* Research and Development, R. J. Reynolds Tobacco Company, Bowman Gray Technical Center, Winston-Salem, North Carolina 27102; and
Center for Thrombosis and Hemostasis, University of North Carolina, Chapel Hill, North Carolina
Received June 22, 2000; accepted October 17, 2000
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key Words: environmental tobacco smoke; platelets; thromboxane; cardiovascular disease.
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Proponents of an adverse association between ETS and CVD have hypothesized that platelet aggregation induced by ETS could explain the highly nonlinear shape of the dose-response relationship (California EPA, 1997; Law et al., 1997; Matthews, 1999
; OSHA, 1994; SCOTH, 1998). In essence, this hypothesis states that a small amount of ETS exposure would induce platelets to aggregate to a quantitative degree similar to that seen by exposure to a relatively larger amount of mainstream smoke (active smoking). Further, it is hypothesized that the induction of platelet aggregation may lead to the development of thrombi in occluded coronary arteries and contribute to myocardial infarction.
Thromboxanes are synthesized in platelets and can cause vasoconstriction and platelet aggregation after release (Hamberg and Samuelsson, 1974). Prostacyclins are potent inhibitors of platelet aggregation, produced by blood vessel walls (DeWitt et al., 1983
) when endothelial cells are exposed to activated platelets. Therefore, thromboxanes and prostacyclins are considered to be antagonistic in their actions (Mayes, 1985
). Several groups have previously shown that an increased urinary level of the thromboxane metabolite 2,3-dinor-thromboxane B2 (Tx-M) is a marker of in vivo platelet activation and that an increased urinary level of the prostacyclin metabolite 2,3-dinor-6-keto-prostaglandin F1
(PGI-M) potentially indicates "activated platelet" vessel wall interaction (Dotevall et al., 1992
; Lassila et al., 1988
; Murray et al., 1985
; Nowak et al., 1987
; Rangemark and Wennmalm, 1991
). These groups have reported that cigarette smokers usually excrete higher levels of Tx-M and sometimes of PGI-M. A meta-analysis of 56 smoking and platelet-aggregation studies conducted by our group also found a strong association between smoking and thromboxane excretion, and a less robust association between smoking and prostacyclin excretion (Smith et al., 1998
). Of 16 studies examining the chronic urinary excretion of thromboxane metabolites in cigarette smokers, all 16 showed an increase in the urinary level of thromboxane metabolite as compared with nonsmokers. In the 17 studies that examined the urinary excretion of prostacyclin metabolites in smokers, 5 studies reported an increased excretion in smokers and 12 studies showed no difference between smokers and nonsmokers.
In the present study, we tested the hypothesis that ETS exposure is associated with increased platelet activation by measuring the concentration of the stable metabolites of thromboxane and prostacyclin in 24-h urinary samples from 3 groups of subjects. The 3 groups were (1) 21 nonsmokers who self-reported minimal or no ETS exposure (non-ETS-exposed); (2) 22 nonsmokers who self-reported at least 5 h of ETS exposure per day (ETS-exposed); and (3) a positive control group of 20 smokers who self-reported an average consumption of 40 cigarettes (2 packs) per day. Personal exposure monitoring of a number of particulate and vapor-phase smoke constituents throughout the 24-h study period, and salivary cotinine values, validated the self-reported exposure. This is the first study conducted on ETS and cardiovascular disease risk that has included both an accurate assessment of the amount of smoke exposure and an integrated measurement of platelet activity. Collecting indicators of platelet activity over a 24-h period accounts for endogenous biological variability (for example, circadian) and differences in postexposure time prior to platelet activity determination.
As an additional indicator of CVD risk, 8-hydroxy-2'-deoxyguanosine (8-OHdG) was measured in the 3 groups of subjects. A principal stable marker of hydroxyl radical damage to DNA, 8-OHdG is excreted in urine as a result of oxidative damage to DNA (Lagorio et al., 1994; Sperati et al., 1999
). Some studies on smokers have shown statistically significant increases in levels of 8-OHdG (van Zeeland et al., 1999
) while others have shown no difference (Howard et al., 1988
). Urinary free cortisol was also measured as a possible marker of "physiological stress."
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
Confidential medical history questionnaire.
Female subjects taking hormone supplements were excluded, as use of supplements such as birth control pills, Norplant, and Depo Provera injection have been reported to influence platelet activation (Schiff et al., 1999). Medical conditions that could potentially affect thrombosis and hemostasis were exclusionary, including: phlebitis, pulmonary embolism, blood clot at another site, hemophilia, sickle cell anemia, and polycythemia. Subjects at elevated risk for an adverse cardiovascular event were excluded, including those with a history of the following: hypertension, diabetes, heart disease, chest pains, intermittent claudication, and restricted arteries in the neck. A history of infectious disease was exclusionary, including hepatitis, AIDS, sexually transmitted diseases, and "other infectious disease." Inflammatory or autoimmune disease, including rheumatoid arthritis, systemic lupus erythematosus, thyroiditis, and Hashimoto's disease, was also exclusionary.
The questionnaire was used to determine tobacco use history for selection of the cigarette smoking group. Current heavy smokers with an average consumption of approximately 40 cigarettes per day were included.
Secondhand smoke exposure survey.
Exposure to ETS was defined as the ability to smell tobacco smoke, as this is a highly sensitive index of exposure (Walker et al., 1997). Nonsmokers who self-reported 5 h per day or more of ETS exposure were considered exposed. Nonsmokers who self-reported 15 min or less per day were classified as non-ETS-exposed. Exposure to wood burning stoves, to occupational diesel or auto exhaust, or to chemical fumes was exclusionary.
Abstention from platelet-inhibiting drugs.
At a subject information session held prior to study initiation, each subject was given a list of over-the-counter medications that contained both reversible and nonreversible platelet-inhibiting agents, including aspirin, ibuprofen, naproxen sodium, and ketoprofen. All subjects abstained from platelet-inhibiting drugs for one week prior to collection of the 24-h urine sample.
Subject Description
Three different groups of subjects were studied: 21 nonsmokers with minimal or no ETS exposure (7 males, 14 females; mean age 28.4 ± 5.7 years); 22 nonsmokers with at least 5 h per day of ETS exposure (11 males, 11 females; mean age 32.8 ± 7.2 years); and 20 cigarette smokers, with an average consumption of 2 packs per day (14 males, 6 females; mean age 40.6 ± 5.2 years). Our meta-analysis of 150 separate platelet aggregation results taken from 56 studies on the relationship between cigarette smoking and platelet aggregation showed a consistent pattern of increased platelet aggregation over a wide age range of smokers (Smith et al., 1998). Therefore, the somewhat older average age of the smoking group as compared with the 2 nonsmoking groups should not be physiologically significant vis à vis the use of this group as a positive control.
Recruiting 3 different subject groups experiencing such disparate levels of exposure to cigarette smoke was problematic. First, smoking restrictions in the U.S. workplace are almost ubiquitous. Therefore, members of the heavily ETS-exposed subject group had to be employed at a workplace that permitted smoking and also, preferably, to live with a smoker. Conversely, the non-ETS-exposed subject group was required to work, recreate, and live in a smoke-free environment. Based on the 2 extremes of ETS-exposure criteria required, the ETS-exposed subjects and the cigarette-smoking subjects were recruited and enrolled from employees of R. J. Reynolds Tobacco Company (Winston-Salem, NC) and their primary relatives and friends. The non-exposed subjects were recruited and enrolled from employees and students at the University of North Carolina School of Medicine, Chapel Hill, NC.
The study was reviewed by the Ethics Committee of the University of North Carolina School of Medicine and was performed in accordance with the ethical standards of the 1964 Declaration of Helsinki. All persons gave written informed consent prior to their inclusion in the study. Subjects were not aware of the study hypothesis prior to completion of the experiment.
Chemical Analyses
Tx-M and PGI-M analyses.
Radioimmunoassay analyses of the stable urinary metabolites of thromboxane (Tx-M) (Kuhn et al., 1993) and prostacyclin (PGI-M) (Demers et al., 1981
) were performed without knowledge of study design or subject group by Clinical Laboratory Services, Department of Pathology, Milton S. Hershey Medical Center, Pennsylvania State University. One ml of straight urine was extracted once using 3 ml of ethyl acetate, 3 ml of saline, and a 3-ml mixture of ethyl acetate, isopropyl alcohol, and 0.1 N hydrochloric acid. The organic layer was taken off by drying down under nitrogen in a water bath. The sample was then brought back up to the original volume of 1 ml, using BGG buffer. Therefore, there was no dilution of the urine samples. The measurements were validated by using internal standards. The detection limits for Tx-M and PGI-M are 5 pg/ml and 9 pg/ml, respectively. PGI-M and Tx-M antiserum were ordered from Advanced Magnetics (Cambridge, MA). Urinary creatinine was determined by the Jaffe method using a commercial test kit (Merck AG, Darmstadt, Germany).
8-Hydroxy-2'-deoxyguanosine (8-OHdG).
ESA Laboratories, Inc. (Chelmsford, MA) determined 8-OHdG in 24-h urine by a liquid chromatography electrochemical (LCEC) column switching system, without knowledge of study design or subject group. This 30-min automated column switching, high performance LCEC method for 8-OHdG was developed based on the unique purine selectivity of integral porous carbon columns. Detection with series coulometric electrodes provides 500 fg sensitivity and qualitative certainty by 8-OHdG/creatinine response ratios.
Urinary free cortisol.
Using the antibody-coated tube method (Diagnostic Products Corporation, Los Angeles, CA), urinary free cortisol was also measured by Clinical Laboratory Services at the Milton S. Hershey Medical Center.
ETS Exposure Assessment
Markers of ETS exposure were assessed for each subject during the 24-h study period by personal monitoring for 3 vapor phase analytes and 4 particulate phase analytes. The vapor phase analytes were nicotine, 3-ethenylpyridine, and myosmine. The 4 particulate phase analytes were gravimetric respirable suspended particulate matter (RSP), ultraviolet particulate matter (UVPM), fluorescent particulate matter (FPM), and solanesol particulate matter (Sol-PM). The vapor phase analytes were collected on XAD-4 solid sorbent tubes (SKC, Inc., Eighty Four, PA), and the respirable (4.0 µm median cutoff) particulate phase analytes were collected on 37-mm diameter, 1.0-µm pore size Fluoropore filters (Millipore Corp., Bedford, MA). Simultaneous collection was achieved by using personal Double Take Samplers (SKC, Inc.). Nicotine and 3-ethenylpyridine analyses were performed by ASTM Method D 507596 (ASTM Method D 507596, 1996; Ogden et al., 1996; Ogden and Nelson, 1994
). Myosmine analyses were performed as reported by Ogden et al. (1996) and Ogden and Nelson (1994). The XAD-4 resin was transferred to an auto-sampler vial, extracted with ethyl acetate modified with 0.0125% triethylamine, and analyzed by capillary gas chromatography with N-thermionic detection using quinoline as an internal standard. Instrumentation used included a Hewlett-Packard Model 5890 A gas chromatograph (Palo Alto, CA) equipped with a Hewlett-Packard Model 7673 auto-sampler, a split/splitless injector, a 30 m x 0.32 mm JDB-5 capillary column (J&W Scientific, Folsom, CA), and a nitrogen-phosphorus detector. Quantification was achieved using EZChrom data software (Scientific Software, Inc., Pleasanton, CA). This method allows for simultaneous determination of nicotine, 3-ethenylpyridine, and myosmine. Gravimetric RSP, UVPM, and FPM analyses were conducted by ASTM Method D 595596 (ASTM Method D 627198, 1998; Ogden et al., 1996
). The pre-weighed filters were conditioned at 50% relative humidity for a minimum of 12 h (static inhibited) and the final weight was determined. After gravimetric determination of RSP, the filters were placed in vials and extracted with methanol. Determinations of UVPM and FPM were performed simultaneously, using standard HPLC equipment (Waters Chromatography Division, Millipore Corp., Milford, MA) without a column installed; Model M-45 Solvent Delivery System; Model 712 WISP autosampler; Model 490E Programmable Wavelength Detector at 325 nm absorbance; and a Hitachi Corporation (Danbury, CT) Model F1000 Fluorescence Spectrophotometer at 300 nm excitation and 420 nm emission. Sol-PM analyses were performed using ASTM Method D 627198 (ASTM Method 627198, 1998; Heavner et al., 1996
; Ogden et al., 1990
, 1996
). Determination of solanesol and Sol-PM was performed on the same methanol extract using a Waters Model 510 pump, Waters Model 712 WISP auto-sampler, and Hitachi Model L-4200 UV-Visible detector at 205 nm absorbance with a Keystone Scientific (Bellefonte, PA) 250 mm x 4.6 mm, 5 µm DELTABOND ODS LC column at isocratic conditions of 95% acetonitrile/5% methanol. Quantification for UVPM, FPM, and solanesol was achieved using Scientific Software, Inc. (Pleasanton, CA) EZChrom data software. The readings were field blank corrected with nominally negative values being assigned a value of zero. A saliva sample was collected from each subject at the beginning of the 24-h study period using Salivettes (Sarstedt Inc., Newton, NC). Salivary cotinine was determined by radioimmunoassay (Langone et al., 1973
; Langone and van Vunakis, 1982
).
Statistical Methods
Thromboxane, prostacyclin, cortisol and 8-OHdG comparisons were all performed in a similar fashion. The primary comparison in each case related to whether the exposure groups (smokers, ETS-exposed nonsmokers, and non-ETS-exposed nonsmokers) were different from one another. All comparisons utilized analysis-of-variance techniques. The potential confounding factors taken into account were gender, creatinine level, and age in a subset of calculations. Creatinine is formed and excreted in constant amounts and serves as an internal standard. The analysis was done using the following combinations of covariates: (1) gender; (2) gender and creatinine; and (3) interactions of gender and creatinine with exposure group. Typically the terms accounting for the interaction of creatinine with exposure group were necessary, but the interaction term with gender was not. Effectively, fitting a model with an interaction between creatinine and exposure group is equivalent to fitting separate regression lines for each exposure group to account for the relationship between creatinine and the various response variables. This model was chosen because the slopes often differed among the different groups. Note that this is not equivalent to the common practice of dividing the response by creatinine. This approach appears to be more appropriate than the common practice, because the regression lines had a non-zero intercept. Terms were retained in the model only when significant.
The mean values for exposure groups are compared after adjusting for other terms in the model. For instance, in the models that include creatinine, the exposure group values are adjusted to the average level of creatinine over all of the subjects.
The air concentrations and salivary cotinine values were compared using an analysis-of-variance model, taking only the exposure group into account. Formal comparison of these data required that the values be transformed to make the variances within the exposure groups comparable. This was done using a power transformation, y, where
, (n = 1, 2...). The value of
was chosen to make the variances similar to one another.
Several of the subjects in the ETS-exposed and non-ETS-exposed groups had salivary cotinine levels below the limit of detection of about 0.40 ng/ml. There are a number of different comparison procedures that are possible with different approaches for handling the values that are below the limit of detection. All of the procedures reach the same conclusions for these data. The approach reported here used all of the readings in their raw form whether they were above or below the limit of detection.
All of the comparisons were conducted using PROC GLM in the SAS® system (SAS Institute, 1990).
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
|
|
Biological Measurements (Table 4)
As noted above, thromboxane was compared using the 3 different analysis-of-variance models, i.e., gender, gender and creatinine, and interactions of gender and creatinine with exposure group. All show that the thromboxane levels differ among exposure groups (p < 0.0001 for each pairwise comparison). The thromboxane levels are highest for the smokers (54.7 ± 2.3 pg/ml), followed by the non-ETS-exposed group (34.5 ± 1.8 pg/ml), followed by the ETS-exposed group (18.6 ± 1.8 pg/ml) (Fig. 2).
|
|
Because of age differences among the exposure groups, separate analyses were performed with an age term included as a covariate in the models for thromboxane and prostacyclin. All of the conclusions with respect to thromboxane and prostacyclin were identical, whether or not age was included in the model.
The method of comparison for 8-OHdG levels among exposure groups was similar to that used for thromboxane and prostacyclin. Gender was statistically significant, and was therefore retained in the model (p = 0.042). Smokers (4.94 ng/ml) were statistically significantly higher than the ETS-exposed group (2.60 ng/ml) (p = 0.001). The non-ETS-exposed group (3.70 ng/ml) was not statistically significantly different from either the smokers or the ETS-exposed group (p = 0.082 and 0.062, respectively).
The above analyses used a statistical model that assumes that the relationship between 8-OHdG and creatinine differs among exposure groups (p = 0.067). A model assuming no difference in the slope of creatinine among the three exposure groups gives slightly different conclusions. The 8-OHdG level in the ETS-exposed group remains significantly lower than in smokers (p = 0.005). The test comparing the ETS-exposed and non-ETS-exposed groups becomes statistically significant (p = 0.029) (non-ETS-exposed group is higher) and the test comparing the smokers and the non-ETS-exposed group remains not statistically significant (p = 0.42).
No statistically significant exposure group differences were found for cortisol.
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Interestingly, the group of non-exposed nonsmokers who were employed at the University of North Carolina School of Medicine showed a statistically significantly higher urinary thromboxane excretion as compared with the ETS-exposed nonsmokers. Whether the UNC Chapel Hill group experienced increased physical (Burghuber et al., 1981; Wang et al., 1994
) or psychological occupational stress, thereby leading to catecholamine-induced platelet priming, remains unknown. One possible marker of "physiological stress," i.e., urinary free cortisol, showed no differences among the 3 groups.
In addition to the meta-analysis of He et al. (1999), two published government reports have also suggested a causal association between ETS and the incidence of CHD (California EPA, 1997; SCOTH, 1998). The California Environmental Protection Agency (CALEPA) and the Scientific Committee on Tobacco and Health (SCOTH) in the United Kingdom have estimated the relative risk for CHD in ETS-exposed nonsmokers at 1.30 and 1.23, respectively. However Bailar (1999) has suggested that the large differential in exposure between ETS and active smoking stands in contrast to the small difference in CVD risk estimates. One hypothesis has been that the platelet response to cigarette smoke is highly nonlinear, with only a tiny "dose" needed to activate platelets. Since ETS-exposed nonsmokers did not demonstrate increased indicators of platelet aggregation, the results from the present study suggest that platelet aggregation is not a plausible or quantitatively consistent mechanism to explain such a nonlinear dose-response anomaly.
Exposure to reactive oxygen species is reportedly a risk factor for chronic diseases including CVD (Mehta and Mehta, 1999; Palace et al., 1999
). Several studies have examined oxidative stress levels in cigarette smokers by measuring urinary 8-OHdG levels (Howard et al., 1988
; van Zeeland et al., 1999
). In the present study, smokers displayed significant increases in urinary 8-OHdG as compared with ETS-exposed nonsmokers. ETS exposure did not result in increases in urinary 8-OHdG.
![]() |
ACKNOWLEDGMENTS |
---|
![]() |
NOTES |
---|
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
ASTM (1996). Standard Test Methods for Estimating the Contribution of Environmental Tobacco Smoke to Respirable Suspended Particles Based on UVPM and FPM: Method D 595596. American Society for Testing and Materials, W. Conshohocken, PA.
ASTM (1998). Standard Test Method for Estimating Contribution of Environmental Tobacco Smoke to Respirable Suspended Particles Based on Solanesol: Method D 627198.American Society for Testing and Materials, W. Conshohocken, PA.
Bailar, J. C. (1999). Passive smoking, coronary heart disease, and meta-analysis. N. Engl. J. Med. 340, 958959.
Burghuber, O., Sinzinger, H., Silberbauer, K., Wolf, C., and Haber, P. (1981). Decreased prostacyclin sensitivity of human platelets after jogging and squash. Prostaglandins Med. 6, 127130.[ISI][Medline]
California Environmental Protection Agency. (1997). Health Effects of Exposure to Environmental Tobacco Smoke: Cardiovascular Health Effects, pp. 174. Office of Environmental Health Hazard Assessment, Sacramento, CA.
Cryer, P. E., Haymond, M. W., Santiago, J. V., and Shah, S. D. (1976). Norepinephrine and epinephrine release and adrenergic mediation of smoking-associated hemodynamic and metabolic events. New Engl. J. Med. 295, 573577.[Abstract]
Demers, L. M., Harrison, T. S., Halbert, D. R., and Santen, R. J. (1981). Effect of prolonged exercise on plasma prostaglandin levels. Prostaglandins Med. 6, 413418.[ISI][Medline]
DeWitt, L., Day, S., Sommenburg, K., and Smith, L. (1983). Concentrations of prostaglandin endoperoxide synthase and prostaglandin I2 synthase in the endothelium and smooth muscle of bovine aorta. J. Clin. Invest. 72, 18821888.[ISI][Medline]
Dotevall, A., Raangemark, C., Eriksson, E., Kutti, J., Wadenvik, H., and Wennmalm, A. (1992). Cigarette smoking increases thromboxane A2 formation without affecting platelet survival in young healthy females. Thromb. Haemost. 68, 583588.[ISI][Medline]
Hamberg, M., and Samuelsson, B. (1974). Prostaglandin endoperoxides: Novel transformations of arachidonic acid in human platelets. Proc. Nat. Acad. Sci. U.S.A. 71, 34003404.[Abstract]
He, J., Vupputuri, S., Allen, K., Prerost, M. R., Hughes, J., and Whelton, P. K. (1999). Passive smoking and the risk of coronary heart diseasea meta-analysis of epidemiologic studies. N. Engl. J. Med. 340, 920926.
Heavner, D. L., Morgan, W. T., and Ogden, M. W. (1996). Determination of volatile organic compounds and respirable suspended particulate matter in New Jersey and Pennsylvania homes and workplaces. Environ. Intl. 22, 159183.
Howard, D. J., Ota, R. B., Briggs, L. A., Hampton, M., and Pritsos, C. A. (1988). Oxidative stress induced by environmental tobacco smoke in the workplace is mitigated by antioxidant supplementation. Cancer Epidemiol. Biomarkers Prev. 7, 981988.[Abstract]
Kuhn, D. C., Stauffer, J. L., Gaydos, L. J., Lacey, S. L., and Demers, L. M. (1993). An inhibitor of thromboxane production attenuates tumor necrosis factor release by activated human alveolar macrophages. Prostaglandins 46, 195205.[Medline]
Lagorio, S., Tagesson, C., Forastiere, F., Iavarone, I., Axelson, O., and Carere, A. (1994). Exposure to benzene and urinary concentrations of 8-hydroxydeoxyguanosine, a biological marker of oxidative damage to DNA. Occup. Environ. Med. 51, 739743.[Abstract]
Langone, J. J., Gjika, H. B., and van Vunakis, H. (1973). Nicotine and its metabolites: Radioimmunoassay for nicotine and cotinine. Biochemistry 12, 50255030.[ISI][Medline]
Langone, J. J., and van Vunakis, H. (1982). Radioimmunoassay of nicotine, cotinine, and gamma-(3-pyridyl)-gamma-oxo-N-methylbutyramide. Methods Enzymol. 84, 628640.[ISI][Medline]
Lassila, R., Seyberth, H. W., Haapanen, A., Schweer, H., Koskenvuo, M., and Laustiola, K. E. (1988). Vasoactive and atherogenic effects of cigarette smoking: A study of monozygotic twins discordant for smoking. Br. Med. J. 297, 955957.[ISI][Medline]
Law, M. R., Morris, J. K., and Wald, N. J. (1997). Environmental tobacco smoke exposure and ischemic heart disease: An evaluation of the evidence. Br. Med. J. 315, 973980.
Matthews, R. (1999). Smoke gets in your eyes. New Sci. 162, 1819.
Mayes, P. A. (1985). Metabolism of lipids: I. Fatty acids. In Harper's Review of Biochemistry, 20th ed. (D.W. Martin, P.A. Mayes, V.W. Rodwell, and D.K. Granner-Lange, Eds.), pp. 208231. Lange Medical Publications, Los Altos, CA.
Mehta, J. L., and Mehta, J. (1999). Antioxidants and vitamins in your cardiac patient: Are they helpful? Cardiol. Rev. 7, 5661.[Medline]
Murray, J. J., Nowak, J., Oates, J. A., and FitzGerald, G. A. (1985). Platelet function during chronic smoking and withdrawal in man. Clin. Res. 33, 350A.
Nowak, J., Murray, J. J., Oates, J. A., and FitzGerald, G. A. (1987). Biochemical evidence of a chronic abnormality in platelet and vascular function in healthy individuals who smoke cigarettes. Circulation 76, 614.[Abstract]
Occupational Safety and Health Administration (OSHA) (1994). Indoor Air Quality 59, 179.
Ogden, M. W., Heavner, D. L., Foster, T. L., Maiolo, K. C., Cash, S. L., Richardson, J. D., Martin, P., Simmons, P. S., Conrad, F. W., and Nelson, P. R (1996). Personal monitoring system for measuring environmental tobacco smoke exposure. Environ. Technol. 17, 239250.[ISI]
Ogden, M. W., Maiolo, K. C., Oldaker, G. B., and Conrad, F. W., Jr. (1990). Evaluation of methods for estimating the contribution of ETS to respirable suspended particles. Proc. 5th International Conference on Indoor Air Quality and Climate. Vol. 2., pp. 415420. International Conference on Indoor Air Quality and Climate, Ottawa, Canada.
Ogden, M. W., and Nelson, P. R. (1994). Detection of alkaloids in environmental tobacco smoke. Modern Methods of Plant Analysis: Alkaloids (H. F. Linskens and J. F. Jackson ,Eds.), Vol. 15, pp. 163189. Springer-Verlag, Berlin.
Palace, V. P., Khaper, N., Qin, Q., and Singal, P. K. (1999). Antioxidant potentials of vitamin A and carotenoids and their relevance to heart disease. Free Radic. Biol. Med. 26, 746761.[ISI][Medline]
Rangemark, C., and Wennmalm, A. (1991). Cigarette smoking and urinary excretion of markers for platelet/vessel wall interaction in healthy women. Clin. Sci. 81, 1115.[ISI][Medline]
Scientific Committee on Tobacco and Health (SCOTH). (1998). Report of the Scientific Committee on Tobacco and Health on Environmental tobacco smoke and ischemic heart disease. The Stationary Office 32, London.
SAS Institute, Inc. (1990). SAS/STAT User's Guide, Volume 2, Version 6, Fourth Edition, Cary, NC.
Schiff, I., Bell, W. R., Davis, V., Kessler, C. M., Meyers, C. Nakajima, S., Sexton, B. J. (1999). Oral contraceptives and smoking, current considerations: Recommendations of a consensus panel. Am. J. Obstet. Gynecol. 180, 383384.
Siess, W., Lorenz, R., Roth, P., and Weber, P. C. (1982). Plasma catecholamines, platelet aggregation, and associated thromboxane formation after physical exercise, smoking, or norepinephrine infusion. Circulation 66, 4448.[Abstract]
Smith, C. J., Fischer, T. H., and Sears, S. B. (2000). Environmental tobacco smoke, cardiovascular disease, and the nonlinear dose-response hypothesis. Toxicol. Sci. 54, 462472.
Smith, C. J., Sears, S. B., Walker, J. C., and DeLuca, P. O. (1992). Environmental tobacco smoke: Current assessment and future directions. Toxicol. Pathol. 20, 289303.[ISI][Medline]
Smith, C. J., Steichen, T. J., and Fischer, T. H. (1998). Platelet aggregation in cigarette smokers: A meta-analysis. Inhal. Tox. 10, 765793.[ISI]
Sperati, A., Abeni, D. D., Tagesson, C., Forastiere, F., Miceli, M., and Axelson, O. (1999). Exposure to indoor background radiation and urinary concentrations of 8-hydroxydeoxyguanosine, a marker of oxidative DNA damage. Environ. Health Perspect. 107, 213215.[ISI][Medline]
Trap-Jensen, J., Carlsen, J. E., Svendsen, T. L., Christensen, N. J. (1979). Cardiovascular and adrenergic effects of cigarette smoking during immediate non-selective and selective beta-adrenoreceptor blockade in humans. Eur. J. Clin. Invest. 9, 181183.[ISI][Medline]
van Zeeland, A. A., de Groot, A. L., Hall, J., and Donato, F. (1999). 8-Hydroxy-deoxyguanosine in DNA from leukocytes of healthy adults: relationship with cigarette smoking, environmental tobacco smoke, alcohol and coffee consumption. Mutat. Res. 439, 249257.[ISI][Medline]
Walker, J. C., Nelson, P. R., and Cain, W. S. (1997). Perceptual and psychophysiological responses of nonsmokers to a range of environmental tobacco smoke concentrations. Indoor Air 7, 173188.[ISI]
Wang, J. S., Jen, C. J., Kung, H. C., Lin, L. J., Hsiue, T. R., and Chen, H. I. (1994). Different effects of strenuous exercise and moderate exercise on platelet function in men. Circulation 90, 28772885.[Abstract]