A5000326101- Inflammatory mediators and NSAID effects
- Asthma and respiratory diseases
- Eicosanoids and Hypertension Pharmacology
- Estrogen and related hormone effects
- Immune Response and Inflammation
- Peroxisome Proliferator-Activated Receptors
- Bioactive Compounds and Antitumor Agents
- Cannabis and Cannabinoid Research
- DNA Repair Mechanisms
- Alcohol Consumption and Health Effects
- Pediatric health and respiratory diseases
- Receptor Mechanisms and Signaling
- Biochemical Analysis and Sensing Techniques
- Synthesis and Biological Evaluation
- DNA and Nucleic Acid Chemistry
- Immune Cell Function and Interaction
- Metabolomics and Mass Spectrometry Studies
- Click Chemistry and Applications
- Computational Drug Discovery Methods
- Respiratory and Cough-Related Research
- Lipid metabolism and biosynthesis
- Carcinogens and Genotoxicity Assessment
- Genomics, phytochemicals, and oxidative stress
- Synthesis and biological activity
- Tea Polyphenols and Effects
Vanderbilt University
2013-2024
Institute of Chemical Biology and Fundamental Medicine
2014
Scripps Research Institute
2008
Vanderbilt-Ingram Cancer Center
2007
Center of Molecular Immunology (Cuba)
2004
McDaniel College
1990-1998
Massachusetts Institute of Technology
1998
Wayne State University
1997
Science Applications International Corporation (United States)
1996
National Institutes of Health
1995
Malondialdehyde (MDA) is an endogenous genotoxic product of enzymatic and oxygen radical-induced lipid peroxidation whose adducts are known to exist in DNA isolated from healthy human beings. To evaluate the mutagenic potential MDA cells, we reacted with pSP189 shuttle vector then transfected them into fibroblasts for replication. induced up a 15-fold increase mutation frequency supF reporter gene compared untreated DNA. Sequence analysis revealed that majority MDA-induced mutations occurred...
A high glucose concentration in vivo or an increased of 6-phosphate vitro has been found to lead the glycosylation epsilon-amino groups lysine residues bovine and rat lens crystallins. In vitro, this imparts susceptibility crystallins sulfhydryl oxidation. Disulfide crosslinks result formation molecular weight aggregates opalescence crystallin solutions. The addition reducing agents prevents as well reverses These phenomena suggest a new interpretation previous results on cataract approach...
L-663,536 (3-[1-(4-chlorobenzyl)-3-t-butyl-thio-5-isopropylindol-2-yl]-2,2-dimethylpropanoic acid) is a potent inhibitor of leukotriene (LT) biosynthesis in intact human polymorphonuclear leukocytes (PMN) (IC 50 , 2.5 nM). Similarly, inhibited A23187-induced LTB 4 formation by rat peripheral blood and elicited PMN. At concentrations where inhibition occurred whole (1.1 μM), no effect was seen on cyclooxygenase or 12-lipoxygenase, an also observed washed platelets. The compound had porcine...
Challenge of human peripheral blood leukocytes with ionophore A23187 resulted in leukotriene (LT) synthesis, a decrease total cellular 5-lipoxygenase activity, and change the subcellular localization enzyme. In homogenates from control cells, greater than 90% activity protein was localized cytosol (100,000 X g supernatant). Ionophore challenge (2 microM) loss approximately 55% enzymatic 35% enzyme cytosol. Concomitantly, there an accumulation inactive membrane pellets) which accounted for at...
Recently, we have shown that ionophore activation of human leukocytes results in leukotriene synthesis and a translocation 5-lipoxygenase from the cytosol to cellular membrane. This membrane was postulated be an important early step for enzyme. 3-[1-(p-Chlorobenzyl)-5-(isopropyl)-3-tert-butylthioindol-2-yl]-2, 2- dimethylpropanoic acid (MK886) is potent specific inhibitor biosynthesis vivo intact cells, but has no direct effect on activity cell-free systems. In this report, show MK886 can...
The activity of leukotriene A4 (LTA4) synthase in crude human leukocyte homogenates was found to have a similar requirement for Ca2+ and ATP as had been noted previously 5-lipoxygenase activity. Purification the using ammonium sulfate fractionation, AcA 44 gel-filtration chromatography, HPLC on anion-exchange hydroxyapatite columns demonstrated that LTA4 copurified with recoveries increases specific Furthermore, two enzymatic activities coeluted exactly three different systems. Maximal...
Arachidonate 5-lipoxygenase was purified 400-fold from homogenates of human peripheral blood leukocytes by a combination ammonium sulfate fractionation, gel filtration chromatography, and HPLC on anion exchange hydroxylapatite columns. NaDodSO4/polyacrylamide electrophoresis the protein revealed presence single major band (apparent Mr, 80,000). Densitometric analysis Coomassie blue staining pattern gels that 90-97% purity had been achieved. As has reported for other mammalian sources,...
The enzyme 5-lipoxygenase (5-LO) catalyzes the first two reactions in pathway leading to formation of leukotrienes from arachidonic acid. Leukotrienes are potent acid metabolites possessing biological activities that suggest a role pathophysiology allergic and inflammatory diseases. To obtain structural information about 5-LO for use developing anti-inflammatory chemotherapeutic agents, was purified human polymorphonuclear leukocytes amino sequences were determined several cyanogen...
Mouse peritoneal macrophages that had ingested zymosan particles released a polar metabolite of arachidonic acid possessing slow-reacting substance activity in the guinea pig ileum assay. The was purified by solvent extraction, Sephadex G-25 column chromatography. absorbed light at 280 nm and contained free amino group. When were preincubated overnight with [3H]arachidonic acid, [35H]cysteine, or [14C]glutamic each radiolabel incorporated into compound. Direct analysis revealed glycine,...
Resident mouse peritoneal macrophages release the slow-reacting substance leukotriene C (LTC) on exposure to particulate IgE immune complexes. Because these cells lose their responsiveness an stimulus after 4 h in culture, maximum of 20:4 metabolites is observed before this time. However, a similar diminution metabolism was not with zymosan stimulus. Freshly explanted are deficient intracellular glutathione (GSH) (12.4 +/- 0.4 pmol/micrograms cell protein), but GSH increases steady state...
Mouse resident pulmonary macrophages were subdivided into alveolar (PAM) and interstitial (PTM) populations on the basis of accessibility to lavage, zymosan-induced arachidonic acid (20:4) metabolism was examined in both labeled with [3H]20:4. Maximal phagocytic doses unopsonized zymosan induced specific release 11% phospholipid 20:4 by PTM 4.6% PAM. Direct fatty analysis [3H]20:4-labeled cultured presence or absence indicated that activity [3H]20:4 cell provided an accurate measure released...
The ingestion of particles by macrophages leads to the prompt induction prostaglandin (PG) synthesis. We have now dissected endocytic process and examined requirements E (PGE) synthesis for particle attachment, membrane interiorization, phagosome-lysosome fusion. Macrophages that were loaded with polyanion dextran sulfate exhibited a greater than 99% inhibition fusion produced normal amounts PGE upon challenge zymosan. Inhibition interiorization cytochalasin D was similarly ineffective in...
Ibuprofen and mefenamic acid are weak, competitive inhibitors of cyclooxygenase-2 (COX-2) oxygenation arachidonic (AA) but potent, noncompetitive 2-arachidonoylglycerol (2-AG) oxygenation. The slow, tight-binding inhibitor, indomethacin, is a potent inhibitor 2-AG AA whereas the rapidly reversible 2'-des-methylindomethacin, poor These observations consistent with model in which bind one subunit COX-2 inhibit binding other homodimeric protein. In contrast, ibuprofen mefenamate must both...
Oxidation of endogenous macromolecules can generate electrophiles capable forming mutagenic adducts in DNA. The lipid peroxidation product malondialdehyde, for example, reacts with DNA to form M 1 G, the pyrimidopurinone adduct deoxyguanosine. In addition free radical attack lipids, is also continuously subjected oxidative damage. Among products damage are base propenals. We hypothesized that these structural analogs malondialdehyde would react G. Consistent this hypothesis, we detected a...
Sulfhydryl reagents such as dithiothreitol stabilized human leukocyte 5‐lipoxygenase (5‐LO) during purification. During enzyme assay, however, these led to irreproducible or unexpectedly low activity. This inconsistency in the assay was eliminated by inclusion of hydroperoxyeicosatetraenoic acids (1–5 μM) reaction which effected a 10–20‐fold stimulation 5‐LO Structural studies indicated that an intact hydroperoxy function, and long‐chain fatty acyl moiety were required for stimulation. These...
A method for the radiochemical assay of LTC production by mouse peritoneal macrophages in vitro is presented. The involves labeling culture with [5,6,8,9,11,12,14,15-3H]20:4 followed stimulation arachidonic acid (20:4) release under experimental conditions desired. Radiolabeled leukotriene C (LTC) recovered from medium extraction and silicic chromatography 40% yield full retention biological activity. Because this radiochemically pure, quantity may be estimated amount radioactivity sample....