Stable lodinated polypeptide hormones prepared by polyacrylamide gel electrophoresis

b-Cell mass, hexokinase/glucokinase (HK/GK) activity, glucose metabolism and insulin secretion were studied in the islets of rats with fructose-induced insulin resistance (IR). Normal male Wistar rats were fed a standard commercial diet and water without (control, C) or with 10% fructose-rich diet (FRD) for 3 weeks. Blood glucose (strips), triglyceride (commercial kit), and insulin (RIA) levels were measured at the time of death. Glucose-induced insulin release, glucose metabolism ( 14 CO 2 andH]-glucose) and HK/GK activity (G-6-P production), transcription (RT-PCR), protein expression (Western blot), and cellular compartmentalization were measured in isolated islets (collagenase digestion). FRD rats presented normoglycemia but impaired glucose tolerance, hypertriglyceridemia, hyperinsulinemia, and increased HOMA-IR index. In these rats, b-cell mass decreased significantly by 33%, with a 44% increase in the percentage of apoptotic cells. Glucoseinduced insulin release and islet glucose metabolism were higher in FRD rats. While GK activity (total and cytosolic fraction) and protein expression were significantly higher in FRD islets, HK showed no change in any of these parameters. Our results demonstrate that the changes induced by dietary-induced IR upon b-cell function and mass are strongly conditional on the nutrient model used. In our model (intact animals with impaired glucose tolerance), GK activity increases through mechanisms previously shown only in vitro or under highly hyperglycemic conditions. Such an increase plays a pivotal role in the adaptive increased release of insulin in response to IR, even in the presence of marked b-cell mass reduction.

Section: Blood Measurementsmentioning

confidence: 99%

Islet adaptive changes to fructose-induced insulin resistance: β-cell mass, glucokinase, glucose metabolism, and insulin secretion

Maiztegui¹,

Borelli²,

Raschia³

et al. 2008

“…When the animals were killed blood samples were collected for glucose determination by the glucose oxidase GOD-PAP method (Roche Diagnostics), and for immunoreactive insulin levels by RIA (Herbert et al 1965), using an antibody against rat insulin, rat insulin standard (Linco Research, St. Charles, MO, USA), and highly purified porcine insulin labeled with 125 I (Linde et al 1980); parallel tracer-displacement curves were obtained using either the rat insulin standard, or partially purified insulin extracted from a pool of hamster pancreases (Massa et al 1997). The pancreas from each animal was removed to isolate islets by collagenase digestion (Lacy & Kostianovsky 1967).…”

Section: Blood Measurements and Pancreas Removalmentioning

confidence: 99%

“…At the end of the incubation period, aliquots were taken from the medium to measure insulin by RIA (Herbert et al 1965, Linde et al 1980.…”

Section: Insulin Secretion In Vitromentioning

confidence: 99%

Enhanced expression of hexokinase I in pancreatic islets induced by sucrose administration

Maiztegui

Borelli²,

Massa³

et al. 2006

Administration of a sucrose-rich diet (SRD) to normal hamsters induces an insulin-resistant state and a significant increase of insulin secretion and -cell mass. Islets isolated from these animals had a marked increase in glucose metabolism and glucose-induced insulin secretion, at both low and high glucose concentrations. They also presented increased hexokinase (HK) activity, without measurable changes in glucokinase (GK) activity. In this study we measured HK and GK activity in homogenates of islets isolated from normal control and SRD-fed hamsters, as well as in their particulate and cytosolic fractions. We also measured transcription rate (mRNA by reverse transcriptase PCR) and expression levels (Western blotting) of both enzymes in these islets. We found an increase in HK activity and expression levels, without measurable changes in HK mRNA level in SRD-fed animals. Whereas a similar GK activity was measured in homogenates of islets isolated from both groups, such activity was significantly higher in the cytosolic fraction of SRD islets. On the other hand, GK transcription rate and expression level were similar in both experimental groups. Our results suggest that the increased -cell secretory response to low glucose can be partly ascribed to an increased activity of islet HK consecutive to an enhanced expression of the enzyme, while the enhanced response to high glucose could be due to changes in GK compartmentalization.

“…Insulin levels were determined by radioimmunoassay (RIA) (Herbert et al 1965), using an antibody against rat insulin, a rat-insulin standard (Linco Research, St Charles, MI, USA) and highly purified porcine insulin labelled with 125 I (Linde et al 1980).…”

Section: Blood Parametersmentioning

confidence: 99%

Islet neogenesis: an apparent key component of long-term pancreas adaptation to increased insulin demand

Zotto

Borelli²,

Flores³

et al. 2004

This study aimed to determine the relative importance of different functional and morphological pancreatic changes induced by the chronic administration of a sucrose-rich diet (SRD) to maintain normal glucose homeostasis. Male Wistar rats were fed either sucrose (SRD) or starch (CD) for 6 and 12 months. At both periods, serum glucose and triacylglycerol levels were significantly higher (P<0·05; paired and unpaired Student's t-test) in SRD rats. Serum insulin levels were significantly lower in SRD only at 12 months. At 6 months, the insulin secretion doseresponse curve in SRD rats showed a shift to the left that was no longer observed at 12 months, when SRD islets decreased their response to 16 mM glucose. At 6 months, SRD rats showed a significant increase in -cell volume density (Vvi) and islet cell replication rate, together with a decrease in -cell apoptotic rate. Changes were not detected in the percentage of PDX-1-and islet neogenesis associated protein (INGAP)-positive cells. Conversely, at 12 months, there was a significant decrease in -cell Vvi and in the percentage of PDX-1-positive cells; the islet cell replication rate was not modified, and the number of apoptotic -cells increased significantly. No signs of increased neogenesis or INGAP-positive cells were recorded at any period in SRD rats. Our results show that SRD rats are unable to develop functional and morphological pancreatic reactive changes sufficient to maintain normal glucose and triacylglycerol levels for a long period. Such failure could be ascribed to their inability to increase the rate of neogenesis and of INGAP production.