bluuebunny/crossref_metadata_2025_split huggingface dataset:
Age-associated loss of lamin-B1 in thymic epithelial cells disrupts the thymic microenvironment for thymopoiesis and leads to thymic involution
Authors: Yue Sibiao 
Published: January 2023

Abstract
Lamins are abundant type V intermediate filaments important for the structure and function of the nucleus. By studying the Drosophila immune organ, the fat body, recent work in our lab has uncovered a mechanism by which lamin-B loss in the fat body upon aging induces age-associated systemic inflammation and gut hyperplasia, suggesting that lamin-B loss triggers a phenomenon known as immunosenescence. A role of B-type lamins in mammalian immune aging, however, remains unexplored. Here, we report an age-related reduction of lamin-B1 inthymic epithelial cells (TECs) in the thymus, the primary immune organ for T-cell generation. We further demonstrate that genetic ablation of lmnb1 in TECs causes remarkably similar phenotypes as these observed in aged thymus. Furthermore, RNA-seq analyses reveal that depletion of lmnb1 accelerates age-related transcriptional changes in TECs, suggesting that loss of lamin-B1 in TECs upon aging may lead to global transcriptome alterations and trigger a degenerative cascade in the thymus. This work provides new insights into the cause and consequence of immunosenescence during mammalian immune aging.
Psmb11 molds the transcriptome of cortical TECs and CD8 thymocytes
Authors: Apavaloaei Anca , Rouette Alexandre , Brochu Sylvie , Perreault Claude 
Published: January 2023

Abstract
Psmb11 is a catalytic subunit of the thymoproteasome, expressed exclusively in cTECs (cortical thymic epithelial cells). It has been shown that Psmb11-mediated positive selection is pivotal for the generation of an immunocompetent T-cell repertoire. We found that lack of Psmb11 results in a suite of gene expression changes in cTECs and not in mTECs, about 200 genes with enrichment in cell-cell interaction and antigen processing and presentation. One yet unsolved question is why the cellularity of CD8+ single-positive (SP8) but not SP4 thymocytes decreases strongly as a consequence of Psmb11-deficiency. Our transcriptomic analyses of pure SM, M1 and M2 subsets of developing SP4 and SP8 T cells show that the maturation of SP4 cells is essentially independent of Psmb11. However SP8 thymocytes present a dysregulated development with impaired migration, survival and lineage commitment in Psmb11 knock-out mice, also detected at the protein level by flow-cytometry. Thymocytes present decreased surface levels of CD127 on SP8 and slightly increased CD5 on all subsets. This indicates a stronger activation of the developing T cells, which alone cannot explain the impact on the SP8 cellularity. We have identified a novel role of Psmb11 in modulating, directly or indirectly, the expression of genes in cTECs, which is necessary to guide and complete the positive selection of SP8 thymocytes.
CCR7 modulates the generation of thymic regulatory T cells by altering the composition of the thymic dendritic cell compartment
Authors: Ehrlich Lauren Ilyse Richie , Hu Zicheng , Li Yu , van Nieuwenhuijze Annemarie , Kessler Rachel A , Liston Adrian 
Published: January 2023

Abstract
Upon recognition of auto-antigens, thymocytes are negatively selected or diverted to a regulatory T cell (Treg) fate. Both dendritic cells (DC) and medullary thymic epithelial cells (mTEC) present antigens to tolerize thymocytes. CCR7 guides thymocytes into the medulla, promoting encounters with medullary DC and mTEC. Thus, in the absence of CCR7 negative selection of auto-reactive thymocytes is impaired and autoimmunity ensues. Here we describe an unanticipated, non-thymocyte intrinsic contribution of CCR7 to intrathymic Treg generation. We find that Ccr7−/− mice have increased Treg cellularity, due to a hematopoietic, but non-T cell autonomous function of CCR7. In addition to thymocytes, thymic DC also express CCR7, which promotes survival of Sirpα−DC. Without CCR7, Sirpα−DC are preferentially lost, altering the composition of the conventional thymic DC compartment to favor intrathymic Treg generation. CCR7 deficiency results in enhanced production of thymic Treg at the neonatal stage and in lymphodeficient adults, when Treg generation is most critical for the establishment of self-tolerance. Together these results reveal a more complex function for CCR7 in thymic tolerance induction, in which CCR7 not only promotes negative selection through its effect on thymocyte localization, but also governs intrathymic Treg generation via non-thymocyte intrinsic mechanisms.
AIF1 expression regulates differentiation of dendritic cells from hematopoietic stem cell multipotent progenitor subsets and regulates antigen presentation capacity
Authors: Elizondo Diana Margarita , Andargie Temesgen Ewunetu , Lee Clarence Matthews , Anderson Winston Anthony , Lipscomb Michael Wheeler 
Published: January 2023

Abstract
Dendritic cells (DC) are indispensable in orchestrating adaptive immunity, serving as the principal messengers between innate and adaptive arms. DC subsets are derived from CD34+ hematopoietic stem cells (HSC). DC development progress through differentiation from HSC multipotent progenitors towards common myeloid progenitors. However, additional proteins involved in this selective differentiation process remain to be identified. For these studies, use of transcriptomic profiling and RNA interference (RNAi) approaches have now identified that Allograft Inflammatory Factor-1 (AIF1) directly contributes to DC differentiation and immunomodulatory processes. HSC multipotent progenitors, identified as CD117+Lin-CD34+SCA1+, were found to not express AIF1. However, CD117+Lin-CD34+SCA1- and CD117+Lin-CD34-SCA1- subsets, known as the oligopotent progenitors, did express AIF1. Importantly, this was restricted to common myeloid progenitors, as the common lymphoid progenitors, defined as CD117+Lin-SCA1-IL-7Rα+ did not express AIF1. RNAi knockdown of AIF1 in sorted Lin-CD34−CD117+SCA1+ cells led to restrained differentiation in DC upon culturing with GM-CSF. Interestingly, knockdown of AIF1 in committed CD11c+ DC resulted in impaired antigen presentation to cognate CD4+ T cells in vitro, suggesting an important immunostimulatory role in committed myeloid lineages. Current studies are now adoptively transferring HSC knocked out for AIF1 into mice to assess in vivo DC differentiation ability and capacity to migrate and populate the splenic compartment. This is the first report showing the novel role of AIF1 in dendritic cell development and antigen presentation.
Ras, Erk, and PI3K signaling pathways in the central selection of B cells
Authors: Greaves Sarah A , Babolin Chiara , Torres Raul M , Pelanda Roberta 
Published: January 2023

Abstract
B cell selection mechanisms in the bone marrow control the nature of the primary B cell repertoire, generating diverse specificities while reducing autoreactivity. Our studies aim to characterize the mechanisms that drive the positive selection of immature B cells in the bone marrow. It has been shown that the B cell receptor (BCR) is able to signal in the absence of ligand (tonic signal) and this signal is required for B cell development. We hypothesize that alterations in tonic signaling could cause aberrant positive selection of autoreactive immature B cells. We have previously shown that low levels of active Ras, Erk, and Akt correlate with levels of BCR tonic signal and that constitutive activation of Ras allows for differentiation of autoreactive immature B cells in vitro. We are currently testing the individual contributions of the Erk and PI3K pathways in this selection process. By using retroviral transduction of primary bone marrow B cells in vitro, we show that Erk activation can overcome a lack of tonic signaling to induce differentiation of nonautoreactive or slightly autoreactive B cells. We have also found that the Dual Specificity Phosphatase 5 (DUSP5), a novel phosphatase, may play a role in Erk regulation and the selection of immature B cells. In order to study these pathways in vivo we generated mouse models in which constitutively active forms of MEK or PI3K are specifically expressed in autoreactive B cells to determine if activation of these molecules is sufficient to bypass central tolerance. Preliminary data suggests that in vivo activation of PI3K can drive development of autoreactive B cells and allow some escape to the periphery.
Foxn1-β5t transcriptional axis controls CD8+ T cell production in the thymus
Authors: Uddin Muhammad Myn , Ohigashi Izumi , Takahama Yousuke 
Published: January 2023

Abstract
The thymus is an organ that produces functionally competent T cells that protect us from pathogens and malignancies. Foxn1 is a transcription factor that is essential for thymus organogenesis; however, the direct target for Foxn1 to actuate thymic T cell production is unknown. Here we show that a Foxn1-binding cis-regulatory element promotes the transcription of β5t, which has an essential role in cortical thymic epithelial cells to induce positive selection of functionally competent CD8+ T cells. A point mutation in this genome element results in a defect in β5t expression and CD8+ T cell production in mice. The results reveal a Foxn1-β5t transcriptional axis that governs CD8+ T cell production in the thymus.
Stroma-free, serum-free differentiation and expansion of hematopoietic stem and progenitor cells to the T cell lineage
Authors: Tabatabaei-Zavareh Nooshin , Le Fevre Tim A , Man Alexander JY , Karas Evan A , Szilvassy Stephen J , Thomas Terry E , Eaves Allen C , Wognum Albertus W 
Published: January 2023

Abstract
T cells are produced by the differentiation of hematopoietic stem and progenitor cells (HSPCs) into Lin− CD34+ CD7+ progenitors that migrate from the bone marrow (BM) to the thymus where further T cell development takes place. The differentiation of HSPCs into T cells can be supported in vitro by using OP9 stromal cells that express a Notch ligand. We have developed a more simple stroma-free culture method that promotes the expansion and differentiation of T cell progenitors from purified CD34+ HSPCs and purified multipotent lymphoid progenitors (MLPs) from cord blood (CB) or BM. CD34+ CB cells isolated using EasySep magnetic separation or CD34+ CD38− CD90− CD45RA+ MLPs further purified by cell sorting were seeded into culture plates coated with an attachment substrate containing Notch ligand and cultured in serum-free StemSpan SFEM II medium in the presence of a cytokine supplement containing SCF, TPO, Flt3L and IL-7. After 3 weeks, CD7+ CD5+ proT cells were detected at a frequency of 84% (range: 63–97%, n=26) with a proT cell yield of 2,130 (142–5,970, n=26) per initial CD34+ cell and 28,110 (4,720–63,930, n=4) per MLP initially seeded. In these cultures 28% (12–43%, n=12) of CD7+ cells also expressed CD1a, indicating further differentiation into preT cells. The pro and preT cells could mature into CD4+ CD8+ double positive T cells (38%; 24–64%, n=6) and TCRαβ+ CD3+ T cells (12%; 5–23%) over two weeks of continued culture in the same medium and in the presence of OP9-DL1 stromal cells. These results demonstrate that HSPCs can expand and differentiate into pro and preT cells under stroma-free and serum-free culture conditions. This novel culture system may benefit research and development of immunotherapies where large numbers of T cells are required.
Influenza virus induces mast cell death independent of mast cell activation
Authors: Temple Rachel Mary , Obar Joshua J 
Published: January 2023

Abstract
Influenza A virus (IAV) infects over a million individuals each year, resulting in significant morbidity, and during pandemic years, significant mortality. Severe IAV infection is characterized by a dysregulated cytokine response, termed a ‘cytokine storm’. Recent evidence indicates mast cells can play a central role in initiating this cytokine storm, though the long-term effects of IAV infection on mast cells is poorly understood. To this end, we infected bone marrow cultured mast cells with IAV at a low multiplicity of infection, allowing us to track IAV infection in mast cell cultures over time. Within hours mast cells express viral proteins, but fail to release appreciable levels of infectious viral particles throughout the course of infection. Despite the abortive nature of this infection, it is highly cytotoxic, with significant cell death occurring between 12–48 hours post infection. This cell death is the result of simultaneous activation of apoptosis and necroptosis pathways. Remarkably, while UV-inactivated virus activates mast cells, it does not induce cell death. Conversely, infection with a neuraminidase mutant is cytotoxic despite the fact this mutant fails to activate mast cells. These data suggest the induction of programmed cell death is dependent on viral replication, but independent of mast cell activation. In support of this hypothesis, the RIG-I and interferon signaling pathways are dispensable for the induction of cell death, as mast cells derived from MAVS−/−, AG129 (IFNAR1/IFNGR DKO), and STAT1−/− mice die at rates comparable to WT cells. Together, these data indicate mast cells detect IAV infection using multiple innate sensors, with each mode of detection resulting in the activation of a distinct response.
Assessment of lymph node stromal cells as an underlying factor in age related immune impairment to influenza infection
Authors: Masters April , Hall Alexxus L. , Haynes Laura 
Published: January 2023

Abstract
It is well established that aging negatively impacts immunity, resulting in inefficient responses to vaccinations and infections, like influenza. Fibroblastic reticular cells (FRCs) are the major stromal cell subset in lymph nodes and play an intricate role in the orchestration and control of adaptive immune responses. Although stromal cells have a major impact on immune responses, the impact of aging on LN stromal cells has yet to be determined. We sought to determine how aging impacts FRC number, phenotype and morphology at steady state and after influenza infection. An enzymatic and mechanical digestion was used to make single cell suspensions of lymph node stromal cells for analysis. Flow cytometry was performed to quantify the number and frequency of stromal cells in lymph nodes from young (13 weeks) and aged (19 months) C57BL/6 mice. To analyze stromal cell morphology, lymph nodes from young and aged mice were cryopreserved, and then sectioned for confocal microscopy. Quantitative analysis of lymph node stromal cells by flow cytometry revealed that there are no significant differences in the number stromal cells in young and aged lymph nodes at steady state, but after influenza infection aged FRCs fail to expand in number. Reduced proliferation and activation of aged FRCs, but not increased death leads to the decreased expansion. Microscopic analysis revealed that young and aged LN T cell zone FRCs have similar morphology at steady state and after infection. These results are the first evidence that aging alters lymph node stromal cells ability to respond to challenge. These changes in aged FRCs may be an underlying cause behind the impaired immune response to influenza infection found in the elderly.
Transcriptional and non-transcriptional functions of IRF3 in host defense
Authors: Chattopadhyay Saurabh , Subramanian Gayatri , Zhang Ying , Veleeparambil Manoj , Sen Ganes C 
Published: January 2023

Abstract
Innate immune response is the first line of host defense against microbial infection. Interferon Regulatory Factor 3 (IRF3), a critical transcription factor, is rapidly activated during virus infection to trigger numerous antiviral genes, including the interferons. Our studies have revealed that in addition to triggering these genes, IRF3 activates direct apoptosis of virus-infected cells by a newly discovered antiviral apoptotic pathway, RIPA (Chattopadhyay et al, Immunity 2016, EMBO J, 2010). In RIPA, IRF3 is differentially modified by linear polyubiquitination of two lysine residues. Moreover, a knock-in mouse strain, without the transcriptional activity of IRF3, can still mount antiviral response by its RIPA branch. Importantly, the use of pathway-specific mutants of IRF3 revealed that both transcriptional, and RIPA, branches contribute to the overall antiviral functions of IRF3. To investigate the contribution of the transcriptional branch of IRF3, we screened a shRNA library of the human ISGs. Our screen identified a small subset of novel IRF3-dependent genes, which exhibit antiviral functions in human and mouse cells. These newly-identified ISGs are protective against a wide range of clinically relevant human viruses, e.g. RSV, HPIV3, HSV-1, HCMV. In-depth investigation revealed that these ISGs regulate cellular autophagy pathway to control virus replication. The presentation will highlight how both pathways of IRF3 mount an optimum host response against viral as well as non-viral pathogenesis.
Distinct innate signaling and activation of dendritic cells during the neonatal period determines their functional response to respiratory syncytial virus infection
Authors: Lau-Kilby Annie W , Kehl Margaret , Malloy Allison MW 
Published: January 2023

Abstract
Dendritic cells (DCs) are among the first to respond to infection, providing the critical link between innate and adaptive immunity. Neonates often have weak innate immune responses that can limit adaptive immunity and contribute to increased susceptibility and more severe infection with intracellular pathogens, such as respiratory syncytial virus (RSV). We have previously shown differential migration and defective activation of neonatal murine respiratory CD103+ DCs, which is associated with diminished RSV-specific CD8 T cell responses in neonatal mice. In order to define the age-dependent regulation of respiratory DC function, we have developed a neonatal mouse model of RSV to track antigen presentation via infection with an ovalbumin-expressing RSV and an antibody against H2kb-OVA257–264. We found adequate transfer of RSV-derived OVA257–264 by neonatal CD103+ DCs to the dLN, suggesting poor CD8 T cell responses are not due to limited antigen presentation. However, neonatal CD103+ DCs expressing H2kb-OVA257–264 failed to upregulate CD80/86, which is critical to T cell stimulation. RNA-seq analysis of H2kb-OVA257–264+ CD103+ DCs implicated defective type I IFN signaling in neonates that may influence reduced activation. In support of this, while we find type I IFN to be diminished in the neonatal lung, treatment of neonates with IFN-α failed to restore the activation of neonatal H2kb-OVA257–264+ CD103+DCs.
Our data demonstrate a distinct developmental regulation of innate immunity in early life and identify type I IFN signaling as contributing to defective DC activation during RSV infection in neonates. Defining age-dependent mechanisms controlling immunity may lead to intelligently designed vaccines for this age group.
Aging of B-1a Cell Derived Natural Antibodies
Authors: Holodick Nichol E , Rothstein Thomas L. 
Published: January 2023

Abstract
Streptococcus pneumoniae is the most common cause of pneumonia, which claims the lives of people over the age of 65 seven times more frequently than those aged 5–49. B-1a cells provide immediate and essential protection from S. pneumoniae through production of natural immunoglobulin, which is germline-like due to minimal insertion of N-region additions. We have previously demonstrated B-1a derived phosphorylcholine (PC)- specific and total IgM moves away from the germline with age as a result of selection. Consequently, the protective capacity of natural serum IgM against S. pneumoniae infection is significantly decreased in aged mice. Anti-phosphatidylcholine (PtC) antibodies, like anti-PC antibodies, contain few N-additions, and have been shown to be essential in protection from bacterial sepsis. Herein, we demonstrate the number of N-additions in PtC-specific B-1a cell antibodies do not change with age. Furthermore, the average percent of B-1a cells binding PtC-liposomes (13–15%) did not change in aged mice. Therefore, in aged mice the majority of natural IgM derives from non-PtC binding B-1a cells producing antibody that has moved away from the germline. In direct contrast, the PtC-binding B-1a cell population preserves its germline status. These results suggest a difference in selection pressures and/or self-renewal upon B cells expressing different specificities. Elucidating the role of different B-1a cell progenitors and cues influencing B-1a cell circulation will be interesting and necessary lines of investigation to further understand how diversification and selection affect the maintenance of B-1a cells and the efficacy of the natural IgM they produce with increasing age.
A Mouse Model of Acute Ethanol and Burkholderia Infection
Authors: Monroy Fernando Porfirio , Jimenez Victor , Settles Erik , Currie Bart , Keim Paul S 
Published: January 2023

Abstract
Alcohol consumption both, acute and chronic has well-documented effects on the immune system. Melioidosis is an infection caused by Burkholderia pseudomallei of public health importance in Southeast Asia and Northern Australia. Alcohol consumption has been described as risk factor associated with B. pseudomallei infections in humans. Evidence supports the importance of alcohol consumption as a risk factor for development of meliodosis, but few experimental studies have focused on ethanol effects on Burkholderia spp. infection. In these studies, we propose the use of the closely related B. thailandensis E264 as a useful BSL-1 model to study the effects of alcohol on host’s immune responses. Female C57BL/6 were administered ethanol (4 g/kg) intraperitoneally 30 min before intra-nasal infection with 1 × 105 B. thailandensis CFU. Animals were euthanized at 24 and 72 hr post-infection (PI), blood and tissues were collected to determine bacterial dissemination. Our results at 24 hr showed bacterial dissemination to the lung, liver, and spleen in the alcohol group. Spleen had the largest number of bacteria present. At 72 hr PI, bacteria were found only in the lungs and the brains of mice administered alcohol. Mice in the alcohol group had significantly larger spleens at both time points. Furthermore, serum and tissue cytokine concentrations (TNF-α, GM-CSF, IL-10) were decreased by alcohol. These results provide insight into the effects of a culturally prevalent meliodosis risk factor, acute alcohol consumption as a predisposing factor for melioidosis. Future work will determine mechanisms of alcohol on bacterial dissemination and how this translates to B. pseudomallei and melioidosis rates of infection.