Two-site assay of intact parathyroid hormone in the investigation of primary hyperparathyroidism and other disorders of calcium metabolism compared with a midregion assay

We compared the utility of measurements of serum intact human PTH-(1-84) and midregion human PTH-(44-68) in patients with disorders of extracellular calcium metabolism. Serum midregion PTH was determined by RIA, and serum intact PTH was measured by a sensitive and specific immunoradiometric two-site assay. The serum intact PTH concentrations in 70 patients with primary hyperparathyroidism were above the normal range in 69, and thus widely separated from the levels in 40 patients with hypercalcemia of malignancy, in whom serum intact PTH values were usually below normal. In contrast, both groups had overlapping serum midregion PTH values. In patients after renal transplantation and those with chronic renal failure, serum intact PTH levels were in the normal range twice as often as were serum midregion PTH values. The intact PTH assay was also superior in detecting venous gradients of the hormone and changes in PTH secretion caused by altered serum calcium concentrations, and serum intact PTH was remarkably low in hepatic venous effluent. We conclude that this new assay for serum intact PTH is superior to the midregion RIA in investigating parathyroid function in several different clinical conditions.     

CONTROL OF SECRETION OF PARATHYROID HORMONE IN SECONDARY HYPERPARATHYROIDISM

The response of the parathyroids to an infusion of calcium has been studied in patients with secondary hyperparathyroidism due to either vitamin D deficiency or to chronic renal failure. Two specific homologous immunoradiometric assays for human parathyroid hormone (PTH) have been used to assess the response, one specific for the amino-terminus (N-PTH), the other specific for the carboxy-terminus (C-PTH) of the molecule.
In both the vitamin D deficient and the uraemic patients, in response to a 4 mg/kg/h infusion of calcium generally for 4 h, suppression of N-PTH was similar, falling to around 20–30% of the initial value. In the uraemic subjects, the degree of suppression was inversely related to the initial plasma calcium ( r = 0·9, P < 0·001). Measured with the C-PTH assay, the response was generally less, particularly in the uraemic subjects in whom it was often preceded by an initial rise in C-PTH.
In both groups of subjects, suppression of N-PTH began as soon as the plasma calcium began to rise and before hypercalcaemia was produced. At the end of the infusion the concentration of N-PTH rose quickly, although the plasma calcium was still high. In four uraemic subjects the rate of infusion of calcium was reduced after 1–1·5 h to limit the rise in plasma calcium. As the calcium approached a plateau, the concentration of N-PTH was found to rise again.
These results indicate the importance of using PTH assays of well defined specificity to evaluate autonomy, and show that the degree of suppression achieved is dependent both on renal function and basal plasma calcium. In addition, these observations suggest that the direction and rate of change of plasma calcium are important in the control of secretion of parathyroid hormone.     

Malignancy hypercalcemia: evaluation of parathyroid function and response to treatment

In 53 patients with malignancy-associated hypercalcemia, we measured serum parathyroid hormone (PTH) with an immunoradiometric assay (IRMA) for intact PTH and a midregion-specific radioimmunoassay (RIA). Values were measured at baseline to detect clinically unrecognized hyperparathyroidism and after treatment with intravenous bisphosphonate to test for a parathyroid response. One patient probably had primary hyperparathyroidism, since his serum PTH values were not appropriately suppressed at baseline and increased markedly during treatment before normocalcemia was achieved. In each of the 51 other evaluable cases, the intact PTH value was suppressed below 25 pg/ml (normal range 10-55 pg/ml), confirming a nonparathyroid hypercalcemia. Midregion PTH values were less fully suppressed but supported the diagnosis in 44 out of 49 cases. The five patients with high midregion PTH values each had renal insufficiency but in four, the intact PTH values were also the highest we observed in these patients. This suggests either a poorly suppressible intact PTH secretion or prolongation of intact PTH half-life by the renal insufficiency. Overall, midregion and intact PTH values were highly correlated (p less than 0.001). Restoration of normocalcemia increased PTH in both assays into or within the normal range, and when posttreatment hypocalcemia occurred, PTH values became elevated in both assays. We conclude the following: (1) both assays are useful for detecting parathyroid hyperfunction but the intact PTH assay is the better diagnostic tool when renal insufficiency is present, (2) hypercalcemia seems to suppress intact PTH secretion more fully than secretion of PTH fragments, and (3) parathyroid glands chronically suppressed by hypercalcemia can increase PTH secretion within 1 or 2 days after the hypercalcemia is corrected.     

Parathyroid hormone and parathyroid hormone-related protein in the investigation of hypercalcaemia in two hospital populations

BACKGROUND AND OBJECTIVE Parathyroid hormone-related protein (PTHrP) produced by cancers has a central role as a humoral mediator of hypercalcaemia in patients with malignancy. Since the prevalences of hypercalcaemia of malignancy and parathyroid disease reflect the population studied, hypercalcaemia patients from a district general hospital (DGH) and an oncology centre (OC) were studied in order to assess the diagnostic role of assays for serum PTH and plasma PTHrP in different clinical settings. DESIGN A prospective study of consecutive patients presenting with their first episode of hypercalcaemia during an 18-month period. PATIENTS A total of 123 patients (DGH, n= 69; OC, n= 54) had corrected serum calcium concentrations > 2·65mmol/l. MEASUREMENTS PTH, PTHrP, calcium and albumin were measured together with urine calcium and creatinine, enabling fractional calcium excretion to be assessed. Urine cyclic adenosine monophosphate was also measured. RESULTS Hypercalcaemia was attributed to malignancy alone in 72 patients (DGH, n= 20; OC, n= 52), benign causes in 42 (DGH, n= 42) and parathyroid disease coexisting with malignancy in 9 (DGH, n = 7; OC, n= 2). Plasma PTHrP levels were increased in 59/72 (82%) patients with hypercalcaemia due to malignancy. Measurements of both analytes contributed to a change in the final diagnosis in 12 patients (10%). Thus serum PTH was Increased In seven patients with parathyroid disease coexisting with malignancy, and plasma PTHrP was increased in five patients with previous undiagnosed malignancy. Median survival for patients with parathyroid disease and coexisting malignancy was 13 months compared with 3 months for those with hypercalcaemia due to malignancy alone (P < 0.02). CONCLUSIONS Since hypercalcaemia was attributable to parathyroid disease In 10% of ail patients with malignancy we advise measurement of serum PTH at the Initial presentation of hypercalcaemia in ail patients, while plasma PTHrP may be useful to identify those patients in whom malignancy may not be clinically apparent, or coexist with primary hyperparathyroidism.     

Overproduction of an amino-terminal form of PTH distinct from human PTH(1-84) in a case of severe primary hyperparathyroidism: influence of medical treatment and surgery

OBJECTIVE: Rare patients with severe primary hyperparathyroidism present with large parathyroid tumours, severe hypercalcaemia, very high PTH levels and osteitis fibrosa cystica. Some of these patients display a large amount of C-PTH fragments in circulation and present with a higher C-PTH/I-PTH ratio than seen in less severe cases of primary hyperparathyroidism. We wanted to determine how PTH levels and circulating PTH high-performance liquid chromatography (HPLC) profiles analysed with PTH assays having different epitopes could be affected by medical and surgical treatment in such patients. DESIGN: A 55-year-old man with severe hypercalcaemia (Ca(2+): 2.01 mmol/l), very high PTH levels (CA-PTH 82.1 and T-PTH 72 pmol/l) caused by a large parathyroid tumour (7.35 g) and accompanied by significant bone involvement (alkaline phosphatase of 185 UI/l and subperiostal bone resorption of hands) was referred to us. Blood was obtained at various time points during his medical treatment, before and after surgery, to measure parameters of calcium and phosphorus metabolism, and of bone turnover. HPLC separations of circulating PTH molecular forms were performed and analysed with PTH assays having 1-4 (CA), 12-18 (T), 26-32 (E) and 65-84 (C) epitopes. RESULTS: Before surgery, serum Ca2+ was nearly normalized with hydratation, intravenous (IV) pamidronate and oral vitamin D administration. Despite a decrease in Ca2+ to 1.31 mmol/l, CA-PTH and T-PTH levels decreased by half in relation to a threefold increase in basal 1,25-dihydroxyvitamin D [1,25(OH)2D] level (94 to 337 pmol/l). After this initial positive response, hypercalcaemia and elevated CA- and T-PTH levels recurred even if 1,25(OH)2D levels remained elevated. The tumour was removed surgically and proved to be poorly differentiated with nuclear atypia and mitosis. After surgery, the Ca2+ level and PTH secretion normalized. The higher CA-PTH level relative to the T-PTH level observed before surgery in this patient was related to the oversecretion of an amino-terminal (N) form of PTH recognized by PTH assays with (1-4) or (26-32) epitopes but not by the T-PTH assay with a (12-18) epitope. This molecular form represented 50% of CA-PTH measured in this patient, but only 7% in less severe cases of primary hyperparathyroidism. It was unaffected by medical therapy and disappeared after surgery. CONCLUSION: The relationship between the overexpression of this N-PTH molecular form and severe primary hyperparathyroidism remains unclear. Further studies will be required in these rare patients to see whether N-PTH is a marker of less well differentiated parathyroid tumours and/or relates to the overproduction of C-PTH fragments in the presence of severe hypercalcaemia.     

Parathyroid hormone-related peptide, measured by a midmolecule radioimmunoassay, in various hypercalcaemic and normocalcaemic conditions.

The diagnosis of humoral hypercalcaemia of malignancy often presents considerable clinical problems. We have studied parathyroid hormone-related peptide (PTHrP) in serum from patients with humoral hypercalcaemia of malignancy (N = 22), hypercalcaemia of malignancy with skeletal metastases (17), histologically confirmed primary hyperparathyroidism (21) and hypercalcaemic patients with various benign diseases (9). PTHrP measurements were also made in normocalcaemic patients with various malignancies (23), endocrine diseases (13), sarcoidosis (22) and chronic renal failure (17). PTHrP was measured by a novel radioimmunoassay using rabbit antibodies directed towards the midregion of the molecule. Immuno- or silica cartridge extraction of serum before radioimmunoassay enabled us to measure PTHrP in all samples, which may add further information about circulating forms of PTHrP. PTHrP was clearly elevated in patients with humoral hypercalcaemia of malignancy (5.0 +/- 4.7 pmol/l) (mean +/- SD, N = 12) and when the kidney function was impaired (4.0 +/- 0.9 pmol/l) (N = 15) (silica cartridge extraction), whether the subject was hypercalcaemic or not. Some patients with endocrine diseases, including two with primary hyperparathyroidism, had slightly elevated serum PTHrP concentrations, while they were normal in sarcoidosis. In healthy subjects the levels were 1.1 +/- 0.5 pmol/l (N = 15) after immunoextraction and 0.8 +/- 0.2 pmol/l (N = 33) after silica cartridge extraction.     

Sodium bicarbonate infusion test: a new method for evaluating parathyroid function

We have developed a new test for estimating the secretory capacity of parathyroid hormone (PTH) from the parathyroid gland. Sodium bicarbonate solution [8.4% (w/v); 35 ml/m(2) body surface area] was infused for 2 min, and blood samples for the determination of plasma ionized calcium, plasma PTH (intact, midregion, carboxy-terminus) and related parameters were serially obtained. In 8 healthy volunteers, the mean (+/-SE) plasma ionized calcium fell promptly and significantly (from 1.21 +/- 0.01 to 1.11 +/- 0.01 mmol/L) after the sodium bicarbonate infusion. The mean (+/-SE) plasma intact PTH increased promptly and significantly, by more than four fold (42.3 +/- 4.2 to 182.4 +/- 34.7 pg/ml), and then gradually returned to basal levels. In patients with partial hypoparathyroidism who have detectable basal plasma levels of PTH, the absolute increment in PTH levels was much less, and in the plasma obtained from patients with complete hypoparathyroidism, absolutely no response was observed. Plasma obtained from patients diagnosed with primary hyperparathyroidism (parathyroid adenoma or hyperplasia) has high basal PTH levels. The response to the sodium bicarbonate infusion in these patients was markedly blunted (less than a two-fold increase in all cases examined). No significant adverse effects were observed during the procedure. Therefore, the sodium bicarbonate infusion test is a simple and sensitive method to stimulate PTH release, and is clinically useful for evaluating parathyroid gland function.     

Comparison of intact, midregion, and carboxy terminal assays of parathyroid hormone for the diagnosis of bone disease in hemodialyzed patients

The predictive value of three different RIAs of PTH for the diagnosis of the histological type of bone disease has been compared in 24 asymptomatic patients on chronic hemodialysis who had never been exposed to aluminum intoxication and who agreed to have a bone biopsy after double tetracycline labeling. The serum concentrations of PTH were measured using a two-site immunoradiometric assay for intact PTH(1-84) and region specific assays directed against the C-terminal (53-84) fragment or the midregion (44-68) of the molecule. The bone histomorphometric analysis showed that six patients had nonaluminic adynamic bone disease with low bone formation rate (BFR), eight had mild hyperparathyroidism characterized by increased bone resorption and normal BFR, nine had severe hyperparathyroidism with increased BFR, and only one had true osteomalacia with increased osteoid seam thickness. All PTH assays correlated with the various parameters of bone resorption and bone formation and were able to differentiate the histological type of bone disease only when groups of patients were considered. For classifying individual patients into severe hyperparathyroidism and adynamic bone disease groups, the intact PTH assay had the best predictive value with a sensitivity of 100% and a specificity of at least 70%. A nonaluminic adynamic bone disease was observed in more than 50% of the patients who had normal intact PTH levels (6/11). It is concluded that the intact PTH measurement is superior to C-terminal and midregion assays for the prediction of the histological type of bone disease in hemodialyzed patients and should be of considerable value to adapt their treatment in order to avoid the emergence of both severe hyperparathyroidism and adynamic bone disease. In the absence of aluminum intoxication it seems that maintaining intact PTH concentrations 1 to 1.5 times the upper limit of normal would correspond to the best bone histology.     

Clinical Studies on Phosphate Handling in Hypercalcaemia

ABSTRACT. Phosphate indices (serum phosphate, tubular reabsorption of phosphate, renal threshold phosphate concentration (TmP/GFR) and index of phosphate excretion) were studied in 88 hypercalcaemic subjects: 64 with primary hyperparathyroidism (HPT) and 24 with hypercalcaemia from other causes, predominantly malignant disease. HPT patients as a group could easily be separated from normal subjects (n = 16) and patients with functional hypoparathyroidism (n =7) by use of the phosphate variables but these indices were of little discriminating value for the differential diagnosis between HPT and hypercalcaemia from other causes. There was no difference in the urinary cyclic adenosine monophosphate (cAMP) excretion between the two hypercalcaemic patient groups, but HPT patients had clearly elevated serum parathyroid hormone (PTH) levels compared with normal PTH concentrations in patients with other causes of hypercalcaemia. A positive correlation between cAMP and serum calcium and an inverse relationship between cAMP and TmP/GFR were found in patients with hypercalcaemic malignant disease. These findings suggest the existence of a humoral factor with PTH-like effects in malignant disease. Since PTH levels were low, the physiological actions were apparently not mediated by circulating PTH. No difference in the values for phosphate variables, PTH, cAMP or serum calcium was found between renal stone-forming and stone-free patients with HPT.     

Lithium treatment increases intact and midregion parathyroid hormone and parathyroid volume

Lithium carbonate is known to alter calcium metabolism by lowering urinary calcium excretion and increasing serum calcium concentrations. Several investigators have reported increases in serum immunoreactive PTH (iPTH) values after a few weeks or months of lithium treatment, and several cases of primary hyperparathyroidism developing during lithium treatment have been reported. To determine whether the increases in serum iPTH might be the result of increased renal retention of inactive PTH fragments rather than stimulation of parathyroid function, we measured plasma intact PTH by immunoradiometric assay and estimated parathyroid size by ultrasonography in men who had received short term (less than 6 months) or long term (greater than 3 yr) lithium treatment and in normal subjects. Serum ionized calcium was higher by 0.03-0.04 mmol/L in subjects treated short term (mean, 1.7 months) with lithium than in normal subjects, but plasma intact PTH and serum midregion iPTH values were not different. The absence of a reciprocal decrease in PTH values is compatible with a lithium-induced shift in the set-point for the inhibition of PTH secretion by calcium toward a higher calcium value. Both plasma intact PTH and serum midregion PTH values were higher in subjects during longer term (mean, 103 months) lithium treatment, and estimated parathyroid volume was 3-fold higher. Serum phosphate was lower, and serum chloride and plasma 1,25-dihydroxy-vitamin D values were higher in those treated with lithium long term, probably from the biological action of the increased PTH. We conclude that long term lithium treatment increases circulating biologically active PTH and causes parathyroid enlargement. Whether this chronic stimulus to parathyroid growth might lead to adenoma formation in certain susceptible individuals and whether a PTH-induced increase in skeletal remodelling occurs that might hasten the appearance of osteopenia remain to be determined.     

Large carboxy-terminal parathyroid hormone (PTH) fragment with a relatively longer half-life than 1-84 PTH is secreted directly from the parathyroid gland in humans

OBJECTIVE: It was discovered that an immunoreactive large carboxy-terminal parathyroid hormone (PTH) fragment (large C-PTH), likely 7-84 PTH, is present in the circulation. However, very little is known about the production and metabolism of this large C-PTH. Combining a whole molecule PTH (whole PTH) immunoradiometric assay (IRMA) specifically for 1-84 PTH and an intact PTH (iPTH) IRMA for the sum of 1-84 PTH and large C-PTH, we were able to assess the circulating level of this large C-PTH as well as the glandular secretion and metabolism of this large C-PTH in primary hyperparathyroidism (pHPT). METHODS: This study consisted of two patient groups consisting of 77 pHPT patients with a single adenoma. Of these, 43 comprised the venous sampling study group and 70 comprised the intra-operative PTH study group. (Seven patients belonged only to the former group, 34 patients to only the latter group, and 36 patients to both groups.) Preoperatively, blood samples were drawn from the bilateral internal jugular vein by ultrasonographic guidance and from the peripheral vein (n=43). During surgery, blood samples were drawn after anesthesia (basal level), before excision (pre-excision level) of one enlarged parathyroid gland, and at 5, 10, and 15 min post-excision (n=70). RESULTS: There were 26 patients whose iPTH assay levels differed by more than 10% between the right and left internal jugular. In 24 of the 26 patients, the large C-PTH levels obtained from the adenoma side were significantly higher than those from the contralateral side (117+/-135 vs 43+/-33 pg/ml, P<0.001). The plasma whole PTH values decreased more rapidly than the iPTH values after parathyroidectomy (P<0.001). CONCLUSIONS: Our study has demonstrated that the large C-PTH, likely 7-84 PTH, is directly released from the parathyroid gland in humans. Since the half-life of 1-84 PTH is much shorter than large C-PTH, likely 7-84 PTH, it would be advantageous to use an assay that specifically measures 1-84 PTH for intra-operative monitoring of parathyroidectomy.     

The diagnostic value of a radioimmunoassay for parathyroid hormone in human serum.

A radioimmunoassay for the measurement of immunoreactive parathyroid hormone (PTH) in human serum is described. The assay is based on the ability of human parathyroid hormone (h-PTH) to compete with 125I-labelled bovine parathyroid hormone (b-PTH) for binding to a guinea-pig antiserum directed against b-PTH. The linear part of the standard curve was parallel with dose response curves for anti-b-PTH serum reacting with dilutions of sera from patients with primary hyperparathyroidism and from h-PTH purified from human parathyroid adenomas, indicating that levels of immunoreactive PTH could be expressed as b-PTH equivalents.The range in 62 healthy blood donors was 1.1-2.5 ng b-PTH Eg./ml. The reproducibility was satisfactory, and the sensitivity permitted the measurement of PTH concentrations down to 0.8 ng b-PTH Eg./ml. No crossreaction with h-CT, h-STH or h-ACTH was observed. The clinical value of the assay has been considered in a number of patients with various disorders of calcium metabolism, diagnosed and treated conventionally. About 80 per cent of patients with primary hyperparathyroidism had elevated PTH levels on one or more occasions before surgery. In patients with chronic renal failure of other aetiology than primary hyperparathyroidism the levels were usually far higher. Patients with primary hyperparathyroidism and increased S-creatinine had higher PTH levels than those with normal S-creatinine. After parathyroidectomy all previously increased PTH levels became normal or low. High PTH concentrations were found in 3 patients with normocalcaemic hyperparathyroidism who at operation were shown to have parathyroid adenomas. However, in normocalcaemic patients there were also some falsely elevated PTH values which limit the diagnostic value of the assay in this group of patients. Low PTH values were observed in patients with hypercalcaemia due to malignant disorders, indicating that PTH determination may be of some value in the diagnosis of patients with hypercalcaemia of unknown origin.     

Studies on the half-life of C-terminal parathyroid hormone in chronic renal failure

Subtotal parathyroidectomy (PTX) was performed for patients with hyperparathyroidism associated with chronic renal failure, and from sequential determination of PTH after PTX, the half-life of 2 different C-PTH was calculated. The half-life (T 1/2) of 46 approximately 84 PTH and 65 approximately 84 PTH was 18.1 +/- 3.9 and 14.0 +/- 2.2 hours (Mean +/- SD) respectively. Half-life of both C-PTH was markedly prolonged compared with that (30 approximately 60 minutes) of healthy subjects. It is considered that the longer half-life of 46 approximately 84 PTH than 65 approximately 84 PTH might be attributed to the difference of molecular weight, degrading enzymes and operative procedures. Determined value of C-PTH in chronic renal failure should be evaluated in consideration of prolonged metabolic clearance of C-PTH.     

Conventional and new diagnostic applications of a two-site immunochemiluminometric assay for parathyroid hormone.

This investigation was carried out to evaluate the clinical utility and diagnostic value of serum intact PTH measurement using a recently introduced immunochemiluminometric assay (ICMA). Studies were carried out in 42 normal subjects, 24 patients with primary hyperparathyroidism, 21 patients on chronic maintenance hemodialysis, 8 patients with postsurgical hypoparathyroidism, 7 patients with cancer hypercalcemia and 6 patients with osteomalacia. A good correlation was found in normal subjects between serum ICMA PTH levels and both intact PTH measured by a two-site immunoradiometric assay (n = 42, r = 0.67, p less than 0.001) and a widely used midmolecule radioimmunoassay (n = 21, r = 0.78; p less than 0.001). Similar good correlations were found in primary hyperparathyroidism patients (IC-MA vs immunoradiometric assay r = 0.74; p less than 0.001; ICMA vs midmolecule assay r = 0.77; p less than 0.001). As far as the hypercalcemic conditions were concerned, in 5 patients with mild primary hyperparathyroidism, ICMA PTH levels were in the upper range of those found in normal subjects, even though they were inappropriately high in respect to serum calcium values. However, serum ICMA PTH levels were clearly suppressed or undetectable in the majority of patients with cancer hypercalcemia or postsurgical hypoparathyroidism. Following calcium and EDTA infusions in patients with primary hyperparathyroidism, the behaviour of ICMA PTH levels in general parallelled that of immunoradiometric PTH assay, thus indirectly suggesting the ability of the method to measure the intact molecule.(ABSTRACT TRUNCATED AT 250 WORDS)     

Renal adenylate cyclase assay for biologically active parathyroid hormone: clinical utility and physiological significance

The stimulation of cyclic AMP production by human renal cortical membranes in the presence of the GTP analogue 5'-guanylimidodiphosphate and a calcium chelator represents a homologous assay system for the evaluation of biologically active parathyroid hormone (bioPTH) in human serum. Bioactive PTH was raised above normal (normal range: undetectable to 4.6 pmol human PTH(1-34) per 1) in 13/17 (76%) patients with primary hyperparathyroidism, in 5/6 (83%) patients with surgically proven hyperparathyroidism secondary to chronic renal failure, in 4/5 (80%) patients with hyperparathyroidism secondary to hypocalcaemia, in all three patients with pseudohypoparathyroidism, in 5/17 (29%) patients with osteoporosis and in 1/9 (11%) patients with renal stones and/or hypercalciuria. Bioactive PTH correlated positively with immunoreactive PTH (iPTH) measured with a radioimmunoassay predominantly recognizing the middle- and carboxyl-terminal region of the PTH molecule (r = 0.503, P less than 0.001). A positive correlation (r = 0.572, P less than 0.05) was found between values of serum calcium and bioPTH in the group with primary hyperparathyroidism. Immunoreactive PTH did not correlate significantly with calcium in this group. In the other patients except those who had chronic renal failure, a negative correlation between serum calcium and both bioPTH and iPTH was observed (P less than 0.01). When alkaline phosphatase was compared with bioPTH in all patients, the correlation was positive (r = 0.390, P less than 0.01); no significant correlation existed between iPTH and alkaline phosphatase in the patients studied.(ABSTRACT TRUNCATED AT 250 WORDS)     

DEVELOPMENT AND APPLICATION OF A MID-REGION SPECIFIC ASSAY FOR HUMAN PARATHYROID HORMONE

A new peptide spanning residues 28-54 of human parathyroid hormone (PTH) was synthesized and used to develop a homologous immunoradiometric assay specific for the mid-region of human PTH. The peptide was coupled to cellulose and used to absorb mid-region antibodies from a goat antiserum against intact human PTH. This assay has been applied to the measurement of circulating PTH in man: in normal subjects the concentration in serum ranged from undetectable (less than 40 pg/ml) to 70 pg/ml, the reference standard being the human PTH 28-54 peptide. In patients with primary hyperparathyroidism concentrations ranged from 120 to 1800 pg/ml. Hormone was not detected in patients with hypoparathyroidism. In normal subjects and in patients with primary hyperparathyroidism the mid-region PTH concentrations were similar to those obtained in an amino-terminal specific assay. By contrast, carboxy-terminal PTH concentrations were markedly higher being 10-fold greater in both groups studied. In patients with primary hyperparathyroidism undergoing parathyroidectomy and in chronic renal failure patients who were infused with calcium, mid-region and amino-terminal PTH disappeared much more rapidly than carboxy-terminal PTH. However, although mid-region PTH was initially cleared as quickly as amino-terminal PTH, it then reached a plateau and remained at a higher level. Thus the mid-region specific assay described here is proving to be of value in the study of the secretion and metabolism of PTH.     

Coincidental occurrence of primary hyperparathyroidism and cancer-associated hypercalcaemia in a middle-aged man

Primary hyperparathyroidism (PHPT) is found not uncommonly in patients with cancer. In this report, however, we describe a patient where both humoral hypercalcaemia of malignancy and PHPT were present coincidentally. A 47-year-old man was found to have PHPT due to parathyroid hyperplasia. Serum parathyroid hormone (PTH) levels, which were elevated before parathyroidectomy, were undetectable post-operatively; however, hypercalcaemia persisted. Nephrogenous cyclic adenosine monophosphate was elevated along with this undetectable PTH, indicative of the presence of a PTH-like factor in the serum. This was confirmed by the finding of an elevated level of PTH-related protein (PTHr P) in plasma (91 pmol/l, normal <2.6 pmoI/I). Secondary carcinoma was identified in a lesion in the region of the manubrium sternii. This stained positively for PTHr P by immunocyto-chemistry and PTHr P messenger RNA was detected by insitu hybridization. This case illustrates the value of sensitive PTH assays in distinguishing PHPT from other causes of hypercalcaemia and also shows the importance of considering primary hyperparathyroidism in the differential diagnosis of the patient with cancer and hypercalcaemia.     

Non-autonomy of parathyroid hormone secretion in acute primary hyperparathyroidism

A patient with acute primary hyperparathyroidism treated with mithramycin preoperatively, underwent neck exploration and two enlarged parathyroid glands were excised: one huge adenoma (6g) and another smaller gland. Mithramycin was administered preoperatively to lower life-threatening hypercalcaemia, and parathyroid slices from the huge adenoma removed at surgery were submitted in vitro to various calcium concentrations in the media to determine the influence of calcium on parathyroid adenoma secretory pattern in acute primary hyperparathyroidism. Mithramycin induced a significant decline in calcium levels and significant elevations of calciotrophic hormones (intact PTH, mid-region specific PTH, calcitonin and calcitriol). Significant suppression in PTH output in vitro was achieved by increasing calcium levels in the media. These results exclude autonomous PTH secretion (non-calcium dependent) as a possible aetiology of acute primary hyperparathyroidism. We suggest that a sudden increase in the set-point of the diseased parathyroid cells in the presence of a huge cell mass accounts, in large part, for both the marked hypercalcaemia and elevated PTH levels in this patient.     

Discordant disappearance of bioactive and immunoreactive parathyroid hormone after parathyroidectomy

The disappearance of plasma PTH after parathyroidectomy was assessed in patients with primary hyperparathyroidism and normal renal function, chronic renal failure or restored renal function (after transplantation). Plasma PTH levels were determined by renal cytochemical bioassay and by midregion and carboxyl-terminal RIAs. Baseline PTH levels were lower in each patient when assessed by bioassay than when determined by RIA, and the rate of hormone disappearance was faster when determined by bioassay than when measured by RIA. This difference was accentuated in chronic renal failure due to prolongation of the disappearance rates of midregion and carboxyl-terminal immunoreactivity. The half-life of bioassayable hormone in patients with chronic renal failure was prolonged less than 2-fold compared to the half-life in patients with normal or restored renal function. The results emphasize the discordance between levels of bioactive and immunoreactive hormone regardless of renal function, demonstrate that this discordance is augmented after acute reduction in circulating hormone, and show that it is further increased when kidney function is impaired. The studies also implicate extrarenal mechanisms as a major factor in the clearance of bioactive hormone in established renal failure.     

Circulating levels of midregional parathyroid hormone-related protein in hypercalcaemia of malignancy

OBJECTIVE: We have developed and evaluated a sensitive radioimmunoassay directed against the midregional part of parathyroid hormone-related protein (PTHrP), which is involved in the syndrome of humoral hypercalcaemia of malignancy. PATIENTS: Midregional PTHrP levels were studied in 41 consecutive inpatients with malignancy and hypercalcaemia, 32 normocalcaemic patients with malignancy, 21 patients with primary hyperparathyroidism, 34 patients with renal failure, and 87 normals. MEASUREMENTS: The assay used an antiserum against the midregional amino acid residues 53-84 of PTHrP and PTHrP(1-86) as label and standard. Midregional PTHrP was stable in serum and plasma and could be measured directly without sample extraction. RESULTS: Normal plasma concentrations ranged from undetectable (< 5 pmol/l) to 21 pmol/l. In renal failure, PTHrP was positively correlated with serum creatinine, but PTHrP elevations of up to 30 pmol/l were found only in severe renal dysfunction with creatinine > 850 mumol/l. In hypercalcaemia caused by solid tumours, midregional PTHrP was elevated in 81% (22 of 27) of patients, ranging from undetectable to 203 pmol/l (median: 40 pmol/l). In these patients serum calcium correlated positively with PTHrP (P < 0.01). Mean PTHrP levels were indistinguishable in subgroups with and without metastatic skeletal disease. The mechanism of hypercalcaemia in 14 patients with haematological malignancy was apparently different, since all but one had normal or only marginally elevated PTHrP levels. In 21 patients with primary hyperparathyroidism midregional PTHrP was normal in 20. The assay was therefore especially useful in distinguishing the latter condition from humoral hypercalcaemia of malignancy as the second major cause of hypercalcaemia. PTHrP was normal in all 32 patients with normocalcaemic malignancy. CONCLUSION: This radioimmunoassay of midregional PTHrP provides high diagnostic sensitivity in the identification of humoral hypercalcaemia of malignancy caused by solid tumours. The assay should therefore be useful in the differential diagnosis of hypercalcaemia.