Cellular stress responses and dysfunctional Mitochondrial–cellular senescence, and therapeutics in chronic respiratory diseases

The abnormal inflammatory responses due to the lung tissue damage and ineffective repair/resolution in response to the inhaled toxicants result in the pathological changes associated with chronic respiratory diseases. Investigation of such pathophysiological mechanisms provides the opportunity to develop the molecular phenotype-specific diagnostic assays and could help in designing the personalized medicine-based therapeutic approaches against these prevalent diseases. As the central hubs of cell metabolism and energetics, mitochondria integrate cellular responses and interorganellar signaling pathways to maintain cellular and extracellular redox status and the cellular senescence that dictate the lung tissue responses. Specifically, as observed in chronic obstructive pulmonary disease (COPD) and pulmonary fibrosis, the mitochondria-endoplasmic reticulum (ER) crosstalk is disrupted by the inhaled toxicants such as the combustible and emerging electronic nicotine-delivery system (ENDS) tobacco products. Thus, the recent research efforts have focused on understanding how the mitochondria-ER dysfunctions and oxidative stress responses can be targeted to improve inflammatory and cellular dysfunctions associated with these pathologic illnesses that are exacerbated by viral infections. The present review assesses the importance of these redox signaling and cellular senescence pathways that describe the role of mitochondria and ER on the development and function of lung epithelial responses, highlighting the cause and effect associations that reflect the disease pathogenesis and possible intervention strategies

A Review of the Toxicity of Ingredients in E-Cigarettes, Including Those Ingredients Having the Fda\u27s generally Recognized As Safe (GRAS) Regulatory Status for Use in Food.

Some firms and marketers of electronic cigarettes (e-cigarettes; a type of electronic nicotine delivery system (ENDS)) and refill liquids (e-liquids) have made claims about the safety of ingredients used in their products based on the term GRAS or Generally Recognized As Safe (GRAS). However, GRAS is a provision within the definition of a food additive under section 201(s) (21 U.S.C. 321(s)) of the U.S. Federal Food Drug and Cosmetic Act (FD&C Act). Food additives and GRAS substances are by the FD&C Act definition intended for use in food, thus safety is based on oral consumption; the term GRAS cannot serve as an indicator of the toxicity of e-cigarette ingredients when aerosolized and inhaled (i.e., vaped). There is no legal or scientific support for labeling e-cigarette product ingredients as GRAS . This review discusses our concerns with the GRAS provision being applied to e-cigarette products and provides examples of chemical compounds that have been used as food ingredients but have been shown to lead to adverse health effects when inhaled. The review provides scientific insight into the toxicological evaluation of e-liquid ingredients and their aerosols to help determine the potential respiratory risks associated with their use in e-cigarettes

Testing of an oral dosing technique for double-crested cormorants, \u3ci\u3ePhalacocorax auritus\u3c/i\u3e, laughing gulls, \u3ci\u3eLeucophaeus atricilla\u3c/i\u3e, homing pigeons, \u3ci\u3eColumba livia\u3c/i\u3e, and western sandpipers, \u3ci\u3eCalidris mauri\u3c/i\u3e, with artificially weather MC252 oil

Scoping studies were designed to determine if double-crested cormorants (Phalacocorax auritus), laughing gulls (Leucophaues atricilla), homing pigeons (Columba livia) and western sandpipers (Calidris mauri) that were gavaged with a mixture of artificially weathered MC252 oil and food for either a single day or 4–5 consecutive days showed signs of oil toxicity. Where volume allowed, samples were collected for hematology, plasma protein electrophoresis, clinical chemistry and electrolytes, oxidative stress and organ weigh changes. Double-crested cormorants, laughing gulls and western sandpipers all excreted oil within 30 min of dose, while pigeons regurgitated within less than one hour of dosing. There were species differences in the effectiveness of the dosing technique, with double-crested cormorants having the greatest number of responsive endpoints at the completion of the trial. Statistically significant changes in packed cell volume, white cell counts, alkaline phosphatase, alanine aminotransferase, creatine phosphokinase, gamma glutamyl transferase, uric acid, chloride, sodium, potassium, calcium, total glutathione, glutathione disulfide, reduced glutathione, spleen and liver weights were measured in double-crested cormorants. Homing pigeons had statistically significant changes in creatine phosphokinase, total glutathione, glutathione disulfide, reduced glutathione and Trolox equivalents. Laughing gulls exhibited statistically significant decreases in spleen and kidney weight, and no changes were observed in any measurement endpoints tested in western sandpipers

Effects of commonly used air filters on secondhand tobacco smoke and the induction of oxidative stress and inflammation in mice.

The annual number of deaths related to secondhand smoke (SHS) exposure exceeds 40,000 in the United States. Exposure to SHS primarily occurs in homes and work places. Public places of exposure to SHS include restaurants, bars and casinos due to lack of smoke-free laws. Approximately 7000 chemicals have been identified as constituents of SHS, of which 250 are known to be toxic and 70 as carcinogens. Exposure to these harmful chemicals can result in cardiovascular diseases and lung cancer. The critics of smoking bans have proposed air filtration systems as a viable solution to eliminate the risk of exposure to SHS. In this project, one of the primary objectives was to assess the effects of commonly used air filters, MERV 4 fiberglass, MERV 8 pleated and MERV 8 pleated activated charcoal on SHS and its biological effects. SHS was generated using a TE-10 smoke machine system, and the total suspended particulates, the smaller respirable particles and the carbon monoxide concentrations were measured in filtered smoke. Our results showed that these filters failed to remove these components of SHS. We also examined the physiological responses caused by exposure to SHS. Using murine models, the oxidative stress and the inflammatory responses caused by SHS exposure were assessed. In order to assess the oxidative stress response caused by the free radicals in SHS, superoxide dismutase (SOD), glutathione peroxidase (GPx), total glutathione (GSH) and lipid peroxidation products (MDA+HAE) in the lung homogenates were quantified. GSH and MDA+HAE in the lung tissues were chosen as the appropriate biomarkers to assess the oxidative stress by SHS. For the assessment of the inflammatory response, interleukin 1 beta (IL-1), interleukin 6 (IL-6), granulocyte macrophage colony stimulating factor (GM-CSF) and tumor necrosis factor alpha (TNF-) cytokines were quantified in the bronchoalveolarlavage (BAL) fluid. TNF-α in the BAL fluid was chosen the appropriate biomarker to assess the inflammatory response due to SHS. Exposure to SHS caused significantly upregulated oxidative stress response and a suppressed cytokine response. The attenuated cytokine response displayed characteristics of endotoxin tolerance/cross tolerance. The tested air filters were unable to remove the oxidative stress response in the lung homogenates. The tested filters also failed to protect against the suppression of cytokine response due to SHS. Our results showed that exposure to SHS causes a complex oxidative stress and an inflammatory response and that these commonly used air filters failed to protect against the SHS exposure

Challenges in current inhalable tobacco toxicity assessment models: A narrative review

Emerging tobacco products such as electronic nicotine delivery systems (ENDS) and heated tobacco products (HTPs) have a dynamic landscape and are becoming widely popular as they claim to offer a low-risk alternative to conventional smoking. Most pre-clinical laboratories currently exploit in vitro , ex vivo , and in vivo experimental models to assess toxicological outcomes as well as to develop risk-estimation models. While most laboratories have produced a wide range of cell culture and mouse model data utilizing current smoke/aerosol generators and standardized puffing profiles, much variation still exists between research studies, hindering the generation of usable data appropriate for the standardization of these tobacco products. In this review, we discuss current state-of-the-art in vitro and in vivo models and their challenges, as well as insights into risk estimation of novel products and recommendations for toxicological parameters for reporting, allowing comparability of the research studies between laboratories, resulting in usable data for regulation of these products before approval by regulatory authorities

Noninvasive systemic biomarkers of e-cigarette or vaping use-associated lung injury: a pilot study

Background Electronic cigarette (e-cigarette) vaping, containing nicotine and/or Δ8, Δ9 or Δ10 or Δo tetrahydrocannabinol (Δn-THC), is associated with an outbreak of e-cigarette, or vaping, product use-associated lung injury (EVALI). Despite thousands being hospitalised with EVALI, much remains unknown about diagnosis, treatment and disease pathogenesis. Biomarkers of inflammation, oxidative stress and lipid mediators may help identify e-cigarette users with EVALI. Methods We collected plasma and urine along with demographic and vaping-related data of EVALI subjects (age 18–35 years) and non-users matched for sex and age in a pilot study. Biomarkers were assessed by ELISA/EIA and Luminex-based assays. Results Elevated levels of THC metabolite (11-nor-9-carboxy-Δ9-THC) were found in plasma from EVALI subjects compared to non-users. Levels of 8-hydroxy-2′-deoxyguanosine (8-OHdG), an oxidative DNA damage biomarker, and 8-isoprostane, an oxidative stress marker, were slightly increased in urine samples from EVALI subjects compared to non-users. Conversely, plasma levels of lipid mediators, including resolvin D1 (RvD1) and prostaglandin E2 (PGE2), were significantly lower in EVALI subjects compared to non-users. Both pro-inflammatory biomarkers, such as tumour necrosis factor-α, macrophage inflammatory protein-1β, RANTES (regulated on activation, normal T-cell expressed and secreted) and granulocyte–macrophage colony-stimulating factor, as well as anti-inflammatory biomarkers, such as interleukin-9 and CC10/16, were decreased in plasma from EVALI subjects compared to non-users, supportive of a possible dysregulated inflammatory response in EVALI subjects. Conclusions Significant elevations in urine and plasma biomarkers of oxidative stress, as well as reductions in lipid mediators, were shown in EVALI subjects. These noninvasive biomarkers (8-OHdG, 8-isoprostane, RvD1 and CC10/16), either individually or collectively, may serve as tools in diagnosing future EVALI subjects

Nose-Only Exposure to Cherry- and Tobacco-Flavored E-Cigarettes Induced Lung Inflammation in Mice in a Sex-Dependent Manner

Flavoring chemicals in electronic nicotine delivery systems have been shown to cause cellular inflammation; meanwhile, the effects of fruit and tobacco flavors on lung inflammation by nose-only exposures to mice are relatively unknown. We hypothesized that exposure to flavored e-cigarettes would cause lung inflammation in C57BL/6 J mice. The mice were exposed to air, propylene glycol/vegetable glycerin, and flavored e-liquids: Apple, Cherry, Strawberry, Wintergreen, and Smooth & Mild Tobacco, one hour per day for three days. Quantification of flavoring chemicals by proton nuclear magnetic resonance spectroscopy (1H NMR), differential cell counts by flow cytometry, pro-inflammatory cytokines/chemokines by ELISA, and matrix metalloproteinase levels by western blot were performed. Exposure to PG/VG increased neutrophil cell count in lung bronchoalveolar lavage fluid (BALF). KC and IL6 levels were increased by PG/VG exposure and female mice exposed to Cherry flavored e-cigarettes, in lung homogenate. Mice exposed to PG/VG, Apple, Cherry, and Wintergreen increased MMP2 levels. Our results revealed flavor- and sex-based e-cigarette effects in female mice exposed to cherry-flavored e-liquids and male mice exposed to tobacco-flavored e-liquids, namely, increased lung inflammation

Waterpipe smoke and e-cigarette vapor differentially affect circadian molecular clock gene expression in mouse lungs.

The use of emerging tobacco products, such as waterpipe or hookah and electronic cigarettes (e-cigs), has gained significant popularity and are promoted as safer alternatives to conventional cigarettes. Circadian systems are internal biological oscillations that are considered important regulators of immune functions in mammals. Tobacco induced inflammatory lung diseases frequently exhibit time-of-day/night variation in lung function and symptom severity. We investigated the impact of inhaled e-cig vapor and waterpipe smoke (WPS) on pulmonary circadian molecular clock disruption by determining the changes in expression levels and abundance of core clock component genes (BMAL1, CLOCK) and clock-controlled output genes (Rev-erbα, Per2, Rev-erbβ, Cry2, Rorα) in mouse lungs. We showed that the expression levels of these pulmonary core clock genes and clock-controlled output genes were altered significantly following exposure to WPS (Bmal1, Clock, and Rev-erbα). We further showed a significant yet differential effect on expression levels of core clock and clock-controlled genes (Bmal1, Per2) in the lungs of mice exposed to e-cig vapor containing nicotine. Thus, acute exposure to WPS and e-cig vapor containing nicotine contributes to altered expression of circadian molecular clock genes in mouse lungs, which may have repercussions on lung cellular and biological functions

Inflammatory and Oxidative Responses Induced by Exposure to Commonly Used e-Cigarette Flavoring Chemicals and Flavored e-Liquids without Nicotine

Background: The respiratory health effects of inhalation exposure to e-cigarette flavoring chemicals are not well understood. We focused our study on the immuno-toxicological and the oxidative stress effects by these e-cigarette flavoring chemicals on two types of human monocytic cell lines, Mono Mac 6 (MM6) and U937. The potential to cause oxidative stress by these flavoring chemicals was assessed by measuring the production of reactive oxygen species (ROS). We hypothesized that the flavoring chemicals used in e-juices/e-liquids induce an inflammatory response, cellular toxicity, and ROS production.Methods: Two monocytic cell types, MM6 and U937 were exposed to commonly used e-cigarette flavoring chemicals; diacetyl, cinnamaldehyde, acetoin, pentanedione, o-vanillin, maltol and coumarin at different doses between 10 and 1,000 μM. Cell viability and the concentrations of the secreted inflammatory cytokine interleukin 8 (IL-8) were measured in the conditioned media. Cell-free ROS produced by these commonly used flavoring chemicals were also measured using a 2′,7′dichlorofluorescein diacetate probe. These DCF fluorescence data were expressed as hydrogen peroxide (H2O2) equivalents. Cytotoxicity due to the exposure to selected e-liquids was assessed by cell viability and the IL-8 inflammatory cytokine response in the conditioned media.Results: Treatment of the cells with flavoring chemicals and flavored e-liquid without nicotine caused cytotoxicity dose-dependently. The exposed monocytic cells secreted interleukin 8 (IL-8) chemokine in a dose-dependent manner compared to the unexposed cell groups depicting a biologically significant inflammatory response. The measurement of cell-free ROS by the flavoring chemicals and e-liquids showed significantly increased levels of H2O2 equivalents in a dose-dependent manner compared to the control reagents. Mixing a variety of flavors resulted in greater cytotoxicity and cell-free ROS levels compared to the treatments with individual flavors, suggesting that mixing of multiple flavors of e-liquids are more harmful to the users.Conclusions: Our data suggest that the flavorings used in e-juices can trigger an inflammatory response in monocytes, mediated by ROS production, providing insights into potential pulmonary toxicity and tissue damage in e-cigarette users

Classification of flavors in cigarillos and little cigars and their variable cellular and acellular oxidative and cytotoxic responses.

Flavored tobacco products are increasing in popularity but remain unregulated, with the exception of the ban on flavored conventional cigarettes. Lack of regulation of cigarillos and little cigars allows vendors to have their own version of popular flavors, each with different chemical components. A new classification system was created for flavored cigars in order to easily communicate our results with the scientific community. To understand the physicochemical characteristics of flavored little cigars and cigarillo smoke, size distribution and concentration of particulate matter in smoke were determined. Acellular reactive oxygen species generation was measured as an indirect measurement of the potential to cause oxidative stress in cells. In addition, cigarillo smoke extract treatment on bronchial epithelial (Beas-2b) cells were assessed to determine the flavor-induced cellular toxicity. Flavored cigars/cigarillos showed significant variability in the tested parameters between flavors as well as brands of the same flavor, but most of the cigars showed higher potential of generating oxidative stress, than research grade cigarettes. Flavored cigars produced maximum particle concentrations at 1.0μm and 4.0 μm compared with 3R4F reference research cigarettes. A differential cytotoxic response was observed with cigarillo smoke extract treatments, with "fruits/candy" and "drinks" being the most toxic, but were not more cytotoxic than smoke from cigarettes. These cigarillos with flavors must be well characterized for toxicity in order to prevent adverse effects caused by exposure to flavor chemicals. Our study provides insight into understanding the potential health effects of flavor-infused cigars/cigarillos and the need for the regulation of those flavoring chemicals in these products. Future research is directed to determine the flavoring toxicity of little cigars and cigarillos in vivo studies