bluuebunny/crossref_metadata_2025_split huggingface dataset:
Cigarette smoke extracts inhibit TLR ligand-induced inflammation via inactivation of NF-κB and activation of histone deacetylase in macrophages (INM7P.353)
Authors: Lee Kyoung-Hee , Jang An-Hee , Lee Chang-Hoon , Yoo Chul-Gyu 
Published: January 2023

Abstract
Cigarette smoke (CS) is the major risk factor of chronic obstructive pulmonary disease (COPD). CS has been reported to inhibit Toll-like receptor (TLR) function and phagocytosis in macrophages. Moreover, alveolar macrophages in COPD have impaired responsiveness to TLR stimulation. However, the molecular mechanism of CS-mediated impairments of macrophage immunity is not fully defined. We investigated the inhibitory mechanism of cigarette smoke extracts (CSE) in macrophages. Lipopolysaccharide (LPS, TLR4 ligand), Pam3CSK4 (TLR2 ligand), and CpG-oligodeoxynucleotides (TLR9 ligand) induced IL-6, TNF-α, and IL-1β production in RAW264.7 macrophage cells, which was significantly suppressed by CSE co-treatment. CSE did not block LPS-induced IκBα degradation, MAP kinase activation, and nuclear translocation of NF-κB subunit. However, it markedly suppressed mRNA expression of IL-6, TNF-α, and IL-1β, which implies that CSE regulate inflammatory cytokine production at the transcriptional level. Therefore, we next investigated the activity and modification of transcription factor, NF-κB, and histone deacetylase (HDAC). CSE significantly suppressed basal and LPS-induced NF-κB activity and reduced the level of acetyl-NF-κB p65. CSE increased HDAC activity and decreased the level of acetyl- and phospho-Histone H3 (Lys9/Ser10) expression. Together, these results suggest that CSE-mediated impairments of macrophage immunity are mediated by inhibition of NF-κB activation and activation of HDAC.
CD13 restricts TLR4 endocytic signal transduction in inflammation (INM7P.350)
Authors: Ghosh Mallika , Subramani Jaganathan , Rahman Mamunur , Shapiro Linda 
Published: January 2023

Abstract
The Toll-like Receptor 4 (TLR4) is the prototypical sensor of infection or injury that orchestrates the innate response via sequential activation of both cell-surface and endocytic signaling pathways that trigger distinct downstream consequences. Positive and negative regulators of this process exist to maintain the critical balance required for resolving inflammation. Myeloid CD13 negatively modulates receptor-mediated antigen uptake to control T cell activation in the adaptive immune response. Here we report that CD13 governs the internalization of TLR4 and subsequent innate signal transduction cascades leading to IRF3 activation independently of CD14, a positive regulator of TLR4. In response to LPS, CD13 is co-internalized with TLR4, CD14 and dynamin into Rab5+ early endosomes. Importantly, pIRF-3 activation and transcription of its targets are enhanced in CD13KO cells, while TLR4 surface signaling remains unaffected resulting in a skewed inflammatory response. Further, ischemic injury in vivo provoked identical TLR4 responses. Finally, CD13KO mice showed significantly enhanced type I Interferon-mediated signal transduction, escalating inducible nitric oxide synthase transcription levels and promoting accumulation of oxidative stress mediators and tissue injury. In response to injury or infection, CD13 negatively regulates TLR4 signal transduction, thereby balancing the innate response by maintaining the inflammatory equilibrium critical to innate immune regulation.
B4GALT6 regulates astrocyte activation during CNS inflammation (INM8P.360)
Authors: Mayo Lior , Trauger Sunia , Blain Manon , Mascanfroni Ivan , Kivisäkk Pia , Yeste Ada , Bakshi Rohit , Alvarez Jorge Iván , Prat Alexandre , Antel Jack , Weiner Howard , Quintana Francisco 
Published: January 2023

Abstract
Astrocytes play complex roles in neuroinflammation. Thus, it is important to characterize the mechanisms regulating astrocyte function, as well as potential targets for the therapeutic modulation of astrocyte activity. Here we report that during the chronic-progressive phase of EAE, astrocytes promote disease pathogenesis by a lipid-dependent signaling pathway. In line with previous studies, depletion of reactive astrocytes during the acute phase of chronic-progressive EAE exacerbated disease. However, depletion of astrocytes in the chronic phase ameliorated the disease. Hence, to understand this dichotomy we compared gene expression profiles of astrocytes from mice in acute and chronic stages of EAE. One gene associated with the chronic stage was B4GALT6, which codes for the β-1,4-galactosyltransferase 6 that catalyzes the synthesis of lactosylceramide (LacCer). We observed that LacCer levels are upregulated in the CNS during chronic EAE, and that LacCer synthesized by the astrocytes acts in an autocrine manner to trigger transcriptional programs that promote the recruitment and activation of infiltrating monocytes and microglia, and neurodegeneration. We also detected increased B4GALT6 expression and LacCer levels in MS lesions. Finally, the inhibition of LacCer synthesis suppressed CNS innate immunity and neurodegeneration, and interfered with the activation of human astrocytes. Thus, B4GALT6 is a potential therapeutic target for MS and neuroinflammatory disorders.
The extracellular adherence protein from Staphylococcus aureus inhibits the classical and lectin pathways of complement by blocking formation of the C3 proconvertase (INM8P.361)
Authors: Woehl Jordan , Stapels Daphne , Garcia Brandon , Ramyar Kasra , Keightley Andrew , Ruyken Maartje , Sfyroera Georgia , Weber Alexander , Zolkiewski Michal , Ricklin Daniel , Lambris John , Rooijakkers Suzan , Geisbrecht Brian 
Published: January 2023

Abstract
The pathogenic bacterium Staphylococcus aureus actively evades many aspects of human innate immunity by expressing a series of small inhibitory proteins. A number of these proteins inhibit the complement system, with the majority of S. aureus acting on the alternative pathway, blocking the amplification loop. Only a few proteins act on the initial recognition cascades that constitute the classical pathway (CP) and lectin pathway (LP). By screening a collection of recombinant, secreted staphylococcal proteins, we identified the extracellular adherence protein (Eap) as a potent, specific inhibitor of both the CP and LP. We found that Eap blocked CP/LP-dependent activation of C3, but not C4, and that Eap likewise inhibited deposition of C3b on the surface of S. aureus cells. In turn, this significantly diminished the extent of S. aureus opsonophagocytosis and killing by neutrophils. This combination of functional properties suggested that Eap acts specifically at the level of the CP/LP C3 convertase (C4b2a). Indeed, we demonstrated a direct, nanomolar-affinity interaction of Eap with C4b. Eap binding to C4b inhibited binding of both full-length C2 and its C2b fragment, which indicated that Eap disrupts formation of the CP/LP C3 proconvertase (C4b2). As a whole, our results demonstrate that S. aureus inhibits two initiation routes of complement by expression of the Eap protein, and thereby define a novel mechanism of immune evasion.
Intranasal or intraperitoneal administration of human tubal mesenchymal stem cells reduces T lymphocytes-associated lung inflammation in cigarette-induced emphysema in mice (IRC10P.415)
Authors: Peron Jean Pierre , de Brito Auriléia , Rigonato Nicole , Vitoretti Luana , Moraes Gabriel , Soares Clariana , Czeresnia Carlos Eduardo , Perin Paulo , Maluf Mariângela , Zatz Mayana , Ligeiro de Oliveira Ana Paula 
Published: January 2023

Abstract
Objective: To study the effectiveness of human tubal Estromal Stem Cells (htESCs) therapy by intra-nasal or intra-peritoneal route in experimental cigarette smoke-induced emphysema in C57BL/6 mice. Design: To achieve that, C57BL/6 mice were exposed to cigarette smoke for 75 days (2 times / day) and received or not two doses of htMSCS either intraperitoneally or intranasally. Experiments were performed on day 76th. Materials and Methods: Briefly, brochoalveolar lavage fluid was obtained and either used to measure cytokine secretion or inflammatory infiltrate. Moreover, lung tissue was also submitted to imunohistochemistry for the transcription factors NF-AT and NF-kB and also IL-10. Results: Our results clearly show that htMSCs greatly reduced lung inflammation, with decreased cellular infiltrate of T CD4 and T CD8 lymphocytes, which was confirmed by reduced NF-AT staining in lung tissues. This was associated with reduced pro-inflammatory cytokine secretion, as TNF-a, IL-1b and IL-6, less mucus and collagen deposition, and interestingly, increased IL-10. Conclusion: Altogether, our data indicate that therapy with htMSCs for the treatment of emphysematous disease may be a promising clinical approach. Moreover, intranasal route is also effective, and must taken in consideration due to its lack of invasiveness.
NKT cell-mediated enhancement of humoral immunity against Clostridium difficile toxin B (IRC10P.406)
Authors: Rampuria Pragya , Devera Thomas , Lang Mark 
Published: January 2023

Abstract
To develop more effective antibody-stimulating vaccines, we must improve our understanding of the mechanisms by which vaccines induce and maintain long-lasting humoral immunity. This is important for the development of new vaccines against emerging pathogens such as C. difficile. It is well-established that Natural Killer T (NKT) cells, activated with the glycolipid, α-galactosylceramide (α-GC) enhance humoral immunity, but the mechanisms by which they do so are poorly understood. Here we show that activated NKT cells enhance the antigen-specific IgG1 response and survival against C. difficile Toxin B (TcdB). We and others have shown that NKT cells may enhance priming of T helper (Th) cells. We therefore hypothesized that T helper and NKT subsets coordinate to enhance the humoral immune response against C. difficile. Flow cytometry analysis demonstrated that NKT cell activation with a-GC did not alter numbers of Th or follicular T helper (Tfh) cells, but increased the number of NKTfh cells. ELISPOT assays indicated that the α-GC enhanced secretion of IL-4 and IL-21 is mainly Th-derived. Adoptive transfer experiments show that α-GC did not alter the ability of Th cells to boost antibody production. This suggests that NKT cells may exert direct and indirect effects (via Th cells) on humoral immunity. Understanding the mechanisms by which NKT cells enhance humoral immunity will help in designing strategies to appropriately activate NKT cells and boost vaccine efficacy.
CTLA-4-mediated suppressor effect by human T follicular regulatory cells (IRC11P.432)
Authors: Sayin Ismail , Herati Ramin , Wherry E. , Betts Michael , Canaday David 
Published: January 2023

Abstract
T follicular helper (Tfh) cells, a subset of CD4 T cells, help B cells produce high affinity class switched antibodies and are essential for generation of germinal centers (GC). T follicular regulatory cells (Tfr) are a newly defined regulatory CD4 T cell subset that suppresses GC formation and B cell function through CTLA-4 dependent mechanisms in murine models. We will test the hypothesis in human systems that Tfr have significant regulatory effects on antibody production in a CTLA-4 dependent manner. Tfh and Tfr cells from human mesenteric lymph nodes were analyzed using flow cytometry. Tfr were defined as CXCR5+, PD-1bright, FoxP3+ cells that were also found to be Bcl6+, CD25+, CD127dim, ICOS+. Flow sorted Tfr and Tfh cells were used to determine the effect of Tfr on Tfh-mediated B cell production of IgG. Tfh cells with mitogen helped B cells produce high amounts of IgG whereas addition of Tfr to this co-culture decreased the IgG levels &gt;90%. Tfh/Tfh ratio directly impacts levels of suppression. The mechanism of the suppressive effect of Tfr cells was investigated by neutralizing IL-10, TGF-β RII and CTLA-4 in the co-culture functional assay. Only anti-CTLA-4 affected IgG production. The present study in the human system suggests that Tfr cells suppressed Tfh-induced antibody production by B cells with CTLA-4 playing an important role in this suppression. These regulatory interactions may play a role in autoimmune diseases and humoral response to vaccines and pathogens.
AhR acts as a Foxp3-independent transcription factor for the suppression of type 1 diabetes in NOD mice (IRC11P.425)
Authors: Ehrlich Allison , Pennington Jamie , Wang Xisheng , Kerkvliet Nancy 
Published: January 2023

Abstract
The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor associated with immunosuppression. We previously reported that suppression of type 1 diabetes by TCDD, a potent AhR ligand, is associated with an increased percentage of Foxp3+ regulatory T cells. Similar associations between AhR activation and increased Foxp3+ cells have been reported for suppression of other autoimmune diseases; thus, AhR is a potential therapeutic target. We recently discovered a novel, rapidly metabolized AhR ligand, 10-chloro-7H-benzimidazo[2,1-a]benzo[de]iso-quinolin-7-one (10-Cl-BBQ), that activates AhR in CD4+ T cells, in a manner similar to TCDD. Here we show oral treatment with 10-Cl-BBQ is highly effective in suppressing pancreatic islet infiltration through 20 weeks of age in NOD mice. When Foxp3+ cells were assessed, a small yet significant increase in the frequency, but not total number, of cells was observed. This raised the yet unanswered question, is the increase in Foxp3+ Tregs driving AhR-mediated immune suppression? To directly assess the requirement of Foxp3+ cells in AhR induced suppression of islet infiltration, transgenic NOD.Foxp3DTR mice were employed. In control mice, Foxp3 depletion led to rapid pancreatic islet infiltration. However, treatment with 10-Cl-BBQ suppressed infiltration even in the absence of Foxp3+ cells. This suggests that AhR functions as a Foxp3-independent transcription factor driving suppression of T cell dependent immune responses.
Genome wide mapping of epigenetic signatures identify novel enhancer elements that underpin virus-specific CD8+ T cell differentiation (IRM14P.445)
Authors: Turner Stephen , Olshansky Moshe , Li Jasmine , Nguyen Michelle , Chen Eddie , Nguyen T.H. , Rao Sudha , Russ Brendan 
Published: January 2023

Abstract
The molecular mechanisms that underpin acquisition and maintenance of lineage specific gene expression by virus-specific CD8+ T cells are not fully delineated. Post-translational modifications (PTMs) of genome associated histone proteins are a key mechanism for regulating gene expression. We mapped genome wide changes in histone PTM deposition during virus-specific CD8+ T cell differentiation. By comparing the location and level of deposition of a combination of H3K4me1, H3K4me2, H3K4me3, H3K27me3, H3K27Ac, chromatin accessibility and binding of the histone acetyltransferase, p300, we could map both known and putative enhancer elements that are differentially regulated between naive, effector and memory virus-specific CD8+ T cells. Using this epigenetic blueprint, we identified putative transcriptional enhancers located upstream of the Ccl5 gene, a key CD8+ T cell effector gene. We demonstrate that acquisition of Ccl5 transcription upon T cell activation is regulated via temporal and transcription factor (TF) dependent chromatin remodelling. Using chromatin conformation capture (3C) to measure enhancer-promoter interactions, we show that that TF dependent looping of these enhancer elements onto the Ccl5 promoter correlated with transcriptional activation and potential. These studies form a platform for more in depth analysis of key non-coding regulatory elements that regulate acquisition of virus-specific T cell gene expression and maintenance into memory.
The immunological clock: the role of cryptochrome in regulatory T cell function (IRM15P.602)
Authors: Mack Lauren , LeBlanc Mathias , Zheng Ye 
Published: January 2023

Abstract
Circadian rhythms are defined as 24 hour oscillations in physiological and behavioral processes. Clinical data suggests circadian involvement in the predisposition and progression of immune-related morbidities such as cancer and autoimmune disease. Recent studies have demonstrated the direct role of core molecular clock components in maintaining the immune system. A subset of CD4+ T cells known as regulatory T cells (Treg) are critical for preserving balance in the immune system and have been shown to be able to prevent a broad range of autoimmune disease. By studying the role of core circadian genes in Treg cells the connection between the circadian clock and autoimmune disease may be further elucidated. We have identified a core circadian clock gene, cryptochrome (Cry), to have a role in Treg cell function both in vitro and in vivo. The deficiency of Cry in Treg cells results in a breakdown in immune tolerance, which results in severe pathological autoimmunity in mice. This is characterized by immune cell infiltration into peripheral tissues as well as increased numbers of cytokine producing CD4 and CD8 T cells. It was also seen that ectopic expression of Cry in Treg cells results in increased suppression of T cell proliferation in vitro. Thus, we have demonstrated a direct role of Cry expression in the function of Treg cells.
Role of miR-27 in controlling T cell immunity (IRM15P.605)
Authors: Cruz Leilani , Cho Sunglim , Wu Chen-Jang , Yasuda Tomoharu , Lin Ling-Li , Rajewsky Klaus , Lu Li-Fan 
Published: January 2023

Abstract
miR-23~27~24 clusters are highly enriched in regulatory cells while expressed at low levels in conventional T cells. However, despite a selective expression pattern of miR-23~27~24 clusters in T cells, to date, studies of this miRNA family have primarily focused on their role in tumorigenesis. Here, we show miR-23~27~24 clusters play a pivotal role in regulating T cell immunity. Particularly, our results have demonstrated that the majority of miR-23~27~24 cluster-dependent phenotypes could be attributed to a single member of this miRNA family, miR-27. Mice with T cell-specific overexpression of miR-27 spontaneously developed autoimmune phenotypes. On the other hand, in vitro polarization studies revealed that T cells with miR-27 overexpression exhibited impaired differentiation and effector function of multiple T helper cell lineages. Finally, our mixed bone marrow chimeras studies further demonstrated that miR-27 could control different T cell responses through either a T cell-intrinsic or -extrinsic manner. Collectively, Our results identify a new "immune regulatory" miRNA that can play multi-faceted roles in regulating T cell immunity.
FOXP3 regulates RNA alternative splicing in regulatory T cells (IRM15P.461)
Authors: Zhou Baohua , Du Jianguang , Ziegler Steven 
Published: January 2023

Abstract
Alternative pre-mRNA splicing is an important mechanism for protein regulation and cellular function, affecting more than 95% of multi-exon genes in humans and contributing to a wide range of physiological and pathophysiological processes. It is well known that FOXP3, the master transcription factor for Treg differentiation and function, directly binds to its target genes to regulate their transcription. Here we report an interaction between FOXP3 and heterogeneous nuclear ribonucleoprotein F (hnRNPF), a protein that regulates pre-mRNA alternative splicing and/or 3’ polyadenylation. FOXP3 through its exon 2 region interacts with hnRNPF to modulate the alternative splicing pattern of hnRNPF targets like BCL-X. Interestingly, the alternatively spliced exon 2 of human FOXP3 contains hnRNPF putative binding sites and point mutations of these sites eliminate the alternative splicing of human FOXP3. These findings suggest human FOXP3 might autoregulate the alternative splicing of FOXP3 transcripts through its interaction with hnRNPF. Finally, overexpression of hnRNPF in mouse iTregs can reduce their ability to suppress CD4+ T cell proliferation. Taken together, our results reveal a novel molecular mechanism through which FOXP3 modulates pre-mRNA alternative splicing to regulate the differentiation and function of Tregs.
B cell differentiation governed by distinct microenvironments (LYM7P.622)
Authors: Cen Selena , Simard Nathalie , Paige Christopher 
Published: January 2023

Abstract
B cell differentiation is regulated by the surrounding microenvironment, which produces cytokines, chemokines, and adhesion molecules that guide B cell commitment and differentiation. The gastrointestinal tract, in constant interaction with microbiota, constitutes a distinctive microenvironment for B cell differentiation. While the bone marrow and spleen contain few IgA+ cells, the gut lamina propria is largely colonized by IgA+ cells, which contain a subset of myeloid-like plasma cells expressing IgA, TNFα, iNOS, CD11c, Ly6C and Ly6G. Whether the bone marrow stroma produces inhibitory factors for IgA+ cell generation or the gut lamina propria stroma produces inducing factors needs further investigation. We developed an in vitro system to co-culture stromal cells with B cells to characterize the signals necessary to generate IgA+ cells. It was found that intestinal microbial components along with the intestinal lamina propria stromal cells played an important role in the emergence of both IgA+ and myeloid-like plasma cells. Interestingly, bone marrow and spleen-derived stroma showed an inhibitory effect on the generation of IgA+ cells. Stromal soluble factors (e.g. IL-1, IL-6, and MCP-1) might play a role in the guidance of B cell IgA switching, proliferation, survival, and differentiation. These experiments will provide a better understanding of the impact of microenvironments in shaping B cell development, and we will identify the molecular mechanisms they use.
Gut microbial dysbiosis due to Helicobacter drives marginal zone B cell development in the absence of IL-10 signaling in macrophages (LYM7P.623)
Authors: Dittel Bonnie , Basu Sreemanti , Gharaibeh Raad , Fordor Anthony , Ray Avijit 
Published: January 2023

Abstract
IL-10 plays an essential role in maintaining homeostasis in the gut in response to the microbiome. However, it is unknown whether IL-10 also facilitates immune homeostasis at distal sites. To address this, we asked whether splenic immune populations were altered in IL-10-deficient mice with differential susceptibility to microbiome-dependent spontaneous enterocolitis. Susceptible mice exhibited a significant increase in splenic macrophages, neutrophils and marginal zone (MZ) B cells that was inhibited by IL-10 signaling in macrophages, but not B cells. The increase in myeloid cells was due to increased proliferation that correlated with increased MZ B cell differentiation. Cohousing and antibiotic treatment suggested that the alteration in splenic immune homeostasis was microbiome-dependent. 16S rRNA sequencing reveled that susceptible mice harbored a more diverse microbiome with a significant increase in the abundance of the bacterial genus Helicobacter. The introduction of H. hepaticus to the gut of nonsusceptible mice was sufficient to drive myeloid cell expansion and MZ B cell development. Given that MZ B cells are part of the first line of defense against blood-borne-pathogens, their increase following a breach in the gut epithelial barrier would be protective. Thus IL-10 is an essential gatekeeper that maintains immune homeostasis at distal sites that can become functionally imbalanced upon the introduction of specific bacteria to the intestinal track.