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Biomedical subjects

P Roos

Publications and source records attributed to P Roos.

At least 73 records · Page 4Linked to original sources
PubMed1987

Biological characterization of purified native 20-kDa human growth hormone.

Because of the propensity of the 20-kDa variant of human growth hormone (GH) to aggregate with itself and with 22-kDa human GH, it has been difficult to prepare monomeric 20-kDa GH in highly purified form. This has been a major complicating factor in determining whether 20-kDa GH has a biological activity profile distinct from that of 22-kDa GH. In the present study, native 20-kDa GH was isolated from a human GH dimer concentrate and purified by a procedure that included column electrophoresis in agarose suspension as a final separation step. This procedure yielded highly purified monomeric 20-kDa GH, which was contaminated to an extent of less than 1% with 22-kDa GH, and which exhibited only a small degree of dimerization upon storage. The native 20-kDa GH was quite active in stimulating growth in hypophysectomized rats, when growth was assessed by body weight gain, longitudinal bone growth, the stimulation of sulfation of cartilage, and the elevation of serum IGF-1 level. However, in all of these growth assays, the 20-kDa GH was somewhat less active than the native 22-kDa GH to which it was compared; e.g., in the body weight gain and longitudinal bone growth assays, it had an estimated potency of 0.6 relative to the 22-kDa GH. The 20-kDa GH exhibited substantial diabetogenic activity when tested for the ability to raise fasting blood glucose concentration and to impair glucose tolerance in ob/ob mice. Also, the native 20-kDa GH had significant in vitro insulin-like activity, although its potency was approximately 20% that of the native 22-kDa GH to which it was compared. Thus, the biological activity profile of native 20-kDa GH differs from that of 22-kDa GH primarily in that insulin-like activity is markedly attenuated.

Animals↗
PubMed1987

Isolation of three electrophoretic variants of rat pituitary growth hormone.

A procedure has been developed for the isolation of rat pituitary growth hormone and for the subsequent resolution of the preparation into three variants by preparative electrophoresis. The starting material was whole frozen glands and the process involved homogenization and extraction at pH 6.2, ammonium sulfate fractionation and molecular-sieve chromatography on Sephadex G-100. The separation into charge variants was achieved by zone electrophoresis in agarose suspension at alkaline pH. The purification was monitored by radioimmunoassay and the specific activities were expressed in terms of the rat growth hormone reference preparation (RP-1) supplied by the NIADDK, Bethesda, U.S.A. The three-component preparation and its constituents all had activities in the same range, exceeding the activity of the reference by a factor up to 20 times. Bioassay of the three-component preparation, based on measurement of longitudinal bone growth in hypophysectomized rats gave a potency of 4-5 IU/mg. The reference was the 1st International Standard (bovine) for growth hormone. The yield of the three-component preparation was 3.3 mg per gram pituitary tissue. Different electrophoretic analyses revealed the efficiency of the preparative procedure in separating the variants. The results of the analyses also support the view that difference in electrophoretic behaviour is due to a difference of a single net charge between adjacent variants. In addition, growth hormone was prepared from two side extracts (at pH 7.0 and pH 9.8, respectively), provided by a procedure developed earlier for rat prolactin. The three preparations gave electrophoretic patterns of equal appearance although the relative proportions of the activity peaks differed.

Animals↗
PubMed1986

Suppression of low-dose streptozotocin-induced diabetes by immunomodulatory lectins.

The effect of immunomodulatory lectins on diabetes development following low-dose streptozotocin treatment in inbred mice was studied. All lectins administered had been shown previously to suppress immune reactivity. Among plant lectins concanavalin A but not Lens culinaris or wheat germ agglutinin partially suppressed hyperglycaemia following low-dose streptozotocin. A similar inhibitory effect was found for the staphylococcal enterotoxin B. Finally, administration of an immunomodulatory lectin from vertebrates, electrolectin, also had a beneficial effect on the course of the disease. These findings indicate that some lectins have a suppressive effect on Type 1 diabetes in an animal model.

Animals↗
PubMed1985

Effects of TRH and a rat TSH preparation on discrete hypothalamic and forebrain catecholamine nerve terminal networks in the hypophysectomized male rat.

A rat pituitary TSH preparation in doses of 10 and 100 micrograms/kg produced rapid and marked increases in dopamine (DA) levels and alpha-methyltyrosine-induced decline consistent with increased DA synthesis and release in the medial and lateral palisade zones (MPZ, LPZ) of the median eminence, and reduced noradrenaline (NA) turnover in the paraventricular hypothalamic nucleus (PA) of the hypophysectomized male rat. The TSH serum levels measured in these rats 2 h after the injection were within the physiological range after the injection of 10 micrograms/kg. TRH given intravenously in a dose of 100 micrograms/kg produced rapid and marked increases of DA release in the MPZ and LPZ of the median eminence and reduction of NA turnover in the PA of the hypophysectomized male rat. The TRH injection did not alter the serum levels of prolactin, TSH, T3 and T4. The results indicate that TRH-TSH-DA interactions take place in the local circuits in the median eminence thus supporting the view that a short and an ultrashort feedback action of rTSH and TRH respectively may exist in the median eminence. The rapid action of the rat TSH preparation as well as of TRH further supports this concept. The rat TSH preparation and TRH produced marked reductions in NA turnover in the PA. These results support the possibility that rat TSH and TRH may, via an action on the hypothalamus, influence the facilitatory noradrenergic mechanism operating at the soma-dendritic level of the TRH immunoreactive neurons projecting to the median eminence. Thus, the existence of a neuronal feedback loop from the medio-basal hypothalamus into the paraventricular hypothalamic nucleus is postulated.

Animals↗
PubMed1985

Comparison between the effects of FSH and LH on the steroidogenic pattern in isolated pre-ovulatory rat follicles.

The effects of purified FSH preparations from three different species (ovine, human, rat) on the steroidogenic pattern in isolated pre-ovulatory follicles of PMSG-treated immature rats were examined and compared to the effects of LH (ovine, human). Steroids were analyzed by RIA. During 6 h incubation, all three FSH preparations in a concentration range from 10-100 ng/ml caused a significant increase in progesterone (P), testosterone (T), and oestradiol (E2) accumulation. With LH a significant increase in steroid accumulation was seen in a concentration of 1 ng/ml. The time-course of stimulation by hFSH and hLH in a concentration of 10 ng/ml showed no difference in steroidogenic response between the two hormones, with a significant stimulation of steroids within 2 h. During prolonged incubation (6-8 h), accumulation of T was measured in the presence or absence of unlabelled 17 alpha-hydroxyprogesterone. Follicles exposed to LH did not increase their T formation, while follicles incubated in plain medium or in the presence of oFSH increased their T formation. In a concentration of 100 ng/ml hFSH showed a tendency to reduce maximal T production. These results suggest that FSH alone in physiological doses may stimulate T and E2 production in the preovulatory follicle.

Animals↗
PubMed1985

Effects of FSH and LH on adenylate cyclase activity in rat granulosa cell membranes during follicular maturation.

Adenylate cyclase activity was measured in membranes prepared of granulosa cells isolated from PMSG-treated immature rats and the effects of ovine, highly purified human and rat gonadotrophins were compared. Furthermore, a comparison of the effects of the human preparations (hLH, hFSH) on the adenylate cyclase activity in membranes prepared from granulosa cells isolated at different stages of follicular maturation, was performed. The adenylate cyclase in membranes of immature granulosa cells was stimulable with FSH but not with LH, while in pre-ovulatory granulosa cell membranes, both gonadotrophins were stimulatory with FSH generally being more effective than LH. Surprisingly, the dose-response curve for ovine LH (oLH) was biphasic with a plateau at a level of adenylate cyclase activity corresponding to the maximal stimulatory effect of hCG. With increasing oLH concentrations the response resumed and the maximal stimulation corresponded to that of oFSH. With highly purified rat gonadotrophins the FSH response was significantly higher than the response to LH at all concentrations tested. Using highly purified human gonadotrophins the maximal FSH response was 50% higher than the maximal LH response and by adding increasing concentrations of hFSH to a maximally stimulatory concentration of hLH it was possible to mimic the biphasic dose-response curve for oLH. When the membranes were prepared from granulosa cells isolated after the pro-oestrus LH/FSH surge there was clear increase in the sensitivity of the adenylate cyclase to stimulation with LH although the maximal response was unaffected.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenylyl Cyclases↗
PubMed1985

Effect of gonadotrophins on progesterone secretion by cultured granulosa cells obtained from human preovulatory follicles.

Granulosa cells were obtained from human preovulatory follicles in 31 women undergoing in vitro fertilization and embryo transfer due to tubal infertility. Follicular maturation was stimulated and synchronized by treatment with Clomiphene or human menopausal gonadotrophin (hMG), or both, plus human chorionic gonadotrophin (hCG). Follicles were aspirated by ultrasound guided puncture approximately 34-36 h after the hCG injection. The granulosa cells were washed and suspended in modified medium 199 containing 10% foetal bovine serum and cultured as monolayers for 6-8 days in the absence and presence of hormones and reactants. Progesterone formation was analyzed by RIA. In general, the cells underwent morphological luteinization and secreted high amount of progesterone. Under basal conditions the secretion of progesterone was highest during the first 2 days in culture and then gradually declined. Progesterone secretion was stimulated by human LH, hCG and the adenylate cyclase stimulator forskolin, with a maximal effect between days 2-6. The beta-adrenergic agonist isoproterenol in preliminary experiments potentiated the stimulatory effect of hCG but had no own stimulatory effect. No clear differences in progesterone secretion or responsiveness to in vitro stimulation relating to the various in vivo stimulation protocols were found.

Cell Count↗
PubMed1985

Stability studies on human pituitary prolactin.

Differently charged isomers of human pituitary prolactin were subjected to stability studies under various conditions. The study was guided by analytical electrophoresis and radioimmunoassay. In alkaline medium (pH 10) it was found that each isohormone was converted into faster-migrating components, probably due to deamidation. A quantitative study of this alteration was made by measuring the rate constant for this conversion assuming first-order kinetics. The result indicated that the rate constant decreased with increased acidity of the examined prolactin components. Furthermore, this alteration was also found to be paralleled by a decreased immunoactivity of the components. At neutral pH the conversion reaction was studied both in 0.9% NaCl and in human serum. For the least acidic isohormone the rate constant in serum was calculated to be about four times higher than that in the saline solution which in turn was comparatively low. It was concluded that the alteration observed at pH 10 might be attributed to non-enzymic deamidation. The conversion of prolactin that occurred in human serum indicated that this reaction could be caused by enzymes. A study of the immunoactivity of human prolactin as a function of time at various storage conditions is also included in this work. The stability of the hormone was found to be strictly concentration dependent and also affected by buffer ionic strength. In the presence of ethylene glycol (50%, v/v) at -20 degrees C and at a protein concentration in the range of 0.1-2 mg/ml the hormone was found to be both immunologically and electrophoretically stable for several years.

Drug Stability↗
PubMed1984

A collaborative evaluation of a rapid automated bacterial identification system: the Autobac IDX.

A collaborative evaluation of the Autobac IDX, a rapid, semi-automated bacterial identification system, was performed in three independent laboratories in three European countries. The system utilises growth inhibition by a series of chemical compounds. Subsequent analysis of the resultant data by quadratic discriminant function automatically results in a bacterial identification. Three sets of 30 strains were examined repeatedly in each of the participating laboratories. The reproducibilities obtained ranged from 85.6% to 96.6%, with an overall average of 91.8%. The accuracy of the system was also determined by examining 1076 isolates from the three participating laboratories. An overall accuracy of 90.3% was calculated by comparing the Autobac result with a reference method. When the results were weighted to represent clinical frequency, the accuracy was 93.6%.

Bacteria↗
PubMed1984

GalNAc/Gal-specific rat liver lectins: their role in cellular recognition.

By investigating the presence and distribution of GalNAc/Gal-specific receptors on liver cells in vitro and in vivo, we provided evidence that the hepatocyte is not the only liver cell expressing receptor activity but that receptors of similar specificity are found on liver macrophages and also on endothelial cells. The receptor distribution in the plasma membrane is strinkingly different between the three cell types, as judged from the binding pattern of colloidal gold particles coated with asialofetuin or lactosylated serum albumin. Binding to hepatocytes occurs as single particles statistically distributed, binding to liver macrophages in a clustered arrangement all over the cell membrane and binding to endothelial cells also in a clustered arrangement but restricted to coated pits only. The different receptor distribution results in different binding and uptake abilities. Whereas hepatocytes bind and take up molecules and small particles (5 nm) only, the clustered receptor arrangement of endothelial cells and macrophages enables them to effectively bind and ingest larger particles. Ligands larger than 35 nm can be taken up by the macrophages only. The different receptor arrangement results also in different capacities of cell contact formation. Although in vitro liver macrophages and hepatocytes can both bind desialylated cells the macrophage needs much less galactosyl groups exposed on erythrocytes to establish stable contacts than the hepatocyte. The contacts formed by hepatocytes stay reversible for 30 min at 37 degrees C, whereas the contacts formed by the liver macrophages become irreversible after 10 min at 37 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1983

Rat growth hormone and hypothalamic catecholamine nerve terminal systems. Evidence for rapid and discrete reductions in dopamine and noradrenaline levels and turnover in the median eminence of the hypophysectomized male rat.

Rat growth hormone (rGH) (100 micrograms/kg) produced 2-4 h after its i.v. injection a rapid reduction of catecholamine stores and turnover in the subependymal layer and in the medial and lateral palisade zone of the median eminence. It is suggested that rGH may inhibit its own secretion partly via reduction of DA synthesis and release in the median eminence leading to increased somatostatin release and partly via reduced noradrenaline synthesis and turnover in the median eminence leading to reduced secretion of a growth hormone releasing factor.

Animals↗
PubMed1983

Thyroid hormone secretion rates: response to endogenous and exogenous TSH in man during surgery.

The thyroid hormone secretory response to TSH was studied in twenty-eight patients undergoing thyroid and parathyroid surgery. Eighteen patients received bovine or human exogenous TSH by injection into a thyroid artery, and 10 received TRH to stimulate endogenous TSH secretion. Thyroid secretion rates of T4, T3, and rT3 were determined directly from measurement of blood flow and the hormone gradient across the gland. A significant secretory response was seen for all three hormones following TSH increase. T3 secretion accelerated more rapidly than that of T4 and rT3, thus reducing the T4/T3 and rT3/T3 ratios. The T4/rT3 ratio fell during the first 30 min but then increased. The responses correlated with the area under the curve of the TSH serum concentration, and were similar after administration of bovine and human exogenous TSH, and TRH. Conclusions regarding preferential secretion ought to be made by comparing ratios of thyroid hormone secretion with those of the hormone content of the gland, but our results indicate that TSH induces preferential secretion of triiodothyronines in man.

Adult↗
PubMed1982

Isolation of rat pituitary prolactin isohormones differing in charge, size, and specific immunological activity.

A method has been described for the isolation of three differently charged isohormones of rat prolactin from a discard fraction obtained after extraction of gonadotropins, thyrotropin and growth hormone from homogenized frozen pituitaries. The procedure involved extraction at pH 9.8, ammonium sulphate fractionation, molecular sieve chromatography on Sephadex G-100, and column electrophoresis in agarose suspension. The purification was monitored by radioimmunoassays and the recovered components were all found to possess a specific immunoactivity exceeding that of the standard preparation (RP-1) supplied by the NIAMDD, Bethesda, U.S.A. Increased acidity among these isohormones was found to be paralleled by significantly decreased immunopotency. Each component showed biological activity in radioreceptor assay. A high degree of purity of the isolated components was shown by analytical electrophoresis in polyacrylamide gel. Sodium dodecyl sulphate electrophoresis in the same medium showed no size heterogeneity and yielded a value of approximately 25 000 for the molecular weight of the isohormones. In addition a large form of prolactin, suggested to represent a dimer, was isolated by a further extraction step (pH 10.5) followed by molecular sieve chromatography on Sephadex G-100 and electrophoresis in agarose suspension. The large form was associated with both biopotency and immunopotency. The electrophoresis resolved the prolactin activity into three or four immunoactive components. This pleomorphism of the large prolactin was confirmed by analytical polyacrylamide gel electrophoresis. Amino acid analyses revealed a close similarity between the three monomers and the major dimeric form of the hormone.

Amino Acids↗
PubMed1982

Comparison between the progestin secretion responsiveness to gonadotrophins of rat cumulus and mural granulosa cells in vitro.

Several studies have shown differences in gonadotrophin receptor content between different granulosa cell (gc) populations within the ovarian follicle, but little is known about the gonadotrophin sensitivity in gc from different parts of the follicle. This study is an investigation of progestin synthesis and responsiveness to highly purified human and partly purified rat gonadotrophins. It involves short-term culture of rat cumulus and mural gc obtained from preovulatory follicles of PMSG-treated immature rats. The responsiveness to FSH in terms of progestin accumulation was similar in the two gc populations, whereas the responsiveness to LH was greater in the mural gc than in the cumulus. The morphological response in the cumulus cells (mucification) correlated with the steroidogenic response. The pattern of progestin synthesis differed between the two gc populations. In the cumulus gc progesterone was dominant, whereas 20 alpha-dihydroprogesterone was the main progestin secreted in the mural gc. The difference in responsiveness to gonadotrophins correlates well with the reported differences in receptor content in the two gc populations.

20-alpha-Dihydroprogesterone↗
PubMed1981

Rat prolactin and hypothalamic catecholamine nerve terminal systems. Evidence for rapid and discrete increases in dopamine and noradrenaline turnover in the hypophysectomized male rat.

Rat prolactin produces in a dose of 100 micrograms/kg a rapid and marked increase of dopamine turnover in the medial palisade zone of the median eminence and of noradrenaline turnover in the posterior periventricular hypothalamic region as well as a possible minor increase of noradrenaline turnover in the magnocellular part of the paraventricular hypothalamic nucleus of the hypophysectomized male rat. The rat prolactin serum levels measured 2 h following injection were within the upper physiological range of lactating rats. The findings suggest that dopamine in the medial palisade zone may be released as a prolactin inhibitory factor and respond rapidly to increases in serum prolactin levels. Furthermore, rat prolactin can acutely modify noradrenergic mechanisms in discrete areas of the hypothalamus indicating that prolactin can participate in the regulation of types of hypothalamic functions other than regulation of its own secretion.

Animals↗
PubMed1981

Human thyroid blood flow response to endogenous, exogenous human, and bovine thyrotrophin measured by electromagnetic flowmetry.

Human thyroid blood flow (TBF) was studied with electromagnetic flowmetry during operation. Measurements were made of the effect on TBF of injections of bovine TSH into one inferior thyroid artery in 6 patients; 6 other patients were given human TSH, and in 10 patients measurements were made both of TBF and endogenous TSH released after administration of TRH in a peripheral vein. The TBF increased after all three types of injection. The mean of the TBF maxima after bovine TSH was 2.26 +/- 0.35 (mean +/- SEM) relative to basal TBF, after human TSH 1.97 +/- 0.28, and after TRH 1.64 +/- 0.20. In the three groups combined it was 1.92 +/- 0.16. The TBF was often increased already during the first recording period 1-10 min after TSH or TRH administration. The mean TBF was approximately doubled at 30-50 min. There were considerable inter-individual variations in the latent time and maximum response of TBF, especially after human TSH, but we found no correlation between the response and the TSH serum concentration in any group. A prompt, but inter-individually varying, increase in TBF was confirmed. This increase is suggested to be secondary to an increased intrafollicular metabolic activity and not primarily regulating thyroid function.

Adult↗
PubMed1980

Human pituitary prolactin: isolation and characterization of three isohormones with different bioassay and radioimmunoassay activities.

Three forms of human pituitary prolactin, separable at alkaline pH in a highly purified preparation, were isolated by means of column electrophoresis in agarose suspension. The most acidic component showed a significantly lower radioimmunological activity but a higher bioactivity than the other two components, which were approximately equipotent in both assays. Consequently, in both assays the most acidic component differed markedly from the other components. Amino acid analysis indicated close similarity between the three components and no size heterogeneity was observed by sodium dodecyl sulphate electrophoresis in polyacrylamide gel. A high degree of purity of the isolated components was demonstrated by analytical electrophoresis in polyacrylamide gel at alkaline pH. Runs in the same medium also revealed that the difference in electrophoretic migration velocity remained after reduction and alkylation of the isohormones. A comparison of the migrations of the S-carboxymethylated and the S-carbamidomethylated derivatives with those of the unmodified components indicated that the diversity in electrophoretical behaviour of two adjacent isohormones was consistent with a difference in one single net charge. By analytical isoelectric focusing the component of intermediate migration velocity was resolved into two distinct bands proposed to contain isohormones differing only in the exchange of one residue of aspartic acid for one of glutamic acid. The isoelectric points of the prolactin isomers were all in the range of 5.7--5.9.

Amino Acids↗