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Biomedical subjects

E Cerasi

Publications and source records attributed to E Cerasi.

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

Calcium-binding proteins and insulin release. Differential effects of phenothiazines on first- and second-phase secretion and on islet cAMP response to glucose.

Calcium and cAMP are interdependent regulators of glucose-induced insulin release. In the present study we investigated the importance of cAMP and calcium-binding proteins for biphasic insulin secretion by assessing the effects of two phenothiazines known to block such proteins, trifluoroperazine (TFP) and promethazine (PMZ). In isolated rat islets, during 60-min incubations with 16.7 mmol/l glucose both agents inhibited the insulin response with ID50 values of 15 mumol/l for TFP and 5 mumol/l for PMZ. Both agents decreased the maximal insulin response without gross changes in the islet sensitivity to glucose. TFP (15 mumol/l), whereas inducing 50% inhibition of second-phase insulin release, totally suppressed the cAMP response to glucose and the accompanying first-phase insulin secretion (5-min incubations); these effects of TFP could be partially reversed by isobutyl methylxanthine (IBMX). In contrast, 5 mumol/l PMZ, which produced 60% inhibition of second-phase insulin release, had no effect on first-phase insulin and cAMP responses to glucose. Furthermore, IBMX did not modify the inhibitory effect of PMZ on second-phase insulin secretion. The following is concluded: 1. TFP acts preferentially on first-phase insulin release and inhibits cAMP formation; this suggests that calmodulin plays a major role in mediating the initial glucose effect on secretion via stimulation of cAMP. 2. The islet probably contains calcium-sensitive proteins other than calmodulin, since the low concentrations of PMZ shown to inhibit second-phase insulin release lack effects on calmodulin. Synexin could be such a protein.(ABSTRACT TRUNCATED AT 250 WORDS)

1-Methyl-3-isobutylxanthine↗
PubMed1986

Substrate regulation of the glucose transport system in rat skeletal muscle. Characterization and kinetic analysis in isolated soleus muscle and skeletal muscle cells in culture.

A self-regulatory mechanism of the glucose transport in rat skeletal muscle cells is described. In isolated rat soleus muscles and rat skeletal myocytes and myotubes in culture, pre-exposure to varying glucose concentrations modulated the rate of 2-deoxyglucose uptake. Maximal uptake was observed at glucose concentrations below 3 mM. Between 2.5 and 4.0 mM glucose it was reduced by 25-35%; further elevation of the glucose concentration resulted in a gradual decrease of the transport rate by approximately 2% for each millimolar glucose. The effect of glucose was time-dependent and fully reversible. Insulin rapidly increased the 2-deoxyglucose uptake in the soleus muscle; however, the insulin effect depended on the glucose concentration of the preincubation. Insulin was totally ineffective in muscles pre-exposed to 1.0-3.0 mM glucose, whereas its stimulatory action increased with increasing glucose concentrations above 4 mM. The effect of low glucose and insulin were not additive, and the maximal 2-deoxyglucose uptake rates induced by both conditions were of identical magnitude. It is postulated that glucose may "up- and down-regulate" its transport by affecting the number of active glucose transporters in the plasma membrane, and that insulin exerts its stimulatory effect only when the extracellular glucose reaches a threshold concentration.

Aminoisobutyric Acids↗
PubMed1986

Induction of cell surface insulin antigenicity in fibroblasts transfected with islet genomic DNA.

Ltk- cells were transfected with total genomic DNA obtained from rat islets or from insulinoma cells mixed with DNA tagged with fluorescent dye. Fluorescence-activated cell sorter (FACS) was used for initial selection of successful transfectants, monitoring fluorescent DNA incorporated into the cell. For subsequent selections the cells were treated with anti-insulin antiserum labeled with fluorescent dye and selected by FACS. Beta-cell DNA, but not DNA from murine leukemic cells, induced the appearance of cell surface insulin antigenicity in fibroblasts. This phenotypic expression was transient. Together with our previous demonstration of the induction of insulin secretion in Ltk- cells [(1986) Biochem. Biophys. Res. Commun. 136, 638-644], these results indicate that beta-cell-specific characteristics can be transfected to non-endocrine cells by genomic DNA transfection.

Animals↗
PubMed1986

Induction of insulin release in fibroblasts transfected with genomic rat islet DNA.

L cells (tk-) were cotransfected with total genomic rat islet DNA and a plasmid containing thymidine kinase gene (ptk). Transfectants were tested for their ability to release insulin into the medium. At least 10% of the colonies contained immunoreactive insulin (IRI) during the initial two weeks. The insulin secreted competed linearly with rat insulin in RIA, the majority of the insulin antigenicity comigrating with rat insulin on G-50 Sephadex chromatography. With continuing propagation the IRI activity diminished; however 3 selected cultures demonstrated increased secretion of IRI following stimulation with glucose. These findings indicate that glucose-induced insulin secretion can be obtained in non-beta cells; however the frequency of success was below one stable transfection for every 5 X 10(8) Ltk- cells exposed to the transfection procedure.

Animals↗
PubMed1985

Correction of diabetic pattern of insulin release from islets of the spiny mouse (Acomys cahirinus) by glucose priming in vitro.

Insulin release kinetics were studied in perifused islets of Langerhans, isolated from mildly hyperglycaemic and from normoglycaemic spiny mice (Acomys cahirinus), a rodent predisposed to develop spontaneously non-ketotic diabetes. In both groups, insulin response to glucose (16.7 mmol/l) was delayed in comparison with that of rat islets, the release kinetics being analogous to that of human Type 2 (non-insulin-dependent) diabetes. Thirty min priming of the isolated Acomys islets with glucose (16.7 mmol/l) resulted in potentiation of the insulin release to a second stimulation. The degree of potentiation decreased exponentially with the time interval between stimulations, showing a t1/2 of 18 min. Induction of potentiation by glucose was time-dependent, giving a maximal effect after 20 min of priming. In addition to overall amplification of the insulin response, priming with glucose accelerated markedly the initial release rates, correcting the dynamics of the response. We conclude that: (1) decreased and delayed insulin secretion is found in Acomys cahirinus before the development of hyperglycaemia; (2) induction of time-dependent potentiation in the islet by priming with glucose corrects the diabetic-type dynamics of insulin release; (3) therefore the deficient insulin release of Acomys is of a functional nature, the mechanism of potentiation bypassing the defect; (4) since insulin release in Acomys resembles that in prediabetic and diabetic man, similar conclusions might apply to the islet dysfunction in Type 2 diabetes.

Animals↗
PubMed1985

Genetic regulation of the kinetics of glucose-induced insulin release in man. Studies in families with diabetic and non-diabetic probands.

Insulin release and sensitivity were estimated from glucose and insulin curves obtained at a glucose infusion test performed on altogether 601 subjects belonging to 155 nuclear families. Ascertainment was through one of the parents, and 96 of the probands had diabetes with clinical onset after the age of 30 years, while 59 were healthy subjects. Three variables obtained by a computer model were analysed, i.e. the glucose regulation of insulin release by a direct stimulatory event (KI) and time-dependent modulatory events (KP) as well as insulin sensitivity (KG). Complex segregation analysis revealed that the variables are genetically regulated, but there was no evidence for a major locus. The children of the diabetics did not differ from those of the non-diabetics as far as insulin release is concerned.

Age Factors↗
PubMed1985

Binding, degradation, and biological activity of insulin in vascular smooth muscle cells.

The interaction of insulin with the vascular smooth muscle was studied using cultures derived from the bovine aortic arch. The cultured cells exhibited specific binding of 125I-insulin that was reversible and dependent on pH. Both insulin and insulinlike growth factor (IGF) I competed for 125I-insulin binding; IGF I, however, was less effective than insulin by at least an order of magnitude. Insulin binding was accompanied by internalization and degradation of the hormone in a temperature- and time-dependent manner. Chloroquine and other lysosomotropic agents elevated the internalized insulin and reduced its degradation. Pre-exposure of cell cultures to insulin resulted in downregulation of cell surface receptors. Insulin stimulated alpha-aminoisobutyric acid transport in confluent smooth muscle cells. The maximal response was observed at 100 ng/ml insulin with a half-maximal effect at 10 ng/ml. Sparse, serum-starved smooth muscle cells responded to insulin with a dose-dependent increase in [3H]-thymidine incorporation into DNA. Although the effect was already apparent at 1 ng/ml insulin, it reached near maximal level only at 10,000 ng/ml. IGF I also stimulated DNA synthesis in smooth muscle cells; however, at low concentrations insulin was more efficient in this respect. Human growth hormone was inactive. The data indicate the presence of specific receptors for insulin in bovine aortic smooth muscle cells. These receptors appear to mediate the metabolic activity as well as part of the mitogenic effect of insulin in these cells.

Aminoisobutyric Acids↗
PubMed1985

Relative potencies of human and pork insulin during i.v. and oral glucose tolerance test in lean and obese persons.

The biological efficacy of biosynthetic human insulin (BHI) of recombinant DNA origin was compared with that of highly purified pork insulin in subjects with normal (lean persons) and decreased (obese persons) sensitivity to insulin. Sequential i.v. administration of glucose with increasing insulin doses under somatostatin coverage was used to establish the dose-response characteristics of insulin-induced glucose clearance [insulin vs. Kg (glucose disappearance rate, %/min). Insulin was also given s.c. prior to an oral glucose tolerance test (OGTT) performed under somatostatin blockade; the appearance of insulin in serum and the modification of glucose levels were followed. The insulin vs. Kg dose-response curve was flatter and shifted to the right in obese persons compared with lean persons, demonstrating insulin resistance. In both lean and obese subjects, BHI and pork insulin had comparable effects on glucose clearance. Similarly, the effect of s.c. insulin on the serum glucose values during OGTT was the same when BHI or pork insulin were used. The absorption characteristics from the injection site were similar for both insulins. Thus, in man, human and pork insulin have similar pharmacokinetic and biological characteristics.

Administration, Oral↗
PubMed1984

Time-dependent inhibition of insulin release: suppression of the arginine effect by hyperglycaemia.

Brief stimulation of the pancreas with arginine causes a refractory state which reduces the insulin response to subsequent stimulations (time-dependent inhibition). In control subjects, a pair of arginine injections (75 mg/kg) at a 30-min interval resulted in 20% reduction of peak and integrated insulin responses to the second injection. In Type 2 (non-insulin-dependent) diabetic patients and in obese subjects, the inhibitory effect of repeated arginine stimuli was abolished. Healthy subjects were made acutely hyperglycaemic (9.3 +/- 0.3 mmol/l) by the glucose clamp technique. This induced a three- to fivefold greater insulin response to arginine. Compared to the response of diabetic subjects with similar hyperglycaemia, the control subjects secreted four to nine times more insulin. When the arginine stimulation was repeated 30 min later, no inhibition was observed, the second insulin response being instead augmented 1.5- to 1.8-fold. We conclude that (1) the insulin response to arginine is markedly reduced in Type 2 diabetes; (2) arginine-induced time-dependent inhibition of insulin release is abolished in patients with minimal to moderate hyperglycaemia; (3) this is probably due to the acute synergistic action of glucose and arginine on the B cell; (4) time-dependent inhibition of insulin release may be a protective mechanism against insulin oversecretion following repetitive stimulation of the pancreas; its abolition in hyperglycaemic states may be a compensatory mechanism, allowing substantial increases in insulin output.

Adult↗
PubMed1984

Time-dependent inhibition of insulin release: glucose-arginine interactions in the perfused rat pancreas.

The isolated perfused rat pancreas was stimulated sequentially with arginine or glucose to analyze the time-dependent modulation of insulin release. A 10-min perfusion with arginine (5.0 mmol/l) induced 75% inhibition of the insulin response to repeated arginine stimulation 10 min later. When glucose (8.3 mmol/l) was given as two pulses, inhibition of the second insulin response was less pronounced. The inhibitory effect generated by arginine also suppressed the insulin response to glucose (27.7 mmol/l), and this inhibitory effect persisted for over 80 min. Stimulation for 30 min with glucose (27.7 mmol/l) strongly potentiated the insulin responses to a pair of arginine stimuli given 20 min later. However, despite augmented secretion rates, the insulin response to the second arginine pulse was still inhibited by 75%. When insulin secretion was strongly amplified by two 10 min pulses of the synergistic mixture of arginine (5.0 mmol/l) and glucose (8.3 mmol/l), there was no inhibition of the second insulin response. If glucose (8.3 mmol/l) was present during the first arginine stimulation only, the response to the second arginine pulse was inhibited as in control experiments. However, when glucose was added to the second arginine pulse only, the inhibition generated by the first arginine pulse did not express itself, insulin release remaining similar to control.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1983

Differential control of insulin secretion and somatostatin-receptor recruitment in isolated pancreatic islets.

Somatostatin receptors appear to be localized to secretory granules in pancreatic islet homogenates. Recruitment of these receptors to the islet-cell surfaces may mark the contact event between secretory granules and plasma membranes before release of insulin by fission. Isethionate, an impermeant anionic replacement for chloride, blocks the release step but does not affect receptor recruitment. By contrast, low concentrations of phenothiazine drugs, such as trifluoperazine and promethazine, inhibit both receptor recruitment and secretion. Scatchard analysis of phenothiazine effects on somatostatin receptors reveals that these drugs reduce the number of receptors but do not affect the affinity of the receptor for somatostatin. These data indicate that membrane contact and fission steps during exocytosis can be biochemically separated.

Animals↗
PubMed1983

Insulin binding and degradation in vascular endothelial cells: modulation by cell growth and culture organization.

The interaction of insulin with the vascular endothelium and its modulation by cell growth and culture organization was studied using bovine aortic endothelial cells in monolayer cultures. Three types of cultures were investigated: 1) confluent nondividing cultures, organized and differentiated as the in vivo tissue; 2) subconfluent, not yet organized cell cultures, representing proliferating endothelium; and 3) endothelial cell cultures modified to lose their property of contact inhibition, growing in multiple layers. All three types of cultures exhibited specific binding of 125I-insulin to high and low affinity cell surface receptor sites, and were capable of degrading 125I-insulin. Preexposure of the cultures to insulin resulted in a time dependent reduction in the availability of cell surface receptors (down-regulation). Insulin binding per cell was 2.4-fold and 10-fold higher in the subconfluent and modified cultures, respectively, as compared to the contact-inhibited confluent cultures. Similarly, the rate of insulin degradation was higher in the subconfluent and modified cultures (2.3-fold and 20-fold, respectively). Subconfluent cultures were more sensitive than confluent cultures to the down-regulatory effect of insulin. They exhibited a 60% decrease in insulin binding as compared to a 40% decrease in confluent cultures after preexposure to 50 ng/ml insulin. The increase in insulin binding and degradation in growing endothelial cells suggests a role for the hormone in the regulation of endothelial cell growth, e.g. in response to injury. This was further supported by the observation of a dose-dependent stimulation of [3H]thymidine incorporation into sparse, serum-starved endothelial cells by physiological concentrations of the hormone.

Animals↗
PubMed1982

Evidence for an autosomal recessive gene regulating the persistence of the insulin response to glucose in man.

The significance of genetic factors for insulin release after glucose infusion was studied in 155 nuclear families of which 59 were control families and 96 had been ascertained through a parent with onset of diabetes after 30 years of age. Fasting insulin and glucose as well as three principal components of the insulin and glucose curves were submitted to path analysis and complex segregation analysis. The three principal components were considered to reflect the magnitude, the degree of response and the persistence of the curves. The genetic heritability of the insulin variables varied between 0.47-0.93 and that of the glucose variables between 0.20-0.54. There were considerable intergenerational differences in the genetic heritability for the persistence of the glucose curve and for the degree of response and persistence of the insulin curve. The cultural heritability was found to be of minor importance, while the non-transmitted sibling environment was large. There was significant evidence for a major locus for the persistence of the insulin curve. The best fit was for a completely recessive autosomal gene with the gene frequency 0.21. The phenotype distribution of this variable showed significant kurtosis which could simulate a major locus. However, the significant evidence for such a locus remained after an analysis using partial quantitation. The diabetics were significantly different from the non-diabetics for all the variables studied, but a complete discrimination between the diabetics and non-diabetics could not be obtained. There was no significant difference between the children of the diabetics and non-diabetics for any of the variables studied.

Adult↗
PubMed1982

Familial hyperthyroidism due to inappropriate thyrotropin secretion successfully treated with triiodothyronine.

A family of three generations is described in which six females had hyperthyroidism secondary to chronic overstimulation of the thyroid by pituitary TSH. In the untreated state, their basal levels of T4 ranged between 14-22 microgram/dl, T3 levels ranged from 205-300 ng/dl, T3 resin uptake ranged from 43-61%, TSH ranged from 5-26 microU/ml, and PRL ranged from 33-75 ng/ml. Basal metabolic rate (BMR) was elevated in all patients (+32 to +100%). There was no evidence of pituitary tumor, In spite of elevated circulating thyroid hormones, TRH stimulated TSH and PRL to 25-57 microU/ml and 120-300 ng/ml, respectively. Serum TSH could be suppressed to normal after 1 week of T3 administration (25 microgram three times per day). Concomitantly, serum T3 and T4 levels fell, and the TSH response to TRH became normal. In contrast, T4 (200 microgram/day) administered for 1 and 4 weeks, respectively, to two patients did not suppress the pituitary-thyroidal axis. A long term therapeutic trial was performed in three patients with T3 and a single morning dose of 25-50 microgram. TSH gradually returned to normal, as did thyroid hormone levels and the BMR. The clinical manifestations of hyperthyroidism regressed, and complete remission was achieved after 2-3 months of T3 therapy, which persists to data as long as medication is continued. The inappropriate TSH secretion of our patients appears to be due to partial unresponsiveness of the thyrotroph to thyroid hormone. It is suggested that either the pituitary T4 monodeiodinase is deficient in our patients, resulting in low intracellular T3 levels, or the thyrotroph has reduced sensitivity to T3 and therefore can shut off TSH only when serum T3 is raised to high levels, albeit intermittently.

Adolescent↗