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Query: UMLS:C0011849 (diabetes)
 277,896 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The expression of a novel regenerating (reg) gene has been reported previously in the regenerating islets of a surgical model of diabetes in rats. We exposed collagenase-isolated rat islets for three days to nutrient and non-nutrient growth factors in minimally supplemented RPMI medium (2.7 mmol/l glucose, 2% fetal calf serum), and investigated the relationship between reg gene expression and islet cell replication. RNA was prepared from half of the islets by homogenisation in guanidinium isothiocyanate followed by phenol/chloroform extraction. Northern/dot blot analyses were used to semi-quantify reg mRNA. Islet cell replication was estimated by culturing the remaining islets in radiolabelled thymidine to determine de novo DNA synthesis. Thymidine uptake was stimulated by the following factors: 11 mmol/l glucose (50% increase); 10% amino acids (126% increase); 10% fetal calf serum (39% increase); 100 ng/ml insulin (45% increase); 250 ng/ml growth hormone (65% increase); 1.5 nmol/l aldosterone (29% increase); 2 U/ml platelet derived growth factor (116% increase). The results are expressed as a percentage of the thymidine incorporated into control islets cultured in minimal RPMI (1118 +/- 100 (SD) cpm/microgram protein, n = 15). Increased islet cell replication was paralleled in each case by a clear rise in reg mRNA expression compared to controls. Furthermore, the rank order for reg gene expression was the same as that for thymidine uptake (r = 0.90). The present findings suggest a clear association between reg gene expression and islet cell replication in vitro, and are the first to demonstrate reg gene expression in response to individual growth factors.
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PMID:Expression of an islet regenerating (reg) gene in isolated rat islets: effects of nutrient and non-nutrient growth factors. 137 94

Before clinical onset of insulin-dependent diabetes mellitus a decreasing pancreatic beta-cell mass maintains glucose homeostasis. We currently aimed to study the function of pancreatic islets isolated 2 weeks after a 60% partial pancreatectomy (P) or after a sham operation (S) on adult rats. Experiments on the islets were subsequently performed acutely (day 0) and after 1 week (day 7) of tissue culture in medium RPMI 1640 (11.1 mM glucose) + 10% calf serum. There was no difference in the body weight 2 weeks after surgery. The pancreatic remnant weight of the P rats was 35% less than the pancreatic weight in the S rats. The islet DNA content was 25% higher in the islets of the P rats on day 0, indicating a stimulated islet growth. However, this difference did not remain after culture for 7 days. Islet proinsulin mRNA content and (pro)insulin biosynthesis rates were slightly increased in the islets of P rats on day 0, which could be due to the increased islet mass. The islet insulin content was not different on day 0, but was higher after culture in the islets of the P rats. The islet rates of glucose oxidation and insulin release were markedly higher in the P rats on day 0, suggesting a selective effect on these processes. A higher glucose oxidation rate was, however, not evident on day 7. The relative fraction of insulin-positive cells was slightly lowered in the islets of the P rats on day 0.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Adaptive response in beta-cell function in pancreatic islets isolated from partially pancreatectomized rats. 151 84

We studied the possible relationships between the functional status of the beta-cell and activities or mRNA contents of enzymes involved in the catabolism of glucose. Three different in vitro models with attenuated insulin response were used: rat islets cultured at a low glucose concentration, rat islets incubated in vitro with streptozocin, and fetal rat islets. The fetal and streptozocin-administered islets were compared with adult islets cultured in RPMI-1640 containing 11 mM glucose, and the effects of the in vitro glucose concentrations (3.3, 11, and 28 mM) were assessed on adult islets only. Cellular mRNA levels for the mitochondrial DNA-encoded cytochrome b and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were determined by Northern-blot analysis. Enzymatic activities of high-Km (glucokinase) and low-Km (hexokinase) glucose-phosphorylating enzymes and succinate-cytochrome c reductase were also determined. Islets cultured at 3.3 mM glucose displayed a decreased activity of glucokinase compared with islets cultured at 28 mM glucose (23.3 +/- 12%), whereas there was no difference in hexokinase activity or the level of GAPDH mRNA. The activity of succinate-cytochrome c reductase was similar in islets cultured at the different glucose concentrations. The level of cytochrome b mRNA increased at 28 mM glucose compared with islets cultured at 11 mM glucose (140 +/- 14%). Islets incubated with streptozocin and subsequently cultured for 7 days at 11 mM glucose exhibited a decreased level of cytochrome b mRNA (65 +/- 5%) and no differences in the activities of glucokinase, hexokinase, succinate-cytochrome c reductase, or the level of GAPDH mRNA.(ABSTRACT TRUNCATED AT 250 WORDS)
Diabetes 1991 Jun
PMID:Exhibition of specific alterations in activities and mRNA levels of rat islet glycolytic and mitochondrial enzymes in three different in vitro model systems for attenuated insulin release. 164 83

An improved rapid cell enzyme-linked immunosorbent assay (CELISA) is described which is suitable for the large scale screening of monoclonal antibodies to islet cell surface antigens. 5 x 10(4) insulin-producing rat insulinoma (RIN) cells were seeded per well in a 96-well flat-bottomed polystyrene plate coated one day before a 0.01% poly-D-lysine solution in PBS. After culture for 4 days in 200 microliters/well RPMI 1640 supplemented with 7.5% heat-inactivated fetal calf serum, the cell number per well was up to 2.1 x 10(5). These monolayer RIN cell cultures were used as a target for the detection of islet cell surface antibodies (ICSA) in the supernatants of hybridomas. The cells were used without fixation to avoid modification of sensitive surface antigens. Poly-D-lysine did not cause non-specific binding of immunoglobulins to the plastic wells as tested with irrelevant monoclonals. The specificity and sensitivity of the method is comparable to indirect immunofluorescence. All mc-ICSA primary screened by indirect immunofluorescence using viable RIN cell suspensions were positive in this CELISA. There was a correlation (r = 0.7; n = 44) between the antibody binding measured by CELISA and the indirect immunofluorescence technique. The advantage of this CELISA is that cell surface structures are well preserved in a viable cell monolayer used as target without chemical fixation. This assay procedure should be generally suitable for the initial screening of monoclonal antibodies to cell surface antigens of cells growing under culture conditions.
Diabetes Res 1991 Jan
PMID:CELISA for rapid screening of monoclonal islet cell surface antibodies using living rat insulinoma cells as target. 181 97

The endocrine pancreas secretes insulin in a pulsatile fashion. This rhythm is generated at a site within the pancreas, although its precise location has not been determined. With an in vitro system, we tested the possibility that beta-cells might generate spontaneous pulsatile insulin secretion in the absence of any external influence. Human insulinoma tissue from five patients was perifused for 7-10 h with RPMI-1640 medium and constant concentrations of glucose (5.5 mM). Insulin, C-peptide, and proinsulin were measured in the effluent collected at 3.3-min intervals. All three peptides demonstrated pulsatility of secretion in a similar, synchronous fashion that was sustained throughout each study. The Clifton cycle detection program demonstrated cycling in all five tumors, with an average period for all tumors of 28, 29, and 26 min for insulin, C-peptide, and proinsulin, respectively. Spectral analysis confirmed the regularity and consistency of the hormonal secretory patterns. Mean hormone concentrations secreted by different tumors varied, but insulin and C-peptide were secreted in a nearly 1:1 ratio. This study demonstrates 1) that beta-cells are able to generate spontaneous pulsatile insulin secretory activity, which is independent of innervation or the presence of other islet cells, and 2) proinsulin secretion from the beta-cell also has an inherent pulsatility. The synchrony observed in the cycles of proinsulin and its peptide products confirms their common secretory pathway in the beta-cell. We conclude that the beta-cell may be the originator of insulin cycling.
Diabetes 1991 Nov
PMID:Sustained pulsatile insulin secretion from adenomatous human beta-cells. Synchronous cycling of insulin, C-peptide, and proinsulin. 193 5

We describe the effects of metabolic diabetic factors and sera from diabetic animals and humans on the development of early pre-implantation mouse embryos. Our studies demonstrated that 20 to 24% of control mouse blastocysts failed to develop successfully when grown for 72 h in RPMI medium supplemented with 10% fetal bovine serum. D-glucose in concentrations greater than 3 mg/ml, insulin at concentrations of 0.5 and 1.0 IU/ml, glucagon in concentrations of greater than or equal to 10 micrograms/ml, beta-hydroxybutyrate in concentrations greater than 5 mg/ml, and acetoacetate at concentrations of greater than or equal to 10 micrograms/ml were all embryotoxic, the number of underdeveloped blastocysts rising to over 50%. The combination of these factors in relatively low concentrations was highly embryotoxic, especially when accompanied by hyperglycemia. The addition, to a control medium, of serum from nondiabetic rats (in concentrations of 20%) or of nondiabetic human serum (in concentrations of 50%) did not significantly change the rate of blastocystic development. Serum from streptozotocin-diabetic rats, in the same concentrations, increased the number of undeveloped embryos to 53%. With human diabetic sera the highest embryotoxic effect was found in type I diabetes with and without ketoacidosis. In type II diabetes, embryotoxic effects, although lower, were observed among all types studied [untreated, treated with insulin or with DAONIL (Hoechst, Germany)]. A high correlation was found between the number of undeveloped embryos and the blood concentrations of metabolic diabetic factors: glucose (in type I diabetes), beta-hydroxybutyrate (in type II diabetes untreated or treated with Daonil), acetoacetate (in insulin-treated type II diabetes), and HbA1c (in both insulin-treated and in Daonil-treated type II diabetes). The possible role of diabetic metabolic factors in causing increased risk of spontaneous abortions and infertility among diabetic women is discussed.
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PMID:Embryotoxic effects of diabetes on pre-implantation embryos. 196 45

Cultured cells derived from hamster insulinoma (In-111 R1 cells) were placed in 1.4 M dimethyl sulfoxide (Me2SO)-containing RPMI 1640 at 20 degrees C for 20 min. They were frozen to -40 degrees C at a cooling rate of 1.0 or 0.5 degrees C/min, subsequently to -80 degrees C at 3 degrees C/min with a programmable freezer. After being maintained at -80 degrees C, they were rapidly thawed to 37 degrees C. Thawed cells were washed with 0.75 M sucrose for removal of Me2SO. Recovered cells were cultured in 2 ml of RPMI 1640 with 1.3 mM theophylline under a gas phase of 95% air -5% CO2 at 37 degrees C for 2 days. In both cooling rates, frozen-thawed cells discharged more insulin than the thawed in the absence of theophylline. However, this released insulin level was higher in the cells frozen at a cooling rate of 0.5 degrees C/min than that at 1.0 degrees C/min. Moreover, insulin released from frozen-thawed hamster insulinoma cells increased significantly with the addition of 1.3 mM theophylline. Considering that the higher insulin release level at 11.1 mM glucose alone might indicate cellular damage, it is suggested that the cooling rate of 1 degree C/min may be better for cryopreservation of the dispersed cells under the present protocol for the assessment of the function of insulin release.
Diabetes Res Clin Pract 1991 Feb
PMID:Effect of the cooling rate on insulin release from frozen-thawed In-111 cells. 202 80

Antigen expression corresponding to anti-islet cell surface monoclonal antibodies IC2 and A2B5 was studied. IC2 is a rat-rat hybridoma autoantibody produced from the BB rat; among islet cells, IC2 is beta-cell specific. A2B5 is an anti-ganglioside antibody described as labeling beta-cells. Islets of Langerhans from Lewis rats were isolated and cultured for 18 h in RPMI-1640 with five different glucose concentrations (2.2, 3.3, 5.5, 11.1, and 18.3 mM). In some experiments, islets were precultured for 2 or 3 days. After isolation of islet cells and antibody labeling, the percent of IC2+ beta-cells in the different groups increased from 33.3, 34.5, 40.9, and 57.2 to 58.6% (P less than 10(-6). For A2B5, the percent of labeled islet cells increased from 37.4, 41.8, 46.7, and 53.8 to 56.2% (P less than 10(-4). Thus, increasing glucose concentration leading to higher beta-cell activity implies an increase in antigen expression. Neither A2B5 nor IC2 reacts with insulin, as shown by absorption experiments and immune electron microscopy of binding sites. Electron microscopy of IC2-gold-labeled islet cells substantiated the beta-cell specificity of IC2. In conclusion, expression of the corresponding antigens to IC2 and A2B5 depends on the functional state of the beta-cells; because this has been shown to be an important factor in the development of insulin-dependent diabetes, our findings may be of potential pathogenetic interest.
Diabetes 1990 Jun
PMID:Dependence of antigen expression on functional state of beta-cells. 218 61

Recently it has been postulated that interleukin-1 (IL-1) locally released by infiltrating mononuclear cells may destroy the pancreatic B cells during the development of insulin-dependent diabetes mellitus. Since IL-1 is a potent inducer of interleukin-6 (IL-6) in various cells, it is conceivable that IL-6 is a second mediator of the IL-1 action. In the present study the effects of IL-6 alone or in combination with IL-1 were studied on pancreatic islet function in vitro after tissue culture and compared with the effects observed after exposure to IL-1 only. Rat pancreatic islets were cultured in medium RPMI 1640 + 10% calf serum with or without the addition of human recombinant IL-6 (500-5000 pg/ml) for 48 h. The medium insulin accumulation was increased by 40-50% after culture with 500-2000 pg/ml IL-6, but was similar to the controls at 5000 pg/ml. When islets were cultured for 18 h only, also 5000 pg/ml IL-6 stimulated the medium insulin accumulation. IL-6 did not affect the islet insulin content and the rates of islet (pro)insulin and total protein biosynthesis. It inconsistently decreased the islet DNA content. In short-term experiments after 48-h culture with IL-6, there was a dose-dependent inhibition of the glucose-stimulated insulin release. On the other hand, islets cultured with IL-6 (5000 pg/ml) exhibited an elevated glucose oxidation and oxygen uptake, but a lower ATP content at 16.7 mM glucose and an unaffected glucose utilization and glutamine oxidation compared to the controls. This raises the possibility that IL-6 had induced a condition with an increased energy expenditure, resulting in an enhanced mitochondrial metabolism of glucose. Islets cultured with human recombinant IL-1 beta (25 units/ml) showed a strong inhibition of the insulin accumulation in the culture medium and of glucose-stimulated insulin release and a marked decrease in the islet DNA and insulin content. A combination of IL-1 (25 U/ml) + IL-6 (1000 pg/ml) did not alter the inhibitory action of IL-1 alone. The present findings thus show that IL-6 induces a dissociation between insulin secretion and glucose oxidation in islets in vitro. This has not been observed in islets exposed to IL-1, which suggests that IL-6 does not solely mediate the inhibitory effects of IL-1 on islet function.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Interleukin-6 affects insulin secretion and glucose metabolism of rat pancreatic islets in vitro. 240 46

The ability of the pancreatic beta-cell to repair itself after a cytotoxic injury and reassume its functional activities may be a key issue in affording protection from insulin-dependent diabetes mellitus. The molecular mechanisms behind the functional responses of the beta-cell after cytotoxic damage are still largely unknown. The present study in an attempt to elucidate this issue. Mouse pancreatic islets were isolated with collagenase and, after overnight culture, exposed for 30 min at 37 C to 2.2 mM streptozotocin (SZ) or vehicle alone (controls). The islets were subsequently cultured for 6 days in medium RPMI-1640 plus 10% calf serum. After the culture they were subjected to light microscopical examinations or different functional tests during short term incubations. The SZ-treated islets showed markedly diminished insulin release after stimulation with the beta-cell nutrients glucose and leucine plus glutamine. Compounds known to increase intracellular cAMP [theophylline and (Bu)2-cAMP] were able to partially counteract the SZ-induced reduction of insulin release. Stimulation with arginine could also slightly restore the impaired insulin release. Glucose-stimulated oxygen uptake, proinsulin biosynthesis, and insulin and insulin mRNA contents were also decreased, with values at about 50% of the controls. However, the cellular contents of DNA and RNA and total protein biosynthesis rates were essentially normal. Besides mild degranulation in some islets, the morphological appearance of the SZ-treated islets did not reveal any obvious differences compared to the control islets. The present observations suggest that after a toxic injury there remains a population of partially damaged beta-cells, which are able to maintain most of their basal metabolic functions, but fail to maintain adequate insulin biosynthesis and release.
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PMID:Preferential reduction of insulin production in mouse pancreatic islets maintained in culture after streptozotocin exposure. 245 14


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