big-gastrin and Zollinger-Ellison-Syndrome

Topics: Gastrins; Humans; Pancreas; Protein Precursors; Protein Processing, Post-Translational; Zollinger-Ellison Syndrome

This study evaluates whether a new analytic principle, processing-independent analysis (PIA), offers better specificity and sensitivity than the conventional gastrin radioimmunoassay in the diagnosis of gastrinomas.. Plasma concentrations of alpha-amidated gastrins and the total progastrin product were measured with radioimmunoassay and with PIA, respectively, in 512 samples taken for gastrin measurement and in a selected group of gastrinoma patients (n=10).. Among the 512 patients were 9 with gastrinomas. In plasma from these patients the median degree of amidation (ratio of alpha-amidated gastrins to total progastrin product) was 75% (range, 25-98%), whereas in the other groups the medians varied from 41% to 86%. In the second group of gastrinoma patients all had a degree of amidation of less than 50%.. In screening for gastrinomas PIA offered no diagnostic advantages in comparison with conventional gastrin radioimmunoassay. However, in selected patients who in spite of normal or slightly increased concentrations of amidated gastrins were still suspected of having gastrinoma, additional measurement of the total progastrin product showed incomplete processing of progastrin and thus proved helpful in establishing the diagnosis.

Topics: Anti-Ulcer Agents; Case-Control Studies; Female; Gastrinoma; Gastrins; Humans; Male; Middle Aged; Pancreatic Neoplasms; Peptic Ulcer; Protein Precursors; Radioimmunoassay; Sensitivity and Specificity; Zollinger-Ellison Syndrome

Posttranslational processing is an important phase of the expression of most eucaryotic genes in terms of functional proteins. Among these, secretory proteins and peptides are of particular interest for clinical chemists, since diagnostic measurements of circulating proteins and peptides constitute a major discipline in clinical chemistry. The posttranslational covalent maturation of secretory proteins and peptides involves multiple enzymatic modifications of the corresponding proproteins along the intracellular secretory pathway. During the eighties, an increasing amount of evidence has indicated that sick secretory cells fail to process their secretory products normally. The diseased cells therefore fail to process their secretory products normally. The diseased cells therefore release also incompletely processed precursors and processing-intermediates. In order to measure the degree of disease, assays that measure proteins and peptides independent of the degree of processing are therefore desirable. We have now designed a new analytical principle, according to which secretory proteins, peptides and their precursors can be accurately quantitated irrespective of the degree of processing. This principle, named processing-independent analysis (PIA), is generally applicable to all cellular synthesized substances. The principle has been applied to and developed first for a well-defined secretory peptide system, progastrin and its products. Using this model, the results obtained so far confirm the diagnostic superiority of processing-independent analysis in comparison with conventional assays for bioactive peptides.

Topics: Amino Acid Sequence; Animals; Chemistry, Clinical; Clinical Laboratory Techniques; Duodenal Ulcer; Gastrinoma; Gastrins; Humans; Molecular Sequence Data; Peptides; Protein Precursors; Protein Processing, Post-Translational; Proteins; Radioimmunoassay; Zollinger-Ellison Syndrome

Processing-independent radioimmunoanalysis for progastrin showed that extracts of normal pancreatic tissue from normal subjects (n = 5) and from patients with adenocarcinoma of the papilla of Vater (n = 4) contain progastrin and its products. The concentrations varied from 0.1 to 5.8 pmol/g tissue, of which carboxyamidated bioactive gastrins constituted 0.03-1.9 pmol/g. In histologically normal and nonneoplastic pancreatic tissue from patients with duodenal (n = 3) and pancreatic (n = 2) gastrinomas the expression of gastrin was significantly higher-14.5 pmol/g (median), of which 28% was bioactive amidated gastrins. Gastrin-17 was the main bioactive product, but its immediate precursor, glycine-extended gastrin-17, constituted the predominant part of the preprogastrin product in pancreatic tissue. Proper gastrinoma tissue contained several precursor forms, including intact unprocessed progastrin. Progastrins were also found in high concentrations in plasma from the gastrinoma patients. The results raise the possibility that increased expression of progastrin and its products in non-neoplastic pancreatic tissue is a primary defect predisposing to neoplasia.

Topics: Adenocarcinoma; Ampulla of Vater; Common Bile Duct Neoplasms; Gastrinoma; Gastrins; Gene Expression; Humans; Pancreas; Pancreatic Neoplasms; Protein Precursors; Protein Processing, Post-Translational; Zollinger-Ellison Syndrome

Antisera were raised against fragment 38-54 of human progastrin. All of eight immunized rabbits responded, but only one (No. 2145) produced high-titer (3.2 x 10(4)) and high-avidity (Keff degrees = 1.2 x 10(12) l/mol) antibodies. A radioimmunoassay based on antiserum 2145 and monoiodinated gastrin-34 was specific for the N-terminal sequence of human gastrin-34. It measured concentrations of 9.7 +/- 1, 18.4 +/- 2 pmol/l (mean +/- SEM) and 1.553 (0.7-476) nmol/l (median (range], respectively, in sera from normal subjects (n = 20), patients with duodenal ulcer (n = 19), and Zollinger-Ellison patients (n = 8). Conventionally measured concentrations of carboxyamidated gastrins in the same sera were 21.4 +/- 1, 23.8 +/- 3 pmol/l (mean +/- SEM) and 0.833 (0.4-214) nmol/l (median (range)), respectively. The results show that radioimmunoassays specific for the N-terminus of human gastrin-34 discriminate between healthy subjects and patients with duodenal ulcer. The improved diagnostic specificity is due to co-measurement of unprocessed and partly processed progastrins that occur in plasma of patients with duodenal ulcer disease and gastrinomas. We suggest that conventional gastrin assays are supplemented with assays specific for the N-terminus of gastrin-34 in studies of duodenal ulcer disease.

Topics: Amino Acid Sequence; Anemia, Pernicious; Animals; Chromatography, Gel; Duodenal Ulcer; Gastrins; Immune Sera; Molecular Sequence Data; Protein Precursors; Rabbits; Radioimmunoassay; Zollinger-Ellison Syndrome

Progastrin and all of its processing products were measured in serum from 48 patients with Zollinger-Ellison syndrome, 42 patients with duodenal ulcers, and 34 normal subjects. A processing-independent gastrin analysis and a conventional radioimmunoassay for the biologically active alpha-amidated gastrins were used. In serum from normal subjects, 87% (median; range, 27%-160%) of all progastrin products were alpha-amidated gastrins, whereas they constituted only 39% (15%-130%) in serum from patients with duodenal ulcers (p less than 0.01) and 46% (16%-100%) in serum from gastrinoma patients (p less than 0.01). A significantly lower percentage of alpha-amidated gastrin was found in patients with hepatic metastases (23%) than in patients with apparently benign tumors (54%). Chromatography of serum showed that large progastrin molecules occurred mainly in patients with malignant tumors, whereas smaller glycine-extended precursors dominated in patients with benign tumors. The results indicate that the total progastrin product reflects tumor synthesis of gastrin better than conventional measurements of alpha-amidated gastrin. Moreover, the results suggest that a low degree of processing of progastrin could serve as a predictor of a malignant clinical course at an early stage of the disease.

Topics: Adolescent; Adult; Aged; Biomarkers, Tumor; Chromatography, Gel; Duodenal Ulcer; Female; Follow-Up Studies; Gastrins; Humans; Male; Middle Aged; Protein Precursors; Radioimmunoassay; Zollinger-Ellison Syndrome

Antibodies to different peptides produced from the gastrin precursor have been used in light microscopic- and electron microscopic-immunogold studies of gastrinoma tissue and normal antral mucosa. Antibodies to the C terminus of progastrin, which are known to react with the intact precursor, revealed immunoreactive material in the rough endoplasmic reticulum, Golgi region, and electron-dense granules in gastrinoma cells. In normal antrum these antibodies again revealed the Golgi region and a population of electron-dense granules. Other antibodies that react with the products of progastrin processing, but not the precursor, e.g., C-terminal and N-terminal gastrin 17 specific antibodies, revealed only granules. In addition to electron-dense granules already mentioned, the latter antibodies also revealed electron-lucent and intermediate granule populations, which in antrum were the major granule types. It is proposed that the intact precursor occurs in rough endoplasmic reticulum and Golgi; thereafter, in the immature electron-dense granules, and subsequently in electron-lucent granules, biosynthetic processing liberates gastrin 17 and gastrin 34, which are the major active products of gastrin gene expression.

Topics: Gastrins; Gold; Histocytochemistry; Humans; Immunoenzyme Techniques; Immunoglobulin G; Protein Precursors; Pyloric Antrum; Zollinger-Ellison Syndrome

We examined the posttranslational modification of gastrin in human gastrointestinal and Zollinger-Ellison tumor tissues using antibodies specific for progastrin and its glycine-extended processing intermediates. Gel filtration of antral and duodenal extracts on Sephadex G-50 revealed multiple molecular forms of immunoreactive glycine-extended processing intermediates corresponding to known molecular forms of gastrin and cholecystokinin. Immunoaffinity chromatography studies of antral mucosal and gastrinoma extracts identified a molecular form of glycine-extended processing intermediates that was characterized by an amino terminus identical to that of gastrin heptadecapeptide. Large differences in the relative contents of precursors and products of gastrin synthesis were found in gastrinoma tissue as compared to gastrointestinal mucosal extracts. These studies suggest the potential importance of glycine-extended peptides as intermediates in the posttranslational processing of gastrin and cholecystokinin in humans and indicate that gastrin processing mechanisms may be altered in Zollinger-Ellison tumors.

Topics: Chromatography, Affinity; Chromatography, Gel; Digestive System; Gastrins; Humans; Protein Precursors; Protein Processing, Post-Translational; Radioimmunoassay; Zollinger-Ellison Syndrome

Antibodies to the extreme C-terminal region of human progastrin have been used to monitor the isolation of high-Mr immunoreactive material in a gastrinoma extract. Microsequence analysis of the product revealed amino acid residues in the first 18 positions corresponding to those predicted from the cDNA sequence for preprogastrin starting at position 22; the sequence and immunochemical data together allow the identification of this material as intact progastrin. Implications for gastrin biosynthesis are discussed.

Topics: Amino Acid Sequence; Gastrins; Humans; Liver Neoplasms; Protein Precursors; Zollinger-Ellison Syndrome

Forty-six patients with the gastrinoma syndrome were divided into 2 categories: 1) benign sporadic gastrinoma (n = 30), and 2) gastrinoma with metastases to liver (n = 16). Thirteen of the 46 patients had multiple endocrine neoplasia type I syndrome. Serum gastrin levels in patients fasted overnight were determined by RIA using antisera directed toward the NH2- and COOH-terminals of heptadecapeptide gastrin (G17) and the NH2-terminus of the triacontatetrapeptide (G34). These results were compared with findings in 50 normal subjects. In the normal subjects, the mean COOH-terminal gastrin-17 level was higher [65 +/- 8 (+/- SEM) pg/ml] than the NH2-terminal gastrin-17 level (11 +/- 0.2 pg/ml) and lower than the NH2-terminal gastrin-34 level (134 +/- 20 pg/ml). The levels of NH2-terminal gastrin-17 were higher in patients with metastatic disease than in those with benign gastrinoma, whereas the COOH-terminal gastrin-17 and the NH2-terminal gastrin-34 levels were similarly high in both groups. The mean ratio of NH2-terminal gastrin-17 to COOH-terminal gastrin-17 was less than 1 in normal subjects (0.22 +/- 0.02) and benign gastrinoma patients (0.2 +/- 0.04), and it was 2.2 +/- 0.41 in the patients with metastatic gastrinoma. An NH2 to COOH gastrin-17 ratio greater than 1 was found in 13 of 16 patients with metastatic gastrinoma, but in none of the patients with benign gastrinoma or normal subjects. Similar results were found in multiple endocrine neoplasia type I patients with benign and metastatic disease. A high NH2 to COOH gastrin-17 ratio is suggestive of metastatic gastrinoma. In 4 patients with metastatic gastrinoma, the NH2 to COOH gastrin-17 ratio fell in parallel with the response to chemotherapy.

Topics: Chromatography, Gel; Gastrins; Humans; Liver Neoplasms; Multiple Endocrine Neoplasia; Protein Precursors; Radioimmunoassay; Zollinger-Ellison Syndrome

Recent studies on the gene sequence encoding the human pyloric antral hormone, gastrin, indicate a precursor of 101 residues. We have now raised antibodies to a synthetic analogue corresponding to (Tyr)-human progastrin COOH-terminal pentapeptide. The antibodies could be used in radioimmunoassay to measure this peptide, but they did not react with corresponding fragments of procholecystokinin, porcine progastrin, or other human progastrin-derived peptides, notably heptadecapeptide gastrin (G17), and 34-residue gastrin (G34). Radioimmunoassay of human antral and duodenal extracts revealed a major peak of activity that corresponded to the native COOH-terminal fragment of progastrin, and occurred in approximately equimolar amounts with COOH-terminal G17 immunoreactivity. In addition, there was a minor peak of apparently higher molecular weight material. In some gastrinomas the latter material was the predominant immunoreactive form, and it occurred in higher molar concentrations than any other form of gastrin. Digestion of this material with trypsin liberated peptides that reacted with antibodies specific for the NH2-terminus of G34, and G17. On this basis the high molecular weight component was identified as a form of gastrin that extended from the COOH-terminus of the precursor to a point at least beyond the NH2-terminus of G34, and probably included the entire progastrin sequence. The results suggest differences in posttranslational processing pathways of progastrin in antrum, duodenum, and gastrinomas. They also indicate that the present experimental approach allows the identification of progastrin-like substances, which should open the way to studying the mechanisms of gastrin biosynthesis.

Topics: Chromatography, High Pressure Liquid; Duodenum; Gastric Mucosa; Gastrins; Humans; Peptide Fragments; Protein Precursors; Pyloric Antrum; Radioimmunoassay; Trypsin; Zollinger-Ellison Syndrome

Topics: Gastrins; Humans; Middle Aged; Protein Precursors; Radioimmunoassay; Zollinger-Ellison Syndrome

Gastrin- 17 (G-17) and gastrin-34-like immunoreactivities of human gastrinoma cells were investigated at light and electron microscope level using N-terminally directed antisera. The procedure includes (a) the 24 hr/immunoperoxidase staining of Bouin-fixed paraffin embedded tissues, (b) the immunoelectron microscopic labelling of aldehyde-fixed Epon-embedded tissues according to the immunogold technique. On light microscopy, a variable number of tumor cells stained for G-34. In contrast, G-17 immunoreactivity was very low or undetectable in the tumor material, although it was easily detected in endocrine cells of similarly processed human pyloric mucosa. On electron microscopy, most of the tumor cell granules belonging to the round compact or dense-cored type exhibited a variable labelling for G-34, whereas the vacuolar/floccular type remained unlabelled. In contrast, the labelling for G-17 occurred over most of the tumor cell granules, whether compact or floccular. Dense granules of varying size and shape, previously shown to store C-terminal gastrin immunoreactivity, were only faintly labelled by the two antisera. When compared to the labelling pattern of human pyloric G-cells, the predominance of round dense granules with G-34 and G-17 immunoreactivity in gastrinoma cells suggests an incomplete or defective post-translational processing of the precursor peptide.

Topics: Colloids; Gastrins; Gold; Humans; Immune Sera; Immunoassay; Immunoglobulin G; Microscopy; Microscopy, Electron; Protein Precursors; Zollinger-Ellison Syndrome

Two peptides which copurified from a human gastrinoma were found to correspond to the amino acid sequence deduced for the amino terminal portion of human and porcine progastrin. The sequence of peptide A is Ser-Trp-Lys-Pro-Arg-Ser-Gln-Gln-Pro-Asp-Ala-Pro-Leu-Gly-Thr-Gly-Ala-Asn- Arg-Asp-Leu-Glu-Leu which is identical to an amino terminal portion of human progastrin. The sequence of peptide. B is identical to that of peptide A except it is missing the first five amino acids. If peptide A corresponds to the amino terminus of progastrin, the signal peptidase cleaves at an Ala-Ser bond.

Topics: Amino Acid Sequence; Animals; Gastrins; Genes; Humans; Peptide Fragments; Protein Precursors; Species Specificity; Swine; Zollinger-Ellison Syndrome

Topics: Gastrins; Humans; Protein Precursors; Radioimmunoassay; Reagent Kits, Diagnostic; Zollinger-Ellison Syndrome

The previously assigned structure of human big gastrin is revised as a result of sequencing and immunological studies on synthetic peptides. A nonadecapeptide has been synthesized and found to have full immunochemical potency compared with natural human G34 in a radioimmunoassay which is specific for the N-terminal sequence. Syntheses of the peptides were achieved using the stepwise procedure with benzyloxycarbonyl-amino acids and fragment couplings mediated mainly by the dicyclohexylcarbodiimide procedure in the presence of either N-hydroxysuccinimide or 1-hydroxybenzotriazole. Purification of the peptide fragments was by Sephadex LH-20 chromatography and removal of protecting groups was effected using 90% trifluoroacetic acid in the presence of scavengers. Purification of the nonadecapeptide was achieved by high performance liquid chromatography.

Topics: Amino Acid Sequence; Amino Acids; Chemical Phenomena; Chemistry; Gastrins; Humans; Protein Precursors; Radioimmunoassay; Zollinger-Ellison Syndrome