ATP and phosphate reciprocally affect subunit association of human recombinant High Km 5'-nucleotidase. Role for the C-terminal polyglutamic acid tract in subunit association and catalytic activity.

IMP-specific, High Km 5'-nucleotidase (EC 3.1.3.

Generation of lipid free radicals by adherent leukocytes from transplanted rat liver.

The production of free radicals in blood correlates with primary nonfunction of transplanted livers, but the source of the free radicals is unknown. The purpose of this study was to determine if adherent leukocytes in the transplanted liver are responsible for the radicals detected in blood. First, a new method to harvest adherent leukocytes from the liver without enzymatic digestion was developed and characterized by transplanting livers from ethanol-treated rats, which increases primary nonfunction, and from saline-treated controls.

Estrogen increases sensitivity of hepatic Kupffer cells to endotoxin.

The relationship among gender, lipopolysaccharide (LPS), and liver disease is complex. Accordingly, the effect of estrogen on activation of Kupffer cells by endotoxin was studied. All rats given estrogen intraperitoneally 24 h before an injection of a sublethal dose of LPS (5 mg/kg) died within 24 h, whereas none of the control rats died.

Kupffer cells contain a glycine-gated chloride channel.

Here the effect of glycine on intracellular Ca2+ concentration ([Ca2+]i) in cultured Kupffer cells stimulated with lipopolysaccharide (LPS) was investigated to assess the possibility that they contain a glycine-gated chloride channel. LPS (10 micrograms/ml) increased [Ca2+]i rapidly, with peak values reaching 307 +/- 29 nM. Glycine (1 mM) prevented this increase nearly completely.

Female rats exhibit greater susceptibility to early alcohol-induced liver injury than males.

It is known that women develop hepatic injury more rapidly and with exposure to less ethanol than men; however, mechanisms remain unclear. The purpose of this study was to determine if an enteral alcohol delivery model could be used to study susceptibility of females to alcohol-induced liver injury. Male and female Wistar rats (age- or weight-matched) were given ethanol (11-12 g.

Role of Kupffer cells, endotoxin and free radicals in hepatotoxicity due to prolonged alcohol consumption: studies in female and male rats.

Alcohol ingestion results in increases in the release of endotoxin from gut bacteria or membrane permeability of the gut to endotoxin, or both. Female rats are more sensitive to these changes. Elevated levels of endotoxin activate Kupffer cells to release substances such as eicosanoids, tumor necrosis factor-alpha and free radicals.

Ras activation in platelets after stimulation of the thrombin receptor, thromboxane A2 receptor or protein kinase C.

Several reports have indicated that the small G-protein Ras is not present immunologically in platelets. However, here we report the identification of Ras in platelets by immunoprecipitation with the Ras-specific monoclonal antibodies Y13-259 or Y13-238, followed by Western blotting. The presence of Ras was not due to contamination of samples with erythrocytes or leucocytes.

Activation and inhibition of reconstituted cardiac L-type calcium channels by palmitoyl-L-carnitine.

We have studied the effect of a palmitoyl-L-carnitine (L-PC) on single cardiac L-type Ca channels incorporated from porcine ventricular sarcolemma into planar lipid bilayers where we could control the concentration, intracellular and/or extracellular location, and duration of L-PC treatment. We found that 1.0 microM L-PC in either the intracellular or extracellular chamber caused an approximately 8 fold increase in channel open probability when measured within the first minute after L-PC addition.

Nimodipine, a dihydropyridine-type calcium channel blocker, prevents alcoholic hepatitis caused by chronic intragastric ethanol exposure in the rat.

It has been shown recently that inactivation of Kupffer cells prevents free radical formation and early alcohol-induced liver injury, and that hypoxia subsequent to a hypermetabolic state caused by activated Kupffer cells is likely involved in the mechanism. Calcium is essential for the activation of Kupffer cells, which contain L-type voltage-dependent Ca2+ channels. Therefore, the purpose of this study was to determine whether a Ca2+ channel blocker, nimodipine, prevents early alcohol-induced liver injury in vivo and to evaluate its effect on intracellular calcium ([Ca2+]i) in Kupffer cells in vitro.

Bacterial artificial chromosome cloning and mapping of a 630-kb human extrachromosomal structure.

We have cloned and mapped a circular 630-kb human extrachromosomal structure (termed amplisome) using the bacterial artificial chromosome (BAC) cloning system. Twenty-one BACs were isolated from an amplisome-enriched library by colony hybridization. The insert sizes range from 25 to 143 kb, with an average size of 82 kb.

Glycoprotein IV-independent adhesion of sickle red blood cells to immobilized thrombospondin under flow conditions.

The abnormal adherence of red blood cells (RBC to the blood vessel wall is believed to contribute to the vascular occlusion observed in patients with sickle call anemia. The cell adhesion receptors GPIV (CD36) and integrin alpha 4 beta 1 (CD49d/CD29) were previously identified on circulating sickle reticulocytes, and shown to mediate sickle RBC adhesion to the endothelium. The presence of damaged endothelium in these patients suggests that exposed extracellular matrix proteins could provide a potential substrate for sickle RBC adhesion.

Localization of a putative second membrane association site in penicillin-binding protein 1B of Escherichia coli.

We have shown previously that the periplasmic domain of penicillin-binding protein 1B (PBP 1Bper; residues 90-844) from Escherichia coli is insoluble in the absence of detergents, and can be reconstituted into liposomes [Nicholas, Lamson and Schultz (1993) J. Biol. Chem.

Kupffer cell prostaglandin E2 stimulates parenchymal cell O2 consumption: alcohol and cell-cell communication.

Several studies have demonstrated that ethanol can increase hepatic O2 uptake (e.g., produce a hypermetabolic state); however, a complete explanation of this important phenomenon remains unclear.

Paradoxical increase in peroxisomal cyanide-insensitive respiration following dietary exposure to WY-14,643 in the perfused liver.

WY-14,643, a lipid-lowering drug, increases basal rates of oxygen uptake in perfused livers. Because peroxisomes consume oxygen for H2O2 production and are induced by WY-14,643 treatment, it is possible that peroxisomal beta-oxidation can account for some of this increase in cellular respiration. Therefore, cyanide, an inhibitor of mitochondrial cytochrome oxidase, was infused into livers of WY-14,643-fed rats (0.

Glycine minimizes reperfusion injury in a low-flow, reflow liver perfusion model in the rat.

This study investigated the effects of glycine on reperfusion injury in a low-flow, reflow liver perfusion model. With this protocol, livers were perfused at low flow rates of approximately 1 ml.g-1.

Phorbol ester enhances integrin alpha IIb beta 3-dependent adhesion of human erythroleukemic cells to activation-dependent monoclonal antibodies.

Following platelet stimulation by agonists, integrin-alpha IIb beta 3 (or glycoprotein IIb-IIIa) is converted to an activated state that can bind soluble fibrinogen and mediate platelet aggregation. However, little is known about modulation of alpha IIb beta 3 in cell lines. In the present study, we show that agonist stimulation modulates alpha IIb beta 3-dependent adhesive properties of a human erythroleukemic (HEL) cell line.

Peroxisome proliferators activate Kupffer cells in vivo.

The mechanism by which peroxisome proliferators increase cell replication and cause liver tumors in rodents remains unknown. When activated, Kupffer cells, the resident hepatic macrophages, release a variety of mitogenic stimuli that could theoretically increase cell proliferation in nearby hepatocytes. Therefore, in the present study we evaluated the effect of two potent peroxisome proliferators, nafenopin and WY-14,643, on Kupffer cell activation in vivo.

A fish oil diet minimizes hepatic reperfusion injury in the low-flow, reflow liver perfusion model.

In this study, the effects of fish oil treatment on hepatic reperfusion injury in a low-flow, reflow perfusion model were investigated. Rats were fed powdered diets containing either 5% corn or 5% encapsulated fish oil for 13 to 15 days. Average daily food intake in both control and fish oil groups was about 20 g per rat, and weight gain averaged 9 g/rat/d.

Food restriction stimulates conjugation of p-nitrophenol in perfused rat liver.

Rates of conjugation of p-nitrophenol were studied in livers from normal and food-restricted rats perfused with either p-nitroanisole or p-nitrophenol. Female Sprague-Dawley rats had ad libitum access to a Purina 5001 nonpurified diet (control) or were given 65% of the intake of controls for 3 weeks. Livers were perfused with oxygenated Krebs-Henseleit buffer using a nonrecirculating system.

Radiolabeling of methylphosphonate and phosphorothioate oligonucleotides and evaluation of their transport in everted rat jejunum sacs.

Purpose: The therapeutic use of antisense oligonucleotides will likely involve their administration over protracted periods of time. The oral route of drug dosing offers many advantages over other possible routes when chronic drug administration is necessary. However, little is known about the potential for oligonucleotide uptake from the gastrointestinal tract.

Single-channel currents from diethylpyrocarbonate-modified NMDA receptors in cultured rat brain cortical neurons.

The role of histidine residues in the function of N-methyl-D-aspartate (NMDA)-activated channels was tested with the histidine-modifying reagent diethylpyrocarbonate (DEP) applied to cells and membrane patches from rat brain cortical neurons in culture. Channels in excised outside-out patches that were treated with 3 mM DEP for 15-30 s (pH 6.5) showed an average 3.