The Asthma Risk Gene, GSDMB, Promotes Mitochondrial DNA-induced ISGs Expression.

Released mitochondrial DNA (mtDNA) in cells activates cGAS-STING pathway, which induces expression of interferon-stimulated genes (ISGs) and thereby promotes inflammation, as frequently seen in asthmatic airways. However, whether the genetic determinant, Gasdermin B (GSDMB), the most replicated asthma risk gene, regulates this pathway remains unknown. We set out to determine whether and how GSDMB regulates mtDNA-activated cGAS-STING pathway and subsequent ISGs induction in human airway epithelial cells. Using qPCR, ELISA, native polyacrylamide gel electrophoresis, co-immunoprecipitation and immunofluorescence assays, we evaluated the regulation of GSDMB on cGAS-STING pathway in both BEAS-2B cells and primary normal human bronchial epithelial cells (nHBEs). mtDNA was extracted in plasma samples from human asthmatics and the correlation between mtDNA levels and eosinophil counts was analyzed. GSDMB is significantly associated with RANTES expression in asthmatic nasal epithelial brushing samples from the Genes-environments and Admixture in Latino Americans (GALA) II study. Over-expression of GSDMB promotes DNA-induced IFN and ISGs expression in bronchial epithelial BEAS-2B cells and nHBEs. Conversely, knockout of GSDMB led to weakened induction of interferon (IFNs) and ISGs in BEAS-2B cells. Mechanistically, GSDMB interacts with the C-terminus of STING, promoting the translocation of STING to Golgi, leading to the phosphorylation of IRF3 and induction of IFNs and ISGs. mtDNA copy number in serum from asthmatics was significantly correlated with blood eosinophil counts especially in male subjects. GSDMB promotes the activation of mtDNA and poly (dA:dT)-induced activation of cGAS-STING pathway in airway epithelial cells, leading to enhanced induction of ISGs.

Alveolar type II epithelial cell FASN maintains lipid homeostasis in experimental COPD.

Alveolar epithelial type II (AEC2) cells strictly regulate lipid metabolism to maintain surfactant synthesis. Loss of AEC2 cell function and surfactant production are implicated in the pathogenesis of the smoking-related lung disease chronic obstructive pulmonary disease (COPD). Whether smoking alters lipid synthesis in AEC2 cells and whether altering lipid metabolism in AEC2 cells contributes to COPD development are unclear. In this study, high-throughput lipidomic analysis revealed increased lipid biosynthesis in AEC2 cells isolated from mice chronically exposed to cigarette smoke (CS). Mice with a targeted deletion of the de novo lipogenesis enzyme, fatty acid synthase (FASN), in AEC2 cells (FasniΔAEC2) exposed to CS exhibited higher bronchoalveolar lavage fluid (BALF) neutrophils, higher BALF protein, and more severe airspace enlargement. FasniΔAEC2 mice exposed to CS had lower levels of key surfactant phospholipids but higher levels of BALF ether phospholipids, sphingomyelins, and polyunsaturated fatty acid-containing phospholipids, as well as increased BALF surface tension. FasniΔAEC2 mice exposed to CS also had higher levels of protective ferroptosis markers in the lung. These data suggest that AEC2 cell FASN modulates the response of the lung to smoke by regulating the composition of the surfactant phospholipidome.

Auto-immunoproteomics analysis of COVID-19 ICU patients revealed increased levels of autoantibodies related to the male reproductive system.

Background: Coronavirus disease (COVID-19) manifests many clinical symptoms, including an exacerbated immune response and cytokine storm. Autoantibodies in COVID-19 may have severe prodromal effects that are poorly understood. The interaction between these autoantibodies and self-antigens can result in systemic inflammation and organ dysfunction. However, the role of autoantibodies in COVID-19 complications has yet to be fully understood. Methods: The current investigation screened two independent cohorts of 97 COVID-19 patients [discovery (Disc) cohort from Qatar (case = 49 vs. control = 48) and replication (Rep) cohort from New York (case = 48 vs. control = 28)] utilizing high-throughput KoRectly Expressed (KREX) Immunome protein-array technology. Total IgG autoantibody responses were evaluated against 1,318 correctly folded and full-length human proteins. Samples were randomly applied on the precoated microarray slides for 2 h. Cy3-labeled secondary antibodies were used to detect IgG autoantibody response. Slides were scanned at a fixed gain setting using the Agilent fluorescence microarray scanner, generating a 16-bit TIFF file. Group comparisons were performed using a linear model and Fisher's exact test. Differentially expressed proteins were used for KEGG and WIKIpathway annotation to determine pathways in which the proteins of interest were significantly over-represented. Results and conclusion: Autoantibody responses to 57 proteins were significantly altered in the COVID-19 Disc cohort compared to healthy controls (p ≤ 0.05). The Rep cohort had altered autoantibody responses against 26 proteins compared to non-COVID-19 ICU patients who served as controls. Both cohorts showed substantial similarities (r 2 = 0.73) and exhibited higher autoantibody responses to numerous transcription factors, immunomodulatory proteins, and human disease markers. Analysis of the combined cohorts revealed elevated autoantibody responses against SPANXN4, STK25, ATF4, PRKD2, and CHMP3 proteins in COVID-19 patients. The sequences for SPANXN4 and STK25 were cross-validated using sequence alignment tools. ELISA and Western blot further verified the autoantigen-autoantibody response of SPANXN4. SPANXN4 is essential for spermiogenesis and male fertility, which may predict a potential role for this protein in COVID-19-associated male reproductive tract complications, and warrants further research.

Mitochondria in health, disease, and aging.

Mitochondria are well known as organelles responsible for the maintenance of cellular bioenergetics through the production of ATP. Although oxidative phosphorylation may be their most important function, mitochondria are also integral for the synthesis of metabolic precursors, calcium regulation, the production of reactive oxygen species, immune signaling, and apoptosis. Considering the breadth of their responsibilities, mitochondria are fundamental for cellular metabolism and homeostasis. Appreciating this significance, translational medicine has begun to investigate how mitochondrial dysfunction can represent a harbinger of disease. In this review, we provide a detailed overview of mitochondrial metabolism, cellular bioenergetics, mitochondrial dynamics, autophagy, mitochondrial damage-associated molecular patterns, mitochondria-mediated cell death pathways, and how mitochondrial dysfunction at any of these levels is associated with disease pathogenesis. Mitochondria-dependent pathways may thereby represent an attractive therapeutic target for ameliorating human disease.

IL-4 drastically decreases deuterosomal and multiciliated cells via alteration in progenitor cell differentiation.

BACKGROUND: Allergic inflammation affects the epithelial cell populations resulting in goblet cell hyperplasia and decreased ciliated cells. Recent advances in single-cell RNA sequencing (scRNAseq) have enabled the identification of new cell subtypes and genomic features of single cells. In this study, we aimed to investigate the effect of allergic inflammation in nasal epithelial cell transcriptomes at the single-cell level. METHODS: We performed scRNAseq in cultured primary human nasal epithelial (HNE) cells and in vivo nasal epithelium. The transcriptomic features and epithelial cell subtypes were determined under IL-4 stimulation, and cell-specific marker genes and proteins were identified. RESULTS: We confirmed that cultured HNE cells were similar to in vivo epithelial cells through scRNAseq. Cell-specific marker genes were utilized to cluster the cell subtypes, and FOXJ1+ -ciliated cells were sub-classified into multiciliated and deuterosomal cells. PLK4 and CDC20B were specific for deuterosomal cells, and SNTN, CPASL, and GSTA2 were specific for multiciliated cells. IL-4 altered the proportions of cell subtypes, resulting in a decrease in multiciliated cells and loss of deuterosomal cells. The trajectory analysis revealed deuterosomal cells as precursor cells of multiciliated cells and deuterosomal cells function as a bridge between club and multiciliated cells. A decrease in deuterosomal cell marker genes was observed in nasal tissue samples with type 2 inflammation. CONCLUSION: The effects of IL-4 appear to be mediated through the loss of the deuterosomal population, resulting in the reduction in multiciliated cells. This study also newly suggests cell-specific markers that might be pivotal for investigating respiratory inflammatory diseases.

Urine-based multi-omic comparative analysis of COVID-19 and bacterial sepsis-induced ARDS.

BACKGROUND: Acute respiratory distress syndrome (ARDS), a life-threatening condition during critical illness, is a common complication of COVID-19. It can originate from various disease etiologies, including severe infections, major injury, or inhalation of irritants. ARDS poses substantial clinical challenges due to a lack of etiology-specific therapies, multisystem involvement, and heterogeneous, poor patient outcomes. A molecular comparison of ARDS groups holds the potential to reveal common and distinct mechanisms underlying ARDS pathogenesis. METHODS: We performed a comparative analysis of urine-based metabolomics and proteomics profiles from COVID-19 ARDS patients (n = 42) and bacterial sepsis-induced ARDS patients (n = 17). To this end, we used two different approaches, first we compared the molecular omics profiles between ARDS groups, and second, we correlated clinical manifestations within each group with the omics profiles. RESULTS: The comparison of the two ARDS etiologies identified 150 metabolites and 70 proteins that were differentially abundant between the two groups. Based on these findings, we interrogated the interplay of cell adhesion/extracellular matrix molecules, inflammation, and mitochondrial dysfunction in ARDS pathogenesis through a multi-omic network approach. Moreover, we identified a proteomic signature associated with mortality in COVID-19 ARDS patients, which contained several proteins that had previously been implicated in clinical manifestations frequently linked with ARDS pathogenesis. CONCLUSION: In summary, our results provide evidence for significant molecular differences in ARDS patients from different etiologies and a potential synergy of extracellular matrix molecules, inflammation, and mitochondrial dysfunction in ARDS pathogenesis. The proteomic mortality signature should be further investigated in future studies to develop prediction models for COVID-19 patient outcomes.

Multi-omic comparative analysis of COVID-19 and bacterial sepsis-induced ARDS.

BACKGROUND: Acute respiratory distress syndrome (ARDS), a life-threatening condition characterized by hypoxemia and poor lung compliance, is associated with high mortality. ARDS induced by COVID-19 has similar clinical presentations and pathological manifestations as non-COVID-19 ARDS. However, COVID-19 ARDS is associated with a more protracted inflammatory respiratory failure compared to traditional ARDS. Therefore, a comprehensive molecular comparison of ARDS of different etiologies groups may pave the way for more specific clinical interventions. METHODS AND FINDINGS: In this study, we compared COVID-19 ARDS (n = 43) and bacterial sepsis-induced (non-COVID-19) ARDS (n = 24) using multi-omic plasma profiles covering 663 metabolites, 1,051 lipids, and 266 proteins. To address both between- and within- ARDS group variabilities we followed two approaches. First, we identified 706 molecules differently abundant between the two ARDS etiologies, revealing more than 40 biological processes differently regulated between the two groups. From these processes, we assembled a cascade of therapeutically relevant pathways downstream of sphingosine metabolism. The analysis suggests a possible overactivation of arginine metabolism involved in long-term sequelae of ARDS and highlights the potential of JAK inhibitors to improve outcomes in bacterial sepsis-induced ARDS. The second part of our study involved the comparison of the two ARDS groups with respect to clinical manifestations. Using a data-driven multi-omic network, we identified signatures of acute kidney injury (AKI) and thrombocytosis within each ARDS group. The AKI-associated network implicated mitochondrial dysregulation which might lead to post-ARDS renal-sequalae. The thrombocytosis-associated network hinted at a synergy between prothrombotic processes, namely IL-17, MAPK, TNF signaling pathways, and cell adhesion molecules. Thus, we speculate that combination therapy targeting two or more of these processes may ameliorate thrombocytosis-mediated hypercoagulation. CONCLUSION: We present a first comprehensive molecular characterization of differences between two ARDS etiologies-COVID-19 and bacterial sepsis. Further investigation into the identified pathways will lead to a better understanding of the pathophysiological processes, potentially enabling novel therapeutic interventions.

Multi-omic comparative analysis of COVID-19 and bacterial sepsis-induced ARDS.

Background: Acute respiratory distress syndrome (ARDS), a life-threatening condition characterized by hypoxemia and poor lung compliance, is associated with high mortality. ARDS induced by COVID-19 has similar clinical presentations and pathological manifestations as non-COVID-19 ARDS. However, COVID-19 ARDS is associated with a more protracted inflammatory respiratory failure compared to traditional ARDS. Therefore, a comprehensive molecular comparison of ARDS of different etiologies groups may pave the way for more specific clinical interventions. Methods and Findings: In this study, we compared COVID-19 ARDS (n=43) and bacterial sepsis-induced (non-COVID-19) ARDS (n=24) using multi-omic plasma profiles covering 663 metabolites, 1,051 lipids, and 266 proteins. To address both between- and within-ARDS group variabilities we followed two approaches. First, we identified 706 molecules differently abundant between the two ARDS etiologies, revealing more than 40 biological processes differently regulated between the two groups. From these processes, we assembled a cascade of therapeutically relevant pathways downstream of sphingosine metabolism. The analysis suggests a possible overactivation of arginine metabolism involved in long-term sequelae of ARDS and highlights the potential of JAK inhibitors to improve outcomes in bacterial sepsis-induced ARDS. The second part of our study involved the comparison of the two ARDS groups with respect to clinical manifestations. Using a data-driven multi-omic network, we identified signatures of acute kidney injury (AKI) and thrombocytosis within each ARDS group. The AKI-associated network implicated mitochondrial dysregulation which might lead to post-ARDS renal-sequalae. The thrombocytosis-associated network hinted at a synergy between prothrombotic processes, namely IL-17, MAPK, TNF signaling pathways, and cell adhesion molecules. Thus, we speculate that combination therapy targeting two or more of these processes may ameliorate thrombocytosis-mediated hypercoagulation. Conclusion: We present a first comprehensive molecular characterization of differences between two ARDS etiologies - COVID-19 and bacterial sepsis. Further investigation into the identified pathways will lead to a better understanding of the pathophysiological processes, potentially enabling novel therapeutic interventions.

Urine-based multi-omic comparative analysis of COVID-19 and bacterial sepsis-induced ARDS.

Acute respiratory distress syndrome (ARDS), a life-threatening condition during critical illness, is a common complication of COVID-19. It can originate from various disease etiologies, including severe infections, major injury, or inhalation of irritants. ARDS poses substantial clinical challenges due to a lack of etiology-specific therapies, multisystem involvement, and heterogeneous, poor patient outcomes. A molecular comparison of ARDS groups holds the potential to reveal common and distinct mechanisms underlying ARDS pathogenesis. In this study, we performed a comparative analysis of urine-based metabolomics and proteomics profiles from COVID-19 ARDS patients (n = 42) and bacterial sepsis-induced ARDS patients (n = 17). The comparison of these ARDS etiologies identified 150 metabolites and 70 proteins that were differentially abundant between the two groups. Based on these findings, we interrogated the interplay of cell adhesion/extracellular matrix molecules, inflammation, and mitochondrial dysfunction in ARDS pathogenesis through a multi-omic network approach. Moreover, we identified a proteomic signature associated with mortality in COVID-19 ARDS patients, which contained several proteins that had previously been implicated in clinical manifestations frequently linked with ARDS pathogenesis. In summary, our results provide evidence for significant molecular differences in ARDS patients from different etiologies and a potential synergy of extracellular matrix molecules, inflammation, and mitochondrial dysfunction in ARDS pathogenesis. The proteomic mortality signature should be further investigated in future studies to develop prediction models for COVID-19 patient outcomes.

Serum proteomic profiling of rheumatoid arthritis-interstitial lung disease with a comparison to idiopathic pulmonary fibrosis.

Although interstitial lung disease (ILD) causes significant morbidity and mortality in rheumatoid arthritis (RA), it is difficult to predict the development or progression of ILD, emphasising the need for improved discovery through minimally invasive diagnostic tests. Aptamer-based proteomic profiling was used to assess 1321 proteins from 159 patients with rheumatoid arthritis with interstitial lung disease (RA-ILD), RA without ILD, idiopathic pulmonary fibrosis and healthy controls. Differential expression and gene set enrichment analyses revealed molecular signatures that are strongly associated with the presence and severity of RA-ILD and provided insight into unexplored pathways of disease. These warrant further study as non-invasive diagnostic tools and future therapeutic targets.

Cell-Type-Specific Expression of Hyaluronan Synthases HAS2 and HAS3 Promotes Goblet Cell Hyperplasia in Allergic Airway Inflammation.

Allergic rhinitis (AR) is a multifactorial airway disease characterized by basal and goblet cell hyperplasia. Hyaluronic acid (HA) is a major component of extracellular matrix and a critical contributor to tissue repair and remodeling after injury. We previously demonstrated that the intermediate progenitor cell (IPC) surface marker CD44v3 is upregulated in the basal and suprabasal layers of well-differentiated primary human nasal epithelial (HNE) cells after stimulation with the Th2 (T-helper cell type 2) cytokine IL-4, and an antibody blocking the CD44v3-HA interaction suppressed IL-4-induced goblet cell hyperplasia. We now show that the expression of HA and two HA synthases, HAS2 and HAS3, was upregulated in both the nasal surface epithelium of subjects with AR and IL-4-stimulated HNE cells. Inhibition of HA synthesis by 4-methylumbelliferone suppressed IL-4-induced goblet cell hyperplasia. Moreover, HAS2 and HAS3 were expressed in IPCs depending on the differentiation events, as follows: the rapid, transient upregulation of HAS2 induced basal IPC proliferation and basal-to-suprabasal transition, whereas the delayed upregulation of HAS3 promoted the transition of suprabasal IPCs to a goblet cell fate. 4-methylumbelliferone treatment in a house dust mite-induced murine AR model attenuated goblet cell metaplasia. Last, HA concentrations in nasal epithelial lining fluids from patients with AR positively correlated with the concentrations of mediators causing allergic inflammation. These data suggest that HA produced after the sequential upregulation of HAS2 and HAS3 contributes to goblet cell hyperplasia in allergic airway inflammation and modulates disease progression.