bluuebunny/crossref_metadata_2025_split huggingface dataset:
Efficacy of selective RAR-related orphan receptor gamma (RORγ) inverse agonists in psoriasis and asthma models (CCR6P.278)
Authors: Nellore Kavitha , Bejugam Mallesham , Damarla Ravi Krishna Babu , Hegde Pragathi , Dodheri Samiulla , Mukherjee Subhendu , Rao Narasimha , Lakshminarasimhan Anirudha , Ramanathan Anuradha , Balasubramanian Wesley , Mattegunta Suresh Babu , Mahalingam Natarajan , Samajdar Susanta , Subramanya Hosahalli 
Published: January 2023

Abstract
Th17 cells play a key pro-inflammatory role in a variety of autoimmune diseases. The nuclear hormone receptor RORγ controls the differentiation of Th17 cells and expression of IL-17. RORγ inverse agonists were designed using a combination of approaches including structure and knowledge based methods. Compounds were screened in a RORγ radio-ligand binding assay using 3H 25-Hydroxycholesterol, as well as in a cell based reporter assay to demonstrate inverse agonism. Compounds from multiple structural classes have been identified, with IC50 in the range of 5 - 500 nM in binding assay. Co-crystal structures of RORγ in complex with known as well as in-house compounds were utilized to optimize the lead compounds. Compounds from the lead series demonstrated good activity (&lt; 500 nM) in reporter assay. Several compounds demonstrated &gt; 10 fold selectivity against RORα as well as other nuclear receptors. Compounds from multiple series have shown significant inhibition of IL-17 release from differentiated mouse and human Th17 cells. Lead compounds have shown good pharmacokinetic properties in mice. Efficacy of lead compounds have been demonstrated in IMQ induced Psoriasis model as well as in intranasal OVA/LPS induced asthma model. In summary, we have identified structurally diverse small molecule inverse agonists of RORγ and have demonstrated efficacy in Th17 mediated efficacy models.
Acute alcohol intoxication suppresses early Il-17 response during pneumococcal pneumonia. (CCR6P.273)
Authors: Trevejo-Nunez Giraldina , Chen Kong , Kolls Jay 
Published: January 2023

Abstract
In animal models, acute alcohol intoxication suppresses the host immune responses against S. pneumoniae. Il-17 plays essential roles in host defense against extracellular pathogens including S. pneumoniae. We demonstrated that Il-17R signaling is critical for the clearance of S. pneumoniae by blocking the Il-17 receptor with monoclonal antibody. We thus hypothesize that alcohol impairs pneumococcal lung clearance by affecting Il-17 production and Il-17R related genes in the lung. We used a model of acute alcohol intoxication in mice followed by S. pneumoniae pneumonia. Alcohol and control group had similar bacterial burden in the lungs at 5 hours, however, the former group failed to clear pneumococci at 18h. Il-17a production in the lung was decreased in the alcohol group at 5 hours, suggesting that the impaired clearance is, at least in part, due to the suppression of Il-17 production. Gene expression analyses showed that both Il17a and Il17f transcripts were down-regulated in alcohol mice, suggesting the suppression is at transcriptional level. This notion is supported by decreased expression of Il-17 upstream genes such as Il1b and Il23. Consequently, downstream genes such as Csf3 and Cxcl1, which are critical in mobilizing and recruiting bacterial phagocytes, were down regulated at 5 hours compared to control mice. Taken together, this study provides a novel mechanism by which alcohol impairs host defense against bacterial infection.
Circulating T helper cells with T follicular helper cell phenotype renders the autoantibody generation in patients with systemic lupus erythematosus (HUM7P.304)
Authors: Zhang Xin , Lindwall Elvira , Gauthier Carl , Chen Minzi , Khan Omar , Choi Yong Sung , Zakem Jerald , Quinet Robert 
Published: January 2023

Abstract
SLE is a chronic autoimmune disease characterized by autoreactive B cells and autoantibody production. Follicular helper T (Tfh) cells, a T cell subset located in the germinal center (GC) of lymphoid organs, is a key participant regulating B cell tolerance and antibody production. To identify the role of circulating Tfh (cTfh) cells in SLE, we examined their phenotype and function on supporting antibody production, and their correlation with autoantibody level and disease activity. Peripheral blood was collected from 29 SLE patients and age/gender-matched healthy donors. Tonsils from non-SLE controls were used to extract GC-Tfh cells. Tfh cells were defined by their signature surface markers (CXCR5, ICOS, CD57, and PD-1) via flow cytometry. IL-21 expression was measured by real-time PCR and intracellular staining. Tfh cells, non-Tfh cells were purified by MACS columns and co-cultured with B cells. IgG in the culture supernatant was detected by ELISA. Our results showed that cTfh cells in SLE patients expressed GC-Tfh signature molecules and IL-21 and were capable of driving B cells differentiate into plasma cells in vitro. The elevated frequency of cTfh cells correlated positively with levels of circulating plasmablasts, serum anti-dsDNA Ab and ANA. Our data suggests that the increased cTfh cells in SLE patients share phenotypic and functional properties with GC-Tfh cells. Tfh cells may serve as perpetrator to the pathogenesis in SLE patients and target for novel treatment.
Ubiquitination modulates expression of the serine arginine-rich splicing factor 1 (SRSF1) in activated T cells and in SLE patients (HUM7P.305)
Authors: Moulton Vaishali , Gillooly Andrew , Tsokos George 
Published: January 2023

Abstract
T cells from patients with systemic lupus erythematosus (SLE) are poor producers of the vital cytokine interleukin (IL)-2. We recently showed that SLE T cells express lower levels of the serine arginine- rich splicing factor 1 (SRSF1), more so in patients with active disease. Forced expression of SRSF1 restores IL-2 production, indicating that SRSF1 is important in T cell function, and its reduced expression is a potential molecular defect in SLE T cells. The goal of this study was to determine the mechanism/s regulating SRSF1 expression in human T cells and in SLE patients. T cell activation increased mRNA expression of SRSF1 by 3- to 4-fold within 24 hours, while protein expression did not alter much. This discrepancy between mRNA and protein expression was not due to reduced RNA stability. Activating T cells in presence of specific proteolysis inhibitors revealed that proteasome- but not lysosome-mediated mechanisms contribute to reduced SRSF1 expression. Accordingly, immunoprecipitation (IP) studies showed increased ubiquitination of SRSF1 in activated T cells compared with resting cells. More interestingly, SRSF1 showed increased ubiquitination in SLE patients compared to healthy controls and rheumatoid arthritis patients. Our results identify a novel mechanism of SRSF1 protein regulation in human T cells and suggest that increased ubiquitination and proteasome degradation represent a molecular mechanism that contributes to reduced SRSF1 expression in SLE T cells.
Acute lymphoblastic leukemia induces caspase-1 activation in human bone marrow monocyte/macrophages and is associated with impaired CD8 function. (HUM7P.315)
Authors: Shand Jessica , Mariano John , Toia Liana 
Published: January 2023

Abstract
B-acute lymphoblastic leukemia (ALL) is a malignancy of bone marrow-derived precursor-B cells and represents the leading cause of pediatric cancer death. Suppressive signals induced in the marrow of ALL patients may explain both the failure of host CD8 surveillance and inconsistent responses to adoptive CD8 therapy. We hypothesize that ALL induces a caspase-1 driven inflammasome in bone marrow monocyte/macrophages (BM-MM) that modulates CD8 function and contributes to relapse. To determine whether BM-MM could produce soluble factors that suppress CD8 function in the setting of ALL, we treated CFSE-labeled CD8 from healthy donor PBMC with supernatants from cocultures of primary human ALL and either CD14+ BM-MM or stromal fibroblast controls. CD8 exposed to supernatants from ALL cultured alone or with stromal fibroblasts maintained 100% proliferation, while those exposed to BM-MM cocultures failed to proliferate, regardless of whether ALL was cultured directly or in transwell with BM-MM. Cleaved caspase-1 was detected, both by qPCR (2.2-fold expression over endogenous control) and Western blot from BM-MM cultured with ALL, but not stromal fibroblasts Similar levels of soluble IL1B were detected from stromal fibroblast and BM-MM cocultures. Current studies are determining whether the reduced proliferative capacity observed in CD8 corresponds to impaired antileukemia killing and identifying specific signals downstream of caspase-1 that may be targeted to reverse this phenomenon.
Stable individual immune response gene expression signatures in peripheral human CD4 cells foreshadow response to stimulation (HUM7P.324)
Authors: Pease Larry , Zhang Yugi , Felts Sara , Scheid Adam , Van Keulen Virginia , Pankratz V. Shane 
Published: January 2023

Abstract
The complexity of molecular networks regulating the immune response presents challenges to measuring immune response potential in human populations. We measured gene expression in CD4+ peripheral blood cells from healthy subjects using RNAseq. RNA expression was assessed twice, one month apart to identify the subset of genes expressed stably within individuals over time yet discriminating individuals in a cohort. This immune response signature gene set then was used to identify and successfully order the immune response phenotypes of independent, unrelated subjects. Following a brief 4 hr stimulation of freshly isolated cells with anti-CD3/C28 antibody-coated beads, changes in gene expression were measured using a similar approach, defining individual response profiles and clustering response patterns. Examination of the RNA expression signatures of unstimulated CD4 cells revealed a gene subset which when analyzed for gene expression levels correctly aligned independent test subjects into the defined response-phenotype clusters following stimulation. Therefore, there exists a subset of genes stably expressed over time in circulating CD4+ cells which discriminate among individuals in a manner associated with immune response potential. Thus a systems approach can be used to describe the complex immune phenotypes of individuals in ways that might be useful in determining how individuals will respond to complex stimuli or in disease and treatment settings.
Evaluation of PBMC processing in the Precision Bioservices laboratory network (HUM7P.325)
Authors: Bahunde Faith , Neidinger Erik , Awoyode Rotimi , Johnson Netasha , Osunsan Tai , Tambwe Celestine , Waddington Michael , Shea Katheryn 
Published: January 2023

Abstract
Cryopreservation of peripheral blood mononuclear cells (PBMC) is a good alternative to testing fresh PBMC in clinical trials as functional immunologic assays may be batched to increase efficiency. Functional assays are essential for accurately evaluating the performance of vaccines, antiviral and anticancer agents. Optimal retention of PBMC functionality during cryopreservation depends on optimized and standardized cell isolation, freezing and storage conditions. Collection and analysis of PBMC is a significant technical challenge in multi-center clinical trials due to pre-analytical variability in processing, freezing and storage. It is vital that all centers participating in a multi-center collaboration operate with consistent protocols. To this end, Precision has developed a Laboratory Network (PLN) to ensure quality PBMC processing across multi-center trials. The current study was performed within the PLN to ensure samples processed at each laboratory achieved similar results. We report the recovery, viability and functionality of PBMC obtained from participating laboratories. Phenotypic characterization of the major cell subset was also analyzed and the percent obtained was comparable across laboratories. This data provides confidence that PBMC processing in a multi-center clinical trial operating with the same standard operating procedures (SOPs) produces comparable data, thus accelerating drug and vaccine development.
ASC inflammasome mediates host protection against rickettsiae via a NLRP3-independent signaling pathway (INC7P.403)
Authors: Fang Rong , Zhao Xuemei , Smalley Claire , Saito Tais , Brouwer Dedeke , Ismail Nahed , Walker David 
Published: January 2023

Abstract
Rickettsiae escape from the vacuole into the cytosol, where they replicate and inflammasomes recognize the cytosolic infectious pathogens via Nod-like receptors leading to secretion of IL-1β and IL-18. However, the interaction of rickettsiae with inflammasomes has never been investigated. In this study, we found that secretion of mature IL-1β and IL-18 from Rickettsia australis-infected macrophages is caspase-1 and ASC-dependent, suggesting that Rickettsia activates ASC inflammasome. All R. australis-infected ASC-/- mice succumbed to infection (100% mortality) while all infected wild type (WT) mice survived (0% mortality). Rickettsial loads in liver in infected ASC-/- mice were significantly increased with less cellular infiltration compared to infected WT mice. In vivo production of IL-18 was completely dependent on ASC in R. australis-infected mice as evidenced by abolished IL-18 production in infected ASC-/- mice compared to infected WT mice. R. australis-induced-IL-1β and IL-18 concentrations were only reduced, but not abolished in macrophages from NLRP3-/- mice compared to WT controls. Host survival and rickettsial loads in lung and liver of NLRP3-/- mice were comparable with those of WT mice, suggesting a negligible role of NLRP3 in host defense. Taken together, our results suggest that R. australis are recognized by ASC inflammasome in vitro and in vivo via a NLRP3-independent pathway. ASC inflammasome mediates host protection against this intracellular bacterium.
Lack of functional selectin ligand interactions enhances inherent neutrophil function leading to increased resistance to systemic Listeria monocytogenes infection (INC7P.408)
Authors: Agbayani Gerard , Sad Subash , Krishnan Lakshmi 
Published: January 2023

Abstract
Leukocyte recruitment towards sites of infection involves coordinated ligand-receptor interactions between leukocytes and the vascular wall. Selectins transiently bind to selectin ligands to enable leukocyte tethering and rolling on the vascular wall prior to leukocyte adhesion and transmigration into tissues. These interactions occur following post-translational modifications of selectin ligands, including terminal fucosylation by Fucosyltransferase-IV and -VII. In mice, deficiency in both enzymes (FtDKO) leads to loss of leukocyte tethering and rolling, increased circulating neutrophils and impaired T cell trafficking. However, the role of functional selectin ligand interactions in modulating immunity to bacterial infections remains unclear. We analyzed the kinetics of infection and modulation of immune responses in FtDKO mice infected with Listeria monocytogenes, an intracellular bacterium. FtDKO mice controlled systemic infection more rapidly relative to wild-type mice. This was correlated to increased number and functionality of neutrophils. Adoptive transfer of bone marrow cells from FtDKO mice into irradiated wild-type mice resulted in expansion of neutrophil populations and increased resistance to infection in a neutrophil-dependent manner. Functional selectin ligand deficiency on neutrophils was correlated to increased resistance to cell death. Thus, functional selectin ligand expression is a potential target for enhancing innate immunity against bacterial infection.
Intratracheal transfer of lung eosinophils reduces influenza virus burden in mice (INC8P.438)
Authors: Samarasinghe Amali , Woolard Stacie , LeMessurier Kim , Melo Rossana , McCullers Jonathan 
Published: January 2023

Abstract
Eosinophils, commonly known for anti-parasitic immunity and a controversial role in allergic asthma, may exhibit antiviral immunity against respiratory viruses. We have shown previously that mice subjected to a fungal asthma model were protected from morbidity associated with influenza A virus (IAV). Furthermore, we have also shown that mice studied in the model following resolution of allergic/eosinophilic inflammation and were more susceptible to influenza. We hypothesized that eosinophils promote viral clearance from the lungs. Lung eosinophils harvested from “allergic” mice were transferred directly into H1N1 influenza (A/CA/04/2009) infected mice. Recipients had improved lung compliance and reduced viral burden, although there was no change in weight compared to controls. The inflammatory response was equivalent between groups at day 3. There were fewer cells in airways of recipients by day 7, however, there were significantly more virus-specific CD8+ T cells present. The enhanced viral clearance was associated with increased IgA content in the airways of recipients. When mice which had achieved resolution of allergic inflammation prior to IAV infection were supplemented with eosinophils, viral burden also decreased, but the inflammatory profile was unaffected. Bone marrow-derived eosinophils killed IAV by a mechanism that appeared to be piecemeal degranulation. Taken together, these data suggest a novel function for eosinophils in altering the immune response to IAV.
Neonatal oxygen supplementation in mice leads to persistent modifications in NK cell functions during influenza A virus infection (INC8P.436)
Authors: Reilly Emma , Martin Kyle , Jin Guangbi , Yee Min , O'Reilly Michael , Lawrence B. Paige 
Published: January 2023

Abstract
Supplemental oxygen is used to treat preterm infants in respiratory distress. However, this treatment can disrupt normal lung development and alter responses to respiratory insults. Similarly, exposure of newborn mice to 100% oxygen leads to alveolar simplification and altered host resistance to respiratory infection. Upon challenge with influenza A virus, adult mice exposed to neonatal hyperoxia exhibit higher morbidity and mortality, and survivors display hallmarks of pulmonary fibrosis, indicative of atypical repair. Natural Killer (NK) cells are key players in antiviral immunity; identifying and killing infected cells. Emerging evidence suggest they also help maintain homeostasis in peripheral tissues, including the lung, by promoting epithelial cell regeneration via IL-22. We tested the hypothesis that adult mice exposed to hyperoxia as neonates have modified NK cell responses to infection. We found that mice exposed to neonatal hyperoxia had fewer IL-22 producing pulmonary NK cells during influenza virus challenge, but an increase in IFN-γ producing NK cells. In reciprocal bone marrow chimera mice, diminished IL-22+ NK cells aligned with hyperoxia-exposed donors, irrespective of the host’s exposure status. Overall, our findings suggest that neonatal hyperoxia leads to long-term changes in the reparative versus cytotoxic nature of NK cells, due in part to intrinsic changes in hematopoietic cells. These differences may contribute to altered host responses to infection.
Liver sulfatide-reactive type II NKT cells recognize endogenous phospholipids. (INC9P.445)
Authors: Kato Shingo , Terabe Masaki , Berzofsky Jay 
Published: January 2023

Abstract
NKT cells are T cells recognizing lipids presented by CD1d. Type I and II subsets recognize distinct antigens. Type I NKT cells recognize alpha-galactosylceramide (aGC). aGC-loaded CD1d tetramers are widely used to study them. Type II NKT cells and their antigens are much less well characterized than type I due to lack of widely available tools to study them. Sulfatide (3-o-sulfo-beta-D-galactosylceramide), an endogenous lipid, is the only lipid proven to be recognized by type II NKT cells in vivo. Recently, phosphatidylglycerol (PG) and phosphatidylinositol (PI) were reported to be recognized by type II NKT cell hybridomas. Here, PG-or PI-loaded CD1d-tetramer-reactive cells were detected in liver mononuclear cells as a distinct population from aGC-loaded CD1d-tetramer-reactive type I NKT cells, consistent with type II NKT cells. Simultaneous staining with PG-, PI-, and sulfatide-loaded CD1d-tetramers indicated, surprisingly, that these three populations were largely identical, even though the phospholipids have structures quite distinct from that of sulfatide. IFN-gamma and IL-4 production by spleen cells was detected after stimulation with sulfatide, but not with PG or PI. These findings suggest that although PG and PI, when loaded onto CD1d, bind the same cells as do sulfatide-CD1d tetramers, their ability to induce cytokine production was distinct. Whether these phospholipids are weak agonists or antagonists or induce other functional activities is under investigation.
Dual variable domain autoantibodies to analyze the distinct role of BCR-TLR7 and BCR-TLR9 co-engagement on B cells (INM9P.455)
Authors: Pawaria Sudesh , Moody Krishna , Marshak-Rothstein Ann 
Published: January 2023

Abstract
Self-constituents can aberrantly engage pattern recognition receptors and result in autoimmune diseases. The activation of autoreactive B cells by DNA- and RNA- immune complexes through TLR9 and TLR7 respectively, has been implicated in the development of systemic lupus erythematosus (SLE). Interestingly, TLR9-deficient autoimmune-prone mice develop more severe disease and TLR7-deficient autoimmune-prone mice develop less severe disease. Previous studies in the lab were limited to using primary B cells from an autoreactive IgG2a-specific BCR transgenic line (AM14) together with monoclonal IgG2a autoantibody-autoantigen complexes to dissect the TLR7/9 dichotomy. We have developed bifunctional dual variable domain autoantibodies (autoDVDs) to analyze the differences between BCR-TLR9 versus BCR-TLR7 signaling in polyclonal (non-transgenic) mouse and human B cell populations. The autoDVDs consist of conventional heavy and light chains fused in tandem by a short linker, and then connected to either murine or human C1 constant region domains. These autoDVDs provide a covalent bridge between an anti-BCR and either a DNA-reactive or RNA-reactive antibody. Initial results suggest that the DNA DVDs are functionally similar to natural DNA immune complexes on non-transgenic B cells from different murine strains. The autoDVDs provide us an important tool to target autoantigens to human BCR and also provide a high-throughput screen for identifying TLR-targeted therapies.
Role of NKG2D in the induction of mouse natural killer cell exhaustion (IRC8P.473)
Authors: Alvarez Maite , Negrin Robert 
Published: January 2023

Abstract
Natural Killer (NK) cells have been used as a cancer therapy against multiple cancers. Despite initial promises, NK-based immunotherapy has resulted in limited success and therefore, new approaches are needed. Our group described the induction of NK cell exhaustion (NCE) upon tumor exposure that resulted in reduced cytotoxic function associated with lower expression of several molecules such as NKG2D and the transcription factors Eomesodermin and T-bet. Here we show that NCE induced by tumor cells is MHC-I independent and is not mediated by the expression of NKG2D ligands (NKG2DL) on tumor cells. However, when NKG2D-deficient NK cells were exposed to tumor cells limited proliferation and higher IFNg production was observed suggesting a role for NKG2D in NCE. Moreover, NCE patters were observed at later stages of in vitro IL2-activation that were reduced in NKG2D-deficient NK cells. Interestingly, concomitant downregulation of NKG2D and upregulation of NKG2DL (MULT1, Rae-1) was also detected. Upregulation of NKGDL expression was particularly seen after the cells were acid-stripped which was shown to disrupt MHC-I-Ly49s cis-interaction complexes. NCE was also observed after prolonged in vivo IL2 stimulation that impaired NK cell maturation. In conclusion, we demonstrated a role for NKG2D in the induction of NCE that could represent a self-regulatory mechanism important in controlling NK cell activation utilized by tumor cells to prevent NK cell anti-tumor responses.