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Biomedical subjects

J R Garrett

Publications and source records attributed to J R Garrett.

At least 109 records · Page 6Linked to original sources
PubMed1987

Selection of a simple protease procedure for identifying mast cells in routinely processed human tissues.

Proteases present in mast cell granules have been harnessed to demonstrate mast cells in human tissues. A number of substrate mixtures were tested. D-Val-Leu-Arg-4-methoxy-2-naphthylamide (MNA) plus Fast Blue B was the best for identifying human mast cells, yielding the most specific and complete staining. The procedure is simple and the results are permanent. Cryostat sections of aldehyde-fixed routine preparations or paraffin sections of Carnoy-fixed tissues give the most satisfactory results. Mast cells are stained a strong red color that stands out distinctly from the surrounding tissues, so that they can be easily identified by simple microscopy. A double-staining technique, first for protease and subsequently using Alcian Blue, showed that as progressive protease staining occurs, the alcianophilia of mast cells is lost. This procedure demonstrated that mast cells in the mucosa of human gut generally required longer incubations to develop protease staining than in other connective tissue sites. In post-mortem tissues, mast cells retain their protease activity well and so can be demonstrated in cryostat sections of aldehyde-fixed material, giving a more complete picture than with Alcian Blue. The synthetic substrate D-Val-Leu-Arg-MNA can be recommended for routine identification of mast cells in human tissues.

Alcian Blue↗
PubMed1986

Lipid accumulation in the major salivary glands of streptozotocin-diabetic rats.

Four-five months after the induction of diabetes, salivary tissues of male Wistar rats were preserved by glutaraldehyde fixation or rapid freezing in dry-ice cooled hexane. Fixed tissues were either processed and embedded for light and electron microscopy, or frozen and, together with unfixed tissues, sectioned and stained with Oil Red 0 or by the calcium-lipase method. All diabetic glands had considerably more intracellular lipid than control ones. Lipid accumulation within parenchymal cells varied with the type of gland, and was more pronounced in animals with the highest serum-glucose levels. Serous cells of parotid and sublingual glands accumulated the greatest amount of lipid; lesser amounts were present in seromucous acinar cells of submandibular glands; little or none in mucous acinar cells of sublingual glands. There was no lipid in striated, granular or excretory ducts. Histochemical staining suggested that the intracellular lipid was mainly triglyceride which may accumulate by increased uptake for use as an energy source, or by decreased use in the synthesis of secretory granule and plasma-membrane material.

Animals↗
PubMed1986

Structural and functional studies of the effects of parasympathetic nerve stimulation on rabbit submandibular salivary glands.

The effects were assessed by combining physiological experiments with morphological examination, using light and electron microscopy. Continuous, parasympathetic nerve stimulation, at frequencies varying from 1 to 10 Hz, caused a copious flow of saliva. Both acinar and granular tubule cells showed extensive degranulation, but the effects on tubule cells were the more dramatic. After stimulation, there was a marked loss of acidic mucosubstances from the acinar cells and an almost complete loss of neutral mucosubstances from the granular tubule cells; this was particularly evident with stimulation at higher frequencies (6-10 Hz).

Animals↗
PubMed1986

Septoplasty for left ventricular outflow obstruction without aortic valve replacement: a new technique.

A new technique is described for relief of diffuse obstruction in the left ventricular outflow tract without aortic valve replacement. Left ventricular septoplasty was performed, preserving the aortic valve. The supra-valve stenosis was repaired using a Y-shaped extension of the aortotomy proximally and a pantaloon-shaped patch of woven Dacron fabric. A 10-year-old girl with "tunnel" or diffuse stenosis obtained striking relief of left ventricular hypertension by this technique.

Aortic Valve Stenosis↗
PubMed1986

Ultrastructural and histochemical observations on intracellular and luminal microcalculi in the feline sublingual salivary gland.

As the genesis of salivary calculi in man has not been established, the fortuitous observation of microcalculi in sublingual glands of cats prompted this study. Microcalculi were seen occasionally within acinar cells and lumina and macrophages, and rarely interstitially. Microcalculi were stained with the periodic-acid/Schiff technique, and acid-phosphatase activity was associated with intracellular microcalculi. Intracellular microcalculi were seen ultrastructurally in acinar cells in membrane-bound vacuoles that also contained debris and sometimes what appeared to be secretory material. X-ray microanalysis showed the microcalculi to consist of crystals containing calcium and phosphorus. The observations suggest that the microcalculi form within autophagosomes in acinar cells by a precipitation of calcium and phosphorus present in degenerate material, and that the microcalculi eventually enter lumina, where they may be discharged in the saliva or retained and enlarge.

Animals↗
PubMed1985

Use of different derivatives of D-Val-Leu-Arg for studying kallikrein activities in cat submandibular glands and saliva.

Glandular kallikrein shows a special selectivity for D-Val-Leu-Arg-4-methoxy-2-naphthylamide in comparison with other potential oligopeptide substrates and it provides a useful histochemical substrate, although the reaction may not always be specific. However, in cat submandibular saliva, a biochemical assay using the closely related D-Val-Leu-Arg-7-amino-4-trifluoromethylcoumarin (AFC) as substrate, which affords more sensitive detection, showed that soya bean trypsin inhibitor causes no inhibition. This indicates that there are unlikely to be contaminating enzymes competing for the substrate in this body fluid. Support for this observation has been gained by the useful new enzyme overlay membrane technique for fluorescent assessment of reactive bands of enzymes after isoelectric focusing, using membranes of cellulose acetate impregnated with D-Val-Leu-Arg-AFC. Comparison of results after isoelectric focusing of purified cat submandibular kallikrein with samples of cat submandibular saliva confirmed that the substrate is monospecific for kallikrein in saliva of the cat. This knowledge has enabled us to start assessing the dynamics of the secretion of kallikrein by the gland. Testing individual drops of saliva has shown that an amazingly rapid mobilization of kallikrein occurs in high concentrations on sympathetic nerve stimulation. The corresponding oligopeptide-based inhibitor D-Val-Leu-Arg-chloromethyl ketone was found to be strongly inhibitory of the amidase reaction by kallikrein but showed a low specificity for kallikrein. Nevertheless, its effects have been tested in vivo by the intravascular route and it caused an increase in the resting salivary vascular resistance whether administered close-arterially or intravenously. Thus, it would seem that a kallikrein-like protease does influence the background tone in the vessels and the source of this enzyme is thought to be mast cells.

Amino Acid Chloromethyl Ketones↗
PubMed1985

Histochemistry of hydrolytic enzymes in resting submandibular glands of rabbits.

The distribution of several hydrolytic enzymes was investigated in rabbit submandibular glands at both the light and electron microscopical levels. Glands were fixed by either immersion or perfusion fixation with a variety of fixatives containing 1-2% glutaraldehyde and 2-4% formaldehyde in 0.1 M cacodylate buffer at pH 7.2. Light microscopically, the acinar cells showed some staining for ATPase, acid phosphatase and nonspecific esterases but showed weak staining for thiamine pyrophosphatase. Acid phosphatase staining occurred most strongly in granular tubule cells. Staining for esteroproteases was confined to the periluminal rims of intercalary and striated ducts. Alkaline phosphatase was very sensitive to glutaraldehyde and was confined to myoepithelial cells. Electron microscopical observations revealed the presence of acid phosphatase reaction product in lysosomes, immature granules and in GERL-like structures, the last being much more conspicuous in the granular tubule cells. ATPase reaction product was localized to the basal and luminal plasma membranes and lumina of both acinar and granular tubule cells. The Golgi complex of these two types of cells exhibited only moderate amounts of reaction product for thiamine pyrophosphatase. Alkaline phosphatase activity, on the other hand, was exclusively localized to myoepithelial cells in their plasma membranes and sometimes in the nuclear envelope.

Acid Phosphatase↗
PubMed1985

Secretion of parotid acinar granules in rats during reflex stimulation after chronic sympathectomy.

After chronic post-ganglionic sympathectomy the ultrastructure of 'resting' parotid acinar cells in rats remains relatively normal, despite the loss of nerve impulses normally causing secretion of acinar granules from this gland. The question whether eating hard chow still induces some acinar degranulation weeks after superior cervical ganglionectomy, has now been tested. It was found that scattered pockets of parotid acinar degranulation did occur. Some of these changes after unilateral denervation may have been attributable to intact sympathetic nerves arising from the contralateral superior cervical ganglion (Alm, Asking, Emmelin & Gjörstrup, 1984), but this cannot explain similar changes that occurred in glands after bilateral ganglionectomy. The possibility that some degranulation may be induced by circulating catecholamines was tested, 9-16 weeks after unilateral sympathectomy, by placing animals in the cold (0-4 degrees C) for 2 h before removal of tissues. Animals not given food during that time showed no evidence of parotid acinar degranulation, but those given hard chow showed extensive acinar degranulation in both parotid glands. On the sympathectomized side the degranulation was, however, less uniform and more patchy than on the side with an intact innervation. These findings agree with he belief that circulating catecholamines can, at times, induce degranulation of parotid acinar cells made supersensitive by chronic sympathetic denervation but, for it to occur, the cells must also be receiving parasympathetic stimulation at the same time. This interaction probably permits a sufficient turnover of granules to maintain a relative homoeostasis of the parotid acinar cells in rats.

Animals↗
PubMed1984

Influence of circulating catecholamines on protein secretion into rat parotid saliva during parasympathetic stimulation.

Secretion of proteins by rat parotid glands in response to parasympathetic nerve stimulation was studied in vivo during pentobarbitone anaesthesia. Parasympathetic stimulation (3-10 Hz) via the auriculotemporal nerve resulted in a copious flow of saliva low in protein. In contrast, sympathetic stimulation (5 Hz) via the cervical sympathetic trunk evoked saliva low in volume but high in protein. Nevertheless, the specific concentrations of amylase and peroxidase (mg/mg protein) and the ratio of amylase to peroxidase remained constant. Sodium dodecyl sulphate (SDS) polyacrylamide gel electrophoresis revealed a single, rapidly migrating protein band of unknown identity in proportionately greater amounts in parasympathetic saliva than in sympathetic saliva. Bilateral adrenalectomy led to reduced amylase and peroxidase secretion in response to parasympathetic stimulation both on a mg/ml and a mg/mg protein basis. SDS gel electrophoresis also demonstrated the decrease in specific amylase concentration following adrenalectomy. The ratio of amylase to peroxidase, however, was not significantly affected. Administration of 6-hydroxydopamine 17-72 h prior to adrenalectomy caused no further reduction in the secretion of amylase and peroxidase. Chronic sympathectomy of 2.5-4 months duration resulted in an increased protein secretion (mg/ml) by the parotid gland in response to parasympathetic stimulation. This increase was only slightly reduced by bilateral adrenalectomy. However, as observed in non-sympathectomized rats, adrenalectomy caused a significant reduction in the specific concentrations of both amylase and peroxidase, but did not affect the amylase to peroxidase ratios. We conclude that parasympathetic nerve stimulation of rat parotid glands after overnight starvation causes secretion of proteins in proportions similar to, but in significantly lower concentrations than those found in sympathetic saliva. Circulating catecholamines, however, influence the amount of amylase and peroxidase secreted by the rat parotid gland in response to parasympathetic nerve stimulation and account for most of the increased secretion of these enzymes following chronic sympathectomy.

Adrenalectomy↗
PubMed1984

Diabetic gastroparesis from autonomic neuropathy: surgical considerations and changes in vagus nerve morphology.

Two cases with intractable vomiting due to gastroparesis, a rare feature of diabetic autonomic neuropathy, are described. Both required surgical treatment. In the first a gastroenterostomy was complicated by reflux gastritis requiring a revision operation; in the second a gastrojejunostomy was successful. Electron microscopic studies of the vagus nerve in one of the cases showed a severe reduction in the density of unmyelinated axons, the surviving axons tending to be of small calibre. The severity of the abnormalities supports the view that diabetic gastroparesis is related to vagal denervation.

Adult↗
PubMed1984

Constipation and congenital disorders of the myenteric plexus.

Full-thickness muscle biopsies have been taken from patients with severe disabling chronic constipation that has not responded to conservative measures. Assessment by neurohistochemical techniques has revealed that a range of neuronal dysplasias of the myenteric plexus are responsible in many cases; these include aganglionosis (Hirschsprung's disease), hypoganglionosis and hyperganglionosis. In cases considered unlikely to be Hirschsprung's disease on clinical grounds, the procedure used has often been anorectal myectomy; this has not only provided tissue for diagnosis but has also been of therapeutic value in most cases of hypoganglionosis and some cases reported as 'normal'.

Adult↗
PubMed1983

Tissue reactions to the separate implantation of individual constituent phases of dental amalgam, including assessment by energy dispersive X-ray microanalysis.

Soft tissue degradation of the 3 principal amalgam phases have been investigated in relation to their role in the formation of the amalgam tattoo. Each phase, finely powdered, was implanted subcutaneously into the submandibular region of guinea-pigs for periods ranging from 1 week to 1 year. The rates of breakdown were assessed radiographically and the final lesions were examined by light and electron microscopy and X-ray microanalysis. gamma 2 (Sn7Hg) phase degraded rapidly, mainly extracellularly, and did not produce a tattoo. Both mercury and tin disappeared from the lesion. gamma 1 (Ag2Hg3) phase degraded less rapidly, both extra and intracellularly, and produced a small tattoo. Mercury was lost from the lesion. gamma (Ag3Sn) phase degraded slowly, intracellularly, and produced a large tattoo. Tattoos always resulted from persistence of minute particles of silver and sulphur associated with basal lamina and connective tissue.

Animals↗
PubMed1983

Atropine-resistant secretion of parotid saliva on stimulation of the auriculo-temporal nerve.

Non-adrenergic non-cholinergic parasympathetic secretory responses in parotid glands of rats have been demonstrated by electrical stimulation of the auriculo-temporal nerve after appropriate blocking drugs and compared with secretion in the absence of these drugs. After atropine +/- adrenergic blocking drugs secretion was greatly reduced but some secretion was always evident at frequencies of 5 Hz and higher. The time taken for secretion to appear was greatly increased and this may relate to an absence of accompanying myoepithelial contraction. Continuous stimulation always produced greater flows of saliva than intermittent bursts of stimulation at higher frequencies both in the absence and in the presence of blocking drugs. Thus it is likely in vivo that secretion is induced by relatively steady nerve-impulse traffic at moderate frequencies rather than by bursts of high frequency stimulation. Possible transmitters contributing to the atropine-resistant secretion are considered.

Adrenergic alpha-Antagonists↗
PubMed1982

Kallikrein-like activity in salivary glands using a new tripeptide substrate, including preliminary secretory studies and observations on mast cells.

The tripeptide substrate D-val-leu-arg-4-methoxy-2-naphthylamine gave a precise localization of reaction product in cryostat sections of aldehyde-fixed salivary glands from a number of species, with Fast Blue B as the capture reagent. In submandibular glands, there was strong staining of the granules in granular tubules of rats and hamsters and somewhat less in mice. Submandibular striated ducts showed variable periluminal staining in a finer granular form; it was abundant in guinea-pigs, strong in cats but somewhat less pronounced in dogs. Parotid glands contained less reactivity with none detectable in hamsters and guinea-pigs. In the rabbit, neither gland showed any reaction. Mast cells were densely stained in glands from cats and dogs; they were less reactive in rats and unstained in the other species. The closely related 7-amino-4-trifluoromethylcoumarin derivative of the tripeptide has been found highly satisfactory for assessing activity in submandibular saliva from cats. Preliminary functional studies indicate that an extensive rapid secretion of enzyme occurs into saliva on sympathetic stimulation, with a corresponding depletion of reactive material from the striated ducts in tissue sections. Far less mobilization of enzyme occurs into saliva on parasympathetic stimulation with no obvious change in the histochemical reaction of striated ducts. The possible significance of these findings in cats is discussed. Extensive qualitative and quantitative studies are required to evaluate enzyme and substrate specificities in each species. Nevertheless, derivatives of D-val-leu-arg offer great promise for the functional testing of kallikrein-like reactivity both by histochemical means on cells and biochemically in their secretions.

Animals↗