Rat Pancreatic Protein Research Articles

Adaptive changes in translation initiation activities for rat pancreatic protein synthesis with feeding of a high-protein diet

We have previously demonstrated that dietary protein induced pancreatic hypergrowth in pancreaticobiliary diverted (PBD) rats. Dietary protein and dietary amino acids stimulate protein synthesis by regulating translation initiation in the rat skeletal muscle and liver. The aim of the present study was to determine whether feeding a high-protein diet induces activation of translation initiation for protein synthesis in the rat pancreas. In PBD rats in which the bile–pancreatic juice was surgically diverted to the upper ileum for 11–13 days, pancreatic dry weight and protein content were doubled compared with those in sham rats and further increased with feeding of a high-protein diet (60% casein diet) for 2 days. These pancreatic growth parameters were maintained at high levels for the next 5 days and were much higher than those of sham rats fed a high-protein diet. In both sham and PBD rats, feeding of a high-protein diet for 2 days induced phosphorylation of eukaryotic initiation factor 4E-binding protein 1 and 70-kDa ribosomal protein S6 kinase, indicating the activation of the initiation phase of translation for pancreatic protein synthesis. However, this increased phosphorylation returned to normal levels on Day 7 in PBD but not in sham rats. We concluded that feeding a high-protein diet induced pancreatic growth with increases in the translation initiation activities for pancreatic protein synthesis within 2 days and that prolonged feeding of a high-protein diet changed the initiation activities differently in sham and PBD rats.

Manganese-stimulated phosphorylation of a rat pancreatic protein: identity with elongation factor 2

To investigate the effect of Mn 2+ on pancreatic protein phosphorylation, we incubated rat pancreatic cytosol in Tris buffer (pH 7.5) with [γ- 32P]ATP. Analysis using sodium dodecyl sulphate polyacrylamide gel electrophoresis and autoradiography revealed a single protein (p98), with an M r of 98 000 and p I of 6.4–6.5, which was phosphorylated in a dose-dependent manner by Mn 2+. A threshold effect was observed at 35 μM, and maximal effect at 1.1 mM Mn 2+. Ca 2+ and calmodulin (CaM) did not cause p98 phosphorylation, but Mg 2+ (10 mM) caused faint non-specific phosphorylation of p98. Ca 2+ (0.03–3 mM) and CaM (1–10 μg/ml) significantly enhanced, whereas trifluoperazine (TFP) and Mg 2+ inhibited Mn 2+-stimulated p98 phosphorylation. Under the above incubation conditions, Mn 2+-stimulated protein phosphorylation of p98 was also observed in isolated pancreatic acini, but not in cytosols from liver or kidney. Partial purification of p98 and amino acid sequencing of the protein band corresponding to p98 indicated complete sequence homology with rat elongation factor 2 (EF-2). Furthermore, the combination of Ca 2+, Mg 2+ and CaM, which is known to induce the phosphorylation of EF-2, mimicked the actions of Mn 2+. Inasmuch as EF-2 is the major substrate for CaM-dependent protein kinase III (CaM-PK III), these studies suggest that in the pancreatic acinar cell Mn 2+/CaM protein kinase activity is mediated via CaM-PK III and that Mn 2+ participates in the regulation of this enzyme in the pancreas.

Inhibition of Polyamine Synthesis by α-Difluoromethylornithine and its Effects on Pancreatic Secretion and Growth in the Rat

The role played by the polyamines in mediating the pancreatic growth and secretory responses to hormonal stimulation is uncertain. The effect of an inhibitor of ornithine decarboxylase (ODC), alpha-difluoromethylornithine (DFMO), on rat pancreatic protein secretion and synthesis and on growth in response to hormonal stimulation was therefore studied. Anesthetized rats were given an intravenous injection of DFMO (50, 100, or 150 mg/kg), followed by a 7-h continuous infusion (15, 25, or 35 mg/kg/h, respectively). After a basal 1-h period an intravenous infusion of 2.5 micrograms/kg/h of the cholecystokinin-like peptide Thr28Nle31CCK25-33 (CCK-LP) was added and continued for 6 h. The control rats received CCK-LP only. The ODC activity in the pancreas was markedly reduced by DFMO, but DFMO did not affect pancreatic juice volume or protein output. In another series conscious rats were given a continuous intravenous infusion of 2.5 micrograms/kg/h of CCK-LP for 8, 24, and 48 h or 5.0 micrograms/kg/h of secretin for 8 and 48 h, with or without DFMO (100 mg/kg as an injection initially and thereafter 25 mg/kg/h). The ODC activity and putrescine concentration in the pancreas were significantly reduced by DFMO at 8 and 24 h but not at 48 h. DFMO also significantly reduced the activities of RNA polymerase, DNA polymerase, and thymidine kinase at 24 h, but not at 48 h. The present study thus indicates that polyamines play a role in the initiation of the growth response to hormonal stimulation but does not support a similar dependence for early pancreatic protein synthetic and secretory responses.

Synthesis and secretion of rat pancreatic proteins by Xenopus laevis oocytes.

An in vivo translation system, the Xenopus laevis oocyte, was employed to study the synthesis and secretion of pancreatic proteins. RNA was purified from normal and diabetic rat pancreas and normal rat liver by use of guanidine isothiocyanate lysis and cesium chloride gradient centrifugation. The presence of functional mRNA was documented by translation in a reticulocyte lysate that yielded precursors of all major secretory proteins, i.e., slightly higher Mr than proteins synthesized in situ by pancreatic acini. Mature X. laevis oocytes were then microinjected with either total RNA or purified mRNA. When oocytes were subsequently incubated with 35S-methionine, pancreatic secretory proteins or hepatic albumin could be immunoprecipitated from oocyte lysate with specific polyclonal antibodies against amylase, trypsin, ribonuclease, and albumin. Amylase was shown to be enzymatically active. Moreover, oocytes released pancreatic secretory proteins into the medium when injected with pancreatic RNA in a time-dependent manner. Only the mature form of amylase was secreted and secretion was not regulated by secretagogues. When a comparison was made after injection of RNA from diabetic pancreas known to contain altered amounts of individual mRNAs, there was a decrease in amylase and an increase in trypsinogen synthesis in oocytes that was comparable to the results of cell free translation. The oocyte expression system, therefore, should be useful not only for studies of protein synthesis but also for processing and secretion.

Evidence in vivo of asynchronous intracellular transport of rat pancreatic secretory proteins

Labeled proteins which appeared in pancreatic juice after the intravenous (i.v.) injection of [35S]methionine into conscious rats with chronic pancreatic duct fistulae were separated by gel electrophoresis and measured by determination of the radioactivity of each of the separated bands. Radioactivity appeared in the secreted proteins 20 min after injection of the label. In the subsequent 10 min, 6.44% of the radioactivity was found in trypsinogen, whereas 100 min later only 3.4% of the radioactivity was associated with this enzyme. The values at 10 and 100 min for amylase were 10.85% and 21%, respectively, showing an earlier appearance of labeled trypsinogen than of amylase. Chymotrypsinogen behaved similarly to trypsinogen. Early secretion of labeled proteases was also demonstrated by separation of pancreatic proteins by two-dimensional gel electrophoresis followed by fluorography. In pancreatic duct cannulated rats, zymogen granules were prepared 30 and 60 min after injection of the labeled methionine. Determination of the radioactivity of the individual proteins demonstrated a similar time course of the labeling pattern in the zymogen granule fraction to that in pancreatic juice. The results of the experiments suggest an asynchronous secretion of newly synthesized rat pancreatic proteins.

Influence of Anticholinergic Treatment on Rat Pancreatic Protein and Amylase Concentrations

Pancreatic protein and amylase concentrations were measured after subcutaneous administration of an anticholinergic drug (Pro-Banthine to rats in different doses. In doses comparable to those given in acute pancreatitis an increase of both pancreatic protein and amylase concentrations was found. If pancreatitis is indeed caused or maintained by proteases released from the acinar cell, anticholinergics--giving rise to an accumulation of enzymes within the pancreas--may do more harm than good in the treatment of acute pancreatitis.

Non-parallel secretion of newly synthesized rat pancreatic proteins after feeding a diet containing raw soybean flour

The relative rate of secretion of rat pancreatic proteins was studied in vivo using a double label method. Rats were injected with [ 3Hleucine and after different time intervals with [ 14C]leucine. At a fixed time after administration of the second precursor the animals were killed, and the pancreatic proteins were separated by polyacrylamide gel electrophoresis. The dpm of tritium to dpm of 14C ratio of several identified enzymes was assessed. The percentage secretion of a newly synthesized secretory protein was derived from the difference between the actual 3H/ 14C ratio and the 3H/ 14C ratio that was found for non-secretory proteins. In pancreata of rats fed with a standard diet several identified proteins, viz. three trypsinogens, chymotrypsinogen and three amylases were secreted in “parallel”. When a diet containing raw soybean flour was fed, the secretory pattern for the amylases differed from that of the other proteins.

Changes in Rat Pancreatic Protein Synthesis After a Single Feeding with Diets Containing Raw or Heated Soybeans

The patterns of protein synthesis following the administration of a single meal containing defatted ground soybeans (RSD) or heated defatted ground soybeans (HSD) were compared. A double label method was used so that the determination of the relative rate of synthesis of an enzyme could not be obscured by a possible alteration of the activity or quantity of the enzyme. Rats were given one meal of RSD or HSD and were subsequently killed at different times after the meal. Eight hours after the meal, the relative rate of synthesis of one of the three trypsinogens was substantially increased with RSD feeding, whereas that of the amylases and one chymotrypsinogen were somewhat lower. The synthetic rate of lipase, ribonclease, proelastase, another chymotrypsinogen and of two trypsinogens was unaffected when feeding RSD is compared to feeding HSD. The relative rate of synthesis of one of the trypsinogens was unaffected 8 hours after RSD feeding, but was increased 16 hours after RSD feeding. Actinomycin D could suppress the effects of RSD feeding on the protein synthetic rate of some, but not of all, secretory proteins.

Effects of atropine on pigeon pancreas.

ARTICLESEffects of atropine on pigeon pancreasJ Morisset, and PD WebsterJ Morisset, and PD WebsterPublished Online:01 Nov 1970https://doi.org/10.1152/ajplegacy.1970.219.5.1286MoreSectionsPDF (1 MB)Download PDF ToolsExport citationAdd to favoritesGet permissionsTrack citations ShareShare onFacebookTwitterLinkedInWeChat Previous Back to Top Next Download PDF FiguresReferencesRelatedInformation Cited ByLife with the pancreas: A personal experienceAdvances in Medical Sciences, Vol. 65, No. 1Hormonal control of pancreatic growth during fetal, neonatal and adult lifeAdvances in Medical Sciences, Vol. 53, No. 2An embryonic chick pancreas organ culture model: characterization and neural control of exocrine releaseAutonomic Neuroscience, Vol. 105, No. 2Conservation of Digestive EnzymesStephen Rothman, Charles Liebow, and Lois Isenman1 January 2002 | Physiological Reviews, Vol. 82, No. 1Comparative inhibitory effects of pirenzepine and atropine on cholinergic stimulation of exocrine and endocrine rat pancreasGastroenterology, Vol. 89, No. 2Surgical Management of Acute PancreatitisMayo Clinic Proceedings, Vol. 59, No. 4Duct, exocrine, and endocrine components of cultured fetal mouse pancreasIn Vitro, Vol. 18, No. 9Exocrine pancreatic secretion in the syrian golden hamster Mesocricetus auratus—I. Basic valuesComparative Biochemistry and Physiology Part A: Physiology, Vol. 66, No. 3Influence of Anticholinergic Treatment on Rat Pancreatic Protein and Amylase Concentrations23 February 2010 | Scandinavian Journal of Gastroenterology, Vol. 14, No. 1Medical treatment of acute pancreatitisGastroenterology, Vol. 74, No. 3Neurohormonal control of pancreatic secretionGastroenterology, Vol. 74, No. 2COMPARATIVE INHIBITORY EFFECTS OF 3-QUINUCLIDINYL BENZILATE (QNB) AND ATROPINE ON AMYLASE RELEASE FROM RAT PANCREAS19 July 2012 | British Journal of Pharmacology, Vol. 61, No. 1Respiratory exchanges and acid-base balance during perfusion ofex vivo isolated pancreasThe American Journal of Digestive Diseases, Vol. 21, No. 5Secretion of Gastric and Pancreatic Juice, pH of Tract, Digestion in Alimentary Canal, Liver and Bile, and AbsorptionEffect of Ethanol on Pancreatic Enzyme SecretionGastroenterology, Vol. 67, No. 5Effect of bethanechol chloride on protein synthesis in rat pancreasThe American Journal of Digestive Diseases, Vol. 19, No. 2AtropineThe neurohumoral control of pancreatic exocrine secretionThe American Journal of Clinical Nutrition, Vol. 26, No. 3Effect of Drugs on Pancreatic Amylase Secretion in VitroGastroenterology, Vol. 63, No. 3Effects of Fasting and Feeding on the PancreasGastroenterology, Vol. 62, No. 4 More from this issue > Volume 219Issue 5November 1970Pages 1286-1291 Copyright & PermissionsCopyright © 1970 by American Physiological Societyhttps://doi.org/10.1152/ajplegacy.1970.219.5.1286PubMed5473111History Published online 1 November 1970 Published in print 1 November 1970 Metrics

The effect of sodium taurodesoxycholate and pH on the gel filtration behaviour of rat pancreatic protein and lipases

Gel filtration of lyophilized rat pancreatic juice has been carried out on Sephadex G-100 at pH 5.8 and 8.0, both in the presence and absence of sodium taurodesoxycholate, to determine whether bile salts affect the gel filtration behaviour of pancreatic juice proteins or the enzymes hydrolyzing emulsified triolein (triolein lipase) and micellar mono-olein (mono-olein lipase). At pH 5.8 and at pH 8.0 sodium taurodesoxycholate, in concentrations above its critical micellar concentration, changed the elution volume of triolein lipase only slightly. It did, however, significantly increase the recovery of applied activity in the column eluent. This increased recovery appeared to be due to the elution of a lipase activator with the lipase in the presence of the bile salts. In the absence of sodium taurodesoxycholate this activator was separated from the lipase with consequent loss of activity. At pH 5.8 sodium taurodesoxycholate changed the protein elution pattern markedly, protein being eluted earlier, as a single peak instead of a multicomponent complex. At least some of this change was probably due to desorption of amylase from the column. Mono-olein lipase was also eluted earlier in the presence of sodium taurodesoxycholate. At pH 8.0, proteins were eluted earlier than at pH 5.8 and the effect of sodium taurodesoxycholate was less marked. Mono-olein lipase was eluted with the same K av at pH 8.0 as at pH 5.8, and showed a similar change in elution volume in the presence of sodium taurodesoxycholate at both pH's. Studies with 35S-labelled sodium taurodesoxycholate failed to demonstrate any binding of this bile salt to proteins or either lipase. It is concluded that sodium taurodesoxycholate does not change the molecular size or configuration of triolein lipase sufficiently for this to be detected by gel filtration. It does, however, promote the elution of a lipase activator together with the enzyme, resulting in an improved yield of lipase after chromatography.