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© Borgis - Postępy Nauk Medycznych 12/2015, s. 816-819
*Anna A. Kasperlik-Załuska1, Lucyna Papierska1, Wojciech Jeske1, Renata Kapuścińska1, Barbara Czarnocka2
Wieloośrodkowe kliniczne i immunologiczne badania u osób z chorobą Addisona w Polsce
Multicenter clinical and immunological study on Addison’s disease in Poland**
1Department of Endocrinology, Centre of Postgraduate Medical Education, Bielański Hospital, Warszawa
Head of Department: prof. Wojciech Zgliczyński, MD, PhD
2Department of Biochemistry and Molecular Biology, Centre of Postgraduate Medical Education, Warszawa
Head of Department: prof. Barbara Czarnocka, MD, PhD
Streszczenie
Wstęp. Pierwotna niedoczynność nadnerczy – choroba Addisona (AD), jest rzadką chorobą rozwijającą się na tle procesu autoimmunologicznego. Autoimmunizacja skierowana przeciwko 21-hydroksylazie, enzymowi uczestniczącemu w steroidogenezie, jest najczęstszym czynnikiem patogenetycznym w tej chorobie.
Cel pracy. Celem pracy było zbadanie częstości występowania przeciwciał przeciwko 21-hydroksylazie oraz częstości współistnienia innych chorób autoimmunologicznych u pacjentów z pierwotną niedoczynnością nadnerczy w Polsce.
Materiał i metody. Zrekrutowano 212 pacjentów (w tym 160 kobiet ), z udokumentowaną badaniami hormonalnymi chorobą Addisona, w wieku 20-84 lat, obserwowanych przez 1-52 lat. Wykonano badanie kliniczne oraz oznaczenie przeciwciał przeciwko 21-hydroksylazie (21-OH) i przeciwciał tarczycowych (aTg i aTPO). Wyniki badań uzyskane w 2012 roku porównano z wynikami badania przeciwciał nadnerczowych wykonanych w 2000 roku (z 62 pacjentów 39 miało wówczas wynik pozytywny i 30 ponownie poddano badaniu w 2012 roku).
Wyniki. Z 212 pacjentów poddanych badaniu w 2012 roku obecność przeciwciał przeciwko 21-OH wykryto u 114 osób (54%). Wśród 30 chorych badanych pierwotnie w 2000 roku 27 miało te przeciwciała, natomiast ponowne badanie 12 lat później wykazało pozytywny wynik jedynie u 13 z nich. Wśród innych immunologicznych odchyleń najczęstsza była obecność przeciwciał tarczycowych, a najczęstszą towarzyszącą chorobą autoimmunologiczną – niedoczynność tarczycy.
Wnioski. 1. Częstość występowania przeciwciał przeciwko 21-hydroksylazie w grupie polskich pacjentów z chorobą Addisona (54%) jest niższa, aniżeli w materiale pacjentów skandynawskich, czego przypuszczalną przyczyną może być dłuższy okres obserwacji w naszej grupie chorych. 2. Konwersja wyniku dodatniego do ujemnego zaobserwowana u części pacjentów badanych ponownie po wielu latach może przemawiać za takim wytłumaczeniem.
Summary
Introduction. Primary adrenal insufficiency – Addison’s disease (AD) is a rare disease, developing mainly due to autoimmune processes. Autoimmunity against 21-hydroxylase, an enzyme participating in steroidogenesis, is the most frequent pathogenic factor in AD.
Aim. Our study aimed at evaluating the incidence of 21-hydroxylase (21-OH) antibodies and presence of other immunological disorders in patients with primary adrenal insufficiency.
Material and methods. 212 patients (160 women) with hormonally documented Addison’s disease, aged 20-84 years, observed for 1-52 years. Clinical examination, 21-hydroxylase and thyroid autoantibodies investigations. The results obtained in 2012 were compared with the results of adrenal autoantibodies measured in 2000 (then of 62 patients, adrenal antibodies were detected in 39 persons and 30 of them were investigated again in 2012).
Results. In 212 patients examined in 2012 antibodies to 21-OH were present in 114 cases (54%). Among 30 patients examined in 2000, 27 cases showed 21-OH positivity, whereas twelve years later antibodies were detected in only 13 cases. Presence of thyroid antibodies was the most frequent immunological finding, while hypothyroidism was the most frequent autoimmune disease.
Conclusions. 1. The incidence of positive results of 21-OH antibodies in 2012 = 54% appeared to be lower than in Scandinavian cohorts, probably due to long term observation in Poland. 2. A conversion from a positive immunological phenotype to a negative one during 12 years period of observation seems to confirm this suggestion.
Introduction
Primary adrenal insufficiency (Addison’s disease – AD) is a rare disease, developing mainly due to autoimmune processes. Autoimmunity against 21-hydroxylase, an enzyme participating in steroidogenesis, is the most frequent pathogenic factor in AD. Clinical, immunological and genetic investigations have been conducted in the european project, Euradrenal, to study more deeply the pathogenesis of AD, its natural course and current possibilities of the therapy improvement. Clinical experience of the members of the Steering Committee resulted in an Expert Consensus Statement concerning diagnosis, treatment and follow-up of patients with AD (1).
At our department a tradition of addisonian patients’ diagnosis, treatment and follow up, initiated in the sixties years of the 20th century, have been continued till now. Our observations were described in 1991 (2) and 2010 (3). In the first series of AD patients, including 180 cases, tuberculosis was diagnosed in 52 persons (29%). In the group of 138 patients, registered between 1990 and 2008, there were only 3 patients with tuberculous destruction of the adrenals.
Within Polish part of the Euradrenal program, combined groups of patients with autoimmune AD (AAD) observed in our department and 7 other endocrinological centers in Poland1 have been studied.
Aim
The aim of this work was evaluation of the presence of 21-hydroxylase and thyroid (aTPO, aTg) autoantibodies as well as evaluation of the coincidence of other autoimmunological disorders.
The current results of 21-OH antibodies measurements were compared with adrenal autoantibodies (abs) determinations performed in 2000. Of 62 patients investigated initially, adrenal antibodies were positive in 39 cases. Thirty of them (five with associated primary ovarian failure – POF) were included in both studies. Such comparison enabled immunological follow-up of the autoimmune processes in Addison’s disease.
Material and methods
Our material included 212 patients, 160 women and 52 men, aged 20-84 years (at the time of the study), observed for 1-52 years. In this number, there were 148 patients from our department, and 64 patients from 7 other endocrinological centers. Diagnostic methods: clinical examination and hormonal determinations, which included: serum cortisol, DHEA-S, fT4, TSH, FSH, estradiol, testosterone, PTH (if necessary), plasma ACTH, urinary 17-hydroxycortisteroids (17-OHCS) or free cortisol and 1-24ACTH stimulation test.
Inclusion criteria: characteristic clinical features (4) and hormonal measurements – low serum cortisol at 9.00 hours, below 209 nmol/l (7.5 μg/dl) combined with high plasma ACTH concentration, over 13.2 pmol/l (60 pg/ml), decreased 17-OHCS urinary excretion, below 6.1 μmol/24 hours (2.2 mg/24 h), low peak serum cortisol level following 1-24ACTH (Synacthen) 0.25 mg i.m. or i.v. injection (1). Normal values: serum cortisol – 209 to 692 nmol/l (7.5-25 μg/dl), morning plasma ACTH – 3.3 to 13.2 pmol/l (15-60 pg/ml), urinary 17-OHCS – 6.1 to 19.3 μmol/24 h (2.2-7.0 mg/24 h).
Immunological studies: serum 21-hydroxylase (21-OH) autoantibodies (abs) determinations, thyroid autoantibodies (aTPO and aTg) measurements and a search for clinical data of other autoimmune disorders. The antibodies against 21-OH at first were determined by a method based on the in vitro transcribed and translated protein as described by Ekwall et al. (5). The upper normal limit of each antibody index was established as the mean value of negative controls plus three standard deviations and values above this cut-off indicated the presence of adrenal antibodies. Additionally, in 5 patients with POF, antibodies against 17-hydroxylase and against side-chain cleavage enzyme were measured (5).
Analytical assays (if necessary): serum glucose, HbA1c levels, calcium and phosphorus, serum and urinary values, sodium and potassium levels.
Results
The 21-OH antibody assays performed in 2012 detected positive responses in 114 out of 212 cases (54%). The time of observation ranged from 1 to 52 years; in 96 cases it exceeded 10 years. A comparison performed in the group of 27 patients with a significant titer of 21-OH autoantibodies detected in 2000 showed that in 2012 positive results remained only in 13 ones. The remaining 14 patients, which were positive in the first assessment became 21-OH antibodies (21-OH abs) negative in the last one. Such a negative conversion has been illustrated in the table 1. Among 5 patients with POF, the 17-OH abs and SCC antibodies were present in four, while negative in one patient (No 110) with premature menopause noted 48 years ago. Only in one patient with POF (No 115) positive 21-OH abs remained in 2012. In one patient (No 94), only significant anti-SCC abs were noted (without 21-OH abs). All patients presented in table 1 manifested other autoimmune disorders, the most frequently hypothyroidism, diabetes mellitus, vitiligo and POF.
Table 1. Immunological data in patients with Addison’s disease, studied in 2000 and 2012 (only patients with positive 21-OH or SCC antibodies in 2000).
PatientsAutoimmune disordersAdrenal antibodies 2000Adrenal antibodies 2012Negative conversion
No#SexAgeAD yrsTypes of disorders21-OH17-OHSCC21-OHYes or not
6F6017T-, DM, V+  Yes
19F6820T-, DM+  Yes
27F5111T++  Yes
39F7711T-, A+  Yes
54F6023T-, POF+++Yes
65F6025T-, V+  Yes
68F4110T+, T-+  Yes
74F8537T-+  Yes
90M5519T-+  Yes
94F5926T-, POF++
110F7611T-, A, V, POF+  Yes
115F5423T-, V, POF++++Not
118F3819T+, T-, POF+++Yes
125F5515T-, DM, V+  Yes
127F4013DM, A+  Yes
132F5917T-+  Yes
# – number on the list from 2012; T- – hyperthyroidism; DM – diabetes mellitus; A – anemia Addison-Biermeri; V – vitiligo; POF – primary ovarian failure
Association of other autoimmune disorders was characteristic for majority of the addisonian patients under study. Thyroid abs, mainly aTPO, present in 180 patients (85%) have been the most frequent autoimmune finding. Table 2 shows the most frequent autoimmune diseases, with timing of their appearance. Thyroid diseases, hypothyroidism and thyrotoxicosis, were the most frequent disorders. In 25 patients thyrotoxicosis was followed by hypothyreosis, mainly in the cases treated by radioiodine. In single cases other autoimmune disorders were noted: rheumatoid arthritis, coeliac disease, Duhring’s disease, chronic hepatitis, pericarditis. The analysis of timing of autoimmune disorders associated with AD did not show any significant differences, except of hypothyroidism, which followed AD in most cases. In 23 cases two autoimmune disorders accompanied AD, in 11 cases – three, while in two patients four other autoimmune disorders were associated with AD.
Table 2. Autoimmune disorders associated with Addison’s disease in 212 patients under study.
Type of disorderNumber%Appearance
BeforeSimult*After
Hypothyroidism10650262555
Hyperthorodism3818131213
T1 Diabetes Mellitus241110311
Vitiligo23111058
Pernicious anemia147554
Premature menopause**105424
Alopecia4222
Moniliasis313
Hypoparathyroidism2~12
*simultaneously
**in women only
Discussion
Most of the studies on the incidence of 21-OH antibodies in AD aims on diagnosis of its autoimmune form (6-8). Our investigations, including patients with long-time duration of AAD, aimed mainly on follow-up through the physiopathology of this disease and age-related modifications of the autoimmune reactivity. In our group there were 96 patients who have been observed at least for 10 years, which could influence the results of our studies.
Owing to the possibility to compare the same group of patients studied in 2000 and 2012 we were able to expose a conversion of a positive autoimmune phenotype of AAD in the first study to a negative one in the last series of immunological investigation. In 2000, in the group of 30 patients under study, 27 ones were positive for 21-OH abs while in 2012 only 13 of them remained positive. In the remaining group of 14 patients a negative immune phenotype appeared, interestingly – also in the women with high titer of 21-OH abs in 2000 (Nos 27, 68 and 132 in table 1). This observation suggests that an intensive autoimmune process could accelerate disappearance of autoimmune responses.
It is well known from the literature that in diabetes mellitus most of the islet-cell cytoplasmic autoantibodies (ICA) disappear within initial 10 years; only in about 5% of cases the antibodies remain in the circulation (9-11). We suppose that in our patients in whom a negative conversion of immune phenotype was noted a similar mechanism took place. The destruction of adrenal tissue, due to autoimmunity, deprives the organism of the tissue antigens and the possibility to produce anti-adrenal autoantibodies. Diabetes mellitus, the most frequent endocrine disease of autoimmune origin could be a model of physiopathology of autoimmune processes.
An exception in our material, it was a 74-yrs old woman, observed for AAD for 52 years, who remained 21-OH abs positive. During that time she temporary suffered of hyperthyroidism. Thus immune reactivity is not age-related in all cases. Her sister’s son (observed 18 years for AAD, and for hypothyroidism – 17 years) appeared to be 21-OH abs negative.
A selective production of anti-SCC abs only might explain a lack of 21-OH abs. In one our patient, with AAD and POF of 26 years duration (No 94 in table 1).
The relatively lower percentage of 21-OH abs presence in AD (54% in our last study), in comparison with the Scandinavian statistics (8) seems to be due to long duration of our observations. Including in this assessment 14 patients who were positive for 21-OH autoimmunity in 2000, and negative in 2012, the total number of 21-OH abs presence would increase up to 128/212 = 60%, which differs still evidently from the Scandinavian findings, probably due to evidently longer time of duration of AD in our series of patients.
Prevalence of thyroid autoimmunity in AAD has been observed by many authors, including our own experience (1-3, 6-8, 12). In our observations an increasing tendency to thyroid autoimmunity development in AAD was found (13). Diabetes mellitus and vitiligo took ex aequo the second place on our list of the incidence frequency (tab. 2), however only vitiligo preceeded AD even for 18 years. Pernicious anemia was the next on this list, usually associated with atrophic gastritis. Moniliasis, characteristic for APS1, was present in all three patients with this syndrome, while hypoparathyroidism only in two patients. A young man with AD, brother of a female patient with AD, hypoparathyroidism, T1DM and pernicious anemia, had only severe moniliasis. Mothers of the APS1 patients, included in the study, appeared to be 21-OH abs negative. In polyglandular insufficiency syndromes the associated disorders have been well tolerated by the patients except of T1DM, which worsened their self-estimation.
Conclusions
1. The incidence of positive results of 21-OH antibodies in 2012 = 54% appeared to be lower than in Scandinavian cohorts, probably due to long term observation in Poland.
2. A conversion from a positive immunological phenotype to a negative one during 12 years period of observation seems to confirm this suggestion.

1Participants of the Euradrenal study from 7 endocrinological centers in Poland: Department of Endocrinology, Metabolic and Internal Diseases, Pomeranian Medical University in Szczecin: Syrenicz Anhelli; Department of Internal Medicine, Subdivision Endocrinology, Provincial Hospital in Opole: Łomna-Bogdanov Elżbieta, Truch Anna, Kowalska Beata, Baran Bogusława, Tabora-Ciszewska Joanna, Sicińska-Werner Teresa; Military Medical Academy, University Teaching Hospital – Central Veterans Hospital in Łódź, Department of Endocrinology: Siejka Agnieszka, Gruszka Anna, Salamon-Krekora Ewa, Zieleniewski Wojciech; Department of Endocrinology, Diabetology and Internal Medicine, Medical University of Białystok: Siewko Katarzyna; Department of Pathophysiology and Endocrinology, Silesian Medical University: Kos-Kudła Beata; The Clinical Department of Endocrinology and Nuclear Medicine, Holycross Cancer Center in Kielce: Mikina Estera; Wrocław Medical University, Department of Endocrinology, Diabetology and Isotope Therapy: Lenarcik Agnieszka, Jawiarczyk Aleksandra, Brona Anna, Stachowska Barbara, Syrycka Joanna.
**This work was supported by EU project FP7 “Euradrenal” and CMKP grant 501-1-08-11/10-14. Authors express their thanks to Professor Olle Kämpe for measurement of adrenal antibodies and thanks to the listed contributing Polish endocrinologists for recruiting c. 30% of the total number of the studied patients.
Piśmiennictwo
1. Husebye ES, Allolio B, Arlt W et al.: Consensus statement on the diagnosis, treatment and follow-up of patients with primary adrenal insufficiency. Journal of Internal Medicine 2014; 275: 104-115.
2. Kasperlik-Załuska AA, Migdalska B, Czarnocka B et al.: Association of Addison’s disease with autoimmune disorders – a long-term observation of 180 patients. Postgraduate Medical Journal 1991; 67: 984-987.
3. Kasperlik-Załuska AA, Czarnocka B, Jeske W, Papierska L: Addison’s disease revisited in Poland: Year 2008 versus year 1990. Autoimmune Dis 2010 Jun 6; 2010:731834. doi: 10.4061/2010/731834.
4. Arlt W, Allolio B: Adrenal insufficiency. The Lancet 2003; 361: 1881-1893.
5. Ekwall O, Hedstrand H, Grimelius L et al.: Identification of tryptophan hydroxylase as an intestinal autoantigen. Lancet 1998; 352: 279-283.
6. Söderbergh A, Winquist O, Norheim I et al.: Adrenal autoantibodies and organ-specific autoimmunity in patients with Addison’s disease. Clinical Endocrinology 1996; 45: 453-460.
7. Betterle C, Dal Pra C, Mantero F, Zanchetta R: Autoimmune adrenal insufficiency and autoimmune polyendocrine syndromes: autoantibodies, autoantigens, and their applicability in diagnosis and disease prediction. Endocrine Reviews 2002; 23: 327-364.
8. Erichsen MM, Løvas K, Skinningsrud B et al.: Clinical, immunological, and genetic features of autoimmune primary adrenal insufficiency: observations from a Norvegian Registry. Journal of Clinical Endocrinology and Metabolism 2009; 94: 4882-4890.
9. Winter WE, Harris N, Schatz D: Immunological markers in the diagnosis and prediction of autoimmune type 1a diabetes. Clinical Diabetes 2002; 20: 183-191.
10. Pihoker C, Gilliam LK, Hampe CS, Lernmark A: Autoantibodies in diabetes. Diabetes 2005; 54 (suppl. 2): 52-61.
11. Winter WE, Schatz D: Autoimmune markers in diabetes. Clinical Chemistry 2011; 57: 168-175.
12. Kasperlik-Załuska AA, Czarnocka B, Czech W: High prevalence of thyroid autoimmunity in idiopathic Addison’s disease. Autoimmunity 1994; 18: 213-216.
13. Kasperlik-Załuska AA, Czarnocka B, Jeske W: Addison’s disease – E-letter. Journal of Clinical Endocrinology and Metabolism 2010, 14 Jan.
otrzymano: 2015-10-30
zaakceptowano do druku: 2015-11-23

Adres do korespondencji:
*Anna A. Kasperlik-Załuska
Department of Endocrinology Centre of Postgraduate Medical Education Bielański Hospital
ul. Cegłowska 80, 01-809 Warszawa
tel. +48 (22) 834-31-31
anna.kasperlik@gmail.com

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