Microcirculatory effects of leukotrienes C4, D4, and E4 in the guinea pig.
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Animals↗
Biomedical subjects
S Hammarström
Publications and source records attributed to S Hammarström.
Transformation of leukotriene A4 methyl ester to leukotriene C4 monomethyl ester by cytosolic rat glutathione transferases.
Six major basic cytosolic glutathione transferases from rat liver catalyzed the conversion of leukotriene A4 methyl ester to the corresponding leukotriene C4 monomethyl ester. Glutathione transferase 4-4, the most active among these enzymes, had a Vmax of 615 nmol X min-1 X mg protein-1 at 30 degrees C in the presence of 5 mM glutathione. It was followed in efficiency by transferase 3-4 which had a Vmax of 160 nmol X min-1 X mg-1 under the same conditions. Transferases 1-1, 1-2, 2-2 and 3-3 had at least 30 times lower Vmax values than transferase 4-4.
Animals↗
Inhibition of proliferative and cytotoxic activities of human T lymphocytes with rabbit antibodies directed against leucoagglutinin-reactive T cell surface components.
Three rabbit antisera (870, 872 and 873) were raised against leucoagglutinin-reactive components from the surface of human T cells. The antibodies reacted with two major glycoproteins of 175 kDa and 105 kDa. None of the antibodies triggered peripheral blood lymphocytes or T cells to proliferation when tested under different culture conditions. All antibodies inhibited the proliferative response to concanavalin A or to allogeneic lymphocytes in mixed lymphocyte culture when whole IgG fractions were used. Complete inhibition of cytotoxic activity was obtained in cell-mediated lympholysis and in natural killer cell cytotoxicity (NK) when fresh peripheral blood lymphocytes were used as effector cells. Weak inhibition was also obtained in NK when mixed lymphocyte culture-activated effector cells were used. The inhibition was stronger, when NK activity was determined against MOLT4 target cells as compared to K562. Whereas F(ab')2 fragments of 873 IgG inhibited cytotoxic T lymphocyte activity completely, Fab fragments of 873 IgG neither inhibited proliferation nor cytotoxic T lymphocyte activity, but gave some inhibition of NK against MOLT4 targets. The results indicate that antibodies against these leucoagglutinin-reactive structures reacted with polypeptides similar to or identical with the human "leukocyte function-associated antigen-1" (LFA-1) considered to be an important mediator of cell-cell interactions and nonspecific adherence.
Agglutinins↗
Action and metabolism of circulating leukotriene C.
The microvascular permeability effects of intravenously administered leukotriene C4 were assessed by fluorometric determinations of extravasated Evans blue dye in various tissues of guinea pigs. The effects were widespread, with intensity differences between different vascular beds, which in part reflected the tissue distribution of tritium-labeled leukotriene C3 in mice. Local tissue specificity favored permeability effects in the urinary tract, pulmonary airways, esophagus, and distal colon. The permeability effects may contribute to the hypotensive actions of leukotriene C4 and enhance airway obstruction by edema formation in addition to smooth muscle contraction.
Animals↗
Fractionation of human lymphocytes on Helix pomatia a hemagglutinin-sepharose and wheat germ agglutinin-sepharose.
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Cell Separation↗
The stimulation of the initiation of DNA synthesis and cell division in Swiss mouse 3T3 cells by prostaglandin F2 alpha requires specific functional groups in the molecule.
Among a number of prostaglandins, PGF2 alpha had the highest specific activity for stimulating the initiation of DNA synthesis in confluent resting Swiss 3T3 cells. At a saturating concentration of 8.5 X 10(-7) M, PGF2 alpha stimulated 21% of the cells to incorporate [ methyl-3H ]thymidine within 28 h. To elicit similar effects, prostaglandins F1 alpha, E1, E2, and D2 were required in 10-fold higher concentrations. Prostaglandins A1, A2, B1, and prostacyclin had no mitogenic activity. Insulin at 10(-8) M enhanced the stimulatory effect of PGF2 alpha and also of prostaglandins F1 alpha, E1, E2, and D2 by increasing the fraction of labeled nuclei. Methyl derivatives of PGF2 alpha were as effective as PGF2 alpha. Epimerization of the hydroxyl group at C-9 abolished the activity of the molecule. In contrast, upon epimerization at C-11 and C-15, some mitogenic activity was retained. In the presence of insulin, the latter molecules were as active as PGF2 alpha. Oxidation of the hydroxyl group at C-15 to a ketone abolished the mitogenic effect, while methyl ether formation led to only a slight loss of activity. Reduction of the delta 13 double bond also led only to a small reduction of activity. Similar differences in the activity of the various prostaglandins and analogues of PGF2 alpha were observed for 2-deoxyglucose uptake and increases in cell number. The relationships between structure and activity of prostaglandins suggest the existence of some specific receptor for PGF2 alpha to confer mitogenic response.
Animals↗
Characteristics of the uptake of cysteine-containing leukotrienes by isolated hepatocytes.
Leukotrienes were transported into rat hepatocytes by a temperature- and energy-dependent mechanism. The uptake was saturable with high- and low-affinity sites (Km values approx. 1 and 17 microM). Competition and kinetic experiments indicated that leukotrienes C4, D4 and E4 were transported by a common mechanism. The maximal velocity of transport was about 50% higher for leukotrienes D4 and E4 than for leukotriene C4. Leukotriene B4, glutathione disulfide, and the glutathione-S-conjugate of acetaminophen did not interfere with the transport of leukotriene C into hepatocytes. This suggests that the process is specific for cysteine-containing leukotrienes. It is likely that the transport mechanism described here participates in biliary excretion of leukotrienes. This route was previously found to be a major one for elimination of leukotriene C3 in mice and guinea-pigs.
Acetaminophen↗
Isolation and characterization of 15-hydroxylated metabolites of leukotriene C4.
A polar metabolite of leukotriene C4 was formed by sequential conversions with soybean lipoxygenase I and liver peroxidase. The structure of this product was found to be 5(S), 15(S)-dihydroxy-6(R)-S-glutathionyl-7,9,13-trans-11-cis-eicosatetraenoic acid (15-hydroxy-delta 13-trans-leukotriene C3. The HPLC behaviour, the molar extinction coefficient and the biological activity of the metabolite are reported. Preliminary evidence suggests that this product is formed by mammalian tissues.
Animals↗
Expression of Helix pomatia (HP) haemagglutinin receptors on cytolytic lymphocytes activated in mixed cultures.
The expression of receptors for Helix pomatia (HP) haemagglutinin, a T cell marker, was assayed on human lymphocytes cultivated with K562 or allogeneic lymphocytes. The receptor was detected on the cells after neuraminidase treatment by reactivity with FITC conjugated HP. By affinity chromatography the lymphocyte populations were separated into 3 subsets: (1) a subset which did not attach to the lectin column; (2) and (3) two subsets attaching with different avidities, and therefore eluting with different concentrations of the lectin-binding sugar hapten. The subsets were characterized for T cell markers, HP and E receptors, the B cell marker SIg and also for Fc receptors, and were tested for lytic potential against K562 and allogeneic blasts. A high proportion of HP receptor positive T blasts did not attach to the lectin column, and thus had low avidity HP receptors, confirming that activation of T lymphocytes is accompanied by decreased expression of T markers. The passed fraction which was enriched in blasts had the strongest cytotoxic function, while the fraction rich in cells with high avidity HP receptors and containing mainly small cells, had weak activity. This was true for both the anti-K562 and allospecific activities. Thus the phenotypic characteristics of the cells exerting the two types of lytic function were similar. The distribution of the lytic potential in the three subsets correlated with the presence of E and EA receptor positive blasts.
Cells, Cultured↗
Stereospecific elimination of hydrogen at C-10 in eicosapentaenoic acid during the conversion to leukotriene C5.
(10L)- and (10D)-[1-14C, 10-3H]5,8,11,14,17-eicosapentaenoic acids were synthesized to investigate mechanistic and stereochemical aspects of leukotriene biosynthesis. Experiments with mastocytoma cells showed that a hydrogen is stereospecifically eliminated from C-10 during the conversion of eicosapentaenoic acid to leukotriene C5. The hydrogen lost has the pro-S (D) configuration. 5-Hydroxy-6,8,11,14,17-eicosapentaenoic acid, formed in the same experiments, was enriched in tritium when the (10D), but not when the (10L), isomer of labeled eicosapentaenoic acid was used. This indicates that oxygenation of the acid at C-5 occurred before the elimination of hydrogen and suggests that removal of the pro-S hydrogen at C-10 in 5-hydroperoxy-6,8,11,14,17-eicosapentaenoic acid initiates its transformation to trans-5(S),6(S)-oxido-7,9-trans-11,14,17-cis-eicosapentaenoic acid (leukotriene A5).
Animals↗
A sensitive and specific radioimmunoassay for leukotriene C4.
A sensitive and specific radioimmunoassay suitable for direct measurement of leukotriene C4 was developed. Acetylated leukotriene C4 was coupled to polyamino bovine serum albumin using 1-ethyl-3-(3-dimethylamino-propyl) carbodiimide hydrochloride as coupling agent. The conjugate in complete or incomplete Freund's adjuvant was injected into New Zealand white rabbits. At a final antiplasma dilution of 1 : 1050 the lowest detection limit of leukotriene C4 was 0.046 pmol. The antiplasma cross-reacted less than 1% with leukotrienes D4, E4 and F4, while high relative cross-reaction (86-100%) was obtained with leukotrienes C3, C5 and 11-trans leukotriene C4. In experiments where known amounts of leukotriene C4 were added to leukocyte suspensions, 67-100% of the added amount was recovered by the method. The radioimmunoassay was used to study leukotriene C4 formation after stimulation of leukocyte suspensions with the ionophore A23187.
Antibody Specificity↗
The effect of prostaglandin I on the contractility of the term pregnant human myometrium.
Small myometrial strips were dissected from the upper and lower segments of the term pregnant human uterus. The specimens were superfused in organ chambers and contractile activity was recorded isometrically. In strips from the upper segment, prostacyclin (PGI2), induced an initial excitatory response followed in the majority of experiments by transient inhibition. In the lower segment the response was generally the same although direct inhibition without initial stimulation occurred in some cases. During the period of inhibition the specimens were refractory to iterated exposure to PGI2. Furthermore, during this period of PGI2-induced inhibition the muscle strip was also refractory to PGE2 but responded to PGF2 alpha and oxytocin by stimulation. After inhibition of spontaneous contractile activity induced by indomethacin PGI2 induced an excitatory response. The results do not indicate any critical change in the myometrial responsiveness of the upper uterine segment to PGI2 during labor. In strips from the lower segment obtained before labor there tended to be a dominance of non-responders and inhibition only as compared to the results during labor. Nevertheless, whether or not PGI2 under physiological or pharmacological conditions has any significant influence on the contractility of the term pregnant human uterus, still remains obscure. As judged from earlier reports from our laboratory and the present study it is evident that the uterine vessels are considerably more sensitive to the action of PGI2 than the myometrium.
Dinoprost↗
Wheal reactions in human skin after injection of leukotrienes B4, C4, D4 and E4.
Intradermal injections of 1-50 pmol leukotriene B4, C4, D4 and E4, (LTB4, LTC4, LTD4 and LTE4) were performed in healthy subjects and patients with recurrent urticaria. A wheal and erythema were seen 15 minutes after injection and were most marked after LTC4 and LTD4. LTE4 also produced a wheal whereas the reaction to LTB4 did not significantly differ from the saline control. When mixed with histamine no potentiation of the wheals was noted and no significant difference in reaction was observed between the healthy controls and those with urticaria.
Drug Hypersensitivity↗
A monoclonal antibody-enzyme immunoassay for serum carcinoembryonic antigen with increased specificity for carcinomas.
A two-site monoclonal antibody-enzyme immunoassay (MEIA) for carcinoembryonic antigen (CEA) was developed that uses two monoclonal anti-CEA antibodies, which recognize two different epitopes in the peptide moiety of CEA. The assay was sensitive to 0.5 micrograms/liter and had a measuring range of 0.5-200 micrograms of CEA per liter. It was highly specific inasmuch as none of three known CEA-related substances, "nonspecific crossreacting antigens 1 and 2" (NCA-1 and NCA-2) and biliary glycoprotein I (BGP I), reacted in the assay. NCA-2 (meconium antigen) is very similar to CEA. None of five commercially available CEA assays were able to differentiate between CEA and NCA-2. With one exception (colon), normal tissue extracts did not react in the MEIA even when tested at very high concentrations. Sera from a total of 180 healthy individuals and patients with malignant and nonmalignant disease were analyzed for CEA levels by using the MEIA and in parallel a conventional radioimmunoassay. A significant increase in specificity for carcinomas was obtained with the MEIA. This was essentially due to a decrease of MEIA CEA values in sera from patients with nonmalignant disease. The CEA values in the group of carcinoma patients (colon, pancreas, lung, and breast) were the same in the two assays.
Antibodies, Monoclonal↗
Mechanisms of leukotriene-induced contractions of guinea pig airways: leukotriene C4 has a potent direct action whereas leukotriene B4 acts indirectly.
The leukotrienes (LT's) are a group of arachidonic acid derivatives implicated as mediators of allergic bronchoconstriction and acute inflammation. Tracheal spirals and strips of lung parenchyma from guinea pigs were used under non-flow conditions to characterize the contractions caused by LTA4, LTB4 and LTC4. Cumulative administrations of leukotrienes desensitized the lung strip, whereas non-cumulative dose-response relationships for the leukotrienes and histamine were reasonably parallel. Half maximal contractions of the lung strips were obtained at a final bath concentration of 1 nM for LTC4 and 300 nM for LTA4 or LTB4, as compared with 6 000 nM for histamine. In the trachea, LTC4 was approximately 100 times more potent than LTA4 and histamine. Leukotrienes B4 and C4, but not acetylcholine or histamine, elicited release of the bronchoconstrictive thromboxane A2 from the lung under non-flow conditions. Indomethacin blocked the contractile response to LTB4, whereas the contractile effect of LTC4 remained unaltered. The beta-adrenoceptor agonist isoproterenol and the LTC4 antagonist FPL 55712 attenuated the contraction, but not the release of thromboxane A2, induced by LTC4. Changing to a perifusion technique rendered the lung strips less sensitive to the direct action of LTC4, and released thromboxane A2 now contributed significantly to the contractile response. In addition, the perifusion experiments indicated that LTB4 released histamine as well. We conclude that the chemoattractant LTB4 is an indirectly acting bronchoconstrictor, whereas the slow reacting substance LTC4 contracts the airway muscle by a predominantly direct mechanism. The exquisite bronchoconstrictive activity of LTC4 may be unrelated to its ability to induce formation of thromboxane A2.
Acetylcholine↗
NCA: a differentiation antigen of myelopoietic cells in humans and hominoid monkeys.
NCA, a normal colonic and granulocytic antigen, could be demonstrated in serum and in myelopoietic, but not lymphopoietic or erythropoietic, cells of Homo sapiens and other Primates. The levels of NCA in both serum and myelopoietic cells of Homo and hominoids were higher than those of more distant relatives of the same order. Thus, the classic phylogenetic differences are reflected also by the distribution of NCA. Hyperimmunization of Macaca irus, in which the NCA content of serum and cells is low, led to occurrence of anti-NCA IgG in all animals. The phylogenetic differences may accordingly have to do with slight antigenic NCA differences between Homo and other Primates rather than differences in amount only. Purified NCA did not affect growth and maturation of myelopoietic stem cells in vitro, whereas anti-NCA inhibited development of the majority of myelopoietic clusters and colonies.
Animals↗
Leukotrienes.
The leukotrienes are a family of biologically active molecules, formed by leukocytes, mastocytoma cells, macrophages, and other tissues and cells in response to immunological and nonimmunological stimuli. They exhibit a number of biological effects such as contraction of bronchial smooth muscles, stimulation of vascular permeability, and attraction and activation of leukocytes. Compared to histamine, which causes constriction of airways and edema formation, the leukotrienes are three to four orders of magnitude more potent and the effects have longer duration. The leukotrienes were discovered in 1938 as a smooth muscle-contracting factor in lung perfusates. It was referred to as "slow reacting substance" (SRS) or "slow reacting substance of anaphylaxis" (SRS-A) until 1979 when its structure was reported. The term "leukotriene" was introduced at that time as a trivial name for the new type of compound. Leukotrienes C4 and D4 are glutathione and cysteinylglycine conjugates, respectively, of arachidonic acid. After hydrolytic release from phospholipids of the cell membrane, arachidonic acid is oxygenated by a lipoxygenase to 5-hydroperoxy-6,8,11,14-eicosatetraenoic acid. This product is further converted to leukotrienes by elimination of the 10-pro-R hydrogen and OH from the hydroperoxy group to give 5,6-oxido-7,9,11, 14-eicosatetraenoic acid (leukotriene A4). Nucleophilic opening of the epoxide at C-6 by the sulfhydryl group of glutathione gives leukotriene C4, which is metabolized to leukotrienes D4 and E4 by sequential elimination of glutamic acid and glycine. The latter reactions are catalyzed by gamma-glutamyl transpeptidase and a particulate dipeptidase from kidney. Alternatively, water may add at C-12 of leukotriene A4, leading also to opening of the epoxide at C-6 with formation of 5,12-dihydroxy-6,8,10,14-eicosatetraenoic acid (leukotriene B4). Leukotriene B4 is metabolized by omega-hydroxylation to 20-hydroxy and 20-carboxy leukotriene B4. Leukotrienes are also formed from eicosatrienoic acid (n-9) and eicosapentaenoic acid (n-3) after oxygenation at C-5 and from eicosatrienoic acid (n-6) and arachidonic acid after oxygenation at C-8 (eicosatrienoic acid) and C-12 or C-15 (arachidonic acid). Although they are formed from the same and additional fatty acids as prostaglandins and thromboxanes [reviewed in this series in (1)], the structures and the reactions involved in biosynthesis and catabolism of leukotrienes are completely separate from those required for prostaglandin formation and metabolism. The leukotrienes seem to provide a new system of biological regulators that are important in many diseases involving inflammatory or immediate hypersensitivity reactions.
Animals↗
Active transport of leukotrienes into rat hepatocytes.
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Animals↗