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Biomedical subjects

R L Jackson

Publications and source records attributed to R L Jackson.

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

Antioxidant activity of probucol and its analogues in hypercholesterolemic Watanabe rabbits.

Probucol (1) and probucol analogues with the substitutions at the disulfide-linked carbon (2, 3) and an additional substitution at a tert-butyl of each phenolic ring (4) were tested for their ability to lower total serum cholesterol and prevent aortic atherosclerosis in modified Watanabe heritable hyperlipidemic (WHHL) rabbits and to inhibit Cu2(+)-induced lipid peroxidation of isolated plasma low-density lipoproteins (LDL). After 84 days of feeding 1% of each compound in rabbit chow, probucol was effective in lowering serum cholesterol, whereas 2-4 were not. The concentration of drug in serum and LDL was 2 greater than 1 greater than 3 greater than 4. Probucol and analogues prevented Cu2(+)-induced oxidation of LDL in vitro to an extent that directly related to their concentrations in LDL. The decrease in lipid oxidation was directly correlated with the inhibition of both oxidized-LDL-induced cholesteryl ester synthesis in cultured macrophages and to the inhibition of aortic atherosclerosis in vivo. These results show that the antioxidant activity of probucol and analogues is directly related to their concentration in LDL, which may explain their pharmacological activity in reducing atherosclerosis.

Animals↗
PubMed1991

Lipid hydroperoxide involvement in copper-dependent and independent oxidation of low density lipoproteins.

The involvement of lipid hydroperoxides in the oxidation of human low density lipoproteins (LDL) induced by Cu++ or azoperoxyl radicals was studied. Concurrent or pretreatment of LDL with the synthetic peroxidase Ebselen and its cofactor glutathione totally prevented Cu+(+)-dependent oxidation, as measured by the formation of thiobarbituric acid reactive substances and LDL mobility on agarose gels. LDL could be oxidized in the absence of metals with peroxy radicals generated via homolysis of azo compounds. With the azo initiator, 2,2'-azobis-2-amidinopropane hydrochloride, Ebselen plus glutathione prevented lipid oxidation when present during oxidation. However, if LDL were first pretreated with Ebselen plus glutathione and these reagents removed before oxidation, they did not prevent oxidation. Analysis by high-performance liquid chromatography of lipids extracted from Cu+(+)- or azoperoxyl radical-modified lipoproteins indicated that the major lipid oxidation products were derived from linoleate and that Ebselen-glutathione addition reduced linoleate hydroperoxides. Oxidation of LDL by human monocyte-derived macrophages was also prevented by coincubation with Ebselen-glutathione or the antioxidant, probucol. These results demonstrate that initiation of Cu+(+)-dependent oxidation absolutely requires the presence of trace amounts of lipid hydroperoxides in the lipoprotein and that hydroperoxides play a major role in the propagation reactions even in the absence of added transition metals.

Azo Compounds↗
PubMed1990

Differential regulation of hepatic triglyceride lipase and 3-hydroxy-3-methylglutaryl-CoA reductase gene expression in a human hepatoma cell line, HepG2.

The level of hepatic triglyceride lipase (H-TGL) synthesis and secretion was examined in response to changes in cholesterol biosynthesis in the human hepatoma cell line HepG2. Cells were first fed a lipoprotein-deficient serum-supplemented medium to eliminate exogenous cholesterol. Mevinolin, a 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase inhibitor, was then added at a concentration (37 microM) which inhibited cholesterol biosynthesis by greater than 85% and decreased total cell cholesterol from 36.1 to 27.4 micrograms/ml of cell protein. Mevinolin treatment caused a 4.9 +/- 0.8-fold increase in the amount of H-TGL activity secreted into the medium, a 1.8 +/- 0.4-fold rise in H-TGL-specific mRNA, and a concurrent 14-fold increase in HMG-CoA reductase mRNA. Addition of 1 mM mevalonic acid to normal or mevinolin-treated cells raised the cellular cholesterol content and decreased the amount of secreted H-TGL activity to levels below control values. Mevalonic acid also prevented mevinolin-induction of H-TGL and HMG-CoA reductase mRNA, suggesting a common regulatory step for H-TGL and HMG-CoA reductase. Exposure of cells to mevinolin and 25-hydroxycholesterol together resulted in a marked repression of HMG-CoA reductase mRNA levels, whereas these conditions further enhanced the secretion of H-TGL activity and the expression of H-TGL mRNA. These results demonstrate a differential role for 25-hydroxycholesterol in the regulation of H-TGL and HMG-CoA reductase expression.

Carcinoma, Hepatocellular↗
PubMed1990

Characterization of a monoclonal antibody specific to the amino terminus of the alpha-chain of human fibrin.

A peptide, Gly-Pro-Arg-Val-Val-Glu, corresponding to the first six residues of the amino terminus of the alpha-chain of human fibrin (desAA-fibrin) was prepared by solid-phase peptide synthesis. The peptide was covalently linked to keyhole-limpet hemocyanin (KLH) and used as an immunogen for preparing monoclonal antibodies. A monoclonal antibody specific to the hexapeptide, but not to KLH or fibrinogen, was produced. The antibody did not bind to thrombin-mediated clots prepared from either plasma or purified fibrinogen. However, immunoreactivity was detected when fibrin (prepared from fibrinogen) was solubilized with 8 M urea. In contrast, a monoclonal antibody specific to the amino terminus (Gly-His-Arg-Pro-Leu-Asp-Lys) of the beta-chain of fibrin recognized the epitope in clots. These results indicate that thrombin cleavage of fibrinogen produces a structural change in the amino terminal domain of the alpha-chain that makes it inaccessible to antibody interaction. In addition, our study suggests that the potential clinical application of monoclonal antibodies to localize fibrin-rich thrombi must take into account the final structure of clots.

Amino Acid Sequence↗
PubMed1990

Ex vivo lipopolysaccharide-induced interleukin-1 secretion from murine peritoneal macrophages inhibited by probucol, a hypocholesterolemic agent with antioxidant properties.

Probucol, 4,4'-(isopropylidenedithio)bis(2,6-di-tert-butyl-phenol), has been shown to inhibit atherogenesis in genetically hypercholesterolemic (Watanabe) rabbits. Since atherosclerotic lesions contain macrophages capable of screting interleukin 1 (IL 1) and other cytokines that could contribute to the pathogenesis of the disease, we have investigated whether probucol affects IL 1 secretion. Resident peritoneal macrophages from mice dosed with probucol secreted 40-80% less IL 1 than macrophages from control animals when stimulated in vitro with lipopolysaccharide (LPS). The inhibitory effect of probucol was observed when IL 1 was assayed by the standard bioassay, the thymocyte proliferation assay, or a competitive IL 1 receptor binding assay. Probucol treatment had no effect on LPS-induced membrane IL 1 expression; secretion of tumor necrosis factor (TNF); Con A-induced splenic interleukin 2 (IL 2) and interleukin 3 (IL 3) release; and prostaglandin- or zymosan-induced secretion of prostacyclin, leukotriene C4, acid phosphatase, or superoxide anion. In contrast to the effect of oral administration, direct addition of probucol to macrophage cultures did not inhibit IL 1 release. Probucol administration did, however, inhibit the fall in serum zinc level induced by intravenous injection of LPS in zymosan-primed mice but had no effect on the LPS-induced increase in serum triglyceride levels, which indirectly confirms that probucol administration inhibits IL 1 but not TNF secretion. Paw granuloma induced in mice by heat-killed mycobacteria was inhibited by oral administration of probucol, an effect that may be attributable to inhibition of IL 1 secretion. Probucol neither reduced zymosan-induced liver granulomata in mice nor inhibited adjuvant-induced arthritis in rats. We suggest that inhibition of IL 1 secretion from macrophages by probucol contributes to its therapeutic effects in atherosclerosis and may also result in beneficial activity in some chronic inflammatory diseases.

Administration, Oral↗
PubMed1990

Probucol does not alter acetylated low density lipoprotein uptake by murine peritoneal macrophages.

It has been suggested that the anti-atherogenic effect of probucol (MDL 11,309) in familial hypercholesterolemic rabbits may be due in part to the inhibition of the uptake of modified low density lipoproteins by macrophages in the arterial wall. To test this hypothesis, mice were treated with dietary probucol (0.25%) for fourteen days, peritoneal macrophages were isolated and the uptake of acetylated low density lipoprotein (ALDL) was studied. In addition, peritoneal macrophages from control animals were treated with probucol (200 micrograms/ml) for 24 h in vitro prior to the ALDL uptake assay. The assay involved a fluorescent ALDL probe (dioctadecyltetramethylindocarbocyanine perchlorate-labeled ALDL), and measurement of uptake with a flow cytometer. No differences in ALDL uptake were detected between the control macrophages and macrophages treated with probucol in vitro or macrophages taken from probucol-treated mice.

Acylation↗
PubMed1990

Relationship between post-heparin plasma lipases, triglycerides and high density lipoproteins in normal subjects.

The relationship between plasma lipids and lipoproteins and the lipolytic activities of post-heparin plasma lipoprotein lipase (LpL) and hepatic-triglyceride lipase (H-TGL) was examined in normal subjects. Seven males and six females were given a high fat diet [15% carbohydrate (CARB), 65% fat, 20% protein] for 2 weeks followed by 4 weeks of a high CARB diet (65% CARB, 15% fat, 20% protein). Changes in plasma triglyceride concentrations associated with diet were negatively correlated with changes in HDL-C (r = -0.533, P less than 0.001) and the HDL subfraction HDL2b (r = -0.308, P less than 0.001). The activity of LpL in post-heparin plasma was positively correlated with changes in plasma HDL-C (r = 0.668, P less than 0.001) and HDL2b (r = 0.457, P less than 0.001), and negatively with plasma triglycerides (r = -0.546, P less than 0.001). Changes in H-TGL activity were negatively correlated with changes in HDL2b (r = -231, P less than 0.05) and positively correlated with HDL-C (r = 0.326, P less than 0.01). These results in normal subjects provide further evidence that LpL and H-TGL are important enzymes in the metabolism of plasma lipoproteins and that changes in their activities contribute to plasma lipid and lipoprotein concentrations.

Adult↗
PubMed1989

Heparin induces the expression of hepatic triglyceride lipase in a human hepatoma (HepG2) cell line.

Hepatic triglyceride lipase (H-TGL) is a key lipolytic enzyme in the metabolism of human plasma high density lipoproteins. The enzyme is bound to glycosaminoglycans on endothelial cells in the liver and is immediately released into the circulation after heparin administration. In addition to releasing H-TGL, heparin-like glycosaminoglycans have also been shown to suppress hepatocyte proliferation and to alter tissue-specific gene expression. In the present study, the effects of heparin exposure on the secretion of H-TGL were examined in a human parenchymal hepatoma (HepG2) cell line. The addition of heparin to serum-supplemented medium induced the secretion of H-TGL in a time- and concentration-dependent manner. At 5.4 micrograms/ml heparin, H-TGL levels, as determined by triacyglycerol hydrolase activity, increased 7-fold after a 44-h incubation. Heparin exposure decreased intracellular H-TGL activity from 21.3 to 4.8 nmol of oleic acids released/h/10(8) cells and increased enzyme activity in the medium from 16.2 to 165.3 nmol of oleic acids released/h/10(8) cells. The heparin-induced secretion of H-TGL was associated with increased levels of H-TGL-specific mRNA. The addition of actinomycin D or cycloheximide reversed the heparin-induced increase in H-TGL activity and mRNA. Heparin treatment did not increase the level of actin mRNA suggesting that elevated H-TGL mRNA is due to enhanced tissue-specific expression of H-TGL. Expression of apolipoprotein E, another protein involved in lipoprotein metabolism, also showed induced levels of mRNA by heparin but to a lesser extent than that for H-TGL. We conclude that heparin stimulates the de novo synthesis of H-TGL in liver parenchymal cells in vivo by influencing both transcriptional and post-transcriptional events.

Carcinoma, Hepatocellular↗
PubMed1989

Human hepatoma (HepG2) cells secrete a single 65 K dalton triglyceride lipase immunologically identical to postheparin plasma hepatic lipase.

A triacylglycerol lipase was isolated from the culture medium of HepG2 human hepatoma cells and its properties were compared to hepatic triglyceride lipase (H-TGL) from human postheparin plasma. The HepG2 cell enzyme bound to heparin-Sepharose, was eluted with 1 M NaCl and was not inhibited by 1 M salt. Western-blotting of the fractions from the heparin-Sepharose column with a monoclonal antibody prepared against postheparin plasma H-TGL and which binds to an epitope in the carboxyl-terminus of H-TGL gave a single immunoreactive protein band of 65 kDa. This finding of immunochemical identity was confirmed with polyclonal antibodies prepared against synthetic peptides of H-TGL corresponding to amino acid residues 82-94 near the amino-terminus and residues 468-477, the carboxyl-terminus of the enzyme. We conclude that HepG2 cells secrete a single triacylglycerol lipase with molecular weight properties and immunological characteristics identical to post-heparin plasma H-TGL.

Antibodies, Monoclonal↗
PubMed1989

Dependence on heparin chain-length of the interaction of heparin with human plasma low density lipoproteins.

Successive rechromatography of commercial bovine lung heparin on human plasma low density lipoproteins (LDL) immobilized to AffiGel-10 yielded four high reactive heparin (HRH-I to IV) fractions and an unreactive fraction (URH). HRH-I was the most sulphated HRH fraction whereas URH had the least sulphation. In the presence of 10 mM Ca2+, LDL were precipitated by these heparins in the following order: HRH-II greater than HRH-III greater than HRH-IV greater than HRH-I greater than URH. The average molecular weight of HRH-I to IV was 8600, 11400, 10,100, and 10,000, respectively. A plot of log molecular weight versus the concentration of HRH required to give half-maximal precipitation of LDL showed a negative correlation (r = -0.880). These results indicate that heparin chain length is an important determinant of heparin binding to LDL in solution and may have relevance to the binding and precipitation of LDL in the arterial wall.

Animals↗
PubMed1989

Biochemical pharmacology of zidovudine in human T-lymphoblastoid cells (CEM).

HIV is the causative agent of AIDS. The purpose of this study was to examine the biochemical pharmacology of the anti-viral agent zidovudine (AZT) in the T-cell origin line (CEM). We have shown that zidovudine is activated by thymidine kinase (TK) in CEM cells to the triphosphate anabolite, which is incorporated into DNA. One microM zidovudine is sufficient for saturation of activation by TK and also of zidovudine monophosphate, by thymidylate kinase, to the diphosphate. Zidovudine triphosphate peaked 4 h after initiation of drug administration in CEM cells and then declined biexponentially. Nucleoside triphosphate (NTP) cellular concentrations declined rapidly in the cells after exposure to zidovudine. Concomitantly phosphorylation of zidovudine to zidovudine monophosphate and zidovudine monophosphate to zidovudine diphosphate declined in a similar manner in the CEM cells. The amount of zidovudine anabolite incorporated into purified DNA peaked 1 h after zidovudine treatment and declined thereafter with first order elimination kinetics. These studies elucidate the cellular activation of zidovudine in a T-cell line, CEM, and enhance our understanding of this important anti-HIV drug.

Cell Division↗
PubMed1989

Interaction of glycosaminoglycans with lipoproteins.

Apolipoproteins B-100 and E are protein constituents of human plasma chylomicrons, very low (VLDL), and low density lipoproteins (LDL). The interaction of lipoproteins with cell receptors is mediated by apoB and E. Lipoproteins also bind to the extracellular matrix, such as glycosaminoglycans (GAG), forming insoluble complexes in the presence of Ca2+. The purpose of this study was to identify the GAG-binding domains in apoB and E. By a combination of fragmentation of the intact proteins, peptide synthesis and quantitative GAG-binding, domains in apoB and apoE were identified and are shown below. These domains contain clusters of basic amino acids that we suggest are required for GAG-binding. table; see text.

Amino Acid Sequence↗
PubMed1989

Concentration-dependent antioxidant activity of probucol in low density lipoproteins in vitro: probucol degradation precedes lipoprotein oxidation.

The ability of probucol, a lipid-lowering drug with antioxidant properties, to prevent the Cu2+-induced oxidation of human plasma low density lipoproteins (LDL) was examined as a function of the concentration of probucol in LDL. In the absence of probucol, 3 microM Cu2+ induced half-maximal LDL lipid oxidation, as determined by the formation of thiobarbituric acid reactive substances (TBARS). Oxidation was associated with a loss of apolipoprotein B-100 and the appearance of higher molecular weight forms of the protein. In the presence of 0.6 mol% probucol (relative to phospholipid) and with 3 microM Cu2+, the time required to obtain half-maximal LDL lipid oxidation increased from 130 to 270 min and was explained by an increase in the lag time prior to LDL lipid oxidation. Once rapid oxidation of LDL had begun, the rate of TBARS formation was similar to that for LDL containing no probucol. At a probucol concentration of 4.2 mol%, the antioxidant prevented the oxidation of LDL-lipids. The delay in Cu2+-induced LDL oxidation with probucol corresponded to the time required for free radical-mediated processes to convert probucol to a spiroquinone and a diphenoquinone. These in vitro findings suggest that the potent antioxidant property of probucol is directly related to the amount of drug in the LDL particle and may have relevance to its antiatherosclerotic effects observed in vivo.

Apolipoprotein B-100↗
PubMed1989

Indomethacin in the treatment of lithium-induced nephrogenic diabetes insipidus.

Nephrogenic diabetes insipidus (NDI) is a frequent complication in patients receiving long-term lithium therapy. Both thiazide diuretics and amiloride may reduce the polyuria, but the use of each is associated with problems. We report the results of a clinical trial using the nonsteroidal anti-inflammatory drug indomethacin to treat a patient with well-documented lithium-induced NDI that persisted following cessation of lithium treatment. The administration of a single dose of indomethacin resulted in a dramatic decrease in urine volume and increase in urine osmolality that persisted for several hours, and was independent of renal hemodynamic changes. Subsequently, the patient experienced a sustained, favorable effect on her polyuria during long-term (3 months) indomethacin therapy without a deleterious effect on her renal function. Indomethacin may be a useful therapeutic tool for the amelioration of lithium-induced NDI.

Adult↗