# NEUROLOGICAL AND PSYCHIATRIC DISORDERS IN ENDOCRINE DISEASES

EDITED BY: Gianluca Tamagno and Jacques Epelbaum PUBLISHED IN: Frontiers in Neuroscience and Frontiers in Endocrinology

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ISSN 1664-8714 ISBN 978-2-88919-589-3 DOI 10.3389/978-2-88919-589-3

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# **NEUROLOGICAL AND PSYCHIATRIC DISORDERS IN ENDOCRINE DISEASES**

Topic Editors:

**Gianluca Tamagno,** Mater Misericordiae University Hospital – University College Dublin, Ireland

**Jacques Epelbaum,** Center for Psychiatry and Neuroscience, UMR 894 – INSERM, Université Paris Descartes, Sorbonne Paris Cité, France

Neurological and psychiatric disorders can occur in endocrine diseases either in the setting of the clinical manifestations of the same (i.e., hyper- or hyposecretion of hormones or peptides from the endocrine glands) or as events secondary to the pathogenetic mechanisms of the endocrinopathy (i.e., autommunity affecting endocrine glands and the brain). Also the medical or surgical treatment of the endocrine disease can sometimes determine the occurrence of neurological or psychiatric abnormalities. Moreover some genetic alterations can lead to syndromes affecting both the endocrine and the nervous system with a variety of possible manifestations. In the last couple of decades a number of associations between dysfunctions of the endocrine system and neurological or psychiatric manifestations have appeared and only in the minority of the cases this link has been fully elucidated. Often the neurological or psychiatric alterations still represent a relevant challenge for clinicians with regard to the management of the patients. The complexity of the topic and the limited availability of laboratory research models for the study of the endocrine system-nervous system cross-interaction are making the scientific progresses intricate and, sometimes, slow. A dedicated focus to such broad and often still obscure topic might help and clarify the current state-of-the-art in the field and direct the goals of future research.

**Citation:** Gianluca Tamagno and Jacques Epelbaum, eds. (2015). Neurological and psychiatric disorders in endocrine diseases. Lausanne: Frontiers Media. doi: 10.3389/978-2-88919-589-3

# Table of Contents

Gianluca Tamagno and Jacques Epelbaum

*08 Irisin levels are not affected by physical activity in patients with anorexia nervosa*

Tobias Hofmann, Ulf Elbelt, Anne Ahnis, Peter Kobelt, Matthias Rose and Andreas Stengel

*16 Personality and psychiatric disorders in women affected by polycystic ovary syndrome*

Elisabetta Scaruffi, Alessandra Gambineri, Stefania Cattaneo, Jenni Turra, Roberto Vettor and Roberto Mioni

Hernan Valdes-Socin, Matilde Rubio Almanza, Mariana Tomé Fernández-Ladreda, François Guillaume Debray, Vincent Bours and Albert Beckers

*37 Health outcomes in acromegaly: depression and anxiety are promising targets for improving reduced quality of life*

Victor Jacobus Geraedts, Christina Dimopoulou, Matthias Auer, Jochen Schopohl, Günter Karl Stalla and Caroline Sievers

*44 Neuropsychiatric disorders in Cushing's syndrome*

Rosario Pivonello, Chiara Simeoli, Maria Cristina De Martino, Alessia Cozzolino, Monica De Leo, Davide Iacuaniello, Claudia Pivonello, Mariarosaria Negri, Maria Teresa Pellecchia, Felice Iasevoli and Annamaria Colao

*50 Neurological complications in thyroid surgery: a surgical point of view on laryngeal nerves*

Emanuela Varaldo, Gian Luca Ansaldo, Matteo Mascherini, Ferdinando Cafiero and Michele N. Minuto

# *71 Ghrelin-derived peptides: a link between appetite/reward, GH axis, and psychiatric disorders?*

Alexandra Labarthe, Oriane Fiquet, Rim Hassouna, Philippe Zizzari, Laurence Lanfumey, Nicolas Ramoz, Dominique Grouselle, Jacques Epelbaum and Virginie Tolle

# *90 Apo-ghrelin receptor (apo-GHSR1a) regulates dopamine signaling in the brain*

Andras Kern, Cristina Grande and Roy G. Smith

# **Editorial: Neurological and psychiatric disorders in endocrine diseases**

#### *Gianluca Tamagno1 \* and Jacques Epelbaum2 \**

*1 Department of Endocrinology/Diabetes, Mater Misericordiae University Hospital, University College Dublin, Dublin, Ireland, 2 UMR 894, Center for Psychiatry and Neuroscience, INSERM, Université Paris Descartes, Sorbonne Paris Cité, Paris, France*

**Keywords: eating disorders, exercise, gender identity, acromegaly, Cushing syndrome, polycystic ovary syndrome, obesity, hypogonadism, thyroid surgery, quality of life**

In the last few decades, a number of reciprocal associations between endocrine system dysfunctions and neurological or psychiatric manifestations have appeared, and only in the minority of the cases this link has been fully elucidated (1). Unfortunately, the complexity, and sometimes also the severity, of the clinical issues and the limited availability of reliable experimental models for the study of the endocrine system–nervous system cross-interactions do not help the fast achievement of scientific progresses.

Isolated neurological or psychiatric symptoms as well as more complex disorders can occur in the setting of most endocrine diseases. Such manifestations can be the direct result of hyper- or hyposecretion of hormones from the endocrine glands or may occur as events secondary to the pathogenic mechanisms of the endocrinopathy, like in the case of autoimmunity affecting both endocrine glands and the brain (2). Moreover, the medical or surgical treatment of endocrine diseases can sometimes determine the occurrence of neurological or psychiatric syndromes. Some genetic aberrations can also lead to conditions affecting both the endocrine and the nervous system with a variety of possible manifestations. On the other side, many psychiatric or neurological diseases, including but not limiting to conditions, which affect the hypothalamus or the pituitary gland, may impair the physiological endocrine functions and demonstrate once again the intricate complexity of the psychoneuroendocrine connection.

Covering all basic, translational, and clinical aspects in a single Research Topic on the "Neurological and psychiatric disorders in endocrine diseases" cannot be realistically feasible. The scope is simply raising a number of interesting points from experts in the field to bring a contribution to the knowledge and maybe direct future research in this very intriguing area. Thus, we have collected reviews and original research articles, which may speak to a wide audience, including neurologists, psychiatrists, endocrinologists, and basic scientists in neuroscience, endocrinology, and metabolism.

In the clinical area, an interesting study assesses the plasma levels of the muscle-derived hormone irisin in anorexic women, without finding any significant correlation between irisin level and physical exercise (3). Another fascinating study analyzes the complexity of psychiatric and personality disorders in women with polycystic ovary syndrome, highlighting the issue of psychological distress in this patient group (4). A comprehensive picture of gender dysphoria, a psychiatric condition, which requires endocrine management, in Ireland is presented (5). In a short review, the most relevant reproductive, neurodevelopmental, and genetic aspects of hypogonadotropic hypogonadal syndromes are outlined (6). A German study indicates that psychopathology significantly predicts quality of life in patients with acromegaly and suggests that depressive symptoms and anxiety, being modifiable factors, may represent relevant targets for a broad treatment intervention in acromegalic patients (7). In a concise review, the psychiatric alterations, the neurocognitive impairment, and the altered quality of life affecting at some extent the majority of the patients with Cushing's syndrome are summarized, and the authors highlight that resolution of hypercortisolism, a challenging and

#### *Edited and reviewed by:*

*Hubert Vaudry, University of Rouen, France*

#### *\*Correspondence:*

*Gianluca Tamagno<EMAIL_ADDRESS>Jacques Epelbaum<EMAIL_ADDRESS>
#### *Specialty section:*

*This article was submitted to Neuroendocrine Science, a section of the journal Frontiers in Endocrinology*

> *Received: 06 April 2015 Accepted: 27 April 2015 Published: 12 May 2015*

#### *Citation:*

*Tamagno G and Epelbaum J (2015) Editorial: Neurological and psychiatric disorders in endocrine diseases. Front. Endocrinol. 6:75. doi: 10.3389/fendo.2015.00075* non-granted achievement, does not always lead to the complete remission of the neuropsychiatric changes or restore the quality of life (8). With an incursion in the surgical field, the complications potentially affecting the laryngeal nerves during thyroid surgery are extensively reviewed, suggesting that still the incidence of laryngeal nerves damage secondary to thyroid surgery cannot be suppressed but may be maintained in a low range with thorough surgical techniques and the use of intraoperative neuromonitoring (9). Three articles address the complex interactions between metabolism and neuropsychiatric symptoms. The first

# **References**

one focuses on biological differences between restrictive anorexia nervosa and constitutional thinness, a controversial concept to describe young girls who follow a normal diet and differ from restrictive anorexia nervosa on a number of endocrine parameters (10). At the opposite of the spectrum, the second one reviews the role of inflammatory processes in the neuropsychiatric comorbidity associated with obesity (11). Finally, the last one summarizes the fascinating link through ghrelin peptides between appetite/reward/growth hormone axis and psychiatric disorders (12).

**Conflict of Interest Statement:** The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

*Copyright © 2015 Tamagno and Epelbaum. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.*

# **Corrigendum: "Editorial: Neurological and psychiatric disorders in endocrine diseases"**

*Gianluca Tamagno1 \* and Jacques Epelbaum2*

*1 Department of Endocrinology/Diabetes, Mater Misericordiae University Hospital, University College Dublin, Dublin, Ireland, 2 UMR 894, Center for Psychiatry and Neuroscience, INSERM, Université Paris Descartes, Sorbonne Paris Cité, Paris, France*

**Keywords: eating disorders, exercise, gender identity, acromegaly, Cushing syndrome, polycystic ovary syndrome, obesity, hypogonadism, thyroid surgery, quality of life**

# **A corrigendum on**

**Editorial: Neurological and psychiatric disorders in endocrine diseases** *by Tamagno G, Epelbaum J (2015). Front Endocrinol 6:75. doi: 10.3389/fendo.2015.00075*

We have added the following correction to the Editorial article, now including a reference to Kern et al. "Apo-ghrelin receptor (apo-GHSR1a) regulates dopamine signaling in the brain."

Four articles address the complex interactions between metabolism and neuropsychiatric symptoms. The first one focuses on biological differences between restrictive anorexia nervosa and constitutional thinness, a controversial concept to describe young girls who follow a normal diet and differ from restrictive anorexia nervosa on a number of endocrine parameters (10). At the opposite of the spectrum, the second one reviews the role of inflammatory processes in the neuropsychiatric comorbidity associated with obesity (11). The third one summarizes the fascinating link through ghrelin peptides between appetite/reward/growth hormone axis and psychiatric disorders (12). Finally, the last one proposes a molecular mechanism through allosteric interactions between dopamine/DRD2 and GHSR1 receptors for controlling appetite and the uncontrollable hyperphagia associated with Prader–Willi syndrome (13).

Addition of reference:

13. Kern A, Grande C, Smith RG. Apo-ghrelin receptor (apo-GHSR1a) regulates dopamine signaling in the brain. *Front Endocrinol* (2014) **5**:129. doi:10.3389/fendo.2014.00129

**Conflict of Interest Statement:** The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

*Copyright © 2015 Tamagno and Epelbaum. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.*

*Edited and reviewed by: Hubert Vaudry, University of Rouen, France*

*\*Correspondence:*

*Gianluca Tamagno<EMAIL_ADDRESS>
#### *Specialty section:*

*This article was submitted to Neuroendocrine Science, a section of the journal Frontiers in Endocrinology*

> *Received: 02 June 2015 Accepted: 05 June 2015 Published: 17 June 2015*

#### *Citation:*

*Tamagno G and Epelbaum J (2015) Corrigendum: "Editorial: Neurological and psychiatric disorders in endocrine diseases". Front. Endocrinol. 6:101. doi: 10.3389/fendo.2015.00101*

# Irisin levels are not affected by physical activity in patients with anorexia nervosa

#### **Tobias Hofmann1\* † , Ulf Elbelt 1,2† , Anne Ahnis 1 , Peter Kobelt 1 , Matthias Rose1 and Andreas Stengel 1**

1 Division for General Internal and Psychosomatic Medicine, Charité Center for Internal Medicine and Dermatology, Charité – Universitätsmedizin Berlin, Berlin, Germany

2 Division for Endocrinology, Diabetes and Nutrition, Charité Center for Internal Medicine with Gastroenterology and Nephrology, Charité – Universitätsmedizin Berlin, Berlin, Germany

#### **Edited by:**

Gianluca Tamagno, St. Columcille's Hospital, Ireland

#### **Reviewed by:**

Francisco Gaytán, University of Cordoba, Spain Günter Karl Stalla, Max-Planck-Institute of Psychiatry, Germany

#### **\*Correspondence:**

Tobias Hofmann, Division for General Internal and Psychosomatic Medicine, Charité Center for Internal Medicine and Dermatology, Charité – Universitätsmedizin Berlin, Hindenburgdamm 30, 12200 Berlin, Germany e-mail<EMAIL_ADDRESS>
†Tobias Hofmann and Ulf Elbelt have contributed equally to this work.

Irisin was recently identified as muscle-derived hormone that increases energy expenditure. Studies in normal weight and obese subjects reported an increased irisin expression following physical activity, although inconsistent results were observed. Increased physical activity in a subgroup of patients with anorexia nervosa (AN) complicates the course of the disease. Since irisin could account for differences in clinical outcomes, we investigated irisin levels in anorexic patients with high and moderate physical activity to evaluate whether irisin differs with increasing physical activity. Hospitalized female anorexic patients (n = 39) were included. Plasma irisin measured by enzymelinked immunosorbent assay and locomotor activity were assessed at the same time. Patients were separated into two groups (n = 19/group; median excluded): moderate and high activity (6331 ± 423 vs. 13743 ± 1047 steps/day, p < 0.001). The groups did not differ in body mass index (14.2 ± 0.4 vs. 15.0 ± 0.4 kg/m2 ), irisin levels (558.2 ± 26.1 vs. 524.9 ± 25.2 ng/ml), and body weight-adjusted resting energy expenditure (17.6 ± 0.3 vs. 18.0 ± 0.3 kcal/kg/day, p > 0.05), whereas body weight-adjusted total energy expenditure (46.0 ± 1.4 vs. 41.1 ± 1.1 kcal/kg/day), metabolic equivalents (METs, 1.9 ± 0.1 vs. 1.7 ± 0.1 METs/day), body weight-adjusted exercise activity thermogenesis (1.8 ± 0.5 vs. 0.6 ± 0.3 kcal/kg/day), duration of exercise (18.6 ± 4.7 vs. 6.2 ± 3.1 min/day), and body weight-adjusted non-exercise activity thermogenesis (21.6 ± 1.0 vs. 18.8 ± 0.8 kcal/kg/day) were higher in the high activity compared to the moderate activity group (p < 0.05). No correlations were observed between irisin and activity parameters in the whole sample (p > 0.05). In conclusion, the current data do not support the concept of irisin being induced by exercise, at least not under conditions of severely reduced body weight like AN.

**Keywords: brown adipose tissue, energy expenditure, exercise, FNDC5, myokine, SenseWear™ armband**

# **INTRODUCTION**

Irisin is a recently identified muscle-derived hormone produced by cleavage from fibronectin type III domain containing 5 (FNDC5) (1). The expression of FNDC5 is induced by the peroxisome proliferator-activated receptor γ (PPARγ) coactivator 1α (PGC-1α) that has been demonstrated to play an important role in the expression of uncoupling protein 1 (UCP1), a key factor of thermogenesis in brown adipose tissue (2).  These beige fat cells have been shown to be a cell type distinct from white or brown fat cells being located in white adipose tissue (3) and exhibiting properties of brown adipose tissue which – for long thought to exist only in newborns – was recently also detected in adult humans (4, 5) and demonstrated to be activated in response to cold exposure (4). Besides its occurrence in skeletal muscle, irisin was detected – although in much smaller quantities – in several human tissues containing smooth muscle as well as heart muscle, and organs including liver, kidney, and lung (6). Additionally, this peptide is also present in adipose tissue (7, 8) and rodent cerebellar Purkinje cells (9). Lastly, irisin was detected in the circulation (1, 6, 10, 11) with decreased levels in patients with type 2 diabetes mellitus (10).

In their initial study, Boström et al. described an increase of circulating irisin levels following physical activity in mice and in male subjects after a 10-week endurance exercise program (1). In a subsequent study, elevated circulating irisin levels were observed following acute exercise consisting of 80-m sprints, while this effect was weakened after a 8-week sprint training program completed by young healthy men (6). However, one study detected a modest 30% higher FNDC5 mRNA expression only in a subgroup of older male subjects after an endurance training program compared to sedentary controls, whereas no changes were observed in a large sample of younger adults (12). While data on an exercisedependent production of irisin in muscle and its secretion into the circulation are inconsistent in humans, an association of plasma irisin levels with body mass index (BMI) seems to be a robust finding. Extending the results of a previous study that showed a trend toward a correlation of circulating irisin with BMI in subjects with a BMI range from 20 to 48 kg/m2 (6), we previously reported a positive correlation of plasma irisin levels with BMI over a very broad spectrum of body weights with BMIs ranging from 8 to 85 kg/m2 as well as positive associations with fat mass (FM), fat-free mass (FFM), and body cell mass (BCM) (11). One previous study on the association of irisin and energy expenditure reported that irisin levels correlated with 24-h energy expenditure measured by indirect calorimetry in overweight and obese postmenopausal women only in a subgroup of subjects whose energy expenditure was higher than predicted by an FFM-based equation (13), whereas it did not hold true when the whole sample was analyzed.

In light of these data, more research is needed in order to characterize the regulation of irisin under conditions of different activity patterns and to establish its possible role as a novel muscle-derived and exercise-induced hormone. In contrast to its potentially beneficial effect as health-promoting hormone in overweight or obese patients, the role as an exercise-induced and energy-dissipating myokine might be an additional biological mechanism explaining worse clinical prognosis and higher relapse rates of hyperactive anorexic subjects. Furthermore, anorexia nervosa (AN) might be viewed as a mirror image of obesity and thus investigation of this disease likely contributes to the understanding of weight disorders in general. Therefore, in the present cross-sectional study we investigated the relationship of circulating irisin levels with measures of physical activity and different aspects of energy expenditure in anorexic patients, since subjects suffering from AN are known to display very different activity patterns encompassing a relevant subgroup of hyperactive in contrast to moderately active patients (14–16).

# **SUBJECTS AND METHODS**

### **SUBJECTS**

We recruited 39 anorexic female inpatients at admission to their treatment in the Division for General Internal and Psychosomatic Medicine at Charité – Universitätsmedizin Berlin. Inclusion criteria were a BMI of <18 kg/m2 and fulfillment of ICD-10 (International Statistical Classification of Diseases and Related Health Problems of the World Health Organization, 10th revision) criteria (17) for typical or atypical AN. Atypical AN is defined as a disorder meeting some criteria of the typical form (BMI below 17.5 kg/m2 , self-induced weight loss, body image distortion, and endocrine disorder most often reflected by secondary amenorrhea) without fulfilling all key symptoms (17). The restricting subtype of AN according to the fourth edition of the Diagnostic and Statistical Manual of Mental Disorders (DSM-IV) differs from the purging subtype by the absence of binge-eating or purging behavior such as self-induced vomiting or misuse of appetite suppressants, thyroid hormones or laxatives, diuretics, and enemas (18).

According to their locomotor activity, anorexic patients were separated into two groups by dividing the patients at the median step count resulting in a moderate activity and a high activity group (*n* = 19/group, one patient with the median value was excluded). Additionally, since the restricting subtype was shown to be associated with excessive exercise (19, 20), calculations were also performed for three subgroups of AN: typical restricting, typical purging, and atypical AN.

Patients with current pregnancy or psychotic disorders and an age of <18 years were excluded. The investigations were conducted according to the Declaration of Helsinki. All patients gave written informed consent and the study was approved by the institutional ethics committee of the Charité – Universitätsmedizin Berlin (protocol number: EA1/114/10).

### **LABORATORY ANALYSIS OF IRISIN PLASMA LEVELS**

All blood samples were collected from a forearm vein between 7 and 8 a.m. after an overnight fast during the period of the data collection on physical activity. Participants were allowed to drink small amounts of water but were advised not to smoke or exercise before blood withdrawal. The blood was collected in pre-cooled EDTA tubes prepared with aprotinin (1.2 Trypsin Inhibitory Unit per 1 ml blood; ICN Pharmaceuticals, Costa Mesa, CA, USA) for peptidase inhibition. The tubes were placed back on ice immediately after blood withdrawal and then centrifuged at 4°C for 10 min at 3000 × *g*. Plasma was separated and stored at −80°C until further processing. At the day of the measurement, samples were diluted 1:10 and irisin plasma levels were analyzed using a commercial enzyme-linked immunosorbent assay (ELISA, catalog # EK-067-16, Phoenix Pharmaceutical, Inc., Burlingame, CA, USA). All samples were processed in one batch (intra-assay variability <5%).

# **MEASUREMENTS AND CALCULATION OF ACTIVITY AND ENERGY EXPENDITURE**

Weight and height were assessed on the day of the blood collection. Weight was measured to the nearest 0.1 kg and height to the nearest 0.5 cm. The BMI was calculated as kilogram per square meter. Physical activity data were continuously measured for 3 days (Friday to Sunday) during inpatient treatment using a portable armband device (SWA; SenseWear™ PRO3 armband; BodyMedia, Inc., Pittsburgh, PA, USA). One day was included in the data analysis if the minimum duration of data acquisition was 20 h and 30 min/day. Patients were not activity restricted during the measurement. Activity displayed by individuals is suggested to be affected only in part deliberately with a high predictive value of the activity level under sedentary conditions for daily life activity (21). Although activity patterns under conditions of hospitalization might differ from those in daily life, activity habits are likely to persist during inpatient treatment if activity is not restricted due to a genetic influence on physical activity (22).

The SWA uses a multisensory array including sensors measuring heat flux, galvanic skin response, skin temperature, near-body ambient temperature, and a two-axis accelerometer. Step counts were measured by the accelerometer and directly taken for analyses. Obtained data were analyzed using a generalized proprietary algorithm developed by the manufacturer (SenseWear™ Software, Version 6.1, BodyMedia, Inc.) and prepared for statistical analysis.

Total energy expenditure (TEE) consists of resting energy expenditure (REE), thermic effect of food (TEF), and activity thermogenesis. Activity thermogenesis can be further separated into the two components exercise-related activity thermogenesis (EAT, deliberate physical exercise, sports) and non-exercise activity thermogenesis (NEAT, spontaneous daily physical activity) (23). Energy expenditure of more than five metabolic equivalents (METs) of a task (a physiological measure expressing the energy cost of physical activities) was classified as EAT, whereas energy expenditure of up to five METs was classified as NEAT adapting the findings of Ainsworth et al. (24) and as described before by our group (25). While TEE, EAT, and duration of exercise were directly received from the proprietary algorithms of the SWA, NEAT was calculated according to the equation: NEAT = TEE − TEF − REE − EAT. TEF was estimated as 10% of TEE and calculated as TEE × 0.1 (26). REE, required for calculation of NEAT, was calculated using weight-/group-specific REE prediction equations provided by Müller et al. (27) since REE cannot be directly determined by the SWA.

#### **STATISTICAL ANALYSIS**

Distribution of the data was determined by using the Kolmogorov– Smirnov test. Data are expressed as mean ± SEM. Differences between groups were calculated using the *t*-test. For the comparison of the subtypes of AN differences between groups were calculated using one-way analysis of variance (ANOVA) followed by Tukey *post hoc* test. Correlations were determined by Pearson's or Spearman's analysis depending on the distribution of the data. Differences between groups were considered significant when *p* < 0.05. All statistical analyses were conducted using SigmaStat 3.1.

# **RESULTS**

### **IRISIN PLASMA LEVELS DO NOT DEPEND ON ACTIVITY AND ENERGY EXPENDITURE**

Per definition, the moderate activity and the high activity group differed significantly in the number of steps performed per day (*p* < 0.001; **Figure 1A**). However, irisin plasma levels were not different between these two groups (*p* = 0.37; **Figure 1B**). Associated with the higher activity, significant differences were observed for METs per day (*p* = 0.03;**Figure 1C**) and body weight-adjusted TEE (*p* = 0.009; **Figure 1D**) with higher values in the high compared to the moderate activity group. No differences were detected for body weight-adjusted REE (*p* = 0.38; **Figure 1E**), whereas duration of exercise (*p* = 0.03; **Figure 1F**) or body weight-adjusted EAT (*p* = 0.03; **Figure 1G**) and body weight-adjusted NEAT (*p* = 0.03; **Figure 1H**) were higher in the high activity compared to the moderate activity group.

In the whole sample no correlation was observed between plasma irisin levels and the number of steps as a measure of activity (**Figure 2A**), the amount of METs (**Figure 2B**) and other parameters of energy expenditure, namely body weightadjusted TEE (**Figure 2C**), body weight-adjusted REE (**Figure 2D**), body weight-adjusted EAT (**Figure 2E**), and body weight-adjusted NEAT (**Figure 2F**).

# **IRISIN PLASMA LEVELS SHOW A NEGATIVE CORRELATION WITH COMPONENTS OF ENERGY EXPENDITURE IN PATIENTS WITH RESTRICTING TYPE ANOREXIA NERVOSA ONLY**

After division of the sample into subgroups according to the type of AN, namely purging type (*n* = 10), restricting type (*n* = 20), and atypical (*n* = 9) AN, the three groups did not differ in terms of age and BMI (*p* > 0.05; **Table 1**). As shown in **Table 1**, no differences among these three groups were observed for irisin, number of steps per day, METs per day, and the different components of energy expenditure (*p* > 0.05).

When investigating the three subgroups separately, irisin levels showed a negative correlation with the number of steps per day, duration of exercise, and body weight-adjusted EAT in patients with restricting type AN (*p* < 0.05; **Table 2**), whereas no correlations of plasma irisin and components of energy expenditure were observed for the subgroups of purging type and atypical AN (*p* > 0.05; **Table 2**).

# **DISCUSSION**

Irisin is a newly defined myokine that increases energy expenditure by stimulating the expression of UCP1 and thus the browning of white adipose tissue (1). In addition, it has been shown that irisin plasma levels are increased in response to different types of exercise (1, 28, 29). Consequently, irisin was proposed to mediate some of the health-promoting effects of physical activity. In turn, irisin could be an additional biological mechanism contributing to the course of the disease and therefore the prognosis in highly active patients suffering from AN (30, 31). However, in rejection of our initial hypothesis, we did not detect differences in irisin plasma levels between anorexic patients with high vs. moderate locomotor activity as determined by average daily step count. When investigating the whole cohort, we also did not observe any correlations of circulating irisin with physical activity, TEE or other parameters of energy expenditure, in particular EAT. Several points could contribute to this lack of correlation of irisin with measures of locomotor activity and energy expenditure in our study.

First, Timmons et al. reported that they failed to detect a strong increase of the irisin precursor, FNDC5 after exercise in a population of younger adults, whereas only in a subgroup of older subjects an increase of FNDC5 mRNA expression after exercise was observed (12). Since our data were collected in a population of young patients with a mean age of 27, the results could underline the hypothesis that exercise-related effects on irisin expression are associated with age and only (or predominantly) observed in elderly subjects.

Second, irisin is a cleavage product of FNDC5 expressed mainly in skeletal muscle.  In addition, using bioelectrical impedance analysis (BIA) in a previous study, we showed a positive correlation of irisin with FFM and BCM (the measure best reflecting muscle mass using BIA) (11). Since patients suffering from AN have greatly diminished muscle mass as a consequence of malnutrition and weight loss, the total muscle mass might be too low in our patient population (BCM 15.54 ± 0.43 kg; FFM 37.02 ± 0.66 kg) to detect differences in the levels of the circulating myokine, irisin.

Third, in line with our finding of a lacking association between energy expenditure and circulating irisin, one recent study did not detect a correlation of plasma irisin levels with 24-h energy expenditure in post-menopausal women when assessed in a metabolic chamber (13). Subjects in this and our study are in a state of downregulated sexual hormones that could contribute to the results. Another explanation could be that only exercise with respect to a defined acute or repetitive physical training program exerts significant effects on irisin muscle expression or plasma levels, whereas chronic changes in exercise do not. This might be due to counterregulatory adaptive changes that have to be further investigated.

In addition, since irisin has been shown to initiate the browning of white adipose tissue and thereby eliciting an additional energydissipating effect (1, 3), the existence of a reasonable amount of beige adipocytes would be crucial to mediate these effects. However, little is known about the existence of brown and beige adipose tissue in fat-deprived anorexic subjects. The activity of brown fat has been reported to be inversely correlated with BMI with reduced activity in overweight or obese compared to lean young male subjects (4). However, a more recent study did not detect brown fat activity in anorexic patients, whereas it was detectable in constitutionally lean (average BMI 16.2 kg/m2 ) subjects (32). The absence of brown (or beige) fat activity could therefore be another factor contributing to the lacking correlation of irisin with components of energy expenditure.

Fourth, parameters of physical activity were measured using the SenseWear™ armband device shown to be the best estimate of energy expenditure when comparing several monitors of physical activity in healthy volunteers under regular daily life

conditions (33). When compared with predictive equations (34), the SenseWear™ armband device and the doubly labeled water method have shown sufficient concordance for the assessment of resting and TEE in healthy volunteers and in overweight subjects with type 2 diabetes mellitus (35, 36). However, this comparison is still pending for subjects with reduced body weight. Therefore, the SenseWear™ armband device might have limitations for the use in anorexic patients. In addition, the measurement method of irisin could contribute to the lacking correlation of physical activity and circulating irisin. Erickson recently commented on the missing validity of the antibodies used to detect irisin in the existing studies since a quantitative western blot analysis has not been conducted yet and thus questioned the results on circulating irisin published so far (37). However, the antibody by Phoenix used in the present study seems to be the most suitable to date since it detects an amino acid sequence that is part of the cleaved

irisin protein (full cross-reactivity with amino acids 42–112 of the 112-amino acid polypeptide irisin, while no cross-reactivity was observed with the C-terminal parts of the irisin precursor FNDC5 amino acids 149–196 and 162–209, manufacturer's information). This contrasts with other antibodies that were raised against the transmembrane sequence of FNDC5 (1, 8) unlikely to occur in the circulation.

Lastly, a recent study suggested that the human FNDC5 gene differs from other species by a mutation in the start codon sequence (38) which may result in a lower translation efficiency (39) and consequently could explain difficulties to translate animal data to humans.

After investigating the whole cohort, we separated the study population according to the type of anorexia (purging type, restricting type, and atypical) since there is evidence for differences in excessive exercise in these subtypes of AN (19, 20, 40,

#### **Table 1 | Parameters of energy expenditure according to the subtype of anorexia nervosa**.

Data are expressed as mean ± SEM, p > 0.05. AN, anorexia nervosa; EAT, exercise activity thermogenesis; MET, metabolic equivalent; NEAT, non-exercise activity thermogenesis; REE, resting energy expenditure; TEE, total energy expenditure.

AN, anorexia nervosa; EAT, exercise activity thermogenesis; MET, metabolic equivalent; NEAT, non-exercise activity thermogenesis; REE, resting energy expenditure; TEE, total energy expenditure. Values for r and p are indicated in each row, \*p < 0.05.

41). However, irisin levels and all other parameters of energy expenditure assessed did not differ between these groups. Similarly to the whole sample, in the subgroups purging and atypical AN, no correlation was observed between irisin levels and any parameter of energy expenditure. However, when investigating the patients with restricting type AN separately, irisin levels negatively correlated with the number of steps per day and body weight-adjusted EAT which may indicate that this subgroup is a more homogeneous population of patients. This negative association is unexpected and in contrast to the reported positive association of exercise and circulating irisin for normal and overweight subjects (1, 6, 28). The present finding may point toward a differential regulation of irisin in patients with restricting type AN (*n* = 20) but should be confirmed in a larger cohort of patients.

In summary, irisin plasma levels do not differ in anorexic patients with moderate activity from those with high locomotor activity. Moreover, no association was observed for plasma irisin levels with various parameters of energy expenditure assessed under unrestricted conditions of physical activity in hospitalized patients. Therefore, the current data do not support the concept of irisin being induced by exercise, at least not under conditions of severely reduced body weight like AN. Thus, it is also unlikely that irisin contributes to exceeding energy expenditure in highly active anorexic patients. Further studies are needed to clarify whether the lacking correlation of irisin and physical activity described here is due to a missing association of irisin and exercise under conditions of reduced body weight *per se* or because of a unique characteristic of anorexic patients.

# **AUTHOR CONTRIBUTIONS**

Tobias Hofmann, Ulf Elbelt, and Andreas Stengel designed the study; Tobias Hofmann and Andreas Stengel coordinated and supervised the data collection, carried out the statistical analyses and drafted the manuscript. Ulf Elbelt, Anne Ahnis, Peter Kobelt, and Matthias Rose discussed the data and reviewed the article. All authors discussed the results, reviewed, and finalized the manuscript.

## **ACKNOWLEDGMENTS**

We would like to thank Reinhard Lommel and Petra Buße for their laboratory work; Karin Johansson and Christina Hentzschel for their help with blood withdrawals; Leoni Gmöhling and Heike Pietz for their help with SenseWear™ armband data collection and Friederice Schröder and Florian Bruckbauer for maintaining the database. **Funding**: This work was supported by the German Research Foundation STE 1765/3-1 (Andreas Stengel) and Charité University Funding UFF 89-441-176 (Andreas Stengel, Tobias Hofmann).

# **REFERENCES**

**Conflict of Interest Statement:** The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

*Received: 31 October 2013; accepted: 18 December 2013; published online: 06 January 2014.*

*Citation: Hofmann T, Elbelt U, Ahnis A, Kobelt P, Rose M and Stengel A (2014) Irisin levels are not affected by physical activity in patients with anorexia nervosa. Front. Endocrinol. 4:202. doi: 10.3389/fendo.2013.00202*

*This article was submitted to Neuroendocrine Science, a section of the journal Frontiers in Endocrinology.*

*Copyright © 2014 Hofmann, Elbelt , Ahnis, Kobelt , Rose and Stengel. This is an openaccess article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.*

# Personality and psychiatric disorders in women affected by polycystic ovary syndrome

#### **Elisabetta Scaruffi1 , Alessandra Gambineri 2 , Stefania Cattaneo3 , JenniTurra4 , Roberto Vettor 4 and Roberto Mioni 4\***

1 Edo ed Elvo Tempia Valenta Foundation, Biella, Italy

4 Clinical Medical 3, Department of Medicine, Hospital of Padua, University of Padua, Padua, Italy

#### **Edited by:**

Gianluca Tamagno, St. Columcille's Hospital, Ireland

#### **Reviewed by:**

Ruijin Shao, Gothenburg University, Sweden Stanko S. Stojilkovic, National Institutes of Health, USA

# **\*Correspondence:**

Roberto Mioni, Clinica Medica 3, Università di Padova, 9th floor Monoblocco, Azienda Ospedaliera di Padova Via Giustiniani 2, Padova 35128, Italy e-mail<EMAIL_ADDRESS>
**Background:** Polycystic ovary syndrome (PCOS) is the most prevalent endocrine disorder among fertile women. Studies show reduced quality of life, anxiety, depression, body dissatisfaction, eating disorder, and sexual dysfunction, but the etiology of these disturbs remains still debated. The aim of our study is to verify whether this hyperandrogenic syndrome characterizes a strong psycho(patho)logical personality.

**Method:** Sixty PCOS subjects (mean age 25.8 ± 4.7 years) were evaluated by anthropometric, metabolic, hormonal, clinical, and psychological parameters. After the certainty of the diagnosis of PCOS, the Rorschach test, according to Exner's comprehensive system (CS) and the Millon Clinical Multiaxial Inventory-III (MCMI-III) were administered to each patient. The control group, on which the comparison was carried out, was composed by 40 healthy and aged compared women who were exclusively administered the Rorschach test according to CS.

**Results:** MCMI-III evidenced axis II DSM-IV personality disorders [4.1% schizoid, depressive, sadistic, negativistic (passive–aggressive), and masochistic, 6.1% avoiding, 12.2% dependent, 20.4% histrionic, 16.3% narcissistic, 2.0% obsessive–compulsive], and axis I DSM-IV psychiatric disorders: 10.2% anxiety, 2.0% somatoform disorder and bipolar disorder, 16.3% major depressive disorder. Finally, we found 44.9% delusional disorder and 4.1% thought disorder. Rorschach test's results show 53.1% reduced coping abilities and social skills, 55.1% depression, 30.6% perceptual distortion and cognitive slippage, 24.5% constantly alert and worry, 8.1% at risk for suicide, and finally about 50% of our patients had chronic stress.

**Conclusion:** PCOS women have relevant personality and psychiatric disorders, when compared with normal subjects.

**Keywords: polycystic ovary syndrome, personality, Rorschach, MCMI-III, psychopathology, personality disorder, psychiatric disorder**

# **INTRODUCTION**

Polycystic ovary syndrome (PCOS) is a heterogeneous gynecological endocrine disorder, characterized by chronic anovulation, hyperandrogenism, and hyperinsulinism with insulin resistance, which affects about 6–10% of women in reproductive age (1–3) and profoundly affects the quality of life of these subjects (4, 5). In fact, this hyperandrogenic syndrome is frequently characterized by hirsutism, acne, alopecia, obesity, and irregular periods with infertility and it is often treated as cosmetic problem. The existence of a linkage between these latter clinical features and reduced quality of life was frequently suggested in several researches (6–8). Other studies evidenced how physical symptoms caused mental disorders, but depression and anxiety seem to be independent of obesity and infertility, the most frequent symptoms of PCOS (9, 10). Also, the relationship between androgen excess and mood

remains still controversial because clinical and biochemical parameters of hyperandrogenism seem not directly cause of depression (11). Again, diagnosis of PCOS was related to negative feelings, such as frustration (67%), anxiety (16%), and sadness (10%), although the (self-reported) level of knowledge about the illness was high and the women were aware that PCOS can be treated and is not a deadly disease (12). Other authors reported reduced scores in the sub-categories of perception about health in general, physical performances, general behavior, and limitations in family activities due to the "girls' disease" (12). At the same time, higher scores were reported in the rating of health conditions, during the diagnostic pathway and above all the treatment period, further supporting a dependent effect between clinical features and psychological aspects. In different manner, several data obtained in PCOS subjects reported that differences for the QoL

2 Division of Endocrinology, Department of Internal Medicine, S. Orsola-Malpighi Hospital, University of Bologna, Bologna, Italy

3 Scuola Universitaria Interfacoltà in Scienze Motorie (SUISM), University of Turin, Turin, Italy

are more related to the subjective perception of the illness's seriousness rather than to the clinical seriousness of the illness itself. These data, therefore, suggest that a subjective feeling of discomfort might be totally unrelated to the clinical picture, although objectively recorded. Again, other psychological aspects, such as depression (8, 13), and/or anxiety (14, 15), have been emphasized in PCOS, but only few articles analyzed women's personality structure (16). In women with PCOS, Sahingoz et al. (17) assessed a high prevalence of psychiatric and personality disorders, about 29.0 and 14.0%, respectively. They also reported 9.6% of mood and 26.0% of anxiety disorders, 13.7% of social phobia, 11.0% of generalized anxiety, and 12.3% of avoidant personality disorders, according to DSM-IV axis I and II Diagnostic Criteria, respectively. Again, Patients with borderline personality disorders, diagnosed by SCID for DSM-IV, about 30.0% was affected by PCOS (18). Personality disorders seem to be characterized by the tendency to exhibit a tenuous stability, or lack of resilience, under conditions of stress, by the inflexibility to adapt and by vicious cycles that repeats once again their pathology. Again, the interaction of psychosocial stressors and personality characteristics leads to the expression of psychological symptoms, that is, axis II and axis IV interact to produce axis I. Since no studies used Rorschach test or Millon Clinical Multiaxial Inventory to analyze the personality structure of subjects with PCOS, we believe it is useful to carefully study and develop the personality aspects related to this hyperandrogenic syndrome, as well as the psychopathological ones. Moreover, to minimize the influence of diagnostic and therapeutic pathways, we only studied young women, with comparable social and school conditions, undergoing for the first time to the endocrine evaluation for clinical features of hyperandrogenism and/or disorders of menstrual cycle.

This study was performed in collaboration with the Division of Endocrinology, Department of Internal Medicine of St. Orsola-Malpighi Hospital, University of Bologna, and with the Clinical Medical 3 of Health Centre/Hospital-University of Padua, on the white patients coming from the northern provinces of Italy.

# **PATIENTS AND METHODS**

#### **SUBJECTS RECRUITMENT**

Sixty Caucasian women, all of Italian origin, affected by certain diagnosis of PCOS, were enrolled to participate in this research study: 30 subjects from the Endocrinology Unit of St. Orsola-Malpighi Hospital – University of Bologna and 30 subjects from the Clinical Medical 3 of Health Centre/Hospital-University of Padua. For all patients, it was the first time they turned to a Public Health Institution to resolve their hyperandrogenic symptoms.

All women were hyperandrogenic (clinical presence of hirsutism with Ferriman–Gallwey score >8, acne or alopecia and/or elevated androgen levels: testosterone (*T*) > 2.1 nmol/L and or androstenedione (A) > 10.4 nmol/L or Free Androgen Index > 5 [FAI = Testosterone(nmol/L)/SHBG(nmol/L) × 100] and met the diagnosis of PCOS, according to the Rotterdam criteria (19). Ovarian ultrasound findings were performed by transvaginal or transrectal pelvic ultrasound (US ESAOTE F.U.5, Probe of 6.5 MHz, Milan, Italy) and PCO diagnosis were considered according to previous criteria (20). All subjects were no-smokers, no dieting, had a normal glucose tolerance test and performed normal physical

activities, none drank alcoholic beverages or took any continuous medications. According to PCOS diagnostic criteria, none had thyroid dysfunction, hyperprolactinemia, type 2 diabetes mellitus or concomitant cardiovascular, renal, and liver dysfunctions. Other causes of hyperandrogenism, such as Cushing syndrome/disease and congenital adrenal hyperplasia, were excluded, after nocturnal fasting, by basal and/or stimulated (0.25 mg Synacthen® iv, Biofutura Pharma S.p.A., Pomezia, Rome) plasma 17-OHprogesterone levels. At the beginning of the study, the insulin resistance and hyperinsulinism were, respectively, evaluated by using the homeostatic model assessment (HOMA-ir) [(fasting glucose (mmol/L) × fasting insulin (mIU/L)/22.5; n.v. <1.9 in our control group of at least 200 normal healthy subjects (data not shown)] and the 3-h oral glucose tolerance test (OGTT; 75 g of glucose) for plasma glucose, insulin, and C-peptide levels. Whole-body insulin sensitivity was calculated by using the insulin sensitivity index derived from the OGTT (ISICOMPOSITE n.v. >6.0) according to Matsuda and DeFronzo (21) considering normo-sensitive subjects with values higher than 6.0 and low-sensitive those with values lower than 4.0, respectively.

All women were studied in the follicular phase (days 1–7) of the cycle, or after 2 or more months of amenorrhea, after a negative pregnancy test. After this diagnostic pathway, but before of any pharmacological treatment, at each subject was administered the Millon Clinical Multiaxial Inventory-III (MCMI-III) (22). The MCMI-III has been adapted to the Italian population by a research group, which has involves the Universities of Milan-Bicocca, Rome-La Sapienza, and Padua, coordinated by Prof. Alessandro Zennaro, University of Turin, Italy (23, 24). We used Millon words to explain personality disorder and clinical syndrome.

In a subsequent day, for the first time in subject affected by PCOS, Rorschach, which is considered a scientific, valid, and reliable projective test to assess personality, was administered according to the Exner's comprehensive system (CS), once adapted to Italian population (24). The protocol was approved by the Local Ethics Committee of both Padua and Bologna Hospital, and all women gave their informed consent approval to collaborate with this research. Eleven women invalidated the Rorschach protocols because they have not given a sufficient number of answers in the first tables, so the original group of the patients was reduced to 49 patients. All subjects aged from 19 to a maximum of 39 years (25.80 ± 4.27 years), with comparable social and school degree (from 13 to 18 years of school). Forty five women, with age (from 19 to 41 years; 25.2 ± 6.0), social, and school degree comparable, were selected from a total of 2.500 records present in our database, and considered as control group. Each control subject had a normal menstrual cycle, was not hirsute, not taking any drugs or abusing alcohol, did not smoke, and was matched to the subjects with PCOS for body weight and age. Anonymity was assured to each patient and names were substituted by an alphanumerical progression.

#### **TESTS AND SELECTION OF VARIABLES**

The tests chosen for the personality analysis of the sample were (a) the *Rorschach Test* administered, scored, and interpreted according to the Exner's CS (23, 24); and (b) The MCMI-III (22) is composed by 175 true/false items with 28 scales of which 14 refer to personality disorders and 10 refer to clinical syndromes and the remaining 4 refer to test validity indices. In MCMI-III scoring system vary from 1 to 115 with a median score of 60 base rate (BR), scores of 85 or above evidence a personality disorder, according to DSM-IV axis II classification and/or the prominence of a clinical syndrome, according to DSM-IV axis I diagnosis (24, 25). Italian version of MCMI-III shows good internal validity (Cronbach's alpha range for subscales = 0.66–0.90) and test-retest reliability (*r* = 0.84–0.96) (23). The Rorschach tables were administered, scored, and interpreted according to the Exner's CS (24, 26, 27). It was decided to select, besides the 27 variables proposed by Exner and Andronikof-Sanglade (28) and by Bihlar and Carlsson (29), further variables that we consider useful FD, SumT, SumC0 , SumV, SumY, Sum Shading, Sum Color Shading Blends, Sum Shading Shading Blends, Wsum6, and An + Xy. Taking a cue from the work of Sultan et al. (30), conducted on patients with mellitus diabetes (IDDM), we wished to also consider part of their proposed indices.

### **LABORATORY AND PLASMA ASSAYS**

Glucose was measured by using the glucose oxidase method (Gluco-Quant®; Roche Diagnostics GmbH, Mannheim Germany), its detection limit was 0.11 mmol/L. Insulin by means of the two-site chemiluminescent immunometric immunoassay (Immulite 2000; Diagnostic Products Corporation, Los Angeles, CA, USA) and its analytic sensibility was 2.0µIU/mL. The intra- and inter-assay coefficients of variation (CVs) were <5.5 and 7.3%, respectively. LH, FSH, and E2 were measured by a competitive immunoassay with the use of an electrochemiluminescence immunoassay (ECLIA; Roche Diagnostics GmbH, Mannheim, Germany). Analytic sensibility of LH and FSH was 0.2 and 0.34 UI/L, respectively, whereas for E2 was 0.05 pmol/L. The intra- and inter-assay CVs were 1.8 and 5.2% for LH, 2.0 and 5.3% for FSH, 5.7 and 6.2% for E2, respectively. Testosterone, DHEA-s, 17-OHP, and A, after celite column chromatographic separation, were assayed as previously described (31). Serum T and DHEA-s were further measured by competitive immunometric chemiluminescent enzyme immunoassay (Immulite 2000; Diagnostic Products Corporation, Los Angeles, CA, USA). The detection limit was 0.35 nmol/L for T and 0.08µmol/L for DHEAs, respectively, the inter- and intra-assay CVs were below 7.4% for T and below 6% for DHEA-s. A and OHP were measured by using an enzyme-linked immunosorbent assay, ELISA (DRG Instruments GmbH, Marburg, Germany) and their detection limit was 0.6 nmol/L for A and 0.18 nmol/L for OHP, respectively, with intra- and inter-assay CVs <9.1 and 12.1% for A and 7.8 and 9.4% for OHP, respectively. SHBG was measured by using two-site chemiluminescent immunometric assay (Immulite 2000; Diagnostic Products Corporation, Los Angeles, CA, USA), its analytic sensibility was 0.8 nmol/L with intra- and inter-assay CVs <5.3 and 6.6%, respectively.

# **ANALYSIS OF THE DATA**

The anthropometric and hormonal data were reported as mean ± SD. All continuous variables were compared using the Student's *t*-test for unpaired data. Glycometabolic and hormonal data from PCOS and control groups were compared and analyzed with repeated measures by ANOVA. Mann–Whitney *U* test and Wilcoxon paired rank test were also used for evaluating the distribution of non-parametric values. Values for *p* < 0.05 were considered significantly different.

For an in-depth approach to the psychological investigation of our samples, we carried out the following analyses – comparison of the BR points obtained from the administration of MCMI-III under the original norms proposed by Millon et al. (22); statistical comparison of the average values of the Rorschach variables selected with the most recent norms proposed by Exner and Erldberg (26) through the use of confidence intervals (CI 95%); statistical percentage comparison of the threshold values of the same variables with those proposed by Exner (25), ambitent subject, non-patient norms. The reference to the descriptive statistics of the ambitent subjects is justified by the confirmation that, on average, this coping style is representative of the M: WSumC relationship present in the PCOS sample (*M* = 2.25: SumC = 4.133). Comparison of Rorschach Indices [suicide constellation (S-Con), coping deficit index (CDI), depression index (DEP-I), perceptualthinking index (PTI), schizophrenia index (SCZI), hypervigilance index (HVI)] obtained from subjects with PCOS were done with those obtained from control group recorded in our database.

# **RESULTS AND DISCUSSION**

Auxological and endocrine–metabolic parameters of our subjects are summarized in **Table 1**. As expected, hyperandrogenic parameters were elevated in women affected by PCOS and significantly different when compared with Control group. In particular, LH/FSH ratio and serum androgens, above all testosterone, were significantly higher in PCOS than in Controls. Also, metabolic parameters and insulin resistance were significantly elevated in patients with PCOS, whereas, as expected by the enrollment criteria, body weight (calculated as BMI) did not differ between the groups.

When we analyzed personality disorders scales, by MCMI-III, in PCOS group, we observed that several diseases, diagnosed in according to DSM-IV axis II, reached percentage significantly higher than in controls. In **Figure 1**, in fact, we summarized the

most important personality disorders in PCOS group compared with those observed in the control group. However, whether this increased incidence of personality disorders in patients affected by PCOS can be considered a link with clinical signs of hyperandrogenism remain at present unclear. In fact, the majority of studies showed a linkage between psychological disorders and major clinical PCOS symptoms, such as excessive body hairs, acne, infertility, or excessive body weight (4–6, 10, 13, 32, 33). Our results, in contrast, indicated that an intrinsic psychopathological characteristic of PCOS can be really consistent. According to our hypothesis, there are only three articles (17, 18, 34) that evidenced personality disorders in patients affected by PCOS, diagnosed by Diagnostic and Statistical Manual of Mental Disorders (DSM-IV). However, these latter studies recorded lower percentage of psychological disorders, when compared with our data, so suggesting that a heterogeneous expression of the same psychopathological diseases can exist in PCOS or it was assessed a heterogeneous pool of subjects affected by PCOS, therefore, further studies that consider the same Diagnostic Criteria for PCOS and a more numerous group

of person are needed. When we considered the results of clinical syndrome scales, diagnosed according to DSM-IV axis I Criteria, we also found several clinical psychopathological disorders, as summarized in **Figure 2**. The most representative disorders were anxiety (about 10.0%) and depression (about 16.0%). These analyses, in fact, seem to confirm recent data obtained in PCOS patients (17), which showed a frequent finding of psychiatric disorders. Interesting to note that in our sample we didn't observe neither alcohol addiction nor drug addiction or post-traumatic stress disorders. Because, we also observed different incidences (from 30.0% to about 70%) of psychopathological diseases, such as anxiety, mood, or thought disorders, using different psychometric test, it can be also suggested that different clinical expressions of psychopathological diseases can exist in PCOS. Also, other data, analyzed by different questionnaires, confirmed a prevalent expression of anxiety and depression, further suggesting that an elevated rate of mental illness in these hyperandrogenic patients (11, 14, 35). Again, Klipstein et al. (36) evidenced a rate of about 30.0% of bipolar disorder types I and II and Rassi (37) of about

11.0% of patients, respectively. Our data are in agreement with these latter results, so further evidencing that women with PCOS had a rate of mental illness several times higher (three to fivefold), when compared with normal population. However, the absence of rigorous diagnostic criteria of PCOS can not exclude an enrollment of heterogeneous groups of patients, so reducing the diagnostic capacity and, of consequent, the percentage of psychopathological diseases in the PCOS *per se*. Results of Rorschach test, obtained from our patients, are summarized in **Table 2** and **Figure 3**. We used as control a group of 45 healthy Italian subjects already recorded in our database and gently provided by Zennaro (23). We used this trick because in Italy, at present, there are no any data about this pathology.

Using a statistical comparison (Student's*t*-test for independent groups) between the samples of patients and the control group, we have evidenced significant differences among several variables, as showed in **Table 2**. Calculating Cohen's *d*, the effect size (statistically significant >0.6) underlined a high effect for 10 of these variables: WSum6, AdjD, ego impairment index (EII), MOR, Popular, Shading Blends, SumT, SumV, Sum Shading e Sum6. Our data show an important deficit in cognitive functions (MEDIA-TION and IDEATION clusters). Cohen's *d* underlines as "high effects" (≥0.6) of the following variables: X + %, WSum6, AdjD, COP, GHR, Popular, SumV, SumY, Sum Shading, and Sum6. The following variables, WSum6, SumV, Sum Shading, Sum6, are significantly higher. The variables, X + %, COP, GHR, Popular, were significantly lower, while AdjD was actually negative. All these data evidenced that in subjects with PCOS exists a massive reality distortion (XA%, X − %, X + %, EII, PTI index, and SCZI index) with incoherent and disorganized thoughts (Sum6 and Wsum6), failures in the ability to judge, bizarre, or magical ideations and with interpersonal communication difficulties. PTI was positive for 30.6% of patients as well as SCZI for 20.4% of our sample, therefore, we can underline as "*daily life*" is really difficult for the majority of our patients affected by PCOS, because misperception leads to behavior often inappropriate in social life. Again, a large proportion (63.3%) of patients (X − % > 0.25) were displayed serious perceptual inaccuracies, stressing therefore how the reality was not adequately perceived because emotions

distorted it (X − %, X + %). Again, 42.9% of subjects (Xu% = 0.2) were anti-conformism, displaying a tendency to originality, but 53.1% (Populars ≤ 4) rejected conventional models, so adopting behaviors not socially adequate and/or accepted. Ninety-two percent of subjects with PCOS (X + % <0.70) showed a perception of the situations and words in an unusual manner. About four (4.1) % (Populars ≥ 8) of our sample was exaggerated in conformism. These latter patients usually need a positive social judgment and they need the approval of other people. In our sample, 26.5% (lambda < 0.30) had an inadequate control over emotions that interfere with logical functions. EII showed serious impairment in 38.8% of the sample, moderate in 18.4%, average in 8.2%, and slight in 24.5% (X − %, Sum6, WSum6 and EII higher, XA%, WDA%, X + % lower). Only 10.0% of women analyzed, had a normal, non-pathological values. According all these results, we evidenced that our patients had really difficulties in organizing the stimulus when they face the reality, so forcing them to use more psychic energy than needed (WDA < XA%). However, in our study about 12.2% of patients (12 subjects) ignored the complexity of the reality and tended to simplistic situations and interpersonal relationships. Again, 40.8% were very self-involved (Egoindex ≥ 0.45) and about 30% built up their own self-esteem, by considering themselves better than others (Fr + Rf = 1). This focus on self could be detrimental in the interpersonal relationship and could contribute to reality distortion. Our data seem to confirm Willmott's results (38) that evidenced as women with PCOS exhibit a lack of control over their own lives' events. Considering the CDI, we found it positive for 53.1% of the patients (vs. 29% of the controls), revealing a deficit in coping abilities, and consequently interpersonal relationships were superficial and rarely maintained. Also, social skills appeared to be reduced (CDI, EA) and interpersonal relationships were perceived as unsatisfying. About thirty (30.6) % of PCOS sample had a damaged and negative interpersonal relationship representation (PHR higher). Interactions of personal situations were perceived as not cooperative, but in a hostile manner. All of this creates, in our patients, a state of chronic stress, which was reflected in a cognitive slippage. More than 50% of our patients resulted depressed (DEP-I positive) and with a percentage significantly higher than observed in controls (55.1 vs. 25.0% controls). Unexpectedly, 8.1% of patients with PCOS (vs. 6% of the controls) were at risk for suicide (S-Con positive, MOR = 2, FD > 2, Shadings). These latter results were in agreement with previous studies that underlined how this psychopathological tendency was more frequently observed in women with PCOS (35). However, at present, whether suicide can be cause or effect of high level of discomfort induced by clinical or psychological signs, or both, remains still unknown. In expected manner, several authors (39, 40) reported a high prevalence of negative self image and poor body image in subjects with PCOS. Also, in our PCOS group, with respect of controls, we evidenced a significant higher, about 18.0%, a self image damaged (MOR = 2). Again, about 45% of patients had an exaggerated introspective behavior (FD ≥ 2) that it could became a pathological self criticism and a ruminative thinking. This point of view is important, because women with PCOS had a negative and pessimistic view of themselves; therefore, they had constant need of reassurance about their own value

### **Table 2 | Comparison between PCOS and control groups using t-test for independent samples**.

and had serious problems of establishing intimate relationship (18.4% Fr + Rf ≥ 2). In our patients, we recorded about 25% of the samples (vs. 7% of healthy and normal subjects) constantly in alert (HVI) and consequently worried, therefore, this mood justifies how the majority part of the subjects examined, about 64% of PCOS, lived with a constant state of anxiety (SumY ≥ 2). It was interesting to underline that similar results were obtained both in depressed patients and in subjects with juvenile diabetes

mellitus type I (30). And again, it was suggested that the observation of Monzani et al. (41) who emphasized how in subjects affected by PCOS scores of somatization, anxiety, and depression variables reached higher values when compared with normal people. At the same time, these authors interestingly did not observe any relevant psychopathological disorders, but differently noticed that the subjects tend to hidden and/or clear behaviors typical of the opposite gender. Moreover, the same women tended to conform themselves to social standards, so probably reducing the perception of their "diversity."

About 60% of PCOS women experienced painful emotions and high sense of insecurity, confusion, and ambivalence in relation to emotions (Color Shading Blends > 1 in 69.4% and Shading Blends > 1 in 61.2%), showed a low self-esteem (36.7% with Egoindex < 0.32), and feel anxious, tense, nervous, and irritable (more than 71.0% showed FQS-higher). All these data evidenced that these women did not tolerate frustration and they were impulsive (D < 0). About 70.0% was feared to affective stimuli and avoid them (Afr lower) and experienced discomfort with introspection, because feelings of inferiority and depressed feelings (SumV ≥ 1). We observed that many of the samples (69.4%) did not use intellectualization (Intell > 3) as a defense mechanism against affects, but tended to avoid (Afr) situations where they were generated. Since several patients affected by PCOS tended to isolate themselves, to refuse interpersonal relationships, to present social phobia, or tended to be worried and keep at a certain emotional distance from others, it can be concluded that these subject are not very interested in interpersonal relationship. However, our results, in different manner, evidenced that about 60% of patients tended to actively research contact and interpersonal relationships (Isoindex < 0.25). Because, we studied a group of young patients, with

the majority of them in a period of university study, where the interpersonal contacts are very high, we cannot exclude that we have enrolled a group of patients with different social or cultural impact.

# **CONCLUSION**

Our results clearly highlight that in our patients, affected by PCOS, exist mood and severe thought disorders with perceptual distortion in about half of subjects. In this hyperandrogenic syndrome, women have elevated dysphoric feelings, chronic emotional stress, and several difficulties in social skills and daily life. In our group of patients, to attenuate an important painful emotional overload, many avoid interpersonal relationships and emotional inputs. Again, intimate relationships can often cause fear and frustration. All these conditions suggest that the characteristics of psychological suffering in PCOS can constitute a distinguishing element, which, for some aspects, appears to be so represented and shared by the sample as to be promoted to a proper diagnostic indicator. We also underline that the knowledge of these psychopathologic aspects may ameliorate the doctor–patient interaction and relationship with a desirable improvement on the medical examination, sensibility, and probably reaching greater adherence to diagnostic and medical therapy.

### **ACKNOWLEDGMENTS**

We thank the Financial Management Committee of SINAPSI (the regional intermediary agency for the administration and management of scholarships and grants) that alongside the Piedmont Regional Government, the Italian Ministry of Labour and Social Politics and the European Union funded our research. We also wish to thank Dr. Antonella Vaccina from St. Orsola-Malpighi Hospital in Bologna, Italy, for her precious collaboration, and Miss Roberta Maoret from the Medical Library AIL (Italian Association against Leukemia) of the Clelio Angelino Onlus Foundation in Biella, Italy, for her kind help in providing the scientific articles. This research was partially supported by a Grant (n. A00000D150 SIC DE PPR/08) of the University of Padua to Prof. Roberto Vettor.

#### **REFERENCES**

**Conflict of Interest Statement:** The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

*Received: 14 June 2014; accepted: 12 October 2014; published online: 12 November 2014.*

*Citation: Scaruffi E, Gambineri A, Cattaneo S, Turra J, Vettor R and Mioni R (2014) Personality and psychiatric disorders in women affected by polycystic ovary syndrome. Front. Endocrinol. 5:185. doi: 10.3389/fendo.2014.00185*

*This article was submitted to Neuroendocrine Science, a section of the journal Frontiers in Endocrinology.*

*Copyright © 2014 Scaruffi, Gambineri, Cattaneo, Turra, Vettor and Mioni. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.*

# Gender dysphoria – prevalence and co-morbidities in an Irish adult population

#### **Ciaran Judge1† , Claire O'Donovan1† , Grainne Callaghan1 , Gadintshware Gaoatswe1 and Donal O'Shea1,2\***

1 Department of Endocrinology, St. Columcille's Hospital, Dublin, Ireland

2 Department of Endocrinology, St. Vincent's University Hospital, Dublin, Ireland

#### **Edited by:**

Jacques Epelbaum, Université Paris Descartes, France

#### **Reviewed by:**

Eugenio De Carlo, University School of Medicine, Italy Maria-Cristina Burlacu, Cliniques Universitaires St-Luc, Belgium

# **\*Correspondence:**

Donal O'Shea, Department of Endocrinology, St. Columcille's Hospital, Loughlinstown and St Vincent's University Hospital, Elm Park, Dublin 4, Ireland e-mail<EMAIL_ADDRESS>
†Ciaran Judge and Claire O'Donovan have contributed equally to this work. **Introduction:** Gender dysphoria (GD) is a condition in which there is a marked incongruence between an individual's psychological perception of his/her sex and their biological phenotype. Gender identity disorder was officially renamed "gender dysphoria" in the DSM-V in 2013. The prevalence and demographics of GD vary according to geographical location and has not been well-documented in Ireland.

**Methods:**We retrospectively reviewed medical records of 218 patients with suspected or confirmed GD referred to our endocrine service for consideration of hormonal therapy (HT) between 2005 and early 2014.We documented their demographics, clinical characteristics, and treatment during the study period.

**Results:** The prevalence of GD in the Irish population was 1:10,154 male-to-female (MTF) and 1:27,668 female-to-male (FTM), similar to reported figures in Western Europe. 159 of the patients were MTF and 59 were FTM, accounting for 72.9% and 27.1% of the cohort, respectively. The rate of referral has increased year-on-year, with 55 patients referred in 2013 versus 6 in 2005. Mean ages were 32.6 years (MTF) and 32.2 years (FTM). 22 of the patients were married and 41 had children, with 2 others having pregnant partners. 37.6% were referred by a psychologist, with the remainder evenly divided between GPs and psychiatric services. There were low rates of coexistent medical illness although psychiatric conditions were more prevalent, depression being a factor in 34.4% of patients. 5.9% of patients did not attend a mental health professional. 74.3% are currently on HT, and 9.17% have had gender reassignment surgery (GRS). Regret following hormonal or surgical treatment was in line with other Western European countries (1.83%).

**Conclusion:**The incidence of diagnosis and referral of GD in Ireland is increasing.This brings with it multiple social, health, and financial implications. Clear and accessible treatment pathways supported by mental health professionals is essential.

**Keywords: gender dysphoria, gender identity disorder, transgender, gender reassignment surgery, gonadectomy, hormone therapy**

# **INTRODUCTION**

Gender dysphoria (GD) is a recognized condition in which there is marked incongruence between an individual's psychological perception of his/her sex and their biological phenotype (1). GD has replaced the term gender identity disorder (GID) in the 2013 publication of the DSM-V, which itself replaced "transexualism" in 1980. The cause is not well understood and is believed to be due to a complex interaction of psychosocial and biological factors during development (2, 3).

Prevalence varies based on geographical location, with higher rates in Western Europe and America (0.001–0.002%) (4–6) compared to lower rates in Japan (0.0009%) (7). In most studies, male-to-female (MTF) GD cases tend to significantly outnumber female-to-male (FTM) cases (7).

The DSM-V has strict criteria for the diagnosis of GD that differ between children and adults. However, both include a persistent and strong conviction that they are the wrong sex, feel discomfort, distress, or anxiety with their gender-role and sexual characteristics, and possess a strong and persistent desire to be a member of the opposite sex (1). These criteria are based on the assumption that GD is a purely psychiatric condition which is a topic of controversy.

The complex nature of GD requires that psychological, social, and medical issues are addressed by a multidisciplinary team.

The Department of Endocrinology in St. Columcille's Hospital (SCH), Dublin, has been managing patients with GD since 2000. Patients are referred for consideration of hormonal therapy (HT) by a mental health professional or GP in the majority of cases. We have previously reported the characteristics of the first 52 patients referred for HT in SCH (8), however, GD in Ireland remains poorly investigated. This paper aims to update the characteristics of GD in Ireland, show the current prevalence of the condition, and discuss the need for developing accessible, well-defined pathways of care.

# **MATERIALS AND METHODS**

We retrospectively reviewed the medical records of 218 patients referred to the GD clinic between 2005 and early 2014. A diagnosis had been made by a MHP based on DSM-IV/V criteria in most instances, however, some patients required a referral from the clinic to a mental health professional if this had not occurred This initial consultation included a full medical and surgical history and examination as well as baseline laboratory investigations; full blood count, renal, liver, and bone profiles, fasting lipids, and glucose as well as hormonal profiling.

Data collection focused on documentation of past medical and surgical history (including hypertension, coronary artery disease, thromboembolic disease, dyslipidemia, diabetes mellitus, etc.), psychiatric background (including depression and autistic spectrum disorder etc.), and family history (including cardiovascular conditions etc.). Smoking and alcohol use were also recorded. GDspecific questioning aimed to establish age at self-diagnosis, time spent in desired role, and hormonal or surgical therapy prior to presentation, etc (**Table 1**).

Consideration for HT required that patients were diagnosed by a minimum of one, preferably two mental health professionals.

In our service, we aimed to initiate HT at the second visit, thus this necessitated that a detailed discussion with the patient regarding the risks and benefits of HT be undertaken at the initial consultation. Providing there were no contraindications and the patient received a definitive diagnosis of GD from an appropriate mental health specialist, HT could commence on the second consultation. We also analyzed the incidence of regret or adverse effects to treatment in those presenting for follow-up. Those wishing to proceed with gender-related surgery were referred to appropriate specialists in Ireland or abroad.

# **RESULTS**

# **DEMOGRAPHICS**

In the study, 218 patients with GD were referred to our clinic between 2005 and early 2014 (**Table 1**). 72.9% of these were MTF and 27.1% FTM, with a MTF:FTM ratio of 2.7:1. Based on the 2011 census reports (9), this indicates a prevalence of GD in 1:14,756 people (0.0067% of the population), categorized into 1:10,154 MTF (0.0098%) and 1:27,668 FTM (0.0036%). Seventy-one percent of the patients were Irish, with Leinster providing the most patients when subdividing the cohort based on region. The numbers of patients being referred increased steadily over the 9 years study period, from 6 referrals in 2005 to 55 in 2013 (**Table 2**; **Figure 1**).

The average age at referral was similar between both gender groups at 32.6 years for MTF and 32.2 years for FTM, ranging from 15 to 69 years. There was a clear trend in the study showing people are being referred at younger ages as time goes on. The average age at referral from 2005 to 2009 inclusive was 36.77 years, compared with the average age between 2010 and 2013, which was 30.14 years. Twenty-one percent of the patients were married at presentation or in the past, and 18.8% reported having children. Two patients reported having pregnant partners, one of whom conceived using banked sperm.

One hundred thirty-five patients (61.9%) declared their age at self-diagnosis of GD as being pre-pubertal, with 14.7% stating

#### **Table 1 | Demographics and co-morbidities**.

SD, standard deviation; MTF, male-to-female; FTM, female-to-male.

#### **Table 2 | Number of referrals per year**.

MTF, male-to-female; FTM, female-to-male.

they realized their issues around gender as adolescents. 50.9% of patients were living in their desired gender-role on a full-time basis, with approximately one quarter of patients not living in role prior to initial presentation.

The majority of referrals were made by a psychologist (37.6%), with the remainder evenly divided between psychiatrists and GPs. Although most patients were seen by at least one MHP, 5.96% of patients did not attend either a psychiatrist or psychologist. Of these, five patients received HT without prior MHP consultation, however, one patient obtained the medications via the internet and three others were on HT and also had genderrelated surgery prior to presentation. Forty-four patients (20.2%) were on HT prior to initial visit to the GD clinic, 9 of whom ordered it via the internet, 16 individuals sourced hormones from practitioners either in Ireland or the UK, 9 sourced it from overseas, and in the case of the remainder the source was unknown.

### **MEDICAL AND PSYCHIATRIC CO-MORBIDITIES**

Several patients had co-existing medical co-morbidities, with dyslipidemia (21.6%), asthma (11.45%), and hypertension (9.17%) being the most prevalent. Twenty-six percent of the patients reported being smokers at the time of presentation, with 23.85% having smoked regularly in the past. Twelve patients consumed more than 15 units of alcohol per week, however, eight patients (3.67%) reported abusing alcohol to a damaging degree in the past. Seventy-five (34.4%) patients had a history of depression, 25.3% of those admitting to acts of deliberate self-harm. Other psychiatric conditions encountered included schizophrenia (3.67%), bipolar disorder (2.29%), and Asperger's syndrome (2.29%) (**Table 1**).

Blood samples were taken as they are important to both assess initial risk and evaluate possible future adverse events, including cardiovascular disease (10). Twenty-five percent were found to have an initial fasting cholesterol of >5.3 mmol/L and 31% had a fasting LDL of >2.6 mmol/L, both of these figures being the upper reference range for the laboratory in SCH. Of these, 47 were known to have dyslipidemia (42 MTF versus 5 FTM), 12 people went on to commence lipid-lowering therapy, with others initiating diet control or having a repeat sample within normal ranges. Five people were found to have a fasting blood glucose of >5.9 mmol/L, however, none were found to have diabetes mellitus on repeat testing (**Table 1**). Nearly 20% of patients had a family history of vascular disease including myocardial infarction (9%), deep venous thrombosis (2%), and cerebrovascular disease (9%).

### **POST PRESENTATION**

To date, 55 patients (22.3%) have undergone surgery, 36 (65.5%) of these have been MTF patients. Mammoplasty was the most common procedure, with 19 (52.8%) of those undergoing surgery electing to have breast augmentation. Twenty (36.4%) surgical patients opted for gender reassignment surgery (GRS), which

#### **Table 3 | Surgical procedures in gender dysphoria patients**.

GRS, gender reassignment surgery.

constitutes 9.17% of the total cohort. A very small number also had facial or laryngeal surgery for voice alteration (**Table 3**). As there are currently no GRS services in Ireland, patients are referred to a variety of centers in the United Kingdom and elsewhere. Additionally, there are currently 11 patients referred to Charing Cross Hospital for consideration of GRS.

Dual energy X-ray absorptiometry (DEXA) scanning is now part of the standard screening protocol of patients presenting to our clinic. In our cohort, 39 patients (17.9%) underwent this investigation at time of last known follow-up, with 20.5% of those studied discovered to have osteopenia, and 7.7% diagnosed with osteoporosis.

Thirty-seven patients (17%) reported adverse effects following hormonal or surgical therapy. Eight patients reported decreased mood or libido, six patients were found to have developed dyslipidemia, five patients had hot flushes/headaches, and one patient developed a fistula post GRS. Four patients (1.8%) expressed regret following treatment. All of the incidences of regret involved MTF, three of who were post GRS with the other in the hormonal phase of treatment.

# **DISCUSSION**

In this retrospective study, we characterized GD patients attending an endocrine unit in the Republic of Ireland. From this study, the prevalence of GD is 1:10,154 MTF (0.0098%) and 1:27,668 FTM (0.0036%). This is comparable to the prevalence of GD reported in Western Europe, which ranges from 1:11,900 to 1:18,000 for MTF and 1:30,400 to 1:54,000 for FTM (4, 10–12). Our analysis echoed similar international studies in that we collated data from a single specialist GD clinic. De Cuypere et al. reviewed 10 such studies spanning 39 years with prevalence very much matching our own (10). The prevalence rate of GD in our cohort is likely to reflect that of the Republic of Ireland as a nation, as our unit has been the main specialist center in the country over the study period.

In the seventh version of the World Professional Association for Transgender Health (WPATH) Standards of Care for the Health of Transsexual, Transgender, and Gender-Non-conforming People (13), it is highlighted that in more recent analyses there appears to be a higher prevalence of GD, possibly reflecting an increase in the number of people seeking help from healthcare professionals (13– 15). This hypothesis is supported by our analysis, which shows 6 referrals to our service in 2005 versus 55 in 2013.

Our MTF/FTM sex ratio of 2.7:1 is similar to the ratio of 3:1 reported in our previous study (8) and the sex ratio reported in Western Europe (10, 11). It is in line international data ranging from 3 to 5:1 (16).

The age of initial assessment was the same for both MTF and FTM (32 years). This is significantly different to our previous figures which indicated FTM patients presented, on average, 8 years younger than MTF (8). Age of presentation for MTF has decreased dramatically (39 versus 32.6) while that of FTM patients has remained approximately the same (31 versus 32.2) (8). The reason for this is unclear but may represent the condition now being more socially acceptable facilitating earlier presentation. This age is in line with a Belgian study, which showed a mean age of initial presentation of ~30 years (10). Over 60% of patients declared their age at self-diagnosis to be pre-pubertal which highlights the need for early recognition and diagnosis of the condition.

21% of the patients were married at presentation or in the past, and 18.8% reported having children. This is lower than the reported rates in similar European studies (10). Two of the patients had pregnant partners, one of whom conceived using banked sperm.

The rate of co-existing medical illness such as hypertension and diabetes were low in GD patients, which could perhaps be expected as the patients are generally young (average age 32 years). It is important to identify and optimize control of these medical co-morbidities prior to initiating the hormonal phase of treatment, as they can be exacerbated by hormones such as estrogen (17–19). Close attention should be paid to the possibility of cardiovascular events, as accruing evidence has suggested that MTF GD patients may be at an increased cardiovascular risk (20–22). Twenty-six percent of patients were smokers at initial presentation which is in line with the national average of 29% (23). Patients were informed that they would not be commenced on HT until they had ceased smoking due to the increased risk of venous thrombosis (24).

Depression was common to 34.4% of patients, while the incidence of other major psychiatric illnesses was relatively low. A survey of Dutch psychiatrists regarding psychiatric co-morbidity in those with GD found that 61% of their cohort had a co-existing psychiatric disorder (25). The presence of such a high rate of comorbid psychiatric illness underlines the need for a well-defined and rigidly adhered-to mental health consultation service. While our rate of patients who did not attend a MHP (5.96%) is low, it outlines the challenges faced by specialist centers in ensuring that patients with GD receive the appropriate mental health support in the setting of increased referrals.

Currently, there are no centers in Ireland which provide GRS, which may have contributed to only 11% of our patients undergoing sex reassignment surgery. However, it is worth noting that there is an agreement between the Irish Health Service Executive and certain GRS centers in other countries to facilitate ease of access for Irish patients wishing to undergo GRS. Unfortunately, this system is somewhat convoluted and frequently causes delays in access to surgery. This is a frustration experienced by many of the patients examined in our study.

The regret rate following both HT and GRS was low with four (2%) patients expressing regret (all MTF). This is slightly higher than previously reported figures of 1–1.5% for MTF (26, 27). Three of these had gender-related surgery and one had received hormones as the only treatment. Although low, the regret rate of 2% illustrates the need for continued input from mental health professionals beyond GRS. Unfortunately, these services are not always readily available, and this is a matter for further discussion.

# **CONCLUSION**

The prevalence of GD in the Republic of Ireland is comparable to that reported in Western Europe. However, the true prevalence may be difficult to assess due to patient worries regarding stigma and societal acceptance of the condition. The number of patients treated by our unit is increasing each year, which will pose considerable challenges with already limited resources. The liaison with mental health services is of critical importance to patients and to the success of treatment. Difficulties in obtaining ongoing mental health care are all too common and lead to the burden of care resting on the endocrinology service. It is vital that a well-defined and accessible treatment pathway is continually developed in order to ensure a gold standard of care for patients with GD.

### **ACKNOWLEDGMENTS**

Ciaran Judge and Claire O'Donovan contributed equally in data collection, writing, and reviewing the manuscript. The remaining authors assisted in data collection, writing, and reviewing the manuscript.

### **REFERENCES**

**Conflict of Interest Statement:** The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

*Received: 02 April 2014; accepted: 28 May 2014; published online: 13 June 2014. Citation: Judge C, O'Donovan C, Callaghan G, Gaoatswe G and O'Shea D (2014) Gender dysphoria – prevalence and co-morbidities in an Irish adult population. Front. Endocrinol. 5:87. doi: 10.3389/fendo.2014.00087*

*This article was submitted to Neuroendocrine Science, a section of the journal Frontiers in Endocrinology.*

*Copyright © 2014 Judge, O'Donovan, Callaghan, Gaoatswe and O'Shea. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.*

# Reproduction, smell, and neurodevelopmental disorders: genetic defects in different hypogonadotropic hypogonadal syndromes

#### **Hernan Valdes-Socin1\*, Matilde Rubio Almanza1 , Mariana Tomé Fernández-Ladreda1 , François Guillaume Debray 2 , Vincent Bours 2 and Albert Beckers 1**

1 Service of Endocrinology, CHU Liège, University of Liège, Liège, Belgium

2 Service of Human Genetics, CHU Liège, University of Liège, Liège, Belgium

#### **Edited by:**

Gianluca Tamagno, St Columcille's Hospital, Ireland

#### **Reviewed by:**

Alexandru Saveanu, Aix Marseille University, France Thomas King, Mater Misericordiae University Hospital, Ireland

#### **\*Correspondence:**

Hernan Valdes-Socin, Service of Endocrinology, Centre Hospitalier Universitaire, Rue de l'Hôpital 1, Liège 4000, Belgium e-mail<EMAIL_ADDRESS>The neuroendocrine control of reproduction in mammals is governed by a neural hypothalamic network of nearly 1500 gonadotropin-releasing hormone (GnRH) secreting neurons that modulate the activity of the reproductive axis across life. Congenital hypogonadotropic hypogonadism (HH) is a clinical syndrome that is characterized by partial or complete pubertal failure. HH may result from inadequate hypothalamic GnRH axis activation, or a failure of pituitary gonadotropin secretion/effects. In man, several genes that participate in olfactory and GnRH neuronal migration are thought to interact during the embryonic life. A growing number of mutations in different genes are responsible for congenital HH. Based on the presence or absence of olfaction dysfunction, HH is divided in two syndromes: HH with olfactory alterations [Kallmann syndrome (KS)] and idiopathic hypogonadotropic hypogonadism (IHH) with normal smell (normosmic IHH). KS is a heterogeneous disorder affecting 1 in 5000 males, with a three to fivefold of males over females. KS is associated with mutations in KAL1, FGFR1/FGF8, FGF17, IL17RD, PROK2/PROKR2, NELF, CHD7, HS6ST1, FLRT3, SPRY4, DUSP6, SEMA3A, NELF, and WDR11 genes that are related to defects in neuronal migration. These reproductive and olfactory deficits include a variable non-reproductive phenotype, including sensorineural deafness, coloboma, bimanual synkinesis, craniofacial abnormalities, and/or renal agenesis. Interestingly, defects in PROKR2, FGFR1, FGF8, CHD7, DUSP6, andWDR11 genes are also associated with normosmic IHH, whereas mutations in KISS1/KISSR, TAC3/TACR3, GNRH1/GNRHR, LEP/LEPR, HESX1, FSHB, and LHB are only present in patients with normosmic IHH. In this paper, we summarize the reproductive, neurodevelopmental, and genetic aspects of HH in human pathology.

**Keywords: reproduction, male, Kallman syndrome, hypogonadotropic hypogonadism, olfaction, kisspeptin, genetics**

# **INTRODUCTION**

Reproductive system development and control in mammals is dependent on specific neurons located in the hypothalamus that secrete gonadotropin-releasing hormone-1 (GnRH-1) and control the pituitary–gonadal axis (**Figure 1**). During embryogenesis, these neurons originate in the nasal placode and migrate into the forebrain along the olfactoryvomeronasal nerves (1–3). Alterations in this migratory process lead to defective GnRH-1 secretion, resulting in heterogeneous genetic disorders such as idiopathic hypogonadotropic hypogonadism (IHH), and other reproductive diseases characterized by the reduction in or failure of sexual maturation and competence. Another consequence of these migratory neuronal defects can be olfactory dysfunction. Depending of the affected genes, other neurological developmental disorders can also be encountered (1–4).

Thus, idiopathic hypogonadotropic hypogonadism (IHH) is a genetic disease that can occur with a normal sense of smell (normosmic IHH) or in association with anosmia (Kallmann syndrome; KS). To date, mutations in many genes have been described in relations to KS and/or normosmic IHH (nIHH) (**Tables 1** and **2**). Hypogonadotropic hypogonadism (HH) can also be found in association with other distinctive clinical syndromic conditions, such as Prader Willi syndrome, that are outside the scope of the current review.

In this review,wefocus on genetic central hypogonadism,which is more frequently encountered in males than in females. Congenital IHH is a clinically and genetically heterogeneous disorder (3, 4). Although sporadic cases predominate, families with congenital IHH have been reported with X-linked, autosomal dominant (AD) or autosomal recessive (AR) inheritance patterns (1–4). In some families, high variability in reproductive and non-reproductive phenotypic features suggests the presence of complex inheritance. In particular, polygenic (digenic or oligogenic) forms and variable forms of transmission can be found in selected cases (6–11). Indeed,further complexity is added by the remarkable observation

of reversibility of the phenotype in some cases of genetically determined hypogonadism (12–16).

 Hypogonadism is defined as the insufficient production of sex hormones with or without disturbed gametogenesis. HH results from a dysfunction of the hypothalamic–pituitary axis interfering with control of gonadotropin secretion (1–3, 16).

During life, the activity of the HPG axis has a tri-phasic pattern of "on-off-on." A first phase of activity occurs from the 16th week of intrauterine life as well as in the period between the 4th and 10th weeks of postnatal life (or "mini-puberty"). Mini-puberty is characterized by an increase in gonadotropin and steroid hormone secretion. Gonadotropins and sex hormones levels rise to a lesser extent than in true puberty. After mini-puberty, the HPG axis is repressed ("off") until puberty, when the system is reactivated ("on"). HPG axis activity is maintained throughout adult life in men whereas in women, menopause intervenes, and low sex steroids and compensatory high gonadotropin levels are characteristic (1–3).

The immediate postnatal period can be a window of opportunity for pediatricians and neonatologists to diagnose certain forms of HH. The congenital gonadotropin deficiency phenotype

**Table 1 | Genes and phenotype related only with normosmic IHH**.

is variable and depends on the gender, the magnitude of the deficit, and the specific genetic abnormalities (**Figure 1**). At the time of puberty, the diagnosis of HH may be suspected due to the absence in the onset of puberty and development of secondary sex characteristics in both sexes. In adulthood, gonadotropin deficiency can be suspected in a woman without breast development or who presents with primary amenorrhea. In adult men, gynecomastia, small testes (<14 mL), penile hypoplasia, and/or oligo-azoospermia raise the clinical suspicion of congenital hypogonadism (1–3).

#### **NORMOSMIC IDIOPATHIC HYPOGONADOTROPIC HYPOGONADISM**

The genetic abnormalities described below are infrequent or rare (see **Table 1**). In contrast to KS, patients with nIHH have a normal sense of smell and tend not to have other clinical signs. From a biological point of view, sex steroids secretion and gametogenesis are compromised, but to varying degrees. As it would be expected, reproductive phenotypes are more pronounced in subjects in whom the receptor is inactivated as compared to those harboring hormone inactivating mutations.

#### **GNRH-1 and GNRHR mutations**

Gonadotropin-releasing hormone (GnRH) is encoded by the *GNRH1* gene, which is located on chromosome 8p21-11.2. GNRH-1 mutations are rare and have been described in only two families (17, 18). A single homozygous mutation (c.18-19insA) affecting the peptide precursor preproGnRH was described in a Romanian family (17). It encoded a truncated and biologically inactive peptide in a male patient and his sister, both of whom had delayed puberty and normal sense of smell. The phenotype was reversed by pulsatile GnRH administration. Another homozygous *GNRH1* mutation was identified in a prepubertal boy from Armenia, with cryptorchidism and microphallus (18).

The *GNRHR* gene (locus on chromosome 4q13.2-3) encodes for the GNRH receptor. There is some variability in clinical expression of *GnRHR* mutations that is due to a partial loss of function. *GNRHR* mutations have been described in about 40– 50% of familial AR nIHH cases, and in around 17% of sporadic nIHH (1–3).

# **KISS1 and GFPR54 mutations**

The gene *KISS1* was described originally as a metastasis suppressor gene but it is a key gene in reproduction. It is localized on chromosome 1q32, encoding a protein called kisspeptin, which is, in turn, processed in four peptides Kp10, Kp13, Kp14, and Kp54. Kisspeptins stimulate GnRH neuronal firing and GnRH secretion, which then triggers an increased release of LH and FSH (**Figure 1**). The *KISS1R* gene (locus 19p.13.3), a G-proteincoupled receptor, is also known as the *GPR54* gene, and it is the receptor for kisspeptins. *GPR54* mutations can be compound heterozygous or homozygous (19–22). Loss of function mutations in *KISS1* (20) and *GPR54* cause HH in mice and men (19, 21, 22). Moreover, higher serum kisspeptins levels are found in obese hypogonadal men and in central hypogonadism than in controls (23). In patients with *GPR54* mutations, GnRH deficiency can be partial or complete, permanent, or reversible, and can have a congenital or adult onset. Six homozygous inactivating mutations have been described in 19 individuals with nIHH: their LH and FSH secretion was blunted but normal secretion was restored after exogenous GnRH stimulation (19, 21, 22). Kisspeptins are highly expressed in placenta during pregnancy; different patterns of spatiotemporal expression of *KISS1* and *KISSR* were described in normal and pathological placentas (24).

# **TAC3R and TAC3 mutations**

The *TACR3* gene (chromosome 4q25) encodes the neurokinin 3 receptor (NK3R) and the *TAC3* gene (chromosome 12q13.3) encodes neurokinin B (NKB),its endogenous ligand. nIHH caused by mutations in *TAC3* and *TAC3R* have an AR heritance (25). As well as the *GPR54/Kisspeptin* system, *TACR3/TAC3* pathway stimulates GnRH neurons. In initial studies, defects in either *TAC3* or *TACR3* were found in 11 patients from 5 of 10 families studied, but in none of 50 sporadic cases (25–27). Francou et al. studied the gonadotropin axis dysfunction associated with nCHH due to TAC3/TACR3 mutations: it was related to a low GnRH pulsatile frequency leading to a low frequency of alpha-subunit pulses and to an elevated FSH/LH ratio (27). They suggested that this ratio might be useful for pre-screening nCHH patients for TAC3/TACR3 mutations. In another broad cohort of normosmic IHH patients, 7 of the 16 males and 5 of the 7 females with *TACR3/TAC3* mutations were assessed after discontinuation of therapy: 6 of the 7 males and 4 of the 5 females demonstrated evidence for reversibility of their hypogonadotropism (14).

# **Leptin (Ob) and leptin receptor mutations**

Leptin is an adipocyte secreted protein that ensures a link between body fat and the reproductive axis. HH and severe obesity are seen in humans and ob/ob mice with genetic leptin deficiency. There are at least 12 patients with leptin deficiency and homozygous mutations. In such cases, recombinant leptin administration restores gonadotropin secretion and dramatically reduces body mass index. Defects in the leptin receptor are more common, being identified in 3% of severe early onset obesity patients. Interestingly, the leptin receptor is expressed on kisspeptin neurons whereas leptin administration induces the expression of *Kiss-1* in *ob/ob* mice (3, 28).

# **LHB mutations**

The *LHB* subunit gene is located at chromosome 19q13.32. Five mutations have been published up to now; clinical and molecular data are summarized in **Table 3**. The syndrome of preserved spermatogenesis with androgenic failure (now known to be due to LH deficiency) was described for the first time by Pasqualini and Bur in 1950 (29). The term "fertile eunuch" was then coined to describe these men.

In affected men, sexual differentiation was normal, but the absence of or significantly reduced LH secretion restrained the induction of puberty and altered Leydig cell proliferation and maturation (30–34). These males have impaired spermatogenesis, ranging from azoospermia to oligospermia, which has been linked to the lack of LH stimulation and low intratesticular testosterone action (5, 30–35). In 2004, we described a man with a homozygous missense mutation (G36D) in the *LHB* subunit gene that abrogated subunit dimerization and rendered LH biologically and immunologically inactive (31). Treatment with human chorionic gonadotropin (hCG) induced near normalization of testicular structure (5). The patient and his wife conceived a child by intracytoplasmic sperm injection from ejaculated sperm. The male heterozygous child had normal LH, FSH, and testosterone levels, at the age of 4 weeks (5, 35).

In women, *LHB* mutations lead to a normal pubertal development but they can have primary amenorrhea and micropolycystic ovaries (32–34).

# **FSHB mutations**

The β subunit of FSH (*FSHB*) is located at chromosome 11p13. Three men and four women with inactivating FSH mutations have been reported. Men have normal pubertal development although they have azoospermia, whereas women have abnormal pubertal maturation; in these patients high level of LH are found whereas FSH is low/undetectable. Estrogen and progesterone concentrations are low (1, 2).

# **Gonadotropins receptor (LHR and FSHR) mutations**

Inactivating mutations affecting the gonadotropin receptors contrast with those affecting their ligands in that they are invariably associated with hypergonadotropic hypogonadism; hence, they are not discussed here.

### **KALLMANN SYNDROME**

Kallmann syndrome, involving the characteristic features of HH and anosmia was noted in the historical literature long before being properly characterized as a genetic disorder. A man with delayed puberty and the lack of olfactory bulbs was reported over 150 years ago by the Spanish doctor Aureliano Maestre de San Juan (1828–1890). The German Franz Kallmann (1897–1965) completed in the 1940s a description of hypogonadism and anosmia in two families, establishing the genetic basis of transmission.

**Table 3 | Clinical, biological, pathological, and genetic studies in patients with LH deficiency**.

All but one patient (Basciani et al.) are homozygotes for an inactivating βLH mutation.

SU α, alpha subunit; inhB, inhibin B; AMH, antimullerian hormone; SPG, spermatogenesis; T2, testosterone; N, normal; Anorm, abnormal; Dim, dimerization. Normal values: FSH (2–14 UI/L), LH (2–10 UI/L), alpha subunit (<1.2 mUI/L).

The Swiss scientist, Georges de Morsier (1894–1982) provided the neuropathological description of the syndrome. KS has a prevalence of 1/5000, with a clear male predominance (1–3, 36, 37). Several mechanisms of inheritance and molecular mutations are described here after and summarized in **Table 2**.

# **Anosmin-1 (KAL-1) mutations**

The *KAL-1* gene is located on the X chromosome at Xp22.3. *KAL-1* encodes anosmin-1, a glycoprotein playing an in important role in kidney, respiratory tract, digestive system, and brain embryogenesis (1–3, 37, 38). Anosmin-1 is an adhesion molecule located on cell surface, consistent with the underlying defect of embryonic neuronal migration in KS. Anosmin-1 is mainly involved in growth and migration of GnRH, mitral olfactory cells, and Purkinje cerebellum neurons. Mutations in *KAL-1* gene cause 14% of familial cases of KS and 11% of cases of sporadic cases (1–3, 38). *KAL-1* mutations lead to HH with or without anosmia and may include synkinesis (mirror movements), unilateral renal aplasia, and mid-line abnormalities such as cleft lip/palate (37, 38).

# **FGF8 (KAL-6), FGF17, and FGFR1 (KAL-2) mutations**

The *FGF8* gene (also known as *KAL-6*) is located on chromosome 10q24. Fibroblast growth factors (FGF) interact with FGF tyrosine kinase receptors to mediate growth and development. *FGF8* participates in gastrulation, regionalization of the brain, and organogenesis of the limb and face as an embryonic epithelial factor. *FGF8* and its receptor *FGFR1* are involved in GnRH neuron migration. *FGF8* inactivating mutations can lead to both KS and nIHH with an AD inheritance. Triallelic inheritance has also been described. In addition, cleft lip or palate and other midline defects have been described in patients with *FGF8* and *FGFR1* mutations. Other features such as corpus callosum hypoplasiaagenesis or nose, ear, and finger abnormalities are more specific of *FGFR1* defects (37, 39, 40).

*FGF17* is located at chromosome 8p2.3 and *FGF17* has a strong sequence identity with *FGF8*. *FGF17* might be implicated in GnRH neuron biology as an alternative to ligand *FGF8b.* Miraoui et al. have identified FGF17 heterozygous mutations in three patients with congenital HH and anosmia and in another individual. In a sporadic male patient with congenital, HH without anosmia (41).

*FGFR1* is located at 8p11.22-p11.23 and *FGFR1* mutations have been identified in 10% of KS. *FGFR1* related KS has an AD inheritance, associated with incomplete penetrance and interfamilial variability. FGFR1 encodes for type 1 FGF receptor, which is expressed in several embryonic tissues. The activation of the FGF– FGFR complex requires two FGF ligands. FLRT3 (Fibronecting like domain containing leucine enrich transmembrane protein 3) also interacts with FGFR (see **Table 2**). In addition, the binding of heparin or HS: heparan sulfate proteoglycan (see HS6ST1 gene later) have been shown to be essential for FGF receptor dimerization and function (38). Mice with Fgfr1−/− mutations and patients with loss-of-function mutations in *FGFR1* have defective GnRH neuron migration (31). Thus, loss of function mutations in *FGFR1* which is involved in the development of the face, lead to the abnormal morphogenesis of the olfactory bulb, while specific gain-of function mutations in *FGFR1* cause craniosynostosis (37, 39–41).

### **PROK2 and PROKR2 mutations**

*PROK2* (locus 3p21.1) and *PROKR2* (locus 20p13) genes encode for prokineticin-2 and its receptor (42). Prok2 and prokr2 gene knockout mice both have agenesis or hypoplasia of the olfactory bulbs, in association with HH. In this model, there is also abnormal GnRH neuron migration (43). Its heritance can be AD or AR. Mutations in these genes are described in up to 6% of KS and 3% of nIHH (1–3, 42). In pituitary deficits associated with septo-optical dysplasia, McCabe et al. (44), described a patient with a heterozygous L173R mutation in *PROKR2* gene, while its healthy mother is homozygous for this mutation. As some controversy exists on the pathogenic role of some PROKR2 mutations, the prevalence given should be interpreted cautiously. Indeed, digenic mutations are encountered (i.e., *PROKR2* and *KAL1*), while heterozygous patients (i.e.,AD transmission) are present in families with healthy relatives presenting the same genotype. There are no reliable accessory pathognomonic features of *PROK2/PROKR2* function loss: patients have been described with obesity, sleep disorder, fibrous dysplasia, epilepsy, and synkinesia (2, 3, 43, 44).

# **NELF mutations**

The *NELF* gene is located at chromosome 9q34.3. This gene encodes the nasal embryonic LHRH factor. The *NELF* gene is detected in olfactory sensory cells and GnRH cells during embryonic development. It constitutes a guidance molecule for the olfactory axon and GnRH neurons across the nasal region (45). *NELF* mutations have been described in patients with KS, in association with mutations in *FGFR1* or *HS6ST1*, indicating digenic inheritance. More studies are necessary to confirm a relationship between *NELF* and any reproductive and olfactory disorders (2).

#### **WDR11 mutations**

The *WDR11* locus is at chromosome 10q26.12 and its heritance is AD. It encodes murine Wdr11 that is expressed in the developing olfactory and GnRH migratory pathway and in the adult hypothalamus.*WDR11* biological function is not well understood: however, Kim et al. identified five different heterozygous mutations in nIHH and KS patients. WDR11 probably also plays an important role in puberty (46).

### **CHD7 mutations**

The *CHD7* gene that encodes a chromatin-remodeling factor is located on chromosome 8q12.1. Mutations (AD inheritance) of this gene can cause CHARGE syndrome (Colobomata, Heart Anomalies, Choanal Atresia, Retardation, Genital, and Ear anomalies). *CHD7* was screened in nearly 200 patients: 7 KS and nIHH patients were found, 3 of them with olfactory abnormalities. CHD7 mutations were identified in 6% of KS and 6% of nIHH, respectively (1, 2, 47, 48). Laittinen et al. described in 2012, a KS patient with a truncating CHD7 mutation that underwent a reversal of central hypogonadism after therapy discontinuation (15).

#### **HS6ST1 mutations**

The *HS6ST1* gene (locus 2q.21) encodes a 6-*O*-sulfation enzyme, which is a member of the heparan sulfate enzyme family. The protein is involved in normal neuronal development and may play a role in limb development. In nematodes, HS 6-*O*-sulfate interacts with anosmin-1 and it is involved in function of FGFR1 and FGF8.

*HS6ST1* shows complex inheritance patterns, not following autosomal or recessive transmission. *HS6ST1* mutations were found in KS patients in combination with mutations affecting the FGFR1 gene. *HS6ST1* mutations were found in patients who had nIHH or variable degrees of olfactory dysfunction (KS) as well as with either normal or abnormal olfactory structures (49).

### **IL17RD, DUSP6, and SPRY4 mutations**

The *IL17RD* gene (locus 3p14.3) encodes a membrane protein belonging to the interleukin-17 receptor (*IL-17R*) protein family. In a study with eight patients with congenital hypogonadism all had KS, 7/8 had absent puberty, 6/8 showed congenital hearing loss. One *IL17RD* allelic defect is likely to be insufficient, meaning that additional affected alleles in the same and/or other genes must be present to create the phenotype of KS with hearing loss (41).

*DUSP6* (locus 12q22-q23) encodes a member of the dual specificity protein phosphatase subfamily. They negatively regulate members of the mitogen-activated protein (MAP) kinase superfamily (25) Three patients were described with DUSP6 and *FGFR1* heterozygous mutation; they were hypogonadic, while one had hearing loss and the two others had abnormal speech. DUSP6−/− mice are however viable and fertile (41).

*SPRY4* (locus 5q.31.3) gene encodes a protein (sprouty homolog 4), which is an inhibitor of the receptor-transduced mitogen-activated protein kinase (MAPK) signaling pathway. It is positioned upstream of RAS gene activation and impairs the formation of active GTP-RAS. Diseases associated with SPRY4 include germ cell cancer, and testicular cancer. Miraoui et al. identified four anosmic patients with congenital HH (three females and one male) with heterozygosity for a c.530A-G transition in exon 3 of the SPRY4 gene. Another female patient had a heterozygosity for a c.910G-A transition in exon 3 of the SPRY4 gene. These mutations were not found in 155 controls. One of the patients also had hearing loss and another one had abnormal dentition (41).

#### **HESX1 mutations**

The *HESX1* gene (locus 3p14.3) encodes a protein that is a transcriptional repressor in the developing forebrain and pituitary gland (27). *HESX1* plays an important role in the temporal and sequential development of the forebrain, hypothalamus, optic nerve, and posterior pituitary (28). Mutations in *HESX1* have also been described in isolated growth hormone deficiency and combined pituitary deficiency (50, 51). *HESX1* mutations have been described in 1.4% of IHH/KS patients (50, 51), but as in *PROKR2* mutations this prevalence should be interpreted cautiously.

# **SEMA3A mutations**

The *SEMA3A* gene (7q21.11) encodes the semaphorin 3A protein, which regulates axonal path finding and participates in GnRH migration. Deletions and mutations of the *SEMA3A* gene validate a role for *SEMA3A* in KS. Moreover, SEMA3A knockout mice exhibit GnRH dependent hypogonadism and abnormal olfactory bulb innervation (52).

# **CONCLUSION AND PERSPECTIVES**

Kallmann syndrome and nIHH have the potential to unravel the processes behind normal embryonic development and reproductive neuroendocrine maturation (2). The complex biological path from childhood to the onset of human puberty is still incompletely understood (1–3). The molecular mechanism behind IHH remains unknown in a large number of cases. Over the past decade, the same genetic mutations have been described in associated with both KS and nIHH. Moreover, a significant clinical heterogeneity is seen in isolated GnRH deficiency, which might be explained to some extent by oligogenicity (6, 7, 9). In addition, over 60% of central hypogonadic patients have no identifiable mutations, suggesting that yet more disease loci remain to be discovered (1–3). These unidentified genes warrant an integrated research including clinicians, geneticists, and biological investigators to pursue further understanding of these fascinating cases.

# **ACKNOWLEDGMENTS**

We acknowledge the National Fund of Scientific Research (FNRS, Brussels, Belgium), for their financial support and Dr AF Daly for kindly reviewing the manuscript.

# **REFERENCES**

**Conflict of Interest Statement:** The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

*Received: 31 March 2014; accepted: 24 June 2014; published online: 09 July 2014.*

*Citation: Valdes-Socin H, Rubio Almanza M, Tomé Fernández-Ladreda M, Debray FG, Bours V and Beckers A (2014) Reproduction, smell, and neurodevelopmental disorders: genetic defects in different hypogonadotropic hypogonadal syndromes. Front. Endocrinol. 5:109. doi: 10.3389/fendo.2014.00109*

*This article was submitted to Neuroendocrine Science, a section of the journal Frontiers in Endocrinology.*

*Copyright © 2014 Valdes-Socin, Rubio Almanza, Tomé Fernández-Ladreda, Debray, Bours and Beckers. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.*

# Health outcomes in acromegaly: depression and anxiety are promising targets for improving reduced quality of life

#### **Victor Jacobus Geraedts 1,2, Christina Dimopoulou1 , Matthias Auer 1 , Jochen Schopohl 3 , Günter Karl Stalla1 and Caroline Sievers 1\***

1 Max Planck Institute of Psychiatry, Munich, Germany

2 Leiden University Medical Center, Leiden, Netherlands

3 Medizinische Klinik Innenstadt, Ludwig-Maximilians University, Munich, Germany

#### **Edited by:**

Gianluca Tamagno, St. Columcille's Hospital, Ireland

#### **Reviewed by:**

Giovanni Vitale, Università degli Studi di Milano, Italy Marek Bolanowski, Wroclaw Medical

University, Poland

#### **\*Correspondence:**

Caroline Sievers, Department of Endocrinology, Max Planck Institute of Psychiatry, Kraepelinstraße 2-10, Munich 80804, Germany e-mail<EMAIL_ADDRESS>
**Introduction:** Remission criteria of acromegaly are based on biochemical variables, i.e., normalization of increased hormone levels. However, the established reduction in Quality of Life (QoL) is suggested to be independent of biochemical control. The aim of this study was to test which aspects predict QoL best in acromegaly.

**Methods/design:** This is a prospective cohort study in 80 acromegalic patients, with a cross-sectional and longitudinal part. The main outcome measure was health-related QoL, measured by a generic and a disease-specific questionnaire (the SF-36 and AcroQoL). Main predictors were age, gender, biochemical control, disease characteristics, treatment modalities, and psychopathology.

**Results:** Our cohort of 80 acromegalics had a mean age 54.7 ± 12.3 years with an average disease duration of 10.8 ± 10.0 years. Ratio macro-/microadenoma was 54/26. In adjusted mixed method models, we found that psychopathology significantly predicts QoL in acromegaly (in models including the variables age, gender, disease duration, tumor size, basal hormone levels, relevant treatment modalities, and relevant comorbidities), with a higher degree of psychopathology indicating a lower QoL (depression vs. AcroQoL: B = −1.175, p < 0.001, depression vs. SF-36: B = −1.648, p < 0.001, anxiety vs. AcroQoL: B = −0.399, p < 0.001, anxiety vs. SF-36: B = −0.661, p < 0.001). The explained variances demonstrate superiority of psychopathology over biochemical control and other variables in predicting QoL in our models.

**Discussion:** Superiority of psychopathology over biochemical control calls for a more extensive approach regarding diagnosing depression and anxiety in pituitary adenomas to improve QoL. Depressive symptoms and anxiety are modifiable factors that might provide valuable targets for possible future treatment interventions.

**Keywords: acromegaly, anxiety, biochemical control, depression, pituitary adenomas, quality of life**

# **INTRODUCTION**

Objectives for health outcomes include reduction of mortality, morbidity, and the improvement of Quality of Life (QoL) (1).

However, in patients with pituitary adenomas, such as acromegaly, this third treatment goal remains often unfulfilled as they report to have a markedly reduced QoL, which often persists under biochemical control/remission (2–7).

Various factors have been suggested to be associated with reduced QoL in pituitary disease, particularly acromegaly; however, there is no clear consensus in this regard.

The current consensus criteria for cure and remission of acromegaly are based on biochemical variables, i.e., normalization of elevated hormonal levels of the biomarkers growth hormone (GH) and/or insulin like growth factor-1 (IGF-1) (8–10). However, the definition of such a "remission" remains insufficient with regard to absolute recovery: a drawback of a purely biochemical approach is that there are strong indications that biochemical control does not necessarily provide complete "cure" in patients' view, since their health-related QoL remains reduced in most patients with acromegaly.

However, there is no consensus on the value of these contributing factors. For instance, several articles reported biochemical control to have no significant association with reduced QoL (11– 13), whereas other articles report exactly the opposite (14–16), indicating a clear need for elucidation.

Psychopathological variables are candidate modifiable factors to link pituitary disease, especially acromegaly, to a lower healthrelated QoL. On the one hand, acromegaly is reported to be associated with neuropsychiatric comorbidities such as depressive symptoms (17) and anxiety (5, 18, 19). On the other hand, there is a clear association between psychopathology and perceived QoL (20–22).