Confounding patient factors affecting the proper interpretation of the periostin level as a biomarker in asthma development

Introduction: The proper use of serum periostin (POSTN) as a biomarker for asthma is hindered by inconsistent performance in different clinical settings. Objective: To explore patient’s factors that may affect POSTN expression locally and systematically and its utility as a biomarker for asthma development. Materials and Methods: Here we used bioinformatics analysis of publicly available transcriptomics data to confirm that POSTN is an asthma specific gene involved in core signaling pathways enriched in the bronchial epithelium during asthma. We then explored a large number of datasets to identify possible confounders that may affect the POSTN gene expression and consequently, its interpretation as a reliable biomarker for asthma. Plasma and saliva levels of POSTN were determined in locally recruited asthmatic patients (mild, moderate and severe) compared to healthy controls to confirm the bioinformatics findings. Results: Our bioinformatics results confirmed that POSTN was consistently upregulated in the bronchial epithelium in asthma, chronic obstructive pulmonary disease (COPD) and idiopathic pulmonary fibrosis (IPF) bronchial epithelium. In asthma, its mRNA expression was affected by gender, sample anatomical site and type, steroid therapy, and smoking. In our cohort, plasma POSTN was upregulated in severe and non-severe asthmatic patients. Saliva POSTN was significantly higher in non-severe asthmatic patients compared to healthy and severe asthmatic patients (specifically those who are not on Xolair (omalizumab)). Patients’ BMI, inhaled steroid use and Xolair treatment affected POSTN plasma levels. Conclusion: Up to our knowledge, this is the first study examining the level of POSTN in the saliva of asthmatic patients. Both plasma and saliva POSTN levels can aid in early diagnosis of asthma. Saliva POSTN level was more sensitive than plasma POSTN in differentiating between severe and non-severe asthmatics. Patients’ characteristics like BMI, the use of inhaled steroids, or Xolair treatment should be carefully reviewed before any meaningful interpretation of POSTN level in clinical practice

Derangement of cell cycle markers in peripheral blood mononuclear cells of asthmatic patients as a reliable biomarker for asthma control

In asthma, most of the identified biomarkers pertain to the Th2 phenotype and no known biomarkers have been verified for severe asthmatics. Therefore, identifying biomarkers using the integrative phenotype-genotype approach in severe asthma is needed. The study aims to identify novel biomarkers as genes or pathways representing the core drivers in asthma development, progression to the severe form, resistance to therapy, and tissue remodeling regardless of the sample cells or tissues examined. Comprehensive reanalysis of publicly available transcriptomic data that later was validated in vitro, and locally recruited patients were used to decipher the molecular basis of asthma. Our in-silicoanalysis revealed a total of 10 genes (GPRC5A, SFN, ABCA1, KRT8, TOP2A, SERPINE1, ANLN, MKI67, NEK2, and RRM2) related to cell cycle and proliferation to be deranged in the severe asthmatic bronchial epithelium and fibroblasts compared to their healthy counterparts. In vitro, RT qPCR results showed that (SERPINE1 and RRM2) were upregulated in severe asthmatic bronchial epithelium and fibroblasts, (SFN, ABCA1, TOP2A, SERPINE1, MKI67, and NEK2) were upregulated in asthmatic bronchial epithelium while (GPRC5A and KRT8) were upregulated only in asthmatic bronchial fibroblasts. Furthermore, MKI76, RRM2, and TOP2A were upregulated in Th2 high epithelium while GPRC5A, SFN, ABCA1 were upregulated in the blood of asthmatic patients. SFN, ABCA1 were higher, while MKI67 was lower in severe asthmatic with wheeze compared to nonasthmatics with wheezes. SERPINE1 and GPRC5A were downregulated in the blood of eosinophilic asthmatics, while RRM2 was upregulated in an acute attack of asthma. Validation of the gene expression in PBMC of locally recruited asthma patients showed that SERPINE1, GPRC5A, SFN, ABCA1, MKI67, and RRM2 were downregulated in severe uncontrolled asthma. We have identified a set of biologically crucial genes to the homeostasis of the lung and in asthma development and progression. This study can help us further understand the complex interplay between the transcriptomic data and the external factors which may deviate our understanding of asthma heterogeneity

Blood and Salivary Amphiregulin Levels as Biomarkers for Asthma

BACKGROUND: mphiregulin (AREG) expression in asthmatic airways and sputum was shown to increase and correlate with asthma. However, no studies were carried out to evaluate the AREG level in blood and saliva of asthmatic patients. OBJECTIVE: To measure circulating AREG mRNA and protein concentrations in blood, saliva, and bronchial biopsies samples from asthmatic patients. METHODS: Plasma and Saliva AREG protein concentrations were measured using ELISA while PBMCs, and Saliva mRNA expression was measured by RT qPCR in non-severe, and severe asthmatic patients compared to healthy controls. Primary asthmatic bronchial epithelial cells and fibroblasts were assessed for AREG mRNA expression and released soluble AREG in their conditioned media. Tissue expression of AREG was evaluated using immunohistochemistry of bronchial biopsies from asthmatic patients and healthy controls. Publicly available transcriptomic databases were explored for the global transcriptomic profile of bronchial epithelium, and PBMCs were explored for AREG expression in asthmatic vs. healthy controls. RESULTS: Asthmatic patients had higher AREG protein levels in blood and saliva compared to control subjects. Higher mRNA expression in saliva and primary bronchial epithelial cells plus higher AREG immunoreactivity in bronchial biopsies were also observed. Both blood and saliva AREG levels showed positive correlations with allergic rhinitis status, atopy status, eczema status, plasma periostin, neutrophilia, Montelukast sodium use, ACT score, FEV1, and FEV1/FVC. In silico analysis showed that severe asthmatic bronchial epithelium with high AREG gene expression is associated with higher neutrophils infiltration. CONCLUSION: AREG levels measured in a minimally invasive blood sample and a non-invasive saliva sample are higher in non-allergic severe asthma

IL-17 Induced Autophagy Regulates Mitochondrial Dysfunction and Fibrosis in Severe Asthmatic Bronchial Fibroblasts

The accumulation of fibroblasts, their synthesis of extracellular matrix (ECM) proteins and their innate resistance to apoptosis are characteristics of subepithelial fibrosis observed in severe asthma. Interleukin-17 (IL-17) is an important regulator of airway remodeling in asthma. However, the contribution of IL-17 to the pro-fibrotic phenotype of bronchial fibroblasts is not well-characterized. In this study, we investigated whether IL-17 induced autophagy regulates mitochondrial and pro-fibrotic function in bronchial fibroblasts. The primary cultured bronchial fibroblasts isolated from non-asthmatic (NHBF) and severe asthmatic (DHBF) subjects were treated with IL-17 in order to ascertain its effect on mitochondrial function, mitochondrial quality control, and apoptosis using immunoblotting and flow cytometric analyses. At baseline, DHBF exhibited higher levels of mitophagy and mitochondrial biogenesis compared to NHBF. Immunohistochemical evaluation of bronchial biopsies showed intense PINK1 immunoreactivity in severe asthma than in control. IL-17 intensified the mitochondrial dysfunction and impaired the mitochondrial quality control machinery in NHBF and DHBF. Moreover, IL-17 augmented a pro-fibrotic and anti-apoptotic response in both group of fibroblasts. Inhibition of autophagy using bafilomycin-A1 reduced PINK1 expression in NHBF and restored the IL-17 mediated changes in PINK1 to their basal levels in DHBF. Bafilomycin-A1 also reversed the IL-17 associated fibrotic response in these fibroblasts, suggesting a role for IL-17 induced autophagy in the induction of fibrosis in bronchial fibroblasts. Taken together, our findings suggest that IL-17 induced autophagy promotes mitochondrial dysfunction and fibrosis in bronchial fibroblasts from both non-asthmatic and severe asthmatic subjects. Our study provides insights into the therapeutic potential of targeting autophagy in ameliorating fibrosis, particularly in severe asthmatic individuals

Metagenomic sequencing and reverse transcriptase PCR reveal that mobile phones and environmental surfaces are reservoirs of Multidrug-Resistant superbugs and SARS-CoV-2

Background: Mobile phones of healthcare workers (HCWs) can act as fomites in the dissemination of microbes. This study was carried out to investigate microbial contamination of mobile phones of HCWs and environmental samples from the hospital unit using a combination of phenotypic and molecular methods. Methods: This point prevalence survey was carried out at the Emergency unit of a tertiary care facility. The emergency unit has two zones, a general zone for non-COVID-19 patients and a dedicated COVID-19 zone for confirmed or suspected COVID-19 patients. Swabs were obtained from the mobile phones of HCWs in both zones for bacterial culture and shotgun metagenomic analysis. Metagenomic sequencing of pooled environmental swabs was conducted. RT-PCR for SARS-CoV-2 detection was carried out. Results: Bacteria contamination on culture was detected from 33 (94.2%) mobile phones with a preponderance of Staphylococcus epidermidis (n/N = 18/35), Staphylococcus hominis (n/N = 13/35), and Staphylococcus haemolyticus (n/N = 7/35). Two methicillin-sensitive and three methicillin-resistant Staphylococcus aureus, and one pan-drug-resistant carbapenemase producer Acinetobacter baumannii were detected. Shotgun metagenomic analysis showed high signature of Pseudomonas aeruginosa in mobile phone and environmental samples with preponderance of P. aeruginosa bacteriophages. Malassezia and Aspergillus spp. were the predominant fungi detected. Fourteen mobile phones and one environmental sample harbored protists. P. aeruginosa antimicrobial resistance genes mostly encoding for efflux pump systems were detected. The P. aeruginosa virulent factor genes detected were related to motility, adherence, aggregation, and biofilms. One mobile phone from the COVID-19 zone (n/N = 1/5; 20%) had positive SARS-CoV-2 detection while all other phone and environmental samples were negative. Conclusion: The findings demonstrate that mobile phones of HCWs are fomites for potentially pathogenic and highly drug-resistant microbes. The presence of these microbes on the mobile phones and hospital environmental surfaces is a concern as it poses a risk of pathogen transfer to patients and dissemination into the community

Logistic regression prediction model identify type 2 diabetes mellitus as a prognostic factor for human papillomavirus-16 associated head and neck squamous cell carcinoma

Background: HPV-16–positive HNSCC and HPV-16–negative HNSCC have different clinical factors, representing distinct forms of cancers. The study aimed to identify patient-specific factors for HPV-16-positive HNSCC based on baseline clinical data. Method: Factors associated with HPV-16-positive HNSCC were identified using the data from 210 patients diagnosed with HNSCC at University College of London Hospital between January 1, 2003, and April 30, 2015, inclusive. A series of models were developed using logistic regression methods, and the overall model fit was compared using Akaike Information Criterion. Survival analysis was carried with Cox proportional hazards model for survival-time outcomes. The survival time for individual patients was defined as the time from diagnosis of HNSCC to the date of death from any cause. For patients who did not die, they were censored at the end of study on April 30, 2015. Results: Of the 210 patients, 151 (72%) were found to have HPV-16-positive HNSCC. The logistic regression model showed that the prevalence of developing HPV-16-positive HNSCC was 3.79 times higher in patients with Type 2 Diabetes Mellitus (T2DM) (odd ratio [OR], 3.79; 95% CI, 1.70–8.44) than in those without T2DM, and 8.84 times higher in patients with history of primary HNSCC (OR, 8.84; 95% CI, 2.30–33.88) than in those without a history of primary HNSCC. HPV-16–positive HNSCC was also observed more in tonsils (OR, 4.02; 95% CL, 1.56–10.36) and less in non-alcohol drinker’s oral cavity (OR, 0.14; 95% CI, 0.03–0.56). Furthermore, individual patients were followed-up for 1 to 13 years (median of 1 year). Patients with HPV-positive HNSCC had a median survival of 5 years (95% CI, 2.6–7.3 years). Among HPV-16–positive HNSCC cohort, T2DM was a risk for poorer prognosis (hazard ratio, 2.57; 95% Cl, 1.09–6.07), and had lower median survival of 3 years (95% CI, 1.8–4.1 years), as compared to 6 years (95% CI, 2.8–9.1 years) in non-T2DM. Conclusions: Patient-specific factors for HPV-positive HNSCC are T2DM, history of primary HNSCC and tonsillar site. T2DM is associated with poorer prognosis. These findings suggest that it might be beneficial if routine HPV-16 screening is carried out in T2DM patients which can provide better therapeutic and management strategies

Characterization of SARS-CoV-2 nucleocapsid protein reveals multiple functional consequences of the C-terminal domain

Nucleocapsid (N) encoded by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) plays key roles in the replication cycle and is a critical serological marker. Here, we characterize essential biochemical properties of N and describe the utility of these insights in serological studies. We define N domains important for oligomerization and RNA binding and show that N oligomerization provides a high-affinity RNA-binding platform. We also map the RNA-binding interface, showing protection in the N-terminal domain and linker region. In addition, phosphorylation causes reduction of RNA binding and redistribution of N from liquid droplets to loose coils, showing how N-RNA accessibility and assembly may be regulated by phosphorylation. Finally, we find that the C-terminal domain of N is the most immunogenic, based on antibody binding to patient samples. Together, we provide a biochemical description of SARS-CoV-2 N and highlight the value of using N domains as highly specific and sensitive diagnostic markers

The magnetic susceptibility of graphite biintercalated with CoCl2 and GaCl3

We present the first results of magnetic susceptibility on CoCl2-GaCl 3 biintercalated in graphite, and compare them with those obtained in second stage CoCl2-graphite. A fit by a power law above the maximum of the susceptibility present two regimes : in the high temperature one, the γ values are high and even higher in the CoCl2-GaCl 3 compound. This indicates that the three-dimensional coupling is lower in the biintercalation compound. In the low temperature regime, near the transition, the susceptibility is driven by the islandic nature of the intercalate layer.Nous présentons les premiers résultats obtenus par susceptibilité magnétique sur le composé de bi-insertion du graphite avec CoCl2-GaCl3 et nous les comparons à ceux du composé d'insertion graphite-CoCl2 de deuxième stade. L'ajustement de la partie haute température du pic de susceptibilité présente deux régimes. Dans le régime haute température, les valeurs de γ obtenues sont élevées, et plus élevées encore dans le biinséré. Ceci indique que le couplage tridimensionnel est encore plus faible dans ce dernier. Dans le régime basse température, près de la transition, la susceptibilité est gouvernée par la nature insulaire des couches insérées

Experimental study of flow around two grooved cylinders arranged in tandem

In the present paper, the influence of longitudinal grooves placed on tubes is investigated for different configurations in tandem for three Reynolds number. Each cylinder has two grooves on the external surface at 90° and 270° degrees. The experiments are carried out using a subsonic wind tunnel. The pressure distributions along the tubes are determined for a variation of the azimuthal angle from 0 to 360 degrees. The drag and lift forces are measured using the TE 44 balance. The results show that the upstream cylinder pressure is greater than for second one. There is a difference in the pressure coefficient as well as the pressure drag for configuration smooth-smooth and grooved-grooved