bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Role of Beta Cell Antigen Presentation in Dendritic Cell-Based Type 1 Diabetes Immunotherapy (B189)
Authors: Xia Chang-Qing , Qiu Yushi , Peng Ruihua , Lo-Dauer Jeannette , Clare-Salzler Michael J 
Published: January 2023

Abstract
Dendritic cells (DCs) are promising cells for type 1 diabetes (T1D) immunotherapy. Most studies have been focusing on how to modulate DCs with tolerogenicity, but little attention has been directed towards the antigenic epitopes presented by DCs. In this study, we evaluated the roles of beta cell antigens presented by immature DCs (iDCs) in T1D prevention. At first, we established that beta cell antigen pulsing was required for iDCs to prevent T1D by using iDC pulsed with whole beta cell (NIT1) lysates. We further evaluated the capacity of different beta cell-derived antigenic peptides (Insulin B9-23, GAD78-97, GAD217-236, proInsulin C3-A19) in priming NOD splenic T cells in vitro. We discovered that GAD217-236 predominantly stimulated Th2 cells. Consistent with the in vitro results, 4 weekly intravenous injections of GAD217-236-pulsed iDCs significantly prevented diabetes in younger mice. Moreover, the same results were achieved when older NOD mice were treated. In the T cell recall experiments, it was noted that the splenocytes from the mice treated with GAD217-236-plused iDCs, in contrast to PBS-treated mice, produced significantly higher IL-4 and IL-10. Of interest, four beta cell antigenic peptides mentioned above stimulated the similar cytokine-producing pattern, suggesting that the immune response induced by GAD217-236-pulsed iDCs spreads to other beta cell antigens through epitope spreading.
Evaluation of cellular immune responses induced in mice by alphavirus replicon particle vaccines for SARS-CoV (B190)
Authors: Reap Elizabeth A , Norberg Pamela K , Watson Aubrey D , Cecil Chad , Wagner Jordan , Kamrud Kurt I , Smith Jonathan 
Published: January 2023

Abstract
An RNA replicon vector system derived from an attenuated strain of VEE virus was used to produce virus-like replicon particles (VRP) expressing SARS-CoV genes. We previously showed in mouse studies that complete protection was conferred by SARS S VRP and partial protection with SARS M VRP. In this study, two assays for cellular immunity were used to characterize the T cell phenotypes and cytokine profiles induced after SARS VRP vaccination. Following SARS M, N, S or E VRP immunization of BALB/c mice, splenocytes were harvested for antigen specific ICS and ELISPOTs. The SARS S VRP induced both CD4 and CD8 responses with the latter being the dominant component as measured by ICS for IFN-γ or TNF-α. Polyfunctional CD8 cells secreting both IFN-γ and TNF-α were also detected. Following SARS N VRP immunization, T cells secreting IFN-γ or TNF-α were readily detected with cells secreting both predominately identified as CD4 cells. Th1 cytokine secretion was not detected in cells from SARS M VRP inoculated mice. SARS E VRP elicited a robust CD8 response as shown by cytokine expression in cells stimulated with a 9mer peptide. IL-4 was not detectable after any SARS VRP vaccination. The IFN-γ ELISPOT results paralleled those from ICS, indicating that the SARS VRP vaccines induced Th1 biased responses, although the cytokines elicited differed depending on the SARS gene expressed.
This study was supported by NIH grant #UC1-AI62582
Evaluation of the immunogenicity of an alphavirus replicon particle vaccine expressing the SARS-CoV spike (S) glycoprotein in non-human primates (B191)
Authors: Reap Elizabeth A , Watson Aubrey D , Norberg Pamela K , Cecil Chad , Dornsife Ronna E , Kamrud Kurt I , Smith Jonathan 
Published: January 2023

Abstract
We previously evaluated the immunogenicity and efficacy of a virus-like replicon particle (VRP) vaccine expressing the SARS CoV S glycoprotein in mice. SARS S VRP were highly immunogenic and conferred protection upon challenge. To extend that work we have evaluated the immune responses induced by SARS S VRP in macaques immunized at 0, 4 and 24 weeks. PBMC were assayed at two week intervals by ELISPOT and ICS. Post prime, both CD4 and CD8 T cells were found that secrete IFN- γ, IL-2 and TNF- α. The percentage of CD4 cells secreting IFN- γ, IL-2 and TNF- α was substantially more than the percentage of CD4 cells secreting IL-4 post prime indicating a Th1 bias. Post boost, the number of CD4 cells secreting all measured cytokines were equivalent indicating that the SARS S VRP induced a balanced Th1/Th2 response after 2 inoculations. By week 8 a measured decline in the percentage of cytokine-secreting CD8 cells was observed. There were no significant differences in the numbers of T cells secreting cytokines with 2 different VRP dosages given as measured by ICS, however a dose response was seen by ELISPOT. Neutralizing antibodies (NAb) to SARS CoV were measured in sera following post prime and boost inoculations. Three of 4 animals in the low dose group had measurable NAb post boost (GMT = 101) whereas all 4 animals in the high dose group had serum NAb after two doses (GMT = 190). These data provide support to further development of a SARS S VRP vaccine.
This study was supported by NIH grant #UC1-AI62582
Development and Qualification of a Human IFN-gamma ELISPOT Assay for Testing T Cell Responses to an HIV gag DNA Vaccine (B203)
Authors: Chen Hong , Cohen Michael , Rappaport Ruth , Wu Hong-Yin 
Published: January 2023

Abstract
Cell-mediated immunity is believed to be critical for the control and clearance of viral infections. However, frequencies of antigen-specific human T cells are rather low, quantitation of these cells requires assays that exhibit robust sensitivity, specificity, and reproducibility. In this study, a peptide-induced human IFN-gamma ELISPOT assay has been developed and qualified for testing T cell responses to an HIV gag DNA vaccine. We optimized the number of cells per well, the concentrations of HIV gag peptide pools used for stimulation, and the incubation time of cells with the stimulants. We found that overnight-cultured PBMCs, rather than immediately thawed PBMCs, yielded a greater number of spots. The assay is specific, as it detected IFN-gamma but not IL-2 or granzyme B secretion by stimulated PBMCs. The assay results were reproducible and robust across a wide range of cell concentrations. Coefficients of variation of 25% or less were observed for intra-assay replicates from samples containing more than 50 spot-forming cells. Furthermore, the use of cryopreserved PBMC samples allowed direct comparison within a single assay run of samples from multiple time points during the course of a vaccination regimen, minimizing potential run-to-run assay variability. These results indicate that the ELISPOT assay was reliable and suitable for direct ex vivo quantitation of antigen-specific T cells in HIV vaccine trials.
IDRs as Novel Immunomodulators (B208)
Authors: Guarna Marta , Yang Haiyan , Glavas Natalie , Deng Yanshen , Dullaghan Edie , Mookherjee Neeloffer , Bishop Jennifer , Waldbrook Matt , Donini Oreola , Scott Monisha , Gold Michael R. , Finlay Brett , Hancock Robert E. W. , North John R. 
Published: January 2023

Abstract
Inimex Innate Defence Regulators (IDRs) are novel, synthetic immunomodulatory peptides that protect against infections by selectively activating the innate immune system while regulating inflammation. The multi-faceted effects of IDRs are mediated primarily by monocytes and macrophages. These cells respond to IDRs by selectively increasing the expression of cell surface receptors, and cytokines and chemokines (MCP-1, MCP-3 and CCL-5) which trigger the recruitment and activation of immune cells to the site of the infection. In addition, IDRs control inflammation by enhancing the expression of the anti-inflammatory cytokine IL-10, and down-regulating the release of pro-inflammatory cytokines TNF-α and IL-6 in response to pathogen-associated stimuli. As a result, IDRs selectively activate the immune system without concomitant up-regulation of inflammatory responses. This combination of IDR effects is a distinctive quality of these agents since they are able to maintain a balance between immunostimulatory and inflammatory responses to an invading pathogen.
Haiyan Yang is the recipient of an NSERC Industrial R&D Fellowship.
Funded in part by a grant from the FNIH and the CIHR through the Grand Challenges in Global Health initiative
Atorvastatin treatment decreases brain Amyloid-β and serum TNF-α levels in Alzheimer’s disease transgenic mice (B214)
Authors: Kao Patricia F. , Gupta Eric K. , O’Barr Stephen A. 
Published: January 2023

Abstract
Alzheimer’s Disease (AD) is the most common form of dementia affecting over 14 million elderly worldwide. AD, which has no single cause, is characterized neuropathologically by deposits of Amyloid-β (Aβ) peptides, neurofibrillary tangles, reactive astrocytosis, activation of microglial cells, and deficits in the cholinergic system. Since inflammatory markers are increased markedly in AD brain, it is likely that they participate directly in the destruction of brain cells, and may predict the development of AD years before clinical symptoms appear. Non-steroidal anti-inflammatory drug (NSAID) therapies have shown clinical benefit in treating AD. Likewise, it has been hypothesized that the anti-inflammatory properties of statins may alter AD pathology, independent of their cholesterol lowering effect.
In this current study, we treat huAPP R1.40 YAC transgenic mice, a model of Alzheimer’s disease, with therapeutic doses of atorvastatin. Mice either receive a control dose of vehicle, a low dose of atorvastatin or a high dose of atorvastatin via oral gavage. After 12 weeks, animals are sacrificed and levels of Aβ and inflammatory markers, such as IL-1β, IL-6 and TNF-α, were measured. We find that Aβ levels in membrane fractions of the brain are reduced while cytosol fractions show no change. In serum, TNF-α levels decrease with atorvastatin treatment.
EVIDENCE FOR THE EXISTENCE OF AN ALTERNATIVE B CELL DEVELOPMENT PATHWAY (B3)
Authors: Kumar Ritu , Fossati Valentina , Deriva Alessandra , Martensson Lill , Snoeck Hans Willem 
Published: January 2023

Abstract
During B-cell development rearrangement of immunoglobulin heavy and light chain genes occurs sequentially, separated by a pre-BCR checkpoint where the quality of the heavy chains is tested by pairing with a surrogate light chain (SLC). Here we demonstrate the existence of a second B-cell development pathway, that is characterized by a high expression of Sca1 and LFA1, is less dependent on SLC expression and does not have a late pre-B stage. This pathway becomes predominant with age and in conditions of lymphopenia, and is biased towards marginal zone B cells. However, cell fate switches occur between marginal zone and follicular B cells, likely to maintain marginal zone homeostasis. FO B cells derived directly from MZ B cells are distinguished by a higher expression of Sca1. Thus, an alternative, and perhaps ancestral, B-cell development pathway exists that is biased toward to the generation of MZ B cells.
Induction of Chemokines through LTB4 Receptors, BLT1 and BLT2, in Primary Human Monocytes (B47)
Authors: Sun Binggang , Huang Jing-Feng , Dai Heng , Lars Karlsson , Fourie Anne 
Published: January 2023

Abstract
Leukotriene B4 (LTB4) is a potent lipid mediator of inflammation. The biological functions of LTB4 are mediated through two homologous receptors with high and low affinities, namely, BLT1 and BLT2. Using human primary monocytes, we are trying to understand the functions of LTB4 in atherosclerosis from a transcriptional activation perspective. We have also examined the contributions from each receptor. Primary human monocytes were treated with LTB4 and total RNA prepared and analyzed by microarray. Of particular interest, among genes upregulated were 4 chemokines including CCL2/MCP-1, CCL7/MCP-3, CXCL2/Groβ and CXCL3/Groγ. Follow up analysis with real-time PCR and ELISA showed time- and dose-dependent induction of these chemokines. The induction of MCP-1 is Gi-coupled since PTX effectively blocked the induction. BLT2 antagonist LY255283 dose-dependently inhibited the induction of MCP-1 gene expression, while BLT1 agonist, U75302 did the opposite. Selective inhibitors of Ca2+ signaling, PI3K, tyrosine kinase and ERK1/2, but not JNK, effectively blocked the LTB4-induced MCP-1 production. To understand the relative contributions of each receptor, we used receptor specific agonist U75302 for BLT1 and 12(S)-HETE for BLT2. We showed that both agonists induced the expression of all four genes, suggesting overlapping roles of the two receptors in induction of gene expression of these chemokines. More interestingly, 12(S)-HETE greatly increased the induction of MCP-1, MCP-3 and CXCL2 by U75302, suggesting synergistic effect between the two receptors.
The Regulation of CCL19 Gene Expression in Human Antigen Presenting Cells (B49)
Authors: Pietila Taija E , Veckman Ville , Lehtonen Anne , Osterlund Pamela , Julkunen Ilkka 
Published: January 2023

Abstract
CCL19 chemokine plays an important role in regulating DC traffic and recruitment of naïve T cells to the vicinity of activated DCs and macrophages. We have analyzed the regulation of CCL19 gene expression in human monocyte-derived DCs/macrophages infected with Salmonella enterica or Sendai virus. Computer analysis of the CCL19 promoter identified two putative NF-κB binding sites and one interferon stimulated response element (ISRE). Transcription factor binding experiments demonstrated that Salmonella or Sendai virus infection increased the binding of NF-κB proteins (p50, p52, p65, and RelB) to both CCL19 promoter NF-κB elements. Salmonella or Sendai virus infection also increased the binding of multiple IRFs, as well as STAT1 and STAT2, to the ISRE site.
The role of NF-κB and various IRF family members in transcriptional control of CCL19 gene was further studied in transfection experiments. The expression of NF-κB dimers readily activated CCL19 promoter. The expression of IRF1, IRF3, or IRF7 proteins was also able to activate the promoter in the presence of Sendai virus infection. CCL19 promoter constructs containing mutated NF-κB and/or ISRE sites were weakly activated. Our data also suggests that c-Jun and c-Fos proteins binding to target AP-1 elements in CCL19 promoter plays a role in transcriptional control of CCL19. Further work focuses on the regulation of CCL19 in human macrophages and myeloid DCs.
A Role for IL-27/IL-27R Signaling in Prevention of Acute Intestinal Inflammation (B53)
Authors: Troy Amy E. , Saris Christiaan J M , Hunter Christopher A , Artis David 
Published: January 2023

Abstract
IL-27 is a member of the IL-12 cytokine family initially implicated in promoting Th1 cell differentiation. IL-27 is elevated in the intestines of inflammatory bowel disease patients and in a murine model of this disease, although the influence of IL-27 on intestinal inflammation remains unclear. To address this, we employed IL-27R−/− mice and the dextran sodium sulfate (DSS) model of acute intestinal inflammation. Following exposure to 5% DSS, IL-27R−/− mice exhibited early and severe disease, as defined by increased weight loss, increased severity of edema and inflammatory lesions in the colon, and increased diarrhea and intestinal bleeding. This pathology lead to high mortality in IL-27R−/− mice during the first five days of DSS treatment. Cytokines IL-6 and IL-17 were elevated in IL-27R−/− mice, while other pro-inflammatory cytokines such as IFNγ remained unchanged. IL-17 was produced both by CD4+ T cells and by non-T cells. Finally, the lamina propria of DSS-treated IL-27R−/− mice contained increased frequencies of innate immune cells following treatment. These results indicate that endogenous IL-27-IL-27R signaling can regulate both innate and adaptive immune cells and contribute to prevention of severe intestinal inflammation.
Assessing function of skin homing T cell selectin-ligands via selective knockdown with siRNA (B62)
Authors: Thieu Khanh P , King Sandy , Fuhlbrigge Robert C 
Published: January 2023

Abstract
The recruitment of T cells to skin is a key feature of inflammatory diseases such as psoriasis and eczema. Skin homing T cells express cutaneous lymphocyte-associated antigen (CLA), a carbohydrate epitope that binds to E-selectin on postcapillary venules and mediates lymphocyte tethering and rolling in shear flow. E-selectin ligand (ESL) activity and CLA epitope expression have been described on P-selectin glycoprotein ligand-1 (PSGL-1) and leukosialin (CD43). CLA+ PSGL-1 serves as both a P-selectin ligand (PSL) and an ESL, whereas CLA+ CD43 is only an ESL. Although their ESL functions appear redundant, the true relative contribution of PSGL-1 and CD43 to ESL activity in human CLA+ T cells has not yet been evaluated.
Using siRNA, we inhibited expression of PSGL-1, CD43, or both on CLA+ T cells by 70–80%, as measured by flow cytometry. PSGL-1 knockdown resulted in a 70% reduction of PSL but relatively unchanged ESL activity, as assessed by shear flow binding assays. CD43 knockdown did not signficantly impact either PSL or ESL activity. Surprisingly, knockdown of both PSGL-1 and CD43 did not noticeably reduce ESL activity nor CLA expression.
As predicted from animal models, PSGL-1 is the major PSL on human CLA+ T cells but serves a redundant role as an ESL. The high residual amounts of CLA expression and ESL activity remaining after knockdown of both PSGL-1 and CD43 suggest that an uncharacterized ESL distinct from PSGL-1 and CD43 exists and may represent a signficant pool of ESL function on skin homing T cells.
Regulation of Toll Like Receptor-3 mediated inflammatory responses by glucocorticoid receptors: Role of MAP Kinases (B66)
Authors: BHATTACHARYYA SANDIP , MUGLIA LOUIS J. 
Published: January 2023

Abstract
Exaggerated activation of toll like receptors (TLRs) often causes chronic inflammatory disorders. Glucocorticoids are a mainstay of therapy for the treatment of an array of inflammatory diseases. Although macrophages are the integral part of TLR-mediated immune response, the crucial mediators present in macrophages to regulate the complex proinflammatory intracellular signaling are yet to be fully delineated. To explore the role of glucocorticoids in this context, we generated mice (MGRKO) with conditional deletion of glucocorticoid receptor (GR) in macrophages. We show that in vivo activation of TLR-3 by Poly (I:C) caused elevated serum TNF-α (2.5 fold, p&lt;0.01), IL-6 (3.5 fold, p&lt;0.01), IL-12 (3 fold, p&lt;0.01) in MGRKO mice as compared to control mice. In vitro studies with cultured macrophages showed that pretreatment with dexamethasone (Dex) significantly inhibited TLR-3 induced expression of TNF-α (3.4 fold), IL-6 (3.5 fold), and IL-12 (4.3 fold) in the control but no effect in GR-deficient macrophages. We observed that among the major MAP kinases JNK was the crucial target for GR after TLR-3 activation. This hypothesis was validated by the use of specific inhibitors to MAP Kinase pathways. In contrast, after TLR-4 activation Dex selectively suppressed p38 MAPK with no effect on JNK. Collectively, we identify JNK as an integral part of GR-driven regulation of TLR-3 mediated inflammatory responses.
This work was supported by grants from NIH/NIAD to Louis J Muglia.
Characterization of a novel, chronic inflammatory response following spinal cord injury with flow cytometry (B67)
Authors: Beck Kevin Douglas , Nguyen Hal X , Galvan Manuel , Salazar Desiree L , Anderson Aileen J 
Published: January 2023

Abstract
Traumatic spinal cord injury results in a breakdown of the blood brain barrier allowing inflammatory cells to enter the cord, potentially mediating both damage and repair. Several studies have described the acute cellular inflammatory response, however our study quantifies inflammatory cells up to 90 days post injury, revealing a previously uncharacterized chronic inflammatory phase. We developed a rapid, debris-minimizing flow cytometry method, allowing for accurate quantification of cellular inflammation in a defined area of the injured spinal cord. We validated the technique by confirming its ability to distinguish differences in cellular inflammation between mild, moderate, and severe injuries. Using this method, we quantified inflammatory cells in segments T8-T10 following a 200 kd contusion injury to T9 in female Sprague-Dawley rats. We confirm and expand upon the published timecourse for both neutrophil and macrophage/microglial infiltration of the injured spinal cord by assaying daily for the first 10 days post injury and over wider intervals up to 90 days post injury. Interestingly, we were able to detect novel chronic inflammation months after injury, with the number of macrophages at 60 days exceeding that at 7 days, the previously published peak of activity. Further experimentation is needed to test whether the chronic inflammatory response aids remyelination or regeneration of damaged axons.
Naïve T cells activated in the liver by hepatocytes express a phenotype consistent with death by neglect (B69)
Authors: Bertolino Patrick Jerome , Holz Lauren , O’Reilly Lorraine , Benseler Volker , McCaughan Geoffrey 
Published: January 2023

Abstract
The liver has unique immunological properties. We have proposed that these properties are associated with the unique ability of the hepatic tissue to support primary activation of CD8+ T cells independently of lymphoid tissues. The fate of T cells activated in the liver however remains a matter of debate. Our results indicate that liver-activated cells die prematurely due to lower expression of cytokines and cell survival genes. In contrast, recent studies in a mouse liver transplantation model indicate that liver-activated T cells mature into full effector cells able to recirculate into lymph nodes (LN) and become tolerised at a latter stage through other mechanisms. In this latter model, it is not clear whether full effector T cells originate from recirculating T cells activated in LN or whether intrahepatic inflammation changes the phenotype of T cells activated locally. To address specifically the recirculation patterns, phenotype and fate of naïve CD8+ lymphocytes activated in the liver and LN in the absence of injury, we used transgenic mouse models in which the antigen is expressed at only one of these sites. Liver-activated cells down-regulated CD62L and thus, were unable to recirculate to LN during the first 3 days. Importantly, these cells were found to express lower levels of CD25 and CD54 than LN-activated cells. Simultaneously anti-apoptotic proteins were upregulated in liver-activated cells whereas this did not occur in LN-activated cells. Thus, liver-activated CD8+ T cells do not recirculate to the LN at early time points and are rather programmed to undergo premature death in vivo thus contributing to the liver tolerance effect.