Natural antibodies targeting LPS in pleural effusions of various etiologies.

Respiratory infection, cancer, and heart failure can cause abnormal accumulation of fluid in the pleural cavity. The immune responses within the cavity are orchestrated by leucocytes that reside in the serosal-associated lymphoid tissue. Natural antibodies (NAbs) are abundant in the serum (S) having a major role in systemic and mucosal immunity; however, their occurrence in pleural fluid (PF) remains an open question. Our aim herein was to detect and measure the levels of NAbs (IgM, IgG, IgA) targeting lipopolysaccharides (LPS) in both the pleural fluid and the serum of 78 patients with pleural effusions (PEs) of various etiologies. The values of anti-LPS NAb activity were extracted through a normalization step regarding the total IgM, IgG, and IgA levels, all determined by in-house ELISA. In addition, the ratios of PF/S values were analyzed further with other critical biochemical parameters from pleural fluids. Anti-LPS NAbs of all Ig classes were detected in most of the samples, while a significant increase of anti-LPS activity was observed in infectious and noninfectious compared with malignant PEs. Multivariate linear regression confirmed a negative correlation of IgM and IgA anti-LPS PF/S ratio with malignancy. Moreover, anti-LPS NAbs PF/S measurements led to increased positive and negative predictive power in ROC curves generated for the discrimination between benign and malignant PEs. Our results highlight the role of anti-LPS NAbs in the pleural cavity and demonstrate the potential translational impact that should be further explored.NEW & NOTEWORTHY Here we describe the detection and quantification of natural antibodies (NAbs) in the human pleural cavity. We show for the first time that IgM, IgG, and IgA anti-LPS natural antibodies are detected and measured in pleural effusions of infectious, noninfectious, and malignant etiologies and provide clinical correlates to demonstrate the translational impact of our findings.

2-Deoxy-glucose ameliorates the peritoneal mesothelial and endothelial barrier function perturbation occurring due to Peritoneal Dialysis fluids exposure.

Peritoneal dialysis (PD) and prolonged exposure to PD fluids (PDF) induce peritoneal membrane (PM) fibrosis and hypervascularity, leading to functional PM degeneration. 2-deoxy-glucose (2-DG) has shown potential as PM antifibrotic by inhibiting hyper-glycolysis induced mesothelial-to-mesenchymal transition (MMT). We investigated whether administration of 2-DG with several PDF affects the permeability of mesothelial and endothelial barrier of the PM. The antifibrotic effect of 2-DG was confirmed by the gel contraction assay with embedded mesothelial (MeT-5A) or endothelial (EA.hy926) cells cultured in Dianeal® 2.5 % (CPDF), BicaVera® 2.3 % (BPDF), Balance® 2.3 % (LPDF) with/without 2-DG addition (0.2 mM), and qPCR for αSMA, CDH2 genes. Moreover, 2-DG effect was tested on the permeability of monolayers of mesothelial and endothelial cells by monitoring the transmembrane resistance (RTM), FITC-dextran (10, 70 kDa) diffusion and mRNA expression levels of CLDN-1 to -5, ZO1, SGLT1, and SGLT2 genes. Contractility of MeT-5A cells in CPDF/2-DG was decreased, accompanied by αSMA (0.17 ± 0.03) and CDH2 (2.92 ± 0.29) gene expression fold changes. Changes in αSMA, CDH2 were found in EA.hy926 cells, though αSMA also decreased under LPDF/2-DG incubation (0.42 ± 0.02). Overall, 2-DG mitigated the PDF-induced alterations in mesothelial and endothelial barrier function as shown by RTM, dextran transport and expression levels of the CLDN-1 to -5, ZO1, and SGLT2. Thus, supplementation of PDF with 2-DG not only reduces MMT but also improves functional permeability characteristics of the PM mesothelial and endothelial barrier.

Differential effects of biocompatible peritoneal dialysis fluids on human mesothelial and endothelial cells in 2D and 3D phenotypes.

INTRODUCTION: Peritoneal dialysis (PD) is a life maintaining treatment in patients with end-stage renal disease. Its chronic application leads to peritoneal mesothelial layer denudation and fibrotic transformation along with vascular activation of inflammatory pathways. The impact of different PD fluids (PDF) on mesothelial and endothelial cell function and repair mechanisms are not comprehensively described. MATERIALS AND METHODS: Mesothelial (MeT-5A) and endothelial cells (EA.hy926) were cultured in 1:1 ratio with cell medium and different PDF (icodextrin-based, amino acid-based, and glucose-based). Cell adhesion, cell migration, and cell proliferation in 2D and spheroid formation and collagen gel contraction assays in 3D cell cultures were performed. RESULTS: Cell proliferation and cell-mediated gel contraction were both significantly decreased in all conditions. 3D spheroid formation was significantly reduced with icodextrin and amino acid PDF, but unchanged with glucose PDF. Adhesion was significantly increased by amino acid PDF in mesothelial cells and decreased by icodextrin and amino acid PDF in endothelial cells. Migration capacity was significantly decreased in mesothelial cells by all three PDF, while endothelial cells remained unaffected. CONCLUSIONS: In 3D phenotypes the effects of PDF are more uniform in both mesothelial and endothelial cells, mitigating spheroid formation and gel contraction. On the contrary, effects on 2D phenotypes are more uniform in the icodextrin and amino acid PDF as opposed to glucose ones and affect mesothelial cells more variably. 2D and 3D comparative assessments of PDF effects on the main peritoneal membrane cell barriers, the mesothelial and endothelial, could provide useful translational information for PD studies.

Asbestos ban policies and mesothelioma mortality in Greece.

BACKGROUND: Malignant mesothelioma is a rare form of cancer that mostly affects the pleura and has a strong link to asbestos exposure. Greece banned the use of asbestos in 2005, however, the public was already aware of this substance in the 1980s. This research aims to present an overview of Greece's mesothelioma age-standardized mortality rates (ASMR) from 1983 to 2019 by age, gender, and geographic region and to determine whether the actions to ban asbestos impacted these rates. METHODS: Data were retrieved by the Hellenic Statistical Authority (HSA) from death certificates that mentioned mesothelioma as the cause of death from 1983 to 2019 with details on the residence, gender, and age. Statistical analysis was performed using PRISM 6.0 software, a two-way ANOVA test, Trend analysis was conducted using Joinpoint Regression Program 5.0 software. The linear and non-linear model was used to calculate the age-standardized rates of annual percentage change (APC) and its 95% confidential interval (95% CI). RESULTS: From 1983 to 2019, 850 total mesothelioma deaths were recorded, the majority of whom were males (634). A rate of 74.6% accounts for males and 25.4% for females, and the ratio of Males: Females was 3:1. Males' ASMR and the whole population's ASMR reached their highest levels in 2011 (0.93/100000person-years and 0.53/100000person-years, respectively). To look for potential changes between the first two decades of the 21st century, we compared the mean ASMR of each geographic region in Greece between two different 10-year subperiods (2000-2009 and 2010-2019). Except for Epirus, all regions of Greece had elevated regional ASMRs, particularly in those with the highest asbestos deposits. Notably, the ASMR in Epirus decreased from 0.54/100000person-years (2000-2009) to 0.31/100000person-years (2010-2019). After 2011, the ASMR for men and the general population stabilized. This stability is important since mesothelioma in men is associated with occupational asbestos exposure. The intriguing discovery of a lower ASMR in Epirus emphasizes the need to raise awareness of the condition and implement effective public health measures. CONCLUSIONS: In Greece, the annual ASMR for males and the whole population reached its highest level in 2011, which is positive and encouraging and may be a sign that the rate will stabilize during the following years. Moreover, this study showed that the actions made in the 1980s regarding public awareness and surveillance directly impacted the decrease in Epirus rates. Future research, continual awareness, information, and recording are needed to monitor the mesothelioma epidemic. The possible benefit of a mesothelioma registry and the epidemiological surveillance of asbestos-related diseases, particularly mesothelioma mortality, need to be addressed. TRIAL REGISTRATION: Not applicable.

Calcimimetic AMG-416 induced short-term changes in calcium concentrations and calcium isotope ratios in rats.

Calcium (Ca) isotopes (δ44/42Ca) in serum and urine have been suggested as novel sensitive markers of bone calcification. The response of δ44/42Ca to acute changes in Ca homeostasis, has not yet been demonstrated. We measured serum Ca and δ44/42Ca in rats maintained on a standard and a 50% Ca reduced diet for 4 weeks, and after injection of 1 mg/kg of the calcimimetic AMG-416, 24 h prior to sacrifice. AMG-416 decreased serum Ca by a maximum of 0.38 ± 0.10 and 0.53 ± 0.35 mmol/l after 12 and 6 h, respectively, in the standard and low-Ca diet groups (p = 0.0006/0.02), while serum δ44/42Ca did not change over 24 h in both groups. Urinary Ca concentrations were higher 24 h after AMG-416 injection in both groups (p = 0.03/0.06), urine δ44/42Ca was not different compared to the untreated control groups. Our data does not show acute changes in δ44/42Ca in response to a single dose of AMG-416 within 24 h after injection, possibly due to a lack of bone calcification.

Effects of pharmacological primary cilium disturbance in the context of in vitro 2D and 3D malignant pleura mesothelioma.

INTRODUCTION: Primary cilium (PC) is a single non-motile antenna-like organelle composed of a microtubule core axon originating from the mother centriole of the centrosome. The PC is universal in all mammalian cells and protrudes to the extracellular environment receiving mechanochemical cues that it transmits in the cell. AIM: To investigate the role of PC in mesothelial malignancy in the context of two-dimensional (2D) and three-dimensional (3D) phenotypes. MATERIALS AND METHODS: The effect of pharmacological deciliation [using ammonium sulphate (AS) or chloral hydrate (CH)] and PC elongation [using lithium chloride (LC)] on cell viability, adhesion, and migration (2D cultures) as well as in mesothelial sphere formation, spheroid invasion and collagen gel contraction (3D cultures) was investigated in benign mesothelial MeT-5A cells and in malignant pleural mesothelioma (MPM) cell lines, M14K (epithelioid) and MSTO (biphasic), and primary malignant pleural mesothelioma cells (pMPM). RESULTS: Pharmacological deciliation or elongation of the PC significantly affected cell viability, adhesion, migration, spheroid formation, spheroid invasion and collagen gel contraction in MeT-5A, M14K, MSTO cell lines and in pMPM cells compared to controls (no drug treatment). CONCLUSIONS: Our findings indicate a pivotal role of the PC in functional phenotypes of benign mesothelial cells and MPM cells.

Glucose Derivative Induced Vasculopathy in Children on Chronic Peritoneal Dialysis.

[Figure: see text].

Methods to Measure Water Permeability.

Water permeability is a key feature of the cell plasma membranes, and it has seminal importance for several cell functions such as cell volume regulation, cell proliferation, cell migration, and angiogenesis to name a few. The transport of water occurs mainly through plasma membrane water channels, aquaporins. Aquaporins have very important function in physiological and pathophysiological states. Due to the above, the experimental assessment of the water permeability of cells and tissues is necessary. The development of new methodologies of measuring water permeability is a vibrant scientific field that constantly develops during the last three decades along with the advances in imaging mainly. In this chapter we describe and critically assess several methods that have been developed for the measurement of water permeability both in living cells and in tissues with a focus in the first category.

Prediction and enrichment analyses of the Homo sapiens-Drosophila melanogaster COPD-related orthologs: potential for modeling of human COPD genomic responses with the fruit fly.

The significant similarities in airway epithelial cells between mammals and the fruit fly Drosophila melanogaster have rendered the latter an important model organism for studies of chronic inflammatory lung diseases. Focusing on the chronic obstructive pulmonary disease (COPD), we here mapped human gene orthologs associated with this disease in D. melanogaster to identify functionally equivalent genes for immediate, further screening with the fruit fly model. The DIOPT-DIST tool was accessed for the prediction of the COPD-associated orthologs between humans and Drosophila. Enrichment analyses with respect to pathways of the retrieved functional homologs were performed using the ToppFun and FlyMine tools, identifying 73 unique human genes as well as 438 fruit fly genes. The ToppFun analysis verified that the human gene list is associated with COPD phenotypes. Furthermore, the FlyMine investigation highlighted that the Drosophila genes are functionally connected mainly with the "ABC-family proteins mediated transport" and the "ß-catenin-independent WNT signaling pathway." These results suggest an evolutionarily conserved role toward responses to inhaled toxicants and CO2 in both species. We reason that the predicted orthologous genes should be further studied in the Drosophila models of cigarette smoke-induced COPD.

Post-COVID-19 Parkinsonism and Parkinson's Disease Pathogenesis: The Exosomal Cargo Hypothesis.

Parkinson's disease (PD) is the second most prevalent neurodegenerative disease after Alzheimer's disease, globally. Dopaminergic neuron degeneration in substantia nigra pars compacta and aggregation of misfolded alpha-synuclein are the PD hallmarks, accompanied by motor and non-motor symptoms. Several viruses have been linked to the appearance of a post-infection parkinsonian phenotype. Coronavirus disease 2019 (COVID-19), caused by emerging severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection, has evolved from a novel pneumonia to a multifaceted syndrome with multiple clinical manifestations, among which neurological sequalae appear insidious and potentially long-lasting. Exosomes are extracellular nanovesicles bearing a complex cargo of active biomolecules and playing crucial roles in intercellular communication under pathophysiological conditions. Exosomes constitute a reliable route for misfolded protein transmission, contributing to PD pathogenesis and diagnosis. Herein, we summarize recent evidence suggesting that SARS-CoV-2 infection shares numerous clinical manifestations and inflammatory and molecular pathways with PD. We carry on hypothesizing that these similarities may be reflected in exosomal cargo modulated by the virus in correlation with disease severity. Travelling from the periphery to the brain, SARS-CoV-2-related exosomal cargo contains SARS-CoV-2 RNA, viral proteins, inflammatory mediators, and modified host proteins that could operate as promoters of neurodegenerative and neuroinflammatory cascades, potentially leading to a future parkinsonism and PD development.

Nutritional Calcium Supply Dependent Calcium Balance, Bone Calcification and Calcium Isotope Ratios in Rats.

Serum calcium isotopes (δ44/42Ca) have been suggested as a non-invasive and sensitive Ca balance marker. Quantitative δ44/42Ca changes associated with Ca flux across body compartment barriers relative to the dietary Ca and the correlation of δ44/42CaSerum with bone histology are unknown. We analyzed Ca and δ44/42Ca by mass-spectrometry in rats after two weeks of standard-Ca-diet (0.5%) and after four subsequent weeks of standard- and of low-Ca-diet (0.25%). In animals on a low-Ca-diet net Ca gain was 61 ± 3% and femur Ca content 68 ± 41% of standard-Ca-diet, bone mineralized area per section area was 68 ± 15% compared to standard-Ca-diet. δ44/42Ca was similar in the diets, and decreased in feces and urine and increased in serum in animals on low-Ca-diet. δ44/42CaBone was higher in animals on low-Ca-diet, lower in the diaphysis than the metaphysis and epiphysis, and unaffected by gender. Independent of diet, δ44/42CaBone was similar in the femora and ribs. At the time of sacrifice, δ44/42CaSerum inversely correlated with intestinal Ca uptake and histological bone mineralization markers, but not with Ca content and bone mineral density by µCT. In conclusion, δ44/42CaBone was bone site specific, but mechanical stress and gender independent. Low-Ca-diet induced marked changes in feces, serum and urine δ44/42Ca in growing rats. δ44/42CaSerum inversely correlated with markers of bone mineralization.

Integrated Deadenylase Genetic Association Network and Transcriptome Analysis in Thoracic Carcinomas.

The poly(A) tail at the 3' end of mRNAs determines their stability, translational efficiency, and fate. The shortening of the poly(A) tail, and its efficient removal, triggers the degradation of mRNAs, thus, regulating gene expression. The process is catalyzed by a family of enzymes, known as deadenylases. As the dysregulation of gene expression is a hallmark of cancer, understanding the role of deadenylases has gained additional interest. Herein, the genetic association network shows that CNOT6 and CNOT7 are the most prevalent and most interconnected nodes in the equilibrated diagram. Subsequent silencing and transcriptomic analysis identifies transcripts possibly regulated by specific deadenylases. Furthermore, several gene ontologies are enriched by common deregulated genes. Given the potential concerted action and overlapping functions of deadenylases, we examined the effect of silencing a deadenylase on the remaining ones. Our results suggest that specific deadenylases target unique subsets of mRNAs, whilst at the same time, multiple deadenylases may affect the same mRNAs with overlapping functions.

In silico investigation of the viroporin E as a vaccine target against SARS-CoV-2.

Viroporins, integral viral membrane ion channel proteins, interact with host-cell proteins deregulating physiological processes and activating inflammasomes. Severity of COVID-19 might be associated with hyperinflammation, thus we aimed at the complete immunoinformatic analysis of the SARS-CoV-2 viroporin E, P0DTC4. We also identified the human proteins interacting with P0DTC4 and the enriched molecular functions of the corresponding genes. The complete sequence of P0DTC4 in FASTA format was processed in 10 databases relative to secondary and tertiary protein structure analyses and prediction of optimal vaccine epitopes. Three more databases were accessed for the retrieval and the molecular functional characterization of the P0DTC4 human interactors. The immunoinformatics analysis resulted in the identification of 4 discontinuous B-cell epitopes along with 1 linear B-cell epitope and 11 T-cell epitopes which were found to be antigenic, immunogenic, nonallergen, nontoxin, and unable to induce autoimmunity thus fulfilling prerequisites for vaccine design. The functional enrichment analysis showed that the predicted host interactors of P0DTC4 target the cellular acetylation network. Two of the identified host-cell proteins - BRD2 and BRD4 - have been shown to be promising targets for antiviral therapy. Thus, our findings have implications for COVID-19 therapy and indicate that viroporin E could serve as a promising vaccine target against SARS-CoV-2. Validation experiments are required to complement these in silico results.

Calcium isotope fractionation by osteoblasts and osteoclasts, across endothelial and epithelial cell barriers, and with binding to proteins.

Timely and accurate diagnosis of osteoporosis is essential for adequate therapy. Calcium isotope ratio (δ44/42Ca) determination has been suggested as a sensitive, noninvasive, and radiation-free biomarker for the diagnosis of osteoporosis, reflecting bone calcium balance. The quantitative diagnostic is based on the calculation of the δ44/42Ca difference between blood, urine, and bone. The underlying cellular processes, however, have not been studied systematically. We quantified calcium transport and δ44/42Ca fractionation during in vitro bone formation and resorption by osteoblasts and osteoclasts and across renal proximal tubular epithelial cells (HK-2), human vein umbilical endothelial cells (HUVECs), and enterocytes (Caco-2) in transwell systems and determined transepithelial electrical resistance characteristics. δ44/42Ca fractionation was furthermore quantified with calcium binding to albumin and collagen. Calcified matrix formed by osteoblasts was isotopically lighter than culture medium by -0.27 ± 0.03‰ within 5 days, while a consistent effect of activated osteoclasts on δ44/42Ca could not be demonstrated. A transient increase in δ44/42Ca in the apical compartment by 0.26‰ occured across HK-2 cells, while δ44/42Ca fractionation was small across the HUVEC barrier and absent with Caco-2 enterocytes, and with binding of calcium to albumin and collagen. In conclusion, δ44/42Ca fractionation follows similar universal principles as during inorganic mineral precipitation; osteoblast activity results in δ44/42Ca fractionation. δ44/42Ca fractionation also occurs across the proximal tubular cell barrier and needs to be considered for in vivo bone mineralization modeling. In contrast, the effect of calcium transport across endothelial and enterocyte barriers on blood δ44/42Ca should be low and is absent with physiochemical binding of calcium to proteins.

HΜGB1/sRAGE levels differ significantly between transudates and exudates.

High mobility group box 1(HMGB1) protein operates as an alarmin with multiple roles in immunity and cell homeostasis. It is highly expressed in epithelial barrier sites and acts via the binding to the receptor for advanced glycation end products (RAGE). Production of HMGB1 and soluble RAGE (sRAGE), a decoy receptor for HMGB1, has been implicated in several pulmonary diseases, but both have been scarcely investigated in pleural diseases. The aim of this study was to determine the levels of HMGB1 and sRAGE in transudative, malignant and parapneumonic pleural effusions (PEs) and to investigate the effect of low and high HMGB1 pleural fluid levels on MeT-5A cell adhesion, migration and spheroid formation, in each group. HMGB1 and sRAGE levels were significantly lower and higher in transudative PEs compared to malignant and parapneumonic PEs, respectively. Patients above 65 years of age had significantly lower HMGB1 and higher sRAGE levels compared to patients below 65 years old. Furthermore, incubation of MeT-5A cells with malignant or parapneumonic PEs bearing low or high levels of HMGB1 yielded significant differential effects on MeT-5A cell adhesion, migration and spheroid formation. In all types of effusions, high HMGB1 levels correlated with more adherence compared to low HMGB1 levels. In transudative and malignant PEs high HMGB1 levels correlated with decreased migration of MeT-5A cells while in parapneumonic ones the effect was the opposite. Only samples from parapneumonic PEs high in HMGB1 achieved uniform spheroid formation. These results reveal a clinical context-dependent effect of the HMGB1/sRAGE axis in PEs.

Cell and extracellular matrix interaction models in benign mesothelial and malignant pleural mesothelioma cells in 2D and 3D in-vitro.

Malignant pleural mesothelioma (MPM) is an aggressive tumour that grows in the pleural cavity. MPM spheroids released in the pleural fluid can form new tumour foci. Cell-cell, cell-extracellular matrix (ECM) interactions in 2D and 3D impact malignant cell behaviour during cell adhesion, migration, proliferation and epithelial-mesenchymal transition (EMT). In this study, epithelioid, biphasic and sarcomatoid MPM cell types as well as benign mesothelial cells were tested with regards to the above phenotypes. Fibronectin (FN) and homologous cell-derived extracellular matrix (hcd-ECM) treated substratum differentially affected the above phenotypes. 3D MPM spheroid invasion was higher in FN-collagen matrices in the epithelioid and biphasic cells, while 3D cell cultures of epithelioid and sarcomatoid MPM cells in FN-collagen showed a higher contractility compared to hcd-ECM-collagen. Cell aggregates demonstrated invasive behaviour in hcd-ECM matrices alone. Our results suggest that ECM and the dimensionality affect malignant cell behaviour during cell culture studies.

An Experimental Workflow for Studying Barrier Integrity, Permeability, and Tight Junction Composition and Localization in a Single Endothelial Cell Monolayer: Proof of Concept.

Endothelial and epithelial barrier function is crucial for the maintenance of physiological processes. The barrier paracellular permeability depends on the composition and spatial distribution of the cell-to-cell tight junctions (TJ). Here, we provide an experimental workflow that yields several layers of physiological data in the setting of a single endothelial cell monolayer. Human umbilical vein endothelial cells were grown on Transwell filters. Transendothelial electrical resistance (TER) and 10 kDa FITC dextran flux were measured using Alanyl-Glutamine (AlaGln) as a paracellular barrier modulator. Single monolayers were immunolabelled for Zonula Occludens-1 (ZO-1) and Claudin-5 (CLDN5) and used for automated immunofluorescence imaging. Finally, the same monolayers were used for single molecule localization microscopy (SMLM) of ZO-1 and CLDN5 at the nanoscale for spatial clustering analysis. The TER increased and the paracellular dextran flux decreased after the application of AlaGln and these functional changes of the monolayer were mediated by an increase in the ZO-1 and CLDN5 abundance in the cell-cell interface. At the nanoscale level, the functional and protein abundance data were accompanied by non-random increased clustering of CLDN5. Our experimental workflow provides multiple data from a single monolayer and has wide applicability in the setting of paracellular studies in endothelia and epithelia.

Interactome networks between the human respiratory syncytial virus (HRSV), the human metapneumovirus (ΗMPV), and their host: In silico investigation and comparative functional enrichment analysis.

BACKGROUND AND OBJECTIVES: Human respiratory syncytial virus (HRSV) and human metapneumovirus (HMPV) are leading causes of upper and lower respiratory tract infections in non-immunocompetent subjects, yet the mechanisms by which they induce their pathogenicity differ significantly and remain elusive. In this study we aimed at identifying the gene interaction networks between the HRSV, HMPV respiratory pathogens and their host along with the different cell-signaling pathways associated with the above interactomes. MATERIALS AND METHODS: The Viruses STRING database (http://viruses.string-db.org/) was used for the identification of the host-viruses interaction networks. The two lists of the predicted functional partners were entered in the FunRich tool (http://www.funrich.org) for the construction of the Venn diagram and the comparative Funcional Enrichment Analysis (FEA) with respect to biological pathways. The sets of the common and unique human genes identified in the two networks were also analyzed. The computational predictions regarding the shared human genes in the host-HRSV and the host-HMPV interactomes were further evaluated via the analysis of the GSE111732 dataset. miRNA transcriptomics data were mapped to gene targets using the miRNomics pipeline of the GeneTrail2 database (https://genetrail2.bioinf.uni-sb.de/). RESULTS: Eleven out of twenty predicted human genes were common in the two interactomes (TLR4, SOCS3, SFXN1, AKT1, SFXN3, LY96, SFXN2, SOCS7, CISH, SOCS6, SOCS1). FEA of these common genes identified the kit receptor and the GH receptor signaling pathways as the most significantly enriched annotations. The remaining nine genes of the host-HRSV and the host-HMPV interaction networks were the IFIH1, DDX58, NCL, IRF3, STAT2, HSPA4, CD209, KLF6, CHKA and the MYD88, SOCS4, SOCS2, SOCS5 AKT2, AKT3, SFXN4, SFXN5 and TLR3 respectively. Distinct cell-signaling pathways were enriched per interactome. The comparative FEA highlighted the association of the host-HRSV functional partners with the negative regulation of RIG-I/MDA5 signaling. The analysis with respect to miRNAs mapping to gene targets of the GSE111732 dataset indicated that nine out of the eleven common host genes are either enriched or depleted in the sample sets (HRSV or HMPV infected) as compared with the reference set (non-infected), although with no significant scores. CONCLUSIONS: We have identified both shared and unique host genes as members of the HRSV and HMPV interaction networks. The disparate human genes likely contribute to distinct responses in airway epithelial cells.

Benign Pleural Mesothelial Cells Have Higher Osmotic Water Permeability than Malignant Pleural Mesothelioma Cells and Differentially Respond to Hyperosmolality.

BACKGROUND/AIMS: Cell volume regulation is a critical mechanism for cell homeostasis and depends on the osmotic water permeability (Pf) of the cell plasma membrane. The Pf of human mesothelial cells is unknown although they contribute to serosal fluid turnover. METHODS: In this study we measured the osmotic water permeability of benign human mesothelial cells (MeT-5A) and of epithelioid (M14K) and sarcomatoid (ZL34) malignant pleural mesothelioma (MPM) cells in response to acute hyperosmotic stress. We also assessed the changes in their Pf after preconditioning with 4% glucose for 24 hours. In both cases we also assessed the role of AQP1 inhibition (0.1 mM HgCl2) on the Pf. Finally, we assessed corresponding changes in the AQP1 plasma membrane availability by immunofluorescence. RESULTS: We report that MeT-5A cells have a significantly higher Pf as compared to M14K and ZL34 MPM cells [4.85E-03±2.37E-03 cm/sec (n=17) versus 2.74E-03±0.74E-03 cm/sec (n=11) and 2.86E-03±0.11E-03 cm/sec (n=11)]. AQP1 inhibition significantly decreased the Pf in all cells lines (p<0.001 in all cases). High glucose preconditioning for 24 hours significantly increased MeT-5A Pf (p<0.001), did not influence M14K Pf (p=0.19) and significantly reduced ZL34 Pf (p=0.02). Comparing cell lines after high glucose preconditioning, MeT-5A Pf was significantly higher than that of M14K and ZL34 MPM cells and the AQP1 inhibition effect was significant in MeT-5A and M14K cells. These results were corroborated by AQP1 immunofluorescence. CONCLUSION: We provide evidence for a differential regulation of Pf in benign and MPM cells that require further mechanistic investigation.