bluuebunny/crossref_metadata_2025 huggingface dataset:
IL-15 differentially affects the migratory capacity of young and aged memory CD8 T cells to mucosal sites. (46.5)
Authors: Marzo Amanda , Sowell Ryan , Rogozinska Magdalena , Lerret Nadine
Published: January 2023

Given that CD4 T cells are targets of depletion by HIV and required for the homing capacity of CD8 T cells, enhancing recruitment of both CD4 and CD8’s could potentially enhance protection against HIV infection. CD8 T cells at the site of infection are critical for clearance of HIV-infected cells. Memory CD8 T cells migrate into the periphery in the absence of “CD4 help”, homeostatically. However, migration to mucosal tissues such as the gut, require up-regulation of chemokine receptors, intergrins and other signals to gain entry. γC cytokines are key components of CD4 and CD8 T cell survival and maintenance. To date, the specific role of γC cytokines on migration of antigen-specific memory CD8 T cells to mucosal tissues is poorly understood. We show that IL-15 complex treatment drives migration of memory CD8 T cells to mucosa tissues such as the lung and lamina propria of the small intestine (SI LP). However, if the memory cells are aged then migration of CD8 T cells to the mucosal tissues is limited to the lung. These data reveal a novel role of for IL-15 for use as a potential vaccine adjuvant to enhance migration of young memory CD8 T cells to mucosal sites of infection and enhance protection against HIV infection.
Cytokines establish T cell memory by regulating mitochondrial metabolism (46.7)
Authors: van der Windt Gerritje , Everts Bart , Curtis Jonathan , Freitas Tori , Amiel Eyal , Pearce Erika
Published: January 2023

CD8 T cells have a crucial role in host defense during infection. Upon activation, antigen-specific effector T cell populations will first expand and then contract, however, a portion of this population will persist as long-lived memory T cells that will mediate immunity to re-infection. Although this predictable pattern of CD8 T cell responses is well established, the underlying cellular mechanisms regulating the transition to memory T cells remain undefined. We speculate that during a primary response to infection, T cells experience metabolic stress, perhaps due to nutrient and/or growth factor limitation as the pathogen is cleared and antigen concentrations decline. Our previous work suggests that during this period, T cells need to switch from glycolysis to forms of catabolic metabolism, like fatty acid oxidation, in order to survive and develop into memory T cells. Using the Seahorse Extracellular Flux Analyzer, which measures cellular oxygen consumption and proton production, we are able to determine T cell metabolism in real-time. Currently we are investigating the alterations in T cell bioenergetics and mitochondrial function that are critical for the generation of memory T cells. Our recent data show that specific common γ chain cytokines regulate this process. These results offer new insight into the molecular mechanisms of how cytokines drive T cell fate during an immune response.
ST6Gal I promotes rapid IL-2Rα expression and early proliferation of terminal effector CD8 T cells (46.8)
Authors: Onami Thandi , Zeng Junwei , Patel Jenish , Rajini Bheemreddy , Kaech Susan , Smith Kendall
Published: January 2023

ST6Gal I is a glycosyltransferase highly expressed by lymphocytes and cancer cells that catalyzes the addition of α2,6 sialic acid to galactose (Siaα2-6Gal) on N-linked glycoproteins. We discovered that ST6Gal I shows dramatic changes in gene expression during memory CD8 T cell differentiation that corresponds with lectin binding, but the significance of this regulated carbohydrate modification remained a mystery. To identify the significance of these regulated glyco-phenotypic changes, we infected ST6Gal I-/- mice and found that viral-specific cell expansion was impaired. Moreover, defective viral-specific CD8 T cell expansion was due to early intrinsic defects in proliferation of terminal effector cells, whereas memory precursor development was enhanced. We provide direct evidence that loss of ST6Gal I expression by CD8 T cells resulted in delayed surface expression of IL-2Rα in vivo due to impaired IL-2/IL-2R signaling. These results suggest that CD8 T cells regulate ST6Gal I expression to promote proliferation induced by cytokine-dependent signaling.
A loss-of-function variant of the antiviral molecule MAVS is associated with a subset of systemic lupus patients (47.10)
Authors: Si-Tahar Mustapha , Niewold Timothy , Vitour Damien , Solhonne Brigitte , Crow Mary , Pothlichet Julien
Published: January 2023

Dysregulation of the antiviral immune response may contribute to autoimmune diseases. Here, we hypothesized that altered expression or function of MAVS, a key molecule downstream of the viral sensors RIG-I and MDA-5, may impair antiviral cell signaling and thereby influence the risk for systemic lupus erythematosus (SLE), the prototype autoimmune disease. We used molecular techniques to screen non-synonymous single-nucleotide polymorphisms (SNPs) in the MAVS gene for functional significance in human cell lines, and identified one critical loss-of-function variant (C79F, rs11905552). This SNP substantially reduced expression of type I interferon and other proinflammatory mediators, and was found almost exclusively in the African-American population. Importantly, in African-American SLE patients, the C79F allele was associated with low type I interferon production and absence of anti-RNA-binding protein antibodies. These serologic associations were not related to a distinct, functionally neutral, MAVS SNP Q198K. Hence, this is the first demonstration that an uncommon genetic variant in the MAVS gene has a functional impact upon the anti-viral interferon pathway in vivo in humans, and is associated with a novel sub-phenotype in SLE.
Protein phosphatase 2A facilitates IL-17 production by promoting accessibility to the IL-17 locus (47.15)
Authors: Apostolidis Sokratis , Crispin Jose , Rauen Thomas , Thai To-Ha , Tsokos George
Published: January 2023

Systemic lupus erythematosus (SLE) is a complex autoimmune disease caused by the combination of several immune abnormalities. T cells from SLE patients exhibit an aberrant phenotype, caused in part by increased levels of the protein phosphatase 2A (PP2A). To study the mechanisms through which PP2A alters T cell behavior and to identify the contribution of this defect to the disease, we generated a transgenic mouse that over-expresses PP2A in T cells. The transgenic mice produced abnormally high levels of IL-17A and F in vivo and in vitro. Remarkably, naïve CD4+ TPP2A cells produced IL-17 as early as 3h after stimulation. This effect was specific for IL-17 as production of type I and type II cytokines was not affected. PP2A over-expression did not promote Th17 differentiation or significant increases in Th17-related transcription factors. Due to the specificity of the phenomenon and its kinetics, we hypothesized that PP2A may facilitate transcription in the IL-17 locus. ChIP experiments revealed increased tri-methylation of H3K4 and decreased tri-methylation of H3K9 at the Il17a and Il17f promoters as well as in local conserved non-coding sequences. These changes are associated with increased chromatin accessibility. We propose that PP2A plays a central role in the transcriptional regulation of IL-17 by promoting chromatin accessibility to the IL-17 locus. This pathway may play a role in the excessive IL-17 production observed in SLE T cells.
HLA and T cell receptor associations in narcolepsy, a disorder associated with the selective loss of hypocretin neurons (47.17)
Authors: Mignot Emmanuel , Lin Ling , Faraco Juliette , Kornum Birgitte , Steinman Lawrence , Hallmayer Joachim
Published: January 2023

Narcolepsy is a life long disorder characterized by severe sleepiness and cataplexy. It affects 1 in 2,000 individuals and is caused by the selective loss of the hypocretin producing neurons in hypothalamus. This cell loss is likely to be autoimmune and recent findings are closing in on some of the key genetic and environmental effects involved. At the genetic level, the disorder is very tightly associated with DQA1*01:02/DQB1*06:02. In addition to the established role of HLA-DQ, a specific T Cell Receptor Alpha J segment polymorphism has been identified as another susceptibility factor. We are currently sequencing the T-cell TCR@ repertoire to explore this further in recent onset narcolepsy cases. At the level of environmental triggers, winter related upper airway infections are believed to be involved, e.g. we have reported an increased presence of antistreptolysin-O antibody, implicating a role for streptococcal infections. In China, studies of disease onset seasonality indicate a 16 fold increase in onset occurrence around spring when compared to early winter. We anticipate that the cause of narcolepsy will likely involve a specific antigen presentation by DQA1*01:02/DQB1*06:02 to a restricted number of pathogenic T-cell subtypes. As immune involvement is increasingly being recognized in neurodegenerative and neuropsychiatric disorders, understanding why the hypocretin neurons in narcolepsy die is likely to increase understanding of these other disorders as well.
CTLA4 1661 A/G polymorphisms in North Indian Celiac disease patients. (47.21)
Authors: Palhora Shilpa , Saikia Biman , Arora Sunil , Thapa B. , Lal Sadhna
Published: January 2023

Celiac disease or Gluten sensitive enteropathy is an immunomediated chronic small bowel disease caused by intolerance to prolamins,the proteic and alcohol soluble fractions of gluten,a major component of some of commonest cereal like wheat,rye and barley. Celiac Disease is an autoimmune disorder, characterised by villous atrophy of the small intestine, which results from a T-cell-mediated response to gluten-derived peptides. The cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) is involved in the regulation of T-cell activation and the CTLA4 1661 A/G polymorphism has been implicated in several autoimmune disorders, including celiac disease. The aim of this study is to determine the Genetic polymorphisms of the CTLA4 gene and genotyping of CTLA4-1661 A/G in celiac disease in the North Indian population. The 1661 A/G polymorphisms was genotyped in a case of 50 patients and 25 controls.The frequency of the 1661 G-allele was increased in celiac disease patients (p=0.013).The genotype distributions shows a significant difference between cases and controls.These results indicate the involvement of the CTLA4 gene in celiac disease development.The haplotype carrying the CTLA 4 1661 G allele was shown to be associated with disease. A larger patients and controls needs to be studied for further conclusive evidence. Statistical analysis was carried out using Microsoft Excel 2002 and SPSS 16 for Windows
Scleroderma signs in Tight Skin 2 mice develop progressively over time. (47.25)
Authors: Long Kristen , Artlett Carol , Blankenhorn Elizabeth
Published: January 2023

Systemic sclerosis (SSc) is an often fatal inflammatory autoimmune disease affecting connective tissue, characterized by excessive collagen deposition in skin and other organs. The Tight Skin (Tsk) 2 mouse model of disease has many features of human disease including tight skin, excessive collagen deposition, alterations in the extracellular matrix, and occurrence of antinuclear antibodies (ANA) with age. Since current treatments only ameliorate disease symptoms, the Tsk2/+ mouse is crucial for the understanding of disease pathology and progression, however, the Tsk2 gene has yet to be elucidated. Our lab has recently narrowed the interval containing the gene to 2.9 megabases. Understanding disease progression in this model is essential, and histological and total protein studies have shown that increased collagen accumulation does not occur until at least 10 weeks of age in Tsk2/+ mice, despite the “tight” phenotype occurring at 2 weeks. mRNA expression levels from the skin of young mice showed that Tsk2/+ mice have an up regulation of Col1a1, Col3a1, and Col5a2, as well as a robust expression of TGF-β signatures and extracellular matrix fibers. Histological studies revealed a significant increase of elastic bundles in the dermis of 2 week old Tsk2/+ mice. Aged Tsk2/+ mice produced ANA specific to severe disease after initial fibrosis, suggesting additional progression of the disease state. Our studies reveal a novel timeline of disease progression in the Tsk2/+ mouse.
Somatic KRAS mutations associated with a human non-malignant syndrome of autoimmunity and abnormal leukocyte homeostasis (47.4)
Authors: Lu Lianghao , Niemela Julie , Fleisher Thomas , Davis Joie , Caminha Iusta , Natter Mark , Beer Laurel , Dowdell Kennichi , Pittaluga Stefania , Raffeld Mark , Rao Koneti , Oliveira João Bosco
Published: January 2023

Autoimmune lymphoproliferative syndrome (ALPS) is a non-malignant disease characterized by early-onset chronic lymphadenopathy, splenomegaly, multilineage cytopenias, polyclonal hypergammaglobulinemia, expansion of circulating TCRαβ+B220+CD4-CD8- T (αβ+-DNT) lymphocytes, and an increased risk of B-cell lymphoma. Most ALPS patients have germline or somatic TNFRSF6 (FAS) mutations, and a small minority of patients have germline mutations in FAS ligand and caspase 10. We recently reported that an ALPS-like phenotype can be caused by a somatic NRAS mutation (G13D) resulting in the defective lymphocyte apoptosis. Here we demonstrate that somatic mutations in the related KRAS gene can also be associated with a non-malignant syndrome of autoimmunity and breakdown of leukocyte homeostasis. The activating KRAS mutation (G13C) impaired cytokine-withdrawal induced T cell apoptosis through the suppression of the pro-apoptotic protein BIM and facilitated proliferation through p27kip1 downregulation. These defects could be corrected by MEK1 or PI3K inhibition in vitro. We suggest the use of the term RAS-associated autoimmune leukoproliferative disease (RALD) to differentiate this disorder from ALPS. In the hematopoietic system, somatic KRAS and NRAS mutations are commonly observed in aggressive tumors such as multiple myeloma or juvenile myelomonocytic leukemia (JMML). From our reports, we think patients with somatic KRAS and NRAS mutations can follow a more benign clinical course.
Immunological synapses do not restrict secretion of interferon γ during antigen-specific lysis of target cells by CD-8+ cytotoxic T lymphocytes (48.12)
Authors: Sanderson Nicholas , Puntel Mariana , Kroeger Kurt , Bondale Niyati , Iranmanesh Niloufar , Swerdlow Mark , Ibrahim Ahmed , Castro Maria , Lowenstein Pedro
Published: January 2023

Cytotoxic T lymphocytes display two main effector mechanisms when interacting with target cells in an antigen-specific manner: (i) destruction of the target cell, and (ii) secretion of cytokines. It is thought that while target cell lysis occurs following brief contact with the T cell, cytokine secretion requires formation of immunological synapses between T cells and target cells. At immunological synapses between CTLs and adenovirus-infected astrocytes in the brain, interferon γ is polarized at the contact, suggesting immunological synapses constrain secretion of interferon γ. To test this hypothesis we developed a live cell imaging system to examine whether individual OT-I T cells interacting with individual ovalbumin-expressing astrocytes (cultured on a monolayer of ovalbumin non-expressing astrocytes) secrete interferon γ in a vectorial or diffuse manner. In this paradigm, OT-I T cells form contacts with targets lasting for several hours, during which interferon γ secretion is always observed, and approximately 50% of targets are lysed. Interferon γ secretion appears to be diffuse, acting equally on targets and non-antigenic neighbors. Our data indicate that secretion of interferon γ is not restricted spatially by immunological synapses.
Tyrosinase reactive T cells can eliminate large established human melanoma tumors in vivo (48.23)
Authors: Martins da Palma Telma , Spivey Natali , Naga Osama , Al-Khami Amir , Graves Michael , Lesher Aaron , Ferrone Soldano , Mehrotra Shikhar , Norell Håkan , Nishimura Michael
Published: January 2023

Objective clinical responses of melanoma can be achieved by infusion of tumor infiltrating lymphocytes or T cell receptor (TCR) gene transduced T cells. To evaluate the therapeutic efficacy of T cells expressing a high affinity TCR specific for the tyrosinase melanoma antigen (epitope Tyr368-376), we have developed a xenogenic model of human melanoma. Established human melanoma tumors (624 MEL) were rejected by administering low numbers of TIL 1383I TCR transgenic (h3T) splenocytes in an antigen specific, HLA-A2 restricted manner. Importantly, this therapeutic effect was observed with both small (30mm3) and large (100mm3) tumors. However, many of the mice that rejected their tumors recurred in time and the recurrent tumor cells were no longer recognized by h3T cells in vitro or in vivo. Phenotypic analysis revealed that all recurrent tumors from mice bearing 624 MEL had specifically lost HLA-A2, but not other MHC class I alleles, and this was not restored by IFN-γ treatment. Interestingly, mice bearing a HLA-A2 positive clone of 624 MEL (624-38 MEL) did not exhibit any tumor recurrences when treated with h3T cells. These results highlight the limitations of using a monoclonal therapeutic approach for melanoma. They also stress the need of models that enable us to evaluate combinatory therapies targeting heterogeneous tumors to develop improved immunotherapeutic strategies.
Enhancing NK cell-mediated tumor rejection by activation of the aryl hydrocarbon receptor (48.31)
Authors: Sunwoo John , Shin June , Zhang Luhua , Murillo Oihana , Kim Junmo
Published: January 2023

Natural killer (NK) cells’ tumoricidal effector activity is regulated by a balance of inhibitory and activating receptors. We demonstrate that murine NK cells express a cytoplasmic receptor called the aryl hydrocarbon receptor (AhR). AhR is upregulated upon NK cell activation and binds a wide range of endogenous and exogenous ligands. Upon binding agonistic ligands, this receptor translocates to the nucleus and regulates transcription. We show that activation of AhR in NK cells enhances IFNγ production by stimulated NK cells. Furthermore, activation of AhR in vivo results in enhanced rejection of RMA-S tumors in an NK cell - dependent and AhR - dependent manner. This rejection is correlated with an increase in tumor-infiltrating NK cells. Importantly, this rejection of RMA-S tumors is not the result of the AhR agonist’s effects on the tumor cells directly because RMA-S cells do not express AhR, and their proliferation rate in vivo is not affected by exposure to the ligands. This function of AhR as a potential enhancer of NK cell activity is likely to be physiologic because we also find that AhR-deficient mice are not able to control RMA-S tumors. We show that this defect is inherent to the NK cells themselves using an adoptive transfer approach. Thus, we conclude that AhR plays an important role in the modulation of NK cell effector functions and that activation of this receptor may be a novel therapeutic strategy for cancer treatment and prevention.
Systemic inhibition of IKKβ for the treatment of malignant melanoma profoundly alters the leukocytic composition within the tumor and lymphoid organs (48.32)
Authors: Hawkins Oriana , Horton Linda , Ayers Gregory , Richmond Ann
Published: January 2023

With over 8500 deaths each year in the USA, malignant melanoma is the deadliest of all skin cancers. Constitutive activation of the nuclear factor kappa B (NF-κB) is common in melanoma, promoting tumor growth, survival and metastasis. In light of its role in tumorigenesis, targeting of the NF-κB pathway is a therapeutic option for the treatment of melanoma. However, NF- κB activation is critical for the development, survival, migration and function of immune cells. We hypothesized that systemic inhibition of NF- κB activation might have a deleterious effect on the tumor immune response. Melanoma bearing C57BL/6 mice received 100 mg/kg of BMS-345541, a selective inhibitor of the inhibitor of κB kinase (IKKβ), twice daily. Tumors, spleens and bone marrow were harvested at different time points and analyzed by flow cytometry for leukocyte composition. There was a profound reduction of B cells and immature myeloid cells in the spleen and bone marrow, as well as an overall reduction in leukocytic infiltration of the tumor. Unexpected was the increase in the proportion of CD8+ T cells at all sites. These data suggest a shift from a pro-inflammatory and pro-tumorigenic leukocytic profile to an anti-tumor profile. The implications of these findings are explored.
Morphine inhibits recruitment of tumor promoting leukocytes (48.33)
Authors: Koodie Lisa , Ramakrishnan Sundaram , Yu Haidong , Charboneau Richard , Liu Xiuiu , Roy Sabita
Published: January 2023

Tumor cells initiate angiogenesis within the tumor micro-environment. The maintenance of these newly formed vessels is highly dependent on the migration and recruitment of immune cells. In previous studies, we have demonstrated that chronic morphine administration suppresses tumor cell induced angiogenesis to inhibit tumor growth in mice. Here we tested the hypothesis that a known immunosuppressant as morphine also regulates immune cell recruitment into- and migration towards a tumor microenvironment. To mimic a tumor microenvironment, matrigel was admixed with conditioned media derived from Lewis lung carcinoma cells (LLCCM) and implanted subcutaneous in WT or Mu-Opioid Receptor knockout mice (MORKO) treated with saline or morphine through pellet implantation. Interestingly, plugs taken from placebo treated mice (day 7) showed significant leukocyte infiltration, greater than that compared to plugs from morphine treated mice (p<0.05) and this effect was abolished in MORKO mice. Similarly, morphine suppressed the recruitment of adoptively transferred luciferase expressing bone marrow derived cells (known to contribute to tumor growth) into PVA sponges-presoaked with LLCCM and implanted subcutaneous in mice- pretreated with placebo or morphine. Taken together, these studies suggest that morphine treatment can potentially decrease solid tumor associated inflammation to reduce angiogenesis associated with increased tumor growth.
Adaptive immune response initiated by high mobility group box 1 promotes tumor progression (48.36)
Authors: Yu Ping , He Ying , Zha Jikun , Wang Yamin , Fu Yangxin , Liu Wenhua
Published: January 2023

Cancers are often observed to be associated with infiltrating lymphocytes. However, the role of lymphocytes in regulating pro-tumor versus anti-tumor immune responses remains controversial. We report that lymphocytes are essential for tumor progression in an autochthonous prostate cancer model. Activated T lymphocytes gradually accumulate inside the tumor during the process of de novo spontaneous tumor development driven by the tissue-specific expression of an oncogene. T cell infiltration is essential for the increased CD11b+Gr-1-F4/80+ macrophages inside the tumor. Intriguingly, blockade of high mobility group box 1 (HMGB1) drastically inhibits tumor-antigen-specific T cell activation, infiltration and tumor progression. Our study has demonstrated that “endogenous danger signals” like HMGB1 produced by a growing tumor are essential to bridge innate and adaptive immune responses, which could further promote tumor progression over time. Our finding may have important implications for future cancer prevention and therapy.
Association of inhibition of tumor growth with intratumoral hematopoiesis induced by IL-33 (48.40)
Authors: Kim Wonyoung , Kim Juyang , Kim Hyun , Cho Hong , Kwon Byungsuk
Published: January 2023

Immune surveillance by a functional immune system is a well-established mechanism for regulation of tumor growth. In this study, we have shown that innate type 2 helper cells (Ih2) could be an important component responsible for tumor immunosurveillance. Local production of IL-33 by lymphoma cells induces recruitment of Ih2 cells into tumor masses. In response to IL-33, Ih2 cells can proliferate, produce high levels of Th2 cytokines, and play an important role in differentiation and expansion of eosinophils in an IL-5-dependent manner. Eosinophils in turn exhibit a potent anti-tumor activity. The ability of IL-33 to induce the emergence of an Ih2 population and eosinophils in tumors identifies a link between the Th2 cytokine family member and extramedullary hematopoiesis, and suggest a previously unrecognized innate immune pathway involving lineage-negative innate cells which mediated type 2 innate immunity leading to antitumor activity
Crossreactive influenza A virus M1- and EBV-BMLF-1-specific CD8 T cells modulate disease severity of Epstein-Barr Virus-induced infectious mononucleosis (49.20)
Authors: Aslan Nuray , Watkin Levi , Luzuriaga Katherine , Selin Liisa
Published: January 2023

During EBV-associated infectious mononucleosis (IM) influenza A-specific crossreactive memory T cells are highly activated and play a role in disease severity. In HLA-A2+ IM patients, influenza M158 (IAV-M1)-specific CD8 memory T cell responses cross-reacted with two different EBV lytic epitopes, BRLF-1(190) (19/20) and BMLF-1(280) (17/29). Furthermore, 11/22 IM patients demonstrated some intra-viral cross-reactivity between EBV-BRLF-1 and -BMLF-1 responses. Disease severity of IM did not correlate with viral load, but instead directly correlated with the percentage of IAV-M1, EBV-BMLF-1 and crossreactive IAV-M1/EBV-BMLF-1-specific CD8 T cells. This was associated with altered TCR Vbeta usage. In all IM patients cross-reactive T cell responses were observed but different patterns of cross-reactivity were seen in each individual. A number of unique features of the TCR of the highly cross-reactive BRLF-1 epitope were noted. BRLF-1 T cell responses were of very high avidity. There was tremendous variation in the Vbeta usage between different individuals and even in the same individual over time. Cross-reactive BRLF-1 responses only became apparent at higher antigen doses in vitro, such as might occur during different phases of infectious mononucleosis.
Pathogenic H1N1 2009 influenza virus-induced pulmonary injury is suppressed by sphingosine analog-mediated inhibition of cytokine storm (49.3)
Authors: Walsh Kevin , Tiejaro John , Wilker Peter , Jatzek Anna , Suresh Marulasiddappa , Kawaoka Yoshihiro , Rosen Hugh , Oldstone Michael
Published: January 2023

Human pandemic H1N1 (2009) influenza virus rapidly infected millions worldwide and caused significant morbidity and mortality that was associated with cytokine storm and virus-induced cytopathology. Antiviral drugs are the primary therapy to treat disease, although there are limitations to their effective use: 1) administration must be early for optimal efficacy; 2) selective pressure is exerted on the virus resulting in the generation of viral progeny that are resistant to treatment; 3) treatment does not directly inhibit immune-mediated tissue damage, a significant component of disease. Here we show in a mouse model of infection with pathogenic pandemic H1N1 (2009) influenza virus that an immunomodulatory drug, AAL-R, provides significantly greater protection than that occurring with the antiviral drug, oseltamivir. Combined administration of both AAL-R and oseltamivir enhances survival over that from either drug alone protecting 94% of infected mice from a lethal inoculation of virus. Mechanistically, AAL-R significantly downregulates host cellular as well as cytokine responses thereby limiting pulmonary damage ultimately enhancing survival without interfering with host control of pandemic H1N1 (2009) influenza virus replication. Suppressing cytokine storm using sphingosine analogs may prove to be a useful therapy against influenza virus as well as other respiratory diseases where immune-mediated tissue injury is a significant component of disease.
Defective CD8+ T cell effector function in pathogenic influenza infection (49.7)
Authors: Rutigliano John , Morris Melissa , Doherty Peter , Thomas Paul
Published: January 2023

Many questions about CD8+ T cell responses during influenza infection remain unanswered. We therefore examined the development and differentiation of CD8+ T cell memory populations based on specificity and effector function in a mouse model system comparing high and low pathological infections of influenza. Our findings reveal that initial or peak viral titer was not the determining factor in the outcome of disease. Instead, differences in CD8+ T cell effector function were observed for each virus type (high and low pathogenic), and these differences correlated with disease outcome. Further investigation has demonstrated that differences in the airway microenvironment after infection with high and low path infections may significantly affect CD8+ T cell activity. Specifically, increased early production of certain cytokines is associated with highly pathogenic infection. We hypothesize that this altered airway microenvironment results in downmodulation of CTL function via established negative feedback pathways.These phenotypic and functional analyses have revealed distinct effector memory CD8+ T cell populations in the context of influenza infection that have profoundly different capabilities for viral clearance.
Memory T cells generated in youth function well in old age (49.8)
Authors: Decman Vilma , Laidlaw Brian , Ertl Hildegund , Wherry E. John
Published: January 2023

Aging-associated changes negatively influence T cell responses. Since we previously observed that the memory CD8 T cells generated in aged mice have substantially reduced proliferative expansion upon secondary infection, we wanted to investigate if observed defects of memory T cells are due to the age of the cell or the age of the host when primed. Briefly, virus-specific memory T cells were generated by infecting young mice, aged mice or by infecting mice when they were young, but then allowing these mice to age for 2 years. T cells from these three groups were adoptively transferred to congenic young mice and these recipient mice were challenged to assess the recall capacity of memory T cells. We used both systemic (vaccinia virus or LCMV) and local (influenza virus) challenge models to investigate whether there were differences in the ability to respond to different types of infections and to assess the recall capacity of virus-specific T cells. Strikingly, the memory CD8 and CD4 T cells that were primed in young mice and then aged for 2 years responded as well as the memory T cells primed recently in young mice. These highly functional memory T cells appear largely impervious to age-associated changes occurring in their environment. As expected, memory CD8 and CD4 T cells recently generated in aged mice fail to mount strong recall responses. To conclude, our studies point to events during priming and the starting of T cell population in aged mice as a key area of focus.
TULA-family proteins are key to the regulation of T-cell-driven inflammatory response. (50.1)
Authors: Newman Tiffanny , Carpino Nicholas , Ganea Doina , Safadi Fayez , Tsygankov Alexander
Published: January 2023

The TULA-family consists of two proteins implicated in cellular regulation. TULA-1 is expressed in T-cells and is pro-apoptotic. TULA-2 is a ubiquitously expressed phosphatase that suppresses receptor-mediated signaling. T-cells from mice lacking TULA-1 and 2 (dKO) are hypersensitive to TCR stimulation. This may be due to these proteins having a similar function working synergistically or dissimilar functions having a convergent effect. To understand functional interaction of these proteins we have characterized TULA-family knockout mice without and during an immune challenge. We show that CD4+ T cells of dKO mice have an atypical CD45RB distribution, and that within this subset memory T-cells are expanded only in dKO not in single TULA knockouts. However, CD4+ T-cells of single knockout and wild-type mice respond differently to TCR stimulation as seen using signaling and responses in vitro. To evaluate consequences of TULA deficiency in vivo, we utilize two mouse models of inflammatory bowel disease: TNBS-induced colitis and colitis induced by the adoptive transfer of CD45RBHigh CD4+ T-cells. Studies utilizing TNBS indicate that deficiency of any TULA-family protein exacerbates TNBS-induced colitis. Likewise, dKO CD45RBHigh CD4+ T cells were significantly more colitogenic than cells from WT mice in the transfer model. Taken together, our data indicate that TULA-family proteins are key to the physiological regulation of T-cell reactivity that drives intestinal inflammation.
Disk large protein 1 promotes the initiation of isotype-switched IgG-BCR signaling (50.15)
Authors: Liu Wanli , Pierce Susan
Published: January 2023

B cell memory is critical for the protection from many infectious diseases and is the basis of many current vaccines. Memory B cells expressing high-affinity, IgG-BCRs give rise to the rapid production of high-affinity IgG antibody recall responses, the mechanisms by which the IgG BCRs function to enhance B cell responses are not understood. Studies in mouse models showed that the cytoplasmic tail of membrane-bound IgG (mIgG) is both required and sufficient for the enhanced IgG antibody recall response. Recently, we showed that the conserved membrane proximal region of the mIgG1 cytoplasmic tail, a region not previously appreciated to play a role in BCR signaling, was both necessary and sufficient to dramatically enhance the early events in the initiation of B cell activation. We now identify Dlg1 as a specific, antigen-induced binding partner of the mIgG1 cytoplasmic tail. We demonstrate that Dlg1 and mIg associate through a mechanism involving the binding of the N-terminal and PDZ domains of Dlg1 to the membrane proximal region of the mIgG1 tail. In response to antigens, Dlg1 is rapidly recruited to IgG1-BCR microcluster and then the IgG1-B cell immune synapse. Using live B cell imaging by fluorescence lifetime imaging microscopy we observed that antigen binding induces a molecular complex of Dlg1 and IgG-BCR. Our further evidence suggests that Dlg1 serves as a BCR adaptor stabilizing the formation of IgG-BCR oligomers and facilitating the assembly of signalosome.
Immunology and the liberal arts: constructing a multi-level immunology curriculum at an undergraduate institution (51.2)
Authors: Chatterjea Devavani
Published: January 2023

At a liberal arts college, immunology is not restricted to a specialized biology course. Its relevance to global public health attracts biology majors and non-majors. Its inherent evolutionary scaffolding and broad focus on cellular/molecular physiology makes it suitable for all levels of the majors’ curriculum. With sophisticated technology and model systems, and translational significance, it appeals to senior biology majors undertaking collaborative research with faculty, participating in journal club-style seminar courses and writing independent capstone papers. At Macalester College, we offer a multi-level immunology curriculum ranging from non-majors global and community health seminars on pandemics and vaccine development, an introductory immunology course, with laboratory and civic engagement components, for juniors, and senior-level seminar and research courses and honors research projects. Our flow cytometry core supports labs across the biology curriculum. With its inherent focus on integrative thinking and application of concepts to solving problems, relevance to global health and lastly, but importantly, room for concepts of identity, organization and collective action, immunology is a perfect liberal art. We present examples of course and laboratory innovations, strategies for funding a research laboratory at a liberal arts college, and ways of getting students across campus engaged in studying the immune system.
Toll-like Receptor 2 polymorphisms, signaling function and human disease (51.5)
Authors: Woodham Hadiya , Rees Steven , Medvedev Andrei
Published: January 2023

To introduce high school students to the molecular mechanisms governing health and disease, I recently developed a course entitled, “The Cell and Molecular Basis of Human Health and Disease”, which offers an exploration of how such fundamental processes as gene regulation, cell communication, and cell death form the basis of health and disease. To enhance the immunology unit within this course, I have developed a curriculum based on my summer research that is focused on current understanding of host antimicrobial innate defense and the impact of polymorphisms within Toll-like receptor (TLR) 2. Using transfection-based complementation of HEK293 cells, we found that R753Q inhibits TLR2-mediated activation of NF-kB and expression of IL-8 but does not affect TLR2 expression. Students will learn the basics of TLR innate defense, study TLR expression/functions and how expression of inflammatory cytokines is controlled by transcription
There’s no time to teach that: integrating the immune system, autoimmunity, literacy, and research into an over-crowded curriculum (51.7)
Authors: Pazol Jon
Published: January 2023

Because of various curricular mandates, teachers are faced with covering an ever-expanding range of topics with little room for study of more interesting aspects of biology. Adding to this problem is the need for many teachers to incorporate literacy strategies to remediate student deficits. In addition, assigning independent research often results in plagiarized reports based on unreliable sources. To manage these issues, I have developed a curriculum framework that utilizes autoimmunity, while simultaneously fostering increased literacy and improved research methods. After a general introduction to the immune system, students will explore the specifics of autoimmunity. Experimental Autoimmune Encephalomyelitis (EAE), the mouse model of Multiple Sclerosis currently being researched by Dr. Stephen D. Miller of Northwestern University, will be used as the basis. Students will read, summarize, and formulate questions about current immune-related news articles. Then they will engage in independent research about an autoimmune disease, utilizing relevant and appropriate web sites. Students’ website searches will not only focus on the disease, but will also require them to identify resources available for patients. Steering them in this direction will also provide them with a model for any type of future disease research. The unit is designed for flexibility in its implementation, and can successfully be transferred to a wide variety of topics.
Enabling in vivo visualization of cellular therapies in the clinic: generation of preclinical data sets (52.2)
Authors: Helfer Brooke , Balducci Anthony , Kalinski Pawel , Ahrens Eric , Wesa Amy
Published: January 2023

Cellular therapies are increasingly being used to modify a patient’s immune function in various areas of medicine, including cancer, infectious disease, and regenerative medicine. A question that remains with such therapies is the dispersion and persistence of cells after administration. Here we utilize a novel 19F MRI tracer agent to label Natural Killer cells (NK), T cells, dendritic cells (DC), and stem cells ex vivo and illustrate preclinical data sets which may be used to support a cellular therapeutic IND. Important therapeutic functions, such as differentiation of stem cells, migration and antigen processing of DC, and cytokine production and proliferation of NK and T cells, are maintained upon 19F labeling. Toxicity studies indicate that this 19F tracer lacks mutagenicity and, in acute toxicity studies, induces no adverse systemic or local effects at 100x the expected patient dose. In vivo results demonstrate the capability of MRI to detect and quantify labeled cells with a high degree of specificity. Since there is little exogenous fluorine present in tissue, labeled cells appear with little background. Pairing this signal with conventional proton MRI from the same imaging session, cells are able to be traced to their precise anatomical location. The MRI tracking capabilities, safety profiles, and use of 19F labeling in preclinical data sets demonstrate the ability of 19F to be used in clinical applications to strengthen the understanding of cellular therapeutics.
Longevity of memory CD8 T cell responses after multiple antigen-stimulations in vivo (52.26)
Authors: Badovinac Vladimir , Rai Deepa
Published: January 2023

Recently it has been shown that proliferative capacity and long term maintenance and survival of CD8 T cell responses (T-cell receptor transgenic - TCR-Tg) are controlled by the number of Ag-stimulations. Here, in a similar experimental design and using an adoptive transfer model we show that repetitive Ag-stimulation of endogenous memory CD8 T cells populations leads to the inability of those cells to undergo homeostatic proliferation and survive. In contrast, repetitive homologous boosting (up to 6 times) of memory CD8 T cells in situ (no adoptive transfer involved) did not lead to loss of memory CD8 T cells or their ability to survive, undergo homeostatic proliferation, and acquire a central memory (ie. CD62L high, CD27 high) phenotype and function (ie. IL-2 production). Interestingly, continuous boosting of Ag-specific CD8 T cells is required to preserve the numbers and increase the complexity of memory CD8 T cell responses. Thus, these results suggest that memory CD8 T cell responses in an individual host can survive multiple rounds of Ag-encounters and that repeated Ag-stimulation might be required for the host to sustain the quantity while maintaining the diversity (quality) of the memory CD8 T cell pool.
Activation of Wnt signaling preserves naive T cell phenotype (52.28)
Authors: Muralidharan Sujatha , Chakraborty Rikhia , Hanley Patrick , Bollard Catherine , Rodgers John , Savoldo Barbara , Dotti Gianpietro
Published: January 2023

The study of canonical Wnt-β catenin signaling in mature T cells in mouse models has shown that activation of Wnt signaling led to an arrest in effector T cell differentiation and promoted memory T cell formation. This project extends this investigation into human T cells and examines the effect of this signaling pathway on naïve-effector-memory development in mature human T cells. Recapitulating the results seen in mouse models may help to overcome limitations in clinical treatments such as adoptive T cell therapy for cancer since studies have shown that infusion of naïve or memory T cells improves in vivo persistence of these T cells. Wnt signaling was induced in polyclonally activated human T cells and after a week’s culture, there was a significant increase in naïve CD45RA+CD62L+ T cells (4 vs. 32%) and a corresponding decrease in effector CD45RO+ T cells (86 vs. 63%) as measured by flow cytometry. Cell expansion was significantly reduced as a result of an arrest in cell proliferation (88% vs. 27%). Treating Wnt-activated cells with IL-15, IL-7 or IL-2 restored proliferation but led to a corresponding loss of the naïve phenotype. Wnt signaling also led to impaired T cell effector function, in terms of degranulation (60 vs. 11%) and IFN-γ production (24% vs. 5%). Therefore, activation of Wnt signaling in mature human T cells preserves a subset of naïve T cells by arresting proliferation and impairs T cell effector function.
Qualitative and quantitative parameters predict functional humoral response to alum-adjuvanted immunization. (52.4)
Authors: Lynch Heather , Stewart Shelley , Riebe Kristina , Kepler Thomas , Alam S. , Sempowski Gregory
Published: January 2023

Adjuvants are crucial vaccine components for enhancing immune response to immunogens. Many potential adjuvant materials have been studied for their ability to augment the response to antigens while maintaining low toxicity. A US FDA approved adjuvant routinely used in humans is alum. The aim of this study was to comprehensively measure the ways in which innate immune signals induced by alum manifest in terms of the effect on the adaptive immune response to recombinant protective antigen (PA) from Bacillus anthracis. C57Bl/6 mice were given a single subcutaneous immunization with PA +/- alum at the base of each limb. Serum and draining lymph nodes were collected after immunization (days 1-70). On day 71, saline or PA only boost was administered, followed by additional serum and tissue collection (days 78-85). Using multi-color immunohistochemistry it was shown beginning day 10 that alum induces a rapid and strong germinal center response in the draining lymph node. This observation correlates with quantitative and qualitative changes in PA-specific antibody assessed by ELISA and Biacore (SPR). In addition, augmented antibody response to PA+alum resulted in increased antibody functionality, as determined by anthrax toxin neutralization assays. This study demonstrates a novel multi-parameter approach to evaluation of antigen-specific responses to immunization and the impact of adjuvants. This approach will be useful in the development and testing of novel adjuvants.
Induction of long-lasting T cell memory against M. tuberculosis on vaccination with promiscuous peptide of 16 kDa antigen linked to Pam2Cys (53.11)
Authors: Agrewala Javed , Gowthaman Uthaman , Singh Vijender , Zeng Weiguang , Gupta Umesh , Jackson David
Published: January 2023

Tuberculosis (TB) is one of the major killer diseases, especially in developing countries. In TB-endemic areas, one of the main reasons considered for the failure of BCG (Bacillus Calmette-Guérin) is the impediment caused by environmental mycobacteria in its processing and presentation. In general, peptides do not require antigen processing since they can directly bind to major histocompatibilty complex (MHC) molecules and be presented to T cells. Consequently, vaccines constructed employing peptides can overcome the problem responsible for BCG failure. In the present study, we have developed a unique lipopeptide by linking the promiscuous peptide of 16 kDa antigen of Mycobacterium tuberculosis to Pam2Cys, a toll like receptor-2 (TLR2) agonist. This peptide elicits immune response irrespective of HLA polymorphism. Further, it does not require extensive antigen processing, can bind directly to MHC molecules and activate T cells. Most interestingly it has self-adjuvanting properties and activates both innate and adaptive arms of immunity. Further, it activates dendritic cells (DCs) and imparts enduring Th1 memory response. Furthermore, it surmounts the barrier of MHC polymorphism and induces better protection than BCG. This lipopeptide can be a potent future vaccine candidate against TB.
Selection of conserved HIV-1 vaccine epitopes based on cross-reactivity to feline immunodeficiency virus (53.17)
Authors: Sanou Missa , Coleman James , Levy Jay , Yamamoto Janet
Published: January 2023

Recent reports suggest that CTLs against the conserved epitopes on HIV-1 are required for an effective HIV-1 vaccine, and that the vaccine should induce CTLs to conserved epitopes mimicking those generated by HIV-1+ long-term survivors (LTS). The CTL epitopes needed for vaccine are still unclear. Our current study identifies the evolutionarily-conserved CTL epitopes preserved between HIV-1 and FIV reverse transcriptases (RT) most frequently recognized by HIV+ subjects, including LTS. The IFNγ-ELISpot, CFSE-proliferation, and intracellular staining (ICS) analyses of PBMC upon stimulation with HIV or FIV RT peptide pools or peptides (overlapping 15-mer peptides of whole RT) were used to map the anti-FIV and anti-HIV responses made by HIV+ group (n=14 includes 7 LTS) and HIV- controls (n=5). The HIV+ subjects had HLA alleles from 8 supertypes (mainly A2 supertype) and some with B57 or B27 allele for AIDS nonprogression. Seven of 14 HIV+ subjects (includes 4 LTS) reacted to 4 FIV RT pools and to their HIV counterparts, and none by the controls. Peptides from these pools induced the proliferation of CD3+CD8+ or CD3+CD4+ T cells, which were ICS positive for IFNγ, perforin, granzyme-A, or granzyme-B in various combinations. The existence of evolutionarily-conserved CTL epitopes was determined using HIV/FIV epitope mapping. This approach can select for highly conserved epitopes that are less likely to mutate and as a result can be useful in the development of an HIV-1 vaccine.