A novel experimental workflow to determine the impact of storage parameters on the mass spectrometric profiling and assessment of representative phosphatidylethanolamine lipids in mouse tissues.

Evaluation of signaling lipids is essential for measuring biological processes. There is a lack of experimental data regarding the proper storage of extracts for signaling lipid analysis, potentially impacting the procedures that can lead to accurate and reproducible evaluation. In this study, the importance of pre-analytical conditions for analyzing ion transitions for phosphatidylethanolamines (PEs), an abundant signaling phospholipid, was systematically assessed. A novel workflow was utilized involving an MRM-based experimental approach followed by statistical analysis. Specifically, lipids were extracted from the brain, heart, lungs, and serum of C57BL/6 mice. Extract subsets were resuspended in organic solvents prior to storage in various temperature conditions. Mass spectrometry analysis by multiple reaction monitoring (MRM) profiling was performed at four time points (1 day, 2 weeks, 2 months, or 6 months) to measure relative amounts of PEs in distinct lipid extract aliquots. We introduce an innovative statistical workflow to measure the changes in relative amounts of PEs in the profiles over time to determine lipid extract storage conditions in which fewer profile changes occur. Results demonstrated that time is the most significant factor affecting the changes in lipid samples, with temperature and solvent having comparatively minor effects. We conclude that for lipid extracts obtained by Bligh & Dyer extraction, storage at - 80.0 °C without solvent for less than 2 weeks before analysis is ideal. By considering the data generated by this study, lipid extract storage practices may be optimized and standardized, enhancing the validity and reproducibility of lipid assessments.

Comparison of silver nanoparticle-induced inflammatory responses between healthy and metabolic syndrome mouse models.

Silver nanoparticles (AgNPs) are utilized in surgical implants and medical textiles, thus providing access to the circulation. While research has been conducted primarily in healthy models, AgNP-induced toxicity evaluations in disease conditions are critical, as many individuals have preexisting conditions. Specifically, over 20% of United States adults suffer from metabolic syndrome (MetS). It was hypothesized that MetS may increase susceptibility to AgNP-mediated toxicity due to induction of differential inflammation and altered biodistribution. Mice were injected with 2 mg/kg AgNPs, and organs assessed for inflammatory gene expression (TNF-α, CXCL1, CXCL2, CCL2, TGF-ß, HO-1, IL-4, IL-13), and Ag content. AgNPs were determined to induce differential inflammation in healthy and MetS mice. While AgNP exposure increased TNF-α, CXCL1, TGF-ß, HO-1, and IL-4 expression within healthy mouse spleens, MetS-treated animals demonstrated decreased CXCL1, IL-4, and IL-13 expression. Healthy and MetS mice livers exhibited similar inflammatory responses to one another. AgNPs localized primarily to the liver and spleen, although Ag was present in all examined organs. In organs of minor AgNP deposition, such as kidney, gene expression was variable. Induction of inflammatory genes did not correspond with biodistribution, suggesting disease-related variations in AgNP-mediated adverse responses. These findings indicate that disease may influence inflammation and biodistribution, impacting AgNP clinical applications.

In vitro toxicity assessment of emitted materials collected during the manufacture of water pipe plastic linings.

Objectives: US water infrastructure is in need of widespread repair due to age-related deterioration. Currently, the cured-in-place (CIPP) procedure is the most common method for water pipe repair. This method involves the on-site manufacture of a new polymer composite plastic liner within the damaged pipe. The CIPP process can release materials resulting in occupational and public health concerns. To understand hazards associated with CIPP-related emission exposures, an in vitro toxicity assessment was performed. Materials and Methods: Mouse alveolar epithelial and alveolar macrophage cell lines and condensates collected at 3 worksites utilizing styrene-based resins were utilized for evaluations. All condensate samples were normalized based on the major emission component, styrene. Further, a styrene-only exposure group was used as a control to determine mixture related toxicity. Results: Cytotoxicity differences were observed between worksite samples, with the CIPP worksite 4 sample inducing the most cell death. A proteomic evaluation was performed, which demonstrated styrene-, worksite-, and cell-specific alterations. This examination of protein expression changes determined potential biomarkers of exposure including transglutaminase 2, advillin, collagen type 1, perilipin-2, and others. Pathway analysis of exposure-induced proteomic alterations identified MYC and p53 to be regulators of cellular responses. Protein changes were also related to pathways involved in cell damage, immune response, and cancer. Conclusions: Together these findings demonstrate potential risks associated with the CIPP procedure as well as variations between worksites regarding emissions and toxicity. Our evaluation identified biological pathways that require a future evaluation and also demonstrates that exposure assessment of CIPP worksites should examine multiple chemical components beyond styrene, as many cellular responses were styrene-independent.

Characterization of Cleaning and Disinfection Product Use, Glove Use, and Skin Disorders by Healthcare Occupations in a Midwestern Healthcare Facility.

Healthcare facility staff use a wide variety of cleaning and disinfecting products during their daily operations, many of which are associated with respiratory or skin irritation or sensitization with repeated exposure. The objective of this study was to characterize the prevalence of cleaning and disinfection product use, glove use during cleaning and disinfection, and skin/allergy symptoms by occupation and identify the factors influencing glove use among the healthcare facility staff. A questionnaire was administered to the current employees at a midwestern Veterans Affairs healthcare facility that elicited information on cleaning and disinfection product use, glove use during cleaning and disinfection, skin/allergy symptoms, and other demographic characteristics, which were summarized by occupation. The central supply/environmental service workers (2% of the total survey population), nurses (26%,), nurse assistants (3%), and laboratory technicians (5%) had the highest prevalence of using cleaning or disinfecting products, specifically quaternary ammonium compounds, bleach, and alcohol. Glove use while using products was common in both patient care and non-patient care occupations. The factors associated with glove use included using bleach or quaternary ammonium compounds and using cleaning products 2-3 or 4-5 days per week. A high frequency of glove use (≥75%) was reported by workers in most occupations when using quaternary ammonium compounds or bleach. The use of alcohol, bleach, and quaternary ammonium compounds was associated with skin disorders (p < 0.05). These research findings indicate that although the workers from most occupations report a high frequency of glove use when using cleaning and disinfection products, there is room for improvement, especially among administrative, maintenance, and nursing workers. These groups may represent populations which could benefit from the implementation of workplace interventions and further training regarding the use of personal protective equipment and the potential health hazards of exposure to cleaning and disinfecting chemicals.

Particulate matter inhalation and the exacerbation of cardiopulmonary toxicity due to metabolic disease.

Particulate matter is a significant public health issue in the United States and globally. Inhalation of particulate matter is associated with a number of systemic and organ-specific adverse health outcomes, with the pulmonary and cardiovascular systems being particularly vulnerable. Certain subpopulations are well-recognized as being more susceptible to inhalation exposures, such as the elderly and those with pre-existing respiratory disease. Metabolic syndrome is becoming increasingly prevalent in our society and has known adverse effects on the heart, lungs, and vascular systems. The limited evaluations of individuals with metabolic syndromehave demonstrated that theymay compose a sensitive subpopulation to particulate exposures. However, the toxicological mechanisms responsible for this increased vulnerability are not fully understood. This review evaluates the currently available literature regarding how the response of an individual's pulmonary and cardiovascular systems is influenced by metabolic syndrome and metabolic syndrome-associated conditions such as hypertension, dyslipidemia, and diabetes. Further, we will discuss potential therapeutic agents and targets for the alleviation and treatment of particulate-matter induced metabolic illness. The information reviewed here may contribute to the understanding of metabolic illness as a risk factor for particulate matter exposure and further the development of therapeutic approaches to treat vulnerable subpopulations, such as those with metabolic diseases.

Biocorona-induced modifications in engineered nanomaterial-cellular interactions impacting biomedical applications.

When nanoparticles (NPs) enter a physiological environment, a complex coating of biomolecules is absorbed onto their surface, known as the biocorona (BC). This coating alters nanomaterial physical properties, modulating cellular viability, internalization, and immune responses. To safely utilize NPs within medical settings, it is necessary to understand the influence of the BC on cellular responses. Due to the variety of cell types, NPs, and physiological environments, responses are variable; though trends do exist. This review article critically evaluates the currently available literature regarding the influence of the BC on NP interactions with prominent cell types that they are likely to encounter during biomedical applications. Specifically, we will examine responses related to interactions with endothelial cells, macrophages, and epithelial cells of the digestive tract and lung. Further, we will evaluate how the BC may influence interactions with bacteria and fungi, as NPs have been proposed as antimicrobial agents in medical settings. The information reviewed and discussed here may enhance the development of effective of NP-based therapeutics and diagnostic tools. This article is categorized under: Therapeutic Approaches and Drug Discovery > Emerging Technologies Toxicology and Regulatory Issues in Nanomedicine > Toxicology of Nanomaterials Diagnostic Tools > Diagnostic Nanodevices.

Exacerbation of Nanoparticle-Induced Acute Pulmonary Inflammation in a Mouse Model of Metabolic Syndrome.

Nanotechnology has the capacity to revolutionize numerous fields and processes, however, exposure-induced health effects are of concern. The majority of nanoparticle (NP) safety evaluations have been performed utilizing healthy models and have demonstrated the potential for pulmonary toxicity. A growing proportion of individuals suffer diseases that may enhance their susceptibility to exposures. Specifically, metabolic syndrome (MetS) is increasingly prevalent and is a risk factor for the development of chronic diseases including type-2 diabetes, cardiovascular disease, and cancer. MetS is a combination of conditions which includes dyslipidemia, obesity, hypertension, and insulin resistance. Due to the role of lipids in inflammatory signaling, we hypothesize that MetS-associated dyslipidemia may modulate NP-induced immune responses. To examine this hypothesis, mice were fed either a control diet or a high-fat western diet (HFWD) for 14-weeks. A subset of mice were treated with atorvastatin for the final 7-weeks to modulate lipids. Mice were exposed to silver NPs (AgNPs) via oropharyngeal aspiration and acute toxicity endpoints were evaluated 24-h post-exposure. Mice on the HFWD demonstrated MetS-associated alterations such as increased body weight and cholesterol compared to control-diet mice. Cytometry analysis of bronchoalveolar lavage fluid (BALF) demonstrated exacerbation of AgNP-induced neutrophilic influx in MetS mice compared to healthy. Additionally, enhanced proinflammatory mRNA expression and protein levels of monocyte chemoattractant protein-1, macrophage inflammatory protein-2, and interleukin-6 were observed in MetS mice compared to healthy following exposure. AgNP exposure reduced mRNA expression of enzymes involved in lipid metabolism, such as arachidonate 5-lipoxygenase and arachidonate 15-lipoxygenase in both mouse models. Exposure to AgNPs decreased inducible nitric oxide synthase gene expression in MetS mice. An exploratory lipidomic profiling approach was utilized to screen lipid mediators involved in pulmonary inflammation. This assessment indicates the potential for reduced levels of lipids mediators of inflammatory resolution (LMIR) in the MetS model compared to healthy mice following AgNP exposure. Statin treatment inhibited enhanced inflammatory responses as well as alterations in LMIR observed in the MetS model due to AgNP exposure. Taken together our data suggests that MetS exacerbates the acute toxicity induced by AgNPs exposure possibly via a disruption of LMIR leading to enhanced pulmonary inflammation.

An Integrative Proteomic/Lipidomic Analysis of the Gold Nanoparticle Biocorona in Healthy and Obese Conditions.

Introduction: When nanoparticles (NPs) enter a physiological environment, a coating of biomolecules or biocorona (BC) forms on the surface. Formation of the NP-BC is dependent on NP properties, the physiological environment, and time. The BC influences NP properties and biological interactions such as cellular internalization, immune responses, biodistribution, and others, leading to pharmacological and toxicological consequences. To date, examination of the NP-BC has focused primarily on protein components and healthy conditions. Therefore, we evaluated the protein and lipid content of BCs that formed on physicochemically distinct gold nanoparticles (AuNPs) under healthy and obese conditions. A comprehensive understanding of the NP-BC is necessary for the translation of in vitro toxicity assessments to clinical applications. Materials and Methods: AuNPs with two coatings (poly-N-vinylpyrrolidone [PVP] or citrate) and diameters (20 or 100 nm) were incubated in pooled human serum, and an integrated proteomic/lipidomic approach was used to evaluate BC composition. Macrophages were utilized to evaluate differential immune responses due to variations in the AuNP-BC. Results: AuNPs form distinct BCs based on physicochemical properties and the surrounding environment, with the obese BC containing more proteins and fewer lipids than the healthy BC. Differential macrophage inflammatory responses were observed based on AuNP properties and BC composition. Discussion and Conclusion: Overall, these findings demonstrate that AuNP size and coating, as well as physiological environment, influence the protein and lipid composition of the BC, which impacts cellular responses following exposure. These findings demonstrate that incorporation of BCs representing distinct physiological conditions may enhance the translatability of nanosafety in vitro studies.

Experimental challenges regarding the in vitro investigation of the nanoparticle-biocorona in disease states.

Toxicological evaluation of nanoparticles (NPs) requires the utilization of in vitro techniques due to their number and diverse properties. Cell culture systems are often lacking in their ability to perform comparative toxicity assessment due to dosimetry issues and capacity to simulate in vivo environments. Upon encountering a physiological environment, NPs become coated with biomolecules forming a biocorona (BC), influencing function, biodistribution, and toxicity. Disease-induced alterations in the biological milieu can alter BC formation. This study evaluates the role of low-density lipoprotein (LDL) in altering macrophage responses to iron oxide (Fe3O4) NPs. BCs were formed by incubating Fe3O4 NPs in serum-free media, or 10% fetal bovine serum with or without LDL present. Following exposures to a normalized dose (25 µg/mL), macrophage association of Fe3O4 NPs with a LDL-BC was enhanced. TNF-α mRNA expression and protein levels were differentially induced due to BCs. Cell surface expression of SR-B1 was reduced following all Fe3O4 NPs exposures, while only NPs with an LDL-BC enhanced mitochondrial membrane potential. These findings suggest that elevations in LDL may contribute to distinct BC formation thereby influencing NP-cellular interactions and response. Further, our study highlights challenges that may arise during the in vitro evaluation of disease-related variations in the NP-BC.

Altered formation of the iron oxide nanoparticle-biocorona due to individual variability and exercise.

Nanoparticles (NPs), introduced into a biological environment, accumulate a coating of biomolecules or biocorona (BC). Although the BC has toxicological and pharmacological consequences, the effects of inter-individual variability and exercise on NP-BC formation are unknown. We hypothesized that NPs incubated in plasma form distinct BCs between individuals, and exercise causes additional intra-individual alterations. 20 nm iron oxide (Fe3O4) NPs were incubated in pre- or post-exercise plasma ex vivo, and proteomics was utilized to evaluate BC components. Analysis demonstrated distinct BC formation between individuals, while exercise was found to enhance NP-BC complexity. Abundance differences of NP-BC proteins were determined between individuals and resulting from exercise. Differential human macrophage response was identified due to NP-BC variability. These findings demonstrate that individuals form unique BCs and that exercise influences NP-biomolecule interactions. An understanding of NP-biomolecule interactions is necessary for elucidation of mechanisms responsible for variations in human responses to NP exposures and/or nano-based therapies.