Anesthetics as teratogens: nitrous oxide is fetotoxic, xenon is not.

Exposure of pregnant rats to the anesthetic nitrous oxide on the ninth day of gestation causes fetal resorption, skeletal anomalies, and macroscopic lesions including encephalocele, anophthalmia, microphthalmia, and gastroschisis. The inert gas xenon, which has anesthetic properties similar to those of nitrous oxide, does not cause teratogenic effects under the same experimental conditions.

[Fatty acid composition and phospholipid pattern in auxotrophs for unsaturated fatty acids (author's transl)]. / Composition en acides gras et profil des phospholipides chez des auxotrophes pour les acides gras insaturés

The relationship between fatty acid composition and phospholipid pattern has been studied in Escherichia coli auxotrophs for unsaturated fatty acids. 1. The presence of a regulatory mechanism which enables the organism to maintain a given fluidity of the lipids has been corroborated using exogenous fatty acids which cause dramatic changes in fatty acid composition. 2. The fatty composition of phosphatidic acid is different from that of the other classes of phospholipids. 3. Changes in fatty acid composition are concomittant with the alteration of the phospholipid pattern. The ratio of phosphatidylglycerol to diphosphatidylglycerol is particularly sensitive to the physical characteristics of the exogenous unsaturated fatty acid. The relative increase in diphosphatidylglycerol is associated with membrane alterations.

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.

Newly synthesized secretory proteins from pig pancreas are not released from a homogeneous granule compartment.

The pancreatic secretion of anesthetized pigs was collected by cannulation after pulse labeling with [3H]leucine. Collection at 5 min intervals started immediately post-pulse labeling up to 85 min. The volume, the protein content and the trichloroacetic acid-precipitable radioactivity of the juice were measured. The specific radioactivity of the secretory proteins was compared to that of a zymogen granule fraction isolated from the same animal. The latter was very much higher. Caerulein stimulation for 5 min at 80 min post-pulse caused a sharp drop in the specific activity of secretory proteins in the juice, to a level lower than that of the zymogen granule content. These data support the concept of more than one pool of secretory proteins in the pancreas and are incompatible with the concept that secretory proteins derive from an homogeneous granule compartment in a functionally homogeneous population of cells. To explain our results the hypothesis of a second intracellular route for the secretory proteins in proposed.

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.

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.

Kinetic properties of type-II ATP diphosphohydrolase from the tunica media of the bovine aorta.

The kinetic properties of type-II ATP diphosphohydrolase are described in this work. The enzyme preparation from the inner layer of the bovine aorta, mostly composed of smooth muscle cells, shows an optimum at pH 7.5. It catalyzes the hydrolysis of tri- and diphosphonucleosides and it requires either Ca2+ or Mg2+ for activity. It is insensitive to ouabain (3 mM), an inhibitor of Na+/K(+)-ATPase, to tetramisole (5 mM), an inhibitor of alkaline phosphatase, and to Ap5A (100 microM), an inhibitor of adenylate kinase. In contrast, sodium azide (10 mM), a known inhibitor for ATPDases and mitochondrial ATPase, is an effective inhibitor. Mercuric chloride (10 microM) and 5'-p-fluorosulfonylbenzoyl adenosine are also powerful inhibitors, both with ATP and ADP as substrates. The inhibition patterns are similar for ATP and DP, thereby, supporting the concept of a common catalytic site for these substrates. Apparent Km and Vmax, obtained with ATP as the substrate, were evaluated at 23 +/- 3 microM and 1.09 mumol Pi/min per mg protein, respectively. The kinetic properties of this enzyme and its localization as an ectoenzyme on bovine aorta smooth muscle cells suggest that it may play a major role in regulating the relative concentrations of extracellular nucleotides in blood vessels.

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.

Purification of the blood vessel ATP diphosphohydrolase, identification and localisation by immunological techniques.

ATP diphosphohydrolase (ATPDase) or apyrase (EC 3.6.1.5), an enzyme that hydrolyses the gamma and beta phosphate residues of triphospho- and diphosphonucleosides, has been purified from the bovine aorta media. A particulate fraction was isolated by differential, and sucrose cushion centrifugations, producing a 33-fold enrichment in ADPase activity. Solubilization of the enzyme from the particulate fraction with Triton X-100 caused a partial loss of activity. The solubilized enzyme was purified by DEAE-agarose, Affi-Gel blue and Concanavalin A column chromatographies yielding an additional 138-fold enrichment of the enzyme. The enzyme preparation was further purified by PAGE under non-denaturing conditions, followed by its detection on the gel. The active band was cut out and separated by SDS/PAGE. Overstaining with silver nitrate revealed a single band corresponding to a molecular mass of 78000. Presence of an ATP binding site on the latter protein was demonstrated by labelling with 5'-p-fluorosulfonylbenzoyladenosine (FSBA), an analogue of ATP, followed by its detection by a Western blot technique. Labelling specificity was demonstrated by competition experiments with Ca-ATP and Ca-ADP. An antiserum directed against the N-terminal sequence of the pig pancreas ATPDase (54 kDa) cross-reacted with the bovine aorta ATPDase at 78 kDa. Digestion of the ATPDase with N-glycosidase F caused a marked shift of the molecular mass, thereby showing multiple N-oligosaccharide chains. Immunohistochemical localisation confirmed the presence of ATPDase on both endothelial and smooth muscle cells.

Characterization and immunohistochemical localization of nucleoside triphosphate diphosphohydrolase (NTPDase) in pig adrenal glands (presence of a non-sedimentable isoform).

Considering that adrenal glands possess a variety of purinoceptors associated with various cell types and that some of these cells (chromaffin cells) secrete large amounts of adenine nucleotides, it was of interest to localize nucleoside triphosphate diphosphohydrolase (NTPDase) in these glands and to define the biochemical characteristics of this ectonucleotidase. Immunolocalization produced a moderate reaction in capsula and medulla, with no signal in zona glomerulosa and zona reticularis. In contrast, a very strong reaction was found in zona fasciculata. Biochemical analysis of particulate fractions isolated from whole glands revealed high levels of ATPase and ADPase activities. This appeared to be attributable to the NTPDase since the level of ADPase was as high as ATPase. Both ATPase and ADPase activities were similarly inhibited by sodium azide. Additionally electrophoretograms with these two substrates showed comparable patterns. Western blots with 'Ringo', an antibody that recognizes the different isoforms of mammalian NTPDases, showed the presence of isoforms of NTPDases at 54 and 78 kDa, respectively. Interestingly, the 54 kDa isoform remains in the supernatant of a chromaffin granule lysate after ultracentrifugation. Up until now little interest has been given to the relationship between adrenal medulla and cortex. Presence of purinoceptors and ectonucleotidases in both these regions together with the effects of ATP in vivo and in vitro in different species indicate that purines play a significant role in adrenal glands.

Demonstration of an ectoATP-diphosphohydrolase (E.C.3.6.1.5.) in non-vascular smooth muscles of the bovine trachea.

An ectoATP-diphosphohydrolase (ATPDase) is put in evidence in non-vascular smooth muscles of the bovine trachea. The enzyme has an optimum pH of 7.0 and catalyzes the hydrolysis of the gamma- and beta-phosphate residues from extracellular triphospho- and diphosphonucleosides. It requires either Ca2+ or Mg2+ and is insensitive to ouabain, oligomycin and Ap5A. Sodium azide (20 mM), mercuric chloride (10 microM) and gossypol (35 microM) inhibit the enzyme activity by more than 45%. Polyacrylamide gel electrophoresis under non-denaturing conditions and kinetic properties, namely pH dependency profiles, heat inactivation and 60Co gamma-irradiation-inactivation curves, support the view that the same catalytic site is responsible for the hydrolysis of ATP and ADP to AMP. Accordingly, when both ATP and ADP were combined, reaction rates were not additive. With ATP, Km,app and Vmax,app were estimated at 15 +/- 2 microM and 1.9 +/- 0.1 mumol inorganic phosphate/min per mg of protein, respectively. From 60Co gamma-irradiation-inactivation curves, the molecular mass of the enzyme was estimated at 71 +/- 5 kDa. Enzyme markers indicate that the ATPDase is associated with the plasma membrane. Enzyme assays on trachea smooth muscle cells in suspension confirm that the catalytic site of this ATPDase is localized on the outer surface of the plasma membrane. Analysis of the biochemical properties shows many points of similarity between the tracheal ATPDase and the ATPDase recently described in the bovine lung.

Identification and localization of ATP-diphosphohydrolase (apyrase) in bovine aorta: relevance to vascular tone and platelet aggregation.

In this work, we confirm the existence of an ATP-diphosphohydrolase (apyrase) in bovine aorta and we show that its properties are different from the previously described pancreas ATP-diphosphohydrolase. Hence the aorta enzyme should be considered as a novel type of apyrase. The demonstration is based on pH dependency profiles, heat denaturation curves, 60Co irradiation-inactivation curves and enzyme localization after polyacrylamide gel electrophoresis under non-denaturing conditions. In addition, the irradiation-inactivation curves clearly showed that for both pancreas and aorta enzymes preparations, the same catalytic site is responsible for the hydrolysis of ATP and ADP. The molecular masses of enzymes calculated with this method are 132 +/- 19 kDa (mean +/- S.D.) and 189 +/- 30 kDa (mean +/- S.D.) for the pancreas and aorta enzymes, respectively. Preliminary observations on isolated bovine brain capillaries revealed a high level of enzyme activity strongly suggesting that an ATP-diphosphohydrolase is associated with endothelial cells. The presence of the enzyme on this type of cells was confirmed with pulmonary endothelial cells in culture. Considering the high proportions of smooth muscle cells relative to endothelial cells and the high level of enzyme activity in the aorta preparation, an ATP-diphosphohydrolase activity is definitely present in smooth muscle cells. The ATP-diphosphohydrolase activities described above could regulate the relative concentrations of purine nucleotides both in the plasma and within the vascular wall and hence could play a role both in platelet aggregation and in the control of vascular tone.

Characterization of ATP-diphosphohydrolase activities in the intima and media of the bovine aorta: evidence for a regulatory role in platelet activation in vitro.

The inner layer of the aorta contains the enzyme ATP diphosphohydrolase (ATPDase: EC 3.6.1.5) which catalyzes the sequential phosphorolysis of ATP----ADP----AMP. Two zones of the inner layer, the intima and media, were separated and both were shown to contain ATPDase activity of similar specific activity (0.08 and 0.10 U/mg protein, respectively). However, the media exhibited about 100-times more enzyme activity than the intima. Both preparations were virtually identical with respect to pH optima (7.5), migration patterns after electrophoresis under non-denaturing conditions, relative rates of ATP and ADP hydrolysis and potency to inhibit ADP-induced platelet aggregation in both human platelet-rich plasma and whole blood. The IC50 values for ADP (2 microM)-induced aggregation were 6.8 and 12.9 mU/ml in platelet-rich plasma and whole blood, respectively. Addition of ATPDase to platelets pre-aggregated with ADP resulted in a dose-dependent disaggregation in platelet-rich plasma (IC50 4.9 mU/ml), but not in whole blood. When both ATPDase (5.6-58.7 mU/ml) and ATP (0.5-10 microM) were added to platelet-rich plasma, there was an immediate dose-dependent aggregation of platelets followed by a slowly developing disaggregation. These data show that ATPDase is present in both the intima and media layers of bovine aorta and suggest a dual role for this enzyme in platelet activation. By converting ATP released from damaged cells into ADP, the enzyme could facilitate platelet aggregation at the site of vascular injury, whereas the subsequent conversion of ADP to AMP could inhibit or reverse platelet aggregation. The consequence of these activities would be to control the growth of a platelet thrombus.

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.

Cytochemical and immunocytochemical characterization of a fibrillar network (GP2) in pancreatic juice: possible role as a sieve in the pancreatic ductal system.

The secretory product of the exocrine pancreas contains sedimentable and non-sedimentable materials. Electron microscopy of the pellet obtained after ultracentrifugation reveals two major components: microvesicles (pancreasomes) and a fibrillar network of small mesh size. Negative staining of an unfixed pellet demonstrated that these structures are not fixation artifacts. Cytochemical analysis showed that pancreasomes are reactive to osmication and uranyl acetate staining, whereas the fibrillar network was unreactive thereby indicating that the latter does not contain lipids; however, lead citrate staining reveals the network. Alcian blue, known to bind sulfate groups of mucosubstances, reacted strongly with the fibrillar network. The pellet was also characterized by immunocytochemistry with specific antibodies to amylase and glycoprotein 2 (GP2). Both antibodies were located only on the fibrillar network. Washing of the pellet with 100 mM KCl-250 mM NaBr had little effect on GP2 content, but reduced considerably alpha-amylase associated with the reticular matrix. It appeared that GP2 was the major component of the scaffolding that gives rise to the fibrillar network and that other proteins such as alpha-amylase could reversibly bind to it. When double-labeling immunocytochemistry was carried out on the unwashed pellet, labeling of the first antigen reduced the labeling of the second. Removal of amylase by washing the pellet increased the GP2 signal. These results indicate that amylase is bound on the GP2 network. Although the function of the GP2 network is still not clearly defined several possibilities could be envisaged at the level of the pancreatic duct system: 1) The network could drain off any aggregates or precipitates forming in small ducts. 2) The small mesh of the network would present a physical barrier to infecting bacteria that could enter into the duct system from the intestine, especially in conditions of low flow rates. 3) The network may exert a mechanical pressure on the membranes bordering the acinar lumen and small ducts thereby preventing their collapse in basal conditions.

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.

Isolation of zymogen granules from rat pancreas and characterization of their membrane proteins.

A zymogen granule fraction has been isolated from rat pancreas, and its purity has been assessed by biochemical and morphological criteria. Specific activities of two marker enzymes, amylase and chymotrypsin, are increased by 4.6 and 5.4-fold, respectively, as compared to the homogenate. The purified fraction is devoid of detectable RNA, DNA and 5'-nucleotidase, glucose-6-phosphatase, and cytochrome c oxidase activities. Electron micrographs confirm the absence of mitochondria, lysosomes, and rough endoplasmic reticulum fragments. Zymogen granule membranes were isolated from this fraction on a sucrose gradient following lysis in alkaline buffer. Secretory contaminants were efficiently removed from the membranes as indicated by experiments in which labeled secretory proteins were added during the isolation procedure and secondly by measuring residual levels of amylase and chymotrypsin. Three enzyme activities were found in the membranes: thiamine pyrophosphatase, ATP-diphosphohydrolase, and low levels of acid phosphatase. Membrane proteins were solubilized by urea-Triton X-100 and separated in double-dimension (isoelectric focusing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis). Isoelectric point and molecular weight of each protein band were determined.

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)

The origin of the zymogen granule membrane of the pancreatic acinar cell as examined by ultrastructural cytochemistry of acid phosphatase, thiamine pyrophosphatase, and ATP-diphosphohydrolase activities.

Cytochemical distributions of acid phosphatase, thiamine pyrophosphatase, and ATP-diphosphohydrolase activities have been examined on thin sections of rat pancreas and on isolated zymogen-granule membranes. Acid phosphatase was found in the rigid lamellae separated from the Golgi stacked cisternae, in condensing vacuoles, and in the trans-saccules of Golgi apparatus; it was not detected in purified zymogen-granule membranes. Thiamine pyrophosphatase was detected in trans-saccules of the Golgi apparatus, in purified zymogen-granule membranes, and in the plasmalemma of the acinar cell. It was absent in condensing vacuoles. The ATP-diphosphohydrolase activity has a distribution similar to thiamine pyrophosphatase. These observations illustrate the similarity between the trans-saccules of the Golgi apparatus and the membrane of mature zymogen granules and the disparity between the latter membrane and the membrane of the condensing vacuole. They suggest that the condensing vacuole might not be the immediate precursor of the zymogen granule as commonly assumed. An alternative possibility would be that condensing vacuoles would fuse with the trans-saccule (transition) of the Golgi apparatus which in turn would form mature zymogen granules.

Marked differences in immunocytological localization of [3H]estradiol-binding protein in rat pancreatic acinar tumor cells compared to normal acinar cells.

[3H]Estradiol can bind to a specific protein in normal rat pancreatic acinar cells. Electron microscopic immunocytochemical analysis has shown this protein to be localized primarily in the rough endoplasmic reticulum and mitochondria. Rat exocrine pancreatic tumor cell lines, whether grown in tissue culture (AR42J) or as a tumor mass after sc injection into rats (DSL-2), lacked detectable amounts of this [3H]estradiol-binding protein (EBP), as determined by the dextran-coated charcoal assay. Furthermore, primary exocrine pancreatic neoplasms induced with the carcinogen azaserine contained little or no detectable [3H]estradiol-binding activity. However, electron immunocytochemical studies of transformed cells indicated the presence of material that cross-reacted with antibodies prepared against the [3H]EBP. The immunopositive reaction in transformed cells was localized almost exclusively in lipid granules. Such lipid organelles in normal acinar cells, although present less frequently than in transformed cells, have never been observed to contain EBP-like immunopositive material. Presumably, the aberrant localization of EBP in these acinar tumor cells results in loss of function of this protein, which in normal pancreatic acinar cells appears to exert a modulating influence on zymogen granule formation and the process of secretion.