D e r m a t o l o g i c a 1991;183:100-105
© 1991 Karger A G , Ba^
0011-9075/91/1832-0100 $2.75.
Elevated Glucocorticoid Receptor Concentrations before and after Glucocorticoid Therapy in Peripheral Mononuclear Leukocytes of Patients with Atopic Dermatitis
M. Rupprecht, R. Rupprecht, J. Kornhuber, N. Wodan, H.U. Koch, P. Riederer, O.P. Hornstein
D e p a r t m e n t of D e r m a t o l o g y , U n i v e r s i t y o f E r l a n g e n - N u r n b e r g ; D e p a r t m e n t o f P s y c h i a t r y , U n i v e r s i t y o f W l i r z b u r g ; M a x P l a n c k Institute of P s y c h i a t r y , M u n c h e n , F R G
Key Words. Glucocorticoids • Glucocorticoid receptors • Atopic dermatitis
Abstract, The number and affinity of glucocorticoid binding sites in peripheral mononuclear leukocytes of patients with atopic dermatitis ( A D ) and healthy controls were determined under baseline conditions and after a defined oral glucocorticoid treatment. Patients with A D (n = 15) exhibited significantly more glucocorticoid receptors ( G R ) per cell than the control group (n = 22), while the G R affinity did not differ. Methylprednisolone treatment resulted in a signif- icant reduction of the G R sites per cell in the steroid-treated control group (n = 10) in contrast to the patients. The dis- sociation constant was not affected by methylprednisolone treatment in either group. In view of the therapeutic effi- ciency of glucocorticoids in A D and findings of abnormal c A M P and cAMP-phosphodiesterase activity, the elevated G R concentrations in A D lend support to the hypothesis of a compensatory G R upregulation due to an insufficient action of endogenous Cortisol or to altered cAMP-induced G R expression.
Introduction
Studies on patients with atopic dermatitis ( A D ) have revealed a variety of immunological, hormonal and psy- chological [1,2] derangements including elevations of IgE levels [3, 4], disordered cell-mediated immunity [5-7], abnormal c A M P and cAMP-phosphodiesterase activity [8, 9] and an impairment of p-adrenoceptor function [10, 11]. Recent investigations pointed to a possible hormonal dysregulation of the hypothalamic-pituitary-adrenal ( H P A ) system in A D . Slightly lowered baseline values of Cortisol [12] in face of a normal adrenomedullar function [13], minor imbalances in the circadian rhythm of Cortisol serum levels [14] as well as a reduced urine excretion rate of steroid metabolites contrasting with normal Cortisol blood levels [15] have been noted.
Topical and systemic administration of glucocorticoids is of major importance in the treatment of A D . The effects of glucocorticoids are mainly mediated via specific
intracellular receptors which have been identified in vari- ous mammalian and human tissues, e.g. the skin [16-18], lung [19], brain [20] and mononuclear leukocytes [21-23].
A s the glucocorticoid receptor ( G R ) has been shown to underlie an autoregulatory control in skin fibroblasts [17, 18] and in human leukocytes [21-26], the present study was designed to explore the potential role of G R autoregulation for the alteration of the H P A system in A D .
Material and Methods
Subjects
Fifteen inpatients (3 men and 12 w o m e n ) suffering from A D and 22 healthy controls (11 m e n , 11 women) w i t h o u t personal history o f asth- m a , hay fever or A D participated in the study v o l u n t a r i l y w i t h i n f o r m e d consent. T h e patients were aged between 18 and 36 years ( m e a n ± S D : 23.4 ± 5.6) and the controls for baseline studies between 19 and 42 years ( m e a n ± S D : 28.5 ± 6 . 3 ) . T e n o f the healthy subjects (4 m e n , 6 w o m e n ;
mean age ± S D : 24.3 ± 2.8 years), who had the same age as the patients, agreed to further participate in the study and received the same steroid dosage as the patients for the autoregulation study.
A l l p a r t i c i p a n t s w e r e w i t h d r a w n from any systemic therapy with corticoids o r a d r e n o c o r t i c o t r o p i c h o r m o n e at least 2 months p r i o r to the study. M e d i c a t i o n , i f a n y , w i t h (3-blocking and/or sleep-inducing drugs was s t o p p e d 3 o r m o r e days before entering the study. Cutaneous lesions o f A D w e r e treated o n l y w i t h corticoid-free ointments at least 1 week before onset a n d d u r i n g the study.
E x c l u s i o n c r i t e r i a w e r e prevalence o f severe internal illness and a l c o h o l , d r u g o r n i c o t i n e abuse. Seven o f the patients and 4 of the 10 healthy c o n t r o l s w e r e t a k i n g b i r t h c o n t r o l pills.
T h e diagnosis o f A D was based on the history of recurrent flexura!
eczema, p e r s o n a l and/or family history of atopy and clinical criteria of A D as established by H a n i f i n and R a j k a [27]. T h e severity of cutaneous i n v o l v e m e n t was e v a l u a t e d a c c o r d i n g to C o s t a ct a l . [28] and revealed moderate to m a r k e d disease activity. T h e score ranged between 37 and 6 9 p o i n t s ( m e a n ± S D : 4 7 . 6 ± 10.6) before therapy and between 11 and 55 points ( m e a n ± S D : 2 7 . 9 + 1 0 . 7 ) at the end of the study. N o n e of the patients suffered f r o m actual b r o n c h i a l asthma.
Study Design
A f t e r o b t a i n i n g the baseline G R and h o r m o n e parameters, all patients a n d the steroid-treated controls underwent a 1-mg dcxametha- sone suppression test ( D S T ) a n d subsequent oral methylprednisolone therapy as s h o w n i n table 1. T h e D S T was used to evaluate the integrity of the feedback r e g u l a t i o n o f the H P A system.
F o r d e t e r m i n a t i o n o f G R b i n d i n g characteristics and hormone data p e r i p h e r a l venous b l o o d samples were collected into E D T A - c o n t a i n - ing plastic tubes (Sarstedt, F R G ) .
Cell Preparation and Binding Assay
M o n o n u c l e a r c e l l fraction was prepared by s o d i u m metrizoate- F i c o l l ( S i g m a , St. L o u i s , M o . , U S A ) density gradient cent! if ligation [29]. C e l l s w e r e w a s h e d twice i n phosphate-buffered saline ( P B S , p i I 7.4; B o e h r i n g e r , M a n n h e i m , F R G ) for 10 m i n , incubated for 60 min at 37 ° C in P B S and then washed again to a l l o w sufficient dissociation of endogenous h o r m o n e . T h e final concentration of cells was determined using a C o u l t e r c o u n t e r ( M o d e l S 5 , C o u l t e r E l e c t r o n i c s L t d . U K ) . V i a - bility o f cells e x c e e d e d 9 5 % , as j u d g e d from their ability to exclude try- pan b l u e . C o n t a m i n a t i o n by erythrocytes was < 1 0 % , by granulocytes and m o n o c y t e s < 8 % .
B i n d i n g e x p e r i m e n t s as d e s c r i b e d by Rupprecht et a l . [25] were c a r r i e d out at 37 ° C in plastic m i c r o t i t e r plates in a total volume of 0.25 m l . T h e d i s p l a c i n g c o m p o u n d in a final concentration of 10 \iM unla- b e l e d dexamethasone ( S i g m a , St. L o u i s , M o . . U S A ) solved in 5% ctha- n o l , c o n t a i n i n g 5 m M D - g l u c o s e , was added to half of the samples to d e t e r m i n e nonspecific b i n d i n g i m m e d i a t e l y p r i o r to the addition of [3H ] d e x a m e t h a s o n e ( A m e r s h a m , U K ) . Increasing concentrations of [3H ] d e x a m e t h a s o n e (specific activity: 84 C i / m m o l ) from 1 to 40 nA/
were used. S a t u r a t i o n experiments were carried out at e q u i l i b r i u m after 90 m i n i n c u b a t i o n in P B S with 5 vaM D-glucose at 37 C . A f t e r i n c u b a t i o n , b o u n d ligand was separated from free ligand by rapid filtra- tion t h r o u g h S c a t r o n filters ( N o . 11731) with a Titertek cell harvester by t w o 5-second washes w i t h P B S at r o o m temperature. T h e filters were transferred i n t o plastic v i a l s , 5 m l of a toluene-based scintillation c o c k t a i l was added ( R o t i s z i n t 22: R o t h . K a r l s r u h e . F R G ) and they were m o n i t o r e d for t r i t i u m in a B e c k m a n L S 1801 counter at about 54%
efficiency. A l l samples were assayed in triplicate with a variation coef- ficient < 7 % .
5 10 15 Bound, M~1 2/ 1 06 cells
20
F i g . 1. Seatehard plot of | \ I|dexamethasone b i n d i n g to h u m a n mononuclear leukocytes. Specifically bound/free ((H | d e x a m e t h a s o n e is plotted versus specifically h o u n d f H )dexamethasone.
Table 1. Study design
Day Time C J1 ucoeoriicoid doses
1 7 a.m.
4 p . m .
11 p . m . 1 mg dexamethasone 2 7 a.m.
4 p . m .
3--6 7 a . m . 8 mg methylprednisolone 7 - S 7 a . m . 4 mg methylprednisolone 8 4 p . m .
Table 2. C o r t i s o l mean 4 S D values ( controls (n 10)
A B
7 a ,m. 4 p . m . 7 a
B l o o d samples
hormone data
hormone -f receptor data
hormone data h o r m o n e data
h o r m o n e -f receptor data
C
4 p . m . 4 p . m .
Patients 27.5 + 13.8 13.4 + 11.3 1.6 + 1.4 4.1 + 4.0 5.6 ± h.9
Controls 25.1 t 8.6 14.6 + 6.9 2.3 + 1.5 3.3 + 3.3 7.7 i 5.7
A Baseline levels. K after oral administration of 1 mg dexame- thasone: C after oral methylprednisolone treatment.
Table 3. G R sites in female patients and female controls t a k i n g and not taking birth control pills under baseline conditions and after methyl- prednisolone treatment
Patients Baseline Prednisolone C o n t r o l s
Baseline Prednisolone
Pill
n - 7 3.783 + 1.658 3.353 +. 1.105 n - 4 2.642 * 567 2.030 + 470
N o pill
n - 5 3.886+ 1.537 3.250 ± 1.643 n - 2 2.764 + 85<>
2.087+ 14
5,000 -I
Baseline Corticoid treatment
F i g . 2. G R sites p e r c e l l . M e a n s ± S E , i n patients w i t h A D ( • , n = 15), the c o r t i c o i d - t r e a t e d c o n t r o l s ( H , n = 10) a n d the w h o l e c o n t r o l g r o u p ( • , n = 22) before ( b a s e l i n e ) a n d after g l u c o c o r t i c o i d t r e a t m e n t . T h e asterisks i n d i c a t e significant differences ( p < 0 . 0 2 ) .
Baseline Corticoid treatment
F i g . 3. K d . M e a n s ± S E , i n patients w i t h A D ( • . n = 15), the c o r t i - c o i d - t r e a t e d c o n t r o l s ( ^ , n = 10) a n d the w h o l e c o n t r o l g r o u p ( • , n —22) before ( b a s e l i n e ) a n d after g l u c o c o r t i c o i d t r e a t m e n t .
R e c e p t o r l e v e l s a n d the d i s s o c i a t i o n constant ( Kd) were d e t e r m i n e d b y S c a t c h a r d plots [30]. A t y p i c a l S c a t c h a r d p l o t o f [3H ] d e x a m e t h a s o n e is s h o w n i n figure 1.
S p e c i f i c b i n d i n g was d e t e r m i n e d b y s u b t r a c t i n g the n o n s p e c i f i c f r o m t o t a l b i n d i n g . N o n s p e c i f i c b i n d i n g ( i n percent o f t o t a l b i n d i n g ) at a c o n c e n t r a t i o n o f 10 nM [3H ] d e x a m e t h a s o n e was 28 ± 1 8 % i n patients a n d 26 ± 1 5 % i n c o n t r o l s .
Cortisol Assay
C o r t i s o l was m e a s u r e d b y a n e n z y m e - l i n k e d i m m u n o a s s a y ( E l i a s , F r e i b u r g , F R G ) . T h e l o w e r d e t e c t i o n l i m i t was 22 nmol/1, a n d the i n t r a - a n d i n t e r a s s a y coefficients o f v a r i a t i o n w e r e 6 % for b o t h .
C o r t i s o l n o n s u p p r e s s i o n was d e f i n e d as the i n a b i l i t y to suppress b o t h p o s t d e x a m e t h a s o n e Cortisol l e v e l s t o b e l o w 5 ug/dl [31].
Statistical Analysis
P r e l i m i n a r y e s t i m a t e s o f b i n d i n g p a r a m e t e r s f r o m s a t u r a t i o n e x p e r i m e n t s w e r e p r o v i d e d by the E B D A p r o g r a m . F i n a l e s t i m a t e s o f b i n d i n g p a r a m e t e r s w e r e d e t e r m i n e d w i t h a c o m p u t e r i z e d n o n l i n e a r , least-square r e g r e s s i o n a n a l y s i s [32]. T h i s w e i g h t e d c u r v e f i t t i n g p r o - g r a m assumes b i n d i n g a c c o r d i n g to the l a w o f mass a c t i o n to i n d e p e n - dent classes o f b i n d i n g sites. T h e results are e x p r e s s e d as the m e a n
± S D , a n d as the m e a n ± S E i n the f i g u r e s .
D a t a w e r e a n a l y z e d u s i n g the t-test for p a i r e d o r g r o u p s a m p l e s a n d P e a r s o n ' s p r o d u c t m o m e n t c o r r e l a t i o n . A l l s i g n i f i c a n c e l e v e l s are t w o - t a i l e d .
Results
Cortisol mean values of the patients and the controls did not differ significantly at any of the blood sampling time points during the 1-mg D S T (table 2). Three of the patients and 2 of the controls fulfilled the criterion for Cor- tisol nonsuppression.
Comparison of the postmethylprednisolone Cortisol levels to the 4 p.m. baseline levels revealed significant reduction in patients and controls (p<0.03).
The patients had significantly (p<0.02) more G R sites per cell than controls (fig. 2). After methylprednisolone therapy the patients revealed a slightly smaller mean number of G R sites per cell in relation to the baseline val- ues (mean baseline: 3,894± 1,521; after treatment:
3,394± 1,189, not significant), whereas in the controls (n= 10) a significant reduction of the G R sites was found (mean baseline: 2,695 ± 5 8 2 ; after treatment: 2,036 ± 4 1 0 , p<0.02).
The significantly more G R binding sites per cell per- sisted after therapy in the patient group when compared with the steroid-treated control group (p<0.002).
There was no difference in baseline or methylpredniso- lone-treated G R sites between female subjects taking birth control pills and those who did not (table 3).
The K
ddid not differ at baseline condition and after methylprednisolone treatment in both groups (fig. 3). N o relations were found between age, sex, Cortisol levels, severity of the skin disorder and G R binding parameters under the conditions studied.
Discussion
Although slightly lowered baseline Cortisol serum con-
centrations in patients with A D have been observed [12],
our study is in accordance with reports on normal Cortisol
levels in A D [15]. Twenty percent of the patients with A D
and 20% of the controls showed no suppression of Cortisol
to below 5 [xg/dl after 1 mg dexamethasone. However, abnormal D S T data may not only occur in various dis- eases but can also be induced by multiple interference fac- tors such as stress and weight loss in healthy subjects [33, 34]. Therefore, the abnormal D S T results obtained in A D and healthy controls may represent an unspecific finding.
A s no data are available on G R binding characteristics in A D we used peripheral blood mononuclear leukocytes to study G R pharmacology, partly for their easy accessi- bility but also for their relevance to the pathogenesis in A D [35] as emphasized by the finding that bone marrow transplantation for other reasons could either confer the state of atopy to former nonatopics [36, 37] or may clear A D in patients with a concomitant immune deficiency disorder [38].
The elevated G R concentrations found in the patients with A D are probably not representing a causal factor, otherwise worsening of the skin lesions would be expected at the beginning of glucocorticoid treatment due to an increased number of activated hormone-receptor com- plexes. A n upregulation of the glucocorticoid binding sites as a compensatory mechanism is more likely. Possi- ble hypotheses for such an upregulation may be altera- tions in cAMP-induced G R expression [39], activation of the G R synthesis by unspecific mediators [40] or a relative biological inefficiency of endogenous corticoids. The lat- ter hypothesis might be supported by the well-known ben- efit of glucocorticoids for the treatment of A D and also by the frequent occurrence of a rebound phenomenon after discontinuation of glucocorticoid administration. A n increase in receptor protein half-life [39, 41] due to reduced cortisol-receptor interactions as supposed after depletion of endogenous Cortisol following metyrapone administration [26] or adrenalectomy [42], or an enhanced transcription rate for the G R [43] may also be responsible for the elevated G R sites in A D .
N o correlations were found between endogenous Cor- tisol levels and G R binding sites in the control and patient group. This result is in accordance with previous studies in healthy and asthmatic subjects [26, 44, 45] and indicates that the plasma concentrations of endogenous glucocorti- coids at the time of the G R assay are of minor importance for the estimation of G R number.
However, prolonged glucocorticoid administration results in downregulation of the number of G R [18, 21, 24] by influencing G R gene transcription [46] without sig- nificantly affecting the binding affinity. In the controls methylprednisolone administration was followed by a sig- nificant decrease in G R binding, whereas in the patients with A D only a slight reduction of G R binding was
observed. This finding might be interpreted as a dimin- ished downregulation of G R by exogenous glucocorti- coids which could be due to the heterogeneity of the patient group shown by the greater standard deviation and might also be compatible with the hypothesis of a cAMP-induced overdrive of G R expression [39] or a rela- tive biological inefficiency of endogenous Cortisol which would require a greater amount of exogenous glucocorti- coids to downregulate the G R . However, it cannot be excluded that an eventual displacement of receptor- bound methylprednisolone by dexamethasone during the assay may underlie different kinetics in patients with A D which might affect the estimation of K
dand B
m a xvalues.
The use of monoclonal antibodies for quantification of G R expression [47] might overcome the difficulties with ligand exchange assays and furnish a direct proof for the hypothesis of a decreased G R downregulation in A D .
A s no data are available on G R in other inflammatory skin diseases so far, the specificity of the present findings is still questionable. However, although A D and bron- chial asthma are often closely associated [48] and gluco- corticoid therapy is efficient in both diseases [44], normal lymphocyte G R content and affinity has been described in asthmatic subjects [44, 45].
In line with observations that patients with A D exhibit a minor derangement of the H P A system shown by imbal- ances of the circadian rhythm of Cortisol [14] and may have a reduced excretion of glucocorticoids [15], the here re- ported alterations in G R binding characteristics on mono- nuclear leukocytes lend further support to the view of A D as being a systemic rather than a mere cutaneous disease.
Future perspectives might be the concomitant analysis of G R behavior in mononuclear leukocytes and cells of both involved and uninvolved skin areas of patients with A D .
Acknowledgements
W e thank M i s s N i s t e l w e c k for e x c e l l e n t t e c h n i c a l assistance. T h e J o h a n n e s a n d F r i e d a M a r o h n Stiftung o f the U n i v e r s i t y o f E r l a n g e n - N u r n b e r g p r o v i d e d f i n a n c i a l s u p p o r t .
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18 O i k a r i n e n A , O i k a r i n e n H , M e e k e r C A , T a n E M L , U i t t o J : G l u c o - c o r t i c o i d r e c e p t o r s i n h u m a n s k i n f i b r o b l a s t s : E v i d e n c e for d o w n - r e g u l a t i o n o f r e c e p t o r by g l u c o c o r t i c o i d h o r m o n e . A c t a D e r m V e n e r e o l ( S t o c k h ) 1987;67:461-468.
19 B a l l a r d P L , B a l l a r d R A : C y t o p l a s m i c r e c e p t o r s for g l u c o c o r t i c o i d s i n l u n g o f the h u m a n fetus a n d n e o n a t e . J C l i n Invest 1974;53:
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20 F u x e K , W i k s t r o m A C , O k r e t S, A g n a t i L F , H a r f s t r a n d A , Y u Z Y , G r a n h o l L , Z o l i M , V a l e W , G u s t a f s s o n J A : M a p p i n g o f g l u c o c o r t i - c o i d r e c e p t o r i m m u n o r e a c t i v e n e u r o n s i n the rat t e l - a n d d i e n - c e p h a l o n u s i n g a m o n o c l o n a l a n t i b o d y against the rat l i v e r g l u c o - c o r t i c o i d r e c e p t o r . E n d o c r i n o l o g y 1985;117:1803-1812.
21 S h i p m a n G F , B l o o m f i e l d C D , G a j l - P e c z a l s k a K , M u n c k A U , S m i t h K A : G l u c o c o r t i c o i d s a n d l y m p h o c y t e s . I I I . Effects o f g l u c o - c o r t i c o i d a d m i n i s t r a t i o n o n l y m p h o c y t e g l u c o c o r t i c o i d r e c e p t o r s . B l o o d 1 9 8 3 ; 6 1 : 1 0 8 6 - 1 0 9 0 .
22 L i p p m a n M , B a r r R : G l u c o c o r t i c o i d r e c e p t o r s i n p u r i f i e d s u b p o p - u l a t i o n s o f h u m a n p e r i p h e r a l b l o o d l y m p h o c y t e s . J I m m u n o l 1977;
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23 B r e n t a n i M M , W a j c h e n b e r g B L , C e s a r F P , M a r t i n s V R : R e g u l a - t i o n o f the g l u c o c o r t i c o i d r e c e p t o r b y g l u c o c o r t i c o i d s i n h u m a n m o n o n u c l e a r l e u c o c y t e s . H o r m R e s 1 9 8 6 ; 2 4 : 9 - 1 7 .
24 S c h l e c h t e J A , G i n s b e r g B H , S h e r m a n B M : R e g u l a t i o n o f the g l u - c o c o r t i c o i d r e c e p t o r i n h u m a n l y m p h o c y t e s . J S t e r o i d B i o c h e m 1 9 8 2 ; 1 6 : 6 9 - 7 4 .
25 R u p p r e c h t R , K o r n h u b e r J , W o d a r z N , G o b e l C , L u g a u e r J , S i n z - ger C , B e c k m a n n H , R i e d e r e r P , M i i l l e r O A : A u t o r e g u l a t i o n o f the g l u c o c o r t i c o i d r e c e p t o r i n m a n : U p - r e g u l a t i o n b y m e t y r a p o n e is a v o i d e d b y d e x a m e t h a s o n e p r e t r e a t m e n t . J N e u r o e n d o c r i n o l 1990;
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26 R u p p r e c h t R , K o r n h u b e r J , W o d a r z N , L u g a u e r J , G o b e l C , H a a c k D , M u l l e r O A , R i e d e r e r P , B e c k m a n n H : D i s t u r b e d g l u c o c o r t i c o i d r e c e p t o r a u t o r e g u l a t i o n a n d c o r t i c o t r o p i n r e s p o n s e to d e x a m e t h a - sone i n d e p r e s s i v e s p r e t r e a t e d w i t h m e t y r a p o n e . B i o l P s y c h i a t r y , i n press.
27 H a n i f i n J M , R a j k a G : D i a g n o s t i c features o f a t o p i c d e r m a t i t i s . A c t a D e r m V e n e r e o l S u p p l ( S t o c k h ) 1 9 8 0 ; 9 2 : 4 4 - 4 7 .
28 C o s t a C , R i l l i e t A , N i c o l e t M , Saurat J H : S c o r i n g a t o p i c d e r m a t i t i s : T h e s i m p l e r the b e t t e r ? A c t a D e r m V e n e r e o l ( S t o c k h ) 1989;69:
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29 B o y u m A : S e p a r a t i o n o f l e u k o c y t e s f r o m b l o o d a n d b o n e m a r r o w . S c a n d J C l i n L a b Invest 1 9 6 8 ; 2 1 : 9 7 - 1 0 7 .
30 S c a t c h a r d G : T h e a t t r a c t i o n o f p r o t e i n s for s m a l l m o l e c u l e s a n d i o n s . A n n N Y A c a d S c i 1 9 4 9 ; 5 1 : 6 6 0 - 6 7 2 .
31 C a r r o l l B J , M a r t i n F I , D a v i s B M : R e s i s t a n c e t o s u p p r e s s i o n by d e x a m e t h a s o n e o f p l a s m a 1 1 - O H C S l e v e l s i n s e v e r e d e p r e s s i v e i l l - ness. B r M e d J 1 9 6 8 ; i i i : 2 8 5 - 2 8 7 .
32 M c P e r s h o n G A : A p r a c t i c a l c o m p u t e r - b a s e d a p p r o a c h to the a n a l - ysis o f r a d i o l i g a n d b i n d i n g e x p e r i m e n t s . C o m p u t P r o g r a m s B i o - m e d 1 9 8 3 ; 1 7 : 1 0 7 - 1 1 4 .
33 B e r g e r M , P i r k e K M , D o e r r P , K r i e g J C , Z e r s s e n D : T h e l i m i t e d u t i l i t y o f the d e x a m e t h a s o n e s u p p r e s s i o n test for the d i a g n o s t i c p r o - cess i n p s y c h i a t r y . B r J P s y c h i a t r y 1 9 8 4 ; 1 4 5 : 3 7 2 - 3 8 2 .
34 R u p p r e c h t R . L e s c h K P : P s y c h o n e u r o e n d o c r i n e r e s e a r c h i n d e p r e s s i o n . I. H o r m o n e l e v e l s o f different n e u r o e n d o c r i n e axes a n d the d e x a m e t h a s o n e s u p p r e s s i o n test. J N e u r a l T r a n s m 1989;
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35 A r c h e r C B : C y c l i c n u c l e o t i d e m e t a b o l i s m i n a t o p i c d e r m a t i t i s . C l i n E x p D e r m a t o l 1 9 8 7 ; 1 2 : 4 2 4 - 4 3 1 .
36 S a a r i n e n U M : T r a n s f e r o f latent a t o p y b y b o n e m a r r o w t r a n s p l a n - t a t i o n ? J C l i n A l l e r g y I m m u n o l 1 9 8 4 ; 7 4 : 1 9 6 - 2 0 0 .
37 T u c k e r J , B a r n e t s o n R , E d e n O B : A t o p y after b o n e m a r r o w trans- p l a n t a t i o n . B r M e d J 1 9 8 5 ; 2 9 0 : 1 1 6 - 1 1 7 .
38 S a u r a t J H : E c z e m a i n p r i m a r y i m m u n e d e f i c i e n c i e s . C l u e to the p a t h o g e n e s i s o f a t o p i c d e r m a t i t i s w i t h s p e c i a l r e f e r e n c e to the W i s - k o t t - A l d r i c h s y n d r o m e . A c t a D e r m V e n e r e o l S u p p l ( S t o c k h ) 1985;
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39 D o n g Y , A r o n s s o n J A , O k r e t S: T h e m e c h a n i s m o f c A M P - i n d u c e d g l u c o c o r t i c o i d r e c e p t o r e x p r e s s i o n . J B i o l C h e m 1989;23:
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40 D a m o n M , R a b i e n M , L o u b a t i e r e J , B l o t m a n F , C r a s t e s de P a u l e t A : G l u c o c o r t i c o i d r e c e p t o r s i n f i b r o b l a s t s f r o m s y n o v i a l tissue.
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41 D i s t e l h o s t C W : R e c e n t insight i n t o the s t r u c t u r e a n d f u n c t i o n o f the g l u c o c o r t i c o i d r e c e p t o r . J L a b C l i n M e d 1 9 8 9 ; 1 1 3 : 4 0 4 - 4 1 2 . 42 T u r n e r B B : T i s s u e d i f f e r e n t i a t i o n i n the u p - r e g u l a t i o n o f g l u c o c o r -
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44 T s a i B S , W a t t G , K o e n s n a d i K , T o w n l e y R G . L y m p h o c y t e g l u c o - c o r t i c o i d r e c e p t o r s i n a s t h m a t i c a n d c o n t r o l subjects. C l i n A l l e r g y 1 9 8 4 ; 1 4 : 3 6 3 - 3 7 1 .
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48 A a s K : C o m m o n i m m u n o c h e m i s t r y i n a t o p i c d e r m a t i t i s a n d b r o n - c h i a l a s t h m a . A c t a D e r m V e n e r e o l S u p p l ( S t o c k h ) 1980;92:64-66.
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