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T Walker

Publications and source records attributed to T Walker.

At least 37 records · Page 2Linked to original sources
PubMed1996

Apoptotic human SH-SY5Y neuroblastoma cells have regularly spaced single strand DNA breaks and increased DNA-dependent protein kinase activity.

SH-SY5Y human neuroblastoma cells died by apoptosis when treated with staurosporine or ceramide. The treated cells had both the nuclear morphology and patterns of DNA fragmentation which are characteristic of apoptosis. Higher order DNA fragments separable by pulse field gel electrophoresis were shown to contain regularly spaced single-strand nicks by producing a laddered pattern upon alkali treatment. Further evidence of DNA damage in treated cells was shown by increased activity of DNA-dependent protein kinase. This human cell model may prove useful in delineating the role of a cellular repair response to DNA damage prior to the irreversible steps of the cell death program.

Apoptosis↗
PubMed1995

A constant current source for extracellular microiontophoresis.

A sophisticated constant-current source suitable for extracellular microiontophoresis of tract-tracing substances, such as Phaseolus vulgaris leucoagglutinin, Biocytin or Fluoro-Gold, is described. This design uses a flyback switched-mode power supply to generate controllable high-voltage and operational amplifier circuitry to regulate current and provide instrumentation. Design features include a fast rise time, +/- 2000 V supply (stable output in < 250 ms), simultaneous load current and voltage monitoring, and separate pumping and holding current settings. Three features of this constant-current source make it especially useful for extracellular microiontophoresis. First, the output voltage monitor permits one to follow changes in the microelectrode resistance during current injection. Second, the voltage-limit (or out-of-compliance) indicator circuitry will sound an alarm when the iontophoretic pump is unable to generate the desired current, such as when the micropipette is blocked. Third, the high-compliance voltage power supply insures up to +/- 20 microA of current through 100 M omega resistance. This device has proven itself to be a reliable constant-current source for extracellular microiontophoresis in the laboratory.

Brain↗
PubMed1995

Differences in DNA fragmentation following transient cerebral or decapitation ischemia in rats.

The time course of appearance of cells with DNA damage was studied in rats following transient severe forebrain ischemia. This DNA damage could be detected by in situ end-labeling on brain sections. The breaks in DNA appeared selectively by day 1 in the striatum and later in the CA1 region of the hippocampus. It was possible by double labeling to show that there was no DNA damage in astrocytes. The DNA breaks consisted of laddered DNA fragments indicative of an ordered apoptotic type of internucleosomal cleavage, which persisted without smearing for up to 7 days of reperfusion. In contrast, the DNA breaks following ischemia induced by decapitation were random and, after gel electrophoresis, consisted of smeared fragments of multiple sizes. There was some early regional cellular death, restricted to the dentate of the hippocampus, prior to the pannecrotic degeneration. It is concluded that transient forebrain ischemia leads to a type of neuronal destruction that is not random necrosis but that shares some component of the apoptotic cell death pathway.

Animals↗
PubMed1995

Multiple roles of phosphatidylinositol 3-kinase in regulation of glucose transport, amino acid transport, and glucose transporters in L6 skeletal muscle cells.

Phosphatidylinositol 3-kinase (PI3k) activity is required for the insulin stimulation of glucose transport in adipocytes and Chinese hamster ovary cells. Wortmannin (WM), an inhibitor of PI3k, inhibits the stimulation of glucose transport by insulin and the gain of glucose transporters at the cell surface. However, the effect of inhibition of PI3k on the maintenance of the basal and the insulin-stimulated glucose transport and on the intracellular donor pool of glucose transporters has not been clarified. Here we show that in L6 skeletal muscle cells in culture WM significantly inhibits the basal PI3k activity (by 40%), decreases the levels of phosphatidylinositol 3,4-phosphate and 3,4,5-phosphate (by about 50%) and abolishes the activation of the enzyme by insulin. WM inhibited the basal rate of transport of glucose (by 45%) and of amino acids through system A (by 25%) and abolished their stimulation by insulin. Insulin caused a transient increase in PI3k activity and PI3k products that returned to basal levels within 40 min, whereas glucose and amino acid transport remained elevated. Under these conditions, WM reduced the rate of glucose and amino acid transport back to basal levels. In unstimulated cells, WM decreased significantly the GLUT4 glucose transporter content at the plasma membrane and prevented the ability of insulin to recruit transporters to this membrane. Interestingly, the intracellular pools of the GLUT3 and GLUT4 glucose transporters were significantly reduced in response to WM treatment alone. We conclude that in muscle cells PI3k activity is required to maintain basal and insulin-stimulated glucose and amino acid transport, as well as to develop the stimulation of the two transport processes in response to the hormone. We hypothesize that PI3k, likely through production of phosphatidylinositol 3,4-phosphate and 3,4,5-phosphate, regulates the basal plasma membrane glucose transporter recycling and the organization of the transporter intracellular pool, in addition to being an insulin signal.

Amino Acids↗
PubMed1995

Thoracoscopy: anesthetic considerations.

Thoracoscopy is surgical procedure which was performed in the early 1900s primarily for the treatment of tuberculosis until antibiotics became the treatment of choice. Over the last two decades, thoracoscopy has experienced a rebirth as the indications for this endoscopic procedure have become numerous. Although less invasive than the thoracotomy, thoracoscopy is not without potential complications. The anesthesia provider must be alert for problems related to one-lung ventilation, lateral positioning, and pleural cavity insufflation. The anesthetic technique must be planned with the physiological changes which occur during thoracoscopy in mind.

Anesthesia↗
PubMed1994

Fc gamma receptor II stimulated formation of inositol phosphates in human platelets is blocked by tyrosine kinase inhibitors and associated with tyrosine phosphorylation of the receptor.

We report that activation of phospholipase C (PLC) by cross-linking of the platelet low-affinity Fc gamma receptor II (Fc gamma RII) is inhibited by two structurally distinct tyrosine kinase inhibitors, staurosporine and ST271. This contrasts with PLC activation induced by thrombin and U46619, a thromboxane mimetic, whose receptors have seven transmembrane domains characteristic of G-protein coupled receptors. Several proteins undergo phosphorylation on tyrosine on Fc gamma RII cross-linking upstream of protein kinase C (PKC), Ca2+ and aggregation, including the Fc gamma RII itself. The role of Fc gamma RII phosphorylation in the regulation of PLC is discussed.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1994

Medium-induced fragility of Schwanniomyces.

Schwanniomyces occidentalis has attracted interest because of its ability to metabolize starch and similar complex carbohydrates. Studies have been undertaken, mostly using defined media, to ascertain conditions for optimal production and secretion of hydrolytic enzymes. Here we demonstrate the fragility of Schw. occidentalis in many defined media. We especially examined viability in YNB (Yeast Nitrogen Base) plus 1% glucose. Without phosphate supplementation, viability was routinely very low at stationary phase (usually less than 37%), whereas viability of stationary-phase cultures in phosphate-supplemented YNB usually exceeded 97%. The negative implications of having many, presumably permeabilized, dead cells present in assays for secretion of enzymes by living yeast cells are discussed.

Culture Media↗
PubMed1993

[Differentiation of schistosome species and strains by DNA hybridization].

Genomic DNA from Schistosoma mansoni and S. japonicum adult worms was hybridized to a 32p-labelled pSM 889 probe after cleavage by restriction endonuclease BglII, BamHI, XbaI and EcoRI, or to a 32p-labelled pSM 389 probe after cleavage by EcoRI. The resulting hybridization banding patterns were significantly different between the two species. Genomic DNA from S. japonicum adult worms of Hunan, Hubei, Jiangxi, Zhejiang and Yunnan isolates was hybridized to a 32p-labelled pSM 389 probe after cleavage by EcoRI. The major hybridization bands were identical while the minor bands were more or less different among the isolates. Of them, isolates Hunan, Hubei and Zhejiang have similar minor bands while isolates Jiangxi and Yunnan have minor bands significantly different from those of the above three isolates and also markedly distinct from those of each other. These results indicate that the major hybridization banding patterns are unique to schistosome species while the minor banding patterns may serve as the basis for strain differentiation.

Animals↗
PubMed1992

Differentiation of HT-29 human colonic adenocarcinoma cells correlates with increased expression of mitochondrial RNA: effects of trehalose on cell growth and maturation.

The HT-29 human adenocarcinoma cell line has been used extensively in the study of colonic cell differentiation and colon cancer. We report here that substitution of glucose with trehalose (alpha-D-glucopyranosyl-alpha-D-glucopyranoside) depresses growth and promotes mucin-producing, goblet-like maturation of HT-29. An initial characterization of this process was made by analyzing several cDNA clones whose RNA templates were differentially expressed at elevated levels in cells grown in trehalose-containing medium. Seven of the 9 clones examined corresponded to 6 mitochondrial genes whose expression levels, relative to those from glucose-grown cells, ranged from approximately 3-fold for 16S rRNA to 8-23-fold for NADH dehydrogenase subunit 4. On the other hand, levels of mitochondrial DNA copy, measured by using NADH dehydrogenase subunit 4 cDNA as probe, were shown to be unaffected by trehalose treatment. Elevation of cellular NADH dehydrogenase subunit 4 RNA in HT-29 cultures grown in medium containing different components (sodium butyrate, galactose, no-sugar, glucose, cellobiose) generally correlated with depressed growth levels and specifically with increased numbers of mucin-producing cells present. Like butyrate, the sugar, trehalose, is an effective inducer of HT-29 differentiation, and may prove useful as a dietary therapeutic, and as a probe for elucidating mitochondrial involvement in colonic cell differentiation and transformation.

Adenocarcinoma↗
PubMed1991

The diacylglycerol kinase inhibitor, R59949, potentiates secretion but not increased phosphorylation of a 47 kDalton protein in human platelets.

We have investigated the action of a novel inhibitor of DG-kinase, R59949. This agent was found to produce partial inhibition of formation of phosphatidic acid in human platelets challenged with thrombin, DC8 or OAG. However, this effect was not associated with enhanced phosphorylation of a 47 kDa protein, a known substrate for protein kinase C. We therefore believe that this compound does not represent a major advance on its earlier prototype, R59022.

Blood Platelets↗
PubMed1991

Reperfusion modification with a simplified blood cardioplegia system compared with oxygenated crystalloid cardioplegia.

A simplified system was developed for administration of blood cardioplegia with reperfusion modification. This system utilizes a single pass stainless steel coil to eliminate the need for a separate heat exchanger circuit. This system was compared with an oxygenated crystalloid cardioplegia system which was utilized in a manner which allowed warm blood perfusion of the heart for the last three minutes of the crossclamp interval. Both of these systems were compared with regard to mortality, spontaneous defibrillation, myocardial temperature, blood usage and peak CK-MB levels. In this series of patients, no significant advantage of either system could be identified.

Blood↗