LC-MS/MS reduces interference by high levels of 25(OH)D and its metabolites on measured 1,25(OH)2D.

BACKGROUND: High-dose vitamin D supplementation has been recommended as treatment to several conditions; however, potential side effects such as hypercalcemia should be considered, plus the fact that high levels of 25(OH)D may interfere with potential 1,25(OH)2D measurements. Our study compared two methods of measuring 1,25-dihydroxyvitamin D [1,25(OH)2D] in samples with 25-hydroxyvitamin D [25(OH)D] levels above 150 ng/mL (375 nmol/L). METHODS: We studied serum samples referred to 25(OH)D and 1,25(OH)2D quantification. The concentrations of 25(OH)D and 1,25(OH)2D were measured using DiaSorin chemiluminescent assays (CLIA) in 213 samples (CLIA group), whereas in 357 samples 25(OH)D and 1,25(OH)2D were measured by DiaSorin CLIA and liquid chromatography-tandem mass spectrometry (LC-MS/MS), respectively (CLIA + MS group). RESULTS: Median concentrations of 25(OH)D and 1,25(OH)2D in the CLIA group were 371 ng/mL (928 nmol/L, range 154-856 ng/mL) and 350 pg/mL (875 pmol/L, range 41-1280 pg/mL), respectively, and correlated significantly (Spearman correlation coefficient rs = 0.8469, P < 0.001). In the CLIA + MS group, the median concentrations of 25(OH)D and 1,25(OH)2D were, respectively, 344 ng/mL (860 nmol/L, range 152-756 ng/mL) and 56 pg/mL (140 pmol/L, range 17-151 pg/mL), and were not correlated (rs = 0.0218, P = 0.6811). No significant difference was found in calcium, creatinine, and PTH serum values between the groups. CONCLUSION: Methods for measuring 1,25(OH)2D in patients with high levels of 25(OH)D may be susceptible to interference by 25(OH)D and its metabolites and should be validated carefully. In such cases, measurement of 1,25(OH)2D using LC-MS/MS is preferred.

Effect of soft tissue noise on trabecular bone score in postmenopausal women with diabetes: A cross sectional study.

BACKGROUND: Type 2 diabetes (T2D) is associated with increased fracture risk, despite similar or greater BMD compared to nondiabetics. TBS predicts fracture risk in T2D and nondiabetics. However, increased abdominal thickness, a common feature in T2D, may reduce TBS values. AIM: To study the relationship among glycemic status, BMD and TBS, considering abdominal soft tissue thickness (STT) interference. METHODS: Cross-sectional analysis of 493 women ≥65 years, with simultaneous DXA scans and HbA1c measures. STT and TBS (iNsight Software, v3.0) were derived from lumbar spine (LS) scans. Subjects were divided according to HbA1c levels: 1 (≥6.5%; n = 116), 2 (5.7-6.4%; n = 217) and 3 (≤5.6%; n = 160). Group 1 was further divided based on HbA1c and/or disease duration: 1a (HbA1c ≥ 7.5%; n = 42), 1b (HbA1c ≥ 6.5% and disease duration ≥5 years; n = 63) and 1c (HbA1c ≥ 7.5% and disease duration ≥5 years; n = 30). FINDINGS: For the entire cohort, mean age, TBS, BMI and STT were 71.8 ± 6.0 years, 1.299 ± 0.101, 26.9 ± 4.1 kg/m2, and 21.4 ± 2.9 cm, respectively. LS-BMD was similar among groups. BMD in hip sites and STT were higher in group 1. TBS was lower in patients with higher HbA1c (P = 0.020), with a mean TBS in groups 1, 2, and 3 of 1.280, 1.299 and 1.314, respectively. This difference remained after adjusting for age, LS-BMD and BMI (P = 0.010). After replacing BMI with STT, TBS differences were no longer significant (P = 0.270). The same was observed when subgroups 1a and 1b were compared to group 3. However, for subgroup 1c, TBS remained lower compared to group 3, even after adjusting for age, LS-BMD and STT, with a borderline P-value (1.275 vs. 1.308; P = 0.047). CONCLUSION: Higher HbA1c levels were associated with greater BMD in hip sites, higher abdominal STT and lower TBS values. However, after including the STT in the adjustment, TBS differences among groups disappeared, except in women with higher HbA1c levels and longer disease duration.

Basal serum thyroglobulin measured by a second-generation assay is equivalent to stimulated thyroglobulin in identifying metastases in patients with differentiated thyroid cancer with low or intermediate risk of recurrence.

BACKGROUND: Guidelines for the follow-up of differentiated thyroid cancer (DTC) recommend the measurement of TSH-stimulated thyroglobulin (s-Tg) instead of basal Tg on T4 therapy (b-Tg). However, these guidelines were established using first-generation Tg assays with a functional sensitivity (FS) of 0.5-1.0 ng/ml. Current more sensitive second-generation Tg assays (Tg2G; FS 0.05-0.10 ng/ml) have shown that low-risk DTC patients with undetectable b-Tg rarely have recurrences. OBJECTIVES: This study was undertaken to compare b-Tg using a chemiluminescent Tg2G assay (Tg2GICMA; FS 0.1 ng/ml) with s-Tg in DTC patients with an intermediate risk of recurrence. METHODS: We evaluated 168 DTC patients with a low (n = 101) and intermediate (n = 67) risk of recurrence treated by total thyroidectomy (147 also treated with radioiodine), with a mean follow-up of 5 years. RESULTS: b-Tg was undetectable with the Tg2GICMA in 142 of 168 patients. s-Tg was <2 ng/ml in 138 of these 142 patients, and only 3 of these 138 (2%) presented metastases on cervical ultrasound (US). Of the 4 of 142 patients with s-Tg >2 ng/ml, 1 had cervical metastases seen after radioiodine. Furthermore, 26 of 168 patients presented detectable b-Tg with the Tg2GICMA; 17 of these 26 patients also presented s-Tg >2 ng/ml. In 10 of these 17 patients, metastases were detected. Cervical US or b-Tg were positive in 14 of 15 patients with recurrent disease. Globally, the sensitivity and negative predictive value of the Tg2GICMA plus US were 93 and 99%, respectively. CONCLUSION: b-Tg measured with a Tg2GICMA and cervical US, used together, are equivalent to s-Tg in identifying metastases in patients with DTC with a low or intermediate risk of recurrence.

Acromegaly and pregnancy: a prospective study.

CONTEXT AND OBJECTIVE: The interaction between pregnancy and acromegaly has been studied only retrospectively. We used prospective data to assess those interactions. DESIGN: Prospective, interventional, multicentric study. PATIENTS: TEN PREGNANCIES IN EIGHT ACROMEGALIC PATIENTS WERE INCLUDED ACCORDING TO THE FOLLOWING CRITERIA: previous diagnosis of acromegaly; and active acromegaly before pregnancy. Sellar magnetic resonance image (MRI), GH, and IGF1 measurements were carried out before pregnancy. The exclusion criterion was radiotherapy. INTERVENTION: Withdrawal of pharmacological treatment (octreotide and/or cabergoline and/or pegvisomant) following pregnancy diagnosis. MAIN OUTCOME MEASURES: Clinical/biochemical evaluations throughout pregnancy/puerperium and sellar MRI after delivery; and GH and IGF1 measurements before pregnancy. GH was measured by an interference-free IFMA assay during pregnancy and IGF1 by measured by Immulite 2000 assay in patients and 64 control pregnancies. RESULTS: No tumor growth was observed. Nine deliveries were at term and one at 35 weeks (preeclampsia). All newborns were healthy. Mean IGF1 levels before and during pregnancy were similar, but increased significantly during puerperium. As IGF1 in controls increased after midgestation, the prevalence of controlled IGF1 rose significantly from 2/10 (<20 weeks) to 9/10 (>30 weeks). Diabetes mellitus and hypertension/preeclampsia developed in one patient in each group; both complications were nonsignificantly (P=0.06) associated with IGF1 >1.3 ULN before pregnancy. CONCLUSIONS: Acromegaly control usually improved and tumor growth was not stimulated during pregnancy in spite of withdrawal of drug treatment. Drug treatment can be discontinued in most patients. Uncontrolled disease before pregnancy may pose a higher risk for diabetes and hypertension.

Measurement of calcitonin and calcitonin gene-related peptide mRNA refines the management of patients with medullary thyroid cancer and may replace calcitonin-stimulation tests.

BACKGROUND: Serum calcitonin (sCT) is the main tumor marker for medullary thyroid cancer (MTC), but it has certain limitations. Various sCT assays may have important intra-assay or interassay variation and may yield different and sometimes conflicting results. A pentagastrin- or calcium-stimulation calcitonin (CT) test may be desirable in some situations. Alternatively, or in the absence of the stimulation test, mRNA detection offers the advantages of being more comfortable and less invasive; it only requires blood collection and has no side effects. The objective of this study was to investigate the applicability of measuring calcitonin-related polypeptide alpha (CALCA) gene transcripts (CT-CALCA and calcitonin gene-related peptide [CGRP]-CALCA) in patients with MTC and in relatives diagnosed with a RET mutation and to test mRNA as an alternative diagnostic tool for the calcitonin-stimulation test. METHODS: Twenty-three healthy controls and 26 individuals evaluated for MTC were selected, including patients with sporadic or hereditary MTC and RET mutation-carrying relatives. For molecular analysis, RNA was extracted from peripheral blood, followed by cDNA synthesis using 3.5 µg of total RNA. Quantitative real-time polymerase chain reaction (RT-qPCR) was performed with SYBR Green and 200 nM of each primer for the two specific mRNA targets (CT-CALCA or CGRP-CALCA) and normalized with the ribosomal protein S8 as the reference gene. RESULTS: We detected CALCA transcripts in the blood samples and observed a positive correlation between them (r=0.946, p<0.0001). Both mRNAs also correlated with sCT (CT-CALCA, r=0.713, p<0.0001; CGRP-CALCA, r=0.714, p<0.0001). The relative expression of CT-CALCA and CGRP-CALCA presented higher clinical sensitivity (86.67 and 100, respectively), specificity (97.06 and 97.06), positive predictive value (92.86 and 93.75), and negative predictive value (94.29 and 100), than did sCT (73.33, 82.35, 64.71, and 87.50, respectively). In addition, the CALCA transcript measurement mirrored the response to the pentagastrin test. CONCLUSION: We demonstrate that the measurement of CALCA gene transcripts in the bloodstream is feasible and may refine the management of patients with MTC and RET mutation-carrying relatives. We propose considering the application of this diagnostic tool as an alternative to the calcitonin-stimulation test.

Association of ADIPOQ variants, total and high molecular weight adiponectin levels with coronary artery disease in diabetic and non-diabetic Brazilian subjects.

OBJECTIVE: To investigate the association of ADIPOQ variants, total and high molecular weight adiponectin (HMW) adiponectin levels with the prevalence of diabetes mellitus and coronary artery disease (CAD) diagnosed by coronary angiography in Brazilian subjects with high cardiovascular risk. METHODS: 603 subjects undergoing coronary angiography were studied in regard to their glycemic status and presence of CAD (lesions >0%). We evaluated baseline concentrations of total and HMW adiponectin and three ADIPOQ variants: -11391G>A (rs17300539), +45T>G (rs2241766) and+276G>T (rs1501299). RESULTS: The G-allele of rs2241766 was associated with higher levels of total and HMW adiponectin, and the A-allele of rs17300539 was associated with higher levels of HMW adiponectin. Lower levels of total and HMW adiponectin were independently associated with CAD. The G-allele of rs2241766 (OR 2.45, 95% C.I. 1.05-6.04, p=0.04) and the G-allele of rs1501299 (OR 1.89, 95% C.I. 1.04-3.45, p=0.03) were associated with CAD, and these associations were independent of circulating levels of adiponectin. CONCLUSIONS: In Brazilian subjects with high cardiovascular risk, CAD was associated with lower total and HMW adiponectin levels. The rs2241766 and rs1501299 polymorphisms were associated with CAD. The rs2241766 variant was associated with total and HMW adiponectin levels, while rs17300539 was associated with HMW adiponectin levels.

Superior discriminating value of ACTH-stimulated serum 21-deoxycortisol in identifying heterozygote carriers for 21-hydroxylase deficiency.

BACKGROUND: Congenital adrenal hyperplasia caused by classic 21-hydroxylase deficiency (21OHD) is an autosomal recessive disorder with a high prevalence of asymptomatic heterozygote carriers (HTZ) in the general population, making case detection desirable by routine methodology. HTZ for classic and nonclassic (NC) forms have basal and ACTH-stimulated values of 17-hydroxyprogesterone (17OHP) that fail to discriminate them from the general population. 21-Deoxycortisol (21DF), an 11-hydroxylated derivative of 17OHP, is an alternative approach to identify 21OHD HTZ. OBJECTIVE: To determine the discriminating value of basal and ACTH-stimulated serum levels of 21DF in comparison with 17OHP in a population of HTZ for 21OHD (n = 60), as well as in NC patients (n = 16) and in genotypically normal control subjects (CS, n = 30), using fourth generation tandem mass spectrometry after HPLC separation (LC-MS/MS). RESULTS: Basal 21DF levels were not different between HTZ and CS, but stimulated values were increased in the former and virtually nonresponsive in CS. Only 17·7% of the ACTH-stimulated 21DF levels overlapped with CS, when compared to 46·8% for 17OHP. For 100% specificity, the sensitivities achieved for ACTH-stimulated 21DF, 17OHP and the quotient [(21DF + 17OHP)/F] were 82·3%, 53·2% and 87%, using cut-offs of 40, 300 ng/dl and 46 (unitless), respectively. Similar to 17OHP, ACTH-stimulated 21DF levels did not overlap between HTZ and NC patients. A positive and highly significant correlation (r = 0·846; P < 0·001) was observed between 21DF and 17OHP pairs of values from NC and HTZ. CONCLUSION: This study confirms the superiority of ACTH-stimulated 21DF, when compared to 17OHP, both measured by LC-MS/MS, in identifying carriers for 21OHD. Serum 21DF is a useful tool in genetic counselling to screen carriers among relatives in families with affected subjects, giving support to molecular results.

Adipokine profile and urinary albumin excretion in isolated growth hormone deficiency.

BACKGROUND: GH deficiency (GHD) is often associated with cardiovascular risk factors, including abdominal fat accumulation, hypercholesterolemia, and increased C-reactive protein. Despite the presence of these risk factors, adults with congenital lifetime isolated GHD (IGHD) due to an inactivating mutation in the GHRH receptor gene do not have premature atherosclerosis. OBJECTIVE: The aim was to study the serum levels of adiponectin and leptin (antiatherogenic and atherogenic adipokine, respectively), and the urinary albumin excretion (UAE) in these IGHD individuals. DESIGN AND PATIENTS: We conducted a cross-sectional study of 20 IGHD individuals (seven males; age, 50.8 +/- 14.6 yr) and 22 control subjects (eight males; age, 49.9 +/- 11.5 yr). MAIN OUTCOME MEASURES: Anthropometric factors, body composition, blood pressure, serum adiponectin, leptin, and UAE were measured. RESULTS: Adiponectin was higher [12.8 (7.1) vs. 9.7 (5) ng/ml; P = 0.041] in IGHD subjects, whereas no difference was observed in leptin [7.3 (6.3) vs. 9.3 (18.7 ng/ml] and UAE [8.6 (13.8) vs. 8.5 (11.1) microg/min]. CONCLUSIONS: Subjects with lifetime untreated IGHD have an adipokine profile with high adiponectin and normal leptin levels that may delay vascular damage and lesions of the renal endothelium.

Adrenocorticotrophic hormone (ACTH) responsiveness to ghrelin increases after 6 months of ketoconazole use in patients with Cushing's disease: comparison with GH-releasing peptide-6 (GHRP-6).

BACKGROUND: In Cushing's disease (CD), adrenocorticotrophic hormone (ACTH)/cortisol responses to growth hormone secretagogues (GHS), such as ghrelin and GHRP-6, are exaggerated. The effect of clinical treatment of hypercortisolism with ketoconazole on ACTH secretion in CD is controversial. There are no studies evaluating ACTH/cortisol responses to GHS after prolonged ketoconazole use in these patients. OBJECTIVE: To compare ghrelin- and GHRP-6-induced ACTH/cortisol release before and after ketoconazole treatment in patients with CD. DESIGN/PATIENTS: Eight untreated patients with CD (BMI: 28.5 +/- 0.8 kg/m(2)) were evaluated before and after 3 and 6 months of ketoconazole treatment and compared with 11 controls (BMI: 25.0 +/- 0.8). RESULTS: After ketoconazole use, mean urinary free cortisol values decreased significantly (before: 613.6 +/- 95.2 nmol/24 h; 3rd month: 170.0 +/- 27.9; 6th month: 107.9 +/- 30.1). The same was observed with basal serum cortisol (before: 612.5 +/- 69.0 nmol/l; 3rd month: 463.5 +/- 44.1; 6th month: 402.8 +/- 44.1) and ghrelin- and GHRP-6-stimulated peak cortisol levels (before: 1183.6 +/- 137.9 and 1045.7 +/- 132.4; 3rd month: 637.3 +/- 69.0 and 767.0 +/- 91.0; 6th month: 689.8 +/- 74.5 and 571.1 +/- 71.7 respectively). An increase in basal ACTH (before: 11.2 +/- 1.6 pmol/l; 6th month: 19.4 +/- 2.7) and in ghrelin-stimulated peak ACTH values occurred after 6 months (before: 59.8 +/- 15.4; 6th month: 112.0 +/- 11.2). GHRP-6-induced ACTH release also increased (before: 60.7 +/- 17.2; 6th month: 78.5 +/- 12.1), although not significantly. CONCLUSIONS: The rise in basal ACTH levels during ketoconazole treatment in CD could be because of the activation of normal corticotrophs, which were earlier suppressed by hypercortisolism. The enhanced ACTH responses to ghrelin after ketoconazole in CD could also be due to activation of the hypothalamic-pituitary-adrenal axis and/or to an increase in GHS-receptors expression in the corticotroph adenoma, consequent to reductions in circulating glucocorticoids.

Discordant results in Tc-99m tetrofosmin and Tc-99m sestamibi parathyroid scintigraphies.

Parathyroid scintigraphies have been used to detect pathological parathyroid glands either before as well as after the parathyroid resection surgery in patients with hyperparathyroidism. One of the most utilized techniques to perform the studies is the double-phase images with Tc-99m sestamibi, which has been shown to be very accurate in the localization of enlarged parathyroid glands. Similar to Tc-99m sestamibi, Tc-99m tetrofosmin is a radiopharmaceutical initially developed to perform myocardial perfusion study that has been used to perform parathyroid scintigraphies. Although most of the papers suggest that the overall sensitivities of both radiopharmaceuticals are similar, there are some papers questioning the accuracy of Tc-99m tetrofosmin to detect abnormal parathyroid glands. In the present article, we report a case with discordant results by both methods.

[Brown tumor of the sternum--a potential cause of false-positive result on parathyroid scintigraphy with Tc-99m sestamibi]. / Tumor marrom em esterno--uma causa potencial de resultado falso-positivo na cintilografia das paratireóides com sestamibi-99mTc.

Parathyroid scintigraphies have been used to detect pathological parathyroid glands either before as well as after the parathyroid resection surgery in patients with hyperparathyroidism. Although this test presents high specificity for detection of increased parathyroid glands, there exist causes of false positive results. In the present article, we report a case of a renal transplanted patient, with multiple lytic lesions on pelvic bones reported as brown tumors, who presented a focal uptake in the anterior portion of the superior mediastinum on Tc-99m sestamibi scintigraphy. This focal uptake, initially thought to be an ectopic parathyroid gland, after a more detailed analysis and the performance of other imaging diagnostic tests was demonstrated to be a brown tumor of the sternum.

Discordant results in Tc-99m tetrofosmin and Tc-99m sestamibi parathyroid scintigraphies

Parathyroid scintigraphies have been used to detect pathological parathyroid glands either before as well as after the parathyroid resection surgery in patients with hyperparathyroidism. One of the most utilized techniques to perform the studies is the double-phase images with Tc-99m sestamibi, which has been shown to be very accurate in the localization of enlarged parathyroid glands. Similar to Tc-99m sestamibi, Tc-99m tetrofosmin is a radiopharmaceutical initially developed to perform myocardial perfusion study that has been used to perform parathyroid scintigraphies. Although most of the papers suggest that the overall sensitivities of both radiopharmaceuticals are similar, there are some papers questioning the accuracy of Tc-99m tetrofosmin to detect abnormal parathyroid glands. In the present article, we report a case with discordant results by both methods.

A cintilografia das paratireóides tem sido utilizada para detectar glândulas paratireóides patológicas tanto antes quanto após (em caso de insucesso) a cirurgia de ressecção em pacientes com hiperparatireoidismo. Uma das técnicas mais utilizadas para realizar este exame é a de duas fases utilizando como radiofármaco o sestamibi-99mTc, a qual tem se mostrado acurada na localização de glândulas paratireóides aumentadas. Similarmente ao sestamibi-99mTc, o tetrofosmin-99mTc é um radiofármaco que foi inicialmente desenvolvido para a realização de cintilografia de perfusão do miocárdio e que tem sido utilizado para a realização de cintilografia das paratireóides. Apesar de muitos artigos sugerirem que as sensibilidades dos dois radiofármacos são idênticas, alguns poucos trabalhos questionam a acurácia do tetrofosmin-99mTc para a localização de glândulas paratireóides anômalas. No presente artigo, relatamos um caso em que foi observado resultado discordante pelos dois métodos.

Tumor marrom em esterno: uma causa potencial de resultado falso-positivo na cintilografia das paratireóides com sestamibi-99mTc / Brown tumor of the sternum: a potential cause of false-positive result on parathyroid scintigraphy with Tc-99m sestamibi

A cintilografia das paratireóides tem sido utilizada para detectar glândulas patológicas em pacientes com hiperparatireoidismo, tanto antes quanto após a cirurgia de paratireoidectomia. Apesar da elevada especificidade, a cintilografia das paratireóides pode apresentar resultados falso-positivos. Neste artigo, relatamos o caso de uma paciente transplantada renal, com múltiplas lesões osteolíticas na bacia, interpretadas como tumores marrons, que à cintilografia das paratireóides com sestamibi-99mTc apresentou hipercaptação focal do radiofármaco em topografia de porção anterior do mediastino superior. Esta área hipercaptante foi inicialmente interpretada como glândula paratireóide ectópica, porém, após realização de outros exames de imagem e análise mais detalhada, mostrou-se corresponder a tumor marrom em esterno.

Parathyroid scintigraphies have been used to detect pathological parathyroid glands either before as well as after the parathyroid resection surgery in patients with hyperparathyroidism. Although this test presents high specificity for detection of increased parathyroid glands, there exist causes of false positive results. In the present article, we report a case of a renal transplanted patient, with multiple lytic lesions on pelvic bones reported as brown tumors, who presented a focal uptake in the anterior portion of the superior mediastinum on Tc-99m sestamibi scintigraphy. This focal uptake, initially thought to be an ectopic parathyroid gland, after a more detailed analysis and the performance of other imaging diagnostic tests was demonstrated to be a brown tumor of the sternum.

Fasting insulin concentration is highly correlated with quantitative insulin sensitivity check index.

Given that the "gold standard" method for evaluating insulin sensitivity in vivo (hyperinsulinemic euglycemic glucose clamp technique) cannot be routinely applied because of technical reasons, simple methods and indexes were developed and are currently available to assess insulin sensitivity in vivo. Quantitative insulin sensitivity check index (QUICKI) has recently been described and is able to accurately estimate insulin sensitivity from a fasting blood sample. We demonstrated that fasting insulin levels strongly inversely correlated with QUICKI in three different groups: 215 healthy nondiabetic nonobese subjects, 62 nondiabetic obese subjects, and 44 patients with glucose intolerance or type 2 diabetes mellitus. Fasting insulin measurement is a simple way of assessing insulin sensitivity in obese and nonobese humans, with or without glucose intolerance or type 2 diabetes mellitus.

Reversed-phase high-performance liquid chromatography separation of adrenal steroids prior to radioimmunoassay: application in congenital adrenal hyperplasia.

21-Hydroxylase deficiency (21-OHD) is the most common form of congenital adrenal hyperplasia (CAH), followed by 11beta-hydroxylase deficiency (11beta-OHD). Diagnostic serum markers for these conditions are 17-hydroxyprogesterone (17-OHP) and 11-desoxycortisol (S), respectively. In 21-OHD, the large amounts of 17-OHP are further 11beta-hydroxylated to form 21-deoxycortisol (21-DF), making it also an excellent marker of this disease. These steroids can be measured in blood by radioimmunoassay (RIA). In this paper, we report the use of high-performance liquid chromatography (HPLC) for steroid purification, prior to RIA determinations of 21-DF, S, 17-OHP, and testosterone (T) in ether-extracted serum. The chromatographic separation is developed in a BDS-Hypersil column using water-methanol (53:47, v/v) as the mobile phase. The method is applied to 35 patients with the classic form of 21-OHD (18 females, 17 males, 5.1-14.2 years old) and 2 with 11beta-OHD (1 female, 1 male, 9.5 and 12.6 years old). Thirteen control children (5 females and 8 males, 5.2-15.2 years) are also studied. The results obtained for all measured steroids are compatible with those reported in the literature. The method is precise, and recovery is adequate. The HPLC technique proved to be of value for the purification of several steroids from single serum samples prior to RIA in patients with CAH.

Hereditary corticosteroid-binding globulin deficiency due to a missense mutation (Asp367Asn, CBG Lyon) in a Brazilian kindred.

OBJECTIVE: Abnormal corticosteroid-binding globulin (CBG) is an extremely rare condition and only three mutations have been described in four families. The molecular basis of an abnormal CBG in a Brazilian family was studied and correlations between genotype and serum cortisol, cortisol binding capacity (CBC) and CBG levels were determined. SUBJECTS: All 10 family members, comprising three generations, and nine healthy volunteers were studied. MEASUREMENTS: Genomic DNA was extracted from white blood cells from all family members. The human cbg exons 2-5 were amplified by PCR, submitted to automatic sequencing. Cortisol and CBG levels in serum were measured by radioimmunoassay (RIA). CBC in serum was determined using tritiated cortisol and other cortisol binding parameters were calculated through Scatchard analysis. RESULTS: A missense mutation in exon 5 of cbg (1254G --> A; Asp367Asn), recently described as CBG Lyon, was found in all family members. The proband and one sister were homozygous whereas all other family members, including parents, were heterozygous for this mutation. Cortisol levels in the only two homozygotes were lower than in heterozygotes and both were significantly lower as compared to controls (69 and 182 nmol/l vs. 267 +/- 129 nmol/l vs. 459 +/- 195 nmol/l, respectively, P < 0.05). CBC was decreased in the two homozygotes as compared to heterozygotes and in both groups as compared to controls (< 90 and 114 nmol/l vs. 305.0 +/- 81.4 nmol/l vs. 594.8 +/- 59.5 nmol/l, respectively, P < 0.05). CBG levels were lower in homozygotes as compared to heterozygotes and in both as compared to controls (325 and 375 nmol/l vs. 496.75 +/- 50.75 nmol/l vs. 647.25 +/- 87.50 nmol/l, respectively, P < 0.05). CONCLUSIONS: An abnormal CBG resulting from a missense mutation and known as CBG Lyon was found in this Brazilian kindred. This abnormal CBG has decreased affinity for cortisol and results in low or low normal serum cortisol levels in homozygous and heterozygous subjects. Although relative hypotension and fatigue have recently been associated with CBG deficiency in a family with two CBG mutations (null and Lyon), the two homozygous subjects in this kindred were both normotensive and only the proband presented with fatigue.

Relação dos níveis séricos de cálcio e paratormônio com a positividade da cintiligrafia das paratiróides / Relation of serum calcium and parathormone levels and positivity of parathyroid scintigraphy

O hiperparatiroidismo primário (HPP) é caracterizado pelo aumento da secreção de PTH, com conseqüente aumento da concentração sérica de cálcio. O diagnóstico é realizado pela dosagem de cálcio e PTH. A cintilografia das paratiróides é solicitada, classicamente, em pacientes com recorrência de HPP após paratiroidectomia, na tentativa de detectar glândulas patológicas ectópicas ou remanescentes. Algumas vezes este exame tem sido solicitado antes do primeiro ato cirúrgico, na tentativa de localizar as glândulas comprometidas e abreviar a duração da cirurgia; na nossa casuística, no entanto, a maioria dos exames solicitados com esta indicação resulta negativa. Comparamos os níveis séricos de cálcio e PTH nos pacientes com cintilografia das paratiróides positiva, com aqueles dos pacientes com cintilografia negativa, a fim de tentar definir níveis com índice maior de positividade na cintilografia. Foram estudados retrospectivamente 74 pacientes consecutivos submetidos à cintilografia das paratiróides. Avaliou-se a utilização dos valores mais baixos de PTH (79pg/mL) e cálcio (l0mg/dL) registrados no grupo com cintilografia positiva como referência para a indicação do exame. No grupo total de pacientes, 17 (23 por cento) apresentaram cintilografia positiva. No subgrupo com valores de PTH >79pg/mL e de cálcio >_10 mg/dL a porcentagem de exames positivos foi de 49 por cento. Concluímos que cintilografias das paratiróides realizadas antes da cirurgia de paratiroidectomia em pacientes com níveis de cálcio sérico abaixo do limite superior da normalidade ou níveis de PTH pouco aumentados são, na maioria das vezes, negativas.