Richard I. Dorin

Diagnosis of adrenal insufficiency

Adrenal insufficiency is an uncommon clinical disorder that results from an inadequate basal or stress level of plasma cortisol. It is important to diagnose adrenal insufficiency because the disorder may be fatal if left unrecognized or untreated. With diagnosis and appropriate adrenocortical hormone replacement, normal quality of life and longevity can be achieved. The presentation of adrenal insufficiency may be insidious and thus difficult to recognize. Once suspected, however, the definitive diagnosis can be confirmed by laboratory evaluation of adrenocortical function. Although many different tests for adrenal insufficiency have been developed, few have been adequately studied and many are inconvenient for testing in the outpatient clinical setting. By contrast, the cosyntropin stimulation test is widely used in many different clinical settings and is easy to perform. In addition, data on test performance in various clinical settings are plentiful. The cosyntropin stimulation test has therefore emerged as the initial test used to evaluate patients for both primary and secondary adrenal insufficiency. Methods We reviewed all English-language studies in humans identified in the MEDLINE database (1966 to 2002) through the Ovid search service. Search terms were adrenal gland hypofunction restricted to diagnosis. For the normal response to high-dose cosyntropin, we selected studies with 10 or more participants. For the diagnosis of primary adrenal insufficiency, we selected studies with 5 or more participants. For evaluation of the sensitivity and specificity of cosyntropin tests in secondary adrenal insufficiency, we selected only studies that stratified all participants with suspected adrenal insufficiency by integrated tests of adrenal function (insulin tolerance or metyrapone tests). Summary receiver-operating characteristic (ROC) curves were developed from sensitivity and specificity values derived from individual studies, as described by Littenberg, Moses, and colleagues (1, 2) (see the Appendix for detailed formulas). Summary ROC curves were compared by using area under the curves (AUCs), as described by Walter (3). For our data sets, we verify the condition (B 0; see the Appendix) that yields explicit formulas for AUC and its CI for the summary ROC curves. The slope parameter (B) did not differ significantly from 0 for all data sets used to generate summary ROC curves. We compared ROC curves for data paired by individual participants using likelihood methods with a program (ROCKIT 0.9B) developed by Metz and colleagues (4) (available at www-radiology.uchicago.edu/cgi-bin/software.cgi). The funding source had no role in the design, conduct, or reporting of the study or in the decision to submit the manuscript for publication. Data Synthesis High-Dose Cosyntropin Stimulation Test The standard cosyntropin test is performed by administering one ampule (250 g) of cosyntropin intramuscularly or intravenously and measuring serum or plasma cortisol levels 30 to 60 minutes later. With a normal (negative) test result, the serum cortisol level after cosyntropin stimulation is generally greater than 500 nmol/L. A subnormal cortisol response (<500 nmol/L) is defined as a positive test result and indicates an increased probability of either primary or secondary adrenal insufficiency. The cosyntropin test may be performed at any time of the day. In patients with suspected adrenal insufficiency, a basal plasma cortisol level is not usually necessary because neither the absolute nor the percentage change from the basal level is useful as a diagnostic criterion for the cosyntropin test (5). However, in the absence of corticosteroid-binding globulin deficiency, an unstimulated serum cortisol level, determined between 6:00 and 8:00 a.m., may be helpful because a level less than 80 nmol/L strongly suggests adrenal insufficiency (5). Normal Response to the High-Dose Cosyntropin Test In healthy persons without evidence of adrenal insufficiency, serum cortisol response 30 or 60 minutes after 250 g of cosyntropin is administered intramuscularly or intravenously has been studied extensively (6-22). The responses to intramuscular and intravenous injections are similar, and the responses among normal persons vary. In 10 studies that included a total of 288 participants and that reported the entire range of postcosyntropin serum cortisol levels, the levels ranged from 415 to 2200 nmol/L (9, 10, 12-15, 17, 19-21). The broad range of normal responses to cosyntropin stimulation reflects various factors, including differences in hypothalamicpituitaryadrenal axis set point, serum corticosteroid-binding globulin level, stress level, body composition, time of testing, and performance characteristics of the cortisol assay used. In one detailed study of 100 healthy persons, the distribution curves of serum cortisol levels obtained 30 and 60 minutes after a 250-g intramuscular injection of cosyntropin displayed a non-Gaussian configuration for each of four separate cortisol assays, with the distribution skewed to the right toward higher cortisol levels (22). The 5th percentile lower cortisol cutoff limit for these four assays ranged from 510 to 615 nmol/L at 30 minutes and from 620 to 675 nmol/L at 60 minutes. Other studies also show increases in the cortisol response at 60 minutes compared with 30 minutes (16, 18, 20). In 11 studies involving 340 healthy participants, the data presented as the mean 2 SDs show lower limits ranging from 390 to 620 nmol/L at 30 minutes (6-10, 16-20) and from 500 to 725 nmol/L at 60 minutes (11, 16, 18, 20). Because the distribution curve is non-Gaussian, no conclusion can be drawn from these studies about the percentage of healthy persons with serum cortisol levels less than the lower cutoff limit. The studies described show that an appreciable number of normal persons will have a postcosyntropin cortisol level less than a cutoff limit of 500 nmol/L. However, none of the 288 participants in the 10 studies described earlier (in which the entire range of cortisol responses was reported) had a cortisol level less than 415 nmol/L. Diagnosis of Primary Adrenal Insufficiency Primary adrenal insufficiency (often called Addison disease) is an uncommon disorder that often presents with a slowly progressive increase in nonspecific symptoms. The prevalence of this disorder in the community is approximately 100 cases per 1 million people (23-26); the incidence is 5 cases per year per 1 million people (26). The prevalence of primary adrenal insufficiency is higher (although not precisely known) in persons with HIV disease, family histories of adrenoleukodystrophy, autoimmune endocrine disorders, metastatic cancer, and granulomatous disease. The prevalence among persons with nonspecific symptoms, such as tiredness, fatigue, weakness, listlessness, weight loss, nausea, and anorexia, is not known. More specific symptoms, such as unexplained darkening of the skin, orthostatic dizziness, and salt-craving, may not be among presenting symptoms. Cosyntropin Stimulation Tests in Primary Adrenal Insufficiency Table 1 summarizes the results of 8 studies in which 122 patients with primary adrenal insufficiency and controls were given 250 g of cosyntropin intravenously or intramuscularly and the serum cortisol levels were measured 30 or 60 minutes later. None of the patients in these studies underwent consecutive prospective evaluation for adrenal insufficiency; rather, they were selected for study either because previous evaluation showed that they had typical Addison disease (13, 14, 20, 27-29) or because their cosyntropin tests were compared with historical controls in retrospective surveys (23, 30). Controls in these studies varied from healthy volunteers (13, 14, 23) to participants with nonendocrine illness (14, 27) or suspected adrenal insufficiency (29). Thus, case-patients and controls were not recruited from the same setting. In general, the case-patients with Addison disease in these studies were selected on the basis of typical clinical and nonendocrine laboratory criteria, such as hyperkalemia, supplemented in many cases with elevated plasma adrenocorticotropic hormone (ACTH) levels and low urine steroid responses to intravenous ACTH infusions. In several retrospective analyses using historical controls, cosyntropin tests may have contributed to the diagnosis of Addison disease, but several patients with Addison disease in each of these surveys had normal cosyntropin test results. None of the studies indicated that patients with borderline cosyntropin test results were selectively excluded. However, it is clear that the cases of Addison disease selected in these studies were more advanced and easily recognized by well-established clinical and laboratory criteria. Thus, in most cases in these studies, the diagnosis of Addison disease was based on clinical evidence supported by serum electrolyte, plasma ACTH, and urine steroid levels. Cosyntropin tests were then performed in these patients, and the results were interpreted independently of the original diagnostic criteria. Table 1. The 250-g Cosyntropin Stimulation Test in Patients with Primary Adrenal Insufficiency For the summary ROC curve, which is based on four of the studies in Table 1 (14, 20, 27, 29), the point on the summary ROC where sensitivity and specificity are equal was 96.5% (95% CI, 94.5% to 98.5%) for the diagnosis of primary adrenal insufficiency. When specificity is set at 95%, this summary ROC curve yields a sensitivity of 97.5% (CI, 95% to 100%), with a corresponding positive likelihood ratio of 19.5 (CI, 19.0 to 20.0) and a negative likelihood ratio of 0.026 (CI, 0 to 0.6). The AUC for this summary ROC curve was 0.99 (CI, 0.985 to 1.000), indicating excellent test discrimination. As a result of the selection bias in these studies toward patients with severe Addison disease, the cosyntropin test performance characteristics derived from Table 1 are most

Dexras1/AGS-1 Inhibits Signal Transduction from the Gi-coupled Formyl Peptide Receptor to Erk-1/2 MAP Kinases

Dexras1 is a novel GTP-binding protein (G protein) that was recently discovered on the basis of rapid mRNA up-regulation by glucocorticoids in murine AtT-20 corticotroph cells and in several primary tissues. The human homologue of Dexras1, termed activator of G protein signaling-1 (AGS-1), has been reported to stimulate signaling by Gi heterotrimeric G proteins independently of receptor activation. The effects of Dexras1/AGS-1 on receptor-initiated signaling by Gi have not been examined. Here we report that Dexras1 inhibits ligand-dependent signaling by the Gi-coupled N-formyl peptide receptor (FPR). Dexras1 and FPR were transiently co-expressed in both COS-7 and HEK-293 cells. Activation of FPR by ligand (N-formyl-methionine-leucine-phenylalanine (f-MLF)) caused phosphorylation of endogenous Erk-1/2 that was reduced by co-expression of Dexras1. Direct effects of Dexras1 on the activity of co-expressed, epitope-tagged Erk-2 (hemagglutinin (HA)-Erk-2) were measured by immune complex in vitro kinase assay. Expression of Dexras1 alone resulted in a 1.9- to 4.9-fold increase in HA-Erk-2 activity; expression of the unliganded FPR alone resulted in a 6.2- to 8.1-fold increase in HA-Erk-2 activity. Stimulation of FPR by f-MLF produced a further 8- to 10-fold increase in HA-Erk-2 activity over the basal (non-ligand-stimulated) state, and this ligand-dependent activity was attenuated at the time points of maximal activity by co-expression of Dexras1 (reduced 31 ± 6.8% in COS-7 at 10 min and 86 ± 9.2% in HEK-293 at 5 min, p < 0.01 for each). Expression of Dexras1 did not influence protein expression of FPR or Erk, suggesting that the inhibitory effects of Dexras1 reflect a functional alteration in the signaling cascade from FPR to Erk. Expression of Dexras1 had no effect on expression of Giα species, but significantly impaired pertussis toxin-catalyzed ADP-ribosylation of membrane-associated Giα. Expression of Dexras1 also significantly decreasedin vitro binding of GTPγS in f-MLF-stimulated membranes of cells co-transfected with FPR. These data suggest that Dexras1 inhibits signal transduction from FPR to Erk-1/2 through an effect that is very proximal to receptor-Gi coupling. While Dexras1 weakly activates Erk in the resting state, more potent effects are evident in the modulation of ligand-stimulated receptor signal transduction, where Dexras1 functions as an inhibitor rather than activator of the Erk mitogen-activated protein kinase signaling cascade.

To Breathe, Perchance to Sleep: Sleep-Disordered Breathing and Chronic Insomnia Among Trauma Survivors

Standard psychiatric classification (DSM-IV-TR) traditionally attributes post-traumatic sleep disturbance to a secondary or symptomatic feature of a primary psychiatric disorder. The DSM-IV-TR paradigm, however, has not been validated with objective sleep assessment technology, incorporated nosological constructs from the field of sleep disorders medicine, or adequately addressed the potential for post-traumatic stress disorder (PTSD) sleep problems to manifest as primary, physical disorders, requiring independent medical assessments and therapies. This paradigm may limit understanding of sleep problems in PTSD by promulgating such terms as “insomnia related to another mental disorder,” a.k.a. “psychiatric insomnia.” Emerging evidence invites a broader comorbidity perspective, based on recent findings that post-traumatic sleep disturbance frequently manifests with the combination of insomnia and a higher-than-expected prevalence of sleep-disordered breathing (SDB). In this model of complex sleep disturbance, the underlying sleep pathophysiology interacts with PTSD and related psychiatric distress; and this relationship appears very important as demonstrated by improvement in insomnia, nightmares, and post-traumatic stress with successful SDB treatment, independent of psychiatric interventions. Continuous positive airway pressure treatment in PTSD patients with SDB reduced electroencephalographic arousals and sleep fragmentation, which are usually attributed to central nervous system or psychophysiological processes. Related findings and clinical experience suggest that other types of chronic insomnia may also be related to SDB. We hypothesize that an arousal-based mechanism, perhaps initiated by post-traumatic stress and/or chronic insomnia, may promote the development of SDB in a trauma survivor and perhaps other patients with chronic insomnia. We discuss potential neurohormonal pathways and neuroanatomatical sites that may be involved in this proposed interaction between insomnia and SDB.

Regulation of aldose reductase gene expression in renal cortex and medulla of rats

SummaryA role for aldose reductase-mediated production of polyol in the aetiology of diabetic nephropathy has been supported by both animal and clinical studies. In the renal medulla, the rate of polyol production is influenced in part by regulated changes in the level of aldose reductase gene expression. However, little is known about the expression of aldose reductase in the renal cortex. In this study, we evaluated the regulation of aldose reductase gene expression in the renal cortex and medulla in response to galactose feeding. Four groups of rats (n=6) were treated for 9 weeks with control or galactose diet in the presence or absence of sorbinil, an aldose reductase inhibitor. In the renal medulla, galactose treatment produced a significant (p<0.01) decrease in aldose reductase mRNA, to approximately 10% of control levels. Coadministration of sorbinil partially prevented the effect of galactose feeding on medullary aldose reductase mRNA (to 43% of control). Under basal conditions, the concentration of aldose reductase mRNA in the cortex was only 1% that of the renal medulla. Galactose feeding significantly reduced cortical aldose reductase mRNA by 29% relative to control (p<0.01), and this was completely reversed by addition of sorbinil. Sorbinil administration to rats fed a control diet also decreased aldose reductase expression in the renal medulla and cortex. These results demonstrate that galactose feeding results in dynamic, polyol-dependent regulation of aldose reductase gene expression in the renal cortex as well as the medulla. We also describe a method for quantitative analysis of low abundance renal cortical mRNA. [Diabetologia (1995) 38: 46–54]

Transcriptional regulation of human corticotropin releasing factor gene expression by cyclic adenosine 3',5'-monophosphate: differential effects at proximal and distal promoter elements

cAMP participates in the regulation of endogenous hypothalamic and placental CRF by increasing levels of both peptide secretion and mRNA expression. In previous studies we have shown that stimulation of the protein kinase A-dependent pathway by cAMP analogues or forskolin produced a dose-dependent increase in levels of CRF mRNA when the intact hCRF gene was stably transfected and expressed in the mouse corticotroph AtT20 cell line. In the present study, we explored the mechanism of the cAMP-dependent increase in CRF gene expression in the stably transfected AtT20 cell line using pharmacologic, slot-blot, and RNase mapping methodologies. Following incubation with cAMP, there was a rapid increase in CRF mRNA which was completely blocked by pre-treatment with actinomycin D, an inhibitor of transcription. Cycloheximide, an inhibitor of protein synthesis, produced an independent increase in CRF mRNA, but did not change the relative induction of CRF mRNA produced by cAMP. Solution hybridization studies using intron- and exon-specific hCRF probes demonstrated a rapid rise in nuclear CRF hnRNA, which was apparent within 15 min of cAMP incubation and preceded the rise in cytoplasmic CRF mRNA. RNase mapping studies demonstrated that CRF transcription was initiated at discrete promoter sites in CRF-AtT20 cells, and that this pattern of promoter utilization was similar to that observed in mRNA derived from sites of endogenous CRF expression, human placenta and human hepatoma NPLC cell line. Treatment with cAMP selectively increased CRF mRNA transcripts initiated at the proximal promoter site, but had little or no effect on transcripts initiated at the distal promoters. We conclude that cAMP effects on CRF gene expression occur rapidly, do not require new protein synthesis, and are initiated within the nuclear compartment, consistent with a direct effect on CRF gene transcription. This effect is mediated predominantly through the proximal promoter element, while more distal promoters are less sensitive to transcriptional activation by cAMP.

High serum levels of free cortisol indicate severity of cirrhosis in hemodynamically stable patients

Background and Aim:  We investigated: (i) the association between severity of cirrhosis and serum levels of free cortisol (SFC) and total cortisol (STC), measured before and 30 min after (T30) the low‐dose 1‐µg short synacthen test (LD‐SST); and (ii) the prognostic value of SFC and STC.

Comparison of the functional properties of three different truncated thyroid hormone receptors identified in subjects with resistance to thyroid hormone.

The tau4 domain in the extreme carboxyl (C) terminal region of thyroid hormone receptor (TR) is important to transactivation. We identified three truncated TRbeta1s with 11 (F451X), 13 (E449X) and 16 (C446X) amino acid deletions within this domain in subjects with resistance to thyroid hormone (RTH). F451X and C446X were found in a 6-year-old Japanese girl and a 31-year-old American male, respectively, who had both severe mental retardation. E449X was identified in a 16-year-old Japanese boy with no remarkable clinical symptoms except for goiter. Transient expression study revealed that all three mutants had negligible T3 binding and transcriptional activities. Each mutant TRbeta1 exhibited not only very strong dominant negative activity against wild TRbeta1, but also marked silencing activity. Interestingly, the dominant negative activity and silencing activity were significantly stronger in F451X than in E449X and C446X (P < 0.05). Gel-shift experiments revealed no apparent differences in homodimer formations of wild-type or mutant TRbeta1 proteins and in heterodimer formations with retinoid X receptor (RXR). These observations indicate that the tau4 domain affects diverse TR functions, and that the region between 11 and 13 C-terminal amino acids influences ligand-independent TR functions, including dominant negative and silencing activities. The central nervous system involvement is not necessarily determined by the dominant negative potency of the mutant TRbeta1 and other environmental or genetic factors may influence the RTH manifestations.

A Commentary on 10 Years of Aldose Reductase Inhibition for Limited Joint Mobility in Diabetes

This investigation examines the clinical response to long-term treatment of the diabetic syndrome of limited joint mobility (LJM) using an aldose reductase inhibitor (ARI) in comparison to historical controls, and proposes a potential role of aldose reductase (AR) genotype and expression in the clinical response to ARI. Clinical parameters, including quantitative hand movement and electromyogram, were followed over a decade of continuous ARI treatment with sorbinil (400 mg/day) in two patients with insulin-dependent diabetes mellitus (IDDM) and severe compromising LJM, and compared to the published 10-year prospective investigation of untreated IDDM diabetic patients with LJM. Both subjects were homozygous for the Z-2 AR allele (A-C)23 that has been linked with microvascular complications of DM. Cellular AR mRNA/beta-actin ratios for both treated patients while on ARI therapy were approximately one-half the value observed in untreated patients with the complications of nephropathy or neuropathy. This is the longest reported experience of ARI intervention for any diabetic complication, documenting sustained correction of LJM, lack of side effects, and a potential molecular basis for the therapeutic response.

Estimation of Maximal Cortisol Secretion Rate in Healthy Humans

CONTEXT Cortisol secretion is related to ACTH concentration by a sigmoidal dose-response curve, in which high ACTH concentrations drive maximal cortisol secretion rates (CSR(max)). OBJECTIVE We sought to estimate CSR(max) and free cortisol half-life in healthy humans (n = 21) using numerical methods applied to data acquired during cosyntropin (250 μg) stimulation. We also evaluated the effect of overnight dexamethasone (DEX; 1 mg) vs. placebo on estimates of CSR(max) and free cortisol half-life. DESIGN This study was a double-blind, placebo-controlled, randomized order of overnight DEX vs. placebo, cosyntropin (250 μg) stimulation with frequent serum cortisol sampling and computer-assisted numerical analysis. SETTING The study was conducted at a single academic medical center. PARTICIPANTS Twenty-one healthy adult subjects (15 females and six males), mean aged 46 yr, participated in the study. INTERVENTION Intervention in the study included DEX vs. placebo pretreatment, cosyntropin (250 μg) iv with frequent cortisol sampling. MAIN OUTCOME MEASURES CSR(max) and free cortisol half-life estimates, R² for goodness of fit, were measured. RESULTS Mean ± sd CSR(max) was 0.44 ± 0.13 nm/second, with free cortisol half-life of 2.2 ± 1.1 min. DEX did not significantly affect estimates of CSR(max) or free cortisol half-life. Our model accounts for most of the variability of measured cortisol concentrations (overall R² = 90.9 ±11.0%) and was more accurate (P = 0.004) during DEX suppression (R² = 94.6 ± 4.6%) compared with placebo (R² = 87.2 ± 8.7%). CONCLUSIONS Application of a mass-action model under conditions of cosyntropin stimulation provides a relatively simple method for estimation CSR(max) that accurately predicts measured cortisol concentrations. DEX administration did not significantly affect estimates of CSR(max) or free cortisol half-life.

Generalized thyroid hormone resistance due to a deletion of the carboxy terminus of the c-erbA β receptor

Generalized resistance to thyroid hormone (GRTH) is a disorder of thyroid hormone action which has been linked to the beta thyroid hormone receptor (TR beta) gene. A diverse array of TR beta mutations have been characterized, and these distinct genotypes have been associated with characteristic patterns of severity and tissue distribution of clinical thyroid resistance. In this report, we describe a patient with GRTH caused by a single C-->A base mutation (nucleotide 1623) in one allele of TR beta (exon 10). The mutation produces a premature translation termination signal (UGA) at codon 446 and predicts expression of a mutant TR beta which is truncated by 16 carboxyl-terminal amino acids (TR beta delta 446-461). This sequence was absent in both parents, indicative of a de novo mutation in the proband. To our knowledge, this case represents the first description of a mutation producing premature translation termination of TR beta in association with the syndrome GRTH, and emphasizes the critical role of the carboxyl terminal region of TR beta in mediating both positive and negative regulation of thyroid-responsive target genes in many tissues.