bluuebunny/crossref_metadata_2025_split huggingface dataset:
SPA4 peptide suppresses the lipopolysaccharide (LPS)-stimulated lactate secretion in lung epithelial cells through modulation of Toll-like receptor-4 (TLR4)
Authors: Godbole Nachiket Madan , Awasthi Shanjana 
Published: August 2023

Abstract
Lung infection and inflammation can cause severe injury leading to acute respiratory distress syndrome. An increased expression and activity of the TLR4 is associated with inflammatory response. Simultaneous elevated level of lactate is used as the marker of disease severity and is associated with poor outcomes in critically ill patients. Therapeutic treatment with TLR4-interacting SPA4 peptide (amino acid sequence: GDFRYSDGTPVNYTNWYRGE) suppresses inflammation in immune cells and in mouse models of lung infection and inflammation. In this study, we evaluated the effect of SPA4 peptide on lactate production by H441 human lung epithelial cells against Pseudomonas aeruginosa-derived LPS. Briefly, the H441 cells were challenged with LPS (1 μg/ml) and treated with SPA4 peptide (100 μM) after 1 h of LPS challenge. Next day, the cell-free supernatants and cell lysates were harvested. We also transfected the cells with TLR4 siRNA to achieve suppression of TLR4. Levels of protein and lactate were quantitated in cell lysates and in filtered supernatants using biochemical assays. The lactate levels were normalized with cell protein. The unchallenged, vehicle-treated cells served as control. Our results demonstrated reduced secretion of lactate by SPA4 peptide-treated cells (187.5% of control) versus vehicle-treated cells (324.7% of control) against LPS stimuli (p&lt;0.05). The immunostaining and confocal imaging showed knockdown of TLR4 in up to 50% cells. The LPS-stimulated lactate secretion was reduced and SPA4 peptide treatment did not further affect the lactate levels in TLR4 siRNA transfected cells. Together, our results suggest that the SPA4 peptide suppresses lactate secretion through modulation of TLR4 in lung epithelial cells.
Supported by grant from NIH (R01HL136325) and 2021 American Association of Immunologists Careers in Immunology Fellowship Program.
The BNT162b2 vaccine’s empty lipid nanoparticle is able to induce an NF-κB response
Authors: Zelkoski Amanda E , Ennis Katherine H E , Lu Zhongyan , Malloy Allison M W 
Published: August 2023

Abstract
Lipid nanoparticles (LNP) are essential components of messenger RNA (mRNA) vaccines that have been a cornerstone of the COVID-19 public health response. These LNPs are used for cell entry and protection of the mRNA. Others have shown that the LNP of the BNT162b2 vaccine is able to augment the immune response against the vaccine target. However, it is unknown how this LNP stimulates innate immune cells such as monocytes that participate in the engagement of the adaptive immune response.
We used a THP-1 monocyte reporter cell line for nuclear factor kappa B (NF-κB) and interferon regulatory factor (IRF) activation to understand if empty LNPs could stimulate innate cells in comparison with toll-like receptor (TLR) agonists. With this cell line, we show that the BNT162b2’s LNP is able to initiate activation of NF-κB, but not IRF in a dose-dependent manner. The NF-κB response was similar to that of low doses of R848, a TLR 7/8 agonist, as well as LPS, a TLR4 agonist. Next, we evaluated NF-kB activation in the absence of MyD88 or TRIF using knockout reporter cell lines. We demonstrate that NF-κB activation is reduced in both knockout cell lines. In comparison, LPS primarily stimulated NF-kB activation through MyD88, and R848 relies on both pathways.
Our data show that LNPs can stimulate key innate immune cells through NF-kB pathways but not through IRF suggesting that this master regulator may participate in the immune activation induced by the current mRNA vaccine platform. Moreover, our data suggest that LNPs can use both MyD88 and TRIF signaling cascades to induce activation. Understanding the mechanisms behind the immunostimulatory capacity of LNPs can shed light on modulating these vaccine components to enhance vaccine design.
Defense Health Program, HU00012120094
The transcriptional repressor HIC1 is required for the maintenance of tissue-resident ILC1
Authors: Rodrigues Grace , Doan Anh , Camilleri Tania , Huntington Nicholas , Scheer Sebastian , Schuster Iona , Degli-Esposti Mariapia A , Zaph Colby 
Published: August 2023

Abstract
The group 1 innate lymphoid cell compartment is comprised of conventional natural killer (cNK) and type 1 innate lymphoid cells (ILC1). Within the innate lymphoid cell pool, these cells are delineated by expression of the transcription factor TBET, and both subtypes express cell surface receptors NK1.1 and NKp46. Functionally, ILC1 and cNK cells have been shown to mediate pathogen clearance and tumour control through release of type 1 cytokines, particularly IFN-□. While being closely related, a key feature distinguishing ILC1 from cNK cells is their propensity to reside within a tissue niche rather than enter the circulation. However, the precise mechanisms that regulate tissue residency of ILC1s remain unknown. Here, we show that the transcription factor Hypermethylated in Cancer 1 (HIC1) is a critical control point in the development and maintenance of ILC1. In the liver, HIC1 expression is restricted to the ILC1 compartment. Deletion of HIC1 in NKp46+ cells (Hic1ΔNcr1 mice) resulted in a specific and significant reduction in liver-resident ILC1, while cNK cells were unaffected. Additionally, we show that cNK cells isolated from Hic1ΔNcr1 mice are incapable of restoring the ILC1 compartment in lymphocyte-deficient hosts, unlike wild type donors. Thus, our results identify an important role for HIC1 in the development and maintenance of peripheral ILC1.
The role of MATRIN3 (MATR3) in innate immune response
Authors: Islam Zohirul , Chim Bryan , Suzawa Masataka , Cutrona Nicholas , Smith Patrick T , Foroushani Amir K , Hafner Markus , Muljo Stefan A 
Published: August 2023

Abstract
MATR3, one of the most abundant inner nuclear matrix proteins, is implicated in amyotrophic lateral sclerosis (ALS) and distal myopathy. MATR3 has been reported to have both DNA- and RNA-binding abilities. However, it is not understood mechanistically how mutation of MATR3 leads to ALS pathogenesis. To understand its loss of function effects, we mutated MATR3 in the human HAP1 cell line using CRISPR/Cas9 technology. Using two different RNA sequencing methods, Oxford Nanopore Technologies and Illumina, we discovered the upregulation of many interferon-stimulated genes (ISGs) in MATR3-deficient HAP1 cells. Furthermore, we have validated these results by Nanostring technology. Next, we hypothesized that HAP1 cells may be responding to endogenous double-stranded RNAs. Interestingly, we observed similar amounts of dsRNA in both MATR3 knock-out (KO) and wildtype (WT) cells which suggests that indeed intracellular dsRNAs are present and could be stimulating HAP1 cells; however, it is not the case that MATR3 KO cells are stimulated because they are producing significantly more dsRNAs. Currently, we are investigating whether and which RNA sensors are activated in MATR3 KO cells. Also, we are investigating whether MATR3’s DNA- and/or RNA-binding abilities are responsible for the innate immune response. Accordingly, we plan to perform CUT & RUN and photoactivatable ribonucleoside enhanced crosslinking and immunoprecipitation (PAR-CLIP) respectively, in HAP1 cells. Altogether, our results illustrate that MATR3 is a novel negative regulator of the innate immune response. This work was supported by the Intramural Research Program of NIAID, NIH.
This work was supported by the Intramural Research Program of NIAID, NIH.
BCG component mycolic acid protects against monocyte exhaustion in vitro
Authors: Wu Yajun , Wang Jing , Li Liwu 
Published: August 2023

Abstract
Monocyte exhaustion reflected in pathogenic inflammation and immune suppression underlies the pathogenesis of sepsis. The objective of this study is to define novel approaches in alleviating monocyte exhaustion, based on intriguing clinical reports that BCG components may be beneficial in reducing polymicrobial infection-triggered sepsis severity. We recapitulated in vitro the development of exhausted murine monocytes through prolonged culture with high dose bacterial lipopolysaccharide (LPS), measured by the expansion of immature Ly6Chi population with elevated expression of pathogenic mediator CD38 and immune suppressor PD-L1, and reduced expression of CD86. Intriguingly, we observed that co-culture of murine bone marrow monocytes with Mycobacterium tuberculosis (BCG) component mycolic acid (MA) can significantly alleviate the development of exhausted monocytes. We found that monocytes co-stimulated with MA and LPS exhibited significantly reduced induction of CD38 and PD-L1 as compared to monocytes challenged with LPS alone. On the other hand, monocytes co-treated with MA and LPS exhibited enhanced expression of CD86 as compared to cells treated with LPS. In the monocyte-T cell coculture system, we found that T cells co-cultured with MA and LPS-treated monocytes had a significantly higher proliferation potential as compared to the T cells co-cultured with LPS-treated monocytes. Mechanically, we demonstrated that MA exerts its protective effect by selectively activating P-AKT and inhibiting STAT1/P-STAT1. In conclusion, we demonstrate that the BCG component mycolic acid can potently alleviate monocyte exhaustion and may serve as a future promising agent in treating sepsis.
Supported by grants from NIH (AI136386)
The glucose transporter GLUT4 tethers RIG-I-like receptors to suppress antiviral immunity
Authors: Harrison Andrew G , Yang Duomeng , Cahoon Jason G , Geng Tingting , Karginov Timofey A , Torrance Blake L , Chiari Conner C , Haynes Laura , Vella Anthony T , Witczak Carol A , Bogan Jonathan S , Wang Penghua 
Published: August 2023

Abstract
The current dogma of RIG-I-like receptor (RLR) regulation states that dysregulated interferon (IFN) production is predominantly repressed by reversible posttranslational modifications (e.g., ubiquitination). However, RLR activation has profound outcomes on cellular metabolism, which governs the nature of these inflammatory immune responses. Here, we reveal a novel mechanism of immunometabolic regulation of RLR signaling by the glucose transporter 4 (GLUT4). In addition to its canonical role in maintaining organismal glucose homeostasis, GLUT4 attenuates innate immune signaling through sequestering RLRs into the plasma membrane. Translocation of GLUT4 from intracellular compartments to the plasma membrane is tightly regulated by UBXN9, a ubiquitin-domain containing protein (UBXN). Disruption of UBXN9 releases GLUT4, suppresses RLR signaling/IFN responses and enhances viral replication, while genetic ablation of GLUT4 improves antiviral immunity. Strikingly, these distinct phenotypes are independent of glycolysis, but dictated by the mobilization of GLUT4 to the plasma membrane. Further, GLUT4 colocalizes with RIG-I after insulin treatment (which stimulates GLUT4 trafficking) or viral infection to blunt RLR activation. Fine-tuning of this UBXN9-GLUT4 axis is critical for antiviral immunity, but dysregulation of these components may underlie inflammatory myopathies that are characterized by a hyperactive RLR-IFN pathway. Together, this study reveals a novel link between GLUTs and cytosolic immune sensors, and underscores the intersection of vesicular trafficking, glucose metabolism and antiviral immunity.
Supported by grants from NIH (NIH R01AI132526, R21AI155820)
Upping the “Anti-” for arthritis: Antimicrobial defenses through anti-inflammatory NOD-like Receptor NLRX1 during Lyme disease
Authors: Tupik Juselyn D , McClune Mecaila E , Camp Hailey W , Gregory Julia A , Dressler Jules M , Markov Madanick Justin W , Nagai-Singer Margaret A , Jutras Brandon L , Allen Irving C 
Published: August 2023

Abstract
Lyme disease, caused by the bacterium Borrelia burgdorferi, is an emerging infectious disease of global concern. Roughly 60% of untreated patients will develop synovitis of the joints termed Lyme arthritis, resulting from sustained cytokine and chemokine production by the innate immune system. Currently, there are limited therapeutics for antibiotic-refractory Lyme arthritis, warranting investigation into how our innate immune system can mitigate this inflammation. Here, we studied how the anti-inflammatory NOD-like Receptor (NLR) NLRX1 could play a role in controlling Borrelia infection. Because NLRX1 modulates inflammatory signaling, cell death, autophagy, and metabolism, we hypothesized that NLRX1 could have unique antimicrobial effects against Lyme disease. Using novel Nlrx1−/− mice, we found that NLRX1 significantly decreased arthritis severity in wildtype mice compared to knockouts and modulated bacterial load in vivo. We next determined in Nlrx1−/− murine BMDMs that NLRX1 may control infection by promoting Reactive Oxygen Species (ROS) mediated-cell death and autophagy, while decreasing cell proliferation and modulating inflammasome activation in vitro. Finally, by infecting novel NLRX1 overexpression human monocytes, we found that elevated NLRX1 significantly decreased pro-inflammatory NF-κB-mediated cytokine secretion. Ultimately, these results indicate that NLRX1 plays a protective role in mitigating Lyme arthritis in murine and human models. Further, this protection may occur through NLRX1’s modulation of cell death and attenuation of NF-κB signaling. Consequently, these results warrant further exploration of Borrelia regulation by NLRX1 for development of new treatments for Lyme arthritis.
Supported by grants from the NIH/NIAID (R21AI159800) and the Cohen Foundation
Biphasic JNK signaling reveals distinct MAP3K complexes licensing inflammasome formation and pyroptosis
Authors: Bradfield Clinton J , Liang Jonathan J , Ernst Orna , John Sinu P , Sun Jing , Ganesan Sundar , de Jesus Adriana A , Bryant Clare E , Goldbach-Mansky Raphaela , Fraser Iain D C 
Published: August 2023

Abstract
Kinase signaling in the tiered activation of inflammasomes and associated pyroptosis is a prime therapeutic target for inflammatory diseases. While MAPKs subsume pivotal roles during inflammasome priming, specifically the MAP3K7/JNK1/NLRP3 licensing axis, their involvement in successive steps of inflammasome activation is poorly defined. Using live–cell MAPK biosensors to focus on the inflammasome triggering event allowed us to identify a subsequent biphasic JNK activation. We find that post–trigger JNK signaling distinctly facilitates both mitochondrial reactive oxygen species generation needed to support core inflammasome formation, and gasdermin–mediated cell permeation leading to release of active IL–1β from human macrophages. We further identify and characterize a xanthine oxidase–ROS activated MAP3K5/JNK2 substrate licensing complex as a novel regulator of GSDMD mobilization preceding pyroptosis. We find that inhibitors targeting this MAP3K5 cascade alleviate morbidity in mouse models of colitis and dampen both augmented IL–1β release and cell permeation in monocytes derived from patients with gain–of–function inflammasomopathies.
This work was supported by the Intramural Research Program of NIAID, NIH.
Characterization of Neuro-Immune Communication in the White Pulp Region of the Spleen
Authors: Keller Alexandra , Ross Ashley 
Published: August 2023

Abstract
Norepinephrine (NE) is a ubiquitous neurotransmitter that facilitates cellular communication in the sympathetic nervous system. Recent advancements have shown sympathetic neurons innervate secondary immune organs, such as the spleen, to control immune physiology. Fast-scan cyclic voltammetry (FSCV) is an electrochemical technique that varies an applied potential at a very fast scan rate with micro-electrodes to detect neurotransmitters with high temporal and spatial resolution. FSCV can be used to detect the sub-second release of NE in an ex vivo murine slice model to study the mechanisms related to neuro-immune communication. Our lab is the first to characterize and quantify this spontaneous transient signaling of catecholamines in the white pulp region of the murine spleen with FSCV. The average concentration of catecholamines released is 581 ± 20 nM, with an average inter-event time of 23.4 ± 0.7 s and a duration of 6.6 ± 0.1 s. Norepinephrine transporter (NET) is known to transport NE back into cells after endogenous release. We blocked NET with the inhibitor nomifensine to provide further evidence of NE release and study the mechanisms involved in NE uptake. To study the mechanisms of NE release in the spleen, we blocked α2-adrenoceptor, a known autoreceptor only found on neurons. Yohimbine is an α2-adrenoceptor antagonist, and when applied to slices, it increased catecholamine concentration and duration, demonstrating that we are detecting neuronal release of NE. Furthermore, we used ELISAs to quantify the extent to which the α2-adrenoceptor affects cytokine production, providing further evidence neuronal-regulated immune signaling in the white pulp region of the spleen.
Supported by grants from NIH (R01AI151552, R01NS121426)
Bi-directional epithelial-immune communication amplifies inflammasome activation and IL-6 release during SARS-CoV-2 infection
Authors: Barnett Katherine Camille , Xie Yuying , Asakura Takanori , Song Dingka , Liang Kaixin , Taft-Benz Sharon A , Guo Haitao , Yang Shuangshuang , Okuda Kenichi , Gilmore Rodney C , Loome Jennifer F , Oguin Thomas , Sempowski Gregory D , Randell Scott H , Heise Mark T , Lei Yu Leo , Boucher Richard C , Ting Jenny P 
Published: August 2023

Abstract
Increased levels of the pro-inflammatory cytokines IL-1β and IL-6 are linked to severe COVID-19. Here, we investigate the contributions of viral host cells, primary human airway epithelia (HAE), and immune bystanders, peripheral blood mononuclear cells (PBMCs), in the production of these cytokines. Despite having functional inflammasomes and supporting viral replication, HAE do not secrete IL-1β upon SARS-CoV-2 infection. PBMCs do not support productive viral replication, but the SARS-CoV-2 envelope (E) protein primes inflammasomes through stimulation of TLR2. To activate inflammasomes in PBMCs, SARS-CoV-2-infected HAE provide a second signal in the form of damage-associated molecular patterns (DAMPs), which includes mitochondrial and genomic DNA. Co-culture of PBMCs with infected epithelial cells stimulates IL-1β release, while addition of benzonase, STING, or caspase-1 inhibitors, but not NLRP3 inhibitors, abrogates this response. In turn, IL-1β stimulates IL-6 secretion from both populations with a robust response from HAE. Thus, while infection in either cell type in isolation does not yield IL-1β or IL-6, immune bystander detection of SARS-CoV-2 infection in the epithelium stimulates both IL-1β and IL-6 production, amplifying pro-inflammatory cytokine production through cell-cell communication. This circuit is reflected in patient data, as COVID-19 autopsy lungs show hallmarks of inflammasome priming and activation in myeloid populations lacking SARS-CoV-2 antigens. Meta-analysis of single cell RNA sequencing (scRNAseq) data reveals discordant, exacerbated priming of inflammasomes in myeloid cells from severe but not mild COVID-19 patients, indicating this circuit is intensified in severe disease.
Supported by grants from NIH R56 AI158314, R01 AI158314, AI029564, AI141333 and CA232109 to J.P.-Y.T; NCI T32CA071341 to K.C.B.; UH3 HL123645, R01 HL136961, P30 DK 065988, P01 HL108808, and from the Cystic Fibrosis Foundation (CFF) BOUCHE19R0 to T.A. and R.C.B; CFF grant BOUCHE19XX0 to R.C.B. and K.O.; CFF grant OKUDA20G0 KO; NC Policy Collaboratory, NIH R01AI157253, and U19 AI100625 to M. T. H.; NSF GRFP 2019255197 to J.F.L. T.A. is a Japan Society for the Promotion of Science Overseas Research Fellow. Y.X. is supported by NSF DMS-191290 and NSF IOS 2107215. Y.X and Y.L.L. are supported by NIH grants R01DE026728 and R01DE031951. Some BSL3 work was performed in the Duke Regional Biocontainment Laboratory (Duke RBL) which received partial support for construction from NIH/NIAID (UC6AI058607).
Scavenger Receptor CD36 Regulates Inflammatory and Type-I Interferon Responses in Tumor-Associated Macrophages
Authors: Xu Ziyan Zoe , Kaech Susan 
Published: August 2023

Abstract
Macrophages are plastic cells of innate immune system with diverse functions in tissue homeostasis. Despite their abundance in the tumor microenvironment (TME), the functions of tumor-associated macrophages (TAMs) and their regulation mechanisms remain largely unknown. Macrophages are also key players in metabolic processes including lipid metabolism. As TME has been known as a “lipid-rich” environment, we are interested in understanding how TAMs’ metabolism and function are regulated by lipid receptors.
In this research, we firstly characterized the lipid metabolic phenotype of TAMs in mouse tumor models. We found the tolerogenic subset of TAM (high expression of F4/80 and PD-L1) is lipid-laden and has greater ability to import lipids. We found F4/80hi TAMs highly express scavenger receptor CD36 which binds to oxidized low-density lipoprotein (oxLDL) in TME. Using myeloid-specific knockout (Cd36flox/flox x Csf1r-Cre) mouse models, we observed that CD36-deficient TAMs lost binding to oxLDL, down-regulated expression of immunosuppressive molecules PD-L1 and CD206, while up-regulated secretion of inflammatory cytokines (TNF, IL-12, IL-1b) and IFNb. Overall, this reprogrammed TAM functional state leads to better anti-tumor immunity. Mechanistically, we conducted single-cell RNA-sequencing and found inflammatory response and type-I interferon response pathways were up-regulated in CD36-deficient TAMs. Furthermore, we showed that IFNb and P38 play critical roles in CD36-dependent immune regulations.
In summary, our data suggests that scavenger receptor CD36 plays critical roles in functional reprogramming of TAMs; and blocking CD36 may have multi-faceted beneficial effects for future cancer immunotherapies.
Supported by fellowship from American Heart Association
Infection with LCMV delays the progression of demyelinating disease in EAE
Authors: Essaghir Ibtissam , Cosby Jennifer , Rase Viva Jeanne , Evavold Brian D 
Published: August 2023

Abstract
Multiple sclerosis is a chronic autoimmune demyelinating disease that affects approximately 2.8 million people worldwide. While several preventative treatments for multiple sclerosis, including Natalizumab and Mitoxantrone exist, there currently is no cure. It is also unknown what provides the initial trigger of disease episodes or exacerbation of the ongoing disease although viral infections might play a role. In contrast to their role as a driver of demyelination, here we investigate whether viral infections play a potential protective role. Using a well-established animal model of chronic-progressive autoimmune disease known as Experimental Autoimmune Encephalomyelitis, we found that infection with lymphocytic choriomeningitis virus (LCMV) infection delayed onset of paralysis as compared to uninfected controls. Consistent with the reduction in EAE disease scores, we identified reduced numbers of myelin-specific CD4 T cells in the CNS. The myelin-specific CD4 T cells were found in the periphery suggesting that the virus impeded their trafficking to the CNS. Additionally, we analyzed that virus-specific CD8 T cells were found in the CNS in preference to the myelin-specific CD4 T cells. Thus, in contrast to the prevailing paradigm of infections driving autoimmune disease progression, we find that viral infection can lessen acute demyelinating disease.
R01NS071518 R01AI147641