Stored Secrets Research Articles

Genetic differences in the histochemically defined structure of oligosaccharides in mice.

A wide range of tissues from three interfertile species of mice and an interspecific hybrid was examined with lectins conjugated to peroxidase to localize specifically glycoconjugates containing terminal alpha-N-acetylgalactosamine, alpha-galactose, and alpha-fucose, and the terminal disaccharide galactose-(beta 1----3)-N-acetylgalactosamine. This battery of lectins disclosed marked heterogeneity of glycoconjugates in different histological sites in a given animal and even between cells in a presumably homogeneous cell population within an organ. No variation with any lectin was observed between individuals of two closely related inbred strains of Mus domesticus at any specific histological or cytological site. In contrast, littermates of an outbred strain of Mus castaneus differed in binding of certain lectins at various sites, attesting to a genetic basis for individual variation. Hybrids between castaneus and domesticus mice also showed individual variation. Moreover, extensive differences between the mouse species were demonstrable with every lectin in glycoconjugates of stored secretions, Golgi cisternae, and apical or basolateral plasmalemma in many cell types. Totaling the differences in tabulated staining intensities for each possible species pair gave a measure of the overall extent of difference at 53 histological sites. According to this measure, the three species are about equally divergent from one another. Some differences between species appeared to depend on histological rather than histochemical variation, as, for example, a greater abundance of granular duct cells in the sublingual and submandibular glands in Mus hortulanus. Other differences were apparently derived from pathological change, as exemplified by casts and lymphoid infiltrates in kidney and structurally atypical submandibular gland lobules in Mus castaneus, and possibly by infiltrating cells in intestinal lamina propria and epithelium in Mus castaneus and hortulanus.

Synthesis of seminal ribonuclease in isolated lobules of bull seminal vesicles.

A procedure is described for preparing and maintaining in culture isolated lobules of bovine seminal vesicles, consisting of glandular acini, surrounded by little connective tissue and with free access to the external medium, in which secreted material can be collected. After 48 h in culture, the isolated lobules appeared indistinguishable, by morphological and biochemical criteria, from freshly isolated lobules. After much longer culture times about one third of the glandular cells were still capable of effective protein synthesis. Studying the biosynthesis of seminal ribonuclease with preparations of isolated lobules we found that the enzyme was synthesized and secreted; only the fully amidated isoenzyme was synthesized and secreted, indicating that production of the selectively deamidated isoenzymic forms occurred after secretion, newly synthesized protein was rapidly exported, indicating that the high levels of enzyme previously reported for the seminal vesicle tissue were essentially due to its content of stored secretion.

Effects of rat growth hormone (rGH)-releasing factor and somatostatin on the release and synthesis of rGH in dispersed pituitary cells.

The effects of rat hypothalamic GH-releasing factor (GRF) and somatostatin (SRIF) on the release and biosynthesis of rat GH were studied by RIA and quantitative immunoprecipitation using monolayer cultures of rat anterior pituitary cells. In kinetic studies, GRF stimulation of GH release appeared at the first sampling time (20-min incubation) and the effect began to diminish after 2-h incubation with GRF. On the other hand, total (cell plus medium) content of GH significantly increased only after 24-h incubation. To examine the GH-synthesizing effect of GRF more directly, newly synthesized GH labeled by [35S]methionine during incubation with GRF was quantified by immunoprecipitation. The amount of immunoprecipitable GH increased significantly and specifically (compared with the total amount of labeled proteins) also only after 24-h incubation. When GH pools were labeled with [35S]methionine under different schedules, the basal release of newly synthesized GH, which was labeled for 1 h immediately before chase incubation was lower during the first 15 min than stored GH which had been labeled earlier. Basal newly synthesized GH secretion exceeded stored GH secretion after 30 min. GRF stimulated the release of GH from both pools but the stimulation of stored GH was greater. In this system, SRIF suppressed both the basal and stimulated release of GH but did not modify GH biosynthesis under either condition. Newly synthesized GH showed significant degradation during 24-h incubation; neither GRF nor SRIF affected the rate of GH degradation during the same incubation period. These results indicate that 1) GRF stimulates both release and synthesis of GH; 2) these two effects have different kinetics and different sensitivities to SRIF; and 3) GRF stimulates the release of GH from heterogeneous pools disproportionally.

Fate of stored secretion following ligation of the seminal vesicle-coagulating gland in the rat.

Unilateral ligation of the seminal vesicle-coagulating gland complex in castrated and testes intact sexually mature rats resulted in a significant asymmetry in weight after 14 days, with ligated glands heavier in both groups. Differences in weight between glands resulted from ligation, and the primary route by which stored secretion is eliminated from these glands is by spontaneous ejaculation rather than resorption via the glandular epithelia.

GLYCOPROTEINS OF HUMAN JEJUNAL CELLS: LOCATION, MATURAT10NAL CHANGES AND INDIVIDUAL VARIATION

A battery of 12 lectins conjugated to horseradish peroxidase was used to characterize secretory and cell surface glycoproteins (GP) in secretions from small bowel biopsies of 9 children. The glycoproteins found differed from site to site, among individuals at a given site and from crypt to villar tip. In goblet cells stored secretions showed individual variation in intensity of staining and increasing fucose content from crypt to villus. Terminal N-acetylglucosamine was seen in the Golgi cisternae but not in stored secretion, suggesting capping of this sugar by other residues during GP biosynthesis. Terminal α-N-acetylgalactosamine was present in Golgi cisternae of all and in the stored secretion of two-thirds of the specimens. Immature cryptal columnar cells had fucose-rich GP presumably in secretory granules. Mature villar cells did not. Columnar cells showed mannose-rich GP, presumably lysosomal enzymes, in apical punctate bodies. In some specimens the basolateral plasmalemma of columnar cells stained selectively for fucose and β-galactose. Brush border staining generally paralleled that of goblet cell secretions with similar maturational changes. Staining for terminal galactose highlighted cells, presumably leukocytes, infiltrating the epithelium and lamina propria. The diversity and variability of the GP in human jejunal mucosa suggest a relationship between specific chemical structures and biologic function at the cytologic loci.

Elimination of stored material from the seminal vesicles of the guinea pig following castration.

Experiments were carried out to determine what factors are involved in the disappearance of stored secretion from the seminal vesicles of the guinea pig and in the concurrent decline in weight of these glands following castration. Seminal vesicles of sexually mature males were unilaterally ligated, and the males were either castrated or left testes intact for a 14-day period. At necropsy, the seminal vesicles were removed and weighed, and biopsies were collected for histological inspection. The weight of nonligated vesicles from castrated males declined to less than half that of ligated sides. In testes-intact males, the weight of ligated vesicles was nearly double that of nonligated vesicles. Epithelial cell heights remained unchanged during the study. The rapid decline in weight of the guinea pig seminal vesicles following castration was attributed to the elimination of stored secretion by way of the urethra rather than by absorption.

Histochemical reactivity of peanut lectin-horseradish peroxidase conjugate.

A peanut lectin-horseradish peroxidase (PL-HRP) conjugate has been applied to histochemical staining of paraffin sections of various mouse organs. The PL-HRP conjugate has selectively reacted with secretory bodies, the Golgi zone, and the apical cell surface in various cell types. Some positive sites, including lingual and tracheal serous glands, Brunner's glands, and the brush border of the proximal straight nephron, contained periodic acid-Schiff (PAS)-positive glycoconjugate with no affinity for basic reagents. The stored secretion in these sites was interpreted as containing neutral glycoprotein with terminal galactose residues which could, in part at least, account for the PAS reactivity. Duodenal goblet cells, which exhibited basophilia attributable to sulfate esters, also bound PL-HRP. As the binding was affected by prior sialidase digestion, the secretory glycoprotein in the duodenal goblet cells was judged to contain oligosaccharides with sulfate esters and terminal galactose uncapped by sialic acid. All sites known from their basophilia to form sialomucin failed to stain with the PL-HRP conjugate, but consistently gained reactivity following sialidase digestion and were inferred, therefore, to possess glycoproteins with oligosaccharide side chains containing subterminal galactose and terminal sialic acid. Lingual mucous glands, known to secrete a mucosubstance with basophilic properties indicative of the presence of sulfate esters but not sialic acid, stained with PL-HRP only after sialidase digestion and, accordingly, were reinterpreted as containing both sulfate esters and terminal galactose-sialic acid dimers. Staining of gastric surface epithelium demonstrated a srongly PAS-reactive neutral glycoprotein, and that of goblet cells in the cecum disclosed PAS-positive sulfated glycoprotein. The latter two sites lacked PL-HRP affinity without or with prior sialidase treatment and apparently possessed neither terminal galactose residues nor galactose-sialic acid dimers. PL-HRP affinity was observed exclusively in the Golgi cisternae of some epithelial cells, thus indicating that galactose occurs transiently as a terminal residue in this site. A few histologic sites, such as pancreatic and gastric zymogen cells and renal tubules, were devoid of both PAS reactivity and basophilia indicative of the presence of complex carbohydrate but stained strongly with the PL-HRP conjugate by means which are not understood. Galactose in the PL-HRP solution blocked or reversed the PL-HRP binding in most of the structures with an affinity for the conjugate, supporting the conclusions that the reagent is specific for galactosyl residues.

Serum acid phosphatase levels following prostatic massage (author's transl)

We find an elevation of serum acid phosphatase levels 5 min after prostatic massage in only 10% of patients with prostatic adenoma. This increase is caused by stored prostatic secretion being pressed into the blood vessels. 60 min later these serum levels decrease. With some other patients a slower increase of phosphatase levels occurs; this increase, however, lasts for hours. This kind of increase is caused by prostatic fluid being forced into the interstitium where it is slowly absorbed. A combination of both kinds leads to a curve with two peaks; this could be demonstrated in two cases. No difference was seen in reaction of total acid phosphatase and prostatic phosphatase levels. The increase of phosphatase levels following prostatic massage was no sign of prostatic carcinoma.

A randomly generated program for automatic identity checking

A method for making secret programs for automatic identity checking with very high demands for security, is described. The consistency of the sequence of digits, which identifies the person, is tested by a sequence of transformations, chosen by means of a stored secret random table. The method has been implemented for a system of unattended cash dispenser terminals, equipped with microprocessors.

Caerulein secretion by dermal glands in Xenopus laevis.

Since gastrin and its related peptides are secreted by a minority population of widely dispersed cells in mamamalian tissues it has, in the past, been difficult to study the subcellular aspects of their secretion. From published reports (1, 2) it seemed possible that a satisfactory system for such studies might be provided by the skin of certain amphibians such as Xenopus laevis since in these tissues high concentrations of peptides such as caerulein exist, and there is some indication (3) that this, or a similar gastrin-like peptide, may be a dermal gland secretory product. We have therefore explored this possibility by studying the structure, secretory process, and secretory product of the most prominent non mucous type of gland in the skin of X. laevis. These studies clearly demonstrate that most, if not all, of the caerulein in which the skin is contained in secretion granules within the dermal glands and that its release can be specifically evoked by adrenergic stimulation. The release process by a holocrine mechanism expels all of the stored secretion onto the skin surface and thus for biosynthetic studies it should now be possible to synchronize the processes which lead to the replenishment of the peptide.

Function of coated membranes and multivesicular bodies during membrane regulation in stimulated exocrine pancreas cells.

Stimulation of secretion by pilocarpine results in a 70% loss of zymogen granules from pancreatic acinar cell during the first hr after injection of the drug. In previous work (Geuze and Poort, 1973), we found that the amount of membrane stored in the surface of the microvilli and of the numerous infoldings present in highly stimulated cells, increases during the first 2 hr and then decreases again during the 3rd hr after stimulation, concurrently with maximal endocytosis of sorbitol-[su14C]. Further observations on the fine structure of stimulated cells at various time intervals after injection of pilocarpine showed that during the first hr numerous smooth vesicles and multivesicular bodies (mvb's) appear in the apical cytoplasm, while the number of coated vesicles and their relative total volume increase significantly 3 hr after stimulation. By infusion of ferritin in the pancreatic duct system in vivo and application of cytochemical techniques (osmium impregnation, electron microscope autoradiography and acid phosphatase cytochemistry) it could be established that after stimulated exocytotic secretion, redundant apical cell membrane is withdrawn by at least two routes: 1) During the initial rapid increase of the amount of apical cell membrane, withdrawal is accomplished by interiorization of luminal invaginations into smooth endocytotic vesicles, which in turn give rise to mvb's by infolding and subsequent fission of their limiting membrane. 2) Once the bulk of stored secretion granules has been discharged, endocytotic coated vesicles become gradually more prominent as carriers for redundant cell membrane. The contents of endocytotic structures ultimately become incorporated in residual bodies, suggesting lysosomal degradation of cell membrane prior to eventual reutilization. Coated vesicles also originate by pinching off from mature Golgi cisternae and condensing vacuoles. A possible function of the coated membranes in the concentration of exportable protein within forming secretory granules is discussed.