bluuebunny/crossref_metadata_2025_split huggingface dataset:
Antibacterial resistance of severely burned mice treated with an RORα selective inverse agonist
Authors: Ito Ichiaki , Kobayashi Makiko , Finnerty Celeste C. , Herndon David N. , Suzuki Fujio 
Published: January 2023

Abstract
In our previous studies, an antibacterial resistance of severely burned mice to enterococcal translocation has been improved by treatment with RORα selective agonist SR3335 (an inhibitor of ILC2 activation). Therefore, the effect of SR3335 against infections with P. aeruginosa (PA) and MRSA was studied in severely burned mice. Severely burned mice (30% TBSA burn) infected with PA (103 CFU/mouse, i.d.) or MRSA (106 CFU/mouse, i.v.) 3 days post burn injury were treated i.p. with 100 μg/mouse of SR3335 simultaneously, 1 and 2 days after the infection. The antibacterial effect of SR3335 was evaluated by bacterial growth in organs of these mice 3-5 days post-infection. In some experiments, lamina propria mononuclear cells (LPMC) were isolated from burned mice treated with or without SR3335, and analyzed for lin−CD127+ICOS+Sca-1+ cells (ILC2), F4/80+CD11b+IL-12+CD38+ cells (M1Mϕ), F4/80+CD11b+CD206+ cells (M2Mϕ), and Gr-1+CD11b+ cells (MDSC) by flow cytometry. In the results, PA and MRSA vigorously grew in various organs of burned mice. However, both of the pathogens did not grow in organs of the same mice treated with SR3335. The number of ILC2 increased greatly in the LPMC of burned mice, while this increase was not shown in these mice treated with SR3335. The numbers of M2Mϕ and MDSC increased in the LPMC of burned mice, while the numbers of these cells were not increased in the same mice treated with the compound. M1Mϕ were induced by CpG DNA in cultures of LPMC isolated from burned mice treated with SR3335. These results suggest that SR3335 improves the antibacterial resistance of severely burned mice against PA and MRSA through the modification of burn-associated appearance of ILC2, M2Mϕ, and MDSC.
Defective innate immune responses lead to hepatic damage in CD154-deficient mice following infection with attenuated Salmonella typhimurium
Authors: Fernandez-Cabezudo Maria J. , Bashir Ghada , Al-Sbiei Ashraf , Cabral-Marques Otavio , al-Ramadi Basel K. 
Published: January 2023

Abstract
In addition to its established role in immunoglobulin isotype switching, CD40–CD154 interaction plays a role in the regulation of multiple innate immune system cell types. Of note, we have shown that CD154-deficient (CD154−/−) mice are hyper-susceptible to intraperitoneal infection by attenuated strains of Salmonella enterica serovar Typhimurium. Compared to their normal counterparts, CD154−/− mice exhibited progressive morbidity during the early phases of disease and succumbed to infection 3–4 weeks after inoculation. Since the liver is a key target organ in Salmonella infections, we carried out a comparative analysis of infection-associated hepatic changes at different time-points. Bacterial loads in the liver were significantly higher in CD154−/− compared to WT mice, and this correlated with progressively worsening hepatic damage and rising levels of liver enzymes. Liver histomorphology revealed focal necrosis early in the infection that was self-limiting in WT mice. In contrast, infected CD154−/− mice developed large and generalized necrosis coupled with the appearance of large venous thrombi. Immunohistochemical analysis showed a retarded inflammatory response affecting recruitment of myeloid cells in the liver of CD154−/− animals. Moreover, neutrophil recruitment into the peritoneal cavity, as well as the activation of the respiratory burst, was defective in CD154−/− mice. Taken together, our findings demonstrate that CD154-deficiency results in a failure to mount an appropriate innate immune response to contain bacterial proliferation during the acute phase of disease. This renders the animals incapable of limiting the extent of histopathological damage in the liver resulting, ultimately, in their demise.
Blocking CXCL1-dependent neutrophil recruitment prevents immune damage and reduces pulmonary bacterial infection after inhalation injury.
Authors: Maile Robert , Dunn Julia L. , Kartchner Laurel B. , Jones Samuel W. , Cairns Bruce A. 
Published: January 2023

Abstract
Smoke inhalation associated with structural fires, wildfires, or explosions leads to lung injury. Clinically relevant animal models are needed to develop therapeutics. We have previously reported that damage associated molecular patterns (DAMPs) and anti-inflammatory cytokines correlate with infectious complications in patients diagnosed with inhalational injury. We describe a novel translational murine model of acute inhalational injury characterized by an accumulation of protein and neutrophils in the bronchoalveolar space, and histological evidence of tissue damage. Mice were anesthetized, a cannula placed in the trachea, and exposed to smoldering plywood smoke three times for two minute intervals. Here we demonstrate that this model recapitulates clinically relevant phenotypes including early release of dsDNA, IL-10, MCP-1, and CXCL1 along with neutrophilia early after injury, accompanied by subsequent susceptibility to opportunistic infection with Pseudomonas aeruginosa. The mouse model, and a re-analysis of patient samples, revealed a late release of the DAMP hyaluronic acid (HA) from the lung. Using nitric oxide synthase deficient (Nos2−/−) mice, we found that Nos2 was required for increases in IL-10, MCP-1, and HA following injury but not release of dsDNA, CXCL1 expression, early neutrophilia, nor susceptibility to opportunistic infection. Depletion of CXCL1 attenuated early neutrophil recruitment, leading to decreased histopathology scores and improved bacterial clearance in this model of smoke inhalation compared to untreated injured mice. These data highlight the potential therapeutic benefit of attenuating neutrophil recruitment in the first 24h after injury in patients.
IL-21 regulates NK cell responses during Mycobacterium tuberculosis infection. Padmaja Paidipally, Deepak Tripathi, Abhinav Van, Rajesh Kumar Radhakrishnan, Ramakrishna Vankayalapati1
Authors: Paidipally Padmaja 
Published: January 2023

Abstract
In the current study, we determined the effects of interleukin (IL)-21 on human natural killer (NK) cells and monocyte responses during Mycobacterium tuberculosis (Mtb) infection. We found that Mtb stimulated CD4+ and NKT cells from healthy individuals with latent tuberculosis infection (LTBI+) are major sources of IL-21. CD4+ cells from tuberculosis patients secreted less IL-21 than did CD4+ cells from healthy LTBI+ individuals. IL-21 had no direct effect on Mtb stimulated monocytes. IL-21 activated NK cells produced IFN-γ, perforin, granzyme B and granulysin; lysed Mtb infected monocytes; and reduced Mtb growth. IL-21 activated NK cells also enhanced IL-1β, IL-18, CCL4/MIP-1β production and reduced IL-10 production by Mtb stimulated monocytes. Recombinant IL-21 inhibited Mtb growth, enhanced IFN-γ, IL-1β, IL-18 and MIP-1β; and reduced IL-10 expression in the lungs of Mtb-infected Rag2 knockout (Rag2 KO) mice. These findings suggest that activated T cells enhance NK cell responses to lyse Mtb infected human monocytes and restrict Mtb growth in monocytes through IL-21 production. IL-21 activated NK cells also enhances the immune response by augmenting IL-1β, IL-18 and MIP-1β production and reducing IL-10 production by monocytes in response to an intracellular pathogen.
ASC phosphorylation at Y146 regulates inflammasome activation and pyroptosis
Authors: McAndrew Christian C. , Napolitano Nina M. , Vompe Alex D. , Wewers Mark D. , Gavrilin Mikhail A. 
Published: January 2023

Abstract

Rationale
Inflammasome assembly and activation is a complex process regulated, in part, by posttranslational modification of inflammasome proteins. ASC is a universal adaptor in virtually all inflammasome platforms and its modification is critical in regulation of inflammasome function. In particular, PTK inhibitor AG126 greatly inhibits inflammasome activation.

Methods
We generated stable THP-1 cells with either overexpression of ASC (YFP-ASC) or knock down of ASC (siRNA and CRISPR) followed by ASC knock in. To test the role of ASC phosphorylation in inflammasome assembly and activity, we mutated the highly conserved tyrosine 146 to Y146A. Cells were stimulated with LPS or live bacteria and inflammasome activity was measured by fluorescent microscopy, ELISA, and immunoblotting.

Results
Upon inflammasome activation ASC is released from the cell together with IL-1β, IL-18 and other inflammasome proteins. Inhibition of select tyrosine kinases severely reduces inflammasome activation, pyroptosis and ASC release. Elimination of the potential phosphorylation site Y146 dramatically changes visible ASC complex formation – from a single round speck to a filamentous structure. Cells with the Y146A ASC mutation show significantly abrogated inflammasome activation as compared to a wild type. Screening of a PTK inhibitor library (Selleckchem) linked several tyrosine kinases to regulating ASC phosphorylation and inflammasome activity.

Conclusion
ASC phosphorylation at Y146 by tyrosine kinases is an important regulator of inflammasome activity and pyroptosis.
Lymphocyte-specific protein tyrosine kinase (LCK) is involved in the aryl hydrocarbon receptor (AHR)-mediated impairment of Immunoglobulin secretion in human primary B cells
Authors: Zhou Jiajun , Henriquz Joseph , Crawford Robert , Kaminski Norbert E. 
Published: January 2023

Abstract
AHR activation by the high affinity ligand, TCDD, is widely established to suppress the immunoglobulin M (IgM) response in virtually every animal species tested, and extensively investigated in various mouse models. In mice, activation of AHR is known to impair B cell to plasma cell differentiation and IgM synthesis. In contrast to mouse B cells, activation of AHR in human primary B cells impairs immunoglobulin secretion in the absence of suppressing IgM synthesis. In recent studies, we have identified the putative involvement of LCK in impaired immunoglobulin secretion by human B cells. LCK is a well-characterized tyrosine kinase that phosphorylates known critical signaling proteins involved in vesicular secretion by T cells. Specifically, phosphorylation of tyrosine residue 505 inhibits the activity of LCK. By contrast, little is known concerning the role of LCK in human primary B cells. For the first time, our studies show that activation of the AHR by TCDD upregulates LCK protein expression, which then leads to an impairment of IgM secretion. Treatment with a LCK specific inhibitor restores IgM secretion by human primary B cells. Additionally, the presence of AHR antagonist reverses the AHR-mediated increase of LCK and the impairment of IgM secretion. We also observe a significant increase in phosphorylation of Tyr-505 LCK with TCDD treatment, indicating that AHR activation increases the level of inhibitory LCK. Taken together, our studies revealed a novel and species-dependent mechanism involving the AHR-mediated impairment of IgM secretion and an increase in total as well as inhibitory LCK in human but not mouse primary B cells.
IL-1 and IL-33 differentially regulate the functional specialization of mucosal Foxp3+ regulatory T cells.
Authors: Alvarez Fernando , Fritz Jorg H. , Piccirillo Ciriaco A. 
Published: January 2023

Abstract
CD4+ regulatory T (TREG) cells are critical mediators of peripheral immune tolerance and homeostasis, and express the forkhead box p3 (Foxp3) transcription factor, the master-regulator driving the programming of the TREG cell suppressive phenotype. TREG cells are abundant at mucosal surfaces, where they acquire tissue-specific adaptations. The biological consequences of these adaptations on the stability of thymic-derived TREG (tTreg) cells remain largely unknown. To determine the signals that drive the fate of TREG cells, we isolated and compared stable from unstable TREG cells using a TREG transfer model where we previously observed the functional reprogramming of Foxp3+ TREG cells into Th1/Th17 effector T cells. We identified the expression of the IL-33 receptor (IL-33R, ST2) on stable tTREG cells and the IL-1 receptor (IL1R1) on unstable exTREG cells undergoing functional reprogramming. We show that both TREG cell populations represent competing subsets in inflammatory conditions. This is further underlined by the fact that the absence of IL1R1 expression (IL1R1−/−) leads to the accumulation of ST2+ TREG cells at mucosal sites in vivo. In two distinct lung infection models, we demonstrate that ST2-expressing TREG cells express GATA3 and resist production of inflammatory cytokines, whereas IL1R1-expressing TREG cells express RORγT and lose Foxp3 expression in vivo. While IL-1 signaling impairs TREG cell suppressive function, ST2 is required for the maintenance of the lineage identity and suppressive function of TREG cells. These observations demonstrate that IL-1 and IL-33 produced during immune challenge exert distinct roles on the functional adaptation of Foxp3+ tTREG cells at mucosal surfaces.
c-Maf and STAT3 are essential but distinct regulators of RORγt+ and follicular regulatory T cells
Authors: Wheaton Joshua D. , Yeh Chen-Hao , Ciofani Maria 
Published: January 2023

Abstract
Foxp3+ regulatory T cells (Tregs) must adapt to function within a wide range of tissue environments and types of immune response. This flexibility is achieved through specialization, whereby context-dependent transcriptional changes integrate unique functionality into general Treg programming. In the most well-studied examples, this involves the co-expression of Foxp3 and another “lineage-defining” transcription factor (TF) which confers expression of specific functional genes. Two such specialized populations are RORγt+ Tregs and Bcl-6+ T follicular regulatory (Tfr) cells, which have important functions in regulating colitis and humoral immune responses, respectively. Although the importance of Treg specialization is now clear, the underlying network of signals and TFs driving this process is largely undefined. Here, we identify the TF c-Maf as an essential regulator of Treg specialization for both RORγt+ Tregs and Tfr cells. Importantly, the role of c-Maf in these subsets is highly specific, because deletion of Maf does not affect general Treg development, generation of other specialized Tregs, or expression of suppressive molecules. In vitro, we show that addition of IL-6 to Treg-promoting conditions is sufficient to drive both c-Maf expression and c-Maf-dependent conversion of naïve T cells to RORγt+ Tregs, suggesting that IL-6 might drive c-Maf in vivo. Surprisingly, although STAT3 – the primary downstream effector of IL-6 – is required for development of both RORγt+ Tregs and Tfr cells, we find that Stat3 deletion in vivo does not ablate c-Maf expression and results in Treg defects distinct from Maf deletion. Therefore, c-Maf and STAT3 are cooperative, but distinct regulators of RORγt+ and follicular Treg development.
A C-reactive protein-based strategy to reduce antibiotic dosing
Authors: Ngwa Donald Neba , Singh Sanjay K. , Gang Toh B. , Agrawal Alok 
Published: January 2023

Abstract
C-reactive protein (CRP) binds to Streptococcus pneumoniae through the phosphocholine groups present in the cell wall and subsequently activates the complement system to kill the pathogen. To escape the attack of complement, pneumococci recruit a complement inhibitory protein, factor H, on their surface. CRP protects mice against pneumococcal infection only when injected within 2 hours after administering pneumococci. We hypothesized that CRP is not protective when injected at later times because, by then, factor H is recruited by pneumococci. In the current study, we evaluated the protective effects of an engineered CRP molecule (E-CRP) which does not bind to factor H in fluid phase but binds to factor H-coated pneumococci. We found that E-CRP, unlike native CRP, protected mice regardless of the timing of administering E-CRP by drastically reducing bacteremia and increasing survival of mice. Next, we established another murine model of pneumococcal infection using the antibiotic clarithromycin. We found that the protective ability of 0.2 mg/dose of antibiotic was equivalent to that of 0.02 mg/dose of antibiotic combined with E-CRP; thus, there was 90% reduction in antibiotic dosing by including E-CRP in the treatment. Our data indicate that administration of E-CRP is therapeutically beneficial to reduce antibiotic dosing to decrease bacteremia and enhance survival. These results have implications for infections with antibiotic-resistant pneumococcal strains and possibly for other bacterial species that use factor H to evade the attack of complement. Additionally, the strategy to include E-CRP in the treatment using antibiotics might resolve the problem of developing antibiotic resistance.
IRF3 inhibits IFNγ-mediated restriction of intracellular pathogens in macrophages independently of type I interferons
Authors: Maciag Karolina , Raychowdhury Raktima , Smith Karen , Coers Jörn , Mumbach Maxwell R. , Schwartz Schraga , Polidoro Rafael B. , Gazzinelli Ricardo , Hacohen Nir 
Published: January 2023

Abstract
Macrophages use an array of innate immune sensors to detect intracellular pathogens and to tailor effective antimicrobial responses. In addition, extrinsic activation with the cytokine interferon gamma (IFNγ) is often required as well to tip the scales of the host-pathogen balance toward pathogen restriction. However, little is known about how host pathogen sensing impacts the anti-microbial IFNγ-activated state. We observed that in the absence of IRF3, a key downstream component of pathogen sensing pathways, IFNγ-primed macrophages more efficiently restricted the intracellular bacterium Legionella pneumophila and the intracellular protozoan parasite Trypanosoma cruzi. This effect was not mediated by Type I interferons, the key cytokines known to be induced by IRF3, or by the sensing adaptors MyD88/TRIF, MAVS, and STING. However, IRF3-deficient macrophages displayed a significantly altered IFNγ-induced gene expression program, with upregulation of microbial restriction factors such as Nos2. Furthermore, Nos2 activity was required for the enhanced IFNγ-mediated restriction of T. cruzi observed in IRF3-deficient macrophages. Finally, we found that IFNγ-primed but not resting macrophages largely excluded the activated form of IRF3 from the nucleus. These data are consistent with a relationship of mutual inhibition between IRF3 and IFNγ-activated programs, possibly as a component of a partially reversible mechanism for modulating the activity of potent innate immune effectors such as Nos2 in the context of intracellular infection.
Cannabinoid receptor 1 blockade attenuates metabolic inflammation and gut microbial dysbiosis during high-fat diet-induced obesity.
Authors: Miranda Kathryn , Becker William J. , Busbee Philip Brandon , Menzel Lorenzo , Dopkins Nicholas , Nagarkatti Prakash S. , Nagarkatti Mitzi 
Published: January 2023

Abstract
The endocannabinoid (eCB) system regulates a variety of physiological processes including inflammation and metabolism. Obesity is characterized by overactivation of the eCB system, chronic low-grade inflammation, metabolic dysfunction, and gut microbial dysbiosis. Blockade of cannabinoid receptor 1 (CB1) ameliorates obesity, however CB1-mediated immune-microbial interactions have not been extensively explored. In the current study, antagonists and genetic ablation of CB1 were used in a mouse model of diet-induced obesity to determine the role of CB1 in modulating inflammation, metabolism, and the gut microbiome. As expected, CB1−/− mice were resistant to high-fat diet (HFD)-induced weight gain and metabolic inflammation was significantly less in CB1−/− mice when compared to wild-type (WT) controls. Lamina propria lymphocytes from CB1−/− mice did not display a Th1 induction after HFD feeding whereas WT mice fed HFD had elevated CD4+T-bet+ cells compared to WT mice fed low-fat diet. Decreased colonic inflammation indicated CB1-dependent alterations in the gut microbiome may contribute to a decreased obese phenotype. Similarly, WT obese mice treated with the CB1 antagonist AM251 experienced weight loss, decreased inflammation, and improvement in metabolic parameters. Interrogation of the gut microbiome by 16S metagenomics revealed CB1-dependent alterations in abundance of bacteria belonging to the Clostridiales order. In addition, functional predictions by PICRUSt analysis revealed decreased abundance of operational taxonomic units belonging to bacterial metabolism and membrane transport pathways. Together these data suggest CB1 regulates metabolism by shaping the immune response and gut microbial community.
Random mutagenesis unveils novel host-pathogen interactions during colonic bacterial infections in immunocompromised hosts
Authors: Liu Yue , Xia Xue , Wan Fengyi 
Published: January 2023

Abstract
Attaching/Effacing (A/E) pathogens including enteropathogenic Escherichia coli (EPEC), enterohemorrhagic E. coli (EHEC), and the rodent equivalent Citrobacter rodentium (CR) are important causative agents of foodborne diseases. A/E pathogen infections cause severe morbidity and mortality in immunocompromised hosts with low interleukin-22 (IL-22); however, the crucial host-pathogen interactions and the pivotal A/E virulence proteins (effectors) under immunocompromised conditions, remain elusive. We conducted a functional screening of a CR mutant library consisting of ~2,000 mutant strains, generated via transposon-mediated random mutagenesis, and identified several avirulent strains, which abolish CR-induced severe morbidity and mortality in IL-22 knockout (Il22−/−) mice. Recapitulating the attenuated virulence phenotype by the transposon-interrupted mutant strains, genomic deletion of each identified effector in CR dramatically attenuates CR-caused lethality in Il22−/− mice. Interestingly, genetic deletion of each identified effector gene in CR genome exhibits no impact on the colonization and proliferation of CR in the infected animals; however, the deficiencies in these effectors substantially impede the colonic inflammatory response in the CR-infected Il22−/− mice, through distinct cellular and molecular mechanisms. Hence, our findings unveil a set of novel A/E pathogen effectors pathologically relevant to the severe morbidity and mortality in Il22−/− animals, which could provide new strategies to control A/E pathogen infections under the low IL-22 immunocompromised conditions, which are frequently associated with chronic HIV infection, organ transplantation, and other diseases.