Detection of adrenocortical autoantibodies in Addison's disease with a peroxidase-labelled protein A technique

Adrenocortical autoantibodies (ACA), present in 60-80 percent of patients with idiopathic Addison's disease, are conventionally detected by indirect immunofluorescence (IIF) on frozen sections of adrenal glands. The large-scale use of IIF is limited in part by the need for a fluorescence microscope and the fact that histological sections cannot be stored for long periods of time. To circumvent these restrictions we developed a novel peroxidase-labelled protein A (PLPA) technique for the detection of ACA in patients with Addison's disease and compared the results with those obtained with the classical IIF assay. We studied serum samples from 90 healthy control subjects and 22 patients with Addison's disease, who had been clinically classified into two groups: idiopathic (N = 13) and granulomatous (N = 9). ACA-PLPA were detected in 10/22 (45 percent) patients: 9/13 (69 percent) with the idiopathic form and 1/9 (11 percent) with the granulomatous form, whereas ACA-IIF were detected in 11/22 patients (50 percent): 10/13 (77 percent) with the idiopathic form and 1/9 (11 percent) with the granulomatous form. Twelve of the 13 idiopathic addisonians (92 percent) were positive for either ACA-PLPA or ACA-IIF, but only 7 were positive by both methods. In contrast, none of 90 healthy subjects was found to be positive for ACA. Thus, our study shows that the PLPA-based technique is useful, has technical advantages over the IIF method (by not requiring the use of a fluorescence microscope and by permitting section storage for long periods of time). However, since it is only 60 percent concordant with the ACA-IIF method, it should be considered complementary instead of an alternative method to IIF for the detection of ACA in human sera

Detection of adrenocortical autoantibodies in Addison's disease with a peroxidase-labelled protein A technique.

Adrenocortical autoantibodies (ACA), present in 60-80% of patients with idiopathic Addison's disease, are conventionally detected by indirect immunofluorescence (IIF) on frozen sections of adrenal glands. The large-scale use of IIF is limited in part by the need for a fluorescence microscope and the fact that histological sections cannot be stored for long periods of time. To circumvent these restrictions we developed a novel peroxidase-labelled protein A (PLPA) technique for the detection of ACA in patients with Addison's disease and compared the results with those obtained with the classical IIF assay. We studied serum samples from 90 healthy control subjects and 22 patients with Addison's disease, who had been clinically classified into two groups: idiopathic (N = 13) and granulomatous (N = 9). ACA-PLPA were detected in 10/22 (45%) patients: 9/13 (69%) with the idiopathic form and 1/9 (11%) with the granulomatous form, whereas ACA-IIF were detected in 11/22 patients (50%): 10/13 (77%) with the idiopathic form and 1/9 (11%) with the granulomatous form. Twelve of the 13 idiopathic addisonians (92%) were positive for either ACA-PLPA or ACA-IIF, but only 7 were positive by both methods. In contrast, none of 90 healthy subjects was found to be positive for ACA. Thus, our study shows that the PLPA-based technique is useful, has technical advantages over the IIF method (by not requiring the use of a fluorescence microscope and by permitting section storage for long periods of time). However, since it is only 60% concordant with the ACA-IIF method, it should be considered complementary instead of an alternative method to IIF for the detection of ACA in human sera.

Acute adrenal haemorrhage: diagnosis, treatment and follow-up.

Acute adrenal haemorrhage (AAH) is a rare disorder with different aetiologies. Aiming to discuss this condition, this report deals with four different cases that will be analysed and examined below, each one of them confirmed by biopsy or surgery and followed clinically and radiologically. In these cases it was found that the patients suffered from localized abdominal pain (4/4) and fever (2/4); one patient had adrenal insufficiency due to bilateral massive AAH. Therefore we concluded that AAH is an uncommon condition with variable clinical manifestations.

Systematic evaluation of the adrenocortical function in patients with paracoccidioidomycosis.

Limited cortisol response to ACTH stimulation has been documented in 22 to 48% of patients with paracoccidioidomycosis (PM). Different approaches to interpret the test and inadequate selection of patients preclude an accurate appraisal of the actual incidence of adrenal insufficiency in PM. Rapid cosyntropin (ACTH) stimulation tests were performed in 38 consecutive patients (9 with the localized and 29 with the disseminated form of PM) and 40 normal controls. Subnormal cortisol responses to ACTH (60 minutes post-ACTH values below 455 nmol/l, 95% confidence limits) were found in only 4 patients (14%) with disseminated PM. If a retrospective sample of 6 patients studied previously (in whom tests were indicated due to clinical suspicion of Addison's disease) were included, or if the absolute cortisol increment above baseline was used for interpretation, we would find figures closer to those previously reported (23 and 24%, respectively). These data reflect that non-systematic evaluation or selection of a substandard criterion to interpret the test overestimates the frequency of adrenocortical insufficiency in PM.

[Standardization and clinical applications of the rapid and prolonged ACTH stimulation tests in patients with primary and secondary adrenal insufficiency]. / Padronização e aplicação dos testes rápido e prolongado de estímulo com ACTH em pacientes com insuficiência adrenocortical primária e secundária.

Some methodological aspects of the rapid ACTH stimulation test (RST) and the prolonged ACTH stimulation test (PST) remain heterogeneous, e.g. administration routes, time of the day, duration, interpretation criteria and clinical indications. To standardize both tests we studied the serum cortisol responses to ACTH in 16 normal subjects, 16 patients with Addison's disease (AD), 14 with hypopituitarism (HP) of different etiologies and 13 patients on chronic glucocorticoid (CG) treatment (doses equivalent to 5-20 mg/d of prednisone for at least 6 months). For the RST, 250mcg of Cortrosina Organon were injected as an IV bolus and blood collected before and 60 minutes later, whereas for the PST, 250mcg of Cortrosina-Depot Organon were injected IM, every 8 hours for 3 days, and blood drawn daily between 8 and 9 o'clock AM. The post-ACTH cortisol value was the single parameter chosen to interpret both tests (the absolute or percent increase was of no further value). The 95% lower confidence limits (mean minus 1.64 SD) for the RST and the 3rd day of PST were 19.1 and 49.0mcg/dL, respectively. AD patients did not increase cortisol in response to both the RST (1.1 +/- 1.2 to 1.2 +/- 1.1mcg/dL) or the PST (1.2 +/- 1.3 to 2.0 +/- 1.7mcg/dL). However, whereas 22 out of 27 patients with HP and CG showed a modest cortisol increase to the RST (1.6 +/- 2.3 to 5.6 +/- 4.7 mcg/dL), all had a gradual and nearly normal response on the 3rd day of the PST (1.6 +/- 2.1 to 29.6 +/- 16.6mcg/dL).(ABSTRACT TRUNCATED AT 250 WORDS)

[Importance of percutaneous venous catheterization in the identification of the origin of hyperandrogenism]. / Importância do cateterismo venoso percutâneo na identificação da origem do hiperandrogenismo.

Selective percutaneous venous catheterization (SPVC) was performed to ascertain the source of androgen excess (AE) in 3 virilized women (12, 50 and 58y) in whom hormonal and radiologic investigation (ultrasonography and computerized tomography) was inconclusive. In all cases, elevated testosterone levels remained unsuppressed with dexamethasone, but were adequately suppressed by normal basal DHEA-S and cortisol. SPVC for T and F determinations (and the T/F ratio) were performed before surgery in all cases and during surgery in case 3. A T/F ratio of 152.4 and 71.8 (with an ovary-peripheral vein gradient [OPG] of 637 and 1173 ng/dl) identified the right and left ovary as the source of androgen excess respectively in cases 1 and 2, whereas the T/F ratio (87.4 and 97.2) and OPG (749 and 1162 ng/dl) were increased in both ovaries in patient 3. Thus, lateralization was evident in cases 1 (R ovary Leydig cell tumor) and 2 (L ovary hemangioma and stromal luteinization) whereas bilateral production in case 3 was confirmed by biopsy (PCO). Pre-operative identification of the origin of AE by SPVC is essential in defining proper treatment for patients suspected of harboring a neoplasia. Because it is not widely available nor devoid of complications, SPVC should only be indicated in hirsute/virilized women whose basal plasma testosterone and DHEA-S levels are in the tumoral range, i.e. higher than 150 ng/dl and 700 micrograms/dl, respectively.

Continuous adrenocorticotropin administration in hypopituitarism produces asynchronous increases of deoxycorticosterone and 11-deoxycortisol relative to other reduced zona fasciculata steroids.

Short term suppression of ACTH by dexamethasone effects limited reduction in plasma deoxycorticosterone (DOC) while cortisol levels are almost completely suppressed in normal control subjects. The zona fasciculata (ZF) microsomal cytochrome P-450(21) appeared less influenced by lack of ACTH than mitochondrial cytochrome P-450(11 beta-18). Eleven patients with hypopituitarism were studied to quantitate basal ZF microsomal and mitochondrial derived steroids and their acute and extended responses to ACTH. Basal levels of 11-deoxycortisol (S) and DOC were modestly reduced (70% and 53%, respectively), while other ZF steroids were almost completely absent. Acute and prolonged ACTH treatment amplified the discrepancy in both plasma levels and production rates. DOC and S demonstrated prompt and sustained increases similar to those in normal controls, while cortisol, 18-hydroxydeoxycorticosterone, and corticosterone showed a slow subnormal recovery of steroid production. The preservation of microsomal cytochrome P-450(21) and P-450(17 alpha) to maintain DOC and S levels contrasts the reduced and delayed responses of steroids dependent on mitochondrial cytochrome P-450(11 beta-18), cortisol, corticosterone, and 18-hydroxydeoxycorticosterone. A greater effect of ACTH deficiency on mitochondrial over microsomal cytochrome P-450 activity is demonstrated, and in addition, the possibility is raised that other non-ACTH regulators sustain microsomal cytochrome P-450(21) and P-450(17 alpha) in a setting of reduced ACTH-stimulated factors.