bluuebunny/crossref_metadata_2025_split huggingface dataset:
The C3-like molecule CD109 controls Th1 versus Th17 induction in CD4+ T cells
Authors: Merle Nicolas S , Kolev Martin , Rahman Jubayer , West Erin , Yan Bingyu , Kazemian Majid , Afzali Behdad , Kemper Claudia 
Published: February 2023

Abstract
Recent workdefined an unexpected and critical role for intracellular/autocrine active complement in human Th1 biology. Specifically, autocrine stimulation of CD46via TCR-induced C3b generation is needed for IFN-γ production by human CD4+ T cells. Thus, CD46-deficient patients are unable to generate normal Th1 responses. Analysis of the CD46-driven gene signature in Th1 cells revealed CD109 as a direct target of CD46. Indeed, T cells from CD46 deficient patients and from healthy donors in which CD46 expression was ablated via CRISPR-Cas9 technology failed to upregulate CD109. CD109 is aGPI-linked surface protein, mostly expressed on non-immune cells, and belongs to the complement C3 protein family. Though CD109 has been reported as a major negative regulator of TGF-β receptor signaling, direct functional activity for of CD109 on T cells remains unexplored.
Knock-out of CD109 in human CD4+ T cells with CRISPR-Cas9 technology induced uncontrolled Th1 and Th17 activation upon in vitro stimulation under non-skewing conditions. Similarly, CD4+T cells from Cd109−/− mice displayed augmented Th1 and Th17 in vitro responses and caused significantly more tissue pathology and weight loss in a T cell-transfer colitis model. Surprisingly, on T cells, CD109 does not restrain IFN-γ and/or IL-17 induction via modulation of TGF-β receptor activity. Instead, CD109 engages with a non-canonical costimulatory molecule and controls stemness- and metabolism-related signaling pathways.
Together, these data suggest that the complement-related protein CD109 serves as an important and novel molecular switch on CD4+ T-cells, where it regulates the balance between Th1 and Th17-induction pathways.
Time- and tissue-dependent roles of regulatory T cells in the immune response to HSV-2
Authors: Traxinger Brianna , Vick Sarah , Voillet Valentin , Erickson Jami , Woodward-Davis Amanda , Prlic Martin , Lund Jennifer M 
Published: February 2023

Abstract
Herpes simplex virus 2 (HSV-2) is a chronic sexually transmitted infection that causes recurrent genital ulcers and increases HIV susceptibility. Efforts to design a vaccine have assumed regulatory T cells (Tregs) restrain immune activation and impede viral clearance. However, we have previously shown that in a mouse model of vaginal primary HSV-2 infection, Treg removal leads to impaired T cell priming and migration. This suggests an alternative model where Treg function is location- and context-dependent. Additionally, research suggests that tissue-resident memory T cells (Trm) localized in the vaginal tract (VT) are crucial for efficient viral control, and Trm are likely regulated by local Tregs. These findings underscore the importance of understanding the specific contributions of VT Tregs during viral infection and vaccination.
We performed RNA sequencing of VT and lymph node (LN) Tregs after HSV-2 infection to reveal a highly activated transcriptional profile in VT Tregs compared to LN Tregs. Using flow cytometry, we confirmed an elevated activation and tissue residency phenotype in VT Tregs, exacerbated after HSV-2 infection. Notably, we found that VT Tregs express Granzyme B (GzmB) by both RNA-sequencing and flow cytometry. These Tregs persist in the VT up to 90 days, suggesting that they remain poised to respond to HSV-2 reencounter by restraining memory Trm to prevent tissue damage, possibly through GzmB-mediated toxicity. Tregs from healthy human VT corroborated mouse VT Treg phenotypes. These results suggest that VT Tregs are distinct and uniquely poised to respond to viral infection. Studies are ongoing to interrogate the signals that drive Treg migration into the VT and that lead to the tissue-specific Treg phenotype.
Sexual dimorphism in the antigen-specific T cell response in S. Typhimurium infections within extra-lymphoid tissues
Authors: D’Souza Shaina J , Godwin Matthew , Immethun Vicki E , Bauer David L , McLachlan James B 
Published: February 2023

Abstract
Differences in the immune response between the sexes, particularly in the susceptibility to and severity of infectious diseases, are well established. Females commonly generate a more robust response that is beneficial when combatting infections but leads to an increased risk of developing autoimmune disorders. While the sex-related hormone, estradiol (E2), is known to affect CD4 T cell cytokine release in vitro, less is known about how T cells differ in non-lymphoid tissues in vivo or how lymphoid tissues might influence T cell responses to infection. To address this, we intravenously infected lymphotoxin-alpha (LTa) male and female mice that genetically lack lymphoid tissues save for a small, disorganized spleen that was either left intact or removed surgically, with attenuated Salmonella enterica sp. Typhimurium (S. Tm). In infected mice lacking all lymphoid tissues, rates of survival between the sexes were similar. In sharp contrast, females with intact spleens had a significantly lower rate of survival in comparison to their male counterparts. We show that, in mice with or without intact spleens, the number of S. Tm-specific CD4 T cells are higher in male livers on day 10 post infection; however, by day 15, female mice with intact spleens had significantly increased CD4 T cell expansion in comparison to male mice with intact spleens, correlating to lower survival. This difference in T cell expansion, and the commensurate effect on survival, in the presence or absence of lymphoid tissue indicate that females may have evolved a separate ability to regulate immune responses outside of traditional lymphoid organs. These studies will provide insights into sex-based differences seen in cellular immunity against bacterial pathogens in vivo.
Impact of early like trauma on murine multiple sclerosis phenotype and development via dysregulated β1 adrenergic signaling
Authors: Khaw Yee Ming , Inoue Makoto 
Published: February 2023

Abstract
Environmental triggers have important functions in multiple sclerosis (MS) susceptibility, phenotype, and trajectory. Exposure to early life trauma (ELT) has been associated with higher relapse rates in MS patients; however, the underlying mechanisms are not well-defined. Using C57BL/6J mice, we show that ELT induces mechanistic and phenotypical alterations during experimental autoimmune encephalitis (EAE). ELT induces not only increases in severity and susceptibility in EAE but also induces a phenotype shift characterized by interferon-β insensitivity, severe neuronal damage in the spinal cord, in addition to membrane-bound lymphotoxin and CXCR2 involvement. To understand how ELT induces phenotype change in EAE development, we assessed sympathetic adrenergic neuron activity in immune cells of ELT mice. Using flow cytometry, we found that immune cell adrenergic receptors are chronically downregulated in ELT mice, attributable to chronic norepinephrine circulation. ELT-mediated EAE phenotypic changes are observed in control EAE mice treated with β1 adrenergic receptor antagonist. Conversely, β1 adrenergic receptor agonist treatment to ELT mice abrogates phenotype changes via restoration of immune cell β1 adrenergic receptor function. Our results indicate that ELT alters EAE phenotype via downregulation of β1 adrenergic signaling in immune cells. These results have implications for the effect of environmental factors in provoking disease heterogeneity and may enable prediction of outcomes in MS.
Cooperative ectodomain interaction among TCRαβ, CD3δɛ and CD3γɛ
Authors: Yuan Zhou , Cong Peiwen , Ge Chenghao , Natarajan Aswin , Travaglino Stefano , Krogsgaard Michelle , Zhu Cheng 
Published: February 2023

Abstract
The T-cell receptor (TCR) complex comprises of the ligand-binding subunit TCRαβ, and the signaling subunits CD3δɛ, CD3γɛ and CD3ζζ, with the Cα/Cβ in proximity to both CD3δɛ and CD3γɛ extracellularly. Direct measurements of the ectodomain interactions had not been successful, although they are believed to be important for TCR stability and functionality. Mechanical force has been shown to modulate TCR–ligand interactions. The TCR mechanosensor hypothesis predicts that force-encoded information may transmit from pMHC to CD3 via TCR-CD3 interaction. Evaluating ectodomain interactions among TCRαβ, CD3δɛ and CD3γɛ can help elucidate the TCR triggering mechanism and further guide the design of TCR-based immunotherapy.
Using two mechanical based assays, we were able to measure the weak two-dimensional (2D) affinities among ectodomains of human TCRαβ (2B4-LC13), and human CD3δɛ or CD3γɛ and showed catch bond formation where lifetimes of TCRαβ–CD3δɛ and TCRαβ–CD3γɛ bonds are prolonged by forces &lt;15 pN. Remarkably, CD3δɛ and CD3γɛ bind TCRαβ cooperatively, forming more bonds that last longer when both CD3s interact with TCRαβ as a whole than the sum of either CD3 interacting with TCRαβ individually. Interestingly, these measurements are comparable to 2D affinity and force-dependent bond lifetime of the 2B4 TCR interaction with its cognate ligand K5:I-Ek, supporting their relevance to TCR function. Using molecular dynamics simulations based on a published Cryo-EM structure, we identified the formation of long-lasting CD3δɛ–TCRαβ–CD3γɛ trimolecular bonds as the structural mechanisms of the cooperativity. Our work helps explain TCR function under force and suggests strategies for engineering of TCR for immunotherapy applications.
Inhibiting Yap for improved T cell activation and anti-tumor immunity.
Authors: Cheng Nan , Stampouloglou Eleni , Federico Anthony , Monti Stefano , Szeto Gregory Lee , Varelas Xaralabos 
Published: February 2023

Abstract
Immunotherapies have shown promise in the treatment of cancers, but challenges such as sustained T cell activation and recruitment into tumors have prevented their wide-spread success. We have identified Yes-associated-protein (Yap), a transcriptional regulator that plays key roles in mechanotransduction, as an important immunoinhibitory protein. Yap is rapidly induced at the mRNA and protein levels in CD4-posititve and CD8-positive T cells upon activation with mechanical stimuli impacting Yap nuclear-cytoplasmic dynamics. We found that T cell specific deletion of Yap results in enhanced T cell activation, differentiation, and function, which translates in vivo to an improved ability for Yap-deleted T cells to repress solid tumor growth. Strikingly, Yap-deleted T cells showed an enhanced ability to infiltrate solid tumors. Gene expression analyses of Yap-deleted tumor-infiltrating T cells demonstrates Yap as a mediator of T-cell responses in the local tumor microenvironment, and implicates Yap as a negative regulator of T-cell tumor infiltration and patient survival in diverse human cancers. Our observations indicate that Yap plays critical roles in T-cell biology and suggest that Yap inhibition may offer a method for improving T cell recruitment and activation in therapies for solid tumors.
CD5 regulates the heterogenous depot of NFκB available for T cell activation
Authors: Matson Courtney Alexandra , Choi Seeyoung , Livak Ferenc , Love Paul E , Singh Nevil J 
Published: February 2023

Abstract
T cells bearing receptors to the same antigen often differ considerably in their ability to respond. We hypothesized that variations in intracellular signaling molecules in T cells may underlie these cell-to-cell heterogeneities. We found that the expression levels of PLCγ and IκBα, varied considerably between individual peripheral T cells. The ~1000-fold variation in IκBα levels, but that not of PLCγ, strictly correlated with surface levels of CD5. CD5 is typically used as a marker for T cell self-reactivity and differences in responses between CD5-high and -low cells is attributed to the self-awareness of the TCR. Our data suggested that CD5 directly regulates the NFκB pathway to modulate T cell responses. Accordingly, ablating CD5 using germline or conditional knock-out mice resulted in a significant reduction in IκBα. This regulation was independent of SHP-1 and casein kinase II and not evident in mRNA levels, suggesting a previously unknown post-translational mechanism. Overexpression of CD5 in TCR-deficient cells also increased basal IκBα levels indicating independence from TCR signaling. Intriguingly, cells with higher CD5 and IκBα expression maintained a proportionally greater amount of NFκBp65 in a cellular reservoir. This suggests that CD5-high T cells with a greater NFκB reservoir might allow for a more robust NFκB-dependent gene expression. Consistent with this postulate, we found that CD5-high thymocytes survived ex vivo better than CD5-low ones, but only when NFκB signaling was intact. These data suggest a key role for CD5 in the control of T cell heterogeneity, independent of the self-reactivity of a TCR itself.
T cell stemness and ST6Gal-I mediated sialylation
Authors: Harrington Laurie E , Shin Boyoung , Anderson Ashlyn E , Ingram Jennifer T , Bellis Susan , Zajac Allan J 
Published: February 2023

Abstract
Populations of both CD4 and CD8 T cells with stem-like properties have been documented in the setting of autoimmunity, chronic infection, and cancer. These cells exhibit the capacity to self renew, as well as serve as progenitors to more differentiated T cell phenotypes, but the cell intrinsic factors that promote the development and maintenance of T cell stemness are not well known. In a mouse model of chronic intestinal inflammation, we observed that a subset of effector CD4 T cells displayed many of the attributes consistent with T cell stemness; these cells expressed the key stemness transcription factor TCF1, could persist long term in vivo, and further transitioned into terminal effector cells. Importantly, these stem-like effector CD4 T cells were able to both sustain and confer intestinal inflammation. To discern molecules that contribute to the stemness phenotype, we employed gene set enrichment analyses with multiple data sets and identified the glycosyltransferase ST6Gal-I, which facilitates the addition of a2,6-linked sialic acid moieties to surface N-glycans. We found that this sialyltransferase was preferentially expressed in stem-like CD4 T cells during colitis and CD8 T cells during chronic lymphocytic choriomeningitis virus infection, and that these cell populations displayed a2,6-linked sialic acids on the cell surface. Moreover, in the colitis model, we observed that genetic deletion of ST6Gal-I in T cells correlated with diminished TCF1 expression and reduced inflammation. Furthermore, constitutive expression of ST6Gal-I by T cells was associated with the ability to promote intestinal inflammation. Together, these data highlight ST6Gal-I as a potential regulator of the T cell stemness phenotype.
Serum- and feeder-free culture expansion of human peripheral blood NK cells
Authors: Le Fevre Tim , Ang Elaine , Wognum Albertus W , Szilvassy Stephen J , Louis Sharon A , Eaves Allen C , Tabatabaei-Zavareh Nooshin 
Published: February 2023

Abstract
Natural killer (NK) cells are critical effectors of innate immunity that secrete proinflammatory cytokines and kill tumor cells and virus-infected cells. The ability to expand human NK cells in culture provides a ready source of cells for disease modeling, development of immunotherapies, and basic research. We have developed a culture system that supports the expansion of primary NK cells in the absence of feeder cells and serum. NK cells were enriched from peripheral blood using EasySep™ negative selection and cultured in serum-free medium with interleukin-2 in plates coated with a NK cell ‘activation coating matrix’ for 14 days. Cells were passaged with medium changes on days 7 and 11, and were harvested on day 14 for assessment or functional characterization. The average frequency of CD56+CD3− NK cells was 92% (range 80 – 97%, n=16) with an average expansion of 121-fold (range 10 – 274). On average 79% (48–92%) of expanded NK cells expressed CD16. The ability of expanded NK cells to produce interferon-γ (IFN-γ) and to degranulate were tested after stimulation with phorbol 12-myristate 13-acetate (PMA) + ionomycin or co-culture with K562 cells. As detected by intracellular flow cytometry, the average frequency of IFN-γ+ NK cells was 56% (range 41 – 63%, n = 4) and 40% (range 12 – 55%) when stimulated by PMA/ionomycin or K562 cells, respectively. Similarly, the average frequency of degranulated NK cells as detected by surface expression of CD107a, a lysosomal-associated membrane protein, was 69% (range 35 – 84%, n = 4) and 53% (range 18 – 75%), respectively. These results show that NK cells can be expanded and stimulated under serum- and feeder-free conditions to generate large numbers of functional NK cells for basic and translational research.
A unique multiplex assay to profile growth factors and cytokines in CSF of MS and ALS patients
Authors: Sastry Amani , Madiraju Charitha , Krajewska Maryla , Krajewski Stan , Tomik Barbara , Szczudlik Andrzej , Matson Robert 
Published: February 2023

Abstract
Growth factors and cytokines play a critical role in the pathogenesis of multiple sclerosis (MS) and amyotrophic lateral sclerosis (ALS). The shift in the balance between pro-inflammatory and anti-inflammatory cytokines contributes toward autoimmunity. Epidermal growth factor (EGF) and fibroblast growth factor (FGF) are elevated in the cerebrospinal fluid (CSF) of MS and ALS patients. Identification of relevant biomarkers helps in the understanding of the disease pathogenesis. The goal is to establish a sandwich ELISA-based multiplex assay to assess growth factor and cytokine levels in CSF samples from MS and ALS patients. Capture antibodies against EGF, FGF, IFN-gamma, IL-6, IL-1beta and IL-4 imprinted in a multiplex array format into each well of a 96-well plate. Arrays subjected to multiplex sandwich ELISA of CSF samples. Test samples included CSF from Relapsing-Remitting MS (RRMS, N = 10), Secondary Progressive MS (SPMS, N = 10) and ALS patients [N=10]. Control samples included CSF from healthy subjects [N=10]. Colorimetric read-outs from multiplex ELISA are quantified using ImageJ analysis of standards and samples. Results suggest that multiplex array designed for ELISA is a unique, robust, and a cost-effective method for profiling growth factors and cytokines present in biological matrices.
[*In memoriam: abstract submitted as a tribute to honor &lt;underline&gt;Dr. Barbara Tomik&lt;/underline&gt; for her valuable contributions towards the collection and preparation of patient samples].
Linking immune receptor sequence to transcriptional states with clonotype neighbor graph analysis (CoNGA)
Authors: Schattgen Stefan A , Guion Kate , Crawford Jeremy Chase , Souquette Aisha , Barrio Alvaro Martinez , Stubbington Michael , Thomas Paul Glyndwr , Bradley Philip 
Published: February 2023

Abstract
Recent advances now allow for deep simultaneous profiling of T cell clonotypes, defined by T cell receptor (TCR) sequence, and phenotype, as reflected in gene expression (GEX) profile, surface protein expression, and epitope binding at the single-cell-level. However, there currently few tools available for unsupervised discovery of relationships between TCR sequence and cell phenotype. We hypothesized that by identifying correlations between “TCR neighborhoods”, defined by shared TCR sequence and GEX features, we could move beyond simply measuring GEX variation within clonal descendants and identify novel associations between T cell specificities and states. Previously, we introduced TCRdist, a measure for assessing inter-TCR similarity capable of identifying closely related clonotypes based on shared sequence features. Using TCRdist to quantify TCR similarity, we developed a graph-theoretic approach—clonotype neighbor-graph analysis or “CoNGA”—that identifies correlations between GEX profile and TCR sequence in an unbiased and automated manner through statistical analysis of GEX and TCR similarity graphs. Applying CoNGA, we uncovered novel associations between TCR and GEX space including a previously undescribed “natural lymphocyte” population of human blood CD8+ T cells; an association between TRBV gene usage and EPHB6 expression; and TCR sequence determinants of differentiation in thymocytes. These examples demonstrate that CoNGA is able to effectively deconvolve complex relationships between TCR sequence and cellular state. Conceptually, CoNGA could be extended to other clonally-related populations (e.g. B cells, tumors), and can easily incorporate other measurable features (e.g. ATAC-Seq).
Development of a 30-parameter flow cytometry panel for functional characterization of unconventional T cells and innate lymphoid cells
Authors: Iftikhar Yaser , Liechti Thomas , Roederer Mario 
Published: February 2023

Abstract
Unconventional T cells, which include mucosal-associated invariant T (MAIT) cells, invariant natural killer T (iNKT) cells, and γδ T cells, are defined by their recognition of antigens and microbial products, such as vitamin B metabolites, presented on non-classical MHC molecules. These “innate-like” cells have garnered attention for their unique potential for bridging the gap between innate and adaptive immunity, although their exact roles during infection is still not well-understood.
To evaluate the phenotypic and functional characteristics of these cells, we developed and optimized a comprehensive, 30-parameter flow cytometry panel which delineates these cell subsets on the basis of surface marker expression. In addition, we simultaneously assess effector functions through the measurement of intracellular markers for cytokines, activation molecules, and cytotoxic molecules, following stimulation. We also included markers for the evaluation of natural killer (NK) cells and innate lymphoid cells (ILCs), as they play distinct roles at this junction of innate and adaptive immunity. We optimized our panel in combination with innate, pan-T cell (IL-12/IL-18) and subset-specific (e.g. HMBPP for Vγ9 T cells) stimulation reagents to evaluate responses to distinct innate-like and adaptive immune stimulation modes.
We will use this flow cytometry panel to assess the immune profiles of various cohorts, including individuals who are cytomegalovirus (CMV) positive and human immunodeficiency virus (HIV) positive, in order to further elucidate how different acute and chronic infections can alter the frequencies, phenotypes, and functional characteristics of these cells.
Analysis of the structure and dynamics of synaptic interface revealed functional anomaly of naive CD8+ T cells during HIV infection
Authors: Anikeeva Nadia , Steblyanko Maria , Kuri-Cervantes Leticia , Buggert Marcus , Betts Michael , Sykulev Yuri 
Published: February 2023

Abstract
While it is thought that HIV infection leads to a global defect in T cells, effect of the infection on the T cells at various differentiation stages has not been studied. We have exposed freshly isolated polyclonal CD8 T cells at various differentiation stages to planar lipid bilayers that display anti-CD3 antibody and soluble ICAM-1 and analyzed morphology and dynamics of the synaptic interface formation by the T cells, parameters that are linked to their functional activities. We have found that a large fraction of purified naïve CD8 T cells from HIV-infected people developed mature synapses exhibiting atypical degranulation capacity. These data provide evidence that even at the earlier stages of differentiation CD8 T cells derived from HIV-infected patients reveal functional anomaly causing changes in the ability to degranulate. The latter mediated by aberrant functioning of integrins and actin cytoskeleton resulting from continuous inflammation during chronic HIV infection. Functional anomaly of naïve T cells further impairs functions of CD8 T cells at later differentiation stages contributing to a global defect in T cells and failure to control the infection.