Hypercalcaemia secondary to secretion of parathyroid hormone related protein from a somatostatinoma of the pancreas.
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Adult↗
Biomedical subjects
W A Ratcliffe
Publications and source records attributed to W A Ratcliffe.
Production of parathyroid hormone-related protein by the mammary gland of the goat.
Parathyroid hormone-related protein (PTHRP) has been quantified by sensitive specific immunoassays in mammary venous blood and milk from 7 days before to 7 days after parturition in the goat. A significant venous-arterial concentration gradient in plasma PTHRP 1-86 concentrations was demonstrated across the mammary gland, indicating that PTHRP enters the maternal circulation and may have a role in calcium homoeostasis during lactation. Significant and sustained increases in mammary venous and milk PTHRP 1-86 concentrations were found from 1 day before parturition to 7 days afterwards, with peak concentrations of 1.57 +/- 0.58 pmol/l (plasma) and 8.69 +/- 2.95 nmol/l (milk) (mean +/- S.E.M.) occurring on day -1 and the day of parturition respectively. Estimates of the mammary output of PTHRP into plasma in four goats averaged 9% (range 1-25%) of that secreted into milk. Suppression of maternal prolactin concentrations by bromocriptine significantly reduced milk yield and the mammary venous PTHRP concentration, without affecting the concentration of PTHRP in milk. In conclusion, parturition in the goat is associated with a sustained increase in secretion of PTHRP into both plasma and milk; the former may be involved in maternal calcium homoeostasis, whereas the latter may have a role in the neonate.
Animals↗
Parathyroid hormone related protein and hypercalcaemia in breast cancer.
OBJECTIVE: To see whether parathyroid hormone related protein has a humoral role in breast cancer. DESIGN: Plasma concentrations and tumour expression of parathyroid hormone related protein were determined (by two site immunoradiometric assay and immunohistochemistry respectively) in women with breast cancer and related to the presence of bone metastases and serum calcium concentrations. SUBJECTS: Plasma concentrations of parathyroid hormone related protein were measured in 57 women with early breast cancer without apparent bone metastases, 28 women with bone metastases, and 13 women with bone metastases and hypercalcaemia. Tissue positivity for parathyroid hormone related protein was determined retrospectively in 106 primary breast tumours from women without apparent bone metastases and 72 tumours from women with bone metastases, 25 of whom subsequently developed hypercalcaemia. RESULTS: Plasma parathyroid hormone related protein concentrations were detectable (greater than 0.23 pmol/l) in 12 (92%) of the 13 hypercalcaemic patients with bone metastases compared with 10 (36%) of the 28 normocalcaemic patients with bone metastases and five (9%) of the 57 normocalcaemic patients without bone metastases. Parathyroid hormone related protein concentrations were significantly higher in hypercalcaemic than normocalcaemic patients with bone metastases. Tumour staining was positive for parathyroid hormone related protein in 22 (88%) of the 25 primary breast cancers from patients with bone metastases. Tumour staining was positive for parathyroid hormone related protein in 22 (88%) of the 25 primary breast cancers from patients with bone metastases who later developed hypercalcaemia compared with 25 (53%) of the 47 from women in this group who remained normocalcaemic and 55 (52%) of the 106 early breast cancers from women without known metastases. CONCLUSION: Tumour derived parathyroid hormone related protein may have an important humoral role in hypercalcaemia associated with metastatic breast cancer.
Bone Neoplasms↗
Production of parathyroid-hormone-related protein by cholesteatoma cells in culture.
Specimens of cholesteatoma were removed at surgery from five patients who had evidence of bone resorption. Parathyroid-hormone-related protein (PTHrP) was detected, by radioimmunoassay, in conditioned media from keratinocyte cultures derived from all five samples. Concentrations of PTHrP in conditioned media from secondary cultures were higher for the cholesteatoma cells than for normal keratinocytes from controls matched for age and sex. Thus, production of PTHrP by cholesteatoma may be a contributory factor in the bone destruction commonly associated with this disorder.
Adolescent↗
Expression of parathyroid hormone-related protein in abnormal human parathyroids.
The expression of parathyroid hormone-related protein (PTHrP) in abnormal human parathyroids was investigated. Northern blot analysis of RNA extracted from human benign parathyroid adenomata (n = 4) revealed multiple PTHrP mRNA species ranging in size from 1.8 to 4 kb. The relative abundance of PTHrP mRNA expressed in two of the adenomata was similar to that of a tumour (DAF) associated with humoral hypercalcaemia of malignancy, whereas PTHrP mRNA was of low abundance in a third and was undetectable in the fourth. PTHrP-like immunoreactivity was detected in extracts of abnormal parathyroid tissue (benign adenoma (n = 7), hyperplasia (n = 5) and parathyroid carcinoma (n = 2] using a sensitive specific two-site immunoradiometric assay for human (h) PTHrP(1-86) and a radioimmunoassay for hPTHrP(1-34). Ratios of hPTHrP(1-86)- and hPTHrP(1-34)-like immunoreactivities relative to hPTH(1-84)-like immunoreactivity in the parathyroid tissue extracts were, on average, less than 1%. PTHrP bioactivity in the extracts could not be distinguished from that of PTH, by an osteosarcoma cell bioassay. We conclude that, despite reports of over-expression of PTHrP mRNA in parathyroid adenomata, the potential contribution of PTHrP to the total PTH-like activity of adenomata and other abnormal parathyroid tissue may be insignificant relative to PTH.
Adenoma↗
Immunoreactivity of plasma parathyrin-related peptide: three region-specific radioimmunoassays and a two-site immunoradiometric assay compared.
We measured parathyrin (parathyroid hormone)-related peptide (PTHRP) in plasma by three region-specific RIAs and compared them with an established two-site immunoradiometric assay (IRMA) of PTHRP1-86 in samples from control subjects and from patients with primary hyperparathyroidism (PH) and humoral hypercalcemia of malignancy (HHM). The two direct RIAs of PTHRP1-34 and PTHRP37-67 were specific for regions 9-18 and 52-61, respectively. In the extraction RIA of PTHRP1-34 we used an affinity gel containing a monoclonal antibody specific for the 17-27 sequence; cross-reacting PTHRP species eluted from the gel were assayed by the RIA of PTHRP1-34. PTHRP1-86 plasma concentrations by IRMA were less than 0.23 pmol/L in control subjects and patients with PH, and were significantly increased in patients with HHM (mean 6.1 pmol/L, P less than 0.001). In contrast, plasma PTHRP1-34 concentrations were not significantly different in the three groups; in HHM patients, the mean was 190 pmol/L. Plasma PTHRP37-67 concentrations were similar in control and PH groups and, although significantly increased in HHM patients (mean 440 pmol/L, P less than 0.002), showed poor diagnostic discrimination. PTHRP1-34 concentrations after affinity extraction of plasma were also significantly higher in HHM patients (mean 10.7 pmol/L, P less than 0.001), but showed incomplete diagnostic discrimination. We conclude that the diagnostic utility of the direct RIAs for quantifying PTHRP is markedly inferior to the IRMA of PTHRP1-86.
Humans↗
Development and validation of an immunoradiometric assay of parathyrin-related protein in unextracted plasma.
This two-site immunoradiometric assay for human parathyrin-related protein 1-86 (PTHRP1-86) in plasma uses a mouse monoclonal antibody to PTHRP1-34 coupled to cellulose particles for immunoextraction of N-terminal immunoreactivity, and a rabbit antiserum to PTHRP37-67 that is indirectly labeled with 125I-labeled PTHRP37-67 for quantifying the bound analyte. The detection limit of the assay is 0.23 pmol/L, corresponding to 0.4 pg (0.04 fmol) per tube, for a sample volume of 200 microL. Recovery of PTHRP1-86 added to serum is essentially quantitative, and within- and between-batch precision is 4.4% and 11.1%, respectively. PTH1-84, PTHRP18-34, PTHRP9-34, PTHRP1-34, and PTHRP37-67 do not cross-react in the assay at concentrations up to 2 nmol/L. Plasma concentrations of PTHRP1-86 were below or close to the detection limit of the assay in normal subjects and in patients with primary hyperparathyroidism, hypoparathyroidism, chronic renal failure, and normocalcemic malignancy. In 37 hypercalcemic patients with various malignancies, we found detectable PTHRP1-86 concentrations in 35 (95%, mean 7.4 pmol/L, range 0.46-24.7). The data support the proposed humoral role of PTHRP in cancer-associated hypercalcemia and suggest that the assay has clinical utility in the differential diagnosis of hypercalcemia.
Antibodies, Monoclonal↗
Treatment of a malignant pancreatic endocrine tumour secreting parathyroid hormone related protein.
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Adenoma, Islet Cell↗
Production and characterisation of monoclonal antibodies to parathyroid hormone-related protein.
The production and characterisation of 17 monoclonal antibodies to human parathyroid hormone-related protein (PTH-rP) 1-34 is described. Five of the antibodies were shown to be of high avidity (Ka 4 X 10(10)-1.9 X 10(11) L/M) and able to detect 15-100 pg PTH-rP 1-34 per tube by RIA. None cross-reacted with PTH 1-34, and inhibition studies with peptide subfragments of PTH-rP 1-34 indicated that all recognise a central region extending from residues 9-18 to between residues 23 and 34. All antibodies tested cross-reacted with native PTH-rP in culture fluids from keratinocytes and squamous cancer cell lines and in human and bovine milk. The concentrations of PTH-rP 1-34 (ng/ml) in these fluids as determined by RIA were: keratinocytes 1-3, squamous cancer 0.2-2.5, human milk, up to 80. Selected antibodies coupled to Sepharose 4B were used to extract PTH-rP from biological fluids with high yields.
Animals↗
Characterization of the molecular forms of parathyroid hormone-related protein in milk.
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Animals↗
Patterns of ACTH and cortisol pulsatility over twenty-four hours in normal males and females.
Patterns of plasma ACTH and cortisol concentrations were studied in 10 healthy subjects (five male, five female in the early follicular phase, overall age range 21-32 years) by sampling through an indwelling cannula every 15 min for 24 h. The subjects were in hospital, ambulant, and taking normal meals. Plasma ACTH was measured by a two-site immunoradiometric assay with a detection limit of 3.9 ng/l (0.9 pmol/l). Pulses were identified by the method of Clayton et al. (1987) using stringent criteria to minimize false positive peaks. All subjects showed a circadian rhythm of ACTH, the acrophase occurring between 0615 and 0920 h in all but one subject and the mesor value was between 9.2 and 18.6 ng/l (2.0 and 4.1 pmol/l). There were significantly fewer pulses between 1800 and 2400 h compared with the other three 6-h periods. The pattern of ACTH differed between males and females in several respects: more pulses (18 vs 10), greater mean peak amplitude (16.8 vs 10.3 ng/l), greater area under the 24-h profile (350.9 vs 206.6 ng/l h) and higher mean level (14.7 vs 8.6 ng/l) in the males. In contrast, the cortisol pattern did not show statistically different sex differences. The sex differences suggest greater sensitivity to, or availability of, ACTH to the female adrenal cortex, or different set points in cortisol feedback.
Adrenocorticotropic Hormone↗
Non-isotopic, two-step free thyroxine immunoassay: a better measure of free thyroxine than analogue radioimmunoassay.
Plasma or serum free thyroxine (T4) was measured by a novel non-isotopic, two-step immunoassay in 373 consecutive patients attending a thyroid clinic, in whom thyroid status was categorized according to clinical findings, supported by routine thyroid function tests. The 95% confidence limit of free T4 in the euthyroid patients (n = 112) was 7-20 pmol/L. Free T4 concentrations within the reference range were found in six of 40 patients with primary hypothyroidism and nine of 182 patients with overt thyrotoxicosis, six of whom had T3 toxicosis. Serum or plasma free T4 measured by the two-step method showed improved diagnostic specificity over an analogue RIA in selected groups of euthyroid patients in whom abnormal binding of analogue T4 can affect the validity of the result. Free T4 results found by analogue RIA and the two-step method in 58 patients who were receiving thyroxine replacement therapy were similar. The between-assay precision of the two-step method was poor ranging from a coefficient of variation of 9.7% to 19.3% over a free T4 concentration range of 5.0 to 46.0 pmol/L. We conclude that the two-step methodology offers diagnostic advantages for a laboratory which receives specimens from such patients for exclusion of thyroid disease but that improved assay precision is required before it could be used in a routine situation.
Autoantibodies↗
Identification and partial characterization of parathyroid hormone-related protein in human and bovine milk.
Parathyroid hormone-related protein (PTHrP) was measured in human and bovine milk by radioimmunoassay (RIA) and bioassay, and the molecular forms characterized by gel chromatography and immunoblotting of affinity-purified PTHrP. Mean immunoreactive PTHrP(1-34) concentrations were 23 and 87 micrograms/l in human and bovine milk respectively. Bioactive (BIO) PTHrP concentrations determined by cyclic AMP production by ROS 17/2.8 cells correlated significantly (P less than 0.001) with those obtained by RIA (BIO = 1.04RIA--3.4, r = 0.939). Gel filtration of human and bovine milk identified several peaks with immunoactivity and bioactivity. Immunoblotting of affinity-purified PTHrP revealed multiple molecular species including components with mobilities similar to those of PTHrP and its subfragments. These studies confirm the presence of immuno- and bioactive PTHrP in milk and suggest that post-translational processing is complex and variable.
Animals↗
Development and validation of a two-site immunoradiometric assay for human atrial natriuretic factor in unextracted plasma.
This two-site immunoradiometric assay (IRMA) for human atrial natriuretic factor (ANF)99-126 in plasma utilizes a mouse monoclonal antibody raised against rat (r) ANF103-125 with specificity directed towards the ring structure of ANF, and a rabbit antiserum to human (h) ANF99-126. The monoclonal antibody is radioiodinated, and the IgG fraction of the antiserum is coated onto wells of a microtiter plate. Plasma or standard hANF99-126 (150 microL) is incubated with the radioligand in coated wells for 24 h. The detection limit is 0.9 pg per well, corresponding to 2 pmol/L, with a working range (CV less than 10%) from 4.5 to 540 pmol/L. Intra- and interassay precision are 7% and 9%, respectively, and the assay is unaffected by plasma matrix. In humans, the IRMA is specific for hANF99-126, the major circulating form of ANF, and does not cross-react with metabolites having deletions at the carboxy terminus. Plasma IRMA values in normal seated subjects were 6.6 +/- 1.5 (SEM) pmol/L and results correlated with those of an extraction RIA (r = 0.81, P less than 0.001).
Antibodies, Monoclonal↗
Performance and diagnostic application of a two-site immunoradiometric assay for parathyrin in serum.
The "N-tact" immunoradiometric assay (IRMA) from INCSTAR for parathyrin (PTH) in serum involves a 125I-labeled affinity-purified antiserum to PTH 1-34 and an affinity-purified antiserum to PTH 39-84, the latter bound to a polystyrene bead. The mean detection limit, determined in six consecutive assays, was 4 ng/L. The within-batch CV was less than 7% in the range 15 to 2135 ng/L. The between-batch CV was 11.7% and 5.3% at 30 and 371 ng/L, respectively. Serum PTH in 14 proven cases of primary hyperparathyroidism was 49-808 (median 111) ng/L, undetectable (less than 5 ng/L) in 10 cases of primary hypoparathyroidism and in 10 cases of hypercalcemia associated with malignancy, compared with 7-39 ng/L in 45 normal subjects. PTH was 9 to 19 ng/L in four patients with familial benign hypercalcemia. In 39 patients with renal failure, apparent concentrations were 14 to 857 (median 133) ng/L, but sera from these patients pre-diluted with zero standard did not parallel dilutions of the standard, PTH 1-84. PTH concentrations were not significantly decreased in blood or serum kept at 20 degrees C for up to 6 h. After successful removal of a parathyroid adenoma, the mean half-time for disappearance of PTH in vivo in five hyperparathyroid patients was 3.3 min.
Adult↗
Adrenocorticotrophin (ACTH) deficiency undetected by standard dynamic tests of the hypothalamic-pituitary-adrenal axis.
Six patients (four females, two males; aged 18-65 years), previously treated by external pituitary irradiation (2000-4000 cGY in 8-15 fractions over 10-20 days) for pituitary tumours, presented with the symptoms of excessive and inappropriate tiredness suggestive of ACTH deficiency, despite a normal peak cortisol response to an insulin tolerance test (four cases) or to a glucagon stimulation test (two cases). These six patients were found to have significantly lower mean 24 h urinary free cortisol levels (100 +/- 40 nmol; mean +/- SD) compared with the mean value of 31 normal controls (210 +/- 70.8 nmol; P less than 0.01). In addition serum cortisol profiles based on a series of four timed samples between 0900-2300 h were subnormal (mean 130 nmol/l) in comparison with profiles obtained from 12 normal controls (mean 270 nmol/l) (P less than 0.001). Glucocorticoid replacement therapy promptly abolished their symptoms. These results suggest that a discordance between ACTH secretion under basal circumstances and ACTH response to pharmacological tests may exist in patients with ACTH deficiency. We speculate that defective endogenous corticotrophin-releasing hormone (CRF) secretion, due to radiation-induced damage at hypothalamic level, is one cause of this phenomenon.
Adolescent↗
Oestradiol assays: applications and guidelines for the provision of a clinical biochemistry service.
This paper reports on the provision of a clinical biochemistry service for serum oestradiol. The pathophysiology and recognised applications of oestradiol assays are discussed and the current availability of assay reagents and methodologies reviewed. Data are presented on the analytical performance of assays for serum oestradiol in the UK External Quality Assessment Scheme (UKEQAS) and general guidance is offered to laboratories providing a diagnostic service for this analyte.
Adult↗
Trial of 17-hydroxyprogesterone caproate (Proluton Depot) in women with long-standing infertility; failure of estrogen positive feedback the following cycle.
In open and double-blind studies 40 women with long-standing unexplained infertility were investigated and treated with 17 alpha-hydroxyprogesterone caproate (17HPC, Proluton Depot). In the open study, 16 women with a high index of suspicious abortions were given 500 mg 17HPC imtramuscularly (i.m.) weekly for 6-16 weeks. Six of the women received the drug from a time preceding the expected date of a period; 2 of these conceived that cycle and their pregnancies continued to term, while 4 were not pregnant. Ten women (9 with definite previous abortions) were given 17HPC when they suspected (correctly) that they were pregnant. Their pregnancies continued to term in all but 1, who had a premature delivery (still-birth) at 34 weeks. In the double-blind study 24 women were given injections of 17HPC or placebo i.m. at weekly intervals, from about 4 days before the expected period (day -4), provided that the level of progesterone (Prog) (on days -9 to -7) was greater than 10 nmol/l. After placebo no delay in menstruation or disruption of the succeeding cycle was observed. In 14 of 16 cycles in 14 women given 500 mg 17HPC the withdrawal period was delayed by a few days, and then followed by highly erratic bleeding over the next 1-3 months. The dose was therefore reduced to 250 mg 17HPC but the same problem resulted in 8 of 29 cycles in 16 women (including the above studied in later cycles). Regular cycles were eventually restored in all cases but in 3 this necessitated treatment with the contraceptive pill (Microgynon). In most of the post-treatment cycles there was a progressive and prolonged estradiol (E2) rise, which was not preceded by changes either in serum FSH concentration or in the LH/FSH ratio nor associated with the expected positive feedback rise in LH. We conclude that 17HPC disrupts the following cycle, probably by allowing follicular development while interfering with positive LH feedback. None of the patients of the double-blind study had conceived (as evidenced by undetectable hCG levels). Our study confirmed that this progestogen exerts no direct luteolytic effect. However, in order to establish the efficacy or otherwise of 17HPC given before the end of the cycle, women should be selected with a very high index of suspicion of recurrent early implantation failure or abortion.
17 alpha-Hydroxyprogesterone Caproate↗