Pharmacodynamic effects of chronic cigarette smoke exposure in spontaneously hypertensive rats

In investigating the influence of chronic cigarette smoke exposure on hypertension, we compared the pharmacodynamic effects of enforced exposure to smoke on spontaneously hypertensive rats (SHR) with those on Wistar-Kyoto (WKY) rats. Chronic cigarette smoke exposure for 8 weeks decreased the elevated heart rate of mature male SHR to approximately the rate in WKY rats 24 h after smoke exposure. Both systolic and diastolic blood pressures also decreased slightly. However, WKY rats showed a marked rise in heart rate soon after exposure to cigarette smoke began, with no change in blood pressure, while the heart rate of SHR in the early stage remained similar to that of animals without exposure, although their blood pressure was clearly reduced. The body weight of both strains tended to decrease during smoke exposure, but the effect was more severe in SHR. Moreover, the effects of chronic smoke exposure were observed using retired, aged female SHR breeders. A decrease in body weight and heart rate, but not in blood pressure, was also recognized even in these mature animals. These effects gradually recovered after withdrawal from exposure. On the basis of these results, a profile of chronic cigarette smoke exposure under hypertension is discussed in this study.     

Pharmacokinetics of nicotine in rats after multiple-cigarette smoke exposure

The pharmacokinetics of nicotine and its major metabolites was evaluated in male rats after multiple-cigarette smoke exposure. A smoke-exposure apparatus was used to deliver cigarette smoke to the exposure chamber. The rats were exposed to smoke from a single cigarette every 8 hr for 14 days and to the smoke of a cigarette spiked with radiolabeled nicotine on the 15th day. Blood and urine samples were collected at timed intervals during the 10-min smoke-exposure period of the last cigarette and up to 48 hr thereafter. Nicotine, cotinine, and other polar metabolites were separated by thin-layer chromatography and quantified by liquid scintillation counting. The data were analyzed by computer fitting, and the derived pharmacokinetic parameters were compared to those observed after a single iv injection of nicotine and after a single-cigarette smoke exposure. The results indicated that the amount of nicotine absorbed from multiple-cigarette smoke was approximately 10-fold greater than that absorbed from a single cigarette. Also, unlike the single-cigarette smoke exposure experiment, nicotine plasma levels did not decay monotonically but increased after the 5th hr, and high plasma concentrations persisted for 30 hr. The rate and extent of the formation of cotinine, the major metabolite of nicotine, were decreased as compared with their values following a single-cigarette smoke exposure. It was concluded that nicotine or a constituent of tobacco smoke inhibits the formation of cotinine and may affect the biotransformation of other metabolites. Urinary excretion tended to support the conclusions that the pharmacokinetic parameters of nicotine and its metabolites were altered upon multiple as compared to single dose exposure.     

Influences of exposure to cigarette smoke on concentration of nicorandil in plasma of rats.

Influences of acute exposure to cigarette smoke on plasma concentrations of nicorandil administered orally and parenterally were investigated in rats by HPLC. The animals were exposed to tobacco smoke of two kinds of cigarettes using a smoking machine (i.e., the cigarette smoke contained either low or high nicotine and tar). The plasma concentration of nicorandil administered orally at a dose of 10 mg/kg had a lower absorption phase in two cigarette smoke-exposed groups, particularly in the high nicotine and tar-containing cigarette smoke-exposed group, compared with the nonsmoking control group. The AUC and MRT values in a high nicotine and tar-containing cigarette smoke-exposed group were lower and higher, respectively, than in the nonsmoking control group. However, there was no marked difference in nicorandil plasma concentrations between the cigarette smoke-exposed group and the nonsmoking control group when nicorandil was administered ip or iv at a dose of 5 mg/kg. These results suggest that cigarette smoke exposure causes the suppression or delay of absorption of nicorandil from the gastrointestinal tract.     

The effect of Bitis gabonica (Gaboon viper) snake venom on external iliac and mesenteric arterial circulation in the dog

The effects of Gaboon viper (Bitis gabonica) venom on external iliac and mesenteric arterial blood flow and resistance were investigated in eight anaesthetized, close-chest dogs. Venom doses in the range 0.125 – 0.5 mg/kg produced a profound fall in external iliac and mesenteric arterial resistance, which recovered to control values after 30 min. After a third dose of venom, the mean arterial blood pressure failed to recover and the animals died after a period of severe hypotension. External iliac arterial blood flow rose concomitantly with the fall in external iliac resistance and decreased to a value significantly below control after 30 min. Paradoxically, mesenteric blood flow fell during the period of vasodilation. The results suggest that widespread vasodilation of muscle vascular beds (of which the external iliac circulation is representative) leads to shunting of blood away from the less-dilated mesenteric circulation. Venom-induced peritoneal haemorrhage caused a fall in blood volume and increase in viscosity. These undoubtedly contributed to the severe haemodynamic deterioration of the preparations after the third injection of the venom.     

Nicotine effect on cardiovascular system and ion channels

Smoking is a leading cause of cardiovascular disease, hypertension, myocardial infarction, and stroke. Nicotine is one of the components of cigarette smoke. Nicotine effects on the cardiovascular system reflect the activity of the nicotine receptors centrally and on peripheral autonomic ganglia. It has been found that cigarette smoke extract-induced contraction of porcine coronary arteries is related to superoxide anion-mediated degradation of nitric oxide. Treatment of rabbit aortas with an oxygen free radicals scavenger attenuated cigarette smoke impairment of arterial relaxation. Treatment of smokers with vitamin C, an antioxidant, improved impaired endothelium-dependent reactivity of large peripheral arteries. Thus it appears that chronic smoking and acute exposure to cigarette smoke extract may alter endothelium-dependent reactivity via the production of oxygen derived free radicals. This review discusses the effects of nicotine on resistance arterioles, compliance arteries, smooth muscle cells, and ion channels in the cardiovascular system. We discuss studies performed on humans, nicotine-exposed animals, and cell cultures yielding varying and inconsistent results that may be due to differences in experimental design, species, and the dose of exposure. Nicotine exposure appears to induce a combination of free radical production, vascular wall adhesion, and a reduction of fibrinolytic activity in the plasma.     

Up-regulation of nicotinic acetylcholine receptors following chronic exposure of rats to mainstream cigarette smoke or α4β2 receptors to nicotine

Smokers are reported to have a higher density of central nicotinic acetylcholine receptors (nAChRs) that non-smokers at autopsy. Whether this increased receptor density is a response to smoking or a result of genetic variability is not known. While sub-chronic treatment of rats and mice with nicotine results in upregulation of central nAChRs, changes in receptor density in response to cigarette smoke have not been studied previously. In this study, male Sprague-Dawley rats were exposed nose-only for 13 weeks to mainstream cigarette smoke followed by assessment of [³H]nicotine binding in five brain regions of smoke- and sham-exposed animals. In smoke-exposed animals, there was a significant increase in nAChR density in the cortex, striatum, and cerebellum (35, 25, and 31% increases, respectively), while there was no significant change in receptor density in the thalamus and hippocampus. Smoke exposure did not alter markedly the affinity of the receptor for nicotine in these brain regions. Furthermore, up-regulation of nAChRs did not alter the biphasic binding properties by which nicotine binds to its receptor. There were no changes in the association (fast phase) or isomerization (slow phase) rate constants, and the percent contribution of slow and fast phase binding to nAChRs was not altered in the up-regulated receptor population compared with control. Similar results were observed following chronic nicotine exposure of cultured cortical cells from fetal rat brain or cells transfected with the α4β2 nAChR subtype. These results show that the up-regulation following smoke exposure in the rat is phenomenologically similar to that observed in vitro. These data provide preliminary evidence for a relationship between cigarette smoking and nAChR up-regulation in vivo and suggest that similar mechanisms of upregulation may underlie chronic smoke exposure of live animals and nicotine exposure of artificially expressed α4β2 receptors in vitro.     

Effect of cigarette smoking on hepatic biotransformations in rats

Adult male rats were nose-only exposed to cigarette smoke for 20 min each day for 6 months. Smoke inhalation was confirmed by urinary excretion of nicotine (4.5 μg/rat/day) and elevated blood carboxyhemoglobin (13.3%). Phenobarbital- and 3-methylcholanthrene-treated rats were included as positive controls for assessing hepatic enzyme induction. Cigarette smoke did not induce statistically significant alterations in hepatic enzyme activity when measured in vivo (pentobarbital sleep time and zoxazolamine paralysis time) or in vitro (oxidation, hydrolysis, glucuronidation, and glutathione conjugation). In some cases, the smoke-exposed rats did exhibit higher microsomal enzyme activity than did the controls, but this increase was also evident in the sham-control group. Therefore, these increases were attributed to stress and not to smoking per se. Additional evidence of stress associated with manipulation of the animals was the smaller percentage weight gains of the smoke-exposed and sham control rats over the 6-month period as compared to the controls (12, 35, and 50%, respectively). Smoking reduced hepatic glutathione by as much as 15%, but the glutathione transferase activity was not affected. These studies showed that chronic exposure of rats to cigarette smoke did not alter hepatic biotransformation processes, but do suggest that smokers may be less efficient than nonsmokers in the deactivation of xenobiotics by glutathione conjugation mechanisms.     

Histopathology,Urine Mutacenicity,and Bone Marrow Cytocenetics of Mice Exposed Nose-Only to Smoke from Cigarettes that Burn or Heat Tobacco

Abstract

Male and female B6C3F₁ mice were exposed nose-only to smoke from a test cigarette that heats tobacco, or from a reference cigarette that burns tobacco. Cigarette smoke was generated by a smoking machine, and the concentrations of wet total particulate matter (WTPM) were adjusted to 0, 0.16, 0.32, or 0.64 mg/l. Exposures were performed 1 h/day for 14 consecutive days. Urine mutagenicity was assessed by a modified Ames bacterial assay Clastogenesis (sister-chromatid exchanges, chromosome aberrations, and micronuclei) was evaluated in bone marrow cells. Respiratory rate was depressed significantly by exposure to smoke from the reference cigarette, but not the test. Blood carboxyhemoglobin, plasma nicotine, and plasma cotinine showed exposure-dependent increases in the smoke-exposed animals. Histopathological changes similar to those noted previously in smoke-exposed rats were noted, with fewer and less pronounced changes in the animals exposed to smoke from the test cigarette when compared with the reference. Positive urine mutagenicity and clastogenic responses were observed in the animals treated with positive control chemicals. However the urine mutagenicity and clastogenic responses of smoke-exposed animals (both cigarette types) were not different from those of sham-exposed animals, except for the micronucleus assay where animals exposed to high concentrations of reference cigarette smoke showed a significant increase over controls.     

Sequential changes in the structure of the rat respiratory system during and after exposure to cigarette smoke

The effect of twice daily exposure to diluted cigarette smoke on the structure of the respiratory system was examined in rats exposed for up to 84 days. Changes in respiratory tract structure were also determined in animals which were exposed for 42 days and then left untreated for an equal length of time. Daily food consumption and growth rate were reduced in sham-smoked and in smoke-exposed rats compared with cage controls. When both these treatments were stopped, food consumption and growth rate increased. Goblet cell hyperplasia of tracheal and bronchial epithelia, increased numbers of alveolar macrophages, squamous metaplasia, and hyperplasia of the larynx were seen in rats after 2 weeks of exposure to smoke. Except for tracheal goblet cell hyperplasia, within 14 to 42 days of exposure commencing all these changes showed a maximal observed response which was subsequently maintained as exposures continued up to 84 days. Tracheal goblet cell hyperplasia and alveolar metaplasia increased progressively during this extended exposure period. When exposure to smoke was discontinued after 42 days, larynx, trachea, and bronchus all reverted to normal at varying rates. The incidence, but not the severity, of the alveolar metaplasia induced during the smoke-exposure period continued to increase when animals were not being exposed to smoke.     

Chronic cardiovascular effects of bepridil in anesthetized and conscious dogs

The chronic cardiovascular effects of bepridil were investigated in pentobarbital-anesthetized dogs following chronic administration for 2 weeks and in awake instrumented dogs for 5 days. Prolonged administration of bepridil, 10 mg/kg, p.o., twice a day for 14 days produced a significant reduction in heart rate, cardiac output, and left ventricular minute work in dogs anesthetized approximately 12 hr following the last dose of the compound. Bepridil significantly increased coronary blood flow, whereas renal and mesenteric flows were not altered. Myocardial contractility was not significantly altered; however, left ventricular stroke work curves obtained during acute volume loading shifted to the left, indicating enhanced myocardial performance. Oral administration of bepridil twice daily produced a significant reduction in mean arterial blood pressure, a significant decrease in left ventricular stroke work and minute work, and myocardial oxygen consumption in awake animals. In addition, the compound decreased total peripheral resistance (TPR) in awake animals, whereas TPR was increased in anesthetized dogs.     

Plasma nicotine and cotinine in tobacco smoke exposed beagle dogs

Nicotine and cotinine have been determined in plasma samples from 87 beagle dogs chronically exposed to cigarette smoke with three different levels of nicotine. An additional 18 sham-exposed animals were included in the study as controls. Smoke was administered to the animals through permanent tracheostomas via cuffed tracheostomy tubes and was generated from reference cigarettes under standard puffing parameters by ADL-II smoking machines. The dogs were exposed for an average of 2 years prior to sample collection. The results from blood samples collected at specific intervals in the daily exposure schedules indicate that nicotine may be useful as a relative index of smoke exposure. At elevated exposure levels, average blood concentrations were related to the number of cigarettes smoked as well as the nitocine delivery of the cigarette. Cotinine was found to increase more slowly than nicotine and was also metabolized more rapidly than in humans. Overall, the study affords an examination of the relationship of plasma nicotine and cotinine with estimated nicotine exposure.     

Cigarette smoke causes inhibition of the immune response to intratracheally administered antigens

Chronic inhalation of cigarette smoke in rats preferentially inhibited the plaque-forming cell (PFC) response of lung-associated lymph nodes (LALN) to sheep red blood cells (SRBC), compared to anatomically distant lymph nodes. Inhibition of the antibody response in LALN of smoke-exposed animals was first detected at 21 weeks of smoke inhalation and was well established by the 27th week of smoke exposure. After prolonged exposure (greater than 34 weeks) to cigarette smoke, similar smoke-induced changes in PFC response took place in other lymphoid tissues as well. Cigarette smoke affected the response of LALN cells to a T cell-dependent antigen (SRBC). Exposure to cigarette smoke, however, did not alter the relative percentages of W3/13-positive (T cells) or Ig-positive (B cells) cells, nor did it alter the relative percentages of T cell subsets as scored by their surface phenotypes, i.e., T helper (W3/25+) or T suppressor/cytotoxic (OX-8+) cells. The percentage of phagocytic cells and the accessory cell functions of macrophages remained comparable between sham and smoke-exposed animals. Exposure to cigarette smoke did not significantly alter the response of LALN cells to T cell mitogens (concanavalin A and phytohemagglutinin). However, response to a T cell-independent antigen trinitrophenyl Brucella abortus was also significantly reduced. These results show that cigarette exposure in the rat results in a decreased antibody response and this exposure to cigarette smoke may primarily affect the B cell function.     

Subchronic inhalation by rats of mainstream smoke from a cigarette that primarily heats tobacco compared to a cigarette that burns tobacco

A subchronic, nose-only inhalation study comparing the potential biological activity of mainstream smoke from a cigarette that primarily heats tobacco (Eclipse) to mainstream smoke from a 1R4F reference cigarette was conducted using Sprague-Dawley rats of each gender. Smoke exposures were for 1 h/day, 5 days/wk for 13 wk, at concentrations of 0, 0.16, 0.32, or 0.64 mg wet total particulate matter (WTPM)/L air. Smoke was generated at the Federal Trade Commission standard of a 2-s puff of 35 ml, taken once per minute. Clinical signs, body and organ weights, clinical chemistry, hematology, carboxyhemoglobin, serum nicotine, plethysmography, gross pathology, and histopathology were determined. Plethysmography indicated that respiratory rate was decreased at all concentrations of 1R4F smoke, but only at the high concentration of Eclipse smoke. Tidal volume was depressed and minute volume was lower for all smoke-exposed rats. Rats exposed to Eclipse smoke inhaled more smoke at the low and mid-concentration exposures than rats exposed to equivalent concentrations 1R4F smoke. Carboxyhemoglobin and serum nicotine were directly related to the exposure concentrations of carbon monoxide (CO) and nicotine in an exposure-dependent manner. Body weights were slightly lower in smoke-exposed rats, while no treatment-related effects were seen in clinical signs, clinical chemistry, hematology, or gross changes at necropsy. The only treatment-related effect seen in organ weights was an increase in heart weight in females in the Eclipse high-concentration exposure group, attributed to higher CO in the Eclipse exposure atmosphere. Higher CO resulted from the lower dilution of Eclipse smoke required to maintain WTPM concentrations equal to those of the 1R4F smoke, and not from a higher CO yield from Eclipse cigarettes. Nasal epithelial hyperplasia and ventral laryngeal squamous metaplasia were noted after exposure to either the 1R4F or Eclipse smoke. The degree of change was less in Eclipse smoke-exposed rats. Lung macrophages were increased to a similar extent in the Eclipse and 1R4F smoke-exposed groups. Brown/gold pigmented macrophages were detected in the lungs of rats exposed to 1R4F smoke, but not those exposed to Eclipse smoke. Subsets of rats from each group were maintained for an additional 13 wk without smoke exposures. Most of the changes noted at the end of the smoke exposures had disappeared, while those that remained were regressing toward normal. Evaluation of these findings indicated the overall biological activity of Eclipse smoke was less than 1R4F smoke at comparable exposure concentrations.     

Free lung cell response of mice and rats to mainstream cigarette smoke exposure

Male C57BL mice and F-344 rats were exposed through nose only to fresh mainstream smoke from one University of Kentucky Reference cigarette (2R1) daily under standardized conditions. At different exposure points, the lungs of room control (RM), sham control (SH), and smoke-exposed (SM) animals were lavaged and the number, composition, and properties of bronchoalveolar lavage (BAL) cells were studied. Significantly elevated levels of blood COHb and pulmonary aryl hydrocarbon hydroxylase activity indicated effective inhalation of smoke by animals. The BAL cell analysis showed that cigarette smoke induced a five- to sevenfold increase in the number of BAL cells in mice following 10- to 12-week exposure. The proportion of neutrophils (PMN) increased to about 18 +/- 3% in SM mice as compared to less than 1% in controls. Cessation of smoke treatments returned the PMN levels to those of controls within 5 weeks. Unlike mice, smoke exposure for up to 32 weeks failed to induce appreciable changes in the number and proportion of macrophages and neutrophils in rats. Large brown macrophages were observed in SM groups of both species. Functional analysis demonstrated that the BAL cells from SM mice but not rats released greater amounts of superoxides than controls under resting and phagocytically stimulated conditions. Enzymatic analysis of macrophages showed that the activity of N-acetylglucosaminidase was increased in SM groups of both species. The activity of 5'nucleotidase was significantly reduced in macrophages from SM mice but not rats. Activity of leucine aminopeptidase remained unaltered in both species. These results demonstrate distinct differences in the response of mice and rats to identically generated cigarette smoke.     

Biochemical markers of cardiovascular damage from tobacco smoke

There is growing evidence that several biochemical constituents of cigarette smoking play a significant role in the development and progression of heart and blood vessel damage, especially atherosclerotic lesions. Some biochemical markers of tobacco smoke may be determined in blood and urine samples. They are also the main responsible factors of cardiovascular harm. Nicotine and its major metabolite, cotinine, carbon monoxide, and thiocyanate seem to be specific markers. Ischaemic heart disease, cardiac arrhythmias, and endothelial dysfunction are the most common evidence of both active and passive smoking exposure. Dosage of cotinine in urine is of easier determination than that of other metabolites in assessing exposure to smoking, although carboxyhaemoglobin levels seem to be a qualitative, but not quantitative factor to estimate either the degree of cardiovascular damage or the level of exposure. Cigarette smoking is addictive because of nicotine, and it is nicotine withdrawal that causes many side effects of quitting smoking as well as the nicotine itself may exacerbate cardiac lesions. Also haematologic changes are a consequence of cigarette smoking exposure. Increased white blood cells, platelet aggregation and adhesiveness, fibrinogen level, and changes in serum lipids characterise the response to smoking. Anatomical and ultrastructural alterations of the heart and blood vessels are also described as a consequence of negative effects of biochemical markers of cigarette smoking. These alterations are known as "Smoke cardiomyopathy" in experimental pathology.     

Intranasal benzo[<Emphasis Type="Italic">a</Emphasis>]pyrene alters circadian blood pressure patterns and causes lung inflammation in rats

Polycyclic aromatic hydrocarbons, including benzo[a]pyrene (BaP), are environmental contaminants formed during organic material combustion (e.g. burning fossil fuels and cigarette smoke). BaP toxicity is mediated, in part, by activation of the aryl hydrocarbon receptor and formation of reactive metabolites, both of which lead to increased oxidative stress. Since air pollution and cigarette smoking are known to increase cardiovascular disease in humans, the objective of this study was to determine the effects of 7-day intranasal BaP exposure on circadian blood pressure patterns, arterial stiffness, and possible sources of oxidative stress in radiotelemetry-implanted rats. Arterial pulse wave dP/dt was used an indicator of arterial stiffness and was compared to both functional (nitric oxide production and bioactivity, endothelin-1 levels) and structural (wall thickness) features of the arterial wall. In addition, histology of lung, heart, and liver were examined as well as pulmonary and hepatic cytochrome P450 1A1 (CYP1A1) activity. BaP exposure altered the circadian pattern of blood pressure, with a reduction in the normal dipping pattern during sleep. This was associated with increased neutrophil recruitment in the lungs of BaP-exposed rats. In contrast, BaP had no effect on cardiovascular tissue histology, arterial stiffness, oxidative stress or lung and liver CYP1A1 activity. Thus, the current study does not support the hypothesis that BaP reactive metabolites increase oxidative stress leading to reduced vascular NO bioactivity and increased blood pressure. Instead, the current study suggests that inflammation, detected only in the lung, is associated with altered circadian rhythm of blood pressure.     

Chronic treatment with the angiotensin I converting enzyme inhibitor, perindopril, protects in vitro carbachol-induced vasorelaxation in a rat model of vascular calcium overload.

1. Treatment of young rats with vitamin D3 plus nicotine produced 31 and 4 fold increases in the calcium content of the aorta and the mesenteric arterial bed, respectively. 2. Aortic rings and perfused mesenteric arterial beds from vitamin D3/nicotine-treated animals showed a diminished contractile response to noradrenaline in vitro. 3. In vascular preparations from vitamin D3/nicotine-treated animals, precontracted with noradrenaline, relaxation by the endothelium-dependent vasodilator, carbachol, was attenuated but responses to sodium nitroprusside were not modified. 4. Prolonged treatment with the angiotensin I converting enzyme inhibitor, perindopril, at a dose (1 mg kg-1) which did not significantly modify blood pressure, failed to prevent vascular calcium overload. 5. Perindopril treatment diminished noradrenaline-evoked vasoconstrictor responses of aortic rings in both groups, but restored responses in mesenteric arterial beds of vitamin D3/nicotine-treated rats. 6. Perindopril treatment also restored the maximal responses to carbachol of both aortic rings and mesenteric arterial beds of vitamin D3/nicotine-treated rats. 7. In conclusion, in the vitamin D3 plus nicotine model of calcium overload, reduced endothelial-mediated relaxation can be prevented by perindopril treatment.     

Cigarette smoke inhalation specifically inhibits depressor responses to prostacyclin in the rat.

Studies have been made of the effects of prior exposure to cigarette smoke on cardiovascular responses of the anaesthetized rat to arachidonic acid, prostaglandin E2 (PGE2), PGF2 alpha and PGI2. When compared with a control group, falls in systolic and diastolic blood pressure and tachycardia following intravenous PGI2 were significantly reduced in those animals exposed to smoke 1 and 24 h previously. Responses 48 h after exposure were not significantly different. Pressor effects of PGF2 alpha and depressor responses to arachidonic acid and PGE2 were not significantly affected these times. It is suggested that the specific and long lasting attenuation of the effects of PGI2 which occurs following cigarette smoke inhalation could contribute to both acute cardiovascular changes and the circulatory diseases associated with smoking.     

Toxicologic evaluation of humectants added to cigarette tobacco: 13-week smoke inhalation study of glycerin and propylene glycol in Fischer 344 rats

Glycerin (CAS no. 56-81-5) and propylene glycol (CAS no. 57-55-6) are commonly used as humectant ingredients in manufactured cigarettes to control and maintain the moisture content of the cut tobacco filler. The potential of these added humectants to affect the toxicity of cigarette smoke was investigated in a subchronic nose-only smoke inhalation study in rats. Filtered test cigarettes were prepared from an American-style tobacco blend containing either glycerin added at 5.1% w/w tobacco, propylene glycol at 2.2% w/w tobacco, or combinations of these humectants totaling 2.3%, 3.9%, and 7.2% w/w tobacco. Other groups of animals were exposed similarly to the smoke of reference cigarettes without added humectants, or to filtered air (sham control). Smoke exposures were conducted for 1 h/day, 5 days/wk for 13 wk, at target smoke particulate concentrations of 350 mg/m(3). All smoke-exposed groups had equivalent increases in blood carboxyhemoglobin, serum nicotine, and serum cotinine relative to the air controls. Smoke-associated reductions in body weights and occasional increases in heart and lung weights were generally similar among the various exposure conditions at necropsy. Increases in serum alkaline phosphatase and decreases in serum glucose and cholesterol were observed among smoke-exposed females relative to air controls. However, no significant differences in these parameters were evident between the humectant-containing and reference cigarette smoke exposure groups. Assessments of respiration conducted after 3, 6, 9, and 12 wk of smoke exposure indicated an initial smoke-related but not humectant-related decrease in respiratory rate, tidal volume, and minute volume during the first 20 min of each smoke exposure. Respiratory-tract histopathology was consistent across sexes and smoke groups, comprising (1) diffuse and focal alveolar pigmented macrophages and chronic interstitial inflammation in the lung, (2) laryngeal epithelial hyperplasia, squamous metaplasia, and scab formation, and (3) epithelial hyperplasia in the anterior nose. Smoke-related histopathology resolved substantially during a 6-wk postexposure recovery period. Addition of the tested humectants to cigarettes, singly or in combination, had no meaningful effect on the site, occurrence, or severity of respiratory-tract changes or on the measured indices of pulmonary function. It was concluded that the addition of glycerin and propylene glycol to cigarettes does not significantly affect the biological activity of inhaled cigarette smoke in this rat model.     

TOXICOLOGIC EVALUATION OF HUMECTANTS ADDED TO CIGARETTE TOBACCO: 13-WEEK SMOKE INHALATION STUDY OF GLYCERIN AND PROPYLENE GLYCOL IN FISCHER 344 RATS

Glycerin (CAS no. 56-81-5) and propylene glycol (CAS no. 57-55-6) are commonly used as humectant ingredients in manufactured cigarettes to control and maintain the moisture content of the cut tobacco filler. The potential of these added humectants to affect the toxicity of cigarette smoke was investigated in a subchronic nose-only smoke inhalation study in rats. Filtered test cigarettes were prepared from an American-style tobacco blend containing either glycerin added at 5.1% w/w tobacco, propylene glycol at 2.2% w/w tobacco, or combinations of these humectants totaling 2.3%, 3.9%, and 7.2% w/w tobacco. Other groups of animals were exposed similarly to the smoke of reference cigarettes without added humectants, or to filtered air (sham control). Smoke exposures were conducted for 1 h/ day, 5 days/wk for 13 wk, at target smoke particulate concentrations of 350 mg/m 3. All smoke-exposed groups had equivalent increases in blood carboxyhemoglobin, serum nicotine, and serum cotinine relative to the air controls. Smoke-associated reductions in body weights and occasional increases in heart and lung weights were generally similar among the various exposure conditions at necropsy. Increases in serum alkaline phosphatase and decreases in serum glucose and cholesterol were observed among smoke-exposed females relative to air controls. However, no significant differences in these parameters were evident between the humectant-containing and reference cigarette smoke exposure groups. Assessments of respiration conducted after 3, 6, 9, and 12 wk of smoke exposure indicated an initial smoke-related but not humectant-related decrease in respiratory rate, tidal volume, and minute volume during the first 20 min of each smoke exposure. Respiratory-tract histopathology was consistent across sexes and smoke groups, comprising (1) diffuse and focal alveolar pigmented macrophages and chronic interstitial inflammation in the lung, (2) laryngeal epithelial hyperplasia, squamous metaplasia, and scab formation, and (3) epithelial hyperplasia in the anterior nose. Smoke-related histopathology resolved substantially during a 6-wk postexposure recovery period. Addition of the tested humectants to cigarettes, singly or in combination, had no meaningful effect on the site, occurrence, or severity of respiratory-tract changes or on the measured indices of pulmonary function. It was concluded that the addition of glycerin and propylene glycol to cigarettes does not significantly affect the biological activity of inhaled cigarette smoke in this rat model.