Influence of Cigarette Aerosol in Alpha-Synuclein Oligomerization and Cell Viability in SH-SY5Y: Implications for Parkinson's Disease.

Although cigarette aerosol exposure is associated with various adverse health issues, its impact on Parkinson's disease (PD) remains elusive. Here, we investigated the effect of cigarette aerosol extract (CAE) on SH-SY5Y cells for the first time, both with and without α-synuclein (α-Syn) overexpression. We found that α-Syn aggravates CAE-induced cell death, oxidative stress, and mitochondrial dysfunction. Fluorescence cross-correlation spectroscopy (FCCS) revealed a dual distribution of α-Syn within the cells, with homogeneous regions indicative of monomeric α-Syn and punctated regions, suggesting the formation of oligomers. Moreover, we observed colocalization of α-Syn oligomers with lysosomes along with a reduction in autophagy activity. These findings suggest that α-Syn overexpression exacerbates CAE-induced intracellular cytotoxicity, mitochondrial dysfunction, and autophagy dysregulation, leading to elevated cell mortality. Our findings provide new insights into the pathogenic mechanisms linking exposure to cigarette aerosols with neurodegenerative diseases.

A Comparative Study of the Effects of Electronic Cigarette and Traditional Cigarette on the Pulmonary Functions of C57BL/6 Male Mice.

INTRODUCTION: Electronic cigarettes (E-cigs) are in a controversial state. Although E-cig aerosol generally contains fewer harmful substances than smoke from burned traditional cigarettes, aerosol along with other compounds of the E-cigs may also affect lung functions and promote the development of lung-related diseases. We investigated the effects of E-cig on the pulmonary functions of male C57BL/6 mice and reveal the potential underlying mechanisms. METHODS: A total of 60 male C57BL/6 mice were randomly divided into four groups. They were exposed to fresh-air, traditional cigarette smoke, E-cig vapor with 12 mg/mL of nicotine, and E-cig with no nicotine for 8 weeks. Lung functions were evaluated by using quantitative analysis of the whole body plethysmograph, FlexiVent system, lung tissue histological and morphometric analysis, and RT-PCR analysis of mRNA expression of inflammation-related genes. In addition, the effects of nicotine and acrolein on the survival rate and DNA damage were investigated using cultured human alveolar basal epithelial cells. RESULTS: Exposure to E-cig vapor led to significant changes in lung functions and structures including the rupture of the alveolar cavity and enlarged alveolar space. The pathological changes were also accompanied by increased expression of interleukin-6 and tumor necrosis factor-α. CONCLUSIONS: The findings of the present study indicate that the safety of E-cig should be further evaluated. IMPLICATIONS: Some people currently believe that using nicotine-free E-cigs is a safe way to smoke. However, our research shows that E-cigs can cause lung damage regardless of whether they contain nicotine.

Comparative assessment of electronic nicotine delivery systems aerosol and cigarette smoke on endothelial cell migration: The Replica Project.

Cigarette smoking is associated with impairment of repair mechanisms necessary for vascular endothelium homeostasis. Reducing the exposure to smoke toxicants may result in the mitigation of the harmful effect on the endothelium and cardiovascular disease development. Previous investigations evaluated in vitro the effect of electronic cigarette (EC) compared with cigarette smoke demonstrating a significant reduction in human umbilical vein endothelial cells (HUVECs) migration inhibition following EC aerosol exposure. In the present study, we replicated one of these studies, evaluating the effects of cigarette smoke on endothelial cell migration compared with aerosol from EC and heated tobacco products (HTPs). We performed an in vitro scratch wound assay on endothelial cells with a multi-center approach (ring-study) to verify the robustness and reliability of the results obtained in the replicated study, also testing the effect of aerosol from two HTPs on endothelial cells. Consistently with the original study, we observed a substantial reduction of the effects of aerosol from EC and HTPs on endothelial cell migration compared with cigarette smoke. While cigarette smoke reduced endothelial wound healing ability already at low concentrations (12.5%) and in a concentration-dependent manner, EC and HTPs aerosol showed no effect on endothelial cells until 80%-100% concentrations. In conclusion, our study further confirms the importance of EC and tobacco heated products as a possible harm reduction strategy for cardiovascular diseases development in smokers.

The pulmonary pharmacokinetics and anti-inflammatory effects after intratracheal and intravenous administration of Chuankezhi injection.

BACKGROUND: Chuankezhi injection (CKZ) is a traditional Chinese medicine for the treatment of respiratory diseases and has been often used off-label as a nebulization therapy. However, little is known about the aerosolization performance and pulmonary fate of the inhaled CKZ. This study aimed to evaluate the aerodynamic characteristics of nebulizer generated aerosols and to compare the properties of pharmacokinetics, lung distribution and anti-inflammation effects of CKZ after intratracheal and intravenous administration. METHODS: The nebulization performance was evaluated in vitro based on the aerodynamic particle size distribution and aerosol output. The concentrations of epimedins A, B, C and icariin, the main active ingredients of CKZ, in plasma, bronchoalveolar lavage fluids (BALF) and lung tissues were measured by LC-MS/MS analysis. The pulmonary anti-inflammatory efficacy were tested using LPS-induced pulmonary inflammation mice model as indicated by the total cells counts, and the levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in BALF. RESULTS: The aerosols of CKZ generated by a commercial nebulizer showed excellent aerodynamic properties and delivery output. Following intratracheal instillation of CKZ, epidemins A, B and C, and icariin, were absorbed into the bloodstream with the mean absorption time varying from 101.8 min to 271.8 min, and their absolute bioavailabilities ranging from 26.4 % to 104 %. The instillation of CKZ increased the lung to plasma concentration ratios by 25.5-718 folds compared to intravenous administration, leading to improved and prolonged local anti-inflammatory effects. CONCLUSION: Nebulization therapy of CKZ could be a promising alternative to the injectable counterpart.

Aerosol delivery, but not intramuscular injection, of adenovirus-vectored tuberculosis vaccine induces respiratory-mucosal immunity in humans.

BackgroundAdenovirus-vectored (Ad-vectored) vaccines are typically administered via i.m. injection to humans and are incapable of inducing respiratory mucosal immunity. However, aerosol delivery of Ad-vectored vaccines remains poorly characterized, and its ability to induce mucosal immunity in humans is unknown. This phase Ib trial evaluated the safety and immunogenicity of human serotype-5 Ad-vectored tuberculosis (TB) vaccine (AdHu5Ag85A) delivered to humans via inhaled aerosol or i.m. injection.MethodsThirty-one healthy, previously BCG-vaccinated adults were enrolled. AdHu5Ag85A was administered by single-dose aerosol using Aeroneb Solo Nebulizer or by i.m. injection. The study consisted of the low-dose (LD) aerosol, high-dose (HD) aerosol, and i.m. groups. The adverse events were assessed at various times after vaccination. Immunogenicity data were collected from the peripheral blood and bronchoalveolar lavage samples at baseline, as well as at select time points after vaccination.ResultsThe nebulized aerosol droplets were < 5.39 µm in size. Both LD and HD of AdHu5Ag85A administered by aerosol inhalation and i.m. injection were safe and well tolerated. Both aerosol doses, particularly LD, but not i.m., vaccination markedly induced airway tissue-resident memory CD4+ and CD8+ T cells of polyfunctionality. While as expected, i.m. vaccination induced Ag85A-specific T cell responses in the blood, the LD aerosol vaccination also elicited such T cells in the blood. Furthermore, the LD aerosol vaccination induced persisting transcriptional changes in alveolar macrophages.ConclusionInhaled aerosol delivery of Ad-vectored vaccine is a safe and superior way to elicit respiratory mucosal immunity. This study warrants further development of aerosol vaccine strategies against respiratory pathogens, including TB and COVID-19.Trial registrationClinicalTrial.gov, NCT02337270.FundingThe Canadian Institutes for Health Research (CIHR) and the Natural Sciences and Engineering Research Council of Canada funded this work.

Targeted Drug Delivery Technologies Potentiate the Overall Therapeutic Efficacy of an Indole Derivative in a Mouse Cystic Fibrosis Setting.

Inflammation plays a major role in the pathophysiology of cystic fibrosis (CF), a multisystem disease. Anti-inflammatory therapies are, therefore, of interest in CF, provided that the inhibition of inflammation does not compromise the ability to fight pathogens. Here, we assess whether indole-3-aldehyde (3-IAld), a ligand of the aryl hydrocarbon receptor (AhR), may encompass such an activity. We resorted to biopharmaceutical technologies in order to deliver 3-IAld directly into the lung, via dry powder inhalation, or into the gut, via enteric microparticles, in murine models of CF infection and inflammation. We found the site-specific delivery of 3-IAld to be an efficient strategy to restore immune and microbial homeostasis in CF organs, and mitigate lung and gut inflammatory pathology in response to fungal infections, in the relative absence of local and systemic inflammatory toxicity. Thus, enhanced delivery to target organs of AhR agonists, such as 3-IAld, may pave the way for the development of safe and effective anti-inflammatory agents in CF.

Beneficial impact of cathelicidin on hypersensitivity pneumonitis treatment-In vivo studies.

Cathelicidin (CRAMP) is a defence peptide with a wide range of biological responses including antimicrobial, immunomodulatory and wound healing. Due to its original properties the usefulness of CRAMP in the treatment of pulmonary fibrosis was assessed in a murine model of hypersensitivity pneumonitis (HP). The studies were conducted on mouse strain C57BL/6J exposed to a saline extract of Pantoea agglomerans cells (HP inducer). Cathelicidin was administered in the form of an aerosol during and after HP development. Changes in the composition of immune cell populations (NK cells, macrophages, lymphocytes: Tc, Th, Treg, B), were monitored in lung tissue by flow cytometry. Extracellular matrix deposition (collagens, hydroxyproline), the concentration of cytokines involved in inflammatory and the fibrosis process (IFNγ, TNFα, TGFß1, IL1ß, IL4, IL5, IL10, IL12α, IL13) were examined in lung homogenates by the ELISA method. Alterations in lung tissue morphology were examined in mouse lung sections stained with haematoxylin and eosin as well as Masson trichrome dyes. The performed studies revealed that cathelicidin did not cause any negative changes in lung morphology/structure, immune cell composition or cytokines production. At the same time, CRAMP attenuated the immune reaction induced by mice chronic exposure to P. agglomerans and inhibited hydroxyproline and collagen deposition in the lung tissue of mice treated with bacteria extract. The beneficial effect of CRAMP on HP treatment was associated with restoring the balance in quantity of immune cells, cytokines production and synthesis of extracellular matrix components. The presented study suggests the usefulness of cathelicidin in preventing lung fibrosis; however, cathelicidin was not able to reverse pathological changes completely.

Integrative Analysis of lncRNA-mRNA Coexpression in Human Lung Epithelial Cells Exposed to Dimethyl Selenide-Derived Secondary Organic Aerosols.

Dimethyl selenide (DMSe) is one of the major volatile organoselenium compounds released into the atmosphere through plant metabolism and microbial methylation. DMSe has been recently revealed as a precursor of secondary organic aerosol (SOA), and its resultant SOA possesses strong oxidizing capability toward thiol groups that can perturb several major biological pathways in human airway epithelial cells and is linked to genotoxicity, DNA damage, and p53-mediated stress responses. Mounting evidence has suggested that long noncoding RNAs (lncRNAs) are involved in stress responses to internal and environmental stimuli. However, the underlying molecular interactions remain to be elucidated. In this study, we performed integrative analyses of lncRNA-mRNA coexpression in the transformed human bronchial epithelial BEAS-2B cell line exposed to DMSe-derived SOA. We identified a total of 971 differentially expressed lncRNAs in BEAS-2B cells exposed to SOA derived from O3 and OH oxidation of DMSe. Gene ontology (GO) network analysis of cis-targeted genes showed significant enrichment of DNA damage, apoptosis, and p53-mediated stress response pathways. trans-Acting lncRNAs, including PINCR, PICART1, DLGAP1-AS2, and LINC01629, known to be associated with human carcinogenesis, also showed altered expression in cell treated with DMSe-SOA. Overall, this study highlights the regulatory role of lncRNAs in altered gene expression induced by DMSe-SOA exposure.

Aerosol 1,25-dihydroxyvitamin D3 supplementation: A strategy to boost anti-tumor innate immune activity.

BACKGROUND: 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] plays a role in calcium homeostasis but can also exert immunomodulatory effects. In lungs, characterized by a particular immunosuppressive environment primarily due to the presence of alveolar macrophages (AM), 1,25(OH)2D3 has been shown to favor the immune response against pathogens. Here, we explored the ability of aerosolized 1,25(OH)2D3 to locally promote an anti-tumor phenotype in alveolar macrophages (AM) in the treatment of lung metastases. METHODS: Cytotoxicity assay has been used to assess the capability of AM, in vitro treated of not with 1,25(OH)2D3, to stimulate NK cells. Sulforhodamine B (SRB) assay has been used to assess the effect of 1,25(OH)2D3 on MC-38 and B16 tumor cells in vitro growth. 1,25(OH)2D3 was aerosolized in immunocompetent mouse models to evaluate the effect of local administration of 1,25(OH)2D3 on in vivo growth of MC-38 and B16 tumor cells within lungs and on infiltrating immune cells. RESULTS: In vitro incubation of naïve AM with 1,25(OH)2D3 improved their ability to stimulate NK cell cytotoxicity. In vivo aerosolized 1,25(OH)2D3 significantly reduced the metastatic growth of MC-38 colon carcinoma, a tumor histotype that frequently metastasizes to lung in human. Immune infiltrate obtained from digested lungs of 1,25(OH)2D3-treated mice bearing MC-38 metastases revealed an increased expression of MHCII and CD80 on AM and an up-modulation of CD69 expression on effector cells that paralleled a strong increased ability of these cells to kill MC-38 tumor in vitro. CONCLUSIONS: Together, these data show that aerosol delivery can represent a feasible and novel approach to supplement 1,25(OH)2D3 directly to the lungs promoting the activation of local immunity against cancer.

Comparison between microcatheter and nebulizer for generating Pressurized IntraPeritoneal Aerosol Chemotherapy (PIPAC).

BACKGROUND: This study compares an endoscopic microcatheter and a nebulizer for delivering Pressurized IntraPeritoneal Aerosol Chemotherapy (PIPAC). METHODS: This is an in vitro and ex vivo study in an established model (inverted bovine urinary bladder). Four parameters were compared to determine the performance of a micro-perforated endoscopic spray catheter vs. state-of-the art, nozzle technology: (1) surface coverage and pattern with methylene blue on blotting paper at three different distances; (2) median aerodynamic diameter (MAD) of aerosol droplets with three different solutions (H2O, Glc 5% and silicon oil); (3) depth of tissue penetration of doxorubicin (DOX) and (4) tissue concentration of cisplatin (CIS) and DOX using standard clinical solutions. RESULTS: The spray area covered by the microcatheter was larger (p < 0.001) but its pattern was inhomogenous than with the nozzle technology. We found that aerosol droplets were larger in the test group than in the control group for all three solutions tested. Median tissue penetration of DOX was lower (980 µm) with the microcatheter than with the nebulizer (1235 µm) and distribution was more heterogeneous ( = 0.003) with the microcatheter. The median tissue concentration of DOX and CIS was lower and concentration of DOX was more heterogeneous with the microcatheter (p = 0.002). CONCLUSIONS: This investigation has revealed that microcatheter technology generates larger aerosol droplet size, less drug tissue penetration and lower drug tissue concentration than the current nozzle technology. In the absence of clinical studies, use of microcatheters for delivering PIPAC can not be recommended at this stage.

Kuanxiong aerosol inhibits apoptosis and attenuates isoproterenol-induced myocardial injury through the mitogen-activated protein kinase pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Kuanxiong aerosol (KXA) is a common clinical drug based on Fangxiang Wentong (FXWT) therapy in the treatment of angina pectoris. However, the pharmacological mechanism of KXA in the prevention and treatment of myocardial injury (MI) is not clear. AIM OF THE STUDY: The purpose of this study was to explore the protective effect of KXA on isoproterenol (ISO)-induced MI in rats. MATERIALS AND METHODS: The study included male Wistar Kyoto rats (age: 6 weeks). The rats were randomly divided into the following 5 groups (n = 6 per group): control group, ISO group, isosorbide mononitrate (ISMN) group (5 mg/kg), KXA-L group (0.1 mL/kg), and KXA-H group (0.3 mL/kg). The rats in the last three groups were given intragastric administration for 14 days, and rats in control group and ISO group were given the same amount of normal saline daily. ISO (120 mg/kg) was used to induce MI on the 13th and 14th days. We assessed electrocardiograms (ECGs), myocardial specific enzymes, histopathological changes, and apoptosis. RESULTS: We found that KXA reduced the increase in the ST-segment amplitude (elevation or depression) and the levels of myocardial marker enzymes induced by ISO in MI rats, improved the pathological changes in myocardial tissue, and reduced cardiomyocyte apoptosis. At the same time, KXA significantly inhibited the up-regulation of caspase-3 and Bax expression and down-regulation of Bcl-2 expression induced by ISO. RNA sequencing showed that 90 up-regulated genes induced by ISO were down-regulated after KXA treatment, whereas 27 down-regulated genes induced by ISO were up-regulated after KXA treatment. In addition, KEGG pathway enrichment analysis showed that the mitogen-activated protein kinase (MAPK) signaling pathway may be an important target of KXA in the treatment of ISO-induced MI in rats. The results of RNA sequencing verified by Western blot analysis showed that KXA significantly inhibited the activation of the ISO-induced MAPK pathway. CONCLUSIONS: KXA improves cardiac function in MI rats by inhibiting apoptosis mediated by the MAPK signaling pathway.

Chemical Elements in Electronic Cigarette Solvents and Aerosols Inhibit Mitochondrial Reductases and Induce Oxidative Stress.

INTRODUCTION: Chemical elements and their toxicity were evaluated in electronic cigarette (EC) solvents, fluids, and aerosols. AIMS AND METHODS: Element identification and quantification in propylene glycol (PG), glycerin (G), refill fluids before and after use, and aerosols was done using inductively coupled plasma optical emission spectrometry. Cytotoxicity and oxidative stress were evaluated using in vitro assays. RESULTS: Seven elements were present in PG, G, and popular refill fluids, and they transferred to aerosols made with ECs. Selenium was in all products (0.125-0.292 mg/L), while arsenic, aluminum, and tin were frequently in solvent and refill fluid samples at lower concentrations. Iron, chromium, copper, nickel, zinc, and lead were only detected in fluid after EC use, indicating they came from heated atomizers. Elements transferred most efficiently to aerosols made with second-/third-generation ECs. Of the elements in fluid, selenium and arsenic were the most cytotoxic to human bronchial epithelial cells (BEAS-2B) and pulmonary fibroblasts in the 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide assay. Selenium increased superoxide production in mitochondria and nucleoli and elevated selenoprotein H in nucleoli of BEAS-2B cells at concentrations found in EC aerosols (10 nM or 0.002 mg/L). CONCLUSIONS: Elements in EC aerosols came from both e-fluids and atomizing units. Within second-/third-generation products, transfer became more efficient as power increased. In vitro responses occurred at concentrations of selenium found in some EC aerosols. Human exposure to chemical elements in ECs could be reduced by regulating (decreasing) allowable EC power and by improving the purity of PG and G. IMPLICATIONS: PG, G, refill fluids, and e-fluids contained potentially toxic chemical elements that transferred to aerosols. Transfer was more efficient in second- and third-generation EC products and increased as power increased. Selenium and arsenic were the most cytotoxic of the elements tested in the 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide assay. Selenium tetrachloride-induced oxidative stress in BEAS-2B cells, but not in human pulmonary fibroblasts. All fluids contained selenium above the concentration that induced oxidative stress in human bronchial epithelial cells. Selenium increased superoxide in mitochondria and nucleoli and increased selenoprotein H, a redox responsive DNA-binding protein that is upregulated by superoxide and an indicator of nucleolar stress. EC users are exposed to elements in aerosols, which may with chronic exposure contribute to diseases associated with oxidative stress.

An experimental aerosol air-agar interface mouse lymphoma assay methodology.

This work investigates a completely novel and experimental concept of exposing L5178Y cells at the air-agar-interface to mainstream cigarette smoke aerosol (Kentucky reference 3R4F). This study highlights the associated challenges of combining a suspension cell line alongside an in vitro aerosol exposure system. To achieve a monolayer, cells were 'seeded' in a concentrated cell super-mix suspension onto an RPMI/agar-matrix -base. The resulting cell suspension media was adsorbed into the agar base leaving the L5178Y cells lightly suspended on the agar surface, approximating a monolayer. Cells were deemed supportable on the agar-matrix, viable and recoverable. Using Vitrocell VC 10 exposure system and the Ames 4 exposure module, L5178Y cells were successfully exposed to a dynamic cigarette smoke aerosol, recovered and assessed for mutant frequencies, using standard assay procedures. Method development included assessment of flowing air conditions, plating efficiency and recovery of L5178Y cells from the agar-matrix surface. Positive controls MMS and B[a]P were successfully incorporated into the agar-matrix and metabolic activation was achieved by S-9 incorporation into the same agar-base-matrix. B[a]P demonstrated metabolic activation and positive response, suggesting a clear cellular interaction with the agar-matrix. Whole smoke exposed cells in the presence of metabolic activation showed a clear dose response and increasing mutant frequencies, well in excess of the controls (air and incubator) and the global evaluation factor following a 2 or 3 day expression period. This experimental concept demonstrates that L5178Y cells can be exposed to cigarette smoke aerosol, using a completely novel and a previously untested approach. Although this work successfully demonstrates the approach is viable and cells can be plated and maintained on an agar-matrix, more optimisation and robustness assessment is required before it can be considered fully adapted and used alongside other whole aerosol methodologies for the assessment of cigarette smoke and other inhaled aerosols.

Prenatal Exposure to Electronic-Cigarette Aerosols Leads to Sex-Dependent Pulmonary Extracellular-Matrix Remodeling and Myogenesis in Offspring Mice.

Electronic-cigarette (e-cig) vaping is a serious concern, as many pregnant women who vape consider it safe. However, little is known about the harmful effects of prenatal e-cig exposure on adult offspring, especially on extracellular-matrix (ECM) deposition and myogenesis in the lungs of offspring. We evaluated the biochemical and molecular implications of maternal exposure during pregnancy to e-cig aerosols on the adult offspring of both sexes, with a particular focus on pulmonary ECM remodeling and myogenesis. Pregnant CD-1 mice were exposed to e-cig aerosols with or without nicotine, throughout gestation, and lungs were collected from adult male and female offspring. Compared with the air-exposed control group, female mice exposed to e-cig aerosols, with or without nicotine, demonstrated increased lung protein abundance of LEF-1 (lymphoid enhancer-binding factor 1), fibronectin, and E-cadherin, whereas altered E-cadherin and PPARγ (peroxisome proliferator-activated receptor γ) levels were observed only in males exposed to e-cig aerosols with nicotine. Moreover, lipogenic and myogenic mRNAs were dysregulated in adult offspring in a sex-dependent manner. PAI-1 (plasminogen activator inhibitor-1), one of the ECM regulators, was significantly increased in females exposed prenatally to e-cig aerosols with nicotine and in males exposed to e-cig aerosols compared with control animals exposed to air. MMP9 (matrix metalloproteinase 9), a downstream target of PAI-1, was downregulated in both sexes exposed to e-cig aerosols with nicotine. No differences in lung histology were observed among any of the treatment groups. Overall, adult mice exposed prenatally to e-cig aerosols could be predisposed to developing pulmonary disease later in life. Thus, these findings suggest that vaping during pregnancy is unsafe and increases the propensity for later-life interstitial lung diseases.

Inhalable Nanocomposite Microparticles with Enhanced Dissolution and Superior Aerosol Performance.

Previous studies have shown that combining colistin (Col), a cationic polypeptide antibiotic, with ivacaftor (Iva), a cystic fibrosis (CF) drug, could achieve synergistic antibacterial effects against Pseudomonas aeruginosa. The purpose of this study was to develop dry powder inhaler formulations for co-delivery of Col and Iva, aiming to treat CF and lung infection simultaneously. In order to improve solubility and dissolution for the water-insoluble Iva, Iva was encapsulated into bovine serum albumin (BSA) nanoparticles (Iva-BSA-NPs). Inhalable composite microparticles of Iva-BSA-NPs were produced by spray-freeze-drying using water-soluble Col as the matrix material and l-leucine as an aerosol enhancer. The optimal formulation showed an irregularly shaped morphology with fine particle fraction (FPF) values of 73.8 ± 5.2% for Col and 80.9 ± 4.1% for Iva. Correlations between "D×ρtapped" and FPF were established for both Iva and Col. The amorphous solubility of Iva is 66 times higher than the crystalline solubility in the buffer. Iva-BSA-NPs were amorphous and remained in the amorphous state after spray-freeze-drying, as examined by powder X-ray diffraction. In vitro dissolution profiles of the selected DPI formulation indicated that Col and Iva were almost completely released within 3 h, which was substantially faster regarding Iva release than the jet-milled physical mixture of the two drugs. In summary, this study developed a novel inhalable nanocomposite microparticle using a synergistic water-soluble drug as the matrix material, which achieved reduced use of excipients for high-dose medications, improved dissolution rate for the water-insoluble drug, and superior aerosol performance.

Electronic Nicotine Delivery System Aerosol-induced Cell Death and Dysfunction in Macrophages and Lung Epithelial Cells.

Electronic nicotine delivery system (ENDS) use is outpacing our understanding of its potential harmful effects. Homeostasis of the lung is maintained through proper balance of cell death, efferocytic clearance, and phagocytosis of pathogens. To investigate whether ENDS use has the potential to alter this balance, we developed physiologically relevant ENDS exposure paradigms for lung epithelial cells and primary macrophages. In our studies, cells were exposed directly to aerosol made from carefully controlled components with and without nicotine. We found that ENDS aerosol exposure led to apoptosis, secondary necrosis, and necrosis in lung epithelial cell models. In contrast, macrophages died mostly by apoptosis and inflammatory caspase-mediated cell death when exposed to ENDS aerosol. The clearance of dead cells and pathogens by efferocytosis and phagocytosis, respectively, is an important process in maintaining a healthy lung. To investigate the impact of ENDS aerosol on macrophage function independent of general toxicity, we used an exposure time that did not induce cell death in primary macrophages. Exposure to ENDS aerosol containing nicotine inhibited nearly all phagocytic and greatly reduced the efferocytic abilities of primary macrophages. When challenged with a bacterial pathogen, there was decreased bacterial clearance. The presence of nicotine in the ENDS aerosol increased its toxicity and functional impact; however, nicotine exposure alone did not have any deleterious effects. These data demonstrate that ENDS aerosol exposure could lead to increased epithelial cell and macrophage death in the lung and impair important macrophage functions that are essential for maintenance of lung function.

In Vitro Toxicity and Chemical Characterization of Aerosol Derived from Electronic Cigarette Humectants Using a Newly Developed Exposure System.

In the United States, the recent surge of electronic cigarette (e-cig) use has raised questions concerning the safety of these devices. This study seeks to assess the pro-inflammatory and cellular stress effects of the vaped humectants propylene glycol (PG) and glycerol (GLY) on airway epithelial cells (16HBE cells and differentiated human bronchial epithelial cells) with a newly developed aerosol exposure system. This system allows for chemical characterization of e-cig generated aerosol particles as well as in vitro exposures of 16HBE cells at an air-liquid interface to vaped PG and GLY aerosol. Our data demonstrate that the process of vaping results in the formation of PG- and GLY-derived oligomers in the aerosol particles. Our in vitro data demonstrate an increase in pro-inflammatory cytokines IL-6 and IL-8 levels in response to vaped PG and GLY exposures. Vaped GLY also causes an increase in cellular stress signals HMOX1, NQO1, and carbonylated proteins when the e-cig device is operated at high wattages. Additionally, we find that the exposure of vaped PG causes elevated IL-6 expression, while the exposure of vaped GLY increases HMOX1 expression in human bronchial epithelial cells when the device is operated at high wattages. These findings suggest that vaporizing PG and GLY results in the formation of novel compounds and the exposure of vaped PG and GLY are detrimental to airway cells. Since PG and/or GLY is universally contained in all e-cig liquids, we conclude that these components alone can cause harm to the airway epithelium.

Development of inhalable quinacrine loaded bovine serum albumin modified cationic nanoparticles: Repurposing quinacrine for lung cancer therapeutics.

Drug repurposing is on the rise as an atypical strategy for discovery of new molecules, involving use of pre-existing molecules for a different therapeutic application than the approved indication. Using this strategy, the current study aims to leverage effects of quinacrine (QA), a well-known anti-malarial drug, for treatment of non-small cell lung cancer (NSCLC). For respiratory diseases, designing a QA loaded inhalable delivery system has multiple advantages over invasive delivery. QA-loaded nanoparticles (NPs) were thus prepared using polyethyleneimine (PEI) as a cationic stabilizer. While the use of PEI provided cationic charge on the particles, it also mediated a burst release of QA and demonstrated potential particle toxicity. These concerns were circumvented by coating nanoparticles with bovine serum albumin (BSA), which retained the cationic charge, reduced NP toxicity and modulated QA release. Prepared nanoparticles were characterized for physicochemical properties along with their aerosolization potential. Therapeutic efficacy of the formulations was tested in different NSCLC cells. Mechanism of higher anti-proliferation was evaluated by studying cell cycle profile, apoptosis and molecular markers involved in the progression of lung cancer. BSA coated QA nanoparticles demonstrated good aerosolization potential with a mass median aerodynamic diameter of significantly less than 5 µm. Nanoparticles also demonstrated improved therapeutic efficacy against NSCLC cells in terms of low IC50 values, cell cycle arrest at G2/M phase and autophagy inhibition leading to increased apoptosis. BSA coated QA NPs also demonstrated enhanced therapeutic efficacy in a 3D cell culture model. The present study thus lays solid groundwork for pre-clinical and eventual clinical studies as a standalone therapy and in combination with existing chemotherapeutics.

Isoprene-Derived Secondary Organic Aerosol Induces the Expression of MicroRNAs Associated with Inflammatory/Oxidative Stress Response in Lung Cells.

Exposure to fine particulate matter (PM2.5), of which secondary organic aerosol (SOA) is a major constituent, is linked to adverse health outcomes, including cardiovascular disease, lung cancer, and preterm birth. Atmospheric oxidation of isoprene, the most abundant nonmethane hydrocarbon emitted into Earth's atmosphere primarily from vegetation, contributes to SOA formation. Isoprene-derived SOA has previously been found to alter inflammatory/oxidative stress genes. MicroRNAs (miRNAs) are epigenetic regulators that serve as post-transcriptional modifiers and key mediators of gene expression. To assess whether isoprene-derived SOA alters miRNA expression, BEAS-2B lung cells were exposed to laboratory-generated isoprene-derived SOA constituents derived from the acid-driven multiphase chemistry of authentic methacrylic acid epoxide (MAE) or isomeric isoprene epoxydiols (IEPOX) with acidic sulfate aerosol particles. These IEPOX- and MAE-derived SOA constituents have been shown to be measured in large quantities within PM2.5 collected from isoprene-rich areas affected by acidic sulfate aerosol particles derived from human activities. A total of 29 miRNAs were identified as differentially expressed when exposed to IEPOX-derived SOA and 2 when exposed to MAE-derived SOA, a number of which are inflammatory/oxidative stress associated. These results suggest that miRNAs may modulate the inflammatory/oxidative stress response to SOA exposure, thereby advancing the understanding of airway cell epigenetic response to SOA.