Diagnostic characteristics of late-night salivary cortisol using liquid chromatography-tandem mass spectrometry.

OBJECTIVE: The objective of the study was to describe the diagnostic performance of a commercially available late-night salivary cortisol (NSC) assay using liquid chromatography tandem mass spectrometry. METHODS: We retrospectively identified 90 patients who had one or more NSC determinations: 52 patients in whom Cushing syndrome (CS) was excluded or could not be confirmed [group 1 (G1)] and 38 patients in whom CS was confirmed [group 2 (G2)]. Eighteen healthy volunteers served as controls. RESULTS: Baseline demographics in all groups were similar with regards to age, ethnicity, gender, and body mass index. NSC levels [median (range)] were higher in G2, 381 (64-13,500) ng/dl [10.51 (1.77-372.46) nmol/liter], compared with controls, 19.3 (2.1-416) ng/dl [0.53 (0.06-11.48) nmol/liter], and G1, 26 (4-176) ng/dl [0.72 (0.11-4.86) nmol/liter, P < 0.001]. The highest combined sensitivity (92%) and specificity (92%) was achieved at a cut point of 107 ng/dl (2.95 nmol/liter). Two or more NSCs were done in 32 of 52 G1 and 31 of 38 G2 patients. In G1 eight of 32 (25%) had at least one elevated [>100 ng/dl (2.76 nmol/liter)] NSC including two in whom both NSCs were elevated. In contrast, four of 31 (13%) in G2 had at least one normal NSC including one with four of five normal NSC values. None of the patients with CS had a NSC less than 60 ng/dl (<1.66 nmol/liter). Comparing G1 and G2, obtaining more than one saliva sample did not improve the diagnostic accuracy of NSC measurement (P = 0.64). CONCLUSION: The liquid chromatography tandem mass spectrometry assay to measure NSC is a simple and reliable test to screen patients suspected to have CS. Clinicians should be aware of appropriate cutoff values for proper interpretation of NSC and use additional tests when necessary.

Cinacalcet hydrochloride reduces the serum calcium concentration in inoperable parathyroid carcinoma.

BACKGROUND: Management of inoperable parathyroid carcinoma presents a challenge because until recently, effective medical therapy was not available. Morbidity and mortality result primarily from severe hypercalcemia. We assessed the ability of the calcimimetic cinacalcet HCl to reduce serum calcium in patients with parathyroid carcinoma as well as its effect on PTH concentrations, bone turnover markers, safety, and health-related quality of life variables. METHODS: Twenty-nine patients with parathyroid carcinoma were enrolled in this open-label, single-arm study consisting of titration and maintenance phases. Cinacalcet doses were titrated (30 mg twice daily to 90 mg four times daily) for 16 wk or until serum calcium was no more than 10.0 mg/dl. The study endpoint was the proportion of patients with at least a 1 mg/dl reduction in serum calcium at the end of the titration phase (responders). RESULTS: Mean (+/- se) serum calcium (14.1 +/- 0.4 mg/dl) and PTH (697 +/- 94 pg/ml) were markedly elevated at baseline. At the end of the titration period, serum calcium was reduced by at least 1 mg/dl in 62% of patients (mean decline to 12.4 +/- 0.5 mg/dl). In the 18 responders, serum calcium fell from 15.0 +/- 0.5 to 11.2 +/- 0.3 mg/dl (P < 0.001). The greatest reductions in serum calcium were observed in patients with highest baseline calcium levels. PTH levels decreased, but not significantly, to 635 +/- 73 pg/ml (-4.6%). Adverse events included nausea, vomiting, headache, and fracture. CONCLUSIONS: Cinacalcet effectively reduces hypercalcemia in approximately two thirds of patients with inoperable parathyroid carcinoma and may represent an important new treatment option for these patients.

Effect of low-dose testosterone treatment on androgen regulated proteins prostate specific antigen and sex hormone binding globulin in short prepubertal boys: lack of initiation of puberty.

The efficacy of testosterone undecanoate (TU) treatment in constitutional delay of growth (CHD) is well recognized. We investigated its role in initiating puberty. Sera taken prior to, just after 6 months on and after 6 months off treatment with TU (20 mg daily) were analyzed from eight boys and compared to results from eight boys receiving placebo. Prostate specific antigen (PSA) and sex hormone binding globulin (SHBG), sleep-entrained pulsatility and mean overnight luteinizing hormone (mLH), and morning testosterone (T) levels were measured. Free androgen index (FAI) was calculated. Testicular volume (TV) and growth parameters were assessed. During treatment, there was a significant increase in height velocity in boys taking TU vs placebo (mean +/- SD: 5.7 +/- 2.0 vs 3.2 +/- 0.9 cm/year, p = 0.008) but no significant differences were observed in regard to LH pulsatility, mLH, T, SHBG, FAI, PSA and TV values. PSA was detectable in four patients (two each in the TU and placebo groups) at 6 months off treatment indicating pubertal progression. Among the hormones measured, only pretreatment mLH levels were significantly higher in the PSA-positive patients compared to 12 PSA-negative patients (mean +/- SEM: 1.5 +/- 0.39 vs 0.37 +/- 0.06 IU/l, p < 0.001). In conclusion, TU treatment shows no significant effect on initiation or advancement of puberty despite its resultant growth acceleration. Among the hormonal changes studied, mLH levels were the earliest indicator of pubertal initiation.

Effect of 131 iodine therapy on the course of Graves' ophthalmopathy: a quantitative analysis of extraocular muscle volumes using orbital magnetic resonance imaging.

There remains uncertainty as to the effect of radioactive iodine (131I) therapy on the associated ophthalmopathy (GO). Twenty newly diagnosed patients with Graves' hyperthyroidism treated with 131I (median dose, 15.5 mCi) were followed with ophthalmologic evaluations (OE) and magnetic resonance imaging (MRI) at baseline, 2, and 6 months, and with OE alone at 3 years. For MRI, the superior, inferior, and medial rectus muscle volumes and total muscle volumes (TMV) were measured. Replacement levothyroxine was initiated as low thyroxine (T4) levels were noted. At baseline, 10 patients (50%) showed evidence of mild GO by OE and/or MRI. There was a significant difference in TMV between the 20 patients with Graves' hyperthyroidism and 10 controls (mean +/- standard error [SE]; 2,652 +/- 118 vs. 2,046 +/- 96 mm3; P = 0.002) and between the 10 patients with and 10 without GO (3,006 +/- 96 vs. 2,298 +/- 61 mm3; P = 0.001). TMV correlated with the Hertel score (r = 0.56, P = 0.01). TMV showed no significant change at 2 or 6 months posttreatment. The inferior rectus volume increased slightly at 2 months posttreatment (P = 0.03) but remained stable at 6 months. Furthermore, no significant changes occurred in Hertel scores or in clinical assessments up to 3 years posttreatment and none showed worsening or new development of GO. In conclusion, our results show no significant risk for radioiodine-induced initiation or progression of mild GO.

Autoantibodies to IA-2 and GAD65 in patients with type 2 diabetes mellitus of varied duration: prevalence and correlation with clinical features.

OBJECTIVE: To determine the prevalence of autoantibodies to IA-2 (IA-2Ab) and glutamic acid decarboxylase (GADAb) in type 2 diabetes, their relationship to disease duration, and their importance in management decisions. METHODS: We undertook a study of 101 patients with type 2 diabetes (defined as nonketotic hyperglycemia at diagnosis) of varied duration (median, 4 years). Results were compared with those from 36 patients with type 1 diabetes also of varied duration (median, 2 years). IA-2Ab and GADAb were measured by radioligand-binding assays with use of in vitro-synthesized, 35S-labeled antigens. RESULTS: Of the 101 patients with type 2 diabetes, 20 (20%) were positive for GADAb; only 4 of these 20 were positive for IA-2Ab. In comparison, 75% of patients with type 1 diabetes were positive for GADAb, IA-2Ab, or both (P<0.0001). The coincidence of IA-2Ab positivity in GADAb-positive patients with type 2 diabetes was significantly lower than in patients with type 1 diabetes (20% versus 73%, respectively; P = 0.002). All four IA-2Ab- and GADAb-positive patients with type 2 diabetes required insulin and were younger than those positive for GADAb alone (P = 0.018). GADAb positivity in patients with type 2 diabetes was highly associated with insulin requirement (P = 0.004), with an odds ratio of 5.8 in predicting insulin dependence. Among patients with type 2 diabetes receiving insulin therapy, disease duration was significantly shorter (P = 0.025) and body mass index was significantly lower (P<0.001) in GADAb-positive versus GADAb-negative patients. In contrast to type 1 diabetes, in which GADAb values were negatively correlated with disease duration (r = -0.34; P = 0.044), no significant correlation with disease duration was observed in type 2 diabetes (r = -0.166; P = 0.48). CONCLUSION: Irrespective of duration of disease, measurement of IA-2Ab and GADAb can help to identify those patients with type 2 diabetes most likely to require insulin therapy.

Leiomyosarcoma of the main renal artery treated by laparoscopic radical nephrectomy.

Renal leiomyosarcomas are rare mesenchymal sarcomas. Although such tumors arising from the renal vein or kidney have been previously reported, we present the first case of a leiomyosarcoma arising from the main renal artery managed by laparoscopic radical nephrectomy.

Contamination of a bracelet following iodine-131 therapy: a case report.

A 73-y-old patient who had thyroid carcinoma had a post-treatment, whole-body 131I scan. The scan demonstrated an artifact caused by a bracelet contaminated with radioactive perspiration. This finding resulted in an artifact on the scan and had potential radiation safety implications.

Normalization of hyperhomocysteinemia with L-thyroxine in hypothyroidism.

BACKGROUND: Hyperhomocysteinemia is an independent risk factor for coronary, peripheral, and cerebrovascular disease. Elevated plasma homocysteine levels were described in a preliminary report on primary hypothyroidism. OBJECTIVE: To determine whether restoration of euthyroidism by L-thyroxine replacement therapy would reduce or normalize plasma homocysteine levels. DESIGN: Prospective cohort study. SETTING: Outpatient endocrinology department of a tertiary center. PATIENTS: 14 patients (10 women and 4 men; 25 to 77 years of age): 4 with newly diagnosed chronic (Hashimoto) hypothyroidism and 10 who had been rendered acutely hypothyroid (thyroid-stimulating hormone level > 25 mU/L) by total thyroidectomy for thyroid carcinoma. MEASUREMENTS: Total plasma homocysteine levels were measured at baseline and 3 to 9 months later, after euthyroidism had been attained by L-thyroxine replacement therapy. RESULTS: Median baseline plasma homocysteine levels in both sexes (women, 11.65 micromol/L [range, 7.2 to 26.5 micromol/L]; men, 15.1 micromol/L [range, 14.1 to 16.3 micromol/L]) were higher (P = 0.002) than those in healthy female (n = 35) and male (n = 36) volunteers (women, 7.52 micromol/L [range, 4.3 to 14.0 micromol/L]; men, 8.72 micromol/L [range, 5.94 to 14.98 micromol/L]). Eight patients (57%) had baseline plasma homocysteine levels that exceeded the upper limit of sex-specific reference ranges. Upon attainment of euthyroidism, all patients had a diminution in plasma homocysteine levels. The median overall change of -5.5 micromol/L (range, -15.4 to -1.8 micromol/L) corresponds to a difference of -44% (range, -58% to -13%) (P < 0.001). Homocysteine levels returned to normal in 7 of the 8 patients with elevated pretreatment values. CONCLUSIONS: Hypothyroidism may be a treatable cause of hyperhomocysteinemia, and elevated plasma homocysteine levels may be an independent risk factor for the accelerated atherosclerosis seen in primary hypothyroidism.

Serum prostate specific antigen, sex hormone binding globulin and free androgen index as markers of pubertal development in boys.

OBJECTIVE: Prostate specific antigen (PSA) expression in the prostate gland is regulated by androgens. Serum levels of PSA are undetectable by routine assays in normal boys. Measurable values could serve as a marker for pubertal development. In order to explore this question, we measured serum PSA levels in normal boys throughout puberty and examined the interrelationships with various hormonal and physical developmental changes. DESIGN: Sera from 77 normal boys in Tanner stages I to V (T-I to T-V) were analysed for PSA levels by a sensitive time-resolved fluoro-immunometric assay (sensitivity: 0.012 microgram/l). In addition, sex hormone binding globulin (SHBG), insulin like growth factor I (IGF-I), IGF binding protein 3(IGFBP-3) and testosterone were measured. RESULTS: PSA was detectable in 0% of Stage T-I (n = 16), 33% of T-II (n = 18), 65% of T-III (n = 17) and 100% of T-IV (n = 10) and T-V (n = 16) boys. PSA levels rose significantly according to stage (P < 0.05). Also, there were significant (P < 0.05) increments in serum testosterone, IGF-I and IGFBP-3 levels from stages T-I to T-IV. PSA showed a positive correlation with testosterone (r = 0.86, P < 0.001), IGF-I (r = 0.66, P < 0.001), and IGFBP-3 (r = 0.34, P = 0.004) levels. Both PSA and these analytes, however, showed significant overlap between stages T-I and T-II with only 6/18 (33%) and 12/18 (66%) of T-II subjects having PSA and testosterone levels, respectively, above the T-I range. In contrast, serum SHBG levels decreased markedly from stages I to II (P < 0.001). At the calculated best cut-off point for SHBG of 50 nmol/l, 16/18 T-II subjects had values below the T-I range (sensitivity = 89%). Because of this decrement of SHBG and the increasing testosterone secretion in early puberty, the Free Androgen Index (FAI = Testosterone/SHBG) could even better differentiate the onset of puberty with all except one of the T-II subjects having FAI levels above the T-I range (sensitivity = 94.4%). The decrease of SHBG in T-II subjects coincided with an increase in total body weight (P = 0.001) and body mass index (BMI, P = 0.0003). Despite the continuing pubertal rise in testosterone, SHBG levels showed a rebound increment from T-II-T-III subjects (P = 0.02) with a concomitant decrease in BMI (P = 0.0014). CONCLUSIONS: Prostate specific antigen closely reflects serum free androgen activity during puberty. However, it was unable to differentiate the earliest pubertal development. In comparison, SHBG levels and Free Androgen Index are more sensitive markers for the onset of puberty in boys. The inverse association between SHBG levels and BMI in pubertal stages Tanner stages, I to III suggests that body fatness, via its effect on insulin sensitivity, may play an important role in the regulation of SHBG production during early pubertal development.

Epitope heterogeneity of thyrotropin receptor-blocking antibodies in Graves' patients as detected with wild-type versus chimeric thyrotropin receptors.

The stable transfectants of wild-type (W25) and mutant thyrotropin-receptor (TSH-R) allow detection of the bioactivities of TSH-R antibodies in Graves' patients. A mutant Chinese hamster ovary (CHO) cell line (Mc1+2) transfected with a chimeric construct, where residues 8 to 165 of the TSH-R are replaced with residues 10 to 166 of the lutropin/choriogonadotropin (LH/CGR) receptor, lacks the cyclic adenosine monophosphate (cAMP) response to most thyrotropin stimulating antibodies (TSAb), yet retains the response to TSH and acquires the response to LH/CG. We compared Mc1+2 cells with wild-type W25 cells for their ability to detect TSAb as well as thyrotropin-blocking antibodies (TBAb) in Graves' sera. Eighteen normal and 39 Graves' sera were tested for TSAb and TBAb levels by in vitro bioassays using W25 and Mc1+2 cells. In addition, these sera were also tested for thyrotropin-binding inhibitory activity (TBII) by a radioreceptor assay. Eighteen (47%) Graves' sera had TBAb activity measured with W25 cells but not with Mc1+2 cells. These TBAbs were, therefore, a population of antibodies with functional epitopes on the N-terminus of the extracellular domain. This TBAb activity by W25 cells exhibited a high degree of correlation with TBII levels by a radioreceptor assay (r = 0.70, p = 0.001). Ten (25.6%) Graves' sera had positive TBAb activity in both W25 and Mc1+2 cells; moreover, their activity in both assays was similar (r = 0.83, p < 0.001). The TBAb activity in these sera, however, did not correlate with TBII activity. Eleven (28%) Graves' sera had no TBAb activity. Overall, thyroid-stimulating antibodies were detected in 87% and 28% of the 39 Graves' sera by W25 and Mc1 +2 cells, respectively. Thus, using the 2 cell lines, at least 2 distinct populations of TBAbs were detected. One is detected in a similar fashion by both W25 and Mc1+2 cell lines and likely interacts with the epitopes residing in the unaltered C-terminus of the TSH-R. The other is reactive in W25 cells only, indicating the loss of TBAb epitope in the chimeric receptor located in the N-terminus of the TSH-R. Furthermore, our results indicate that the TBAb binding epitope in 8-165 residues of the native TSH-R is highly associated with TBII activity in Graves' disease. These results indicate that patients with Graves' disease harbor TBAbs with epitope heterogeneity and favor the notion that there are different sites and mechanisms by which TBAbs act in Graves' patients. It remains to be determined whether or not TBAb subtyping will have a useful predictive role in the management of patients with Graves' disease.

United Kingdom Prospective Diabetes Study (UKPDS). Effects of glucose and blood pressure control on complications of type 2 diabetes mellitus.

The United Kingdom Prospective Diabetes Study (UKPDS) showed that intensive treatment (i.e., glucose-lowering drugs, with a goal fasting blood glucose level of 108 mg/dL) decreases the microvascular complications of type 2 diabetes mellitus. We summarize the key study results and their implications for clinical management of type 2 diabetes mellitus.

Bone mineral density, sex hormones, and long-term use of neuroleptic agents in men.

OBJECTIVE: To study the prevalence and severity of bone mineral loss and its relationship to sex hormone levels in men on long-term neuroleptic therapy. METHODS: Sixteen men with schizophrenia who were from 19 to 62 years old and had taken neuroleptic medication for 1 to 30 years had bone mineral density (BMD) of the lumbar spine and proximal femur measured by dual-energy x-ray absorptiometry. Results were compared with those from 16 age-matched control subjects. Serum testosterone, sex hormone-binding globulin, free testosterone index (FTI), prolactin, and pituitary gonadotropins were assayed and compared with age-matched Red Cross blood donor controls (N = 23 to 90 for the various assays). RESULTS: At the lumbar spine and at two of the three sites in the proximal femur (trochanter and Ward's triangle but not femoral neck), the BMD was lower in patients than in controls. A statistically significant increase in prolactin and sex hormone-binding globulin and a significantly decreased luteinizing hormone and FTI were found in the treated versus the control group. In the patient group, a significant inverse relationship existed between age and BMD at all sites. Lumbar spine density was related directly to FTI (r = 0.607; P=0.05) and inversely to duration of treatment (r = -0.767; P<0.001), although both correlations were accounted for mainly by age associations in multiple stepwise linear regression. Prolactin values did not correlate with either BMD or FTI. CONCLUSION: BMD was significantly lower with long-term neuroleptic use. In the lumbar spine, these changes may be related to the associated findings of decreased free testosterone and hyperprolactinemia, although the significance of other factors such as an independent drug effect, the psychiatric disease itself, and lifestyle cannot be excluded.

Insulin antibodies and hypoglycemia in diabetic patients. Can a quantitative analysis of antibody binding predict the risk of hypoglycemia?

We report a noninsulin-dependent diabetes mellitus (NIDDM) patient with spontaneous, severe hypoglycemic reactions and the presence of insulin antibodies. He had a remote antecedent history of beef-pork insulin therapy as well as exposure to hydralazine. Detailed insulin binding kinetic studies were performed in this patient as well as in six other insulin-treated diabetic patients with anti-insulin antibodies (three with and three without an obvious cause of hypoglycemia). Sera from the current patient and five of the six other diabetic patients (one NIDDM, four IDDM) revealed two types of binding sites: high-affinity with low capacity (Kd, 0.4-12.4 x 10(-9) mol/L; binding capacity, 0.6-659 mU/L) and low-affinity with high capacity (Kd, 0.3 to 35.7 x 10(-7) mol/L; binding capacity; 202-113,680 mU/L). One NIDDM patient had only high-affinity antibodies (Kd, 22.9 x 10(-9) mol/L; binding capacity of 78 mU/L). Type of diabetes mellitus, insulin antibody titers or their binding capacities, insulin levels (total, bound, or free), and bioavailable insulin were not related to hypoglycemic reactions. Two calculated values by the method described tended to discriminate patients with and without hypoglycemia. The calculated amount of low-affinity antibody bound insulin ranged from 69.4-2090 mU/L vs < 4-70.6 mU/L in patients with and without hypoglycemia, respectively. The best discrimination was afford by the percent saturation of low-affinity binding sites; values were clearly higher in the patients with hypoglycemia (2.5-34.4%) than in those without hypoglycemia (not detectable, 0.06, 0.15%). Consideration of the possible drug-associated insulin antibody formation in insulin-treated diabetics and the novel quantitative analysis of the antibody binding kinetics should prove helpful in evaluating patients with high insulin antibody titers and assessing the risk of hypoglycemia.

Thyroid-stimulating antibody assay using a human thyrotropin receptor transfected cell line: relationship to clinical features of graves' disease.

OBJECTIVE: To evaluate a Chinese hamster ovary cell line transfected with the human thyrotropin (thyroid-stimulating hormone or TSH) receptor (CHO-TSHR) for its use in routine clinical testing and compare it with the currently used Fisher rat thyroid cell line (FRTL-5) relative to the cyclic adenosine monophosphate (cAMP) responses to thyroid-stimulating antibodies (TSAb). METHODS: We analyzed 112 serum samples prospectively. TSAb were measured concurrently by both CHO-TSHR cells and FRTL-5 cells. In addition, thyrotropin-binding inhibitory immunoglobulin (TBII) was measured by a radioreceptor assay. Among the study subjects, 51 had active Graves' disease, 40 had other thyroid disease, and 21 had no thyroid disease. RESULTS: Of 51 patients with Graves' disease, 38 (74%) had positive results in both cell assays, 4 (8%) had negative results in both assays, and 9 (18%) had positive responses with CHO-TSHR cells but negative results with FRTL-5 cells. No patient had negative findings with CHO-TSHR cells and positive results by FRTL-5 cells. All 61 patients in the other two groups had negative results by both assays (diagnostic specificity = 100%). The diagnostic sensitivity and accuracy of TSAb activity by the two cell lines (CHO-TSHR versus FRTL-5) were 92.2% versus 74.5% and 96.4% versus 88.4%, respectively. A significant correlation was noted between the TSAb activities in active Graves' sera by the two assays (r = 0.58; P<0.0001); however, values by the CHO-TSHR method were significantly higher (P<0.003). TSAb activities determined with both cell lines were positively correlated with the 24-hour thyroidal 131 I uptake (r = 0.62 and P<0.0005 in CHO-TSHR cells; r = 0.41 and P<0.05 in FRTL-5 cells). The difference in TSAb activity between patients with ophthalmopathy and those without ophthalmopathy was statistically significant with both FRTL-5 (P<0.01) and CHO-TSHR (P = 0.05) assays. The correlation between the severity of eye disease and TSAb activities was significant only with the FRTL-5 assay (P = 0.009) but not with the CHO-TSHR assay (P = 0.16). TSAb activity showed a significant positive correlation with serum thyroxine, triiodothyronine, and TBII by both cell assays. Neither cell line showed a significant correlation between TSAb activity and antimicrosomal antibody titer. CONCLUSION: The CHO-TSHR cell line is superior to the standard FRTL-5 cell line in the cAMP bioassay for TSAb with reference to diagnostic sensitivity and accuracy, correlation with disease activity, simplicity, and cost.

Absence of serologic evidence for hepatitis C virus infection in patients with Hashimoto's thyroiditis.

BACKGROUND/AIMS: This paper explores the reported association between Hashimoto's thyroiditis and hepatitis C virus (HCV) infections. MATERIALS AND METHODS: The results of serologic testing for anti-HCV antibody levels were documented in 28 patients (19 female, 9 male) with Hashimoto's thyroiditis and 23 age and gender matched controls with non-Hashimoto's thyroid disorders. RESULTS: The anti-HCV results were negative in both groups. CONCLUSIONS: These findings suggest that despite a reported high prevalence of Hashimoto's thyroiditis in patients with chronic HCV infections, the reverse is not true and hence routine anti-HCV screening cannot be advocated for patients with Hashimoto's thyroiditis.

Immunoradiometric assay measurements of insulin-like growth factor-I (IGF-I): comparison with radioimmunoassays using native or des (1-3) IGF-I as radioligands.

Insulin-like growth-factor-binding proteins (BPs) in serum interfere with the measurement of insulin-like growth factor-I (IGF-I). Various assays have been developed to overcome this interference. We evaluated an immunoradiometric (IRMA) assay and compared it with the radioimmunoassay (RIA) using both native IGF-I and a truncated form of IGF-I [des (1-3) IGF-I] as radioligands. The IRMA was simpler (one step assay) and faster (3 hr incubation) than RIA(s) (overnight incubation). Sera were extracted with acid ethanol (AE) before all three assays. Analysis of serum samples (n = 78) performed by use of the two different radioligands in the RIA assays were highly correlated (r = 0.967, P < 0.0001). Measurements of serum IGF-I by IRMA in same samples were also highly correlated with those of the RIA assays (r = 0.952 for RIA and 0.947 for triGF-I RIA, P < 0.0001 for both). To assess the effect of binding protein-3 (BP-3) levels (after AE extraction) on these assays, BP-3 levels were measured in sera from 36 healthy women. The mean BP-3 level was 3.6 +/- 0.79 (S.D.) mg/L (range 1.3-5.0), and there was no significant difference in IGF-I levels measured by the three assays. Also, BP-3 levels were inversely correlated with IGF-I levels as measured by all three methods (r = 0.73 for IRMA, 0.71 for triGF-I, and 0.75 for IGF-I RIA). To assess the effect of binding protein-1 (BP-1) levels on these assays, IGF-I was also measured by IRMA and trlGF-I RIA in 19 women with advanced breast cancer. Women with breast cancer had significantly higher (P < 0.001) BP-1 levels than age matched healthy women. IGF-I levels measured by IGF-I IRMA were slightly lower than those measured by trlGF-I RIA in breast cancer patients. However, this difference was not statistically significant (P = 0.56). These findings suggest that variations in BP-3 or BP-1 levels after AE extraction have no significant effect in any of these assays. "We conclude that trlGF-I as a radioligand provides no added advantage over the standard IGF-I RIA. We also conclude that the IRMA assay is valid for measuring IGF-I and is faster and more convenient than RIA.

Characterization of 24-h cortisol release in obese and non-obese hyperandrogenic women.

Excessive androgen output is a well-recognized feature of adrenocortical oversecretion in women with ovarian hyperandrogenism, or polycystic ovary disease (PCOD). However, evidence of a concomitant alteration of cortisol secretion is lacking even though obesity per se, a common clinical feature of PCOD, has been shown to be associated with cortisol oversecretion. To clarify whether a subtle alteration in cortisol secretion exists, a study of 24-h episodic cortisol release and post-prandial cortisol responses was undertaken in eight women with PCOD and eight normal women comprising equal numbers of obese and non-obese subjects. All four groups showed normal biphasic 24-h cortisol secretion profiles but cortisol pulse frequency was increased in the PCOD groups. Independently, both hyperandrogenism and obesity were associated with an accelerated cortisol clearance rate. These changes, together with normal or only slightly elevated 24-h cortisol integrated area under the curve, suggest an increased compensatory cortisol production in women with PCOD. Furthermore, subjects with PCOD and subjects with obesity showed different post-prandial cortisol responses to normal non-obese women. In conclusion, these subtle cortisol abnormalities may be a manifestation of altered central regulation of the hypothalamic-pituitary-adrenal axis and peripheral metabolic abnormalities, and may be linked to the pathophysiology of PCOD.

Measurement of human urinary prolactin as a noninvasive study tool.

We used a time-resolved solid-phase fluoroimmunoassay with a sensitivity of 25 ng/L on 40-fold-concentrated urines to measure urine prolactin (PRL) excretion. The nature of the immunoreactive material was verified to be PRL by: (a) column chromatography showing a monomeric 23-kDa peak; (b) similarity between fluoroimmunoassay and bioassay (Nb2 lymphoma cell) results; and (c) Western blot identification. In 20 normal subjects [serum PRL 6.8 (3.8-14.0) micrograms/L, median (and range)], urine PRL excretion was 0.15 (0.07-0.23) ng/h and 0.24 (0.15-0.54) micrograms/mol of creatinine. Urine values in seven hyperprolactinemic patients were all greater than the upper limit of normal. The correlation between urinary excretion rate and serum values was highly significant (r = 0.979; P < 0.001). These results indicate that a monomeric, immunologically reactive, biologically active form of PRL can be measured in urine at concentrations approximately 0.0005 that in serum. This urine PRL method may provide a practical tool for the repetitive, noninvasive study of PRL dynamics in field studies and in patients with reproductive disorders.

Differential effects of compounds that act at strychnine-insensitive glycine receptors in a punishment procedure.

The anxiolytic and memory-impairing effects of compounds that act at strychnine-insensitive (SI) glycine receptors were examined and compared with those of a competitive N-methyl-D-aspartate antagonist, 2-amino-7-phosphonoheptanoic acid (AP7); a use-dependent channel blocker, dizocilpine; and a benzodiazepine agonist, diazepam (DZP). Mice were trained to avoid a dark compartment and their latencies to step through were measured either within 1 hr after training in the presence of the drug (to assess the anxiolytic effects) or 24 hr after pre- or post-training treatment (to assess the effects on learning and memory). Post-training administration of the glycinergic compounds 1-aminocyclopropanecarboxylic acid, 7-chlorokynurenic acid and D-cycloserine reduced step-through latencies when testing was performed 30 min after drug treatment and within 1 h after training. Latencies were unaltered by these glycinergic compounds when testing was performed 24 hr later. Similar results were obtained with AP7 and DZP. In contrast, an amnesic dose of pentylenetetrazole reduced latencies both within 1 and 24 hr after training. Pretreatment with glycine abolished the reduction in latencies observed with SI glycine receptor ligands 1 hr after training but did not antagonize the reduction produced by AP7. Pretraining administration of SI glycine receptor ligands did not alter step-through latencies measured 24 hr later. In contrast, under these same conditions, AP7, dizocilpine and DZP produced a significant reduction in latencies. These results demonstrate that compounds that act at SI glycine receptors do not impair learning and memory at doses that are anxiolytic in a single-trial punishment paradigm.