bluuebunny/crossref_metadata_2025 huggingface dataset:
Modulation of C3a Activity: Internalization of the Human C3a Receptor and its Inhibition by C5a
Authors: Settmacher Britta , Bock Daniel , Saad Henry , Gärtner Sören , Rheinheimer Claudia , Köhl Jörg , Bautsch Wilfried , Klos Andreas
Published: December 2022

The C3a receptor (C3aR) is expressed on most human peripheral blood leukocytes with the exception of resting lymphocytes, implying a much higher pathophysiological relevance of the anaphylatoxin C3a as a proinflammatory mediator than previously thought. The response to this complement split product must be tightly regulated in situations with sustained complement activation to avoid deleterious effects caused by overactivated inflammatory cells. Receptor internalization, an important control mechanism described for G protein-coupled receptors, was investigated. Using rabbit polyclonal anti-serum directed against the C3aR second extracellular loop, a flow cytometry-based receptor internalization assay was developed. Within minutes of C3a addition to human granulocytes, C3aR almost completely disappeared from the cell surface. C3aR internalization could also be induced by PMA, an activator of protein kinase C. Similarly, monocytes, the human mast cell line HMC-1, and differentiated monocyte/macrophage-like U937-cells exhibited rapid agonist-dependent receptor internalization. Neither C5a nor FMLP stimulated any cross-internalization of the C3aR. On the contrary, costimulation of granulocytes with C5a, but not FMLP, drastically decreased C3aR internalization. This effect could be blocked by a C5aR-neutralizing mAb. HEK293-cells transfected with the C3aR, with or without Gα16, a pertussis toxin-resistant G protein α subunit required for C3aR signal transduction in these cells, did not exhibit agonist-dependent C3aR internalization. Additionally, preincubation with pertussis toxin had no effect on C3a-induced internalization on PMNs. C3aR internalization is a rapid negative control mechanism and is influenced by the C5aR pathway.
Regulation of IFN Consensus Sequence Binding Protein Expression in Murine Macrophages
Authors: Kantakamalakul Wannee , Politis Alexander D. , Marecki Sylvia , Sullivan Teri , Ozato Keiko , Fenton Matthew J. , Vogel Stefanie N.
Published: December 2022

Recent work has demonstrated that the transcription factor, IFN consensus sequence binding protein (ICSBP), plays a critical role in the capacity of mice to control infection with Toxoplasma gondii and Leishmania major, agents that require highly activated macrophages for their elimination. In this report the regulation of ICSBP mRNA and protein were analyzed in murine macrophages stimulated with LPS and/or IFN-γ. Like induction of leishmaniacidal activity, LPS and IFN-γ synergize to induce ICSBP mRNA and protein. Deletion analysis of the ICSBP promoter identified regions that were IFN-γ responsive, regions that mediate the ability of LPS and IFN-γ to activate this promoter synergistically, as well as regions that normally repress ICSBP transcription. Finally, exogenous expression of ICSBP, found in previous studies to down-regulate MHC I gene expression, failed to repress basal or IFN-γ-induced ICSBP transcription. This demonstrates that ICSBP can selectively suppress the expression of IFN-responsive genes. These findings extend in a significant way our understanding of the regulation of ICSBP by LPS and IFN-γ and provide important clues as to its role in macrophage activation.
Biased TCR Repertoire in HIV-1-Infected Patients Due to Clonal Expansion of HIV-1-Reverse Transcriptase-Specific CTL Clones
Authors: Kolowos W. , Schmitt M. , Herrman M. , Harrer E. , Löw P. , Kalden J. R. , Harrer T.
Published: December 2022

To study whether an expansion of HIV-1-specific CTL is contributing to the skewed TCR repertoire in HIV-1-infection, we characterized the TCR usage of CTL clones specific for a conserved epitope in HIV-1 reverse transcriptase (RT/476-484). CTL clones from three HIV-1-infected patients displayed highly similar TCR usage and used the identical Vβ6.1 and Vα2.5 gene segments. CTL clones from two patients showed a very high degree of similarity within the TCR complementarity-determining region-3 (CDR-3). In accordance with the similar molecular structure, all three CTL clones also exhibited a similar functional activity with regard to recognition of variant peptides and cytokine secretion pattern. In one subject clonal expansion of a single CTL specificity could be shown over a 10-mo period. TCR spectratyping of PBMC from two patients revealed a marked expansion of CDR-3 segments of a certain length within the Vβ6-family. Sequence analysis of these CDR-3 yielded sequences identical to the RT/476-484-specific CTL previously isolated from the same patients. This analysis demonstrates that clonal expansion of HIV-1-specific CTL is contributing to the skewed TCR repertoire in HIV-1-infected patients.
Characterization of Paired Ig-Like Receptors in Rats
Authors: Dennis Glynn , Stephan Robert P. , Kubagawa Hiromi , Cooper Max D.
Published: December 2022

To explore the phylogenetic history of the murine paired Ig-like receptors of activating (PIR-A) and inhibitory (PIR-B) types, we isolated PIR homologues from a rat splenocyte cDNA library. The rat (ra) PIR-A and raPIR-B cDNA sequences predict transmembrane proteins with six highly conserved extracellular Ig-like domains and distinctive membrane proximal, transmembrane, and cytoplasmic regions. The raPIR-B cytoplasmic region contains prototypic inhibitory motifs, whereas raPIR-A features a charged transmembrane region and a short cytoplasmic tail. Southern blot analysis predicts the presence of multiple Pira genes and a single Pirb gene in the rat genome. Although raPIR-A and raPIR-B are coordinately expressed by myeloid cells, analysis of mRNA detected unpaired expression of raPIR-A by B cells and raPIR-B by NK cells. Collectively, these findings indicate that the structural hallmarks of the Pir gene family are conserved in rats and mice, yet suggest divergence of PIR regulatory elements during rodent speciation.
T Suppressor Lymphocytes Inhibit NF-κB-Mediated Transcription of CD86 Gene in APC
Authors: Li Jianfeng , Liu Zhuoru , Jiang Shuiping , Cortesini Raffaello , Lederman Seth , Suciu-Foca Nicole
Published: December 2022

CD8+CD28− human T suppressor cells (Ts) act on APC, inhibiting their ability to elicit Th activation and proliferation. This effect is due to inhibition of the CD40 pathway which normally leads to CD80 and CD86 up-regulation. To determine whether Ts inhibit expression of B7 molecules by blocking transcription, we cloned and characterized the CD86 promoter. Mutational analysis revealed that Ts inhibit transcription driven by the CD86 promoter. The NF-κB binding site, at −612 of the CD86 promoter, is essential for Th-induced transcription. In cultures containing Th and Ts, Ts inhibit Th-induced NF-κB activation in APC. Together, these findings indicate that Ts inhibition of NF-κB activation in APC is a means by which they regulate the activation and proliferation of Th.
Th1 and Th2 Deviation of Myelin-Autoreactive T Cells by Altered Peptide Ligands Is Associated with Reciprocal Regulation of Lck, Fyn, and ZAP-70
Authors: Singh Rana A. K. , Zang Ying C. Q. , Shrivastava Anju , Hong Jian , Wang George T. , Li Sufang , Tejada-Simon Maria V. , Kozovska Milena , Rivera Victor M. , Zhang Jingwu Z.
Published: December 2022

Th0 clones recognizing an immunodominant peptide of myelin basic protein (residues 83–99) were derived from patients with multiple sclerosis. We demonstrate that analogue peptides with alanine substitution at Val86 and His88 had a unique partial agonistic property in inducing Th0 →Th1 and Th0 →Th2 deviation of the myelin basic protein-reactive T cell clones, respectively. Th0 to Th1 deviation induced by peptide 86V→A correlated with up-regulation of Fyn and ZAP-70 kinase activities. Conversely, Th0 to Th2 deviation induced by peptide 88H→A was associated with complete failure to activate Fyn and ZAP-70 kinases. The observed Th1 and Th2 shift also correlated, to a lesser extent, with Lck kinase activity that was down-regulated with Th1 deviation and increased with Th2 deviation in some T cell clones. We demonstrated that the Th1 and Th2 shift induced by the analogue peptides was a reversible process, as the T cell clones previously exposed to either 86V→A or 88H→A peptide could revert to an opposite phenotype when rechallenged reciprocally with a different analogue peptide. The study has important implications in our understanding of regulation of TCR-associated tyrosine kinases by altered peptide ligands and its role in cytokine regulation of autoreactive T cells.
Extracellular Signal-Related Kinase (ERK) and p38 Mitogen-Activated Protein (MAP) Kinases Differentially Regulate the Lipopolysaccharide-Mediated Induction of Inducible Nitric Oxide Synthase and IL-12 in Macrophages:LeishmaniaPhosphoglycans Subvert Macrophage IL-12 Production by Targeting ERK MAP Kinase
Authors: Feng Gui-Jie , Goodridge Helen S. , Harnett Margaret M. , Wei Xiao-Qing , Nikolaev Andrei V. , Higson Adrian P. , Liew Foo-Y.
Published: December 2022

Macrophage activation by cytokines or microbial products such as LPS results in the induction and release of several key immune effector molecules including NO and IL-12. These have been shown to play crucial roles in the development of immunity to intracellular pathogens such as Leishmania. The molecular mechanisms underlying the induction of these effector molecules are not fully understood. We now show that the extracellular signal-related kinase (ERK) and p38 mitogen-activated protein (MAP) kinases play differential roles in the regulation of LPS-stimulated inducible NO synthase and IL-12 gene expression. In macrophages, LPS stimulates the simultaneous activation of all three classes of MAP kinases, ERK, c-jun N-terminal kinase, and p38, albeit with differential activation kinetics. However, studies using inhibitors selective for ERK (PD98059) and p38 (SB203580) show that while p38 plays an essential role in the induction of inducible NO synthase, ERK MAP kinases play only a minor role in promoting NO generation. In contrast, while p38 promotes induction of IL-12 (p40) mRNA, ERK activation suppresses LPS-mediated IL-12 transcription. The biological relevance of these regulatory signals is demonstrated by our finding that Leishmania lipophosphoglycans, which promote parasite survival, act by stimulating ERK MAP kinase to inhibit macrophage IL-12 production. Thus, as ERK and p38 MAP kinases differentially regulate the induction of the macrophage effector molecules, inducible NO synthase and IL-12, these kinases are potential targets not only for the development of novel strategies to combat intracellular pathogens but also for therapeutic immunomodulation.
Role of Immunoregulatory Donor T Cells in Suppression of Graft-Versus-Host Disease Following Donor Leukocyte Infusion Therapy
Authors: Johnson Bryon D. , Becker Emily E. , LaBelle James L. , Truitt Robert L.
Published: December 2022

In murine models of allogeneic bone marrow transplantation (BMT), MHC-mismatched recipients given a delayed infusion of donor leukocytes (DLI) at 21 days posttransplant develop significant GVHD whereas MHC-matched recipients do not. The current study was initially designed to test the hypothesis that small numbers of T cells in the MHC-mismatched donor bone marrow (BM) graft exacerbated graft-vs-host disease (GVHD) when DLI was administered at 21 days after BMT. Ex vivo depletion of Thy1+ cells from the donor BM had no impact on the severity of GVHD after DLI. However, depletion of donor T cells in vivo with a Thy1 allele-specific mAb given after BMT resulted in significantly more severe GVHD after DLI. Similar results were obtained in a MHC-matched model of allogeneic BMT, indicating that this was a general phenomenon and not model dependent. These results indicated that a population of donor-derived Thy1+ cells suppressed graft-vs-host reactivity after DLI. Results of experiments with thymectomized recipients demonstrated that an intact thymus was required for generation of the immunoregulatory donor cells. Experiments using TCR β-chain knockout mice as BM donors indicated that the immunosuppressive Thy1+ cells coexpressed αβTCR heterodimers. Similar experiments with CD4 and CD8 knockout donor BM suggested that the immunoregulatory Thy1+αβTCR+ cells consisted of two subpopulations: a CD4+CD8− subpopulation and a CD4−CD8− subpopulation. Together, these results show that thymus-derived, Thy1+αβTCR+ donor cells generated early after allogeneic BMT suppress the graft-vs-host reactivity of T cells given as DLI. These cells may mediate dominant peripheral tolerance after allogeneic BMT.
IFN-γ Production and Cytotoxicity of IL-2-Activated Murine NK Cells Are Differentially Regulated by MHC Class I Molecules
Authors: Kubota Akira , Lian Rebecca H. , Lohwasser Stefan , Salcedo Margarita , Takei Fumio
Published: December 2022

Activation of NK cells by target cells leads to cytotoxicity as well as production of various cytokines including IFN-γ. MHC class I molecules on target cells regulate NK cytotoxicity. However, little is known about the regulation of IFN-γ production by NK cells. We examined the production of IFN-γ in individual murine NK cells stimulated with tumor cell lines by flow cytometric analysis of intracellular IFN-γ. Among several tumor lines tested, the rat basophilic leukemia line RBL-1 induced particularly high level of IFN-γ production in IL-2-activated NK cells, whereas other lines, including the prototypic NK target YAC-1, induced very low or no IFN-γ production. Transfection of murine classical MHC class I molecules into RBL-1 cells substantially inhibited IFN-γ production. This inhibition of IFN-γ production by MHC class I was independent of Ly-49 or CD94/NKG2A expression on NK cells. These results indicate that some target cells directly stimulate IL-2-activated NK cells and induce IFN-γ production, but the requirements for the induction of IFN-γ production seem different from those for NK cytotoxicity. Furthermore, similar to NK cytotoxicity, induction of IFN-γ production is inhibited by MHC class I on stimulating cells. However, the MHC class I-specific receptors inhibiting IFN-γ production are different from those for NK cytotoxicity.
Tolerance to Maternal Immunoglobulins: Resilience of the Specific T Cell Repertoire in Spite of Long-Lasting Perturbations
Authors: Faure Mathias , Calbo Sébastien , Kanellopoulos Jean , Drapier Anne-Marie , Cazenave Pierre-André , Rueff-Juy Dominique
Published: December 2022

T cell tolerance is established and maintained through various mechanisms, the critical component being the persistence of the specific Ag. However, at the molecular level, the nature of the recovering TCR repertoire following breakdown of tolerance is unknown. We address this important question by following κ light chain constant region (Cκ)-specific CD4+ T cells of κ light chain knock-out (κ−/−) mice born to κ+/− mothers. These cells, which were in contact with maternal κ+ Igs from early ontogeny until weaning, were strongly tolerized. Tolerance was reversible and waned with the disappearance of peptide Cκ134–148 presentation in lymphoid organs, including the thymus. Whereas three specific Vβ-Jβ rearrangements emerged in the peptide Cκ134–148-specific CD4+ T cell response of all regular κ−/− mice, soon after breakdown of tolerance only one of these rearrangements was detected. The two others displayed a significant delay in reappearance and were still rare at 26 wk of age, while the control proliferative response had already recovered 3 mo earlier. At 52 wk of age, a complete recovery of the three canonical Vβ-Jβ rearrangements was observed. Thus, although profoundly perturbed for several months, the T cell repertoire returns to equilibrium, highlighting the resilient nature of this system.
Robust B Cell Immunity but Impaired T Cell Proliferation in the Absence of CD134 (OX40)
Authors: Pippig Susanne D. , Peña-Rossi Claudia , Long James , Godfrey Wayne R. , Fowell Deborah J. , Reiner Steven L. , Birkeland Marian L. , Locksley Richard M. , Barclay A. Neil , Killeen Nigel
Published: December 2022

CD134 (OX40) is a member of the TNF receptor family that is expressed on activated T lymphocytes. T cells from mice that lack expression of CD134 made strong responses to a range of challenges, but they showed impaired proliferation in response to direct stimulation through the TCR with monoclonal anti-CD3ε Ab. CD134-deficient mice controlled infection with Leishmania major, Nippostrongylus brasiliensis, and Theiler’s murine encephalomyelitis virus, and they made overtly normal Ab responses to a variety of antigens. Thus, CD134 is not essential for many T cell responses in vivo, nor is it required for the provision of help to B cells. Nonetheless, a subtle role in the regulation of T cell reactivity is suggested by the effect of CD134 deficiency on in vitro T cell responses.
Distinct Activities of p52/NF-κB Required for Proper Secondary Lymphoid Organ Microarchitecture: Functions Enhanced by Bcl-3
Authors: Poljak Ljiljana , Carlson Louise , Cunningham Kirk , Kosco-Vilbois Marie H. , Siebenlist Ulrich
Published: December 2022

Mice rendered deficient in p52, a subunit of NF-κB, or in Bcl-3, an IκB-related regulator that associates with p52 homodimers, share defects in the microarchitecture of secondary lymphoid organs. The mutant mice are impaired in formation of B cell follicles and are unable to form proper follicular dendritic cell (FDC) networks upon antigenic challenge. The defects in formation of B cell follicles may be attributed, at least in part, to impaired production of the B lymphocyte chemoattractant (BLC) chemokine, possibly a result of defective FDCs. The p52- and Bcl-3-deficient mice exhibit additional defects within the splenic marginal zone, including reduced numbers of metallophilic macrophages, reduced deposition of the laminin-β2 chain and impaired expression of a mucosal addressin marker on sinus-lining cells. Whereas p52-deficient mice are severely defective in all of these aspects, Bcl-3-deficient mice are only partially defective. We determined that FDCs or other non-hemopoietic cells that underlie FDCs are intrinsically impaired in p52-deficient mice. Adoptive transfers of wild-type bone marrow into p52-deficient mice failed to restore FDC networks or follicles. The transfers did restore metallophilic macrophages to the marginal zone, however. Together, the results suggest that p52 carries out functions essential for a proper splenic microarchitecture in both hemopoietic and non-hemopoietic cells and that Bcl-3 is important in enhancing these essential activities of p52.
Activation and Targeting of Immunoglobulin Switch Recombination by Activities Induced by EBV Infection
Authors: Li Ming-jie , Maizels Nancy
Published: December 2022

EBV is strongly associated with Burkitt’s lymphoma, a B cell malignancy. In certain types of Burkitt’s lymphoma, the c-myc gene has undergone translocation to the S regions associated with heavy chain switch recombination. It has not been established whether EBV infection induces recombination activities, which in turn promote translocation of c-myc, or whether translocation precedes viral infection and provides a growth advantage that is further enhanced by factors encoded or induced by the virus. To distinguish between these possibilities, we have compared the level of switch recombination activities in the EBV-negative lymphoma, BJAB, and in its EBV-infected derivative, BJAB-B1, in experiments that assayed recombination of an extrachromosomal switch substrate during transient transfection. We have found that BJAB-B1 and other EBV-positive B cell lines supported high levels of recombination of switch substrates, to produce junctions like those found in products of chromosomal switch recombination. In contrast, BJAB did not support comparable levels of switch substrate recombination. In EBV-positive B cell lines, the ability to support switch substrate recombination correlated with levels of LR1, a B cell-specific factor which is a transcriptional regulator of c-myc and which also appears to function in switch recombination. Our observations support the hypothesis that EBV infection can induce activities that affect switch recombination and thus contribute to the translocations of c-myc to the S regions that characterize certain classes of lymphomas.
Kupffer Cells fromSchistosoma mansoni-Infected Mice Participate in the Prompt Type 2 Differentiation of Hepatic T Cells in Response to Worm Antigens
Authors: Hayashi Nobuki , Matsui Kiyoshi , Tsutsui Hiroko , Osada Yoshio , Mohamed Raafat T. , Nakano Hiroki , Kashiwamura Shin-ichiro , Hyodo Yasuko , Takeda Kiyoshi , Akira Shizuo , Hada Toshikazu , Higashino Kazuya , Kojima Somei , Nakanishi3 Kenji
Published: December 2022

Infection with Schistosoma mansoni, a portal vein-residing helminth, is well known to generate life cycle-dependent, systemic immune responses in the host, type 1 deviation during the prepatent period, and type 2 polarization after oviposition. Here we investigated local immunological changes in the liver after infection. Unlike splenocytes, hepatic lymphocytes from infected mice during the prepatent period already produced a higher amount of IL-4 and a lesser amount of IFN-γ than those from uninfected mice. Hepatic lymphocytes, particularly conventional T cells, but not NK1.1+ T cells, promptly produced IL-4 in response to worm products, soluble worm Ag preparation (SWAP), whenever presented by Kupffer cells from infected mice. The hepatic lymphocytes that had been stimulated with SWAP presented by infected mice-derived Kupffer cells produced a huge amount of IL-4, IL-13, and IL-5 as well as little IFN-γ in response to immobilized anti-CD3 mAb. Kupffer cells from uninfected mice produced IL-6 and IL-10, but not IL-12 or IL-18, in response to SWAP stimulation and gained the potential to additionally produce IL-4 and IL-13 after the infection. These results suggested that prompt type 2 deviation in the liver after the infection might be due to the alteration of Kupffer cells that induces SWAP-mediated type 2-development of hepatic T cells.
NF-κB/Rel Transcription Factors: c-Rel Promotes Airway Hyperresponsiveness and Allergic Pulmonary Inflammation
Authors: Donovan Carolyn E. , Mark David A. , He Hong Zhen , Liou Hsiou-Chi , Kobzik Lester , Wang Yunsheng , De Sanctis George T. , Perkins David L. , Finn Patricia W.
Published: December 2022

The NF-κB/Rel family of transcription factors induces many genes involved in immune and inflammatory responses. Mice with germline deletions of individual NF-κB/Rel subunits have different phenotypes, suggesting that the NF-κB/Rel transcription factors have different functions. We tested whether c-Rel promotes allergic asthma using a murine model of allergen-induced pulmonary inflammation and airway hyperresponsiveness. Our investigation focused on c-Rel, which is expressed in lymphoid cells and is important for lymphocyte activation. In response to allergen sensitization and challenge, c-Rel-deficient mice did not develop increases in pulmonary inflammation, bronchoalveolar lavage fluid eosinophilia, or total serum IgE. c-Rel deficiency also prevented the induction of airway hyperresponsiveness. Allergen-treated wild-type mice had increased DNA binding to an NF-κB consensus site. Chemokine expression was altered in allergen-treated c-Rel-deficient mice. Monocyte chemoattractant protein-1, which is regulated by NF-κB, was decreased in allergen-treated c-Rel-deficient mice relative to wild-type controls. The increase in NF-κB/Rel transcription factors after allergen challenge in wild-type mice and the decrease in allergen reactivity found in c-Rel-deficient mice indicate that c-Rel promotes allergic inflammation. Alteration of pulmonary chemokine expression in c-Rel-deficient mice may inhibit allergen-induced pulmonary inflammation and airway hyperresponsiveness.
The Rat and Mouse Homologues of MASP-2 and MAp19, components of the Lectin Activation Pathway of Complement
Authors: Stover Cordula M. , Thiel Steffen , Lynch Nicholas J. , Schwaeble Wilhelm J.
Published: December 2022

Recently, we described two novel constituents of the multimolecular initiation complex of the mannan-binding lectin (MBL) pathway of complement activation, a serine protease of 76 kDa, termed MASP-2, and a MASP-2 related plasma protein of 19 kDa, termed MAp19. Upon activation of the MBL/MASPs/MAp19 complex, MASP-2 cleaves the fourth complement component C4, while the role of MAp19 within the MBL/MASP-1/MASP-2/MAp19 complex remains to be clarified. In humans, the mRNA species encoding MASP-2 (2.6 kb) and MAp19 (1.0 kb) arise by an alternative polyadenylation/splicing mechanism from a single structural MASP-2 gene. Here, we report the complete primary structures of the rat homologue of MASP-2 and of rat and mouse MAp19. We show that both MASP-2 and MAp19 are part of the rat MBL pathway activation complex and demonstrate their exclusively hepatic biosynthesis. Southern blot and PCR analyses of rat genomic DNA indicate that as in humans, rat MASP-2 and MAp19 are encoded by a single structural gene.
Persistent T Cell Anergy in Human Type 1 Diabetes
Authors: Dosch H.-Michael , Cheung Roy K. , Karges Wolfram , Pietropaolo Massimo , Becker Dorothy J.
Published: December 2022

An anergic phenotype has been observed in nonobese diabetic (NOD) mice and some autoreactive T cells from patients with type I diabetes. To better understand this phenomenon, we measured T cell proliferative responses to 10 diabetes-associated and up to 9 control Ags/peptides in 148 new diabetic children, 51 age- and MHC (DQ)-matched siblings (sibs), 31 patients with longstanding diabetes, and 40 healthy controls. Most (78–91%) patient and sib responses to glutamate decarboxylase of 65 kDa (GAD65), islet cell cytoplasmic autoantibody (ICA) 69, diabetes-associated T cell epitopes in ICA69 (Tep69), and heat shock protein (Hsp) 60 involved anergic T cells that required exogenous IL-2 to proliferate. Responses to proinsulin, IA-2 (and tetanus toxoid) required no IL-2 and generated sufficient cytokine to rescue anergic T cell responses. Most new patients (85%) had autoreactive T cells, three quarters targeting more than half of the diabetes Ags. Only 7.8% of the sibs and none of the controls had such multiple T cell autoreactivities, which thus characterize overt disease. Multiple anergic and nonanergic T cell autoreactivities were sustained during 2 yr follow-up after onset and in patients with longstanding (3–26 yr) diabetes. Activated patient T cells survived severe IL-2 deprivation, requiring 20–100 times less IL-2 than normal T cells to escape apoptosis. Diabetic T cell anergy thus persists for decades and is Ag and host specific but not related to disease course. Rescue by IL-2 from bystander T cells and high resistance to apoptosis may contribute to this persistence. These data explain some of the difficulties in the routine detection of disease-associated T cells, and they emphasize challenges for immunotherapy and islet transplantation.
IV. The Deterioration of Isoagglutinins in Stored Bovine Serum. The Reactivation of the Agglutinins by the Addition of Normal Cow Serum
Authors: Little Ralph B.
Published: December 2022

Discussion and Summary

In stored serum the isoagglutinins deteriorate rapidly. There is apparently no set rule for the rate of depletion for this varied with each individual serum tried. With the most powerful serum of Group I the agglutinative properties were inactivated after nine to sixteen days storage in the ice chest. After a serum had been stored only a few days the reaction generally was not equal to that with fresh sera.

When the agglutinative properties in stored sera were inactivated at refrigerator temperature these properties were again restored by the addition of fresh normal cow serum obtained from Subgroup IIIa.

From the results of the work presented on the reactivation of the agglutinins in both heated and stored sera the following explanation is offered tentatively. In a number of bloods the sera fail to agglutinate at different periods. Therefore, in these sera component A (thermostable substance present in heated sera) under natural conditions is depleted or altered. If concentration of component A had been unchanged, such sera should have agglutinated regularly. However, in such sera the heat labile substance is present in normal concentration for the blood always reactivates. The blood withdrawn from Group I shows that the serum component B is labile. After a serum has been heated at 60°C. for one hour or stored in the ice chest for varied periods agglutination can not be demonstrated. When mixed with an equal quantity of normal serum of Subgroup IIIa such sera again strongly agglutinate. Component A, on the other hand, under these same conditions is stable fora serum stored fourteen months in the refrigerator strongly agglutinated when an equal quantity of fresh Subgroup IIIa sera containing substance B was added.
On the Natural Immunity to Scarlet Fever of the Japanese and Chinese Residing in South Manchuria
Authors: Ando K. , Nishimura H. , Ozaki K.
Published: December 2022

Southern Manchuria, where the Chinese, as the native race and the Japanese, as immigrants, live under the same environment affords an appropriate field to study racial differences of natural immunity.

Experienced physicians here believe that the morbidity in scarlet fever of the Japanese is far greater than that of the Chinese. According to the investigation of antiscarlatinal committee of the Kwantung Government the morbidity of the Japanese residents is 3.06 per cent while that of the Chinese is only 0.12 per cent. These figures were obtained by dividing the number of scarlet fever cases occurring between 1921 and 1925 and treated by hospital and civil physicians attached to the S. M. R. Company by the number of the population living during the same period within the area under the jurisdiction of the Local Affairs Department of the S. M. R. Company. While these figures generally seem to be correct, the fact that there are some scarlet fever cases among the Chinese failing to make returns, should be taken into consideration.
Mechanisms of induction of paralysed dendritic cells with poor antigen presentation function following systemic inflammatory response syndrome (APP3P.103)
Authors: Villadangos Jose
Published: December 2022

Severe infections, trauma, burns or hemorrhage trigger SIRS, which is characterized by the release of immune-activating cytokines but is followed by weeks-long immunosuppression. Opportunistic infections post-SIRS are the main cause of mortality and morbidity in critical care patients. Immunosuppression post-SIRS may also contribute to poor vaccination outcomes in areas with high incidence of infection with malaria or other pathogens. We are characterizing the contribution of Dendritic Cell (DC) functional impairment (paralysis) to immunosuppresion in mice where SIRS is triggered by TLR ligands or by infection with viruses, bacteria or the malaria parasite. We show that SIRS changes the local environment where DC undergo final differentiation, leading to weeks-long formation of paralyzed DC with poor antigen presentation function. We have (i) characterized the mechanisms responsible for this defect; (ii) identified a unique genetic program induced in paralyzed DC; (iii) devised strategies to overcome the impaired antigen presenting function of paralyzed DC; (iv) discovered cytokines that induce local formation of paralyzed DC; and (v) blocked these cytokines with mAb to reduce paralysis. Our studies might lead to new therapies to overcome DC paralysis and restore immunocompetence post-SIRS, preventing secondary infections and death in intensive care patients. They may also lead to improved vaccination outcomes in areas with high incidence of pathogen infection.
Double-edged sword: self-acquiring Batf3-dependent dendritic cell required for anti-tumor immunity and graft rejection (APP3P.104)
Authors: Jakubzick Claudia , Atif Shaihk , Gibbings Sophie , Nelsen Michelle , Gill Ronald , Kedl Ross , Marrack Philippa
Published: December 2022

One of the major obstacles for the graft acceptance of tissue and organs is the mismatch of major and minor histocompatibility (H) antigens. Minor H antigens are peptides derived from polymorphic proteins and presented by antigen-presenting cells on major histocompatibility complex (MHC) molecules. Here we hypothesized that the self-acquiring, cross-presenting Batf3-dependent dendritic cell (DC) is responsible for minor-antigen graft rejection. Using three models of minor-antigen graft rejection: 1) male antigen 2) non-sex complex minor antigen and 3) skin organ transplant, we demonstrate that Batf3-dependent DCs are responsible for minor-antigen graft rejection. However, in the presence of a TLR7 ligand, the requirement for Batf3-dependent DCs is bypassed by shifting the functionality of Batf3-independent DCs to elicit minor-antigen graft rejection. Overall our study identifies the DC subtype responsible for minor-antigen graft rejection.
Transport of microspheres from the lung to the draining lymph nodes by inflammatory monocytes. (APP3P.106)
Authors: Samilo Dane , Xiong Huizhong , Carter Rebecca , Leiner Ingrid , Pamer Eric
Published: December 2022

Inflammatory monocytes (Ly6Chi CCR2hi) play a critical role in steering the course of responses against microbial infections of the lung. Lung monocytes can take up live fungi or bacteria and transport them to draining mediastinal lymph nodes (mLN). How microbes are taken up and trafficked by monocytes from the pulmonary site of infection is poorly understood. To study monocyte-mediated microbial trafficking, we developed a system to trace monocytes that carry particles from the lung to mLNs. Herein, we used mice expressing GFP or the diphtheria toxin receptor under the control of the CCR2 promoter to determine the impact of intratracheally instilled toll-like receptor (TLR) agonists on trafficking of with fluorescent, polystyrene microspheres from the lung to mLNs. This system recapitulates monocyte recruitment and trafficking seen during in vivo infections. Monocytes infiltrated the interstitial space in response to TLR2 agonists. And while neutrophils were the primary cell population associated with instilled microspheres in the lung, they did not deliver them independently to mLNs. Conversely, adoptively transferred monocytes successfully delivered microspheres to mLNs of CCR2+-cell-depleted mice. Our current system and findings empower us to further dissect the process of monocyte uptake of inhaled particles and antigen delivery to lymphocytes of mLNs, which may in turn inform novel vaccine strategies in an age of increasing emergence of antibiotic-resistant pathogens.
FasL-expressing B cells are major players in type 1 diabetes (BA6P.129)
Authors: Saxena Ankit , Xiao Zuoxiang , Yagita Hideo , Larman Ben , Donner Thomas , Hamad Abdel Rahim
Published: December 2022

B cells serve both pathogenic and regulatory roles in Type 1 diabetes (T1D). Whereas the regulatory role is mainly limited to IL-10-producing (IL-10pos) B cells, little is known about the characteristics of pathogenic B cells. Here we identified a novel population of FasLhiCD5+ B cells that is significantly increased in T1D as compared to non-diabetic autoantibody positive (Ab+) subjects. Both the FasLhi and IL-10pos sets are localized within the CD5+ B subpopulation. However, FasLhiCD5+ B cells were significantly expanded and IL-10posCD5+ B cells decreased in T1D as compared to Ab+ subjects. FasLhiCD5+ B cells do not produce IL-10 and are largely spared from activation-induced apoptosis, whereas IL-10posCD5+ B cells are prone to Fas-mediated apoptosis. Causally, using NOD mouse model, we identified FasL-expressing B cells as negative regulators of IL-10pos B cells and facilitators of islet infiltration by diabetogenic T cells. These results identify FasLhiCD5+ B cells as an elevated subpopulation in T1D subjects and initiator of the diabetogenic process in NOD mice.
The transcriptional regulator Bcl-3 impaires the function of regulatory T cells leading to the development of intestinal inflammation (BA6P.131)
Authors: Reissig Sonja , Nikolaev Alexei , Waisman Ari , Hoevelmeyer Nadine
Published: December 2022

The oncoprotein and transcription factor Bcl-3 (B cell leukemia-3) is a member of the IkB family of NF-κB inhibitors and was originally identified as a component of a chromosomal rearrangement in some B cell lymphomas. In contrast to other IkB proteins, Bcl-3 associates with p50 and p52 homodimers and is predominantly a nuclear protein involved in regulating nuclear NF-kB activity. To understand more about the effects of Bcl-3 on T cells, we have generated a new mouse model, where Bcl-3 is overexpressed specifically in T cells. We could show that mice overexpressing Bcl-3 solely in T cells developed spontaneous colitis accompanied by a massive cell infiltration into the colon. Interestingly, the TCRgd+ T cell population was dramatically increased in the colonic IEL population of the Bcl-3 overexpressing mice. By examing Tregs in these mice, we found that overexpression of Bcl-3 strongly impairs Treg cell function, which was accompanied by decreased expression of CD25, Foxp3 and the co-inhibitory molecule CTLA-4. Hence, the loss of suppression of Bcl-3 overexpressing Tregs might result in the development of severe spontaneous colitis. Our data emphasize a novel pathway in the maintenance of viability and suppressive function of Tregs in the gut that critically involves Bcl-3 as a negative regulator, qualifying this protein as an interesting pharmacological target for the treatment of IBD.
Specific variations in the protocol for the induction of collagen-induced arthritis in mice improve reproducibility of disease severity and incidence in an SPF facility (BA6P.136)
Authors: Strait Richard , Thornton Sherry
Published: December 2022

Collagen-induced arthritis (CIA) is a widely used model for studying inflammatory arthritis (IA). CIA incidence and severity (I&S) varies between labs. Factors effecting induction include; adjuvant, # of collagen type II (CII) immunizations (imm), use of LPS and environmental hygiene. A better understanding of the effects of variations in protocols/environment on the I&S of CIA would improve the quality use this model. Method: Mice in conventional (con) and specific-pathogen free (SPF) facilities were compared. Mice were imm with 2 doses CII in complete (CFA) or incomplete (IFA) Freund’s adjuvant. LPS and/or CII were given at specific intervals following the 2nd imm. IA was evaluted using established scoring systems. Result: Following CII/CFA imm, con-housed had higher I&S of IA than SPF-housed mice. For SPF-housed mice, LPS 3 d after the 2nd CII/CFA imm modestly increased IA I&S. Unlike LPS or CII alone, additional LPS+CII 14 and 28 d later resulted in severe IA in all mice. Substitution of IFA for CFA along with LPS/CII at 14 and 28 d allowed for development of moderately severe IA in 80% of mice. Conclusion: Specific variations in use of CII, CFA, IFA and LPS can induce a range of IA I&S in a SPF facility . Thus, distinct experimental settings may be designed for robust assessment of factors that either exacerbate or inhibit arthritis pathogenesis. These protocols provide a practical and humane benefit of reducing cost, time and mice necessary for experimental assessment.
TNFR2 regulates gut commensal microbiota tocontrol sex-biased spontaneous experimental autoimmune encephalomyelitis (BA6P.138)
Authors: McKarns Susan , Miller Patrick
Published: December 2022

Host-commensal interactions influence the development and plasticity of gut-associated T helper (Th) and T regulatory (Treg) cells. In mice, the gut commensal segmented filamentous bacteria (SFB) modulate TH17 and FoxP3+ Treg differentiation to enhance host protection against external pathogens. However, in the context of a susceptible genetic background, SFB has the potential to augment the development of autoimmunity, including multiple sclerosis (MS). We have previously demonstrated that greater than 90% of female, but few male, tumor necrosis alpha receptor type 2 (TNFR2)-deficient mice crossed onto the myelin oligodendrocyte glycoprotein peptide fragment 35-55 (MOG35-55)-specific T cell receptor (TCR) transgenic background rapidly develop spontaneous experimental autoimmune encephalomyelitis (EAE). We now report that oral administration of trimethoprim/sulfamethoxazole, an antibiotic which has broad spectrum of activity against a variety of bacteria prevents the development of spontaneous EAE in TNFR2-deficient 2D2 TCR Tg mice. We have further combined Illumina MiSeq sequencing and RT-PCR to demonstrate a sex-biased microbiome profile in TNFR2-deficient MOG35-55-specific TCR mice, including increased SFB expression in the females relative to their male cohorts. Collectively, these findings suggest a role for TNFR2 in regulating commensal microbiota in the context of myelin autoantigen to control autoreactive T cells and autoimmune demyelination.
The gut microbiota regulates K/BxN autoimmune arthritis independent of Th17 and IL-17 (BA6P.121)
Authors: Block Katharine , Huang Haochu
Published: December 2022

The bacterial community that colonizes mucosal surfaces helps shape the development and function of the immune system. The effect of the microbiota on autoimmunity has important implications, although the mechanisms are not well understood. The K/BxN mouse model of rheumatoid arthritis is dependent on the microbiota, and particularly on segmented filamentous bacteria (SFB), for the autoimmune phenotype. Because SFB potently induce Th17 cell differentiation, previous studies have suggested Th17 cells as the link between microbiota and arthritis. In this study we sought to test directly the functional significance of Th17 and IL-17 in arthritis development by genetic approaches. We utilized mice deficient for either transcription factor RORγt (which controls the Th17 program), or the cytokine IL-17A (the major cytokine of the IL-17 family) in both cell transfer and spontaneous disease models. Unexpectedly, mice without Th17 or IL-17 were not protected from disease. Germinal center B and follicular helper T cell (Tfh) populations were normal, as were levels of serum IgG against self antigen. We found no evidence for compensation in production of the related cytokine IL-17F in IL-17A deficient mice. However, antibiotic treatment prevented disease in both IL-17 deficient and sufficient K/BxN mice, and resulted in defective Tfh and germinal center B cell differentiation. We conclude that the gut microbiota promotes autoimmune development by mechanisms independent of Th17 or IL-17.
The role of susceptible HLA shared epitopes in rheumatoid arthritis: a proclivity for citrullinated and self peptides (BA7P.158)
Authors: Anderson Kirsten , Roark Christina , Portas Mary , Aubrey Michael , Freed Brian
Published: December 2022

Rheumatoid Arthritis (RA) is an autoimmune disorder most strongly linked to the HLA-DRB1 gene locus and associated with antibodies to joint proteins, of which many are citrullinated. Polymorphic positions in the peptide-binding groove of HLA-DRB1 correlate with RA susceptibility, suggesting that these residues mediate binding of arthritogenic peptides. We measured binding of type II collagen259-273 and native and citrullinated forms of vimentin66-78 and α-enolase11-25 to 88 HLA Class II alleles including susceptible and resistant DRB1 alleles. The most susceptible allele, DRB1*04:01, has a strong preference for citrullinated forms of vimentin66-78 and α-enolase11-25. Resistant alleles DRB1*04:02 and *08:01 had either no preference or preferred the native form of these peptides. Mutations at positions 67, 70, 71, 74 or 86 in DRB1*04:01 to the corresponding residues in DRB1*04:02 or *08:01 reduced citrullinated peptide specificity, by increased binding of the native form or decreased binding of the citrullinated. Using DRB1*04:01 restricted collagen259-273 specific T cell hybridomas, we tested the effect of the mutations on T cell response. A good correlation exists between collagen binding and the T cell response, with reduced binding also showing a reduced T cell response. However, while mutation at position 70 from a positively charged amino acid to negative still bound collagen259-273, the T cell response was eliminated.
Single-cell RNA sequencing reveals regulatory patterns of self-antigen expression by medullary thymic epithelial cells (BA8P.162)
Authors: Lonnberg Tapio , Miragaia Ricardo , Zhang Xiuwei
Published: December 2022

The crucial capability of T cells for discrimination between self and non-self peptides is based on rigorous negative selection of developing thymocytes by medullary thymic epithelial cells (mTECs). The mTECs purge autoreactive T cells by apparently stochastic expression of cell-type specific genes referred to as tissue-restricted antigens (TRAs). The expression of many TRAs is promoted by the autoimmune regulator (AIRE) protein, while others are induced by yet unknown factors. Despite its tremendous immunological significance, the regulation of TRA expression in mTECs is incompletely understood. Recently developed single-cell RNA-sequencing enables systematic dissection of such genome-wide patterns for the first time. Here we present analysis of 152 murine mTEC transcriptomes, revealing a regulatory hierarchy within the highly diverse population of TRA genes. We find that TRAs dependent on AIRE exhibit distinct transcriptional dynamics, with high degree of genomic clustering and characteristic transcriptional burst profiles. We show that expression of other, AIRE-independent TRAs, is likely to be secured by regulatory pathways promoting differentiation of the mTEC cells from the TEC progenitors. The TRA repertoire is further shaped by the maturation of the mTECs to MHCIIhigh state. We further show that certain subsets of TRAs are neglected by the mTEC population, and that mTEC-driven negative selection of thymocytes may have intrinsic leakiness.
Expression and function of IL-1R complex governing human Th17 responses (BA8P.167)
Authors: Kim Dong Hyun , Cho Sunjung , Chung Yeon-Ho , Lee Won-Woo
Published: December 2022

The IL-1 receptor signaling is essential for differentiation, commitment, and maintenance of Th17 cells by induction of RORγt, a Th17 lineage-specific transcriptional factor. Due to its immunological significance, IL-1 receptor signaling is tightly regulated by expression of a functional IL-1 receptor type I (IL-1RI), a decoy IL-1RII, and IL-1R antagonist. However, underlying mechanism for their expressions still remains to be clarified. We aimed to investigate how the expression of IL-1RI and II is modulated by CD4 T cells and how their expression affects human Th17 responses. IL-1RI expression was identified by a part of CD4 T cells, whereas no CD4 T cells in resting state express IL-1RII. Upon TCR stimulation, IL-1RII was however induced by CD4 T cells but the underlying mechanism seemed to be different in naive and memory subsets. In naive CD4 T cells, IL-6 was a critical cytokine for synergizing TCR-induced IL-1RII expression. However, its expression in CD4 memory T cells was largely dependent upon TCR signaling and cytokine effect was minimal. Of note, the IL-1RI and II expressions were higher in Foxp3+ CD4 T cells in response to TCR stimulation as compared with Foxp3- T cells, suggesting a possible role of IL-1R signaling in newly identified IL-17-producing Foxp3+ T cells. Our findings suggest that the regulatory mechanism for IL-1RI and IL-1RII expression is important for fine-tuning IL-1 signal and Th17 responses thereby affecting the pathogenesis of autoimmunity.