Search

The majority of lung diseases occur with a sex bias in terms of prevalence and/or severity. Previous studies demonstrated that, compared with males, female mice develop greater eosinophilic inflammation in the airways after multiwalled carbon nanotube (MWCNT) exposure. However, the mechanism by which this sex bias occurs is unknown. Two immune cells that could account for the sex bias are type II innate lymphoid cells (ILC2s) and alveolar macrophages (AMs). In order to determine which immune cell type was responsible for MWCNT-induced airway eosinophil recruitment and subsequent sex differences in inflammation and disease, male and female C57BL/6 mice were exposed to MWCNTs (2 mg/kg) via oropharyngeal aspiration, and the respiratory immune response was assessed 7 d later. Greater eosinophilia and eotaxin 2 levels were observed in MWCNT-treated females and corresponded with greater changes in airway hyperresponsiveness than those in MWCNT-treated males. In MWCNT-treated females, there was a significant increase in the frequency of ILC2s within the lungs compared with control animals. However, depletion of ILC2s via α-CD90.2 administration did not decrease eosinophil recruitment 24 h and 7 d after MWCNT exposure. AMs isolated from control and MWCNT-treated animals demonstrated that M2a macrophage phenotype gene expression, ex vivo cytokine production, and activation of (p)STAT6 were upregulated to a significantly greater degree in MWCNT-treated females than in males. Our findings suggest that sex differences in AM phenotype development, not ILC2 signaling, are responsible for the observed female bias in eosinophilic inflammation after MWCNT inhalation.

Allergic asthma is a chronic inflammatory disorder of the airway associated with bronchial obstruction, airway hyper-reactivity (AHR), and mucus production. The epithelium may direct and propagate asthmatic-like responses. Central to this theory is the observation that viruses, air pollution, and allergens promote epithelial damage and trigger the generation of IL-25, IL-33, and TSLP via innate pathways such as TLRs and purinergic receptors. Similarly, engineered nanomaterials promote a Th2-associated pathophysiology. In this study, we tested the hypothesis that instillation of multi-walled carbon nanotubes (MWCNT) impair pulmonary function in C57Bl/6 mice due to the development of IL-33-dependent Th2-associated inflammation. MWCNT exposure resulted in elevated levels of IL-33 in the lavage fluid (likely originating from airway epithelial cells), enhanced AHR, eosinophil recruitment, and production of Th2-associated cytokines and chemokines. Moreover, these events were dependent on IL-13 signaling and the IL-33/ST2 axis, but independent of T and B cells. Finally, MWCNT exposure resulted in the recruitment of innate lymphoid cells. Collectively, our data suggest that MWCNT induce epithelial damage that results in release of IL-33, which in turn promotes innate lymphoid cell recruitment and the development of IL-13-dependent inflammatory response.

Acute and chronic inflammation are vital contributing factors to pulmonary diseases which can be triggered by exposure to occupational and man-made particles; however, there are no established treatments. One potential treatment shown to have anti-inflammatory capabilities is the dietary supplement docosahexaenoic acid (DHA), an omega-3 polyunsaturated fatty acid found in fish oil. DHA’s anti-inflammatory mechanisms are unclear for particle-induced inflammation; therefore, this study evaluated DHA as a prophylactic treatment for semi-acute and chronic particle-induced inflammation in vivo. Balb/c mice were fed a control or 1% DHA diet and exposed to dispersion media, an inflammatory multi-walled carbon nanotube (MWCNT), or crystalline silica (SiO(2)) either once (semi-acute) or once a week for 4 weeks (chronic). The hypothesis was that DHA will decrease pulmonary inflammatory markers in response to particle-induced inflammation. Results indicated that DHA had a trending anti-inflammatory effect in mice exposed to MWCNT. There was a general decrease in inflammatory signals within the lung lavage fluid and upregulation of M2c macrophage gene expression in the spleen tissue. In contrast, mice exposed to SiO(2) while on the DHA diet significantly increased most inflammatory markers. However, DHA stabilized the phagolysosomal membrane upon prolonged treatment. This indicated that DHA treatment may depend upon certain inflammatory particle exposures as well as the length of the exposure.

OBJECTIVE AND DESIGN: The omega-3 polyunsaturated fatty acid docosahexaenoic acid (DHA) has been reported to suppress inflammation. Pulmonary inflammation can be directly linked to exposure of various occupational and man-made particles leading to pulmonary diseases. Therapeutic treatments are lacking for particle-induced pulmonary inflammation. These studies evaluated DHA as a therapeutic treatment for semi-acute and chronic particle-induced pulmonary inflammation. METHODS: Balb/c mice were oropharyngeal instilled with hydrophobic multi-walled carbon nanotube (MWCNT) or hydrophilic crystalline silica (SiO(2)) either as one instillation (semi-acute) or once a week for 4 weeks (chronic). One week later, the mice were placed on either a control or 1% DHA-containing diet for 3 weeks (semi-acute) or 12 weeks (chronic). Mice were assessed for inflammatory signaling within the lung lavage fluid, impact on phagolysosomal membrane permeability, shifts of macrophage phenotype gene expression (M1, M2a, M2b, and M2c), and pulmonary histopathology. RESULTS: DHA increased pulmonary inflammatory markers and lung pathology when mice were exposed to SiO(2). There were trending decreases of inflammatory markers for MWCNT-exposed mice with DHA treatment, however, mostly not statistically significant. CONCLUSION: The anti-inflammatory benefits of DHA treatment depend upon the type of inflammatory particle, magnitude of inflammation, and duration of treatment.

Lysosomes offer a unique arrangement of degradative, exocytic, and signaling capabilities that make their continued function critical to cellular homeostasis. Lysosomes owe their function to the activity of lysosomal ion channels and transporters, which maintain concentration gradients of H+, K+, Ca2+, Na+, and Cl- across the lysosomal membrane. This review examines the contributions of lysosomal ion channels to lysosome function, showing how ion channel function is integral to degradation and autophagy, maintaining lysosomal membrane potential, controlling Ca2+ signaling, and facilitating exocytosis. Evidence of lysosome dysfunction in a variety of disease pathologies creates a need to understand how lysosomal ion channels contribute to lysosome dysfunction. For example, the loss of function of the TRPML1 Ca2+ lysosome channel in multiple lysosome storage diseases leads to lysosome dysfunction and disease pathogenesis while neurodegenerative diseases are marked by lysosome dysfunction caused by changes in ion channel activity through the TRPML1, TPC, and TMEM175 ion channels. Autoimmune disease is marked by dysregulated autophagy, which is dependent on the function of multiple lysosomal ion channels. Understanding the role of lysosomal ion channel activity in lysosome membrane permeability and NLRP3 inflammasome activation could provide valuable mechanistic insight into NLRP3 inflammasome-mediated diseases. Finally, this review seeks to show that understanding the role of lysosomal ion channels in lysosome dysfunction could give mechanistic insight into the efficacy of certain drug classes, specifically those that target the lysosome, such as cationic amphiphilic drugs.

Respirable crystalline silica (RCS) is the leading cause of occupational respiratory disease worldwide. RCS is associated with silicosis, cancer, and autoimmune diseases [1]. Silica (SiO2) is a simple, yet structurally very complex oxide and tens of variable crystalline and amorphous forms exist with different structures and surfaces. Structural heterogeneity is reflected in variable toxic effects, and this in turn generates one of the most intriguing enigmas in particle toxicology, i.e., deciphering the exact molecular nature of the interaction between silica and biological matter.A large set of synthetic and natural, crystalline and amorphous, micrometric and nanometric silica particles were prepared, modified, and characterized. The interaction of these surface-modified silicas with membrane systems of decreasing molecular complexity, e.g., red blood cells, liposomes, and phospholipid supramolecular structures (PLS), was investigated and compared with the results of in vitro and in vivo particle toxicity assessment.A specific silanol (≡Si-OH) sub-group at silica surface, the “nearly free silanols” (NFS), was evidenced as the cause for the membranolytic and inflammatory effect of silica [2]. Silica powders with NFS-rich surfaces caused RBC membrane lysis, and selectively perturbed liposomes and adsorbed PLS. Specific amino groups exposed at the membrane surface are proposed as recognition epitopes for the selective interaction with NFS, which are in turn proposed as the molecular pattern that defines the interaction of silica with biomembranes [3]. Our findings open a new perspective for tailoring less toxic silica particles and for designing improved technological applications of silica. NFS and hydroxylated surface moieties may be also relevant for the toxicity of other respirable mineral dusts, suggesting a new paradigm for particle toxicity mechanism.References[1] Leung et al., Lancet 2012, 379, 2008; [2] Pavan et al., Proc. Natl. Acad. Sci USA 2020, 117, 27836; [3] Pavan et al., sumbitted to ACS Central Science

Inhalation of respirable particles such as silica, has been documented to induce chronic lung diseases such as silicosis, which has been affecting humans since antiquity. Other, smaller materials such as engineered nano-materials (ENM) may trigger lung disease similar to silicosis, as seen in murine models. Within innate immune cells, such as alveolar macrophages, lysosomal membrane permeability (LMP) is proposed as a key and rate-limiting step in silica and ENM-induced inflammation. LMP activates the NLRP3 inflammasome and leads to the maturation and secretion of the pro-inflammatory cytokines IL-1ß and IL-18. In order to better study the membranolytic effects of particles, simpler, non-phagocytic cells like red blood cells (RBC) have been used. In this work, human RBC were treated with crystalline silica (CS), ZnO (ENM) or TiO2 (ENM), and changes to RBC membrane order were analyzed by fluorescence lifetime imaging microscopy of the membrane incorporated probe, Di-4-ANEPPDHQ. RBC treated with methyl-ß-cyclodextrin to extract out cholesterol had a lower fluorescence lifetime of Di-4-ANEPPDHQ, which indicates a decrease in membrane order. Treatment with CS and TiO2 resulted in a significant increase in fluorescence lifetime, indicating that these particles cause localized increases to lipid order. On the other hand, ZnO caused a decrease to the lipid order with a lower fluorescence lifetime. These same three particles were used to treat liposomes composed of either phosphatidylcholine or phosphatidylserine lipids. All three materials induced an increase in lipid order measured by the time-resolved anisotropy of Di-4-ANEPPDHQ. CS and TiO2 changed the order of PC liposomes, while ZnO treatment changed the lipid order of PS liposomes. Addition of cholesterol to these liposomes reduced the effects caused by TiO2 and ZnO treatments. Taken together these results demonstrate a particle-membrane interaction that can disrupt lipid order and that this disruption can be reduced by the presence of cholesterol in the membrane. Selective modulation of lysosomal cholesterol content may be a potential therapeutic intervention for particle-induced inflammatory disease. The research within was funded by the National Institutes of Health grants R01ES023209, 1F32ES027324, P20GM103546, and P30GM103338. It was also funded by the M J Murdock Charitable Trust and the National Science Foundation grant CHE-1531520.

Inhaled crystalline silica causes inflammatory lung diseases, but the mechanism for its unique activity compared to other oxides remains unclear, preventing the development of potential therapeutics. Here, the molecular recognition mechanism between membrane epitopes and "nearly free silanols" (NFS), a specific subgroup of surface silanols, is identified and proposed as a novel broad explanation for particle toxicity in general. Silica samples having different bulk and surface properties, specifically different amounts of NFS, are tested with a set of membrane systems of decreasing molecular complexity and different charge. The results demonstrate that NFS content is the primary determinant of membrane disruption causing red blood cell lysis and changes in lipid order in zwitterionic, but not in negatively charged liposomes. NFS-rich silica strongly and irreversibly adsorbs zwitterionic self-assembled phospholipid structures. This selective interaction is corroborated by density functional theory and supports the hypothesis that NFS recognize membrane epitopes that exhibit a positive quaternary amino and negative phosphate group. These new findings define a new paradigm for deciphering particle-biomembrane interactions that will support safer design of materials and what types of treatments might interrupt particle-biomembrane interactions.

Inhalation of particles results in pulmonary inflammation; however, treatments are currently lacking. Docosahexaenoic acid (DHA) is an omega-3 polyunsaturated fatty acid shown to exhibit anti-inflammatory capabilities. The impact of DHA on particle-induced inflammation is unclear, therefore, the aim of this study was to examine the hypothesis that DHA downregulates macrophage inflammatory responses by altering phagolysosomal membrane permeability (LMP) and shifting macrophage phenotype. Isolated Balb/c alveolar macrophages (AM) were polarized into M1, M2a, M2b, or M2c phenotypes in vitro, treated with DHA, and exposed to a multi-walled carbon nanotube (MWNCT) or crystalline silica (SiO(2)). Results showed minimal cytotoxicity, robust effects for silica particle uptake, and LMP differences between phenotypes. Docosahexaenoic acid prevented these effects to the greatest extent in M2c phenotype. To determine if DHA affected inflammation similarly in vivo, Balb/c mice were placed on a control or 1% DHA diet for 3 weeks, instilled with the same particles, and assessed 24hr following instillation. Data demonstrated that in contrast to in vitro findings, DHA increased pulmonary inflammation and LMP. These results suggest that pulmonary responses in vivo may not necessarily be predicted from single cell responses in vitro.

Electrospun fiber meshes with controlled drug delivery properties have great potential for applications such as wound dressings, tissue engineering, and cancer treatment. However, controlling the d...

Background: While the role of lysosomal membrane permeabilization (LMP) in NP-induced inflammatory responses has been recognized, the underlying mechanism of LMP is still unclear. The assumption ha...

Objective: In this study, we compared in vitro and in vivo bioactivity of nitrogen-doped multi-walled carbon nanotubes (NDMWCNT) to MWCNT to test the hypothesis that nitrogen doping would alter bioactivity.Materials and Methods: High-resolution transmission electron microscopy (TEM) confirmed the multilayer structure of MWCNT with an average layer distance of 0.36 nm, which was not altered by nitrogen doping: the nanomaterials had similar widths and lengths. In vitro studies with THP-1 cells and alveolar macrophages from C57BL/6 mice demonstrated that NDMWCNT were less cytotoxic and stimulated less IL-1β release compared to MWCNT. For in vivo studies, male C57BL/6J mice received a single dose of dispersion medium (DM), 2.5, 10 or 40 µg/mouse of NDMWCNT, or 40 µg/mouse of MWCNT by oropharyngeal aspiration. Animals were euthanized between 1 and 7 days post-exposure for whole lung lavage (WLL) studies.Results and Discussion: NDMWCNT caused time- and dose-dependent pulmonary inflammation. However, it was less than that caused by MWCNT. Activation of the NLRP3 inflammasome was assessed in particle-exposed mice by determining cytokine production in WLL fluid at 1 day post-exposure. Compared to DM-exposed mice, IL-1β and IL-18 were significantly increased in MWCNT- and NDMWCNT-exposed mice, but the increase caused by NDMWCNT was less than MWCNT. At 56 days post-exposure, histopathology determined lung fibrosis in MWCNT-exposed mice was greater than NDMWCNT-exposed mice.Conclusions: These data indicate nitrogen doping of MWCNT decreases their bioactivity, as reflected with lower in vitro and in vivo toxicity inflammation and lung disease. The lower activation of the NLRP3 inflammasome may be responsible. Abbreviations: NDMWCNT: nitrogen-doped multi-walled carbon nanotubes; MWCNT: multi-walled carbon nanotubes; TEM: transmission electron microscopy; HRTEM: high resolution transmission electron microscopy; IL-1s: interleukin-1s; DM: dispersion medium; WLL: whole lung lavage; IL-18: interleukin-18; GSD: geometric standard deviation; XPS: X-ray photoelectron spectroscopy; SEM: standard error of the mean; PMA: phorbol 12-myristate 13-acetate; LPS: lipopolysacharride; LDH: lactate dehydrogenase; AM: alveolar macrophage; PMN: polymorphonuclear leukocyte.

Inhalation is a significant route of exposure to toxic chemicals for electronic waste (e-waste) workers, especially for those whose activities take place in the informal sector. However, there remains a dearth of research on the health effects produced by the hazardous dismantling of e-waste and associated outcomes and biological mechanisms that occur as a result of inhalation exposure. This contemporary review highlights a number of the toxicological and epidemiological studies published on this topic to bring to light the many knowledge gaps that require further research, including in vitro and ex vivo investigations to address the health outcomes and underlying mechanisms of inhaled e-waste-associated pulmonary disease.

The Clean Air and Healthy Homes Program (CAHHP) is a science education outreach program that involves students in research of their own design related to indoor and outdoor air pollution and links with respiratory health. The program, which provides equipment, lesson plans, and support to middle and high school classrooms and professional development for teachers, is an excellent model of how to engage students in relevant and authentic science research and learning. This article describes the current program, how it promotes authentic science learning in secondary science education, and the positive impact it has had on student learning and attitudes.’

Background: As the need to involve more students in STEM learning and future careers becomes more pressing, identifying successful methods of engaging students in meaningful scientific learning that increases their interest in science is essential. Student self-efficacy (their confidence or belief in their ability to accomplish tasks) is closely tied to student interest in science, as is student future scientific possible selves. Material and Methods: This manuscript presents the findings of a study that evaluated the Clean Air and Healthy Homes Program (CAHHP), which provides students the opportunity to design and implement authentic scientific research on indoor air quality issues. The program’s influence on student self-efficacy, scientific research and experimental design skills, and future scientific possible selves was examined. Students (n=169) from six schools completed a pre- and post-assessment at the beginning and end of the program. Results: Results showed the greatest impact on student research self-efficacy, along with improvement in student research and experimental design skills. Conclusions: We conclude that programs promoting authentic learning opportunities aligned with the most recent national science standards show great promise in improving both student interest and skills in science.

BACKGROUND: Respiratory disease is a leading cause of death and disability worldwide. These diseases frequently present with a sex-bias in occurrence and severity, yet the mechanisms responsible for these sex-biases is a critically understudied area of basic research. METHODS: Male and female C57BL/6 mice were exposed to multi-walled carbon nanotubes (MWCNTs) or crystalline silica (cSiO(2)) via oropharyngeal aspiration. Acute assessments were conducted 24-hours and 7-days after a single exposure. In chronic experiments mice were exposed to respective particles once per week for 4 weeks and sacrificed 8 weeks after the last exposure. Lung lavage fluid (LLF) was assessed for markers of injury and inflammation. Immune cell populations were analyzed by flow cytometry and histopathology assessment was performed on lung tissue from chronically exposed mice. RESULTS: Female mice exposed to a single dose of MWCNTs generated a greater eosinophilic response than males 24-hours and 7-days post-exposure. Eosinophilia was accompanied by elevated type 2 cytokine production in LLF. The exaggerated acute response in females was consistent with lung pathology observed in the chronic model: females had greater alveolitis and epithelial cell hyperplasia compared to males. There were no sex-differences 24-hours after cSiO(2) exposure, but by 7-days post-exposure female mice had greater airspace neutrophilia and inflammatory cytokine levels compared to males. However, following repeated exposure to cSiO(2), male mice had worse alveolitis and greater dendritic cell presence within the lungs. CONCLUSIONS: Female mice are more susceptible to acute and chronic MWCNT-induced inflammation, but male mice are more susceptible to chronic cSiO(2)-induced lung pathology.

Exposure to inhaled crystalline silica (cSiO2) is common in occupations where there is cutting, milling, or grinding of cSiO2 containing material. The Occupational Safety and Health Administration estimates that over 2 million workers may be exposed to inhaled cSiO2 in the United States. Inhalation of cSiO2 causes acute and chronic inflammation and may lead pulmonary diseases such as silicosis, as well as an increased risk of developing autoimmune diseases.Unfortunately, treatment of cSiO2-induced lung diseases is limited and primarily focused on supportive care. Inflammation caused by cSiO2 begins when, cSiO2 particles are phagocytized by alveolar macrophages. Interaction between the particle and lysosomal membrane results in damage to the phagolysosomal membrane; a state known as lysosomal membrane permeability (LMP). Leakage of lysosomal contents into the cytoplasm induces NLRP3 inflammasome activation leading to cell death and systemic inflammation. There are currently no pharmaceutical treatments that are directed at this mechanism of disease. Many existing pharmaceuticals become sequestered in the lysosome through an ion-trapping mechanism, and our laboratory aims to determine if these pharmaceuticals are capable of blocking permeabilization of the lysosomal membrane. Previously, our laboratory has shown that the tricyclic antidepressant, imipramine, blocks inflammatory cytokine production and toxicity in alveolar macrophages after exposure to cSiO2. The objective of this research is to determine whether another pharmaceutical, hydroxychloroquine, prevents cSiO2-induced toxicity by blocking LMP in alveolar macrophages. The ability to target the mechanism responsible for initiating particle-induced inflammation may lead to potential treatments and prevention strategies for people exposed to cSiO2.

With the use of engineered nano-materials (ENM) becoming more prevalent, it is essential to determine potential human health impacts. Specifically, the effects on biological lipid membranes will be important for determining molecular events that may contribute to both toxicity and suitable biomedical applications. To better understand the mechanisms of ENM-induced hemolysis and membrane permeability, fluorescence lifetime imaging microscopy (FLIM) was performed on human red blood cells (RBC) exposed to titanium dioxide ENM, zinc oxide ENM, or micron-sized crystalline silica. In the FLIM images, changes in the intensity-weighted fluorescence lifetime of the lipophilic fluorescence probe Di-4-ANEPPDHQ were used to identify localized changes to membrane. Time-resolved fluorescence anisotropy and FLIM of RBC treated with methyl-ß-cyclodextrin was performed to aid in interpreting how changes to membrane order influence changes in the fluorescence lifetime of the probe. Treatment of RBC with methyl-ß-cyclodextrin caused an increase in the wobble-in-a-cone angle and shorter fluorescence lifetimes of di-4-ANEPPDHQ. Treatment of RBC with titanium dioxide caused a significant increase in fluorescence lifetime compared to non-treated samples, indicating increased membrane order. Crystalline silica also increased the fluorescence lifetime compared to control levels. In contrast, zinc oxide decreased the fluorescence lifetime, representing decreased membrane order. However, treatment with soluble zinc sulfate resulted in no significant change in fluorescence lifetime, indicating that the decrease in order of the RBC membranes caused by zinc oxide ENM was not due to zinc ions formed during potential dissolution of the nanoparticles. These results give insight into mechanisms for how these three materials might disrupt RBC membranes and membranes of other cells. The results also provide evidence for a direct correlation between the size, interaction-available surface area of the nano-material and cell membrane disruption.

Background Multinucleated giant cells (MGC) are formed by fusion of macrophages in pathological conditions. These are often studied in the context of the foreign body response to biomaterial implants, but MGC formation is rarely assessed in response to inorganic particles in the lungs. Therefore, a major objective of this study was to quantitatively compare in vivo macrophage fusion resulting from exposure to a spectrum of micron- and nano-sized particles from both environmental and engineered origin, including crystalline silica, multiwalled carbon nanotubes, titanium nanobelts, and crocidolite asbestos. Methods Groups of C57Bl/6 mice were instilled with inorganic particles or PBS control. Lung cells were collected by lavage after one week for cell differentials, quantification of macrophage fusion, and microscopic observation of particle uptake. Results MGC were present in lungs of all mice exposed to particles; no MGC were found in control mice. Asbestos exposure resulted in significant macrophage fusion, which coincided with significantly increased total lavage cells and percent neutrophils. Microscopic observations show particle internalization in MGC and a unique case of potential heterotypic fusion of macrophages with neutrophils. Conclusion MGC can form in the lungs of mice within a relatively short one-week time period after particle exposure. The number of MGC was sufficient for quantification and statistical analysis, indicating that MGC formation was more than simply a rare chance occurrence. Observations of particles within MGC warrants further investigation of MGC involvement in inflammation and particle clearance.

OBJECTIVES: Inhalation of crystalline silica (cSiO2) has been linked to the pathogenesis of human autoimmunity such as systemic lupus erythematosus. Alveolar macrophages (MΦ) are a major site of inflammasome activated release of pro-inflammatory cytokines, thus plays a vital role in the onset and progression of lupus. Diet supplemented with probiotics have been shown to decrease disease severity. The model used was the murine macrophage RAW 264.7 cell line (WT) and RAW 264.7 stably transfected with the inflammasome adapter protein ASC (RAW-ASC) as WT lacks a functional inflammasome. cSiO2 elicits robust inflammasome activation and IL-1β release in lipopolysaccharide (LPS)-primed RAW-ASC cells. We hypothesize that probiotic-associated cell-free fraction (postbiotics) could decrease cSiO2-induced cytotoxicity in MΦ and suppress pro-inflammatory IL-1 cytokines release. METHODS: Lactobacillus rhamnosus (LGG), L. reuteri (REU), and Bifidobacterium lactis (BB12) were grown in de Man, Rogosa and Sharpe (MRS) broth. Postbiotics were collected and filtered between OD 1.0–3.0. RAW-ASC and WT cells were primed with LPS, stimulated with cSiO2 and treated with postbiotics or MRS control. Cytotoxicity and IL-1 cytokine release was measured by an Lactate dehydrogenase (LDH) assay and ELISA, respectively. RESULTS: Postbiotics obtained at OD 3.0 exhibited greatest protective effect on cSiO2-induced cytotoxicity compared to OD 1.0 and 2.0. RAW-ASC stimulated with cSiO2 produced robust IL-1β release (272.8 ± 20.9 pg/mL) compared to WT (70.67 ± 9.6 pg/mL). Postbiotics obtained from LGG, REU, and BB12 grown to OD 1.0–3.0 decreased IL-1β release from RAW-ASC. LGG-associated postbiotics from all ODs reduced IL-1β release in WT. However, postbiotics obtained from BB12 at OD 3.0 but not OD 1.0 and 2.0 decreased IL-1β release. Interestingly, only REU postbiotics reduced IL-1α release in RAW-ASC while LGG and BB12 did not alter IL-1α production in both cell lines. CONCLUSIONS: Dietary postbiotics can protect against cSiO2-induced cytotoxicity in MΦ. Postbiotics from LGG and BB12 down-regulated release of IL-1β but not IL-1α. We propose that the RAW-ASC cell model is a robust model to study mechanisms by which dietary constituents can activate or suppress inflammasome function. FUNDING SOURCES: This work was supported by the USDA National Institute of Food and Federal Appropriations.

Background A very pure multi-walled carbon nanotube (MWCNT) that was shown to have very low toxicity in vitro, was evaluated for lung and systemic effects and distribution following inhalation exposure. Methods B6C3F1/N mice were exposed to varying doses (0, 0.06, 0.2, and 0.6 mg/m3) of the (99.1% carbon) MWCNT by inhalation for 30 days (excluding weekends). Ten days following the last exposure, the lungs and spleen were harvested and processed for histology and immune cell population assessment. In addition, lung lavage cells and fluid were analyzed. Stimulated Raman scattering (SRS) was used to identify particles in the lungs, spleen, kidneys, liver, mediastinal and brachial lymph nodes, and olfactory bulb. Splenic tissue sections were stained with hematoxylin and eosin (H&E) for light microscopic histopathology assessment. Blood plasma was analyzed for cytokines and cathepsins. A section of the spleen was processed for RNA isolation and relative gene expression for 84 inflammation-related cytokines/chemokines. Results Following MWCNT exposure, particles were clearly evident in the lungs, spleens, lymph nodes and olfactory bulbs, (but not livers or kidneys) of exposed mice in a dose-dependent manner. Examination of the lavaged lung cells was unremarkable with no significant inflammation indicated at all particle doses. In contrast, histological examination of the spleen indicated the presence of apoptotic bodies within T cells regions of the white pulp area. Isolated splenic leukocytes had significant changes in various cells including an increased number of proinflammatory CD11b+Ly6C+ splenic cells. The gene expression studies confirmed this observation as several inflammation-related genes were upregulated particularly in the high dose exposure (0.6 mg/m3). Blood plasma evaluations showed a systemic down-regulation of inflammatory cytokines and a dose-dependent up-regulation of lysosomal cathepsins. Conclusions The findings in the lungs were consistent with our hypothesis that this MWCNT exposure would result in minimal lung inflammation and injury. However, the low toxicity of the MWCNT to lung macrophages may have contributed to enhanced migration of the MWCNT to the spleen through the lymph nodes, resulting in splenic toxicity and systemic changes in inflammatory mediators.

The potential health effects associated with the widespread use of engineered nanomaterials are broadly concerning to developers, researchers, and policy-makers alike. While extensive studies have been undertaken to understand the immediate consequences of exposure to these materials, the long-term effects associated with potential translocation to secondary and even tertiary organs are less well understood. Further, mechanisms underlying the toxic properties of nanomaterials after translocating to distal organs, if any, are largely unknown at this time. Here, we describe the current state of knowledge regarding translocation of commonly studied and primarily nonmedical engineered nanomaterials following exposure and discuss potential mechanisms and consequences to health outcomes. Current evidence suggests that nanomaterials are redistributed, at least in part, to secondary organs throughout the body. While the properties associated with any given nanomaterial may influence its redistribution, the most likely explanation for adverse effects following translocation involves increased inflammation leading to pathology. Whether more persistent nanomaterials will eventually be cleared from the body after exposure remains an important question. Further, information regarding the effects of systemic trafficking on the basic characteristics of nanomaterials would help predict whether nanomaterials will accumulate in specific organs and/or cause pathologies in secondary or even tertiary locations.

Biomaterials have a great potential to improve human health, however in vitro and in vivo studies are necessary to provide information on their efficacy and safety. This study reports on a comprehensive evaluation of core-shell electrospun fibers loaded with silver nanoparticles (Ag NP) where the delivery rate was controlled by different sizes of Ag NP and thermoresponsive poly(n-isopropylacrylamide) (PNIPAM) hydrogel particles. Fiber meshes also contain zinc oxide nanoparticles (ZnO NP), to improve pore structure for controlled release of Ag NP. In vitro cytotoxicity studies using cultured human A549 epithelial cells demonstrated that the ZnO NP component, which is known to cause cytotoxicity, of the fiber meshes did cause measurable cell death. In vitro antibacterial efficacy of the fiber meshes was shown with rapid and efficient growth inhibition in E. coli bacterial culture. Fiber meshes were implanted subcutaneously for up to 27 days in male and female C57BL/6 mice to evaluate the in vivo drug release and biocompatibility. Hyperspectral microscopy was used as an advanced tool to determine precise location of released Ag NP into the skin compared to the conventional tissue staining methods. Results suggested that Ag NP were continuously released over 27 days of implantation in mice. Hyperspectral imaging revealed that released Ag NP dispersed in the dermis of male mice, however, Ag NP accumulated in the hair follicles of female mice (Figure). Mice implanted with fiber meshes containing ZnO NP had better hair regrowth and wound healing, which was in contrast to in vitro cytotoxicity results. These findings suggest that these newly developed fiber meshes can have unique long-term release of drugs loaded in the fiber core and appear to be biocompatible. The differences in the sex-bias outcome suggest the opportunity for development of sex-specific drug delivery systems.

OBJECTIVE: The toxicity of silver nanomaterials in various forms has been extensively evaluated, but the toxicity of silver nanocarbon composites is less well understood. Therefore, silver-carbon nanotube composites (Ag-MWCNT-COOH) and silver-graphene oxide composites (Ag-GO) were synthesized by microwave irradiation and evaluated in two in vitro cell models. MATERIALS/METHODS: Toxicity of silver nanosphere (Ag), Ag-MWCNT-COOH and Ag-GO were analyzed by MTS assay and LDH assay in primary C57BL/6 murine alveolar macrophages and human THP-1 cells. Activation of NLRP3 inflammasome by particle variants in these models was done by proxy using LPS co-culture and IL-1β release. RESULTS: The results depended on the model, as the amount of Ag on the modified carbon resulted in slightly increased toxicity for the murine cells, but did not appear to affect toxicity in the human cell model. IL-1β release from carbon particle-exposures was decreased by the presence of Ag in both cell models. Suspensions of Ag-MWCNT-COOH, Ag-GO and Ag in artificial lysosomal fluid were prepared and ICP-MS was used to detect Ag ions concentration in three silver suspension/solutions. The amount of Ag ions released from Ag-MWCNT-COOH and Ag-GO were similar, which were both lower than that of Ag nanospheres. CONCLUSION: The results suggest the bioactivity of silver composites may be related to the amount of Ag ions released, which can be dependent on the cell model under investigation.

Allergic asthma remains an inadequately understood disease. In utero exposure to environmental tobacco smoke (ETS) has been identified as an environmental exposure that can increase an individual's asthma risk. To improve our understanding of asthma onset and development, we examined the effect of in utero ETS exposure on allergic disease susceptibility in an asthmatic phenotype using a house dust mite (HDM) allergen-induced murine model. Pregnant C57BL/6 mice were exposed to either filtered air or ETS during gestation, and their offspring were further exposed to HDM at 6-7 weeks old to induce allergic inflammation. Methylation in the promoter regions of allergic inflammation-related genes and genomic DNA was quantified. Exposure to HDM resulted in the onset of allergic lung inflammation, with an increased presence of inflammatory cells, Th2 cytokines (IL-4, IL-5, and IL-13), and airway remodeling. These asthmatic phenotypes were significantly enhanced when the mice had been exposed to in utero ETS. Furthermore, prenatal ETS exposure and subsequent HDM (ETS/HDM)-induced asthmatic phenotypes agree with methylation changes in the selected asthma-related genes, including IL-4, IL-5, IL-13, INF-γ, and FOXP3. Global DNA methylation was significantly lower in ETS/HDM-exposed mice than that of controls, which coincides with the results observed in lung, spleen, and blood DNAs. Prenatal ETS exposure resulted in a severe increase in allergic inflammatory responses after an HDM challenge, with corresponding methylation changes. Prenatal ETS exposure may influence developmental plasticity and result in altered epigenetic programming, leading to an increased susceptibility to asthma. Environ. Mol. Mutagen. 58:423-433, 2017. © 2017 Wiley Periodicals, Inc.