bluuebunny/crossref_metadata_2025 huggingface dataset:
Counterregulatory Hormone Responses Preserved After Long-Term Intravenous Insulin Infusion Compared to Continuous Subcutaneous Insulin Infusion
Authors: Gulan Mirjane , Perlman Kusiel , Sole Michael , Albisser A Michael , Zinman Bernard
Published: September 2013

The counterregulatory hormone responses of cortisol, growth hormone, glucagon, epinephrine, norepinephrine, and dopamine to a fixed hypoglycemic stimulus (50 mg/dl for 1 h) were studied in five type I (insulin-dependent) diabetic subjects during conventional insulin therapy (CT), after 3 mo of continuous subcutaneous insulin infusion (SC), and after 3 mo of continuous intravenous insulin infusion (IV). During the two infusion periods, the overall mean levels of preprandial blood glucose (116 ± 6 SC vs. 114 ± 5 mg/dl IV) and glycosylated hemoglobin (6.1 ± 2 SC vs. 5.9 ± 2% IV) were virtually identical, but there were more hypoglycemic episodes and greater variability of preprandial blood glucose levels during SC than with IV. During the last 30 min of the hypoglycemic clamps, the mean levels of epinephrine and cortisol were significantly lower after 3 mo of SC (epinephrine, 268 ± 80 pg/ml; cortisol, 14 ± 1 μg/dl) than with both CT (epinephrine 485 ± 80 pg/ml; cortisol, 20 ± 2 μg/dl) and IV (epinephrine, 443 ± 62 pg/ml; cortisol, 19 ± 2 μg/dl)(P < .05). The mean growth hormone level was significantly (P < .05) lower after SC (37 ± 9 ng/ml) than after IV (79 ± 12 ng/ml), but it did not reach statistical significance compared with CT (66 ± 12 ng/ml). The mean glucagon, dopamine, and norepinephrine levels during the same period of hypoglycemia were not different when all treatment regimens were compared. We conclude that intensified insulin therapy with SC leads to significant blunting of the counterregulatory hormone response to hypoglycemia, whereas IV does not. This may partly result from less frequent hypoglycemia with IV, and it may represent a therapeutic advantage of this route of insulin administration.
Insulin Deficiency Decreases Lipoprotein Lipase Secretion by Murine Macrophages
Authors: Behr Stephen R , Kraemer Fredric B
Published: September 2013

We investigated the effects of insulin deficiency and insulin treatment on the secretion of lipoprotein lipase (LPL) by murine macrophages. Streptozocin-induced insulin deficiency caused hyperglycemia and hypertriglyceridemia in mice. Peritoneal macrophages isolated from insulin-deficient mice secreted 70% less LPL activity than control mice. A 65% decrease in LPL activity in epididymal adipose tissue, without any changes in heart LPL activity, was also seen with insulin deficiency. One week of insulin treatment lowered plasma glucose and triglyceride levels in insulin-deficient mice. Additionally, 1 wk of insulin treatment increased LPL secretion by macrophages, but to only one-half of control, while normalizing adipose tissue LPL activity. One injection of insulin also increased LPL secretion by macrophages to one-half of control and normalized adipose tissue LPL activity, even though plasma glucose and triglyceride levels were not affected. In vitro insulin treatment of macrophages isolated from control or insulin-deficient mice had no effect on LPL secretion. The results suggest that insulin does not exert a direct effect on the LPL secretion by macrophages but that deficiency of insulin indirectly causes a profound decrease in macrophage LPL secretion. These changes in macrophage LPL secretion may contribute to the atherosclerotic process in diabetes mellitus.
Interrelationships Among Insulin's Antilipolytic and Glucoregulatory Effects and Plasma Triglycerides in Nondiabetic and Diabetic Patients With Endogenous Hypertriglyceridemia
Authors: Yki-Järvinen Hannele , Taskinen Marja-Riitta
Published: September 2013

We tested the hypothesis that the previously observed association among hypertriglyceridemia, hyperinsulinemia, and insulin resistance could be explained by a defect in insulin's antilipolytic effect. Insulin action was measured in 10 nondiabetic and 8 diabetic patients with hypertriglyceridemia (fasting plasma triglyceride 800 ± 154 and 1105 ± 445 mg/dl, respectively, P NS; fasting plasma glucose 99 ± 3 and 161 ± 12 mg/dl, respectively, P < .001) and in 8 weight-matched normolipemic nondiabetic individuals (fasting plasma triglyceride and glucose 110 ± 21 and 91 ± 3 mg/dl). The slope of the decay in plasma free fatty acid (FFA) during insulin infusion was used as an index of insulin's antilipolytic effect. Insulin stimulation of glucose uptake in vivo during intravenous hyperinsulinemic clamp and in vitro in adipocytes were measures of insulin's glucoregulatory action. Both glucoregulatory and antilipolytic effects were similarly reduced in both hypertriglyceridemic groups compared with normal subjects. The plasma triglyceride concentration correlated positively with the slope of FFA suppression by insulin (r = .81, P < .0001) and the fasting FFA concentration (r = .65, P < .0001). In multiple linear regression analysis, insulin's antilipolytic effect and the fasting FFA concentration explained 83% of the variation in the plasma triglyceride concentration. These associations were independent of insulin's glucoregulatory effect and the fasting plasma insulin concentration. The data indicate that patients with endogenous hypertriglyceridemia are resistant to both the antilipolytic and glucoregulatory actions of insulin and that increased flux of FFA as a result of the latter, rather than hyperinsulinemia, is responsible for elevation of very-low-density lipoprotein production. Finally, we suggest that previous inconsistent results regarding insulin's antilipolytic effect in obesity and non-insulin-dependent diabetes mellitus could be related to variation in the triglyceride concentrations of the study populations.
Calmodulin as Substrate for Insulin-Receptor Kinase: Phosphorylation by Receptors From Rat Skeletal Muscle
Authors: Sacks David B , McDonald Jay M
Published: September 2013

Calmodulin is a substrate for insulin-receptor kinase obtained from rat adipoyctes and hepatocytes and human placenta. In this study, we demonstrate that insulin stimulates the phosphorylation of calmodulin via insulin receptors partially purified from rat skeletal muscle. Phosphorylation of calmodulin was maximal in the presence of Mg2+ and insulin and the absence of Ca2+. Free-Ca2+ concentrations >0.1 μM progressively inhibited phosphorylation with almost total inhibition at 200 μM Ca2+. Insulin-stimulated phosphorylation of calmodulin was dose dependent and saturable with half-maximal effect obtained at ∼5 × 10 −10 M insulin. There was an absolute requirement for certain basic proteins, e.g., polylysine or protamine sulfate, to obtain phosphate incorporation into calmodulin. Polylysine stimulated the phosphorylation of calmodulin independently of insulin, but this was increased up to sixfold by the addition of insulin. Phosphate incorporation into calmodulin increased with increasing concentration of the substrate up to a saturating concentration of 2.4 μM. The Km for calmodulin was ∼0.2 μM. Up to 0.15 mol of phosphate was incorporated per mole of calmodulin with tyrosine the predominant amino acid phosphorylated. The observations that calmodulin is phosphorylated by insulin-receptor kinase from all three classic target organs for insulin confirm that calmodulin is a general substrate for this kinase and suggest that Ca2+ and calmodulin may be components of the insulin-signaling mechanism.
Contribution of Insulin Resistance to Catabolic Effect of Prednisone on Leucine Metabolism in Humans
Authors: Zimmerman Teresa , Horber Fritz , Rodriguez Nancy , Schwenk W Frederick , Haymond Morey W
Published: September 2013

To determine the role insulin resistance may play in the catabolic effect of high-dose prednisone therapy, healthy volunteers were studied on four occasions with the hormone-clamp technique at two insulin infusion rates. Subjects were studied after 5 days of prednisone (60 mg/day) or no steroid treatment and were infused with somatostatin, glucagon, growth hormone, [3H]glucose, [14C]leucine, and insulin (0.1 or 0.2 mil · kg1 · min1). At each rate of insulin infusion, the rate of leucine oxidation was increased (P < .001) after steroid treatment. Leucine flux, an indicator of whole-body proteolysis, was similar in the presence or absence of steroid treatment (2.26 ± 0.08 vs. 2.13 ± 0.04 μmol · kg1 · min1 respectively) at the lower rate of insulin infusion but was higher during steroid treatment (2.18 ± 0.06 vs. 1.84 ± 0.13 μmol · kg1 · min1) at the 0.2-mU · kg1 · min1 insulin infusion. Steroid pretreatment had no significant effect on the nonoxidative rates of leucine disappearance. These data provide strong evidence that the protein wasting associated with glucocorticosteroid therapy is in part the result of steroid-induced resistance to the antiproteolytic effect of insulin and an increase in the oxidation (and thus wasting) of one essential amino acid, leucine.
Prevention of Cyclophosphamide-Induced Diabetes by Anti-Vβ8 T-Lymphocyte–Receptor Monoclonal Antibody Therapy in NOD/Wehi Mice
Authors: Bacelj Angela , Charlton Brett , Mandel Thomas E
Published: September 2013

Walter & Eliza Hall Institute nonobese diabetic (NOD/Wehi) mice exhibit a low incidence of spontaneous diabetes mellitus, but one large dose of cyclophosphamide (CY) can lead to a rapid progression to overt diabetes. Macrophages and Lyt-2+ and L3T4+ cells have been demonstrated to be involved in β-cell destruction in this model. The role of a specific subset of T-lymphocytes expressing a particular T-lymphocyte–receptor segment was examined in CY-induced diabetic NOD mice with a mouse anti-Vβ8 T-lymphocyte–receptor monoclonal antibody (F23.1). After administration of CY, only 4 of 51 treated mice became hyperglycemie compared to 23 of 47 untreated mice, 13 of 26 mice treated with an isotype-matched control ascites, and 4 of 6 mice given antibody-negative ascites. Insulitis was significantly reduced in the F23.1-treated group, and immunocytochemistry revealed the absence of Vβ8 expression on cells in the lymphoid organs and insulitis of these mice. This investigation revealed that Vβ8+ cells were implicated in CY-induced diabetes in NOD/ Wehi mice.
Beneficial Effect of Org 2766 in Treatment of Peripheral Neuropathy in Streptozocin-Induced Diabetic Rats
Authors: Van Der Zee Catherina E E M , Van Der Hoop Roland Gerritsen , Gispen Willem H
Published: September 2013

An injection of streptozocin (STZ) was used to study diabetes-induced peripheral neuropathy inrats. In such rats the values of motor nerve conduction velocity and sensory nerve conduction velocity were decreased compared with the values obtained in nondiabetic controls from 3 wk after STZ injection onward. In recent years it has been extensively documented that peptides related to ACTHand MSH exert a neurotrophic effect on the nervous system that results in enhanced recovery of function after mechanical nerve damage. This article documents the beneficial effect of the peptide Org 2766, an ACTH-(4–9) analogue, in diabetic peripheral neuropathy. Chronic subcutaneous treatmentof diabetic rats with Org 2766 results in a significant enhancement of both motor and sensory nerve conduction velocity compared with saline-treated diabetic rats. Histological analysis of cross sections of the sural nerve showed no difference in the total number of nerve fibers in saline- or peptide-treated diabetic rats. In contrast, a difference in fiber size distribution was demonstrated; i.e., the sural nerves of diabetic rats contained fewer thick myelinated fibers. Treatment with Org 2766 resulted in a normal distribution. Apparently, the peptide Org 2766 has a protective action on nerve fibers and nerve function during STZ-induced diabetes.
Effect of Cyclosporin on Interleukin 2–Related T-Lymphocyte Parameters in IDDM Patients
Authors: Chatenoud Lucienne , Feutren Gilles , Nelson David L , Boitard Christian , Bach Jean-Francois
Published: September 2013

Seventy patients aged 15–40 yr with recent-onset insulin-dependent diabetes mellitus (IDDM) were entered into a double-blind trial, in which they were randomly assigned to either cyclosporin (7.5 mg · kg−1 · day−1) or to placebo and were monitored for 1 yr for various phenotypic and functional parameters of T-lymphocyte-mediated immunity. Before treatment, the proportions of total T-lymphocytes (CD3+) and helper-inducer T-lymphocytes (CD4+) were normal, whereas significantly decreased values of suppressor/ cytotoxic T-lymphocytes (CD8+), as compared with normal controls, were found in 31% of the patients. The interleukin 2 (IL-2)—receptor expression was significantly increased in IDDM patients compared with control subjects, although the single values were low: patients, 2.02 ± 0.41%; controls, 0.88 ± 0.25% (means ± SE). Circulating levels of soluble IL-2 receptor were also significantly increased in IDDM patients compared with controls: patients, 372.3 ± 25.4 U/ml; controls, 235.5 ± 29.3 U/ml (means ± SE). However, no major abnormalities were found in mitogen (phytohemagglutinin)-induced IL-2 production, cell proliferation, or IL-2-receptor expression. After 6 mo of cyclosporin treatment, no major modifications of any of the parameters analyzed were noted, even in patients who had cyclosporin blood trough levels >300 ng/ml, i.e., the threshold value associated with clinical efficacy. One explanation for the absence of a major effect of cyclosporin, in contrast with its demonstrated clinical effectiveness, is thereversibility of its activity. Our results preclude the use of the described tests to reliably monitor IDDM patients undergoing immunosuppressive therapy.
Linkage Studies on NIDDM and the Insulin and Insulin-Receptor Genes
Authors: Cox Nancy J , Epstein Paul A , Spielman Richard S
Published: September 2013

Twenty Black families in which at least two siblings had non-insulin-dependent diabetes mellitus (NIDDM) were typed for restriction-fragment-length polymorphisms at the insulin (INS), insulin-receptor (INSR), and HLA-DRβ loci. Evidence for linkage between NIDDM and these loci was assessed with various genetic models for the transmission of NIDDM and with the affected-sib-pair approach, which does not require assumptions concerning a genetic model for NIDDM. Tight linkage between NIDDM and any of the loci was unlikely under all of the genetic models examined. Similarly, for all three of the loci, the distribution of affected sib pairs sharing 2, 1, or 0 genes identical by descent was not significantly different from (and was very similar to) that expected if the locus were unrelated to disease susceptibility. There was no evidence for linkage heterogeneity for any of the loci when families were grouped according to obesity or age at onset or when considering families individually. We conclude that the INS and INSR loci can be ruled out as major susceptibility loci for NIDDM in most Black families segregating this disorder, but we recognize that defects at either of these loci may cause or contribute to NIDDM in some patients. In addition, it is possible that variation at the INS and/or INSR loci may contribute to NIDDM susceptibility by modifying susceptibility due primarily to another major gene(s) or as part of an overall polygenic component to NIDDM.
Effect of Aldose Reductase Inhibition and Insulin Treatment on Retinal Capillary Basement Membrane Thickening in BB Rats
Authors: Chakrabarti Subrata , Sima Anders A F
Published: September 2013

This study compared the effect of glycemie control with insulin to that of the aldose reductase inhibitor ponalrestat on capillary basement membrane thickening in the retinas of diabetic BB rats. Diabetic animals with or without ponalrestat treatment were compared with diabetic rats subjected to vigorous insulin therapy or nondiabetic control rats with or without ponalrestat treatment after 6 mo of follow-up. Untreated diabetic animals showed the characteristic capillary basement membrane thickening in both the superficial and deep capillary beds of the retina. Vigorous blood glucose control with insulin therapy was accompanied by a complete prevention of capillary basement membrane thickening in both capillary beds, whereas aldose reductase inhibitor treatment achieved a complete prevention of basement membrane thickening in the deep capillary bed but not in the superficial capillary bed of the diabetic retina. These findings suggest that the mechanisms responsible for capillary basement membrane thickening in diabetes may be varied and modulated by topographical peculiarities in various capillary beds.
Anti-Islet Cell Antibodies From NOD Mice
Authors: Supon Patrick , Stecha Peter , Haskins Kathryn
Published: September 2013

We produced a panel of monoclonal autoantibodies from the spleen cells of prediabetic nonobese diabetic mice. The antibodies were selected on the basis of their ability to bind to the surface of nondiabetic mouse islet cells, and from >4000 hybridomas screened, 35 islet-reactive antibodies were isolated. Most of these reagents also bind to nondiabetic rat and human islet cells and β-cell tumor lines. A few of the antibodies were found to be reactive with insulin. When tested for cross-reactivity with other cell types, most of the antibodies were found to be islet specific.
Pattern of Postprandial Carbohydrate Metabolism and Effects of Portal and Peripheral Insulin Delivery
Authors: Kryshak Edward J , Butler Peter C , Marsh Christopher , Miller Alexander , Barr Darlene , Polonsky Kenneth , Perkins James D , Rizza Robert A
Published: September 2013

The importance of portal insulin delivery in the regulation of postprandial carbohydrate metabolism is uncertain. To address this question, three groups of dogs were studied: one group in which pancreatic venous drainage was transected and reanastomosed (portal insulin delivery), one in which the pancreatic drainage was transected and anastomosed to the inferior vena cava (peripheral insulin delivery), and one that received only a sham operation. Plasma insulin was greater (P < 0.05) during peripheral insulin delivery than in either the portal or sham groups, respectively, before and after meal ingestion. On the other hand, C-peptide concentrations did not differ between groups, resulting in a higher (P < 0.001) insulin to C-peptide ratio in the peripheral group. This indicated that the hyperinsulinemia in the peripheral group was due to decreased insulin clearance rather than increased insulin secretion. Isotopically determined splanchnic uptake of ingested glucose, postprandial suppression of hepatic glucose release, incorporation of CO2 into glucose (a qualitative measure of gluconeogenesis), and total-body glucose uptake were virtually identical in all groups. Similarly, plasma lipid, β-hydroxybutyrate, and lactate concentrations did not differ between groups. Our data indicate that, despite differences in systemic insulin concentration, portal and peripheral insulin delivery comparably regulate hepatic and extrahepatic carbohydrate metabolism after meal ingestion.
Increased Basal Glucose Production and Utilization in Nondiabetic First-Degree Relatives of Patients With NIDDM
Authors: Osei Kwame
Published: September 2013

To characterize the abnormalities in basal glucose homeostasis in people who are at increased risk for non-insulin-dependent diabetes mellitus (NIDDM), we measured the rates of basal hepatic glucose output (HGO), glucose disappearance, and metabolic clearance of glucose (MCR) in 27 nondiabetic first-degree relatives of NIDDM patients and 16 age-, sex-, and weight-matched healthy control subjects with no family history of NIDDM. Mean fasting plasma glucose was significantly lower (P < 0.05) in control subjects (mean ± SE 77 ± 2 mg/dl) than in relatives (84 ± 2 mg/dl). Mean basal insulin levels were not significantly different between relatives and control subjects (10.0 ± 1.5 vs. 7.7 ± 1.0 μU/ml). Mean basal HGO was significantly lower in control subjects compared with relatives (1.83 ± 0.07 vs. 2.20 ± 0.10 mg · kg−1 · min−1 P < 0.05). Mean MCR was similar in relatives (2.58 ± 0.12 mg · kg−1 · min−1) and control subjects (2.35 ± 0.09 mg · kg−1 · min−1). In summary, this study demonstrates that basal hepatic glucose production and glucose utilization are increased in glucose-tolerant first-degree relatives compared with healthy control subjects. We conclude that impaired basal hepatic glucose regulation rather than glucose disposal is present as an early defect in glucose-tolerant first-degree relatives of NIDDM patients.
Islet Cell Antibodies as Predictive Markers for IDDM in Children With High Background Incidence of Disease
Authors: Karjalainen Jukka K
Published: September 2013

To determine the prevalence and predictive value of islet cell antibodies (ICAs) for the development of insulin-dependent diabetes mellitus (IDDM) in 1212 Finnish children aged 3–18 yr, samples for ICA determination were taken in 1980, and subsequent analyses were performed in originally ICA+ children and in 296 initially ICA− children in 1983 and 1986. All 1212 subjects were followed for 8 yr for the development of IDDM. Fifty children (4.1%) were positive for conventional ICAs (IF-ICAs) in 1980 (range 3–80 Juvenile Diabetes Foundation units [JDF U]; median 30 JDF U), of which 12 (1.0%) had complementfixing ICAs (CF-ICAs) in their serums (range 3–30 JDF U; median 8 JDF U). None were exclusively CF-ICA+ Boys were CF-ICA+ more often than girls (9 of 563 [1.6%] vs. 3 of 649 [0.5%], respectively; P < 0.05). Over the next 6 yr, 4 of 39 subjects lost their IF-ICAs, and 4 of 12 lost their CF-ICAs without progressing to diabetes. The initial IF-ICA levels in these subjects were lower (range 3–8 JDF U; median 7 JDF U; P < 0.05) than those in the persistent cases. In the initially ICA− subgroup (n = 296), 7 subjects (2.4%) later became IF-ICA+, and 4 (1.4%) became CF-ICA+. The levels of ICA in these subjects were lower than in the originally ICA+ ones (P < 0.05), and 3 IF-ICA+ and 2 CF-ICA+ subjects again became ICA− before 1986. Three initially IF-ICA+ subjects (2 also with CF-ICA) presented with IDDM after 3 yr, compared to no cases among IF-ICA− children. High persistent ICA levels (≥18 JDF U) had the highest positive predictive value for the development of IDDM. The results reveal a high ICA prevalence in unaffected Finnish children. However, only high and persistent levels, especially when they are complement fixing, seem to predict the development of IDDM in healthy children.
Diabetes in NOD Mice Does not Require T Lymphocytes Expressing Vβ8 or Vβ5
Authors: McDuffie Marcia
Published: September 2013

The incidence of destructive pancreatic infiltrates and overt diabetes in animal models of insulin-dependent (type I) diabetes mellitus can be greatly reduced by inactivating or eliminating most T lymphocytes early in life. Because of theoretical and practical concerns about inducing long-term pan–T-lymphocyte inactivation for prevention or treatment of type I diabetes in humans, we hoped that more selective suppression of only the diabetogenic T lymphocyte population might be possible. To this end, two groups suggested that diabetogenic subpopulations of T lymphocytes in NOD mice could be identified by the protein sequence of their T-lymphocyte receptors. This assertion was based on experimental elimination of candidate T-lymphocyte subpopulations in two different short-term models of diabetes induction in NOD mice. For these experiments, identification and elimination of T-lymphocyte subsets were accomplished with monoclonal antibodies that bind specifically to the variable region of the β-chain (Vβ) of the T-lymphocyte antigen receptor and divide the T-lymphocyte pool of the NOD mouse into ∼20 Vβ subsets. To test the relationship between the two T-lymphocyte Vβ subsets implicated in these studies and pancreatic β-cell destruction in unmanipulated animals, both T-lymphocyte subpopulations identified were genetically eliminated from NOD-derived mice by introduction of a mutant T-lymphocyte receptor Vβ gene, from which these sequences are genomically deleted. Histological evidence of severe β-cell destruction and overt diabetes was found in mice homozygous for the deleted Vβ gene, indicating that neither Vβ gene segment identified in previous studies is required for diabetogenesis in unmanipulated diabetes-prone mice.
Pioglitazone hydrochloride inhibits cholesterol absorption and lowers plasma cholesterol concentrations in cholesterol-fed rats
Authors: Colca Jerry R , Dailey Charlene F , Palazuk Barbara J , Hillman Roberta M , Dinh Dac M , Melchior George W , Spilman Charles H
Published: September 2013

Diabetes is associated with altered cholesterol metabolism that may contribute to cardiovascular complications. Treatment of rats with pioglitazone hydrochloride, a novel antidiabetic compound that improves the general response of target cells to insulin, significantly lowered cholesterol levels in rats fed a hypercholesterolemic diet and produced a significant reduction in cholesterol absorption. Drug treatment was ineffective in rats that were not given dietary cholesterol. To determine whether these effects of pioglitazone hydrochloride might be related to the known ability of this compound to improve the response to circulating insulin, similar studies were conducted in streptozocin-induced diabetic rats with and without insulin replacement. Diabetic rats absorbed a greater percentage of dietary cholesterol than control rats. Treatment of insulin-deficient diabetic rats with pioglitazone alone did not affect cholesterol absorption; however, the combination of insulin and pioglitazone was synergistic to lower absorption of cholesterol and circulating cholesterol and triglycerides. Treatment of either normal rats or diabetic rats receiving insulin with pioglitazone hydrochloride produced a twofold decrease in the ratio of total cholesterol to high-density lipoprotein cholesterol. These results suggest that treatments that improve insulin sensitivity may also have a positive impact on coronary artery disease associated with diabetes.
Presence of Liver CGRP/Amylin Receptors in Only Nonparenchymal Cells and Absence of Direct Regulation of Rat Liver Glucose Metabolism by CGRP/Amylin
Authors: Stephens Thomas W , Heath William F , Hermeling Ronald N
Published: September 2013

Human calcitonin gene–related peptide (hCGRP-1) and human amylin (hA) have been reported to increase hepatic glucose output in vivo and to bind with high affinity to rat liver plasma membranes, resulting in increased cAMP production. These observations have led to the hypothesis that CGRP or amylin may be physiological regulators of liver glucose metabolism. Liver plasma membranes are derived from several cell types, including parenchymal (hepatocyte), Kupffer, endothelial, lipid storage, and smooth muscle cells. Because the parenchymal cell is responsible for the contribution of the liver to whole-body glucose homeostasis, it is important to verify the location and activity of the CGRP/amylin receptor to this cell. These studies separate liver cells prepared by collagenase digestion into parenchymal and nonparenchymal fractions by metrizamide gradient and differential centrifugation. 125I-labeled [Tyr-0]hCGRP-1 bound with high affinity to nonparenchymal cell fraction and was displaced by both hCGRP-1 and hA. hCGRP-1 bound with greater affinity than hA (Kd = 2.1 ± 1.6 × 10−11 vs. 2.6 ± 1.2 × 10−8 M) in a manner similar to the binding reported for liver plasma membrane fraction. Linear regression of receptor concentration against nonparenchymal cell count per milliliter was significant (r = 0.999, P = 0.026), leading to an estimate of 3000 receptors/cell. The parenchymal cell fraction bound very little 125I-[Tyr-0]hCGRP-1, and regression of receptor concentration against parenchymal cell count per milliliter was not significant (r = −0.708, P = 0.29), suggesting that binding was not due to parenchymal cells but instead to contamination by nonparenchymal cells. 125I-labeled rat amylin (125I-rA) binding gave similar results for the distribution of receptors but higher apparent affinity for hA (Kd = 2.0 ± 0.9 × 109 M). The following studies were then performed to determine the direct effect of CGRP and amylin on liver glucose metabolism. Of hCGRP-1, hA, human deamidated amylin, and rA, none alter glycogen synthesis (basal or insulin stimulated) or net glucose production (basal or glucagon stimulated) by isolated rat liver parenchymal cells. Glucagon, but not hCGRP-1 or hA, stimulated perfused liver glucose output. The lack of effect by hCGRP-1 and hA on isolated hepatocyte glucose metabolism is therefore not merely a result of the collagenase digestion procedure. These data indicate that CGRP/amylin receptors occur only on the nonparenchymal cells of rat liver and do not directly regulate liver glucose metabolism.
Oxygen Free Radical Effects in Sciatic Nerve in Experimental Diabetes
Authors: Low Phillip A , Nickander Kim K
Published: September 2013

We previously reported the presence of endoneurial hypoxia, ischemia, impairment of the blood-nerve barrier, and reduction of norepinephrine and 6-ketoprostaglandin F1α in chronic streptozocin-induced diabetic neuropathy (SDN) and interpreted these findings as suggesting the involvement of oxygen free radicals (OFRs) but did not directly measure indices of OFR activity. In this study, we report on sciatic nerve conjugated dienes, hydroperoxides, norepinephrine, and malondialdehyde in SDN at 1, 4, and 12 mo in male Sprague-Dawley rats. Severe hyperglycemia was present throughout in diabetic rats. Conjugated dienes were consistently increased at all time points, hydroperoxides were consistently reduced, and malondialdehyde was not significantly different in diabetes compared with controls. These findings are consistent with increased OFR activity in experimental diabetes. It is necessary to monitor several indices of OFR activity in a metabolically active tissue such as the peripheral nerve.
Liver and Kidney Tissue Membranes As Tissue Markers for Nonenzymatic Glycosylation
Authors: Cefalu William T , Wang Zhong Q , Bell-Farrow Audrey , Ralapati Suresh
Published: September 2013

We investigated the relationship of serum protein glycosylation to peripheral tissue membrane glycosylation. We studied 27 Sprague-Dawley rats and induced diabetes in 20 of them. Blood glucose levels were treated in 10 of the diabetic animals with daily subcutaneous insulin. After 8 wk, liver and kidney tissue was removed, purified membranes were prepared, and the percentage of glycosylated membrane protein was determined for the liver and kidney membranes by boronate-affinity methods. The percentage of glycosylated membrane protein for both liver and kidney tissue was found to correlate significantly with the glycemic state of the animal as assessed with glycosylated serum albumin, total glycosylated serum proteins, and glycosylated hemoglobin determinations (P < 0.001 for each glycosylated protein parameter). In addition, the percentage of glycosylated membrane protein in the liver tissue correlated significantly with the measured level in the corresponding kidney tissue (r = 0.78, P < 0.001). To identify the nature of the glycosylated membrane proteins, boronate-affinity methods were used to separate the glycosylated and nonglycosylated membrane proteins. It was determined that two major glycosylated protein bands exist for the liver membrane (78,000 and 58,700 Mr) and four for the kidney membranes (ranging from 48,700 to 74,000 Mr). The ultrastructural location and identification of these glycosylated membrane proteins are not known. This study demonstrates that measurement of clinical glycemic state, as reflected in glycosylated blood protein parameters such as glycosylated serum albumin and glycosylated hemoglobin, correlates significantly with ongoing tissue membrane accumulation of glucose.
Prognostically Significant Heterogeneity of Cytoplasmic Islet Cell Antibodies in Relatives of Patients with Type I Diabetes
Authors: Gianani Roberto , Pugliese Alberto , Bonner-Weir Susan , Shiffrin Alicia J , Soeldner J Stewart , Erlich Henry , Awdeh Zuhier , Alper Chester A , Jackson Richard A , Eisenbarth George S
Published: September 2013

A significant proportion of relatives of patients with insulin-dependent (type I) diabetes with high titers of cytoplasmic islet cell autoantibodies (ICAs) do not progress to overt diabetes with up to 8 yr of follow-up. This may reflect that follow-up of such relatives has not been long enough to observe diabetes, that despite expression of identical ICAs, some relatives will not progress to diabetes; or that there is heterogeneity in what is identified as ICA. We identified a subset of ICA that was restricted in its species (not reacting with mouse islets) and cell-type reactivity within islets (β-cell specific). Only one of eight relatives whose sera had the restricted pattern of reactivity progressed to overt diabetes, and on sequential evaluation, all but the one relative who progressed to diabetes have maintained normal first-phase insulin secretion to intravenous glucose. In contrast, by life-table analysis, 70% of relatives expressing nonrestricted ICA became diabetic within 5 yr of follow-up (1 of 8 vs. 16 of 25 diabetic at last follow-up, P < 0.02). Moreover, preliminary data suggest a significant association of the human leukocyte antigen DQB1*0602 allele of DR2 haplotypes with the restricted ICA pattern (4 of 5 DQB1*0602 restricted vs. 0 nonrestricted ICA, P = 0.006). We propose that expression of a genetically determined restricted ICA pattern confers a markedly lower risk for progression to diabetes. Our studies suggest that relatives with restricted ICA may comprise most ICA+ relatives who do not develop diabetes or abnormal first-phase insulin secretion on follow-up of >5 yr and that expression of such autoantibodies may be associated with the ‘protective’ DQB1*0602 allele.
Estimation of Insulin Secretion Rates from C-Peptide Levels: Comparison of Individual and Standard Kinetic Parameters for C-Peptide Clearance
Authors: Cauter Eve Van , Mestrez Fabienne , Sturis Jeppe , Polonsky Kenneth S
Published: September 2013

Insulin secretion rates can be accurately estimated from plasma C-peptide levels with a two-compartment model for C-peptide distribution and degradation. In previous studies, the kinetic parameters of C-peptide clearance were derived in each subject from the decay curve observed after bolus intravenous injection of biosynthetic human C-peptide. To determine whether standard parameters for C-peptide clearance could be defined and used to calculate insulin secretion without obtaining a decay curve in each subject, we analyzed 200 decay curves of biosynthetic human C-peptide obtained in normal, obese, and non-insulindependent diabetes mellitus subjects studied in ourlaboratory. This analysis showed that the volume of distribution and kinetic parameters of C-peptide distribution and metabolism vary by <30% in a population highly heterogenous in terms of age, sex, degree of obesity, and degree of glucose tolerance. The volume of distribution correlated with the degree of obesity as quantified by body surface area (BSA). This dependence of C-peptide distribution volume on BSA was more marked in men than in women. The long half-life was slightly longer in elderly subjects than in younger adults. When effects of BSA, sex, and age were taken into account, the parameters of C-peptide kinetics were very similar in normal, obese, and diabetic subjects. Based on these findings, a simple procedure to derive standard parameters for C-peptide clearance taking into account degree of obesity, sex, and age was defined. These standard parameters resulted in estimations of mean insulin secretion rates, which differed in each subject by only 10-12% from those obtained with individual parameters. The approach of using standard ratherthan individual parameters did not systematically underestimate or overestimate insulin secretion so that group values for the fasting secretion rate, the mean 24-h secretion rate, and the number and the amplitude of secretory pulses obtained with standard parameters differed by only 1–2% from the values obtained with individual parameters. Furthermore, the accuracy of measurements based on standard parameters was not different from that associated with replicate determinations of the parameters of C-peptide clearance in the same subject. We conclude that it is possible to estimate insulin secretion rates from plasma C-peptide levels with standard parameters for C-peptide clearance rather than individually derived parameters without significant loss of accuracy.
Structure and Segmental Localization of Glycogen in the Diabetic Rat Kidney
Authors: Holck Palle , Rasch Ruth
Published: September 2013

Pathological accumulations of glycogen were studied in the kidney tubular epithelium in untreated STZ-induced diabetic rats of 50 days diabetes duration. Blood glucose concentrations were ∼17 mM, and the animals had no ketonuria. At the termination of the experiment, the kidneys were perfusion-fixed, and serial sections were cut from the renal capsule to the tip of the papilla and stained with toluidine blue and periodic acid Schiff. By tracing tubular profiles from section to section in a light microscope, the outlines of nephrons were reconstructed, and abnormal glycogen accumulations were mapped in accordance with the outlines. The exact segmental localization, character, and extension of the glycogen accumulations were determined. The predominant location of the pathological glycogen accumulations was in the thick ascending limb of Henle's loop. Dot-shaped and diffuse-appearing glycogen accumulations were discretely distributed throughout the segment, and large confluent cytoplasmic accumulations of glycogen were also present. On a continuous basis, glycogen was present only in the cortical thick ascending limb of Henle's loop and the macula densa segment excluding the macula densa cells. In the distal convoluted tubule and the cortical collecting duct system, scattered dot-shaped and diffuse glycogen accumulations were discretely distributed. Furthermore, glycogen appeared as confluent, cytoplasmic accumulations in the initial part of the descending thin limb of Henle's loop. In addition, glycogen accumulations were found in nuclei within a narrow stripe of the outer stripe of the outer medulla in the medullary thick ascending limb of Henle's loop.
Arginine Vasopressin and Oxytocin Effects in Mouse Pancreatic β-cells: Receptors Involved in Stimulation of Insulin Release
Authors: Gao Zhi-Yong , Henquin Jean-Claude
Published: September 2013

Recently AVP and OT were suggested to increase insulin release by stimulating phosphoinositide metabolism in pancreatic β-cells. In this study, mouse islets were used to identify the receptors involved in the effects of AVP and OT. Both agents caused concentration-dependent (0.1 nM to 10 microM) increases in insulin release and IP levels. The potencies of AVP and OT were similar, and their maximal effects were not additive. Synthetic agonists selective for AVP (V1) and OT receptors in other tissues were equipotent with the natural agonists on insulin release and almost equipotent on IPs. Antagonists selective for V1 and OT receptors inhibited both types of effects with a potency that varied with the nature of the agonist. Both AVP and OT were ∼10-fold potent on insulin release (EC50 ≈2 nM) than on IPs (EC50 ≈25 nM). Moreover, the antagonists more readily inhibited the effects of all agonists on IPs than on insulin release. Pancreatic β-cells may possess either a single type of receptor that is distinct from classic V1 or OT receptors, but is closer to the latter, or both V1 and OT receptors coupled to common effectors. The acceleration of phosphoinositide turnover might not be the sole mechanism involved in the stimulation of insulin release by AVP and OT.
Increased Expression of T-Cell Markers of Immunological Memory Associated with Protection from Type I Diabetes: A study of Identical Twins
Authors: Peakman Mark , Alviggi Lorenzo , Hussain Munther J , Lo Simon S S , Hawa Mohammed , Leslie R David G , Vergani Diego
Published: September 2013

Disturbances in the balance of CD4+ helper T-lymphocytes expressing the surface molecules CD45RA and CD45R0, which define naive and memory populations, respectively, are present at diagnosis of type I diabetes. In a prospective study over 10 years, these subsets were analyzed in samples obtained from 18 identical twins of patients with type I diabetes, 8 of whom became diabetic (prediabetic twins), whereas the rest remained nondiabetic after at least 8 years follow-up and are now unlikely to develop the disease (diabetes-protected twins). At the beginning of the study, percentage levels of naive (CD45RA+) CD4+ lymphocytes were significantly elevated in prediabetic twins compared with diabetes-protected twins (P < 0.05) and remained so throughout the study (P < 0.01). Percentage levels of naive cells in diabetes-protected twins were significantly reduced compared with control subjects both at the beginning and throughout the study (P < 0.05, P < 0.01, respectively). In contrast, diabetes-protected twins at the beginning of the study had elevated percentage levels of memory (CD45R0+) CD4+ lymphocytes that persisted throughout the study compared with prediabetic twins (P < 0.05 for both). Percentage levels of memory cells in prediabetic twins were significantly reduced compared with control subjects both at the beginning and throughout the study (P < 0.01, P < 0.05, respectively). Increased percentage levels of a population of CD4+ lymphocytes coexpressing CD45RA and CD45R0 were seen in both twin groups compared with control subjects at entry into and during the study (P < 0.05 for all), but persisted only in the prediabetic twins. In this study, we identify for the first time a distinct immune abnormality—overexpression of memory helper T-lymphocytes—that is present in those twins who, despite genetic susceptibility to type I diabetes, do not develop the disease.
Time Course for Effects of Hypoglycemia on Insulin Gene Transcription In Vivo
Authors: Shalwitz Robert A , Herbst Todd , Carnaghi Lynn R , Giddings Stephen J
Published: September 2013

The purpose of these studies was to determine the time course for onset of effects of hypoglycemia on insulin gene transcription in vivo. Using insulin infusions, we found that insulin-induced hypoglycemia decreased levels of precursors for insulin mRNA, reflecting changes in new mRNA synthesis, to new steady-state values within 100 min. These changes were followed by declines in processed insulin mRNA. An alternate infusion technique was developed to lower plasma glucose levels from a constant level of 120–130 to 50–60 mg/dl in <10 min without changing insulin levels from those maintained during a preceding 1-h control period. Using this protocol, we found that levels of precursors for insulin mRNA remained constant for the first 20 min of hypoglycemia, then decreased rapidly at 40 and 60 min. The initial delay followed by rapid decline suggests that the decrease of insulin gene transcription in response to hypoglycemia is an active process requiring one or more inductive events before implementation.
Mechanisms of Increased Skeletal Muscle Glucose Transport Activity After an Oral Glucose Load in Rats
Authors: Napoli Raffaele , Hirshman Michael F , Horton Edward S
Published: September 2013

It is not known whether the insulin-induced changes in the skeletal muscle glucose transport system occur under physiological circumstances. To clarify whether, by which mechanisms, and for howlong skeletal muscle glucose transport activity is increased after an oral glucose load (OGL), we prepared plasma membrane (PM) and microsomal membrane (MsM) vesicles from hindlimb muscles of Sprague-Dawley rats either in the fasting state or 30, 60, 90, or 120 min after an OGL (2 g/kg body wt). In both PM and MsM, we measured the total number of glucose transporters (Ro), GLUT4, and GLUT1. In the PM, we also determined glucose influx (Vmax) and carrier turnover number (TN), an index of average transporter intrinsic activity, (TN = Vmax/Ro). The Vmax significantly increased after OGL, was maximal at 30 min, and returned to baseline at 90 min. The Ro and GLUT4 in the PM also increased significantly, with themaximum level reached at 60 min. The TN was increased only at 30 min. The changes in Ro andGLUT4 in the MsM were opposite to those in the PM, consistent with translocation of GLUT4 from an intracellular pool to the PM. In conclusion, an OGL induces an increase in the skeletal muscle glucose transport activity. This increase is associated with the translocation of GLUT4 from the MsM to the PM and a more transient increase in the average transporter TN. Our results show that transporter translocation and activation occur under physiological circumstances.
Role for Transforming Growth Factor β in Thromboxane-Induced Increases in Mesangial Cell Fibronectin Synthesis
Authors: Negrete Hilmer , Studer Rebecca K , Craven Patricia A , DeRubertis Frederick R
Published: September 2013

Thromboxane A2 (TXA2) has been implicated in the pathogenesis of progressive glomerulosclerosis and stimulates the synthesis of matrix protein by mesangial cells (MCs). This study examined the role of transforming growth factor β (TGF-β) in the mediation of the action of the stable TXA2/prostaglandin (PG) endoperoxide analog U-46619 to stimulate fibronectin (Fn) synthesis in cultured rat MC. Exogenous TGF-β increased Fn synthesis by MC in a concentration- and time-dependent fashion, as reflected by incorporation of [35S]methionine into immunoprecipitable Fn. Submaximal concentrations of TGF-β (1–2.5 pmol/l) increased Fn synthesis two- to threefold, a response comparable in magnitude to that observed with a maximal stimulatory concentration (1 μmol/l) of U-46619. Anti-TGF-β antibody, but not isotypic IgG, blocked the increases in Fn synthesis induced by both U-46619 and exogenous TGF-β. Endogenous TGF-β bioactivity in MC culture media, assessed by the mink lung epithelial cell system, was significantly increased by 1 μmol/l U-46619 (1.7 ± 0.3 pmol/l) compared with that of control media (0.6 ± 0.1 pmol/l, P < 0.05). Total (active plus latent) TGF-β bioactivity, assayed after heat activation of latent TGF-β, was also significantly higher in media of MCs cultured with U-46619 (45 ± 4 pmol/l) compared with control (24 ± 4 pmol/l). Thus, U-46619 increased endogenous TGF-β bioactivity to a level sufficient to account for the enhancement of Fn synthesis observed with U-46619, as reflected by the Fn synthetic response to exogenous TGF-β. This finding, together with the inhibitory effect of anti-TGF-β antibody on Fn synthesis, implicates TGF-β in expression of this action of U-46619.
An (A-C)n Dinucleotide Repeat Polymorphic Marker at the 5′ End of the Aldose Reductase Gene Is Associated With Early-Onset Diabetic Retinopathy in NIDDM Patients
Authors: Karen Ben C B , Lam Karen S L , Wat Nelson M S , Chung Stephen S M
Published: September 2013

To study the relationship between the aldose reductase gene and diabetic complications, an (A-C)n dinucleotide repeat sequence 2.1 kb upstream of the transcription start site of this gene was identified and studied. There are seven alleles at this locus with a polymorphism information content of 0.73 and a heterozygosity of 0.77 among the Chinese population in Hong Kong. One of the alleles (Z–2) was found to be associated with early onset of retinopathy in patients with non-insulin-dependent diabetes (P = 0.007), suggesting that aldose reductase or a gene in the close vicinity may be involved in the pathogenesis of this diabetic complication.