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Biomedical subjects

W R Robertson

Publications and source records attributed to W R Robertson.

At least 91 records · Page 5Linked to original sources
PubMed1988

Changes in the qualitative and quantitative secretion of luteinizing hormone (LH) following orchidectomy in man.

The effect of orchidectomy and thus withdrawal of testicular hormones on the biological and immunological properties of plasma LH was studied. Plasma samples were obtained from five men (mean age 71, range 65-81 years) with advanced carcinoma of the prostate, before orchidectomy and 1, 4, 8 and 16 weeks after surgery. LH bioactivity was estimated by a mouse Leydig cell bioassay and immunoreactivity by radioimmunoassay, using the same human pituitary LH standard 68/40. FSH and testosterone were measured by radioimmunoassay. Similar baseline data were obtained from a group (n = 17) of normal adult men (26, 19-36 years). Baseline bioactive (40 IU/l, median) and immunoreactive (10.8 IU/l) LH levels in the patients were higher (P less than 0.01) than in the controls (15.1 and 5.7 IU/l respectively), but bioactive to immunoreactive (B:I) LH ratios (3.4 +/- 0.2 versus 2.8 +/- 0.7) and testosterone levels (15.3 vs 18.7 nmol/l) were no different, consistent with compensated Leydig cell failure in the elderly men. After orchidectomy there was a greater increase in immunoreactive (46.6 IU/l at 16 weeks) than bioactive (80.3 IU/l) LH levels i.e. a fourfold vs twofold increase from baseline values. Consequently the B:I LH ratio decreased significantly (1.8 +/- 0.4 at 16 weeks) from the baseline ratio (P less than 0.0001) and that of the controls (P less than 0.01). These data indicate that acute withdrawal of testicular sex steroids results not only in quantitative change in LH secretion but also in qualitative change that decreases the biopotency of the LH molecules.

Aged↗
PubMed1988

Bioactive LH in women with polycystic ovaries and the effect of gonadotrophin suppression.

Discrepancies between levels of bioactive LH (B-LH) and immunoreactive LH (I-LH) in polycystic ovarian syndrome (PCO) have been reported previously. Serum levels of I-LH, B-LH (by dispersed Leydig cell assay), FSH, oestradiol (E2) and progesterone (Prog) were measured once to three times weekly over 4 weeks in 13 women with classical clinical, ultrasound and endocrine features of PCO. Eleven women attending for infertility but whose profiles when studied three times weekly by combined endocrine and ultrasound assessment were normal and ovulatory served as controls. Seven of the women with PCO were evaluated during and after 3 weeks suppression with ethinyloestradiol (30 micrograms) plus 150 micrograms either of desogestrel or levonorgestrel; two were given both treatments. Both I-LH and B-LH levels were higher in PCO patients (20 +/- SD 5 U/l and 46 +/- 9 U/l respectively, P less than 0.0001), compared with all phases of the normal cycles except the mid-cycle peak. The B-LH to I-LH (B:I LH) ratio in PCO patients (2.5 +/- 0.7) was higher than in all the control cycle phases (P less than 0.05). I-LH, B-LH, B:I LH ratio, FSH and E2 were all suppressed from the second week of oestrogen-progestogen treatment (P less than 0.01) and returned gradually to pretreatment levels by the third or fourth week after suppression. The LH and FSH levels and B:I LH ratio in PCO patients during suppression were comparable with levels in the early and mid-follicular phases of control cycles but the LH/FSH ratio remained significantly raised (P less than 0.01) at 2.3 +/- 0.7.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1988

Plasma inhibin levels in men with chemotherapy-induced severe damage to the seminiferous epithelium.

Immunoreactive plasma inhibin levels and free testosterone index (FTI) were estimated in 17 patients who had previously received combination chemotherapy for Hodgkin's disease and in 16 age-matched controls. In the same patients we had previously found significantly raised FSH and LH levels in the presence of normal basal and HCG-stimulated total testosterone levels. Mean plasma inhibin levels were not different between the patients (601 +/- 321 U/l) and controls (530 +/- 174 U/l) nor were FTI values (81.5 +/- 35 vs 91 +/- 47 respectively). There was a positive correlation (r = 0.53, P less than 0.05) between FSH and inhibin levels and a negative correlation between FSH and FTI (r = -0.51, P less than 0.05) in the patients but not in the controls. No such correlations with inhibin or FTI existed for LH but there was a positive correlation between LH and FSH levels in the patients. In four patients inhibin levels were pathologically raised and in this group mean FSH values (21.7 +/- 4.7 IU/l) were higher (P less than 0.001) and mean FTI (59.1 +/- 22.6) lower (P less than 0.001) than respective values (13.6 +/- 5.3 IU/l and 88.4 +/- 35) for the remainder of the patients. These data are not compatible with the hypothesis that inhibin is the major negative feedback signal for the control of FSH secretion.

Adolescent↗
PubMed1988

Acute stimulation of thyroidal NAD+ kinase, NADPH reoxidation, and peroxidase activities by physiological concentrations of thyroid stimulating hormone acting in vitro: a quantitative cytochemical study.

Quantitative cytochemical techniques have been employed in a study of some of the acute effects of low doses (0.01----1 mU/liter) of TSH on the metabolism of guinea pig thyroid segments maintained in nonproliferative organ culture. The enzymes involved in the synthesis of NADP+ (NAD+ kinase), its reduction by the pentose-shunt (glucose 6-phosphate dehydrogenase), and its reoxidation both by the microsomal electron chain (diaphorase activity) and by participation in other cellular processes, have been examined. The effect of TSH on peroxidase activity has also been studied. After 10 min stimulation with TSH (1 mU/liter) there was a 60% increase in NAD+ kinase activity which preceded changes in the microsomal reoxidation of NADPH (up 33% by 30 min). There were no changes in the activity of glucose 6-phosphate dehydrogenase. There was a sustained rise in peroxidase activity which reached 129% over control after 30 min. This is the first in vitro demonstration of an acute stimulation of peroxidase and kinase activities by physiological concentrations of TSH. NADPH reoxidation after stimulation with TSH was such that the ratio of NADPH reoxidized via the microsomal respiratory pathway (diaphorase, hydrogen pathway 1) relative to that available for cytosolic utilization (hydrogen pathway 2) increased compared to the unstimulated controls. We suggest that increased NADP+ production (via NAD+ kinase activity) and the preferential shuttling of the NADPH for reoxidation via the microsomal respiratory pathway, coupled with greatly stimulated peroxidase activity, may be important regulators of the control of thyroglobulin iodination and hence thyroid hormone production.

Animals↗
PubMed1988

Characterization of the regenerated Leydig cell population of the rat after destruction by ethylene-1,2-dimethanesulphonate.

The Leydig cells repopulating the adult rat testis after destruction by a single injection of the cytotoxic ethylene-1,2-dimethanesulphonate (EDS) were investigated. After 14 days, serum concentrations of LH and FSH were significantly raised and concentrations of testosterone in the serum and testis reduced. At 21 days, hormone concentrations had returned to within the normal range. Binding of 125I-labelled human chorionic gonadotrophin (hCG) to testis homogenate, however, was still less than 10% of normal. After 21 or 28 days the 125I-labelled hCG binding profiles of isolated Leydig cells from EDS-treated rats, separated on a Percoll gradient, showed a single peak similar to that of immature (25 days old) rats. After 49 days, 125I-labelled hCG binding resolved into two peaks more like that of normal adult rats. Using a quantitative cytochemical method, 3 beta-hydroxysteroid dehydrogenase activity in individual Leydig cells of unfixed testis sections was determined. Activity was increased by 70% (P less than 0.05) in repopulating Leydig cells 21 days after EDS treatment compared with cells from vehicle-treated rats. In addition, Leydig cells were still capable of further 'in-vivo' stimulation by pharmacological doses of hCG. These data indicate that Leydig cells repopulating the testis are homogeneous. Fewer cells from the newly formed population are capable of maintaining normal serum concentrations of testosterone and must thus be individually more active in secreting testosterone. In these respects, the Leydig cells repopulating the adult rat testis after EDS treatment more closely resemble those of the fetal rat testis.

3-Hydroxysteroid Dehydrogenases↗
PubMed1988

The stability in vitro of bioactive and immunoreactive LH in human blood and plasma.

The degree of stability in vitro of bioactive and immunoreactive LH in human blood, plasma and serum was examined. Bioactivity and immunoreactivity of LH were assayed by the dispersed mouse Leydig cell assay and by standard radioimmunoassay respectively, using the same reference preparation (first international reference preparation for human pituitary LH 68/40 for immunoassay). Bioactive and immunoreactive LH were stable in blood and plasma at 22 degrees C for up to 4 and 24 h respectively, and in blood at 4 degrees C for up to 24 h. There was no loss of biological or immunological LH activity in plasma which had been either snap-frozen and stored at -70 degrees C, allowed to freeze at -20 degrees C and stored at that temperature or kept at 4 degrees C for 24 h and then stored at -70 degrees C. Likewise, the levels of LH in plasma and serum which had been stored at either -20 or -70 degrees C and then thawed and refrozen up to four times remained unchanged. In addition, the biological and immunological activity of LH was not affected after vortexing samples of plasma or serum for up to 60 s. Bioactive LH was also stable in plasma samples after prolonged storage (up to 9 months) at either -70 or -20 degrees C. We conclude that LH bioactivity and immunoreactivity are stable in blood and plasma following a variety of treatments commonly experienced during normal handling of a blood sample after venepuncture.

Biological Assay↗
PubMed1987

Mechanisms leading to hypogonadism in men with burns injuries.

A profound and persistent depression of serum testosterone concentrations was found in 19 men with burns injuries. This could not be explained by changes in sex hormone binding globulin capacity, hyperprolactinaemia, classical primary testicular failure, or a hypogonadotrophic state. Pulsatile release of luteinising hormone was found in control subjects but was absent or diminished in burnt patients with low serum testosterone concentrations. In addition, these patients showed reduced biological activity of luteinising hormone as measured by bioassay even though normal concentrations of luteinising hormone were detected by radioimmunoassay. The temporary hypogonadism after burns injury and possibly in other clinical states may be related to hypothalamic dysfunction, which leads to abnormal generation of luteinising hormone releasing hormone and non-pulsatile secretion of luteinising hormone of reduced biological activity.

Adult↗
PubMed1987

Assessment of the biopotency of anti-thyroid drugs using porcine thyroid cells.

The effect of six drugs on the uptake and organification of iodide by porcine thyroid cells stimulated with bovine TSH (10 miU/L) has been investigated. The drugs fall into two categories: the peroxidase inhibitors, methimazole (MMI), 2-thiouracil (2-TU) and 3-amino 1,2,4 triazole (3-ATA) and the ionic inhibitors, lithium chloride (LiCl), potassium perchlorate (KC10(4], and sodium iodide (NaI). All the drugs led to a dose-related inhibition of iodide metabolism. The most potent effect on iodide uptake was seen with NaI which inhibited this function by 20% even at 10(-8) mol/l. In contrast, the most potent effect on iodide organification was observed with methimazole which led to a 25% inhibition at 10(-8) mol/l. The concentrations of drug which gave rise to a 50% inhibition of iodide uptake were (mumol/l) 0.26 (NaI), 3.5 (KClO4), 9.7 (2-TU), 15 (MMI), 26 (3-ATA), and 1500 (LiCl). The comparable figures for organification were 0.13 (MMI), 0.18 (2-TU), 0.23 (NaI), 1.2 (3-ATA), 15 (KClO4) and 1300 (LiCl). We conclude that this in vitro system has considerable potential for the assessment of potency and possible bioassay of anti-thyroid drugs of varying structures and sites of action.

Animals↗
PubMed1987

Delta (5)3 beta hydroxysteroid dehydrogenase activity of the rat corpus luteum exhibits positive substrate-binding co-operativity: a continuous monitoring microdensitometric study with unfixed ovarian tissue sections.

delta (5)3 beta hydroxysteroid dehydrogenase activity transforms biologically inactive delta (5)3 beta hydroxy steroids into the active delta (4)3-keto products (e.g. pregnenolone to progesterone). Using a cytochemical procedure which allows for the continuous microdensitometric monitoring of an enzyme reaction as it proceeds and a well described cytochemical assay for delta (5)3 beta HSD we have analysed the initial velocity rates (Vo) for dehydroepiandrosterone (DHEA) binding to this enzyme in regressing (i.e. 20 alpha hydroxy steroid dehydrogenase positive) corpus luteum (CL) cells in unfixed tissue sections (5 micron) of the dioestrous and proestrous rat ovary. The results are mean +/- S.E.M. The relationship between DHEA concentration (0 to 50 microM) and delta (5)3 beta HSD activity in the dioestrous corpora lutea was sigmoidal and had an atypical 1/Vo versus 1/S plot, the x intercept being positive. Using a 1/Vo versus 1/S2 plot the Vmax was determined to be 1.0 +/- 0.08 mumol min-1 mg-1 CL (n = 6). The Hill constant was 2.7 +/- 0.02 (n = 6) suggesting a high degree of positive co-operativity for DHEA binding. The S concentration for half maximal activity was 17 +/- 1 mumoles (n = 6). In the corpora lutea cells of the proestrous ovary, the Vmax for DHEA transformation was unchanged (0.95 +/- 0.04 mumol min-1 mg-1, n = 3) whilst the S0.5 was significantly increased to 27 +/- 0.1 (p less than 0.01, n = 3). The Hill constant remained positive being 2.9 +/- 0.2 (n = 3).(ABSTRACT TRUNCATED AT 250 WORDS)

3-Hydroxysteroid Dehydrogenases↗
PubMed1987

The pituitary-Leydig cell axis in men with severe damage to the germinal epithelium.

Eighteen men (mean age 27, range 18-30 years) treated for Hodgkin's disease with 6-8 courses of MVPP (Mustine, Vinblastine, Procarbazine and Prednisolone) have had Leydig cell function assessed by their steroidogenic responses to stimulation by a single bolus dose of HCG (1000 units intramuscularly). Normal age-matched men (n = 16) acted as controls. Baseline immunoreactive FSH was markedly raised in the patients (mean 18.1 +/- SD 6.9 vs 2.0 +/- 1.5 IU/l, P less than 0.0001) reflecting damage to the germinal epithelium. Immunoreactive LH was also greater in patients (10.3 +/- 3.9 IU/l) than in controls (3.9 +/- 1.9 IU/l, P less than 0.0001). There were no differences between the baseline testosterone, androstenedione, oestradiol, oestrone and sex hormone binding globulin (SHBG) concentrations. The testosterone/SHBG ratios were similar in the two groups and there was no correlation between baseline LH and testosterone concentrations or testosterone/SHBG ratios. Testosterone, androstenedione, oestradiol and oestrone secretion in response to HCG stimulation were similar at 24 h and 96 h in both groups. In order to explain the paradox of elevated immunoreactive LH in the face of normal testicular steroidogenesis in such patients, LH biological activity (B) as well as LH immunoreactivity (I) and FSH and testosterone were estimated in a second similar group of patients (n = 17, mean age 27, range 17-43 years) and in a further age-matched control group (n = 17). Bioactive and immunoreactive LH levels were significantly increased (P less than 0.005 and P less than 0.001, respectively) in the patient group.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1987

Biopotency and site of action of drugs affecting testicular steroidogenesis.

The effect of etomidate (an anaesthetic), epostane (WIN 32729; an inhibitor of ovarian and adrenal steroidogenesis) and cyproterone acetate (an antiandrogen) on testosterone secretion from mouse Leydig cells stimulated with LH (5 i.u./l) was tested. The concentration of drug which inhibited testosterone secretion by 50% was 11.5 +/- 1.1 (S.E.M.) mumol/l for cyproterone acetate, 1.2 +/- 0.2 mumol/l for etomidate and 0.23 +/- 0.03 mumol/l for epostane. The effect of all three drugs on testicular steroidogenesis was completely reversible. Thus testicular cells which had been washed after exposure to a greater than 95% inhibitory dose of drug responded in a similar manner to hormone stimulation as cells similarly washed and which had not been exposed to the drug. The sites of the antisteroidogenic effect of epostane, etomidate and cyproterone acetate were established using a method based on the sequential stimulation by the exogenous precursor steroids of the various steps leading to the biosynthesis of testosterone. It was concluded that etomidate acts at the sequence between LH binding and pregnenolone production, epostane acts at 3 beta-hydroxysteroid dehydrogenase and cyproterone acetate inhibits 3 beta-hydroxysteroid dehydrogenase and C17,20-lyase.

Androgen Antagonists↗
PubMed1986

The effect of ketoconazole on adrenal and testicular steroidogenesis in vitro.

The effect of the anti-fungal agent, ketoconazole, on cortisol secretion from adrenal cells stimulated with ACTH (1-24, 50 ng/l) and on testosterone secretion from Leydig cells stimulated with LH (5 i.u./l), has been tested. The concentration of drug which inhibited cortisol and testosterone secretion by 50% (ED50) was 3.6 +/- 0.7 mumol/l and 0.61 +/- 0.03 mumol/l, respectively. The effect of ketoconazole on adrenal and testicular steroidogenesis was completely reversible. Thus, adrenal and testicular cells which had been washed after exposure to greater than 95% inhibitory dose of ketoconazole responded in a similar manner to hormone stimulation as cells similarly washed and which had not been exposed to drug. The sites of the anti-steroidogenic effect of ketoconazole have been established using a method based upon the sequential stimulation by the exogenous precursor steroids of the various steps leading to the biosynthesis of cortisol by adrenal cells and testosterone by Leydig cells. We conclude that ketoconazole reversibly inhibits the sequence between ACTH/LH binding and pregnenolone production and also inhibits testicular C-17-C-20, lyase activity.

Adrenal Glands↗
PubMed1986

The application of continuous monitoring microdensitometry to an analysis of NAD+ binding and 3 beta-hydroxy-delta 5-steroid dehydrogenase activity in the regressing corpus luteum of the pro-oestrous rat ovary.

A technique which allows for the continuous microdensitometric monitoring (at 3.3 s intervals) of an enzyme reaction has been applied to a study of 3 beta-hydroxy-delta 5-steroid dehydrogenase activity in regressing corpora lutea cells of unfixed tissue sections (5 micron thick) of the pro-oestrous rat ovary. Initial reaction rates for NAD+ reduction by the activity of the enzyme were maintained for less than 3 min at all the concentrations (0-500 mumol/l) of co-factor employed. The relationship between NAD+ concentration and enzyme activity under initial velocity rate conditions was hyperbolic and maximum enzyme activity was seen when the NAD+ concentration was greater than 250 mumol/l. The apparent Michaelis-Menten constant (Km) was 29 mumol/l and Vmax 0.63 mumol NAD+ reduced/min/cm3 corpora lutea.

3-Hydroxysteroid Dehydrogenases↗
PubMed1986

Protein-A purified human immunoglobulins: a comparison of thyroid stimulating and thyrotrophin receptor binding activities in thyrotoxicosis.

Protein-A purified human thyroid stimulating immunoglobulins (TSIg) and thyrotrophin binding inhibiting immunoglobulins (TBIIg) were measured in euthyroid subjects and thyrotoxic patients by bioassay and TSH radioligand receptor assay respectively. Unextracted sera from euthyroid and thyrotoxic subjects inhibited both basal and TSH stimulated iodide uptake in the bioassay, which was based on iodide uptake in porcine thyrocytes. Similar effects were seen with Ig and TSIg extracted from sera using either polyethylene glycol or ammonium sulphate. However IgG and TSIg prepared using Protein-A Sepharose CL-4B from sera of euthyroid subjects had little effect in this system. The majority of Protein-A purified TSIg preparations from sera of thyrotoxic patients stimulated iodide uptake in procine thyrocytes in a dose-dependent manner and most (85%) diluted parallel to both bovine and human TSH. TSIg and TBIIg from 73 patients with thyrotoxicosis were assessed using the bioassay and receptor assay and compared to a control group of 35 euthyroid subjects. The median (and range) values for TSIg and TBIIg in the euthyroid group were 4.35 (0.8 to 7.5, % stimulation over control) and 2.7 (-9.3 to 8.6, TBII index) for the bioassay and radioreceptor assay respectively. A value of greater than 10.0 in both assays was taken as a positive result. Of the thyrotoxic patients 61 out of 73 were positive in the bioassay (83.6%) compared to 60 in the radioreceptor assay (82.2%). There was a positive correlation between the two assays (r = 0.821, P less than 0.001). Of the 73 thyrotoxic patients 40 were untreated, 18 had received carbimazole and 15 had been previously treated with iodine-131. TSIg levels in the untreated thyrotoxics were similar to those in either group of treated patients. However they were higher (P less than 0.05) in the iodine-131 group than in the patients treated with carbimazole. Similar results were obtained for TBIIg. The coupling of a specific extraction method for human serum IgG with a bioassay for TSIg has demonstrated a high prevalence of these immunoglobulins in patients with thyrotoxicosis. The agreement between this assay and a radioreceptor assay was good, indicating that TSH displacing and thyroid stimulating activities of these immunoglobulins are closely related.

Adult↗
PubMed1986

On the assessment of the in vitro biopotency and site(s) of action of drugs affecting adrenal steroidogenesis.

Dispersed guinea-pig adrenal cells have been employed in the in vitro estimation of the biological potency and sites of action of drugs acting against the adrenal. The effect of 12 drugs on cortisol secretion from cells stimulated with adrenocorticotrophin (ACTH, 50 ng/L, a 95% saturating dose) has been tested. All the drugs depressed cortisol output in a dose-related fashion. The concentration of drug which inhibited secretion by 50% was (mumol/L, mean +/- SEM): etomidate 0.1 +/- 0.002; epostane 0.44 +/- 0.02: 17-ketotrilostane 0.55 +/- 0.04: trilostane 1.3 +/- 0.1: metyrapone 3.5 +/- 0.6: cyproterone acetate 4.6 +/- 0.2: megestrol acetate 11 +/- 2: danazol 22 +/- 2: aminoglutethimide 41 +/- 5: stanozolol 50 +/- 4: thiopentone 160 +/- 18: propofol 170 +/- 18. The sites of the anti-steroidogenic effect of seven of these drugs have also been established using a method based upon the sequential stimulation by the exogenous precursor steroids of the various steps leading to the biosynthesis of cortisol by adrenal cells. Propofol acts between ACTH binding and pregnenolone production, trilostane, megestrol acetate and cyproterone acetate are 3 beta-hydroxysteroid dehydrogenase inhibitors whereas metyrapone, etomidate and thiopentone act at 11 beta-hydroxylase.

Adrenal Cortex↗
PubMed1986

Effect of adrenocorticotrophin on cortisol and androstenedione secretion from dispersed cells of guinea-pig adrenal zonae fasciculata and reticularis.

We have studied cortisol and androstenedione secretion by dispersed cells of the outer zona fasciculata (ZF) plus zona glomerulosa, and the inner zona reticularis (ZR) plus medulla of the guinea-pig adrenal. The ZF and ZR were microdissected apart, the cells dispersed and incubated (200 000 cells/ml) for 90 min in the presence of adrenocorticotrophin (ACTH; 500 ng/l), dibutyryl cyclic AMP (dbcAMP; 1 mmol/l), pregnenolone, 17-hydroxypregnenolone, 17-hydroxyprogesterone, 11-deoxycortisol and 21-deoxycortisol. The steroid concentrations were 5-25 mumol/l. Cortisol secretion was assayed by radioimmunoassay. There was no detectable cortisol secretion (less than 50 nmol/l) from the ZR in the controls (no additive) or after dbcAMP stimulation. Adrenocorticotrophin-stimulated cortisol secretion was also low (range less than 50-340 nmol/l). In contrast the ZF secreted 177-379 (control), 828-2052 (dbcAMP) and 2863-9735 (ACTH) nmol cortisol/l. There was no detectable (i.e. less than 2 nmol/l) cAMP production by ZR or ZF either basally (no ACTH) or after ACTH stimulation (500 ng/l). Challenge of the ZR cells with each cortisol precursor steroid (5 mumol/l) increased (P less than 0.05) cortisol secretion over that seen with the corresponding basal and ACTH-stimulated controls. Thus pregnenolone, 17-hydroxypregnenolone, 17-hydroxyprogesterone, 11-deoxycortisol and 21-deoxycortisol (converted directly to cortisol by 21-hydroxylase) gave rise to (mean +/- S.D., n = 4) 406 +/- 86, 680 +/- 180, 1307 +/- 111, 1141 +/- 234 and 3160 +/- 419 nmol cortisol/l respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

17-alpha-Hydroxypregnenolone↗