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Pure Human Big Gastrin IMMUNOCHEMICAL PROPERTIES, DISAPPEARANCE HALF TIME, and ACID-STIMULATING ACTION in DOGS

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Pure Human Big Gastrin IMMUNOCHEMICAL PROPERTIES, DISAPPEARANCE HALF TIME, and ACID-STIMULATING ACTION in DOGS Pure Human Big Gastrin IMMUNOCHEMICAL PROPERTIES, DISAPPEARANCE HALF TIME, AND ACID-STIMULATING ACTION IN DOGS John H. Walsh, … , Haile T. Debas, Morton I. Grossman J Clin Invest. 1974;54(2):477-485. https://doi.org/10.1172/JCI107783. Research Article Biological properties of pure natural human “big gastrin” (designated G-34 because it contains 34 amino acid residues) were compared with those of pure natural heptadecapeptide gastrins (G-17) from human and porcine sources. Radioimmunoassay inhibition curves indicated that G-17 was nearly 1.5 times more potent than G-34 with the antibody used in this study. This difference was confirmed by demonstration of increased immunoreactivity generated when G-34 was converted to G-17 by trypsinization. When infused intravenously into dogs with gastric fistulas and Heidenhain pouches in equimolar doses, G-34 produced slightly higher acid secretory responses than G-17. Responses to sulfated and nonsulfated forms were not significantly different, nor were responses to human and porcine G-17. During infusion of equimolar doses, steady-state serum gastrin concentrations were more than fivefold higher with G-34 than with G-17. The difference in steady-state blood concentrations could be accounted for by a corresponding difference in removal rates. The half times of the G-34 preparations averaged 15.8 min and the half times of the G-17 preparations averaged 3.2 min. The calculated spaces of distribution for G-17 and G-34 were similar, about 25% of body weight. When the increment in serum gastrin was plotted against acid secretory response it was found that nearly five times greater increments in molar concentrations of G-34 than of G-17 were required to produce the same […] Find the latest version: https://jci.me/107783/pdf Pure Human Big Gastrin IMMUNOCHEMICAL PROPERTIES, DISAPPEARANCE HALF TIME, AND ACID-STIMULATING ACTION IN DOGS JOHN H. WALSH, HAILE T. DEBAS, and MORTON I. GROSSMAN From the Veterans Administration Wadsworth, Hospital Center, and the University of California at Los Angeles School of Medicine, Los Angeles, California 90073 A B S T R A C T Biological properties of pure natural rate of acid secretion. The potency of these two mole- human "big gastrin" (designated G-34 because it con- cular forms of gastrin can be expressed in two different tains 34 amino acid residues) were compared with those ways. Based on exogenous molar doses, the potencies of of pure natural heptadecapeptide gastrins (G-17) from G-34 and G-17 were similar. However, based on molar human and porcine sources. Radioimmunoassay inhibi- increments in serum gastrin concentration, G-17 was tion curves indicated that G-17 was nearly 1.5 times more approximately five times more potent than G-34. Hence, potent than G-34 with the antibody used in this study. fractionation of these gastrin components may be im- This difference was confirmed by demonstration of in- portant in estimation of the acid-stimulating action creased immunoreactivity generated when G-34 was represented by total serum gastrin as measured by radio- converted to G-17 by trypsinization. immunoassay. When infused intravenously into dogs with gastric fistulas and Heidenhain pouches in equimolar doses, INTRODUCTION G-34 produced slightly higher acid secretory responses Radioimmunoassay systems currently employed for mea- than G-17. Responses to sulfated and nonsulfated forms surement of gastrin all use heptadecapeptide gastrin were not significantly different, nor were responses to (G-17)' as the standard, most commonly nonsulfated hu- human and porcine G-17. man gastrin I (HG-17-I), porcine gastrin I (PG-17-I), During infusion of equimolar doses, steady-state serum or sulfated porcine gastrin II (PG-17-II) (1-3). Yalow gastrin concentrations were more than fivefold higher and Berson recently reported that the principal circulat- with G-34 than with G-17. The difference in steady-state ing form of gastrin in hypergastrinemic humans was a blood concentrations could be accounted for by a cor- larger molecule which they named "big gastrin" (4). responding difference in removal rates. The half times This observation has been extended by them (5, 6) and of the G-34 preparations averaged 15.8 min and the half has been confirmed by others (7, 8). times of the G-17 preparations averaged 3.2 min. The Gregory and Tracy extracted human big gastrin from calculated spaces of distribution for G-17 and G-34 were gastrin-secreting tumors of the Zollinger-Ellison variety similar, about 25% of body weight. and have showed that it contains 34 amino acid resi- When the increment in serum gastrin was plotted dues (9). Both sulfated and nonsulfated forms were against acid secretory response it was found that nearly identified. Throughout the remainder of this paper five times greater increments in molar concentrations of HG-34-I and HG-34-II will be used to designate the G-34 than of G-17 were required to produce the same nonsulfated and sulfated forms of human big gastrin. The purpose of the present investigation was to com- This work was presented in part at the Annual Meeting of the American Gastroenterological Association, New pare G-34 and G-17 preparations in regard to acid stim- York, May 1973, and a preliminary report appeared in ulation, rates of elimination from the circulation, and abstract form' 1973. Gastroenterology. 64: 873. Dr. Grossman is a Veterans Administration Senior 1 Abbreviations used in this paper: G-17, little gastrin; Medical Investigator and Dr. Walsh is a Veterans Ad- G-34, big gastrin; HG-17-I, nonsulfated human little gas- ministration Clinical Investigator. trin; HG-34-I, nonsulfated human big gastrin; HG-34-II, Received for publication 21 Novenmber 1973 and in revised sulfated human big gastrin; PG-17-I, nonsulfated porcine form 18 February 1974. little gastrin; PG-17-II, sulfated porcine little gastrin. The Journal of Clinical Investigation Volume 54 August 1974 477-485 477 the relation between acid secretion rates and change in samples were obtained for serum gastrin determination. gastrin. Acid secretion was measured in consecutive 10-min periods serum immunoreactive during infusions of gastrin and afterward until acid secre- tion had returned to basal rates. 5-ml blood samples were METHODS obtained every 15-30 min during infusion and at 2- to 5-min Gastrins. All gastrin preparations were generous gifts intervals after infusion was discontinued. As many as 22 from Professor R. A. Gregory and Dr. Hilda Tracy, Uni- samples were obtained from each dog. Serum was separated versity of Liverpool. Porcine gastrins, PG-17-I and PG- and stored at -20'C. All samples obtained during a study 17-II, were obtained from extracts of hog antral mucosa with a pair of peptides were measured by radioimmunoassay (10). Natural human gastrins, HG-34-I, HG-34-II, and at the same time. Acid responses were taken as the highest HG-17-I were obtained from gastrin-secreting tumors of the three consecutive 10-min outputs for each dose and each Zollinger-Ellison variety. Each peptide was purified by output was normalized by dividing by the peak 30-min AE-cellulose chromatography and column electrophoresis pentagastrin output and multiplying the result by 100. (11). The purity of each of the gastrin preparations was Steady-state serum gastrin values were taken as the mean proved by homogeneity in these systems and by quantitative of the last two values obtained during infusion. For deter- amino acid analysis. mination of half time, basal gastrin concentration was sub- Animals. Four mongrel dogs weighing between 20 and tracted from all values. All values for each dog were 26 kg were prepared with gastric fistulas drained by normalized by expressing them as a percentage of steady- Thomas cannulas (12) and Heidenhain pouches drained by state value; then the mean of normalized values of the four Gregory cannulas (13). Experiments were performed at dogs was taken for each time period, converted to the least 3 mo after operation. The dogs were deprived of natural logarithm, and regression of ln serum gastrin vs. food but not water for 18 h before each test. Experiments time was computed to yield the slope k. from which the were performed no more than twice per week. disappearance half time was determined by dividing into The maximal acid secretory capacity of each pouch was 0.693. Clearance rate was determined by dividing the in- determined by stimulation with pentagastrin (Ayerst Labo- fusion rate in picomoles per kilogram per minute by the ratories, New York) at doses of 8 and 16 ,tg/kg-h. Each steady-state increment in serum gastrin in picomoles per dose was given on separate days and each dog was studied liter for the two or three highest doses and expressed as twice. The highest rate of acid secretion obtained for three an average in liters per kilogram per minute. consecutive 10-min periods was taken as the maximal penta- Space of distribution was calculated by use of the plateau gastrin response. Acid secretory rates were determined as principle described by Goldstein, Aronow, and Kalman (17) previously described (14) by titration to pH 7 on an auto- from the formula: titrator (Autoburette, Radiometer Co., Copenhagen). All V = D/[ (G) (k.)], other acid secretion rates were expressed as percent maxi- mal pentagastrin-stimulated secretion to minimize variations where V = volume of distribution as fraction of body weight, due to differences in the sizes of the pouches. D = dose of gastrin expressed as picomoles per kilogram Gastrin solutions were prepared in 0.05 M NH4HCO3. per minute, G = steady-state blood level of gastrin expressed Absorbance at 280 nm was measured in a Zeiss PMQ-II as picomoles per liter, and k.
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