bluuebunny/crossref_metadata_2025 huggingface dataset:
Allelic Variants at Insulin-Receptor and Insulin Gene Loci and Susceptibility to NIDDM in Welsh Population
Authors: Morgan Ruth , Bishop Alison , Owens David R , Luzio Stephen D , Peters John R , Rees Alan
Published: September 2013

A cohort of 132 well-documented White Welsh non-insulin-dependent diabetic (NIDDM) subjects were genotyped for 5 restriction-fragment–length polymorphisms (RFLPs) at the insulin-receptor gene (IRG) locus and a polymorphic locus 5' to the insulin gene. There was no significant difference in RFLP frequencies between the NIDDM subjects and a group of 87 matched White control subjects. Paired haplotype analysis of the IRG RFLPs suggested a difference between NIDDM and control groups for the endonuclease combinations Bgl II-Rsa I and Bgl II-Xba I. Analysis of implied haplotypes defined by the endonucleases Bgl II, Rsa I, and Xba I revealed one haplotype to be more prevalent in the NIDDM group; whereas, another haplotype was associated with the control group (P < 0.02). Subset analysis within the NIDDM cohort compared the metabolic response of NIDDM subjects with the differing IRG haplotypes to a standard meal tolerance test. Both groups showed equivalent basal and postprandial glucose excursions, but one group revealed a significantly exaggerated plasma insulin response compared with the other (P < 0.05). This may reflect the influence of genetic variation at the IRG locus on insulin sensitivity in patients with NIDDM.
Isolation of Surface Binding Protein Specific for Advanced Glycosylation End Products From Mouse Macrophage-Derived Cell Line RAW 264.7
Authors: Radoff Steven , Cerami Anthony , Vlassara Helen
Published: September 2013

Macrophages internalize and degrade proteins modified by advanced glycosylation end products (AGEs) via a specific receptor (AGE-R). Chemical cross-linking studies with AGE-bovine serum albumin have demonstrated that the molecular weight of this receptor is ∼90,000. We previously established that the binding constant (Ka) of this receptor site for the chemically synthesized model AGE, 2-(2-furoyl)-4(5)-(2-furanyl)-1H-imidazole-butyric acid (FFI-BA), on cells of the mouse macrophagelike cell line RAW 264.7 is identical to that for AGE proteins. Therefore, FFI was used as an affinity matrix in the first purification step of the AGE-R. The membranes of RAW 264.7 cells were solubilized in octyl-β-glucoside and subjected to affinity chromatography on FFI-sepharose and gel permeation on Superose 6 fast protein liquid chromatography. Sodium dodecyl sulfate (SDS)–polyacrylamide gel electrophoresis analysis of this material revealed a high enrichment of a 90,000-Mr protein that had AGE binding activity. Approximately 25% of the protein at this step was the 90,000-Mr protein. The 90,000-Mr membrane protein was purified to homogeneity by rechromatographing the material on Superose 12 in the presence of SDS before and after reduction with 2-mercaptoethanol. After these harsh conditions, the 90,000-Mr protein lost AGE binding activity. Additional cross-linking studies on human peripheral monocytes revealed an AGE-R protein of identical size to that on RAW 264.7 cells, suggesting the relatively highly conserved nature of this molecule. Given the large heterogeneity of the AGE structures, this AGE-R may represent one of a family of receptors present on a diverse group of different cell types having multiple functions. The implications of this macrophage receptor system in normal tissue remodeling and in proliferative/degenerative diseases are discussed.
Metabolic Effects of Hyperglycemia and Hyperinsulinemia on Fate of Intracellular Glucose in NIDDM
Authors: Henry Robert R , Gumbiner Barry , Flynn Therese , Thorburn Anne W
Published: September 2013

Hyperglycemia in non-insulin-dependent diabetes mellitus (NIDDM) stimulates peripheral glucose uptake, which tends to compensate for impaired insulinmediated glucose uptake. The metabolic fate of glucose and suppression of fat oxidation may differ, however, when glucose uptake is stimulated primarily by insulin or hyperglycemia. To address this issue, three hyperinsulinemic glucose-clamp studies were performed in combination with indirect calorimetry in seven nonobese subjects with NIDDM. In the first two experiments, when glucose uptake was matched at ∼8 mg · kg−1 fat-free mass (FFM) · min1 with primarily hyperinsulinemia (1350 ± 445 pM) or hyperglycemia (20.8 ± 1.8 mM), identical rates of glucose oxidation (3.21 ± 0.29 and 3.10 ± 0.23 mg · kg−1 FFM · min−1, NS) and nonoxidative glucose metabolism (5.19 ± 0.75 and 5.46 ± 0.61 mg · kg−1 FFM · min−1, NS) were achieved. When glucose uptake was increased further to 11.11 ± 0.36 mg · kg−1 FFM · min−1 with less insulin (625 ± 70 pM) and hyperglycemia, glucose oxidation (3.85 ± 0.26 mg · kg−1 FFM · min−1) and nonoxidative glucose metabolism (7.26 ± 0.51 mg · kg−1 FFM min−1) rose significantly (both P < 0.05 from matched studies at lower rates of glucose uptake). During all glucose-clamp studies, free fatty acids were comparably suppressed by 40–46% (all P < 0.005 vs. basal values), whereas fat oxidation was suppressed by 70–80% (all P < 0.005 vs. basal values). A strong negative correlation was observed between rates of glucose and fat oxidation (r = −0.88, P < 0.001) when all studies were combined. These results demonstrate that, when hyperglycemia compensates for impaired insulin-mediated glucose uptake in NIDDM, the ultimate intracellular movement of glucose into oxidative and nonoxidative pathways remains unchanged and the rate of glucose uptake rather than the level of serum insulin determines the disposition of infused glucose. The development of hyperglycemia in NIDDM appears to function as a regulatory mechanism compensating not only for impaired glucose uptake but also for the pathways of intracellular glucose metabolism.
Calcitonin Gene—Related Peptide and Induction of Hyperglycemia in Conscious Rats In Vivo
Authors: Yamaguchi Akinori , Chiba Tsutomu , Morishita Tomoyuki , Nakamura Akira , Inui Tetsuya , Yamatani Toshiyuki , Kadowaki Seizo , Chihara Kazuo , Fukase Masaaki , Fujita Takuo
Published: September 2013

The effect of calcitonin gene-related peptide (CGRP) on glucose metabolism was investigated in conscious and unrestrained rats in vivo. Intravenous injection of rat CGRP (5.67 and 0.567 nmol/kg) caused a significant, dose-dependent increase in plasma glucose concentration and a simultaneous dose-dependent increase in plasma insulin level. In contrast, plasma glucagon level was not changed. On the other hand, intravenous infusion of CGRP (46.6 pmol · kg−1 · min−1) decreased tolerance to intragastric administration of glucose (IGGTT). Plasma insulin response to IGGTT, however, was not affected by CGRP infusion. Moreover, although intravenous injection of CGRP (5.67 nmol/kg) elicited a significant increase in plasma epinephrine and norepinephrine concentrations, concomitant administration of epinephrine and norepinephrine, inducing a more prominent rise in plasma catecholamines than those induced by CGRP, affected neither plasma glucose nor insulin levels. Finally, plasma insulin levels obtained by simulating CGRP-induced changes in plasma glucose or glucose plus catecholamine levels by infusion of glucose or glucose plus catecholamines were not different from those induced by CGRP injection. These results suggest that CGRP has a hyperglycemic action that is not mediated by sympathetic outflow in conscious rats, and inhibition of insulin secretion, if any, does not play a major role in this hyperglycemic action of CGRP. We have demonstrated specific CGRP receptors linked to adenylate cyclase activation in rat liver plasma membranes; this hyperglycemic effect of CGRP in vivo may be partly due to its direct action on the liver.
Effects of Maternal Diabetes on Placental Transfer of Glucose in Rats
Authors: Thomas Chris R , Eriksson Gina L , Eriksson Ulf J
Published: September 2013

In situ perfusion of the fetal side of the anesthetized rat placenta was used to monitor glucose fluxes in nondiabetic, streptozocin-induced diabetic (STZ-D), acutely hyperglycemie nondiabetic, and acutely normoglycemic STZ-D rats. STZ-D resulted in increased accumulation of glucose in the perfusate during a single passage through the fetal vasculature compared with nondiabetic rats, and this increase was maintained in normoglycemic STZ-D rats, indicating glucose release from placental stores. The fractional clearance of 3-O-[14C]methylglucose, a nonmetabolizable glucose analogue, across the placenta was decreased in both STZ-D groups compared with nondiabetic rats but unchanged in hyperglycemic nondiabetic rats, implying a reduction in glucose transporters in diabetic placentas. The difference between the transfer of D-[3H]glucose and 3-O-[14C]methylglucose indicated that 17% of the glucose was retained while traversing the placenta of nondiabetic rats, whereas a smaller percentage (8%) but a larger absolute amount (9 vs. 6 μmol/h) of glucose was retained by the placentas of the severely STZ-D rats. This retention was markedly enhanced in hyperglycemic nondiabetic rats and STZ-D rats when rendered normoglycemic. The net accumulation of perfusate glucose was less than that predicted from radiolabeled transfer data, indicating that glucose is also back transferred from the perfusate to the mother's placenta. We conclude that maternal diabetes markedly affects placental glucose flux.
Effects of Islet Isografts on Hemodynamic and Vascular Filtration Changes in Diabetic Rats
Authors: Pugliese Giuseppe , Tilton Ronald G , Chang Katherine , Speedy Amanda , Province Michael , Eades Donald M , Lacy Paul E , Kilo Charles , Williamson Joseph R
Published: September 2013

To assess the reversibility of diabetes-induced increases in regional vascular albumin permeation and blood flow and changes in kidney filtration islet isografts were given via the portal vein after 2 mo of streptozocin-induced diabetes in male Lewis rats. One month later, vascular function was assessed in control rats, islet-transplanted diabetic rats, and untreated diabetic rats (6–9 rats/group). Untreated diabetic rats were markedly hyperglycemic, hyperphagic, and polyuric. Transplanted rats were euglycemic within 6 days; 24-h urine volumes were virtually normalized by 2 wk and food consumption was normalized 4 wk after transplantation. Vascular albumin permeation in diabetic rats was significantly increased 1.4- to 1.7-fold in anterior uvea, choroid, retina, sciatic nerve, new granulation tissue, and kidney and was increased 1.1- to 1.3-fold in diaphragm, cecum, and optic nerve. Albumin permeation was not increased in aorta, brain, heart, or forelimb skeletal muscle. Islet transplants significantly reduced but did not completely normalize vascular albumin permeation in most tissues in which it was increased by diabetes but had no effect onalbumin permeation in optic nerve, sciatic nerve, and diaphragm. Urinary excretion of endogenous albumin and IgG in diabetic rats was significantly increased 19- and 14-fold, respectively, and wasvirtually normalized 4 days after islet transplantation. Marked (1.8-fold) increases in glomerular filtration rate (GFR) in diabetic rats were also substantially reduced by islet transplants butremained elevated 1.4-fold control values. Likewise, diabetes-induced increases in regional blood flow were reduced in general but not normalized by islet transplants. These observations indicate that 1) diabetes-induced hemodynamic changes and alterations in vascular filtration function are not rapidly reversed by euglycemia after islet transplantation, 2) diabetes-induced increases inurinary albumin and IgG excretion are more readily normalized by euglycemia than increases in GFR and renal 125I-labeled bovine serum albumin (125I-BSA) filtration, and 3) significant increases in GFR and renal 125I-BSA filtration may not be manifested by albuminuria.
Proglucagon Processing Similar to Normal Islets in Pancreatic α-Like Cell Line Derived From Transgenic Mouse Tumor
Authors: Powers Alvin C , Efrat Shimon , Mojsov Svetlana , Spector David , Habener Joel F , Hanahan Douglas
Published: September 2013

A pancreatic α-like cell line has been established from a glucagonoma arising in transgenic mice expressing a hybrid gene consisting of the rat glucagon-promoter sequence fused to the sequence encoding the SV40 T-antigen oncoprotein. The α-tumor cell 1 (αTC1) line maintained many characteristics of differentiated αcells for >40 passages in culture and expressed levels of glucagon mRNA 5- to 10-fold higher than those reported previously in rat and hamster islet cell lines. By radioimmunoassay, the cells synthesized considerable amounts of glucagon, glucagonlike peptide I (GLP-I), the major proglucagon fragment, and small amounts of unprocessed proglucagon but no free GLP-II. This distribution of peptides is similar to that found in extracts of rodent pancreases and is distinct from that seen with other islet cell lines, which process proglucagon in patterns more characteristic of intestinal cells. The GLP-I peptide in the αTC1 cell line was in the form of GLP-I-(1–37), which is inactive as a stimulator of insulin secretion, and not GLP-I-(7–37) or -(7–36)-amide peptides, both of which are potent insulin secretogogues. The αTC1 cell line produced glucagon-related peptides in a relatively uniform pattern by immunocytochemistry, and electron microscopy revealed typical αtype (glucagon) secretory granules. Although the cell line was derived from an islet tumor producing only glucagon, the αTC1 cell line also produced insulin in addition to the glucagon peptides. This and other αTC lines derived from primary αcell tumors that heritably arise in transgenic mice will probably be useful in studies on regulation of glucagon gene expression and αcell physiology.
Systemic Venous Drainage of Pancreas Allografts as Independent Cause of Hyperinsulinemia in Type I Diabetic Recipients
Authors: Diem Peter , Abid Munir , Redmon J Bruce , Sutherland David E R , Robertson R Paul
Published: September 2013

To evaluate the metabolic consequences of pancreas transplantation with systemic venous drainage on β-cell function, we examined insulin and C-peptide responses to glucose and arginine in type I (insulin-dependent) diabetic pancreas recipients (n = 30), nondiabetic kidney recipients (n = 8), and nondiabetic control subjects (n = 28). Basal insulin levels were 66 ± 5 pM in control subjects, 204 ± 18 pM in pancreas recipients (P < 0.0001 vs. control), and 77 ± 17 pM in kidney recipients. Acute insulin responses to glucose were 416 ± 44 pM in control subjects, 763 ± 91 pM in pancreas recipients (P < 0.01 vs. control), and 589 ± 113 pM in kidney recipients (NS vs. control). Basal and stimulated insulin levels in two pancreas recipients with portal venous drainage were normal. Integrated acute C-peptide responses were not statistically different (25.3 ± 4.3 nM/min in pancreas recipients, 34.2 ± 5.5 nM/min in kidney recipients, and 23.7 ± 2.1 nM/min in control subjects). Similar insulin and C-peptide results were obtained with arginine stimulation, and both basal and glucose-stimulated insulin–C-peptide ratios in pancreas recipients were significantly greater than in control subjects. We conclude that recipients of pancreas allografts with systemic venous drainage have elevated basal and stimulated insulin levels and that these alterations are primarily due to alterations of first-pass hepatic insulin clearance, although insulin resistance secondary to immunosuppressive therapy (including prednisone) probably plays a contributing role. To avoid hyperinsulinemia and its possible long-term adverse consequences, transplantation of pancreas allografts into sites with portal rather than systemic venous drainage should be considered.
Uptake of myo-Inositol by Early-Somite Rat Conceptus: Transport Kinetics and Effects of Hyperglycemia
Authors: Weigensberg Marc J , Garcia-Palmer Francisco-José , Freinkel Norbert
Published: September 2013

We have shown that myo-inositol in the cultured rat embryo is diminished whenever malformations are induced by hyperglycemia and that the malformations and reductions of tissue myo-inositol content are not corrected by aldose reductase inhibitors. This study was designed to evaluate the kinetics of myo-[3H]inositol uptake in vitro during 1-, 3-, and 24-h intervals in the 10.5-day rat conceptus (10–12 somites). We found that the equilibration between tissue and medium is relatively slow and that the concentration of free myo-inositol in tissue is only approximately threefold greater than in the medium even after 24 h. The integrated uptake of free myo-inositol by the intact 10.5-day conceptus is a saturable process with a Km(246 ±16 μM) consistent with a low-affinity system. The net rate of accumulation into the tissue pool of free myo-inositol exceeds the rate of incorporation of the accumulated myo-inositol into lipid components. Ambient glucose inhibits net myo-inositol uptake in a concentration-dependent fashion, and the inhibition is competitive in nature. The glucose-mediated inhibitions of myo-inositol transport also compromise the concurrent incorporation of myo-[3H]inositol into lipid components, although to a lesser extent. These inhibitory effects are relatively specific for D-glucose and not replicated by equimolar additions of D-mannose or D-galactose. myo-inositol accumulation by the 10.5-day rat conceptus is also impaired by relatively specific inhibitors of D-glucose transport such as phloridzin or ouabain. Thus, our findings indicate that the concentrative uptake of myo-inositol by the early postimplantation rat conceptus occurs via a comparatively slow transport that may be Na+ dependent and is competitively affected by extracellular glucose. We suggest that the impairment of free myo-inositol uptake and retention by high glucose and the attendant diminution in the incorporation of the myo-inositol into phosphoinositides may contribute to the embryopathic potential of hyperglycemia.
Prediction of Clinical Diabetic Nephropathy in IDDM Patients: Alternatives to Microalbuminuria?
Authors: Mogensen Carl Erik
Published: September 2013

This perspective deals with prediction of overt diabetic nephropathy in patients with insulin-dependent diabetes mellitus (IDDM). The role of elevated urinary albumin excretion rate (microalbuminuria) in predicting diabetic nephropathy has been emphasized by new follow-up studies. Development of severe kidney impairment was seen in a large percentage of patients with microalbuminuria, but with more intensive care for diabetic patients, this percentage may be falling. Herein, I analyzed alternatives to microalbuminuria in predicting kidney disease in diabetes. 1) Parental predisposition to hypertension is not seen in all studies and therefore may not be a decisive factor, and it cannot be used in prediction of nephropathy. 2) Prediabetic blood pressure may predict nephropathy in certain non-insulin-dependent diabetic patients, but elevated blood pressure seems to develop after early microalbuminuria and is likely to be an aggravating factor in established microalbuminuria in IDDM patients. 3) At the clinical diagnosis of IDDM, diabetic nephropathy cannot be predicted. 4) Glycemic control is poor in normoalbuminuric patients with later development of microalbuminuria, and multiple glycosylated hemoglobin measurements are therefore important. 5) In diabetes, glomerular hyperfiltration is associated with late nephropathy, but it alone cannot be the decisive factor, because hyperfiltration in nondiabetic individuals does not produce kidney disease, according to new long-term follow-up studies. 6) Studies of glomerular structure and ultrastructure have not yet documented predictive values for overt nephropathy, but further studies are in progress. 7) Isolated blood pressure elevation without microabuminuria (probably representing essential hypertension in diabetes) has not been predictive. 8) It is clear that elevation of serum creatinine is a very late and insensitive parameter, occurring only with pronounced proteinuria. Creatinine clearance is a poor index for glomerular filtration rate. Therefore, it can be concluded that microalbuminuria currently is the most simple and sensitive parameter for detection of the patient at risk of kidney disease in diabetes. The abnormality probably reflects an early renal lesion rather than being a susceptibility factor. Long-term follow-up of microalbuminuria is recommended for early detection of diabetic kidney disease in IDDM patients, and proposals regarding intervention are discussed.
Synergistic Effects of Male Sex and Obesity on Hepatic Insulin Dynamics in SHR/Mcc-cp Rat
Authors: Hennes Magda M I , McCune Sylvia A , Shrago Earl , Kissebah Ahmed H
Published: September 2013

The effects of obesity and sex on hepatic insulin metabolism were evaluated in the SHR/Mcc-cp rat. During in situ liver perfusion, insulin clearance rate (CLR) expressed per gram of liver tissue was reduced by 58 and 68% in obese females and males, respectively, compared with lean controls. Male sex resulted in CLR reductions of 46% in lean and 59% in obese animals. Obesity resulted in 50% reduction of insulin-receptor binding to isolated hepatocytes. In both lean and obese animals, male sex also resulted in a decrease of −34% in insulin binding. Scatchard plots indicated that the reduction in insulin binding was primarily due to a decrease in number of cell surface receptors. Receptor-mediated insulin degradation was 40% less in obese than lean animals. Male sex also resulted in 27% less insulin degradation relative to females. Receptor-mediated insulin partitioning between four compartments (cell surface bound, internalized and/or cryptic, degraded, and dissociated or released intact), expressed as a percentage of the initial membrane-bound hormone, did not differ between the animal groups. Thus, male sex and obesity are independently and additively associated with a reduction in hepatic insulin clearance and a decrease in the number of cell surface insulin receptors with a proportional decrease in insulin compartmentalization and degradation. This mechanism may partly account for the synergistic effects of male sex and obesity on the degree of hyperinsulinemia and insulin resistance and the predisposition to diabetes.
Relationship of Endoneurial Capillary Abnormalities to Type and Severity of Diabetic Polyneuropathy
Authors: Britland Stephen T , Young Robert J , Sharma Ashutosh K , Clarke Basil F
Published: September 2013

Endoneurial capillary abnormalities have been assessed quantitatively in sural nerve biopsies from diabetic patients with different syndromes of sensory polyneuropathy: chronic painful neuropathy, newly presenting painful neuropathy, and painless neuropathy associated with neurotrophic foot ulceration. Comparisons were made with age-matched nondiabetic control subjects. The diabetic groups showed no abnormality in capillary density or mean endoneurial area per fascicle. Compared with control subjects, all diabetic patients had an increase in mean capillary diameter, capillary wall thickness, and outer tunic (basement membrane and pericytes) thickness. The increase in wall thickness was most pronounced in patients with painless neuropathy (200%) and less marked in similar patients with painful neuropathy (100%). The pericyte volume fraction of the outer tunic was reduced in all diabetic patients, implying that basement membrane hypertrophy and reduplication were responsible for outer tunic thickening. There was evidence of endothelial cell hyperplasia rather than hypertrophy. There was a correlation between the degree of basement membrane thickening and the severity of myelinated fiber abnormality assessed neurophysiologically and morphologically. This study shows a link between the degree of endoneurial capillary basement membrane thickening, the type of neuropathology, and the clinical expression of neuropathy in diabetes mellitus.
Effect of Insulin Therapy on Metabolic Fate of Apolipoprotein B–Containing Lipoproteins in NIDDM
Authors: Taskinen Marja-Riitta , Packard C J , Shepherd J
Published: September 2013

Non-insulin-dependent diabetic (NIDDM) subjects exhibit abnormalities in their plasma lipid and lipoprotein profiles that increase the risk of ischemic heart disease. This study was designed to examine the metabolic behavior of very-low-density (VLDL), intermediate-density (IDL), and low-density (LDL) lipoproteins in NIDDM patients before treatment and after 4 wk of insulin therapy. Basal turnover studies of 131I-labeled VLDL1 (svedberg units [Sf] 60–400) and 131I-labeled VLDL1 (Sf 20–60) apolipoprotein B (apoB) were conducted in a group of seven NIDDM patients who had been off oral therapy for 1 wk. The subjects exhibited higher than normal transport rates for VLDL1 and a diminished input of apoB into the VLDL2 density range. These observations are concordant with the hypothesis that NIDDM patients overproduce VLDL triglyceride but not apoB. VLDL1 and VLDL2 were converted to IDL and ultimately to LDL at approximately normal rates, although the delipidation pathway by which apoB-containing particles were processed exhibited different properties from that seen in control subjects. Insulin therapy reduced plasma triglyceride by 38%, and this was associated with a 41% fall in VLDL1 mass (P < 0.01). VLDL2 was less affected (19% reduction, P < 0.05), IDL was unchanged, and LDL fell 17% (P < 0.05). Repeat metabolic studies revealed that the major effects of insulin were to reduce VLDL1-apoB transport (from 811 to 488 mg/day) and increase the direct input of VLDL2 into the plasma (from 182 to 533 mg/day, P < 0.05). These alterations in VLDL production led to normalization of apoB kinetics in IDL and LDL. The fractional catabolic rate of LDL increased 19% (P < 0.05), whereas direct input into this fraction, which had been high before treatment, was reduced. Postheparin plasma lipoprotein lipase (LPL) and hepatic lipase levels were unaffected by insulin, although the hormone did increase LPL in adipose tissue. This lack of effect on lipase activities correlated well with the observation that the rates of catabolism of apoB in VLDL1, VLDL2, and IDL were not significantly affected by insulin therapy.
Increase in [3H]PN 200-110 Binding to Cardiac Muscle Membrane in Streptozocin-Induced Diabetic Rats
Authors: Nishio Yoshihiko , Kashiwagi Atsunori , Ogawa Tsutomu , Asahina Takayuki , Ikebuchi Motoyoshi , Kodama Mitsuaki , Shigeta Yukio
Published: September 2013

Voltage-sensitve Ca2+ channels in cardiac left ventricular muscle membranes isolated from nondiabetic control and diabetic rats were measured with [3H]PN 200–110, a dihydropyridine derivative, as a ligand. The binding site (Bmax) of [3H]PN 200–110 in cardiac membranes isolated from streptozocin-induced diabetic (STZ-D) rats (128 ± 10 fmol/mg protein) significantly (P < 0.01) increased by 64% compared with that of control rats (78 ± 4 fmol/mg protein) 10 wk after STZ administration without a significant change in Kd. However, the significant increase in Bmax of [3H]PN 200-110 binding in diabetic rats depended on the duration of diabetes such that the increase was not found until 6 wk after STZ injection. An 8-wk intensive insulin treatment, which was initiated 2 wk after STZ injection, normalized the increase in [3H]PN 200–110 binding in STZ-D rats to control levels (85 ± 4 fmol/mg protein). Furthermore, [3H]PN 200–110 binding to control cardiac membranes was dose-dependently inhibited in the presence of verapamil, a phenylalkylamine Ca2+ antagonist, but that was not the case in cardiac membranes isolated from STZ-D rats. These results indicate that voltage-sensitive Ca2+ channels in cardiac muscle isolated from STZ-D rats are quantitatively and qualitatively altered, because the course of diabetes and the increase in the channels can be prevented by treatment with insulin.
Age as Independent Determinant of Glucose Tolerance
Authors: Shimokata Hiroshi , Muller Denis C , Fleg Jerome L , Sorkin John , Ziemba Andrzej W , Andres Reubin
Published: September 2013

It has been proposed that the decline in glucose tolerance with age is not a primary aging effect but is secondary to a combination of other age-associated characteristics, i.e., disease, medication, obesity, central and upper-body fat deposition, and inactivity. To test this hypothesis, we first eliminated from analysis the Baltimore Longitudinal Study of Aging participants with identifiable diseases or medications known to influence glucose tolerance. Seven hundred forty-three men and women, aged 17–92 yr, remained for analysis. As indices of fatness, body mass index and percent body fat were determined. As indices of body fat distribution, waist-hip ratio and subscapular triceps skin-fold ratio were calculated. As indices of fitness, physical activity level, determined by detailed questionnaire, and maximum 02 consumption were calculated. We tested whether the effect of age on glucose tolerance remains when data were adjusted for fatness, fitness, and fat distribution; 2-h glucose values were 6.61, 6.78, and 7.83 mM for young (17–39 yr), middle-aged (40–59 yr), and old (60–92 yr) men and 6.22, 6.22, and 7.28 mM for the three groups of women, respectively. The differences between the young and middle-aged groups were not significant, but the old groups had significantly higher values than young or middle-aged groups. Fatness, fitness, and fat distribution can account for the decline in glucose tolerance from the young adult to the middle-aged years. However, age remains a significant determinant of the further decline in glucose tolerance of healthy old subjects.
Effects of Media Conditions, Insulin, and Dexamethasone on Insulin-Receptor mRNA and Promoter Activity in HepG2 Cells
Authors: Levy James R , Krystal Geoffrey , Glickman Penny , Dastvan Frank
Published: September 2013

Numerous physiological agents and conditions modulate cellular insulin sensitivity by downregulating or upregulating total cellular insulin receptors. In this study, we examined the effects of replacing complete medium in the absence or presence of insulin on the regulation of insulin-receptor gene expression in cultured human hepatoma cells (HepG2). Failure to replace complete medium resulted in growth arrest of HepG2 cells and a six- to sevenfold increase in insulin-receptor mRNA due to the prolongation of insulin-receptor mRNA half-life. Northern analysis revealed multiple insulin-receptor mRNA species; the largest species (11 kilobases) was disproportionately increased in growth-arrested cells. High concentrations of insulin (500 ng/ml) induced a 33.8% decrease in the abundance of insulin-receptor mRNA (n = 14). At lower concentrations, a trend of inhibition was observed but was not statistically significant. Insulin (500 ng/ml) did not affect insulin-receptor mRNA stability. The effect of conditioned media, insulin, and dexamethasone on insulin-receptor promoter activity was also examined. Various constructs of the 5-flanking region of the insulin-receptor gene were attached immediately upstream to a chloramphenicol acetyltransferase (CAT) reporter gene and transiently transfected into HepG2 cells via a pBR322-derived plasmid (pCAT). In cells replaced with complete medium, 12 and 118% of the promoter activity was contained within 578 and 877 base pairs, respectively, from the major translational initiation site. Conditioned media from growth-arrested cells in culture for 7 days increased promoter activity approximately twofold in 48 h. However, this increase failed to localize to any specific region on the insulin-receptor promoter. In addition, CAT activity doubled in cells transiently transfected with pCAT containing no 5-flanking region of the insulin-receptor gene, suggesting that conditioned media increased CAT activity posttranscriptionally. Insulin (500 ng/ml) and dexamethasone (10−6 M) had little effect on promoter activity.
No Reduction in Total Hepatic Glucose Output by Inhibition of Gluconeogenesis With Ethanol in NIDDM Patients
Authors: Puhakainen Ilpo , Koivisto Veikko A , Yki-Järvinen Hannele
Published: September 2013

Increased gluconeogenesis has been suggested to account for all of the increase in basal glucose production in patients with non-insulin-dependent diabetes mellitus (NIDDM). We studied the effect of inhibition of gluconeogenesis with ethanol on total hepatic glucose output (HGO) in patients with NIDDM. Fourteen patients with NIDDM (mean ± SE age 61 ± 2 yr, fasting plasma glucose 11.4 ± 0.8 mM; body mass index 27 ± 1 kg/m2) were studied twice after an overnight fast, once during ethanol administration (blood ethanol ∼12 mM) and once during saline administration. Total HGO rate was measured with [3H]glucose. Inhibition of gluconeogenesis by ethanol was followed qualitatively with [U-14C]lactate (n = 8) and [U-14C]glycerol (n = 6) as tracers. Ethanol inhibited gluconeogenesis from lactate by 71 ± 5% (0.5 ± 0.2 vs. 1.8 ± 0.1 μmol glucose · kg−1 · min−1, 240–300 min, P < 0.001; ethanol vs. saline, P < 0.001) and from glycerol by 65 ± 6% (0.8 ± 0.2 vs. 2.3 ± 0.6 μmol glucose · kg · min−1, P < 0.001). Total HGO rate remained unchanged and averaged 12.8 ± 1.8 and 11.8 ± 2.1 μmol · kg−1 · min−1 in the saline and ethanol studies, respectively (NS). We concluded that inhibition of gluconeogenesis by ethanol does not decrease total HGO in patients with NIDDM. Our results suggest the existence of a regulatory mechanism in the liver that maintains constant total HGO despite inhibition of gluconeogenesis.
Evolution of Insulin Resistance in New Zealand Obese Mice
Authors: Veroni Marta C , Proietto Joseph , Larkins Richard G
Published: September 2013

The etiology of non-insulin-dependent diabetes mellitus (NIDDM) is not known. Hyperglycemia is due to increased hepatic glucose production (HGP), decreased glucose uptake, and impaired insulin secretion. It is unknown if these defects are coinherited or if one precedes and causes the others. The aim of this study was to determine the earliest defects in the evolution of the syndrome in the New Zealand obese (NZO) mouse, a polygenic model of NIDDM. NZO and control NZC mice were studied at 4–5 and 20 wk of age. Glucose turnover and glucose uptake in individual tissues were measured basally and during a hyperinsulinemic clamp. First-phase insulin secretion was measured after an intravenous glucose load. HGP was higher in the NZO mice both basally and during the clamp at bothages. At 4–5 wk of age, there was evidence of insulin insensitivity in brown adipose tissue, soleus, diaphragm, red quadriceps, and red gastrocnemius but not in heart, white quadriceps, and white gastrocnemius. In 20-wk-old mice, insulin responsiveness was decreased in white and brown adipose tissue and soleus muscle but not in heart, diaphragm, red and white quadriceps, and red and white gastrocnemius. First-phase insulin secretion (percentage rise above basal) 3 min after the glucose bolus was impaired in NZO miceat both ages. We conclude that hepatic glucose overproduction, brown adipose tissue and skeletal muscle insulin resistance, and impaired first-phase insulin secretion are all early abnormalities in the NZO mouse.
Differential Effects of Diabetes on Adipocyte and Liver Phosphotyrosine and Phosphoserine Phosphatase Activities
Authors: Begum Najma , Sussman Karl E , Draznin Boris
Published: September 2013

We examined the activities of particulate and cytosolic phosphotyrosine phosphatase (PTPase) and phosphoserine phosphatase (PSPase) in adipocytes and livers of diabetic rats. PTPase activity was assessed with [32P]tyrosine-phosphorylated insulin receptor (IR), whereas PSPase activity was assayed with [32P]serine-phosphorylated glycogen synthase. Diabetes increased adipocyte particulate PTPase activity and enhanced IR dephosphorylation by 75% on the 2nd, 93% on the 14th, and 108% on the 30th day. In contrast, cytosolic PTPase activity decreased by 78% on the 14th and 45% on the 30th day (no change on the 2nd day). Similar changes were observed with PSPase (increased activity in particulate and decreased in cytosolic). Insulin therapy for 14 or 30 days restored PTPase and PSPase activities in both fractions. Vanadate, despite rapid normalization of glycemia, restored these activities only after 30 days of therapy. Diabetes-related changes in liver PTPase activity were observed on the 14th day only. At this time, it was increased in both particulate and cytosolic fractions. There was spontaneous normalization of the liver PTPase activity at 30 days of diabetes. In contrast, liver cytosolic PSPase activity was significantly inhibited and not normalized by the 30th day of disease without therapy. In summary, diabetes appears to induce tissue-specific changes in PTPase and PSPase activities resulting in significant alterations in dephosphorylation of IR and glycogen synthase. Moreover, there appears to be a differential regulation of PTPase and PSPase activities in diabetes, particularly in the liver.
Association of Elevated IGF-I Levels With Increased Retinopathy in Late-Onset Diabetes
Authors: Dills Diana G , Moss Scot E , Klein Ronald , Klein Barbara E K
Published: September 2013

Insulinlike growth factor I (IGF-I) has been suggested to play a role in the pathogenesis of proliferative diabetic retinopathy (PDR). We determined IGF-I levels in subjects in a large population-based study of 928 people with diabetes diagnosed at 30 yr of age or older. PDR was found in 15.7% of the insulin-using group (n = 517) and in 2.8% of those not using insulin (n = 397). The mean serum level of IGF-I was 208 μg/L in individuals using insulin and 222 μg/L in those not using insulin, both significantly lower than in a nondiabetic comparison group (278 μg/L, P < 0.0001). Logistic regression analysis was used to examine the relationship between IGF-I and PDR while controlling for other factors associated with the presence of PDR. After controlling for duration of diabetes, glycosylated hemoglobin, systolic blood pressure, presence of proteinuria, and age at diagnosis, higher levels of IGF-I were significantly associated with an increased frequency of PDR (P = 0.025) in the group using insulin. In individuals not using insulin, higher levels of IGF-I were associated with an increased frequency of PDR or moderate non-PDR (P = 0.08). These data suggest that higher IGF-I levels may be a risk factor for the development of severe retinopathy in people with diabetes diagnosed at 30 yr of age or older.
Influence of Environmental Viral Agents on Frequency and Tempo of Diabetes Mellitus in BB/Wor Rats
Authors: Like Arthur A , Guberski Dennis L , Butler Leonard
Published: September 2013

Elimination of environmental viruses by cesarean derivation of the University of Massachusetts colony of BB/Wor rats increased the frequency and accelerated the tempo of spontaneous diabetes among diabetes-prone (DP) rats. In contrast, the viral-antibody–free (VAF) environment did not alter the resistance of pre-VAF diabetes-resistant (DR) rats to spontaneous and RT6+ T-lymphocyte–depletion–induced diabetes. Pre-VAF and VAF rats have essentially the same lymphocyte subsets, and VAF-DP rats are susceptible to the adoptive transfer of diabetes and to the diabetes-accelerating effects of polyinosinic-polycytidylic acid injections. These results suggest that the presence of environmental viral pathogens may act to inhibit effector cell function in lymphopenic DP rats while enhancing effector cell activity in nonlymphopenic DR rats.
Effects of Nonenzymatic Glycosylation of Mesangial Matrix on Proliferation of Mesangial Cells
Authors: Crowley Susan T , Brownlee Michael , Edelstein Diane , Satriano Joseph A , Mori Tetsuo , Singhal Pravin C , Schlondorff Detlef O
Published: September 2013

Cross-linking of cell matrix components by nonenzymatic glycosylation may contribute to diabetic glomerulopathy. We examined the effects of modification of matrix by nonenzymatic glycosylation on mesangial cell function. Matrix was generated by growing mesangial cells in tissue culture for 2 wk and removing the cells with a detergent cell-lysis solution. By indirect immunofluorescence and Northern-blot analysis, the remaining matrix contained laminin, fibronectin, and collagens type I and IV. The matrix was modified by incubation for 24 h with 50 mM glycolaldehyde, a highly reactive cross-linking nonenzymatic glycosylation product, or for 2 wk with 200 mM glucose-6-phosphate (G6P). Modification was carried out with or without equimolar aminoguanidine, an inhibitor of cross-link formation. Nonenzymatic glycosylation of the matrix by glycolaldehyde or G6P was confirmed by fluorometry and [14C]G6P incorporation and was prevented by aminoguanidine. [3H]thymidine incorporation for 24 h by mesangial cells plated onto unmodified or modified matrix was then performed. Modification of matrix had no effect on attachment of mesangial cells, determined 4 h after plating. Nonenzymatic glycosylation of matrix by glycolaldehyde or G6P significantly inhibited thymidine incorporation by mesangial cells. This effect was partially reversible by aminoguanidine. Aminoguanidine-modified matrix had no effect on thymidine incorporation. Thymidine-incorporation results were confirmed by direct cell counting. We conclude that modification of matrix by nonenzymatic glycosylation influences growth of mesangial cells, which could contribute to the mesangial abnormalities of diabetic glomerulopathy.
Persistent Reversal of Diabetes by Transplantation of Fetal Pig Proislets Into Nude Mice
Authors: Liu Xuemei , Federlin Konrad F , Bretzel Reinhard G , Hering Bernhard J , Brendel Mathias D
Published: September 2013

Facing the limited availability of human adult and fetal pancreases, fetal pig proislets (pancreatic islet precursors) were investigated in view of several inherent advantages. Six litters of fetuses of mean ± SE gestational age 75 ± 3 days were obtained from commercially available farm pigs. Pancreatic tissue was gently digested with collagenase, then a 10-day culture was performed. During culture, fetal proislets showed no insulin response to glucose alone but a significant response to glucose plus theophylline. The insulin content per microgram of DNA in the cultured proislets continuously increased. Histological examination by immunoperoxidase staining showed that, apart from single insulin- and glucagon-positive cells, there were no discrete islets in the pancreatic tissue and the cultured proislets. Diabetes was induced with streptozocin (STZ) in eight nude mice 3–4 wk after proislet transplantation and in another eight nude mice without transplantation. During the initial week, blood glucose levels of mice in both groups increased rapidly. The mean ± SE peak value of blood glucose levels in the transplanted group was 20.4 ± 2.0 mM and was 20.1 ± 1.3 mM in the group without transplantation. Simultaneously, body weight decreased from 29.5 ± 0.7 to 21.5 ± 0.9 g and from 27.9 ± 0.7 to 19 ± 1 g in the groups, respectively. Afterward, blood glucose levels of mice in the transplanted group gradually decreased, and normoglycemia was achieved in all mice within 50 ± 13 days after injection of STZ, i.e., 74 ± 13 days after transplantation. The group without transplantation persistently maintained blood glucose levels >16.7 mM. An intraperitoneal glucose tolerance test in five transplanted diabetic mice with normalized blood glucose levels showed a flat response curve. Grafts from two diabetic mice that had reverted to normoglycemia for >3 wk showed a significant response to 16.7 mM glucose alone and to 16.7 mM glucose plus 10 mM theophylline during perifusion. Abundant ducts, β-cell nests, mature islets, and vascularization were found with immunoperoxidase staining in sections of grafts derived from nondiabetic nude mice 2 mo after transplantation and also from diabetic mice that became normoglycemic by transplantation. Our results show that the tissue-culture method effectively produces and purifies fetal pig proislets. The purified proislets develop and differentiate in vivo but not in vitro. This study demonstrates for the first time that STZ is not toxic to fetal pig pancreatic α- and β-cells and that, with this new procedure, fetal pig proislets persistently reverse diabetes in nude mice.
Transplantation of Purified Islet Cells in Diabetic Rats: I. Standardization of Islet Cell Grafts
Authors: Pipeleers Daniel G , Pipeleers-Marichal Miriam , Hannaert Jean-Claude , Berghmans Marleen , In't Veld Peter A , Rozing Jan , van de Winkel Marnix , Gepts Willy
Published: September 2013

A standardized procedure was developed for the preparation of rat islet cell grafts with selected cell number and composition. After collagenase digestion of pancreases and elutriation of tissue fragments, islets were isolated and dissociated, and cells were purified by autofluorescence-activated cell sorting. Approximately 30% of the initial β-cell mass was lost during digestion and elimination of small mostly exocrine particles. Fifty percent was recovered in isolated islet preparations and 30% in the purified β-cell suspensions of >95% purity and viability. Sorting according to cellular flavin adenine dinucleotide content discriminated islet β-cells from islet endocrine non–beta-cells, fibroblasts, leukocytes, and exocrine cells. Purified endocrine islet cell grafts were prepared by aggregating 106 pure β-cells with or without 8 × 105 pure endocrine non–²-cells. In contrast to intact islets, the purified aggregates were devoid of nonendocrine and damaged cells. Intraportal implantation of a pure β-cell graft rapidly and permanently normalized the diabetic state of streptozocin-administered animals. The standardized preparation of purified β-cell grafts allows us to address several metabolic and immunological questions concerning islet cell transplantation in diabetes.
Abnormal Regulation of Protein Tyrosine Phosphatase Activities in Skeletal Muscle of Insulin-Resistant Humans
Authors: McGuire Mary C , Fields Rose M , Nyomba Bulangu L , Raz Itamar , Bogardus Clifton , Tonks Nicholas K , Sommercorn James
Published: September 2013

Insulin resistance in skeletal muscle may be an expression of the genetic basis of a common form of non-insulin-dependent diabetes mellitus (NIDDM) in humans. Impaired insulin action results from an apparent postreceptor defect in insulin signal transduction that limits the influence of the hormone on various protein serine/threonine kinases and phosphatases that are thought to contribute to the mechanism by which insulin affects intracellular events. The fact that numerous responses to insulin are affected suggests that the cause of insulin resistance involves an early step in insulin action. Therefore, we examined the influence of insulin on protein tyrosine phosphatase (PTPase) activities, which may counteract the protein tyrosine kinase activity of the insulin receptor in skeletal muscle of insulin-sensitive and insulin-resistant humans. Insulin infusion in vivo produced a rapid 25% suppression of soluble-PTPase activity in muscle of insulin-sensitive subjects, but this response was severely impaired in subjects who were insulin resistant. Insulin did not affect PTPase activity in the particulate fraction of muscle from either group, but basal particulate activity was 33% higher in resistant subjects than in sensitive subjects. Either or both of these abnormal characteristics of PTPase activities could be central to the causes of insulin resistance and NIDDM.