Features Of Acute Lung Injury Research Articles

Neutrophil Reduction Attenuates the Severity of Lung Injury in the Early Phase of Pneumococcal Pneumonia in Mice.

Neutrophils are the first leukocytes to be recruited to sites of inflammation in response to chemotactic factors released by activated macrophages and pulmonary epithelial and endothelial cells in bacterial pneumonia, a common cause of acute respiratory distress syndrome (ARDS). Although neutrophilic inflammation facilitates the elimination of pathogens, neutrophils also may cause bystander tissue injury. Even though neutrophils in alveolar spaces is a key feature of acute lung injury and ARDS especially from pneumonia, their contribution to the pathogenesis of lung injury is uncertain. The goal of this study was to elucidate the role of neutrophils in a clinically relevant model of bacterial pneumonia. We investigated the effect of reducing neutrophils in a mouse model of pneumococcal pneumonia treated with antibiotics. Neutrophils were reduced with anti-Ly6G monoclonal antibody 24 hours before and immediately preceding infection. Mice were inoculated intranasally with Streptococcus pneumoniae and received ceftriaxone 12 hours after bacterial inoculation. Neutrophil reduction in mice treated with ceftriaxone attenuated hypoxemia, alveolar permeability, epithelial injury, pulmonary edema, and inflammatory biomarker release induced by bacterial pneumonia, even though bacterial loads in the distal air spaces of the lung were modestly increased as compared to antibiotic treatment alone. Thus, when appropriate antibiotics are administered, lung injury in the early phase of bacterial pneumonia is mediated in part by neutrophils. In the early phase of bacterial pneumonia, neutrophils contribute to the severity of lung injury, although they also participate in host defense.

Hemoglobin-Mediated Oxidation of Low-Density Lipoprotein (oxLDL) Contributes to Lung Microvascular Endothelial Dysfunction Via Lectin-Like Oxidized LDL Receptor 1 (LOX-1)

Introduction: Lung microvascular endothelial barrier disruption is a critical feature of acute lung injury during several pathologies, including sepsis. Plasma cell-free hemoglobin (CFH) is elevated in approximately 80% of patients with sepsis and is independently associated with development of acute respiratory distress syndrome (ARDS) and mortality. However, the precise signaling mechanisms involved in CFH-mediated lung microvascular barrier dysfunction are incompletely understood. One potential mechanism by which CFH might disrupt the microvascular endothelial barrier is through its known ability to oxidize lipids and lipoproteins, including low-density lipoprotein (LDL). Oxidized LDL (oxLDL) is a known endothelial barrier disruptor, primarily signaling through scavenger receptors such as lectin-like oxidized LDL receptor 1 (LOX-1). We hypothesized that CFH-mediated oxidation of LDL exacerbates lung microvascular endothelial barrier dysfunction via LOX-1. Methods: For experimental studies, LDL was oxidized by combining LDL with CFH in a test tube overnight at 37°C and oxidation of LDL was quantified by TBARS assay. In primary human lung microvascular endothelial cells (HLMVEC), barrier dysfunction was assessed with expert permeability testing (XPerT) assay. HLMVEC were stimulated with vehicle control (PBS), LDL (0.05 mg/mL), CFH (0.5 mg/mL), or LDL oxidized by CFH (LDL+CFH) with or without one-hour pre-treatment of hemoglobin scavenger haptoglobin (1:1) or LOX-1 inhibitor BI-0115 (Boehringer Ingelheim, 10 μM). Cleaved caspase-3 (CST 9661, 1:500) was assessed with immunofluorescence microscopy. To test whether CFH and oxLDL are associated in clinical sepsis, circulating levels of CFH and oxLDL were measured in 60 sepsis patients via ELISA. Results: Oxidation of LDL by CFH (p=0.0006 by TBARS) exacerbated HLMVEC barrier dysfunction (p<0.0001 by XPerT) but was partially prevented by scavenging CFH with haptoglobin (p<0.0001 by XPerT) or blocking LOX-1 receptor (p<0.0001 by XPerT). Oxidation of LDL by CFH stimulation of HLMVEC also increased caspase-3 activation (p<0.0001 by immunofluorescence of cleaved caspase-3). In sepsis patients, circulating oxLDL levels correlated with CFH (r=0.4, p=0.002). Conclusions: Oxidation of LDL by CFH contributes to lung microvascular endothelial injury via LOX-1 receptor and activation of caspase-3; targeting this pathway may hold therapeutic potential to treat pathologies in which CFH is elevated, including sepsis-induced acute lung injury/ARDS. Funding: Parker B. Francis Fellowship (JEM); NIH K99HL166865 (JEM), R01HL158906 (LBW), R01HL164937 (LBW), R21GM144915 (LBW, JAB), R35HL150783 (JAB), RF1AG075341 (JAB). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

ACE2 and a Traditional Chinese Medicine Formula NRICM101 Could Alleviate the Inflammation and Pathogenic Process of Acute Lung Injury

The coronavirus disease 2019 (COVID-19) is a highly transmittable respiratory illness caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and acute lung injury (ALI) is the major complication of COVID-19. The challenge in studying SARS-CoV-2 pathogenicity is the limited availability of animal model. Therefore, it is necessary to establish animal models that can reproduce multiple characteristics of ALI to study therapeutic application. The present study established a mouse model that has features of ALI that are similar to COVID-19 syndrome to investigate the role of human angiotensin-converting enzyme type II (ACE2) and the administration of the Chinese herbal prescription Chingguan Yihau (NRICM101) in ALI. The mice with ACE2 genetic modifications, including the overexpression of human ACE2 (K18-hACE2 TG) and the absence of ACE2 (mACE2 KO), were intratracheally instillated with hydrochloric acid to induce ALI. The acid intratracheal instillation induced the imbalance between angiotensin-converting enzyme (ACE) and ACE2 (i.e., ACE/ACE2), severe immune cell infiltration, cytokine storms, inflammatory effect, and pulmonary disease in the mice. Compared with K18-hACE2 TG mice, mACE2 KO mice exhibited dramatically increased levels of multiple inflammatory cytokines Interleukin-6 (IL-6) and Tumor necrosis factor alpha (TNF-α) in bronchoalveolar lavage fluid, histological evidence of lung injury, the production of the ACE expression, and the activation of matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) expression. The results indicate that overproduction of ACE2 and induction of the ACE2-Ang-(1-7)/Mas axis could counteract the proinflammatory effect of acid-induced ALI and effectively alleviate the pathogenic process of acid-induced ALI by regulating MAPK, p-ERK1/2, and p-STAT3 pathways and dysregulation of MMP-2/MMP-9 activation. In addition, NRICM101, which is an effective TCM that has both antiviral and anti-inflammatory effects against COVID-19, shows potential for treating acid-induced ALI. NRICM101 could alleviate the disease progression of acid-induced ALI by suppressing IL-6 and TNF-α production in alveoli. In conclusion, the established mouse model provided an effective platform for researchers to investigate the role of ACE2 on pathological mechanisms and develop therapeutic strategies for ALI, including COVID-19-related pulmonary diseases. The effects of NRICM101 administration is a potential strategy for curing acid-induced ALI evaluated in the mouse model in this study. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Zinc-Dependent Histone Deacetylases in Lung Endothelial Pathobiology.

A monolayer of endothelial cells (ECs) lines the lumen of blood vessels and, as such, provides a semi-selective barrier between the blood and the interstitial space. Compromise of the lung EC barrier due to inflammatory or toxic events may result in pulmonary edema, which is a cardinal feature of acute lung injury (ALI) and its more severe form, acute respiratory distress syndrome (ARDS). The EC functions are controlled, at least in part, via epigenetic mechanisms mediated by histone deacetylases (HDACs). Zinc-dependent HDACs represent the largest group of HDACs and are activated by Zn2+. Members of this HDAC group are involved in epigenetic regulation primarily by modifying the structure of chromatin upon removal of acetyl groups from histones. In addition, they can deacetylate many non-histone histone proteins, including those located in extranuclear compartments. Recently, the therapeutic potential of inhibiting zinc-dependent HDACs for EC barrier preservation has gained momentum. However, the role of specific HDAC subtypes in EC barrier regulation remains largely unknown. This review aims to provide an update on the role of zinc-dependent HDACs in endothelial dysfunction and its related diseases. We will broadly focus on biological contributions, signaling pathways and transcriptional roles of HDACs in endothelial pathobiology associated mainly with lung diseases, and we will discuss the potential of their inhibitors for lung injury prevention.

Progression of Acute Lung Injury in Intratracheal LPS Rat Model: Efficacy of Fluticasone, Dexamethasone, and Pirfenidone

Introduction: We investigated the potential of LPS (10–300 µg/rat) administered intratracheally (i.t.) to induce reproducible features of acute lung injury (ALI) and compared the pharmacological efficacy of anti-inflammatory glucocorticoids and antifibrotic drugs to reduce the disease. Additionally, we studied the time-dependent progression of ALI in this LPS rat model. Methods: We conducted (1) dose effect studies of LPS administered i.t. at 10, 30, 100, and 300 μg/rat on ALI at 4 h timepoint; (2) pharmacological interventions using i.t. fluticasone (100 and 300 μg/rat), i.t. pirfenidone (4,000 μg/rat), and peroral dexamethasone (1 mg/kg) at 4 h timepoint; (3) kinetic studies at 0, 2, 4, 6, 8, 10, and 24 h post-LPS challenge. Phenotype or pharmacological efficacy was assessed using predetermined ALI features such as pulmonary inflammation, edema, and inflammatory mediators. Results: All LPS doses induced a similar increase of inflammation, edema, and inflammatory mediators, e.g., IL6, IL1β, TNFα, and CINC-1. In pharmacological intervention studies, we showed fluticasone and dexamethasone ameliorated ALI by inhibiting inflammation (>60–80%), edema (>70–100%), and the increase of cytokines IL6, IL1β, and TNFα (≥70–90%). We also noticed some inhibition of CINC-1 (25–35%) and TIMP1 (57%) increase with fluticasone and dexamethasone. Conversely, pirfenidone failed to inhibit inflammation, edema, and mediators of inflammation. Last, in ALI kinetic studies, we observed progressive pulmonary inflammation and TIMP1 levels, which peaked at 6 h and remained elevated up to 24 h. Progressive pulmonary edema started between 2 and 4 h and was sustained at later timepoints. On average, levels of IL6 (peak at 6–8 h), IL1β (peak at 2–10 h), TNFα (peak at 2 h), CINC-1 (peak at 2–6 h), and TGFβ1 (peak at 8 h) were elevated between 2 and 10 h and declined toward 24 h post-LPS challenge. Conclusion: Our data show that 10 μg/rat LPS achieved a robust, profound, and reproducible experimental ALI phenotype. Glucocorticoids ameliorated key ALI features at the 4-h timepoint, but the antifibrotic pirfenidone failed. Progressive inflammation and sustained pulmonary edema were present up to 24 h, whereas levels of inflammatory mediators were dynamic during ALI progression. This study’s data might be helpful in designing appropriate experiments to test the potential of new therapeutics to cure ALI.

ACE2 and a Traditional Chinese Medicine Formula NRICM101 Could Alleviate the Inflammation and Pathogenic Process of Acute Lung Injury.

COVID-19 is a highly transmittable respiratory illness caused by SARS-CoV-2, and acute lung injury (ALI) is the major complication of COVID-19. The challenge in studying SARS-CoV-2 pathogenicity is the limited availability of animal models. Therefore, it is necessary to establish animal models that can reproduce multiple characteristics of ALI to study therapeutic applications. The present study established a mouse model that has features of ALI that are similar to COVID-19 syndrome to investigate the role of ACE2 and the administration of the Chinese herbal prescription NRICM101 in ALI. Mice with genetic modifications, including overexpression of human ACE2 (K18-hACE2 TG) and absence of ACE2 (mACE2 KO), were intratracheally instillated with hydrochloric acid. The acid intratracheal instillation induced severe immune cell infiltration, cytokine storms, and pulmonary disease in mice. Compared with K18-hACE2 TG mice, mACE2 KO mice exhibited dramatically increased levels of multiple inflammatory cytokines (IL-6 and TNF-α) in bronchoalveolar lavage fluid, histological evidence of lung injury, and dysregulation of MAPK and MMP activation. In mACE2 KO mice, NRICM101 could ameliorate the disease progression of acid-induced ALI. In conclusion, the established mouse model provided an effective platform for researchers to investigate pathological mechanisms and develop therapeutic strategies for ALI, including COVID-19-related ALI.

Serine/Threonine Protein Phosphatases 1 and 2A in Lung Endothelial Barrier Regulation.

Vascular barrier dysfunction is characterized by increased permeability and inflammation of endothelial cells (ECs), which are prominent features of acute lung injury (ALI), acute respiratory distress syndrome (ARDS), and sepsis, and a major complication of the SARS-CoV-2 infection and COVID-19. Functional impairment of the EC barrier and accompanying inflammation arises due to microbial toxins and from white blood cells of the lung as part of a defensive action against pathogens, ischemia-reperfusion or blood product transfusions, and aspiration syndromes-based injury. A loss of barrier function results in the excessive movement of fluid and macromolecules from the vasculature into the interstitium and alveolae resulting in pulmonary edema and collapse of the architecture and function of the lungs, and eventually culminates in respiratory failure. Therefore, EC barrier integrity, which is heavily dependent on cytoskeletal elements (mainly actin filaments, microtubules (MTs), cell-matrix focal adhesions, and intercellular junctions) to maintain cellular contacts, is a critical requirement for the preservation of lung function. EC cytoskeletal remodeling is regulated, at least in part, by Ser/Thr phosphorylation/dephosphorylation of key cytoskeletal proteins. While a large body of literature describes the role of phosphorylation of cytoskeletal proteins on Ser/Thr residues in the context of EC barrier regulation, the role of Ser/Thr dephosphorylation catalyzed by Ser/Thr protein phosphatases (PPases) in EC barrier regulation is less documented. Ser/Thr PPases have been proposed to act as a counter-regulatory mechanism that preserves the EC barrier and opposes EC contraction. Despite the importance of PPases, our knowledge of the catalytic and regulatory subunits involved, as well as their cellular targets, is limited and under-appreciated. Therefore, the goal of this review is to discuss the role of Ser/Thr PPases in the regulation of lung EC cytoskeleton and permeability with special emphasis on the role of protein phosphatase 1 (PP1) and protein phosphatase 2A (PP2A) as major mammalian Ser/Thr PPases. Importantly, we integrate the role of PPases with the structural dynamics of the cytoskeleton and signaling cascades that regulate endothelial cell permeability and inflammation.

The Effect and Mechanism of Lipoxin A4 on Neutrophil Function in LPS-Induced Lung Injury.

Excessive inflammatory response caused by infiltration of a large number of neutrophils is one of the important features of acute lung injury (ALI)/acute respiratory distress syndrome (ARDS). Lipoxin A4 (LXA4) is an important endogenous mediator in the process of inflammation resolution, which has a strong role in promoting inflammation resolution. In this study, we examined the impact of LXA4 on the pulmonary inflammatory response and the neutrophil function in ARDS rats. Our results indicated that exogenous administration of LXA4 could reduce the degree of lung injury in ARDS rats and inhibit the release of pro-inflammatory factors TNF-α and IL-1β in lung tissue homogenate. However, LXA4 has no lung protective effect on ARDS rats of neutropenia, nor can it inhibit the levels of pro-inflammatory factors TNF-α and IL-1β in lung tissue homogenate. LXA4 can inhibit the production of reactive oxygen species (ROS) and neutrophil extracellular traps (NETs) in peripheral blood neutrophils of ARDS rats. At the same time, LXA4 can promote the phagocytosis of neutrophils in ARDS rats in vitro and can also promote the apoptosis of neutrophils in ARDS rats. In addition, the effect of LXA4 on the function of neutrophils in ARDS rats is mediated by its receptor ALX. LXA4 can inhibit the release of NE and MPO from neutrophils, thereby reducing the production of NETs. In summary, these findings indicate that LXA4 has a protective effect on LPS-induced ARDS rats by affecting the function of neutrophils.

Beneficial effects of aloperine on inflammation and oxidative stress by suppressing necroptosis in lipopolysaccharide-induced acute lung injury mouse model

RationaleAlveolar epithelial cell death, inflammation, and oxidative stress are typical features of acute lung injury (ALI). Aloperine (Alo), an alkaloid isolated from Sophora alopecuroides, has been reported to display various biological effects, such as anti-inflammatory, immunoregulatory, and anti-oxidant properties. In this study, we investigated the effects and mechanisms of Alo in treating a lipopolysaccharide (LPS)-induced ALI in a murine model. MethodsThe effects of Alo in LPS-induced ALI were investigated in C57BL/6 mice. The RIPK1 inhibitor (Nec-1) and the RIPK3 inhibitor (GSK’872) were used to evaluate the relationship of necroptosis, NF-κB activation, and PDC subunits in LPS-treated mouse alveolar epithelial cells (MLE-12). Then the effects of Alo on necroptosis, inflammation, and oxidative stress of LPS-stimulated MLE-12 cells were evaluated. ResultsAlo significantly attenuated histopathological lung injuries and reduced lung wet/dry ratio in LPS-induced ALI mice. Alo also remarkedly reduced total protein and neutrophils recruitment in bronchoalveolar lavage fluid of ALI mice. Meanwhile, Alo ameliorated the LPS-induced necroptosis in the lungs of ALI mice. The RIPK3 inhibitor GSK’872, but not the RIPK1 inhibitor Nec-1, reversed LPS-induced p65 phosphorylation and translocation to the nucleus in MLE-12 cells. GSK’872 also reversed the LPS-induced increase in ROS and binding of RIPK3 and PDC subunits in MLE-12 cells. Moreover, Alo down-regulated the levels of p-RIPK1, p-RIPK3, p-MLKL, p-p65, the translocation of p65 to the nucleus, and reduced the expression of IL-6 and IL-8 in LPS-stimulated MLE-12 cells. Alo also inhibited the binding of RIPK3 and PDC-E1α, PDC-E1β, PDC-E2, and PDC-E3 and the ROS production in LPS-treated MLE-12 cells. ConclusionThe present study validated the beneficial effects of Alo on LPS-induced ALI , suggesting Alo may be a new drug candidate against ALI.

Late Complications of COVID-19.

Studies of lungs in patients with COVID-19 have focused on early findings. To systematically study histopathologic and imaging features and presence of SARS-CoV-2 RNA in lung tissue from patients in later stages of COVID-19. Autopsies, explants, surgical lung biopsies, transbronchial biopsies, cryobiopsies, and needle biopsies from patients with COVID-19 whose onset of symptoms/confirmed diagnosis was more than 28 days before the procedure were studied. Available images were reviewed. Reverse transcription droplet digital polymerase chain reaction for SARS-CoV-2 RNA was performed on lung tissue. Of 44 specimens (43 patients; median age, 59.3 years; 26 [60.5%] male) features of acute lung injury (ALI) were seen in 39 (88.6%), predominantly organizing pneumonia and diffuse alveolar damage, up to 298 days after onset of COVID-19. Fibrotic changes were found in 33 specimens (75%), most commonly fibrotic diffuse alveolar damage (n = 22) and cicatricial organizing pneumonia (n = 12). Time between acquiring COVID-19 and specimen was shorter in patients with diffuse ALI (median, 61.5 days) compared with patients with focal (140 days) or no ALI (130 days) (P = .009). Sixteen (of 20; 80%) SARS-CoV-2 reverse transcription droplet digital polymerase chain reaction tests were positive, up to 174 days after COVID-19 onset. Time between COVID-19 onset and most recent computed tomography in patients with consolidation on imaging was shorter (median, 43.0 days) versus in patients without consolidation (87.5 days; P = .02). Reticulations were associated with longer time to computed tomography after COVID-19 onset (median, 82 versus 23.5 days; P = .006). ALI and SARS-CoV-2 RNA can be detected in patients with COVID-19 for many months. ALI may evolve into fibrotic interstitial lung disease.

Knockdown of receptor interacting protein 140 (RIP140) alleviated lipopolysaccharide-induced inflammation, apoptosis and permeability in pulmonary microvascular endothelial cells by regulating C-terminal binding protein 2 (CTBP2)

ABSTRACT The main pathological feature of acute lung injury (ALI) is pulmonary edema caused by increased permeability of pulmonary microvascular endothelial cells (PMVECs). LPS was has been confirmed to lead to cell damage and barrier dysfunction in PMVECs. Furthermore, receptor interacting protein 140 (RIP140) was discovered to be increased in LPS-induced human pulmonary microvascular endothelial cells (HPMECs), but the mechanism of RIP140 on LPS-induced HPMECs has not been investigated. In this study, an acute lung injury model was constructed in LPS-induced HPMECs. After RIP140 was downregulated, inflammation, apoptosis and cell permeability levels were detected by RT-qPCR, TUNEL staining and FITC-Dextran, respectively. Western blotting was used to detect the protein levels of related factors. The binding of RIP140 and C-terminal binding protein 2 (CTBP2) was predicted by database and verified by Co-IP. Subsequently, CTBP2 overexpression was transfected into cells and the above experiments were performed again. The results showed that inflammation, apoptosis and permeability levels of LPS-induced HPMECs were remarkably increased compared to the untreated control group. However, these levels were suppressed after RIP140 was silenced compared to the LPS-induced HPMECs group. Notably, the Co-IP study demonstrated that RIP140 and CTBP2 interacted with each other. Moreover, CTBP2 overexpression reversed the inhibitory effects of RIP140 silencing on LPS-induced inflammation, apoptosis and permeability levels in HPMECs. Together, the study found that interference of RIP140 could alleviate LPS-induced inflammation, apoptosis and permeability in HPMECs by regulating CTBP2.

Endotoxin stabilizes protein arginine methyltransferase 4 (PRMT4) protein triggering death of lung epithelia

Lung epithelial cell death is a prominent feature of acute lung injury and acute respiratory distress syndrome (ALI/ARDS), which results from severe pulmonary infection leading to respiratory failure. Multiple mechanisms are believed to contribute to the death of epithelia; however, limited data propose a role for epigenetic modifiers. In this study, we report that a chromatin modulator protein arginine N-methyltransferase 4/coactivator-associated arginine methyltransferase 1 (PRMT4/CARM1) is elevated in human lung tissues with pneumonia and in experimental lung injury models. Here PRMT4 is normally targeted for its degradation by an E3 ubiquitin ligase, SCFFBXO9, that interacts with PRMT4 via a phosphodegron to ubiquitinate the chromatin modulator at K228 leading to its proteasomal degradation. Bacterial-derived endotoxin reduced levels of SCFFBXO9 thus increasing PRMT4 cellular concentrations linked to epithelial cell death. Elevated PRMT4 protein caused substantial epithelial cell death via caspase 3-mediated cell death signaling, and depletion of PRMT4 abolished LPS-mediated epithelial cell death both in cellular and murine injury models. These findings implicate a unique molecular interaction between SCFFBXO9 and PRMT4 and its regulation by endotoxin that impacts the life span of lung epithelia, which may play a key role in the pathobiology of tissue injury observed during critical respiratory illness.

Comparative Study of Acute Lung Injury in COVID-19 and Non-COVID-19 Patients

Background: Amid the coronavirus disease 2019 (COVID-19) pandemic, we analyzed clinical characteristics of acute lung injury (ALI) in COVID-19 patients and reported their similarity and dissimilarity to those of non-COVID-19 patients in the intensive care unit (ICU).Methods: We reported on 90 COVID-19 and 130 non-COVID-19 ALI patients in the ICUs of multiple centers. Demographic data, medical histories, laboratory findings, and radiological images were analyzed and compared between the two cohorts and within each cohort between survivors and non-survivors. For ALI survivors, clinical characteristics before and after treatment were also compared.Findings: Aberrations in blood parameters, such as leukocytosis, neutrophilia, and thrombocytopenia, were observed in both cohorts. More characteristic abnormalities, including significantly higher red cell distribution width (RDW), C-reactive proteins, and lactic dehydrogenase (LDH) but lower troponin (TnT) and procalcitonin, were observed in the COVID-19 cohort than in the non-COVID-19 cohort, whereas D-dimer levels showed a similar elevation in both cohorts. The COVID-19 cohort also showed more diversified CT patterns where severe features such as consolidations and crazy paving patterns were more frequently observed. Multivariate analysis indicated that age, fever symptom, prothrombin time, procalcitonin, partial pressure of carbon dioxide, oxygenated hemoglobin, and crazy paving patterns in CT scans were independent risk factors associated with COVID-19.Interpretation: Comparison of ALI characteristics between COVID-19 and non-COVID-19 patients in the ICU setting provided insight into the pathogenesis of ALI induced by different risk factors, suggesting distinct treatment plans.

Group V Phospholipase A2 Mediates Endothelial Dysfunction and Acute Lung Injury Caused by Methicillin-Resistant Staphylococcus Aureus.

Lung endothelial dysfunction is a key feature of acute lung injury (ALI) and clinical acute respiratory distress syndrome (ARDS). Previous studies have identified the lipid-generating enzyme, group V phospholipase A2 (gVPLA2), as a mediator of lung endothelial barrier disruption and inflammation. The current study aimed to determine the role of gVPLA2 in mediating lung endothelial responses to methicillin-resistant Staphylococcus aureus (MRSA, USA300 strain), a major cause of ALI/ARDS. In vitro studies assessed the effects of gVPLA2 inhibition on lung endothelial cell (EC) permeability after exposure to heat-killed (HK) MRSA. In vivo studies assessed the effects of intratracheal live or HK-MRSA on multiple indices of ALI in wild-type (WT) and gVPLA2-deficient (KO) mice. In vitro, HK-MRSA increased gVPLA2 expression and permeability in human lung EC. Inhibition of gVPLA2 with either the PLA2 inhibitor, LY311727, or with a specific monoclonal antibody, attenuated the barrier disruption caused by HK-MRSA. LY311727 also reduced HK-MRSA-induced permeability in mouse lung EC isolated from WT but not gVPLA2-KO mice. In vivo, live MRSA caused significantly less ALI in gVPLA2 KO mice compared to WT, findings confirmed by intravital microscopy assessment in HK-MRSA-treated mice. After targeted delivery of gVPLA2 plasmid to lung endothelium using ACE antibody-conjugated liposomes, MRSA-induced ALI was significantly increased in gVPLA2-KO mice, indicating that lung endothelial expression of gVPLA2 is critical in vivo. In summary, these results demonstrate an important role for gVPLA2 in mediating MRSA-induced lung EC permeability and ALI. Thus, gVPLA2 may represent a novel therapeutic target in ALI/ARDS caused by bacterial infection.

Conditioned Medium of Bone Marrow Mesenchymal Stem Cells Involved in Acute Lung Injury by Regulating Epithelial Sodium Channels via miR-34c

BackgroundOne of the characteristics of acute lung injury (ALI) is severe pulmonary edema, which is closely related to alveolar fluid clearance (AFC). Mesenchymal stem cells (MSCs) secrete a wide range of cytokines, growth factors, and microRNA (miRNAs) through paracrine action to participate in the mechanism of pulmonary inflammatory response, which increase the clearance of edema fluid and promote the repair process of ALI. The epithelial sodium channel (ENaC) is the rate-limiting step in the sodium–water transport and edema clearance in the alveolar cavity; the role of bone marrow-derived MSC-conditioned medium (BMSC-CM) in edema clearance and how miRNAs affect ENaC are still seldom known.MethodsCCK-8 cell proliferation assay was used to detect the effect of BMSC-CM on the survival of alveolar type 2 epithelial (AT2) cells. Real-time polymerase chain reaction (RT-PCR) and western blot were used to detect the expression of ENaC in AT2 cells. The effects of miR-34c on lung fluid absorption were observed in LPS-treated mice in vivo, and the transepithelial short-circuit currents in the monolayer of H441 cells were examined by the Ussing chamber setup. Dual luciferase reporter gene assay was used to detect the target gene of miR-34c.ResultsBMSC-CM could increase the viability of mouse AT2 cells. RT-PCR and western blot results showed that BMSC-CM significantly increased the expression of the γ-ENaC subunit in mouse AT2 cells. MiR-34c could restore the AFC and lung wet/dry weight ratio in the ALI animal model, and Ussing chamber assay revealed that miR-34c enhanced the amiloride-sensitive currents associated with ENaC activity in intact H441 cell monolayers. In addition, we observed a higher expression of miR-34c in mouse AT2 cells administrated with BMSC-CM, and the overexpression or inhibition of miR-34c could regulate the expression of ENaC protein and alter the function of ENaC. Finally, we detected that myristoylated alanine-rich C kinase substrate (MARCKS) may be one of the target genes of miR-34c.ConclusionOur results indicate that BMSC-CM may alleviate LPS-induced ALI through miR-34c targeting MARCKS and regulate ENaC indirectly, which further explores the benefit of paracrine effects of bone marrow-derived MSCs on edematous ALI.

Actin-binding protein, IQGAP1, regulates LPS-induced RPMVECs hyperpermeability and ICAM-1 upregulation via Rap1/Src signalling pathway

Pulmonary microvascular barrier dysfunction is a hallmark feature of acute lung injury (ALI). IQGAP1 is a ubiquitously expressed scaffolding protein known to regulate cancer metastasis, angiogenesis, and barrier stability. However, the function of IQGAP1 in lipopolysaccharide (LPS)-induced microvascular endothelial hyperpermeability remains poorly understood. In the present study, we demonstrated that IQGAP1 was markedly upregulated in LPS-induced ALI models and rat pulmonary microvascular endothelial cells (RPMVECs). Lentivirus-mediated knockdown of IQGAP1 significantly attenuated the formation of actin stress fibers, phosphorylation of myosin light chain (MLC), and disruption of VE-cadherin, thereby protecting the RPMVECs barrier failure from LPS damage. In addition, IQGAP1 depletion reduced the reactive oxygen species (ROS)-mediated increase in intracellular adhesion molecule-1 (ICAM-1) in RPMVECs stimulated with LPS. Mechanistically, we found that the upregulation of IQGAP1 affected the activity of Rap1 and the downstream phosphorylation of Src. In conclusion, these findings reveal an essential mechanism by which increased IQGAP1 in LPS-treated RPMVECs promotes barrier dysfunction and ICAM-1 upregulation, at least in part by regulating Rap1/Src signalling, indicating that IQGAP1 may be a potential therapeutic target to prevent endothelial hyperpermeability and inflammation in ALI.

Aldosterone alleviates lipopolysaccharide-induced acute lung injury by regulating epithelial sodium channel through PI3K/Akt/SGK1 signaling pathway

Reduced alveolar fluid clearance (AFC) is a major pathological feature of acute lung injury (ALI). Epithelial sodium channel (ENaC) plays a key role in regulating the transport of Na+ and clearing alveolar edema fluid effectively. ENaC has been reported to be regulated by aldosterone in the distal collecting tube of the kidney. We hypothesized whether aldosterone regulated ENaC in alveolar epithelium and correspondingly played a role in ALI. In this study we found that the expression of aldosterone synthesis encoding gene, CYP11B2, and ENaC were decreased in the lung tissue of LPS-induced ALI mice. Furthermore, aldosterone alleviated ALI by increasing the expression of ENaC-α and relieving pulmonary edema. Besides, we found that aldosterone upregulated ENaC-α through PI3K/Akt/SGK1 pathway. In conclusion, our study demonstrated that aldosterone attenuated pulmonary edema by upregulating ENaC-α through the PI3K/Akt/SGK1 pathway in LPS-induced ALI, indicating that aldosterone might be a promising adjuvant drug for ALI treatment.

UCHL1, a deubiquitinating enzyme, regulates lung endothelial cell permeability in vitro and in vivo.

Increasing evidence suggests an important role for deubiquitinating enzymes (DUBs) in modulating a variety of biological functions and diseases. We previously identified the upregulation of the DUB ubiquitin carboxyl terminal hydrolase 1 (UCHL1) in murine ventilator-induced lung injury (VILI). However, the role of UCHL1 in modulating vascular permeability, a cardinal feature of acute lung injury (ALI) in general, remains unclear. We investigated the role of UCHL1 in pulmonary endothelial cell (EC) barrier function in vitro and in vivo and examined the effects of UCHL1 on VE-cadherin and claudin-5 regulation, important adherens and tight junctional components, respectively. Measurements of transendothelial electrical resistance confirmed decreased barrier enhancement induced by hepatocyte growth factor (HGF) and increased thrombin-induced permeability in both UCHL1-silenced ECs and in ECs pretreated with LDN-57444 (LDN), a pharmacological UCHL1 inhibitor. In addition, UCHL1 knockdown (siRNA) was associated with decreased expression of VE-cadherin and claudin-5, whereas silencing of the transcription factor FoxO1 restored claudin-5 levels. Finally, UCHL1 inhibition in vivo via LDN was associated with increased VILI in a murine model. These findings support a prominent functional role of UCHL1 in regulating lung vascular permeability via alterations in adherens and tight junctions and implicate UCHL1 as an important mediator of ALI.

Role of chest radiography in the management of COVID-19 pneumonia: An overview and correlation with pathophysiologic changes.

Background:Chest radiography (CXR) is a widely available baseline radiological modality in evaluating symptomatic patients with suspected or confirmed Covid-19 disease. Serial changes can help in monitoring the patients in conjunction with the clinical status of these patients in a hospital setting.Purpose:The purpose of this study was to analyse the patterns of radiological findings on chest radiograph (CXR) for suspected and confirmed COVID-19 patients on initial presentation to the emergency medical services (EMS) on admission and to assess the progression and resolution.Materials and Methods:In this study, patients who presented to EMS of a multispeciality hospital as suspected or confirmed Covid-19 on consecutive reverse transcriptase polymerase chain reaction (RT-PCR). CXR was examined for findings of haziness, patterns and distribution of opacities. Progression and regression of findings in serial CXR were studied and evaluated with the clinical and laboratory parameters. High resolution CT (HRCT) chest was performed initially for some patients.Results:756 RT-PCR confirmed COVID-19 patients were included in our study who had initial CXR. 510 (67.46%) of our patients with positive initial RT-PCR showed abnormal baseline CXR. The abnormal findings were described as haziness akin to ground glass opacities (GGO) on CT, peripheral opacities, patchy parenchymal opacities and consolidation. Peripheral opacities and lower zone distribution were the commonest pattern of CXR abnormalities with bilateral involvement. The severity of findings on serial CXR and radiographic regression was studied along with follow-up to assess response to treatment. Forty-six patients showed features of acute lung injury (ALI). Complications and new CXR findings were reported for patients who were given ventilator support.Conclusion:CXR is a valuable baseline radiological investigation on hospital admission in symptomatic patients with suspected or confirmed Covid-19 presenting to the EMS as it helps to monitor the progress and regression of the disease in conjunction with clinical findings.

Role of Dual Oxidases in Ventilator-induced Lung Injury.

Positive-pressure ventilation results in ventilator-induced lung injury, and few therapeutic modalities have been successful at limiting the degree of injury to the lungs. Understanding the primary drivers of ventilator-induced lung injury will aid in the development of specific treatments to ameliorate the progression of this syndrome. There are conflicting data for the role of neutrophils in acute respiratory distress syndrome pathogenesis. Here, we specifically examined the importance of neutrophils as a primary driver of ventilator-induced lung injury in a mouse model known to have impaired ability to recruit neutrophils in previous models of inflammation. We exposed Duoxa+/+ and Duoxa-/- mice to low- or high-tidal volume ventilation with or without positive end-expiratory pressure (PEEP) and recruitment maneuvers for 4 hours. Absolute neutrophils in BAL fluid were significantly reduced in Duoxa-/- mice compared with Duoxa+/+ mice (6.7 cells/μl; 16.4 cells/μl; P = 0.003), consistent with our hypothesis that neutrophil translocation across the capillary endothelium is reduced in the absence of DUOX1 or DUOX2 in response to ventilator-induced lung injury. Reduced lung neutrophilia was not associated with a reduction in overall lung injury in this study, suggesting that neutrophils do not play an important role in early features of acute lung injury. Surprisingly, Duoxa-/- mice exhibited significant hypoxemia, as measured by the arterial oxygen tension/fraction of inspired oxygen ratio and arterial oxygen content, which was out of proportion with that seen in the Duoxa+/+ mice (141, 257, P = 0.012). These findings suggest a role for dual oxidases to limit physiologic impairment during early ventilator-induced lung injury.