Distribution of oxyntomodulin and glucagon in the gastrointestinal tract and the plasma of the rat

Oxyntomodulin (OXM), an intestinal glucagon-containing peptide extended at its C-terminal end by an octa-peptide, is one of the gut glucagon-like immunoreactants (GLI) or enteroglucagon. The distribution of OXM and glucagon was determined in the gastrointestinal tract and in the plasma of the rat. Reversed-phase HPLC, associated with RRA or RIA, performed with an N-terminally directed glucagon antiserum (GOL), was used. HPLC of intestinal extracts or plasma separated the GOL immunoreactivity into three peaks: two major peaks coeluting with a preparation of rat glicentin (peak I, partially purified from rat intestine) and porcine or rat OXM, respectively, and a smaller peak coeluting with glucagon. The behavior of the three peaks in the analytical systems matched that of glicentin, OXM, and glucagon, respectively, allowing their identification. The concentrations of OXM picomoles per g of tissue) gradually increased from the duodenum (9 +/- 1) to ileum (93 +/- 4), thereafter decreasing in cecum and colon (22 +/- 3). In the gut, OXM, glucagon, and peak I averaged 40%, 1%, and 59% of the total GLI, respectively. OXM was present in significant amounts in the pancreas (18% of GLI) and stomach (27% of GLI), two tissues in which it accounted, together with glucagon, for almost the totality of GLI. In 24 h-fasted rats, plasma concentrations of OXM, glucagon, and peak I, determined after HPLC with GOL antiserum, were 15.1 pM, 8.6 pM, and 12.3 pM, respectively. Two hours after refeeding, both OXM and peak I were significantly increased (P less than 0.05 and P less than 0.02) by a similar factor (2-fold), while glucagon remained unchanged. When the HPLC results were compared with RIA measurement of GLI (GOL antiserum) and glucagon (with a C-terminal glucagon antiserum) in plasma, enteroglucagon (GOL--C-terminal glucagon antiserum immunoreactivities) correlated well with the sum of OXM plus peak I. The combination of HPLC and RRA or RIA allows the unambiguous determination of OXM, glucagon, and glicentin (peak I) in tissues and plasma. In the rat intestine and in the plasma, OXM and glicentin appear roughly in the same ratio and seem to be the major components, if not the totality, of enteroglucagon.     

Gut glucagon, enteroglucagon, gut glucagonlike immunoreactivity, glicentin--current status

Glucagonlike substances in extracts of intestinal mucosa were already described in 1948 by Sutherland and deDuve (1), who used a bioassay technique for the identification. After the development of the first glucagon radioimmunoassays, Unger and co-workers (2,3) confirmed that intestinal extracts contained peptides that "crossreacted" in the glucagon radioimmunoassay [hence gut "glucagonlike immunoreactivity" (GLI)]. In 1968, the same group discovered that the gut GLIs consisted of at least two peptides, GLI I and II (4), both of which differed immunochemically from pancreatic glucagon and, therefore, necessarily had different chemical structures (4,5). Developments during the last decade in the field of peptide chemistry, particularly improved purification and sequencing techniques, have greatly advanced our knowledge of gut peptides, including the enteroglucagons, and the chemical structure of several of the members of this heterogenous group of peptides is now known. Furthermore, progress in the field of nucleotide and gene technology has also spread to this area of research, and although many problems remain unresolved, the progress made has sufficiently important implications to justify a review of the most recent advances.     

Glucagon-like immunoreactive peptides in a rat ileal epithelial cell line (IEC-18)

The presence of cells containing glucagon-like immunoreactive (GLI) peptides was demonstrated in a rat ileal epithelial cell line (IEC-18) by both immunofluorescence and radioimmunoassay. When cell extracts were subjected to gel filtration chromatography, the cells were found to contain 3.5 Kd glucagon in addition to significant quantities of large molecular weight GLI peptides (apparent molecular weights of 4, 6, 8 and 10 Kd) and a 9 Kd peptide with apparent glucagon immunoreactivity. This was in contrast to extracts of adult rat ileum, which contained only large molecular weight GLI peptides (apparent molecular weights of 6 and 12 Kd). Production of GLI peptides by the IEC-18 cells was stimulated by glucose (p less than 0.02) and inhibited by insulin (p less than 0.01). In conclusion, these results demonstrate that the IEC-18 cells produce both GLI peptides and glucagon, and thus support the notion that proglucagon processing is cell-specific. IEC-18 cells may therefore provide a tool for investigations of some aspects of GLI peptide and glucagon synthesis.     

The effect of phenformin upon the plasma pancreatic and gut glucagon-like immunoreactivity in diabetics

Five patients with mild maturity-onset diabetes were given 250 ml of a 20% glucose solution by intraduodenal infusion and eight other patients similarly received an amino acid solution in a dose of 0.5 g amino acids per kg body weight. The pancreatic and gut glucagon-like immunoreactivity (pancreatic GLI and gut GLI) in plasma were measured before and after the application of the two stimuli. Each person was tested twice; the first (control) test was followed by a second test after three days of treatment with phenformin 150 mg daily, plus the same 150 mg dose taken 60 min before the intubation. The plasma pancreatic GLI increased slightly during both infusions, but was not affected by phenformin. Intraduodenal infusion of both glucose and the amino acid solution induced a greater rise in plasma gut GLI. After treatment with phenformin, the fasting plasma gut GLI was higher than the control value in eleven of thirteen patients. In most cases higher gut GLI plasma levels were also found after duodenal administration of glucose and amino acids. These data furnish further evidence of the local action of antidiabetic biguanides on the intestinal wall, including its hormonal activity. The hypothesis is advanced that the phenformin-induced increase in gut GLI secretion may bring about competition of the latter with pancreatic glucagon for receptors in liver cell membranes, reducing the effect of glucagon on the liver, and thus contributing to a decrease in glycaemia.     

Synthesis and release of glucagon by human salivary glands

Summary Pieces of human salivary glands were homogenised with acid-ethanol or acid-saline solutions immediately after surgical removal. With both extraction procedures the immunoreactive glucagon (IRG) content in the submaxillary glands was greater than in parotid glands as determined with a C-terminal reactive glucagon antiserum (30K). Higher amounts of IRG were determined in acid-saline extracts of submaxillary (18.5±2.5 vs 8.9±1.2 ng/g wet weight) and parotid (3.5±0.3 vs 2.9±0.3 ng/g wet weight) glands compared with concentrations obtained with acid-ethanol extracts. IRG material extracted with the latter procedure has similar immunological and biological characteristics as pancreatic glucagon. After fractionation of the acid-ethanol extracts on P-30 columns or gel electrophoresis, an immunoreactive peak of 3500 daltons was always obtained. Arginine, ephinephrine and low glucose concentrations stimulated glucagon release from both salivary glands. Active glucagon biosynthesis by these glands was established by the incorporation of ³H-L-tryptophan into a 3500 daltons polypeptide with specific immune reaction with 30K antiserum. These findings indicate that human salivary glands represent a source of extrapancreatic glucagon in man and may therefore contribute to the circulating levels of this hormone.     

Oxyntomodulin and glicentin: brain-gut peptides in the rat

Glucagon-like materials and glucagon have been identified by immunoassay and immunocytochemistry in the mammalian central nervous system. However, the molecular forms relevant to brain glucagon-like immunoreactivity (GLI) have not been precisely defined. In the rat small intestine, more than 90% of GLI is constituted by two peptides: oxyntomodulin (OXM) and glicentin. This work was initiated to characterize and determine the concentrations of these two peptides and glucagon in the rat central nervous system and to compare their relative proportions with those found in the gut. Different regions from the adult rat brain were analyzed by HPLC in association with RIA, using a central glucagon antiserum and an antibody directed toward the C-terminal end of OXM and glicentin. The elution profiles of hypothalamus extracts were constituted by two main peaks, both detected by the two antibodies used and displaying the same retention times as glicentin and OXM, respectively. A third small peak, which coeluted with glucagon, was constantly recorded with the central glucagon antiserum. The percentages of glicentin, OXM, and glucagon in 10 hypothalami were 37 +/- 1%, 55 +/- 1%, and 8 +/- 2%, respectively (n = 8). This distribution was quite similar to that in small intestinal extracts (38 +/- 1%, 59 +/- 1%, and 1.3 +/- 0.1%, respectively; n = 7); however, the peptide concentrations were almost 50-fold greater in intestine than in hypothalamus. In the medulla oblongata, the same peptide ratio was observed, with 10-fold lower concentrations compared to those in hypothalamus. In olfactory bulb, cerebellum, and cortex the concentrations were close the the detection limit, whereas they could be not detected in the pituitary. The combination of HPLC and specific RIAs allowed us to unambiguously characterize OXM and glicentin as the major components of GLI in the rat hypothalamus and medulla oblongata. The same proportion of these two peptides in the central nervous system and the gut indicates that a similar posttranslational processing exists in these rat tissues, another example of the brain-gut axis.     

Secretion of immunoreactive insulin and glucagon in hamsters bearing a transplantable insuloma.

Glucose, insulin (IRI), pancreatic (IRG) and total (GLI) immunoreactive glucagon were measured in the serum of normal hamsters and of hamsters with an insulin- and glucagon-secreting, transplantable insuloma. The tumor-bearing animals were hypoglycemic, hyperinsulinemic and hyperglucagonemic. The pancreatic islets of tumor-bearing animals secreted less glucagon and insulin in response to arginine or to changes in the glucose concentration of the medium, than did the islets of control hamsters. In addition, the introduction of glucose into the gastro-intestinal tract, which caused a significant rise in the serum GLI concentration of normal hamsters, failed to do so in the tumor-bearing animals. The results suggest that the high levels of serum glucagon and insulin induced by the tumor, suppressed IRI, IRG and GLI secretion in these animals.     

Pancreatic polypeptide and other hormones in pancreas of obese (ob/ob) mice

Summary The insulin, glucagon, somatostatin and pancreatic polypeptide content of acid-ethanol extracts of pancreas from lean and obese (ob/ob) mice of various ages was determined by radioimmunoassay. Rat and mouse pancreatic polypeptide react weakly with antibodies to avian, bovine and canine pancreatic polypeptide, but immunoassay for this peptide was possible using an antibody to the carboxyl-terminal hexapeptide of bovine pancreatic polypeptide and amino-terminal labelled bovine pancreatic polypeptide as tracer. The insulin, glucagon and pancreatic polypeptide content of pancreas from obese mice was greater than that of lean controls. The increase in insulin content uniformly involved ventral, dorsal and splenic lobes. Glucagon content was elevated primarily in the splenic lobe. Pancreatic polypeptide content was most significantly elevated in the splenic lobe where pancreatic polypeptide cells are infrequent in normal lean mice and this was accompanied by increased numbers of pancreatic polypeptide cells per islet in this lobe.     

Control of glucagon-like immunoreactive peptide secretion from fetal rat intestinal cultures

Some of the mechanisms underlying intestinal glucagon-like immunoreactive (GLI) peptide secretion from cultured fetal rat intestinal cells were investigated using modulators of the adenylate cyclase pathway [(Bu)2cAMP, theophylline, isobutylmethylxanthine], calcium fluxes (ionomycin, A23187), and protein kinase-C (phorbol ester). All of these agents were found to stimulate GLI peptide release, to 120-230% of paired control values (P less than 0.05-0.001). (Bu)2cAMP, but not the phorbol ester, also increased the total cell content of GLI peptides over the 2-h incubation period (P less than 0.05). No synergism between any of the three pathways was detected. When the mol wt distribution of the stored and secreted GLI peptides was determined in control and (Bu)2 cAMP-stimulated samples, 68 +/- 2% of the peptide corresponded to glicentin, while the remainder eluted with the same distribution coefficient as oxyntomodulin. No 3.5K glucagon was detected in any of the extracts. GLI peptide secretion by the cells was not altered by several pancreatic glucagon secretagogues (cortisol, bombesin, and prostaglandins E1 and D2), but was stimulated by the opioid peptide beta-endorphin (1 microM; P less than 0.02). These studies have indicated that the control of secretion of fetal rat intestinal GLI peptides is complex, involving activation of any one or a combination of the three major second messenger systems. A role for the adenylate cyclase pathway in regulating GLI peptide biosynthesis is also suggested.     

The significance of plasma gastrointestinal glucagon in endotoxemia

Hyperglucagonemia has been reported during endotoxemia. Increases in plasma glucagon concentration may serve as an important mediator and be influential in modifying metabolism during endotoxin shock. Increases in the plasma concentration of glucagon are brought about by both pancreatic and gastrointestinal release of glucagon during endotoxemia. The purpose of the present study was to determine the importance of gastrointestinal-derived glucagon (glucagon-like immunoreactivity; GLI) during endotoxin shock. LD100 endotoxin (3.0 mg/kg) was injected via the auricular vein into fasted rabbits. Emphasis was focused on total glucagon and gastrointestinal glucagon (GLI) concentrations, but measurements of these parameters were supplemented by determinations of glucose and triglyceride. The results were as follows: 1) gastrointestinal glucagon concentration increased threefold over that of pancreatic glucagon concentration and 27-fold over that of the control group at 6 hr after endotoxin administration and 2) significant increases of both serum glucose and triglyceride concentrations were observed after endotoxin was injected. Findings suggest that gastrointestinal glucagon may be one of the factors responsible for evoking the abnormal metabolism caused by endotoxin shock. In hyperglucagonemia resulting from endotoxemia, it is suggested that attention should be paid not only to the glucagon originating from the pancreas but also to that derived from the intestine.     

Changes in glucagon processing occurring in the intestines of surgically stressed patients

BACKGROUND/AIMS: The kinetics of the pancreatic hormone glucagon in surgically stressed patients has not been investigated as thoroughly as that of insulin, despite its significant influence on energy metabolism in surgically stressed conditions. In the present study, we examined the kinetics of glucagon and glucagon-related peptides assessed by radioimmunoassay, and the molecular forms of these peptides using gel filtration chromatography, and in addition discuss glucagon processes in the pancreas and intestine in surgically stressed patients. METHODOLOGY: Ten patients who had undergone abdominal surgery for acute abdominal disorders were enrolled in this study (group S). Ten healthy volunteers were also enrolled as normal controls (group C). The serum level of glucagon and glucagon-related peptides were assessed in the early morning fasting state in both groups, on the second postoperative day in group S, using glucagon nonspecific N-terminal (glucagon-like immunoreactivity: GLI) and specific C-terminal (immunoreactive glucagon: IRG) radioimmunoassays. The molecular forms of these peptides were also estimated using the gel filtration chromatography method. RESULTS: Serum IRG in group S was significantly higher than that of group C (P < 0.05). Serum GLI was not significantly different between the two groups. In all patients except one in group S, a peculiar glicentin-like peptide (GLLP: MW about 8000) other than pancreatic glucagon was seen in gel filtration chromatography, which was not seen in group C. CONCLUSIONS: The kinetics and processing of glucagon in surgically stressed patients were quite different from those of healthy subjects. In surgically stressed patients, peculiar processing of glucagon occurred in the intestine, which was quite different from ordinary glucagon processing either in the pancreas or the intestine, generating GLLP.     

Extraction,gel filtration pattern,and receptor binding of porcine gastrointestinal glucagon-like immunoreactivity

Summary Different techniques for the extraction and initial purification of porcine gastrointestinal glucagon-like immunoreactivity (GLI) were compared with reference to yield, and preservation of number and pattern of GLI components. The conventional acid-ethanol technique combined with ethanol-ether purification gave high yields and a reproducible pattern of components. Large amounts of tissue were more easily extracted using another technique, based on extraction by boiling, extraction and precipitation with acetone, and — if necessary — salting out. — By means of the latter two techniques mucosal tissue from all of the porcine gastrointestinal tract was extracted and subjected to gel filtration. Glucagon-like peptides were searched for using: — 1. a radioimmunoassay which quantifies gut type glucagon (GTG), as well as pancreatic type glucagon (PTG), 2. a radioimmunoassay highly specific for pancreatic type glucagon (PTG), and 3. a radioreceptor assay based on binding of glucagon to porcine liver cell membranes. — The oesophageal, the fundic, and the antro-pyloric parts of the gastric mucosa contained very small amounts of GLI. The cardiac gland region contained small amounts of a peptide indistinguishable from true glucagon. The duodenal mucosa contained small amounts of true glucagon and may be a smaller, glucagon-like peptide. The mucosa of the small intestine contained large amounts of both high and low molecular weight GTG and, in addition, PTG of high molecular weight and true glucagon. The colon also contained these components with true glucagon in high concentrations. Only small GTG and true glucagon were receptor-active, the former with less than its immunometric potency.     

Prothymosin alpha: isolation and properties of the major immunoreactive form of thymosin alpha 1 in rat thymus.

A polypeptide containing approximately equal to 112 amino acid residues, with the thymosin alpha 1 sequence at its NH2 terminus, has been isolated from rat thymus by using a radioimmunoassay with an antibody prepared against synthetic thymosin alpha 1. The new polypeptide, named "prothymosin alpha," was found to be the major substance crossreacting with thymosin alpha 1 antiserum in rat thymus extracts; peptides corresponding to thymosin alpha 1 or thymosin alpha 11 were not detected. In gel filtration at pH 2.8, prothymosin alpha emerged as a single symmetrical peak corresponding to an apparent molecular weight of 32,000, approximately 3 times larger than the minimum molecular weight calculated from its amino acid composition. On the same gel filtration columns, synthetic thymosin alpha 1 (calculated Mr = 3108) emerged at a position corresponding to a molecular weight of 10,000-11,000. Thus, both prothymosin alpha and thymosin alpha 1 appear to exist in solution as oligomers, possibly as trimers. Prothymosin alpha and synthetic thymosin alpha 1 also were separated readily in reverse-phase HPLC and in isoelectric focusing; the isoelectric point of prothymosin alpha determined by the latter procedure was found to be 3.55, consistent with an unusually high content of glutamic and aspartic acids based on amino acid analyses. Prothymosin alpha appears to represent the native polypeptide from which thymosin alpha 1 and other fragments are generated during the isolation of thymosin fraction 5.     

Silent human pancreatic glucagonoma and "A" nesidioblastosis

Surgical fragments of healthy and tumor-bearing pancreas from a patient with pancreatic tumor were studied by electron or light microscopy, histochemistry, and immunocytochemistry (human insulin, glucagon, somatostatin, gastrin, and bovine pancreatic polypeptide). Histological results were compared to those obtained by radioimmunoassay, both in tumor and serum. The tumor was identified as a glucagonoma because reactions for Grimelius' silver impregnation and immunoreaction with an antiserum against glucagon were positive and because a very high level of glucagon in the tumor was observed. Insulin, somatostatin, and gastrin levels remained normal, both in tumor and serum, but the glucagon level was normal in serum. Associated with this silent glucagonoma, an uncommon nesidioblastosis was also diagnosed with many A cells irregularly mixed with acinar cells, isolated or clustered in small groups. Acinar "intermediate" cells of "A" type were also observed. Such associative histopathological processes evoked possible development of an endocrine tumor from nesidioblastic-like tissue. Its embryogenic origin remained uncertain.     

A method for measuring the release of gut glucagon-like immunoreactivity from rat jejunum „In Vitro“

Summary A convenient method for measuring the release of gut glucagon-like immunoreactivity (G-LI) from rat jejunal pieces incubated in vitro is described. The method utilises Trasylol, EDTA, and excessive glucagon antibody to prevent GLI breakdown. There was satisfactory recovery of added pancreatic glucagon to the incubates, and a highly significant release of GLI was recorded when high glucose concentrations (5–40 g%) were incubated with the pieces.

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Methode zur Bestimmung der Freisetzung von glucagon-ähnlicher immunologischer Aktivität des Darmes aus dem Jejunum der Ratte in vitro.

Zusammenfassung Es wird eine brauchbare Methode zur Bestimmung der Freisetzung von glucagonähnlicher immunologischer Aktivität (GLI) des Darmes beschrieben. Die Methode benutzt Trasylol, EDTA und einen Überschuß von Glucagonantikörpern, um den Abbau von GLI zu verhindern. Die Rückgewinnung von zugefügtem Pankreasglucagon aus den Inkubaten war zufriedenstellend und eine hoch signifikante Freisetzung von GLI wurde beobachtet, wenn hohe Glucosekonzentrationen (5–40%) mit den Darmstücken inkubiert wurden.

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Méthode de mesure de la libération de la substance intestinale douée de l'immunoréactivité du glucagon (glucagon-like) à partir du jéjunum de rat in vitro

Résumé Les auteurs décrivent une méthode commode pour mesurer la libération de la substance intestinale immunologiquement glucagon-like (GLI) à partir de fragments de jéjunum de rat, incubés in vitro. La méthode utilise du trasylol, de l'EDTA et un excès d'anticorps anti-glucagon pour empêcher la dégradation du GLI. Il y avait une récupération satisfaisante du glucagon pancréatique ajouté aux incubats, et une libération de GLI hautement significative a été enregistrée lorsque de fortes de concentrations de glucose (5– 40 g%) étaient mises à incuber avec les fragments.

     

Simultaneous recording of the gastro-entero-pancreatic hormonal peptide response to food in man

The serum or plasma concentrations of gastrin, gastric inhibitory polypeptide (GIP), gut glucagon-like-immunoreactivity (gut GLI), secretin, vasoactive intestinal polypeptide (VIP), insulin, glucagon, and pancreatic polypeptide (PP) were recorded simultaneously following the ingestion of a normal, mixed meal in seven healthy, normal weight men. The concentrations of PP and gastrin increased within 10 min. Subsequently GIP, insulin, glucagon, and gut GLI increased in the order mentioned. The mean concentrations of secretin and VIP were not affected by the meal, although transient decreases in secretin concentrations could be detected in all subjects. The concentrations of the other hormones remained elevated for 4 hr or more. Perhaps the period of observation following food stimulation of gastroentero-pancreatic hormones should be extended.     

Does somatostatin in the gut inhibit the GLI release locally?

Summary Based on the assumption that somatostatin may inhibit peptide release through junctional complexes or through local circulation, an immunofluorescent technique for somatostatin and GLI in the gut was applied in order to investigate whether suppression of GLI release by i.v. administration of somatostatin was a physiological effect of somatostatin or not. Somatostatin-immunoreactive cells (GIF-cells) in the human and canine intestine had no direct cellular contacts with GLI-immunoreactive cells (GLI-cells). This finding suggests that somatostatin in the intestine does not inhibit GLI release through junctional complexes between GIF- and GLI-cells. As to the local circulation, most of GIF-cells in the canine intestine were distributed in the deeper portion of the intestinal gland which corresponds to the upstream sides of the local blood supply of the intestinal gland, as reported byReynold et al. The ratio of GIF-cells to total cells (GIF-cells + GLI-cells) was 68% in the duodenum and 25% in the ileum. In contrast, a limited number of GIF-cells was found in the human duodenum where a few GLI-cells were distributed and a few GIF-cells were seen in the human ileum where a large number of GLI-cells were located. Findings in the dog suggest the possibility that somatostatin inhibits GLI release from GLI-cells through the local circulation system of intestinal glands. However, findings in humans suggest that the same possibility does not apply to the human gut. Differences of population density of intestinal GIF-cells between humans and dogs indicate that the functional meaning of GIF-cells may vary from one species to another. GLI = glucagon like immunoreactivity.     

Changes in glucagon kinetics and processing in patients with acute pancreatitis

BACKGROUND/AIMS: The kinetics of the pancreatic hormone glucagon in patients with acute pancreatitis have not been investigated as carefully as those of insulin, in spite of its crucial influence on energy metabolism. In the present study, we studied the kinetics of glucagon and glucagon-related peptides assessed by radioimmunoassay. Furthermore, the molecular forms of these peptides were examined using gel filtration chromatography, and the glucagon processes in the pancreas and intestine in the early stage in patients with acute pancreatitis were investigated. METHODOLOGY: Fourteen patients with acute pancreatitis were enrolled in this study. Eight had severe pancreatitis (group S) and six had mild pancreatitis (group M). Ten healthy volunteers were also enrolled as the normal control (group C). Serum levels of glucagon and glucagon-related peptides were assessed on the second admission day in groups S and M, and in an early morning fasting state in group C, using glucagon non-specific N-terminal (glucagon-like immunoreactivity: GLI) and specific C-terminal (immunoreactive glucagon: IRG) radioimmunoassays. The molecular forms of these peptides were also estimated using gel filtration chromatography. We then discuss the glucagon processes based on these findings. RESULTS: Serum GLI and IRG in groups S and M were significantly higher than those of group C (P < 0.01), while those in group S were also significantly higher than those in group M (P < 0.05). In all patients in groups S and M, except for only three in group S, a peculiar glicentin-like peptide (GLLP: MW about 8000) other than pancreatic glucagon was observed in IRG gel filtration chromatography, which was clearly absent from group C. CONCLUSIONS: The kinetics and processing of glucagon in patients with acute pancreatitis were quite different from those of healthy subjects. In patients with acute pancreatitis, the peculiar processing of glucagon proceeded in the intestine quite differently from ordinary glucagon processing either in the pancreas or in the intestine, generating a peculiar GLLP.     

The normal range of the plasma immunoreactive glucagon level during a 75 g oral glucose tolerance test

In order to establish a normal value of plasma glucagon immunoreactivity (GI) and glucagon-like immunoreactivity (GLI) during a newly adopted 75 g OGTT, 50 normals (N), 102 individuals with IGT and 20 diabetics (D) were subjected to the OGTT, and their plasma GI and GLI levels were determined at various intervals by radioimmunoassay using 2 kinds of the C-terminal region specific antibody, OAL123 and 30K, and of the antibody specific for the N-terminal and/or central region of glucagon, OAL196, respectively. The basal levels of OAL123-GI and 30K-GI and OAL196-GLI in the 3 groups were as follows; N, 114.3, 80.8, and 335.5; IGT, 107.6, 76.1, and 338.5; and D, 135.7, 76.9, and 342.2 pg/ml. After glucose administration, a significant decrease in plasma GI and increase in plasma GLI were observed in the 3 groups, although their changes from the basal levels were variable. The plasma samples of inexplicably high GI concentration were chromatographed to clarify the nature of the hyperglucagonemia. The apparent GI was mostly eluted in the Vo component, but negligibly at the 3500 mol.wt. glucagon fraction. There was a marked difference in the Vo peak depending upon the antiserum used. These facts suggest that plasma GI values are dependent on the amount of BPG present in particular samples, and the antibody used.