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1.
Part Fibre Toxicol ; 21(1): 41, 2024 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-39380034

RESUMO

BACKGROUND: Exposure to military burn pit smoke during deployment is associated with different respiratory and non-respiratory diseases. However, information linking smoke exposure to human pulmonary health is lacking. This study examined the effects of simulated burn pit smoke condensates on human airway epithelial cells (HAECs) from twelve donors (smokers/non-smokers, biological female/male) cultured at an air-liquid interface and exposed to condensates from three simulated burn pit waste materials (cardboard, plywood, and plastic) incinerated at two combustion conditions: smoldering and flaming. Cellular gene expression was analyzed using bulk RNA sequencing, and basolateral media cytokine levels were assessed using multiplex immunoassay. RESULTS: Flaming smoke condensates caused more significant differentially expressed genes (DEGs) with plywood flaming smoke being the most potent in altering gene expression and modulating cytokine release. Cardboard and plywood flaming condensates primarily activated detoxification pathways, whereas plastic flaming affected genes related to anti-microbial and inflammatory responses. Correlation analysis between smoke condensate chemicals and gene expression to understand the underlying mechanism revealed crucial role of oxygenated polycyclic aromatic hydrocarbons (PAHs) and aluminum, molybdenum, and silicon elements; IL6 expression was positively correlated with most PAHs. Stratification of data based on HAEC donor demographics suggests that these affect gene expression changes. Enrichment analysis indicated similarity with several deployment-related presumptive and reported diseases, including asthma, emphysema, and cancer of different organs. CONCLUSIONS: This study highlights that simulated burn pit smoke exposure of HAECs causes gene expression changes indicative of deployment-related diseases with more pronounced effects seen in smokers and females. Future studies are needed to further characterize how sex and smoking status affect deployment-related diseases.


Assuntos
Células Epiteliais , Fumaça , Humanos , Fumaça/efeitos adversos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Feminino , Masculino , Citocinas/metabolismo , Incineração , Militares , Células Cultivadas , Adulto , Queima de Resíduos a Céu Aberto
2.
Chem Res Toxicol ; 37(5): 791-803, 2024 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-38652897

RESUMO

Burn pits are a method of open-air waste management that was common during military operations in Iraq, Afghanistan, and other regions in Southwest Asia. Veterans returning from deployment have reported respiratory symptoms, potentially from exposure to burn pit smoke, yet comprehensive assessment of such exposure on pulmonary health is lacking. We have previously shown that exposure to condensates from burn pit smoke emissions causes inflammation and cytotoxicity in mice. In this study, we explored the effects of burn pit smoke condensates on human airway epithelial cells (HAECs) to understand their impact on cellular targets in the human lung. HAECs were cultured at the air-liquid interface (ALI) and exposed to burn pit waste smoke condensates (plywood, cardboard, plastic, mixed, and mixed with diesel) generated under smoldering and flaming conditions. Cytotoxicity was evaluated by measuring transepithelial electrical resistance (TEER) and lactate dehydrogenase (LDH) release; toxicity scores (TSs) were quantified for each exposure. Pro-inflammatory cytokine release and modulation of gene expression were examined for cardboard and plastic condensate exposures. Burn pit smoke condensates generated under flaming conditions affected cell viability, with flaming mixed waste and plywood exhibiting the highest toxicity scores. Cardboard and plastic smoke condensates modulated cytokine secretion, with GM-CSF and IL-1ß altered in more than one exposure group. Gene expression of detoxifying enzymes (ALDH1A3, ALDH3A1, CYP1A1, CYP1B1, NQO1, etc.), mucins (MUC5AC, MUC5B), and cytokines was affected by several smoke condensates. Particularly, expression of IL6 was elevated following exposure to all burn pit smoke condensates, and polycyclic aromatic hydrocarbon acenaphthene was positively associated with the IL-6 level in the basolateral media of HAECs. These observations demonstrate that exposure to smoke condensates of materials present in burn pits adversely affects HAECs and that aberrant cytokine secretion and altered gene expression profiles following burn pit material smoke exposure could contribute to the development of airway disease.


Assuntos
Células Epiteliais , Fumaça , Humanos , Fumaça/efeitos adversos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Células Cultivadas , Sobrevivência Celular/efeitos dos fármacos , Citocinas/metabolismo , Linhagem Celular , Queima de Resíduos a Céu Aberto
3.
Sci Rep ; 14(1): 4029, 2024 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-38369546

RESUMO

Fall armyworm (FAW), Spodoptera frugiperda (J.E. Smith) has significantly affected maize crop yields, production efficiency, and farmers' incomes in the Indian Eastern Gangetic Plains region since it was first observed in India in 2018. A lack of awareness by maize growers of the appropriate selection, method, and timing of insecticide application not only creates a barrier to sustainable FAW control but also contributes to increased environmental pollution, reduced human health and increased production costs. We demonstrated that FAW inflicted the most damage in early whorl growth stage of maize, regardless of whether chemical insecticides were applied. FAW egg masses and larvae collected from maize fields in which no insecticides had been sprayed showed high parasitism rates by parasitoid wasps; in contrast fields that had been sprayed had much lower rates of parasitism on FAW. Ten hymenopteran parasitoids were observed in maize fields across the study region, suggesting a diversity of natural methods to suppress FAW in maize at different growth stages. These included two FAW egg parasitoids and eight FAW larval parasitoids. Microplitis manilae Ashmead was the most abundant FAW larval parasitoid species, and Telenomus cf. remus was the dominant FAW egg parasitoid species. Endemic FAW parasitoids such as those observed in this study have great potential as part of a sustainable, cost-effective agroecological management strategy, which can be integrated with other methods to achieve effective control of FAW.


Assuntos
Inseticidas , Vespas , Animais , Humanos , Spodoptera , Inseticidas/farmacologia , Larva , Zea mays
4.
Am J Respir Crit Care Med ; 209(6): 703-715, 2024 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-37972349

RESUMO

Rationale: Acute respiratory distress syndrome (ARDS) has an unacceptably high mortality rate (35%) and is without effective therapy. Orai1 is a Ca2+ channel involved in store-operated Ca2+ entry (SOCE), a process that exquisitely regulates inflammation. Orai1 is considered a druggable target, but no Orai1-specific inhibitors exist to date. Objectives: To evaluate whether ELD607, a first-in-class Orai1 antagonist, can treat ARDS caused by bacterial pneumonia in preclinical models. Methods: ELD607 pharmacology was evaluated in HEK293T cells and freshly isolated immune cells from patients with ARDS. A murine acute lung injury model caused by bacterial pneumonia was then used: mice were infected with Pseudomonas aeruginosa, Staphylococcus aureus, methicillin-resistant S. aureus, or multidrug-resistant P. aeruginosa and then treated with ELD607 intranasally. Measurements and Main Results: ELD607 specifically inhibited SOCE in HEK293T cells with a half-maximal inhibitory concentration of 9 nM. ELD607 was stable in ARDS airway secretions and inhibited SOCE in ARDS immune cells. In vivo, inhaled ELD607 significantly reduced neutrophilia and improved survival. Surprisingly, Orai1 inhibition by ELD607 caused a significant reduction in lung bacteria, including methicillin-resistant S. aureus. ELD607 worked as an immunomodulator that reduced cytokine levels, reduced neutrophilia, and promoted macrophage-mediated resolution of inflammation and clearance of bacteria. Indeed, when alveolar macrophages were depleted with inhaled clodronate, ELD607 was no longer able to resolve inflammation or clear bacteria. Conclusions: These data indicate that specific Orai1 inhibition by ELD607 may be a novel approach to reduce multiorgan inflammation and treat antibiotic-resistant bacteria.


Assuntos
Staphylococcus aureus Resistente à Meticilina , Pneumonia Bacteriana , Síndrome do Desconforto Respiratório , Humanos , Camundongos , Animais , Canais de Cálcio/metabolismo , Canais de Cálcio/farmacologia , Cálcio/metabolismo , Células HEK293 , Staphylococcus aureus Resistente à Meticilina/metabolismo , Sinalização do Cálcio , Inflamação/tratamento farmacológico , Pulmão/metabolismo , Síndrome do Desconforto Respiratório/tratamento farmacológico , Pneumonia Bacteriana/tratamento farmacológico , Proteína ORAI1/metabolismo , Proteína ORAI1/farmacologia
5.
Int J Mol Sci ; 24(20)2023 Oct 19.
Artigo em Inglês | MEDLINE | ID: mdl-37895029

RESUMO

Proteases such as neutrophil elastase cleave and activate the epithelial sodium channel (ENaC), causing airway dehydration. Our current study explores the impact of increased protease levels in vapers' airways on ENaC activity and airway dehydration. Human bronchial epithelial cultures (HBECs) were exposed to bronchoalveolar lavage fluid (BALF) from non-smokers, smokers and vapers. Airway surface liquid (ASL) height was measured by confocal microscopy as a marker of hydration. ENaC cleavage was measured by Western blotting. Human peripheral blood neutrophils were treated with a menthol-flavored e-liquid (Juul), and the resulting secretions were added to HBECs. BALF from smokers and vapers significantly and equally increased ENaC activity and decreased ASL height. The ASL height decrease was attenuated by protease inhibitors. Non-smokers' BALF had no effect on ENaC or ASL height. BALF from smokers and vapers, but not non-smokers, induced ENaC cleavage. E-liquid-treated neutrophil secretions cleaved ENaC and decreased ASL height. Our study demonstrated that elevated protease levels in vapers' airways have functional significance since they can activate ENaC, resulting in airway dehydration. Lung dehydration contributes to diseases like cystic fibrosis (CF), chronic obstructive pulmonary disease (COPD) and asthma. Thus, our data predict that vaping, like smoking, will cause airway surface dehydration that likely leads to lung disease.


Assuntos
Vaping , Humanos , Vaping/efeitos adversos , Proteólise , Desidratação/metabolismo , Mucosa Respiratória/metabolismo , Pulmão/metabolismo , Canais Epiteliais de Sódio/metabolismo
6.
Indian J Nephrol ; 33(5): 387-391, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37881736

RESUMO

Hypercalcemia in infants presents with a variety of clinical features and the etiology of hypercalcemia varies with age. Here we present a case of hypercalcemia in an infant presenting with nephrocalcinosis and nephrolithiasis. Our investigations led us to a diagnosis of primary hyperoxaluria (PH) type 2, a rare metabolic disorder, along with hypercalcemia, a never before reported association. A 9-month-old female presented with urinary tract infection and systemic features requiring hospitalization and parenteral antibiotics. Investigations revealed bilateral medullary nephrocalcinosis. Genetic testing revealed a diagnosis of Primary hyperoxaluria type 2 with two possible mutations. Sanger sequencing of the parents identified the pathogenic mutation in the mother. This is the first report of a genetically proven case of primary hyperoxaluria type 2 associated with hypercalcemia.

7.
J Indian Assoc Pediatr Surg ; 28(2): 154-159, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37197249

RESUMO

46, XX testicular differences of sex development (DSD) is a rare cause of DSD presenting as a phenotypical male with chromosomal sex of 46, XX. Sex-determining region of the Y chromosome (SRY)-positive 46, XX DSDs have a well-characterized pathogenetic mechanism, whereas in SRY-negative 46, XX DSDs, the pathogenesis is not clearly delineated. Herein, we present a case of a 3½-year-old child who presented with ambiguous genitalia and bilateral palpable gonads. On the basis of a karyotype and fluorescent in situ hybridization, we arrived at a diagnosis of SRY-negative 46, XX testicular DSD. Basal serum estradiol and human menopausal gonadotrophin stimulated estradiol levels and inhibin A blood levels were against the presence of any ovarian tissue. Imaging of the gonads showed bilateral normal-looking testis. A clinical exome sequencing revealed a heterozygous missense variant NR5A1:c275G>A (p. Arg92gln) located at exon 4 in the affected child. Protein structure analysis was further performed, and the variant was found to be highly conserved. Sanger's sequencing showed that the mother was heterozygous for the variant detected in the child. This case highlights the rarity of SRY-negative 46, XX testicular DSD with a unique variant. Largely under characterized, this group of DSDs needs to be reported and analyzed to add to the spectrum of presentation and genetic characteristics. Our case is expected to add to the database, knowledge, and approach to cases of 46, XX testicular DSD.

8.
J Appl Toxicol ; 43(6): 862-873, 2023 06.
Artigo em Inglês | MEDLINE | ID: mdl-36594405

RESUMO

Cigarette smoke (CS) exposure induces both cytotoxicity and inflammation, and often causes COPD, a growing cause of morbidity and mortality. CS also inhibits the CFTR Cl- channel, leading to airway surface liquid dehydration, which is predicated to impair clearance of inhaled pathogens and toxicants. Numerous in vitro studies have been performed that utilize acute (≤24 h) CS exposures. However, CS exposure is typically chronic. We evaluated the feasibility of using British-American Tobacco (BAT)-designed CS exposure chambers for chronically exposing human bronchial epithelial cultures (HBECs) to CS. HBECs are polarized and contain mucosal and serosal sides. In vivo, inhaled CS interacts with mucosal membranes, and BAT chambers are designed to direct CS to HBEC mucosal surfaces while keeping CS away from serosal surfaces via a perfusion system. We found that serosal perfusion was absolutely required to maintain HBEC viability over time following chronic CS exposure. Indeed, with this system, we found that CS increased inflammation and mucin levels, while decreasing CFTR function. Without this serosal perfusion, CS was extremely toxic within 24 h. We therefore propose that 5- and 10-day CS exposures with serosal perfusion are suitable for measuring chronic CS exposure and can be used for monitoring new and emerging tobacco products.


Assuntos
Fumar Cigarros , Regulador de Condutância Transmembrana em Fibrose Cística , Humanos , Brônquios , Nicotiana/toxicidade , Inflamação , Células Epiteliais
9.
J Endocr Soc ; 7(1): bvac166, 2022 Nov 17.
Artigo em Inglês | MEDLINE | ID: mdl-36405867

RESUMO

Hypoparathyroidism is a common encounter in endocrinology practice. A thorough search for the etiology is generally futile, and most cases are labeled as idiopathic. Familial idiopathic hypoparathyroidism is a large chunk of these idiopathic cases. Here we present 2 cases who presented with features of hypocalcemia and were eventually diagnosed with hypoparathyroidism. Our first case is that of a middle-age woman who presented with spontaneous tetany and perioral numbness. She had very low serum calcium values, low serum magnesium, hypokalemia, hypercalciuria, and undetectable parathormone levels. She was initially managed with parenteral calcium, magnesium, and oral potassium chloride, which was shifted to oral replacements once stabilized. Focused exome sequencing for causes of hypoparathyroidism and hypocalcemia revealed a frameshift mutation in glial cell missing homolog 2 (GCM2) (NM_004752.4) on chromosome 6, c737dupA variant (p. Asp246Glufs*25) located at exon 5. The second case presented is that of a 1-month-old infant presenting with hypocalcemic seizures, severe hypocalcemia, hyperphosphatemia, and low parathormone levels. The infant was stabilized with parenteral calcium and trial of subcutaneous teriparatide for further improvement. Oral calcium and calcitriol were instituted once stabilized, and teriparatide was tapered off. Focused exome sequencing revealed a homozygous mutation involving GCM2 (ENST0000379491.5) on chromosome 6, variant CM2 chr6:10876558_10877139insT located on exon1-2. Both of these mutations are novel and underscore the profound effect of GCM2 on parathyroid gland development in infants and maintenance in adults.

10.
Physiol Rep ; 10(10): e15306, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-35581745

RESUMO

Orai1 is a ubiquitously-expressed plasma membrane Ca2+ channel that is involved in store-operated Ca2+ entry (SOCE): a fundamental biological process that regulates gene expression, the onset of inflammation, secretion, and the contraction of airway smooth muscle (ASM). During SOCE, Ca2+ leaves the endoplasmic reticulum, which then stimulates a second, amplifying wave of Ca2+ influx through Orai1 into the cytoplasm. Short Palate LUng and Nasal epithelial Clone 1 (SPLUNC1; gene name BPIFA1) is a multi-functional, innate defense protein that is highly abundant in the lung. We have previously reported that SPLUNC1 was secreted from epithelia, where it bound to and inhibited Orai1, leading to reduced SOCE and ASM relaxation. However, the underlying mechanism of action is unknown. Here, we probed the SPLUNC1-Orai1 interactions in ASM and HEK293T cells using biochemical and imaging techniques. We observed that SPLUNC1 caused a conformational change in Orai1, as measured using Forster resonance energy transfer (FRET). SPLUNC1 binding also led to Nedd4-2 dependent ubiquitination of Orai1. Moreover, SPLUNC1 internalized Orai1 to lysosomes, leading to Orai1 degradation. Thus, we conclude that SPLUNC1 is an allosteric regulator of Orai1. Our data indicate that SPLUNC1-mediated Orai1 inhibition could be utilized as a therapeutic strategy to reduce SOCE.


Assuntos
Glicoproteínas/metabolismo , Pulmão , Músculo Liso , Fosfoproteínas/metabolismo , Cálcio/metabolismo , Sinalização do Cálcio/fisiologia , Membrana Celular/metabolismo , Células HEK293 , Humanos , Pulmão/metabolismo , Músculo Liso/metabolismo , Proteína ORAI1/genética , Proteína ORAI1/metabolismo
11.
Circulation ; 145(3): 219-232, 2022 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-35041473

RESUMO

Electronic cigarettes (e-cigarettes) are battery powered electronic nicotine delivery systems that use a propylene glycol/vegetable glycerin base to deliver vaporized nicotine and flavorings to the body. E-cigarettes became commercially available without evidence regarding their risks, long-term safety, or utility in smoking cessation. Recent clinical trials suggest that e-cigarette use with counseling may be effective in reducing cigarette use but not nicotine dependence. However, meta-analyses of observational studies demonstrate that e-cigarette use is not associated with smoking cessation. Cardiovascular studies reported sympathetic activation, vascular stiffening, and endothelial dysfunction, which are associated with adverse cardiovascular events. The majority of pulmonary clinical trials in e-cigarette users included standard spirometry as the primary outcome measure, reporting no change in lung function. However, studies reported increased biomarkers of pulmonary disease in e-cigarette users. These studies were conducted in adults, but >30% of high school-age adolescents reported e-cigarette use. The effects of e-cigarette use on cardiopulmonary endpoints in adolescents and young adults remain unstudied. Because of adverse clinical findings and associations between e-cigarette use and increased incidence of respiratory diseases in people who have never smoked, large longitudinal studies are needed to understand the risk profile of e-cigarettes. Consistent with the Centers for Disease Control and Prevention recommendations, clinicians should monitor the health risks of e-cigarette use, discourage nonsmokers and adolescents from using e-cigarettes, and discourage smokers from engaging in dual use without cigarette reduction or cessation.


Assuntos
Fumar Cigarros/efeitos adversos , Sistemas Eletrônicos de Liberação de Nicotina , Papel do Médico , Vaping/efeitos adversos , Humanos , Fumar/epidemiologia , Tabagismo/prevenção & controle
12.
Nicotine Tob Res ; 24(3): 395-399, 2022 02 14.
Artigo em Inglês | MEDLINE | ID: mdl-34519792

RESUMO

INTRODUCTION: Alveolar macrophages (AMs) are lung-resident immune cells that phagocytose inhaled particles and pathogens, and help coordinate the lung's immune response to infection. Little is known about the impact of chronic e-cigarette use (ie, vaping) on this important pulmonary cell type. Thus, we determined the effect of vaping on AM phenotype and gene expression. AIMS AND METHODS: We recruited never-smokers, smokers, and e-cigarette users (vapers) and performed research bronchoscopies to isolate AMs from bronchoalveolar lavage fluid samples and epithelial cells from bronchial brushings. We then performed morphological analyses and used the Nanostring platform to look for changes in gene expression. RESULTS: AMs obtained from smokers and vapers were phenotypically distinct from those obtained from nonsmokers, and from each other. Immunocytochemistry revealed that vapers AMs had significantly elevated inducible nitric oxide synthase (M1) expression and significantly reduced CD301a (M2) expression compared with nonsmokers or smokers. Vapers' AMs and bronchial epithelia exhibited unique changes in gene expression compared with nonsmokers or smokers. Moreover, vapers' AMs were the most affected of all groups and had 124 genes uniquely downregulated. Gene ontology analysis revealed that vapers and smokers had opposing changes in biological processes. CONCLUSIONS: These data indicate that vaping causes unique changes to AMs and bronchial epithelia compared with nonsmokers and smokers which may impact pulmonary host defense. IMPLICATIONS: These data indicate that normal "healthy" vapers have altered AMs and may be at risk of developing abnormal immune responses to inflammatory stimuli.


Assuntos
Sistemas Eletrônicos de Liberação de Nicotina , Produtos do Tabaco , Vaping , Expressão Gênica , Humanos , Macrófagos Alveolares , Vaping/efeitos adversos
13.
Am J Respir Cell Mol Biol ; 66(3): 271-282, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-34807800

RESUMO

Orai1 is a plasma membrane Ca2+ channel that mediates store-operated Ca2+ entry (SOCE) and regulates inflammation. Short palate lung and nasal epithelial clone 1 (SPLUNC1) is an asthma gene modifier that inhibits Orai1 and SOCE via its C-terminal α6 region. SPLUNC1 levels are diminished in asthma patient airways. Thus, we hypothesized that inhaled α6 peptidomimetics could inhibit Orai1 and reduce airway inflammation in a murine asthma model. To evaluate α6-Orai1 interactions, we used fluorescent assays to measure Ca2+ signaling, Förster resonance energy transfer, fluorescent recovery after photobleaching, immunostaining, total internal reflection microscopy, and Western blotting. To test whether α6 peptidomimetics inhibited SOCE and decreased inflammation in vivo, wild-type and SPLUNC1-/- mice were exposed to house dust mite (HDM) extract with or without α6 peptide. We also performed nebulization, jet milling, and scanning electron microscopy to evaluate α6 for inhalation. SPLUNC1-/- mice had an exaggerated response to HDM. In BAL-derived immune cells, Orai1 levels increased after HDM exposure in SPLUNC1-/- but not wild-type mice. Inhaled α6 reduced Orai1 levels in mice regardless of genotype. In HDM-exposed mice, α6 dose-dependently reduced eosinophilia and neutrophilia. In vitro, α6 inhibited SOCE in multiple immune cell types, and α6 could be nebulized or jet milled without loss of function. These data suggest that α6 peptidomimetics may be a novel, effective antiinflammatory therapy for patients with asthma.


Assuntos
Asma , Peptidomiméticos , Animais , Asma/tratamento farmacológico , Cálcio/metabolismo , Glicoproteínas , Humanos , Inflamação , Pulmão/metabolismo , Camundongos , Proteína ORAI1/genética , Proteína ORAI1/metabolismo , Fosfoproteínas
15.
Sci Signal ; 14(706): eabg4747, 2021 Oct 26.
Artigo em Inglês | MEDLINE | ID: mdl-34699252

RESUMO

Astrocytes are a type of glial cell that are activated in the brain tissue of patients with Alzheimer's disease to induce the accumulation of amyloid (Aß). We previously found that a combination of low-dose gemfibrozil (GFB; a drug approved to treat high cholesterol) and retinoic acid (RA; a vitamin A derivative) induces lysosomal bio-genesis through peroxisome proliferator­activated receptor α (PPARα)­mediated transcription of the gene encoding transcription factor EB (TFEB), a master regulator of lysosomal biogenesis and autophagy. Here, we found that the same combination (GFB-RA) enhanced the uptake of Aß from the extracellular space and its subsequent degradation in astrocytes through a PPARα-dependent pathway. GFB-RA stimulated the abundance of both low-density lipoprotein receptor (LDLR) and TFEB in astrocytes through PPARα. LDLR was critical for Aß uptake, whereas TFEB was critical for its degradation. GFB-RA treatment also increased autophagic flux and lysosomal activity in astrocytes. Consistent with these effects and in a manner dependent on astroglial PPARα, oral administration of GFB-RA switched astroglial activation to a neuroprotective state, lowered Aß burden in the brain, and improved spatial learning and memory in the 5XFAD mouse model of Alzheimer's disease. These findings uncover a new function of PPARα in stimulating astroglial uptake and degradation of Aß and suggest possible repurposing of GFB-RA combination therapy for AD.


Assuntos
Peptídeos beta-Amiloides , PPAR alfa , Peptídeos beta-Amiloides/metabolismo , Astrócitos/metabolismo , Transporte Biológico , PPAR alfa/metabolismo
16.
J Appl Toxicol ; 41(3): 493-505, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33034066

RESUMO

"Pod-based" e-cigarettes such as JUUL are currently the most prevalent electronic nicotine delivery systems (ENDS) in the United States. JUUL-type ENDS utilize nicotine salts protonated with benzoic acid rather than freebase nicotine. However, limited information is available on the cellular effects of these products. Cytoplasmic Ca2+ is a universal second messenger that controls many cellular functions including cell growth and cell death. Of note, dysregulation of cell Ca2+ homeostasis has been linked with several disease processes including autoimmune disease and several types of cancer. We exposed HEK293T cells and THP-1 macrophage-like cells to different JUUL e-liquids. We evaluated their effects on cellular viability and Ca2+ signaling by measuring fluorescence from calcein-AM/propidium iodide and Fluo-4, respectively. E-liquid autofluorescence was used to look for e-liquid permeation into cells. To identify the mechanisms behind the Ca2+ responses, different inhibitors of Ca2+ channels and phospholipase C signaling were used. JUUL e-liquids caused significant cytotoxic effects, with "Mint" flavor being the most cytotoxic. The Mint flavored e-liquid also caused a significant elevation in cytoplasmic Ca2+ . Using autofluorescence, the permeation of JUUL e-liquids into live cells was confirmed, indicating that intracellular organelles are directly exposed to e-liquids. Further studies identified the endoplasmic reticulum as being the source of e-liquid-induced changes in cytoplasmic Ca2+ . Nicotine salt-based e-liquids cause cytotoxicity and elevate cytoplasmic Ca2+ , indicating that they can exert biological effects beyond what would be expected with nicotine alone. These effects are flavor-dependent, and we propose that flavored e-liquids be reassessed for potential lung toxicity.


Assuntos
Morte Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas/efeitos dos fármacos , Vapor do Cigarro Eletrônico/toxicidade , Sistemas Eletrônicos de Liberação de Nicotina , Aromatizantes/toxicidade , Nicotina/toxicidade , Humanos , Estados Unidos
18.
Am J Respir Cell Mol Biol ; 63(6): 767-779, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32877614

RESUMO

Smoking remains a leading cause of preventable morbidity and mortality worldwide. Despite a downward trend in cigarette use, less-regulated tobacco products, such as cigarillos, which are often flavored to appeal to specific demographics, such as younger people, are becoming increasingly popular. Cigar/cigarillo smoking has been considered a safer alternative to cigarettes; however, the health risks associated with cigar in comparison with cigarette smoking are not well understood. To address this knowledge gap, we characterized the effects of multiple brands of cigarillos on the airway epithelium using ex vivo and in vivo models. To analyze these effects, we assessed the cellular viability and integrity of smoke-exposed primary airway cell cultures. We also investigated the protein compositions of apical secretions from cigarillo-exposed airway epithelial cultures and BAL fluid of cigarillo-exposed mice through label-free quantitative proteomics and determined the chemical composition of smoke collected from the investigated cigarillo products. We found that cigarillo smoke exerts similar or greater effects than cigarette smoke in terms of reduced cell viability; altered protein levels, including those of innate immune proteins; induced oxidative-stress markers; and greater nicotine delivery to cells. The analysis of the chemical composition of the investigated cigarillo products revealed differences that might be linked to the differential effects of these products on cell viability and protein abundance profiles, which have been associated with a range of health risks in the context of airway biology. These findings contradict the assumption that cigarillos might be safer and less harmful than cigarettes. Instead, our results indicate that cigarillo smoke is associated with equal or greater health risks and the same or increased airway toxicity compared with cigarette smoke.


Assuntos
Epitélio/efeitos dos fármacos , Epitélio/metabolismo , Nicotina/farmacologia , Sistema Respiratório/metabolismo , Animais , Fumar Cigarros/efeitos adversos , Aromatizantes/farmacologia , Humanos , Camundongos Endogâmicos C57BL , Sistema Respiratório/efeitos dos fármacos , Fumar/efeitos adversos , Nicotiana/efeitos adversos , Produtos do Tabaco/efeitos adversos
19.
Am J Respir Crit Care Med ; 200(11): 1392-1401, 2019 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-31390877

RESUMO

Rationale: Proteolysis is a key aspect of the lung's innate immune system. Proteases, including neutrophil elastase and MMPs (matrix metalloproteases), modulate cell signaling, inflammation, tissue remodeling, and leukocyte recruitment via cleavage of their target proteins. Excessive proteolysis occurs with chronic tobacco use and is causative for bronchiectasis and emphysema. The effect of e-cigarettes (vaping) on proteolysis is unknown.Objectives: We used protease levels as biomarkers of harm to determine the impact of vaping on the lung.Methods: We performed research bronchoscopies on healthy nonsmokers, cigarette smokers, and e-cigarette users (vapers), and determined protease levels in BAL. In parallel, we studied the effects of e-cigarette components on protease secretion in isolated human blood neutrophils and BAL-derived macrophages. We also analyzed the nicotine concentration in induced sputum and BAL.Measurements and Main Results: Neutrophil elastase, MMP-2, and MMP-9 activities and protein levels were equally elevated in both vapers' and smokers' BAL relative to nonsmokers. In contrast, antiprotease levels were unchanged. We also found that exposure of isolated neutrophils and macrophages to nicotine elicited dose-dependent increases in protease release. After vaping, measurable levels of nicotine were detectable in sputum and BAL, which corresponded to the half-maximal effective concentration values for protease release seen in immune cells.Conclusions: We conclude that vaping induces nicotine-dependent protease release from resident pulmonary immune cells. Thus, chronic vaping disrupts the protease-antiprotease balance by increasing proteolysis in lung, which may place vapers at risk of developing chronic lung disease. These data indicate that vaping may not be safer than tobacco smoking.


Assuntos
Elastase de Leucócito/metabolismo , Pulmão/efeitos dos fármacos , Metaloproteinases da Matriz/metabolismo , Vaping/efeitos adversos , Adulto , Biomarcadores/análise , Líquido da Lavagem Broncoalveolar/química , Cotinina/análogos & derivados , Cotinina/análise , Feminino , Humanos , Pulmão/química , Pulmão/enzimologia , Macrófagos/efeitos dos fármacos , Macrófagos/enzimologia , Masculino , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Neutrófilos/efeitos dos fármacos , Neutrófilos/enzimologia , Nicotina/análise , Nicotina/farmacologia
20.
Eur Respir J ; 52(4)2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30190268

RESUMO

The multi-organ disease cystic fibrosis (CF) is caused by mutations in the cystic fibrosis transmembrane regulator gene (CFTR) that lead to diminished transepithelial anion transport. CF lungs are characterised by airway surface liquid (ASL) dehydration, chronic infection/inflammation and neutrophilia. Dysfunctional CFTR may upregulate the epithelial Na+ channel (ENaC), further exacerbating dehydration. We previously demonstrated that short palate lung and nasal epithelial clone 1 (SPLUNC1) negatively regulates ENaC in normal airway epithelia.Here, we used pulmonary tissue samples, sputum and human bronchial epithelial cells (HBECs) to determine whether SPLUNC1 could regulate ENaC in a CF-like environment.We found reduced endogenous SPLUNC1 in CF secretions, and rapid degradation of recombinant SPLUNC1 (rSPLUNC1) by CF secretions. Normal sputum, containing SPLUNC1 and SPLUNC1-derived peptides, inhibited ENaC in both normal and CF HBECs. Conversely, CF sputum activated ENaC, and rSPLUNC1 could not reverse this phenomenon. Additionally, we observed upregulation of ENaC protein levels in human CF bronchi. Unlike SPLUNC1, the novel SPLUNC1-derived peptide SPX-101 resisted protease degradation, bound apically to HBECs, inhibited ENaC and prevented ASL dehydration following extended pre-incubation with CF sputum.Our data indicate that CF mucosal secretions drive ASL hyperabsorption and that protease-resistant peptides, e.g. SPX-101, can reverse this effect to rehydrate CF ASL.


Assuntos
Fibrose Cística/metabolismo , Desidratação/patologia , Células Epiteliais/metabolismo , Glicoproteínas/metabolismo , Fosfoproteínas/metabolismo , Células Cultivadas , Canais Epiteliais de Sódio/metabolismo , Glicoproteínas/genética , Humanos , Transporte de Íons , Pulmão/metabolismo , Fosfoproteínas/genética , Mucosa Respiratória/metabolismo
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