IFNγ and perforin cooperate to control infection and prevent fatal pathology during persistent gammaherpesvirus infection in mice.

Infection with murine gammaherpesvirus 68 has become an accepted model for studying the virus/host interactions with regard to gammaherpesvirus infections. Previous studies using gene-deficient mice have revealed that neither IFNγ nor perforin is essential in controlling the outcome of infection or the virus load during chronic infection in C57BL/6 mice. However, pronounced multiorgan fibrosis and splenic atrophy are observed in mice lacking IFNγ or the IFNγ receptor. To study the interplay between perforin and IFNγ in controlling the virus-induced pathology and the viral load during chronic gammaherpesvirus infection, we infected IFNγ/perforin double-deficient C57BL/6 mice and followed the course of infection. While absence of perforin prevented the splenic atrophy in IFNγ-deficient mice, fibrosis did not disappear. Moreover, double-deficient mice developed extreme splenomegaly, were unable to control the viral load and displayed chronic immune activation. Thus, IFNγ and perforin act in concert to minimize pathology and control the viral load in mice chronically infected with MHV68. Furthermore, while certain aspect of the virus-induced pathology in IFNγ-deficient mice may be alleviated in double-deficient mice, other aspects are exaggerated, and the normal architecture of the spleen is completely destroyed. We believe that these findings add to the understanding of the virus/host interaction during chronic gammaherpes virus infection.

Long-term exendin-4 treatment delays natural deterioration of glycaemic control in diabetic Goto-Kakizaki rats.

AIM: The glucagon-like peptide-1 (GLP-1) receptor agonist, exendin-4, has previously been shown to delay the onset of diabetes when administered to Goto-Kakizaki (GK) rats in the prediabetic period. The present study aimed to evaluate whether long-term administration of exendin-4 to GK rats in the diabetic period would improve their diabetes and how glycaemic control was affected following drug wash-out. METHODS: Glycaemic control was assessed in diabetic GK rats during 12 weeks of exendin-4 or vehicle treatment. Moreover, some animals were followed for an additional 9 weeks without treatment. RESULTS: Glycaemic control was seen to deteriorate in vehicle-treated animals, as assessed by increased glycated haemoglobin A1c (HbA1c), whereas HbA1c improved in exendin-4-treated animals. Following an additional 9 weeks without treatment, glycaemic control in exendin-4-treated animals remained below baseline value and thus remained significantly lower than that of vehicle-treated animals. Following exendin-4 administration, oral glucose tolerance tests revealed greatly reduced glucose and insulin excursions compared with vehicle-treated animals, whereas following overnight drug wash-out, only little difference was seen, suggesting that the improvement in glycaemic control may have been obtained primarily by increased postprandial control. No significant differences were observed in pancreatic islet morphology or islet hormone content. CONCLUSIONS: Exendin-4 treatment improved glycaemic control in diabetic GK rats, independent of changes in beta-cell mass. Additionally, progression of the disease seemed to be delayed because the improvement in HbA1c was still apparent 9 weeks after cessation of treatment.

Targets and tactics: the relative importance of HbA, fasting and postprandial plasma glucose levels to glycaemic control in type 2 diabetes.

BACKGROUND: The incidence of type 2 diabetes is reaching pandemic proportions, impacting patients and healthcare systems across the globe. Evidence suggests that a majority of patients are not achieving recommended blood glucose targets resulting in an increased risk of micro- and macro-vascular complications. AIM: To review literature on the significance of glycosylated haemoglobin (HbA(1c)), fasting plasma glucose (FPG) and postprandial plasma glucose (PPG), their inter-relationships and relative importance in the treatment of diabetes, and to provide practical guidance on effective monitoring of patients. METHODS: Clinical guidelines on diabetes management and clinical and preclinical studies of glycaemic control identified through a publications database search were reviewed. RESULTS: Glycaemic control remains fundamental to the successful management of diabetes. HbA(1c) is the gold standard measure of glycaemic control but recent evidence suggests that postmeal hyperglycaemia also plays an important role in the aetiology of diabetes-associated complications and control of PPG levels is vital to the achievement of recommended HbA(1c) targets. CONCLUSIONS: The call for action on type 2 diabetes has never been more compelling; with a clear focus on strategies for glycaemic control, the impact of the diabetes pandemic can be limited.

Proglucagon products in plasma of noninsulin-dependent diabetics and nondiabetic controls in the fasting state and after oral glucose and intravenous arginine.

We investigated the major products of proglucagon (PG) processing in plasma in the fasting state, after intravenous arginine and after an oral glucose load in noninsulin-dependent diabetics (NIDDM) and in weight matched controls using specific radioimmunoassays and analytical gel filtration. In the fasting state the glucagonlike peptide-1 (GLP-1) immunoreactivity was significantly elevated in the NIDDM group compared with the control group. Both after intravenous arginine and after an oral glucose load a rise in the plasma concentrations of all immunoreactive moieties measured was seen. All integrated incremental responses after intravenous arginine were identical in the two groups. After oral glucose the insulin concentrations in plasma were lower and the concentrations of all proglucagon products were higher in the NIDDM group compared to the control group. The gel filtration analysis showed that arginine stimulated the secretion of pancreatic glucagon (PG 33-61), major proglucagon fragment (PG 72-158) and probably GLP-1 (PG 72-107 amide) in both groups, whereas oral glucose stimulated the secretion of glicentin (PG 1-69) and intestinal GLP-1 (PG 78-107 amide), an insulinotropic hormone. The elevated levels of immunoreactive GLP-1 in diabetics in the fasting state were mainly due to an increased concentration of major proglucagon fragment.

Preserved incretin activity of glucagon-like peptide 1 [7-36 amide] but not of synthetic human gastric inhibitory polypeptide in patients with type-2 diabetes mellitus.

In type-2 diabetes, the overall incretin effect is reduced. The present investigation was designed to compare insulinotropic actions of exogenous incretin hormones (gastric inhibitory peptide [GIP] and glucagon-like peptide 1 [GLP-1] [7-36 amide]) in nine type-2 diabetic patients (fasting plasma glucose 7.8 mmol/liter; hemoglobin A1c 6.3 +/- 0.6%) and in nine age- and weight-matched normal subjects. Synthetic human GIP (0.8 and 2.4 pmol/kg.min over 1 h each), GLP-1 [7-36 amide] (0.4 and 1.2 pmol/kg.min over 1 h each), and placebo were administered under hyperglycemic clamp conditions (8.75 mmol/liter) in separate experiments. Plasma GIP and GLP-1 [7-36 amide] concentrations (radioimmunoassay) were comparable to those after oral glucose with the low, and clearly supraphysiological with the high infusion rates. Both GIP and GLP-1 [7-36 amide] dose-dependently augmented insulin secretion (insulin, C-peptide) in both groups (P < 0.05). With GIP, the maximum effect in type-2 diabetic patients was significantly lower (by 54%; P < 0.05) than in normal subjects. With GLP-1 [7-36 amide] type-2 diabetic patients reached 71% of the increments in C-peptide of normal subjects (difference not significant). Glucagon was lowered during hyperglycemic clamps in normal subjects, but not in type-2 diabetic patients, and further by GLP-1 [7-36 amide] in both groups (P < 0.05), but not by GIP. In conclusion, in mild type-2 diabetes, GLP-1 [7-36 amide], in contrast to GIP, retains much of its insulinotropic activity. It also lowers glucagon concentrations.

A processing enzyme cleaving avian progastrin at post-Phe bonds.

Neuroendocrine peptides mature partly through endoproteolytic processing of long precursor forms. Best characterised is cleavage at mono- and dibasic residues, but additional sites also exist. Among these is post-Phe cleavage, first suggested to participate in the processing of chicken progastrin. In order to characterise this new mechanism, antibodies recognising the processing products of post-Phe cleavage of chicken progastrin were produced for radioimmunoassay measurements and immunocytochemistry. High concentrations of the carboxyamidated C-terminus and the N-terminus of gastrin-53 were measured in extracts of the antrum. In addition, significant amounts were detected using an assay specific for the N-terminus of gastrin-30 and with another assay for the C-terminus of the corresponding peptide, gastrin-53(1-23), obtained after cleavage at the Phe(23)-Ala(24) bond of gastrin-53. Colocalisation in antral G-cells of the N-termini of gastrin-53 and gastrin-30 and of the C-terminus of gastrin-53(1-23) was confirmed by immunohistochemistry. Finally, we identified the intact N-terminal 1-23 fragment of gastrin-53 complementary to gastrin-30, verifying endoproteolytic cleavage at the Phe(23)-Ala(24) bond. Taken together, the results support the existence of vertebrate endoprotease cleaving hormone precursors at post-Phe sites.

Glucagonlike peptide-I-(7-36)-amide receptors only in islets of Langerhans. Autoradiographic survey of extracerebral tissues in rats.

We demonstrated specific binding of the insulin-releasing hormone glucagonlike peptide (GLP)-I-(7-36)-amide, an intestinal product of proglucagon, to pancreatic islet cells by autoradiography using 125I-labeled GLP-I-(7-36)-amide incubated with tissue specimens of extracerebral rat organs. We also found binding of 125I-GLP-I to insulin-, glucagon-, and somatostatin-immunoreactive cells by combined autoradiographic and immunohistochemical analysis of pancreatic specimens using antisera against insulin, glucagon, and somatostatin. An accumulation of radioactivity was also observed in the stomach surface epithelium but could not be prevented by excess unlabeled peptide. No binding was found in other tissues investigated, including the lungs and the small intestinal mucosa. Localization of the binding sites identifies the pancreatic islets as the prime target for GLP-I-(7-36)-amide and suggests that it exerts direct effects on islet cells.

Biological effects and metabolic rates of glucagonlike peptide-1 7-36 amide and glucagonlike peptide-1 7-37 in healthy subjects are indistinguishable.

The biological effects and the metabolism of the intestinal hormone glucagonlike peptide-1 7-36 amide and glucagonlike peptide-1 7-37 were studied in normal healthy subjects. GLP-1 7-36 amide and GLP-1 7-37 equipotently stimulated insulin secretion (integrated hormone response 0-60 min, 631 +/- 211 vs. 483 +/- 177 pmol/h x L-1) and C-peptide secretion (integrated hormone response 9064 +/- 1804 vs. 9954 +/- 2031 pmol/h x L-1) and equipotently lowered plasma glucose (integrated decrease 48.3 +/- 5.7 vs. 46.2 +/- 8.4 mmol/h x L-1) and plasma glucagon (integrated decrease 80.4 +/- 24.3 vs. 156.0 +/- 34.6 pmol/h x L-1). Both GLP-1 7-36 amide and GLP-1 7-37 lowered the plasma concentration of free fatty acids significantly. The plasma half-lives of GLP-1 7-36 amide and GLP-1 7-37 were 5.3 +/- 0.4 vs. 6.1 +/- 0.8 min, and the metabolic clearance rates of the two peptides also were similar (14.6 +/- 2.4 vs. 12.2 +/- 1.0 pmol/kg x min). In conclusion, COOH-terminal amidation is neither important for the metabolism of GLP-1 nor for its effects on the endocrine pancreas.

Tissue and plasma concentrations of amidated and glycine-extended glucagon-like peptide I in humans.

Using specific radioimmunoassays, we studied the occurrence of amidated and glycine-extended glucagon-like peptide I (GLP-I) molecules in the human small intestine and pancreas and in the circulation system in response to a breakfast meal. Through gel permeation chromatography of extracts of the human pancreas (n = 5), we found that 71% of the GLP-I immunoreactivity eluted as a large molecule corresponding to the major proglucagon fragment, 24% corresponded to GLP-I 1-36 amide, and 5% to GLP-I 1-37. By gel permeation chromatography of extracts of human small intestine (n = 6), we found that all immunoreactivity eluted in one peak at the common elution position of the two insulin-releasing peptides, GLP-I 7-36 amide and GLP-I 7-37. Of the GLP-I immunoreactivity, 80% corresponded to GLP-I 7-36 amide and 20% to GLP-I 7-37. The mean concentrations of amidated GLP-I and glycine-extended GLP-I in fasting plasma were 7 +/- 1 and 6 +/- 1 pM, respectively (n = 6). In response to a breakfast meal, the concentration of amidated GLP-I rose significantly amounting to 41 +/- 5 pM 90 min after the meal ingestion, whereas the concentration of glycine-extended GLP-I only rose slightly to a maximum of 10 +/- 1 pM. Thus, both amidated and glycine-extended GLP-I molecules are produced in the small intestine and in the pancreas in humans. Both amidated and glycine-extended GLP-I are measurable in fasting plasma.(ABSTRACT TRUNCATED AT 250 WORDS)

Glucagon-like peptide I receptors in the subfornical organ and the area postrema are accessible to circulating glucagon-like peptide I.

The intestinal incretin hormone glucagon-like peptide I (GLP-I) inhibits gastric motility and secretion in normal, but not in vagotomized subjects, pointing to a centrally mediated effect. Therefore, our aim was to study the availability of rat brain GLP-I receptors to peripherally injected 125I-labeled GLP-I. The specificity of the binding was tested by co-injection of excess amounts of unlabeled GLP-I. Using light microscopical autoradiography of rat brain sections, we found specific 125I-GLP-I binding exclusively in the subfornical organ and the area postrema. This binding was abolished when an excess amount of unlabeled GLP-I was co-injected with the labeled GLP-I. We conclude that cells in the subfornical organ and the area postrema could be responsive to blood-borne GLP-I. The observed binding of peripherally administered GLP-I to the subfornical organ and the area postrema, which both have close neuroanatomical connections with hypothalamic areas involved in water and appetite homeostasis, is consistent with the potential roles of circulating GLP-I in the central regulation of appetite and autonomic functions.

Constitutive activity of glucagon receptor mutants.

Increased constitutive activity has been observed in the PTH receptor in association with naturally occurring mutations of two residues that are conserved between members of the glucagon/vasoactive intestinal peptide/calcitonin 7TM receptor family. Here, the corresponding residues of the glucagon receptor, His178 and Thr352, were probed by mutagenesis. An elevated level of basal cAMP production was observed after the exchange of His178 into Arg, but not for the exchange into Lys, Ala, or Glu. However, for all of these His178 substitutions, an increased binding affinity for glucagon was observed [dissociation constant (Kd) ranging from 1.1-6.4 nM, wild type: Kd = 12.0 nM]. A further increase in cAMP production was observed for the [H178R] construct upon stimulation with glucagon, albeit the EC50 surprisingly was increased approximately 10-fold relative to the wild-type receptor. Substitution of Thr352, located at the intracellular end of transmembrane segment VI, with Ala led to a slightly elevated basal cAMP level, while the introduction of Pro or Ser at this position affected rather the binding affinity of glucagon or the EC50 for stimulation of cAMP production. The large extracellular segment, which is essential for glucagon binding, was not required for constitutive activation of the glucagon receptor as the introduction of the [H178R] mutation into an N-terminally truncated construct exhibited an elevated basal level of cAMP production. The analog des-His1-[Glu9]glucagon amide, which in vivo is a glucagon antagonist, was an agonist on both the wild-type and the [H178R] receptor and did not display any activity as an inverse agonist. It is concluded that the various phenotypes displayed by the constitutively active glucagon receptor mutants reflect the existence of multiple agonist-preferring receptor conformers, which include functionally active as well as inactive states. This view agrees with a recent multi-state extension of the ternary complex model for 7TM receptor activation.

Glucagonostatic actions and reduction of fasting hyperglycemia by exogenous glucagon-like peptide I(7-36) amide in type I diabetic patients.

OBJECTIVE: Glucagon-like peptide I(7-36) amide (GLP-1) is a physiological incretin hormone that, in slightly supraphysiological doses, stimulates insulin secretion, lowers glucagon concentrations, and thereby normalizes elevated fasting plasma glucose concentrations in type II diabetic patients. It is not known whether GLP-1 has effects also in fasting type I diabetic patients. RESEARCH DESIGN AND METHODS: In 11 type I diabetic patients (HbA1c 9.1 +/- 2.1%; normal, 4.2-6.3%), fasting hyperglycemia was provoked by halving their usual evening NPH insulin dose. In random order on two occasions, 1.2 pmol . kg-1 . min-1 GLP-1 or placebo was infused intravenously in the morning (plasma glucose 13.7 +/- 0.9 mmol/l; plasma insulin 26 +/- 4 pmol/l). Glucose (glucose oxidase method), insulin, C-peptide, glucagon, GLP-1, cortisol, growth hormone (immunoassays), triglycerides, cholesterol, and nonesterified fatty acids (enzymatic tests) were measured. RESULTS: Glucagon was reduced from approximately 8 to 4 pmol/l, and plasma glucose was lowered from 13.4 +/- 1.0 to 10.0 +/- 1.2 mmol/l with GLP-1 administration (plasma concentrations approximately 100 pmol, P < 0.0001), but not with placebo (14.2 +/- 0.7 to 13.2 +/- 1.0). Transiently, C-peptide was stimulated from basal 0.09 +/- 0.02 to 0.19 +/- 0.06 nmol/l by GLP-1 (P < 0.0001), but not by placebo (0.07 +/- 0.02 to 0.07 +/- 0.02). There was no significant effect on nonesterified fatty acids (P = 0.34), triglycerides (P = 0.57), cholesterol (P = 0.64), cortisol (P = 0.40), or growth hormone (P = 0.53). CONCLUSIONS: Therefore, exogenous GLP-1 is able to lower fasting glycemia also in type I diabetic patients, mainly by reducing glucagon concentrations. However, this alone is not sufficient to normalize fasting plasma glucose concentrations, as was previously observed in type II diabetic patients, in whom insulin secretion (C-peptide response) was stimulated 20-fold.

Effect of truncated glucagon-like peptide-1 [proglucagon-(78-107) amide] on endocrine secretion from pig pancreas, antrum, and nonantral stomach.

We studied the effect of truncated glucagon-like peptide-1 [naturally occurring GLP-1; proglucagon-(78-107) amide], a potent insulinotropic peptide from the pig ileum, on endocrine and exocrine secretion of potential gastrointestinal target organs using isolated perfused preparations of the porcine pancreas, antrum, and nonantral part of the stomach. Truncated GLP-1 significantly increased somatostatin secretion from the pancreas at 10(-10) mol/liter and more than doubled the secretion at 10(-9) mol/liter, but had no effect on either somatostatin or gastrin secretion from the antrum or on somatostatin secretion from the nonantral stomach in concentrations up to 10(-8) mol/liter. Insulin secretion from the pancreas (with 7 mmol/liter glucose in the perfusate) increased 2-fold with truncated GLP-1 at 10(-10) mol/liter and almost 5-fold at 10(-9) mol/liter. Pancreatic glucagon secretion was inhibited by 50% at 10(-10) mol/liter and by 70-80% at 10(-9) mol/liter. Full-length GLP-1 [proglucagon-(72-107)] and GLP-2 [proglucagon-(126-159)] had no effect on hormone secretion from any of the perfused organs. It is concluded that truncated GLP-1 may participate in an entero-insular control of pancreatic endocrine secretion.

Short-term insulin treatment prevents the diabetogenic action of streptozotocin in rats.

Streptozotocin, which induces diabetes mellitus in experimental animals, has been reported to be taken up by beta-cells by means of the glucose transporter 2 (GLUT2) and then reduce the cellular level of NAD+, leading to necrosis of the beta-cells. We investigated the effect of insulin pretreatment on the diabetogenic action of streptozotocin (60 mg/kg). Four groups of rats were studied: 1) a group that received streptozotocin (STZ), 2) a group that received insulin pretreatment and streptozotocin (INS + STZ), 3) a group that received insulin (INS), and 4) a control group (CTRL). Insulin treatment reduced the beta-cell immunoreactivity (IR) of insulin and GLUT2, which, thus, was reduced in INS + STZ rats at the time of streptozotocin injection. In STZ rats, plasma insulin concentrations after 3 weeks as well as insulin concentrations in pancreatic tissue samples were significantly lower than those in CTRL rats [plasma, 274.3 +/- 101.9 vs. 1078.8 +/- 254.9 pmol/liter (P < 0.05); tissue, 0.46 +/- 0.02 vs. 117.0 +/- 28.4 nmol/g (P < 0.01)]. INS + STZ rats did not become hyperglycemic, and the plasma and tissue levels of insulin were higher than those in STZ rats [plasma, 538.3 +/- 80.1 vs. 274.3 +/- 101.9 pmol/liter (P = 0.08); tissue, 0.46 +/- 0.02 vs. 37.90 +/- 2.13 nmol/g (P < 0.05)]. The immunohistochemical findings of insulin IR in the pancreatic tissues were in accordance with the results obtained by RIA. We conclude that exogenous insulin suppresses the expression of GLUT2 and insulin in beta-cells, and this may prevent the diabetogenic effect of streptozotocin.

Glucagon-like peptide-1-(7-36)amide is transformed to glucagon-like peptide-1-(9-36)amide by dipeptidyl peptidase IV in the capillaries supplying the L cells of the porcine intestine.

The insulinotropic hormone glucagon-like peptide-1 (GLP-1) is stored in the intestinal L cell in an active form, GLP-1-(7-36)amide, but more than half of the endogenous peptide circulates in an inactive, N-terminally truncated form, GLP-1-(9-36)amide. This study examined the GLP-1 newly secreted from the porcine ileum, in vitro (isolated perfused preparation) and in vivo (anesthetized pig), to determine where this conversion occurs. Although the GLP-1 extractable from the porcine ileum is predominantly the intact peptide (94.6+/-1.7%), a large proportion of the GLP-1 that is secreted has already been degraded to the truncated form both in vitro (53.8+/-0.9% intact) and in vivo (32.9+/-10.8% intact). In the presence of a specific dipeptidyl peptidase IV (DPP IV) inhibitor (valine-pyrrolidide), the proportion of intact GLP-1 released from the perfused ileum was increased under both basal (99% intact; P < 0.05) and stimulated (86-101% intact; P < 0.05) conditions. Immunohistochemical and histochemical studies revealed specific DPP IV staining in the brush border epithelium as well as in the capillary endothelium. Double staining showed juxtapositioning of DPP IV-positive capillaries and GLP-1-containing L cells. From these results, we suggest that GLP-1 is degraded as it enters the DPP IV containing blood vessels draining the intestinal mucosa.

Glucagon-like peptides GLP-1 and GLP-2, predicted products of the glucagon gene, are secreted separately from pig small intestine but not pancreas.

We developed specific antibodies and RIAs for glucagon-like peptides 1 and 2 (GLP-1 and GLP-2), two predicted products of the glucagon gene, and studied the occurrence, nature, and secretion of immunoreactive GLP-1 and GLP-2 in pig pancreas and small intestine. Immunoreactive GLP-1 and GLP-2 were identified in glucagon-producing cells of the pancreatic islets, and in glicentin-producing cells of the small intestine. Immunoreactive GLP-1 and 2 in intestinal extracts corresponded in molecular size to peptides synthesized according to the predicted structure. By reverse phase HPLC, intestinal and synthetic GLP-1 behaved similarly, whereas synthetic and intestinal GLP-2 differed. Pancreatic extracts contained a large peptide with both GLP-1 and GLP-2 immunoreactivity. Secretion was studied using isolated perfused pig pancreas during arginine stimulation, and isolated perfused pig ileum during either luminal glucose stimulation or vascular administration of the neuropeptide, gastrin-releasing peptide (GRP). Immunoreactive GLP-1 and GLP-2 were secreted in parallel with pancreatic glucagon and intestinal glicentin. The molecular forms of secreted immunoreactive GLP-1 and 2 corresponded to those identified in the tissue extracts.

Dipeptidyl peptidase IV inhibition enhances the intestinotrophic effect of glucagon-like peptide-2 in rats and mice.

Glucagon-like peptide-2 (GLP-2) induces intestinal growth in mice; but in normal rats, it seems less potent, possibly because of degradation of GLP-2 by the enzyme dipeptidyl peptidase IV (DPP-IV). The purpose of this study was to investigate the survival and effect of GLP-2 in rats and mice after s.c. injection of GLP-2 with or without the specific DPP-IV inhibitor, valine-pyrrolidide (VP). Rats were injected s.c. with 40 microg GLP-2 or 40 microg GLP-2+15 mg VP. Plasma was collected at different time points and analyzed, by RIA, for intact GLP-2. Rats were treated for 14 days with: saline; 15 mg VP; 40 microg GLP-2, 40 microg GLP-2+15 mg VP; 40 microg GLP-2 (3-33). Mice were treated for 10 days with: saline; 5 microg GLP-2; 5 microg GLP-2+1.5 mg VP; 25 microg GLP-2; 25 microg GLP-2 (3-33). In both cases, body weight, intestinal weight, length, and morphometric data were measured. After s.c. injection, the plasma concentration of GLP-2 reached a maximum after 15 min, and elevated concentrations persisted for 4-8 h. With VP, the concentration of intact GLP-2 was about 2-fold higher for at least the initial 60 min. Rats treated with GLP-2+VP had increased (P < 0.01) small-bowel weight (4.68 +/- 0.11%, relative to body weight), compared with the two control groups, [3.01 +/- 0.06% (VP) and 2.94 +/- 0.07% (NaCl)] and GLP-2 alone (3.52 +/- 0.10%). In mice, the growth effect of 5 microg GLP-2+VP was comparable with that of 25 microg GLP-2. GLP-2 (3-33) had no effect in rats, but it had a weak effect on intestinal growth in mice. The extensive GLP-2 degradation in rats can be reduced by VP, and DPP-IV inhibition markedly enhances the intestinotrophic effect of GLP-2 in both rats and mice. We propose that DPP-IV inhibition may be considered to enhance the efficacy of GLP-2 as a therapeutic agent.

All products of proglucagon are elevated in plasma from uremic patients.

We measured and characterized the proglucagon (PG) products in plasma obtained from nine uremic patients and six control subjects in the fasting state. The concentrations of PG products measured by direct RIAs were significantly higher in the uremic patients than in the normal subjects; the concentration of total glucagon immunoreactivity in plasma was 209 +/- 20 vs. 70 +/- 11 pmol/L, the glucagon-like peptide-1 immunoreactivity was 154 +/- 33 vs. 41 +/- 13 pmol/L, and the concentration of pancreatic-type glucagon immunoreactivity was 53 +/- 6 vs. 30 +/- 7 pmol/L. By chromatography, the predominating PG products in both uremic and normal plasma were shown to be glicentin [corresponding to PG-(1-69)] and the major PG fragment [presumably corresponding to PG-(72-158)]. In addition, glucagon-like peptide-1 [presumably corresponding to PG-(72-107) amide] was a major product in uremic plasma. Our results suggest that the kidneys play an important role in the removal from plasma of these products of PG.

Additive insulinotropic effects of exogenous synthetic human gastric inhibitory polypeptide and glucagon-like peptide-1-(7-36) amide infused at near-physiological insulinotropic hormone and glucose concentrations.

Gastric inhibitory polypeptide (GIP) and glucagon-like peptide-1-(7-36) amide (GLP-1) are glucose-dependent insulinotropic gut hormones that may explain the greater insulin secretory response with oral compared to i.v. glucose (incretin effect). To study their individual and combined contributions, in eight healthy volunteers, on separate occasions, synthetic human GIP (1 pmol/kg.min) and/or GLP-1 (0.3 pmol/kg.min) or placebo were infused i.v. (-30 to 120 min), while at 0 min, a glucose infusion "isoglycemic" to the profile after an oral glucose load of 50 g/400 mL was started. After the administration of 50 g oral glucose, immunoreactive GIP rose several-fold to 337 +/- 43 pmol/L, while there was only a transient (10-30 min) and moderate increment in immunoreactive GLP-1 (from basal, 25-30, to 41 +/- 4 pmol/L). Isoglycemic i.v. glucose infusions led to smaller B-cell responses (estimated incretin effect, 41 +/- 5%). With single infusions of GIP or GLP-1 (circulating concentrations, 464 +/- 73 and 54 +/- 3 pmol/L, respectively), B-cell responses were significantly augmented compared to i.v. glucose alone and were no longer significantly different from those after oral glucose. The combination of GIP and GLP-1 led to B-cell responses that were significantly higher than those with either hormone alone (additive mode of cooperation). Plasma GIP concentrations were similar after endogenous secretion (oral glucose) and i.v. infusion, while exogenously administered GLP-1 led to plasma levels that were maintained at an elevated level for a longer period during exogenous infusion than after stimulation by oral glucose. When, in seven volunteers, a lower dose (0.15 pmol/kg.min) of GLP-1 was infused during isoglycemic glucose infusion experiments only for the duration of elevated plasma levels in the oral glucose challenges (0-30 min), a significant, but transient, increment in insulin and C-peptide concentrations was observed, which was equivalent to 26 +/- 10% of the estimated incretin effect. Therefore, in conclusion, circulating GIP seems to make a major contribution to the incretin effect after oral glucose, and GLP-1 appears to mediate a smaller proportion. GIP and GLP-1 can interact in an additive manner in normal man.

Gastric emptying, glucose responses, and insulin secretion after a liquid test meal: effects of exogenous glucagon-like peptide-1 (GLP-1)-(7-36) amide in type 2 (noninsulin-dependent) diabetic patients.

The aim of the study was to investigate whether inhibition of gastric emptying of meals plays a role in the mechanism of the blood glucose-lowering action of glucagon-like peptide-1-(7-36) amide [GLP-1-(7-36) amide] in type 2 diabetes. Eight poorly controlled type 2 diabetic patients (age, 58 +/- 6 yr; body mass index, 30.0 +/- 5.2 kg/m2; hemoglobin A1c, 10.5 +/- 1.2%) were studied in the fasting state (plasma glucose, 11.1 +/- 1.1 mmol/L). A liquid meal of 400 mL containing 8% amino acids and 50 g sucrose (327 Kcal) was administered at time zero by a nasogastric tube. Gastric volume was determined by a dye dilution technique using phenol red. In randomized order, GLP-1-(7-36) amide (1.2 pmol/kg.min; Saxon Biochemicals) or placebo (0.9% NaCl with 1% human serum albumin) was infused between -30 and 240 min. In the control experiment, gastric emptying was completed within 120 min, and plasma glucose, insulin, C-peptide, GLP-1-(7-36) amide, and glucagon concentrations transiently increased. With exogenous GLP-1-(7-36) amide (plasma level, approximately 70 pmol/L), gastric volume remained constant over the period it was measured (120 min; P < 0.0001 vs. placebo), and plasma glucose fell to normal fasting values (5.4 +/- 0.7 mmol/L) within 3-4 h, whereas insulin was stimulated in most, but not all, patients, and glucagon remained at the basal level or was slightly suppressed. In conclusion, GLP-1-(7-36) amide inhibits gastric emptying in type 2 diabetic patients. Together with the stimulation of insulin and the inhibition of glucagon secretion, this effect probably contributes to the blood glucose-lowering action of GLP-1-(7-36) amide in type 2-diabetic patients when studied after meal ingestion. At the degree observed, inhibition of gastric emptying, however, must be overcome by tachyphylaxis, reduction in dose, or pharmacological interventions so as not to interfere with the therapeutic use of GLP-1-(7-36) amide in type 2 diabetic patients.