bluuebunny/crossref_metadata_2025_split huggingface dataset:
X-chromosome inactivation maintenance is perturbed in T cells from lupus mouse models and human lupus patients
Authors: Anguera Montserrat 
Published: January 2023

Abstract
Sex differences exist with immune responses, yet females are more susceptible to autoimmune and inflammatory disorders. One important biological factor underlying this sex bias is the X-chromosome, which is enriched for immunity-related genes. Female mammals use X-Chromosome Inactivation (XCI) to equalize gene expression between the sexes, which generates a transcriptionally silent inactive X-chromosome enriched with heterochromatic modifications and the long noncoding RNA Xist. We have discovered that mature naïve T cells are missing Xist RNA and heterochromatin marks on the inactive X (Xi), and these marks return to most cells with in vitro stimulation. During T cell development in the thymus, Xist RNA is not localized to the Xi and heterochromatin marks are progressively lost. Xist RNA returns to the Xi in mature T cell subsets between days 2 and 3 of in vitro activation. During lupus disease progression in the female-biased mouse model NZB/W F1, Xist RNA signals aberrantly appear in naïve T cells and Xist RNA becomes mis-localized from the Xi in activated T cells. These abnormal XIST RNA localization patterns are also observed in activated T cells from pediatric lupus patients in disease remission. Using next-generation sequencing, we found significant overexpression of X-linked genes across the X in female SLE T cells compared to healthy females. We propose that the dynamic mechanism of XCI maintenance female lymphocytes predisposes the Xi to become partially reactivated and to overexpress immunity-related genes. Our work provides the first mechanistic evidence for the female-specific enhanced immune responses and increased susceptibility for autoimmunity.
Germinal center organization mediated by T-bet dependent expression of CXCR3 and CCR6
Authors: Peel Jessica Nicole , Stone Sara L , Scharer Christopher D. , Boss Jeremy , Lund Frances E 
Published: January 2023

Abstract
The germinal center (GC) is composed of distinct regions with proliferation and affinity maturation of GC B cells (GCB) occurring in the dark zone (DZ) and antigen-mediated selection and commitment of the long-lived plasma cell (LL-PCs) precursors occurring in the light zone (LZ). Despite the key role for GCs in establishing enduring humoral immunity, we know little about the dynamics of the GCB cell response. We showed that the transcription factor T-bet is expressed by GCB cells and that B cell intrinsic T-bet is required for the LL-PC response following influenza (flu) infection. To test whether the loss of LL-PCs in these mice was due to a role for T-bet in regulating the GCB response, we infected 50:50 Tbx21−/−: pepboy chimeric mice with flu and enumerated GCB cells. We found that Tbx21−/− B cells were overrepresented in the GC relative to the pepboy cells and that Tbx21−/− GCB cells were enriched in the LZ. Partitioning of GCB cells in the LZ/DZ is regulated by chemokines and we showed that T-bet controls transcription of two chemokine receptors, CXCR3 and CCR6, which are expressed by GCB cells. Using 50:50 Tbx21−/−: pepboy chimeras, we tested whether T-bet regulates GCB cell positioning by inducing CXCR3 and repressing CCR6. We observed that the CCR6+ GCB cell subset, which is known to localize in the LZ, was very enriched in Tbx21−/− GCB cells. By contrast, the CXCR3+ GCB subset, which was dominated by WT cells, was enriched in the DZ. To determine the role for CXCR3 in GCB cell localization, we infected 50:50 Cxcr3−/− :pepboy chimeras and found that the Cxcr3−/− GCB cells are enriched in the LZ. Therefore, T-bet regulates GCB cell localization and potentially the dynamics of the GC response by controlling CXCR3 and CCR6 expression.
Epithelial cell-derived chemokines induce DC recruitment to the gastric epithelium upon H. pylori infection
Authors: Hashimi Marziah , Seberll Thomas A , Sidar Barkan , Wilking James N , Bimczok Diane 
Published: January 2023

Abstract
H. pylori is a non-invasive bacterial pathogen that infects the lumen of the stomach and interacts with epithelial cells. In tissues from H. pylori infected and non-infected patients, DCs are found in close association with the gastric epithelium, which likely promotes DC uptake of luminal H. pylori. We hypothesized that DCs are recruited to the gastric epithelium in a chemokine-dependent manner. The aim of this study was to identify the epithelial chemokines that recruit DCs under steady state conditions and upon H. pylori infection. In chemotaxis assays, supernatants from human gastric organoids microinjected with H. pylori induced a 2-fold higher recruitment of monocyte-derived DCs compared to mock-infected organoids. Microscopic analysis of DC organoid-co-cultures similarly revealed DC recruitment to the epithelium of non-infected organoids and significantly increased recruitment upon H. pylori infection. DCs showed only low chemotactic activity towards H. pylori or bacterial supernatants, pointing to a role for epithelial rather than bacterial factors. DC migration towards the supernatants of both mock-infected and H. pylori-infected organoids was abrogated by pertussis toxin, indicating that recruitment was chemokine dependent. Using a chemokine array, we showed that organoid gastric organoids secreted increased amounts CXCL1, 5, 8 and CCL20 upon H. pylori infection, while CXCL16 and 17 remained constant. Chemotaxis assays confirmed that human DCs show chemotactic responses to CXCL1, 5, 8, 16, 17 and CCL20. In summary, our study indicates that human gastric epithelial cells actively recruit DCs to the mucosal interface by secreting chemokines, with increased recruitment induced upon H. pylori infection.
Exit of human cutaneous resident memory CD4 T cells that enter the circulation and seed distant skin sites
Authors: Morawski Peter A , Klicznik Maria , Hoellbacher Barbara , Varkhande Suraj , Motley Samantha , Duhen Thomas , Gratz Iris , Campbell Daniel J. 
Published: January 2023

Abstract
Tissue-resident memory T cells (TRM) persist locally in non-lymphoid tissues, providing front-line defense against recurring insults. However, strict tissue compartmentalization of memory may pose certain disadvantages for a large barrier organ like the skin, and the long-term migratory behavior of human TRM and their contribution to the memory pool have not been fully elucidated. TRM at barrier surfaces are defined in part by expression of the markers CD103 and/or CD69 which function to retain TRM in epithelial tissues. Here, we found that CD4+CD69+CD103+ TRM in human skin can downregulate CD69, exit the tissue and be identified as a phenotypically unique population in the circulation of healthy individuals. These circulating TRM produce the cytokines IL-22 and IL-13, and express genes consistent with a role in host-defense and tissue-repair responses. RNA- and TCR-sequencing demonstrated that CD103+ TRM in the blood are transcriptionally and clonally related to CD69+CD103+ TRM in the skin. Furthermore, using a skin xenograft model, we confirmed that human cutaneous CD103+ TRM can exit the skin, enter the circulation, and recirculate to secondary human skin sites where they re-assume a TRM phenotype. Thus, although as a population CD4+ TRM in the skin are largely sessile, recirculation of cutaneous CD4+CD103+ TRM does occur in the steady state in humans, and these recirculating (rc)TRM cells can promote the spread of this functionally specialized T cell population throughout the skin.
EBI3 impacts Leishmania braziliensis-induced inflammation in localized cutaneous leishmaniasis
Authors: DE SANTANA ALYNNE KAREN M , Gupta Gaurav , Gomes Ciro , Turatti Aline , Filho Roberto Bueno , Carregaro Vanessa , Roselino Ana , Almeida Roque , Silva João 
Published: January 2023

Abstract
Localized Cutaneous Leishmaniasis (LCL) can ultimately progresses to chronic ulcerated lesion with strong local inflammatory reaction. We hypothesized that regulatory cytokines, such as Interleukin-27 (IL-27) and interleukin-35 (IL-35) can play a role in susceptibility to Leishmania braziliensis infection. IL-27 and IL-35 are members of the interleukin-12 (IL-12) family structurally related by sharing Epstein–Barr virus-induced gene 3 (EBI3), which forms a heterodimer either with the p28 subunit to build IL-27 or with IL-12p35 to form IL-35. We used RT-PCR and Immunohistochemistry analysis to detect high expression of IL-27 and IL-35 subunits in the patients biopsies compared to healthy controls, suggesting that these cytokines can be involved in immunopathology induced by the infection. Moreover, L. braziliensis infection also induced IL-27/35 mRNA expression both in vitro and in vivo. We here analysed the function of EBI3 during infection with the intracellular parasite Leishmania braziliensis. We showed that EBI3 defficient mice displayed bigger lesion and an elevated inflammatory immune response in the ear compared to Wild-type mice with similar parasites levels in the ear and lymph nodes. The lack of EBI3 led to an increased expression of T helper type 1(Th1-), Th2- and Th17-derived cytokines in the infected ears. So far, our results demonstrate that EBI3 acts as a regulator of inflammatory immune responses in experimental cutaneous leishmaniasis, although, it is not required for control of L. braziliensis infection.
Financial Support: CAPES, FAPESP, CRID.
RGC-32 regulates expression of insulin-like growth factor binding proteins in astrocytes
Authors: Tatomir Alexandru , Boodhoo Dallas , Nguyen Vinh , Cudrici Cornelia , Badea Tudor , Rus Violeta , Rus Horea 
Published: January 2023

Abstract
Response gene to complement (RGC)-32 plays an important role in the mediation of TGF-β downstream effects and activation of cell cycle. During experimental autoimmune encephalomyelitis (EAE), RGC-32 knock-out (KO) mouse astrocytes displayed an elongated, bipolar phenotype, resembling immature astrocytes and glial progenitors whereas those from wild-type (WT) mice had a reactive, hypertrophied phenotype, suggesting a role for RGC-32 in astrocyte differentiation. To gain more insight into the role played by RGC-32 in astrocytes, we investigated the transcriptional profile induced by TGF-β in astrocytes using qPCR arrays. The majority of genes differentially expressed were related to cell growth, extracellular matrix remodeling and signal transduction. Among them, a group of genes belonging to the family of insulin-like growth factor binding proteins (IGFBPs) and insulin-like growth factors (IGFs) were found to be most significantly regulated by RGC-32. The IGFBP category includes IGFBP2, IGFBP3, IGFBP6 and connective tissue growth factor (CTGF). Using Real-time PCR and Western Blotting, we confirmed the differential expression of these genes in RGC-32 KO astrocytes after TGF-β stimulation. When glial fibrillary acidic protein (GFAP) and CTGF levels were compared between WT and KO astrocytes after TGF-β stimulation, significantly lower levels of mRNA and protein expression were found in RGC-32 KO astrocytes. These data suggest that TGF-β stimulation of CTGF is RGC-32 dependent and that RGC-32 plays a role in the regulation of CTGF downstream effects. The differential regulation of IGFBPs and GFAP suggests that they might be responsible for the lack of astrocyte differentiation seen in RGC-32 KO mice during EAE.
Multivariate analysis of genes expressed in the colorectal tumor immune microenvironment driven by CXCL13
Authors: Nunez Sha’Kayla K , Griffen Tiara L , Young Corey , Dill Courtney D. , Adebayo Olayinka , Carey Kaylin , Lillard James W 
Published: January 2023

Abstract
Colorectal cancer (CRC) is a heterogenous disease having numerous molecular phenotypes. Key tumor progression markers such as microsatellite instability, BRAF, and KRAS mutation status have been widely studied. Less is known regarding the influence of chemokines and chemokine receptors in CRC. CXCL13 and CXCR5 have been shown to be associated with poor prognosis of advanced CRC. The central hypothesis tested in this study was CXCR5-CXCL13 signaling mediates gene expressions associated with CRC progression. RNAseq patient control and matched primary tumor samples were acquired from The Cancer Genome Atlas (TCGA). DESeq analysis identified differentially expressed genes between normal tissue and primary tumor groups. Weighted Gene Co-expression Network Analysis (WGCNA) elucidated gene networks that correlated with factors influencing CRC prognosis. Canonical pathway, upstream regulator, and protein-protein interaction analyses were performed using Ingenuity Pathway Analysis (IPA). CXCL13 was identified in the WGCNA-designated green module, which also contained XCL1, TNFRSF17, FCRN1, CCL19, IL-17A and IFNG. These genes are associated with immunoregulatory signals found in lymphoid and non-lymphoid cells, BCR-signaling and poor prognosis of CRC. Genes associated with energy metabolism, apoptosis, hypoxia and angiogenesis, including GNG4, PRKCG, WNT11, and KNG1, were found in the WCGNA-designated yellow module. These factors are involved in Wnt/CTNNNB1 and PI3K/AKT signaling, which have previously shown to be driven by CXCR5-CXCL13 interactions. Taken together, our data suggest a role of the CXCL13-CXCR5 axis in shaping the tumor immune microenvironment of CRC.
The association between the statistical significance of the AUC and measure of stochastic superiority when modeling cytokines: an example from colon cancer
Authors: Qeadan Fares , Stalter Lily , Beswick Ellen 
Published: January 2023

Abstract
Early successes in clinical trials suggest that immunomodulation through cytokine blockade may be an effective approach for developing new colorectal cancer (CRC) immunotherapies. It has been shown that certain cytokines shape a pro-tumorigneic tumor microenvironment and are linked with metastasis. Thus, identifying differentials in the cytokine response with respect to CRC metastatic stage is needed for further exploration to identify potential new therapeutic targets. In this study, we (a) use the area under the receiving operating characteristics curve (AUC) from logistic regression to classify metastasis and non-metastasis cases for CRC patients, (b) use the measure of stochastic superiority (denoted by A using the Wilcoxon-Mann-Whitney test), to give the probability of concordance between predicted and observed outcomes of metastasis and non-metastasis, and (c) examine the association between the statistical significance of the two measures (i.e. p-value for AUC vs. p-value for A) noting that, while it was shown that AUC is the same as A, limited studies were done examining the relationship between their statistical significance.
We obtained the AUC and A measures on a panel of 44 cytokines and chemokines identified in 69 subjects (35 with Metastasis and 34 without Metastasis) with CRC, and revealed that 7 cytokines (GCSF, IL-6, IL-10, MCP-1, IL-8, IL-1a, and IL-17A) were statistically significant in discriminating metastasis and non-metastasis patients according to the p-values of the A measure while only 5 cytokines were significant according to the p-values of the AUC measure (GCSF, IL-6, IL-10, MCP-1, and IL-8). This observation indicates that, while the two measures agree, the AUC test is subject to more type II errors.
Cell-targeted steroid signaling in the thymus determines T cell repertoire strength
Authors: Taves Matthew Donald , Mittelstadt Paul , Presman Diego , Hager Gordon , Ashwell Jonathan D 
Published: January 2023

Abstract
Thymocyte selection is critical for generation of a competent and tolerant T cell repertoire, and glucocorticoids (GCs) antagonize T cell receptor (TCR) signaling and negative selection to strengthen the TCR repertoire. While circulating GCs from the adrenals coordinate systemic responses, the thymus contains all the enzymes needed to independently synthesize GCs. However, the biological relevance, source, and targets of locally-synthesized GCs is unclear. To test the source and importance of local GC synthesis in vivo, we generated mice with targeted deletion of the GC-synthetic enzyme Cyp11b1 in thymic epithelial cells (Cyp11b1foxn1-Cre) or thymocytes (Cyp11b1lck-Cre). We found that expression of GC-responsive genes was reduced in Cyp11b1foxn1-Cre but not Cyp11b1lck-Cre thymocytes, confirming that epithelial cells are a major source of GCs in the mouse thymus. Cyp11b1foxn1-Cre T cells had reduced alloreactivity and antiviral responses, demonstrating a weakened TCR repertoire. To identify targets of epithelial GCs we used chromatin-associated GC receptors as a cell-specific biosensor of endogenous GC exposure. With this approach we detected and quantified targeted GC delivery to activated CD4+8+TCRhi thymocytes undergoing antigen-specific selection. Negative selection, as measured by PD-1 and Bim expression, was specifically increased in CD4+8+TCRhi thymocytes of Cyp11b1foxn1-Cre mice. These findings 1) demonstrate the biological importance of local GC synthesis, 2) show cell-specific targeting of paracrine GCs, and 3) quantitate intra-organ heterogeneity in GC levels within the thymus. GCs may similarly function as paracrine immunoregulatory signals across a range of tissues and contexts.
Advancing threespine stickleback as an outbred immunogenetics model by pinpointing the onset of adaptive immunity
Authors: Niebergall Emily R , Beck Emily A , Bassham Susan , Cresko William A. 
Published: January 2023

Abstract
The development of an outbred immunogenetics model system is needed to understand how genetic variation impacts phenotypic variation of disease states in humans. Threespine stickleback fish (Gasterosteus aculeatus) provide just such a model. Stickleback are genetically tractable laboratory organisms with a well-annotated genome, but with lines drawn from populations inhabiting vastly different habitats. Individuals from different populations show high levels of genetic variation. The onset of the adaptive immune system is currently unknown in stickleback, a significant hindrance in studies of immunodeficiency diseases. To characterize the early development of adaptive immunity, we will analyze the expression of known early indicators of adaptive immunity maturation. These include recombination activating genes, rag1 and rag2, which are essential to the maturation of T and B lymphocytes, and T cell receptor genes, tcr-β and tcr-γ, which are expressed in mature T lymphocytes. To analyze the expression of rag1, rag2, tcr-β, and tcr-γ, we will perform whole mount in situ hybridization throughout a developmental time series to detect when and where the genes are first expressed, with special focus on the head kidney and the thymus, followed by qPCR to quantify the expression of the early adaptive immunity genes. Knowing when adaptive immunity onset occurs in threespine stickleback advances threespine stickleback as an outbred disease model in immunogenetics studies, allowing manipulative studies of immunological disease phenotypes in the context of genetic variation.
mTORC1 signaling mediates T cell lineage choices
Authors: Bastardo Blanco Daniel E. , Yang Kai , Chi Hongbo 
Published: January 2023

Abstract
The thymus supports and guides the generation of a diverse repertoire of mature T cells from precursors derived from the bone marrow. The interaction between extrinsic factors and intrinsic signal strength governs thymocyte development, but the mechanisms linking these two aspects of T cell development remain elusive. Capitalizing on genetic deletion of RAPTOR, here we report that RAPTOR-dependent mTORC1 signaling couples microenvironmental cues with metabolic programs in orchestrating reciprocal development of two fundamentally distinct T cell lineages: αβ and γδ T cells. Loss of RAPTOR impairs αβ but promotes γδ T cell development while disrupting metabolic remodeling of oxidative and glycolytic metabolism. Mechanistically, we identify mTORC1-dependent control of reactive oxygen species (ROS) production as a key metabolic signal that, upon perturbation of redox homeostasis, impinges upon T cell fate decision. Our results establish mTORC1-driven metabolic signaling as a fundamental mechanism underlying thymocyte lineage choices.
Skin microbiome regulates SCF level in Keratinocytes and defines mast cell maturation
Authors: WU CHIA-CHI , Wang Zhenping , Di Nardo Anna 
Published: January 2023

Abstract

Background
Mast cell (MC) progenitors enter the skin and become mature in the surrounding microenvironment during embryogenesis and they fully mature 8 to 15 days after birth (in mouse). We published that skin commensal bacteria work together with skin resident cells (especially keratinocytes), in order to retain and differentiate mast cells into the skin. In the present study, we aim to demonstrate that this effect is due to TLR2 sensing of microbiome.

Methods
we used germ free mice (GF), hairless (hr/hr) mice and hr/hr/Tlr2−/− mice. Analyses were performed by immunohistochemistry, qPCR and FACS. IgE cross-linking and 48/80 have been used to test in vivo MCs degranulation.

Results
we found that GF mice present with less mature skin MCs than pathogen free mice (PFM). MCs immaturity correlated with a low level of stem cell factor (SCF) in keratinocytes, a critical MC differentiation and growth factor. Furthermore, compound 48/80, a known degranulation agent, induced less edema in the paws of GF mice than in those of conventional mice. A smaller population of c-Kit positive MCs in the GF mice normalized after being co-housed with PFM mice for 5 weeks to reconstitute their microbiome. On the other hand, hr/hr mice, have a skin increased number of mature and proliferating MCs that correlates with a more gram positive microbiome than their littermates. When hr/hr were backcrossed into Tlr2−/− (so they cannot sense the gram positive commensals) the number of MCs in skin dramatically decreased.

Conclusions
While baseline keratinocyte SCF expression is essential to recruit MCs into the skin, TLR2 stimulation by gram positive skin microbiome, promotes SCF keratinocyte production, which then enhances the maturation and proliferation of skin MCs.
Silencing of PD-L2/B7-DC by Topical Application of Small Interfering RNA Inhibits Elicitation of Contact Hypersensitivity
Authors: Furusawa Emi , Ohno Tatsukuni , Nagai Shigenori , Noda Taisei , Komiyama Takuya , Kobayashi Katsunori , Hamamoto Hidetoshi , Miyashin Michiyo , Yokozeki Hiroo , Azuma Miyuki 
Published: January 2023

Abstract
PD-L2 is a ligand for the immune checkpoint receptor PD-1, however, its regulatory function is unclear. We previously reported that silencing of CD86 in cutaneous dendritic cells (DCs) by topical siRNA application inhibits the elicitation of contact hypersensitivity (CHS). Here we investigated the effects of topical PD-L2 siRNA application on allergic skin disease. PD-L2 was induced in DCs concurrently with the elevation of MHC class II and CD86 expression. Topical PD-L2 siRNA application inhibited elicitation of CHS by suppressing early proinflammatory cytokine expression and migration of hapten-carrying DCs into lymph nodes. Local injection of neutralizing anti-PD-L2 mAb comparably inhibited CHS. PD-L2 siRNA treatment inhibited the CHS in PD-1/PD-L1 double-knockout mice and in the sensitized T-cell-transferred skin. These results suggest that the effects of PD-L2 silencing are independent of PD-1 but dependent on local memory T cells. Most of the inhibitory effects of PD-L2 and CD86 silencing on CHS were comparable, but PD-L2 siRNA treatment did not inhibit atopic disease-like manifestations and Th2 responses in NC/Nga mice. Our results suggest that PD-L2 in cutaneous DCs acts as a costimulator rather than a regulator. Local PD-L2 silencing by topical siRNA application represents a new therapeutic approach for contact allergy.