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Biomedical subjects

A E Clark

Publications and source records attributed to A E Clark.

At least 55 records · Page 3Linked to original sources
PubMed1991

Determination of the rates of appearance and loss of glucose transporters at the cell surface of rat adipose cells.

We have used an impermeant bis-mannose compound (2-N-[4-(1-azi-2,2,2-trifluoroethyl)benzoyl]-1,3-bis-(D-mannos+ ++- 4-yloxy)-2- propylamine; ATB-BMPA) to photolabel the glucose transporter isoforms GLUT4 and GLUT1 that are present in rat adipose cells. Plasma-membrane fractions and light-microsome membrane fractions were both labelled by ATB-BMPA. The labelling of GLUT4 in the plasma membrane fraction from insulin-treated cells was approximately 3-fold higher than that of basal cells and corresponded with a decrease in the labelling of the light-microsome fraction. In contrast with this, the cell-surface labelling of GLUT4 from insulin-treated intact adipose cells was increased approximately 15-fold above basal levels. In these adipose cell preparations, insulin stimulated glucose transport activity approximately 30-fold. Thus the cell-surface labelling, but not the labelling of membrane fractions, closely corresponded with the stimulation of transport. The remaining discrepancy may be due to an approx. 2-fold activation of GLUT4 intrinsic transport activity. We have studied the kinetics of trafficking of transporters and found the following. (1) Lowering the temperature to 18 degrees C increased basal glucose transport and levels of cell-surface glucose transporters by approximately 3-fold. This net increase in transporters probably occurs because the process of recruitment of transporters is less temperature-sensitive than the process involved in internalization of cell-surface transporters. (2) The time course for insulin stimulation of glucose transport activity occurred with a slight lag period of 47 s and a t 1/2 3.2 min. The time course of GLUT4 and GLUT1 appearance at the cell surface showed no lag and a t 1/2 of approximately 2.3 min for both isoforms. Thus at early times after insulin stimulation there was a discrepancy between transporter abundance and transport activity. The lag period in the stimulation of transport activity may represent the time required for the approximately 2-fold stimulation of transporter intrinsic activity. (3) The decrease in transport activity after insulin removal occurred with a very high activation energy of 159 kJ.mol-1. There was thus no significant decrease in transport or less of cell-surface transporters over 60 min at 18 degrees C. The decrease in transport activity occurred with a t1/2 of 9-11 min at 37 degrees C.(ABSTRACT TRUNCATED AT 400 WORDS)

3-O-Methylglucose↗
PubMed1991

Chronic treatment with insulin selectively down-regulates cell-surface GLUT4 glucose transporters in 3T3-L1 adipocytes.

A new method for photoaffinity labeling of glucose transporters has been used to compare the effects of glucose-starvation, acute-insulin, and chronic-insulin treatments on the cell-surface glucose transporters in 3T3-L1 adipocytes. Starvation alone increased the cell-surface levels of GLUT1 and GLUT4 by approximately 4- and approximately 2-fold, respectively. As shown by Calderhead, D, M., Kitagawa, K., Tanner, L.T., Holman, G.D., and Lienhard, G.E. (1990) J. Biol. Chem. 265, 13800-13808) acute-insulin treatment increased cell-surface GLUT1 and GLUT4 by approximately 5- and approximately 15-fold respectively. In contrast to this, chronic-insulin treatment gave a further 3-4-fold increase in both cell-surface and total cellular GLUT1, but availability of GLUT4 at the cell-surface was down-regulated to half the level found in the acute treatment but with no change in the total cellular level. This effect occurred in starved and non-starved cells and suggests that starvation, acute-insulin, and chronic-insulin treatments regulate glucose transporter availability through independent mechanisms. The down-regulation of GLUT4 reached a maximally reduced cell-surface level in 6 h while the rise in GLUT1 reached a maximum after 24-48 h. The rise in GLUT1 appeared to compensate for the decline in cell-surface GLUT4 as glucose transport activity was further increased during the long term treatment with insulin. The down-regulation of GLUT4 due to the chronic-insulin treatment is associated with a marked resistance of the cells to restimulate glucose transport and particularly to recruit further GLUT4 to the cell-surface following an additional insulin treatment. The defect appears to be in the signaling mechanism that is responsible for translocation.

Adipose Tissue↗
PubMed1991

An investigation of bioactive glass powders by sol-gel processing.

Bioactive glass powders, with a composition of SiO 2-CaO-P 2O 5, have been successfully synthesized via a sol-gel process at considerably lower temperatures than required for conventional melting methods. Bioactive glass powders made via conventional methods form an interfacial bond with bone when they are implanted. Bonding is correlated with the formation of a surface hydroxyapatite layer. This study examined the formation of a hydroxyapatite layer in Tris-buffered solution as a function of SiO 2 content of sol-gel derived powders. A FT-IRRS technique was used to monitor the formation of the hydroxyapatite on the surface of the powders. X-ray diffraction analysis and BET were also used to characterize the chemical and physical properties of the sol-gel derived bioactive powders. It was discovered that: (a) the rate of hydroxyapatite formation decreased with increasing SiO 2 content for powders whose SiO 2 content was less than 90 mol%; (b) a hydroxyapatite film does not form for the powders whose SiO 2 content is more than 90 mol%; (c) the SiO 2 limit, beyond which the powders lost their bioactivity, was much higher for bioactive glass powders made through sol-gel process (90%) than those made by conventional melting methods (60%). These results indicate that it is possible to significantly expand the bioactive composition range through microstructural control made possible by sol-gel processing techniques.

Glass↗
PubMed1990

Cell surface labeling of glucose transporter isoform GLUT4 by bis-mannose photolabel. Correlation with stimulation of glucose transport in rat adipose cells by insulin and phorbol ester.

A new impermeant photoaffinity label has been used for identifying cell surface glucose transporters in isolated rat adipose cells. This compound is 2-N-4(1-azi-2,2,2-trifluoroethyl)benzoyl-1,3-bis(D-mannos-4- yloxy)-2- propylamine. We have used this reagent in combination with immunoprecipitation by specific antibodies against the GLUT4 and GLUT1 glucose transporter isoforms to estimate the relative abundance of these two transporters on the surface of the intact adipose cell following stimulation by insulin and phorbol 12-myristate 13-acetate (PMA). In the basal state, GLUT4 and GLUT1 are both present at the cell surface but GLUT4 is more abundant than GLUT1. In response to insulin, GLUT4 increases 15-20-fold and GLUT1 increases approximately 5-fold while 3-O-methyl-D-glucose transport is stimulated 20-30-fold. By contrast, PMA only induces a approximately 4-fold increase in GLUT4 while GLUT1 increases approximately 5-fold to the same level as seen with insulin. In addition, PMA stimulates 3-O-methyl-D-glucose transport approximately 3-fold to only 13% of the insulin-stimulated state. Thus GLUT4 is the major glucose transporter isoform under all conditions, and it is selectively and markedly enriched in response to insulin but not PMA which increases GLUT1 and GLUT4 equally. Furthermore, stimulation of glucose transport activity correlates closely with the appearance of GLUT4 on the cell surface in response to both insulin and PMA but does not correlate with the sum of GLUT1 and GLUT4 appearance. These results suggest that GLUT4 may be inherently more active than GLUT1 due to a higher TK (turnover/Km).

Adipose Tissue↗
PubMed1990

Exofacial photolabelling of the human erythrocyte glucose transporter with an azitrifluoroethylbenzoyl-substituted bismannose.

The synthesis of 2-N-[4-(1'-azitrifluoroethyl)benzoyl]-1,3-bis-(D-mannos-4-++ +yloxy)-2- propylamine (ATB-BMPA) is described. This compound was used as an exofacial probe for the human erythrocyte glucose-transport system. A new method is described for directly estimating the affinity for exofacial ligands which bind to the erythrocyte glucose transporter. By using this equilibrium-binding method, the Ki for ATB-BMPA was found to be 338 +/- 37 microM at 0 degrees C and 368 +/- 59 microM at 20 degrees C. This was similar to the concentration of ATB-BMPA required to half-maximally inhibit D-galactose uptake (Ki = 297 +/- 53 microM). The new photoaffinity reagent labelled the glucose transporter in intact cells but, because of its improved selectivity, was also used to label the glucose transporter in isolated erythrocyte membranes. The ATB-BMPA-labelled glucose transporter was 80% immunoprecipitated by anti-(GLUT1-C-terminal peptide) antibody, which shows that the GLUT1 glucose transporter is the major isoform present in erythrocytes. The labelling of the glucose transporter at its exofacial site, and the adoption of an outward-facing conformation, renders the transport system resistant to thermolysin and trypsin treatment. Trypsin treatment of the unlabelled glucose transporter in erythrocyte membranes produced an 18 kDa fragment which was subsequently labelled by ATB-BMPA, but had low affinity for this exofacial ligand. This suggests that the trypsin-treated transporter adopts an inward-facing conformation. The ability of D-glucose to displace ATB-BMPA from the native transporter and from the 18 kDa trypsin fragment have been compared. The D-glucose concentration which was required to obtain half-maximal inhibition of ATB-BMPA labelling was 6-fold lower for the 18 kDa tryptic fragment.

Azides↗
PubMed1989

An investigation of Bioglass powders: quality assurance test procedure and test criteria.

Bioglass, in bulk form, has been used to bond to the body's hard and soft tissues in many surgical and dental applications over the past 18 years. There is also a large potential use of Bioglass, in powder or paste from, to fill irregular-shaped bony defects caused by local or systemic disease, congenital malformations, or trauma in the fields of otolaryngology and dental sciences. A quality assurance test procedure to analyze powdered Bioglass to guarantee that the bioactivity of the glass has been maintained during manufacturing was developed. Herein states the quality assurance test procedure, test variables, and test criteria to assure the bioactivity of Bioglass powders made from known bioactive bulk Bioglass samples. Also described herein is the path taken to develop the test procedure and test criteria and experiments performed to study the test variables.

Bone Cements↗
PubMed1987

Residual alveolar ridge maintenance with a new endosseous implant material.

In an effort to maintain the height of the residual alveolar ridge, Bioglass cones were placed in fresh sockets after tooth extraction. Today the emphasis is on maintaining the residual alveolar ridge instead of letting it resorb and subsequently resorting to heroic efforts for augmentation. To date, 242 cone implants have been placed in 29 patients. Implant follow-up observation time ranged from 12 to 32 months with an average of 19.9 months. Twenty-seven patients have been fitted with dentures. To date seven implants (2.9%) have been lost. The Bioglass implants appear to be highly biocompatible as evidenced by absence of infection, normal mucosal healing, and radiographic evidence of bone healing around the implants. Evaluation of several implants reexposed surgically revealed new bone in direct contact with the immobile implants circumferentially. The problem of implant dehiscence that started to occur after approximately 10 months in other investigations had not manifested itself in the Bioglass implant patients who have been wearing their dentures for at least 12 months.

Alveolar Process↗
PubMed1987

An unusual presentation of temporal arteritis.

Serious ocular and systemic complications may result from inflammatory processes of the orbit. Temporal arteritis is one such process that can affect the eye. The patient described herein is unusual because she presented initially with signs and symptoms similar to orbital cellulitis. As her disease evolved, she displayed marked, hemorrhagic chemosis with some limitation of abduction and resistance to retropulsion on the involved side. Laboratory and radiologic findings eliminated orbital cellulitis, limited Wegener's granulomatosis, and other systemic inflammatory disorders and led to the diagnosis of temporal arteritis. To our knowledge, this complex of presenting signs has not been previously reported.

Aged↗
PubMed1985

DNA repeat length in chromatin from murine bone marrow and L1210 leukaemia cells.

Previous studies have suggested that 1-(4-amino-2-methylpyrimidine-5-yl)-methyl-3-(2-chloroethyl) -3-nitrosoureahydrochloride (ACNU) and 1,(2-chloroethyl)-3-cyclohexyl-1-nitrosourea (CCNU) bind specifically to the nucleosomal DNA of murine bone marrow and L1210 leukaemia cells whereas the glucose nitrosoureas, 2-(3-(2-chloroethyl)-3-nitrosoureido)-2-deoxy-D-glucopyranose, (chlorozotocin, CLZ) and 1-(2-chloroethyl)-3-(-D-glucopyranosyl)-1-nitrosourea (GANU), bind preferentially to the linker DNA of bone marrow but not tumour cell chromatin. In order to provide an explanation for this differential, the DNA repeat and linker lengths in murine bone marrow and L1210 leukaemia cells were measured using electrophoresis of micrococcal nuclease-digested DNA. The linker length of bone marrow chromatin was approximately 22% longer than that in L1210 leukaemia cells from mouse ascites. The linker length of L1210 cells maintained in suspension culture was 27% less than in those from ascites fluid. The tissue-specific toxicity of sugar nitrosoureas and the differential binding of these drugs to chromatin does not appear to correlate quantitatively with differences in DNA linker length.

Animals↗
PubMed1983

Studies on the efficacy of implants as orthodontic anchorage.

The abilities of two types of small cylindrical endosseous implant to resist movement when loaded with constant forces of orthodontic magnitudes were tested. Pairs of Bioglass-coated implants and pairs of Vitallium implants of the same size were implanted into the femurs of twelve rabbits. After a 28-day healing period, these implants were loaded with forces of 60 grams, 120 grams, and 180 grams. Analysis of implant movement after 28 days revealed that no statistically significant movement occurred at any of the three force levels for either type of implant. Histologic evaluation revealed a connective tissue encapsulation with the Vitallium implant and an implant-bone bond with the Bioglass implant. No histologic evidence of implant movement was observed for either implant type at any force level.

Animals↗
PubMed1983

Treatment of herpes labialis.

Untreated herpes labialis may take up to 10 days or more to heal and antiviral therapy must be initiated early if it is to be effective. Acyclovir ointment has been reported to have an antiviral effect but no clinical benefit and a newer formulation, acyclovir cream, has been developed in an attempt to enhance skin penetration. In placebo-controlled trials, acyclovir cream shortened the duration of lesions significantly and when therapy was started before vesicles developed, the number of lesions aborting was increased significantly. This therapy was well tolerated by most patients. Topical acyclovir offers hope for the successful management of herpes labialis where systemic therapy is considered inappropriate.

Acyclovir↗
PubMed1981

Ultrastructural localisation of lactoferrin and glycoprotein in human bronchial glands.

The distribution of lactoferrin and glycoprotein in human bronchial glands has been studied by electron microscopy using an immunoperoxidase method to stain the former and a periodic acid-chromic acid-silver methenamine sequence for the latter, each applied to ultrathin Epon sections. The distribution of lactoferrin corresponds to that of lysozyme. Lactoferrin and lysozyme are both confined to serous acini where the granules show a variable pattern of staining. Some serous granules are filled uniformly with lactoferrin, some lack lactoferrin in a small central core or a thin peripheral rim, and some are completely devoid of lactoferrin. Glycoprotein is present in all mucous granules but only certain serous granules. The latter may be filled uniformly with glycoprotein or glycoprotein may form a thin peripheral coat about centrally located lactoferrin and lysozyme. An electron-dense central core found in some serous granules contains neither glycoprotein, lactoferrin, nor lysozyme.

Bronchi↗