-Lopez Genaro , Shaw Stephen , Santos-Argumedo Leopoldo
Published: January 2023

Myosin 1c (Myo1c) is a member of the unconventional class I myosins of vertebrates, which directly link the plasma membrane with the microfilament cortical web. Although this molecular motor has been implicated in cell functions such as cytoskeleton organization, cell motility, nuclear transcription and endocytosis; its role in the hematopoietic cells is largely unknown. We found that Myo1c is abundantly expressed in murine B lymphocytes and is preferentially located at the plasma membrane, especially in peripheral processes such as microvilli. It was observed not only that this motor concentrates at the growing membrane protrusions generated during B cell spreading, but also that is actively recruited to the immune synapse. Interestingly, Myo1c was detected in the lipid rafts of B cells and showed a strong colocalization with MHC-II, particularly after crosslinking of these molecules. By transfecting a dominant negative form of Myo1c, we also detected alterations in spreading and antigen presentation ability of these cells. All these data suggest that Myo1c could be involved in the B lymphocyte cytoskeleton dynamics and membrane protein anchoring and/or sorting, during cell-cell contacts, including those related with CD4+ T cell activation.
Superimposed viral infection recruits CD8+ T cells to the CNS that exacerbate EAE (101.13)
Authors: Rainey-Barger Emily , Rao Praveen , Segal Benjamin , Irani David
Published: January 2023

Epidemiological data support a link between viral infection and multiple sclerosis (MS), but the underlying cellular and molecular mechanisms through which such infections might initiate or alter the disease remain unknown. Likewise, few studies have examined the effects of a superimposed viral infection on experimental autoimmune encephalomyelitis (EAE), an important mouse model of MS. We find that an otherwise asymptomatic alphavirus infection introduced directly into the brains of mice during the pre-clinical phase of active immunization EAE significantly accelerates disease progression compared to sham-inoculated controls. While peripheral T cell responses and the proportions of CD4+ T cells in spinal cords of infected and sham-inoculated EAE mice are similar, the CD8+:CD4+ ratio and the total number of CD8+ T cells increase in the spinal cords of infected EAE hosts. Furthermore, this infection does not exacerbate EAE symptoms compared to sham inoculation in CD8-deficient hosts. These findings suggest that CD8+ T cells activated by a superimposed alphavirus infection promote autoimmune mechanisms that drive EAE pathogenesis, possibly via production of pro-inflammatory cytokines or cytotoxic molecules. Our dual disease paradigm will allow us to determine how acute viral infection influences peripheral and central nervous system (CNS) immune responses during EAE as well as to clarify the role of CD8+ T cells in autoimmune CNS demyelination.
Monocytes promote the formation of autoreactive Th17 cells in inflammatory arthritis (101.23)
Authors: Simons Donald , Oh Soyoung , Garcia Victoria , Mroz Lori , Caton Andrew
Published: January 2023

Th17 cells have been implicated in a number of autoimmune diseases and their animal models including inflammatory arthritis. How these cells arise in the context of autoimmunity is not well understood. We recently described a mouse model of autoimmune arthritis in which mice expressing influenza hemagglutinin (HA) as a self-antigen under the control of a MHC Class II promoter (HACII), and co-expressing an HA-specific CD4+ TCR (TS1), spontaneously develop inflammatory arthritis by an IL-17-dependent mechanism. Here we have used these TS1xHACII mice to investigate the role of specific cell types in the activation of autoreactive CD4+ Th17 cells. We have found that monocytes are selectively enriched in the spleens and LNs of arthritic TS1xHACII mice relative to mice expressing the HACII transgene alone. Moreover, monocytes purified from the spleens of arthritic TS1xHACII and HACII mice had profound differences in their abilities to activate HA-specific T cells in vitro: monocytes from HACII mice were potent inducers of Th1 cells, whereas those from TS1xHACII mice were significantly more efficient at inducing Th17 cells. Monocytes from TS1xHACII mice also had an enhanced capacity to induce Th17 cells relative to B cells and cDCs isolated in parallel from the same organs. Collectively, these data suggest that monocytes can play an important role in the development of Th17 cells in vivo, and can participate in an autoimmune response that results in inflammatory arthritis.
Pathogenesis of rheumatoid arthritis in a fibrinogen-induced mouse model of autoimmunity (101.8)
Authors: Cordova Kristen , Baker Rocky , Barbour Gene , Haskins Kathryn , Holers V.
Published: January 2023

Citrullinated proteins, derived from the conversion of peptidyl-arginine to peptidyl-citrulline, are present in the joints of rheumatoid arthritis (RA) patients who also produce high levels of anti-citrullinated protein antibodies (ACPAs). T cells also have a role in RA and are thought to be involved in initiating, controlling and driving antigen-specific immune responses. We and others found that mice immunized with human fibrinogen (HF) or human citrullinated fibrinogen (HCF) develop mild inflammatory arthritis and peripheral T cells proliferate in response to HCF and HF. To determine if HCF-reactive T cells contribute to disease, DBA1/J mice were immunized with HCF and at various time points, T cells were assessed directly ex vivo or cultured in vitro. T cells were evaluated ex vivo for the presence of activation markers and cytokine production after antigen stimulation; T cell lines were cultured in vitro with antigen and IL-2 and evaluated in the same manner. Following two immunizations, mice produced CD4 T cells with an activated phenotype (CD44hi+CD62L-); TNFα and IL-17 were produced by CD4 T cells in response to HCF. T cell lines produced high levels of TNFα, IL-17, and IL-6 when stimulated by HCF, and one line also produced IFNγ. In sum, CD4 T cells are activated during disease progression and exhibit an inflammatory phenotype that may be pathogenic. T cell lines are being evaluated in vivo to determine if they can precipitate or accelerate disease.
Type I IFN signaling controls T lymphocyte transmigration into the airways (102.10)
Authors: McNally Beth , Ye Fang , Willette Meredith , Partida-Sanchez Santiago , Durbin Joan , Flano Emilio
Published: January 2023

Our current understanding of how T lymphocytes enter the airways in response to respiratory tract infections is limited. Here we show a role for type I interferon (IFN) signaling in regulating T lymphocyte transmigration into the airway mucosa. IFNα receptor (IFNAR) deficiency led to a reduction in the number of T lymphocytes that entered the airways but not the lung parenchyma. This effect was mediated both at the T lymphocyte and epithelial cell level. IFNAR-deficient airway epithelial cells failed to express the chemoattractants CCL3-5, CCL8 and CXCL9-11, and did not up-regulate the adhesion molecules ICAM1 and VCAM1. Lack of IFNAR signaling on T lymphocytes impaired Ly-6C expression and lymphocyte function-associated antigen-1 (LFA-1) clustering, which resulted in reduced cell adhesion. Finally, we show that IFNAR-deficiency inhibited T lymphocyte recruitment to the airways during respiratory syncytial virus (RSV) infection.
Shear forces and high affinity LFA-1 drive PMN recruitment (102.18)
Authors: Dixit Neha , Yamayoshi Itsukyo , Nazarian Ari , Simon Scott
Published: January 2023

Acute inflammation triggers the innate immune response of neutrophils that efficiently traffic from the blood stream to concentrate at high numbers at the site of tissue infection or wounding. A gatekeeper in this process is activation of β2-integrins, which forms bond clusters with ICAM-1 on the endothelial surface. These bond clusters serve dual functions of providing adhesive strength to anchor neutrophils under the shear forces of blood flow and directional guidance for cell polarization and subsequent transmigration on inflamed endothelium. We hypothesized that shear forces transmitted through high affinity LFA-1 facilitates their cooperation with the calcium release activated channel (CRAC) Orai1 in directing localized cytoskeletal activation and directed migration. Employing vascular mimetic microfluidic channels we observed neutrophils arresting on a substrate of either ICAM-1 or allosteric antibodies that stabilize a high or low affinity conformation of LFA-1. Neutrophils captured via low affinity LFA-1 did not exhibit intracellular calcium flux, F-actin polymerization, cell polarization, or directional migration under shear flow. In contrast, high affinity LFA-1 provided orientation along a uropod-pseudopod axis that required calcium flux through Orai1. We demonstrate how the shear stress of blood flow can transduce distinct outside-in signals at focal sites of high affinity LFA-1 that provides contact mediated guidance for neutrophil emigration.
CD8+ T cell trafficking post primary Influenza A virus infection (102.26)
Authors: Miao Hongyu , Topham David , Wu Hulin , Zand Martin
Published: January 2023

An important model for studying influenza A virus (IAV) infection is the acute antiviral immune responses. It is of a great interest to understand the growth and migration of virus-specific CD8+ T cells post IAV infection. However, the biological parameters that determine the kinetics of immune cells currently cannot be directly measured by experiments techniques. In this work, we developed mechanistic and semi-mechanistic mathematical models to infer the expansion, trafficking, and disappearance of CD8+ T cells in lymph node, spleen and lung from mice with primary influenza A infection. It was found that the spleen is a major contributor of effector CD8+ T cells to the lung in comparison with the lymph node. In addition, it is veryfied that the death of CD8+ T cells in lung follows a time-varying pattern. Tissues other than the lung and lymphatic organs notably contribute to the response in the lung, but not dominate the contributions from spleen. Our efforts provide a better understanding of the primary CD8+ T cell response to influenza infection.
Immunologic profile changes associated with aging in rhesus macaques (104.2)
Authors: Didier Elizabeth , Sugimoto Chie , Bowers Lisa , Bohm Rudolf , Gilbert Margaret , Barnes Mary , Pattison Melissa , Slisarenko Nadia , Falkenstein Katherine , Kuroda Marcelo
Published: January 2023

Immunosenescence is a public health concern. This study was to characterize the peripheral immune system in rhesus macaques of varying ages. Flow cytometry and multiplex cytokine levels were evaluated in blood from 35 monkeys (3.2-20 years old). Linear regression was used to define statistically significant changes (i.e. P<0.05) with age. Significant declines were observed in numbers of neutrophils, eosinophils, basophils, and lymphocytes in blood with increasing age. No changes in numbers of monocytes, dendritic cells (DC), RBCs, or platelets were noted. Among lymphocytes, B cell numbers declined while NK cells and T cells were unchanged with increasing age. Within the T cell population, however, significant declines in naive CD4+ and CD8+ T cells, and γδ+T cells were seen with increasing age, whereas the number of CD8+ effector-memory T cells increased with age. Numbers of DCs did not change with advancing age, but the turnover rate of pDCs was higher in the 18-20 year-old group compared to younger groups. Of the cytokines tested in plasma using the Milliplex non-human primate cytokine (23-mer) kit, TNF-α, IL-2, MCP-1, sCD40L, TGF-α, VEGF, and IL-18 levels significantly increased with increasing age whereas IL-5 was the only cytokine tested that decreased with increasing age. These early results provide a basis to further characterize immunosenescence in a non-human primate model.
Early signaling events regulate Fas-induced apoptosis in CD4+ T cells (104.5)
Authors: Ramaswamy Madhu , Do Thao , Cora-Morges Adrian , Cruz Anthony , Siegel Richard
Published: January 2023

Autoreactive CD4+ T cells can be eliminated by the TNF superfamily member Fas through a process termed Restimulation Induced Cell Death (RICD). Previously, we have shown that effector memory CD4 T cells were the most Fas-induced apoptosis sensitive CD4 subset. This was due to efficient recruitment and activation of components of the death-inducing signaling complex (DISC) indicating a role for early signaling events in apoptosis regulation. To discover novel proteins recruited to the DISC, we used 2D-DIGE and Mass Spectrometry on Fas sensitive and resistant CTCL lines to identify differential binding partners in the Fas-DISC. We have identified a number of proteins including a SNARE family protein previously found to be involved in exocytosis. In transfection experiments, this protein interacted with Fas in a manner independent of the Fas death domain as shown by co-immunoprecipitation and Fluorescence Resonance Energy Transfer (FRET) between constructs fused to CFP and YFP. The endogenous protein preferentially interacted with Fas in cell lines and T cell subsets susceptible to Fas-induced cell death. These results suggest that that this protein is a novel component of the DISC and may regulate early events in Fas signaling, possibly through affecting receptor endocytosis or recycling to the plasma membrane. The mechanism by which this protein affects assembly and activation of the Fas DISC, and other proteins found in this proteomic screen are currently being investigated.
Defective CD8 T cell memory generated during latent gammaherpesvirus infection (105.11)
Authors: Freeman Michael , Burkum Claire , Jensen Meghan , Woodland David , Blackman Marcia
Published: January 2023

Antiviral CD8 T cells are essential for protection during murine gammaheresvirus-68 (γHV68) infection. We recently identified a large panel of epitope-specific CD8 T cell responses that were differentially regulated by antigen presentation during the establishment of viral latency, but how these T cell responses are maintained throughout latent infection is unclear. Here we show that recent thymic emigrants are primed during latency, accumulate over time, and can proliferate in response to experimentally-induced viral reactivation. We found that in contrast to ORF6487- and ORF61524-specific responses, CD8 T cells specific for the ORF75c940 epitope primed during latency were deficient in their ability to respond during reactivation. This defect was not a result of impaired generation of ORF75c940-specific cells during latency or a lack of antigen presentation during reactivation, but rather appears to be a consequence of reduced proliferation following stimulation. The finding of epitope-specific differences in proliferative capacity suggests that the functionality of latency-primed responses may be regulated by differential antigen presentation or viral gene expression during latency. These findings have important implications for the development of therapeutic vaccine strategies against latent gammaherpesvirus infections.