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A R Beaudoin

Publications and source records attributed to A R Beaudoin.

At least 73 records · Page 4Linked to original sources
PubMed1988

Heterogeneity of the zymogen granule membranes in rat pancreas.

Zymogen granules (ZG) of rat pancreas have been isolated by the procedure of Pâquet et al. The granules lysed when exposed to alkaline pH (pH 8.2), and their membranes could be subfractionated by centrifugation on a sucrose gradient. Four discrete types of membranes corresponding to densities of 1.105, 1.085, 1.075, and 1.020 were obtained, designated types A, B, C, and D, respectively and characterized both by morphological and biochemical criteria. Electrophoretic profiles showed that they contain the same protein bands but in different proportions. Type A membranes are comprised of four major bands corresponding to molecular weights of 80, 69, 54, and 20 kDa, being in higher concentration than the others. Types B and C contain three major bands at 80, 54 and 20 kDa whereas type D is comprised of only two major bands at 69 and 54 kDa, the latter polypeptide corresponding to ATP-diphosphohydrolase activity which is present in all four membrane types. Freeze-fracture of rapidly frozen membranes, followed by transmission electron microscopy (TEM) showed that type A are large superimposed sheets of membranes with amorphous material between sheets. The surface area of these sheets corresponds grossly to the surface of an intact ZG with a few intramembrane particles (IMP) distributed at random or in small aggregates on large smooth fracture planes. Types B and C exhibit a totally different aspect, forming closed vesicles about the size of a small ZG with few IMP distributed at random or in small aggregates on smooth fracture planes. Type D membranes are very small vesicles with no detectable IMP on relatively smooth fracture planes.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

Lipid analysis of a novel type of cell secretion in the exocrine pancreas: the pancreasomes.

A novel type of cell secretion termed 'microvesicular secretion' has been described recently in the exocrine pancreas. According to this process, microvesicles are released by acinocytes in the pancreas acinar lumen. These microvesicles, identified as 'pancreasomes', were characterized by the presence of a major glycoprotein component originating in the exocrine acinar cell. In the present work, phospholipids of pancreasomes have been identified. Five classes of phospholipid were found: phosphatidylcholines, phosphatidylethanolamines, lysophosphatidylcholines, sphingomyelins and another minor class of ninhydrin-positive phospholipid (phosphatidylserines or lysophosphatidylethanolamines). The ratios of neutral lipids to phospholipids were particularly high (3:1), as estimated by GLC of their fatty acid content. Analysis of fatty acid composition of pancreasomes lipids revealed a very high proportion of two saturated fatty acids, palmitic (40%) and stearic (24%), whereas two main unsaturated fatty acids, oleic (17%) and linoleic (8%), were found in smaller proportions. Differential scanning calorimeter studies on washed pancreasomes indicated that there was no lipid phase transition in their membrane, despite the absence of cholesterol. Our observations show that pancreasomes have an unusual lipid composition and confirm our previous conclusion based on protein analysis that the release of pancreasomes occurs according to an hitherto undescribed type of secretion, in which a glycoprotein is released associated with specific domains of the luminal plasma membrane.

Animals↗
PubMed1987

Pathways of secretion in the exocrine pancreas: the status of resting secretion.

In the last decade the concept of two distinct pathways of secretion in the exocrine pancreas has slowly emerged. According to this concept, one pathway is involved in stimulated (regulated) conditions and another under resting (constitutive) conditions. This hypothesis was elaborated at first from the comparison of the specific radioactivities of secretory proteins released by the gland under resting and stimulated conditions. Analysis of the protein composition of the juice released under these two physiological conditions further supported that hypothesis. More recent studies compared the kinetic of accumulation of newly synthesized proteins in zymogen granule and their release in the gland lumen. The latter results are in agreement with a model in which secretory proteins are channelled in two separate pathways, one regulated, and one constitutive. Essentially, the constitutive pathway would correspond to a paragranular route in which the proteins would be immediately secreted instead of being stored in zymogen granules. In addition, some of the proteins released in the juice under "resting" conditions are associated to microvesicles. The term "microvesicular secretion" is used to designate that type of secretion.

Animals↗
PubMed1986

Calcium binding properties of purified zymogen granule membrane of pig pancreas. Evidence for calcium binding proteins.

Ca2+ binding properties of purified zymogen granule membranes of pig pancreas have been measured: Binding increased linearly with Ca2+ concentration in the medium up to the micromolar range; in the millimolar range a sharp rise in binding capacity was observed. Binding increased with pH both at low and high concentrations of Ca2+. It was insensitive to Na+ and K+ ions at concentrations up to 100 mM. Mg2+ was inhibitory in the millimolar range whereas La2+ and Tb3+ were inhibitory in the micromolar range. The Ca2+ binding components of zymogen granule membranes were identified by two methods: (1) by measuring 45Ca2+ binding after counter-ion electrophoresis and (2) by Stain's-all (forms a complex with Ca2+ binding proteins absorbing maximally at 600 nm), after SDS-polyacrylamide gel electrophoresis. The first method, counter-ion electrophoresis, indicated that most of the 45Ca2+ was associated with an acidic band which could be subsequently subfractionated by SDS-polyacrylamide gel electrophoresis in five bands: 66, 57, 30, 27 and 22.5 kDa. The second method, Stain's-all, revealed six positive polypeptides after SDS-polyacrylamide gel electrophoresis of native zymogen granule membranes' two were unreactive after neuraminidase treatment (130 and 92 kDa, respectively), whereas four other bands were still reactive (66, 57, 43, 30 kDa, respectively.) Ca2+ binding was also measured on intact zymogen granules: the binding capacity was higher than for zymogen granule membranes. Among the Ca2+ binding proteins of the zymogen granule membrane only one is apparently located on the granule external surface: the 30 kDa polypeptide. If Ca2+ directly facilitates fusion of zymogen granules with plasma membrane by a Ca2+-protein interaction, then this protein is a presumptive candidate to play such a key role.

Animals↗
PubMed1986

Asynchronism between amylase secretion and packaging in the zymogen granules of pig pancreas.

By pulse and chase experiments with [3H]leucine we have followed the time-course packaging of newly synthesized amylase in the zymogen granules of pig pancreas in vivo. In parallel, with a second group of pigs, we have followed the time-course release of newly synthesized amylase. The ratios of specific radioactivities of amylase in the zymogen granule over that of the homogenate indicated that substantial amounts of newly synthesized amylase had already reached the zymogen granule compartment at 50-min postpulse. Specific radioactivity increased up to 180 min, then a plateau was obtained. In the juice with the other group of pigs, starting at 40 min postpulse, amylase specific radioactivity increased up to about 100 min, then slowly decreased in the next hours. These results show an asynchronism between packaging of newly synthesized amylase in the zymogen granule compartment and its release in the juice under resting conditions, supporting the concept of a partial bypass of the zymogen granule compartment.

Amylases↗
PubMed1986

Detection and characterization of microvesicles in the acinar lumen and in juice of unstimulated rat pancreas.

Small vesicles were visualized in the lumen of rat pancreas acini by freeze-substitution and conventional electron microscopy. Microvesicles were subsequently isolated from pancreatic juice. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that these vesicles contain only one major protein. The major protein was identified by an immunoblot technique as GP-2, an 80 kD glycoprotein also found in the zymogen granule membrane. The immunocytochemical localization of rabbit anti-GP-2 and anti-amylase by the protein A-gold technique confirmed that GP-2 was associated with clusters of microvesicles, whereas amylase was virtually excluded. Freeze-fracture of the microvesicles revealed that their membrane was devoid of intramembrane particles. Biochemical analysis indicated also that the membrane did not contain any detectable cholesterol. These results demonstrate that GP-2 is released from the acinar cell in the gland lumen within microvesicles by a hitherto undescribed mode of secretion.

Animals↗
PubMed1986

Steroids and the secretory function of the exocrine pancreas.

The influence of steroids on the exocrine pancreas of male rats was examined by removing steroid producing tissue and by introducing individual steroids into these animals at a later date. Castration had no demonstrable morphological effect on acinar cells, whereas castration combined with adrenalectomy caused a marked depletion of zymogen granules as well as widening of peri- and interlobular spaces. Treatment of castrated-adrenalectomized rats with estradiol restored a normal appearance of the pancreas within about 9 h. Triamcinolone acetonide produced similar results. These morphological changes were accompanied by significant alterations of the relative amounts of digestive enzymes present in zymogen granules. A marked reduction of amylase occurred in the castrated-adrenalectomized group. Neither estradiol nor triamcinolone could reverse these effects within 9 h. Castration alone had no significant effect on the relative proportion of amylase; however, it increased the relative amount of proteases. This effect was reversed by estradiol treatment. Estradiol also induced significant changes in the proportion of proelastase in castrated-adrenalectomized animals. Replacement therapy in castrated-adrenalectomized rats with dexamethasone or triamcinolone partially restored the level of amylase in the pancreas, whereas estradiol did not cause any significant effect. At the ultrastructural level, castration and adrenalectomy caused swelling of the Golgi apparatus and accumulation of condensing vacuoles. These effects were reversed by either estradiol or triamcinolone. Also, after acute treatment of these animals with estradiol, two unusual features of acini were noted. In some acini, a type of granule that we termed halo granule appeared. These halo granules were either distinctly separated organelles or formed composite structures that did not appear to be associated with the luminal membrane. Freeze-fracture studies revealed that secretory vesicles in apparently normal acini adhered to each other at specific contact sites characterized by aggregates of intramembrane particles. Multiple sites of contact could be seen in the same vesicle. From our observations it is clear that steroids from the testes and adrenals exert major effects on the secretory apparatus of pancreas; more specifically on the mechanisms that determine the proportions of the different digestive enzyme and on their packaging in the zymogen granules.

Adrenalectomy↗
PubMed1985

Different patterns of proteins are secreted by the pig pancreas when stimulated by secretin alone or in combination with caerulein.

The protein compositions of pig pancreatic secretions collected under stimulation by secretin alone or in combination with caerulein were compared by SDS polyacrylamide gel electrophoresis. Different sets of proteins were observed in these two different conditions. One of the major proteins secreted under secretin alone was immunologically similar to the 92 kDa glycoprotein characteristic of the pig zymogen granule membrane. Since its proportion in the two secretions was drastically different and since this protein is exclusively found in the acinar cell, these observations support the view that the proteins released by the pig pancreas under secretin stimulation alone, and under the combination of secretin + caerulein do not originate from the same intracellular pool of the acinar cell and that the secretin-induced secretion does not derive from zymogen granules.

Animals↗
PubMed1985

The embryotoxicity of gossypol.

In utero development was analyzed in pregnancies that resulted from matings between gossypol-treated male rats and untreated female rats and in pregnancies in which gossypol was administered to the pregnant rat only. Gossypol treatment of males had no effect on the outcome of pregnancy. There was no significant effect on resorption, fetal growth, or malformation rate. Similarly, gossypol administered to pregnant dams at stages during organogenesis had no observable effect on pregnancy. Under the conditions of this experiment, gossypol administered to either the breeding male rat or the pregnant female rat had no demonstrable adverse effect on development in utero.

Abnormalities, Drug-Induced↗
PubMed1985

Electrophoretic and cytological evidence for heterogeneity of pancreatic acinar cell responsiveness to carbachol, caerulein and secretin.

Incubation of rat pancreatic lobules for 90 min with optimal concentrations of caerulein, carbachol or secretin caused the release of about 30% of the amylase content. Combination of secretin with carbachol or caerulein increased the amylase output to about 40%. With secretin, as with carbachol or caerulein, heterogeneity of cellular responsiveness was observed, some acini being partially or completely depleted of their zymogen granules, whereas others appeared to be resting. When secretin was combined with carbachol or caerulein, granule depletion, originally confined to small groups of neighbouring acini, spread to form large areas of degranulated cells, sometimes comprising a whole section of a lobule. In dispersed acini, under the same conditions, carbachol caused the release of about 60% of the amylase content, and secretin 40%. When both secretagogues were combined, a significant increase to 78% was observed. Under these conditions, there was some important cellular damage, as indicated by the release of 20% of the amylase content and between 6 and 12% of lactate dehydrogenase into the media, in the absence of stimulus. These results were corroborated by cytological observations. On the basis of their secretory response two groups of acini can be distinguished, those that respond to carbachol, caerulein or secretin and those that respond to the combination of secretin with carbachol or caerulein. Electrophoretic patterns of secretory proteins released by lobules stimulated by these different types of secretagogues were essentially similar. The pattern was quite different, however, in the absence of a stimulus.(ABSTRACT TRUNCATED AT 250 WORDS)

Amylases↗
PubMed1985

beta-NADPHase- and TMPase-positive "snake-like tubules" in the exocrine pancreas: cytochemical and immunocytochemical studies.

Using five different protocols, two enzymes, nicotinamide adenine dinucleotide phosphate phosphohydrolase (beta-NADPHase) and sodium trimetaphosphatase (TMPase), were localized in the acinar cell of rat pancreas by ultrastructural cytochemistry. The beta-NADPHase cytochemical localization was realized at pH 4.8 and pH 3.9. At pH 4.8, the beta-NADPHase activity was found in the Golgi intermediate saccules, lysosomes, gland lumen, and tubular structures, described as snake-like tubules (Beaudoin AR, Grondin G, Lord A: Eur J Cell Biol 33:275, 1984; Beaudoin AR, Grondin G, Lord A, Pelletier M: In Proc 42nd Ann Meeting Electron Microscopy Soc Am. San Francisco Press, CA, 1984). There was no detectable beta-NADPHase activity at pH 3.9. The TMPase cytochemistry was done at pH 3.9 according to Oliver (J Histochem Cytochem 28:78, 1980) and at pH 3.9 and 4.8 with the medium described by Berg (J Histochem Cytochem 8:92, 1960). TMPase localization varied according to the protocols. It was found in tubular structures described as "basal lysosomes," lysosomes, and zymogen granules, whereas Golgi saccules were generally negative. Our observations showed that the structures identified as "basal elongated lysosomes" and revealed by TMPase (Oliver C: J Histochem Cytochem 28:78, 1980; J Histochem Cytochem 31:1209, 1983) were morphologically similar to snake-like tubules (SLT) revealed by beta-NADPHase. Relationships between SLT and mitochondria as well as lysosomes and plasma membranes were observed. Using amylase-specific antibodies, it was also shown, by the protein A-gold immunocytochemical technique, that SLT do not contain amylase and, in fasting conditions, would not be involved in the transport of secretory proteins.

Acid Anhydride Hydrolases↗
PubMed1984

Cytological effects of ionophore-induced stimulation on the exocrine pancreas of the rat.

Rat-pancreas lobules were incubated with the ionophore A-23187 in the presence of Ca2+. After 90 min, some of the acini were partially or almost completely depleted of their zymogen granules while others had the appearance of resting acini. With few exceptions, the cells of a given acinus were degranulated to a comparable level. Slight dispersion of the zymogen granules was noticed in cells incubated in a Ca2+-free medium containing EGTA with or without A-23187. In the presence of Ca2+ the secretory response obtained with the ionophore was comparable to that observed with 10(-5)M urecholine. The results obtained provide cytological evidence that the secretory response is only partially determined at the membrane-receptor level and that other mechanisms intervene between cytosol Ca2+ increase and exocytosis.

Amylases↗
PubMed1984

Secretagogues cause a preferential discharge of large size granules in rat pancreas.

We have compared the diameters of zymogen granules in the exocrine pancreas of fasted or stimulated rats. The average granule diameter was 0.73 microns +/- 0.18 SD (sample size, 1460) in fasted animals and significantly reduced to 0.68 microns +/- 0.17 SD (sample size, 860) after 120 min and two intraperitoneal injections of urecholine. Reduction of granule size was attributable to the preferential discharge of large size granules. The range between first and third quartiles of the distribution curve was identical in the two groups at 0.24 microns. There was no significant increase in the proportion of granules smaller than 0.40 microns after urecholine stimulation. A third group of animals was "hyperstimulated" by an infusion of a mixture of caerulein, secretin, and urecholine. After 210 min, the average granule diameter was reduced to 0.43 microns +/- 0.14 SD (sample size, 786). The range between first and third quartiles of the distribution curve was 0.16 microns. In this group, 43.5% of the granules was smaller than 0.40 microns. Granule size reduction was accompanied by the appearance of numerous pleiomorphic condensing vacuoles. The present results strongly support the views that secretagogues cause the preferential release of large size granules and favor the formation of small size granules.

Animals↗
PubMed1984

Ultrastructural localization of nicotinamide adenine dinucleotide phosphatase (NADPase) activity in the exocrine pancreas of rat: identification of a novel structure.

NADPase activity has been localized in the exocrine pancreas of rat, by cytochemistry according to the procedure of Smith as modified by Clermont et al. With NADP or NADPH as substrate, an intense reaction was detected in one or two intermediary saccules of the Golgi stack. Reaction product was also present in lysosomes, dense bodies and the gland lumen. It was absent from condensing vacuoles and zymogen granules. A very intense reaction was found over a "snake-like" structure not previously reported. These are elongated tubules located in basal and central portions of the acinar cell where they are frequently seen close to the Golgi stack or the basolateral cell surface.

Animals↗
PubMed1983

Evidence that amylase is released from two distinct pools of secretory proteins in the pancreas.

Previous experiments demonstrated the existence of at least two pools of secretory proteins in the exocrine pancreas. We have measured the specific activities of amylase released under resting conditions and of amylase in the zymogen granules. Specific activity of resting secretion was twice that found under stimulated conditions or in zymogen granules. Secretory proteins were pulse-labeled and amylase was measured after precipitation of the enzyme with glycogen. Pancreatic juice collected at 45-50 min post-pulse contained 10-25-times the amylase activity found in zymogen granules. These results confirm the existence of at least two distinct pools of secretory proteins in the exocrine pancreas and suggest the existence of an intracellular route of secretory proteins which would bypass the zymogen granule compartment.

Amylases↗
PubMed1983

Sequential hydrolysis of the gamma- and beta-phosphate groups of ATP by the ATP diphosphohydrolase from pig pancreas.

The ATP diphosphohydrolase (EC 3.6.1.5) from pig pancreas hydrolyzes triphospho- and diphosphonucleosides. The reaction products of ATP hydrolysis are ADP, AMP and orthophosphate, but AMP accumulates at a faster rate than ADP. A time-course study showed a simultaneous breakdown of ATP and ADP with initial rates for ATP and ADP hydrolysis of 2.1 and 3.8 mumol/min per mg protein, respectively. However, the rates reached similar values toward the end of the incubation period. According to double reciprocal plots and Dixon plots, the Km values for ATP and ADP are similar, Vmax for ADP hydrolysis is twice the Vmax for ATP hydrolysis and both nucleotides are competitive inhibitors of the other with their Ki values similar to their Km. These results are consistent with a sequential hydrolysis of the two diphosphoester bonds of ATP: ATP first binds to the enzyme, its gamma-phosphate group is hydrolyzed and released, resulting in an enzyme-ADP complex which either breaks down to free enzyme and ADP or is further processed via hydrolysis of the beta-phosphate group, releasing free enzyme, AMP and Pi. The experimental data showed that the processing step is favored.

Adenosine Triphosphate↗
PubMed1983

The effect of aminothiadiazole on glycogenesis and glycogenolysis in fetal and neonatal rat liver.

The effect of the teratogen 2-amino-1,3,4-thiadiazole on glycogenesis and glycogenolysis was investigated in the fetal and neonatal rat liver. At day 15, 16, or 17 of gestation (sperm day = day 0) pregnant Sprague-Dawley rats received a single IP injection of an aqueous solution of aminothiadiazole. Dosages used were teratogenic (100 mg/kg maternal body weight) and nonteratogenic (10 mg/kg). At day 16 some rats received nicotinamide (100 mg/rat) in addition to a teratogenic dose of aminothiadiazole. Livers were recovered for assay at fetal day 20 and postnatal day 1. Only at day 16 did a teratogenic dose induce a significant depression in the fetal activity of glycogen synthetase (to 49.6% of control activity) and glucose-6-phosphatase (to 72.2% of control activity), and in glycogen accumulation (to 72.6% of control accumulation). At day 15, a teratogenic dose significantly depressed glucose-6-phosphatase activity but not glycogen synthetase activity or glycogen accumulation. Nicotinamide, given immediately after aminothiadiazole, was effective in blocking the inhibition. Teratogenic treatment had no effect on the postnatal activity of glucose-6-phosphatase. Apparently some event associated with birth releases the enzyme from its prenatal inhibition. These results demonstrate a parallelism between the perturbing effect of aminothiadiazole on biochemical development and morphological development with respect to time of insult, dose response, and protection with its antiteratogen. The mechanism of action whereby aminothiadiazole depresses the activity of glycogen synthetase and glucose-6-phosphatase remains to be determined.

Animals↗