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At least 253 records · Page 14

A data-independent acquisition-based global phosphoproteomics system enables deep profiling

Phosphoproteomics can provide insights into cellular signaling dynamics. To achieve deep and robust quantitative phosphoproteomics profiling for minute amounts of sample, we here develop a global phosphoproteomics strategy based on data-independent acquisition (DIA) mass spectrometry and hybrid spectral libraries derived from data-dependent acquisition (DDA) and DIA data. Benchmarking the method using 166 synthetic phosphopeptides shows high sensitivity (<0.1 ng), accurate site localization and reproducible quantification (~5% median coefficient of variation). As a proof-of-concept, we use lung cancer cell lines and patient-derived tissue to construct a hybrid phosphoproteome spectral library covering 159,524 phosphopeptides (88,107 phosphosites). Based on this library, our single-shot streamlined DIA workflow quantifies 36,350 phosphosites (19,755 class 1) in cell line samples within two hours. Application to drug-resistant cells and patient-derived lung cancer tissues delineates site-specific phosphorylation events associated with resistance and tumor progression, showing that our workflow enables the characterization of phosphorylation signaling with deep coverage, high sensitivity and low between-run missing values.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

Characterization of SARS-CoV-2 Omicron BA.2.75 clinical isolates

The prevalence of the Omicron subvariant BA.2.75 rapidly increased in India and Nepal during the summer of 2022, and spread globally. However, the virological features of BA.2.75 are largely unknown. Here, we evaluated the replicative ability and pathogenicity of BA.2.75 clinical isolates in Syrian hamsters. Although we found no substantial differences in weight change among hamsters infected with BA.2, BA.5, or BA.2.75, the replicative ability of BA.2.75 in the lungs is higher than that of BA.2 and BA.5. Of note, BA.2.75 causes focal viral pneumonia in hamsters, characterized by patchy inflammation interspersed in alveolar regions, which is not observed in BA.5-infected hamsters. Moreover, in competition assays, BA.2.75 replicates better than BA.5 in the lungs of hamsters. These results suggest that BA.2.75 can cause more severe respiratory disease than BA.5 and BA.2 in a hamster model and should be closely monitored.

59 BASIC BIOLOGICAL SCIENCES↗

Genome-wide association study of long COVID

Abstract Infections can lead to persistent symptoms and diseases such as shingles after varicella zoster or rheumatic fever after streptococcal infections. Similarly, severe acute respiratory syndrome coronavirus 2 (SARS‑CoV‑2) infection can result in long coronavirus disease (COVID), typically manifesting as fatigue, pulmonary symptoms and cognitive dysfunction. The biological mechanisms behind long COVID remain unclear. We performed a genome-wide association study for long COVID including up to 6,450 long COVID cases and 1,093,995 population controls from 24 studies across 16 countries. We discovered an association ofFOXP4with long COVID, independent of its previously identified association with severe COVID-19. The signal was replicated in 9,500 long COVID cases and 798,835 population controls. Given the transcription factor FOXP4’s role in lung physiology and pathology, our findings highlight the importance of lung function in the pathophysiology of long COVID.

Genetics & Heredity↗

Concomitant immunity to M. tuberculosis infection

Some persistent infections provide a level of immunity that protects against reinfection with the same pathogen, a process referred to as concomitant immunity. To explore the phenomenon of concomitant immunity during Mycobacterium tuberculosis infection, we utilized HostSim, a previously published virtual host model of the immune response following Mtb infection. By simulating reinfection scenarios and comparing with data from non-human primate studies, we propose a hypothesis that the durability of a concomitant immune response against Mtb is intrinsically tied to levels of tissue resident memory T cells (Trms) during primary infection, with a secondary but important role for circulating Mtb-specific T cells. Further, we compare HostSim reinfection experiments to observational TB studies from the pre-antibiotic era to predict that the upper bound of the lifespan of resident memory T cells in human lung tissue is likely 2–3 years. To the authors’ knowledge, this is the first estimate of resident memory T-cell lifespan in humans. Our findings are a first step towards demonstrating the important role of Trms in preventing disease and suggest that the induction of lung Trms is likely critical for vaccine success.

60 APPLIED LIFE SCIENCES↗

Cosmic ray radiography of a human phantom

Cosmic ray muons that reach the earth's surface provide a natural source of radiation that is used for radiography. In this paper, we show that radiography using the cosmic radiation background provides a method that can be used to monitor bulk aspects of human anatomy. We describe a method that can be used to measure changes in patients as a function of time by cosmic ray muon radiography. Modeling shows muon tomography could provide hourly readouts of parameters such as lung density with sufficient sensitivity to detect the time changes in the inflammation of the lungs in, e.g., COVID patients.

60 APPLIED LIFE SCIENCES↗

Swine acute diarrhea syndrome coronavirus replication in primary human cells reveals potential susceptibility to infection

Zoonotic coronaviruses represent an ongoing threat, yet the myriads of circulating animal viruses complicate the identification of higher-risk isolates that threaten human health. Swine acute diarrhea syndrome coronavirus (SADS-CoV) is a newly discovered, highly pathogenic virus that likely evolved from closely related HKU2 bat coronaviruses, circulating in Rhinolophus spp. bats in China and elsewhere. As coronaviruses cause severe economic losses in the pork industry and swine are key intermediate hosts of human disease outbreaks, we synthetically resurrected a recombinant virus (rSADS-CoV) as well as a derivative encoding tomato red fluorescent protein (tRFP) in place of ORF3. rSADS-CoV replicated efficiently in a variety of continuous animal and primate cell lines, including human liver and rectal carcinoma cell lines. Of concern, rSADS-CoV also replicated efficiently in several different primary human lung cell types, as well as primary human intestinal cells. rSADS-CoV did not use human coronavirus ACE-2, DPP4, or CD13 receptors for docking and entry. Contemporary human donor sera neutralized the group I human coronavirus NL63, but not rSADS-CoV, suggesting limited human group I coronavirus cross protective herd immunity. Importantly, remdesivir, a broad-spectrum nucleoside analog that is effective against other group 1 and 2 coronaviruses, efficiently blocked rSADS-CoV replication in vitro. rSADS-CoV demonstrated little, if any, replicative capacity in either immune-competent or immunodeficient mice, indicating a critical need for improved animal models. Efficient growth in primary human lung and intestinal cells implicate SADS-CoV as a potential higher-risk emerging coronavirus pathogen that could negatively impact the global economy and human health.

59 BASIC BIOLOGICAL SCIENCES↗

Adrenal Stress Hormone Regulation of Hepatic Homeostatic Function After an Acute Ozone Exposure in Wistar-Kyoto Male Rats

Ozone-induced lung injury, inflammation, and pulmonary/hypothalamus gene expression changes are diminished in adrenalectomized (AD) rats. Acute ozone exposure induces metabolic alterations concomitant with increases in epinephrine and corticosterone. We hypothesized that adrenal hormones are responsible for observed hepatic ozone effects, and in AD rats, these changes would be diminished. In total, 5–7 days after sham (SH) or AD surgeries, male Wistar-Kyoto rats were exposed to air or 0.8-ppm ozone for 4 h. Serum samples were analyzed for metabolites and liver for transcriptional changes immediately post-exposure. Ozone increased circulating triglycerides, cholesterol, free fatty-acids, and leptin in SH but not AD rats. Ozone-induced inhibition of glucose-mediated insulin release was absent in AD rats. Unlike diminution of ozone-induced hypothalamus and lung mRNA expression changes, AD in air-exposed rats (AD-air/SH-air) caused differential hepatic expression of ~1000 genes. Likewise, ozone in AD rats caused differential expression of ~1000 genes (AD-ozone/AD-air). Ozone-induced hepatic changes in SH rats reflected enrichment for pathways involving metabolic processes, including acetyl-CoA biosynthesis, TCA cycle, and sirtuins. Upstream predictor analysis identified similarity to responses produced by glucocorticoids and pathways involving forskolin. These changes were absent in AD rats exposed to ozone. However, ozone caused unique changes in AD liver mRNA reflecting activation of synaptogenesis, neurovascular coupling, neuroinflammation, and insulin signaling with inhibition of senescence pathways. In these rats, upstream predictor analysis identified numerous microRNAs likely involved in glucocorticoid insufficiency. These data demonstrate the critical role of adrenal stress hormones in ozone-induced hepatic homeostasis and necessitate further research elucidating their role in propagating environmentally driven diseases.

Toxicology↗

Clinical characteristics and aetiological analysis of 133 patients for pulmonary embolism combined with haemoptysis

Abstract Background In the patients with pulmonary embolism (PE), PE itself can cause haemoptysis and other reasons can also cause haemoptysis. Therefore, the clinical characteristics and the causes of haemoptysis are lacking. Methods A retrospective analysis was performed that involved screening 583 PE patients and determining that haemoptysis occurred in 141 cases. Of these, eight cases were omitted due to anticoagulation‐related haemoptysis or unavailable data, leaving 133 cases that were enrolled in final analysis (127 acute and 6 chronic case of PE). We classified the acute PE patients who combined with diseases which can cause haemoptysis to non‐simple group (n = 61) and those without these diseases to simple group (n = 66). Results The incidence of haemoptysis in PE patients was 23.75%. In the simple group, the amount of haemoptysis ≤ 5 mL was 80.30% (53/66) and ≤ 20 mL was 90.91% (60/66). In the non‐simple group who combined with lung cancer, the amount of haemoptysis ≤ 5 mL was 68.4% (26/38) and ≤ 20 mL was 86.8% (33/38). Further analyses revealed that the amount of haemoptysis in the non‐simple group was larger than that in the simple group (median 5 [5‐125] vs. 5 [5‐5], p < 0.001; volume ≥ 100 mL: 29.5% vs. 6.1%, p < 0.001). Among all the PE patients, chronic thromboembolic pulmonary hypertension (CTEPH), PE combined with tuberculosis (TB) and PE combined with bronchiectasis were independent risk factors for the amount of haemoptysis ≥ 100 mL (OR = 15.00, (95% CI: 2.235‐100.652); 12.00, (3.101‐46.437); 60.00, (6.552‐549.441), respectively). Conclusions The haemoptysis caused by acute PE or PE combined with lung cancer was mild and was characterised by blood in sputum. PE combined with TB, bronchiectasis and CTEPH are associated with moderate to massive haemoptysis, with a greater risk of haemoptysis ≥ 100 mL.

Zou, Yong Wei↗

3C-like protease inhibitors block coronavirus replication in vitro and improve survival in MERS-CoV–infected mice

Pathogenic coronaviruses are a major threat to global public health, as exemplified by severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), and the newly emerged SARS-CoV-2, the causative agent of coronavirus disease 2019 (COVID-19). We describe herein the structure-guided optimization of a series of inhibitors of the coronavirus 3C-like protease (3CLpro), an enzyme essential for viral replication. The optimized compounds were effective against several human coronaviruses including MERS-CoV, SARS-CoV, and SARS-CoV-2 in an enzyme assay and in cell-based assays using Huh-7 and Vero E6 cell lines. Two selected compounds showed antiviral effects against SARS-CoV-2 in cultured primary human airway epithelial cells. In a mouse model of MERS-CoV infection, administration of a lead compound 1 day after virus infection increased survival from 0 to 100% and reduced lung viral titers and lung histopathology. These results suggest that this series of compounds has the potential to be developed further as antiviral drugs against human coronaviruses.

60 APPLIED LIFE SCIENCES↗

Characterization of a 2016-2017 Human Seasonal H3 Influenza A Virus Spillover Now Endemic to U.S. Swine

In 2017, the Iowa State University Veterinary Diagnostic Laboratory detected a reverse-zoonotic transmission of a human seasonal H3 influenza A virus into swine (IAV-S) in Oklahoma. Pairwise comparison between the recently characterized human seasonal H3 IAV-S (H3.2010.2) hemagglutinin (HA) sequences detected in swine and the most similar 2016–2017 human seasonal H3 revealed 99.9% nucleotide identity. To elucidate the origin of H3.2010.2 IAV-S, 45 HA and 27 neuraminidase (NA) sequences from 2017 to 2020 as well as 11 whole-genome sequences (WGS) were genetically characterized. Time to most recent common human ancestor was estimated between August and September 2016. The N2 NA was of human origin in all but one strain from diagnostic submissions with NA sequences, and the internal gene segments from WGS consisted of matrix genes originating from the 2009 pandemic H1N1 and another 5 internal genes of triple reassortant swine origin (TTTTPT). Pigs experimentally infected with H3.2010.2 demonstrated efficient nasal shedding and replication in the lungs, mild pneumonia, and minimal microscopic lung lesions and transmitted the virus to indirect contact swine. Antigenically, H3.2010.2 viruses were closer to a human seasonal vaccine strain, A/Hong Kong/4801/2014, than to the H3.2010.1 human seasonal H3 viruses detected in swine in 2012. This was the second sustained transmission of a human seasonal IAV into swine from the 2010 decade after H3.2010.1. Monitoring the spillover and detection of novel IAV from humans to swine may help vaccine antigen selection and could impact pandemic preparedness.

59 BASIC BIOLOGICAL SCIENCES↗

Early immune response to Coccidioides is characterized by robust neutrophil and fibrotic macrophage recruitment and differentiation

Coccidioidomycosis, or Valley fever, is an emerging respiratory disease caused by soil-dwelling fungi of the Coccidioides genus that is expected to spread from the southwest into the central U.S. by 2050. While 60% of infections are asymptomatic, the other 40% of patients experience a range of symptoms, from self-limiting pneumonia to life-threatening disseminated disease. The immunological events that underlie the progression to severe disease remain underdefined. Here, we probed the early immune response to Coccidioides using a high dose of an attenuated strain of Coccidioides posadasii in a mouse model of infection coupled with single-cell RNA sequencing. At 24 h post-infection, robust immune infiltration is detected in the lung, marked by high levels of inflammatory PD-L1 + neutrophils and fungal-contact-dependent pro-fibrotic Spp1 + macrophages. These findings elucidate the early dynamics of the host response to Coccidioides and provide a deeper understanding of host-pathogen interactions in the lung.

Coccidioides↗

Rat bronchoalveolar lavage proteome changes following e-cigarette aerosol exposures

E-cigarette liquids are complex mixtures of chemicals consisting of humectants, such as propylene glycol (PG) and vegetable glycerin (VG), with nicotine or flavorings added. Published literature emphasizes the toxicity of e-cigarette aerosols with flavorings whereas much less attention has been given to the biologic effects of humectants. The purpose of the current study was to provide a comprehensive view of the acute biologic effects of e-cigarette aerosols on rat bronchoalveolar lavage (BAL) using mass spectrometry-based global proteomics. Sprague–Dawley rats were exposed to e-cigarette aerosol for 3 h/day for three consecutive days. Groups included: PG/VG alone, PG/VG + 2.5% nicotine (N), or PG/VG + N + 3.3% vanillin (V). Right lung lobes were lavaged for BAL and supernatants prepared for proteomics. Extracellular BAL S100A9 concentrations and BAL cell staining for citrullinated histone H3 (citH3) were also performed. From global proteomics, ~2,100 proteins were identified from rat BAL. Overall, the greatest change in number of BAL proteins occurred with PG/VG exposures alone compared with controls with biological pathways enriched for acute phase responses, extracellular trap formation, and coagulation. Extracellular BAL S100A9 concentrations and the number of citH3 + BAL cells also increased significantly in PG/VG and PG/VG + 2.5% N. In contrast to PG/VG or PG/VG + N, the addition of vanillin to PG/VG + N increased BAL neutrophilia and downregulated lipid transport proteins. In summary, global proteomics support e-cigarette aerosol exposures to PG/VG alone as having a significant biologic effect on the lung independent of nicotine or flavoring with increased markers of extracellular trap formation.

59 BASIC BIOLOGICAL SCIENCES↗

Effect of Electroacupuncture at Wushu Acupoints of the Cardiopulmonary Meridian on the Autophagy in Rats with Acute Myocardial Ischemia

Wushu acupoints are the five acupoints distributed below the human elbow and knee joint. They are all located on the same meridian and divided into five categories: Jing, Ying, Shu, Jing, and He. It has been shown that electroacupuncture (EA) at Shenmen point of heart meridian can improve acute myocardial ischemia (AMI) early. However, it is still unclear if all the Wushu acupoints of the heart meridian can improve AMI. Hence, this study emphasizes Wushu acupoints of heart meridian, compares them with Wushu acupoints of lung meridian, and studies the therapeutic effect of EA at Wushu acupoints on AMI and its possible mechanism. It also discusses the specificity of the heart meridian to heart disease. The AMI model is established by ligation of the left anterior descending coronary artery. The detection methods like the physiological recorder, TTC staining, ELISA, and so forth were used to determine the ECG, myocardial infarct size, serum myocardial enzymes, and myocardial tissue-related protein expression in rats. The heart rate (HR) and ST segment along with creatine kinase (CK), creatine kinase isoenzymes (CK-MB), lactate dehydrogenase (LDH), and myocardial infarctions increased after the induction with AMI. Furthermore, the expressions of PINK1 and Parkin protein also showed an increase. However, EA at Wushu acupoints in the heart meridian can reverse the above changes, whereas EA at the lung meridian exhibits limited effect. It is depicted that the heart meridian has a relatively specific relationship with the heart in a diseased state.

Zhu, Chao↗

MIP-4 is Induced by Bleomycin and Stimulates Cell Migration Partially via Nir-1 Receptor

Background. CC-chemokine ligand 18 also known as MIP-4 is a chemokine with roles in inflammation and immune responses. It has been shown that MIP-4 is involved in the development of several diseases including lung fibrosis and cancer. How exactly MIP-4 is regulated and exerts its role in lung fibrosis remains unclear. Therefore, in the present study, we examined how MIP-4 is regulated and whether it acts via its potential receptor Nir-1. Materials and Methods. A549 cells were grown and maintained in DMEM : F12 (1 : 1) and supplemented with 10% FBS and 1000 U of penicillin/streptomycin and maintained as recommended by the manufacturer (ATCC). Cell migration and invasion, immunohistochemistry (IHC), Western blot, qPCR, and siRNA Nir-1 were used to determine MIP-4 regulation and its role in cell migration. Results. Cell migration was increased following stimulation of cells with recombinant (r) MIP-4 and bleomycin (BLM), whereas quenching rMIP-4 with its antibody (Ab) or addition of the Ab to BLM or H 2 O 2 diminished rMIP-4-induced cell migration. Along with cell migration, rMIP-4, BLM, and H 2 O 2 induced the formation of actin filaments dynamic structures whereas costimulation with MIP-4 Ab limited BLM- and H 2 O 2 -induced effects. MIP-4 mRNA and protein were increased by BLM and H 2 O 2 , and the addition of its Ab significantly reduced treatments effect. Experiments with siRNA investigating whether Nir-1 is a potential MIR-4 receptor indicated that the inhibition of Nir-1 decreased cell migration/invasion but did not totally inhibit rMIP-4-induced cell migration. Conclusion. Therefore, our data indicate that MIP-4 is regulated by BLM and H 2 O 2 and costimulation with its Ab limits the effects on MIP-4 and that the Nir-1 receptor partially mediates MIP-4’s effects on increased cell migration. These data also evidenced that MIP-4 is regulated by fibrotic and oxidative stimuli and that quenching MIP-4 with its Ab or therapeutically targeting the Nir-1 receptor may partially limit MIP-4 effects under fibrotic or oxidative stimulation.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

multiBreath.py

This is a research code which is intended to be made available to researchers to the password-protected simtk.org website which hosts the lung deposition simulation project "In-silico LADDER: Lung Aerosol Dosimetry for Drug and Environmental Research" headed by NIH PI Prof. Chantal Darquenne (UCSD). Industrial applications are more accurate deposition predictions of medical aerosols, including potentially subject-specific exposures. Accessing the code requires registration.

Kuprat, Andrew [Pacific Northwest National Laborat↗

COVID-19 Outcomes by Cancer Status, Site, Treatment, and Vaccination

Background: Studies have shown an increased risk of severe SARS-CoV-2–related (COVID-19) disease outcome and mortality for patients with cancer, but it is not well understood whether associations vary by cancer site, cancer treatment, and vaccination status. Methods: Using electronic health record data from an academic medical center, we identified a retrospective cohort of 260,757 individuals tested for or diagnosed with COVID-19 from March 10, 2020, to August 1, 2022. Of these, 52,019 tested positive for COVID-19 of whom 13,752 had a cancer diagnosis. We conducted Firth-corrected logistic regression to assess the association between cancer status, site, treatment, vaccination, and four COVID-19 outcomes: hospitalization, intensive care unit admission, mortality, and a composite “severe COVID” outcome. Results: Cancer diagnosis was significantly associated with higher rates of severe COVID, hospitalization, and mortality. These associations were driven by patients whose most recent initial cancer diagnosis was within the past 3 years. Chemotherapy receipt, colorectal cancer, hematologic malignancies, kidney cancer, and lung cancer were significantly associated with higher rates of worse COVID-19 outcomes. Vaccinations were significantly associated with lower rates of worse COVID-19 outcomes regardless of cancer status. Conclusions: Patients with colorectal cancer, hematologic malignancies, kidney cancer, or lung cancer or who receive chemotherapy for treatment should be cautious because of their increased risk of worse COVID-19 outcomes, even after vaccination. Impact: Additional COVID-19 precautions are warranted for people with certain cancer types and treatments. Further, significant benefit from vaccination is noted for both cancer and cancer-free patients.

60 APPLIED LIFE SCIENCES↗

Broadly neutralizing humanized SARS-CoV-2 antibody binds to a conserved epitope on Spike and provides antiviral protection through inhalation-based delivery in non-human primates

The COVID-19 pandemic represents a global challenge that has impacted and is expected to continue to impact the lives and health of people across the world for the foreseeable future. The rollout of vaccines has provided highly anticipated relief, but effective therapeutics are required to further reduce the risk and severity of infections. Monoclonal antibodies have been shown to be effective as therapeutics for SARS-CoV-2, but as new variants of concern (VoC) continue to emerge, their utility and use have waned due to limited or no efficacy against these variants. Furthermore, cumbersome systemic administration limits easy and broad access to such drugs. As well, concentrations of systemically administered antibodies in the mucosal epithelium, a primary site of initial infection, are dependent on neonatal Fc receptor mediated transport and require high drug concentrations. To reduce the viral load more effectively in the lung, we developed an inhalable formulation of a SARS-CoV-2 neutralizing antibody binding to a conserved epitope on the Spike protein, ensuring pan-neutralizing properties. Administration of this antibody via a vibrating mesh nebulization device retained antibody integrity and resulted in effective distribution of the antibody in the upper and lower respiratory tract of non-human primates (NHP). In comparison with intravenous administration, significantly higher antibody concentrations can be obtained in the lung, resulting in highly effective reduction in viral load post SARS-CoV-2 challenge. This approach may reduce the barriers of access and uptake of antibody therapeutics in real-world clinical settings and provide a more effective blueprint for targeting existing and potentially emerging respiratory tract viruses.

60 APPLIED LIFE SCIENCES↗

PNNL DataHub Project: Omics Lethal Human Viruses Project Profiling of the Host Response to Influenza A Virus Infection, Processed Experimental Dataset Catalog

Influenza A virus (IAV) is a high risk biological agent, classified as a Category C priority pathogen (Orthomyxoviridae) by the National Institute of Allergy and Infectious Diseases (NIAID), and is known to cause severe respiratory disease with high mortality rates in humans. Lethal host-pathogen invasion mechanisms and the cellular intricacies behind these fatal infections still remain unclear. The NIAID Modeling Host Responses to Understand Severe Human Virus Infections Research Program project (2013-2018) aimed to develop an improved comprehensive understanding of the host response to a suite of viruses causing lethal infections leveraging a systems biology approach. Herein, PNNL sub-projects provide a never before released comprehensive infectious disease collection of primary and secondary transformation multi-Omics data profiling a series of priority pathogen primary experimental studies for enhanced open-access to viral Omics datasets and project lifecycle metadata. Secondary host-pathogen viral dataset downloads contain one or more statistically processed (normalization data transformation) quantitative dataset collections resulting in qualitative expression analyses of primary host-pathogen experimental study designs. Leveraging unique high-resolution Omics capabilities for proteomics (P), metabolomics (M), lipidomics (L), and transcriptomics (T) dataset downloads each have a direct relationship to a primary sample submission corresponding to a specific Influenza A virus [NCBITAXON:11320] experimental infection study. Host sample types include human lung adenocarcinoma cells ["Calu-3", BTO:0002750] and whole mouse lung [BTO:0000763] tissue collections.

59 BASIC BIOLOGICAL SCIENCES↗