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Serum Retinol-Binding Protein 4 Is Reduced after Weight Loss in Morbidly Obese Subjects

Departments of Clinical Pharmacology (D.G.H., M.W.) and Surgery (G.P., A.B.), and Division of Endocrinology and Metabolism, Department of Internal Medicine (K.S., A.L., M.W., B.L.), Medical University of Vienna, 1090 Vienna, Austria

Address all correspondence and requests for reprints to: Bernhard Ludvik, M.D., Medical University of Vienna, Department of Internal Medicine III, Division of Endocrinology and Metabolism, Allgemeines Krankenhaus Wien, Währinger Gürtel 18-20, 1090 Vienna, Austria. E-mail: bernhard.ludvik{at}meduniwien.ac.at.

	Abstract

Top Abstract Introduction Subjects and Methods Results Discussion References

 
Context: Administration of retinol-binding protein 4 (RBP-4) impairs insulin sensitivity in animals, and elevated serum concentrations have been associated with insulin resistance in humans.

Objective: We have studied whether weight loss influences RBP-4.

Patients and Methods: Fasting serum concentrations of RBP-4 were measured before and 6 months after gastric banding surgery in 33 morbidly obese patients aged 40 ± 11 yr with a body mass index (BMI) of 46 ± 5 kg/m². Fourteen healthy subjects aged 29 ± 5 yr with a BMI less than 25 kg/m² served as controls. To characterize the association of weight loss with central and peripheral appetite regulation, the signaling protein agouti-related protein (AGRP), the orexigenic hormone ghrelin, and its recently identified antagonist obestatin were determined.

Results: At baseline, RBP-4 levels were markedly higher in obese than in lean subjects (2.7 ± 0.5 vs. 0.9 ± 0.5 µg/ml; P < 0.001). In contrast, AGRP and obestatin were lower in obese subjects compared with lean controls (all P < 0.001). Six months after gastric banding, BMI was reduced to 40 ± 5 kg/m², RBP-4 was reduced to 2.0 ± 0.7 µg/ml, AGRP increased from 1.8 ± 1.1 to 3.4 ± 1.1 ng/ml, ghrelin increased from 93 ± 58 to 131 ± 70 pg/ml, and obestatin increased from 131 ± 52 to 173 ± 35 pg/ml (all P < 0.05). Individual changes of RBP-4 were associated with changes of BMI (r = 0.72), the homeostasis model assessment insulin resistance-index (r = 0.53), and total cholesterol (r = 0.42, for all P < 0.05).

Conclusion: Reductions in circulating RBP-4 may contribute to improved insulin resistance in morbidly obese subjects after weight loss. This is accompanied by favorable changes in appetite-regulating hormones, which might support the sustained weight loss after obesity surgery.

	Introduction

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OBESITY IS A MAJOR cause of insulin resistance (1). However, insulin resistance does not develop in all subjects, and genetic background contributes strongly to impaired insulin sensitivity (2). In insulin-resistant states, the expression of the glucose-transporter 4 is down-regulated in adipocytes but not in skeletal muscle (3); this results in impaired insulin-stimulated glucose transport in adipocytes (3). Genetic knockout of glucose-transporter 4 in adipocytes of mice (4) is associated with increased serum levels of retinol-binding protein 4 (RBP-4) (5). Injection of RBP-4 or transgenic overexpression in mice impairs insulin signaling in skeletal muscle and induces the expression of the gluconeogenic enzyme phosphoenolpyruvate carboxykinase in the liver (5). Consistently, elevated serum concentrations of RBP-4 have been reported in subjects with obesity and type 2 diabetes and appear to identify insulin resistance and associated cardiovascular risk factors (6). However, it is unclear whether RBP-4 concentrations may be affected by weight normalization.

Adipocyte mass and associated changes in adipocytokines are regulated and influenced by peptides of the gut-brain axis mediating satiety and food intake. Ghrelin, as one of these peptides, is predominantly produced by the stomach and acts on GH secretagogue receptors to increase GH release from the pituitary gland (7). Ghrelin administration increases appetite via up-regulation of agouti-related protein (AGRP) release (8, 9, 10, 11, 12). Plasma ghrelin concentrations are reduced in obese subjects (13), and weight loss increases circulating ghrelin concentrations (14, 15). The identification of obestatin, a peptide hormone derived from the same gene as ghrelin, has recently added further complexity to ghrelin physiology. Obestatin appears to antagonize the actions of ghrelin on energy homeostasis and gastrointestinal function (16).

We hypothesized that weight loss in morbidly obese subjects may be associated with reduced serum RBP-4, because fat mass has been proposed to regulate concentrations of this adipocytokine. We tested this hypothesis in a prospective, open-labeled, cohort study in subjects undergoing elective gastric banding surgery. An unmatched group of healthy subjects was recruited as controls. To assess the relationship of weight reduction with peptides related to satiety and food intake, circulating AGRP, ghrelin, and obestatin were also determined.

	Subjects and Methods

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Subjects

Thirty-three subjects (29 females and four males) aged 40 ± 11 yr (range, 19–59 yr) with a body mass index (BMI) of more than 40 kg/m² were consecutively recruited among patients scheduled for bariatric surgery. Standardized laparoscopic adjustable gastric banding was performed by the same team of surgeons. Patient characteristics are shown in Table 1. Subjects with the following conditions were excluded from the study: type 2 diabetes mellitus, myocardial infarction during the last 6 months, malignancy, chronic kidney or liver disease, history of seizures, obesity caused by an endocrine disorder, psychiatric diseases, current pregnancy, or breastfeeding. Seven male and seven female (29 ± 5 yr; range, 24–42 yr) healthy subjects who were not matched with the intervention group for age or sex and who had a BMI of less than 25 kg/m² served as controls. This study was approved by the Ethics Committee of the Medical University of Vienna. All subjects gave written informed consent before entry into the study.

Measurement of RBP-4, AGRP, ghrelin, and obestatin

Serum samples were stored at –70 C until batch analysis. RBP-4 and AGRP were analyzed using commercially available ELISA kits (Phoenix Peptides, Karlsruhe, Germany), and total ghrelin and obestatin were determined by RIAs (human ghrelin RIA kit, Linco Research, St. Charles, MO; human obestatin, Phoenix Peptides) with detection limits between 50–6400 pg/ml. Intraassay and interassay coefficients of variation were less than 5% and less than 6% for all analytes, respectively. Detection of plasma RBP-4 concentrations is linear between 0.5 and 100 ng/ml, which covers the physiological range. For values below the limit of quantification (LOQ), LOQ/2 was used for statistics. Due to limited serum available, ghrelin concentrations could not be determined in lean subjects.

Statistical analyses

Nonparametric tests were performed for comparisons between time points using the Statistica software package (release 6.1; Statsoft Inc., Tulsa, OK). Spearman’s correlation was used for calculation of associations between variables and multiple linear regression analysis with RBP-4 as dependent variable and changes in BMI, HOMA, cholesterol, and high-density lipoprotein cholesterol as independent variables for determination of independent predictors for changes in RBP-4. The Wilcoxon matched-pairs test and the Mann-Whitney U test were used for comparisons within and between groups, respectively. P < 0.05 was considered significant. Values are expressed as means ± SD unless indicated otherwise.

	Results

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RBP-4 concentrations were markedly higher in morbidly obese patients than in lean controls (P < 0.001, Table 2). In contrast, AGRP and obestatin were lower in obese subjects compared with lean controls (Table 2, all P < 0.001). Obestatin was below the LOQ in one patient.

Weight loss decreased RBP-4 concentrations by –0.7 ± 0.6 µg/ml (P < 0.01 vs. baseline; Table 2 and Fig. 1), increased AGRP by 1.6 ± 1.2 ng/ml, increased ghrelin by 32 ± 62 pg/ml, and increased obestatin by 42 ± 44 pg/ml (all P < 0.01; Fig. 2). After 6 months, triglycerides and C-reactive protein concentrations were lowered, but no change was detectable for fasting glucose, insulin (Table 1), or the HOMA-insulin resistance index.

View larger version (35K): [in this window] [in a new window]   FIG. 1. Individual serum RBP-4 concentrations before and 6 months after weight loss surgery in morbidly obese patients.

View larger version (33K): [in this window] [in a new window]   FIG. 2. Serum concentrations of AGRP (A) and obestatin (B) before and 6 months after weight loss surgery.

	Discussion

Top Abstract Introduction Subjects and Methods Results Discussion References

 
This study demonstrates that RBP-4 concentrations are markedly increased in morbidly obese patients. Weight loss after gastric banding is associated with reduced serum RBP-4 levels and was accompanied by increased circulating concentrations of AGRP, ghrelin, and obestatin in these patients.

There is preliminary evidence that elevated RBP-4, which is predominantly secreted by adipocytes and the liver, may identify patients at risk to develop type 2 diabetes (6). A previous study has shown that RBP-4 levels correlate with BMI, which is compatible with our findings. In particular, a lack of RBP-4 reduction in this study was only noted in a patient who did not reduce body weight after 6 months, consistent with the correlation between changes in BMI and RBP-4. It has been reported that diet-induced changes in body weight by approximately 5% are associated with decreased adipose tissue RBP-4 expression in the absence of serum RBP-4 changes (17). The comparatively greater weight loss in our cohort of approximately 13% was associated with decreased serum RBP-4. Nevertheless, these effects are small compared with healthy controls, and a more pronounced reduction in weight might be required to normalize RBP-4 serum concentrations. No time control experiments were conducted and the nearly 2-fold increase in RBP-4 in one patient may also indicate substantial scatter among data, despite the homogeneity of serum RBP-4 concentrations in the other subjects under study. Although blood sampling was standardized regarding fasting status and daytime, fluctuations due to diurnal pattern or feeding status may affect hormone concentrations.

In a heterogenous cohort of subjects with obesity and impaired glucose tolerance, RBP-4 correlated with the magnitude of insulin resistance (6). This result was not reproducible in the morbidly obese subjects under study, which might be due to the small sample size, a selection bias, but also to the HOMA method employed, which is known to be subject to variation and limited sensitivity. However, changes in HOMA over time were again related with changes in RBP-4, confirming these previous reports. However, it is unclear whether RBP-4 exerts a direct effect on improved insulin resistance and may, therefore, be used as a therapeutic target rather than an early marker of impaired glucose homeostasis. Animal experiments at least suggest a pharmacological activity and consequently target function of RBP-4 regulation.

As an additional finding, this trial demonstrates that appetite-regulating hormones of the brain-gut axis are influenced by weight loss, which indicates an important role of these endocrine mechanisms for long-term weight loss. AGRP, ghrelin, and obestatin concentrations were suppressed in morbidly obese subjects. In this study, monoclonal antibodies were used, which explains different hormone concentrations compared with other studies that used polyclonal antibody assays. However, AGRP concentrations are at variance with other reports (18, 19) where increased AGRP levels were detectable in obese patients. Method differences may be attributable for this discrepancy, but the reason for this difference cannot be derived from the present study. In contrast, plasma AGRP was reduced in healthy subjects after a breakfast meal (20) and increased by prolonged fasting (18, 20). These short-term observation could be compatible with results seen in morbidly obese patients, but remain to be confirmed in other studies.

As expected, ghrelin increased during weight reduction in obese patients. Interestingly, this was paralleled by increases in circulating obestatin concentrations. A possible explanation for these findings could be that normalization of central and peripheral satiety regulation by weight loss affects both opposing hormones, although no relationship could be detected between both parameters. In contrast, obestatin is rapidly degraded and might reflect more rapid changes in food intake, glucose levels, or insulin action than ghrelin. Alternatively, the simultaneous obestatin increase over time might be required to counterbalance the endocrine action of ghrelin and weight regain. The interaction between these peptides needs to be characterized in future experiments.

In summary, RBP-4 is elevated in morbidly obese patients compared with lean controls, reduced after weight loss, and is associated with BMI and HOMA. Thus, a decrease in RBP-4 might contribute to improvement in insulin resistance. The paralleled normalization of the appetite-regulating hormones AGRP, ghrelin, and obestatin might support the continued weight loss after gastric banding surgery and favorably affect satiety and food intake.

	Footnotes

 
The authors state that they have no conflict of interest.

First Published Online December 12, 2006

Abbreviations: AGRP, Agouti-related protein; BMI, body mass index; HOMA, homeostasis model assessment; LOQ, limit of quantification; RBP-4, retinol-binding protein 4.

Received August 22, 2006.

Accepted December 6, 2006.

	References

Top Abstract Introduction Subjects and Methods Results Discussion References

 

1. Kahn BB, Flier JS 2000 Obesity and insulin resistance. J Clin Invest 106:473–481[Medline]
2. Perseghin G, Ghosh S, Gerow K, Shulman GI 1997 Metabolic defects in lean nondiabetic offspring of NIDDM parents: a cross-sectional study. Diabetes 46:1001–1009[Abstract]
3. Shepherd PR, Kahn BB 1999 Glucose transporters and insulin action—implications for insulin resistance and diabetes mellitus. N Engl J Med 341:248–257[Free Full Text]
4. Abel ED, Peroni O, Kim JK, Kim YB, Boss O, Hadro E, Minnemann T, Shulman GI, Kahn BB 2001 Adipose-selective targeting of the GLUT4 gene impairs insulin action in muscle and liver. Nature 409:729–733[CrossRef][Medline]
5. Yang Q, Graham TE, Mody N, Preitner F, Peroni OD, Zabolotny JM, Kotani K, Quadro L, Kahn BB 2005 Serum retinol binding protein contributes to insulin resistance in obesity and type 2 diabetes. Nature 436:356–362[CrossRef][Medline]
6. Graham TE, Yang Q, Bluher M, Hammarstedt A, Ciaraldi TP, Henry RR, Wason CJ, Oberbach A, Jansson PA, Smith U, Kahn BB 2006 Retinol-binding protein 4 and insulin resistance in lean, obese, and diabetic subjects. N Engl J Med 354:2552–2563[Abstract/Free Full Text]
7. Kojima M, Hosoda H, Date Y, Nakanzato M, Matsuo H, Kangawa K 1999 Ghrelin is a growth-hormone-releasing acylated peptide from stomach. Nature 402:656–660[CrossRef][Medline]
8. Kamegai J, Tamura H, Shimizu T, Ishii S, Sugihara H, Wakabayashi I 2001 Chronic central infusion of ghrelin increases hypothalamic neuropeptide Y and agouti-related protein mRNA levels and body weight in rats. Diabetes 50:2438–2443[Abstract/Free Full Text]
9. Nakazato M, Murakami N, Date Y, Kojima M, Matsuo H, Kangawa K, Matsukura S 2001 A role for ghrelin in the central regulation of feeding. Nature 409:194–198[CrossRef][Medline]
10. Tschop M, Smiley DL, Heiman ML 2000 Ghrelin induces adiposity in rodents. Nature 407:908–913[CrossRef][Medline]
11. Egecioglu E, Bjursell M, Ljungberg A, Dickson SL, Kopchick JJ, Bergstrom G, Svensson L, Oscarsson J, Tornell J, Bohlooly Y-M 2006 Growth hormone receptor deficiency results in blunted ghrelin feeding response, obesity, and hypolipidemia in mice. Am J Physiol Endocrinol Metab 290:E317–E325
12. Wren AM, Seal LJ, Cohen MA, Brynes AE, Frost GS, Murphy KG, Dhillo WS, Ghatei MA, Bloom SR 2001 Ghrelin enhances appetite and increases food intake in humans. J Clin Endocrinol Metab 86:5992
13. Shiiya T, Nakazato M, Mizuta M, Date Y, Mondal MS, Tanaka M, Nozoe S, Hosoda H, Kangawa K, Matsukura S 2002 Plasma ghrelin levels in lean and obese humans and the effect of glucose on ghrelin secretion. J Clin Endocrinol Metab 87:240–244[Abstract/Free Full Text]
14. Hansen TK, Dall R, Hosoda H, Kojima M, Kangawa K, Christiansen JS, Jorgensen JO 2002 Weight loss increases circulating levels of ghrelin in human obesity. Clin Endocrinol (Oxf) 56:203–206[CrossRef][Medline]
15. Cummings DE, Weigle DS, Frayo RS, Breen PA, Ma MK, Dellinger EP, Purnell JQ 2002 Plasma ghrelin levels after diet-induced weight loss or gastric bypass surgery. N Engl J Med 346:1623–1630[Abstract/Free Full Text]
16. Zhang JV, Ren PG, Avsian-Kretchmer O, Luo CW, Rauch R, Klein C, Hsueh AJ 2005 Obestatin, a peptide encoded by the ghrelin gene, opposes ghrelin’s effects on food intake. Science 310:996–999[Abstract/Free Full Text]
17. Janke J, Engeli S, Boschmann M, Adams F, Bohnke J, Luft FC, Sharma AM, Jordan J 2006 Retinol-binding protein 4 in human obesity. Diabetes 55:2805–2810[Abstract/Free Full Text]
18. Hoggard N, Johnstone AM, Faber P, Gibney ER, Elia M, Lobley G, Rayner V, Horgan G, Hunter L, Bashir S, Stubbs RJ 2004 Plasma concentrations of -MSH, AgRP and leptin in lean and obese men and their relationship to differing states of energy balance perturbation. Clin Endocrinol (Oxf) 61:31–39[CrossRef][Medline]
19. Katsuki A, Sumida Y, Gabazza EC, Murashima S, Tanaka T, Furuta M, Araki-Sasaki R, Hori Y, Nakatani K, Yano Y, Adachi Y 2001 Plasma levels of agouti-related protein are increased in obese men. J Clin Endocrinol Metab 86:1921–1924[Abstract/Free Full Text]
20. Shen CP, Wu KK, Shearman LP, Camacho R, Tota MR, Fong TM, Van der Ploeg LH 2002 Plasma agouti-related protein level: a possible correlation with fasted and fed states in humans and rats. J Neuroendocrinol 14:607–610[CrossRef][Medline]

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This Article Abstract Full Text (PDF) All Versions of this Article: 92/3/1168    most recent Author Manuscript (PDF) Submit a related Letter to the Editor Purchase Article View Shopping Cart Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Copyright Permission Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Haider, D. G. Articles by Ludvik, B. Search for Related Content PubMed PubMed Citation Articles by Haider, D. G. Articles by Ludvik, B. Related Collections Diabetes and Insulin Obesity