Neuropeptide Y and Leptin in Patients with Obstructive Sleep Apnea Syndrome
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美国呼吸和危急护理医学 2005年第1期
Serveis de Aneisis Cliniques and Pneumologia, Hospital Universitari Son Dureta, Institut Universitari de Ciencies de la Salut, Palma de Mallorca; and Servicio de Neumologia, Hospital Txagorritxu, Vitoria-Gasteiz, Spain
ABSTRACT
Neuropeptide Y (NPY) and leptin are two peptides involved in the regulation of body weight, energy balance, and sympathetic tone. This study investigates the independent role of apneas and obesity on NPY and leptin plasma levels in patients with obstructive sleep apnea syndrome (OSAS). To this end we compared their values in 23 obese (body mass index > 30 kg/m2) and 24 nonobese (body mass index < 27 kg/m2) patients with OSAS, and in 19 obese and 18 nonobese control subjects without OSAS. Patients who used continuous positive airway pressure for more than 4 hours/night were reexamined 3 and 12 months later. We found that NPY levels were increased (p < 0.01) in patients with OSAS independently of obesity. Leptin levels were also increased in OSAS but this was mostly associated to obesity. Continuous positive airway pressure treatment reduced NPY levels in all patients and leptin levels only in nonobese patients (p < 0.01). We concluded that NPY and leptin plasma levels are increased in patients with OSAS. Yet, whereas the former appear independent of obesity, the latter are mostly associated with obesity.
Key Words: cardiovascular risk continuous positive airway pressure obesity sympathetic nervous system
Obstructive sleep apnea syndrome (OSAS) is a common disorder characterized by excessive daytime sleepiness and repeated episodes of upper airway obstruction during sleep and nocturnal hypoxemia (1). Metabolic abnormalities and increased sympathetic activity are frequent in patients with OSAS (2eC5). However, the role of OSAS in their pathogenesis is unclear because they also occur in obese patients (6eC8) and many patients with OSAS are obese (9, 10). This study sought to delineate the independent role of OSAS and obesity on two key peptides (leptin and neuropeptide Y [NPY]) involved in the regulation of body weight, energy balance, and sympathetic tone (11eC14).
Leptin is a circulating hormone produced by adipocytes (13) whose plasma levels are increased in obese individuals (15). Leptin levels also appear increased in patients with OSAS (4, 16). It has been suggested that these abnormal leptin levels are related to the abnormal sympathetic activity that characterize OSAS (2, 3, 5), but the potential confounding effect of obesity has not been formally excluded. NPY is a neurotransmitter that interacts with leptin in the regulation of sympathetic activity, body weight, and energy balance (11, 13). Potential abnormalities of NPY in patients with OSAS have been seldom studied (17), and its interaction with leptin levels and obesity has not been directly assessed.
To investigate the independent contribution of sleep apnea and obesity to plasma levels of NPY and leptin, we compared their concentrations in obese (body mass index [BMI] > 30 kg/m2) and nonobese patients with OSAS (BMI < 27 kg/m2), as well as in a control group of subjects without OSAS (with and without obesity). To assess the biological effects derived from the therapeutic suppression of the apneic events, measurements were repeated in patients with OSAS 3 and 12 months after treatment with continuous positive airway pressure (CPAP) had been started. Some of the results of this study have been previously reported in the form of an abstract (18).
METHODS
Subjects and Ethics
Patients were recruited prospectively from those who attended our sleep unit from March 2001 to April 2002. All patients were male, had an apnea-hypopnea index of 20/hour or greater, and were eligible for CPAP treatment (19). Patients were considered obese when their BMI was higher than 30 kg/m2 and nonobese when it was lower than 27 kg/m2 (20, 21). Obese patients with OSAS (n = 23) were recruited consecutively, whereas nonobese patients with OSAS (n = 24) were matched to obese patients for age (± 5 years), apnea-hypopnea index (±10/hour), and subjective daytime sleepiness (Epworth scale ± 2) (22). We excluded patients who suffered from any chronic disease (chronic obstructive pulmonary disease, diabetes mellitus, liver cirrhosis, thyroid dysfunction, rheumatoid arthritis, chronic renal failure and/or psychiatric disorders) or were taking any type of medication. This information was based on physician-adjudicated condition. We screened 85 eligible patients. Thirty-eight were excluded due to the presence of chronic obstructive pulmonary disease (n = 11), ischemic heart disease (n = 11), diabetes (n = 6), cancer (n = 3), refusal to sign informed consent (n = 3), chronic renal failure (n = 2), liver cirrhosis (n = 1), alcoholism (n = 1), depression (n = 1), amyotrophic lateral sclerosis (n = 1), and pulmonary fibrosis (n = 1). Some patients met more than one exclusion criterion. Thus, we finally studied 47 male patients with OSAS whose systemic inflammatory profile has been published recently (21) . The diagnosis of OSAS was established by full polysomnography (Ultrasom, Nicolett, Madison,WI), which included recording of oronasal flow, thoracoabdominal movements, electrocardiography, submental and pretibial electromyography, electrooculography, electroencefalography, and trancutaneous measurement of arterial oxygen saturation. Apnea was defined by the absence of airflow for more than 10 seconds. Hypopnea was defined as any airflow reduction that lasted for more than 10 seconds and resulted in arousal or oxygen desaturation. We considered desaturation a decrease in SaO2 greater than 4% (23). The apnea-hypopnea index was defined as the sum of the number of apneas plus hypopneas per hour of sleep. Blood pressure was measured between 9:00 and 10:00 A.M. with a mercury sphygmomanometer with the subject seated using international recommendations (24). The mean value of two measurements was used for analysis. Arterial hypertension was diagnosed if systolic blood pressure was greater than 140 mm Hg or diastolic pressure was greater than 90 mm Hg (24). Patients were studied three times: at diagnosis and after having been treated effectively with CPAP (REM Star; Respironics, Murrysville, PA) during 3 and 12 months. Compliance with treatment was checked by the timer built into the CPAP device. Twenty patients (10 obese and 10 nonobese) who did not use the device for a minimum of 4 hours/night on average were excluded from the follow-up analysis.
We also included in the study 37 nonsmoker males of similar age without a personal or family history of cardiovascular disease or diabetes. Nonobese healthy control subjects were nonsnorers recruited from nonmedical staff of our hospital. In these subjects, the diagnosis of OSAS was excluded by a cardiorespiratory sleep study that recorded nasal flow, thoracic movements, heart rate, snoring, body position, and transcutaneus oxyhemoglobin saturation (EdenTec, Eden Prairie, MN). We screened 19 potential nonobese control subjects. One subject was excluded due to the presence of respiratory abnormalities in the sleep study, so we finally included 18 nonobese control subjects. Obese control subjects were recruited from those obese subjects who attended our sleep unit and in whom OSAS was excluded by full polysomnography, as described previously. We screened 25 subjects but we had to exclude six for the following reasons: renal dysfunction (n = 1), periodic leg movement (n = 1), suspicion of narcolepsy (n = 1), diabetes (n = 1) or hypothyroidism (n = 1), and one subject refused to sign informed consent and was therefore excluded from the study. We finally included 19 obese control subjects.
The study was approved by the Ethics Committee of our institution, and all participants signed their consent after being fully informed of its goal and characteristics.
Measurements
After fasting overnight, venous blood samples were obtained between 8:00 and 10:00 A.M. Blood was centrifuged, and serum was immediately separated in aliquots and stored at eC80°C until analysis.
Plasma concentrations of NPY and leptin were determined by radioimmunoassay following the instructions of the manufacturer: NPY radioimmunoassay (Euro-Diagnostica, Malm, Sweden) and human leptin immunoradiometric assays (Diagnostic Systems Laboratories Inc.,Webster, TX and Diagnostic Products Corporation, Los Angeles, CA). Measurements were always done in duplicate and mean values were used for analysis. The coefficients of variation were 2.6% (intraassay coefficient of variation) and 7.2% (interassay coefficients of variation) for NPY determinations, and 3.7% and 5.3%, respectively, for leptin determinations.
To characterize the hormone profile of participants, we also quantified at baseline the plasma concentration of thyrotrophin (TSH), cortisol, growth hormone (GH), and insulin by commercial chemiluminiscent assays: TSH and cortisol on a Immulite 2,000 analyser (Diagnostic Products Corporation) and GH and insulin on an Advia Centaur analyzer (Bayer, NY). Likewise, we determined the concentration of insulin-like growth factor-I by an immunoradioimmumoassay (IGF-I IRMA; Nichols Diagnostics, San Juan Capistrano, CA).
Statistical Analysis
Based on previously published results of plasma leptin levels in both obese (25) and nonobese subjects (16), and assuming an error of 0.5 and a error of 0.1, we calculated a minimum sample size of 18 individuals per group. Because we expected a dropout rate during CPAP treatment of approximately 25%, we increased the number of patients recruited accordingly. Results are shown as mean ± SEM. All variables were tested for normality (Kolgomorov-Smirnov test). One-way analysis of variance, followed by post hoc contrast (least significant difference [LSD]) if appropriate, and chi-square test for proportions were used to assess the statistical significance of differences between groups and the effects of CPAP therapy within each group of patients. The correlation between variables was explored using the Spearman test. A p value lower than 0.05 was considered statistically significant.
RESULTS
Clinical Data
Table 1 shows the main clinical characteristics and hormone profile of all subjects studied. Age was similar in patients and control subjects. BMI values of obese patients were not different from those of obese control subjects; likewise, BMI values were not different between nonobese patients and nonobese control subjects (Table 1). Obese patients showed higher blood pressure values than nonobese patients or control subjects, but the prevalence of hypertension was not significantly different in the two OSAS groups (39 vs. 29%, p = 0.547). The percentage of smokers was not significantly different in the two groups of patients (43.5 vs. 37.5%, p = 0.770). Likewise, disease severity (as assessed by the apnea-hypopnea index ) was similar between obese and nonobese patients (Table 1).
Effects of Obesity
To investigate the effects of obesity, we compared the results obtained in obese and nonobese control subjects. NPY plasma levels were similar in both groups (57.6 ± 4.5 vs. 59.9 ± 5.6 pmol/L; mean difference: 3.4 pmol/L; 95% confidence interval [CI]: eC13.9 to +18.7 pmol/L) (Figure 1A). In contrast, obese control subjects showed higher leptin levels (24.7 ± 3.5 vs. 5.5 ± 0.5 ng/ml, p < 0.001, mean difference: eC19.2 ng/ml; 95% CI: eC25.6 to eC12.7 ng/ml) (Figure 1B). Cortisol levels were also increased in obese control subjects (Table 1). TSH, human GH, and insulin-like growth factor-I values were not different between both groups (Table 1).
Effects of OSAS
To investigate the effects of OSAS, we compared the results obtained in nonobese control subjects and nonobese patients. The latter showed higher plasma levels of both NPY (85.5 ± 5.1 vs. 59.9 ± 5.6 pmol/L, p < 0.01; mean difference: eC25.5 pmol/L; 95% CI: eC41.2 to eC9.8 pmol/L) and leptin (11.5 ± 1.6 vs. 5.5 ± 0.5 ng/ml, p < 0.01; mean difference: eC6.0 ng/ml; 95% CI: eC12.2 to +0.1 ng/ml) (Figure 1). Plasma levels of TSH, cortisol, human GH, and insulin-like growth factor-I in these two groups were similar (Table 1).
Combined Effects of OSAS and Obesity
To assess the effects of OSAS when combined with obesity, we compared the results of obese patients with those obtained in obese control subjects. NPY levels were significantly higher in the former (78.0 ± 5.9 vs. 57.6 ± 4.5 pmol/L, p < 0.005; mean difference: eC20.4 pmol/L; 95% CI: eC35.2 to eC5.5 pmol/L). In contrast, leptin levels were similar in both groups (25.4 ± 1.7 vs. 24.7 ± 3.5 ng/ml; mean difference: eC0.6 ng/ml; 95% CI: eC6.5 to +5.3 ng/ml) (Figure 1). Obese control subjects showed higher cortisol plasma levels than obese patients, but no differences were observed in TSH, human GH, and insulin-like growth factor-I levels between these two groups (Table 1).
Effects of CPAP
Fourteen nonobese and 12 obese patients used CPAP for more than 4 hours/night on average (mean usage, 5.7 ± 1.4 hours/night). These patients were studied again 3 and 12 months after having started treatment. BMI values did not change in obese (3 months: 33.6 ± 0.9 kg/m2; 12 months: 33.5 ± 0.9 kg/m2) or nonobese patients (3 months: 26.3 ± 0.3 kg/m2; 12 months: 26.3 ± 0.3 kg/m2). As shown in Figure 2A, NPY levels decreased significantly after 12 months under CPAP both in obese (baseline: 73.5 ± 5.5 pmol/L; 3 months: 69.6 ± 6.5 pmol/L; 12 months: 65.9 ± 6.6 pmol/L; p < 0.01) and nonobese patients (baseline: 85.1 ± 6.9 pmol/L; 3 months: 78.1 ± 6.7 pmol/L; 12 months: 67.6 ± 6.6 pmol/L; p < 0.01). CPAP treatment reduced leptin levels, but changes reached statistical significance only in nonobese patients (baseline: 11.0 ± 1.9 ng/ml; 3 months: 10.5 ± 1.8 ng/ml; 12 months: 9.2 ± 1.5 ng/ml; p < 0.01) (Figure 2B). CPAP treatment did not modify the plasma levels of TSH, cortisol, human GH, and insulin-like growth factor-1, either in obese or nonobese patients (data not shown). The changes in leptin or NPY levels after treatment with CPAP were not related to age, weight, blood pressure, or the levels of these peptides determined at baseline. Systolic blood pressure decreased significantly after 1 year on CPAP in nonobese patients (127 ± 3 to 116 ± 3 mm Hg, p < 0.005) and showed a similar trend in obese patients (135 ± 3 to 127 ± 6 mm Hg), although changes did not reach the level of statistical significance. Diastolic blood pressure did not change in any group.
DISCUSSION
This study provides two main observations of interest. First, NPY levels are increased in OSAS independently of obesity (Figure 1A). In keeping with this observation, we found that treatment with CPAP decreased NPY levels both in obese and nonobese patients (Figure 2A). Second, leptin plasma concentration is also increased in OSAS. However, this is mostly associated with obesity and only to a smaller degree with sleep apnea (Figure 1B). Accordingly, we found that CPAP therapy had a small (albeit significant) effect on leptin plasma levels only in nonobese patients (Figure 2B).
To our knowledge only one previous study has investigated NPY levels in OSAS and it failed to demonstrate that they were increased in these patients (17). This is in contrast to our observations (Figure 1). Several factors can contribute to explaining this difference. First, the methodology used to quantify NYP levels in both studies was different. Second, our study included a substantially higher number of patients (n = 47) than the previous one (n = 11) (17). Finally, as acknowledged by the authors of the previous study, the fact that they were not able to find increased NPY levels in their patients with OSAS was likely related to the low sympathetic activity of the subjects included (17).
In contrast, we found that NPY plasma levels were increased in patients with OSAS. Furthermore, our results suggest that these increased levels are associated with the presence of apneas during sleep and not to obesity because, first, they were increased both in obese and nonobese patients (Figure 1), second, they were similar in control subjects independent of obesity (Figure 1), and, finally, they decreased significantly after CPAP therapy (Figure 2) whereas BMI values did not change.
NPY has multiple cardiovascular effects, including vasoconstriction, stimulation of vascular smooth muscle proliferation, and vascular hypertrophy (26, 27). Thus, the increased NPY levels observed in OSAS can potentially contribute to the pathogenesis of cardiovascular disease in these patients (28, 29). In fact, we observed a decrease of both NPY levels and systolic blood pressure after CPAP treatment (Figure 2A). In turn, this might be related to the effects of CPAP on hypoxemia, arousals, and/or sympathetic drive (30, 31).
Leptin is a circulating hormone produced by adipocytes. It is a key physiologic regulator of both energy intake and expenditure (thus, body weight) and sympathetic activity (12, 13). Several previous studies have reported increased plasma leptin levels in patients with OSAS (3, 4, 16, 32). However, the relationship between leptin and OSAS is far from being resolved mainly because the potential confounding role of obesity has not been specifically considered before. Our results can help to better delineate this relationship because we found that: (1) leptin levels in nonobese patients with OSAS were higher than in nonobese control subjects (Figure 1), an observation not previously reported; (2) leptin levels in obese patients with OSAS were not different from obese control subjects (Figure 1), which is in contrast with some (3, 4, 16, 32) but not all previous studies (33); (3) leptin levels in nonobese patients with OSAS were significantly lower than in obese ones (Figure 1), an observation that reproduces what occurs in control subjects (Figure 1) but that has not been reported before in patients with OSAS; and finally, (4) leptin levels did not decrease with CPAP therapy in obese patients with OSAS and changed only marginally in nonobese patients (Figure 2). This may seem in contrast to previous studies that have generally shown that leptin levels decrease significantly with CPAP (3eC5, 34). However, these changes were generally small in absolute amount (3eC5, 34) and not very different from those determined here. In fact, we cannot exclude that changes in leptin levels after CPAP in obese patients with OSAS may have reached statistical significance if a higher number of patients had been studied. In summary, taking all these observations into account our findings suggest that the increased leptin levels described so far in patients with OSAS is mostly associated with obesity and not with the disease itself.
Some potential limitations of our study deserve comment. First, patients and control subjects were matched for BMI. However, this does not exclude potential differences in body fat distribution. Second, OSAS was excluded in nonobese control subjects by respiratory (not full) polysomnography. This can potentially misclassify these subjects. However, we believe that it is highly unlikely that a nonobese, nonsnorer subject without daytime sleepiness and without evidence of respiratory disturbances during sleep suffers from OSAS. Finally, we had to exclude a significant number of potential candidates in the study due to the presence of comorbid conditions. Although this was essential to dissect the independent effects of OSAS and obesity, this may limit the generalization of our results.
In summary, this study shows that NPY and leptin plasma levels are increased in patients with OSAS. Yet, whereas the former appears independent of obesity and treatable with CPAP, the latter is mostly associated with obesity and, accordingly, treatment with CPAP has a small effect. Overall, these results contribute to better delineation of the independent effects of OSAS and obesity on two peptides involved in the regulation of body weight, energy balance, and sympathetic tone.
Acknowledgments
The authors thank Dr. Felipe Aizpuru for his assistance with data analysis.
REFERENCES
Douglas NJ, Polo O. Pathogenesis of sleep apnoea/hypopnoea syndrome. Lancet 1994;344:653eC655.
Brown LK. A waist is a terrible thing to mind: central obesity, the metabolic syndrome, and sleep apnea hypopnea syndrome. Chest 2002;122:774eC778.
Phillips BG, Kato M, Narkiewicz K, Choe I, Somers VK. Increases in leptin levels, sympathetic drive, and weight gain in obstructive sleep apnea. Am J Physiol Heart Circ Physiol 2000;279:H234eCH237.
Chin K, Shimizu K, Nakamura T, Narai N, Masuzaki H, Ogawa Y, Mishima M, Nakamura T, Nakao K, Ohi M. Changes in intra-abdominal visceral fat and serum leptin levels in patients with obstructive sleep apnea syndrome following nasal continuous positive airway pressure therapy. Circulation 1999;100:706eC712.
Shimizu K, Chin K, Nakamura T, Masuzaki H, Ogawa Y, Hosokawa R, Niimi A, Hattori N, Nohara R, Sasayama S, et al. Plasma leptin levels and cardiac sympathetic function in patients with obstructive sleep apnoea-hypopnoea syndrome. Thorax 2002;57:429eC434.
Flier JS, Maratos-Flier E. Obesity and the hypothalamus: novel peptides for new pathways. Cell 1998;92:437eC440.
Mark AL, Correia M, Morgan DA, Shaffer RA, Haynes WG. Obesity-induced hypertension: new concepts from the emerging biology of obesity. Hypertension 1999;33:537eC541.
Hall JE, Hildebrandt DA, Kuo J. Obesity hypertension: role of leptin and sympathetic nervous system. Am J Hypertens 2001;14:103SeC115S.
Young T, Palta M, Dempsey J, Skatrud J, Weber S, Badr S. The occurrence of sleep-disordered breathing among middle-aged adults. N Engl J Med 1993;328:1230eC1235.
Rajala R, Partinen M, Sane T, Pelkonen R, Huikuri K, Seppalainen AM. Obstructive sleep apnoea syndrome in morbidly obese patients. J Intern Med 1991;230:125eC129.
Matsumura K, Tsuchihashi T, Abe I. Central cardiovascular action of neuropeptide Y in conscious rabbits. Hypertension 2000;36:1040eC1044.
Aizawa-Abe M, Ogawa Y, Masuzaki H, Ebihara K, Satoh N, Iwai H, Matsuoka N, Hayashi T, Hosoda K, Inove G, et al. Pathophysiological role of leptin in obesity-related hypertension. J Clin Invest 2000;105:1243eC1252.
Friedman J, Halaas J. Leptin and the regulation of body weight in mammals. Nature 1998;395:763eC770.
Stanley BG. Neuropeptide Y in multiple hypothalamic sites control eating behavior, endocrine and autonomic systems for energy balance. In: Colmers WF, Wahlestedt C, editors. The biology of neuropeptide Y and related peptides. Totowa, NJ: Humana Press; 1993. pp. 457eC509.
Lonnquist F, Arner P, Nordfors L, Schalling M. Overexpression of the obese (ob) gene in adipose tissue of human obese subjects. Nat Med 1995;1:950eC953.
Ip MSM, Lam KSL, Ho C, Tsang KWT, Lam W. Serum leptin and vascular risk factors in obstructive sleep apnea. Chest 2000;118:580eC586.
Carlson JA, Hedner JA, Elam M, Ejnell H, Sellgren J, Gunar Wallin B. Augmented resting sympathetic activity in awake patients with obstructive sleep apnea. Chest 1993;103:1763eC1768.
Barcelo A, Barbe F, Llompart E, Mayoralas LR, Vila M, Ladaria A, Bosch M, Agusti AGN. Influencia de la obesidad en los niveles plasmeicos de neuropeeptido Y en pacientes con SAS . Arch Bronconeumol 2002;38:21.
Montserrat JM, Amilibia J, Barbe F, Capote F, Duran J, Mangado NG, Jimenez A, Marin JM, Masa F, Teran J. Tratamiento del se猲drome de las apneas-hipoapneas durante el sueo. Arch Bronconeumol 1998;24:206eC208.
U.S. Public Health Service. Clinical guidelines on the identification, evaluation and treatment of overweight and obesity in adults. Bethesda, MD; 1998. National Institutes of Health Publication No. 98-4083.
Barcelo A, Barbe F, Llompart E, Mayoralas LR, Ladaria A, Bosch M, Aguste?AGN. Effects of obesity on C-reactive protein level and metabolic disturbances in male patients with obstructive sleep apnea. Am J Med 2004;117:118eC121.
Johns MW. A new method for measuring daytime sleepiness: the Epworth sleepiness scale. Sleep 1991;14:540eC545.
Carrera M, Barbe F, Agusti AGN. Does the number of hypopnoeas influence therapy in patients with obstructive sleep apnoea Respir Med 1998;92:1028eC1031.
Perloff D, Grim C, Flack J, Frohlich ED, Hill M, McDonald M, Morgensterm BZ. Human blood pressure determination by sphygmomanometry. Circulation 1993;88:2460eC2470.
Vgontzas AN, Papanicolaou DA, Bixler EO, Hopper K, Lotsikas A, Lin HM. Sleep apnea and daytime sleepiness and fatigue: relation to visceral obesity, insulin resistance and hypercytokinemia. J Clin Endocrinol Metab 2000;85:1151eC1158.
Zukowska-Grojec Z, Wahlestedtt C. Origin and actions of neuropeptide Y in the cardiovascular system. In: Colmers WF, Wahlestedt C, editors. The biology of neuropeptide Y and related peptides. Totowa, NJ: Humana Press; 1993. pp. 315eC388.
Zukowska-Grojec Z, Karwatowska-Prokopczuk E, Rose W, Rone J, Movafagh S, Ji H, Yeh Y, Chen WT, Kleinman HK, Grouzmann E. Neuropeptide Y: a novel angiogenic factor from the sympathetic nerves and endothelium. Circ Res 1998;83:187eC195.
Gullestad L, Jorgensen B, Thorvald B, Pernow J, Lundberg JM, Dota CD, Hall C, Simonsen S, Ablad B. Postexercise ischemia is associated with increased neuropeptide Y in patients with coronary artery disease. Circulation 2000;102:987eC993.
Malmstrom RE. Pharmacology of neuropeptide Y receptor antagonists: focus on cardiovascular functions. Eur J Pharmacol 2002;447:11eC30.
Kaijser L, Pernow J, Berglund B, Grubbstrom J, Lundberg JM. Neuropeptide Y release from human heart is enhanced during prolonged exercise in hypoxia. J Appl Physiol 1994;76:1346eC1349.
Kask A, Harro J, von horsten S, Redrobe JP, Dumont Y, Quirion R. The neurocircuitry and receptor subtypes mediating anxiolytic-like effects of neuropeptide Y. Neurosci Biobehav Rev 2002;26:259eC283.
Harsch IA, Konturek PC, Koebnick C, Kuehnlein PP, Fuchs FS, Pour Schahin S, Wiest GH, Hahn EG, Lohmann T, Ficker JH. Leptin and ghrelin levels in patients with obstructive sleep apnea. Eur Respir J 2003;22:251eC257.
Schfer H, Pauleit D, Sudhop T, Gouni-Berthold I, Ewig S, Berthold HK. Body fat distribution, serum leptin, and cardiovascular risk factors in men with obstructive sleep apnea. Chest 2002;122:829eC839.
Sanner BM, Kollhosser P, Buechner N, Zidek W, Tepel M. Influence of treatment on leptin levels in patients with obstructive sleep apnoea. Eur Respir J 2004;23:601eC604.(Antonia Barcele? Ferran B)
ABSTRACT
Neuropeptide Y (NPY) and leptin are two peptides involved in the regulation of body weight, energy balance, and sympathetic tone. This study investigates the independent role of apneas and obesity on NPY and leptin plasma levels in patients with obstructive sleep apnea syndrome (OSAS). To this end we compared their values in 23 obese (body mass index > 30 kg/m2) and 24 nonobese (body mass index < 27 kg/m2) patients with OSAS, and in 19 obese and 18 nonobese control subjects without OSAS. Patients who used continuous positive airway pressure for more than 4 hours/night were reexamined 3 and 12 months later. We found that NPY levels were increased (p < 0.01) in patients with OSAS independently of obesity. Leptin levels were also increased in OSAS but this was mostly associated to obesity. Continuous positive airway pressure treatment reduced NPY levels in all patients and leptin levels only in nonobese patients (p < 0.01). We concluded that NPY and leptin plasma levels are increased in patients with OSAS. Yet, whereas the former appear independent of obesity, the latter are mostly associated with obesity.
Key Words: cardiovascular risk continuous positive airway pressure obesity sympathetic nervous system
Obstructive sleep apnea syndrome (OSAS) is a common disorder characterized by excessive daytime sleepiness and repeated episodes of upper airway obstruction during sleep and nocturnal hypoxemia (1). Metabolic abnormalities and increased sympathetic activity are frequent in patients with OSAS (2eC5). However, the role of OSAS in their pathogenesis is unclear because they also occur in obese patients (6eC8) and many patients with OSAS are obese (9, 10). This study sought to delineate the independent role of OSAS and obesity on two key peptides (leptin and neuropeptide Y [NPY]) involved in the regulation of body weight, energy balance, and sympathetic tone (11eC14).
Leptin is a circulating hormone produced by adipocytes (13) whose plasma levels are increased in obese individuals (15). Leptin levels also appear increased in patients with OSAS (4, 16). It has been suggested that these abnormal leptin levels are related to the abnormal sympathetic activity that characterize OSAS (2, 3, 5), but the potential confounding effect of obesity has not been formally excluded. NPY is a neurotransmitter that interacts with leptin in the regulation of sympathetic activity, body weight, and energy balance (11, 13). Potential abnormalities of NPY in patients with OSAS have been seldom studied (17), and its interaction with leptin levels and obesity has not been directly assessed.
To investigate the independent contribution of sleep apnea and obesity to plasma levels of NPY and leptin, we compared their concentrations in obese (body mass index [BMI] > 30 kg/m2) and nonobese patients with OSAS (BMI < 27 kg/m2), as well as in a control group of subjects without OSAS (with and without obesity). To assess the biological effects derived from the therapeutic suppression of the apneic events, measurements were repeated in patients with OSAS 3 and 12 months after treatment with continuous positive airway pressure (CPAP) had been started. Some of the results of this study have been previously reported in the form of an abstract (18).
METHODS
Subjects and Ethics
Patients were recruited prospectively from those who attended our sleep unit from March 2001 to April 2002. All patients were male, had an apnea-hypopnea index of 20/hour or greater, and were eligible for CPAP treatment (19). Patients were considered obese when their BMI was higher than 30 kg/m2 and nonobese when it was lower than 27 kg/m2 (20, 21). Obese patients with OSAS (n = 23) were recruited consecutively, whereas nonobese patients with OSAS (n = 24) were matched to obese patients for age (± 5 years), apnea-hypopnea index (±10/hour), and subjective daytime sleepiness (Epworth scale ± 2) (22). We excluded patients who suffered from any chronic disease (chronic obstructive pulmonary disease, diabetes mellitus, liver cirrhosis, thyroid dysfunction, rheumatoid arthritis, chronic renal failure and/or psychiatric disorders) or were taking any type of medication. This information was based on physician-adjudicated condition. We screened 85 eligible patients. Thirty-eight were excluded due to the presence of chronic obstructive pulmonary disease (n = 11), ischemic heart disease (n = 11), diabetes (n = 6), cancer (n = 3), refusal to sign informed consent (n = 3), chronic renal failure (n = 2), liver cirrhosis (n = 1), alcoholism (n = 1), depression (n = 1), amyotrophic lateral sclerosis (n = 1), and pulmonary fibrosis (n = 1). Some patients met more than one exclusion criterion. Thus, we finally studied 47 male patients with OSAS whose systemic inflammatory profile has been published recently (21) . The diagnosis of OSAS was established by full polysomnography (Ultrasom, Nicolett, Madison,WI), which included recording of oronasal flow, thoracoabdominal movements, electrocardiography, submental and pretibial electromyography, electrooculography, electroencefalography, and trancutaneous measurement of arterial oxygen saturation. Apnea was defined by the absence of airflow for more than 10 seconds. Hypopnea was defined as any airflow reduction that lasted for more than 10 seconds and resulted in arousal or oxygen desaturation. We considered desaturation a decrease in SaO2 greater than 4% (23). The apnea-hypopnea index was defined as the sum of the number of apneas plus hypopneas per hour of sleep. Blood pressure was measured between 9:00 and 10:00 A.M. with a mercury sphygmomanometer with the subject seated using international recommendations (24). The mean value of two measurements was used for analysis. Arterial hypertension was diagnosed if systolic blood pressure was greater than 140 mm Hg or diastolic pressure was greater than 90 mm Hg (24). Patients were studied three times: at diagnosis and after having been treated effectively with CPAP (REM Star; Respironics, Murrysville, PA) during 3 and 12 months. Compliance with treatment was checked by the timer built into the CPAP device. Twenty patients (10 obese and 10 nonobese) who did not use the device for a minimum of 4 hours/night on average were excluded from the follow-up analysis.
We also included in the study 37 nonsmoker males of similar age without a personal or family history of cardiovascular disease or diabetes. Nonobese healthy control subjects were nonsnorers recruited from nonmedical staff of our hospital. In these subjects, the diagnosis of OSAS was excluded by a cardiorespiratory sleep study that recorded nasal flow, thoracic movements, heart rate, snoring, body position, and transcutaneus oxyhemoglobin saturation (EdenTec, Eden Prairie, MN). We screened 19 potential nonobese control subjects. One subject was excluded due to the presence of respiratory abnormalities in the sleep study, so we finally included 18 nonobese control subjects. Obese control subjects were recruited from those obese subjects who attended our sleep unit and in whom OSAS was excluded by full polysomnography, as described previously. We screened 25 subjects but we had to exclude six for the following reasons: renal dysfunction (n = 1), periodic leg movement (n = 1), suspicion of narcolepsy (n = 1), diabetes (n = 1) or hypothyroidism (n = 1), and one subject refused to sign informed consent and was therefore excluded from the study. We finally included 19 obese control subjects.
The study was approved by the Ethics Committee of our institution, and all participants signed their consent after being fully informed of its goal and characteristics.
Measurements
After fasting overnight, venous blood samples were obtained between 8:00 and 10:00 A.M. Blood was centrifuged, and serum was immediately separated in aliquots and stored at eC80°C until analysis.
Plasma concentrations of NPY and leptin were determined by radioimmunoassay following the instructions of the manufacturer: NPY radioimmunoassay (Euro-Diagnostica, Malm, Sweden) and human leptin immunoradiometric assays (Diagnostic Systems Laboratories Inc.,Webster, TX and Diagnostic Products Corporation, Los Angeles, CA). Measurements were always done in duplicate and mean values were used for analysis. The coefficients of variation were 2.6% (intraassay coefficient of variation) and 7.2% (interassay coefficients of variation) for NPY determinations, and 3.7% and 5.3%, respectively, for leptin determinations.
To characterize the hormone profile of participants, we also quantified at baseline the plasma concentration of thyrotrophin (TSH), cortisol, growth hormone (GH), and insulin by commercial chemiluminiscent assays: TSH and cortisol on a Immulite 2,000 analyser (Diagnostic Products Corporation) and GH and insulin on an Advia Centaur analyzer (Bayer, NY). Likewise, we determined the concentration of insulin-like growth factor-I by an immunoradioimmumoassay (IGF-I IRMA; Nichols Diagnostics, San Juan Capistrano, CA).
Statistical Analysis
Based on previously published results of plasma leptin levels in both obese (25) and nonobese subjects (16), and assuming an error of 0.5 and a error of 0.1, we calculated a minimum sample size of 18 individuals per group. Because we expected a dropout rate during CPAP treatment of approximately 25%, we increased the number of patients recruited accordingly. Results are shown as mean ± SEM. All variables were tested for normality (Kolgomorov-Smirnov test). One-way analysis of variance, followed by post hoc contrast (least significant difference [LSD]) if appropriate, and chi-square test for proportions were used to assess the statistical significance of differences between groups and the effects of CPAP therapy within each group of patients. The correlation between variables was explored using the Spearman test. A p value lower than 0.05 was considered statistically significant.
RESULTS
Clinical Data
Table 1 shows the main clinical characteristics and hormone profile of all subjects studied. Age was similar in patients and control subjects. BMI values of obese patients were not different from those of obese control subjects; likewise, BMI values were not different between nonobese patients and nonobese control subjects (Table 1). Obese patients showed higher blood pressure values than nonobese patients or control subjects, but the prevalence of hypertension was not significantly different in the two OSAS groups (39 vs. 29%, p = 0.547). The percentage of smokers was not significantly different in the two groups of patients (43.5 vs. 37.5%, p = 0.770). Likewise, disease severity (as assessed by the apnea-hypopnea index ) was similar between obese and nonobese patients (Table 1).
Effects of Obesity
To investigate the effects of obesity, we compared the results obtained in obese and nonobese control subjects. NPY plasma levels were similar in both groups (57.6 ± 4.5 vs. 59.9 ± 5.6 pmol/L; mean difference: 3.4 pmol/L; 95% confidence interval [CI]: eC13.9 to +18.7 pmol/L) (Figure 1A). In contrast, obese control subjects showed higher leptin levels (24.7 ± 3.5 vs. 5.5 ± 0.5 ng/ml, p < 0.001, mean difference: eC19.2 ng/ml; 95% CI: eC25.6 to eC12.7 ng/ml) (Figure 1B). Cortisol levels were also increased in obese control subjects (Table 1). TSH, human GH, and insulin-like growth factor-I values were not different between both groups (Table 1).
Effects of OSAS
To investigate the effects of OSAS, we compared the results obtained in nonobese control subjects and nonobese patients. The latter showed higher plasma levels of both NPY (85.5 ± 5.1 vs. 59.9 ± 5.6 pmol/L, p < 0.01; mean difference: eC25.5 pmol/L; 95% CI: eC41.2 to eC9.8 pmol/L) and leptin (11.5 ± 1.6 vs. 5.5 ± 0.5 ng/ml, p < 0.01; mean difference: eC6.0 ng/ml; 95% CI: eC12.2 to +0.1 ng/ml) (Figure 1). Plasma levels of TSH, cortisol, human GH, and insulin-like growth factor-I in these two groups were similar (Table 1).
Combined Effects of OSAS and Obesity
To assess the effects of OSAS when combined with obesity, we compared the results of obese patients with those obtained in obese control subjects. NPY levels were significantly higher in the former (78.0 ± 5.9 vs. 57.6 ± 4.5 pmol/L, p < 0.005; mean difference: eC20.4 pmol/L; 95% CI: eC35.2 to eC5.5 pmol/L). In contrast, leptin levels were similar in both groups (25.4 ± 1.7 vs. 24.7 ± 3.5 ng/ml; mean difference: eC0.6 ng/ml; 95% CI: eC6.5 to +5.3 ng/ml) (Figure 1). Obese control subjects showed higher cortisol plasma levels than obese patients, but no differences were observed in TSH, human GH, and insulin-like growth factor-I levels between these two groups (Table 1).
Effects of CPAP
Fourteen nonobese and 12 obese patients used CPAP for more than 4 hours/night on average (mean usage, 5.7 ± 1.4 hours/night). These patients were studied again 3 and 12 months after having started treatment. BMI values did not change in obese (3 months: 33.6 ± 0.9 kg/m2; 12 months: 33.5 ± 0.9 kg/m2) or nonobese patients (3 months: 26.3 ± 0.3 kg/m2; 12 months: 26.3 ± 0.3 kg/m2). As shown in Figure 2A, NPY levels decreased significantly after 12 months under CPAP both in obese (baseline: 73.5 ± 5.5 pmol/L; 3 months: 69.6 ± 6.5 pmol/L; 12 months: 65.9 ± 6.6 pmol/L; p < 0.01) and nonobese patients (baseline: 85.1 ± 6.9 pmol/L; 3 months: 78.1 ± 6.7 pmol/L; 12 months: 67.6 ± 6.6 pmol/L; p < 0.01). CPAP treatment reduced leptin levels, but changes reached statistical significance only in nonobese patients (baseline: 11.0 ± 1.9 ng/ml; 3 months: 10.5 ± 1.8 ng/ml; 12 months: 9.2 ± 1.5 ng/ml; p < 0.01) (Figure 2B). CPAP treatment did not modify the plasma levels of TSH, cortisol, human GH, and insulin-like growth factor-1, either in obese or nonobese patients (data not shown). The changes in leptin or NPY levels after treatment with CPAP were not related to age, weight, blood pressure, or the levels of these peptides determined at baseline. Systolic blood pressure decreased significantly after 1 year on CPAP in nonobese patients (127 ± 3 to 116 ± 3 mm Hg, p < 0.005) and showed a similar trend in obese patients (135 ± 3 to 127 ± 6 mm Hg), although changes did not reach the level of statistical significance. Diastolic blood pressure did not change in any group.
DISCUSSION
This study provides two main observations of interest. First, NPY levels are increased in OSAS independently of obesity (Figure 1A). In keeping with this observation, we found that treatment with CPAP decreased NPY levels both in obese and nonobese patients (Figure 2A). Second, leptin plasma concentration is also increased in OSAS. However, this is mostly associated with obesity and only to a smaller degree with sleep apnea (Figure 1B). Accordingly, we found that CPAP therapy had a small (albeit significant) effect on leptin plasma levels only in nonobese patients (Figure 2B).
To our knowledge only one previous study has investigated NPY levels in OSAS and it failed to demonstrate that they were increased in these patients (17). This is in contrast to our observations (Figure 1). Several factors can contribute to explaining this difference. First, the methodology used to quantify NYP levels in both studies was different. Second, our study included a substantially higher number of patients (n = 47) than the previous one (n = 11) (17). Finally, as acknowledged by the authors of the previous study, the fact that they were not able to find increased NPY levels in their patients with OSAS was likely related to the low sympathetic activity of the subjects included (17).
In contrast, we found that NPY plasma levels were increased in patients with OSAS. Furthermore, our results suggest that these increased levels are associated with the presence of apneas during sleep and not to obesity because, first, they were increased both in obese and nonobese patients (Figure 1), second, they were similar in control subjects independent of obesity (Figure 1), and, finally, they decreased significantly after CPAP therapy (Figure 2) whereas BMI values did not change.
NPY has multiple cardiovascular effects, including vasoconstriction, stimulation of vascular smooth muscle proliferation, and vascular hypertrophy (26, 27). Thus, the increased NPY levels observed in OSAS can potentially contribute to the pathogenesis of cardiovascular disease in these patients (28, 29). In fact, we observed a decrease of both NPY levels and systolic blood pressure after CPAP treatment (Figure 2A). In turn, this might be related to the effects of CPAP on hypoxemia, arousals, and/or sympathetic drive (30, 31).
Leptin is a circulating hormone produced by adipocytes. It is a key physiologic regulator of both energy intake and expenditure (thus, body weight) and sympathetic activity (12, 13). Several previous studies have reported increased plasma leptin levels in patients with OSAS (3, 4, 16, 32). However, the relationship between leptin and OSAS is far from being resolved mainly because the potential confounding role of obesity has not been specifically considered before. Our results can help to better delineate this relationship because we found that: (1) leptin levels in nonobese patients with OSAS were higher than in nonobese control subjects (Figure 1), an observation not previously reported; (2) leptin levels in obese patients with OSAS were not different from obese control subjects (Figure 1), which is in contrast with some (3, 4, 16, 32) but not all previous studies (33); (3) leptin levels in nonobese patients with OSAS were significantly lower than in obese ones (Figure 1), an observation that reproduces what occurs in control subjects (Figure 1) but that has not been reported before in patients with OSAS; and finally, (4) leptin levels did not decrease with CPAP therapy in obese patients with OSAS and changed only marginally in nonobese patients (Figure 2). This may seem in contrast to previous studies that have generally shown that leptin levels decrease significantly with CPAP (3eC5, 34). However, these changes were generally small in absolute amount (3eC5, 34) and not very different from those determined here. In fact, we cannot exclude that changes in leptin levels after CPAP in obese patients with OSAS may have reached statistical significance if a higher number of patients had been studied. In summary, taking all these observations into account our findings suggest that the increased leptin levels described so far in patients with OSAS is mostly associated with obesity and not with the disease itself.
Some potential limitations of our study deserve comment. First, patients and control subjects were matched for BMI. However, this does not exclude potential differences in body fat distribution. Second, OSAS was excluded in nonobese control subjects by respiratory (not full) polysomnography. This can potentially misclassify these subjects. However, we believe that it is highly unlikely that a nonobese, nonsnorer subject without daytime sleepiness and without evidence of respiratory disturbances during sleep suffers from OSAS. Finally, we had to exclude a significant number of potential candidates in the study due to the presence of comorbid conditions. Although this was essential to dissect the independent effects of OSAS and obesity, this may limit the generalization of our results.
In summary, this study shows that NPY and leptin plasma levels are increased in patients with OSAS. Yet, whereas the former appears independent of obesity and treatable with CPAP, the latter is mostly associated with obesity and, accordingly, treatment with CPAP has a small effect. Overall, these results contribute to better delineation of the independent effects of OSAS and obesity on two peptides involved in the regulation of body weight, energy balance, and sympathetic tone.
Acknowledgments
The authors thank Dr. Felipe Aizpuru for his assistance with data analysis.
REFERENCES
Douglas NJ, Polo O. Pathogenesis of sleep apnoea/hypopnoea syndrome. Lancet 1994;344:653eC655.
Brown LK. A waist is a terrible thing to mind: central obesity, the metabolic syndrome, and sleep apnea hypopnea syndrome. Chest 2002;122:774eC778.
Phillips BG, Kato M, Narkiewicz K, Choe I, Somers VK. Increases in leptin levels, sympathetic drive, and weight gain in obstructive sleep apnea. Am J Physiol Heart Circ Physiol 2000;279:H234eCH237.
Chin K, Shimizu K, Nakamura T, Narai N, Masuzaki H, Ogawa Y, Mishima M, Nakamura T, Nakao K, Ohi M. Changes in intra-abdominal visceral fat and serum leptin levels in patients with obstructive sleep apnea syndrome following nasal continuous positive airway pressure therapy. Circulation 1999;100:706eC712.
Shimizu K, Chin K, Nakamura T, Masuzaki H, Ogawa Y, Hosokawa R, Niimi A, Hattori N, Nohara R, Sasayama S, et al. Plasma leptin levels and cardiac sympathetic function in patients with obstructive sleep apnoea-hypopnoea syndrome. Thorax 2002;57:429eC434.
Flier JS, Maratos-Flier E. Obesity and the hypothalamus: novel peptides for new pathways. Cell 1998;92:437eC440.
Mark AL, Correia M, Morgan DA, Shaffer RA, Haynes WG. Obesity-induced hypertension: new concepts from the emerging biology of obesity. Hypertension 1999;33:537eC541.
Hall JE, Hildebrandt DA, Kuo J. Obesity hypertension: role of leptin and sympathetic nervous system. Am J Hypertens 2001;14:103SeC115S.
Young T, Palta M, Dempsey J, Skatrud J, Weber S, Badr S. The occurrence of sleep-disordered breathing among middle-aged adults. N Engl J Med 1993;328:1230eC1235.
Rajala R, Partinen M, Sane T, Pelkonen R, Huikuri K, Seppalainen AM. Obstructive sleep apnoea syndrome in morbidly obese patients. J Intern Med 1991;230:125eC129.
Matsumura K, Tsuchihashi T, Abe I. Central cardiovascular action of neuropeptide Y in conscious rabbits. Hypertension 2000;36:1040eC1044.
Aizawa-Abe M, Ogawa Y, Masuzaki H, Ebihara K, Satoh N, Iwai H, Matsuoka N, Hayashi T, Hosoda K, Inove G, et al. Pathophysiological role of leptin in obesity-related hypertension. J Clin Invest 2000;105:1243eC1252.
Friedman J, Halaas J. Leptin and the regulation of body weight in mammals. Nature 1998;395:763eC770.
Stanley BG. Neuropeptide Y in multiple hypothalamic sites control eating behavior, endocrine and autonomic systems for energy balance. In: Colmers WF, Wahlestedt C, editors. The biology of neuropeptide Y and related peptides. Totowa, NJ: Humana Press; 1993. pp. 457eC509.
Lonnquist F, Arner P, Nordfors L, Schalling M. Overexpression of the obese (ob) gene in adipose tissue of human obese subjects. Nat Med 1995;1:950eC953.
Ip MSM, Lam KSL, Ho C, Tsang KWT, Lam W. Serum leptin and vascular risk factors in obstructive sleep apnea. Chest 2000;118:580eC586.
Carlson JA, Hedner JA, Elam M, Ejnell H, Sellgren J, Gunar Wallin B. Augmented resting sympathetic activity in awake patients with obstructive sleep apnea. Chest 1993;103:1763eC1768.
Barcelo A, Barbe F, Llompart E, Mayoralas LR, Vila M, Ladaria A, Bosch M, Agusti AGN. Influencia de la obesidad en los niveles plasmeicos de neuropeeptido Y en pacientes con SAS . Arch Bronconeumol 2002;38:21.
Montserrat JM, Amilibia J, Barbe F, Capote F, Duran J, Mangado NG, Jimenez A, Marin JM, Masa F, Teran J. Tratamiento del se猲drome de las apneas-hipoapneas durante el sueo. Arch Bronconeumol 1998;24:206eC208.
U.S. Public Health Service. Clinical guidelines on the identification, evaluation and treatment of overweight and obesity in adults. Bethesda, MD; 1998. National Institutes of Health Publication No. 98-4083.
Barcelo A, Barbe F, Llompart E, Mayoralas LR, Ladaria A, Bosch M, Aguste?AGN. Effects of obesity on C-reactive protein level and metabolic disturbances in male patients with obstructive sleep apnea. Am J Med 2004;117:118eC121.
Johns MW. A new method for measuring daytime sleepiness: the Epworth sleepiness scale. Sleep 1991;14:540eC545.
Carrera M, Barbe F, Agusti AGN. Does the number of hypopnoeas influence therapy in patients with obstructive sleep apnoea Respir Med 1998;92:1028eC1031.
Perloff D, Grim C, Flack J, Frohlich ED, Hill M, McDonald M, Morgensterm BZ. Human blood pressure determination by sphygmomanometry. Circulation 1993;88:2460eC2470.
Vgontzas AN, Papanicolaou DA, Bixler EO, Hopper K, Lotsikas A, Lin HM. Sleep apnea and daytime sleepiness and fatigue: relation to visceral obesity, insulin resistance and hypercytokinemia. J Clin Endocrinol Metab 2000;85:1151eC1158.
Zukowska-Grojec Z, Wahlestedtt C. Origin and actions of neuropeptide Y in the cardiovascular system. In: Colmers WF, Wahlestedt C, editors. The biology of neuropeptide Y and related peptides. Totowa, NJ: Humana Press; 1993. pp. 315eC388.
Zukowska-Grojec Z, Karwatowska-Prokopczuk E, Rose W, Rone J, Movafagh S, Ji H, Yeh Y, Chen WT, Kleinman HK, Grouzmann E. Neuropeptide Y: a novel angiogenic factor from the sympathetic nerves and endothelium. Circ Res 1998;83:187eC195.
Gullestad L, Jorgensen B, Thorvald B, Pernow J, Lundberg JM, Dota CD, Hall C, Simonsen S, Ablad B. Postexercise ischemia is associated with increased neuropeptide Y in patients with coronary artery disease. Circulation 2000;102:987eC993.
Malmstrom RE. Pharmacology of neuropeptide Y receptor antagonists: focus on cardiovascular functions. Eur J Pharmacol 2002;447:11eC30.
Kaijser L, Pernow J, Berglund B, Grubbstrom J, Lundberg JM. Neuropeptide Y release from human heart is enhanced during prolonged exercise in hypoxia. J Appl Physiol 1994;76:1346eC1349.
Kask A, Harro J, von horsten S, Redrobe JP, Dumont Y, Quirion R. The neurocircuitry and receptor subtypes mediating anxiolytic-like effects of neuropeptide Y. Neurosci Biobehav Rev 2002;26:259eC283.
Harsch IA, Konturek PC, Koebnick C, Kuehnlein PP, Fuchs FS, Pour Schahin S, Wiest GH, Hahn EG, Lohmann T, Ficker JH. Leptin and ghrelin levels in patients with obstructive sleep apnea. Eur Respir J 2003;22:251eC257.
Schfer H, Pauleit D, Sudhop T, Gouni-Berthold I, Ewig S, Berthold HK. Body fat distribution, serum leptin, and cardiovascular risk factors in men with obstructive sleep apnea. Chest 2002;122:829eC839.
Sanner BM, Kollhosser P, Buechner N, Zidek W, Tepel M. Influence of treatment on leptin levels in patients with obstructive sleep apnoea. Eur Respir J 2004;23:601eC604.(Antonia Barcele? Ferran B)