bluuebunny/crossref_metadata_2025_split huggingface dataset:
Possible link between cystic fibrosis specific lipid A adaptations by Pseudomonas aeruginosa and disease associated hyperinflammation.
Authors: SenGupta Shuvasree , Uriarte Silvia M. , Ernst Robert K. , Mitchell Thomas C 
Published: January 2023

Abstract
Pseudomonas aeruginosa infection in cystic fibrosis (CF) lung disease causes airway neutrophilia and hyperinflammation without being cleared effectively. We evaluated the immunostimulatory activities of lipid A variants synthesized by P. aeruginosa (PA) exclusively in CF patients to determine if they correlate with CF disease severity and progression. One third of the PA isolates from CF patients with late severe stage express a unique hepta–acylated lipid A; hepta-1855 (m/z=1855) isoform. In primary cell cultures, we found that hepta-1855 functioned as a potent TLR4 agonist by priming neutrophil respiratory burst and stimulating strong chemokine response (interleukin-8 or IL-8) from monocytes and neutrophils. Hepta-1855 also had a potent survival effect on neutrophils. However, it was less efficient in inducing neutrophil granule exocytosis and was also less potent in triggering pro-inflammatory TNF-α response from monocytes. A precursor hexa-1616, found in hepta-1855 containing LPS mixtures, did not have direct inflammatory activity on neutrophils but induced moderate IL-8 response. Together, our data suggest a potential contribution of hepta-1855 to late CF stage associated excessive inflammatory burden by recruiting neutrophils via IL-8 and promoting their maintenance through its survival effect. Being a chemoattractant stimulus, hexa-1616, may serve as an accomplice to hepta-1855. Moreover, the relative inefficiency of hepta-1855 in triggering degranulation may partly explain the persistence of PA in CF disease in spite of neutrophil dominated inflammation.
HLA-DR3 transgenic mice expressing Nur77-GFP reveal early and robust activation of T cells by superantigens during Staphylococcus aureus pneumonia.
Authors: Rajagopalan Govindarajan , Karau Melissa , Krogman Ashton , Patel Robin , David Chella S 
Published: January 2023

Abstract
The role of superantigens (SAg) in the immunopathogenesis Staphylococcus aureus pneumonia has been shown by several indirect means. To directly demonstrate activation of T cells in vivo during staphylococcal pneumonia by SAg and to study the kinetics of T cell activation, HLA-DR3+.Nur77-eGFP+ mice were used. In these mice, rapid expression of green fluorescent protein (GFP) in T cells occurs only upon activation through the TCR, but not following other inflammatory stimuli. Experimental pneumonia was induced using isogenic strains of S. aureus either producing the SAg staphylococcal enterotoxin B (SEB), or not producing any SAg. Mice were sacrificed at different time points and expression of GFP was determined by flow cytometry. As SEB primarily activates CD4+ and CD8+ T cells bearing TCR Vβ8, the expression of GFP within the TCR Vβ8+ sub-populations was examined. Strong upregulation of GFP in the TCR Vβ8+ sub-population was appreciable in both CD4+ and CD8+ T cell subsets (20–25%) even as early as 4 hours only in mice infected with S. aureus strain producing SEB (SEB+SA), but not those infected with the isogenic strain that does not produce SEB (SEB-SA). Serum cytokine/chemokine levels mirrored this finding. By 24 hours, 80–90% of the CD4+ and CD8+ T cells bearing TCR Vβ8+ expressed GFP in mice infected with SEB+SA, compared to only 1–2% in mice infected with SEB-SA. The pattern of expression of CD69 expression, an early T cell activation marker, was distinct from that of GFP. Our novel study directly demonstrated T cell activation and established their activation kinetics in vivo during staphylococcal pneumonia. These results might be useful for timing of administration of novel drugs aimed at suppressing T cell activation during pneumonia.
A multi-prong global study provides insights to gut microbiota and mucosal immunity in dusp6-deficiency conferred obesity-resistance.
Authors: Ruan JW , Statt S , Huang CT , Tsai YT , Chan HL , Tan TH , Kao CY 
Published: January 2023

Abstract
Obesity is now a plague in developed countries and more than one third of U.S. adults are obese. Gut microbiota plays profound roles in host energy harvest, metabolism and inflammatory response associated with the development of obesity. Dusp6-deficient mice have been shown to be resistant to diet-induced-obesity (DIO) but the mechanism is still unclear. Using a gnotobiotic mouse model, we found that the transplantation of fecal/gut microbiota derived from dusp6-deficient mice can significantly increase energy expenditure and reduce weight-gain of the recipient mice. By analyzing fecal 16s rRNA genes, dusp6-deficient mice have different composition of gut microbiota when compared with wild-type mice. We also demonstrated that dusp6-deficient mice were resistant to DIO-mediated dysbiosis of gut microbiota. Upon further analysis of the intestinal transcriptome, we found that dusp6 deficiency is an important regulator of several metabolic pathways and genes related to intestinal barrier. Moreover, our analyses have shown that dusp6-deficient mice could alleviate DIO-induced inflammation.. This study demonstrates that dusp6 deficiency is a strong genetic factor that dominates diet effect on shaping gut microbiota and it remodels mucosal immunity to retain HFD-resistant homeostasis of gut microbiota. Our findings shed lights on obesity and obesity-derived metabolic disease treatment with novel therapeutic candidates/strategies such as DUSP6 pharmacological inhibitors and the development of microbiota-based therapeutics.
The presence of microbiota protects against Pseudomonas aeruginosa induced keratitis
Authors: Kugadas Abirami , Gadjeva Mihaela 
Published: January 2023

Abstract
The impact of microbiota on ocular immunity is unclear. We report that in health, the presence of microbiota strengthens the ocular innate immune barrier function and promotes resistance to infection. Consistent with this view, Germ-Free mice (GF) on a SW background that are typically resistant to P. aeruginosa-induced keratitis were rendered susceptible by the lack of microbiota. This was exemplified by increased corneal bacterial burden and elevated pathology of the GF when compared to conventionally maintained specific pathogen free SW mice (SPF). The elevated susceptibility correlated with significant decrease in the antimicrobial capacity of neutrophils derived from GF SW against P. aeruginosa (PA). Oral antibiotic treatment of SPF SW reduced the ability of neutrophils to kill PA in vitro and in vivo, demonstrating that microbiota-derived cues maintain neutrophil priming in the bone marrow against PA. In contrast, topical antibiotic treatment reduced ocular commensal presence at the conjunctival surface, but did not the resistance to infection, suggesting that microbiota resident at a site different from the ocular mucosal tissue are responsible for neutrophil priming. Consistent with this reconstitutions of GF with mouse or human-derived gut microbiota restored resistance to infectious ocular challenge. To determine the mechanims of commensal priming, RNAseq experiments comparing the phenotype of GF and SPF-derived neutrophils were carried out and showed alterations in type I interferon-dependent gene expression profiles, implicating IFN-dependent pathways in microbiota-dependent neutrophil maturation. Indeed, type I interferon priming of GF mice reconstituted neutrophil bactericidal activities against PA.
Microbiota-derived butyrate negatively regulates RORγt+ ILC3 in Peyer’s patch of the terminal ileum
Authors: Jang Yong-Suk , Kim Sae-Hae , Kim Ju , Kim Yu Na 
Published: January 2023

Abstract
In the intestinal immune system, anatomical and physiological distinctions in the gut contribute to its regional specialization by microbiota and immunomodulatory agents from the diet. Given that the ileal Peyer’s patch (PP) belongs anatomically to the small intestine, while it is physiologically exposed to an environment similar to the large intestine with respect to microbes and microbial metabolites, its characteristics may differ from those in PPs in the jejunum. As the terminal ileal PP is a key mucosal organ where host defense and oral immune tolerance in the gut develop, identification of the specific factors regionally specialized in the tissue is essential to understand mucosal homeostasis. Among the various cell types present in PPs, group 3 innate lymphoid cells (ILC3s) are closely associated with the regulation of commensal bacteria through the suppression of commensal bacteria-specific CD4+ T cells, although the regulation of ILCs in ileal PPs is poorly defined. In this study, we found that butyrate plays a role as a regional specific factor involved in the repression of ILC3s in PPs of the terminal ileum. This butyrate-mediated negative regulation of ILC3s alleviates the tolerogenic mucosal microenvironment by suppressing regulatory T cells in PPs. Collectively, we conclude that the inhibition of ILC3s by microbiota-derived butyrate can confer the functional ability to induce antigen-specific immunity, and that this network contributes to homeostatic regulation of the mucosal immune inductive site.
Discovery of a novel subset of lung resident macrophages that regulate pathogen clearance and immune homeostasis after respiratory infection
Authors: Cicek Basak B , Khanna Kamal Mohan 
Published: January 2023

Abstract
The importance of tissue resident macrophages (TRMϕ) for pathogen clearance and homeostasis is beginning to emerge. However, our understanding of the multifaceted roles these macrophages play in mucosal tissues such as the lung remains incomplete. Currently the major population of TRMϕs in the lung is known to be alveolar macrophages (AMϕ) that express CD11c and CD169. Here we report a novel subset of CD169+ lung resident macrophages that are phenotypically and developmentally distinct from the alveolar or interstitial macrophages. These novel TRMϕ were CD169+CD11C−CX3CR1+Ly6C−CSF−1R+. A remarkable quality of these TRMϕs was their geographical location, which was important for their function. They were exclusively localized around the airways of the lung close to the airway epithelial cells. These airway-associated macrophages (AAMϕ) were radio-resistant, self-renewing and failed to recirculate. To determine the function of lung TRMϕs, we used genetic mouse models and bone marrow chimeric mice to exclusively deplete AMϕ or AAMϕ following influenza infection. We show that AAMϕ cell numbers dramatically increased during infection due to local proliferation and differentiation of infiltrating monocytes. Depletion of CD169+ Mϕ in mice during influenza infection resulted in elevated morbidity. In the absence of CD169+ Mϕ, we observed dramatic changes in the recruitment of innate immune cells such as neutrophils and monocytes in the lungs following infection. Overall, our study relates structure to function at a fundamental level and provides essential insights into a novel subset of AAMϕ in the lung that along with alveolar Mϕ are crucial in mediating protection and maintaining immune homeostasis during influenza infection.
The impact of obesity onset on intraepithelial lymphocyte homeostasis at different ages.
Authors: Jameson Julie M , Cheung Kitty , Limon Natalie , Yee Christine , Costanzo Anne , Park Christa S 
Published: January 2023

Abstract
According to the Centers for Disease Control and Prevention, one in six children and adolescents in the United States are obese. The obesity epidemic is caused by chronic excess energy intake and low energy expenditure. This energy imbalance creates excess adipose tissue that produces systemic inflammation in the body. Obesity is detrimental to barrier tissues that protect the body from foreign pathogens and environmental stressors. This metabolic disease disrupts normal homeostasis within the epithelium, which then results in complications that include more severe inflammatory bowel disease. We have identified that as obesity progresses, T cell numbers are reduced within the epithelial layer of the intestine and proliferation of these immune cells in the tissue becomes dysfunctional. Interestingly enough, we show that these same T cells are partially restored in number with a low-fat diet. We examined the relationship between obesity onset and the ability of intraepithelial lymphocytes to seed and proliferate in the intestine to maintain homeostasis in the barrier tissue. Here we demonstrate that obesity is particularly devastating in young mice that are still establishing lymphocyte populations in their epithelia. In addition, the long-term effects of chronic obesity from childhood to adulthood shows a dramatic reduction in the number of intraepithelial lymphocytes in the intestine. This study helps shed light on how epithelial barrier maintenance differs in early versus late onset of obesity and will direct research to explore alternative approaches to restore intestinal intraepithelial T cells and promote optimal barrier integrity in obese patients.
Continuous, antigen independent re-seeding of lung resident memory CD8+ T-cells after Influenza A virus infection.
Authors: Slutter Bram , Budimir Natalija , Varga Steven M , Harty John T 
Published: January 2023

Abstract
Resident memory CD8+ T-cells (Trm) are a subset of memory T-cells, defined by their permanent localization in non-lymphoid tissues and are associated with potent protection against local infections. Lung Trm can mediate rapid control of respiratory pathogens like influenza A Virus (IAV), but in contrast to other non-lymphoid tissues like the skin, the number of lung Trm is not maintained long term. Here we observed through intranasal CFSE labeling that CD69+/CD103+ memory CD8 T-cell reside in the lung parenchyma longer then their CD69−/CD103− counterparts, but the average residence time of a Trm cell in the lung parenchyma is less than a week. We show that, in contrast to skin, Trm in the lung are prone to apoptosis and require continuous recruitment of memory CD8 T-cells from the circulation to maintain their numbers. Of note, the lung microenvironment drives the continuous conversion of circulating memory CD8+ T-cells into Trm, but does not require the presence of antigen in the lung. Moreover, intrinsic changes to the circulating memory CD8 T-cell population over time decrease their capacity to adopt a Trm phenotype. Thus our data could provide an explanation as to why lung Trm are not maintained long term.
Validating impedance-based methodology for in vitro dynamic analysis of reagents and effector cells efficacy in cancer immunotherapy protocols
Authors: Cerignoli Fabio , Guenther Garret , Xi Biao , Zhu Jenny , Zhao Leyna , Radecke Chris , Abassi Yama 
Published: January 2023

Abstract
In vitro characterization of the efficacy of reagents for cancer immunotherapy is a necessary step before moving to more expensive animal models and clinical studies. However, current in vitro assays are difficult to implement in high throughput environment and are mainly based on end point methodologies that are unable to capture the full dynamic of the immune response. Here we present the adaptation of an impedance-based platform to monitoring cytotoxic activity of immune cells activated trough different means. The technology detects cell death and proliferation of adherent cells by measuring changes in conductance of microelectrodes embedded in 96 and 384-wells cell culture plates. We utilized adherent cell lines and primary tumor cells with different expression levels of the EpCAM antigen, which is currently under evaluation as a target for cancer immunotherapy. Using PBMCs and NK92 as effector cells and combinations of EpCAM/CD3 BiTE and anti-EpCAM antibodies, we demonstrated the suitability of the impedance-based approach to quantitatively monitor the efficacy of immune cells-mediated cancer cell killing both under different effector:target ratios and antibody concentrations. We also validated the same approach with suspension cells immobilized on the surface of the electrodes. Overall, our results demonstrate the value of such approach in measuring the cytotoxic response across the temporal scale, an aspect that is otherwise very difficult to assess with more canonical end point assays. Furthermore, the availability of 384-wells format and minimal sample handling place the technology in an ideal spot for applications in personalized medicine, like therapeutic protocol validation directly on patient samples.
Flow Cytometry Multiplexing as a High Throughput Screening Method to Analyze Metabolic Activity for Pharmaceutical Drugs
Authors: Low Quentin , Anderson April , Calderon Veronica , Clark Scott , DeMarco Carolyn , Finnessy Carmen , Kihn Kathy , O’Grady Michael , Wickett Marcy , Yan Michelle 
Published: January 2023

Abstract
High throughput screening (HTS) is an extremely effective method that allows researchers to identify putative active compounds for therapeutics. Recently, flow cytometry has emerged as a powerful HTS tool with the added benefit of cell-by-cell analysis. Flow cytometry is a fast and sensitive platform aiding in the study of protein-protein interactions, metabolic activities, as well as DNA content in a single or multi-parametric assay format. For this study, we screened several compound libraries and compared the results using a plate reader with flow cytometer multiplexing.
Peripheral blood mononuclear (PBMCs) and Jurkat cells were used as cell models. Cells were cultured under standard conditions and either at normoxic (≤5%) or hyperoxic (≥19%) oxygen levels for varied lengths of time; 24, 48, or 72 hrs. Membrane integrity and metabolic activity were measured as an output for evaluating cellular viability. To further assess cellular activity, post-screening analysis was implemented to establish a drug dose-response and EC50s of “hits” from compound library.
Screening analysis identified target drugs that were taken on to further tests using different mechanisms of action to assess compound toxicity. Results indicate that compound “hits” and potency differed in the screen depending on cell type, the mechanistic readout, length of compound exposure, and mode of readout used to perform the HTS experiments.
Early diagnostics of septicemia using microRNA flow cytometry
Authors: Stein Erica Vanessa , He Hua-Jun , Cole Kenneth D , Wang Lili 
Published: January 2023

Abstract
From 2000 to 2008 the incidence of sepsis in hospitals increased by 83%, but we still lack effective diagnostic methods. Recent advances in probe chemistry have enabled scientists to better detect RNA using flow cytometry. This technique could potentially be used for acute and efficient detection of sepsis. However, it still requires substantial validation before it reaches full potential in the clinic.
We are in the process of creating a robust microRNA flow cytometry assay that can be used in the clinic for early detection of sepsis. In order to do this, first a reference control and diagnostic microRNA biomarkers, specific for detection of septicemia, must be identified and validated. Next, a microRNA flow cytometry assay will be developed and validated with the use of the reference control and diagnostic microRNAs both intracellularly and in serum. Lastly, the developed flow cytometry assay, including controls and diagnostic biomarkers will be verified against blood samples of clinical patients both positive and negative for sepsis.
An effective sepsis diagnostic will save lives, aid in drug discovery, and decrease health care costs. This project provides a unique opportunity to create a robust flow cytometry assay to measure expression differences in microRNAs to detect septicemia in the clinic. We are hopeful that our continued efforts to validate and standardize previously published microRNA detection assays and expand the possible new targets will lead to quicker and more robust methods to survey for sepsis in clinics.
A Framework for Meta-Analysis of Cytometry Data
Authors: Jujjavarapu Chethan , Hughey Jacob , Gherardini Federico , Bruggner Robert , Nolan Garry , Bhattacharya Sanchita , Butte Atul 
Published: January 2023

Abstract
Flow cytometry has been used to analyze cell populations with a certain condition, but because of the limitations of a single flow cytometry experiment, results only present a small portion of cell populations. Meta-analysis has been used to compare and contrast multiple datasets in hopes of discovering new insights. Scientists can then identify more cell populations linked to the condition. Therefore, we are developing a method to standardize and analyze multiple flow cytometry datasets.
89Zr positron emission tomography detects migration of donor cells to the ileum in a mouse bone marrow transplantation model.
Authors: Sato Noriko , Asiedu Kingsley O. , Choyke Peter L. 
Published: January 2023

Abstract
In hematopoietic stem cell (HSC) transplantation, the efficacy of bone marrow homing and engraftment of HSCs directly correlates with the success of treatment. At the same time, reduction of graft-versus-host-disease (GVHD) incidence would be critical. Development of a non-invasive, real time monitoring system for the donor cells would be important for optimizing the HSC transplantation conditions. Recently, we have developed a 89Zr-oxine complex cell labeling technique that enables visualization of cell trafficking at a high sensitivity using positron emission tomography (PET)/computed tomography (CT) imaging. We employed this technique to monitor bone marrow transplantations in mice. PET/CT imaging visualized that 89Zr-labeled donor bone marrow cells rapidly migrated to the bone marrow of the recipient, as well as to the spleen and liver. More than 20% of the donor cells homed to the bone marrow within 4 hours, irrespective of prior bone marrow ablation. While monitoring the donor cell homing, we have found that a small fraction of cells migrating to the intestine. This was observed only when the recipient mice had undergone a conditioning by a whole-body irradiation and the cells targeted the distal part of the small intestine. The most of the cells expressed CD45 and CD103. These cells could be the cause of GVHD in case of allogeneic transplantation. 89Zr PET/CT would be useful to monitor donor cell trafficking in HSC transplantations.