Receptor-Mediated Tobacco Toxicity (original) (raw)
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The FASEB Journal, 2007
Tobacco products and nicotine alter the cell cycle and lead to squamatization of oral keratinocytes (KCs) and squamous cell carcinoma. Activation of nicotinic acetylcholine receptors (nAChRs) elicits Ca 2؉ influx that varies in magnitude between different nAChR subtypes. Normal differentiation of KCs is associated with sequential expression of the nAChR subtypes with increasing Ca 2؉ permeability, such as ␣5-containing ␣3 nAChR and ␣7 nAChR. Exposure to environmental tobacco smoke (ETS) or an equivalent concentration of nicotine accelerated by severalfold the ␣5 and ␣7 expression in KCs, which could be abolished by mecamylamine and ␣-bungarotoxin with different efficacies, suggesting the following sequence of autoregulation of the expression of nAChR subtypes: ␣3(2/4) > ␣3(2/4)␣5 > ␣7 > ␣7. This conjecture was corroborated by results of quantitative assays of subunit mRNA and protein levels, using nAChRspecific pharmacologic antagonists and small interfering RNAs. The genomic effects of ETS and nicotine involved the transcription factor GATA-2 that showed a multifold increase in quantity and activity in exposed KCs. Using protein kinase inhibitors and dominant negative and constitutively active constructs, we characterized the principal signaling cascades mediating a switch in the nAChR subtype. Cumulative results indicated that the ␣3(2/4) to ␣3(2/4)␣5 nAChR transition predominantly involved protein kinase C, ␣3(2/ 4)␣5 to ␣7 nAChR transition-Ca 2؉ /calmodulindependent protein kinase II and p38 MAPK, and ␣7 self-up-regulation-the p38 MAPK/Akt pathway, and JAK-2. These results provide a mechanistic insight into the genomic effects of ETS and nicotine on KCs and characterize signaling pathways mediating autoregulation of stepwise overexpression of nAChR subtypes with increasing Ca 2؉ permeability in exposed cells. These observations have salient clinical implications, because a switch in the nAChR subunit composition can bring about a corresponding switch in receptor function, leading to profound pathobiologic effects observed in KCs exposed to tobacco products. Arre
Faseb Journal, 2007
Tobacco products and nicotine alter the cell cycle and lead to squamatization of oral keratinocytes (KCs) and squamous cell carcinoma. Activation of nicotinic acetylcholine receptors (nAChRs) elicits Ca 2؉ influx that varies in magnitude between different nAChR subtypes. Normal differentiation of KCs is associated with sequential expression of the nAChR subtypes with increasing Ca 2؉ permeability, such as ␣5-containing ␣3 nAChR and ␣7 nAChR. Exposure to environmental tobacco smoke (ETS) or an equivalent concentration of nicotine accelerated by severalfold the ␣5 and ␣7 expression in KCs, which could be abolished by mecamylamine and ␣-bungarotoxin with different efficacies, suggesting the following sequence of autoregulation of the expression of nAChR subtypes: ␣3(2/4) > ␣3(2/4)␣5 > ␣7 > ␣7. This conjecture was corroborated by results of quantitative assays of subunit mRNA and protein levels, using nAChRspecific pharmacologic antagonists and small interfering RNAs. The genomic effects of ETS and nicotine involved the transcription factor GATA-2 that showed a multifold increase in quantity and activity in exposed KCs. Using protein kinase inhibitors and dominant negative and constitutively active constructs, we characterized the principal signaling cascades mediating a switch in the nAChR subtype. Cumulative results indicated that the ␣3(2/4) to ␣3(2/4)␣5 nAChR transition predominantly involved protein kinase C, ␣3(2/ 4)␣5 to ␣7 nAChR transition-Ca 2؉ /calmodulindependent protein kinase II and p38 MAPK, and ␣7 self-up-regulation-the p38 MAPK/Akt pathway, and JAK-2. These results provide a mechanistic insight into the genomic effects of ETS and nicotine on KCs and characterize signaling pathways mediating autoregulation of stepwise overexpression of nAChR subtypes with increasing Ca 2؉ permeability in exposed cells. These observations have salient clinical implications, because a switch in the nAChR subunit composition can bring about a corresponding switch in receptor function, leading to profound pathobiologic effects observed in KCs exposed to tobacco products. Arre-dondo, J., Chernyavsky, A. I., Jolkovsky, D. L., Pinkerton, K. E., Grando, S. A. Receptor-mediated tobacco toxicity: acceleration of sequential expression of ␣5 and ␣7 nicotinic receptor subunits in oral keratinocytes exposed to cigarette smoke. FASEB J. 22, 1356 -1368 (2008)
The FASEB Journal, 2006
The use of tobacco products is associated with an increased incidence of periodontal disease, poor response to periodontal therapy, and a high risk for developing head and neck cancer. Nicotine and tobacco-derived nitrosamines have been shown to exhibit their pathobiologic effects due in part to activation of the nicotinic acetylcholine (ACh) receptors (nAChRs), mainly ␣7 nAChR, expressed by oral keratinocytes (KCs). This study was designed to gain mechanistic insight into ␣7-mediated morbidity of tobacco products in the oral cavity. We investigated the signaling pathways downstream of ␣7 nAChR in monolayers of oral KCs exposed for 24 h to aged and diluted sidestream cigarette smoke (ADSS) or an equivalent concentration of pure nicotine. By both real-time polymerase chain reaction (PCR) and In-cell Western, the KCs stimulated with ADSS or nicotine showed multifold increases of STAT-3. These effects could be completely blocked or significantly (P<0.05) diminished if the cells were pretreated with the ␣7 antagonist ␣-bungarotoxin (␣BTX) or transfected with anti-␣7 small interfering RNA (siRNA-␣7). The use of pathway inhibitors revealed that signaling through the Ras/Raf-1/MEK1/ ERK steps mediated ␣7-dependent up-regulation of STAT-3. Targeted mutation of the ␣7 gene prevented ERK1/2 activation by nicotine. Using the gel mobility shift assay, we demonstrated that an increased protein binding activity of STAT-3 caused by ADSS or pure nicotine was mediated by janus-activated kinase (JAK)-2. Activation of JAK-2/STAT-3 pathway could be prevented by ␣BTX or siRNA-␣7. Thus, nuclear transactivation of STAT-3 in KCs exposed to tobacco products is mediated via intracellular signaling downstream from ␣7, which proceeds via two complementary pathways. The Ras/Raf-1/MEK1/ERK cascade culminates in up-regulated expression of the gene encoding STAT-3, whereas recruitment and activation of tyrosine kinase JAK-2 phosphorylates it. Elucidation of this novel mechanism of nicotine-dependent nuclear transactivation of STAT-3 identifies oral ␣7 nAChR as a promising molecular target to prevent, reverse, or retard tobacco-related periodontal disease and progression of head and neck cancer by receptor inhibitors.-Arredondo, J., Chernyavsky, A. I., Jolkovsky, D. L., Pinkerton, K. E., Grando, S. A. Receptor-mediated tobacco toxicity: cooperation of the Ras/Raf-1/MEK1/ ERK and JAK-2/STAT-3 pathways downstream of ␣7 nicotinic receptor in oral keratinocytes.
Effect of tobacco compounds on gene expression profiles in human epithelial cells
Environmental Toxicology and Pharmacology, 2009
This study was carried out to investigate the effects of the tobacco compounds (TC), nicotine, B(a)P, and 2-naphthylamine, on gene expression profiles in a human epithelial cells (A549). We treated A549 with the TC and analyzed gene expression using microarray and real-time PCR (RTP). Gene expression varied according to the TC used. By microarray, we found that apoptosis-related genes such as apoptosisassociated tyrosine kinase, interleukin 10 receptor beta, caspase 1 and DNA fragmentation factor beta subunit (40 kDa) were down-regulated in TC-treated A549 cells. RTP showed significant increases in the expression of Ahr, Arnt, CYP1A1, and CYP1B1 in TC-treated A549 cells. From these results, we suggest that tobacco compounds can influence apoptosis, inflammation, immunity, and the cell cycle in A549 cells. Also, our study demonstrates that a microarray-based genomic survey is a suitable high-throughput approach for the evaluation of gene expression and for the characterization of TC-induced toxicity.
Involvement of Various Molecular Events in Cellular Injury Induced by Smokeless Tobacco
Chemical Research in Toxicology, 2010
Smokeless tobacco (ST) consumption is implicated in the pathogenesis of oral diseases, including cancer. However, its pathological effect in other organs is not well understood. In the present study, the effect of aqueous extract of smokeless tobacco (AEST) prepared from "gutkha" (a form of ST) on the xenobiotic drug-metabolizing enzymes, histopathological changes, and damage to the genetic material in lung, liver, and kidney of rats was evaluated. Animals were orally administered AEST at a low dose (L-AEST, 96 mg/kg body wt/day) for 2 (L-AEST 2 ) and 28 weeks (L-AEST 28 ) and at a high dose (H-AEST, 960 mg/kg body wt/day) for 2 weeks (H-AEST 2 ). Real-time PCR and immunohistological studies showed that administration of L-AEST 2 did not induce the expression of phase I cytochrome P450s (CYP1A1, 1A2, and 2E1) and phase II µ-glutathione-s-transferase (GST-µ) drug-metabolizing enzymes in lung, liver, and kidney. Although H-AEST 2 administration significantly induced both gene and protein expression of CYP1A1, 1A2, and 2E1 in all of the above organs, it mildly expressed the phase II detoxifying enzyme, GST-µ, in type I and type II epithelial cells of lung and in proximal tubular cells of kidney. L-AEST 28 enhanced the gene and protein expression of CYP1A1, 1A2, and 2E1 in lung, liver, and kidney in a differential manner and induced the expression of GST-µ in lung and kidney. L-AEST 28 induced the micronuclei formation in the peripheral blood mononuclear cells, TNF-R in plasma, and myeloperoxidase activity in the organs. L-AEST 28 significantly enhanced Bax, p53, and NF-κB and decreased Bcl-2 gene expressions differentially in an organ-specific manner. The differential changes in these organs due to AEST might be due to their different physiological functions and variable sensitivities toward the metabolites of AEST, which create a microenvironment favorable for AEST-induced pathogenesis. This study broadens the insight into the different molecular mechanisms in various organs, which appear to be deregulated due to AEST. Understanding these processes may help in clinical treatment planning strategies for tobacco-related diseases.
A Receptor-Mediated Mechanism of Nicotine Toxicity in Oral Keratinocytes
Laboratory Investigation, 2001
Smoking and smokeless tobacco cause morbidity that originates from the epithelium lining of the skin and upper digestive tract. Oral keratinocytes (OKC) express nicotinic acetylcholine receptors (nAChRs) that bind nicotine (Nic). We studied the mechanism of the receptor-mediated toxicity of tobacco products on OKC. Preincubation of normal human OKC with Nic altered the ligand-binding kinetics of their nAChRs, suggesting that
Expression Changes of Apoptotic Genes in Tissues from Mice Exposed to Nicotine
Asian Pacific journal of cancer prevention : APJCP, 2017
Objective: Smoking is the leading preventable cause of various diseases such as lung cancer, chronic obstructive pulmonary disease and cardiovascular disease. Nicotine, one of the major toxic components of tobacco, contributes to the pathogenesis of different diseases. Methods: Given the controversy about nicotine toxicity, the present study was conducted to determine apoptotic effects of nicotine on the heart, kidney, lung and liver of male mice. Real-time PCR was performed to identify mRNA expression changes in apoptotic-related genes between nicotine treated and control mice. Result: In the heart and lung, nicotine caused significant decrease in P53, Bax and Caspase-3 mRNA expression levels compared to the control group. However, in the kidney and liver, the result was significant increase in Bax, Caspase-2, Caspase-3 and a significant decrease in P53 mRNA expression (p<0.01). DNA fragmentation assays indicated no fragmentation in the heart and lung, but in the kidney and live...