bluuebunny/crossref_metadata_2025_split huggingface dataset:
Dissecting the pathogenic versus protective roles of IFN-γ and IL-17 in staphylococcal toxic shock syndrome and pneumonia using gene targeted HLA-DR3 transgenic mice
Authors: Rajagopalan Govindarajan , Krogman Ashton , Chowdhary Vaidehi 
Published: January 2023

Abstract
The classical, adaptive T cell response to infectious agents is delayed and is antigen-specific. However, the superantigen (SAg) family of exotoxins, which are readily produced in vivo during Staphylococcus aureus infections, cause an immediate and non-specific activation of T cells, resulting in systemic elevation in many cytokines including IFN-γ and IL-17. While both IFN-γ and IL-17 are pro-inflammatory, they play distinct roles in mediating tissue injury and in immunity to infections. Therefore, to precisely delineate the roles of IFN-γ and IL-17 in immunopathology and immunity to diseases caused by S. aureus and its SAg, we first assessed the expression profiles of IFN-γ, IL-17 family members and their receptors in the spleens of HLA-DR3 transgenic mice challenged with a purified SAg using Illumina microarrays. The expression of ifng gene in spleens of HLA-DR3 transgenic mice was significantly higher than Il17a, Il17d and IL17f at all time-points tested (3, 6, 9, 24, 48 and 72 hrs, 3–4 mice per time point). Serum levels of these cytokines correlated with the gene expression data. Subsequently, HLA-DR3, HLA-DR3.IFN-γ−/− and HLA-DR3.IL-17−/− mice were generated and challenged with purified SAg or infected intratracheally with a SAg-producing strain of S. aureus. While both HLA-DR3 (mortality - 11/19) and HLA-DR3.IL-17−/− (mortality - 15/21) mice succumbed to SAg-induced toxic shock syndrome as well as pneumonia, HLA-DR.IFN-γ−/− mice (mortality - 1/11) were significantly protected (p&lt;0.01) in both disease models. Bacterial loads were not different. In conclusion, our novel study using gene targeted humanized mice reveals that while both Th1 and Th17 cells are robustly activated by SAg, Th1 cells and IFN-γ play a lethal role.
A blood-based transcriptional signature in a novel murine tuberculosis model predicts risk of human tuberculosis progression.
Authors: Plumlee Courtney R. , Gern Benjamin H. , Duffy Fergal J. , Delahaye Jared , Rustad Tige R. , Aitchison John D. , Sherman David R. , Zak Daniel E. , Gerner Michael Y. , Urdahl Kevin B. 
Published: January 2023

Abstract
Tuberculosis (TB) is a highly heterogeneous human disease that develops in a subset of individuals who inhale Mycobacterium tuberculosis (Mtb). Using advanced machine-learning algorithms, we discovered a blood transcriptional signature that identifies Mtb-exposed individuals in the process of progressing to active TB up to 18 months before they exhibit clinical symptoms. This signature may help prevent TB disease by identifying individuals who could benefit from early intervention. However, it fails to identify ~30% of progressors and does not reveal insights into the diverse outcome-governing pathways within the infected lung that underlie TB progression. To develop a tractable system that enables blood-based signatures to be linked to mechanistic pathways within the lung, we pioneered a novel murine TB model that recapitulates key aspects of human Mtb infection. We find that mice infected with a physiologic, ultra-low dose (i.e., 1–3 CFUs) of aerosolized Mtb exhibit a broad range of outcomes, with bacterial burdens ranging from less than 10 to ~106 CFUs within individual lungs. Mice that contain Mtb exhibit well-circumscribed granulomatous structures that share many features with human Mtb granulomas. In addition, we have identified a blood transcriptional signature that distinguishes “controller” and “progressor” mice. Remarkably, this mouse-derived signature is equally as effective as our previously identified human-derived signature at predicting TB risk in humans, confirming the model’s relevance to human disease. This finding enables us to use the tools of the tractable mouse system to address questions central to TB pathogenesis in a clinically relevant model.
Commensal Neisseria Modulates Host IL-6 to Promote Oral Colonization
Authors: Powell Daniel A. , Ma Mancheong , So Magdalene , Frelinger Jeffrey A. 
Published: January 2023

Abstract
Neisseria musculi, isolated from the oral cavity of wild caught mice, weakly (50%) colonizes C57BL/6J (B6) mice but readily colonizes CAST/EiJ (CAST) mice. In vitro stimulation of B6 or CAST splenocytes with WT Neisseria or E. coli LPS showed that CAST mice had a blunted inflammatory response producing significantly lower levels of IL-6 than B6 mice. The use of specific genetic knockouts highlighted a need for an intact innate immune system to prevent colonization. B6-RAG-1−/− were colonized at a similar rate to WT B6 while B6-MyD88−/− and TLR4−/− mice were readily colonized like CAST (100%). In vitro stimulation of B6 BMDM or splenocytes with WT Neisseria yielded only low levels of IL-6. A series of Neisseria mutants in the Type IV Pilus (TFP), a key component of bacterial-host interaction, failed to colonize susceptible CAST mice and showed increased levels of IL-6 compared to WT Neisseria stimulation, indicating that the TFP plays a role in down regulating host IL-6 production. Surprisingly, UV-inactivated Neisseria induced high levels of IL-6 regardless of TFP status supporting that this is an active bacterial process and the presence of TFP alone was unable to down regulate IL-6 production. Consistent with a critical role for IL-6 is preventing colonization, mice deficient for the IL-6 receptor were colonized at a rate of 100% indicating host IL-6 signaling plays a critical role in determining host colonization susceptibility.
Circulating memory is sufficient for protection against Chlamydia muridarum in the murine female reproductive tract
Authors: Labuda Jasmine C. , Pham Oanh , McSorley Stephen J. 
Published: January 2023

Abstract
Chlamydia trachomatis is the most prevalent bacterial STI worldwide and yet no vaccine currently exists to protect the public from this reproductive tract pathogen. Recent studies show that Tissue Resident Memory T cells (Trm) are essential for vaccine-induced protection against Chlamydia trachomatis in mice. Moreover, Trm have been implicated in protective immunity to other reproductive tract pathogens including Herpes Simplex Virus. Here, we examine the requirement of Trm T cells in protection against the murine pathogen Chlamydia muridarum (Cm), commonly used to model human Chlamydia infection. Using parabiosis to test whether circulating memory from mice immune to Cm infection can protect naïve mice from reproductive tract challenge with Cm, we found that naïve mice receiving circulating immunity were fully protected from Cm challenge. Ongoing studies with MHC class-II tetramers will examine the phenotype of protective circulating Chlamydia-specific CD4 memory T cells and whether migration to the reproductive tract is a requirement for protection against secondary challenge.
MicroRNA-10a regulates intestinal mucosal dendritic cell development and function by targeting CD11c
Authors: Huang Xiangsheng , Chen Feidi , Xue Xiaochang , Chen Liang , Yang Wenjing , Xu Leiqi , Bilotta Anthony James , Yao Suxia , Cong Yingzi 
Published: January 2023

Abstract
We have previously demonstrated that microRNA-10a (miR-10a), highly expressed in intestine DC and epithelial cells, and negatively regulated by microbiota, regulated T cell function and pathogenesis of inflammatory bowel diseases by targeting IL-12p40. However, whether and how miR-10a regulates mucosal DC development and function are largely unknown. In the present study, we found that intestinal lamina propria (LP) DC expressed higher levels of miR10a than that in mesenteric lymph node (MLN) and spleen DC. Retinoic acid (RA), a metabolite of Vitamin A (VitA), has been implicated in regulation of mucosal DC development. Interestingly, expression of ALDH1a, a gene encoding an enzyme converting VitA to RA, was positively associated with miR-10a expression in DC from LPDC, MLN DC and splenic DC. Treatment with RA up-regulated DC expression of miR-10a in vitro. Feeding mice diets with high levels of VitA (VAH) promoted, while feeding VitA deficient diets (VAD) decreased, LPDC expression of miR-10a. By using the luciferase assay, we identified CD11c as a target of miR-10a. In DC specific miR-10a deficient (CD11cCre miR10afl/fl, cKO) mice, CD11c+ DC, especially CD11c+CD11b+ DC, were significantly decreased in intestinal LP. When coculture with microbiota antigen specific T cells, cKO DC promoted the differentiation of T cell to IFNg+ Th1 cells but inhibited Foxp3+ Treg cell development. Collectively, our data demonstrated that miR-10a regulates mucosal DC development by targeting CD11c, and DC expression of miR-10a differentially regulates Th1 and Treg cell differentiation.
Dynamic changes in immune populations at the fetomaternal interface during term and preterm birth
Authors: Lewis Emma L. , Barila Guillermo , Brown Amy G. , Porrett Paige M. , Elovitz Michal A. 
Published: January 2023

Abstract
The biologic mechanisms by which birth is initiated are unknown. Recent evidence suggests that the immune system may modulate gestational length. We previously established that maternal immune cells enter the placenta by mid gestation and change in composition near term. We hypothesize that preterm birth also involves immunologic alterations at the fetomaternal interface.
To test this hypothesis, we used an established model of inflammation-induced preterm birth. Pregnant mice received intrauterine injections (IUI) of lipopolysaccharide (LPS) (250μg/100μl) (N=8) or saline (100μl) (N=7) on embryonic day 15 (E15). Control mice, without IUI, were sacrificed on E15 (N=7) and E19 (N=7). (Term gestation is E19.5.) Maternal blood, spleen, uterine implantation sites, and placentae were collected, processed, and analyzed for immune cells by flow cytometry (CD45, Ly6G, CD11c, CD11b, CD103, F4/80, IA/IE, CD3, CD4, CD8, CD19, CD49b, TCR γ/δ, FoxP3, IL-17).
At E19, term gestation, the frequencies of multiple immune cell types decrease in the placenta: regulatory T cells (1.6 to 0.8%; p=0.0004), NK cells (2.6 to 1.5%; p=0.009), CD11c+ myeloid cells (8.6 to 5.5%; p=0.013), and CD103+ dendritic cells (0.43 to 0.25%; p=0.013). Only NK cells decrease near term in the uterus (9.5 to 1.6%; p=0.011). In contrast, during inflammation-induced preterm birth, macrophages decrease in the placenta (4.4 to 2.5%; p=0.042) and neutrophils increase in the uterus (26 to 52%; p=0.0006). These findings demonstrate that the immune composition of the fetomaternal interface shifts prior to birth. However, both the cell types and location of this shift differ between term and preterm birth, suggesting distinct immunologic mechanisms contribute to these processes.
Surveying the TCR landscape of intestinal intraepithelial lymphocytes
Authors: Schattgen Stefan A. , Crawford Jeremy C. , Thomas Paul G. 
Published: January 2023

Abstract
The mucosal epithelial surface of the gastrointestinal tract (GIT) provides an important barrier against microbes, toxins, and dietary matter, with disruption of epithelial integrity underlying the pathologies of a number of autoimmune conditions. Residing in the gut mucosa, intestinal intraepithelial lymphocytes (IELs) expressing rearranged αβ T cell receptors (TCRs) are able to rapidly respond to antigens and mediate repair of damaged epithelium. IELs are continuously surveying a stream of microbial, food, and self-antigens, and while antigen-recognition via the TCR of IELs is presumed the major mechanism governing their functions, little is known about the TCR repertoire of IELs or its specificity. Even less is known about how the IEL repertoire is formed under “normal” conditions and whether the IEL repertoire is dynamically edited in response to changes in the gut. Here, we perform a comprehensive survey of the IEL landscape of mice for the three distinct subpopulations of TCRαβ+IELs (DN-nIELs, CD8-iIELs, and CD4-iIELs) by leveraging single cell and bulk level TCR repertoire analyses. Our data show the IEL repertoire contains a striking amount of diversity, landing between oligoclonal and polyclonal on the continuum of TCR diversity. We have identified numerous public TCR chains with distinct restrictions and found the repertoires within each subpopulation are close in distance between individuals. Long-term antibiotic treatment and synthetic diets led to an increase in the number, TCR diversity, and activation of the IELs, suggesting the abundance of microbial and dietary antigens are important, though not the only, factors that shape the IEL repertoire.
Obesity and Intraepithelial Lymphocyte Dysfunction
Authors: Jameson Julie M. , Park Christa S. , Limon Natalie 
Published: January 2023

Abstract
Obesity has become prevalent in the United States with over 30% of adults and 17% of children having a BMI over 30. Complications associated with obesity include chronic nonhealing wounds in the skin and increased susceptibility to more severe colitis in the intestine suggesting that obesity leads to barrier dysfunction. Intraepithelial lymphocytes regulate epithelial homeostasis through the production of growth factors and cytokines. Our laboratory has shown that obese mice exhibit reduced numbers of intraepithelial lymphocytes in both the skin and the intestine. The remaining intraepithelial lymphocytes have reduced growth factor and cytokine production resulting in dysregulation of the barrier. To better understand the mechanism of T cell dysfunction, we placed mice on high-fat diet immediately post-weaning when intraepithelial lymphocytes are seeding the intestine. After 7 weeks, mice fed normal chow had normal numbers of intraepithelial lymphocytes seeding the intestine, however mice fed a high-fat diet had almost 50% fewer cells. CCR9 and CD103 were both reduced on the lymphocytes suggesting problems with homing and retention. In addition, mice administered a high-fat diet were more susceptible to DSS-induced colitis. To determine whether intestinal epithelial cells could be restored to normal numbers in obese mice, obese mice were administered normal chow for 7 weeks. Indeed, reducing fat intake was sufficient to restore intraepithelial lymphocytes to normal levels and rescue mice from more severe colitis suggesting that dieting is effective in improving intestinal barrier function in obesity.
Osteopontin provides a survival signal for intestinal intraepithelial lymphocytes in mice
Authors: Nazmi Ali , Olivares-Villagomez Danyvid , Piazuelo M. Blanca 
Published: January 2023

Abstract
Intraepithelial lymphocytes (IEL) are a diverse immune population residing in between intestinal epithelial cells, which possess important roles during mucosal immune responses. The IEL family comprises cells such as TCRαb+CD4+, TCRαb+CD8+, TCRgd+, TCRab+CD8aa+, iCD8a and iCD3+, among others. Osteopontin (OPN), a pleiotropic cytokine encoded by the Spp-1 gene, is well known for its function in tissue remodeling, modulation of Th1 responses development, maintenance of Th17 function, and homeostasis of NK cells. Herein, we investigated the impact of OPN in the IEL compartment.
We compared the cell numbers of IEL, lamina propria (LP) lymphocytes and splenocytes from Spp-1−/− and wild-type (WT) mice. OPN deficiency reduced the number of TCRγδ+, TCRβ+CD4+, TCRβ+CD8α+ and TCRβ+CD4+CD8α+ IEL, but had no effect on LP and spleen lymphocytes. In in vitro studies, IEL from WT mice cultured in the presence of rOPN showed better survival rates than IEL incubated alone. The increased survival was blunted to control levels when CD44, a receptor for OPN, was blocked with anti-CD44 antibody.
When total T cells (including Tregs) from WT splenocytes were transferred into Spp-1−/−Rag-2−/− and Rag2−/− mice, we observed that although OPN deficiency did not affect donor-derived IEL reconstitution at day 7 post-transfer, it decreased the number of CD4+ and CD4+CD8α+ IEL at day 28 post-transfer. As expected, Rag-2−/− mice remained healthy throught the experiment; however, Spp-1−/−Rag-2−/− recipient mice presented a remarkably loss of body weight and colon inflammation.
Together, our results indicate that OPN is a critical survival signal for IEL via CD44, and that OPN-deficiency promotes development of colitis even in the presence of Tregs.
Reciprocal activation of B cells and group 2 innate lymphoid cells
Authors: Feldman Scott , Kasjanski Richard , Norton James E. , Kern Robert C. , Conley David B. , Welch Kevin , Tan Bruce K. , Peters Anju T. , Grammer Leslie C. , Harris Kathleen E. , Carter Roderick G. , Suh Lydia , Stevens Whitney W. , Kato Atsushi , Schleimer Robert P. , Hulse Kathryn E. 
Published: January 2023

Abstract
B cells can be activated at peripheral sites in chronic inflammatory disease, but the mechanisms that drive this, especially in humans, are unclear. We have reported elevated frequencies of Epstein-Barr virus-induced molecule 2 (EBI2) expressing B cells in tissues from patients with chronic airway inflammation. EBI2 is a marker of extrafollicular plasmablasts, which are activated antibody-secreting cells (ASC). We sought to determine the mechanisms of extrafollicular B cell activation during chronic airway inflammation. ELISpot was used to determine ASC frequency in cells from inflamed nasal tissue or control tonsil tissue. B cells, T cells, and group 2 innate lymphoid cells (ILC2) were isolated from peripheral blood, and B cells were co-cultured with group 2 innate lymphoid cells (ILC2) or T cells. EBI2 expression was assessed by flow cytometry, and gene expression changes were assessed by single cell RNA-seq. Inflamed nasal tissue had a higher frequency of ASC compared to tonsil (p&lt;0.05). The majority of the ASC were contained in the EBI2+ B cell subset in nasal and tonsil tissues. Co-culture of B cells and ILC2 significantly increased the frequency of EBI2+ B cells (p&lt;0.01). B cells co-cultured with ILC2 expressed significantly higher levels of CCL17, CCL22, and FceR2 (&gt;5 fold), compared to freshly isolated B cells, or B cells co-cultured with T cells. ILC2 co-cultured with B cells had significantly increased expression of IL-5, IL-13, and IL-2Ra (&gt;7 fold) compared to freshly isolated ILC2. Our data suggest that not only can ILC2 directly activate B cells, but B cells can also enhance ILC2 function. These findings provide new insights into mechanisms that B cells may play in chronic inflammatory disease.
Novel cell-based high-throughput hybridoma screening method using the Celigo image cytometer for antibody discovery
Authors: Chan Leo , Zhang Haohai , Rice William , Kassam Nasim , Longhi Maria Serena , Zhao Haitao , Robson Simon , Gao Wenda , Wu Yan 
Published: January 2023

Abstract
Hybridoma screening is a highly important process that can identify potential clones from hybridoma cultures against the desired target antigen. Traditional screening methods using ELISA and flow cytometry presented technical issues such as limited accuracy, reduced high-throughput capability, increased time and cost. Here, we demonstrate a novel cell-based high-throughput screening method using the Celigo Image Cytometer, which avoids nonspecific signals by contrasting antibody binding signals directly on living cells, with and without recombinant antigen expression. The image cytometry screening method was optimized by detecting the binding of hybridoma supernatants to the recombinant antigen CD39 expressed on CHO cells. Both target CHO and CFSE-stained wild type were co-cultured to simultaneously detect target antibodies and nonspecific binding, which can eliminate the need to produce a separate control sample for each hybridoma supernatant. Next, the image cytometer was used to screen 672 hybridoma supernatants to develop the screening workflow. The Celigo was used to quickly image and analyze antibody binding using Hoechst, Alexa Fluor 594 and CFSE fluorescence to identify high, medium, and low binding hits. In addition, the Celigo was used to measure the binding affinities of the target antibodies to membrane-bound CD39. Furthermore, measuring direct antibody binding to living cells eliminated both false positive and false negative hits. The image cytometry method was highly sensitive and versatile, and could detect positive antibody in supernatants at concentrations as low as ~5 ng/mL. We propose that this method will greatly improve screening technologies and facilitate more efficient antibody discovery.
Sample multiplexing of peripheral immune populations for high throughput single-cell RNA-sequencing
Authors: Bansal Nidhanjali , Chang Christina , Liang Yuqiong , Shum Eleen Y. , Martin Jody C. , Ghadiali James , Jensen Devon , Hu Jing , Rosenfeld David , Zheng Ye , Fan H. Christina 
Published: January 2023

Abstract
Diverse immune populations reside in non-lymphoid organs and contribute to immune defense and tissue homeostasis. However, these cells are often hard to study due to low cell abundance and high heterogeneity. Recent advancements in single-cell sequencing technology provides a powerful high parameter tool to study these peripheral immune populations. But current single cell experiments are costly and limited by sample throughput. To address these limitations, we have developed a novel sample multiplexing approach for high throughput single-cell sequencing.
In this study, we performed single-cell sequencing analysis of thousands of immune cells isolated from peripheral tissues in mice, and utilized a DNA barcoded universal antibody to sample multiplex up to 12 samples in a single experiment. This allowed us to combine samples from different mice and tissue types into a single pooled sample, significantly reducing experimental scale and cost, while eliminating potential batch effects. The sample pool was captured on the BD Rhapsody™ system and a targeted assay was performed to measure gene expression of ~400 genes. We were able to de-multiplex the pooled samples with high specificity after sequencing. The targeted gene panel provided robust clustering of the major immune cell types, enabling us to perform immuno-profiling and compare gene expression of major cell populations across different tissues. We observed distinct tissue-specific expression profiles of major immune populations, and further investigation of the differentially expressed genes may provide better understanding of the interactions between these tissue immune populations and their local environment.