Plasma Levels of Adiponectin, a Novel Adipocyte-Derived Hormone, in Sleep Apnea

Plasma Levels of Adiponectin, a Novel Adipocyte-Derived Hormone, in Sleep Apnea Robert Wolk, Anna Svatikova, Christy A. Nelson, Apoor S. Gami, Kogilan Govender, Mikolaj Winnicki, and Virend K. Somers Abstract WOLK, ROBERT, ANNA SVATIKOVA, CHRISTY A. NELSON, APOOR S. GAMI, KOGILAN GOVENDER, MIKOLAJ WINNICKI, AND VIREND K. SOMERS. Plasma levels of adiponectin, a novel adipocyte-derived hormone, in sleep apnea. Obes Res. 2005;13:186 190. Objective: Obstructive sleep apnea (OSA) is associated with obesity, sympathetic activation, systemic inflammation, and cardiovascular morbidity. Obesity, -adrenergic agonists, and inflammation are linked to decreased expression and/or secretion of an adipose tissue-derived antiatherogenic hormone, adiponectin. The purpose of the study was to investigate whether OSA affected plasma levels of adiponectin, which might help explain OSA-associated cardiovascular morbidity. Research Methods and Procedures: We randomly selected 68 otherwise healthy male subjects, either with moderate/ severe OSA [apnea-hypopnea index (AHI) 20; n 35] or without OSA (AHI 5; n 33). The diagnosis of OSA was made based on prospective full polysomnography. Adiponectin was measured before polysomnography between 8 and 10 PM. Results: AHI was higher in the OSA group (49.5 4.4 vs. 2.9 0.4 events/h; p 0.001). OSA subjects were also more obese, with greater BMI (33 1 vs. 30 1; p 0.016) and percentage body fat (29 1% vs. 26 1%; p 0.030). Adiponectin levels were 7.67 0.73 and 6.33 0.51 g/ml in the OSA and non-osa groups, respectively, and this difference was significant in covariate analysis (taking into account age, hemodynamic characteristics, Received for review March 19, 2004. Accepted in final form November 11, 2004. The costs of publication of this article were defrayed, in part, by the payment of page charges. This article must, therefore, be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. Department of Internal Medicine, Division of Cardiovascular Diseases, Mayo Clinic, Rochester, Minnesota. Address correspondence to Virend K. Somers, Mayo Foundation, St. Mary s Hospital, DO-4-350, 1216 Second Street SW, Rochester, MN 55902. E-mail: somers.virend@mayo.edu Copyright 2005 NAASO measures of body fat, and OSA severity) (p 0.009). After excluding from both groups the subjects with extreme BMI, such that the OSA and non-osa study cohorts had similar BMI and percentage body fat, subjects with OSA had significantly higher plasma adiponectin (8.49 0.92 vs. 6.32 0.55 g/ml; p 0.042), differences also evident in covariate analysis (p 0.017). Discussion: Plasma adiponectin levels are elevated in otherwise healthy subjects with OSA. Therefore, low adiponectin is unlikely to explain the association between OSA and cardiovascular disease. Key words: adiponectin, sleep apnea Introduction Obstructive sleep apnea (OSA) 1 is associated with cardiovascular morbidity, including an increased risk of endothelial dysfunction and atherosclerosis (1,2). In patients with OSA, atherosclerosis may be promoted by the heightened inflammatory state, as evidenced by elevated C-reactive protein and increased expression of adhesion molecules on leukocytes, with their subsequent adherence to endothelial cells (3 8). There is also evidence that OSA can independently cause various metabolic abnormalities. For example, recent studies have demonstrated an independent association between OSA and insulin resistance, which is seen in both obese and nonobese subjects (9,10). In recent years, a new concept has emerged, which suggests that adipocytes are secretory cells, and adipose tissue is an endocrine organ with important effects on the cardiovascular system (11). Among various adipocyte-derived biologically active peptides, adiponectin has attracted considerable attention due to its association with several cardiovascular risk factors. Specifically, reduced plasma adiponectin levels have been related to endothelial inflammatory responses, the presence of coronary heart disease, 1 Nonstandard abbreviations: OSA, obstructive sleep apnea; AHI, apnea-hypopnea index. 186 OBESITY RESEARCH Vol. 13 No. 1 January 2005

dyslipidemia, insulin resistance, and type 2 diabetes in humans (12 21). The primary aim of the present study was to establish whether OSA affected plasma levels of adiponectin. Our hypothesis was based on the following premises. First, adiponectin expression and/or secretion are decreased by some inflammatory mediators (22 24) and by -adrenergic agonists and camp (22,25,26). Second, OSA is accompanied by systemic inflammation (7,8,27) and persistently increased sympathetic activity (28), which might, therefore, lead to OSA-induced hypoadiponectemia. Plasma adiponectin levels are also decreased in obesity (29,30), which is consistent with increased obesity-related cardiovascular morbidity. However, the finding of low adiponectin levels in obesity is somewhat unexpected and counterintuitive because adiponectin is specifically expressed in adipose tissue and would, therefore, be expected to be elevated in conditions associated with increased body fat. The mechanisms of this paradoxical finding are not known. Because sleep apnea is present in 40% of obese individuals (31), it is conceivable that OSA may influence adiponectin levels in obesity. Research Methods and Procedures Our study population consisted of consecutive subjects recruited in the Mayo Clinic Sleep Disorders Center, who were referred there for clinically indicated sleep study (polysomnography). All study participants were nonsmokers and had never been diagnosed with or treated for OSA. In all subjects, anthropometric measurements were performed, including BMI, body fat, and waist-to-hip ratio. Body fat was measured using the impedance method. Blood pressure was also measured using a sphygmomanometer after a 10- to 15-minute rest in a supine position. (Two consecutive measurements were performed, but only the second reading was recorded and used in the analysis.) All subjects screened for this study underwent prospective overnight polysomnography. An apnea was defined as complete cessation of airflow for at least 10 seconds. Hypopnea was defined as a reduction of respiratory signals for at least 10 seconds associated with oxygen desaturation of 4%. Apnea-hypopnea index (AHI) was calculated as the total number of apnea/hypopnea events per hour of sleep. Subjects with AHI 5 were defined as nonapneics, and in those with AHI 20, the diagnosis of moderate to severe OSA was made. In all study subjects, blood samples were taken before full polysomnography between 8 and 10 PM. Adiponectin was measured by radioimmunoassay using a Human Adiponectin RIA Kit (Linco Research, Inc., St. Charles, MO). The study was approved by the Mayo Clinic Institutional Review Board, and the study was performed in accordance with institutional guidelines. Informed written consent was obtained from each subject. For statistical analysis, a two-way unpaired Student s t test was used. Adiponectin levels were also compared between the OSA and non-osa groups by covariate analysis (using age, systolic and diastolic blood pressure, heart rate, waist-to-hip ratio, BMI, percentage body fat, and AHI as covariates). p 0.05 was considered statistically significant. All data are expressed as mean SEM. Results Overall, 106 consecutive male subjects were recruited for this study. Their OSA status was ascertained prospectively based on the results of overnight polysomnography. Subsequently, we excluded from the study subjects with only mild OSA (6 AHI 20; n 15) and those with cardiovascular or other systemic diseases requiring prescription medication (n 23). Therefore, the final study cohort consisted of 68 male subjects, without cardiovascular disease, newly diagnosed with OSA (n 35), or found to be free of OSA (controls; n 33). The demographic and clinical characteristics of these two groups are shown in Table 1. Compared with non-osa controls, subjects with OSA had significantly higher AHI (48.5 4.4 vs. 2.9 0.4; p 0.001). OSA subjects were also more obese, with greater BMI (33 1 vs. 30 1; p 0.016) and percentage body fat (29 1% vs. 26 1%; p 0.030). Heart rate at rest was also significantly higher in the OSA group (76 2 vs. 69 2 beats/min; p 0.012). Differences between the groups in adiponectin levels did not reach statistical significance (7.67 0.73 vs. 6.33 0.51 g/ml in the OSA and non-osa groups, respectively; p 0.142). However, covariate analysis (taking into account differences between the groups in age, hemodynamic characteristics, measures of body fat, and OSA severity) revealed significantly higher adiponectin levels in those with compared with those without OSA (p 0.009). Because the two study groups differed with respect to BMI and percentage body fat, and because obesity per se is known to affect adiponectin levels (29,30), we performed a subanalysis after excluding from both groups those subjects with extreme BMI values, such that the study cohorts had similar BMI and percentage body fat. The results are shown in Table 2. We found that, at a similar level of obesity, subjects with OSA had significantly elevated plasma adiponectin levels (8.49 0.92 vs. 6.32 0.55 g/ml; p 0.042) (Figure 1). The difference in adiponectin levels between the groups was also significant in covariate analysis (p 0.017). Discussion In the present study, we investigated plasma adiponectin levels in OSA patients and healthy controls. We hypothesized that, by inducing inflammatory responses (such as tumor necrosis factor- or C-reactive protein elevation) OBESITY RESEARCH Vol. 13 No. 1 January 2005 187

Table 1. Characteristics of the study groups Control (n 33) OSA (n 35) p Age (years) 46 2 45 2 0.728 BMI (kg/m 2 ) 30 1 33 1 0.016 Body fat (%) 26 1 29 1 0.030 Waist-to-hip ratio 0.98 0.01 0.99 0.01 0.538 Systolic blood pressure (mm Hg) 131 3 132 2 0.814 Diastolic blood pressure (mm Hg) 78 2 77 1 0.620 Heart rate (beats/min) 69 2 76 2 0.012 AHI (events/h) 2.9 0.4 48.5 4.4 0.001 Lowest O 2 (%) 89 1 78 2 0.001 % of total sleep time with O 2 90% 0.3 0.1 12 3 0.001 Adiponectin ( g/ml) 6.33 0.51 7.67 0.73 0.142* * p 0.009 in covariate analysis. (7,27) and/or by increasing sympathetic activity (28), OSA might decrease plasma levels of adiponectin (22 26). Decreased adiponectin levels in patients with OSA could then partly explain the association between OSA and cardiovascular disease. However, in contrast to our expectations, we found that plasma adiponectin levels were higher in subjects with OSA. Reduced adiponectin may exert detrimental effects on cardiovascular homeostasis (12 21). Therefore, our results suggest that, although OSA is known to be a cardiovascular risk factor (1,2), OSA may also trigger certain compensatory processes, which can be perceived as protective of the cardiovascular system. The mechanisms of this effect of OSA on adiponectin are unknown and remain to be established. However, it is interesting to note that plasma levels of leptin (another biologically active protein released by the adipose tissue) are also elevated in OSA (32,33). These observations suggest that OSA may exert a stimulatory effect on the endocrine function of adipose tissue. An important strength of this study is that only normotensive, nonsmoking, and otherwise healthy male OSA patients were included. Although patients with OSA tend to be more obese, and, therefore, the coexistence of obesity can be an important confounding factor when looking at the Table 2. of obesity Characteristics of the study groups after selection of OSA and non-osa subjects with a similar degree CONTROL (n 29) OSA (n 26) p Age (years) 46 2 46 2 0.976 BMI (kg/m 2 ) 31 1 31 1 0.986 Body fat (%) 27 1 27 1 0.722 Waist-to-hip ratio 0.99 0.01 0.98 0.01 0.495 Systolic blood pressure (mm Hg) 133 3 132 1 0.774 Diastolic blood pressure (mm Hg) 78 2 77 1 0.682 Heart rate (beats/min) 71 2 73 2 0.279 AHI (events/h) 3.0 0.4 44 4 0.001 Lowest O 2 (%) 88 1 79 2 0.001 % of total sleep time with O 2 90% 0.4 0.1 10 2 0.001 Adiponectin ( g/ml) 6.32 0.55 8.49 0.92 0.042* * p 0.017 in covariate analysis. 188 OBESITY RESEARCH Vol. 13 No. 1 January 2005

Figure 1: Plasma adiponectin levels in the OSA (n 26) and control non-osa (n 29) groups. The groups were similar with respect to BMI, body fat, and waist-to-hip ratio. p 0.017 in covariate analysis. effects of OSA, we also confirmed our findings in OSA and non-osa subjects who were similar with respect to various demographic characteristics (including indices of adiposity, such as BMI, body fat, and waist-to-hip ratio) (Table 2). Overnight polysomnographic recordings excluded the presence of occult sleep apnea in the control subjects; hence, any differences in adiponectin between OSA and control subjects were not being masked by undiagnosed OSA in the control group. In conclusion, we have demonstrated that plasma adiponectin levels are elevated in otherwise healthy subjects with OSA, independently of obesity and other variables. Therefore, low adiponectin is unlikely to explain the association between OSA and cardiovascular disease. Furthermore, higher plasma adiponectin in OSA may potentially be a protective mechanism against cardiovascular disease. Acknowledgments This work was supported by NIH Grants HL-61560, HL-65176, HL-70302, and MO1-RR00585. References 1. Leung RS, Bradley TD. Sleep apnea and cardiovascular disease. Am J Respir Crit Care Med. 2001;164:2147 65. 2. Wolk R, Kara T, Somers VK. Sleep-disordered breathing and cardiovascular disease. Circulation. 2003;108:9 12. 3. Ohga E, Nagase T, Tomita T, et al. Increased levels of circulating ICAM-1, VCAM-1, and L-selectin in obstructive sleep apnea syndrome. J Appl Physiol. 1999;87:10 4. 4. Chin K, Nakamura T, Shimizu K, et al. Effects of nasal continuous positive airway pressure on soluble cell adhesion molecules in patients with obstructive sleep apnea syndrome. Am J Med. 2000;109:562 7. 5. Dyugovskaya L, Lavie P, Lavie L. Increased adhesion molecules expression and production of reactive oxygen species in leukocytes of sleep apnea patients. Am J Respir Crit Care Med. 2002;165:934 9. 6. El-Solh AA, Mador MJ, Sikka P, Dhillon RS, Amsterdam D, Grant BJ. Adhesion molecules in patients with coronary artery disease and moderate-to-severe obstructive sleep apnea. Chest. 2002;121:1541 7. 7. Shamsuzzaman AS, Winnicki M, Lanfranchi P, et al. Elevated C-reactive protein in patients with obstructive sleep apnea. Circulation. 2002;105:2462 4. 8. Yokoe T, Minoguchi K, Matsuo H, et al. Elevated levels of C-reactive protein and interleukin-6 in patients with obstructive sleep apnea syndrome are decreased by nasal continuous positive airway pressure. Circulation. 2003;107:1129 34. 9. Ip MS, Lam B, Ng MM, Lam WK, Tsang KW, Lam KS. Obstructive sleep apnea is independently associated with insulin resistance. Am J Respir Crit Care Med. 2002;165:670 6. 10. Punjabi NM, Sorkin JD, Katzel LI, Goldberg AP, Schwartz AR, Smith PL. Sleep-disordered breathing and insulin resistance in middle-aged and overweight men. Am J Respir Crit Care Med. 2002;165:677 82. 11. Wajchenberg BL. Subcutaneous and visceral adipose tissue: their relation to the metabolic syndrome. Endocr Rev. 2000; 21:697 738. 12. Ouchi N, Kihara S, Arita Y, et al. Novel modulator for endothelial adhesion molecules: adipocyte-derived plasma protein adiponectin. Circulation. 1999;100:2473 6. 13. Hotta K, Funahashi T, Arita Y, et al. Plasma concentrations of a novel, adipose-specific protein, adiponectin, in type 2 diabetic patients. Arterioscler Thromb Vasc Biol. 2000;20: 1595 9. 14. Weyer C, Funahashi T, Tanaka S, et al. Hypoadiponectinemia in obesity and type 2 diabetes: close association with insulin resistance and hyperinsulinemia. J Clin Endocrinol Metab. 2001;86:1930 5. 15. Hara K, Boutin P, Mori Y, et al. Genetic variation in the gene encoding adiponectin is associated with an increased risk of type 2 diabetes in the Japanese population. Diabetes. 2002; 51:536 40. 16. Lindsay RS, Funahashi T, Hanson RL, et al. Adiponectin and development of type 2 diabetes in the Pima Indian population. Lancet. 2002;360:57 8. 17. Matsubara M, Maruoka S, Katayose S. Decreased plasma adiponectin concentrations in women with dyslipidemia. J Clin Endocrinol Metab. 2002;87:2764 9. 18. Stefan N, Vozarova B, Funahashi T, et al. Plasma adiponectin concentration is associated with skeletal muscle insulin receptor tyrosine phosphorylation, and low plasma concentration precedes a decrease in whole-body insulin sensitivity in humans. Diabetes. 2002;51:1884 8. 19. Yamamoto Y, Hirose H, Saito I, et al. Correlation of the adipocyte-derived protein adiponectin with insulin resistance index and serum high-density lipoprotein-cholesterol, independent of body mass index, in the Japanese population. Clin Sci. 2002;103:137 42. 20. Zoccali C, Mallamaci F, Tripepi G, et al. Adiponectin, metabolic risk factors, and cardiovascular events among patients with end-stage renal disease. J Am Soc Nephrol. 2002; 13:134 41. 21. Ouchi N, Kihara S, Arita Y, et al. Adiponectin, an adipo- OBESITY RESEARCH Vol. 13 No. 1 January 2005 189

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