bluuebunny/crossref_metadata_2025_split huggingface dataset:
Regulation of B Lymphocyte Development by Extracellular ST6Gal-I Sialyltransferase
Authors: Irons Eric Edward , Lau Joseph 
Published: January 2023

Abstract
Post-translational glycosylation of membrane proteins is increasingly appreciated as a means by which cell signaling and transcriptional networks are regulated. Although canonical glycosylation occurs intracellularly, the presence of active glycosyltransferases in the blood has been recognized for some time. We have recently reported that these extracellular enzymes, long thought to be functionless, can in fact remodel glycans by the extrinsic pathway of glycosylation. This axis enables glycosyltransferases to systemically regulate immunologically-relevant processes such as leukocyte production and IgG sialylation. In this study, we report that B cell development is both intrinsically and extrinsically regulated by the α2,6-sialyltransferase ST6Gal-I. We observed that endogenous expression of ST6Gal-I within bone marrow and splenic B cells could not fully account for cell surface sialylation. Bone marrow transplantation experiments between wild-type and ST6Gal-1 KO mice revealed an unexpected contribution of extrinsic ST6Gal-I towards maintenance of type 1 transitional B cells, the target of negative selection in the periphery, whereas intrinsic ST6Gal-I was necessary for marginal zone B cell development. The deficit in transitional B cells upon loss of host ST6Gal-1 was not due to impaired migration from the bone marrow, but a developmental block between the immature and transitional stages. Extrinsic sialylation of immature B cells enhanced BAFF-dependent signaling, CD23 expression, and elevated serum anti-histone auto antibodies in vivo. Collectively, these results highlight a novel glycan-dependent axis of regulation of B lymphopoiesis by extrinsic ST6Gal-I, with implications for autoimmunity.
IL-27 induces autophagy during monocyte-to-macrophage differentiation through a novel LC3-independent pathway
Authors: Laverdure Sylvain , Wang Ziqiu , Nagashima Kunio , Lane H. Clifford , Imamichi Tomozumi 
Published: January 2023

Abstract
IL-27, a member of the IL-12 cytokine family composed of IL-12, IL-23 and IL-35, has previously been shown to be an antiviral cytokine inhibiting infection of several viruses, among which HIV-1, HIV-2, HSV, HCV, HBV and Influenza virus. We have already reported that IL-27 induces HIV-1 inhibition in macrophages and dendritic cells in an IFN-independent manner, and that the cytokine enhances potential reactive oxygen species induction from cells. In this study, we demonstrated for the first time that IL-27 triggers autophagy induction in human AB serum-induced macrophages, while other IL-12 cytokine family members did not have such effect on these cells; IL-27 was able to induce autophagy both in terminally differentiated macrophages, causing a 2 to 5-fold increase in autophagosome accumulation, as well as during monocyte-to-macrophage differentiation, where IL-27 effect was even stronger (generating a 20-fold increase in autophagosome formation). Furthermore, we also determined that two key autophagy proteins, mTOR and LC3, were not involved in this phenomenon: while we were able to assess autophagosome accumulation in IL-27-treated cells via electron microscopy and immunofluorescence, this was not linked to either mTOR dephosphorylation nor LC3 lipidation, implying that IL-27 induces autophagy through a novel non-canonical pathway. Finally, we established that blocking IL-27 signaling during macrophage differentiation using both Ruxoliotinib or Tofacitinib, respectively JAK1/2 and JAK3 inhibitors, successfully inhibited autophagy triggering together with STAT3 phosphorylation, which suggests that the JAK-STAT pathway is responsible for regulating IL-27-induced autophagy.
Identification of a conserved multi-lineage Il9 enhancer
Authors: Koh Byunghee , Qayum Amina Abdul , Fu Yongyao , Kaplan Mark H. 
Published: January 2023

Abstract
The immune system provides resistance to the myriad of pathogens in the environment, but can also respond inappropriately causing allergic inflammation and autoimmune disease. CD4+ T cells, which play a crucial role in adaptive immune system, can be divided into subsets based on their effector functions. T helper 9 (Th9) cells, derived by the IL-4/STAT6 and TGFβ signaling pathways, produce IL-9 as a hallmark cytokine. Through IL-9 production, Th9 cells protect against parasite infection but are also involved in allergic inflammation and autoimmune diseases. In this study, we demonstrate that a conserved noncoding sequence (CNS) located 25kb upstream of the Il9 transcription start site, termed Il9 CNS-25, is critical for regulating Il9 expression in Th cell subsets and IL-9 producing CD8+ T cells (Tc9). Th9 cells derived from Il9 CNS-25 mutant (Il9ΔCNS-25) mice produce significantly less IL-9. Il9 CNS-25 was bound by factors that promote Il9 gene expression including STATs, IRF4 and BATF, promoted chromatin modifications and transcription factor binding at the promoter, and mediated accessibility of the locus. Il9ΔCNS-25 mice showed attenuated airway inflammation compared to control mice. The Il9 CNS-25 region in mice is conserved with an IL9 CNS-18 region in the human genome. We deleted CNS-18 in primary human Th9 cells and observed diminished IL-9 production. Thus, we have demonstrated that the Il9 CNS-25/IL9 CNS-18 elements are respectively critical for Il9/IL9 gene expression in Th9 cells and other IL-9 producing T cell subsets.
Dual T cell receptor expression promotes agonist selection of regulatory T cells
Authors: Balakrishnan Amritha , Jama Burhan , Morris Gerald P. 
Published: January 2023

Abstract
Generation of CD4+FoxP3+ regulatory T cells (Treg) in the thymus is essential for maintenance of self-tolerance. Treg lineage commitment occurs via agonist selection, via TCR high-affinity interaction with self-peptide-MHC (pMHC) ligand. We have previously demonstrated that natural co-expression of dual TCRs, occurring in ~10% of T cells, results in increased ability to recognize self-pMHC ligands and promote positive selection. We hypothesized that dual TCR expression could similarly promote agonist Treg selection. Examination of Treg development in B6.TCRa+/− mice, heterozygous for disruptive mutation of TCRAC and unable to express dual TCRs, demonstrated decreased frequency of Tregs in the thymus (2.76+0.08% of CD4 single-positive thymocytes) compared to age-matched wild-type B6 mice (3.26+0.09%, P &lt; 0.001). To more precisely compare Treg agonist selection, we co-transferred 1:1 ratios of T cell-depleted B6.TCRa+/−.Thy1.1 and B6.Ly5.1 bone marrow to lethally-irradiated B6 recipients and monitored thymic selection. TCRa+/− cells were deficient in Treg selection compared to co-transferred wild-type cells; TCRa+/− cells produced 37.57+4.16% of donor-derived thymic Tregs (P = 0.003). This defect was specific for Treg selection, as TCRa+/− cells comprised 44.68+6.33% of double-positive thymocytes and 45.50+5.77% of CD4 single-positive thymocytes. Tregs generated from TCRa+/− or wild-type cells were phenotypically similar, with no differences in expression of FoxP3 or CD5. We propose that the difference in agonist selection of dual TCR and single TCR Tregs could have important consequences in the TCR repertoire and antigenic specificity of peripheral Tregs, impacting immunologic tolerance.
Expression of Ly6C/6G defines a novel Aire-dependent subset of medullary thymic epithelial cells
Authors: Matsumoto Mitsuru , Matsumoto Minoru , Nishijima Hitoshi , Morimoto Junko 
Published: January 2023

Abstract
Ly6 family proteins are mainly expressed by myeloid-cell lineages such as granulocytes (expressing Ly6G) and monocytes (expressing Ly6C). However, exact function of Ly6 family protein in each cell-type remains unknown. For example, although expression of Ly6C has been used to define functionally distinct subset of monocytes (Ly6C+ monocytes vs. Ly6C− monocytes), molecular basis for the expression of Ly6C and functional difference between the two populations are largely unknown.
Medullary thymic epithelial cells (mTECs) are composed of heterogeneous populations in terms of the spectrum of tissue-restricted antigens (TRAs) expression for ensuring the elimination of autoreactive T-cell repertoire. We have recently identified mTEC subset(s) that express Ly6C and Ly6G both recognized by monoclonal antibody Gr-1 (i.e., Ly6C/6G+ mTECs). Approximately 10% of total mTECs expressed Ly6C/6G and Ly6C/6G+ mTECs were preferentially localized at the cortico-medullary junction. We found that Ly6C/6G+ mTECs and Ly6C/6G− mTECs do not form distinct lineage(s). Instead, Ly6C/6G− mTECs are able to convert into Ly6C/6G+ mTECs and vice versa in a reaggregated thymic organ culture (RTOC). Interestingly, Aire-deficient mice do not possess Ly6C/6G+ mTECs.
RNA-seq analyses have revealed that Ly6C/6G+ mTECs express higher levels of chemokines, integrins and extracellular matrix proteins compared with those from Ly6C/6G− mTECs, suggesting that Ly6C/6G+ mTECs might constitute a unique tolerogenic subset among heterogenous mTECs.
Critical roles for microbiota-mediated regulation of Th17 responses in a maternal immune activation model of autism
Authors: Lukens John R. , Lammert Catherine R. , Frost Elizabeth L. , Bellinger Calli E. 
Published: January 2023

Abstract
Recent studies suggest that autism is often associated with altered microbiota composition and gastrointestinal inflammation. This work has led to growing speculation of a potential role for the microbiome in autism spectrum disorder (ASD). However, how microbiota diversity mechanistically influences the development of autistic phenotypes has not been studied in great detail to date. To formally investigate this, we evaluated how environmentally induced changes in gut microbiota landscape influence the incidence and severity of MIA-induced neurodevelopmental disorders. Herein, we report critical roles for prenatal microbiota composition in the development of behavioral abnormalities in a MIA mouse model of autism. Mechanistically, we show that microflora-dependent calibration of the immune response underlies the effects of the microbiome on inflammation-induced ASD-like phenotypes. Specifically, we find that neutralization of IL-17a ameliorates the ability of the microbiome to affect the development of autism-related behaviors. Our results identify the immune system as a link between gut microbiota and the brain in neurodevelopmental disorders, and suggest that targeting the microbiome and maternal immune responses during gestation may offer strategies to limit autism development in at-risk pregnancies.
Cardiac endothelial cells produce complement C3 in response to hypoxia
Authors: Zhang Ming , Li Xiang , Kahyaoglu Seran , Sironi Marina , Mantovani Alberto 
Published: January 2023

Abstract
Hypoxia takes places in life threatening conditions, such as heart attack. Endothelial cells are on the first line being affected when hypoxia occurs. Complements as innate immune factors have been reported to play an important role in re-oxygenation after hypoxia being corrected, but little is known the complement response in endothelia cells during hypoxia. Using a murine cardiac endothelial line H5V, we investigated this. Under hypoxia condition without serum (thus no exogenous complements provided), these cardiac endothelial cells produced complement C3 for a period time and died. However, when serum was supplemented during hypoxia, these cells did not produce endogenous C3 and maintained a reasonable survival rate. The results indicate that endogenous production of complement C3 is promoted by the hypoxia stress and linked to cell death of cardiac endothelial cells.
Early Inhibition of Fatty Acid Synthesis Reduces Generation of Memory Precursor Effector T cells in Chronic Infection
Authors: Ibitokou Samad , Dillon Brian E. , Sinha Mala , Szczesny Bartosz , Delgadillo Añahi , Abdelrahman Doaa Reda , Szabo Csaba , Abu-Elheiga Lutfi , Porter Craig , Tuvdendorj Demidmaa , Stephens Robin 
Published: January 2023

Abstract
Understanding the mechanisms of CD4 memory T cell (Tmem) differentiation in malaria is critical for vaccine development. However, the metabolic regulation of CD4 Tmem differentiation is not clear, particularly in persistent infections. In this study, we investigated the role of fatty acid synthesis (FAS) in Tmem development in Plasmodium chabaudi chronic mouse malaria infection. We show that T cell-specific deletion, and early pharmaceutical inhibition of acetyl coA carboxylase 1, the rate limiting step of FAS, inhibit generation of early memory precursor effector T cells (MPEC). To compare the role of FAS during early differentiation or survival of Tmem in chronic infection, a specific inhibitor of acetyl coA carboxylase 1, 5-(Tetradecyloxy)-2-furoic acid, was administered at different times postinfection. Strikingly, the number of Tmem was only reduced when FAS was inhibited during T cell priming, but not during the Tmem survival phase. FAS inhibition during priming increased effector T cells (Teff) proliferation, and strongly decreased peak parasitemia, which is consistent with improved Teff function. Conversely, MPEC were decreased, in a T cell-intrinsic manner, upon early FAS inhibition in chronic, but not acute, infection. Early cure of infection also increased mitochondrial volume in Tmem compared to Teff, supporting previous reports in acute infection. We demonstrate that the MPEC-specific effect was due to the higher fatty acid content and synthesis in MPEC compared to terminally differentiated Teff. In conclusion, FAS in CD4 T cells regulates the early divergence of Tmem from Teff in chronic infection.
Correlative Recurrent Expression of Predicted Elements (CREPE): A Novel Computational Approach to Predict LncRNA Function
Authors: Pyfrom Sarah , Koues Olivia , Kowalewski Rodney , Oltz Eugene M. , Payton Jacqueline 
Published: January 2023

Abstract
Long non-coding RNAs (lncRNAs) act as transcriptional regulators, scaffolds, and signaling modulators in development, immune response, and oncogenesis. Our -omics study of &gt;100 human Non-Hodgkin Lymphoma (NHL) and normal B cell samples revealed altered expression of lncRNAs in NHL. LncRNAs have not been characterized in NHL or B cells, and there are few guidelines for functional prediction. To address this gap, we developed a novel computational approach: Correlative Recurrent Expression of Predicted Elements (CREPE). This method calculates and tracks the following for each lncRNA: expression, cell-type specificity, subcellular localization, differential expression (eg, tumor/normal), correlation with neighboring gene transcripts (RNAseq); local enhancer and 3D genome interaction landscapes (ChIPseq, Hi-C, ChIA-PET). From this, CREPE calculates a rankable score to predict the likelihood of potential functions, including 1) transcriptional regulation via lncRNA transcript, 2) enhancer-associated regulation, and 3) scaffold/modulator cytoplasmic function. The likelihood score formula incorporates an if-then logic, correlative regression analysis, and statistical significance. Finally, CREPE enables heuristic ranking of lncRNAs by pathway analysis. CREPE identified lncRNAs that may regulate NHL oncogenes and modulate B-cell receptor signaling. CREPE analysis of data from The Cancer Genome Atlas and a CRISPRi lncRNA growth screen validate its ability to identify lncRNAs with oncogenic or growth-promoting potential in diverse studies. In summary, we developed a novel approach that fills an unmet need for predictive modeling of lncRNA function and identified lncRNAs likely to promote lymphomagenesis.
Type 2 diabetes mellitus induces TNFR1 mediated necroptotic cell death of mice alveolar macrophages infected with Mycobacterium tuberculosis
Authors: Radhakrishnan Rajesh Kumar , Tripathi Deepak , Thandi Ramya Sivangala , Paidipally Padmaja , Vankayalapati Ramakrishna 
Published: January 2023

Abstract
Previously, we developed a mouse model of type 2 diabetes mellitus (T2DM) using streptozotocin and nicotinamide and found that T2DM mice are susceptible to Mycobacterium tuberculosis (Mtb) infection. We also found that alveolar macrophages from T2DM mice were more permissive to Mtb growth ex vivo compared to non-diabetic controls. In the current study, we determined the defective mechanisms that make T2DM mice alveolar macrophages more susceptible to Mtb infection. Mtb infected alveolar macrophages from T2DM mice produced more TNF-α (973.8 ± 13.3 pg/ml vs. 614.6 ± 27.2 pg/ml, p&lt;0.003) and less apoptotic (4.7 ± 1.8% vs. 28.6 ± 2.3%, p&lt;0.001) compared to Mtb infected non-diabetic control mice. Mtb infected alveolar macrophages from T2DM mice expressed higher levels of TNFR1 (12.8 ± 0.5 vs. 2.09 ± 0.01, p&lt;0.001) and markers of necroptosis RIPK1 (11.5 ± 0.57 vs. 2.94 ± 0.20, p&lt;0.002), RIPK3 (16.1 ± 0.58 vs. 2.89 ± 0.2, p&lt;0.001) and MLKL (9.2 ± 0.56 vs. 1.58 ± 0.72, p&lt;0.02) compared to Mtb infected alveolar macrophages from non-diabetic control mice as determined by real-time PCR. This finding was again proved by Western blot and confocal microscopy. Anti-TNFR1 antibody treatment of alveolar macrophages from T2DM mice before or after Mtb infection reduced RIPK1, RIPK3 and MLKL expression as determined by RT-PCR. Our findings demonstrate that T2DM induces necroptosis of alveolar macrophages upon Mtb infection. Enhanced TNFR1 signaling in T2DM mice alveolar macrophages is responsible for enhanced necroptosis. We are also determining the subpopulations of T2DM mice alveolar macrophages that express higher levels of TNFR1 upon Mtb infection. Studies are underway to determine the in vivo relevance of our current findings to Mtb growth in T2DM mice.
Kupffer cells restricts Mycobacterium tuberculosis growth better than alveolar macrophages
Authors: Thandi Ramya Sivangala , Tripathi Deepak , Radhakrishnan Rajesh Kumar , Paidipally Padmaja , Vankayalapati Ramakrishna 
Published: January 2023

Abstract
Kupffer cells protect liver from bacterial infections. In the current study, we compared Mycobacterium tuberculosis (M.tb) growth and cytokine production by mice kupffer cells and alveolar macrophages. M.tb H37Rv infected kupffer cells and alveolar macrophages produced equal amounts of TNF-α, IL-6, IL-10 & IL-1β. In contrast, kupffer cells restricted M.tb growth better than alveolar macrophages (1 ± 0.346×106 CFU vs. 4 ± 0.916×106 CFU, p&lt;0.03). There is no significant difference in the apoptosis of M.tb infected kupffer cells and alveolar macrophages. In contrast, M.tb infected kupffer cells expressed significant higher amounts of autophagy molecules LC-3B, ATG-7, ATG-5 and Beclin-1 (p&lt;0.005, p&lt;0.002, p&lt;0.0032 & p&lt;0.004 respectively) compared to M.tb infected alveolar macrophages as determined by real-time PCR. This was confirmed by Western blot and confocal microscopy. Our results suggest autophagy is involved in better restriction of M.tb growth by kupffer cells. Prime PCR analysis for 35 intracelluar signaling molecules those are involved in autophagy as well as in cytoskeleton indicated that M.tb infected kupffer cells significantly express higher levels of VASP (Vasodilator-stimulated phosphoprotein), RhoA (Ras homolog gene family member A) and Arp3 (Actin–related protein 3) genes (two fold, p&lt;0.01, p&lt;0.02&p&lt;0.02 respectively) compared to M.tb infected alveolar macrophages. Studies are underway 1. To confirm whether autophagy is involved in better restriction of M.tb growth in kupffer cells. 2. Determine the role of cytoskeleton proteins those are involved in enhanced autophagy and M.tb growth inhibition in kupffer cells. 3. Determining in vivo relevance to our current findings using mouse model of M.tb infection.
Lung tissue resident memory T cells coordinate effector T cell dynamics during the protective recall response to influenza
Authors: Paik Daniel H. , Farber Donna L. 
Published: January 2023

Abstract
Influenza remains a serious public health challenge as the current vaccination strategy cannot protect against new strains that emerge every season. Tissue resident memory T cells (TRM) are a non-circulating memory subset that are generated in the lung after influenza infection or intranasal live vaccination, and these lung TRM have been shown to confer broad cross-strain protection. However, the mechanisms by which both CD4+ and CD8+ TRM mediate their protective response are not well defined. We investigated the tissue specific events during an active murine influenza response, including TRM-mediated protection and effector T cell trafficking. To specifically determine the role of TRM, we treated mice with the drug Fingolimod (FTY720), which sequesters circulating lymphocytes into secondary lymphoid organs, enriching the lung for tissue resident memory cells. We found that TRM immediately confer protection, with reduced viral titers evident as early as day 3 after infection. At later time points beginning at days 4–5 post infection, we determined that CD4+ and CD8+ TRM enhance the recruitment of influenza-specific effector T cells into the lung resident niche as well as CD4+ and CD8+ lymphocyte in situ proliferation as measured by BrdU incorporation. Furthermore, total lung RNA sequencing during active infection and FTY720 treatment reveal candidate TRM-associated chemokine pathways for tissue lymphocyte trafficking. These findings indicate that TRM uniquely influence the cellular dynamics of the lung resident niche to mediate cross-strain protection against influenza.
Fast and easy immunomagnetic isolation of untouched human naïve T cells in less than 15 minutes
Authors: Ewen Catherine , MacDonald Neil , de Jong Susan , Kokaji Andy I. , Woodside Steven M. , Eaves Allen C. , Thomas Terry E. 
Published: January 2023

Abstract
Peripheral blood circulating T cells can be subdivided into either antigen experienced effector and memory T cells or naïve T cells that have not yet encountered their cognate antigen. Naïve T cells are valuable tools when studying mechanisms of T cell activation and are required for foundational research into infectious diseases, cancer and transplantation.
We describe here a simple immunomagnetic cell isolation protocol for the isolation of untouched human naïve T cells from fresh or previously frozen PBMC or leukapheresis samples. The column-free, negative selection procedure involves labelling and removing unwanted cells using bispecific antibody complexes that crosslink cell surface antigens to magnetic particles. The labelling cocktail is composed of antibodies that target non-T cells, memory T cells, and an optional cocktail for depleting gamma delta T cells. Briefly, the procedure involves a five minute incubation with the antibody cocktail followed by the addition of magnetic particles and two three minute magnetic separations in a hand-held EasySep™ magnet. Following the second magnetic separation, the untouched naïve T cells are simply poured off into a new tube and are ready for use.
We define naïve T cells phenotypically as CD3+CD45RA+CD45RO-CD197+. Starting from fresh PBMC, we were able to obtain purities of 96.1 ± 2.3% (mean ± SD, n=14) and recoveries of 62.6% ± 24.5% (mean ± SD, n=14). Additionally, isolated naïve T cells are functional and respond by upregulating the expression of CD25 and CD69 upon stimulation. Our EasySep™ Human Naïve Pan T Cell Isolation Kit offers a fast and easy isolation method for naïve T cells that is suitable for downstream applications such as flow cytometry, cell culture or DNA/RNA extraction.