Corrigendum to "The role of synthetic coolants, WS-3 and WS-23, in modulating E-cigarette-induced reactive oxygen species (ROS) in lung epithelial cells" [Toxicol. Rep. 9 (2022) 1700-1709].

[This corrects the article DOI: 10.1016/j.toxrep.2022.08.007.].

From Alpha to Omicron: How Different Variants of Concern of the SARS-Coronavirus-2 Impacted the World.

SARS-CoV-2, the virus that causes COVID-19, is prone to mutations and the generation of genetic variants. Since its first outbreak in 2019, SARS-CoV-2 has continually evolved, resulting in the emergence of several lineages and variants of concern (VOC) that have gained more efficient transmission, severity, and immune evasion properties. The World Health Organization has given these variants names according to the letters of the Greek Alphabet, starting with the Alpha (B.1.1.7) variant, which emerged in 2020, followed by the Beta (B.1.351), Gamma (P.1), Delta (B.1.617.2), and Omicron (B.1.1.529) variants. This review explores the genetic variation among different VOCs of SARS-CoV-2 and how the emergence of variants made a global impact on the pandemic.

Corrigendum to "E-cigarette synthetic cooling agent WS-23 and nicotine aerosols differentially modulate airway epithelial cell responses" [Toxicol. Rep. 9 (2022) 1823-1830].

[This corrects the article DOI: 10.1016/j.toxrep.2022.09.010.].

Indoor-air purification by photoelectrochemical oxidation mitigates allergic airway responses to aerosolized cat dander in a murine model.

Portable air purifiers help improve indoor air quality by neutralizing allergens, including animal dander proteins. However, there are limited in-vivo models to assess the efficacy of these devices. Here, we developed a novel animal model of experimental asthma using aerosolized cat dander extract (CDE) exposure and compared the efficacy of select air purification technologies. Mice were exposed to CDE aerosols for 6 weeks in separate custom-built whole-body exposure chambers equipped with either a photoelectrochemical oxidative (PECO) Molekule filtration device (PFD) or a HEPA-assisted air filtration device (HFD) along with positive (a device with no filtration capability) and negative controls. Compared to the positive control group, the CDE-induced airway resistance, and plasma IgE and IL-13 levels were significantly reduced in both air purifier groups. However, PFD mice showed a better attenuation of lung tissue mucous hyperplasia and eosinophilia than HFD and positive control mice, indicating a better efficacy in managing CDE-induced allergic responses. Cat dander protein destruction was evaluated by LCMS proteomic analysis, which revealed the degradation of 2731 unique peptides on PECO media in 1 h. Thus, allergen protein destruction on filtration media enhances air purifier efficacy that could provide relief from allergy responses compared to traditional HEPA-based filtration alone.

E-cigarette synthetic cooling agent WS-23 and nicotine aerosols differentially modulate airway epithelial cell responses.

Electronic cigarette (e-cig) aerosol exposures are strongly associated with pulmonary dysfunctions, and the airway epithelial cells (AECs) of respiratory passages play a pivotal role in understanding this association. However, not much is known about the effect of synthetic cooling agents such as WS-23 on AECs. WS-23 is a synthetic menthol-like cooling agent widely used to enhance the appeal of e-cigs and to suppress the harshness and bitterness of other e-cig constituents. Using primary human AECs, we compared the effects of aerosolized WS-23 with propylene glycol/vegetable glycerin (PG/VG) vehicle control and nicotine aerosol exposures. AECs treated with 3 % WS-23 aerosols showed a significant increase in viable cell numbers compared to PG/VG-vehicle aerosol exposed cells and cell growth was comparable following 2.5 % nicotine aerosol exposure. AEC inflammatory factors, IL-6 and ICAM-1 levels were significantly suppressed by WS-23 aerosols compared to PG/VG-controls. When differentiated AECs were challenged with WS-23 aerosols, there was a significant increase in secretory mucin MUC5AC expression with no discernible change in airway inflammatory SCGB1A1 expression. Compared to PG/VG-controls, WS-23 or nicotine aerosols presented with increased MUC5AC expression, but there was no synergistic effect of WS-23 + nicotine combination exposure. Thus, WS-23 and nicotine aerosols modulate the AEC responses and induce goblet cell hyperplasia, which could impact the airway physiology and susceptibility to respiratory diseases.

The role of synthetic coolants, WS-3 and WS-23, in modulating E-cigarette-induced reactive oxygen species (ROS) in lung epithelial cells.

There has been a substantial rise in e-cigarette (e-cig) use or vaping in the past decade, prompting growing concerns about their adverse health effects. Recently, e-cig manufacturers have been using synthetic cooling agents, like WS-23 and WS-3, to provide a cooling sensation without the "menthol taste". Studies have shown that aerosols/vapes generated by e-cigs can contain significant levels of reactive oxygen species (ROS). However, studies investigating the role of synthetic coolants in modulating ROS levels generated by e-cigs are lacking. This study seeks to understand how synthetic coolants, e-cig additives that have become increasingly prevalent in e-liquids sold in the United States (US), impact acellular ROS production from e-liquid aerosols as well as cellular ROS levels from pulmonary epithelial cells exposed to these e-liquids. To further explain, our study aims to understand whether the addition of WS-3 and WS-23 to e-liquid base and e-liquid base with nicotine significantly modifies generated acellular ROS levels within aerosolized e-liquids, as well as cellular ROS within BEAS-2B cells treated with these same e-liquids. Aerosols were generated from e-liquids with and without synthetic coolants through a single-puff aerosol generator; subsequently, acellular ROS was semi-quantified in H2O2 equivalents via fluorescence spectroscopy. Our acellular ROS data suggest that adding WS-3 to e-liquid base (PG:VG), regardless of nicotine content, has a minimal impact on modifying e-cig generated acellular ROS levels. Additionally, we also measured cellular ROS in lung epithelial cells using both e-liquids containing and not containing synthetic coolants via the CellROX Green fluorescent sensor. Similar comparable results were found in BEAS2B cells though ROS was increased by WS-3 and WS-23 treated in e-cig nicotine groups. Altogether, our data suggest that neither the addition of WS-23 nor WS-3 to e-liquid base solution, with and without nicotine, significantly modifies e-cig generated acellular ROS levels within aerosolized e-liquids and cellular ROS levels within treated BEAS-2B cells. Together, our data provide insight into whether synthetic coolants added to e-liquids could impact vaping-induced oxidative stress in the lungs.

Health effects and known pathology associated with the use of E-cigarettes.

In recent years, new nicotine delivery methods have emerged, and many users are choosing electronic cigarettes (e-cigarettes) over traditional tobacco cigarettes. E-cigarette use is very popular among adolescents, with more than 3.5 million currently using these products in the US. Despite the increased prevalence of e-cigarette use, there is limited knowledge regarding the health impact of e-cigarettes on the general population. Based on published findings by others, E-cigarette is associated with lung injury outbreak, which increased health and safety concerns related to consuming this product. Different components of e-cigarettes, including food-safe liquid solvents and flavorings, can cause health issues related to pneumonia, pulmonary injury, and bronchiolitis. In addition, e-cigarettes contain alarmingly high levels of carcinogens and toxicants that may have long-lasting effects on other organ systems, including the development of neurological manifestations, lung cancer, cardiovascular disorders, and tooth decay. Despite the well- documented potential for harm, e-cigarettes do not appear to increase susceptibility to SARS-CoV- 2 infection. Furthermore, some studies have found that e-cigarette users experience improvements in lung health and minimal adverse effects. Therefore, more studies are needed to provide a definitive conclusion on the long-term safety of e-cigarettes. The purpose of this review is to inform the readers about the possible health-risks associated with the use of e-cigarettes, especially among the group of young and young-adults, from a molecular biology point of view.

Corrigendum: Increased expression of LASI LncRNA regulates the cigarette smoke and COPD associated airway inflammation and mucous cell hyperplasia.

[This corrects the article DOI: 10.3389/fimmu.2022.803362.].

Immunomodulatory LncRNA on antisense strand of ICAM-1 augments SARS-CoV-2 infection-associated airway mucoinflammatory phenotype.

Noncoding RNAs are important regulators of mucoinflammatory response, but little is known about the contribution of airway long noncoding RNAs (lncRNAs) in COVID-19. RNA-seq analysis showed a more than 4-fold increased expression of IL-6, ICAM-1, CXCL-8, and SCGB1A1 inflammatory factors; MUC5AC and MUC5B mucins; and SPDEF, FOXA3, and FOXJ1 transcription factors in COVID-19 patient nasal samples compared with uninfected controls. A lncRNA on antisense strand to ICAM-1 or LASI was induced 2-fold in COVID-19 patients, and its expression was directly correlated with viral loads. A SARS-CoV-2-infected 3D-airway model largely recapitulated these clinical findings. RNA microscopy and molecular modeling indicated a possible interaction between viral RNA and LASI lncRNA. Notably, blocking LASI lncRNA reduced the SARS-CoV-2 replication and suppressed MUC5AC mucin levels and associated inflammation, and select LASI-dependent miRNAs (e.g., let-7b-5p and miR-200a-5p) were implicated. Thus, LASI lncRNA represents an essential facilitator of SARS-CoV-2 infection and associated airway mucoinflammatory response.

Increased Expression of LASI lncRNA Regulates the Cigarette Smoke and COPD Associated Airway Inflammation and Mucous Cell Hyperplasia.

Research Impact: Cigarette smoke (CS) exposure is strongly associated with chronic obstructive pulmonary disease (COPD). In respiratory airways, CS exposure disrupts airway barrier functions, mucous/phlegm production, and basic immune responses of airway epithelial cells. Based on our recent identification of a specific immunomodulatory long noncoding RNA (lncRNA), we investigated its role in CS-induced responses in bronchial airways of cynomolgus macaque model of CS-induced COPD and in former smokers with and without COPD. The lncRNA was significantly upregulated in CS-induced macaque airways and in COPD airways that exhibited higher mucus expression and goblet cell hyperplasia. Experimental models of cells derived from COPD subjects recapitulated the augmented inflammation and mucus expression following the smoke challenge. Blocking of lncRNA expression in cell culture setting suppressed the smoke-induced and COPD-associated dysregulated mucoinflammatory response suggesting that this airway specific immunomodulatory lncRNA may represent a novel target to mitigate the smoke-mediated inflammation and mucus hyperexpression. Rationale: In conducting airways, CS disrupts airway epithelial functions, mucociliary clearances, and innate immune responses that are primarily orchestrated by human bronchial epithelial cells (HBECs). Mucus hypersecretion and dysregulated immune response are the hallmarks of chronic bronchitis (CB) that is often exacerbated by CS. Notably, we recently identified a long noncoding RNA (lncRNA) antisense to ICAM-1 (LASI) that mediates airway epithelial responses. Objective: To investigate the role of LASI lncRNA in CS-induced airway inflammation and mucin hyperexpression in an animal model of COPD, and in HBECs and lung tissues from former smokers with and without COPD. To interrogate LASI lncRNA role in CS-mediated airway mucoinflammatory responses by targeted gene editing. Methods: Small airway tissue sections from cynomolgus macaques exposed to long-term mainstream CS, and those from former smokers with and without COPD were analyzed. The structured-illumination imaging, RNA fluorescence in-situ hybridization (FISH), and qRT-PCR were used to characterize lncRNA expression and the expression of inflammatory factors and airway mucins in a cell culture model of CS extract (CSE) exposure using HBECs from COPD (CHBEs) in comparison with cells from normal control (NHBEs) subjects. The protein levels of mucin MUC5AC, and inflammatory factors ICAM-1, and IL-6 were determined using specific ELISAs. RNA silencing was used to block LASI lncRNA expression and lentivirus encoding LASI lncRNA was used to achieve LASI overexpression (LASI-OE). Results: Compared to controls, LASI lncRNA was upregulated in CS-exposed macaques and in COPD smoker airways, correlating with mucus hyperexpression and mucus cell hyperplasia in severe COPD airways. At baseline, the unstimulated CHBEs showed increased LASI lncRNA expression with higher expression of secretory mucin MUC5AC, and inflammatory factors, ICAM-1, and IL-6 compared to NHBEs. CSE exposure of CHBEs resulted in augmented inflammation and mucus expression compared to controls. While RNA silencing-mediated LASI knockdown suppressed the mucoinflammatory response, cells overexpressing LASI lncRNA showed elevated mRNA levels of inflammatory factors. Conclusions: Altogether, LASI lncRNA may represent a novel target to control the smoke-mediated dysregulation in airway responses and COPD exacerbations.

Distinct Mucoinflammatory Phenotype and the Immunomodulatory Long Noncoding Transcripts Associated with SARS-CoV-2 Airway Infection.

Respiratory epithelial cells are the primary target for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We investigated the 3D human airway tissue model to evaluate innate epithelial cell responses to SARS-CoV-2 infection. A SARS-CoV-2 clinical isolate productively infected the 3D-airway model with a time-dependent increase in viral load (VL) and concurrent upregulation of airway immunomodulatory factors ( IL-6, ICAM-1 , and SCGB1A1 ) and respiratory mucins ( MUC5AC, MUC5B, MUC2 , and MUC4) , and differential modulation of select long noncoding RNAs (lncRNAs i.e., LASI, TOSL, NEAT1 , and MALAT1 ). Next, we examined these immunomodulators in the COVID-19 patient nasopharyngeal swab samples collected from subjects with high- or low-VLs (∻100-fold difference). As compared to low-VL, high-VL patients had prominent mucoinflammatory signature with elevated expression of IL-6, ICAM-1, SCGB1A1, SPDEF, MUC5AC, MUC5B , and MUC4 . Interestingly, LASI, TOSL , and NEAT1 lncRNA expressions were also markedly elevated in high-VL patients with no change in MALAT1 expression. In addition, dual-staining of LASI and SARS-CoV-2 nucleocapsid N1 RNA showed predominantly nuclear/perinuclear localization at 24 hpi in 3D-airway model as well as in high-VL COVID-19 patient nasopharyngeal cells, which exhibited high MUC5AC immunopositivity. Collectively, these findings suggest SARS-CoV-2 induced lncRNAs may play a role in acute mucoinflammatory response observed in symptomatic COVID-19 patients.

A long noncoding RNA antisense to ICAM-1 is involved in allergic asthma associated hyperreactive response of airway epithelial cells.

Epithelial cells of the conducting airways are a pivotal first line of defense against airborne pathogens and allergens that orchestrate inflammatory responses and mucociliary clearance. Nonetheless, the molecular mechanisms responsible for epithelial hyperreactivity associated with allergic asthma are not completely understood. Transcriptomic analysis of human airway epithelial cells (HAECs), differentiated in-vitro at air-liquid interface (ALI), showed 725 differentially expressed immediate-early transcripts, including putative long noncoding RNAs (lncRNAs). A novel lncRNA on the antisense strand of ICAM-1 or LASI was identified, which was induced in LPS-primed HAECs along with mucin MUC5AC and its transcriptional regulator SPDEF. LPS-primed expression of LASI, MUC5AC, and SPDEF transcripts were higher in ex-vivo cultured asthmatic HAECs that were further augmented by LPS treatment. Airway sections from asthmatics with increased mucus load showed higher LASI expression in MUC5AC+ goblet cells following multi-fluorescent in-situ hybridization and immunostaining. LPS- or IL-13-induced LASI transcripts were mostly enriched in the nuclear/perinuclear region and were associated with increased ICAM-1, IL-6, and CXCL-8 expression. Blocking LASI expression reduced the LPS or IL-13-induced epithelial inflammatory factors and MUC5AC expression, suggesting that the novel lncRNA LASI could play a key role in LPS-primed trained airway epithelial responses that are dysregulated in allergic asthma.

The Impact of CRISPR-Cas9 on Age-related Disorders: From Pathology to Therapy.

With advances in medical technology, the number of people over the age of 60 is on the rise, and thus, increasing the prevalence of age-related pathologies within the aging population. Neurodegenerative disorders, cancers, metabolic and inflammatory diseases are some of the most prevalent age-related pathologies affecting the growing population. It is imperative that a new treatment to combat these pathologies be developed. Although, still in its infancy, the CRISPR-Cas9 system has become a potent gene-editing tool capable of correcting gene-mediated age-related pathology, and therefore ameliorating or eliminating disease symptoms. Deleting target genes using the CRISPR-Cas9 system or correcting for gene mutations may ameliorate many different neurodegenerative disorders detected in the aging population. Cancer cells targeted by the CRISPR-Cas9 system may result in an increased sensitivity to chemotherapeutics, lower proliferation, and higher cancer cell death. Finally, reducing gene targeting inflammatory molecules production through microRNA knockout holds promise as a therapeutic strategy for both arthritis and inflammation. Here we present a review based on how the expanding world of genome editing can be applied to disorders and diseases affecting the aging population.

Gestational Exposure to Cigarette Smoke Suppresses the Gasotransmitter H2S Biogenesis and the Effects Are Transmitted Transgenerationally.

Rationale: Gestational cigarette smoke (CS) impairs lung angiogenesis and alveolarization, promoting transgenerational development of asthma and bronchopulmonary dysplasia (BPD). Hydrogen sulfide (H2S), a proangiogenic, pro-alveolarization, and anti-asthmatic gasotransmitter is synthesized by cystathionine-γ-lyase (CSE), cystathionine-ß-synthase (CBS), and 3-mercaptopyruvate sulfur transferase (3MST). Objective: Determine if gestational CS exposure affected the expression of H2S synthesizing enzymes in the mouse lung and human placenta. Methods: Mice were exposed throughout gestational period to secondhand CS (SS) at approximating the dose of CS received by a pregnant woman sitting in a smoking bar for 3 h/days during pregnancy. Lungs from 7-days old control and SS-exposed pups and human placenta from mothers who were either non-smokers or smokers during pregnancy were analyzed for expression of the enzymes. Measurements: Mouse lungs and human placentas were examined for the expression of CSE, CBS, and 3MST by immunohistochemical staining, qRT-PCR and/or Western blot (WB) analyses. Results: Compared to controls, mouse lung exposed gestationally to SS had significantly lower levels of CSE, CBS, and 3MST. Moreover, the SS-induced suppression of CSE and CBS in F1 lungs was transmitted to the F2 generation without significant change in the magnitude of the suppression. These changes were associated with impaired epithelial-mesenchymal transition (EMT)-a process required for normal lung angiogenesis and alveolarization. Additionally, the placentas from mothers who smoked during pregnancy, expressed significantly lower levels of CSE, CBS, and 3MST, and the effects were partially moderated by quitting smoking during the first trimester. Conclusions: Lung H2S synthesizing enzymes are downregulated by gestational CS and the effects are transmitted to F2 progeny. Smoking during pregnancy decreases H2S synthesizing enzymes is human placentas, which may correlate with the increased risk of asthma/BPD in children.

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.

HIV-1 Productively Infects and Integrates in Bronchial Epithelial Cells.

Background: The role of lung epithelial cells in HIV-1-related lung comorbidities remains unclear, and the major hurdle in curing HIV is the persistence of latent HIV reservoirs in people living with HIV (PLWH). The advent of combined antiretroviral therapy has considerably increased the life span; however, the incidence of chronic lung diseases is significantly higher among PLWH. Lung epithelial cells orchestrate the respiratory immune responses and whether these cells are productively infected by HIV-1 is debatable. Methods: Normal human bronchial epithelial cells (NHBEs) grown on air-liquid interface were infected with X4-tropic HIV-1LAV and examined for latency using latency-reversing agents (LRAs). The role of CD4 and CXCR4 HIV coreceptors in NHBEs were tested, and DNA sequencing analysis was used to analyze the genomic integration of HIV proviral genes, Alu-HIVgag-pol, HIV-nef, and HIV-LTR. Lung epithelial sections from HIV-infected humans and SHIV-infected macaques were analyzed by FISH for HIV-gag-pol RNA and epithelial cell-specific immunostaining. Results and Discussion: NHBEs express CD4 and CXCR4 at higher levels than A549 cells. NHBEs are infected with HIV-1 basolaterally, but not apically, by X4-tropic HIV-1LAV in a CXCR4/CD4-dependent manner leading to HIV-p24 antigen production; however, NHBEs are induced to express CCR5 by IL-13 treatment. In the presence of cART, HIV-1 induces latency and integration of HIV provirus in the cellular DNA, which is rescued by the LRAs (endotoxin/vorinostat). Furthermore, lung epithelial cells from HIV-infected humans and SHIV-infected macaques contain HIV-specific RNA transcripts. Thus, lung epithelial cells are targeted by HIV-1 and could serve as potential HIV reservoirs that may contribute to the respiratory comorbidities in PLWH.

Development and Challenges of Nanotherapeutic Formulations for Targeting Mitochondrial Cell Death Pathways in Lung and Brain Degenerative Diseases.

Mitochondria are among the most dynamic organelles regulating a wide array of cellular processes. They are the cellular hub for oxidative phosphorylation, energy production, and cellular metabolism, and they are important determinants of cell fate, as they control cell death/survival pathways. The mitochondrial network plays a critical role in cellular inflammatory responses, and mitochondria are central in many pathologic conditions such as chronic inflammatory and aging-associated degenerative diseases. Recent advancements in our understanding of the pathogenic pathways and the role of mitochondria therein have identified highly specific therapeutic targets in order to develop personalized nanomedicine approaches for treatment. A wide array of nanoparticle-based formulations has been employed for potential usage in both diagnosing and treating chronic and fatal conditions, with gold nanoparticles and liposomal encapsulation being of particular interest. In this review, we highlight and summarize the advantages and challenges of developing these nanoformulations for targeted and spatiotemporally controlled drug delivery. We discuss the potential of nanotherapy in neoplasms to target the mitochondrial regulated cell death pathways and recent seminal developments in liposomal nanotherapy against chronic inflammatory lung diseases. The need for further development of nanoparticle-based treatment options for neuroinflammatory and neurodegenerative conditions, such as Alzheimer's disease (AD), is also discussed.

Long Noncoding Transcriptome in Chronic Obstructive Pulmonary Disease.

Chronic airway inflammation from recurring exposures to noxious environmental stimuli results in a progressive and irreversible airflow limitation and the lung parenchymal damage that characterizes chronic obstructive pulmonary disease (COPD). The large variability observed in the onset and progression of COPD is primarily driven by complex gene-environment interactions. The transcriptomic and epigenetic memory potential of lung epithelial and innate immune cells drive responses, such as mucus hyperreactivity and airway remodeling, that are tightly regulated by various molecular mechanisms, for which several candidate susceptibility genes have been described. However, the recently described noncoding RNA species, in particular the long noncoding RNAs, may also have an important role in modulating pulmonary responses to chronic inhalation of toxic substances and the development of COPD. This review outlines the features of long noncoding RNAs that have been implicated in regulating the airway inflammatory responses to cigarette smoke exposure and their possible association with COPD pathogenesis. As COPD continues to debilitate the increasingly aging population and contribute to higher morbidity and mortality rates worldwide, the search for better biomarkers and alternative therapeutic options is pivotal.