Regulation of Bacteroides acidifaciens by the aryl hydrocarbon receptor in IL-22-producing immune cells has sex-dependent consequential impact on colitis.

Introduction: Colitis is an inflammatory bowel disease (IBD) characterized by immune cell dysregulation and alterations in the gut microbiome. In our previous report, we showed a natural product in cruciferous vegetables and ligand of the aryl hydrocarbon receptor (AhR), indole-3-carbinol (I3C), was able to reduce colitis-induced disease severity and microbial dysbiosis in an interleukin-22 (IL-22) dependent manner. Methods: In the current study, we performed single-cell RNA sequencing (scRNAseq) from colonocytes during colitis induction and supplementation with I3C and show how this treatment alters expression of genes involved in IL-22 signaling. To further define the role of IL-22 signaling in I3C-mediated protection during colitis and disease-associated microbial dysbiosis, we generated mice with AhR deficiency in RAR-related orphan receptor c (Rorc)-expressing cells (AhR ΔRorc ) which depletes this receptor in immune cells involved in production of IL-22. Colitis was induced in wildtype (WT), AhR ΔRorc , and littermate (LM) mice with or without I3C treatment. Results: Results showed AhR ΔRorc mice lost the efficacy effects of I3C treatment which correlated with a loss of ability to increase IL-22 by innate lymphoid type 3 (ILC3s), not T helper 22 (Th22) cells. 16S rRNA microbiome profiling studies showed AhR ΔRorc mice were unable to regulate disease-associated increases in Bacteroides, which differed between males and females. Lastly, inoculation with a specific disease-associated Bacteroides species, Bacteroides acidifaciens (B. acidifaciens), was shown to exacerbate colitis in females, but not males. Discussion: Collectively, this report highlights the cell and sex-specific role of AhR in regulating microbes that can impact colitis disease.

Delta-9-Tetrahydrocannabinol Blocks Bone Marrow-Derived Macrophage Differentiation through Elimination of Reactive Oxygen Species.

Macrophages are vital components of the immune system and serve as the first line of defense against pathogens. Macrophage colony-stimulating factor (M-CSF) induces macrophage differentiation from bone marrow-derived cells (BMDCs). Δ9-tetrahydrocannabiol (THC), a phytocannabinoid from the Cannabis plant, has profound anti-inflammatory properties with significant effects on myeloid cells. To investigate the effect of THC on macrophage differentiation, we cultured BMDCs with M-CSF in the presence of THC. Interestingly, THC markedly blocked the differentiation of BMDCs into CD45 + CD11b + F4/80+ macrophages. The effect of THC was independent of cannabinoid receptors CB1, and CB2, as well as other potential receptors such as GPR18, GPR55, and Adenosine 2A Receptor. RNA-seq analysis revealed that the THC-treated BMDCs displayed a significant increase in the expression of NRF2-ARE-related genes. KEGG pathway analysis revealed that the expression profiles of THC-treated cells correlated with ferroptosis and glutathione metabolism pathways. Fluorescence-based labile iron assays showed that the THC-treated BMDCs had significantly increased iron levels. Finally, THC-exposed BMDCs showed decreased levels of intracellular ROS. THC has the unique molecular property to block the Fenton Reaction, thus preventing the increase in intracellular ROS that is normally induced by high iron levels. Together, these studies demonstrated that THC blocks M-CSF-induced macrophage differentiation by inhibiting ROS production through both the induction of NRF2-ARE-related gene expression and the prevention of ROS formation via the Fenton Reaction.

Studying the cellular basis of small bowel enteropathy using high-parameter flow cytometry in mouse models of primary antibody deficiency.

Background: Primary immunodeficiencies are heritable defects in immune system function. Antibody deficiency is the most common form of primary immunodeficiency in humans, can be caused by abnormalities in both the development and activation of B cells, and may result from B-cell-intrinsic defects or defective responses by other cells relevant to humoral immunity. Inflammatory gastrointestinal complications are commonly observed in antibody-deficient patients, but the underlying immune mechanisms driving this are largely undefined. Methods: In this study, several mouse strains reflecting a spectrum of primary antibody deficiency (IgA-/-, Aicda-/-, CD19-/- and JH -/-) were used to generate a functional small-bowel-specific cellular atlas using a novel high-parameter flow cytometry approach that allows for the enumeration of 59 unique cell subsets. Using this cellular atlas, we generated a direct and quantifiable estimate of immune dysregulation. This estimate was then used to identify specific immune factors most predictive of the severity of inflammatory disease of the small bowel (small bowel enteropathy). Results: Results from our experiments indicate that the severity of primary antibody deficiency positively correlates with the degree of immune dysregulation that can be expected to develop in an individual. In the SI of mice, immune dysregulation is primarily explained by defective homeostatic responses in T cell and invariant natural killer-like T (iNKT) cell subsets. These defects are strongly correlated with abnormalities in the balance between protein (MHCII-mediated) versus lipid (CD1d-mediated) antigen presentation by intestinal epithelial cells (IECs) and intestinal stem cells (ISCs), respectively. Conclusions: Multivariate statistical approaches can be used to obtain quantifiable estimates of immune dysregulation based on high-parameter flow cytometry readouts of immune function. Using one such estimate, we reveal a previously unrecognized tradeoff between iNKT cell activation and type 1 immunity that underlies disease in the small bowel. The balance between protein/lipid antigen presentation by ISCs may play a crucial role in regulating this balance and thereby suppressing inflammatory disease in the small bowel.

Identification of miRNAs that target Fcγ receptor-mediated phagocytosis during macrophage activation syndrome.

Macrophage activation syndrome (MAS) is a life-threatening complication of systemic juvenile arthritis, accompanied by cytokine storm and hemophagocytosis. In addition, COVID-19-related hyperinflammation shares clinical features of MAS. Mechanisms that activate macrophages in MAS remain unclear. Here, we identify the role of miRNA in increased phagocytosis and interleukin-12 (IL-12) production by macrophages in a murine model of MAS. MAS significantly increased F4/80+ macrophages and phagocytosis in the mouse liver. Gene expression profile revealed the induction of Fcγ receptor-mediated phagocytosis (FGRP) and IL-12 production in the liver. Phagocytosis pathways such as High-affinity IgE receptor is known as Fc epsilon RI -signaling and pattern recognition receptors involved in the recognition of bacteria and viruses and phagosome formation were also significantly upregulated. In MAS, miR-136-5p and miR-501-3p targeted and caused increased expression of Fcgr3, Fcgr4, and Fcgr1 genes in FGRP pathway and consequent increase in phagocytosis by macrophages, whereas miR-129-1-3p and miR-150-3p targeted and induced Il-12. Transcriptome analysis of patients with MAS revealed the upregulation of FGRP and FCGR gene expression. A target analysis of gene expression data from a patient with MAS discovered that miR-136-5p targets FCGR2A and FCGR3A/3B, the human orthologs of mouse Fcgr3 and Fcgr4, and miR-501-3p targets FCGR1A, the human ortholog of mouse Fcgr1. Together, we demonstrate the novel role of miRNAs during MAS pathogenesis, thereby suggesting miRNA mimic-based therapy to control the hyperactivation of macrophages in patients with MAS as well as use overexpression of FCGR genes as a marker for MAS classification.

Indole-3-carbinol attenuates lipopolysaccharide-induced acute respiratory distress syndrome through activation of AhR: role of CCR2+ monocyte activation and recruitment in the regulation of CXCR2+ neutrophils in the lungs.

Introduction: Indole-3-carbinol (I3C) is found in cruciferous vegetables and used as a dietary supplement. It is known to act as a ligand for aryl hydrocarbon receptor (AhR). In the current study, we investigated the role of AhR and the ability of I3C to attenuate LPS-induced Acute Respiratory Distress Syndrome (ARDS). Methods: To that end, we induced ARDS in wild-type C57BL/6 mice, Ccr2gfp/gfp KI/KO mice (mice deficient in the CCR2 receptor), and LyZcreAhRfl/fl mice (mice deficient in the AhR on myeloid linage cells). Additionally, mice were treated with I3C (65 mg/kg) or vehicle to investigate its efficacy to treat ARDS. Results: I3C decreased the neutrophils expressing CXCR2, a receptor associated with neutrophil recruitment in the lungs. In addition, LPS-exposed mice treated with I3C revealed downregulation of CCR2+ monocytes in the lungs and lowered CCL2 (MCP-1) protein levels in serum and bronchoalveolar lavage fluid. Loss of CCR2 on monocytes blocked the recruitment of CXCR2+ neutrophils and decreased the total number of immune cells in the lungs during ARDS. In addition, loss of the AhR on myeloid linage cells ablated I3C-mediated attenuation of CXCR2+ neutrophils and CCR2+ monocytes in the lungs from ARDS animals. Interestingly, scRNASeq showed that in macrophage/monocyte cell clusters of LPS-exposed mice, I3C reduced the expression of CXCL2 and CXCL3, which bind to CXCR2 and are involved in neutrophil recruitment to the disease site. Discussion: These findings suggest that CCR2+ monocytes are involved in the migration and recruitment of CXCR2+ neutrophils during ARDS, and the AhR ligand, I3C, can suppress ARDS through the regulation of immune cell trafficking.

6-Formylindolo[3,2-b]carbazole, a potent ligand for the aryl hydrocarbon receptor, attenuates concanavalin-induced hepatitis by limiting T-cell activation and infiltration of proinflammatory CD11b+ Kupffer cells.

FICZ (6-formylindolo[3,2-b]carbazole) is a potent aryl hydrocarbon receptor agonist that has a poorly understood function in the regulation of inflammation. In this study, we investigated the effect of aryl hydrocarbon receptor activation by FICZ in a murine model of autoimmune hepatitis induced by concanavalin A. High-throughput sequencing techniques such as single-cell RNA sequencing and assay for transposase accessible chromatin sequencing were used to explore the mechanisms through which FICZ induces its effects. FICZ treatment attenuated concanavalin A-induced hepatitis, evidenced by decreased T-cell infiltration, decreased circulating alanine transaminase levels, and suppression of proinflammatory cytokines. Concanavalin A revealed an increase in natural killer T cells, T cells, and mature B cells upon concanavalin A injection while FICZ treatment reversed the presence of these subsets. Surprisingly, concanavalin A depleted a subset of CD55+ B cells, while FICZ partially protected this subset. The immune cells showed significant dysregulation in the gene expression profiles, including diverse expression of migratory markers such as CCL4, CCL5, and CXCL2 and critical regulatory markers such as Junb. Assay for transposase accessible chromatin sequencing showed more accessible chromatin in the CD3e promoter in the concanavalin A-only group as compared to the naive and concanavalin A-exposed, FICZ-treated group. While there was overall more accessible chromatin of the Adgre1 (F4/80) promoter in the FICZ-treated group, we observed less open chromatin in the Itgam (CD11b) promoter in Kupffer cells, supporting the ability of FICZ to reduce the infiltration of proinflammatory cytokine producing CD11b+ Kupffer cells. Taken together, these data demonstrate that aryl hydrocarbon receptor activation by FICZ suppresses liver injury through the limitation of CD3+ T-cell activation and CD11b+ Kupffer cell infiltration.

Aryl Hydrocarbon Receptor Regulates Muc2 Production Independently of IL-22 during Colitis.

We previously reported that an aryl hydrocarbon receptor (AhR) ligand, indole-3-carbinol (I3C), was effective at reducing colitis severity through immune cell-mediated interleukin-22 (IL-22) production. Intestinal epithelial cells (IECs) are also involved in regulating colitis, so we investigated their AhR-mediated mechanisms in the current report. A transcriptome analysis of IECs in wildtype (WT) mice revealed that during colitis, I3C regulated select mucin proteins, which could be attributed to goblet cell development. To address this, experiments under in vivo colitis (mice) or in vitro colon organoid conditions were undertaken to determine how select mucin proteins were altered in the absence or presence of AhR in IECs during I3C treatment. Comparing WT to IEC-specific AhR knockout mice (AhRΔIEC), the results showed that AhR expression was essential in IECs for I3C-mediated protection during colitis. AhR-deficiency also impaired mucin protein expression, particularly mucin 2 (Muc2), independently of IL-22. Collectively, this report highlights the important role of AhR in direct regulation of Muc2. These results provide justification for future studies aimed at determining how AhR might regulate select mucins through mechanisms such as direct transcription binding to enhance production.

Studying the cellular basis of small bowel enteropathy using high-parameter flow cytometry in mouse models of primary antibody deficiency.

Background: Primary immunodeficiencies are heritable defects in immune system function. Antibody deficiency is the most common form of primary immunodeficiency in humans, can be caused by abnormalities in both the development and activation of B cells, and may result from B-cell-intrinsic defects or defective responses by other cells relevant to humoral immunity. Inflammatory gastrointestinal complications are commonly observed in antibody-deficient patients, but the underlying immune mechanisms driving this are largely undefined. Methods: In this study, several mouse strains reflecting a spectrum of primary antibody deficiency (IgA -/- , Aicda -/- , CD19 -/- and J H -/- ) were used to generate a functional small-bowel-specific cellular atlas using a novel high-parameter flow cytometry approach that allows for the enumeration of 59 unique cell subsets. Using this cellular atlas, we generated a direct and quantifiable estimate of immune dysregulation. This estimate was then used to identify specific immune factors most predictive of the severity of inflammatory disease of the small bowel (small bowel enteropathy). Results: Results from our experiments indicate that the severity of primary antibody deficiency positively correlates with the degree of immune dysregulation that can be expected to develop in an individual. In the SI of mice, immune dysregulation is primarily explained by defective homeostatic responses in T cell and invariant natural killer-like T (iNKT) cell subsets. These defects are strongly correlated with abnormalities in the balance between protein (MHCII-mediated) versus lipid (CD1d-mediated) antigen presentation by intestinal epithelial cells (IECs) and intestinal stem cells (ISCs), respectively. Conclusions: Multivariate statistical approaches can be used to obtain quantifiable estimates of immune dysregulation based on high-parameter flow cytometry readouts of immune function. Using one such estimate, we reveal a previously unrecognized tradeoff between iNKT cell activation and type 1 immunity that underlies disease in the small bowel. The balance between protein/lipid antigen presentation by ISCs may play a crucial role in regulating this balance and thereby suppressing inflammatory disease in the small bowel.

Estrobolome dysregulation is associated with altered immunometabolism in a mouse model of endometriosis.

Introduction: Endometriosis is a painful disease that affects around 5% of women of reproductive age. In endometriosis, ectopic endometrial cells or seeded endometrial debris grow in abnormal locations including the peritoneal cavity. Common manifestations of endometriosis include dyspareunia, dysmenorrhea, chronic pelvic pain and often infertility and symptomatic relief or surgical removal are mainstays of treatment. Endometriosis both promotes and responds to estrogen imbalance, leading to intestinal bacterial estrobolome dysregulation and a subsequent induction of inflammation. Methods: In the current study, we investigated the linkage between gut dysbiosis and immune metabolic response in endometriotic mice. Ovariectomized BALB/c mice received intraperitoneal transplantation of endometrial tissue from OVX donors (OVX+END). Control groups included naïve mice (Naïve), naïve mice that received endometrial transplants (Naive+END) and OVX mice that received the vehicle (OVX+VEH). Colonic content was collected 2 weeks post-transplantation for 16s rRNA pyrosequencing and peritoneal fluid was collected to determine the phenotype of inflammatory cells by flow cytometry. Results: We noted a significant increase in the number of peritoneal fluid cells, specifically, T cells, natural killer (NK) cells, and NKT cells in OVX+END mice. Phylogenetic taxonomy analysis showed significant dysbiosis in OVX+END mice, with an increase in abundance of Phylum Tenericutes, Class Mollicutes, Order Aneroplasmatales, and Genus Aneroplasma, and a decrease in Order Clostridiales, and Genus Dehalobacterium, when compared to OVX+VEH controls. The metabolomic profile showed an increase in some tricarboxylic acid cycle (TCA)-related metabolites accompanied by a reduction in short-chain fatty acids (SCFA) such as butyric acid in OVX+END mice. Additionally, the mitochondrial and ATP production of immune cells was enforced to a maximal rate in OVX+END mice when compared to OVX+VEH mice. Conclusion: The current study demonstrates that endometriosis alters the gut microbiota and associated immune metabolism.

Resveratrol Attenuates 2,3,7,8-Tetrachlorodibenzo-p-dioxin-Mediated Induction of Myeloid-Derived Suppressor Cells (MDSC) and Their Functions.

Previously, we showed that 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), an aryl hydrocarbon receptor (AhR) ligand and a potent and persistent toxicant and carcinogenic agent, induces high levels of murine myeloid-derived suppressor cell (MDSC) when injected into mice. In the current study, we demonstrate that Resveratrol (3,4,5-trihydroxy-trans-stilbene; RSV), an AhR antagonist, reduces TCDD-mediated MDSC induction. RSV decreased the number of MDSCs induced by TCDD in mice but also mitigated the immunosuppressive function of TCDD-induced MDSCs. TCDD caused a decrease in F4/80+ macrophages and an increase in CD11C+ dendritic cells, while RSV reversed these effects. TCDD caused upregulation in CXCR2, a critical molecule involved in TCDD-mediated induction of MDSCs, and Arginase-1 (ARG-1), involved in the immunosuppressive functions of MDSCs, while RSV reversed this effect. Transcriptome analysis of Gr1+ MDSCs showed an increased gene expression profile involved in the metabolic pathways in mice exposed to TCDD while RSV-treated mice showed a decrease in such pathways. The bio-energetic profile of these cells showed that RSV treatment decreased the energetic demands induced by TCDD. Overall, the data demonstrated that RSV decreased TCDD-induced MDSC induction and function by altering the dynamics of various myeloid cell populations involving their numbers, phenotype, and immunosuppressive potency. Because MDSCs play a critical role in tumor growth and metastasis, our studies also support the potential use of RSV to attenuate the immunosuppressive properties of MDSC.

Effect of TCDD exposure in adult female and male mice on the expression of miRNA in the ovaries and testes and associated reproductive functions.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is an environmental contaminant found widely across the world. While animal and human studies have shown that exposure to TCDD may cause significant alterations in the reproductive tract, the effect of TCDD on the expression of miRNA in the reproductive organs has not been previously tested. In the current study, we exposed adult female or male mice to TCDD or vehicle and bred them to study the impact on reproduction. The data showed that while TCDD treatment of females caused no significant change in litter size, it did alter the survival of the pups. Also, TCDD exposure of either the male or female mice led to an increase in the gestational period. While TCDD did not alter the gross morphology of the ovaries and testes, it induced significant alterations in the miRNA expression. The ovaries showed the differential expression of 426 miRNAs, of which 315 miRNAs were upregulated and 111 miRNA that were downregulated after TCDD exposure when compared to the vehicle controls. In the testes, TCDD caused the differential expression of 433 miRNAs, with 247 miRNAs upregulated and 186 miRNAs downregulated. Pathway analysis showed that several of these dysregulated miRNAs targeted reproductive functions. The current study suggests that the reproductive toxicity of TCDD may result from alterations in the miRNA expression in the reproductive organs. Because miRNAs also represent one of the epigenetic pathways of gene expression, our studies suggest that the transgenerational toxicity of TCDD may also result from dysregulation in the miRNAs.

Membrane-Active Metallopolymers: Repurposing and Rehabilitating Antibiotics to Gram-Negative Superbugs.

Among multiple approaches to combating antimicrobial resistance, a combination therapy of existing antibiotics with bacterial membrane-perturbing agents is promising. A viable platform of metallopolymers as adjuvants in combination with traditional antibiotics is reported in this work to combat both planktonic and stationary cells of Gram-negative superbugs and their biofilms. Antibacterial efficacy, toxicity, antibiofilm activity, bacterial resistance propensity, and mechanisms of action of metallopolymer-antibiotic combinations are investigated. These metallopolymers exhibit 4-16-fold potentiation of antibiotics against Gram-negative bacteria with negligible toxicity toward mammalian cells. More importantly, the lead combinations (polymer-ceftazidime and polymer-rifampicin) eradicate preformed biofilms of MDR E. coli and P. aeruginosa, respectively. Further, ß-lactamase inhibition, outer membrane permeabilization, and membrane depolarization demonstrate synergy of these adjuvants with different antibiotics. Moreover, the membrane-active metallopolymers enable the antibiotics to circumvent bacterial resistance development. Altogether, the results indicate that such non-antibiotic adjuvants bear the promise to revitalize the efficacy of existing antibiotics to tackle Gram-negative bacterial infections.

Facially Amphiphilic Bile Acid-Functionalized Antimicrobials: Combating Pathogenic Bacteria, Fungi, and Their Biofilms.

We report facially amphiphilic bile acid-based antimicrobials with a broad spectrum of activity against both bacterial and fungal pathogens and negligible detrimental effects on mammalian cells. Two lead compounds eliminated dormant subpopulations of various bacterial species, unlike conventional antibiotics. The lead compounds were also effective in eradicating biofilms of methicillin-resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa, and Candida albicans. Additionally, these compounds substantially inhibited the formation of fungal biofilms (C. albicans). Mechanistic investigations revealed the membrane-active nature and endogenous reactive oxygen species (ROS) induction ability of these compounds. Finally, no detectable resistance was developed by the bacterial strains against this class of membrane-targeting antimicrobials.

Facial amphiphilic naphthoic acid-derived antimicrobial polymers against multi-drug resistant gram-negative bacteria and biofilms.

Inspired by the facial amphiphilic nature and antimicrobial efficacy of many antimicrobial peptides, this work reported facial amphiphilic bicyclic naphthoic acid derivatives with different ratios of charges to rings that were installed onto side chains of poly(glycidyl methacrylate). Six quaternary ammonium-charged (QAC) polymers were prepared to investigate the structure-activity relationship. These QAC polymers displayed potent antibacterial activity against various multi-drug resistant (MDR) gram-negative pathogens such as Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, and Acinetobacter baumannii. Polymers demonstrated low hemolysis and high antimicrobial selectivity. Additionally, they were able to eradicate established biofilms and kill metabolically inactive dormant cells. The membrane permeabilization and depolarization results indicated a mechanism of action through membrane disruption. Two lead polymers showed no resistance from MDR-P. aeruginosa and MDR-K. pneumoniae. These facial amphiphiles are potentially a new class of potent antimicrobial agents to tackle the antimicrobial resistance for both planktonic and biofilm-related infections.

The impact of geographic inequality in federal research funding: A comparative longitudinal study of research and scholarly outputs in EPSCoR versus non-EPSCoR states.

Some states in the U.S. have traditionally received less federal research funding than other states. The National Science Foundation (NSF) created a program in 1979, called the Experimental Program to Stimulate Competitive Research (EPSCoR) to enhance the research competitiveness in such states. While the geographic disparity in federal research funding is well known, the overall impact of federal funding on the research performance of EPSCoR and non-EPSCoR has not been previously studied. In the current study, we compared the combined research productivity of Ph.D. granting institutions in EPSCoR versus the non-EPSCoR states to better understand the scientific impact of federal investments in sponsored research across all states. The research outputs we measured included journal articles, books, conference papers, patents, and citation count in academic literature. Unsurprisingly, results indicated that the non-EPSCoR states received significantly more federal research funding than their EPSCoR counterparts, which correlated with a higher number of faculty members in the non-EPSCoR versus EPSCoR states. Also, in the overall research productivity expressed on a per capita, the non-EPSCoR states fared better than EPSCoR states. However, when the research output was measured based on per $1M investment of federal research funding, EPSCoR states performed significantly better than the non-EPSCoR states in many research productivity indicators, with the notable exception of patents. Together, this study found preliminary evidence that EPSCoR states achieved a high degree of research productivity despite receiving significantly fewer federal research dollars. The limitations and next steps of this study are also discussed.

Characterization of Chemotaxis-Associated Gene Dysregulation in Myeloid Cell Populations in the Lungs during Lipopolysaccharide-Mediated Acute Lung Injury.

During endotoxin-induced acute lung injury (ALI), immune cell recruitment resulting from chemotaxis is mediated by CXC and CC chemokines and their receptors. In this study, we investigated the role of chemokines and their receptors in the regulation of myeloid cell populations in the circulation and the lungs of C57BL/6J mice exhibiting LPS-mediated ALI using single-cell RNA sequencing. During ALI, there was an increase in the myeloid cells, M1 macrophages, monocytes, neutrophils, and other granulocytes, whereas there was a decrease in the residential alveolar macrophages and M2 macrophages. Interestingly, LPS triggered the upregulation of CCL3, CCL4, CXCL2/3, and CXCL10 genes associated with cellular migration of various subsets of macrophages, neutrophils, and granulocytes. Furthermore, there was an increase in the frequency of myeloid cells expressing CCR1, CCR3, CCR5, and CXCR2 receptors during ALI. MicroRNA sequencing studies of vehicle versus LPS groups identified several dysregulated microRNAs targeting the upregulated chemokine genes. This study suggests that chemokine ligand-receptors interactions are responsible for myeloid cell heterogenicity and cellular recruitment to the lungs during ALI. The single-cell transcriptomics allowed for an in-depth assessment and characterization of myeloid cells involved in immune cell trafficking during ALI.

Resveratrol Protects against Skin Inflammation through Inhibition of Mast Cell, Sphingosine Kinase-1, Stat3 and NF-κB p65 Signaling Activation in Mice.

Inflammation is pathogenic to skin diseases, including atopic dermatitis (AD) and eczema. Treatment for AD remains mostly symptomatic with newer but costly options, tainted with adverse side effects. There is an unmet need for safe therapeutic and preventative strategies for AD. Resveratrol (R) is a natural compound known for its anti-inflammatory properties. However, animal and human R studies have yielded contrasting results. Mast cells (MCs) are innate immune skin-resident cells that initiate the development of inflammation and progression to overt disease. R's effects on MCs are also controversial. Using a human-like mouse model of AD development consisting of a single topical application of antigen ovalbumin (O) for 7 days, we previously established that the activation of MCs by a bioactive sphingolipid metabolite sphingosine-1-phosphate (S1P) initiated substantial skin remodeling compared to controls. Here, we show that daily R application normalized O-mediated epidermal thickening, ameliorated cell infiltration, and inhibited skin MC activation and chemokine expression. We unraveled R's multiple mechanisms of action, including decreased activation of the S1P-producing enzyme, sphingosine kinase 1 (SphK1), and of transcription factors Signal Transducer and Activator of Transcription 3 (Stat3) and NF-κBp65, involved in chemokine production. Thus, R may be poised for protection against MC-driven pathogenic skin inflammation.

Resveratrol attenuates staphylococcal enterotoxin B-activated immune cell metabolism via upregulation of miR-100 and suppression of mTOR signaling pathway.

Acute Respiratory Distress Syndrome (ARDS) is triggered by a variety of insults, such as bacterial and viral infections, including SARS-CoV-2, leading to high mortality. In the murine model of ARDS induced by Staphylococcal enterotoxin-B (SEB), our previous studies showed that while SEB triggered 100% mortality, treatment with Resveratrol (RES) completely prevented such mortality by attenuating inflammation in the lungs. In the current study, we investigated the metabolic profile of SEB-activated immune cells in the lungs following treatment with RES. RES-treated mice had higher expression of miR-100 in the lung mononuclear cells (MNCs), which targeted mTOR, leading to its decreased expression. Also, Single-cell RNA-seq (scRNA seq) unveiled the decreased expression of mTOR in a variety of immune cells in the lungs. There was also an increase in glycolytic and mitochondrial respiration in the cells from SEB + VEH group in comparison with SEB + RES group. Together these data suggested that RES alters the metabolic reprogramming of SEB-activated immune cells, through suppression of mTOR activation and its down- and upstream effects on energy metabolism. Also, miR-100 could serve as novel potential therapeutic molecule in the amelioration of ARDS.

Biosynthetic Enzyme-guided Disease Correlation Connects Gut Microbial Metabolites Sulfonolipids to Inflammatory Bowel Disease Involving TLR4 Signaling.

The trillions of microorganisms inhabiting the human gut are intricately linked to human health. At the species abundance level, correlational studies have connected specific bacterial taxa to various diseases. While the abundances of these bacteria in the gut serve as good indicators for disease progression, understanding the functional metabolites they produce is critical to decipher how these microbes influence human health. Here, we report a unique biosynthetic enzyme-guided disease correlation approach to uncover microbial functional metabolites as potential molecular mechanisms in human health. We directly connect the expression of gut microbial sulfonolipid (SoL) biosynthetic enzymes to inflammatory bowel disease (IBD) in patients, revealing a negative correlation. This correlation is then corroborated by targeted metabolomics, identifying that SoLs abundance is significantly decreased in IBD patient samples. We experimentally validate our analysis in a mouse model of IBD, showing that SoLs production is indeed decreased while inflammatory markers are increased in diseased mice. In support of this connection, we apply bioactive molecular networking to show that SoLs consistently contribute to the immunoregulatory activity of SoL-producing human microbes. We further reveal that sulfobacins A and B, two representative SoLs, primarily target Toll-like receptor 4 (TLR4) to mediate immunomodulatory activity through blocking TLR4's natural ligand lipopolysaccharide (LPS) binding to myeloid differentiation factor 2, leading to significant suppression of LPS-induced inflammation and macrophage M1 polarization. Together, these results suggest that SoLs mediate a protective effect against IBD through TLR4 signaling and showcase a widely applicable biosynthetic enzyme-guided disease correlation approach to directly link the biosynthesis of gut microbial functional metabolites to human health.