The metabolic syndrome is related to β3-adrenoceptor sensitivity in visceral adipose tissue

Diabetologia, Jul 1996

Summary The metabolic syndrome is a well-known risk for the development of cardiovascular disease. In the present study the possible importance of an altered visceral adipocyte β-adrenoceptor function in this syndrome was investigated. In 65 subjects of both sexes undergoing elective surgery for non-malignant disorders, the metabolic syndrome phenotype and the lipolytic sensitivity for various β-adrenoceptor subtype agonists in omental adipocytes were determined. The study group represented a wide range of abdominal adipose tissue distribution (waist-to-hip ratios 0.76-1.13), but was otherwise apparently healthy. The subjects were divided into three subgroups according to their waist-to-hip (WHR) ratios: 1) WHR < 0.92; 2) WHR 0.92-1.04; 3) WHR > 1.04. The subgroups demonstrated significant differences regarding body mass index, sagittal diameter, systolic and diastolic blood pressures, plasma concentrations of glucose, insulin, triglycerides and HDL-cholesterol (p = 0.005-0.0001). Furthermore, in omental adipocytes β3-adrenoceptor sensitivity, but not β1 and β2-adrenoceptor sensitivities, differed significantly between the WHR subgroups (p = 0.0001). β3-adrenoceptor sensitivity was also related to the other components of the metabolic syndrome, although a strong covariation between WHR and β3-adrenoceptor sensitivity vs blood pressure and the metabolic parameters was found. The present data provide evidence of a relationship between upper-body obesity and its associated metabolic complications and also, an increased visceral fat β3-adrenoceptor sensitivity. We suggest that the latter finding results in an augmented release of non-esterified fatty acids from the visceral fat depot to the portal venous system. This may in turn contribute to the development of the metabolic syndrome.

A PDF file should load here. If you do not see its contents the file may be temporarily unavailable at the journal website or you do not have a PDF plug-in installed and enabled in your browser.

Alternatively, you can download the file locally and open with any standalone PDF reader:

https://link.springer.com/content/pdf/10.1007%2Fs001250050518.pdf

The metabolic syndrome is related to β3-adrenoceptor sensitivity in visceral adipose tissue

Diabetologia The metabolic syndrome is related to fl3-adrenoceptor sensitivity in visceral adipose tissue J. H o f f s t e d t 0 H . W a h r e n b e r g 0 A . T h 0 r n e 0 E Liinnqvist I 0 0 1Department of Medicine, and the Research Center at Huddinge University Hospital, Karolinska Institute, Huddinge, Sweden 2Department of Surgery, Huddinge University Hospital, Karolinska Institute , Huddinge , Sweden Received: 15 September 1995 and in final revised form: 24 January 1996 adipocytes /33-adrenoceptor sensitivity, but not /31and fle-adrenoceptor sensitivities, differed significantly between the W H R subgroups (p = 0.0001). fl3-adrenoceptor sensitivity was also related to the other c o m p o n e n t s of the metabolic syndrome, although a strong covariation between W H R and/33adrenoceptor sensitivity vs blood pressure and the metabolic p a r a m e t e r s was found. The present data provide evidence of a relationship between upperbody obesity and its associated metabolic complications and also, an increased visceral fat fl3-adrenoceptor sensitivity. We suggest that the latter finding results in an a u g m e n t e d release of non-esterified fatty acids from the visceral fat depot to the portal venous system. This m a y in t u r n contribute to the dev e l o p m e n t of the m e t a b o l i c syndrome. [Diabetologia (1996) 39: 838-844] Adipocytes; fl3-adrenoceptors; insulin resistance; metabolic s y n d r o m e; visceral fat - 9 Springer-Verlag 1996 Summary The m e t a b o l i c s y n d r o m e is a well-known risk for the d e v e l o p m e n t of cardiovascular disease. In the present study the possible i m p o r t a n c e of an altered visceral adipocyte f l - a d r e n o c e p t o r function in this s y n d r o m e was investigated. In 65 subjects of both sexes u n d e r g o i n g elective surgery for non-malignant disorders, the metabolic s y n d r o m e p h e n o t y p e and the lipolytic sensitivity for various fl-adrenoceptor subtype agonists in o m e n t a l adipocytes w e r e determined. The study group r e p r e s e n t e d a wide range of abdominal adipose tissue distribution (waist-tohip ratios 0.76-1.13), but was otherwise apparently healthy. The subjects were divided into t h r e e subgroups according to their waist-to-hip ( W H R ) ratios: 1) W H R < 0.92; 2) W H R 0.92-1.04; 3) W H R > 1.04. The subgroups d e m o n s t r a t e d significant differences regarding b o d y mass index, sagittal diameter, systolic and diastolic blood pressures, plasma concentrations of glucose, insulin, triglycerides and HDL-cholesterol (p = 0.005-0.0001). F u r t h e r m o r e , in o m e n t a l The metabolic s y n d r o m e has aroused an increasing interest in recent years as an i m p o r t a n t risk factor for cardiovascular disease, which is one of the major causes of p r e m a t u r e d e a t h in western countries. The metabolic or insulin resistance syndrome consists of several pathological conditions [ 1-4 ], such as abdominal obesity, glucose intolerance, hypertension, hypertriglyceridaemia and low serum high density lipoprotein ( H D L ) - c h o l e s t e r o l levels. In this respect, the a c c u m u l a t i o n of visceral fat m a y be of particular importance. It has been postulated that an increased release of non-esterified fatty acids (NEFA) delivered to the portal venous system from the lipolytically-active visceral adipocytes m a y have adverse m e t a b o l i c effects on the liver resulting in elevated levels of plasma insulin and triglycerides, accompanied by glucose intolerance [ 5-7 ]. However, the underlying m e c h a n i s m s have n o t b e e n clarified. J. Hoffstedt et al.: The metabolic syndrome and fl 3-adrenoceptors In man, it has long b e e n k n o w n that the release of N E F A and glycerol via lipolysis is mainly influenced by catecholamines and is m o d u l a t e d either through s t i m u l a t o r y / 3 1 % and/32-adrenoceptors or inhibitory c~2-adrenoceptors [ 8 ]. R e c e n t studies have related the existence of a gene coding for a third stimulatory f i - a d r e n o c e p t o r in man [ 9 ], which is functionally active principally in omental adipocytes [ 10, 11 ] but also present in m a m m a r y fat [12] and in subcutaneous fat in vivo [13]. Previous studies from our laboratory have shown that both the abdominal subcutaneous/32-adrenoceptor [14] and the visceral fl3-adrenoceptor functions [15] may play a pathogenic role in upper-body obesity. The aim of the present study was therefore to examine further the possible importance of the three visceral adipose tissue/3-adrenoceptor subtypes (131, /32 and/33) for the metabolic syndrome. The phenotype of the metabolic s y n d r o m e and the lipolytic sensitivity to various/3-adrenoceptor agonists in visceral adipocytes w e r e d e t e r m i n e d in subjects of both sexes representing a wide range of abdominal fat distribution. Subjects, materials and methods Subjects. The study group consisted of 65 r a n d o m l y selected Caucasian subjects undergoing weight reduction surgery, elective cholecystectomy or o p e r a t i o n s for hiatus hernia through open or laparoscopic p r o c e d u r e s at H u d d i n g e University Hospital (Table 1). Before surgery, a thorough history was taken and, except for the surgery indication, all subjects were apparently healthy and not on any medication. N o n e had a history of alcohol overconsumption. The subjects were between 19 and 60 years of age. The body mass index (BMI) ranged from 20 to 53 kg/m 2 with a span of values including non-obese, moderately obese and massively obese subjects. The waist-to-hip ratio ( W H R ) , the sagittal d i a m e t e r and the systolic and diastolic b l o o d pressures were m e a s u r e d in the supine position on the d a y before surgery. The sagittal d i a m e t e r was obtained by measuring the distance from the examination table to a horizontal cross-bar placed over the a b d o m e n of the recumbent subject at the level of the crista. The systolic and diastolic blood pressures were d e t e r m i n e d with a mercury sphygmoman o m e t e r using phases I and V of the K o r o t k o f f sounds. Each value is the mean of three consecutive m e a s u r e m e n t s after a 10-min rest. Following an overnight fast, the study subjects rested in bed for 15 min, whereafter venous blood samples were obtained, which were analysed by the hospital's routine chemistry laboratory, except for insulin which was m e a s u r e d with a radioimmunoassay kit (Pharmacia, Uppsala, Sweden). G e n e r a l anaesthesia was induced at 08.00 hours by a shortacting b a r b i t u r a t e and m a i n t a i n e d by fentanyl and a mixture of oxygen and nitrous oxide. Intravenous saline was administ e r e d prior to the fat biopsies, which were t a k e n from the major o m e n t u m , at the beginning of the operation. The study was a p p r o v e d by the ethics committee of K a r o l i n s k a Institute, Stockholm, and all the patients gave i n f o r m e d consent to participate. Lipolysis. The omental adipose tissue biopsies were immediately t r a n s p o r t e d to the l a b o r a t o r y in saline at 37 ~ and isolated fat cells from fat specimens weighing 0.3-1.0 g were prep a r e d by collagenase treatment, as previously described [16]. The cells were kept in an albumin solution, as described below, and the cell density of the fat-cell suspension was kept constant by slow stirring with the aid of a magnet. Direct microscopic d e t e r m i n a t i o n of the fat-cell diameter, p e r f o r m e d according to the m e t h o d of Di G i r o l a m o and co-workers [17], was calculated by using 200 cells from each subject. The m e a n fat cell volume and weight were determined, taking into account the skewness in the distribution of the cell d i a m e t e r and using the m e t h o d described by Hirsch and G a l l i a n [18]. The total lipid content in each incubation was d e t e r m i n e d gravimetrically after organic extraction. Assuming that lipids constitute more than 95 % of the fat-cell weight, the n u m b e r of fat cells can be calculated by dividing the total lipid weight by the m e a n cell weight. A detailed description of the lipolysis assay has been rep o r t e d elsewhere [19]. In brief, 0.2 ml of diluted suspensions of isolated fat cells (5000-10000 cells/ml) were incubated in duplicate for 2 h with or without the selective fl l - a d r e n o c e p t o r agonist d o b u t a m i n e , the selective fi2-adrenoceptor agonist terbutaline or the selective partial fl3-adrenoceptor agonist C G P 12177 [20]. All incubations were p e r f o r m e d at 37 ~ in K r e b s - H e n s e l e i t p h o s p h a t e buffer ( p H 7.4), s u p p l e m e n t e d with glucose (5.6 mmol/1), bovine serum albumin (0.3 mmol/1) and ascorbic acid (0.6 mmol/1), with air as the gas phase. The agents were a d d e d simultaneously at the start of the incubation. The concentration range for each agent ranged overall f r o m 10-12 to 10-3 mol/1. A f t e r 2 h of incubation, a cell-free aliquot was r e m o v e d for d e t e r m i n a t i o n of the glycerol concentration, using a bioluminescence m e t h o d [21]. A l l agonists caused a c o n c e n t r a t i o n - d e p e n d e n t stimulation of glycerol release that reached a p l a t e a u at the highest agonist concentrations. Consequently, it was always possible to determine the concentration of agonist that p r o d u c e d a half-maxim u m effect on glycerol release. These ECs0 values (expressed as log mol/1) were d e t e r m i n e d by linear regression analysis of log-logit transformation of the ascending part of the individual concentration-response curves [22]. Lipolysis rates in the presence of m a x i m u m effective agonist concentrations were related to fat-cell number. Drugs and chemicals. Bovine serum albumin (fraction V, lot 63F-0748), Clostridium histolyticum collagenase type I and glycerol kinase from Escherichia coli (G4509) were obtained J. Hoffstedt et al.: The metabolic syndrome and/33-adrenoceptors from Sigma (St. Louis, Mo., USA). Terbutaline sulphate came from Draco (Lund, Sweden), d o b u t a m i n e hydrochloride from Lilly (Indianapolis, Ind., U S A ) and C G P (_+) 12177 ((-)-4-(3t-butylamino-2-hydroxy-propoxy)benzimidazole-2-one) from Ciba Geigy (Basel, Switzerland). A T P monitoring reagent containing firefly luciferase came from LKB Wallac (Turku, Finland). All other chemicals were of the highest grade of purity commercially available. The same batches of collagenase and a l b u m i n and the same stock solutions of adrenoceptor agonists were employed throughout the study. Statistical analysis Values are given as the m e a n + SEM or if indicated m e a n + SD. ECs0 values were logarithmically transformed to normalize the data and were defined as fl-adrenoceptor subtype sensitivity. Analysis of variance (ANOVA), analysis of covariance, the B o n f e r r o n i / D u n n post-hoc test, single and multiple regression analyses and the Kolmogorov-Smirnoff one-sample test were used for statistical comparisons. All statistical calculations were performed with a software package for statistics (Abacus Concepts Inc., Berkeley, Calif., USA). R e s u l t s The W H R in the study group showed a clear interindividual variation ranging from 0.76 to 1.13. These values were normally and symmetrically distributed a r o u n d the population m e a n of 0.98 + 0.08 (mean + SD) as evaluated with the K o l m o g o r o v - S m i r n o f f one-sample test. The W H R values w e r e f u r t h e r subdivided into three groups with r e f e r e n c e to the frequency distribution. The first group in the lower 25th percentile included subjects with a W H R less than O ~ . , , , ~ O O O 0 O0 ~'~'CL^ 0 o o-~,.q cb o o 0.92 (n = 16). The second or intermediate group, between the 25th and 75th percentiles, consisted of subjects with W H R values ranging from 0.92-1.04 ( n - - 3 3 ) . The third group of subjects with a W H R above 1.04 (n = 16) were in the higher 75th percentile. To find out w h e t h e r the t h r e e subgroups differed in their metabolic phenotype, a n u m b e r of clinical and metabolic parameters were assessed by A N O V A with the B o n f e r r o n i / D u n n test as the post-hoc test. As shown in Table 2, the t h r e e percentile groups were considerably different f r o m each o t h e r regarding most of the parameters, including the anthropometric data, systolic and diastolic blood pressures and metabolic variables. In general, the only parameter m e a s u r e d which was not influenced by the W H R was the plasma cholesterol level. J. Hoffstedt et al.: The metabolic syndrome and fl3-adrenoceptors O O 10 2.4oE 2.2E 2.0a>21 1.6O O The contribution of the W H R values to the variations in ill-, /32- a n d /33-adrenoceptor sensitivities was investigated using the same m e t h o d as above. As d e m o n s t r a t e d in Table 2, only the ECs0 for C G P 12177 (fl3-adrenoceptor sensitivity), but not the ECs0 for d o b u t a m i n e ( f l l - a d r e n o c e p t o r sensitivity) or terbutaline (fl2-adrenoceptor sensitivity), showed a significant d i f f e r e n c e b e t w e e n the subjects with high, i n t e r m e d i a t e or low W H R values. The relative i m p o r t a n c e of the ill-, f12-, and fl3a d r e n o c e p t o r subtypes for the various obesity measures and the clinical p a r a m e t e r s involved in the metabolic s y n d r o m e w e r e f u r t h e r investigated. In linear regression analysis, t h e ECs0 for C G P 12177 was correlated to W H R , r = - 0 . 6 2 , p - - 0 . 0 0 0 1 (Fig. 1), systolic, r = - 0 . 5 1 , p = 0.0001, and diastolic, r = - 0 . 4 7 , p --0.0001, blood pressures (Fig.2) and plasma concentrations of insulin, r = -0.41, p < 0.001, and H D L cholesterol, r = 0.48, p = 0.0001 (Fig. 3). However, the ECs0 for C G P 12177 showed no correlation to the plasma c o n c e n t r a t i o n s of glucose, triglycerides and cholesterol. Since ECs0 for C G P 12177 was also correlated to B M I (r = -0.57, p = 0.0001) and the sagittal d i a m e t e r (r = -0.60, p = 0.0001), the relative importance of W H R for /33-adrenoceptor sensitivity was investigated in a multiple regression analysis. Using ECs0 for C G P 12177 as the d e p e n d e n t variable and W H R , B M ! a n d sagittal d i a m e t e r as i n d e p e n d e n t variables r e v e a l e d that W H R was i n d e p e n d e n t l y corr e l a t e d to /33-adrenoceptor sensitivity (p = 0.0001). As previously s h o w n [15],/33-adrenoceptor sensitivity was also c o r r e l a t e d to fat-cell v o l u m e ( r = 0.70, p = 0.0001). Finally, no correlations were seen bet w e e n ECs0 for d o b u t a m i n e or terbutaline and any of the p a r a m e t e r s m e n t i o n e d . In o r d e r to calculate the relative i m p o r t a n c e of W H R a n d / 3 3 - a d r e n o c e p t o r sensitivity for m e t a b o l i c complications in u p p e r - b o d y obesity, these p a r a m e ters w e r e e n t e r e d as i n d e p e n d e n t variables in a multiple regression analysis as o p p o s e d to the previously m e n t i o n e d m e t a b o l i c p a r a m e t e r s and blood pressures as the d e p e n d e n t variables. In this analysis, only W H R was c o r r e l a t e d to the various clinical parameters, indicating a strong covariation b e t w e e n W H R a n d / 3 3 - a d r e n o c e p t o r sensitivity. Since the rate of lipolysis is also an i m p o r t a n t m e a sure of a d r e n o c e p t o r function, we have also m e a sured lipolysis i n d u c e d by n o r a d r e n a l i n e , which is the most i m p o r t a n t e n d o g e n o u s stimulator of lipolysis. B a s e d on the W H R subgroups, the m a x i m u m nor a d r e n a l i n e - i n d u c e d a m o u n t s of glycerol r e l e a s e d (~mol - 10 7 cells 1 . 2 h -1) w e r e 21.6 + 2.9, 15.8 + 1.8 and 8.8 + 2.1 in the high, i n t e r m e d i a t e and low W H R subgroups, respectively ( m e a n + SEM). As calculated by A N O V A , these values were statistically different (p < 0.01). F u r t h e r m o r e , ECs0 for C G P 12177, in contrast to d o b u t a m i n e - and terbutaline-sensitivity, correlated to the m a x i m u m n o r a d r e n a l i n e glycerol release (r = 0.47, p = 0.0005), which indicates that fi3a d r e n o c e p t o r function m a y also be i m p o r t a n t for lipolysis rate, in analogy with our previous findings [151. The i m p o r t a n c e of s m o k i n g habits and gender were also e x a m i n e d using A N O V A . No significant differences b e t w e e n s m o k e r s and n o n - s m o k e r s were observed. However, the male subjects had significantly higher W H R values (1.03 _+0.01 vs 0.94 _+0.01, p =0.0001) and sagittal d i a m e t e r (27.8+1.22 vs 24.6 + 0.99, p < 0.05) t h a n the female subjects. In order to evaluate the influence of gender on fi 3-adrenoceptor sensitivity, the W H R values were again subdivided, as previously described, for the males and females separately, resulting in high (males > 1.07, n = 7, f e m a l e s > 0.99, n = 10), i n t e r m e d i a t e (males 0.99-1.07, n = 14, females 0.90-0.99, n = 18) and low (males < 0.99, n = 6, females < 0.90, n = 10) W H R subgroups. R e g a r d i n g the males, ECs0 for C G P 12177 were 9.38 + 0.27, 8.88 _+0.27 and 7.93 _+0.47 in the high, i n t e r m e d i a t e and low W H R subgroups, respectively ( m e a n + SEM, p < 0.05, A N O V A ) . The female values were 8.64 _+0.33 (high), 7.69 _+0.30 (intermediate) and 6.91 _+0.36 (low), respectively (p < 0.01, A N O V A ) . The influence of g e n d e r on the clinical variables was further analysed c o m p a r i n g the male and female subgroups with analysis of covariance using gender as a factor, W H R and fi3-adrenoceptor sensitivity as covariates and insulin, HDL-cholesterol, triglycerides, systolic and diastolic blood pressures as depend e n t variables. However, n o significant interaction of sex on the relationships b e t w e e n the covariates and the clinical p a r a m e t e r s was d e m o n s t r a t e d in this patient material. Discussion In the present study, we provide evidence of an association b e t w e e n the m e t a b o l i c complications accompanied by a b d o m i n a l obesity (the metabolic synd r o m e ) and the fi3-adrenoceptor sensitivity in visceral fat. Subjects with high W H R , as an index for abdominal obesity, were f o u n d to have higher blood pressure levels and higher plasma concentrations of insulin, glucose and triglycerides, but lower plasma concentrations of H D L - c h o l e s t e r o l t h a n non-obese subjects. F u r t h e r m o r e , the visceral fat cells of the upper-body obese subjects were m o r e sensitive to lipolytic stimulation by the fl3-adrenoceptor agonist C G P 12177. H o w e v e r , lipolysis i n d u c e d by J.Hoffstedt et al.: The metabolic syndrome and fl3-adrenoceptors d o b u t a m i n e (a f i l - a d r e n o c e p t o r agonist) or terbutaline (a fl2-adrenoceptor agonist) did not differ between upper-body obese a n d non-obese subjects. A l t h o u g h obesity is associated with various metabolic aberrations such as glucose intolerance and dyslipoproteinaemia, n o t all obese subjects present these findings. As reviewed [ 3-6 ], an increase in the intraabdominal or visceral fat mass is strongly related to a cluster of m e t a b o l i c disturbances (hyperinsulinaemia, glucose intolerance, hypertriglyceridaemia, low-plasma H D L - c h o l e s t e r o l ) and hypertension, which together constitute the insulin resistance or metabolic syndrome. Moreover, visceral obesity is a key risk factor for cardiovascular disease and non-insulin-dependent diabetes mellitus [ 3-6 ]. The regional distribution of b o d y fat is thus of importance for the d e v e l o p m e n t of the m e t a b o l i c syndrome. In this regard, the role of N E F A derived from intra-abdominal adipose tissue is thought to be particularly important. It is well k n o w n that visceral fat cells have higher lipolytic activity (hydrolysis of triglycerides into N E F A and glycerol) than subcutaneous adipocytes in normal subjects [ 3, 4, 6 ]. In visceral obesity, characterized by an increase in the intra-abdominal fat mass [23], it has b e e n shown, firstly, that the fasting plasma c o n c e n t r a t i o n of N E F A is elevated [24], secondly, that the lipolysis rate in hypertrophic adipocytes is higher than in normal-sized fat cells [25] and, thirdly, that the inhibitory action of insulin on N E F A release from intra-abdominal adipocytes is lower than in subcutaneous fat cells [26]. Altogether, these observations indicate an a u g m e n t e d release of N E F A from the visceral fat depots in upper-body obesity, although the pathophysiological mechanisms have not yet b e e n fully elucidated. The fl3-adrenoceptor function was found to be strongly related to visceral obesity and its various metabolic disturbances. A n increased fl3-adrenoceptor sensitivity to c a t e c h o l a m i n e stimulation might thus be the underlying m e c h a n i s m linking e n h a n c e d lipolytic activity in visceral fat to the aberrations observed in the metabolic syndrome. Catecholamine stimulation m a y lead to an increased delivery of N E F A into the portal v e n o u s system, with several possible effects on liver metabolism. N E F A are known to stimulate hepatic V L D L secretion and gluconeogenesis and to interfere with hepatic clearance of insulin [ 5 ], resulting in dyslipoproteinaemia, glucose intolerance and hyperinsulinaemia. Since hyperinsulinaemia seems to be causally related to an increase in blood pressure [27, 28], hypertension m a y also ensue. As regards the ill- and fl2-adrenoceptor function, their sensitivities were n o t found to be associated with visceral obesity or any of the metabolic parameters. The current findings contrast with previous studies on abdominal s u b c u t a n e o u s adipocytes, in which catecholamine resistance, due to a reduced expression Non-obese subjects Obese subjects Values are m e a n + SD. T h e y w e r e c o m p a r e d using Student's u n p a i r e d t-test, a = p < 0.05 non-obese Trp64 h o m o g y g o u s w o m e n than in Trp64Arg heterozygous n o n - o b e s e women. However, sagittal diameter and b o d y mass index did not differ significantly b e t w e e n these two groups (data not shown). Genotypes of all subjects were d e t e r m i n e d by P C R amplification and cleavage with the BstNI restriction enzyme which is specific for the C to T transition in the first c o d o n of amino acid 64 of the beta3-adrenergic receptor. All Trp64Arg heterozygotes were analysed twice to ensure that full cleavage was attained of the P C R fragments. The lipolytic response of fat cells to stimulation with either noradrenaline or C G P 12177 is shown in Figure 1. Both agents caused a concentration-depend e n t stimulation of glycerol release. They were more effective in fat cells of obese as c o m p a r e d with nonobese subjects confirming earlier results [ 3 ]. In neit h e r group was there a statistically significant difference b e t w e e n Trp64 h o m o z y g o t e s and Trp64Arg heterozygotes. It is also seen in Figure 1 that C G P 12177 was less effective than n o r a d r e n a l i n e in stimulating glycerol release. This is expected, since C G P 12177 is a partial agonist. The m e a n concentration-response curves for C G P 12177 in Figure i do n o t display a classical sigm o i d shape. This is due to considerable interindividual variation in the sensitivity of adipocytes to this agent flattening m e a n curves that are obtained from different individuals [ 3 ]. However, the individual curves were always steep spanning 4-5 log units of molar concentration. Discussion R e c e n t data have suggested that m o l e c u l a r defects in the beta3-adrenergic r e c e p t o r m a y predispose subjects to obesity, complications to obesity and changes in the regulation of the energy expenditure [ 4-7 ]. However, this study suggests that the Trp64Arg mutation in the beta3-adrenergic r e c e p t o r gene is not a maj or d e t e r m i n a n t of beta3-adrenergic receptor function and obesity w h e n present in a heterozygous form. We observed no influence on clinical characteristics in either obese or n o n - o b e s e subjects except for waist-hip ratio in a subgroup, which probably is a chance finding due to the large n u m b e r of statistical comparisons that were made. In m a n , beta3-adrenergic receptors are mainly located in the visceral adipose tissue [ 2 ]. We f o u n d that visceral fat cells with the Trp64Arg m u t a t i o n displayed n o r m a l lipolytic function of the beta3-adrenergic r e c e p t o r (as assessed by C G P 12177) and r e s p o n d e d in a n o r m a l fashion "7t,"7 O % O 8- Obese >, o IE 4 62 T I I I i i i I I 0 O-11-10-9 -8 -7 -6 -5 -4 O-11-10-9 -8 -7 -6 -5 -4 CGP 12177 (log mol/I) Fig.1. Concentration response curves for noradrenaline and CGP 12177 as regards glycerol release from fat cells obtained from subjects with 9 or without [] the Trp64Arg mutation. Curves were compared using analysis of variance (F < 1.0). Values are mean + SEM to n o r a d r e n a l i n e stimulation w h e t h e r t h e y were o b t a i n e d f r o m obese or non-obese subjects. Furthermore, the allelic frequency of the m u t a t i o n was similar in obese a n d non-obese subjects which confirms previous findings [ 4-7 ]. It should be k e p t in mind that the n u m b e r of subjects with the Trp64Arg mutation studied here was rather small. We c a n n o t therefore exclude type II errors in some of the statistical calculations. U n f o r t u n a t e l y we have no data on subjects who are h o m o z y g o u s for the mutation. N o h o m o z y g o u s subjects were f o u n d in previous investigations on 279 F r e n c h and 335 Finnish subjects [ 5, 6 ]. This may suggest t h a t h o m o z y g o t e s for the m u t a t i o n are rare a m o n g Caucasians. However, in a study of 624 Pima Indians 9 % of subjects were f o u n d to be h o m o z y g o u s [4] and 5 % in an investigation of 191 Japanese subjects. Arg64 h o m o z y g o t e subjects h a d an earlier onset of n o n - i n s u l i n - d e p e n d e n t diabetes, higher s e r u m insulin, increased b o d y mass index and a t e n d e n c y to lower resting m e t a b o l i c rate than Trp64Arg heterozygotes who did n o t differ f r o m n o r m a l Trp64 homozygotes. Thus, it is possible that the beta3-adrenergic receptor gene m u t a t i o n is of clinical and functional importance only in its h o m o z y g o u s form. Nevertheless, the beta 3adrenergic m u t a t i o n per se appears to play a relatively m o d e s t role in the pathophysiology of obesity, since the o b s e r v e d clinical differences b e t w e e n n o r m a l and m u t a n t h o m o z y g o t e s were small [ 4, 7 ]. Mutations in the first cytoplasmatic lope of several other G-protein receptors such as the beta3-adrenergic, 0 T i i i i i i O -11 -10 -9 -8 -7 -6 O -11 -10 -9 -8 -7 -6 Noradrenaline (log mol/I) rhodopsin, a n d vasopressin receptors are reported [ 8 ]. These m u t a t i o n s alter the receptor function. O n the other hand, in n o r m a l weight rats the wild type beta3-adrenergic gene carries Arg instead of Trp in codon 64 [ 9, 10 ]. Some caution should be exercised when comparing similar mutations b e t w e e n receptor subclasses or the same gene in different species. However, it is t e m p t i n g to speculate that the first intracellular loop of the beta3-adrenergic receptor is less important for this r e c e p t o r t h a n for other G - p r o t e i n receptor subtypes. H o w e v e r , such a question can only be answered by site directed m u t a t i o n analysis. In summary, the present data suggest that the Trp64Arg m u t a t i o n is n o t a major d e t e r m i n a n t of the beta3-adrenergic r e c e p t o r function or obesity and its complications at least not when present in the heterozygous form. Acknowledgements. This study was supported by grants from Swedish Medical Research Council, Swedish Diabetes Association, Karolinska Institute, Swedish Heart and Lung Foundation, King Gustaf V and Queen Victoria and the foundations of Golje, Osterman, Ake Wiberg, Tore Nilsson and Novo Nordisk. R e f e r e n c e s 1. Bouchard C , P6russe L ( 1993 ) Genetics of obesity Ann Rev Nutr 13 : 337 - 354 2. Giacobino J-P ( 1995 ) Beta3-adrenoceptor: an update . Eur J Endocrinol 132 : 377 - 385 3. L6nnqvist F , Th6rne A , Nilsell K , Hoffstedt J , Arner P ( 1995 ) A pathogenetic role of visceral fat/33-adrenoceptors in obesity . J Clin Invest 95 : 1109 - 1116 4. Walston J , Silver K , Bogardus C , et al. ( 1995 ) Time of onset of non-insulin-dependent diabetes mellitus and genetic variation in the fl3-adrenergic-receptor gene . N Engl J Med 333 : 343 - 347 5. Wid6n E , Lehto M , Kanninen T , Walston J , Shuldiner AR , Groop LC ( 1995 ) Association of a polymorphism in the /33-adrenergic-receptor gene with features of the insulin resistance syndrome in Finns . N Engl J Med 333 : 348 - 351 6. C16ment K , Vaisse C , Manning BSJ , et al. ( 1995 ) Genetic variation in the 133-adrenergicreceptor and an increased capacity to gain weight in patients with morbid obesity . N Engl J Med 333 : 352 - 354 7. Kadowaki H , Yasuda K , Iwamoto K , et al. ( 1995 ) A mutation in the fl3-adrenergic receptor gene is associated with obesity and hyperinsulinemia in Japanese subjects . Biochem Biophys Res Commun 215 : 555 - 560 8. Birnbaumer M ( 1995 ) Mutations and diseases of G protein coupled receptors . J Rec Signal Trans Res 15 : 131 - 160 9. Fujisawa T , Ikegama H , Yamoto E , et al. ( 1996 ) Association of Trp64Arg mutation of the beta3-adrenergic receptor with NIDDM and body-weight gain . Diabetologia 39 : 349 - 352 10. Granneman JG , Lahners KN , Chaudry A ( 1991 ) Molecular cloning and expression of the rat beta3-adrenergic receptor . Mol Pharmacol 40 : 895 - 899


This is a preview of a remote PDF: https://link.springer.com/content/pdf/10.1007%2Fs001250050518.pdf

J. Hoffstedt, H. Wahrenberg, A. Thörne, F. Lönnqvist. The metabolic syndrome is related to β3-adrenoceptor sensitivity in visceral adipose tissue, Diabetologia, 1996, 838-860, DOI: 10.1007/s001250050518