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G B Proctor

Publications and source records attributed to G B Proctor.

At least 37 records · Page 2Linked to original sources
PubMed1999

O-linked glycosylation occurs on basic parotid salivary proline-rich proteins.

Interactions between salivary glycoproteins and many oral bacteria have been shown to depend on O-linked glycans on salivary glycoproteins. Basic proline-rich proteins form the largest group of proteins within human parotid saliva. In the present study human parotid salivary glycoproteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis or two-dimensional electrophoresis, electroblotted onto nitrocellulose and probed with two biotin-labelled lectins from Maclura pomifera (MPA) and Arachis hypogaea (PNA) which are specific for O-linked (galactose beta 1,3 N-Acetylgalactosamine) glycans. Lectin binding was detected with avidin-biotin complex and enhanced chemiluminescence. Two-dimensional electrophoresis in combination with lectin binding indicated that only basic parotid salivary glycoproteins bind the lectin MPA. Following removal of terminal sialic acid residues by sialidase digestion the same glycoproteins were detected by the lectin PNA. Glycosidase digestion with endo-alpha-N-acetylgalactosaminidase (O-glycanase) in conjunction with sialidase eliminated MPA binding. Taken together these results indicate that many basic parotid salivary glycoproteins contain O-glycans, all of which are sialylated.

Electrophoresis, Gel, Two-Dimensional↗
PubMed1999

Enzyme histochemistry of tryptase in stomach mucosal mast cells of the mouse.

We investigated the histochemical characteristics of mast cell tryptase in different mouse tissues. By use of peptide substrates, tryptase activity could be demonstrated in unfixed connective tissue mast cells in different tissues, including the stomach. Tryptase activity was better localized after aldehyde fixation and frozen sectioning, and under such conditions was also demonstrated in mucosal mast cells of the stomach but not in those of the gut mucosa. Double staining by enzyme histochemistry followed by toluidine blue indicated that the tryptase activity was present only in mast cells and that all mast cells in the stomach mucosa contained the enzyme. The peptide substrates z-Ala-Ala-Lys-4-methoxy-2-naphthylamide and z-Gly-Pro-Arg-4-methoxy-2-naphthlyamide, which are substrates of choice for demonstrating tryptase in other species, were most effective for demonstrating mouse tryptase. The use of protease inhibitors further indicated that activity present in all mast cells was tryptase. Safranin O did not stain stomach mucosal mast cells, suggesting that the tryptase present in these cells was active in the absence of heparin sulfate proteoglycan.

Animals↗
PubMed1998

The influence of nerves on the secretion of immunoglobulin A into submandibular saliva in rats.

1. The influence of sympathetic and parasympathetic nerve stimulations on salivary secretion of immunoglobulin A (IgA) was studied in the submandibular glands of anaesthetized rats by stimulating the nerve supplies with bipolar electrodes. 2. Although the flow of saliva from sympathetically stimulated glands was only 23% of that from parasympathetically stimulated glands the output of IgA was over 2-fold greater. This difference was attributable to influences of the nerves on IgA secretion through the epithelial cell polymeric immunoglobulin receptor-mediated pathway, as Western blotting with specific antibodies to IgA and secretory component revealed that secretory IgA (SIgA) dominated in all saliva samples. 3. Study of saliva secreted in sequential periods of nerve stimulation or following rest pauses suggested that SIgA secretion occurred in the absence of stimulation but this was upregulated 2.6- and 6-fold by parasympathetic and sympathetic nerve stimulations, respectively, compared with the calculated unstimulated rate. 4. The IgA content of extensively stimulated glands was 77% of levels in unstimulated contralateral control glands despite a secretion into saliva equivalent to almost 90% of the glandular IgA content. The IgA may be synthesized and secreted by glandular plasma cells at a rate which exceeds demand and/or such synthesis may be upregulated by nerve impulses. 5. The results indicate that salivary secretion of SIgA is upregulated by nerve impulses and that sympathetic nerves induce a greater effect than parasympathetic nerves.

Animals↗
PubMed1998

Protein secretion from rat submandibular acini and granular ducts: effects of exogenous VIP and substance P during parasympathetic nerve stimulation.

The influences of exogenous vasoactive intestinal peptide (VIP) and substance P on the release of peroxidase from acini and true tissue kallikrein (rK1) from granular ducts of the rat submandibular gland were studied during continuous parasympathetic stimulation. Parasympathetic nerve impulses caused a moderate flow of saliva (mean +/- SD, 108+/-26 microl/g tissue/min) that had a low protein concentration (174+/-88 microg/ml). The outputs of peroxidase and rK1 were minimal (14.3+/-11.8 pmol DCF/g tissue/min and 6.5+/-3.4 nmol AFC/g tissue/min, respectively). When administered intravenously, VIP had no apparent effect on the overall flow rate, but caused a significant increase in the output of peroxidase; 450% at 1 microg/kg and a further 10-fold increase at 10 microg/kg. In contrast, substance P (1 microg/kg) evoked a marked increase in flow rate (68%), and peroxidase secretion increased only 3-fold. The output of rK1 was unaffected by either VIP or substance P. Our results support the hypothesis that acinar, but not granular duct, protein secretion is evoked by non-adrenergic, non-cholinergic peptides released from parasympathetic nerve terminals.

Animals↗
PubMed1998

Enzyme histochemistry of rat mast cell tryptase.

Fixation and staining conditions for rat mast cell tryptase and its histochemical distribution in different rat tissues were investigated. Prostate, skin, lung, gut, stomach and salivary glands were fixed in either aldehyde or Carnoy fixatives and then frozen or embedded in paraffin wax. Preservation of tryptase enzymic activity against peptide substrates required aldehyde fixation and frozen sectioning. Of the peptide substrates examined, z-Ala-Ala-Lys-4-methoxy-2-naphthylamide and z-Gly-Pro-Arg-4-methoxy-2-naphthylamide proved the most effective for the demonstration of tryptase. Double staining by enzyme cytochemistry followed by immunological detection of tryptase showed that, in all tryptase-containing mast cells, the enzyme is at least in part active. Conventional dye-binding histochemistry was used to confirm the identity of mast cells. Aldehyde-fixed mucosal mast cells required a much shorter staining time with Toluidine Blue if tissue sections were washed directly in t-butyl alcohol. Double staining by enzyme cytochemistry and dye binding showed that tryptase is absent from mucosal and subepidermal mast cells, which are also smaller in size and appear to contain fewer granules than connective tissue mast cells. This study demonstrates that rat mast cell tryptase, unlike tryptases in other species, is a soluble enzyme. It is stored in an active form and is absent from some mast cell subpopulations in mucosa, skin and lung.

Animals↗
PubMed1998

Constitutive secretion of kallikreins in vivo from rat submandibular glands.

In parasympathetic saliva from rat submandibular glands the relative proportions of the various tissue kallikreins differ from those in sympathetic saliva. Kallikreins in sympathetic saliva arise from exocytosis of prepackaged granules in granular tubules, so the kallikreins in parasympathetic saliva must come from a non-granular pool, and are likely to be secreted through a constitutive vesicular route. During periods devoid of stimulation in anaesthetised rats, the kallikreins have been found to accumulate progressively in glandular lumina in parasympathetic-like proportions. As this transport of kallikrein into lumina occurs continuously in vivo, independently of any stimulation or any secretion of fluid, it must arise by constitutive vesicular secretion. During parasympathetic stimulation, the kallikreins are secreted into the saliva at a greater rate than in the resting state but their proportions remain the same and the means by which this increase occurs is open to debate. Constitutively secreted true tissue kallikrein (rK1) has been found to have a different molecular form from that in secretory granules. The submandibular glands also contribute to the kallikreins normally circulating in the blood. Serum levels of kallikrein increased equally during either parasympathetic or sympathetic stimulation and were independent of the amounts secreted into the saliva, so are likely to have arisen from constitutive secretion via the basal sides of the cells, morphological evidence for which has been found in the mouse (Penschow & Coghlan, 1993).

Animals↗
PubMed1997

Secretory interactions between the sympathetic and parasympathetic innervations of the submandibular gland in the anaesthetized cat.

Interactions between the sympathetic and parasympathetic innervations of the submandibular gland have been investigated in the anaesthetized cat. At low frequencies of chorda lingual (parasympathetic) stimulation, simultaneous stimulation of the ascending cervical sympathetic nerve in bursts (20 Hz for 1 s at 10 s intervals) increased the flow of submandibular saliva, but the effect was never more than additive. The output of protein was consistently reduced by simultaneous stimulation of both the sympathetic and parasympathetic innervations, below that evoked by stimulation of either alone. Sympathetic stimulation was more effective than parasympathetic stimulation in promoting the secretion of tissue kallikrein and peroxidase in the submandibular saliva. The output of the latter enzyme, in response to sympathetic stimulation, was significantly reduced by simultaneous stimulation of the parasympathetic innervation at frequencies greater than 1 Hz, but nevertheless exceeded the amount secreted during chorda stimulation alone. Thus, this protocol provided no evidence of synergy between the two divisions of the autonomic nervous system with respect to any submandibular secretory function that was recorded. However, following the administration of a small dose of atropine (2-15 micrograms kg-1 i.v.), sufficient to block secretion during chorda stimulation alone, the flow of saliva, in response to sympathetic stimulation, was potentiated when superimposed on a background of parasympathetic stimulation at all frequencies that were employed. This effect was abolished by larger doses of atropine, indicating that it was dependent upon activation of muscarinic receptors, only some of which could have been blocked by the initial dose.

Anesthesia↗
PubMed1997

The enzymic potential of tissue kallikrein (rK1) in rat submandibular saliva depends on whether it was secreted via constitutive or regulated pathways.

The enzymic activity and immunoreactivity of rat tissue kallikrein (rK1) secreted at rest by granular duct cells of unstimulated submandibular glands has been compared with that secreted on autonomic nerve stimulation. Although a direct vesicular, constitutive secretory pathway operates for rK1 secretion from granular duct cells of unstimulated and parasympathetically stimulated glands the rK1 was not present in a pro-form and actually showed a greater enzymic activity per unit immunoreactive protein than the granule-derived rK1 in sympathetically evoked saliva. Constitutively secreted rK1 was found to be in a single chain molecular form by reducing SDS gel electrophoresis. In contrast rK1 secreted from the storage granule pool of granular duct cells on sympathetic nerve stimulation was present in much higher amounts and occurred in both one-chain and two-chain forms as revealed by SDS gel electrophoresis under reducing conditions. The lower enzymic potential of rK1 in sympathetically evoked saliva might be accounted for by its conversion to a two-chain form.

Animals↗
PubMed1996

Exocytosis from rat submandibular granular tubules during cyclocytidine stimulation shows unusual features, including changes in the granule membrane.

Sequential secretory changes in granular tubule cells caused by the secretagogue cyclocytidine (75 mg/kg i.p.) were studied at the ultrastructural level, in perfusion (n = 5 animals) and immersion (n = 8 animals) fixed rat submandibular glands, using the periodic acid-thiocarbohydrazide-silver proteinate technique (PA-TCH-SP). The onset of secretion varied from 45 to 75 minutes after administering the cyclocytidine. During the initial stages of overt secretion, structural changes occurred irregularly in a progressive fashion with: (1) an increase in granule membrane staining with PA-TCH-SP and a parallel alignment of the secretory granules with the adjacent apical plasma membrane, which developed a honeycomb-like appearance; (2) docking of these secretory granules to the apical plasma membrane; (3) early secretion of some secretory granules in a semiclassical exocytotic fashion (but this was rarely witnessed). During stages (1) and (2), the cytochemical characteristics of the membrane of the secretory granules, as well as of the plasma membrane, suggest a priming process is occurring. After these initial preparatory phases, further structural changes occurred in the granule membranes with a gradually progressive formation of microvesicles and granule fusions; secretion continued in an explosive manner with proteinaceous material being transferred to lumina in at least three different ways: (1) by typical exocytosis (but it was infrequent); (2) from granules fused intracellularly into aggregates (compound exocytosis); and (3) some apocrine-type of secretion through bleb formation. The formation of these intracellular aggregations was associated with the microvesicles in the granule membranes and some aggregates became very large. Secretion of their contents into lumina occurred through elongated membrane channels. The material secreted included microvesicular forms that had become interiorised in the granular aggregates, and any cytoplasm that may also have been entrapped.

Ancitabine↗