Trihydroxytetraenes: A novel series of compounds formed from arachidonic acid in human leukocytes

Addition of 15L-hydroperoxy-5,8,11,13-eicosatetraenoic acid (15-HPETE) to human leukocytes led to the formation of a novel series of compounds containing four conjugated double bonds. The yield of tetraenes was increased approx. 100-fold when ionophore A23187 (5 μM) was added simultaneously with 15-HPETE. The structure of the major tetraene was established by physical methods as well as by chemical degradation and found to be 5,6,15L-trihydroxy-7,9,11,13-eicosatetraenoic acid.     

In vitro metabolism of N-methyl-dibenzo [c,g]carbazole a potent sarcomatogen devoid of hepatotoxic and hepatocarcinogenic properties

The metabolism of N-methyl substituted 7H-dibenzo[c,g]carbazole (N-Me DBC) was investigated in vitro using liver microsomes from 3-methylcholanthrene (MC)-, benzo[c]carbazole (BC) and Arochlor-pretreated mice and rats. N-Me DBC is a potent sarcomatogen devoid of hepatotoxicity and liver carcinogenic activity. The ethyl acetate-extractable metabolites were separated by high performance liquid chromatography (HPLC) and most of them were identified by proton magnetic resonance (PMR), mass spectrometry (MS) and comparison with synthetically prepared specimens. Mouse and rat microsomes gave rise to the same metabolites. The major metabolites were 5-OH-N-Me DBC (50%), N-hydroxymethyl (HMe) DBC (25-30%) and 3-OH-N-Me DBC (10%). Addition of 1,1,1-trichloropropene-2,3-oxide (TCPO) to the standard incubation medium permitted the identification of two dihydrodiols among the minor metabolites. No metabolite of DBC was observed after incubation of N-Me DBC, or its major metabolite N-HMe DBC, with either mouse or rat microsomes, but the possibility of a slight demethylation cannot be totally excluded. The lack of biotransformation at the nitrogen atom site may explain the lack of hepatotoxicity and liver carcinogenic activity of N-Me DBC. The modulation of metabolism by epoxide hydrolase, cytosol and glutathione was also investigated. The results are discussed in the light of data previously obtained with hepatotoxic and hepatocarcinogenic DBC.     

In vitro alkylation of calf thymus DNA by acrylonitrile. Isolation of cyanoethyl-adducts of guanine and thymine and carboxyethyl-adducts of adenine and cytosine

Reaction of the rodent carcinogen acrylonitrile (AN) at pH 5.0 and/or pH 7.0 for 10 and/or 40 days with 2'-deoxyadenosine (dAdo), 2'-deoxycytidine (dCyd), 2'-deoxyguanosine (dGuo), 2'-deoxyinosine (dIno), N6-methyl-2'-deoxyadenosine (N6-Me-dAdo) and thymidine (dThd) resulted in the formation of cyanoethyl and carboxyethyl adducts. Adducts were not detected after 4 h. The adducts isolated were 1-(2-carboxyethyl)-dAdo (1-CE-dAdo), N6-CE-dAdo, 3-CE-dCyd, 7-(2-cyanoethyl)-Gua (7-CNE-Gua), 7,9-bis-CNE-Gua, imidazole ring-opened 7,9-bis-CNE-Gua, 1-CNE-dIno, 1-CE-N6-Me-dAdo and 3-CNE-dThd. Structures were assigned on the basis of UV spectra and electron impact (EI), chemical ionization (CI), desorption chemical ionization (DCI) and Californium-252 fission fragment ionization mass spectra. Evidence is presented which strongly suggests that N6-CE-dAdo was formed by Dimroth rearrangement of 1-CE-dAdo during the reaction between AN and dAdo. The carboxyethyl adducts resulted from initial cyanoethylation (by Michael addition) at a ring nitrogen adjacent to an exocyclic nitrogen atom followed by rapid hydrolysis of the nitrile moiety to a carboxylic acid. It was postulated that the facile hydrolysis is an autocatalyzed reaction resulting from the formation of a cyclic intermediate between nitrile carbon and exocyclic nitrogen. AN was reacted with calf thymus DNA (pH 7.0, 37 degrees C, 40 days) and the relative amounts of adducts isolated were 1-CE-Ade (26%), N6-CE-Ade (8%), 3-CE-Cyt (1%), 7-CNE-Gua (26%), 7,9-bis-CNE-Gua (4%), imidazole ring-opened 7,9-bis-CNE-Gua (19%) and 3-CNE-Thy (16%). Thus a carcinogen once adducted to a base in DNA was shown to be subsequently modified resulting in a mixed pattern of cyanoethylated and carboxyethylated AN-DNA adducts. Three of the adducts (1-CE-Ade, N6-CE-Ade and 3-CE-Cyt) were identical to adducts previously reported by us to be formed following in vitro reaction of the carcinogen beta-propiolactone (BPL) and calf thymus DNA. The results demonstrate that AN can directly alkylate DNA in vitro at a physiological pH and temperature.     

狗在运动中的肌糖原消耗与无氧阈的关系

本实验测定了5条狗的无氧阈值,运动耐受时间、衰竭时的血乳酸浓度及运动中的肌糖原消耗量。结果如下:无氧阈值,1.与运动耐受时间呈正相关(r=0.947,P<0.02);2.与运动中肌糖原消耗量呈负相关(r=-0.959,P<0.01);3.与衰竭时的血乳酸浓度呈负相关(r=-0.942,P<0.02)。实验结果提示,无氧阈值是反映机体耐力的可靠指标。而运动中肌糖原消耗少,血乳酸积累程度轻,可能是无氧阈值之所以能够反映机体耐力的物质基础。     

Influences of medium consistency and shoot density on in vitro shoot proliferation of Populus alba × P. grandidentata

The in vitro shoot proliferation of Populus alba × P. grandidentata was affected by the medium consistency and shoot density, but not by three sizes of vessels. After 4 weeks of culture, the fresh weight and number of shoots per explant on liquid medium were significantly greater than those on agar-solidified medium. In particular, 3.2 shoots, 7 mm or longer per explant, were produced on liquid medium compared with 1.6 shoots per explant or agar-solidified medium. The fresh weight per explant after 4 weeks of culture on liquid medium and agar-solidified medium were 0.68 and 0.25 g, respectively. Increasing the number of shoots per vessel slowed the growth of the explants as measured by fresh weight and the number of shoots produced. There was little difference in the number of shoots produced between vessels with 1 or 2 shoots per vessel, but there were many fewer shoots produced when 3 shoots were placed in each vessel.Journal Paper No. J-11977 of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa. Project 2210.     

Multiple conformations of amino acid residues in ribonuclease A

The highly refined 1.26 A structure (R = 0.15) of phosphate-free bovine pancreatic ribonuclease A was modeled with 13 residues having discrete multiple conformations of side chains. These residues are widely distributed over the protein surface, but only one of them, Lys 61, is involved in crystal packing interactions. The discrete conformers have no unusual torsion angles, and their interactions with the solvent and with other atoms of the protein are similar to those residues modeled with a single conformation. For three of the residues--Val 43, Asp 83, and Arg 85--two correlated conformations are found. The observed multiple conformations on the protein surfaces will be of significance in analyzing structure-function relationships and in performing protein engineering.     

The rapid polyphosphoinositide metabolism may be a triggering event for thrombin-mediated stimulation of human platelets

The metabolism of polyphosphoinositides was examined in human platelets activated by thrombin. The addition of thrombin to [3H]glycerol-labeled platelets induced an initial loss and a subsequent increase of the radioactivity in phosphatidylinositol-4,5-bisphosphate (TPI) without any significant change in phosphatidylinositol-4-phosphate (DPI). A marked enhancement of [32P]Pi incorporation into TPI occurred in parallel with an increase in this lipid content, which was accompanied with a conccurent decrease in phosphatidylinositol (PI). The rate of this subsequent increase in TPI was smaller than that observed in [3H]arachidonic acid-labeled platelets, suggesting that formed TPI in activated platelets may contain much greater amount of arachidonate than preexisting TPI in resting platelets. These data indicate that thrombin causes a rapid change in TPI metabolism (initial degradation of preexisting TPI and subsequent production of arachidonate-rich TPI), which might be a primary candidate to modulate thrombin-induced function in human platelets.     

The modulating influence of the fluidity of cell membrane on excision repair of DNA in UV-irradiated Escherichia coli

When the extent of liquid holding recovery (LHR) was measured as a function of the temperature at the time of liquid holding and the Arrhenius plot was made, two distinctive phases for the LHR were demonstrated in UV-irradiated RecA- derivative of E. coli ole28E1, which are unable to synthesize and degrade unsaturated fatty acids. The inflection temperatures were 17-18 degrees C, 23-24 degrees C and 28-30 degrees C for linoleate-, oleate- and elaidate-grown cells, respectively. These temperatures well corresponded to the phase transition temperatures of the cell membrane supplemented with the fatty acid. It is therefore concluded that at least a component involved in in vivo excision repair in E. coli is associated with cell membrane.     

Streptozotocin diabetes results in increased responsiveness of adipocyte lipolysis to glucagon

Adipocytes from streptozotocin-diabetic rats are approximately 50-times more sensitive to the lipolytic action of glucagon. This change is only perceived in the presence of a small quantity of adenosine deaminase which itself has little effect on basal lipolysis. Insulin treatment restores glucagon sensitivity to normal.     

Selective generation of leukotriene B4 by tracheal epithelial cells from dogs

The incubation of suspensions of canine tracheal epithelial cells of greater than 95% purity with arachidonic acid (25-200 micrograms/ml) for 60-120 min resulted in the generation of a maximum of 36.2 +/- 9.1 picomoles of leukotriene B4/10(6) cells, less than 2.0 picomoles of leukotrienes C4, D4, and E4/10(6) cells, and 1030 +/- 463, 767 +/- 500, and 324 +/- 100 picomoles/10(6) cells of 15-, 12-, and 5-hydroxy-eicosatetraenoic acids, respectively (mean +/- SEM, n = 8). The identity of leukotriene B4 was established by chromatographic and spectral properties, by reactivity with mono-specific anti-plasma, and by the chemotactic activity for neutrophils. Thus, the epithelium may be an important source of mediators of inflammation and hypersensitivity of pulmonary airways.