BACKGROUND: Cigarette smoking increases the risk of cardiovascular disease, and is an important preventable cause of death and illness. One major deterrent to smoking cessation is a gain in body weight. Understanding the mechanisms that contribute to this weight gain may maximize the success of long-term smoking cessation. We hypothesized that smoking cessation is associated with an increase in adipose tissue lipoprotein lipase (AT-LPL) activity and/or a decrease in lipolysis, two metabolic factors that determine the balance between fat storage and fat utilization, and thus affect the propensity for weight gain.
MATERIALS AND METHODS: Ten premenopausal women (37.1±6.2 y, 31.7±6.4 kg/m2 body mass index (BMI), mean±s.d.) participated in a 4 week smoking cessation program. Measurements of body weight, waist and hip circumferences, adipose cell metabolism and resting metabolic rate were obtained at baseline and after 4 weeks of smoking cessation.
RESULTS: Of the 10 women who began the intervention, five successfully completed the smoking cessation intervention. After 4 weeks of smoking cessation, there were significant increases in body weight (95.1±13.9–97.7±14.4 kg, P<0.05), with no change in waist and hip circumferences or resting energy expenditure. Gluteal AT-LPL activity significantly increased in all women by 2.8-fold (1.65±1.30–4.72±3.34 nmol/g/min, P<0.05). Abdominal AT-LPL activity increased in four out of the five women, but did not reach statistical significance (1.14±0.88–3.50±3.76 nmol/g/min, P=0.14). The increase in body weight correlated with the increase in gluteal AT-LPL activity (r=0.89, P<0.05), as well as the baseline activity of gluteal AT-LPL (r=0.86, P=0.06). There were no changes in basal or stimulated lipolysis in the gluteal or abdominal fat depots.
CONCLUSIONS: These results suggest that smoking cessation is associated with significant increases in body weight, as well as changes in adipose cell metabolism, in particular increases in AT-LPL activity. This increase in LPL activity may contribute to the increase in body weight associated with smoking cessation.
Cigarette smoking is an independent risk factor for cardiovascular disease, and the single most important preventable cause of death and illness in the USA.1 However, only about 40% of the smokers in the US ever attempt to quit, and only 6% succeed in long-term smoking cessation.2 One of the immediate consequences and a deterrent to smoking cessation is a gain in weight.3,4 Thus, understanding the metabolic mechanisms by which smoking cessation leads to weight gain may be important for the development of specific interventions that prevent or attenuate this weight gain and maximize the success of long-term smoking cessation.
Mechanisms for weight gain include increased energy intake, decreased resting metabolic rate (RMR), and decreased physical activity.5,6,7,8 In addition, changes in adipose tissue metabolism may also contribute to the weight gain following smoking cessation, although the potential mechanisms are unknown. Although smoking or nicotine exposure may result in changes in adipose tissue lipoprotein lipase (AT-LPL) activity and lipolysis,9,10,11,12 no previous studies have examined the effects of smoking cessation on adipose tissue metabolism in premenopausal women. The purpose of this study was to determine the effects of 4 weeks of smoking cessation on AT-LPL activity and lipolysis in healthy premenopausal female smokers. We hypothesized that smoking cessation would be associated with an increase in the activity of AT-LPL, and/or a decrease in lipolysis, two metabolic factors that determine the balance between fat storage and fat utilization, and thus affect the propensity for weight gain.
Premenopausal women smokers were recruited for participation from the Baltimore-Washington, DC metropolitan area. The women provided informed consent to participate in the study according to the guidelines of the University of Maryland Institutional Review Board for Human Research. All women underwent initial screening evaluations, which included a medical history, physical examination, fasting blood profile and 12-lead resting electrocardiogram. Women with evidence of diabetes (fasting plasma glucose level >6.4 mM), hypertension (blood pressure (BP)>160/90 mmHg), hyperlipidemia, cancer, liver, renal, hematological disease or taking any medications were excluded.
Measurements of waist and hip circumferences, fat metabolism, plasma leptin, lipoprotein lipids and RMR were obtained in the morning after a 12 h overnight fast at baseline and after 4 weeks of smoking cessation. Subjects were asked to abstain from smoking for 12 h prior to study. After completion of baseline testing, subjects participated in a smoking cessation program for 4 weeks (2 days/week, 1 h/day). Compliance with smoking cessation was monitored by the measurement of expired carbon monoxide prior to each session to validate self-reported smoking status.
Body circumference measurements
Waist and hip circumferences were measured in duplicate. The minimal waist circumference and the circumference at the maximal gluteal protuberance were used for waist and hip, respectively.
Resting metabolic rate (RMR)
RMR was measured for 30 min by open-circuit dilution technique (model 2900, Sensormedics, Yorba Linda, CA) with calibrations performed before each test. Energy expenditure was calculated by the Weir equation13 and expressed per 24 h. The coefficient of variation of RMR measured under these conditions in our laboratory is 1.1%.
Venous blood samples were taken in the morning after an overnight fast for the measurement of plasma leptin. The samples were transferred into chilled tubes containing 1 g EDTA/l blood, and plasma was separated by centrifugation at 4°C for 15 min at 2000 g. All samples were stored at −70°C until analysis. Leptin was measured in duplicate using radioimmunoassay (Linco, St Louis, MO). The interassay and intrassay CVs were 3.5 and 5.2%, respectively.
Venous blood samples were taken for the measurement of lipoprotein lipids after a 12 h overnight fast. The samples were transferred into chilled tubes containing 1 g EDTA/l blood, and plasma was separated by centrifugation by 4°C for 15 min at 2000 g. The reported blood lipid values are the mean of values obtained from blood samples drawn on two different days. Total cholesterol, triglyceride, HDL-C, and LDL-C were measured as previously described.14
Adipose cell metabolism
For two days prior to the fat biopsy, nutrient intake was controlled by providing each subject with a eucaloric diet composed of 50–55% carbohydrate, 15–20% protein, and 30% fat, with 300–400 mg cholesterol and a polyunsaturated-to-saturated fat ratio of 0.6:0.8. The amount of energy given to each subject was estimated from 7 day food records and from estimates of energy expenditure based on the Harris–Benedict equation.15 After an overnight fast, 2–4 g of subcutaneous adipose tissue was obtained under local anesthesia (1% xylocaine) from both the abdominal and gluteal regions by aspiration with a 16-gage needle. Adipose cells were isolated using a modification of the Rodbell method, as previously described.16,17
AT-LPL activity and lipolysis assay
Heparin-releasable LPL activity was measured as previously described.18 AT-LPL activity was expressed as nmol of FFA produced per gram of tissue in 1 min. Glycerol released from adipose cells was used as the index of lipolysis, since it is not re-utilized by the adipose cell.19 Basal, dibutyryl 3′,5′-cyclic monophosphate (dcAMP)- and adenosine deaminase (ADA)-stimulated lipolysis was performed as described elsewhere.17 Pharmacological agents (1 mM dcAMP and 1 U/ml ADA) were added just before the beginning of the incubation to stimulate lypolysis. After 2 h, the reaction was stopped with perchloric acid. The glycerol concentration was measured in the infranatant using an enzymatic fluorometric technique.20 Lipolysis was expressed as μmol glycerol/106 cells/2 h.
All data analyses were completed using SPSS for Windows.21 Statistically significant changes in waist and hip circumferences, energy expenditure and adipose cell metabolism as a result of smoking cessation were determined by paired t-tests. Pearson correlation coefficients were calculated between selected variables and weight gain. All data are presented as mean±standard deviation of the mean (s.d.), with the level of significance set at P<0.05 for all analyses.
The 10 women (37.1±6.2 y, smoking for 20.2±5.8 y, 0.9±0.3 packs/day) who participated in the smoking cessation program were obese (BMI of 31.7±6.4 kg/m2). Five women successfully completed the intervention. There were no statistical differences in the physical or metabolic characteristics, including AT-LPL activity, between the women who completed the program and those who did not complete the program (data not shown). After 4 weeks of smoking cessation, there were significant increases in body weight, with no change in waist, hip, resting energy expenditure, or plasma leptin (Table 1). In addition, there were no changes in triglyceride, total or LDL cholesterol, but there was a tendency for an increase in HDL cholesterol (11%, P=0.08) after 4 weeks of smoking cessation (Table 2).
Smoking cessation was not associated with a change in adipose cell size (μg triglyceride/cell). Gluteal AT-LPL activity (expressed per gram of tissue) significantly increased in all five women by 2.8-fold (1.65±1.30–4.72±3.34 nmol FFA/g/min, P<0.05, Figure 1). Abdominal AT-LPL activity increased in four of five women, but the 2.6-fold increase did not reach statistical significance (1.14±0.88–3.50±3.76 nmol FFA/g/min, P=0.14, Figure 1). There were no changes in gluteal or abdominal adipose tissue basal, ADA-, or dcAMP-stimulated lipolysis (Table 3).
There was a significant relationship between change in body weight and increase in gluteal AT-LPL activity (r=0.89, P<0.05, Figure 2), but no relationship between change in weight and increase in abdominal AT-LPL activity (r=0.54, P=0.35). There was also a strong relationship between the weight gain following smoking cessation and the baseline gluteal AT-LPL activity (r=0.86, P=0.06).
These results suggest that smoking cessation is associated with substantial increases in AT-LPL activity. The significant relationship between the change in body weight following smoking cessation and gluteal AT-LPL activity suggests that changes in adipose tissue metabolism following smoking cessation may contribute to the increase in body weight.
To our knowledge, this is the first study to examine the acute effects of smoking cessation on adipose cell metabolism in humans. Smoking cessation of 4 weeks duration was accompanied by an increase in body weight and in gluteal AT-LPL activity. Since abdominal AT-LPL activity increased in four of the five women who quit smoking, there also might have been a significant increase in LPL activity in the abdominal site had more of the women successfully stopped smoking. These results are also consistent with previous findings which show that subjects with higher initial LPL activity prior to stopping smoking seem to have a greater rate of weight gain following smoking cessation.10
Because of the effects of AT-LPL on the hydrolysis of circulating triglyceride into free fatty acid for uptake and storage by adipose tissue, it may play a major role in the maintenance of body weight and fat stores. An increase in LPL activity suggests an increased efficiency of energy storage, and potentially weight gain. The weight gain seen in these women following smoking cessation was associated with an increase in gluteal AT-LPL activity. This is similar to findings seen in weight reduced individuals, where the amount of weight regained correlates with the increase in AT-LPL activity.22,23,24
There was no change in adipose tissue lipolysis (basal, ADA- or dcAMP-stimulated) after smoking cessation. This is different from previous findings in rats, where acute nicotine infusion increased basal lipolysis and blunted the effects of lipolytic agonists.11 These differences are probably related to the exposure of the rats to nicotine via acute infusion, compared to cigarette smoking in humans, as well as potential differences in rat compared to human adipose cell metabolism.25,26 The results of the present study also differ from the report that nicotine increased lipolysis measured in vivo by microdialysis, suggesting the presence of nicotinic receptors;12 however, in that study nicotine had no effect on lipolysis in isolated human adipose cells, similar to the findings of the present study.12 In addition, the acute effects of nicotine on lipolysis may have been blunted since the women were asked to abstain from smoking for 12 h prior to study. The small number of subjects, along with these experimental constraints, may have negated our ability to show a difference in lipolysis after smoking cessation. It is also possible that smoking cessation affected other metabolic sites in the lipolytic cascade, such as insulin suppression of lipolysis or isoprenaline-stimulated lipolysis. These are potential areas of future investigation.
In spite of a significant weight gain, there was a slight increase in HDL cholesterol (P=0.08), with no change in triglycerides, total or LDL-cholesterol. These findings are similar to those of an earlier report that showed a significant increase in HDL cholesterol and no change in triglycerides, total or LDL cholesterol following 8 weeks of smoking cessation.27
In conclusion, these results suggest that in pre-menopausal women smoking cessation is associated with an increase in AT-LPL activity. This may predispose these women to gain weight following smoking cessation, and may be a deterrent to successful abstinence. Understanding the mechanisms by which this occurs, and methods for modifying this response of LPL may improve the success of smoking cessation programs.
We thank all the subjects who volunteered and the nursing and dietary staff at the GRECC at the Baltimore VAMC for assistance with research studies. This work was supported by a T32 AG00219 training grant, NIH grants R29 AG14066-02 and AG00608, and the Department of Veterans Affairs Baltimore Geriatric Research, Education and Clinical Center.
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BMC Public Health (2007)