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Biomedical subjects

J Frost

Publications and source records attributed to J Frost.

At least 55 records · Page 3Linked to original sources
PubMed1985

A simple in vitro approach to the estimation of the biopotency of drugs affecting adrenal steroidogenesis.

Dispersed guinea-pig adrenal cells can be maximally stimulated to secrete cortisol by adrenocorticotrophin (ACTH greater than 50 ng/l). Further, this stimulation appears to be specific to ACTH alone, with other naturally occurring chemicals (e.g. steroids, protein hormones) at supra-physiological concentrations being without effect on cortisol production. The effect of drugs of differing structure and therapeutic function (aminogluthethimide, metyrapone, trilostane, 17-ketotrilostane, danazol, epostane, megestrol acetate, stanozolol and etomidate) on ACTH-stimulated (50 ng/l) cortisol production has been tested in this system. All the drugs depressed steroid output in a similar dose-related fashion. The concentration of drug which inhibited cortisol output by 50% was (mumol/l, mean +/- SEM): etomidate 0.097 +/- 0.002: epostane 0.44 +/- 0.02: 17-ketotrilostane 0.55 +/- 0.04: trilostane 1.3 +/- 0.1: metyrapone 3.5 +/- 0.6: megestrol acetate, 11 +/- 2: danazol 22 +/- 2: aminogluthethimide 41 +/- 5: stanozolol 50 +/- 4. Thus, etomidate, an anaesthetic, is more potent than the established anti-steroidogenic drugs metyrapone, aminogluthethimide and trilostane. Further, direct anti-steroidogenic effects have been demonstrated for megestrol acetate and stanozolol for the first time. We conclude that this technique offers a promising new approach to the assessment of biological potency of drugs affecting endocrine tissues.

Adrenal Glands↗
PubMed1985

On the stability in vitro of bioactive human adrenocorticotrophin in blood and plasma.

The disappearance in vitro of ACTH from whole human blood and plasma held at 22 degrees C has been monitored using a dispersed adrenal cell bioassay and an unextracted radioimmunoassay. In three normal subjects after a metyrapone test and two patients with Addison's disease, endogenous bioactive ACTH levels were unchanged for at least 1 h in blood and 2 h in plasma. Moreover greater than 50% of the bioactive plasma ACTH was still present in the plasma samples from the patients with Addison's disease after 24 h incubation at ambient temperatures. Human pituitary ACTH (1-39), spiked into plasma from dexamethasone suppressed subjects to give a concentration of 250 ng/l, was stable by bioassay for at least 2 h. No loss of biological activity was observed on subjecting plasma from a patient with Addison's disease or spiked plasma to two cycles of thawing at 37 degrees C and freezing at -70 degrees C or thawing at 20 degrees C and freezing at -20 degrees C. Some loss of bioactivity (20%) occurred on subjecting the patient's, but not ACTH-spiked plasma to four cycles of thawing at 20 degrees C/freezing at -20 degrees C. We conclude that bioactive ACTH (endogenous or exogenous) may be more stable in vitro in human blood and plasma than has been previously thought. If our studies can be confirmed in a larger series then it may be that conditions for handling blood specimens for ACTH assays could be reappraised.

Addison Disease↗
PubMed1985

Guinea pig striatum as a model of human dopamine deamination: the role of monoamine oxidase isozyme ratio, localization, and affinity for substrate in synaptic dopamine metabolism.

The kinetic properties of type A and type B monoamine oxidase (MAO) were examined in guinea pig striatum, rat striatum, and autopsied human caudate nucleus using 3,4-dihydroxyphenylethylamine (dopamine, DA) as the substrate. MAO isozyme ratio in guinea pig striatum (28% type A/72% type B) was similar to that in human caudate nucleus (25% type A/75% type B) but different from that in rat striatum (76% type A/24% type B). Additional similarities between guinea pig striatum and human caudate nucleus were demonstrated for the affinity constants (Km) of each MAO) isozyme toward DA. Endogenous concentrations of DA, 3-methoxytyramine, 3,4-dihydroxyphenylacetic acid, and homovanillic acid were also measured in guinea pig and rat striatum following selective type A (clorgyline-treated) and type B (deprenyl-treated) MAO inhibition. In guinea pig, DA metabolism was equally but only partially affected by clorgyline or deprenyl alone. Combined treatment with clorgyline and deprenyl was required for maximal alterations in DA metabolism. By contrast, DA metabolism in rat striatum was extensively altered by clorgyline but unaffected by deprenyl alone. Finally, the deamination of DA in synaptosomes from guinea pig striatum was examined following selective MAO isozyme inhibition. Neither clorgyline nor deprenyl alone reduced synaptosomal DA deamination. However, clorgyline and deprenyl together reduced DA deamination by 94%. These results suggest that the isozyme localization and/or isozyme affinity for DA, rather than the absolute isozyme content, determines the relative importance of type A and type B MAO in synaptic DA deamination. Moreover, based on the enzyme kinetic properties of each MAO isozyme, guinea pig striatum may serve as a suitable model of human DA deamination.

Animals↗
PubMed1985

On the biopotency and site of action of drugs affecting endocrine tissues with special reference to the anti-steroidogenic effect of anaesthetic agents.

Dispersed guinea-pig adrenal cells or mouse Leydig cells were stimulated with a saturating dose of adrenocorticotrophin (ACTH, 50 ng/1) or luteinizing hormone (LH, 5IU/1), respectively. The incubations were performed in the presence of increasing concentrations (10(-9) - 5 X 10(-4)mol/l) of the anaesthetic agents propofol, thiopentone and etomidate. At the end of this stimulation period, cortisol (from the adrenal preparation) or testosterone (from the Leydig cell culture) were assayed by radioimmunoassay. Propofol, thiopentone and etomidate all inhibited ACTH-stimulated cortisol secretion in a dose-related fashion. Similar inhibition of LH-stimulated testosterone output was found with propofol and thiopentone whereas etomidate was without effect at any concentration employed, an observation in accordance with its known site of action, 11 beta-hydroxylase, an enzyme which is not involved in the biosynthesis of testosterone. The concentration (mumol/l) of anaesthetics which gave 50% inhibition (ED50) of ACTH-stimulated cortisol secretion was 0.1 +/- 0.002 (n = 7), 160 +/- 18 (n = 3) and 170 +/- 18 (n = 3) (mean +/- s.e.m.) for etomidate, thiopentone and propofol, respectively. The corresponding values for the LH stimulated testosterone output from the Leydig cell preparations were 186 (thiopentone) and 180 (propofol) mumol/l. In a separate series of experiments adrenal cells were stimulated with (a) the cortisol precursor steroids (all at 10(-5)mol/l) pregnenolone, 17-hydroxypregnenolone, progesterone, 17-hydroxyprogesterone and 11-deoxycortisol, (b) dibutyryl cAMP (10(-3)mol/l) or (c) ACTH (100 ng/l) in the presence and absence of either etomidate (5 X 10(-5)mol/l), propofol (2.5 X 10(-4)mol/l) or thiopentone (5 X 10(-4)mol/l). All the stimulators increased cortisol production by > 7-fold over that seen in their absence. Propofol depressed ACTH and dibutyryl cAMP induced cortisol output by > 60% (P < 0.05) but was without effect when the steroid precursors were used, suggestive of an inhibition between the sequence involving ACTH binding -> pregnenolone production. In contrast, etomidate and thiopentone reduced cortisol secretion by > 40% (P < 0.05) regardless of the stimulator used, indicating that at least one site of action was at the level of the final enzymic step of cortisol synthesis, i.e. 11beta-hydroxylase.

Adrenocorticotropic Hormone↗
PubMed1985

Myocardial infarction and cerebral infarction in a Danish suburban community.

A comparison was made of 485 cases of cerebral infarction (CI), registered prospectively in Frederiksberg, Copenhagen, with 495 cases of myocardial infarction (AMI), recorded retrospectively in the same population. The overall annual incidence of AMI was 6.5 per 1,000 population for males, 3.8 for females; the rates for CI were 2.4 and 2.5, respectively. The age-incidence curves, particularly those for AMI, were significantly steeper in the females than in the males. Correspondingly, the male predominance in the AMI-group subsided with age. Age-specific incidence-rates rose exponentially with advancing age; for each sex and diagnosis the relationship can thus be expressed as a simple mathematical formula, which may facilitate comparisons of incidence patterns in different communities. Theories explaining the similarities and differences of the age-incidence curves for AMI and CI are discussed.

Adult↗
PubMed1984

The cytochemical assay of NAD+ kinase activity in the follicular cells of guinea-pig thyroid gland.

A quantitative cytochemical assay for NAD+ kinase-like activity in the guinea-pig thyroid gland is described. The NADP+ produced by the activity of the kinase was used to drive the NADP+-dependent enzyme glucose-6-phosphate dehydrogenase which is endogenous to the tissue. The activity of glucose-6-phosphate dehydrogenase is greatly in excess of that of the kinase and was unaffected by the constituents of the kinase incubation medium (ATP, Mg2+ and NAD+) either alone or in combination. Kinase activity was dependent both on ATP and Mg2+, with maximal activity seen when the Mg-ATP ratio was between 1:1 and 4:1. Free ATP inhibited the activity of the enzyme. Enzyme activity was exhibited over a broad pH range (7-9) with a peak at pH 8.2. The sulphydryl-blocking agents, p-chloromercuribenzoate, iodoacetate and iodoacetamide (at 1 mM), completely abolished kinase activity but were without effect on glucose-6-phosphate dehydrogenase activity. N-ethylmaleimide and citrate (both at 1 mM) had no effect on either kinase or glucose-6-phosphate dehydrogenase activities.

Adenosine Triphosphate↗
PubMed1984

Cortisol production by dispersed guinea-pig adrenal cells; a specific, sensitive and reproducible response to ACTH....and its fragments.

Naturally occurring steroids and peptide hormones, tested at supraphysiological concentrations, were without effect on basal and human (h) 1-39 ACTH (NIBSC code 74/555, 25 ng/l (5.5 X 10(-12) mol/l] stimulated cortisol production. Further, low concentrations of angiotensin II, N-pro-opiocortin (N terminal fragment 1-76) and gamma-MSH all of which have been reported to synergise with ACTH with regard to cortisol production, were without significant effect alone or in combination with ACTH over the range 2.2 X 10(-13) to 5.5 X 10(-12) mol/l. The activity of h 1-39 was compared with that of the ACTH related peptides 1-24, 1-18, 1-17, 1-16, 1-13-NH2 (alpha MSH), 1-10 and 4-10. The dose responses were parallel and the same maximal cortisol output was observed with all the peptides except the 1-10 fragment. Half maximal stimulation occurred at 3.1 X 10(-12) (1-24), 4.4 X 10(-12) (h 1-39), 1.5 X 10(-11) (1-39), 3.3 X 10(-10) (1-18), 5 X 10(-9) (1-13-NH2), 8 X 10(-9) (1-17), 2 X 10(-7) (1-16) and 1 X 10(-5) (4-10) mol/l respectively. Interference by the above ACTH-derived peptides in cortisol secretion by the cells in response to 5.5 X 10(-12) mol/l h 1-39 ACTH was minimal over the range 5.2 X 10(-12)-2.2 X 10(-6) mol/l. The sensitivity of the adrenal cells to h 1-39 ACTH was such that 2 ng/l (4.4 X 10(-13) mol/l) provoked cortisol secretion over the control (P less than 0.05, n = 17). The coefficient of variation within assay for each dose on the full standard curve (2.2 X 10(-13)-1.1 X 10(-10) mol/l) was less than 10% (n = 6). Half maximal stimulation was given by 14.5 ng/l (3.2 X 10(-12) mol/l). Between control and 1.1 X 10(-10) mol/l ACTH there was a 32 +/- 8 (mean +/- SD, n = 9) fold change in cortisol production.

Adrenal Glands↗
PubMed1984

Preliminary experiences with a bioassay for adrenocorticotrophin (ACTH) in unextracted human plasma using dispersed guinea-pig adrenal cells.

A simple, accurate and precise dispersed cell bioassay suitable for the assay of ACTH levels greater than 50 ng/1 in unextracted human plasma is described. Isolated adrenal cells were prepared by tryptic digestion of the guinea-pig adrenal gland and cortisol production by these cells was controlled specifically by ACTH. The cortisol response to 2 ng/1 of human pituitary ACTH (1-39) was always significantly (P less than 0.05, n = 17) different from the control. ACTH concentrations between 1 and 20 ng/1 gave a 14 +/- 3 fold (n = 17) increment in cortisol response. Dilutions of test plasmas were parallel with standard ACTH (1-39) when the plasma concentrations were maintained throughout at 4% (1 in 25 dilution) by the addition of dexamethasone-suppressed plasma ('ACTH free'). Using a 25 fold plasma dilution, the limit of detection of ACTH in unextracted plasma was 50 ng/1. Recovery of ACTH (1-39) spiked into dexamethasone suppressed plasma at concentrations of 250 and 100 ng/1 was 97 +/- 10% (n = 17) and 114 +/- 15% (n = 17) respectively. The within assay coefficient of variation (CV) of both quality control and patients' samples never exceeded 9% while between assay variation was less than or equal to 13%. The sample throughput was 30 plasma samples/week/technician.

Addison Disease↗
PubMed1984

On the site of action of the anti-adrenal steroidogenic effect of etomidate and megestrol acetate.

The sites of action of the anti-steroidogenic action of etomidate and megestrol acetate have been established with a novel in vitro approach based upon the inhibition of cortisol (Co) secretion by dispersed guinea-pig adrenal cells. The cells were challenged with the Co precursor steroids (all at 10(-5) mol/l) pregnenolone (Pe), 17-hydroxy-pregnenolone (17-Pe), progesterone (Po), 17-hydroxyprogesterone (17-Po), 21-deoxycortisol (21-DOC) and 11-deoxycortisol (11-DOC), or 1-24 adrenocorticotrophin (ACTH, 100 ng/l) in the presence or absence of either etomidate, megestrol acetate, metyrapone or trilostane (all at 5 X 10(-5) mol/l). In the absence of drug, the steroid precursors or ACTH provoked a cortisol secretion of greater than 14 times that secreted by cells incubated in their absence. ACTH-stimulated Co secretion was inhibited by greater than 85% by all the drugs employed. In the presence of trilostane and megestrol acetate, Co secretion provoked by the delta 4 3-keto steroids (Po, 17-Po, 21-DOC and 11-DOC) was similar to the controls. However, with the delta 5, 3 beta-hydroxy steroids, 17-Pe and Pe, Co secretion was inhibited by greater than 57% in the presence of these drugs. In contrast, etomidate and metyrapone inhibited Co secretion by greater than 60% when 11-deoxycortisol was employed, indicative of a block at 11 beta-hydroxylase, the final step in the cortisol biosynthetic pathway. Similar results were seen with Pe, 17-Pe, Po and 17-Po, all of which are converted to cortisol via a biosynthetic route which includes catalysis by 11 beta-hydroxylase.(ABSTRACT TRUNCATED AT 250 WORDS)

3-Hydroxysteroid Dehydrogenases↗
PubMed1984

Skin refrigerants.

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Anesthetics, Local↗
PubMed1982

20 alpha-Hydroxysteroid dehydrogenase activity in the rat corpus luteum; a quantitative cytochemical study.

A quantitative cytochemical method for the demonstration of 20 alpha-hydroxysteroid dehydrogenase activity (20 alpha-HSD) in the regressing corpora lutea of the adult rat ovary is described. The method employs unfixed tissue sections and relies upon the oxidation of 20 alpha-hydroxy-4-pregnen-3-one (20 alpha-OH-P) with nitro blue tetrazolium as the hydrogen acceptor. The enzyme was dependent upon NADP+ for its activity and was inactive when 20 beta-hydroxy-4-pregnen-3-one (20 beta-OH-P) was used as a substrate. The apparent Km values for 20 alpha-OH-P and NADP+ were 3 x 10(-4) M and 2.5 x 10(-5) M respectively. Inhibition of 20 alpha-HSD activity by steroids was demonstrable at pH 8. Androstenedione was by far the most potent inhibitor, followed by progesterone (the product of the enzyme activity) 17 alpha-hydroxyprogesterone, Compound S and 20 beta-OH-P. At pH 6.8, a pH more favourable to the progesterone leads to 20 alpha-OH-P reaction, only progesterone and 17 alpha-hydroxyprogesterone were inhibitory. Testosterone was without demonstrable effect at either pH.

20-Hydroxysteroid Dehydrogenases↗
PubMed1981

Motility assay of human sperm by photon correlation spectroscopy.

Microscopic methods of performing motility assays of spermatozoa are slow, subjective, and involve a small number of spermatozoa. Laser light-scattering methods can analyze the motility of many spermatozoa within minutes. The swimming speed distribution of human spermatozoa was investigated by photon correlation spectroscopy. The sperm was diluted in seminal plasma to avoid modifying the viscosity. The swimming speed distribution was reconstructed from the correlation data by Stock's method of splines. When compared with a videomicroscopic assay, the reconstructed swimming speed distribution accurately reflects translational motion between 0 and 80 micrometers per second, while for speeds greater than 80 micrometers per second the distribution is distorted by the effects of rotational motion.

Humans↗