Body Composition After Endogenous (Cushing's Syndrome) and Exogenous (Rheumatoid Arthritis) Exposure to Glucocorticoids

Exposure to chronic glucocorticoid (GC) excess determines changes in body composition. The aim of the study was to compare body composition in women exposed to endogenous hypercortisolism (Cushing's syndrome, CS), exogenous glucocorticoid treatment (rheumatoid arthritis, RA) and controls. Fifty-one CS women, 26 RA women treated with low-dose prednisone (5 mg/day or 10 mg/2 days), and 78 female controls were included. Fourteen CS patients were hypercortisolemic, 37 in remission (10 required hydrocortisone substitution after surgery). Body composition parameters were measured by dual-energy X-ray absorptiometry scanning (DEXA). RA patients had a greater waist-hip ratio (WHR) (p<0.01), less lean body mass (LBM) (p<0.01), and lumbar bone mineral density (BMD) (p<0.01) than controls. CS patients, globally and those with cured disease, had more total fat (both percentage and kg) and trunk fat percentage, and less whole body-BMD than RA patients (p<0.05, p<0.01, p<0.05, respectively). Active CS patients had less whole body-BMD and more LBM than RA patients (p<0.05, p=0.01, respectively). Cured CS patients not taking hydrocortisone had more total fat [both percentage (p<0.05) and kg (p<0.05)], trunk fat percentage (p<0.05), lumbar BMD (p<0.01) than RA patients. Cured CS patients requiring hydrocortisone only differed from RA patients by smaller WHR (p<0.01). All the differences in BMD disappeared when the data were reanalyzed including only the estrogen-deficient groups. Hypercortisoliof CS determines an irreversible increase in body fat, greater than in RA. Endogenous and exogenous exposure to GC negatively affects body composition by increasing the WHR. There appears to be no additional effect on BMD in estrogen-deficient women.

Introduction ▼ Exposure to chronic glucocorticoid (GC) excess produces marked changes in body composition, reducing bone mass and lean body mass, and favoring central fat accumulation [1,2] . Cushing ' s syndrome (CS) is a rare endocrine disease characterized by cortisol hypersecretion, mainly by a pituitary tumor (Cushing ' s disease) or, less frequently, by an adrenal or an ectopic neuroendocrine tumor. Changes in body composition in CS include increased fat mass, decreased bone mass, thinning of the skin, and reduced lean mass. Why these tissues are aff ected so dramatically is unclear [3] . Glucocorticoids (GC) regulate lipid metabolism through promoting lipogenesis in adipose tissue [4] . An increase in visceral fat in both male and female patients with CS has been reported, with the Exposure to chronic glucocorticoid (GC) excess determines changes in body composition. The aim of the study was to compare body composition in women exposed to endogenous hypercortisolism (Cushing ' s syndrome, CS), exogenous glucocorticoid treatment (rheumatoid arthritis, RA) and controls. Fifty-one CS women, 26 RA women treated with low-dose prednisone (5 mg / day or 10 mg / 2 days), and 78 female controls were included. Fourteen CS patients were hypercortisolemic, 37 in remission (10 required hydrocortisone substitution after surgery). Body composition parameters were measured by dualenergy X-ray absorptiometry scanning (DEXA). RA patients had a greater waist-hip ratio (WHR) (p < 0.01), less lean body mass (LBM) (p < 0.01), and lumbar bone mineral density (BMD) (p < 0.01) than controls. CS patients, globally and those with cured disease, had more total fat (both per-centage and kg) and trunk fat percentage, and less whole body-BMD than RA patients (p < 0.05, p < 0.01, p < 0.05, respectively). Active CS patients had less whole body-BMD and more LBM than RA patients (p < 0.05, p = 0.01, respectively). Cured CS patients not taking hydrocortisone had more total fat [both percentage (p < 0.05) and kg (p < 0.05)], trunk fat percentage (p < 0.05), lumbar BMD (p < 0.01) than RA patients. Cured CS patients requiring hydrocortisone only diff ered from RA patients by smaller WHR (p < 0.01). All the diff erences in BMD disappeared when the data were reanalyzed including only the estrogen-defi cient groups. Hypercortisolism of CS determines an irreversible increase in body fat, greater than in RA. Endogenous and exogenous exposure to GC negatively aff ects body composition by increasing the WHR. There appears to be no additional eff ect on BMD in estrogen-deficient women.
lage and ankylosis of the joints. RA can also produce diff use infl ammation in the lungs, pericardium, pleura, and sclera, and also nodular lesions, most common in subcutaneous tissue under the skin. Although the cause of RA is unknown, autoimmunity plays a pivotal role in its chronicity and progression. Various treatments, including nonpharmacological physical therapy and occupational therapy are available. Analgesia (painkillers) and anti-infl ammatory drugs, as well as steroids are used to suppress symptoms, while disease-modifying antirheumatic drugs (DMARDs) are often required to inhibit or halt the underlying immune process and prevent long-term damage [10] . For sustained improvement in a chronic disease such as RA, it appears that GC must be given more or less continuously [11] , but unwanted signs of hormonal excess have been reported in 40 % of cases at some time, during the course of treatment. Most of them are considered mild or transient and disappear or lessen if the doses of GC are reduced, often followed also by a decline in the degree of improvement [11] . Few data are available on body composition parameters in CS and no data are available on the comparison of body composition parameters in endogenous versus exogenous GC exposure. While CS is a rare condition, exposure to exogenous GC is a common situation in clinical practice, for a variety of causes. It can be assumed that no ideal group of healthy women taking GC chronically can exist. Thus, when considering a comparison group for the CS patients we selected those with RA, since both these conditions predominate in females, who are not associated with malignancy and have chronic diseases treated in ambulatory care. The aim of this study was to compare body composition parameters after exposure to endogenous hypercortisolism (due to Cushing ' s syndrome) and exogenous GC treatment (with lowdose prednisone in rheumatoid arthritis) with healthy controls.

Patients and Methods ▼ Patients
A total of 51 women with CS were included (mean age 51.4 ± 13.5 years). At the time of the study, 14 CS patients were hypercortisolemic (i. e., active, 12 of pituitary and 2 of adrenal origin) and 37 were in remission (i. e., cured, 27 of pituitary and 10 of adrenal origin). CS was considered in remission if either adrenal insuffi ciency was demonstrated [basal AM cortisol < 100 nmol / l ( < 4 μ g / dl) and / or undetectable 24-h free urinary cortisol] or morning cortisol suppression ( < 50 nmol / l, < 1.8 μ g / dl) after 1 mg dexamethasone overnight was observed. Of the 37 in remission, 10 were adrenal insuffi cient at the time of the evaluation and required substitution therapy with hydrocortisone (mean of 20 mg every day), while the other 27 had presented transient hypocortisolism after surgery, but did not currently require substitution therapy. The mean time of hormonal cure since normalization of cortisol to study date was 11 ± 6 years (range, 0.7 -22 years). The mean duration of endogenous hypercortisolism was 70 ± 5 months. Duration of hypercortisolism was considered as the period of time between symptoms onset (as referred by the patients) and the diagnosis of CS plus the period of time between diagnosis and remission of hypercortisolism after treatment. Information on pituitary function (GH, IGF-I, TSH, free T4, prolactin) and estrogen status were collected. Seven patients were GH-deficient (3 of them were treated with recombinant human GH), 5 were hypothyroid (all on L-thyroxine replacement), 25 were estrogen-defi cient (17 cured and 8 active, 2 gonadotropin deficient, 23 menopausal), while the other 26 were estrogen-sufficient (20 cured, 6 active). Twenty-six women with RA (mean age 62.0 ± 10.1 years) who required treatment with low doses of prednisone (5 mg every day or 10 mg every 2 days) were included, from the Rheumatology Department of our hospital. Diagnostic criteria for RA are those proposed by the American College of Rheumatology (ACR) for classifi cation of the disease [12] . Mean duration of exogenous GC treatment was 65 ± 6 months. This dose of prednisone was chosen since 5 mg prednisone is considered to be equivalent to 20 mg of hydrocortisone [13] . Information on thyroid function and estrogen status was collected: 3 were hypothyroid (all on L-thyroxine replacement), 22 were estrogen-defi cient (menopausal), and 4 were estrogen-suffi cient ( Table 1 ). Seventy-eight healthy control women, (mean age 53.71 ± 12.97 years), were selected from the blood donors ' database at our hospital. They have routine blood test and a complete medical history at our hospital. Letters were sent to controls, and a phone call was made one week later; the fi rst control to accept was included. Controls that referred GC treatment or malignant diseases were excluded. All patients and controls signed an informed consent after study approval by the hospital ethics committee.

Methods
Lumbar spine and whole body bone mineral density (BMD) and body composition [lean body mass (LBM), whole and trunkal fat mass (FM) and total mass] were measured by dual-energy X-ray absorptiometry scanning (DEXA, Delphi QDR 4500, Hologic, Vil-

Statistical analysis
Quantitative data are expressed as mean and SD (Gaussian distribution

▼ Comparisons between rheumatoid arthritis (RA) patients and controls
Patients with RA had a greater waist-hip ratio (p < 0.01) and less lean body mass (LBM) (p < 0.01) and lumbar BMD (p < 0.01) than controls ( Table 2 ).

Comparisons between Cushing ' s syndrome (CS) patients and controls
Patients with CS as a whole group had more total fat [both in percentage (p < 0.01) and in kg (p = 0.05)] and trunk fat [both in percentage (p < 0.01) and in kg (p < 0.01)], waist (p = 0.02), and a greater waist-hip ratio (p < 0.01) than controls. CS patients had less whole body-BMD (p < 0.05) and lumbar BMD (p < 0.05) than controls ( Table 2 and • ▶ Fig. 1 ) . When CS patients were divided into " cured " and " active " , the above diff erences were maintained with respect to controls; additionally cured CS had less LBM than controls ( Table 3 ). Active CS had a greater waist-hip ratio (p < 0.05) and less lumbar BMD (p < 0.01) than cured CS. Regarding cured CS patients that were adrenal insuffi cient at the time of the DEXA evaluation and required substitution therapy with hydrocortisone, they had less LBM, whole body-BMD and lumbar BMD, more trunk fat in percentage and a greater waisthip ratio than controls (p < 0.05, p < 0.01, p < 0.05, respectively). Additionally they had less lumbar BMD than the cured CS patients (p < 0.01) who did not require hydrocortisone substitution ( Table 4 ) . Cured CS patients who did not require hydrocortisone substitution had more total fat [both in percentage (p < 0.01) and in kg (p < 0.01)] and trunk fat [both in percentage (p < 0.01) and in kg  (p < 0.01)], and a greater waist-hip ratio (p < 0.01) than controls ( Table 3 ). On the other hand they had a greater lumbar BMD (p < 0.01) than CS patients that required substitution therapy with hydrocortisone ( Table 4 ).

Comparisons between CS and RA patients
CS patients had more total fat (both in percentage and in kg) and trunk fat in percentage and less whole body BMD than patients with RA (p < 0.05, p < 0.01, p < 0.05, respectively) ( Table 2 and • ▶ Fig. 1 ).
Cured CS patients, both globally and those cured who did not require hydrocortisone, had more total fat [both in percentage (p < 0.05) and in kg (p < 0.05)], trunk fat in percentage (p < 0.01 and p < 0.05), lumbar BMD (p < 0.05 and p < 0.01) than RA patients. Additionally the global CS group also had less whole body-BMD (p < 0.05) than RA patients. Patients with active CS had less whole body-BMD and more LBM than RA patients (p < 0.05 and p = 0.01 respectively) ( Table 3 ) . Cured CS patients still requiring hydrocortisone substitution therapy only diff ered from RA patients by a smaller waist-hip ratio (p < 0.01), but were otherwise not diff erent in any of the other body composition parameters evaluated ( Table 4 ). No differences in BMI within the 3 studied groups were observed.  1.05 ± 0.1 1.02 ± 0.7 < 0.01 < 0.01 < 0.05 < 0.05 0.12 Lumbar BMD (g / cm 2 ) 1.05 ± 1.1 0.9 ± 0.1 < 0.05 < 0.01 < 0.05 0.72 < 0.01 a p 1 between Cured CS and controls; p 2 between Active CS and controls; p 3 between Cured CS and RA; p 4 between Active CS and RA; p 5 between Active CS and Cured CS A p-value of < 0.05 was considered signifi cant. For controls and RA values see Table 2 Role of GH defi ciency When the data were reanalyzed excluding the CS patients with GH defi ciency without replacement therapy (n = 4) the same results were found.

Role of estrogens
Because estrogens play an important role on bone, CS patients, RA patients and controls were divided into an estrogen-sufficient group (including premenopausal with regular menses and postmenopausal women with hormone replacement therapy) and an estrogen-defi cient group (including postmenopausal and premenopausal women with no replacement for hypogonadism). When data were reanalyzed comparing only the estrogendefi cient groups, all the diff erences in lumbar BMD and whole body-BMD disappeared. The other diff erences in body composition parameters, however, remained the same ( Table 5 ).

Discussion ▼
While exposure to endogenous hypercortisolism due to CS is a rare situation, chronic treatment with exogenous GC is common in clinical practice for a variety of causes. There is a general consensus that the shortest period at the lowest possible dose of GC is desirable to prevent side eff ects, but there is little awareness that low doses may also be harmful. For this reason we compared body composition after exposure to endogenous hypercortisolism (due to Cushing ' s syndrome) and exogenous GC treatment (with low-dose prednisone in rheumatoid arthritis). Increased total and central body fat is a common feature of CS [14,15] . Waist-hip ratio is an abdominal fat distribution index, predictor of cardiovascular risk [16] , and is more sensitive than BMI to evaluate clustering of coronary risk factors among nonobese men and women [16] . Since the women with RA were older than those with CS, a statistical analysis using age as covariant was performed in order to make the groups comparable and avoid the eff ect of age. We found that both patients with CS and RA have a higher waisthip ratio than controls, with no diff erence between them, consequently indicating a higher cardiovascular risk. This would suggest that the waist-hip ratio might be used in clinical practice to assess cardiovascular risk both in CS and RA patients. Moreover, persistence of increased total and central body fat has been reported in patients with cured CS [17,18] . This increase in trunk fat is a common complaint of patients who have suff ered from CS and also those who have been treated with high doses of exogenous GC. Given the increased cardiovascular risk conferred by central obesity, this persistence in trunk fat may contribute to long-term morbidity in these patients, despite endocrine control [17,19] . A possible explanation for the persistent increase in trunk fat may be the eff ect of cortisol on the omental adipose tissue; after hypercortisolism has disappeared, the increased number of fat cells, enhanced by cortisol, remains and can contribute to explain the persistent increase in abdominal fat deposits [20 -22] . A dysregulation in the hypothalamic-pituitary-adrenal (HPA) axis could also be involved [23] . Another novel mechanism that explains the deposition of visceral adipose tissue and consequent central obesity in patients with iatrogenic or endogenous CS is the inhibition of AMPK activity in adipose tissue by GC [24] .
The coexistence of GH defi ciency in patients with Cushing ' s disease after long-term remission of hypercortisolism obtained by surgery, by itself may aff ect body composition [25 -27] . Male gender and length of hypercortisolism are the most signifi cant predictors of postsurgical GH defi ciency; our study only included women and few GH defi ciency, these could be the reasons why no eff ect of GH defi ciency on body composition was found. Estrogens exert an important eff ect on normal bone density and estrogen defi ciency is associated with bone loss [28] as well as an increase in total and trunk fat when compared to healthy controls [29] . In this study we show that the lower values of BMD in CS and RA, when compared to controls, disappeared when only estrogen-defi cient women were compared, highlighting that once the protective eff ect of estrogen is lost, GC exposure is less important for the bone, be it of endogenous or exogenous origin. This is clearly diff erent from the eff ect on the fat, since CS has greater body fat, independently of the estrogen status; this would confer to these patients a persistent higher cardiovascular risk.
The deleterious eff ects of glucocorticoid replacement on bone in estrogen-suffi cient women after long-term remission of Cushing ' s Syndrome has been recently demonstrated; the damage done to bone during endogenous hypercortisolism may have contributed to determine a worse response to subsequent GC replacement [30] . The exact mechanism is not known, in the literature the mevalonate pathway is involved in glucocorticoidinduced osteoblast dysfunction [31] . Patients with RA had less lean body mass (LBM) than controls and less than active CS, even when only estrogen defi cient women were compared. Less lean body mass has been described in RA [32] , probably related to less muscle exercise due to articular pain in these patients. From our study it appears that neither GC exposure nor estrogen status play a further role on LBM in these RA patients. It is not so clear why CS patients have more severe changes in body composition than RA patients. This is the fi rst study to address and compare this issue. Since the length of glucocorticoid exposure is quite similar, we could hypothesize, based on clinical experience and on the literature, that chronic, continuous, throughout 24 h, endogenous exposure over months or years to severe hypercortisolism, has a greater eff ect on body composition than daily prednisone treatment administrated once a day, as occurs in RA. As demonstrated previously [14] , the cumulative exposure to GC in CS does not appear to be a determinant factor.
In conclusion, prior exposure to endogenous hypercortisolism in CS determines an irreversible increase in body fat, greater than that seen in RA treated with low dose prednisone. Endogenous and exogenous exposure to GC negatively aff ects body composition by increasing the waist-hip ratio. There appears to be no additional eff ect on BMD in estrogen-defi cient women.