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Biomedical subjects

L H Rees

Publications and source records attributed to L H Rees.

At least 91 records · Page 5Linked to original sources
PubMed1987

A phaeochromocytoma presenting with Cushing's syndrome associated with increased concentrations of circulating corticotrophin-releasing factor.

The case is described of a 61-year-old male who presented with hypertension and Cushing's syndrome which resolved on excision of a unilateral adrenal mass. Histology of the tumour revealed a benign phaeochromocytoma which immunostained for corticotrophin-releasing factor (CRF-41) but not for ACTH. Preoperative plasma concentrations of immunoreactive CRF-41 were increased, and gradients for both CRF-41 and ACTH were demonstrated across the tumour. Post-operatively, CRF-41 was undetectable in plasma. The tumour contained high concentrations of immunoreactive CRF-41 which co-eluted with synthetic human CRF-41 on reversed-phase high-performance liquid chromatography. Tumour CRF-41 stimulated the release of ACTH in a dose-dependent manner from isolated rat anterior pituitary cells. We conclude that this tumour secreted CRF-41 and ACTH and had the capacity to produce ACTH-dependent Cushing's syndrome directly by secreting ACTH and indirectly by secreting CRF-41 to stimulate ACTH secretion from the anterior pituitary.

Adrenal Gland Neoplasms↗
PubMed1987

Plasma Met-enkephalin and catecholamine responses to insulin-induced hypoglycaemia in greyhounds.

The involvement of endogenous opioid peptides in the stress response was investigated by measuring plasma concentrations of Met-enkephalin-like immunoreactivity (MLI), adrenaline and noradrenaline during insulin-induced hypoglycaemia in conscious greyhounds. Moreover, the molecular forms of circulating MLI were characterized using gel filtration chromatography. In the first group of animals, i.v. administration of insulin (0.3 units/kg) provoked marked hypoglycaemia (blood glucose concentrations fell from 4.4 +/- 0.1 to 1.5 +/- 0.2 mmol/l; mean +/- S.E.M.) which was associated with significant (P less than 0.001) rises in plasma MLI concentrations from a basal concentration of 45 +/- 8 to a peak of 189 +/- 39 ng/l. A within-subject study comparing five different insulin doses ranging from 0.004 to 0.3 units/kg showed dose-related effects on blood glucose with nadir concentrations of 4.1 +/- 0.6 mmol/l (after the smallest dose of insulin) and 0.8 +/- 0.1 mmol/l (after the largest dose of insulin). This was associated with dose-related rises in plasma MLI with peak concentrations of 56 +/- 17 and 558 +/- 35 ng/l, plasma adrenaline with peak concentrations of 0.45 +/- 0.06 and 15.76 +/- 1.33 nmol/l and plasma noradrenaline with peak concentrations of 0.49 +/- 0.07 and 2.27 +/- 0.45 nmol/l following the smallest and largest doses of insulin respectively. These results are the first demonstration of raised plasma MLI concentrations following hypoglycaemia. Moreover, they show that the hormonal responses vary with the degree of hypoglycaemia achieved. Together with reports by other investigators these findings might suggest opioid modulation of the responses of the sympathoadrenal system to hypoglycaemia.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

The use of reversed-phase high-performance liquid chromatography to detect proteolytic activity from human pancreas in a radioimmunoassay for corticotrophin-releasing factor.

Degradation of tracer during a radioimmunoassay (RIA) can result in false-positive concentrations of immunoreactivity being reported in a biological sample. A technique has been developed using reversed-phase high-performance liquid chromatography (HPLC) to detect proteolytic degradation of corticotrophin-releasing factor-41 (CRF-41) during incubation with tissue extracts under RIA conditions. Human pancreatic tissue was extracted in HCl or urea and incubated with 125I-labelled CRF-41 at neutral pH for 18 h. When samples were analysed by HPLC and fractions counted for radioactivity, tracer was extensively degraded. Heating extracts at 85 degrees C or adding lima bean trypsin inhibitor to the medium prevented degradation. Pancreatic tissue extracted in HCl was analysed by gel filtration and HPLC, and fractions were subjected to RIA for CRF-41. A peak of immunoreactivity was detected by both chromatographic methods. However, when this material was incubated with tracer and analysed by HPLC, the tracer was degraded, indicating that proteolytic activity remained after acid extraction and two forms of chromatography.

Chromatography, High Pressure Liquid↗
PubMed1986

Comparison of reversed-phase and anion-exchange high-performance liquid chromatography for the analysis of human growth hormones.

Separation of human growth hormones (hGH) has been studied by three chromatographic methods: gel filtration chromatography, reversed-phase high-performance liquid chromatography (HPLC) and anion-exchange HPLC. Six growth hormone preparations were used to characterize the systems: two pituitary extracts (an old freeze-dried purified extract A, and a freshly extracted frozen pituitary B); two purified forms (hGH20K and dimer); and two chemically modified monomeric forms (reduced and deamidated). Gel filtration chromatography (pH 8) separated the two pituitary extracts A and B into four components (monomeric, dimeric, aggregate and void material), the relative compositions of which were very similar in both extracts. Reversed-phase HPLC under acid dissociating conditions (pH 2) separated the extracts into four peaks (M1, M2, D and A). The first two components are both monomers: M1 contains all those forms where no major conformational change has occurred; M2 comprises forms with substantial conformational alteration (e.g. disulphide bridge cleavage). Component D includes the interchain disulphide dimer, whilst A is an uncharacterized oligomeric form. Anion-exchange HPLC (pH 8) separated extract A into four regions of immunoreactivity. Region 1 contains hGH20K separated from hGH22K; region 2 includes other monomeric charge variants; region 3 is a broad peak which includes true dimers of hGH20K and hGH22K, as well as loosely aggregated monomer. Region 4 is another broad peak, presumably containing a higher-molecular-weight hGH form or forms. All the hGH forms (identified and unidentified) can be separated by sequential use of two out of three chromatographic methods. No two forms found so far co-eluted in all three systems.

Chromatography, Gel↗
PubMed1986

The interaction of growth hormone releasing hormone with other hypothalamic hormones on the release of anterior pituitary hormones.

To determine whether the 29 amino-acid fragment of growth hormone releasing hormone (GHRH) can be combined with other hypothalamic releasing hormones in a single test of anterior pituitary reserve, the responses of anterior pituitary hormones to combinations of an i.v. bolus of GHRH(1-29)NH2 or saline with an i.v. bolus of either LH releasing hormone (LHRH) plus TRH, ovine CRH(oCRH) or saline were studied. Each infusion of GHRH(1-29)NH2 resulted in a rapid increment of the plasma GH value. Infusion of GHRH(1-29)NH2 also caused a small and transient rise in plasma PRL, but no change in the integrated PRL response. The combination of GHRH(1-29)NH2 with LHRH plus TRH caused a larger increment of peak and integrated plasma TSH levels than LHRH plus TRH alone. GHRH(1-29)NH2 did not affect the release of other anterior pituitary hormones after infusion with oCRH or LHRH plus TRH. Because of the finding of potentiation of the TSH-releasing activity of LHRH plus TRH by GHRH(1-29)NH2, the study was extended to the investigation of TSH release after infusion of TRH in combination with either GHRH(1-29)NH2 or GHRH(1-40). In this study the combination of TRH with both GHRH preparations also caused a larger increment of the peak and integrated plasma TSH levels than TRH alone. It is concluded that GHRH(1-29)NH2 possesses moderate PRL-releasing activity apart from GH-releasing activity. In addition, GHRH potentiates the TSH-releasing activity of TRH.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1986

Normal circulating immunoreactive growth hormone releasing factor (hGRF) concentrations in patients with functional hypothalamic hGRF deficiency.

Using a highly specific radioimmunoassay we have measured the concentrations of human growth hormone releasing factor (ir-hGRF) in the peripheral circulation of six individuals with acquired hypothalamic hGRF deficiency. Despite their hypothalamic dysfunction venous plasma ir-hGRF increased normally in every patient after the stimulus of a mixed breakfast, from an average concentration basally of 13.6 +/- 6.0 pg/ml to a maximum of 29.0 +/- 8.4 pg/ml (mean +/- SEM) at 120 min. The findings indicate that circulating hGRF is at least in large part extrahypothalamic in origin, which in turn implies a physiological role for hGRF in the periphery.

Adolescent↗
PubMed1986

The effect of cholinergic blockade on the ACTH, beta-endorphin and cortisol responses to insulin-induced hypoglycaemia.

To assess the effect of cholinergic blockade on the ACTH, beta-endorphin and cortisol responses to insulin-induced hypoglycaemia, six healthy male volunteers each underwent two insulin tolerance tests in random order, separated by at least 1 week with and without atropine. ACTH levels were significantly greater at +45 min (mean +/- SEM, 223 +/- 21 pg/ml vs 148 +/- 15 pg/ml, P less than 0.01) and at +120 min (54 +/- 11 pg/ml vs 29 +/- 10 pg/ml, P less than 0.05). beta-endorphin levels were significantly greater at +30 min (170 +/- 45 pg/ml vs 96 +/- 32 pg/ml, P less than 0.05) and at +105 min (81 +/- 14 pg/ml vs 54 +/- 7 pg/ml, P less than 0.01). Cholinergic blockade had no effect on plasma glucose or cortisol concentrations. This study demonstrates that cholinergic blockade with atropine facilitates the ACTH and beta-endorphin responses to insulin-induced hypoglycaemia without altering the cortisol responses.

Adrenocorticotropic Hormone↗
PubMed1986

Diagnosis and management of ACTH-dependent Cushing's syndrome: comparison of the features in ectopic and pituitary ACTH production.

The clinical features, diagnosis and management of 16 consecutive patients with ectopic ACTH production are described and biochemical data are compared with those of 48 consecutive patients with pituitary-dependent Cushing's disease. In 10 cases the ectopic ACTH secreting tumour was completely occult to routine clinical and radiological investigation, and no basal or dynamic investigation of adrenal-pituitary function was able clearly to differentiate these patients from those with Cushing's disease. High dose dexamethasone suppression testing assessed by plasma cortisol was usually helpful but unexpected responses were seen in both diagnostic groups; the metyrapone test yielded no useful information and should now be abandoned. Hypokalaemia was seen in all patients with ectopic ACTH production but in only 10% of those with Cushing's disease who were not on diuretics at presentation. Successful diagnosis and tumour localization was most frequently achieved by a combination of CT scanning of the chest and abdomen and venous catheter sampling for ACTH. All patients in whom the ectopic ACTH-secreting tumour was obvious at presentation died of their primary tumour within 8 months, whereas seven of the 10 patients with occult tumours at presentation are alive 1.5-16.5 years later, and appear cured. Occult ectopic ACTH secretion may be impossible to distinguish from pituitary Cushing's disease. Multiple and repeated investigations are often required to make this differential diagnosis, essential for appropriate therapy.

ACTH Syndrome, Ectopic↗
PubMed1986

Immunocytochemical study of 18 tumours causing ectopic Cushing's syndrome.

Eighteen cases of Cushing's syndrome caused by ectopic production of peptide hormones were investigated by histological and immunocytochemical methods and the findings correlated with clinical and biochemical observations. Immunocytochemistry showed immunoreactive adrenocorticotrophic hormone (ACTH) or peptides derived from the ACTH precursor (pro-opiomelanocortin (POMC], or both, in a total of 10 cases: five of these also contained immunoreactive-alpha-melanocyte stimulating hormone, indicating more extensive translational processing of POMC than normally occurs in healthy corticotrophs of the anterior pituitary; in two further cases peptides capable of stimulating ACTH release from the anterior pituitary were present. In the remaining six cases immunocytochemistry failed to show the presence of ACTH, other POMC derived peptides, or peptides with ACTH releasing properties. These findings correlate well with the histological and clinical observations, in that the six tumours had been clinically overt, caused rapid death, and histologically seemed to be highly malignant. In contrast, the 12 other tumours were occult to radiological examination, patients had a much improved survival rate, and histologically the tumours seemed to be less aggressive. All but one of the tumours in this series showed a degree of neuroendocrine differentiation, indicated by the presence of neuron specific enolase. These results suggest that one feature of highly malignant tumours, which cause an ectopic endocrine syndrome, is a high secretion of peptide hormones, leaving amounts that are too small to be shown by immunocytochemistry.

ACTH Syndrome, Ectopic↗
PubMed1986

Characterization of pro-opiomelanocortin-derived peptides in pituitary and ectopic adrenocorticotrophin-secreting tumours.

The molecular forms of pro-opiomelanocortin in plasma of normal subjects and plasma and tissue extracts from patients with disorders of the hypothalamic-pituitary-adrenal axis have been characterized using gel filtration chromatography under acid dissociating conditions, and the molecular forms further investigated by high pressure liquid chromatography and affinity chromatography. A large molecular weight MSH/ACTH fragment was observed in all plasma samples chromatographed, although the proportions of this fragment were significantly greater in patients with the ectopic ACTH syndrome. A similar profile was observed in tumour extracts: a greater proportion of large molecular weight precursors was observed. Immunoreactive-gamma-MSH in extracts of ectopic tumours displayed marked and variable heterogeneity, and affinity chromatography with Concanavalin A-Sepharose indicated that this was due to differential glycosylation. High pressure liquid chromatography of a peak of amino terminal pro-opiomelanocortin (N-POC(1-76)) and 22 000 mol. wt MSH/ACTH indicated two peaks in each peptide, again possibly due to differential glycosylation. A possible neurointermediate lobe origin of atypical invasive pituitary tumours is discussed.

ACTH Syndrome, Ectopic↗
PubMed1986

Corticotrophin-releasing factor-like immunoreactivity and bioactivity of human fetal and adult hypothalami.

Corticotrophin releasing factor-like immunoreactivity (CRF-LI) and bioactivity, and arginine vasopressin-like immunoreactivity (AVP-LI) have been measured in extracts of human fetal and adult hypothalamic tissue and their development with the gestational age of the fetuses (12-27 weeks) studied. CRF-LI was measured by a radioimmunoassay developed for ovine corticotrophin-releasing factor (oCRF-41). Corticotrophin-releasing factor bioactivity was measured in a rat isolated anterior pituitary cell perfusion system. CRF-LI and bioactivity and AVP-LI were all detectable in fetal hypothalamic extracts from 12 to 13 weeks of gestational age. CRF-LI was also present in human fetal pituitary glands from 12 weeks of gestational age. The concentration of CRF-LI in the fetal hypothalamic extracts (9.2 +/- 11.4 ng/g, mean +/- S.E.M., n = 33) showed no significant correlation with the gestational age of the fetuses. However the concentration of AVP-LI (25.0-36.8 ng/g, n = 17) did show a positive correlation (r = 0.508, P less than 0.05) with gestational age, as did the concentration of CRF bioactivity (471.3-556.3 ng ACTH released/g tissue, n = 13, r = 0.725, P less than 0.01). The CRF bioactivity of all fetal hypothalamic extracts was potentiated by the addition of synthetic human (h)AVP, but the bioactivity of the adult hypothalamic extracts was not, presumably because of the higher levels of AVP-LI already present in the adult extracts. Pretreatment of tissue extracts with antisera to oCRF-41 and/or hAVP reduced the CRF bioactivity of all hypothalamic extracts. Sephadex chromatography of fractions which co-eluted with synthetic oCRF-41 or hAVP contained CRF bioactivity and this bioactivity was potentiated when synthetic hAVP or oCRF-41, respectively, were added to the fractions. However, a larger molecular weight form of CRF-LI (8000-10 000 daltons), which was observed only in fetuses of 20 weeks of gestational age or less, did not contain any significant CRF bioactivity.

Arginine Vasopressin↗
PubMed1986

Biochemical characterization of circulating Met-enkephalins in canine endotoxin shock.

The alterations in arterial, venous and adrenal vein levels of immunoreactive Met-enkephalins following endotoxin administration have been investigated in dogs by direct measurement and gel filtration chromatography. Animals were anaesthetized with alpha-chloralose and allowed to breath spontaneously. The left lumbar adrenal vein, limb vein and femoral artery were cannulated for blood sampling. Severe shock was produced by the administration of a large bolus of E. coli endotoxin followed by a continuous infusion. The production of endotoxin shock was associated with significant increases in adrenal vein and systemic venous plasma immunoreactive Met-enkephalin levels. Forty-five minutes after induction of endotoxin shock arterial immunoreactive Met-enkephalin levels were generally higher than baseline values. In resting anaesthetized animals a large 31,000 molecular weight form of Met-enkephalin, presumably proenkephalin, was found in plasma obtained from the adrenal vein, systemic and pulmonary circulations. Following endotoxin this enkephalin-containing peptide still predominated in arterial and venous plasma, whereas in the adrenal vein the proportion of Met-enkephalin immunoreactivity attributable to this large peptide fell. This was associated with the appearance of increasing amounts of smaller molecular forms (18,000, 8000, 3-5000 molecular weight and the pentapeptide itself). In this model enkephalin-containing peptides were not biochemically modified by their passage through the lungs.

Adrenal Glands↗
PubMed1986

The distribution of immunoreactive alpha-melanocyte-stimulating hormone cells in the adult human pituitary gland.

It has been suggested that a proportion of the adenomas and the nodular hyperplasia of cells in the pituitary gland in cases of Cushing's disease are derived from cells of the pars intermedia rather than the pars anterior. The evidence can be summarized as follows: the posterior site of adenoma or nodular hyperplasia in the pituitary, the innervation of cells and the suppressive response to the dopamine agonist bromocriptine in vivo or to dopamine in vitro. All these observations infer analogy with cells of the pars intermedia of other species, which are controlled by direct neural tonic dopaminergic inhibition. The adult human pituitary gland, however, does not possess a morphologically distinct pars intermedia, due to regression of the rudimentary fetal pars intermedia after birth, with mixing of cells into the pars anterior and pars nervosa. Since cells of the pars intermedia characteristically synthesize alpha-MSH, we have studied this peptide in order to assess the occurrence and distribution of intermedia-derived cells in the adult human pituitary. Sections from 100 pituitaries, removed at autopsy, were stained by an indirect immunoperoxidase technique using non-cross-reacting antisera specific for alpha-MSH and ACTH. Immunoreactive alpha-MSH (IR-alpha-MSH) cells were found in a total of 97 specimens. Of these, only ten cases showed a marked concentration of IR-alpha-MSH cells in the zona intermedia. In the majority of pituitaries, IR-alpha-MSH cells were more commonly seen in the pars anterior than in the zona intermedia; in 41 cases, IR-alpha-MSH cells were completely absent from the zona intermedia.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenocorticotropic Hormone↗
PubMed1986

Characterization by high-performance liquid chromatography of circulating growth hormone-releasing factors in normal subjects.

Four forms of circulating immunoreactive human GH-releasing factor (ir-hGRF) have been identified in each of four normal subjects, with a mean increase in total ir-hGRF of twofold over basal levels following a mixed meal. Plasma samples (200 ml) from each individual were subjected to large-scale Vycor extraction with initial purification by high-performance liquid chromatography on a reversed-phase C18 column, followed by analytical separation of the ir-hGRF components using a C3 wide-pore reversed-phase column, and subsequent radioimmunoassay of the fractions. The mean recovery of total ir-hGRF from the plasma (fasted and non-fasted) was 76 +/- 16% (2 X S.E.M.). Analytical separation of the ir-hGRF revealed four components which co-eluted with synthetic hGRF-37, hGRF-40 and hGRF-44, and a peak eluting between hGRF-40 and -44 which may represent hGRF-42. The hGRF-40 was shown to be the predominant circulating molecular form in the fasted state in each subject, and in three out of four subjects following a mixed meal. The hGRF-44 showed the greatest percentage increase over basal in all four individuals.

Chromatography, High Pressure Liquid↗