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Leptin Secretion Is Related to Chronicity and Severity of the Illness in Bulimia Nervosa

From the Department of Psychiatry, University of Naples SUN, Naples, Italy.

Address reprint requests to: Dr. Palmiero Monteleone, Department of Psychiatry, University of Naples SUN, Largo Madonna delle Grazie, 80138 Naples, Italy. Email: monteri{at}tin.it

	ABSTRACT

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OBJECTIVE: A reduction in circulating leptin has been reported in some patients with BN, but not in others. The reasons for such diversity have not been explained. Therefore, we measured plasma levels of leptin and other hormones in a large sample of patients with eating disorders and in healthy controls and explored differences between bulimics with low plasma leptin levels and those with normal concentrations of the hormone.

METHOD: One hundred and twenty-seven women participated in the study. They were 33 patients with AN, 56 women with BN, and 38 healthy volunteers. All were drug free for at least 6 weeks. Blood samples were collected in the morning for determination of hormone levels. Eating-related psychopathology and depressive symptoms were rated by using specific psychopathological scales.

RESULTS: As compared with healthy women, both underweight AN and normal-weight BN patients exhibited decreased plasma levels of leptin, prolactin, and 17ß-estradiol. Plasma levels of cortisol were increased in AN but not in BN women. In bulimics, circulating leptin was inversely correlated with the duration of the illness and the frequency of bingeing/vomiting. Moreover, 29 bulimics had anorexic-like plasma leptin concentrations; the remaining 27 had circulating leptin similar to normal controls. The former had a significantly longer duration of the illness and a higher frequency of bingeing/vomiting compared with the latter. Moreover, a higher number of patients with concomitant borderline personality disorder were found in bulimics with low leptin concentrations. No significant differences between the two groups emerged in BW, BMI, mean plasma levels of cortisol, PRL, 17ß-estradiol and other clinical parameters.

CONCLUSIONS: Present data show that leptin production is decreased in the subgroup of bulimic patients with a more chronic disease and with a greater severity of the bingeing/vomiting behavior. These findings support the idea that factors other than body weight changes (especially chronic malnutrition and bingeing behavior) may impair profoundly and persistently leptin secretion in patients with BN.

Key Words: anorexia nervosa, • bulimia nervosa, • cortisol, • leptin.

Abbreviations: AN = anorexia nervosa;; BN = bulimia nervosa;; BMI = body mass index;; BW = body weight;; EDI = Eating Disorder Inventory;; BITE = Bulimic Investigation Test Edinburgh;; HDRS = Hamilton Depression Rating Scale;; PRL = prolactin;; ANOVA = analysis of variance;; 5-HT = serotonin.

	INTRODUCTION

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Anorexia nervosa and bulimia nervosa are psychiatric disorders characterized by abnormal eating behaviors that generally result in severe food restriction with a dramatic loss of BW in AN and in episodes of binge eating and vomiting without significant changes of BW in BN. Because of these altered eating patterns and purging behaviors, malnutrition and/or denutrition occur in both syndromes, with consequent metabolic changes that, in turn, are responsible for several organic alterations, including abnormalities in the reproductive activity and neuroendocrine functions (1).

The discovery of leptin, the adipocyte-derived peptidergic hormone, has opened new opportunities in the investigation of the pathophysiology of AN and BN. Leptin is thought to be a satiety factor that regulates BW through the modulation of feeding behavior and energy expenditure (2). Despite the strong correlation between circulating leptin and adipose stores in both animals and humans (2), factors other than adipocyte size and fat content influence leptin production. In this regard, energy availability and the macro/micronutrient composition of the diet are considered important determinants of leptin production (3).

Given this background, it is not surprising that, in underweight people with AN, plasma leptin levels have been found consistently reduced, with a progressive re-increase during the recovery of BW (4–8). In normal-weight subjects with BN, circulating leptin has been reported to be either decreased, normal, or increased (7–12).

From scatterplots of leptin concentrations included in two previous reports, one from Brewerton et al. (11) and the other from our group (8), it is evident that, while anorexic patients had almost uniformly low plasma leptin concentrations, bulimics exhibited levels of plasma leptin ranging from anorexic-like values to normal controls’ concentrations. The issue why some bulimics have normal values of plasma leptin and others have decreased levels of this hormone has not been addressed, although this phenomenon could be a source of discrepancy among the study results. We hypothesized that bulimics with low leptin secretion differ from those with normal leptin levels in clinical and/or hormonal characteristics that could be related to leptin production. In the attempt to verify this hypothesis, we measured plasma levels of leptin and other hormones in a large sample of patients with eating disorders and in healthy controls; then, a posteriori, we divided bulimic patients into those with normal or anorexic-like leptin levels and explored differences between the two groups.

	METHODS

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Subjects
A total of 127 women were recruited for the study. They were 89 outpatients attending the Eating Disorder Center of our Institute and 38 healthy controls. According to DSM-IV criteria, 33 patients fulfilled the diagnosis of AN and 56 the diagnosis of BN. Diagnostic assessment was made by a trained interviewer (B.C.) using the Structured Clinical Interview for DSM-IV (13). Of the AN women, 21 were exclusively food restrictors and 12 occasionally binged with a frequency no greater than twice a week. At the time of the study, two anorexics had a concomitant diagnosis of major depression and one of borderline personality disorder. Patients with BN were all of the purging subtype, with binge episodes always followed by self-induced vomiting; two of these patients also abused laxatives and six exercised excessively. Four had a positive history of AN and four had a concomitant diagnosis of major depression and five of borderline personality disorder. All of the AN women and 9 bulimics were amenorrheic; 15 bulimics were oligomenorrheic; the remaining patients had normal regular menses.

At the time of the study, some patients had never taken psychotropic medication (20 anorexics, 34 bulimics); the remaining ones had been drug free for more than 6 weeks.

Control women were mentally healthy, as assessed by a clinical interview, and had no positive family history of mental disorders as assessed by the Family History Research Diagnostic Criteria (14). They were regularly menstruating and had normal diets.

Both patients and healthy volunteers had normal physical examinations, normal values of routine blood and urine tests, and a normal electrocardiogram. Female controls and patients who were normally menstruating were tested in the follicular phase of their menstrual cycle (day 5–10 from menses). The follicular phase was chosen to avoid having to monitor the ovulation in our outpatients. No subject was taking oral contraceptives or had a past history of alcohol or drug abuse.

Procedure
The study was approved by the ethics committee of our department. Subjects gave written informed consent before study participation.

Both patients and healthy subjects underwent a psychopathological assessment by a) the EDI (15) and BITE (16) to evaluate the eating-related psychopathology and b) HDRS (17) to measure concomitant depressive symptoms.

On the day before blood collection, subjects were asked to abstain from food consumption after 8:00 PM. This was carefully checked by relatives; hence, no subject ate after the scheduled time, but some of them (especially anorexic women) had their last meal before 8:00 PM. On the day after, each subject underwent a blood sample collection in the morning, between 8:00 and 9:00 AM, when a butterfly needle was inserted into a forearm vein. The catheter was kept patent by a saline infusion that was slowly infused in order to have the same amount of fluid injected in each individual. Subjects rested supine for 30 minutes, then blood was withdrawn and collected in tubes with lithium heparin as anticoagulant. Plasma was separated by centrifugation and stored at -20°C.

Biochemical Analyses
Plasma PRL and 17ß-estradiol levels were determined by an immunometric method (MAIA clone) using commercial kits purchased from Biochem Immuno System (Milan, Italy). The lower detection limits were 0.4 ng/ml for PRL and 5 pg/ml for 17ß-estradiol. Intra- and interassay coefficients of variation were 5.4 and 8.6% for PRL, respectively, and 4.3 and 3.2% for 17ß-estradiol, respectively. Plasma cortisol concentrations were determined by a double-antibody radioimmunoassay method, using commercial kits purchased from Biochem Immuno System. The lower detection limit was 27 nmol/liter. Intra- and interassay coefficients of variation were less than 5 and 8%, respectively.

Plasma leptin values were determined by a sandwich enzyme-linked immunosorbent assay using a commercial kit purchased from Alexis Biochemicals (Laufelfingen, Switzerland). The sensitivity of the method was 0.2 ng/ml; intra- and interassay coefficients of variation were 6.1 and 8.5%, respectively.

Data Analysis
The BMDP statistical software package (18) was used for data analysis. Because there were significant deviations from normality in the data, nonparametric statistical analyses were used. Where the Kruskal-Wallis ANOVA showed significant differences among the groups, the Mann-Whitney U test was used to assess differences between patients and controls. The Spearman rank order correlation test was used to examine the relationships between leptin values on the one hand and demographic, clinical, or hormonal data on the other. The ² test with Yate’s correction was used to investigate differences in the patients’ distributions between the two bulimic subgroups.

	RESULTS

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Demographic and Clinical Data
The Kruskal-Wallis ANOVA showed significant intergroup differences in BW (p < .0001) and BMI (p < .0001) but not in age. With respect to both healthy women and BN subjects, AN patients had significantly lower BW and BMI whereas no significant difference in these parameters was observed between bulimics and healthy women (Table 1).

Hormonal Data
Kruskal-Wallis ANOVA showed significant intergroup differences in plasma levels of leptin (p < .0001), 17ß-estradiol (p < .01), PRL (p < .0001), and cortisol (p < .0001). Compared with healthy women, AN and BN patients exhibited significantly decreased plasma levels of leptin, 17ß-estradiol, and PRL but enhanced levels of cortisol (Table 1). Moreover, compared with anorexic patients, women with BN showed significantly higher plasma levels of leptin, PRL, and 17ß-estradiol but decreased values of plasma cortisol (Table 1).

Leptin plasma levels were positively correlated with both BW and BMI in AN ( = .32, p < .04; = .41, p < .009, respectively) and BN patients ( = .31, p < .008; = .32, p < .008) as well as in healthy women ( = .50, p < .0006; = .60, p < .0001). Moreover, in the bulimic group, circulating leptin was negatively correlated with both the length of the illness ( = -.30, p < .02) and the frequency of binge/vomiting episodes ( = -.28, p < .02).

Scatterplots of plasma leptin concentrations in the three subject groups showed that some bulimics had hormone values similar to those found in anorexic women while others had leptin concentrations in the range of normal women (Figure 1). Therefore, we ran a stepwise discriminant analysis that identified the plasma leptin level of 5.6 ng/ml as the value leading to the best classification of anorexics and healthy subjects. Indeed, all anorexic patients (100%) had plasma leptin levels below this value, whereas 30 (78.9%) of our 38 healthy women had circulating leptin above this value. On the basis of this result, we divided bulimic patients into two groups: 1) women (N = 29) with plasma leptin levels below the value of 5.6 ng/ml and 2) women (N = 27) with plasma leptin concentrations above that value and compared their clinical and demographic characteristics. The first group had mean (±SD) plasma leptin of 2.1 ± 1.4 ng/ml, which did not significantly differ from anorexic patients; the second group had mean plasma leptin of 10.9 ± 4.8 ng/ml, which was not statistically different from normal controls.

View larger version (16K): [in this window] [in a new window]   Fig. 1. Scatterplots of individual plasma leptin concentrations in patients with anorexia nervosa or bulimia nervosa and in healthy subjects.

	DISCUSSION

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This study found that underweight patients with AN and normal weight women with BN had significantly decreased concentrations of plasma leptin compared with age- and sex-matched healthy subjects. These results are consistent with recent reports from both other authors and our group showing that blood levels of leptin are reduced in patients with current symptoms of AN or BN (8–11).

The main finding of this study was that some bulimic patients had plasma leptin levels indistinguishable from those of AN people and others had leptin concentrations absolutely similar to normal controls. This means that there exists a group of BN patients that hyposecretes leptin, although their BW and BMI do not exhibit significant modifications. Therefore, reasons other than current BW values are likely involved in reducing leptin production in these patients.

One contributing factor to low leptin synthesis in this group of bulimics could be a previous history of substantial weight loss because sustained weight decrease has been shown to result in a persistent reduction of circulating leptin in humans (19, 20). However, BN patients with low leptin did not significantly differ from those with normal leptin in both the maximum past and present BW, so that BW changes were not different in the two groups. Similarly, no significant differences could be detected between the two bulimic groups in other clinical and hormonal parameters that could influence leptin secretion such as menstrual function, plasma levels of estrogens or cortisol, previous history of AN, presence of compensatory behaviors other than vomiting, and all but one (maturity fears) psychometric measure. Conversely, in bulimics with low leptin levels, the length of the illness and the number of daily binge/vomiting episodes were significantly higher than in those with normal leptin secretion, suggesting that the chronicity of the illness and the severity of the bingeing/vomiting behavior could be determinants of the leptin hyposecretion. This hypothesis was corroborated by the finding of negative correlations between circulating leptin and the duration of the illness and the frequency of bingeing and vomiting in the overall bulimic group.

Consistent with our results, Jimerson et al. (10) found a trend toward a negative correlation between decreased leptin levels and the frequency of bingeing in a sample of symptomatic BN women. Those authors suggested that this correlation would imply that an impaired leptin function at the hypothalamic level could contribute to the bingeing behavior through a reduction of patients’ satiety responses. Although at present this speculation cannot be excluded, we believe that the negative correlation between leptin levels and binge/vomiting frequency together with the inverse correlation between the hormone and the length of the illness rather reflect changes in subjects’ nutritional parameters and/or eating patterns chronically affecting leptin synthesis. In support of this idea, it is known that bulimics, despite the large caloric ingestion during the binge episodes, regurgitate a relatively large amount of calories by vomiting and engage in prolonged starvation in order to reduce their daily caloric ingestion. A caloric intake below the normal requirements of the respective individual even with no loss of fat mass may have an impact on leptin synthesis. Moreover, although BN is not characterized by denutrition and cachexia as AN, the incorrect food selection of bulimics may lead to the lack of certain macro/micronutrients in the diet that may affect leptin production (3). Finally, it has been recently shown that, in normal individuals, experimental binge-eating dramatically flattens the leptin diurnal rhythm even when total calories are appropriate for weight (21). Therefore, it seems possible to conclude that, in the subgroup of bulimic patients with low plasma leptin, the chronic malnutrition occurring with restrained eating and purging behaviors act in concert with the bingeing behavior to affect profoundly and persistently leptin synthesis.

A further characteristic differentiating the low-leptin bulimic group from the normal-leptin group was a significantly greater occurrence of concomitant borderline personality disorder in the first group of patients, who showed also a higher score on the maturity fears subitem of the EDI. This difference is not easily understandable in terms of leptin secretion.

Some limitations of the present study need to be discussed. First, it must be noted that body fat correlates with leptin levels to a higher degree than does BW, mainly in patients with eating disorders (22). Hence, a shortcoming of the present study is that we did not measure body fat, which may have differed between groups independently of BMI. However, within the same gender and age range, it has been proven that BMI may represent average body fat estimates (23). Moreover, in the study of Jimerson et al. (10), bulimic women exhibited decreased plasma leptin concentrations in spite of no significant changes in their body fat mass.

A further limitation of our study is that our findings refer to a single morning measurement of blood hormones in spite of the well-recognized circadian rhythmicity in the secretion of these substances. However, although we cannot exclude that phase shifts in the circadian rhythms may have affected our results, it is of note that the present findings are in line with previously reported values of morning plasma leptin, cortisol, and PRL in eating disorder patients. Finally, this is a cross-sectional study that leaves unanswered the question whether bulimics with a shorter duration of illness and with a lower binge/vomiting frequency will ultimately develop lower leptin levels if the disease process continues. A longitudinal analysis will help clarify the issue.

In conclusion, our data confirm that circulating leptin is decreased in both symptomatic underweight anorexic patients and normal-weight bulimic subjects. Moreover, in BN, a reduced leptin production is peculiar of a subgroup of patients characterized by a more chronic illness and a greater severity of the bingeing/vomiting behavior. The prevalence of one or the other type of bulimic patients in a study sample may help explain the discrepancies among the different studies.

Received for publication June 4, 2001.

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