Modulation of alveolar macrophage and mitochondrial fitness by medicinal plant-derived nanovesicles to mitigate acute lung injury and viral pneumonia.

Acute lung injury (ALI) is generally caused by severe respiratory infection and characterized by overexuberant inflammatory responses and inefficient pathogens-containing, the two major processes wherein alveolar macrophages (AMs) play a central role. Dysfunctional mitochondria have been linked with distorted macrophages and hence lung disorders, but few treatments are currently available to correct these defects. Plant-derive nanovesicles have gained significant attention because of their therapeutic potential, but the targeting cells and the underlying mechanism remain elusive. We herein prepared the nanovesicles from Artemisia annua, a well-known medicinal plant with multiple attributes involving anti-inflammatory, anti-infection, and metabolism-regulating properties. By applying three mice models of acute lung injury caused by bacterial endotoxin, influenza A virus (IAV) and SARS-CoV-2 pseudovirus respectively, we showed that Artemisia-derived nanovesicles (ADNVs) substantially alleviated lung immunopathology and raised the survival rate of challenged mice. Macrophage depletion and adoptive transfer studies confirmed the requirement of AMs for ADNVs effects. We identified that gamma-aminobutyric acid (GABA) enclosed in the vesicles is a major molecular effector mediating the regulatory roles of ADNVs. Specifically, GABA acts on macrophages through GABA receptors, promoting mitochondrial gene programming and bioenergy generation, reducing oxidative stress and inflammatory signals, thereby enhancing the adaptability of AMs to inflammation resolution. Collectively, this study identifies a promising nanotherapeutics for alleviating lung pathology, and elucidates a mechanism whereby the canonical neurotransmitter modifies AMs and mitochondria to resume tissue homeostasis, which may have broader implications for treating critical pulmonary diseases such as COVID-19.

Favipiravir ameliorates bleomycin-induced pulmonary fibrosis by reprogramming M1/M2 macrophage polarization.

Corona Virus Disease 2019 (COVID-19) is an infectious disease that seriously endangers human life and health. The pathological anatomy results of patients who died of the COVID-19 showed that there was an excessive inflammatory response in the lungs. It is also known that most of the COVID-19 infected patients will cause different degrees of lung damage after infection, and may have pulmonary fibrosis remaining after cure. Macrophages are a type of immune cell population with pluripotency and plasticity. In the early and late stages of infection, the dynamic changes of the balance and function of M1/M2 alveolar macrophages have a significant impact on the inflammatory response of the lungs. In the early stage of pulmonary fibrosis inflammation, the increase in the proportion of M1 type is beneficial to clear pathogenic microorganisms and promote the progress of inflammation; in the later stage of fibrosis, the increase in the number of M2 type macrophages can inhibit the inflammatory response and promote the degradation of fibrosis. As a potential treatment drug for new coronavirus pneumonia, favipiravir is in the process of continuously carried out relevant clinical trials. This study aims to discuss whether the antiviral drug favipiravir can suppress inflammation and immune response by regulating the M1/M2 type of macrophages, thereby alleviating fibrosis. We established a bleomycin-induced pulmonary fibrosis model, using IL-4/13 and LPS/IFN-γ cell stimulating factor to induce macrophage M1 and M2 polarization models, respectively. Our study shows that favipiravir exerts anti-fibrotic effects mainly by reprogramming M1/M2 macrophages polarization, that is, enhancing the expression of anti-fibrotic M1 type, reducing the expression of M2 type pro-fibrotic factors and reprogramming it to anti-fibrotic phenotype. Aspects of pharmacological mechanisms, favipiravir inhibits the activation of JAK2-STAT6 and JAK2-PI3K-AKT signaling by targeting JAK2 protein, thereby inhibiting pro-fibrotic M2 macrophages polarization and M2-induced myofibroblast activation. In summary, favipiravir can reduce the progression of pulmonary fibrosis, we hope to provide a certain reference for the treatment of pulmonary fibrosis.

Recombinant Rod Domain of Vimentin Reduces SARS-CoV-2 Viral Replication by Blocking Spike Protein-ACE2 Interactions.

Although the SARS-CoV-2 vaccination is the primary preventive intervention, there are still few antiviral therapies available, with current drugs decreasing viral replication once the virus is intracellular. Adding novel drugs to target additional points in the viral life cycle is paramount in preventing future pandemics. The purpose of this study was to create and test a novel protein to decrease SARS-CoV-2 replication. We created the recombinant rod domain of vimentin (rhRod) in E. coli and used biolayer interferometry to measure its affinity to the SARS-CoV-2 S1S2 spike protein and the ability to block the SARS-CoV-2-ACE2 interaction. We performed plaque assays to measure rhRod's effect on SARS-CoV-2 replication in Vero E6 cells. Finally, we measured lung inflammation in SARS-CoV-2-exposed K18-hACE transgenic mice given intranasal and intraperitoneal rhRod. We found that rhRod has a high affinity for the S1S2 protein with a strong ability to block S1S2-ACE2 interactions. The daily addition of rhRod decreased viral replication in Vero E6 cells starting at 48 h at concentrations >1 µM. Finally, SARS-CoV-2-infected mice receiving rhRod had decreased lung inflammation compared to mock-treated animals. Based on our data, rhRod decreases SARS-CoV-2 replication in vitro and lung inflammation in vivo. Future studies will need to evaluate the protective effects of rhRod against additional viral variants and identify the optimal dosing scheme that both prevents viral replication and host lung injury.

Lactobacillus plantarum L168 improves hyperoxia-induced pulmonary inflammation and hypoalveolarization in a rat model of bronchopulmonary dysplasia.

Alteration of gut microbiota can affect chronic lung diseases, such as asthma and chronic obstructive pulmonary disease, through abnormal immune and inflammatory responses. Previous studies have shown a feasible connection between gut microbiota and bronchopulmonary dysplasia (BPD) in preterm infants. However, whether BPD can be ameliorated by restoring the gut microbiota remains unclear. In preterm infants with BPD, we found variance in the diversity and structure of gut microbiota. Similarly, BPD rats showed gut dysbiosis, characterized by a deficiency of Lactobacillus, which was abundant in normal rats. We therefore explored the effect and potential mechanism of action of a probiotic strain, Lactobacillus plantarum L168, in improving BPD. The BPD rats were treated with L. plantarum L168 by gavage for 2 weeks, and the effect was evaluated by lung histopathology, lung function, and serum inflammatory markers. Subsequently, we observed reduced lung injury and improved lung development in BPD rats exposed to L. plantarum L168. Further evaluation revealed that L. plantarum L168 improved intestinal permeability in BPD rats. Serum metabolomics showed altered inflammation-associated metabolites following L. plantarum L168 intervention, notably a marked increase in anti-inflammatory metabolites. In agreement with the metabolites analysis, RNA-seq analysis of the intestine and lung showed that inflammation and immune-related genes were down-regulated. Based on the information from RNA-seq, we validated that L. plantarum L168 might improve BPD relating to down-regulation of TLR4 /NF-κB /CCL4 pathway. Together, our findings suggest the potential of L. plantarum L168 to provide probiotic-based therapeutic strategies for BPD.

Pla2g5 contributes to viral-like-induced lung inflammation through macrophage proliferation and LA/Ffar1 lung cell recruitment.

Macrophages expressing group V phospholipase A2 (Pla2g5) release the free fatty acid (FFA) linoleic acid (LA), potentiating lung type 2 inflammation. Although Pla2g5 and LA increase in viral infections, their role remains obscure. We generated Pla2g5flox/flox mice, deleted Pla2g5 by using the Cx3cr1cre transgene, and activated bone marrow-derived macrophages (BM-Macs) with poly:IC, a synthetic double-stranded RNA that triggers a viral-like immune response, known Pla2g5-dependent stimuli (IL-4, LPS + IFNγ, IL-33 + IL-4 + GM-CSF) and poly:IC + LA followed by lipidomic and transcriptomic analysis. Poly:IC-activated Pla2g5flox/flox;Cx3cr1cre/+ BM-Macs had downregulation of major bioactive lipids and critical enzymes producing those bioactive lipids. In addition, AKT phosphorylation was lower in poly:IC-stimulated Pla2g5flox/flox;Cx3cr1cre/+ BM-Macs, which was not restored by adding LA to poly:IC-stimulated BM-Macs. Consistently, Pla2g5flox/flox;Cx3cr1cre/+ mice had diminished poly:IC-induced lung inflammation, including inflammatory macrophage proliferation, while challenging Pla2g5flox/flox;Cx3cr1cre/+ mice with poly:IC + LA partially restored lung inflammation and inflammatory macrophage proliferation. Finally, mice lacking FFA receptor-1 (Ffar1)-null mice had reduced poly:IC-induced lung cell recruitment and tissue macrophage proliferation, not corrected by LA. Thus, Pla2g5 contributes to poly:IC-induced lung inflammation by regulating inflammatory macrophage proliferation and LA/Ffar1-mediated lung cell recruitment and tissue macrophage proliferation.

Qingfei Tongluo Mixture Attenuates Bleomycin-Induced Pulmonary Inflammation and Fibrosis through mTOR-Dependent Autophagy in Rats.

As an interstitial fibrosis disease characterized by diffuse alveolitis and structural alveolar disorders, idiopathic pulmonary fibrosis (IPF) has high lethality but lacks limited therapeutic drugs. A hospital preparation used for the treatment of viral pneumonia, Qingfei Tongluo mixture (QFTL), is rumored to have protective effects against inflammatory and respiratory disease. This study aims to confirm whether it has a therapeutic effect on bleomycin-induced IPF in rats and to elucidate its mechanism of action. Male SD rats were randomly divided into the following groups: control, model, CQ + QFTL (84 mg/kg chloroquine (CQ) + 3.64 g/kg QFTL), QFTL-L, M, H (3.64, 7.28, and 14.56 g/kg, respectively) and pirfenidone (PFD 420 mg/kg). After induction modeling and drug intervention, blood samples and lung tissue were collected for further detection. Body weight and lung coefficient were examined, combined with hematoxylin and eosin (H&E) and Masson staining to observe lung tissue lesions. The enzyme-linked immunosorbent assay (ELISA) and the hydroxyproline (HYP) assay kit were used to detect changes in proinflammatory factors (transforming growth factor-ß (TGF-ß), tumor necrosis factor-α (TNF-α), and interleukin-1ß (IL-1ß)) and HYP. Immunohistochemistry and Western blotting were performed to observe changes in proteins related to pulmonary fibrosis (α-smooth muscle actin (α-SMA) and matrix metalloproteinase 12 (MMP12)) and autophagy (P62 and mechanistic target of rapamycin (mTOR)). Treatment with QFTL significantly improved the adverse effects of bleomycin on body weight, lung coefficient, and pathological changes. Then, QFTL reduced bleomycin-induced increases in proinflammatory mediators and HYP. The expression changes of pulmonary fibrosis and autophagy marker proteins are attenuated by QFTL. Furthermore, the autophagy inhibitor CQ significantly reversed the downward trend in HYP levels and α-SMA protein expression, which QFTL improved in BLM-induced pulmonary fibrosis rats. In conclusion, QFTL could effectively attenuate bleomycin-induced inflammation and pulmonary fibrosis through mTOR-dependent autophagy in rats. Therefore, QFTL has the potential to be an alternative treatment for IPF in clinical practice.

Fatal attractions that trigger inflammation and drive atherosclerotic disease.

BACKGROUND: Atherosclerosis is the salient, underlying cause of cardiovascular diseases, such as arrhythmia, coronary artery disease, cardiomyopathy, pulmonary embolism and myocardial infarction. In recent years, atherosclerosis pathophysiology has evolved from a lipid-based to an inflammation-centric ideology. METHODS: This narrative review is comprised of review and original articles that were found through the PubMed search engine. The following search terms or amalgamation of terms were used: "cardiovascular disease," "atherosclerosis," "inflammation," "GRP78," "Hsp60," "oxidative low-density lipoproteins," "aldehyde dehydrogenase," "ß2-glycoprotein," "lipoprotein lipase A," "human cytomegalovirus." "SARS-CoV-2," "chlamydia pneumonia," "autophagy," "thrombosis" and "therapeutics." RESULTS: Emerging evidence supports the concept that atherosclerosis is associated with the interaction between cell surface expression of stress response chaperones, including GRP78 and Hsp60, and their respective autoantibodies. Moreover, various other autoantigens and their autoantibodies have displayed a compelling connection with the development of atherosclerosis, including oxidative low-density lipoproteins, aldehyde dehydrogenase, ß2-glycoprotein and lipoprotein lipase A. Atherosclerosis progression is also concurrent with viral and bacterial activators of various diseases. This narrative review will focus on the contributions of human cytomegalovirus as well as SARS-CoV-2 and chlamydia pneumonia in atherosclerosis development. Notably, the interaction of an autoantigen with their respective autoantibodies or the presence of a foreign antigen can enhance inflammation development, which leads to atherosclerotic lesion progression. CONCLUSION: We will highlight and discuss the complex role of the interaction between autoantigens and autoantibodies, and the presence of foreign antigens in the development of atherosclerotic lesions in relationship to pro-inflammatory responses.

Impact of pulmonary infection after radical esophagectomy on serum inflammatory markers, pulmonary function indices, and prognosis.

OBJECTIVES: To analyze the influence of pulmonary infection after radical esophagectomy on serum inflammatory markers, pulmonary function, and prognosis. METHODS: We enrolled 278 esophageal cancer patients who underwent radical esophagectomy. Patients were split into the infected (n=51) and uninfected groups (n=227). The inflammatory parameters, complications, and prognosis were compared. RESULTS: In the infected group, interleukin (IL)-6 was 16.19±2.63 ng/L, tumor necrosis factor-α was 19.64±3.07 µg/L, and IL-1ß was 22.49±5.13 ng/L at 7 days postoperatively; white blood cell counts was 12.65±2.14 ×109/L, percentage of neutrophils (NEU%) was 67.04±10.48%, and platelet (PLT) counts was 249.82±63.26 ×109/L; the increasing ranges of the above factors after the operation were much raised compared with the uninfected group (p<0.05). Compared with the uninfected group, forced expiratory volume in one second (FEV1), forced vital capacity (FVC), and FEV1/FVC were greater declines in ranges (p<0.05), and the arrhythmia incidence and the mortality within 60 days postoperatively were greater in the infected group (p<0.05). CONCLUSION: Postoperative pulmonary infection can lead to pulmonary function damage, proinflammatory factor overexpression, and an increased risk of early death.

Impaired STING Activation Due to a Variant in the E3 Ubiquitin Ligase AMFR in a Patient with Severe VZV Infection and Hemophagocytic Lymphohistiocytosis.

Varicella zoster virus (VZV) is a neurotropic alphaherpesvirus exclusively infecting humans, causing two distinct pathologies: varicella (chickenpox) upon primary infection and herpes zoster (shingles) following reactivation. In susceptible individuals, VZV can give rise to more severe clinical manifestations, including disseminated infection, pneumonitis, encephalitis, and vasculopathy with stroke. Here, we describe a 3-year-old boy in whom varicella followed a complicated course with thrombocytopenia, hemorrhagic and necrotic lesions, pneumonitis, and intermittent encephalopathy. Hemophagocytic lymphohistiocytosis (HLH) was strongly suspected and as the condition deteriorated, HLH therapy was initiated. Although the clinical condition improved, longstanding hemophagocytosis followed despite therapy. We found that the patient carries a rare monoallelic variant in autocrine motility factor receptor (AMFR), encoding a ubiquitin ligase involved in innate cytosolic DNA sensing and interferon (IFN) production through the cyclic GMP-AMP synthase-stimulator of IFN genes (cGAS-STING) pathway. Peripheral blood mononuclear cells (PBMCs) from the patient exhibited impaired signaling downstream of STING in response dsDNA and 2'3'-cGAMP, agonists of cGAS and STING, respectively, and fibroblasts from the patient showed impaired type I IFN responses and significantly increased VZV replication. Overexpression of the variant AMFR R594C resulted in decreased K27-linked STING ubiquitination compared to WT AMFR. Moreover, ImageStream technology revealed reduced STING trafficking from ER to Golgi in cells expressing the patient AMFR R594C variant. This was supported by a dose-dependent dominant negative effect of expression of the patient AMFR variant as measured by IFN-ß reporter gene assay. Finally, lentiviral transduction with WT AMFR partially reconstituted 2'3'-cGAMP-induced STING-mediated signaling and ISG expression in patient PBMCs. This work links defective AMFR-STING signaling to severe VZV disease and hyperinflammation and suggests a direct role for cGAS-STING in the control of viral infections in humans. In conclusion, we describe a novel genetic etiology of severe VZV disease in childhood, also representing the first inborn error of immunity related to a defect in the cGAS-STING pathway.

Severe Acute Respiratory Syndrome Coronavirus 2 Pneumonia in Critically Ill Patients: A Cluster Analysis According to Baseline Characteristics, Biological Features, and Chest CT Scan on Admission.

BACKGROUND: Inconsistent results from COVID-19 studies raise the issue of patient heterogeneity. OBJECTIVE: The objective of this study was to identify homogeneous subgroups of patients (clusters) using baseline characteristics including inflammatory biomarkers and the extent of lung parenchymal lesions on CT, and to compare their outcomes. DESIGN: Retrospective single-center study. SETTING: Medical ICU of the University Hospital of Clermont-Ferrand, France. PATIENTS: All consecutive adult patients aged greater than or equal to 18 years, admitted between March 20, 2020, and August 31, 2021, for COVID-19 pneumonia. INTERVENTIONS: Characteristics at baseline, during ICU stay, and outcomes at day 60 were recorded. On the chest CT performed at admission the extent of lung parenchyma lesions was established by artificial intelligence software. MEASUREMENTS AND MAIN RESULTS: Clusters were determined by hierarchical clustering on principal components using principal component analysis of admission characteristics including plasma interleukin-6, human histocompatibility leukocyte antigen-DR expression rate on blood monocytes (HLA-DR) monocytic-expression rate (mHLA-DR), and the extent of lung parenchymal lesions. Factors associated with day 60 mortality were investigated by univariate survival analysis. Two hundred seventy patients were included. Four clusters were identified and three were fully described. Cluster 1 (obese patients, with moderate hypoxemia, moderate extent of lung parenchymal lesions, no inflammation, and no down-regulation of mHLA-DR) had a better prognosis at day 60 (hazard ratio [HR] = 0.27 [0.15-0.46], p < 0.01), whereas cluster 2 (older patients with comorbidities, moderate extent of lung parenchyma lesions but significant hypoxemia, inflammation, and down-regulation of mHLA-DR) and cluster 3 (patients with severe parenchymal disease, hypoxemia, inflammatory reaction, and down-regulation of mHLA-DR) had an increased risk of mortality (HR = 2.07 [1.37-3.13], p < 0.01 and HR = 1.52 [1-2.32], p = 0.05, respectively). In multivariate analysis, only clusters 1 and 2 were independently associated with day 60 death. CONCLUSIONS: Three clusters with distinct characteristics and outcomes were identified. Such clusters could facilitate the identification of targeted populations for the next trials.

The proteolytic airway environment associated with pneumonia acts as a barrier for treatment with anti-infective antibodies.

Like pneumonia, coronavirus disease 2019 (COVID-19) is characterized by a massive infiltration of innate immune cells (such as polymorphonuclear leukocytes) into the airways and alveolar spaces. These cells release proteases that may degrade therapeutic antibodies and thus limit their effectiveness. Here, we investigated the in vitro and ex vivo impact on anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) IgG1s and other IgG subclasses (IgG2 and IgG4) of the neutrophil elastase, proteinase 3 and cathepsin G (the three main neutrophil serine proteases) found in endotracheal aspirates from patients with severe COVID-19. Although the IgGs were sensitive to neutrophil serine proteases, IgG2 was most resistant to proteolytic degradation. The two anti-SARS CoV2 antibodies (casirivimab and imdevimab) were sensitive to the lung's proteolytic environment, although neutrophil serine protease inhibitors only partly limited the degradation. Overall, our results show that the pneumonia-associated imbalance between proteases and their inhibitors in the airways contributes to degradation of antiviral antibodies.

Luteolin-7-O-glucoside promotes macrophage release of IFN-ß by maintaining mitochondrial function and corrects the disorder of glucose metabolism during RSV infection.

Respiratory syncytial virus (RSV) pneumonia is the main cause of acute bronchiolitis in infants. Luteolin-7-O-glucoside (LUT-7G) is a natural flavonoid, which exists in a variety of plants and has the potential to treat viral pneumonia. We established RSV pneumonia mouse models and RSV-infected cell models. Clodronate liposomes were used to deplete macrophages. We used HE staining and immunohistochemistry to determine inflammatory damage and virus replication. We detected the expression levels of inflammatory factors and IFN-ß through qPCR and ELISA. JC-1 kit was used for detecting the cell mitochondrial Membrane potential (MMP). ROS, SOD, and MDA kits were used for detecting intracellular oxidative stress damage. Metabolites of TCA in lung tissue and serum of mice were detected by GC-MS. Pharmacodynamic studies have shown that intervention with LUT-7G can alleviate lung tissue damage caused by RSV infection, inhibit RSV replication, and downregulate TNF-α, IL-1ß, and IL-6 mRNA expression. LUT-7G upregulated the IFN-ß content and the expression of IFN-ß, ISG15, and OAS1 mRNA. In vitro, LUT-7G inhibited RSV-induced cell death, reversed the RSV-induced decrease of MMP and decreased intracellular oxidative stress. Target metabonomics showed that RSV infection upregulated the levels of glycolysis and TCA metabolites in lung tissue and serum, while LUT-7G could improve the disorder of glucose metabolism. The results indicate that LUT-7G can promote the release of IFN-ß in the lung, alleviate inflammatory damage, and inhibit RSV replication during RSV infection. These effects may be achieved by protecting the mitochondrial function of alveolar macrophages and correcting the disorder of glucose metabolism.

Implications of LDH in patients with coronavirus disease 2019 pneumonia.

Objective: The objective of this study was to explore the value of serum lactic dehydrogenase (LDH) in the early diagnosis and prognostic evaluation of pneumonia associated with the novel coronavirus infection. Methods: A total of 101 patients with coronavirus disease 2019 (COVID-19) pneumonia were included in the study. According to the severity of the initial chest computed tomography (CT), the patients were divided into the ordinary pneumonia group and the severe pneumonia group and then divided into the remission group and the nonremission group according to the changes of the chest CT after medication treatment. The differences in general characteristics, underlying diseases, clinical symptoms, laboratory findings, and imaging examination outcomes between groups were observed retrospectively. To analyze the diagnostic performance of LDH, receiver operating characteristic (ROC) curves were constructed and the area under the curve (AUC) was calculated. Results: Compared with ordinary pneumonia patients, patients in the severe group presented with significantly higher LDH, neutrophil count, high-sensitivity troponin T (HS-TnT), C-reactive protein (CRP), human serum amyloid A (SAA), N-terminal pro-brain natriuretic peptide (NTproBNP), and D-dimer. Compared with remission patients, non-remission patients presented with significantly higher LDH, neutrophil count, HS-TnT, CRP, SAA, procalcitonin (PCT), creatine kinase-MB mass (CKMB_M), NTproBNP, and D-dimer. In multivariate logistic regression analysis, we found that LDH [odds ratio (OR), 1.015; 95% confidence interval (CI), 1.006-1024; p = 0.001] and neutrophil count (OR, 1.352; 95% CI, 1.008-1.811; p = 0.044) were independently associated with exacerbation in COVID-19 patients. For ROC analysis, the AUC was 0.833 (95% CI, 0.729-0.936; p < 0.001) when we use the LDH value of 256.69 U/L to discriminate the ordinary pneumonia and severe pneumonia patients. The AUC was 0.759 (95% CI, 0.603-0.914; p = 0.008) and the sensitivity is 92.3% when we combined the LDH (cutoff value 258.46 U/L) and the neutrophil count (cutoff value 6.76 × 109/L) to discriminate remission and non-remission patients. Conclusion: The level of LDH is associated with the severity of COVID-19 pneumonia and can be used as important indicators to evaluate the prognosis of patients.

Metatranscriptome of human lung microbial communities in a cohort of mechanically ventilated COVID-19 Omicron patients.

The Omicron variant of the severe acute respiratory syndrome coronavirus 2 (SARS­CoV­2) infected a substantial proportion of Chinese population, and understanding the factors underlying the severity of the disease and fatality is valuable for future prevention and clinical treatment. We recruited 64 patients with invasive ventilation for COVID-19 and performed metatranscriptomic sequencing to profile host transcriptomic profiles, plus viral, bacterial, and fungal content, as well as virulence factors and examined their relationships to 28-day mortality were examined. In addition, the bronchoalveolar lavage fluid (BALF) samples from invasive ventilated hospital/community-acquired pneumonia patients (HAP/CAP) sampled in 2019 were included for comparison. Genomic analysis revealed that all Omicron strains belong to BA.5 and BF.7 sub-lineages, with no difference in 28-day mortality between them. Compared to HAP/CAP cohort, invasive ventilated COVID-19 patients have distinct host transcriptomic and microbial signatures in the lower respiratory tract; and in the COVID-19 non-survivors, we found significantly lower gene expressions in pathways related viral processes and positive regulation of protein localization to plasma membrane, higher abundance of opportunistic pathogens including bacterial Alloprevotella, Caulobacter, Escherichia-Shigella, Ralstonia and fungal Aspergillus sydowii and Penicillium rubens. Correlational analysis further revealed significant associations between host immune responses and microbial compositions, besides synergy within viral, bacterial, and fungal pathogens. Our study presents the relationships of lower respiratory tract microbiome and transcriptome in invasive ventilated COVID-19 patients, providing the basis for future clinical treatment and reduction of fatality.

Characterization of CCoV-HuPn-2018 spike protein-mediated viral entry.

Canine coronavirus-human pneumonia-2018 (CCoV-HuPn-2018) was recently isolated from a child with pneumonia. This novel human pathogen resulted from cross-species transmission of a canine coronavirus. It has been known that CCoV-HuPn-2018 uses aminopeptidase N (APN) from canines, felines, and porcines, but not humans, as functional receptors for cell entry. The molecular mechanism of cell entry in CCoV-HuPn-2018 remains poorly understood. In this study, we demonstrated that among the nine APN orthologs tested, the APN of the Mexican free-tailed bat could also efficiently support CCoV-HuPn-2018 spike (S) protein-mediated entry, raising the possibility that bats may also be an alternative host epidemiologically important for the transmission of this virus. The glycosylation at residue N747 of canine APN is critical for its receptor activity. The gain of glycosylation at the corresponding residues in human and rabbit APNs converted them to functional receptors for CCoV-HuPn-2018. Interestingly, the CCoV-HuPn-2018 spike protein pseudotyped virus infected multiple human cancer cell lines in a human APN-independent manner, whereas sialic acid appeared to facilitate the entry of the pseudotyped virus into human cancer cells. Moreover, while host cell surface proteases trypsin and TMPRSS2 did not promote the entry of CCoV-HuPn-2018, endosomal proteases cathepsin L and B are required for the entry of CCoV-HuPn-2018 in a pH-dependent manner. IFITMs and LY6E are host restriction factors for the CCoV-HuPn-2018 entry. Our results thus suggest that CCoV-HuPn-2018 has not yet evolved to be an efficient human pathogen. Collectively, this study helps us understand the cell tropism, receptor usage, cross-species transmission, natural reservoir, and pathogenesis of this potential human coronavirus. IMPORTANCE Viral entry is driven by the interaction between the viral spike protein and its specific cellular receptor, which determines cell tropism and host range and is the major constraint to interspecies transmission of coronaviruses. Aminopeptidase N (APN; also called CD13) is a cellular receptor for HCoV-229E, the newly discovered canine coronavirus-human pneumonia-2018 (CCoV-HuPn-2018), and many other animal alphacoronaviruses. We examined the receptor activity of nine APN orthologs and found that CCoV-HuPn-2018 utilizes APN from a broad range of animal species, including bats but not humans, to enter host cells. To our surprise, we found that CCoV-HuPn-2018 spike protein pseudotyped viral particles successfully infected multiple human hepatoma-derived cell lines and a lung cancer cell line, which is independent of the expression of human APN. Our findings thus provide mechanistic insight into the natural hosts and interspecies transmission of CCoV-HuPn-2018-like coronaviruses.

Circulating markers of extracellular matrix remodelling in severe COVID-19 patients.

BACKGROUND: Abnormal remodelling of the extracellular matrix (ECM) has generally been linked to pulmonary inflammation and fibrosis and may also play a role in the pathogenesis of severe COVID-19. To further elucidate the role of ECM remodelling and excessive fibrogenesis in severe COVID-19, we examined circulating levels of mediators involved in various aspects of these processes in COVID-19 patients. METHODS: Serial blood samples were obtained from two cohorts of hospitalised COVID-19 patients (n = 414). Circulating levels of ECM remodelling mediators were quantified by enzyme immunoassays in samples collected during hospitalisation and at 3-month follow-up. Samples were related to disease severity (respiratory failure and/or treatment at the intensive care unit), 60-day total mortality and pulmonary pathology after 3-months. We also evaluated the direct effect of inactivated SARS-CoV-2 on the release of the different ECM mediators in relevant cell lines. RESULTS: Several of the measured markers were associated with adverse outcomes, notably osteopontin (OPN), S100 calcium-binding protein A12 and YKL-40 were associated with disease severity and mortality. High levels of ECM mediators during hospitalisation were associated with computed tomography thorax pathology after 3-months. Some markers (i.e. growth differential factor 15, galectin 3 and matrix metalloproteinase 9) were released from various relevant cell lines (i.e. macrophages and lung cell lines) in vitro after exposure to inactivated SARS-CoV-2 suggesting a direct link between these mediators and the causal agent of COVID-19. CONCLUSION: Our findings highlight changes to ECM remodelling and particularly a possible role of OPN, S100A12 and YKL-40 in the pathogenesis of severe COVID-19.

Inhibitory Effect of Salicin on Staphylococcus aureus Coagulase.

The massive use of antibiotics has resulted in an alarming increase in antibiotic resistance in Staphylococcus aureus (S. aureus). This study aimed to identify the inhibitory effect of salicin on S. aureus. Coagulase (Coa) activity was assessed using in vitro Coa assays and Western blot, thermal shift assay (TSA), fluorescence quenching and molecular docking experiments were conducted to verify the interaction between salicin and Coa. An in vivo mouse pneumonia model demonstrated that salicin can reduce the virulence of S. aureus. In vitro Coa assays elucidated that salicin directly inhibited Coa activity. The Western blot and TSA results suggested that salicin did not block the expression of Coa but affected the thermal stability of the protein by binding to Coa. The fluorescence quenching, molecular docking and molecular dynamics assays have found that the most promising binding site between salicin and Coa was GLN-97. The pneumonia model of mice infected with S. aureus revealed that salicin could not only reduce the content of lung bacteria in mice but also prolong their survival. Salicin was identified as a novel anti-infective candidate compound with the potential to target Coa and inhibit its activity by binding to it, which would facilitate the development of roadmaps for future research.

Influenza H7N9 virus disrupts the monolayer human brain microvascular endothelial cells barrier in vitro.

Influenza H7N9 virus causes human infections with about 40% case fatality rate. The severe cases usually present with pneumonia; however, some present with central nervous system complications. Pneumonia syndrome is attributed to the cytokine storm after infection with H7N9, but the pathogenic mechanism of central nervous system complications has not been clarified. This study used immortalized human brain microvascular endothelial cells hCMEC/D3 to simulate the blood-brain barrier. It demonstrated that H7N9 virus could infect brain microvascular endothelial cells and compromise the blood-brain barrier integrity and permeability by down-regulating the expression of cell junction-related proteins, including claudin-5, occludin, and vascular endothelial (VE)-cadherin. These results suggested that H7N9 could infect the blood-brain barrier in vitro and affect its functions, which could be a potential mechanism for the pathogenesis of H7N9 viral encephalopathy.

Metabolomics Study Revealing Purines as Potential Diagnostic Biomarkers of Acute Respiratory Distress Syndrome in Patients with Community─Acquired Pneumonia.

Community-acquired pneumonia (CAP) is a significant threat to human health and the leading cause of acute respiratory distress syndrome (ARDS). We aimed to reveal the metabolic profiling whether can be used for assessing CAP with or without ARDS (nARDS) and therapeutic effects on CAP patients after treatment. Urine samples were collected at the onset and recovery periods, and metabolomics was employed to identify robust biomarkers. 19 metabolites were significantly changed in the ARDS relative to nARDS, mainly involving purines and fatty acids. After treatment, 7 metabolites in the nARDS and 14 in the ARDS were found to be significantly dysregulated, including fatty acids and amino acids. In the validation cohort, we observed that the biomarker panel consisted of N2,N2-dimethylguanosine, 1-methyladenosine, 3-methylguanine, 1-methyladenosine, and uric acid exhibited better AUCs of 0.900 than pneumonia severity index and acute physiology and chronic health evaluation II (APACHE II) scores between the ARDS and nARDS. Combining L-phenylalanine, phytosphingosine, and N-acetylaspartylglutamate as biomarkers for discriminating the nARDS and ARDS patients after treatment exhibited good AUCs of 0.811 and 0.821, respectively. The metabolic pathway and defined biomarkers may serve as crucial indicators for predicting the development of ARDS in CAP patients and for assessing therapeutic effects.

Th2-dependent disappearance and phenotypic conversion of mouse alveolar macrophages.

Alveolar macrophages (alvMs) play an important role for maintenance of lung function by constant removal of cellular debris in the alveolar space. They further contribute to defense against microbial or viral infections and limit tissue damage during acute lung injury. alvMs arise from embryonic progenitor cells, seed the alveoli before birth, and have life-long self-renewing capacity. However, recruited monocytes may also help to restore the alvM population after depletion caused by toxins or influenza virus infection. At present, the population dynamics and cellular plasticity of alvMs during allergic lung inflammation is poorly defined. To address this point, we used a mouse model of Aspergillus fumigatus-induced allergic lung inflammation and observed that Th2-derived IL-4 and IL-13 caused almost complete disappearance of alvMs. This effect required STAT6 expression in alvMs and also occurred in various other settings of type 2 immunity-mediated lung inflammation or administration of IL-4 complexes to the lung. In addition, Th2 cells promoted conversion of alvMs to alternatively activated macrophages and multinucleated giant cells. Given the well-established role of alvMs for maintenance of lung function, this process may have implications for resolution of inflammation and tissue homeostasis in allergic asthma.