bluuebunny/crossref_metadata_2025_split huggingface dataset:
Depletion of Dead Cells from Primary Tissue in 6 Minutes
Authors: MacDonald Neil , Barr Christine , McQuenn Karina , Woodside Steven M. , Thomas Terry E. , Eaves Allen C. 
Published: January 2023

Abstract
Tissue specific research often requires mechanical and/or enzymatic digestion to isolate and study certain cell types. The digestion can be a harsh process resulting in a significant number of dead cells in the final cell suspension. Subsequent analysis by flow cytometry is difficult to interpret due to non-specific binding of antibodies to dead cells and dead cell auto fluorescence. Factors released by dead cells can also interfere with downstream assays, complicating the study of primary tissues.
During apoptosis, the cell membrane loses its phospholipid asymmetry resulting in exposure of negatively charged phospholipids on the cell surface. Relocation of phosphatidylserine (PS) to the outer leaflet of the cell membrane is a well-established marker of apoptosis. By targeting exposed PS with Annexin V we have developed a rapid method (EasySep™) to immunomagnetically remove dead cells from primary tissue samples.
Performance of this kit was examined on various mouse and human tissue types. Using this method, we were able to improve viability of a single cell suspension of mouse lungs digested with collagenase/hyaluronidase from an initial viability of 39.6 ± 12.3% AnxV−/PI− to 70.9 ± 11.8% AnxV−/PI. From 1×10^8 total start cells, 1.43 ± 0.68 ×10^7 live cells were recovered (n=10). From human polymorphonuclear leukocytes cultured overnight, viability was improved from 23.7 ± 9.8% AnxV−/PI− to 67.7 ± 12.1% AnxV−/PI− with recovery of 1.27 ± 0.52 ×10^7 live cells from 1×10^8 total start cells (n=7). This equates to removal of 88.9 ± 8.3% and 91.8 ± 5.3% dead cells respectively. Since live cells are untouched, subsequent isolation of desired cell types can be performed, resulting in a more viable population of cells for downstream applications.
Integrative analysis reveals CD38 as a therapeutic target for plasma cell-rich rheumatoid arthritis, pre-rheumatoid arthritis and systemic lupus erythematosus
Authors: Cole Suzanne , Walsh Alice , Yin Xuefeng , Wechalekar Mihir , Smith Malcolm , Proudman Susanna M. , Veale Douglas J. , Fearon Ursula , Pitzalis Constantino , Chiu Christopher , Sharp Michael , Alvarez John , Anderson Ian , Madakamutil Loui , Nagpal Sunil , Guo Yanxia 
Published: January 2023

Abstract
Autoantibodies play a significant role in the progression and pathogenesis of many autoimmune diseases such as rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). No studies have analyzed the expression of plasma cell/plasmablast genes during RA disease progression and the potential of an anti-CD38 antibody in depleting plasma cells and plasmablasts for efficacy in autoimmunity. Therefore, we interrogated the rationale of daratumumab, an anti-CD38 monoclonal antibody, as a potential therapeutic in RA and SLE. RNA-Seq analysis of synovial biopsies from various stages of RA disease development shows that plasma cell/plasmablast-related genes CD38, XBP1, IRF4, PRDM1, IGJ and TNFSF13B are significantly up-regulated in synovial biopsies from arthralgia, undifferentiated arthritis, early RA and established RA compared to healthy and osteoarthritis controls. In addition, flow cytometry analysis reveals highest CD38 expression on plasma cells and plasmablasts compared to natural killer cells, classical dendritic cell (DC), plasmacytoid DC, and T cells, in peripheral blood of healthy control, SLE and RA donors. We show that IGJ expression mRNA strongly correlates with the number of plasma cell/plasmablast in SLE patient PBMCs. Most importantly, IGJ mRNA down-regulation correlates with daratumumab-mediated depletion of these cells ex vivo. The data indicates a potential use of IGJ mRNA as a surrogate pharmacodynamic biomarker in clinical testing of daratumumab. Taken together, our data provides rationale for daratumumab in the treatment of RA and SLE.
Novel targeted therapies for African-American and European-American SLE patients identified from E-Genes elucidated by transancestral SNP mapping
Authors: Aidukaitis Bryce N. , Labonte Adam C. , Catalina Michelle D. , Bachali Prathyusha , Rouffa Sean , Ainsworth Hannah C. , Marion Miranda C. , Howard Timothy D. , Langefeld Carl D. , Lipsky Peter E. , Grammer Amrie C. 
Published: January 2023

Abstract
SLE is more prevalent and severe in African-American (AA) compared with European-American (EA) populations but standard-of-care drugs vary in effectiveness between these groups. To gain insight, drugs specific for E-gene containing pathways were investigated using the Immunochip transancestral SNP mapping and a curated database of SLE DE genes. SNPs proxy to SLE-associated SNPs were compared with known eQTLs contained in the GTEx database. Since GTEx contains genotype data as well as gene expression data from cell lines, tissues and whole blood from healthy individuals, an independent query of GTEx for expression of RNAs abnormally expressed in SLE patients compared to healthy individuals was carried out. The results of the eQTL and DE gene queries of GTEx were combined and eQTLs and associated E-Genes were pooled by ancestry. Drugs targeting E-genes/pathways shared by EA and AA include ibrutinib, ruxolitinib and ustekinumab. Predicted EA-specific drugs include antimalarials and cyclosporine, as well as drugs-in-development targeting CD40, CXCR1 and CXCR2; AA-specific drugs include retinoids and HDAC inhibitors. This analysis of the distinct genetic profiles of EA and AA populations indicates that unique sets of drugs may be particularly effective at treating lupus in each ancestral group.
A Low molecular weight BAFF receptor antagonist inhibits differentiation of human peripheral B cells into plasma blasts/plasma cells in vitro.
Authors: Yoshimoto Keiko , Seki Noriyasu , Suzuki Katsuya , Sugahara Kunio , Takeuchi Tsutomu 
Published: January 2023

Abstract

Backgrounds and Purpose
We have demonstrated that the elevated expression of BAFF receptor (BR3) on monocytes is involved in the overproduction of IgG by B cells in patients with primary Sjögren’s syndrome (pSS) which is often accompanied with hypergammaglobulinemia. We discovered a low molecular weight compound, BIK13, which inhibits binding of BAFF to BR3, by our original high-throughput screening system. We found that BIK-13 suppressed IL-6 production by BAFF-stimulated pSS monocytes. In this study, we investigated inhibitory effects of BIK-13 on B cell functions.

Methods
PBMCs were cultured with a B cell-stimulants cocktail, such as anti-IgM and CD40 antibodies, human IL-21 and human BAFF, in the presence of BIK-13. B cell-differentiation was monitored through analysis of the expression levels of CD19/CD38/IgD and activation-induced cytidine deaminase (AID) by FACS and qPCR, respectively. The amounts of IL-6 and IgG produced in vitro by the cells were measured by ELISA.

Results
Increased production of IgG and IL-6 by PBMC upon stimulation with B cell-stimulants was suppressed by BIK13 in a dose dependent manner. FACS analysis indicated that differentiation of B cells into plasma blasts and/or plasma cells was inhibited by BIK-13. In addition, the expression level of AID was also suppressed by the compound, suggesting that the IgG class switching was impaired.

Conclusion
These data collectively suggest that BIK-13 suppresses the B cell functions and may provide a novel therapeutic possibility to treat autoimmune diseases such as pSS.
Neutrophils contribute to T cell activation via reactive oxygen species production in HIV infection
Authors: Dunsmore Garett John , Shahbaz Shima , Houston Stan , Elahi Shokrollah 
Published: January 2023

Abstract
Neutrophils are the most abundant leukocytes in the human immune system contributing to 50–80% of all the white blood cells in the peripheral blood. Interestingly, the role of neutrophils in HIV infection has been poorly investigated. Previous studies have demonstrated that during HIV infection neutrophils are capable of both activating T cells and suppressing T cells. Our preliminary findings suggest that T cell activation could be modulated by neutrophils through various mechanisms. Galectin-9 has been demonstrated to play an integral role in HIV pathogenesis by suppressing CD8+ T cell function through the engagement with TIM-3. Interestingly, we have observed neutrophils have high expression of galectin-9, which is reduced after prolonged activation. In confirmation with previous studies demonstrating that HIV infected patients have hyperimmune activation, galectin-9 expression is decreased on neutrophils from HIV infected individuals, while the expression of the neutrophil activation marker CD32 is increased. Additionally, galectin-9 expression on neutrophils could be involved in suppressing T cell responses via. galectin-9:TIM-3 engagement.
We found that neutrophils from HIV patients enhance T cells activation when co-cultured, whereas this was not the case for healthy controls. Interestingly, these effects could be reversed by inhibiting neutrophil reactive oxygen species (ROS) production. These findings suggest that HIV infection results in hyperimmune activation of neutrophils contributing to T cell activation.
The surprising role of Indoleamine 2,3-dioxygenase (IDO) in supporting the survival of bone marrow resident long lived plasma cells (LLPC)
Authors: Lightman Shivana M , Carlson Louise M , Lee Kelvin P 
Published: January 2023

Abstract
Long lived plasma cells (LLPC) are a key component of protective humoral immunity. The ability of these cells to maintain longevity and a durable antibody response is largely dependent on the complex nature of the bone marrow (BM) microenvironment in which these cells reside. Our lab has demonstrated that the co-stimulatory receptor, CD28, is essential for the survival and function of LLPC. Furthermore, when CD28 on LLPC ligates with CD80/CD86 expressed on DC, it induces upregulation of the enzyme indoleamine 2,3-dioxygenase (IDO), which catabolizes tryptophan (Trp) into kynurenine (kyn), a well-known ligand of the aryl hydrocarbon receptor (AhR). AhR is a nuclear transcription factor that can modulate immune responses and is highly expressed in plasma cells (PC). Our preliminary data reveals that IDO knock-out (KO) mice in comparison to wild type (Wt) mice have a decrease in bone marrow resident PCs and a decrease in overall Ig levels. Additionally, 15 weeks post vaccination IDO KO mice have significantly less antigen-specific long lived antibody titers compared to Wt. It was also observed that interaction with DC both in vitro and in vivo is needed for LLPC survival and function. Moreover, AhR activation by kyn causes an increase in CD28 mRNA in LLPC. This leads us to propose a model where CD28 interaction with CD80/CD86 induces IDO production, which through its metabolite kyn activates AhR which can then regulate CD28 expression- thus creating a mechanism that promotes LLPC survival and function. These findings challenge the traditional idea of IDO as an inhibitory molecule and provide the rationale for investigating its essential role in maintaining LLPC survival and sustained antibody responses.
Infant T cells exhibit increased TCR signaling and proliferation to respiratory infection
Authors: Thapa Puspa , Guyer Rebecca S , Farber Donna L 
Published: January 2023

Abstract
Infants are highly susceptible to respiratory infections and frequently experience recurrent episodes. Our previous study demonstrated that tissue resident memory T cells (TRM), which are critical in mediating long-term protection against pathogens, are reduced in infant mice compared to adult mice after influenza infection. However, the underlying mechanisms for reduced TRM generation in infants is still unknown. Our preliminary data show that infant T cells have enhanced TCR sensitivity leading to increased activation compared to adult T cells and thus are biased towards effector differentiation at the expense of memory formation. We show that infant OVA-specific (OTII) T cells exhibit lower T cell activation threshold for proliferation and enhanced sensitivity for low does of antigen. Infant T cells express higher levels of CD69 and CD25 with a greater induction of Nur77, a direct indicator of TCR signal strength, at peptide doses that fail to fully activate adult T cells. Co-transfer of infant and adult OTII T cells into a congenic host resulted in infant T cells dominating the effector response in the lungs (at a ratio of 3:1) after primary influenza challenge. At the peak of infection, infant effector T cells also expressed lower levels of TCF1 and Bcl-2, indicating transcriptional bias for terminal effector differentiation with reduced survival. Moreover, infant derived effector and memory T cells express lower levels of CD5, suggesting enhanced survival of lower affinity T cells after influenza infection. Taken together, these results show that enhanced sensitivity for antigenic stimulation promotes cell intrinsic mechanisms that govern decreased potential of infants to form durable and functional lung TRM.
Tumor necrosis factor receptor associated factor 3 (TRAF3) regulates B cell antigen receptor (BCR) signaling
Authors: Whillock Amy Louise , Bishop Gail A 
Published: January 2023

Abstract
Tumor necrosis factor receptor associated factor 3 (TRAF3) is an intracellular adaptor protein that plays multiple cell type- and context-dependent roles. In B cells, TRAF3 negatively regulates signaling by several receptors important to survival and activation, including the B cell activating factor receptor (BAFF-R), CD40, toll-like receptors and the IL-6 receptor. Deletions or inactivating mutations of TRAF3 are common in the human B cell malignancies multiple myeloma and B cell lymphoma (BCL). Enhanced homeostatic B cell survival in B cell-specific TRAF3-deficient mice (B-Traf3−/−) causes enlarged secondary lymphoid organs, spontaneous germinal centers, and autoantibody production. Highlighting the role TRAF3 plays as a B cell tumor suppressor, B-Traf3−/− mice develop BCL at an increased rate as they age.
The B cell antigen receptor (BCR) is crucial for B cell antigen recognition, survival, proliferation, and antibody production. Defects in BCR signaling can lead to malignancy, and many BCLs are dependent upon BCR signaling for survival. TRAF3 regulates the signaling of several pathways that interact with the BCR pathway, but the role of TRAF3 in regulating BCR signals is unknown, a knowledge gap we are currently addressing. Our initial studies show that TRAF3 associates with CD79b of the BCR complex. Phosphorylation of Syk, a proximal BCR signaling target, is increased after BCR stimulation in the absence of TRAF3, with increased downstream phosphorylation and activation of Btk, PLCγ2, Akt, Erk, and canonical NF-κB. This suggests that TRAF3 plays an important role as a regulator of BCR signaling. Ongoing studies are examining the mechanisms of this regulation.
A screening platform for human plasma cells (PCs) reveals a selective response to SYK inhibition
Authors: Campbell Michelle , Stephenson Sophie J , Whale Kevin , Rapecki Stephen , Finney Helene , Doody Gina M , Tooze Reuben M 
Published: January 2023

Abstract
Therapeutic targeting of PC populations has potential value in various auto-immune, allergic and neoplastic cell states, but at present few options exist for selective targeting. However, the unique physiology and niche dependence of PCs suggests that such targeting may be achievable. To this end we have developed a sensitive screening platform capable of detecting changes in PC biology. Starting from our published methods for in vitro PC generation we optimised conditions to allow maintenance of PCs in polarised niche environments reflective of homeostatic and inflammatory conditions in microtitre plates. Using this assay format we probed PC biology via small molecule inhibitors (SMIs) targeting BCL2 family members, proteasome, and kinase signalling pathways. We were able to identify SMIs with broad and selective toxicity, as well as SMIs with no effect on PCs. Under all conditions analysed we found consistent divergence in the impact of targeting aspects of B cell receptor (BCR)-associated signalling pathways. Targeting SYK via two distinct SMIs consistently resulted in selective killing of subsets of PCs. By contrast BTK inhibition had no significant effect on PC viability. Whilst the importance of SYK signalling has been well documented at the B cell stage its importance to PCs has to our knowledge not been extensively documented. Retention of functional BCRs has been recently described in BM IgA+ and IgM+ PCs, and examination of expression data indicates selective retention of components of the BCR signalling complex in our assay system. The data presented here demonstrate the potential utility of an in vitro screening platform to evaluate the selective dependency of subsets of PCs on therapeutically targetable pathways.
CD209 activation promotes survival of lymphoblastic human B cells
Authors: Valentin Anibal J , Díaz Glorianne Y 
Published: January 2023

Abstract
The role of B cells in host protection and antibody production depends in part on their ability to recognize antigens and pathogen associated molecular patterns (e.g. carbohydrates and nucleic acids) mostly through membrane bound receptors. CD209 (i.e. DC-SIGN) is a C-type lectin receptor expressed on dendritic cells, macrophages and B cells which binds to different oligosaccharides and plays a role in antigen capture and cell adhesion. CD209 also functions as a receptor for many bacterial, fungal and viral pathogens. In B cells, CD209 expression enhances infections by HHV8 and H5N1 viruses and in HIV positive patients, promotes trans-infection of CD4+ T cells. Besides its role in cell adhesion and pathogen recognition, the role of CD209 in B cell physiology remains elusive. Our goal is to identify and characterize CD209-induced signaling pathways in human B cells and determine how these pathways relate to normal and pathologic B cell responses. Our data shows that CD209+ lymphoblastic B cells have a higher level of basal cell death suggesting that expression sensitizes B cells to apoptosis. Stimulating CD209+ B cells using an anti-CD209 antibody or Mannan, promotes cell survival. Moreover, CD209 stimulation inhibits BCR-mediated programmed cell death and modifies the expression of several genes involved in cell survival and apoptosis (e.g. Bim, BclXL, Xiap). Elucidating the role of CD209 in B cell physiology will have important implications towards understanding the development of the humoral immune response to pathogens such as HHV8, Zika, Dengue and HIV. Moreover, our results will set the groundwork for future studies on vaccine development and biomarker discovery. Supported by NIGMS-INBRE P20 GM103475-14
IL-33 induces a metabolic programming supporting dendritic cells tolerogenicity
Authors: Velayutham Murugesan , Roessing Anna S , Shiva Sruti , Turnquist Heth R 
Published: January 2023

Abstract

Objective
Delivery of the stromal cell-derived cytokine IL-33 protects allografts from rejection by acting on CD11c+ DC that facilitate the expansion of regulatory T cells (Treg). IL-33 administration also permits tolerance induction at doses of co-stimulatory blockade that fails on their own. Based on these data we hypothesized that IL-33-stimulated DC represent a tolerogenic DC subset and their precise examination will define biochemical pathways that contribute to tolerogenic DC functions.

Methods
Mouse bone marrow-derived CD11c+ DCs were treated with lipopolysaccharide (LPS) or IL-33 before nitric oxide (NO) generation, glucose and fatty acid uptake, and mitochondrial function and potential were assessed by EPR spectroscopy, flow cytometry, and Seahorse analysis.

Results
DC responded to the TLR4 ligand LPS by generating high levels of NO and shifting their metabolic activity to aerobic glycolysis. This was in stark contrast to IL-33, which caused a metabolic reprogramming in DC that involved augmented mitochondrial basal respiration, ATP production, and maximum respiration. IL-33-stimulated DC also displayed increase mitochondrial potential and a profound increase in the uptake of fatty acids.

Conclusions
It has emerged that immunogenic DCs undergo metabolic changes that enables their pro-inflammatory functions. The metabolic underpinnings of tolerogenic DCs is less understood. Here we show that IL-33-stimulated DC, which potently expand Treg, rely on oxidative phosphorylation and uptake fatty acids. These data suggest IL-33 initiates metabolic reprogramming matching those found in regulatory myeloid populations, including tumor associated DC and alternatively activated macrophages.
Investigation on the role of temperature in macrophage responses
Authors: Hassan Intisar Abdullah , Durdik Jeannine M , Satyajit Rath , Renfro Zachary , Blake Harrison 
Published: January 2023

Abstract
This study investigated temperature-associated changes in macrophage activation and metabolism across species. Macrophages were isolated from C57BL/6J mice, broiler chicken spleens, and rainbow trout head kidneys, treated with lipopolysaccharide (LPS) and nitric oxide (NO) responses measured. Mouse macrophage NO production was similar at 37°C (normal body temp.) and 35°C. However at 33°C and 28°C responses dropped notably. At 39°C, Mouse bone marrow-derived macrophages (BMDM) and mouse cell line, RAW264.7, showed higher NO production versus 37°C. In chicken splenic macrophages (CSM) LPS-induced NO responses were similar at 41°C (normal for avian) 37°C, and 39°C. A mild fever of 42°C had a large stimulatory effect on NO production compared to 41°C. During infection, trout induce fever by swimming to warmer water. We found NO responses in LPS-treated head kidney macrophages to be higher at 15, 28, and 37°C than at 4°C. Total protein synthesis was evaluated via fluorescence. Without LPS stimulation, peritoneal resident mouse macrophages (PRM) from young mice showed similar protein synthesis at 37°C and 39°C. In both young and aged LPS-stimulated PRM, significantly higher protein synthesis was observed at 39°C compared to 37°C, though the increase was smaller in aged PRM. However, this effect was not uniform across all tissues. LPS stimulated young (BMDM) showed an increased protein synthesis at 37°C but cells from aged did not. A further increase in LPS stimulation was observed at 39°C. Consistent with fever increasing protein synthesis, LPS-stimulated CSM and chicken cell line HTC, showed higher protein synthesis at 42°C compared with 41°C.