bluuebunny/crossref_metadata_2025_split huggingface dataset:
The biological mechanisms of metformin effects on aging-associated inflammation in obesity
Authors: Calvo Sara Santa Cruz , Bharath Leena , Nikolajczyk Barbara 
Published: January 2023

Abstract
Obesity prevalence within the 60+ years-old (yrs) population is drastically increasing. This alarmingly high rate of obesity compels comparative studies to identify common players involved in aging-derived and obesity-associated inflammation. Blood CD4+ T cells from lean older humans show an upregulated Th17 signature accompanied by a defective redox system. Treatment of these cells with the anti-diabetic drug metformin (100μM) promoted a more young-like anti-inflammatory/antioxidant phenotype. Here, we investigate whether the protective effects of metformin observed in CD4+ T cells from lean aging adults persist in obesity.
Blood CD4+ T cells from obese older (60+ yrs) donors secrete twice the amount of the Th17 cytokines IL-17F and IL-21 when compared to their younger counterparts (25–35 yrs). Glutamate production, which supports Th17 cytokines secretion, is 1.5-times higher in cells from older compared to younger subjects. Mitochondrial superoxide production in these cells was 25% higher than in younger cells, with no changes in total peroxide.
Contrary to findings from cells of lean subjects, metformin failed to reduce Th17 cytokines in cells from obese older subjects, perhaps due to metformin’s inability to reduce glutamate production. However, metformin reduced hydrogen peroxide production 1.75-fold, with no effect on superoxide, in cells from obese older subjects.
These results collectively indicate that 1) obesity-associated upregulation of Th17 cytokines is mechanistically linked to glutamine metabolism and mitochondrial superoxide production and 2) metformin’s action targets peroxide production rather than superoxide, which is insufficient for reducing age-associated inflammation in obesity.
The Biological mechanisms of Metformin Effects on Aging-Associated Inflammation 1R56AG069685-01
The ZFP36 family of RNA-binding proteins regulate homeostatic and autoreactive T cell responses
Authors: Cook Melissa Erin , Bradstreet Tara R , Webber Ashlee M , Kim Jongshin , Santeford Andrea , Schwarzkopf Elizabeth A , Apte Rajendra S , Blackshear Perry J , Edelson Brian T 
Published: January 2023

Abstract
The zinc finger 36 (ZFP36) family of RNA-binding proteins, consisting of ZFP36, ZFP36L1, and ZFP36L2, is known to negatively regulate mRNA stability and/or inhibit translation of many transcripts, including cytokines. While there are reports of all three family members regulating T cell cytokine production, delineating the functions of these genes is challenging due to spontaneous phenotypes upon global deletion of single genes and potential redundancy in their functions. To overcome this, we generated Cd4-Cre+ Zfp36fl/fl Zfp36l1fl/fl Zfp36l2fl/fl mice. Only upon triple deletion, but not individual or paired deletions, do mice spontaneously develop an inflammatory disease characterized by early lethality, hypercytokinemia, and immune cell infiltration into some peripheral organs, including the central nervous system. As SNPs in human ZFP36L1 and ZFP36L2 have been linked to susceptibility for multiple sclerosis, we tested ZFP36 family member individual and paired T cell-deficient mice in experimental autoimmune encephalomyelitis (EAE). To our surprise, Cd4-Cre+ Zfp36l1fl/fl Zfp36l2fl/fl mice were markedly protected from EAE clinical disease, while no change in disease severity was seen with any deletion strains involving Zfp36. There was a severe impairment in generating antigen-specific CD4+ T cells in Cd4-Cre+ Zfp36l1fl/fl Zfp36l2fl/fl mice during EAE priming. Our findings demonstrate a novel redundancy of the ZFP family members in regulating T cell homeostasis and a shared role for ZFP36L1 and ZFP36L2 to promote clonal expansion. Understanding the individual and shared functions of the ZFP36 family members may lead to opportunities to target them to suppress T cell-driven autoimmunity.
M.E.C. supported by the National Science Foundation Graduate Research Fellowship program (DGE-1745038) and by grant 5T32AI007163 from the NIAID. B.T.E. supported by the NIAID (R01 AI113118).
Copper homeostasis is critical for T cell activation
Authors: Mandarano Alexandra H , McGargill Maureen A 
Published: January 2023

Abstract
Elevated copper levels are associated with autoimmune disease, infection, and tumors in both humans and mice. Modulating copper availability is a promising potential intervention to treat these diseases. However, the mechanisms underlying the beneficial impact of copper modulation in disease are poorly understood. Moreover, the role of copper homeostasis in immune function, including in T cell responses, is not clear. Copper is a required cofactor for proteins involved in essential T cell processes, including extracellular matrix regulation, signaling pathways, mitochondrial metabolism, autophagy, and maintenance of reactive oxygen species. To investigate the impact of copper levels on T cell function, we examined T cell activation during altered copper availability. We found that copper chelation limited T cell proliferation and reduced expression of activation markers following stimulation. We also found that both activation and copper availability altered copper transporter expression in T cells. Additionally, copper modulation substantially impacted the transcriptional profiles of activated T cells, including expression of transcription factors, co-stimulatory markers, cytokines, and metabolic genes. Overall, our data demonstrate that altered copper levels impair T cell activation. Further, our results reveal new pathways impacted by copper homeostasis in T cells, which will be key to both understanding the role of copper in disease and leveraging this role for therapeutic development. Studies to evaluate the role of copper homeostasis in the immune response in vivo are ongoing.
Supported by funding from ALSAC.
The phosphatidylinositol-transfer protein Nir3 modulates T cell development and function
Authors: Lu Wen , Helou Ynes A , Au-Yeung Byron B , Shrinivas Krishna , Liou Jen , Weiss Arthur 
Published: January 2023

Abstract
Before emerging as functional T lymphocytes, thymocytes transit through multiple selection stages during which T cell antigen receptor (TCR) signaling controls the survival and subsequent maturation. Hydrolysis of phosphatidylinositol 4,5-biphosphate (PIP2) by phospholipase C-γ (PLCγ1) represents a critical step in T cell receptor (TCR) signaling which leads to calcium increases as well as PKC and Ras activation, events that contribute to T cell activation. PIP2 in the plasma membrane (PM) is depleted rapidly upon TCR stimulation and the replenishment of PIP2 levels is dependent on delivery of its precursor phosphatidylinositol (PI) from the endoplasmic reticulum (ER) to the PM. It was not clear how the delivery of PI from ER is regulated in thymocytes and T cells or how the process impacts T cell development and function. Here, we show that a membrane-associated PI transfer protein, Nir3 (Pitpnm2), promotes PIP2 replenishment following TCR stimulation and is important for T cell development. The high expression level of the Nir3 gene in thymocytes suggests its role in thymocyte selection. In Nir3 deficient thymocytes, the replenishment of PIP2 following TCR stimulation is significantly slower. Moreover, Nir3 deficiency attenuates calcium mobilization in DP thymocytes in response to weak TCR stimulation. The impaired TCR signaling led to impaired thymocyte development at the beta-selection and positive selection stages in Nir3 deficient mice. These findings reveal the role of Nir3 in TCR signaling and thymocyte development. This study highlights the importance of PIP2 replenishment and the role of Nir3 in PI transfer through regulated ER-PM interactions during TCR signaling.
This work was supported by in part by the Howard Hughes Medical Institute, the National Institutes of Health (NIH), NIAID R37 AI114575, and DRC Center Grant P30 DK063720 (UCSF Parnassus Flow Cytometry Core, UCSF Diabetes Center).
The Ikaros zinc finger transcription factor Eos as a candidate regulator of TH2 differentiation and function.
Authors: Tuazon Jasmine , Sreekumar Bharath , Read Kaitlin , Yaeger Michael , Varikuti Sanjay , Gowdy Kymberly M , Oestreich Kenneth J 
Published: January 2023

Abstract
Although CD4+ T helper 2 (TH2) cells normally defend against parasitic infection, they play a central role in the pathogenesis of allergic asthma. Therefore, understanding the molecular mechanisms governing TH2 differentiation and function is a top priority for improving asthma outcomes. One pathway of interest for controlling TH2-mediated disease is IL-2/STAT5 signaling, which is essential for TH2 polarization yet boasts a complex regulatory network that is incompletely understood. Here, we identify the Ikaros zinc finger (IkZF) transcription factor Eos as a candidate regulator of IL-2/STAT5 signaling underlying TH2 differentiation and function. To start, we show that Eos gene and protein expression is increased in TH2 cells relative to CD4+ T cell subsets that are negatively regulated by IL-2/STAT5 signaling, such as TH17 and TFH cells. Given these findings, we sought to define the functional effects of Eos in TH2 cells using in vitro and in vivo murine house dust mite asthma models. Intriguingly, Eos deficiency in vitro and in vivo results in reduced expression of key TH2 transcription factors (Gata3, Blimp-1), effector cytokines (IL-4, IL-13), and differentiation receptors (IL-2Rα, IL-2Rβ, IL-4Rα). Mechanistically, our data reveal that Eos interacts with and increases the activity of STAT5, suggesting that Eos enhances TH2 differentiation and effector function through direct STAT5 regulation. These findings in proinflammatory TH2 cells are of high significance, as Eos, to date, has largely been associated with the immunosuppressive functions of TREG cells. Taken together, our data reveal a novel mechanism by which Eos positively regulates IL-2/STAT5 signaling to promote TH2 differentiation and function.
Work supported by grants from the NIH (R01AI134972) and The Ohio State University (Susan Huntington Dean’s Distinguished University Fellowship).
Profound abnormalities in thymic epithelial cells in Rag1 hypomorphic mice: implications for immune reconstitution after stem cell transplantation
Authors: Pala Francesca , Oguz Cihan , Corsino Cristina , Martins Andrew , Lack Justin , Tsang John , Notarangelo Luigi , Bosticardo Marita 
Published: January 2023

Abstract
Thymic epithelial cells (TEC) are a heterogeneous population of stromal cells which orchestrate T cell development and selection. Although recent reports highlighted TEC complexity, their developmental origin, hierarchy and differentiation still remain ill-defined. We compared TEC distribution and gene expression in wild-type (WT) mice and in mice carrying Rag1 hypomorphic mutations (Rag1mut) observed in patients with immunedeficiency and immune dysregulation. Single cell RNA-seq analysis of TECs isolated from Rag1mut mice revealed an excess of cortical TECs (cTECs). Medullary TECs (mTECs), albeit reduced in number, showed a similar distribution in previously described mature subsets. To address whether TEC abnormalities in Rag1mut mice might result from a developmental block, we compared them to TECs from neonate WT mice. Indeed, we observed a similar TEC distribution in adult Rag1mut mice and newborn WT mice, although we identified peculiar features in cTEC subsets of Rag1mut mice. Transplantation of WT hematopoietic stem cells (HSC) into Rag1mut mice was able not only to correct thymocyte development, but also to revert TEC phenotype to normalcy. However, in a competitive transplantation setting where 10% of WT and 90% mutant HSCs were used, only partial restoration of thymocyte subsets was observed, associated with incomplete rescue of TEC phenotype. In summary, we show that abnormalities of TEC composition in Rag1 mutant mice correlate with impaired development of thymocytes, highlighting the role of lymphostromal cross-talk in TEC maturation. Moreover, these data point to the need to achieve full donor chimerism after HSC transplantation in order to rescue the thymic compartment and prevent defects of immune tolerance.
This research was supported by the Intramural Research Program of the NIAID, NIH
Age-associated changes to lymph node fibroblastic reticular cells
Authors: Lancaster Jessica N , Kwok Tina , Medovich Shannon , da Silva Ildefonso Alves 
Published: January 2023

Abstract
The decreased proportion of antigen-inexperienced, naïve T cells is a hallmark of aging in both humans and mice, and contributes to reduced immune responses, particularly against novel and re-emerging pathogens. Naïve T cells depend on survival signals received during their circulation among the lymph nodes by direct contacts with stroma, in particular fibroblastic reticular cells. Gross changes to the architecture of the lymph nodes have been described, but it is unclear how lymph node stroma are altered by aging, and whether these changes contribute to reduced naïve T cell maintenance. Here, we visualized naïve T cell migration in the aged lymph node using 2-photon microscopy and determined that T cell motility is impaired when in proximity to fibrotic deposition. Adoptive transfer of young naïve T cells into young or aged hosts revealed reduced homeostatic turnover in the aged host, supporting the role of T cell-extrinsic mechanisms that regulate their survival. Further, we determined that development of early fibroblastic reticular cells of the lymph node stroma is impaired, which can contribute to declining levels of naïve T cell survival factors. Thus, our study addresses the controversy as to whether aging impacts the composition lymph node stroma and supports a model in which impaired differentiation of lymph node fibroblasts inhibits the interactions necessary for naïve T cell homeostasis.
Spatially Restricted T-cell Activation in Inflamed Tissues
Authors: Bala Noor , Prizant Hen , Hughson Angela , McGurk Alexander , Fowell Deborah J 
Published: January 2023

Abstract
Pathogen control requires T-cells to locate and make direct contact with antigen-presenting cells (APCs) for peripheral reactivation and delivery of effector molecules. The signals that orchestrate T-cell-APC interactions are poorly understood in inflamed peripheral tissues. Using intravital imaging (IVI), we have found that Th1 cell entry into the inflamed dermis is nucleated by perivascular CXCL10+ cellular clusters enriched for APCs. Th1 cells form more stable interactions with APCs within these clusters compared to outer regions. CXCL10+ clusters are amplified in a IFNγ-dependent manner, where CD4+ T-cell IFNγ release increases APC recruitment and CXCL10 expression. We hypothesize that these chemokine clusters serve as CXCL10 Peripheral Activation (PAC-10) niches, where Th1 cells achieve early peripheral activation away from the pathogen modulated milieu. We seek to determine the relationship between these spatially restricted PAC-10 niches, infection foci, and pathogen clearance. Using a combination of IVI, flow cytometry, and transcriptomics we characterized niche specific Th1 cells and niche cellular composition to determine if PAC-10 niches modulate Th1 cells for pathogen control. Initial studies reveal that niche specific Th1 cells have increased CD69 and IFNγ expression compared to non-niche counterparts. PAC-10 niches appear to have a spatially distinct innate cell population, likely with enhanced ability to support Th1 activation. Future studies will characterize the functional status of niche specific Th1 cells and evaluate the necessity of PAC-10 niches for pathogen control. These studies will inform therapeutic strategies to boost T-cell activation in infection or hinder activation in autoimmunity.
Supported by NIH NIAID P01 AI02851 NIH NIAID R01 AI070826
Strong tonic TCR signaling is associated with negative regulation of naive CD4+ T cells
Authors: Au-Yeung Byron B , Eggert Joel , Scharer Christopher , Zinzow-Kramer Wendy 
Published: January 2023

Abstract
Naive T cells constantly experience tonic or basal TCR signals in response to self-peptides presented by MHC (pMHC) in vivo. We aimed to define how tonic TCR signaling impacts naive CD4+ T cells at the functional, transcriptional, and epigenetic levels. The transgenic Nur77-GFP reporter gene is responsive to tonic TCR signals. Naive CD4+ T cells can express a broad range of Nur77-GFP transgene expression at a single cell level, even if they express identical transgenic TCRs. Immature T cells that express high levels of Nur77-GFP following selection in the thymus tend to sustain high Nur77-GFP expression in the periphery. Naive CD4+ T cells with a Nur77-GFPHI Ly6C− phenotype experience the strongest tonic TCR signaling and in the OT-II TCR transgenic system, exhibit less sensitivity to stimulation with low doses of cognate pMHC or to weaker affinity pMHC. Analyses of gene expression in Nur77-GFPHI Ly6C− cells suggest a transcriptional program consistent with T cell activation, but also of negative regulation. Analyses of chromatin accessibility by ATAC-seq revealed over 3000 differentially accessible chromatin regions in Nur77-GFPHI Ly6C− cells, including several loci within or near genes encoding negative regulators such as Ctla-4 and Tim3. Nur77-GFPHI Ly6C− cells exhibited increased accessibility at multiple loci in the gene encoding Tox. A subset of these loci was previously reported to be more highly accessible in exhausted T cells. These findings suggest that persistent strong tonic TCR signaling may induce changes similar to those that occur during exhaustion. We propose that naive CD4+ T cells adapt to strong tonic TCR signaling and as a result, exhibit decreased sensitivity to subsequent TCR stimulations.
Supported by a grant from NIH (K01AR065481)
Naive CD4+ T cell heterogeneity
Authors: Sajani Alia , Schaafsma Evelien , ElTanbouly Mohamed , Lines Janet , Noelle Randolph J 
Published: January 2023

Abstract
Although naive T cells comprise about 90% of circulating and 20% of tissue restricted T cells, most studies on T cells emphasize effector, regulatory or memory functions. The consensus that all naive cells comprise a functionally monomorphic population is challenged by our recent findings. A redefinition of the naive T cell state shows that these cells are heterogeneous, with pre-programed differentiation trajectories. VISTA (V-domain Ig suppressor of T cell activation) is an immune checkpoint restricted to the hematopoietic compartment. Here, using single cell sequencing, we demonstrate unexpected loss of quiescence in VISTA deficient CD4+ T cells compared to Wild Type CD4+ T cells. The quiescence module, defined by Klf2, Klf6, Btg1, Btg2 expression, is lost in VISTA deficient mice. Conversely, there is an expansion of the stem-cell memory-like cluster, defined by Tcf7, Bcl2, and Il7r. This altered subset make-up due to VISTA deficiency demonstrates that naive T cell heterogeneity is tightly regulated. Consequently, we identified 5 subsets in healthy adult mice using single cell sequencing of naive CD4+ T cells. These include a quiescence cluster, memory-like cluster, TCR reactive cluster, IFN responsive cluster and T cell undifferentiated cluster. This heterogeneity is imprinted in the thymus, maintained in the periphery, and is not influenced by the TCR repertoire. Tracking the differentiation trajectory of these naive CD4+ T cells shows biased lineage commitment dependent on progenitor cluster identity. Thus, thymic imprinting and peripheral maintenance of varied naive T cell identities determines T cell fate.
Supported by 5R01AR070760-05
The development of humoral immunity to common dietary antigens in human infants is accompanied by the accumulation of specific plasma cells in the thymus
Authors: Cordero Hector , Hess Jacob , Nitschki Elio , Shihab Ronzon , Roy Poulomi , Kalfa David M , Bacha Emile A , Zorn Emmanuel 
Published: January 2023

Abstract
We have reported on the accumulation of plasma cells (PCs) in the human thymus, starting several months after birth. This thymic PC niche includes clones specific to common viruses and vaccination antigens presumably generated through peripheral responses. Humans acquire humoral immunity to dietary antigens during the first 3 years of life. Here we investigated whether such immunity also resulted in the homing of specific PC to the thymus. Using a combination of dietary antigen microarrays, ELISA and ELISPOT assays, we tested for the presence of serum IgG and thymic antibody-producing cells specific to common food antigens in discarded blood and thymus specimens from 1 day to 2 year-old subjects (N=20). Our studies first confirmed cow’s milk proteins, including casein and lactoferrin, as prevalent targets of humoral responses during infancy. Remarkably, subjects with the highest serum IgG titer to cow’s milk antigens also had high-frequency PC in the thymus specific to the same antigens. Our study demonstrates for the first time the presence of antibody-secreting cells specific to dietary antigens in the thymic of human infants. Their possible contribution to tolerance mechanisms warrants further investigation.
This work was supported by a NIH-NIAID grant U01-AI131339. Dietary antigen arrays (ImmunoCap) used in this study were a gift from Thermo Fisher Scientific.
The role of microRNA-142 in B cell activation and effector functions
Authors: Graham Natalie Marie , Wang Wei-Le , Reyes Estefany , Boldin Mark P 
Published: January 2023

Abstract
Control of gene expression by microRNA (miRNA) has recently emerged as a critical mechanism that regulates B cell activation and function. However, the role of specific miRNAs in this process is unclear. Using germline knockout (KO) mice, we have previously shown that miR-142 ablation impairs humoral immune responses despite significant expansion of the B cell compartment, suggesting that miR-142 is critical for B cell effector function and terminal differentiation. To more precisely dissect the role of miR 142 in B cell effector responses, we developed an activated B cell-specific miR-142 KO mouse by breeding conditional miR-142 KO (miR-142fl/fl) mice with mice expressing Cre recombinase driven by activation-induced cytosine deaminase (AIDcre). To label activated B cells in miR-142fl/fl AIDcre mice, we bred them to mice carrying a loxP-flanked STOP cassette, allowing for the expression of fluorescent Zsgreen1 reporter protein in the presence of Cre recombinase. As expected, B cell development was largely unperturbed in miR-142fl/fl AIDcreZsgreen+/− mice. In contrast, FACS analysis of naive and antigen-challenged miR-142fl/fl AIDcre Zsgreen+/− mice revealed a significant decrease in germinal center (GC) B cells and plasma cells (PCs). RNA sequencing of GC B cells suggests dysregulation of transcription factor networks such as miR-142-3p targets BACH2 and IRF4 may contribute to the phenotypes observed in miR 142fl/fl AIDcre Zsgreen+/− mice. Future experiments will investigate molecular pathways governing terminal B cell differentiation and effector function in miR-142fl/fl AIDcre Zsgreen+/− mice, thereby elucidating the role of miR-142 in promoting a healthy response to antigen.