Sensitization to molecular dog allergens in an adult population: Results from the West Sweden Asthma Study.

BACKGROUND: As the prevalence of dog allergy rises, component resolved diagnosis might improve the diagnosis, understanding of the clinical outcomes and the effectiveness of immunotherapy. Considering the paucity of data in adults, the current study characterized the patterns of sensitization to dog molecular allergens in an adult population. METHODS: Data were derived from the West Sweden Asthma Study, a population-based and representative sample of adults from western Sweden. Of the 2006 subjects clinically examined, 313 participants sensitized to whole dog allergen extract were measured for specific immunoglobulin E (sIgE) levels to Can f 1, Can f 2, Can f 3, Can f 4, Can f 5 and Can f 6 using ImmunoCAP™. Polysensitization was defined as sensitization to ≥3 components. Overlapping sensitization was defined as having concomitant sensitization to at least two dog molecular allergen families (lipocalin, albumin or prostatic kallikrein). RESULTS: Of 313, 218 (70%) subjects tested positive to at least one dog allergen component. Sensitization to Can f 1 (43%) was the most common, followed by Can f 5 (33%) among molecular allergens, while sensitization to lipocalins (56%) was the most common among component families. Polysensitization was found in 22% of all participants and was more common in participants with than in those without asthma. Subjects with asthma were less likely to be monosensitized to Can f 5 than those without asthma. Subjects with asthma had higher IgE levels of Can f 3, Can f 4 and Can f 6 than those without asthma. Overlapping sensitizations also differed between those with asthma and allergic rhinitis and those without. CONCLUSION: Increased knowledge about the sensitization patterns of dog allergen components can aid in defining their role in asthma and rhinitis. In complex clinical cases of dog allergy, a detailed analysis of dog allergen components can provide additional information on the nature of sensitization.

Comparison of clinician diagnosis of COVID-19 with real time polymerase chain reaction in an adult-representative population in Sweden.

BACKGROUND: Due to the high transmissibility of SARS-CoV-2, accurate diagnosis is essential for effective infection control, but the gold standard, real-time reverse transcriptase-polymerase chain reaction (RT-PCR), is costly, slow, and test capacity has at times been insufficient. We compared the accuracy of clinician diagnosis of COVID-19 against RT-PCR in a general adult population. METHODS: COVID-19 diagnosis data by 30th September 2021 for participants in an ongoing population-based cohort study of adults in Western Sweden were retrieved from registers, based on positive RT-PCR and clinician diagnosis using recommended ICD-10 codes. We calculated accuracy measures of clinician diagnosis using RT-PCR as reference for all subjects and stratified by age, gender, BMI, and comorbidity collected pre-COVID-19. RESULTS: Of 42,621 subjects, 3,936 (9.2%) and 5705 (13.4%) had had COVID-19 identified by RT-PCR and clinician diagnosis, respectively. Sensitivity and specificity of clinician diagnosis against RT-PCR were 78% (95%CI 77-80%) and 93% (95%CI 93-93%), respectively. Positive predictive value (PPV) was 54% (95%CI 53-55%), while negative predictive value (NPV) was 98% (95%CI 98-98%) and Youden's index 71% (95%CI 70-72%). These estimates were similar between men and women, across age groups, BMI categories, and between patients with and without asthma. However, while specificity, NPV, and Youden's index were similar between patients with and without chronic obstructive pulmonary disease (COPD), sensitivity was slightly higher in patients with (84% [95%CI 74-90%]) than those without (78% [95%CI 77-79%]) COPD. CONCLUSIONS: The accuracy of clinician diagnosis for COVID-19 is adequate, regardless of gender, age, BMI, and asthma, and thus can be used for screening purposes to supplement RT-PCR.

Detoxified synthetic bacterial membrane vesicles as a vaccine platform against bacteria and SARS-CoV-2.

The development of vaccines based on outer membrane vesicles (OMV) that naturally bud off from bacteria is an evolving field in infectious diseases. However, the inherent inflammatory nature of OMV limits their use as human vaccines. This study employed an engineered vesicle technology to develop synthetic bacterial vesicles (SyBV) that activate the immune system without the severe immunotoxicity of OMV. SyBV were generated from bacterial membranes through treatment with detergent and ionic stress. SyBV induced less inflammatory responses in macrophages and in mice compared to natural OMV. Immunization with SyBV or OMV induced comparable antigen-specific adaptive immunity. Specifically, immunization with Pseudomonas aeruginosa-derived SyBV protected mice against bacterial challenge, and this was accompanied by significant reduction in lung cell infiltration and inflammatory cytokines. Further, immunization with Escherichia coli-derived SyBV protected mice against E. coli sepsis, comparable to OMV-immunized group. The protective activity of SyBV was driven by the stimulation of B-cell and T-cell immunity. Also, SyBV were engineered to display the SARS-CoV-2 S1 protein on their surface, and these vesicles induced specific S1 protein antibody and T-cell responses. Collectively, these results demonstrate that SyBV may be a safe and efficient vaccine platform for the prevention of bacterial and viral infections.

The Anti-Inflammatory Effects of Adipose Tissue Mesenchymal Stem Cell Exosomes in a Mouse Model of Inflammatory Bowel Disease.

Inflammatory bowel disease (IBD) is a group of chronic, relapsing inflammatory disorders that affect the gastrointestinal tract, with the primary subtypes being ulcerative colitis (UC) and Crohn's disease (CD). We aimed to evaluate the therapeutic potential of extracellular vesicles released by adipose-tissue-derived mesenchymal stem cells, which we, in this manuscript, call "exosomes" (ASC-EXOs), in a mouse model of IBD. We specifically aimed to determine the effectiveness of different treatment protocols and compare the effects with that of anti-IL-12 p40 monoclonal antibody. The addition of dextran sulfate sodium (DSS) to drinking water induced multiple signs of IBD, including weight loss, soft stool, and bloody feces. ASC-EXOs given by either intraperitoneal (IP) or intravenous (IV) routes resulted in moderate improvement in these signs of IBD. IV ASC-EXOs resulted in significantly reduced body weight loss, improved histopathological scoring, and suppressed the disease activity index (DAI) compared to the IBD control group. Also, a reduction in PCR for pro-inflammatory cytokines was observed. IV ASC treatment resulted in dose-related reduction in IBD signs, including weight loss. An increasing number of injections with ASC-EXOs reduced histopathological scores as well as DAI. Co-administration of ASC-EXOs with anti-IL-12 p40 significantly decreased DAI scores in the ASC-EXO + anti-IL-12 p40 group. In conclusion, ASC-EXOs have potential as a therapeutic agent for IBD, but the route of administration, number of injections, and dosage need to be considered to optimize the effects of ASC-EXO treatment. This study also highlights the potential benefits of combination therapies of ASC-EXOs and anti-IL-12. Our findings pave the way for further studies to unravel the underlying therapeutic mechanisms of ASC-EXOs in IBD treatment.

Targeting Myd88 using peptide-loaded mesenchymal stem cell membrane-derived synthetic vesicles to treat systemic inflammation.

Mesenchymal stem cells (MSC) secrete extracellular vesicles (EV) with a regenerative profile, and an increasing number of studies have focused on the utilization of MSC-EV for therapeutic drug delivery. However, EV are usually produced by cells in low quantities and are packed with numerous cytoplasmic components, which may be unfavorable for further drug loading. In this study, we developed a simple process for generating membrane vesicles directly from the cells, which we refer to as synthetic eukaryotic vesicles (SyEV). We hypothesized that MSC-derived SyEV can be efficiently loaded with an anti-inflammatory drug and the loaded vesicles can strongly suppress the systemic inflammation induced by bacterial outer membrane vesicles (OMV). SyEV were generated from MSC membranes through serial extrusion of the cells, ionic stress, and subsequent vesiculation of the membrane sheets, leading to high yield and purity of the SyEV with few cytosolic components remaining. When these SyEV were given to macrophages or mice exposed to OMV, the release of pro-inflammatory cytokines was similarly attenuated comparable to treatment with natural EV. We then loaded the SyEV with large numbers of peptides targeting Myd88 and observed enhanced therapeutic potential of the loaded vesicles in OMV-induced macrophages. Further, in vivo experiments showed that the peptide-encapsulated MSC-SyEV suppressed cytokine production synergistically. Taken together, these findings suggest that SyEV-based therapeutics is a highly interesting platform for delivering an advanced therapeutic drug for the treatment of systemic inflammation without severe side effects.

Isolation and characterization of microvesicles from mesenchymal stem cells.

Mesenchymal stem or stromal cells are currently under clinical investigation for multiple diseases. While their mechanism of action is still not fully elucidated, vesicles secreted by MSCs are believed to recapitulate their therapeutic potentials to some extent. Microvesicles (MVs), also called as microparticles or ectosome, are among secreted vesicles that could transfer cytoplasmic cargo, including RNA and proteins, from emitting (source) cells to recipient cells. Given the importance of MVs, we here attempted to establish a method to isolate and characterize MVs secreted from unmodified human bone marrow derived MSCs (referred to as native MSCs, and their microvesicles as Native-MVs) and IFNγ stimulated MSCs (referred to as IFNγ-MSCs, and their microvesicles as IFNγ-MVs). We first describe an ultracentrifugation technique to isolate MVs from the conditioned cell culture media of MSCs. Next, we describe characterization and quality control steps to analyze the protein and RNA content of MVs. Finally, we examined the potential of MVs to exert immunomodulatory effects through induction of regulatory T cells (Tregs). Secretory vesicles from MSCs are promising alternatives for cell therapy with applications in drug delivery, regenerative medicine, and immunotherapy.

Immune-Associated Proteins Are Enriched in Lung Tissue-Derived Extracellular Vesicles during Allergen-Induced Eosinophilic Airway Inflammation.

Studying the proteomes of tissue-derived extracellular vesicles (EVs) can lead to the identification of biomarkers of disease and can provide a better understanding of cell-to-cell communication in both healthy and diseased tissue. The aim of this study was to apply our previously established tissue-derived EV isolation protocol to mouse lungs in order to determine the changes in the proteomes of lung tissue-derived EVs during allergen-induced eosinophilic airway inflammation. A mouse model for allergic airway inflammation was used by sensitizing the mice intraperitoneal with ovalbumin (OVA), and one week after the final sensitization, the mice were challenged intranasal with OVA or PBS. The animals were sacrificed 24 h after the final challenge, and their lungs were removed and sliced into smaller pieces that were incubated in culture media with DNase I and Collagenase D for 30 min at 37 °C. Vesicles were isolated from the medium by ultracentrifugation and bottom-loaded iodixanol density cushions, and the proteomes were determined using quantitative mass spectrometry. More EVs were present in the lungs of the OVA-challenged mice compared to the PBS-challenged control mice. In total, 4510 proteins were quantified in all samples. Among them, over 1000 proteins were significantly altered (fold change >2), with 614 proteins being increased and 425 proteins being decreased in the EVs from OVA-challenged mice compared to EVs from PBS-challenged animals. The associated cellular components and biological processes were analyzed for the altered EV proteins, and the proteins enriched during allergen-induced airway inflammation were mainly associated with gene ontology (GO) terms related to immune responses. In conclusion, EVs can be isolated from mouse lung tissue, and the EVs' proteomes undergo changes in response to allergen-induced airway inflammation. This suggests that the composition of lung-derived EVs is altered in diseases associated with inflammation of the lung, which may have implications in type-2 driven eosinophilic asthma pathogenesis.

Stem Cell Extracellular Vesicles: Extended Messages of Regeneration.

Stem cells are critical to maintaining steady-state organ homeostasis and regenerating injured tissues. Recent intriguing reports implicate extracellular vesicles (EVs) as carriers for the distribution of morphogens and growth and differentiation factors from tissue parenchymal cells to stem cells, and conversely, stem cell-derived EVs carrying certain proteins and nucleic acids can support healing of injured tissues. We describe approaches to make use of engineered EVs as technology platforms in therapeutics and diagnostics in the context of stem cells. For some regenerative therapies, natural and engineered EVs from stem cells may be superior to single-molecule drugs, biologics, whole cells, and synthetic liposome or nanoparticle formulations because of the ease of bioengineering with multiple factors while retaining superior biocompatibility and biostability and posing fewer risks for abnormal differentiation or neoplastic transformation. Finally, we provide an overview of current challenges and future directions of EVs as potential therapeutic alternatives to cells for clinical applications.

Extracellular vesicles from mast cells induce mesenchymal transition in airway epithelial cells.

BACKGROUND: In the airways, mast cells are present in close vicinity to epithelial cells, and they can interact with each other via multiple factors, including extracellular vesicles (EVs). Mast cell-derived EVs have a large repertoire of cargos, including proteins and RNA, as well as surface DNA. In this study, we hypothesized that these EVs can induce epithelial to mesenchymal transition (EMT) in airway epithelial cells. METHODS: In this in-vitro study we systematically determined the effects of mast cell-derived EVs on epithelial A549 cells. We determined the changes that are induced by EVs on A549 cells at both the RNA and protein levels. Moreover, we also analyzed the rapid changes in phosphorylation events in EV-recipient A549 cells using a phosphorylated protein microarray. Some of the phosphorylation-associated events associated with EMT were validated using immunoblotting. RESULTS: Morphological and transcript analysis of epithelial A549 cells indicated that an EMT-like phenotype was induced by the EVs. Transcript analysis indicated the upregulation of genes involved in EMT, including TWIST1, MMP9, TGFB1, and BMP-7. This was accompanied by downregulation of proteins such as E-cadherin and upregulation of Slug-Snail and matrix metalloproteinases. Additionally, our phosphorylated-protein microarray analysis revealed proteins associated with the EMT cascade that were upregulated after EV treatment. We also found that transforming growth factor beta-1, a well-known EMT inducer, is associated with EVs and mediates the EMT cascade induced in the A549 cells. CONCLUSION: Mast cell-derived EVs mediate the induction of EMT in epithelial cells, and our evidence suggests that this is triggered through the induction of protein phosphorylation cascades.

Decreased COPD prevalence in Sweden after decades of decrease in smoking.

BACKGROUND: COPD has increased in prevalence worldwide over several decades until the first decade after the millennium shift. Evidence from a few recent population studies indicate that the prevalence may be levelling or even decreasing in some areas in Europe. Since the 1970s, a substantial and ongoing decrease in smoking prevalence has been observed in several European countries including Sweden. The aim of the current study was to estimate the prevalence, characteristics and risk factors for COPD in the Swedish general population. A further aim was to estimate the prevalence trend of COPD in Northern Sweden from 1994 to 2009. METHODS: Two large random population samples were invited to spirometry with bronchodilator testing and structured interviews in 2009-2012, one in south-western and one in northern Sweden, n = 1839 participants in total. The results from northern Sweden were compared to a study performed 15 years earlier in the same area and age-span. The diagnosis of COPD required both chronic airway obstruction (CAO) and the presence of respiratory symptoms, in line with the GOLD documents since 2017. CAO was defined as post-bronchodilator FEV1/FVC < 0.70, with sensitivity analyses based on the FEV1/FVC < lower limit of normal (LLN) criterion. RESULTS: Based on the fixed ratio definition, the prevalence of COPD was 7.0% (men 8.3%; women 5.8%) in 2009-2012. The prevalence of moderate to severe (GOLD ≥ 2) COPD was 3.5%. The LLN based results were about 30% lower. Smoking, occupational exposures, and older age were risk factors for COPD, whereof smoking was the most dominating risk factor. In northern Sweden the prevalence of COPD, particularly moderate to severe COPD, decreased significantly from 1994 to 2009, and the decrease followed a decrease in smoking. CONCLUSIONS: The prevalence of COPD has decreased in Sweden, and the prevalence of moderate to severe COPD was particularly low. The decrease follows a major decrease in smoking prevalence over several decades, but smoking remained the dominating risk factor for COPD.

International Society for Extracellular Vesicles and International Society for Cell and Gene Therapy statement on extracellular vesicles from mesenchymal stromal cells and other cells: considerations for potential therapeutic agents to suppress coronavirus disease-19.

STATEMENT: The International Society for Cellular and Gene Therapies (ISCT) and the International Society for Extracellular Vesicles (ISEV) recognize the potential of extracellular vesicles (EVs, including exosomes) from mesenchymal stromal cells (MSCs) and possibly other cell sources as treatments for COVID-19. Research and trials in this area are encouraged. However, ISEV and ISCT do not currently endorse the use of EVs or exosomes for any purpose in COVID-19, including but not limited to reducing cytokine storm, exerting regenerative effects or delivering drugs, pending the generation of appropriate manufacturing and quality control provisions, pre-clinical safety and efficacy data, rational clinical trial design and proper regulatory oversight.

BRAFV600 inhibition alters the microRNA cargo in the vesicular secretome of malignant melanoma cells.

The BRAF inhibitors vemurafenib and dabrafenib can be used to treat patients with metastatic melanomas harboring BRAFV600 mutations. Initial antitumoral responses are often seen, but drug-resistant clones with reactivation of the MEK-ERK pathway soon appear. Recently, the secretome of tumor-derived extracellular vesicles (EVs) has been ascribed important functions in cancers. To elucidate the possible functions of EVs in BRAF-mutant melanoma, we determined the RNA content of the EVs, including apoptotic bodies, microvesicles, and exosomes, released from such cancer cells after vemurafenib treatment. We found that vemurafenib significantly increased the total RNA and protein content of the released EVs and caused significant changes in the RNA profiles. RNA sequencing and quantitative PCR show that cells and EVs from vemurafenib-treated cell cultures and tumor tissues harvested from cell-derived and patient-derived xenografts harbor unique miRNAs, especially increased expression of miR-211-5p. Mechanistically, the expression of miR-211-5p as a result of BRAF inhibition was induced by increased expression of MITF that regulates the TRPM1 gene resulting in activation of the survival pathway. In addition, transfection of miR-211 in melanoma cells reduced the sensitivity to vemurafenib treatment, whereas miR-211-5p inhibition in a vemurafenib resistant cell line affected the proliferation negatively. Taken together, our results show that vemurafenib treatment induces miR-211-5p up-regulation in melanoma cells both in vitro and in vivo, as well as in subsets of EVs, suggesting that EVs may provide a tool to understand malignant melanoma progression.

Lung Regulatory T Cells Express Adiponectin Receptor 1: Modulation by Obesity and Airway Allergic Inflammation.

Regulatory T cells (Tregs) decrease in the adipose tissue upon weight gain, contributing to persistent low-grade inflammation in obesity. We previously showed that adipose tissue Tregs express the adiponectin receptor 1 (AdipoR1); however, the expression in lung Tregs is still unknown. Here, we aimed to determine whether Helios+ and Helios- Treg subsets expressed AdipoR1 in the lungs of obese mice and whether different obesity grades affected the expression upon allergic lung inflammation. For diet-induced obesity (DIO), mice were fed a high-fat diet (HFD) for up to 15 weeks (overweight), 21 weeks (obesity), and 26 weeks (morbid obesity). Overweight and morbidly obese mice were sensitized and challenged with ovalbumin (OVA) to induce allergic lung inflammation. The AdipoR1 expression was reduced significantly in the lung Helios+ and Helios- Tregs of obese mice compared with lean mice. Airway allergic inflammation showed reduced AdipoR1 expression in lung Foxp3+ Tregs. Obesity significantly exacerbated the eosinophilic airway inflammation and reduced the number of Helios+ Tregs in lung and adipose tissue in the obesity-associated asthma model. Upon further weight gain, AdipoR1-expressing Tregs in the lungs of allergic mice were increased, whereas AdipoR1-expressing Tregs in adipose tissue were reduced. These data suggest that obesity-associated adipose tissue inflammation may exacerbate allergic inflammation by downregulating the AdipoR1+ Tregs in the lungs.

Characterization of sensitization to furry animal allergen components in an adult population.

BACKGROUND: There are paucity of data on sensitization to furry animal allergen components in adults. Furry animals are major sensitizers and contributors to asthma burden in northern Europe and North America. OBJECTIVE: To characterize sensitization patterns to furry animal allergen components in Swedish adults. METHODS: Based on the West Sweden Asthma Study, a random population (n = 1103) and an asthma sample (n = 769) were tested for allergen sensitization using Phadiatop® . Those with IgE ≥ 0.35 kUA /L were tested for cat (Fel d 1, 2, and 4), dog (Can f 1, 2, 3, and 5), and horse (Equ c 1) allergen component sensitization. We defined allergen component poly-sensitization patterns, identified data-driven sensitization clusters, described component sensitization overlaps, and assessed determinants of sensitization patterns. RESULTS: The prevalence of allergen component sensitization ranged from 0.8% for Fel d 2 and Can f 3 to 8.9% for Fel d 1. The most common dog component was Can f 5 (3.6%); 2.1% were sensitized to Equ c 1. Those sensitized to Fel d 2 and Fel d 4 were commonly sensitized to Fel d 1. The most common dog component overlap was between Can f 1/Can f 2 and Can f 5. Mono-sensitization was 5.6%, double sensitization 1.5% and poly-sensitization 2.1%. Sensitization was always higher in the asthma than in the random sample. Three sensitization clusters were derived, namely non-sensitized (90% in random vs 66% in asthma sample); Fel d 1-driven sensitized (7% vs 19%); and multi-sensitized (3% vs 15%). Key determinants of sensitization were gender, age, raised on a farm, family history of allergy or asthma, smoking, and occupational exposure to dust or fumes. CONCLUSIONS & CLINICAL RELEVANCE: Fel d 1 and Can f 5 are the most common cat and dog components sensitization in this adult Swedish population. Mono-sensitization is more common than poly-sensitization. This detailed characterization highlights the current distribution of furry animal allergen components in Swedish adults, and their impact on clinical outcomes of asthma will be further explored.

Detailed analysis of the plasma extracellular vesicle proteome after separation from lipoproteins.

The isolation of extracellular vesicles (EVs) from blood is of great importance to understand the biological role of circulating EVs and to develop EVs as biomarkers of disease. Due to the concurrent presence of lipoprotein particles, however, blood is one of the most difficult body fluids to isolate EVs from. The aim of this study was to develop a robust method to isolate and characterise EVs from blood with minimal contamination by plasma proteins and lipoprotein particles. Plasma and serum were collected from healthy subjects, and EVs were isolated by size-exclusion chromatography (SEC), with most particles being present in fractions 8-12, while the bulk of the plasma proteins was present in fractions 11-28. Vesicle markers peaked in fractions 7-11; however, the same fractions also contained lipoprotein particles. The purity of EVs was improved by combining a density cushion with SEC to further separate lipoprotein particles from the vesicles, which reduced the contamination of lipoprotein particles by 100-fold. Using this novel isolation procedure, a total of 1187 proteins were identified in plasma EVs by mass spectrometry, of which several proteins are known as EV-associated proteins but have hitherto not been identified in the previous proteomic studies of plasma EVs. This study shows that SEC alone is unable to completely separate plasma EVs from lipoprotein particles. However, combining SEC with a density cushion significantly improved the separation of EVs from lipoproteins and allowed for a detailed analysis of the proteome of plasma EVs, thus making blood a viable source for EV biomarker discovery.

High prevalence of severe asthma in a large random population study.

BACKGROUND: The prevalence of asthma severity is not well described at a population level. OBJECTIVE: We sought to determine the prevalence of phenotypic signs of asthma severity among asthmatic patients in a general population and to describe risk factors for asthma severity. METHODS: We performed an epidemiologic study conducted between 2008 and 2012 (West Sweden Asthma Study). A postal questionnaire was sent to a random population (n = 30,000) in west Sweden, with 18,087 responses. A total of 2,006 subjects were carefully phenotyped. Only subjects with "active asthma" (symptoms or medication in the last year, n = 744) were analyzed in this study to determine the degree of severity of the disease within an asthma cohort. Phenotypes of severity were calculated based on (1) multiple symptoms during the day despite ongoing use of asthma medications, (2) FEV1 of less than 70% of predicted value, (3) daily or almost daily use of rescue medications, (4) nighttime symptoms once a week or more, and (5) oral corticosteroid use/emergency department visits. Asthmatic patients were grouped as having nonsevere disease, 1 sign of severity, or 2 or more signs of severity. RESULTS: A total of 36.2% of asthmatic patients expressed at least 1 sign of asthma severity, and 13.2% had 2 or more signs. The group with 2 or more signs was older in age and had higher body mass index, a higher rate of tobacco smoking, and lower lung function. Bronchial hyperreactivity, airway inflammation, and sensitization were significantly different among the 3 groups. At a population level, the prevalence of asthma severity was 3.1% for 1 sign and 1.3% for at least 2 signs. CONCLUSION: More than 1 in 3 asthmatic patients show at least 1 sign of asthma severity. The phenotypes of asthma severity are highly diverse, which is important to consider when implementing personalized medicine in asthmatic patients.

Effective Refractive Index and Lipid Content of Extracellular Vesicles Revealed Using Optical Waveguide Scattering and Fluorescence Microscopy.

Extracellular vesicles (EVs) are generating a growing interest because of the key roles they play in various biological processes and because of their potential use as biomarkers in clinical diagnostics and as efficient carriers in drug-delivery and gene-therapy applications. Their full exploitation, however, depends critically on the possibility to classify them into different subpopulations, a task that in turn relies on efficient means to identify their unique biomolecular and physical signatures. Because of the large heterogeneity of EV samples, such information remains rather elusive, and there is accordingly a need for new and complementary characterization schemes that can help expand the library of distinct EV features. In this work, we used surface-sensitive waveguide scattering microscopy with single EV resolution to characterize two subsets of similarly sized EVs that were preseparated based on their difference in buoyant density. Unexpectedly, the scattering intensity distribution revealed that the scattering intensity of the high-density (HD) population was on an average a factor of three lower than that of the low-density (LD) population. By further labeling the EV samples with a self-inserting lipid-membrane dye, the scattering and fluorescence intensities from EVs could be simultaneously measured and correlated at the single-particle level. The labeled HD sample exhibited not only lower fluorescence and scattering intensities but also lower effective refractive index ( n ≈ 1.35) compared with the LD EVs ( n ≈ 1.38), indicating that both the lipid and protein contents were indeed lower in the HD EVs. Although separation in density gradients of similarly sized EVs is usually linked to differences in biomolecular content, we suggest based on these observations that the separation rather reflects the ability of the solute of the gradient to penetrate the lipid membrane enclosing the EVs, that is, the two gradient bands are more likely because of the differences in membrane permeability than to differences in biomolecular content of the EVs.

Prevalence, clinical characteristics and morbidity of the Asthma-COPD overlap in a general population sample.

OBJECTIVE: Although asthma and chronic obstructive pulmonary disease (COPD) have been regarded as distinct conditions, emerging literature suggests that overlapping phenotypes, called asthma-COPD overlap (ACO), exists. The aim of this study was to describe prevalence, patient characteristics and morbidity of ACO. METHODS: From a cross-sectional population sample, the West Sweden Asthma Study, subjects with suspected asthma, chronic bronchitis or COPD, and a random sample, were invited to clinical examinations. ACO was defined as doctor-diagnosed asthma, or clear clinical signs of asthma at examination, with a FEV1/FVC < 0.7. RESULTS: Subjects were categorized as ACO (N = 181), COPD only (N = 89), asthma only (N = 651) or healthy (n = 1036) based on clinical examinations. Prevalence of ACO was 3.4% in the random sample (N = 1172) and 18.1% among asthmatics (N = 138) in the random sample. Subjects with ACO (mean age 59 years, 54% women) had an age and gender distribution in between asthma only (45 years, 63% women) and COPD only (62 years, 41% women). Ever-smoking was reported by 71%, 48% and 74% in the ACO, asthma only and COPD only groups, respectively. Subjects with ACO had worse lung function (mean FEV1% of predicted normal 76%) than asthma only (100%) and COPD only (87%) and reported more respiratory symptoms. Also respiratory related emergency visits were more common in ACO compared to asthma only and COPD only, respectively. CONCLUSIONS: ACO is present in 3.4% of the population and common among subjects with both asthma and COPD. Subjects with ACO had worse lung function and more symptoms than subjects with asthma or COPD only.

Upper airway and skin symptoms in allergic and non-allergic asthma: Results from the Swedish GA2LEN study.

BACKGROUND: Allergic and non-allergic asthma are viewed as separate entities, despite sharing similarities. The aims of this study were to determine differences in symptoms from the upper airways and the skin in allergic and non-allergic asthma. The secondary aims were to identify childhood risk factors and to compare quality of life in the two asthma groups. METHODS: This cohort (age 17-76 years) consisted of 575 subjects with allergic or non-allergic asthma and 219 controls. The participants participated in an interview, spirometry, FeNO, skin prick test, and responded to the Mini Asthma Quality of Life Questionnaire. RESULTS: Self-reported allergic rhinitis was significantly more common in both allergic and non-allergic asthma (82.3 and 40.7%) groups compared with the controls. The prevalence of chronic rhinosinusitis (CRS) was similar in both asthma groups. Eczema was significantly more common in both asthmatic groups (72.3 and 59.8%) than controls (47.0%) (p < 0.001 and p = 0.012). Severe respiratory infection in childhood and parental allergy were risk factors for both allergic and non-allergic asthma groups. Quality of life was significantly lower in non-allergic than allergic asthma groups (p = 0.01). CONCLUSION: Concomitant symptoms from the upper airways and the skin were significantly more common in both allergic and non-allergic asthma. This indicates that non-allergic asthma has a systemic component with similarities to what is found in allergic asthma. There were similarities in the childhood risk factor pattern between the two types of asthma but asthma-related quality of life was lower in the non-allergic asthma group.

EVpedia: A community web resource for prokaryotic and eukaryotic extracellular vesicles research.

For cell-to-cell communication, all living cells including archaea, bacteria, and eukaryotes secrete nano-sized membrane vesicles into the extracellular space. These extracellular vesicles harbor specific subsets of proteins, mRNAs, miRNAs, lipids, and metabolites that represent their cellular status. These vesicle-specific cargos are considered as novel diagnostic biomarkers as well as therapeutic targets. With the advancement in high-throughput technologies on multiomics studies and improvements in bioinformatics approaches, a huge number of vesicular proteins, mRNAs, miRNAs, lipids, and metabolites have been identified, and our understanding of these complex extracellular organelles has considerably increased during these past years. In this review, we highlight EVpedia (http://evpedia.info), a community web portal for systematic analyses of prokaryotic and eukaryotic extracellular vesicles research.