Cigarette Smoke Constituents Research Articles

Cadmium Impairs p53 Activity in HepG2 Cells.

Cadmium and cadmium compounds are contaminants of the environment, food, and drinking water and are important constituents of cigarette smoke. Cd exposure has also been associated with airborne particulate CdO and with Cd-containing quantum dots in medical therapy. Adverse cadmium effects reported in the literature have stimulated during recent years an ongoing discussion to better elucidate cadmium outcomes at cell and molecular level. The present work is designed to gain an insight into the mechanism of p53 impairment at gene and protein level to understand Cd-induced resistance to apoptosis. We used a hepatoma cell line (HepG2) derived from liver, known to be metal responsive. At genotoxic cadmium concentrations no cell cycle arrest was observed. The p53 at gene and protein level was not regulated. Fluorescence images showed that p53 was correctly translocated into the nucleus but that the p21Cip1/WAF-1, a downstream protein of p53 network involved in cell cycle regulation, was not activated at the highest cadmium concentrations used. The miRNAs analysis revealed an upregulation of mir-372, an miRNA able to affect p21Cip1/WAF-1 expression and promote cell cycle progression and proliferation. The role of metallothioneins and possible conformational changes of p53 are discussed.

Low-dose nicotine self-administration is reduced in adult male rats naïve to high doses of nicotine: implications for nicotine product standards.

Product standards that greatly reduce the content of nicotine within cigarettes may result in improved public health. The study presented here used an animal model to investigate whether individuals who start smoking after implementation of regulation may be affected differently from current smokers who form the basis of most clinical studies. One group of adult male rats (n = 14/group) acquired nicotine self-administration at a high nicotine dose (60 μg/kg/infusion) before experiencing a reduction to one to three low doses of nicotine (3.75, 7.5, or 15 μg/kg/infusion) or vehicle. Their self-administration behavior at the low doses was compared with a group of adult male rats given the opportunity to acquire nicotine self-administration at one of the same low doses or vehicle (n = 7-14/group). Second, the self-administration behavior of the acquisition group of rats was compared with their own self-administration behavior after experience self-administering a high dose of nicotine. A cocktail of non-nicotine cigarette smoke constituents was included in the vehicle for all rats across all phases of the study. Rats with a history of self-administering a high dose of nicotine had a higher rate of self-administration across the low doses than rats with no history. In addition, the number of earned infusions increased after rats experienced self-administration of a higher dose of nicotine. These data show that low-dose nicotine self-administration is higher after a dose reduction than during acquisition. If a nicotine reduction policy were implemented, then this policy may be especially effective at reducing acquisition of smoking.

Comparison of Smoke Yield Data Collected from Different Laboratories

Abstract In the context of increasing tobacco product regulations, more requirements are observed for the reporting of smoke constituent yield data and its variability e.g., US Food and Drug Administration (FDA). The objective of this work was to evaluate the relevance of the short term standard deviation to describe the variability of measurements using the dataset of the CORESTA 2006 Joint Experiment which included a number of cigarette smoke constituents more recently identified by FDA for reporting. Their testing protocol required the analysis of Kentucky Reference cigarettes 2R4F and 1R5F performing five replicates run over consecutive days, repeated during three different time periods. This dataset provided access to different sources of smoke yield variability across measurements: short term and medium term within-laboratory variability and among-laboratory variability. For each reference cigarette, analysis of variance on one factor (laboratory) combined with the Newman-Keuls multiple range test was performed to compare data generated across laboratories. Results showed that the expression of yield variability as an individual standard deviation (describing repeatability) gives erroneous conclusions due to the major contribution of amonglaboratory variability not being taken into account. The different sources of variability can be taken into account in the comparison using the critical difference, as described in the ISO Standard 5725 part 6. This paper shows the importance of having i) the appropriate statistical methods to compare results from different laboratories in order to avoid erroneous conclusions, and ii) validated and standardized methods with known precision across laboratories. Moreover, it was demonstrated that the number of replicates had only a minor effect on product comparison on the basis of the critical difference as a function of repeatability and reproducibility of the methods.

A comprehensive toxicological evaluation of three adhesives using experimental cigarettes

ABSTRACTContext: Adhesives are used in several different manufacturing operations in the production of cigarettes. The use of new, “high-speed-manufacture” adhesives (e.g. vinyl acetate based) could affect the smoke chemistry and toxicology of cigarettes, compared with older “low-speed-manufacture” adhesives (e.g. starch based).Objective: This study was conducted to determine whether the inclusion of different levels of three adhesives (ethylene vinyl acetate, polyvinyl acetate and starch) in experimental cigarettes results in different smoke chemistry and toxicological responses in in vitro and in vivo assays.Materials and methods: A battery of tests (analytical chemistry, in vitro and in vivo assays) was used to compare the chemistry and toxicology of smoke from experimental cigarettes made with different combinations of the three adhesives. Varying levels of the different side-seam adhesives, as well as the transfer of adhesives from packaging materials, were tested.Results: There were differences in some mainstream cigarette smoke constituents as a function of the level of adhesive added to experimental cigarettes and between the tested adhesives. None of these differences translated into statistically significant differences in the in vitro or in vivo assays.Conclusion: The use of newer “high-speed-manufacture” vinyl acetate-based adhesives in cigarettes does not produce toxicological profiles that prevent the adhesives from replacing the older “low-speed-manufacture” adhesives (such as starch).

The detox strategy in smoking comprising nutraceutical formulas of non-hydrolyzed carnosine or carcinine used to protect human health

The increased oxidative stress in patients with smoking-associated disease, such as chronic obstructive pulmonary disease, is the result of an increased burden of inhaled oxidants as well as increased amounts of reactive oxygen species generated by various inflammatory, immune and epithelial cells of the airways. Nicotine sustains tobacco addiction, a major cause of disability and premature death. In addition to the neurochemical effects of nicotine, behavioural factors also affect the severity of nicotine withdrawal symptoms. For some people, the feel, smell and sight of a cigarette and the ritual of obtaining, handling, lighting and smoking a cigarette are all associated with the pleasurable effects of smoking. For individuals who are motivated to quit smoking, a combination of pharmacotherapy and behavioural therapy has been shown to be most effective in controlling the symptoms of nicotine withdrawal. In the previous studies, we proposed the viability and versatility of the imidazole-containing dipeptide-based compounds in the nutritional compositions as the telomere protection targeted therapeutic system for smokers in combination with in vitro cellular culture techniques being an investigative tool to study telomere attrition in cells induced by cigarette smoke (CS) and smoke constituents. Our working therapeutic concept is that imidazole-containing dipeptide-based compounds (non-hydrolyzed carnosine and carcinine) can modulate the telomerase activity in the normal cells and can provide the redox regulation of the cellular function under the terms of environmental and oxidative stress and in this way protect the length and the structure of telomeres from attrition. The detoxifying system of non-hydrolyzed carnosine or carcinine can be applied in the therapeutic nutrition formulations or installed in the cigarette filter. Patented specific oral formulations of non-hydrolyzed carnosine and carcinine provide a powerful manipulation tool for targeted therapeutic inhibition of cumulative oxidative stress and inflammation and protection from telomere attrition associated with smoking. It is demonstrated in this work that both non-hydrolyzed carnosine and carcinine are characterized by greater bioavailability than pure l-carnosine subjected to enzymatic hydrolysis with carnosinase, and perform the detoxification of the α,β-unsaturated carbonyl compounds present in tobacco smoke. We argue that while an array of factors has shaped the history of the 'safer' cigarette, it is the current understanding of the industry's past deceptions and continuing avoidance of the moral implications of the sale of products that cause the enormous suffering and death of millions that makes reconsideration of 'safer' cigarettes challenging. In contrast to this, the data presented in the article show that recommended oral forms of non-hydrolyzed carnosine and carcinine protect against CS-induced disease and inflammation, and synergistic agents with the actions of imidazole-containing dipeptide compounds in developed formulations may have therapeutic utility in inflammatory lung diseases where CS plays a role.

High throughput liquid chromatography-tandem mass spectrometry assay for mercapturic acids of acrolein and crotonaldehyde in cigarette smokers' urine.

3-Hydroxypropylmercapturic acid (3-HPMA) and 3-hydroxy-1-methylpropylmercapturic acid (HMPMA) are urinary metabolites of the toxicants acrolein and crotonaldehyde, respectively. Virtually all human urine samples contain these metabolites, resulting from the action of glutathione-S-transferases on acrolein and crotonaldehyde, which are lipid peroxidation products, environmental and dietary contaminants, and constituents of cigarette smoke. We have developed a high throughput liquid chromatography-tandem mass spectrometry method for quantitative analysis of 3-HPMA and HMPMA in large numbers of small urine samples, as would be required in molecular epidemiology and clinical studies relating levels of these metabolites to cancer risk. Solid-phase extraction on mixed mode reverse phase-anion exchange 96-well plates provided sufficient purification for LC-MS/MS analysis, which was performed by auto-injection using a 96-well format, and resulted in clean, readily interpretable chromatograms, with detection limits of 4.5pmol/mL urine for 3-HPMA and 3.5pmol/mL urine for HMPMA. Accuracy was 92% for 3-HPMA and 97% for HMPMA while inter-day precision was 9.1% (coefficient of variation) for 3-HPMA and 11.0% for HMPMA. The method was applied to more than 2600 urine samples from smokers; mean levels of 3-HPMA and HMPMA were 4800±5358 (S.D.)pmol/mL and 3302±3341pmol/mL, respectively.

Tobacco-related cardiovascular diseases in the 21st century.

HomeArteriosclerosis, Thrombosis, and Vascular BiologyVol. 33, No. 7Tobacco-Related Cardiovascular Diseases in the 21st Century Free AccessResearch ArticlePDF/EPUBAboutView PDFView EPUBSections ToolsAdd to favoritesDownload citationsTrack citationsPermissions ShareShare onFacebookTwitterLinked InMendeleyReddit Jump toFree AccessResearch ArticlePDF/EPUBTobacco-Related Cardiovascular Diseases in the 21st Century Muredach P. Reilly Muredach P. ReillyMuredach P. Reilly From the Cardiovascular Institute, The Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA. Search for more papers by this author Originally published1 Jul 2013https://doi.org/10.1161/ATVBAHA.113.301632Arteriosclerosis, Thrombosis, and Vascular Biology. 2013;33:1458–1459Cigarette smoking activates platelets, induces endothelial inflammation, and promotes vascular dysfunction leading to accelerated atherosclerosis, plaque rupture, acute coronary syndromes, and death. Over several decades, notable advances in public policy in the United States have reduced initiation and increased cessation of cigarette smoking, thus lowering the prevalence of tobacco product consumption and related cardiovascular disease (CVD). Indeed, most people in developed countries understand the harm of cigarette smoking and the health and economic benefits of abstinence and cessation. However, these public health successes have slowed recently,1 reflecting perhaps a higher degree of addiction in remaining smokers and also a degree of fatigue in further advancing preventive and treatment strategies, as well as scientific knowledge more generally.Several realities underscore the current challenges to further reducing tobacco-related CVD. First, rates of tobacco product use continue to increase in developing nations and, in combination with dietary and additional lifestyle changes, will drive a surge of CVD in the coming decades in these societies. Second, even in developed countries, tobacco use continues to be stubbornly prevalent particularly in minorities, in patients with psychiatric disorders, and in economically disadvantaged populations and represents a major cause of preventable morbidity and mortality. Last, and in some respects of most direct concern to our research community, mechanistic insights into how tobacco consumption causes CVDs and how this affects individual risk have not kept pace with epidemiological or public health advances. Although we have gained some knowledge of the genetic basis for cigarette smoking behavior and nicotine addiction,2 we have an incomplete knowledge at the molecular level of the processes that promote CVD, have learned little regarding individualized and genetic risk of CVD complications in those exposed, and have surprisingly limited knowledge of the specific constituents of cigarette smoke that lead to CVD.3 Thus, we are poorly positioned to apply mechanism-based strategies to treat smoking-related CVD or to execute preventive strategies in those at greatest risk.In this miniseries, we focus on mechanistic aspects of tobacco-related CVD as a focus for galvanizing scientific efforts for the 21st century global epidemic challenge. Norman and Curci highlight how cigarette smoking dominates as a risk factor for abdominal aortic aneurysm and emphasize recent experimental insights but acknowledge our incomplete mechanistic understanding of abdominal aortic aneurysm in smokers. Barau and Ambrose provide an update on the role and potential mechanisms of cigarette smoking in platelet activation, thrombosis, acute cardiovascular events, and modulation of therapeutic interventions in coronary thrombosis. Breitling emphasizes the paucity of contemporary genetic studies in understanding the interindividual risk of developing CVD in cigarette smokers, highlights promising epigenetic discoveries, and reviews the priorities for large-scale and rigorous genomic approaches to smoking-related CVD. In a future issue of the journal, additional reviews in this series will focus on tobacco-related endothelial dysfunction and atherosclerosis, as well as present an integrated view of CVD in the context of nicotine addiction, public policies, and global health.We are now at a critical juncture in the battle against tobacco-related CVD. A striking observation in editing this miniseries was the relative dearth of recent mechanistic and genomic studies of smoking-related CVD. This failure to advance mechanistic and genomic understanding of smoking-related CVD at an individual level coupled with unchallenged marketing and an increasing surge in tobacco product consumption in developing countries may severely challenge our ability to mount effective preventive strategies and manage the global epidemic of related CVD. This series marks current knowledge, highlights deficiencies, and defines the challenge for future mechanistic, therapeutic, and public policy requirement for prevention and treatment of tobacco-related CVDs in the 21st century.Sources of FundingDr Reilly is supported by R01-HL-111694, R01-HL-113147, U01-HL-108636, and K24-HL-107643 from the National Institutes of Health.DisclosuresNone.FootnotesCorrespondence to Muredach P. Reilly, MB, MS, Cardiovascular Institute, The Perelman School of Medicine, University of Pennsylvania, 11-136 Smilow Center for Translational Research, 3400 Civic Center Blvd, Bldg 421, Philadelphia, PA 19104. E-mail [email protected]

Preparation of phenyl group-functionalized magnetic mesoporous silica microspheres for fast extraction and analysis of acetaldehyde in mainstream cigarette smoke by gas chromatography–mass spectrometry

Acetaldehyde is regarded as a toxic mainstream cigarette smoke constituent, and measurement of acetaldehyde in complex real samples is difficult owing to its high volatility and reactivity. In this work, phenyl group-functionalized magnetic mesoporous microspheres were developed as the solid-phase extraction sorbents for enrichment and analysis of acetaldehyde in mainstream cigarette smoke. The functional magnetic microspheres were first synthesized through a facile one-pot co-condensation approach. The prepared nanomaterials possessed abundant silanol groups in the exterior surface and numerous phenyl groups in the interior pore-walls, as well as a large surface area (273.5m2/g), strong superparamagnetism and uniform mesopores (3.3nm). Acetaldehyde in mainstream cigarette smoke was collected in water and derivatizated with O-2,3,4,5,6-(pentafluorobenzyl)hydroxylamine. The formed acetaldehyde oximes were extracted and enriched by the prepared adsorbents via π–π interactions and subsequently analyzed using GC–MS. Extraction conditions such as amounts of sorbents, eluting solvent, adsorption and desorption time were investigated and optimized to achieve the best efficiency. Method validations including linearity, recovery, repeatability, and limit of detection were also studied. It was found that the suggested methodology provided low detection limit of 0.04mg/mL, good recovery of 88–92%, intra-day and inter-day RSD values of 4.5% and 10.1%, and linear range of 0.25–4mg/mL (R2=0.999). The results indicated that the proposed method based on phenyl-functionalized magnetic mesoporous microspheres was rapid, efficient and convenient for the enrichment and analysis of acetaldehyde in tobacco.

The cigarette smoke constituent benzo[a]pyrene disrupts metabolic enzyme, and apoptosis pathway member gene expression in ovarian follicles

Benzo[a]pyrene (B[a]P) is a prototypical polycyclic aromatic hydrocarbon (PAH) present in cigarette smoke. We previously showed that B[a]P adversely affects follicular development and survival. The objective of this study was to identify the key molecular pathways underlying B[a]P-induced abnormal follicular development. Isolated follicles (100–130μm) from ovaries of F1 hybrid (C57BL/6j×CBA/Ca) mice were cultured for 8 (preantral/antral follicles) and 12 (preovulatory follicles) days in increasing concentrations of B[a]P (0ng/mL [control] to 45ng/mL). Expression of aryl hydrocarbon receptor (AhR), aryl hydroxylase steroidogenic enzyme, cell-cycle, and apoptotic genes were quantified. B[a]P exposure significantly (P<0.05) increased mRNA expression of Cyp1a1 in preantral/antral follicles and Cyp1b1, Bax and Hsp90ab1 in preovulatory follicles. No significant effect on mRNA expression of StAR, Cyp11a1, aromatase, Cdk4, Cdk2, Ccnd2, cIAP2, and survivin was observed. In conclusion, this study suggests that B[a]P exposure significantly affects the phase I enzymes and cell death genes during preantral/antral and preovulatory growth, and thus highlight the AhR signaling and apoptotis pathways in delayed follicle growth and decreased viability.

Airway Vascular Endothelial Function in Healthy Smokers Without Systemic Endothelial Dysfunction

Cigarette smoking can lead to systemic endothelial dysfunction. Since the airway circulation is exposed to a high concentration of cigarette smoke constituents, we reasoned that airway vascular endothelial dysfunction could be present in healthy smokers without systemic endothelial dysfunction. The purpose of this study was to compare airway and systemic endothelial function and measure markers of systemic inflammation in lung-healthy current smokers. Since endothelial dysfunction in smokers has been related to systemic inflammation, we also investigated its response to an inhaled glucocorticosteroid (ICS). Fifteen healthy, current smokers and 17 healthy, lifetime nonsmokers were enrolled. Smokers were randomly assigned to 3-week treatments with inhaled fluticasone propionate or placebo in a crossover design. Vascular endothelial function was assessed in the airway by the airway blood-flow response to inhaled albuterol (ΔQaw) and in the extrapulmonary circulation by brachial arterial flow-mediated vasodilation (FMD). Venous blood was collected for C-reactive protein and IL-6. Baseline parameters did not differ between groups except for ΔQaw, which was greater in nonsmokers (45% ± 12%) than smokers (1% ± 12%) (P = .001). In the smokers, ICS treatment increased Qaw to 41% ± 7% (P &lt; .001), but had no effect on FMD or inflammatory markers. There was an inverse relationship between baseline and ICS-induced changes in ΔQaw. Healthy smokers with no signs of systemic inflammation or endothelial dysfunction display impaired airway vascular endothelial function, possibly preceding systemic endothelial dysfunction. Airway endothelial function was restored with an ICS, and the response was directly related to the severity of endothelial dysfunction.

Methodologies for the quantitative estimation of toxicant dose to cigarette smokers using physical, chemical and bioanalytical data

Methodologies have been developed, described and demonstrated that convert mouth exposure estimates of cigarette smoke constituents to dose by accounting for smoke spilled from the mouth prior to inhalation (mouth-spill (MS)) and the respiratory retention (RR) during the inhalation cycle. The methodologies are applicable to just about any chemical compound in cigarette smoke that can be measured analytically and can be used with ambulatory population studies. Conversion of exposure to dose improves the relevancy for risk assessment paradigms. Except for urinary nicotine plus metabolites, biomarkers generally do not provide quantitative exposure or dose estimates. In addition, many smoke constituents have no reliable biomarkers. We describe methods to estimate the RR of chemical compounds in smoke based on their vapor pressure (VP) and to estimate the MS for a given subject. Data from two clinical studies were used to demonstrate dose estimation for 13 compounds, of which only 3 have urinary biomarkers. Compounds with VP > 10−5 Pa generally have RRs of 88% or greater, which do not vary appreciably with inhalation volume (IV). Compounds with VP < 10−7 Pa generally have RRs dependent on IV and lung exposure time. For MS, mean subject values from both studies were slightly greater than 30%. For constituents with urinary biomarkers, correlations with the calculated dose were significantly improved over correlations with mouth exposure. Of toxicological importance is that the dose correlations provide an estimate of the metabolic conversion of a constituent to its respective biomarker.

Abstract 1297: SETD6-mediated deregulation of NF- κB signaling in 4-ABP-induced bladder carcinogenesis.

Abstract Epidemiological data support an etiological role for chemicals from occupational exposure and cigarette smoke with increased risk of bladder cancer. However, epigenetic effects of these chemicals remain to be elucidated. Protein methyltransferases (PMTs) methylate protein lysine and arginine residues and play an important role in regulating gene transcription. Deregulation of SET-domain-containing PMTs has a crucial role in carcinogenesis. SETD6 has been shown to monomethylate RelA (a principal subunit of NF-κB at Lys310). Monomethylated RelA recruits GLP (G9A-like protein), a histone dimethyl transferase to the NF-κB target genes, which results in transcriptional repression. The role of SETD6 in bladder cancer is unknown. We used qPCR and immunoblotting to determine changes in SETD6 at the mRNA and protein level in bladder cancer cells relative to primary urothelial cells. We also found that knockdown of SETD6 significantly decreases the viability and proliferation of bladder cancer cells suggesting that SETD6 may function as an oncogene in bladder cancer. Furthermore, we also found that message and protein levels of SETD6 are induced in primary urothelial cells following treatment of 4-aminobiphenyl (4-ABP), an important constituent of cigarette smoke. Preliminary results indicate that induction may be mediated by 4-ABP-induced ROS. RelAK310Me1 level also increased with 4-ABP treatment showing the increase in SETD6 functional activity with 4-ABP exposure. We also found the decrease in expression of RelB, one of the target genes of RelA. Overall our results show that SETD6 may function as an oncogene in 4-ABP induced bladder cancer by deregulating transactivation property of RelA. These results show for the first time the relevance of PKMTs and epigenetic regulation of NF-κB in carcinogen induced bladder cancer. Supported by RO3 CA136058 (RG). Citation Format: Neelam Mukherjee, Rita Ghosh. SETD6-mediated deregulation of NF- κB signaling in 4-ABP-induced bladder carcinogenesis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1297. doi:10.1158/1538-7445.AM2013-1297

β-Cryptoxanthin Restores Nicotine-Reduced Lung SIRT1 to Normal Levels and Inhibits Nicotine-Promoted Lung Tumorigenesis and Emphysema in A/J Mice

Nicotine, a large constituent of cigarette smoke, is associated with an increased risk of lung cancer, but the data supporting this relationship are inconsistent. Here, we found that nicotine treatment not only induced emphysema but also increased both lung tumor multiplicity and volume in 4-nitrosamino-1-(3-pyridyl)-1-butanone (NNK)-initiated lung cancer in A/J mice. This tumor-promoting effect of nicotine was accompanied by significant reductions in survival probability and lung Sirtuin 1 (SIRT1) expression, which has been proposed as a tumor suppressor. The decreased level of SIRT1 was associated with increased levels of AKT phosphorylation and interleukin (il)-6 mRNA but decreased tumor suppressor p53 and retinoic acid receptor (RAR)-β mRNA levels in the lungs. Using this mouse model, we then determined whether β-cryptoxanthin (BCX), a xanthophyll that is strongly associated with a reduced risk of lung cancer in several cohort studies, can inhibit nicotine-induced emphysema and lung tumorigenesis. We found that BCX supplementation at two different doses was associated with reductions of the nicotine-promoted lung tumor multiplicity and volume, as well as emphysema in mice treated with both NNK and nicotine. Moreover, BCX supplementation restored the nicotine-suppressed expression of lung SIRT1, p53, and RAR-β to that of the control group, increased survival probability, and decreased the levels of lung il-6 mRNA and phosphorylation of AKT. The present study indicates that BCX is a preventive agent against emphysema and lung cancer with SIRT1 as a potential target. In addition, our study establishes a relevant animal lung cancer model for studying tumor growth within emphysematous microenvironments.

PP007. Nicotinic acetylcholine receptor subunits in the pre-eclamptic and cigarette smoke exposed human placenta

Cigarette smoking during pregnancy is associated with low birth weight, prematurity and neonatal morbidity and mortality. Nicotine, a major constituent of cigarette smoke, binds to nicotinic acetylcholine receptors (nAChRs). To date, 16 mammalian nAChR subunits have been identified. The effect of smoking on these subunits in human placenta has not yet been determined. Smoking is also associated with reduced pre-eclampsia (PE) risk and its protective effects may occur via changes in nAChRs. This study aimed to determine which nAChR subunits are present in the normal human placenta, and whether any changes are occur from smoking or PE. Using RT-qPCR, all 16 nAChR subunits were investigated in normal, healthy human placentas, and mRNA expression compared between controls (n=8), smokers (n=8) and PE (n=7). Results All 16 nAChR subunits were expressed in healthy placentas. Smoke exposure significantly increased α2 (p=0.006) and α9 (p=0.038), and decreased δ (p=0.013), subunit mRNA expression. In PE placentas, β1 (p=0.048) and β2 (p=0.031) subunit mRNA expression was increased. Nicotine exposure in pregnancy increases nAChR subunit mRNAs that play a role in vasoconstriction and amino acid uptake, possibly contributing to abnormalities in placentas from smoking mothers. Different subunits were affected in PE placentas, thus the hypoxic environment may induce changes in endogenous ACh levels, yielding compensatory increases in β1 and β2 subunits.

Paracrine potential of fibroblasts exposed to cigarette smoke extract with vascular growth factor induction

Nicotine, a major constituent of cigarette smoke, can activate the cholinergic anti-inflammatory pathway by binding to α7-nicotinic acetylcholine receptor (α7nAChR) expressed on the surface of certain cells. Here, we ask whether cigarette smoke extract induced different paracrine factors compared to the in vivo regulator of inflammation, tumor necrosis factor-α, in human vocal fold fibroblasts (hVFFs) shown to express low levels of α7nAChR. In vitro. α7nAChR was detected by nested polymerase chain reaction and immunohistochemistry. γH2AX, a marker for DNA double-stand breaks, was measured by immunofluorescence. Cigarette smoke extract was prepared in accordance with investigators studying effects of cigarette smoke. hVFFs treated for 3 hours had media replaced for an additional 24 hours. Cytokine, chemokine, and growth factor levels in media were assessed by multiplex analysis. α7nAChR expression levels decreased with the passage number of fibroblasts. Tumor necrosis factor-α induced a significantly different profile of cytokines, chemokines, and growth factor compared to cigarette smoke extract exposure. Cigarette smoke extract at a concentration not associated with induction of γH2AX nuclear foci significantly increased vascular endothelial growth factor. Cigarette smoke extract elicited a response important for regulation of angiogenesis and vascular permeability during inflammation, without evidence of DNA double-stand breaks associated with carcinogenesis. hVFFs are capable of participating in paracrine regulation of pathological blood vessel formation associated with cigarette smoking-related diseases (ie, Reinke edema). These cells express α7nAChR, an essential component of the cholinergic anti-inflammatory pathway regulated by the vagus nerve in certain tissues and a target of therapeutic agents.

Quantitative Determination of Menthol in a Single Puff of Mainstream Cigarette Smoke

Abstract A method for the determination of menthol in a single puff of mainstream cigarette smoke is reported. A rotary smoking machine with a twin filter interface was used to smoke cigarettes with varying menthol and “tar” deliveries determined based on the Cambridge filter method. The twin filter interface mechanically switches to a new filter pad for collection of smoke from each cigarette puff. The individual filter pad extracts were analyzed by gas chromatography-mass spectrometry (GC-MS) to determine menthol puff-by-puff deliveries. Menthol puff-by-puff profiles show an increase in menthol smoke delivery with increasing puff count, a trend consistent with total particulate matter and smoke delivery profiles of other mainstream smoke constituents. The sum of single puff menthol deliveries is comparable to the whole cigarette menthol smoke delivery as collected on a single filter pad. This method can also determine quantitative puff-by-puff deliveries of other mainstream cigarette smoke constituents.

The biology behind the atherothrombotic effects of cigarette smoke

Cigarette smoke is an aerosol that contains >4,000 chemicals, including nicotine, carbon monoxide, acrolein, and oxidant compounds. Exposure to cigarette smoke induces multiple pathological effects in the endothelium, several of which are the result of oxidative stress initiated by reactive oxygen species, reactive nitrogen species, and other oxidant constituents of cigarette smoke. Cigarette-smoke exposure interferes adversely with the control of all stages of plaque formation and development and pathological thrombus formation. The reactive oxygen species in cigarette smoke contribute to oxidative stress, upregulation of inflammatory cytokines, and endothelial dysfunction, by reducing the bioavailability of nitric oxide. Plaque formation and the development of vulnerable plaques also result from exposure to cigarette smoke via the enhancement of inflammatory processes and the activation of matrix metalloproteases. Moreover, exposure to cigarette smoke results in platelet activation, stimulation of the coagulation cascade, and impairment of anticoagulative fibrinolysis. Many cigarette-smoke-mediated prothrombotic changes are quickly reversible upon smoking cessation. Public health efforts should urgently promote our understanding of current cigarette-smoke-induced cardiovascular pathology to encourage individuals to reduce their exposure to cigarette smoke and, therefore, the detrimental consequences of associated atherothrombotic disease.

Formation of mainstream cigarette smoke constituents prioritized by the World Health Organization – Yield patterns observed in market surveys, clustering and inverse correlations

The WHO TobReg proposed mandating ceilings on selected smoke constituents determined from the market-specific median of nicotine-normalized yield distributions. Data validating this regulatory concept were obtained from essentially single-blend surveys. This process is strongly impacted by inverse correlations among yields. In the present study, 18 priority WHO smoke constituent yields (nicotine-normalized) were determined (using two smoking regimens) from 262 commercial brands including American, Virginia and local blends from 13 countries. Principal Component Analysis was used to identify yields patterns, clustering of blend types and the inverse correlations causing these clusters. Three principal components explain about 75% of total data variability. PC1 was sensitive to the relative levels of gas- and particle-phase compounds. PC2 and PC3 cluster American- and Virginia-blends, revealing inverse correlations: Nitrogen oxides and amino- or nitroso-aromatic compounds inversely correlate to either formaldehyde and acrolein, or benzo(a)pyrene and di-hydroxybenzenes. These results can be explained by reviewing the processes determining each components smoke delivery. Regulatory initiatives simultaneously targeting selected smoke constituents in markets with mixed blend styles will be strongly impacted by the inverse correlations described. It is difficult to predict the ultimate impact of such regulations on public health, considering the complex chemistry of cigarette smoke formation.

Development of microwave-assisted headspace solid-phase microextraction followed by gas chromatography-mass spectrometry for the analysis of phenol in a cigarette pad

Phenol is a mainstream cigarette smoke constituent and has been classified as a main toxic component directly related to environmental and health issues. Analysis of phenol in mainstream cigarette smoke is of great importance. In this work, microwave-assisted headspace solid-phase microextraction (HS-SPME) was developed for the fast analysis of phenol in a cigarette pad. Analyte in a Cambridge cigarette pad, activated by a short period of microwave irradiation, was headspace extracted by SPME and analyzed by GC-MS. Extraction conditions such as microwave irradiation time, SPME fiber coating, adsorption time and desorption time were investigated and optimized to achieve the best effect. The method was validated through the investigation of linearity, detection limit, recovery and precision. The linearity was in a wide range of 0.005–1 μg mL−1 with a correlation coefficient R2 = 0.999. The limit of detection was as low as 0.5 ng mL−1, and acceptable recovery and RSD values of 86.5% and 11.5% were achieved. The proposed method was successfully applied to the analysis of phenol in cigarette pad samples. The experimental results have demonstrated that microwave-assisted HS-SPME followed by GC-MS was a rapid, efficient and convenient method for the determination of phenol in tobacco products.

Synthesis of Vegetable-Based Activated Carbons with Mixed Micro- and Mesoporosity for Use in Cigarette Filters

Activated carbons with micropores for adsorption and filtration of the volatile constituents of mainstream cigarette smoke, together with mesopores for enhanced mass transport were prepared by a novel route. Treatment of coconut shell or other lignocellulosic precursors with aqueous NaOH, followed by thorough washing, charring and steam activation produced carbons with enhanced adsorption characteristics in smoking trials, compared with their microporous analogues. The mechanism of formation of these carbons is explained in terms of initial partial dissolution of the precursor in an aqueous alkali solution, followed by catalytic gasification of carbon in steam involving residual sodium.