Plasma levels of chromogranin A (CgA) were measured by ELISA in 22 patients with pheochromocytoma (18 non-metastatic, 3 metastatic, and 1 mixed neuroendocrine-neural tumor), 9 patients with primary hyperparathyroidism, and 9 patients with pituitary adenoma. The plasma levels of CgA were compared with norepinephrine, epinephrine, parathyroid hormone and pituitary hormones, i.e., growth hormone and prolactin. In pheochromocytoma, CgA in preoperative plasma of the patients without metastasis was 228 +/- 38 U/L (mean +/- SEM) and significantly higher than healthy controls (30 +/- 11 U/L, n = 40). Plasma CgA was decreased after removal of the tumors (28 +/- 6.0 U/L), except in three patients with metastatic pheochromocytoma and a mixed neuroendocrine neural tumor. The concentration of CgA in the patients with non-metastatic pheochromocytoma was significantly correlated with that of plasma norepinephrine (P < 0.005, r = 0.68) and urinary norepinephrine (P < 0.05, r = 0.65), but not with that of epinephrine. There was an exceptional case in which CgA was extremely high, but the CA level was normal. This tumor was a highly malignant pheochromocytoma with extensive metastases composed of small tumor cells which were occasionally positive for tyrosine hydroxylase immunohistochemically. These cells were considered to be poorly differentiated tumor cells and synthesized a very small amount of norepinephrine. Plasma levels of the patients with primary hyperparathyroidism and the patients with pituitary adenoma were 44 +/- 4 U/L and 48 +/- 8 U/L, respectively. Only one patient with a growth hormone-producing pituitary adenoma had a high level of CgA. Plasma CgA is a useful tumor marker for pheochromocytoma, even for malignant pheochromocytoma without elevated CA level, but not for hyperparathyroidism, or pituitary adenoma.

Urinary excretion of calcium (Ca) was measured in 9 patients with pseudohypoparathyroidism (PHP) type I—3 with Albright's hereditary osteodystrophy (AHO): AHO(+) and 6 without AHO: AHO(−)—and in 13 with idiopathic hypoparathyroidism (IHP), treated with active vitamin D3 (1,25(OH)2D3 or 1αOHD3) to maintain serum Ca levels at 8.4–9.5 mg/dl. Fasting urinary excretion of Ca in PHP was significantly lower than that in IHP. Moreover, fasting urinary excretion of Ca in PHP AHO(+) was lower than that in PHP AHO(−). This difference was also seen in the urine after oral loading of Ca. Urinary excretion, of sodium (Na) was not different between PHP AHP(+) and PHP AHO(−). Serum levels of immunoreactive PTH in PHP AHO(+) were higher than those in PHP AHO(−). The difference in urinary excretion of Ca between PHP AHO(+) and PHP AHO(−) may come from the difference in the circulating levels of PTH.

Serum osteocalcin was measured in patients with idiopathic hypoparathyroidism or pseudohypoparathyroidism, before or during the treatment with active vitamin D3 (1,25(OH)2D3 or 1 alpha OHD3). Serum osteocalcin and plasma 1,25(OH)2D were decreased in 11 patients with idiopathic hypoparathyroidism before treatment (2.8 +/- 1.27 ng/ml, P less than 0.001 and 14.3 +/- 4.27 pg/ml, P less than 0.001, respectively). In 24 patients with idiopathic hypoparathyroidism during the treatment, serum osteocalcin and plasma 1,25(OH)2D were within the normal range (4.5 +/- 0.74 ng/ml and 25.7 +/- 5.69 pg/ml, respectively). In five patients with pseudohypoparathyroidism before treatment, plasma 1,25(OH)2D was decreased (15.6 +/- 10.6 pg/ml, P less than 0.001) but serum osteocalcin was normal (7.8 +/- 1.66 ng/ml). In nine patients with pseudohypoparathyroidism during the treatment with active vitamin D3, serum osteocalcin and plasma 1,25(OH)2D were normal (6.8 +/- 1.47 ng/ml and 27.2 +/- 6.0 pg/ml, respectively). Serum PTH in pseudohypoparathyroidism was increased before treatment (0.70 +/- 0.34 ng/ml, P less than 0.05) and was normal during the treatment (0.50 +/- 0.13 ng/ml). In idiopathic hypoparathyroidism, the active vitamin D3 increased serum osteocalcin without PTH. In pseudohypoparathyroidism, PTH may increase serum osteocalcin or modulate the effect of active vitamin D3 on serum osteocalcin.

Plasma 1,25-dihydroxyvitamin D (1,25-(OH)2D) level, which is considered to be an indicator of parathyroid function, is possibly modified by the level of vitamin D. In the present study, we have investigated parathyroid function in terms of enhancement of the plasma levels of 1,25-(OH)2D after oral administration of 100 micrograms of 25-hydroxyvitamin D3 (25OHD3) in 9 cases of primary hyperparathyroidism (1 degree HPT), 7 cases of hypoparathyroidism (HP), 2 cases of pseudohypoparathyroidism (PHP) and 6 normal subjects. The plasma levels of 25-hydroxyvitamin D (25OHD) increased and reached a peak at 6-12 hours after the administration of 25OHD3. The plasma levels of 1,25-(OH)2D slightly increased but remained within the normal range after 25OHD3 administration in 3 of the normal subjects whose basal levels were rather low, but the increase in plasma 1,25-(OH)2D in control subjects was not statistically significant. In cases of 1 degrees HPT, the plasma 1,25-(OH)2D level rose significantly in all cases (P less than 0.05), although the pattern of the increase was not uniform. These increases were remarkable in the patients whose basal levels were low. On the other hand, an increase in the level was rarely observed in any of the cases of HP and in one of the cases of PHP. In another case, normocalcemic PHP, the plasma 1,25-(OH)2D level rose.(ABSTRACT TRUNCATED AT 250 WORDS)

Three cases from two families with idiopathic hypoparathyroidism and progressive sensorineural deafness are described. Cases 1 and 2 were siblings. Case 3 was one of four siblings from another family. All of them had both idiopathic hypoparathyroidism and progressive sensorineural hearing loss. There was no evidence to suggest involvement of autoimmune mechanism in these cases except for the associated Graves' hyperthyroidism in case 3. Human leukocyte antigen A9 and A11 were positive in both families. The sensorineural hearing loss was progressive even after the treatment for hypoparathyroidism. As the familial idiopathic hypoparathyroidism is a very rare entity, it is unlikely that this disease is associated with familial progressive sensorineural deafness by chance. The combination of these two diseases may compose a new syndrome.

Several problems in the measurement of plasma cyclic AMP (PcAMP) and nephrogenous cyclic AMP were studied using a YAMASA RIA Kit (YAMASA Shoyu, Choshi, Japan). In this assay method, cAMP in plasma is directly succinylated without prior deproteinization, and then it is bound to antibody in an imidazole buffer. So far as the blood samples were obtained with EDTA-4Na at least 5.0 mM in the final concentration, PcAMP was not reduced until 16 hours after the blood samples were drawn. Even without EDTA, the reductions in PcAMP were not detected within 1 hour after the blood samples were drawn. This assay method for PcAMP showed parallelism in the dilution curve. Recovery was almost complete. Intra- and interassay variations were low. When plasma was incubated at 37 degrees C for 24 hours, PcAMP became negligible. Furthermore, the values of PcAMP measured with this direct assay system almost agreed with those obtained after the purification by deproteinization and Dowex column chromatography through an anion-exchange resin. The normal values of PcAMP were 13.6 +/- 3.62 pmol/ml [mean +/- SD, n = 43]. Nephrogenous cAMP expressed as a function of GFR never did show any negative values in various clinical situations. From the data of basal levels and the oral calcium tolerance test, nephrogenous cAMP appeared to be more useful than total urinary cAMP in the diagnosis of parathyroid disorders, especially hyperparathyroidism.

The parameters of calcium metabolism were determined in 22 patients with untreated hyperthyroidism (5 males and 17 females) and 5 control subjects. Hypercalcemia was found in the patients with hyperthyroidism in comparison with the control subjects (serum Ca: 10.0 +/- 0.56 vs. 9.0 +/- 0.18, p less than 0.001 and Ca++: 5.1 +/- 0.28 vs. 4.6 +/- 0.15 mg/dl, p less than 0.001, mean +/- SD). Although the urinary excretion of calcium was decreased in many patients, abnormalities of phosphate metabolism were not found in this study. The parameters of bone resorption, urinary hydroxyproline, serum alkaline phosphatase and acid phosphatase, were increased in all patients with hyperthyroidism. Serum immunoreactive PTH was decreased (0.23 +/- 0.05 vs. 0.29 +/- 0.05 ngEq/ml, p less than 0.05). In vitamin D metabolites, 25-OH-D did not differ from the control (16.9 +/- 7.76 vs. 17.9 +/- 5.52 ng/ml), 1,25-(OH)2D showed a tendency to decrease (32.6 +/- 19.53 vs. 37.2 +/- 13.75 pg/ml) and 24,25-(OH)2D was obviously increased (5.57 +/- 3.582 vs. 1.73 +/- 0.619 ng/ml, p less than 0.001) in the hyperthyroid patients. Thus, the parathyroid function was suppressed in the patients with hyperthyroidism, and hypercalcemia in hyperthyroidism was suggested to be due to the direct action of thyroid hormone upon the bone.

The present study was undertaken to establish a standard method to perform the Ellsworth-Howard test using human PTH-1-34). For this purpose we made a survey of literature concerning the Ellsworth-Howard test and then examined the data of the Ellsworth-Howard tests performed on 178 hypoparathyroid patients using human PTH-(1-34). The main items of investigation were: (i) to determine the appropriate dose of PTH for administration to adults and children; and (ii) to define the criteria of practical usefulness for the differential diagnosis of the types of hypoparathyroidism. From the analysis of the data, the following findings and conclusions were obtained. The dose of human PTH-(1-34) appropriate for diagnostic use is 100 U per person for adults and 100 U per body surface area of one square meter (100 U/m2) for children. The criteria of positive response in the Ellsworth-Howard test are defined as follows. a) phosphaturic response: (U4 + U5) - (U2 + U3) = more than 35 mg/2 h b) cyclic AMP response: U4 - U3 = more than 1 mumol/h, and U4/U3 = more than 10 times. In the above formula, U2 - U5 represent the urine samples collected hourly in order. PTH is injected at the time between U3 and U4. For the application of the criteria in children, one should use the values corrected for body surface area of one square meter. It is necessary to confirm the following conditions before the application of the criteria: the presence of hypocalcemia and hyperphosphatemia; the lack of phosphate deficiency (basal urinary phosphate excretion more than 10 mg/2 h); the accuracy of timed urine collections (ratio of creatinine excretion during 2 hours before PTH to that after PTH administration in the range from 0.8 to 1.2); and the absence of marked diurnal variation in phosphate excretion (difference in phosphate excretion between the two basal hourly urine less than 17.5 mg/h). To ensure the above conditions, medications such as phosphate-binding antacids should be withheld for at least 1 week before the test, and the test should be performed according to the standard procedure described in this paper. The diagnosis of pseudohypoparathyroidism Type II should be done cautiously. It is necessary to take account of the high basal urinary cyclic AMP excretion and the elevated serum PTH level along with the results of the Ellsworth-Howard test (positive cyclic AMP response and negative phosphaturic response) for a definite diagnosis of this entity.

The effect of synthetic 1-34 fragment of human parathyroid hormone (hPTH(1-34] on plasma adenosine 3',5'-monophosphate (cAMP) in human subjects and the diagnostic criteria for the plasma cAMP response in an Ellsworth-Howard test were studied. 20 or 30 micrograms hPTH(1-34) and 200 USP Parathormone (Eli Lilly & Co.), infused intravenously over 5 min, produced very similar patterns of response in plasma cAMP, peak values being observed within 5 or 10 min after the end of the infusion. The maximum levels of plasma cAMP were over 111.5 pmol/ml in all of the normal subjects (n = 5) and patients with idiopathic hypoparathyroidism (n = 22), including those of children, but the plasma cAMP did not rise above 65.0 pmol/ml in pseudohypoparathyroidism (n = 7). There existed a significant correlation between the maximum plasma cAMP concentrations and increases in urinary cAMP excretion after infusions of both hPTH(1-34) and Parathormone. These results suggest that hPTH(1-34) has effects essentially identical to those of native PTH on plasma cAMP. We would like to propose a new diagnostic criterion in the Ellsworth-Howard test: a peak value of plasma cAMP over 100 pmol/ml after 30 micrograms hPTH(1-34) infusion is regarded as a normal response.

A 29-year-old man came to our hospital with tetany and sensorineural deafness. Routine laboratory testing revealed a serum calcium of 5.5 mg/dl and a serum phosphate of 6.1 mg/dl. The level of circulating immunoreactive parathyroid hormone was undetectable, and he showed hyperresponsiveness to the exogenous parathyroid hormone (synthetic hPTH (1-34), 100U). Thus, he was diagnosed as having idiopathic hypoparathyroidism. The other endocrine functions were normal. The parathyroid antibody was negative and no moniliasis was found. On the examination of his family, it was found that his male sibling and one of his female siblings also had hypocalcemia (6.4 mg/dl and 6.0 mg/dl) and undetectable levels of circulating immunoreactive parathyroid hormone, and all his siblings, his father and one of his nieces had sensorineural deafness, so he was diagnosed as as having familial idiopathic hypoparathyroidism and familial progressive sensorineural deafness. In addition to this case, we have experienced two siblings of another family having the same combination of these disorders (unpublished observation). Since familial idiopathic hypoparathyroidism is not common, it seems that the present combination of these disorders is not a chance association.

UNAKAMI, H., FURUKAWA, Y., SOHN, H.E., YUMITA, S., MIURA, R., SASAKI, A., KOKUBUN, M., MIURA, Y., YOSHINAGA, K. and NAKANOME, C. 1, 25-Dihydroxyvitamin D Production Stimulated by Dibutyryl 3', 5'-Cyclic AMP in Normal Subjects and a Patient with Pseudohypoparathyroidism. Tohoku J. exp. Med., 1982, 138 (4), 411-418-Dibutyryl 3', 5'-cyclic AMP (DBcAMP) was infused in 4 normal subjects and a patient with pseudohypoparathyroidism (PHP) to study its effect on the production of 1, 25-dihydroxyvitamin D [1, 25(OH)2D]. In normal subjects, 2.5mg/kg of DBcAMP increased plasma 1, 25(OH)2D. reaching the peak at 6hr after infusion, while, 6.0mg/kg of DBcAMP delayed the peak to 12hr. The maximal increment of plasma 1, 25(OH)2D was 19.7±2.8pg/ml (mean±S.D.) in normals with 6.0mg/kg of DBcAMP and 12.2±1.0pg/ml (mean±S.D.) with 2.5 mg/kg, respectively. There was a significant difference between the doses (p<0.01). Plasma 24, 25-dihydroxyvitamin D [24, 25(OH)2D] showed no significant change after DBcAMP infusion in normal subjects. In a patients with PHP, however, 2.5mg/kg of DBcAMP markedly increased plasma 1, 25(OH)2D to reach the peak (48.9pg/ml) at 14hr and plasma 24, 25(OH)2D was decreased reciprocally. After DBcAMP infusion, depression of serum P, slight decrease in %TRP and the elevation of IRI were proved in normal subjects, while in a patient with PHP, there was a marked depression of %TRP. It is suggsted that the increments of plasma 1, 25(OH)2D by infusion of DBcAMP is dose-dependent in normal subjects, and that the 1, 25(OH)2D productivity by DBcAMP may be accelerated in a patient with PHP.

Parathyroid extract (PTE) or synthetic 1-34 human parathyroid hormone (1-34 hPTH) was injected intravenously as a bolus in 4 normal subjects, 4 patients with PTH deficient hypoparathyroidism (HP) and 3 patients with pseudohypoparathyroidism (PHP). In normal subjects and HP, plasma 1,25(OH)2D was markedly increased at 6 hr and reached the peak at 12 or 14 hr after administration of 200 units of PTE or 20 to 30 micrograms of 1-34hPTH. On the other hand, 500 units of PTE or 20 micrograms of 1-34hPTH failed to increase plasma 1,25(OH)2D in PHP. However, 2.5 mg/kg of dibutyryl cAMP remarkably increased plasma 1,25(OH)2D in a patient with PHP. Maximal increments of plasma 1,25(OH)2D in 3 patients with HP(21.7 +/- 5.6 pg/ml, mean +/- S.D.) were nearly as high as in normal subjects (20.6 +/- 7.0 pg/ml), whereas those in 3 patients with PHP (2.3 +/- 2.3 pg/ml) were distinctly lower than in normal subjects or HP. It is suggested that 1,25(OH)2D production by PTH is intact in HP, but is impaired in PHP mainly due to a defect in the activation of adenylate cyclase system.

The effects of calcium injection (3 mg/Kg/10 min) or oral calcium administration (calcium lactate 7.7 g) on plasma iPTH and Nephrogenous cyclic AMP (NcAMP) were studied in 6 normal controls and 13 patients with primary hyperparathyroidism. In the control subjects, plasma iPTH determined by a predominantly carboxyl-terminal antiserum was less than 0.3 ng/ml before and after both calcium loads, whereas 41 approximately 98% (mean 67%) of NcAMP was rapidly and uniformly suppressed to a level lower than the normal value. In 2 patients with primary hyperparathyroidism, iPTH was clearly reduced from 8.0 to 4.6 ng/ml and 1.6 to 0.96 ng/ml, respectively, by the calcium load. However, in the other 7 patients with primary hyperparathyroidism who showed only a slight elevation of iPTH: less than 0.3 approximately 0.9 ng/ml, the reductions in iPTH were not detected after the calcium load: less than 0.3 approximately 0.7 ng/ml. In contrast, 30 approximately 54% (1.02 approximately 3.85 nmol/dl GF) of NcAMP, which was greater than the diurnal variation, was suppressed after calcium injection in 5 patients with primary hyperparathyroidism (2 of 4 patients with urological, and 3 of 5 patients with chemical hyperparathyroidism). But NcAMP was not suppressed in all 4 patients with skeletal hyperparathyroidism including one with proximal renal tubular dysfunction whose basal iPTH was elevated markedly but reduced clearly by the calcium load. In general, suppression of NcAMP was followed by a decrease of phosphate excretion. On the other hand, even in a patient with primary hyperparathyroidism whose NcAMP was not suppressed at all after the calcium injection, calcium infusion (15 mg/Kg/3h) resulted in some (23%) decrease in NcAMP. Oral calcium administration resulted in responses which were almost the same as those produced by calcium injection. These results suggest that NcAMP provides a useful index in the parathyroid suppression test in patients with primary hyperparathyroidism, especially those who display a rather mild elevation of iPTH. This is not the case, however, in a few patients who show a marked elevation of iPTH and/or proximal renal tubular dysfunction.