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Density gradient analysis of secretions produced in vitro by human and canine airway mucosa: identification of lipids and proteoglycans in such secretions.

Human and canine airway mucosal explants synthesize and secrete high molecular weight glycoconjugates, incorporating 14C-glucosamine, a radioactive precursor to epithelial glycoprotein. Our examination of secretions produced by several individual specimens, however, did not reveal epithelial glycoprotein of typical buoyant density (1.5 g/ml in CsBr); only a high-density component with features of glycoprotein and proteoglycan. To provide sufficient material for characterization, secretions from several specimens of human and canine explants were separately pooled and subjected to DGU in CsBr. After removal of lipids and proteins, the glycoconjugates were recovered into five fractions of different density. 14C-glucosamine had been incorporated in all five fractions. Fractions 1-4 together accounted for 88% of the radiolabel but gas chromatography indicated that none of these contained epithelial glycoprotein. Their amino acid compositions were similar to those of proteoglycans and electrophoresis confirmed the presence of chondroitin sulfates A, B, C, heparan sulfate and hyaluronic acid. Sugars typical of epithelial glycoprotein were identified only in the glycoconjugate subfraction 5 of lowest density (and also lowest in yield) in which glycosaminoglycans were also identified. By addition of radioactive precursors, 14C acetate, 14C palmitate and 14C mevalonic acid to the culture medium and autoradiography of the secreted lipids we have shown that the tracheal explants actively synthesize lipids. Lipids accounted for a high proportion, almost half by weight, of the explant secretion. While neutral and phospholipids predominate, glycolipids were also identified.

Amino Acids↗

[Cryptal origin of the intestinal secretion induced in rats by prostaglandin El. Evolution of secretion after a single injection of 5-fluorouracil].

The intestinal secretions induced by an intrajejunal perfusion of prostaglandin E1 (PGE1) at the rate of 24 gamma/min in anaesthetized male Wistar rats, weighing 200-200 g, were studied during 6 days after a single injection of an antimitotic agent, the 5-fluorouracil (5-FU, 40 mg/kg). 5-FU induced changes in the structure of the intestinal mucosa. The modifications observed in the intestinal secretions of water and sodium induced by PGE1 suggest that these secretions might (a) originate from crypts; (b) be partly reabsorbed by villi. Concerning the endogenous secretions, the intestinal mucosa might act as a secretion-reabsorption system.

Alprostadil↗

CD11b+ Peyer's patch dendritic cells secrete IL-6 and induce IgA secretion from naive B cells.

Peyer's patch (PP) dendritic cells (DCs) have been shown to exhibit a distinct capacity to induce cytokine secretion from CD4(+) T cells compared with DCs in other lymphoid organs such as the spleen (SP). In this study, we investigated whether PP DCs are functionally different from DCs in the SP in their ability to induce Ab production from B cells. Compared with SP DCs, freshly isolated PP DCs induced higher levels of IgA secretion from naive B cells in DC-T cell-B cell coculture system in vitro. The IgA production induced by PP DCs was attenuated by neutralization of IL-6. In addition, the induction of IgA secretion by SP DCs, but not PP DCs, was further enhanced by the addition of exogenous IL-6. Finally, we demonstrated that only PP CD11b(+) DC subset secreted higher levels of IL-6 compared with other DC subsets in the PP and all SP DC populations, and that PP CD11b(+) DC induced naive B cells to produce higher levels of IgA compared with SP CD11b(+) DC. These results suggest a unique role of PP CD11b(+) DCs in enhancing IgA production from B cells via secretion of IL-6.

Animals↗

FSH secretion patterns during pregnant and nonpregnant luteal periods and 24 h secretion patterns in male and female dogs.

The aim of this study was to investigate the secretion patterns of FSH: (i) during the canine oestrous cycle with particular attention to the luteal period; and (ii) during 24 h in male and female dogs. Plasma FSH concentrations were measured by a highly specific homologous canine immunoradiometrical (IRMA) assay using monoclonal antibodies specific for canine FSH. In the first study, FSH concentrations were measured daily from +/- 15 days before until 150 days after the LH surge in groups of pregnant (n = 5) and nonpregnant (n = 5) Beagle bitches. Day 0 of the cycle was the day of the preovulatory LH peak as measured by LH radioimmunoassay. In the second study, FSH concentrations were measured at 1 h intervals for 24 h in five bitches at day 120 after the LH surge and in five males. From day 35 to day 40 after the LH peak, FSH concentrations were higher in pregnant than in nonpregnant luteal phases. A significant and abrupt decrease in FSH concentrations was observed at about the time of parturition (day 65) and was followed by lower FSH concentrations during lactation compared with nonpregnant bitches. FSH concentrations in nonpregnant animals were fairly constant from the end of oestrus to mid-anoestrus. In both post-lactation and anoestrous bitches, plasma concentrations of FSH increased consistently as anoestrus progressed. No significant differences in the mean hour-to-hour patterns of FSH secretion were observed over 24 h and no differences were detected between male and female dogs. However, a clear pulsatile pattern of secretion was observed in all individuals, both males and females, with an apparent 4.8 h interval between peaks. The results of this study demonstrate major differences in FSH secretion between pregnant and nonpregnant bitches. This finding indicates that there are differences in the regulation of the hypothalamo-pituitary-ovarian axis during pregnancy, possibly related to changes in activity of the corpus luteum. The results of this study also demonstrate a pulsatile pattern of FSH secretion in both male and female bitches, with a 4.8 h interval between peaks, and confirms the progressive increase in plasma FSH concentrations observed during anoestrus.

Animals↗

ApoA-I secretion from HepG2 cells: evidence for the secretion of both lipid-poor apoA-I and intracellularly assembled nascent HDL.

The goal of this study was to determine whether apolipoprotein A-I (apoA-I) is lipidated before secretion by HepG2 cells. ApoA-I was extracted from microsomes after radiolabeling with [(35)S]Met/Cys. After ultracentrifugal flotation, d < 1.25 g/ml and d > 1.25 g/ml fractions were immunoprecipitated and analyzed by SDS-PAGE. Under steady state radiolabeling conditions, 20% of extracted microsomal apoA-I floated at d < 1.25 g/ml. Pulse-chase experiments demonstrated that the percentage of microsomal apoA-I associated with lipid peaked between 2 and 8 min postsynthesis. Density gradient ultracentrifugation, and nondenaturing gradient gel electrophoresis of HepG2 cell medium, indicated that 50% of secretory apoA-I existed as small HDL particles with a diameter of approximately 7.5 nm. These and additional data suggested that approximately 20% of newly secreted apoA-I is lipidated intracellularly and another 30% is secreted in lipid-free or lipid-poor form but acquires sufficient lipid to become small HDL within 1 h of secretion, with little further maturation over the time course of the incubation (2 h).These results indicate that a process exists for the presecretory intracellular assembly of apoA-I with lipid in HepG2 cells and that apoA-I is secreted in both lipid-poor and lipidated forms.

Apolipoprotein A-I↗

[Functional distinction of secreting pools of the pancreas and participation of the pancreatic ductal system in the development of the pancreatic secret properties].

Acute experiments on dogs with separate secretion of the pancreatic enzymes, either stimulated or inhibited, from two lobes of pancreas, and investigation into the character of kinetics of the secretion from 5-7 ductal pools of the pancreas showed the functional specifics, with the device of ductal valves and microdepot of a secret of the pancreas ductal system playing an important role. The conclusion is made that the final secret of pancreas removed to the duodenum is a product of a secret-motor activation of non-equipotentional microregions of the gland, being the components of an intervisceral dynamical mosaic.

Animals↗

Prepubertal rat thymus secretes a factor that modulates gonadotropin secretion in cultured rat pituitary cells.

Thymus development and function are influenced by endocrine glands. On the other hand, the absence of thymus results in endocrine disturbances. The aim of the present paper is to investigate the direct influence of thymus products on the gonadotropin secretion of rat pituitary cells in culture. In this study, a chromatographic fraction obtained from the acetonic powder of thymus from 14-15 day-old rats (TF) increased the GnRH stimulated LH and FSH secretion from cultured pituitary cells, but had no effect on the basal secretion of these gonadotropins. The addition of incubation medium from thymus (TIM) to cultured pituitary cells further enhanced GnRH stimulated FSH and LH production, the effect was dose related. The GnRH potentiating effect of thymus incubation medium was also present in a chromatographic fraction of around 28 Kd obtained from this medium. These data suggest that the prepubertal thymus contains and secretes a factor that modulates pituitary cells secretion potentiating the Gn-RH effect.

Age Factors↗

Effect of estradiol treatment on the secretion and structure of gastric H+ and pepsin-secreting cells in the rat.

Gastrin-stimulated secretory rate of gastric H+ and pepsin were markedly inhibited (P less than 0.001) in ovariectomized rat after 7 days of estradiol treatment (80 micrograms/kg/day) in comparison to ovariectomized untreated rat and sham control. In correlation to the secretions, the majority of parietal cells failed to show the typical ultrastructural transformation associated with active secretion. Pyknotic nuclei with dilatation of nuclear envelope and degenerative changes were observed in the parietal cells. Meanwhile, the chief cell showed inactive characteristics in that a large number of pepsinogen granules were retained in the cells. In contrast, in ovariectomized untreated rat, the rate of gastrin-stimulated H+ and pepsin secretion were not significantly different from the sham (P greater than 0.05). Consistent with secretion, after gastrin stimulation, the morphology and ultrastructure of the parietal cell and chief cell showed active secretory features in a pattern similar to changes observed in sham. The results suggest that impairment in the structure of the functional parts of secretory cells by estrogen treatment was a factor contributing to the inhibition of the gastric secretions.

Animals↗

[A global study of gastric secretion in duodenal ulcers: gastrin, pepsinogen and acid secretion].

AIM: To examine gastric secretion in duodenal ulcer patients; simultaneous assessment of serum gastrin and pepsinogen I levels and acid secretion. METHODS: 32 patients with duodenal ulcer disease were studied (mean age: 44.7 +/- 14 years, 24 males (75%). At endoscopy, biopsy specimens from duodenal bulb, antrum, body and gastric fundus were taken. Basal levels of acid secretion, pepsinogen I and gastrin were measured, as well as stimulated levels (with pentagastrin and beef meal). RESULTS: Mean (+/- SD) BAO and MAO was 5.5 +/- 4 and 27.7 +/- 10 mEq/h, respectively. BAO in males were higher than in females (p < 0.05). A positive correlation (p < 0.01) was observed between MAO, and basal and stimulated pepsinogen I. Basal and stimulated (integrated-90 min) gastrin levels were 43.4 +/- 12 pg/ml and 5,260 pg/ml min, respectively. No correlation between such levels and acid secretion or pepsinogen I was observed. Mean basal and stimulated (integrated-120 min) pepsinogen levels were 109.3 +/- 35 ng/ml and 4,950 +/- 160 ng/ml min, respectively, and they were higher in males (p < 0.01), and in smokers (p < 0.05). CONCLUSION: It was confirmed, in a group of duodenal ulcer patients, that BAO is higher in males than in females. There is a positive correlation between MAO and pepsinogen I levels, although this correlation is not present between gastrin levels and acid secretion or pepsinogen I. Pepsinogen I levels are higher in males and smokers.

Adult↗

Secretion of matrix metalloproteinases and their inhibitors (tissue inhibitor of metalloproteinases) by human prostate in explant cultures: reduced tissue inhibitor of metalloproteinase secretion by malignant tissues.

Unregulated secretion of matrix metalloproteinases (MMPs) or their endogenous protein inhibitors (tissue inhibitor of metalloproteinases, TIMPs) has been implicated in tumor invasion and metastasis. Species of MMPs and TIMPs secreted by epithelial cultures of normal, benign, and malignant prostate were identified and their levels were compared. Fragments of fresh tissue were cultured in a serum-free medium that supported the outgrowth of prostatic epithelial cells. Biochemical analysis of the conditioned media by gelatin zymography and enzyme assays showed that both normal and neoplastic tissues secreted latent and active forms of both M(r) 72,000 type IV collagenase (MMP-2) and M(r) 92,000 gelatinase (MMP-9). However, conditioned media from malignant prostate explants contained a higher proportion of the active form of MMP-2. Significant amounts of free TIMPs were secreted by normal juvenile and adult prostates, but they were either markedly reduced or not detectable in conditioned media from neoplastic tissues. These findings suggest that there is an imbalance of secretion between MMPs and TIMPs in prostatic carcinoma.

Adult↗

[Saliva secretion, concentration and secretion of human salivary amylase in the 1st year of life].

This work show the levels of the salivary secretion volume in 5 min., the salivary amylase concentration in U/ml and the salivary amylase secretion in U/kg/5 min. in 10 children at 4, 15, 30, 45, 60, 90, 120, 150, 180 days of life and at 9 and 12 months. All the children had the same nourishment orientation and showed a good weight and neuromotor development during the period of the observation. The datas were compared with datas of 18 children with 20-36 months of age and 18 healthy young man. It was standardized the technic and the time-table to collect the salivary secretion. It was observed that the salivary secretion is higher between 90 and 180 days of age preceding theeth eruption. At 12 months of life the levels of the amylase salivary concentration reach the levels of infants and adults. The salivary amylase secretion is high after the 60 days of life. Probably this is responsible by the good starch tolerance at this time.

Adult↗

Vasoactive intestinal peptide-associated inhibition of stimulated gastric secretion. II. Inhibition of pentagastrin-stimulated gastric secretion.

The possible role of vasoactive intestinal peptide as an inhibitor of stimulated gastric secretion was investigated. Awake dogs with Heidenhain pouches were stimulated with pentagastrin alone and in combination with intraduodenal instillation of various potential inhibitors of gastrointestinal secretion including isotonic phenylalanine, 3 percent hypertonic saline solution and 0.16 normal hydrochloric acid. Plasma vasoactive intestinal peptide is significantly increased by pentagastrin, and this release is enhanced by intraduodenal hydrochloric acid. Simultaneously, there is significant inhibition of pentagastrin-stimulated Heidenhain pouch secretion. These findings provide evidence for a possible physiologic role of vasoactive intestinal peptide as an inhibitor of gastric secretion, at least in dogs.

Animals↗

Effect of type-selective inhibitors on cyclic nucleotide phosphodiesterase activity and insulin secretion in the clonal insulin secreting cell line BRIN-BD11.

1. The cyclic nucleotide phosphodiesterases (PDEs) present in an insulin secreting cell line, BRIN - BD11, were characterized using calcium/calmodulin, IGF-1, isoenzyme-selective PDE inhibitors and RT - PCR. 2. Calmodulin activated cyclic AMP or cyclic GMP PDE activity in pellet and was 3 fold (P=0.002) more potent in activating cyclic nucleotide hydrolysis in pellet compared with supernatant fractions. 3. The PDE1/PDE5 inhibitor zaprinast inhibited both cyclic AMP and cyclic GMP PDE activity in both pellet and supernatant fractions of cell homogenates by a maximum of around 25% (IC(50) 1 - 5 microM), while rolipram (PDE4 selective) inhibited only cyclic AMP hydrolysis. 4. The PDE3-selective inhibitors Org 9935 (0.02 - 10 microM) and siguazodan (0.1 - 10 microM) inhibited cyclic AMP PDE activity in the pellet but not the supernatant fractions of cell homogenates, with a maximum inhibition of about 30%. IGF-1 (2 - 7.5 ng ml(-1)) potently augmented this PDE activity. 5. RT - PCR using specific primers for PDE3B, but not for PDE3A, amplified, from BRIN - BD11 cell total RNA, a 351 base pair product that was >97% homologous with rat adipose tissue PDE3B. 6. IBMX, Org 9935, siguazodan and rolipram (1 - 50 microM), but not zaprinast, each augmented glucose-induced insulin secretion in the presence of 16.7 mM but not 1 mM glucose. 7. These findings, in a clonal insulin secreting cell line, are consistent with an important role for PDE3B in regulating the pool of cyclic AMP relevant to the modulation of glucose-induced insulin secretion.

1-Methyl-3-isobutylxanthine↗

Influence of residual insulin secretion and duration of diabetes mellitus on the control of luteinizing hormone secretion in women.

BACKGROUND: The aim of the present study was to establish whether the persistence of residual beta-cell activity after long-term diabetes mellitus (DM) exerts a protective role on luteinizing hormone (LH) secretion. METHODS: The LH responses to stimulation with gonadotropin-releasing hormone (Gn-RH) (100 microg in an i.v. bolus) or naloxone (4 mg injected in an i.v. bolus, followed by the constant infusion of 8 mg in 2 h) were measured in C-peptide-positive (CpP) and C-peptide-negative (CpN) normally menstruating women with short-term (group 1 < 3 years, CpP n = 11, CpN n = 11) or long-term (group 2 > 10 years, CpP n = 11, CpN n = 11) DM and in age-matched normal control subjects (n = 11). RESULTS: Gn-RH induced significant increments in LH secretion in all groups. Significant LH responses to naloxone were observed in all groups, except in group 2 CpN patients. However, the LH response to either Gn-RH or naloxone was significantly lower in group 1 CpN, group 2 CpP and group 2 CpN patients than in the normal control subjects. Furthermore, the LH response was significantly lower in group 2 CpP than in group 1 CpP patients and in group 2 CpN than in group 1 CpN subjects. CONCLUSIONS: These results indicate a role for both deficiency in residual endogenous insulin secretion and duration of diabetes in the derangement of LH secretory control. The data suggest that the protective role exerted by residual beta-cell activity on LH secretion during the early years of DM diminishes with time elapsed after the onset of diabetes mellitus.

Adult↗

The effect of artificial sweetener on insulin secretion. 1. The effect of acesulfame K on insulin secretion in the rat (studies in vivo).

Acesulfame K is an artificial sweetener which has been used in the food industry for some years. As yet no metabolic effects have been reported. It was reported that the sweetener can induce a cephalic phase of insulin secretion. To analyse the mechanism of this phenomenon, we studied the effect of Acesulfame K on insulin secretion in vivo. Male Wistar rats, weighing 250-300 g were fasted overnight and anaesthetized with phenobarbital. A silicon catheter was inserted into the right cervical vein for injection of test substances and for obtaining blood samples. In some experiments, another catheter was inserted into the left cervical vein for continuous infusion. Blood samples were drawn at 0, 5, 10, 15, 30 and 60 min after injection, and at -10, 0, 10, 20, 30, 40, 60, 80, 100 and 120 min after the infusion started. Injection of Acesulfame K (150 mg/kg body weight) increased the plasma insulin concentration at 5 min from 27.3 +/- 3.0 microU/ml to 58.6 +/- 4.2 microU/ml without any significant change in the blood glucose. Infusion of Acesulfame K (20 mg/kg body weight/min) for one hour maintained the insulin concentration at a high level (about 85-100 microU/ml) during this period, and at the same time blood glucose was gradually reduced from 103.0 +/- 7.3 to 72.0 +/- 7.2 mg/dl. When using different amounts of Acesulfame K, the insulin secretion was stimulated in a dose-dependent fashion. The effect of Acesulfame K on insulin secretion was similar to that observed by injecting or infusing the same doses of glucose (150 mg/kg) body weight for injection and 20 mg/kg body weight/min for infusion), except that no hyperglycemia was observed with Acesulfame K.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Regulatory role of CD38 (ADP-ribosyl cyclase/cyclic ADP-ribose hydrolase) in insulin secretion by glucose in pancreatic beta cells. Enhanced insulin secretion in CD38-expressing transgenic mice.

Cyclic ADP-ribose (cADPR) serves as a second messenger for Ca2+ mobilization in insulin secretion, and CD38 has both ADP-ribosyl cyclase and cADPR hydrolase activities (Takasawa, S., Tohgo, A., Noguchi, N., Koguma, T., Nata, K., Sugimoto, T., Yonekura, H., and Okamoto, H. (1993) J. Biol. Chem. 268, 26052-26054). Here, we produced transgenic mice overexpressing human CD38 in pancreatic beta cells. The enzymatic activity of CD38 in transgenic islets was greatly increased, and ATP efficiently inhibited the cADPR hydrolase activity. The Ca2+ mobilizing activity of cell extracts from transgenic islets incubated in high glucose was 3-fold higher than that of the control, suggesting that ATP produced by glucose metabolism increased cADPR accumulation in transgenic islets. Glucose- and ketoisocaproate-induced but not tolbutamide- nor KCl-induced insulin secretions from transgenic islets were 1.7-2.3-fold higher than that of control. In glucose-tolerance tests, the transgenic serum insulin level was higher than that of control. The present study provides the first evidence that CD38 has a regulatory role in insulin secretion by glucose in beta cells, suggesting that the Ca2+ release from intracellular cADPR-sensitive Ca2+ stores as well as the Ca2+ influx from extracellular sources play important roles in insulin secretion.

ADP-ribosyl Cyclase↗

Engineering of glycerol-stimulated insulin secretion in islet beta cells. Differential metabolic fates of glucose and glycerol provide insight into mechanisms of stimulus-secretion coupling.

Insulin secretion from beta cells in the islets of Langerhans can be stimulated by a number of metabolic fuels, including glucose and glyceraldehyde, and is thought to be mediated by metabolism of the secretagogues and an attendant increase in the ATP:ADP ratio. Curiously, glycerol fails to stimulate insulin secretion, even though it has been reported that islets contain abundant glycerol kinase activity and oxidize glycerol efficiently. We have reinvestigated this point and find that rat islets and the well differentiated insulinoma cell line INS-1 contain negligible glycerol kinase activity. A recombinant adenovirus containing the bacterial glycerol kinase gene (AdCMV-GlpK) was constructed and used to express the enzyme in islets and INS-1 cells, resulting in insulin secretion in response to glycerol. In AdCMV-GlpK-treated INS-1 cells a greater proportion of glycerol is converted to lactate and a lesser proportion is oxidized compared with glucose. The two fuels are equally potent as insulin secretagogues, despite the fact that oxidation of glycerol at its maximally effective dose (2-5 mM) occurs at a rate that is similar to the rate of glucose oxidation at its basal, nonstimulatory concentration (3 mM). We also investigated the possibility that glycerol may signal via expansion of the glycerol phosphate pool to allow enhanced fatty acid esterification and formation of complex lipids. Addition of Triacsin-C, an inhibitor of long-chain acyl-CoA synthetase, to AdCMV-GlpK-treated INS-1 cells did not inhibit glycerol-stimulated insulin secretion despite the fact that it blocked glycerol incorporation into cellular lipids. We conclude from these studies that glycerol kinase expression is sufficient to activate glycerol signaling in beta cells, showing that the failure of normal islets to respond to this substrate is due to a lack of this enzyme activity. Further, our studies show that glycerol signaling is not linked to esterification or oxidation of the substrate, but is likely mediated by its metabolism in the glycerol phosphate shuttle and/or the distal portion of the glycolytic pathway, either of which can lead to production of ATP and an increased ATP:ADP ratio.

Adenoviridae↗

Disruption of the WFS1 gene in mice causes progressive beta-cell loss and impaired stimulus-secretion coupling in insulin secretion.

Wolfram syndrome, an autosomal recessive disorder characterized by juvenile-onset diabetes mellitus and optic atrophy, is caused by mutations in the WFS1 gene. In order to gain insight into the pathophysiology of this disease, we disrupted the wfs1 gene in mice. The mutant mice developed glucose intolerance or overt diabetes due to insufficient insulin secretion in vivo. Islets isolated from mutant mice exhibited a decrease in insulin secretion in response to glucose. The defective insulin secretion was accompanied by reduced cellular calcium responses to the secretagogue. Immunohistochemical analyses with morphometry and measurement of whole-pancreas insulin content demonstrated progressive beta-cell loss in mutant mice, while the alpha-cell, which barely expresses WFS1 protein, was preserved. Furthermore, isolated islets from mutant mice exhibited increased apoptosis, as assessed by DNA fragment formation, at high concentration of glucose or with exposure to endoplasmic reticulum-stress inducers. These results strongly suggest that WFS1 protein plays an important role in both stimulus-secretion coupling for insulin exocytosis and maintenance of beta-cell mass, deterioration of which leads to impaired glucose homeostasis. These WFS1 mutant mice provide a valuable tool for understanding better the pathophysiology of Wolfram syndrome as well as WFS1 function.

Adenoviridae↗