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Biomedical subjects

J Zapf

Publications and source records attributed to J Zapf.

At least 199 records · Page 11Linked to original sources
PubMed1979

The effect of different extraction procedures on two different molecular weight species of serum NSILA and on the carrier protein of small molecular weight NSILA (NSILA-S).

The influence of Dowex-50 adsorption chromatography on the recovery of two different forms of serum NSILA, large and small mol. wt. NSILA, and on the recovery of the binding protein of the small mol. wt. form was studied and compared with another extraction procedure, gel filtration on Sephadex G-50 in 1 M acetic acid. Partially purified NSILA-S is adsorbed to Dowex-50 at pH 6.8. It can be eluted with 20 mM NH4OH and appears unchanged with regard to its biological activity and molecular weight. Adsorption of 125I-labelled NSILA-S to Dowex-50 does not change its binding characteristics to serum. When serum is chromatographed on Sephedex G-50 in 1 M acetic acid, NSILA is obtained in a large and in a small molecular weight form (NSILA-S). After recombination of the small molecular weight NSILA fraction with the "stripped" serum fraction, which contains large mol. wt. NSILA and a specific carrier protein for NSILA-S, re-chromatography of this mixutre on Sephadex G-50 at neutral pH yields NSILA mostly in the void volume. It adsorbs to Dowex-50. After elution from Dowex, acidic gel filtration on Sephadex G-50 results in an elution pattern which is completely different from that of NSILA-S. Adsorption of serum to Dowex-50 results in a dramatic decrease of the NSILA-S binding activity. It is concluded that Dowex-50 adsorption chromatography of serum 1) inactivates most of the serum NSILA-S binding protein 2) leads to the loss of acid dissociable small mol. wt. NSILA (NSILA-S). Therefore, Dowex-50 adsorption chromatography is not suitable for the subsequent determination or further purification of NSILA-S from whole serum.

Adsorption↗
PubMed1978

NSILA-carrier protein abolishes the action of nonsuppressible insulin-like activity (NSILA-S) on perfused rat heart.

Human serum in a concentration of 10% in the perfusion medium failed to increase glucose uptake by the isolated perfused rat heart, indicating that nonsuppressible insulin-like activity (NSILA) in whole serum was inactive in this system. When NSILA-carrier protein was added to partially purified NSILA-S, its biological activity on the rat heart disappeared. In contrast, the action of insulin was not affected by the presence of NSILA-carrier protein. Binding of 125I-labelled NSILA-S to rat heart was inhibited by NSILA-carrier protein. 125I-labelled insulin binding was not inhibited. These results support the hypothesis that NSILA-S bound to serum carrier protein is a large molecular compound which does not readily diffuse out of the capillary bed and therefore does not exert insulin-like effects in vivo.

Animals↗
PubMed1977

Effects of insulin and NSILA on adipocytes of normal and diabetic rats: receptor binding, glucose transport and glucose metabolism.

Isolated fat cells from normal and streptozotocin-diabetic rats were compared with respect to metabolic indices (glucose-uptake, 3-0-methyl-glucose efflux) with and without stimulation by insulin and nonsuppressible insulin-like activity (NSI-LA). In addition, binding studies were carried out with these two hormones. Basal 14C-glucose oxidation and incorporation into lipids was decreased in diabetic cells and their response to insulin and NSILA was greatly reduced. Basal efflux of 3-0-methylglucose from diabetic cells was somewhat faster than from normal cells. The response to insulin and NSILA was less than in normal cells and it was delayed. The apparent number of insulin binding sites as well as their affinity for insulin was increased in diabetic cells. In contrast, the apparent number of binding sites for NSILA was decreased in diabetic cells and their affinity for NSILA was increased. In normal cells insulin enhanced binding of 125I-NSILA more markedly than in diabetic cells. These findings show that the rate-limiting step of impaired glucose metabolism (oxidation and lipogenesis) in diabetic fat cells is beyond the interaction of the hormone with the receptor. They suggest that the apparent number of hormone receptors (insulin, NSILA) on the cell membrane is regulated individually for each binding site.

Adipose Tissue↗
PubMed1977

Demonstration of a specific serum carrier protein of nonsuppressible insulin-like activity in vivo.

The fate of nonsuppressible insulin-like activity (NSILA-S) was studied by injecting a tracer of 125I-NSILA-S iv into rats. Ten minutes after injection of 125I-NSILA-S alone, 20% of the label is found in serum, whereas after the injection of 125-I-insulin or 125I-NSILA-S together with an excess of cold NSILA-S only 8% of the label are recovered. Sephadex G-200 chromatography at neutral pH of serum after injection of 125I-NSILA-S reveals 2 peaks of radioactivity in the high molecular weight region at 67 and 47% bed volume. Five minutes after injection the peak at 67% starts to disappear, whereas the one at 47% persists with a half-life of 3 h. The latter peak was submitted to Sephadex G-200 chromatography at acidic pH which dissociates NSILA-S from its binding protein. The labeled material obtained by this procedure still exhibits the same binding characteristics to chick embryo fibroblasts as standard 125 I-NSILA-S. A third peak at 90% bed volume corresponding to low molecular NSILA-S is no longer detectable 20 min after injection. A fourth peak at 100% bed volume becomes apparent after 1 h. The half-life and chromatographic pattern of iv injected 125 I-NSILA-S are not changed by the simultaneous administration of insulin or growth hormone. These findings confirm the existence of a specific serum carrier protein for NSILA-S and may explain why endogenous NSILA-S does not exert insulin-like effects under physiological conditions in vivo.

Animals↗
PubMed1977

Non-suppressible insulin-like activity during acute metabolic and endocrine changes in dogs.

Non-suppressible insulin-like activity (NSILA-S) was determined in dogs after acute changes in the blood sugar and after injection of growth hormone. Serum was chromatographed over Sephadex G-50 columns equilibrated in 1 M acetic acid and NSILA-S was determined in the fractions between 55 and 85% column volume using a protein binding assay and the conventional fat pad assay. The results obtained with these two methods correlated rather well (r = 0.74). Hyperglycaemia induced by an intravenous glucose load, by intravenous administration of mannoheptulose or both was not followed by an increase in NSILA-S levels. The injection of insulin and human growth hormone did not lead to alterations of the NSILA-S levels. It is concluded that total NSILA-S levels in the dog do not change acutely following manipulations of the blood sugar and that, in all likelihood, NSILA-S plays no role in the regulation of blood glucose.

Animals↗
PubMed1977

Decrease of non-suppressible insulin-like activity after pancreatectomy and normalization by insulin therapy.

Non-suppressible insulin-like activity (NSILA-S) was determined in 5 dogs before and after pancreatectomy and again during insulin therapy. All NSILA-S determinations were carried out on serum samples which were passed over Sephadex G-50 columns equilibrated with 1 M acetic acid. The levels of NSILA-S decreased drastically shortly after pancreatectomy and rose slowly after institution of insulin therapy, to normal levels. During the period of severe diabetes after pancreatectomy the concentration of growth hormone was elevated. These findings indicate that 1) the pancreas cannot be the site of synthesis and release of NSILA-S, 2) NSILA-S levels do not always parallel growth hormone levels and 3) the synthesis and secretion of NSILA-S among other factors is under the control of insulin.

Animals↗
PubMed1977

The influence of hypophysectomy on NSILA concentrations in the dog: evidence for partially pituitary-independent regulation.

Non-suppressible insulin-like activity (NSILA) was determined in 5 dogs before and after hypophysectomy. All NSILA determinations were carried out on serum samples after acidic Sephadex G-50 chromatography by two different assay systems, i.e. a bioassay and a protein binding assay. The levels of NSILA decreased significantly after hypophysectomy and returned to near normal levels after 2 weeks. T3-, T4- and cortisol levels were drastically reduced during the entire period of the experiment. Several GH determinations after hypophysectomy revealed very low levels. Insulin-induced hypoglycaemia failed to provoke a rise of GH levels as late as 4 months after hypophysectomy. These findings indicate that: 1) The pituitary gland cannot be the site of synthesis of NSILA. 2) NSILA concentrations in the dog are maintained at a near normal level in the presence of very low growth hormone and thyroid hormone concentrations, so that these latter hormones do not appear to be the only regulatory factors concerned in NSILA synthesis.

Animals↗
PubMed1977

Determination of nonsuppressible insulin-like activity in human serum by a sensitive protein-binding assay.

We describe a sensitive protein-binding assay for non-suppressible insulin-like activity in human serum. It can detect as little as 0.2 microunits (corresponding to 0.5 ng) of the activity in 0.4 ml of the assay mixture. It is measured in a low-molecular-weight fraction (termed "biological material") obtained by chromatography of serum on Sephadex G-50 in 1 mol/liter acetic acid. This fraction has been shown earlier to contain nearly all this billogically active material that is present in serum. A partially purified carrier protein from human serum is used as the binding protein; different concentrations of a partially purified preparation of material with the activity serve as standards, which compete with 125l-labeled tracer for binding. Biological material dilutes more or less in parallel with the standard over a 10-fold concentration range. In the chromatographed serum fractions, displacing activity appears between 50 and 80% bed volume, with the peak at 60%, and coincides with the distribution and the peak of radioactivity obtained by chromatography of tracer. A good correlation (r = 0.88) is observed between the values determined for this activity in the rat fat-pad assay and the protein-binding assay, although the latter yields about two fold higher results (190 +/- 37 milliunits/liter vs. 345+- 65 milliunits/liter, mean values for 18 normal sera). Values determined in the protein-binding assay are decreased in hypopituitary patients (183 +/- 27 milliunits/liter) and increased in acromegalics (486 +/- 88 milliunits/liter), in accord with the results of the bioassay (68 +/- 21 milliunits/liter for hypopituitary patients, 293 +/- 53 for acromegalics).

Adolescent↗
PubMed1976

[Malignant hyperthermia: attempt at an early diagnosis by means of determination of creatine phosphokinase (CPK) and its isoenzymes].

Creatinphosphokinase (CPK) was elevated in sera of patients with malignant hyperthermia and in sera of some of their relatives. Only the MM-isoenzyme (but not the MB- or the BB-isoenzyme) could be detected by paperchromatographic analysis. In some of the patients elevation of muscle aldolase was also observed. Thus, the appearance of the BB-isoenzyme in sera of patients with malignant hyperthermia, as described by another group of investigators, was not confirmed. No specific screening method exists as yet to detect patients with a high risk of developing this often lethal reaction to anesthesia. However, in patients without muscle disease or trauma and without prior i.m. injections, myocardial infarction or major physical strain, elevation of CPK in serum should be interpreted as meaning that malignant hyperthermia may develop during anesthesia. The pathophysiology of malignant hyperthermia is discussed.

Creatine Kinase↗
PubMed1976

Nonsuppressible insulin-like activity and thyroid hormones: major pituitary-dependent sulfation factors for chick embryo cartilage.

Serum from hypothyroid hypophysectomized rats did not stimulate sulfation or incorporation of amino acids into chick embryo sterna. When such rats were treated for a short time with growth hormone (somatotropin), their serum stimulated incorporation both of sulfate and of amino acids. The different actions of the two types of sera were not due to changes in thyroid state. The results support the existence in serum of a sulfation factor for chick embryo cartilage that is dependent upon growth hormone. Highly purified preparations of nonsuppressible insulin-like activity from human serum stimulated incorporation of amino acids, and of uridine into RNA, in chick embryo sterna in vitro; chondrocytes prepared from this tissue had specific high-affinity binding sites for this insulin-like activity. However, sulfate incorporation was stimulated very little, unless serum from hypothyroid hypophysectomized rats was also present. When L-3,5,3'-triiodothyronine was added as well, the stimulation was enhanced further. From these and other experiments, we conclude that (i) nonsuppressible insulin-like activity or a closely related peptide is the growth-hormone-dependent growth and sulfation factor for chick embryo cartilage: (ii) a second, unidentified factor must be present for the insulin-like activity to stimulate sulfation; and (iii) stimulation of sulfation by thyroid hormones in vitro is additive to that of nonsuppressible insulin-like activity.

Animals↗
PubMed1976

Non-suppressible insulin-like activity and sulphaton activity in serum extracts of normal subjects, acromegalics and pituitary dwarfs.

Non-suppressible insulin-like activity (NSILA) is a term used for a variety of substances in serum, excluding insulin, which promote glucose uptake of adipose tissue and diaphragm in vitro. NSILA-S is a peptide with a molecular weight of 7000 which is soluble in acid ethanol and which has been purified on a large scale from human serum. This study describes a simple chromatographic one step procedure by which NSILA-S can be extracted and quantitatively measured in individual sera. Using Sephadex G-75 equilibrated with 1 M acetic acid, NSILA-S was detected only in one peak containing small molecular peptides. NSILA-S obtained with this one step chromatographic procedure exerted all the effects of purified NSILA-S including sulphation activity on the rat cartilage. All chromatographic fractions with NSILA-S also had sulphation activity. Both, NSILA-S and sulphation activity were increased in acromegalics and decreased in pituitary dwarfs. It is suggested that one molecule in serum is responsible for both activities which are, at least in part, under the control of growth hormone.

Acromegaly↗