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E Granström

Publications and source records attributed to E Granström.

At least 19 recordsLinked to original sources
PubMed1999

Optimization of an enzyme immunoassay for 11-dehydro-thromboxane B(2) in urine: comparison with GC-MS.

The urinary excretion of stable metabolites of thromboxane A2, such as 11-dehydro-thromboxane B2, reflects platelet activity in vivo. Efficient sample purification is required before analysis of thromboxane metabolites, due to the presence of large amounts of interfering material in urine. Analysis by gas chromatography-mass spectrometry after extensive sample work-up procedures provides the most reliable data, but detection by enzyme immunoassay may be reliable if sample cleanup is adequate. We describe an improved immunoassay procedure for 11-dehydro-thromboxane B2, which is based on a simple one-step solid phase extraction, by using Bond-Elut Certify II columns, followed by enzyme immunoassay by using commercially available reagents. 11-Dehydro-thromboxane B2 exists in two forms, with different chemical and immunological characteristics, which are in pH-dependent equilibrium. We kept 11-dehydrothromboxane B2 in its open ring form throughout the assay, by incubating and handling samples at pH 8.6. The extraction step achieved a recovery of 83% (95% confidence interval 74-92%), the sensitivity of the enzyme immunoassay was doubled, and the reproducibility of the assay improved under these conditions. Intra- and interassay coefficients of variation were 3 and 13.8%, respectively. A single 500-mg dose of aspirin reduced the excretion of 11-dehydro-thromboxane B2 by 77+/-14%, suggesting good specificity. Comparison with gas chromatography-mass spectrometry in 28 urine samples showed excellent agreement between the two methods (r2 = 0.94; p<0.0001), and a regression line with a slope close to 1.0. The presently modified enzyme immunoassay for 11-dehydro-thromboxane B2 is suitable for clinical studies evaluating platelet function in vivo and has the advantage of being simpler and less expensive to use than gas chromatography-mass spectrometry.

Aspirin↗
PubMed1998

12-hydroxyeicosatetraenoic acid is a long-lived substance in the rabbit circulation.

12-Hydroxyeicosatetraenoic acid (12-HETE) is one of the major metabolites formed from arachidonic acid in platelets. We have recently shown that the in vitro metabolism of 12-HETE by human leukocytes, with and without stimulation, is effectively inhibited by the addition of physiological concentrations of albumin, probably by sequestration of the compound. In the present paper, we have studied the in vivo metabolism of 12-HETE in the rabbit, using either [1-14C]- or [14C(U)]12-HETE. Distribution of radioactivity was followed in urine, plasma, and bile, as well as in a number of tissues. In most of the tissues examined, the hydrophilic radioactivity constituted more than 50% of the total radioactivity after 20 min. When the lipophilic fraction was analyzed, around 15% of the radioactivity was shown to be unesterified 12-HETE, and only a very minor part could be detected as metabolites. The dominating lipophilic compound in the circulation after i.v. administration of radiolabeled 12-HETE was at all time points (1-60 min.) the parent compound, as analyzed by HPTLC and HPLC. A comparison of the plasma metabolite profiles obtained when [1-14C]- and [14C(U)]12-HETE were used displayed almost identical patterns, thus indicating that beta-oxidized metabolites either were not formed or were rapidly removed from the circulation. The appearance of large amounts of water-soluble radioactivity with time supported the latter conclusion. Several minor metabolites were seen that chromatographed in the dihydroxy acid region as judged by HPLC and TLC. The major one of these compounds represented about 10% of the lipophilic plasma radioactivity after 60 min., while unmetabolized 12-HETE at this stage still represented about 30%. The metabolite had a polarity similar to 12,20-dihydroxyeicosatetraenoic acid; however, when chromatographed together, these two compounds separated, indicating a different structure of the metabolite. Our findings are in agreement with in vitro data concerning the protective effect of albumin on the metabolism of 12-HETE and is the first extensive metabolic study of 12-HETE in vivo covering all metabolic possibilities involving the carbon skeleton.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1996

Albumin prevents metabolism of 12-hydroxyeicosatetraenoic acid by leukocytes in vitro.

In the present paper we studied the influence of albumin on the in vitro metabolism of 12-hydroxyeicosatetraenoic acid (12-HETE) and arachidonic acid in leukocytes and aspirin-treated platelets. In the presence of physiological concentrations of albumin, the metabolism of both 12-HETE and arachidonic acid was substantially altered, implicating the importance fatty acid binding proteins might have on the profile of products formed both in vitro and in vivo. The results clearly showed that albumin effectively withdraws arachidonic acid and 12-HETE from further metabolism by the leukocytes but does not influence the conversion of arachidonic acid to 12-HETE by the platelets. Thus, some of the hypotheses concerning transcellular metabolism raised from in vitro data within the eicosanoid field might have little relevance for the in vivo situation.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1995

Effects of oxygen radicals on cysteinyl leukotriene metabolism and pulmonary circulation in young pigs.

The effects of oxygen radicals, generated by the hypoxanthine-xanthine oxidase (XO) system, on pulmonary circulation and release of cysteinyl-containing leukotrienes (LTs) were studied in pigs after XO infusion into the right atrium. A 2.3-fold increase in pulmonary vascular resistance (PVR) (p < 0.05 vs. baseline) and a 2.1-fold increase in LT release (p < 0.05 vs. baseline) was observed. Pretreatment with indomethacin and allopurinol attenauted the vascular response (p < 0.01 and p < 0.05 vs. XO), and the LT release was inhibited by allopurinol and catalase (p < 0.01 and p < 0.02 vs. XO). We conclude that oxygen radicals stimulate lipoxygenase metabolism. This coincides with the observed increase in PVR, however, no causal relationship can be derived from the data presented.

Animals↗
PubMed1992

Metoprolol does not reduce platelet aggregability during sympatho-adrenal stimulation.

The possibility that beta-adrenoceptor blockers, especially beta 1-selective agents might inhibit platelet function is of considerable interest, as this might be of pathophysiological importance in cardiovascular diseases. Platelet function, however, is difficult to assess and in vivo related data are scarce. The effect of one week of treatment with metoprolol 200 mg/day on platelet aggregability during mental stress (colour word conflict test; CWT) and low and high dose adrenaline infusions has been evaluated in a double-blind, placebo-controlled, cross-over study in 10 healthy male volunteers. Platelet function in vivo was assessed using ex vivo filtragometry, and the urinary excretions of beta-thromboglobulin (HMW beta-TG) and 11-dehydro-TxB2 (a thromboxane metabolite). Conventional in vitro aggregometry and the urinary levels of 2,3-dinor-6-keto-PGF1 alpha (a prostacyclin metabolite) were also studied. During the interventions there was increased platelet aggregability in vivo, as filtragometry readings were shortened by 41 +/- 11% during high dose adrenaline infusion, urinary HMW beta-TG levels increased and urinary 11-dehydro-TxB2 tended to increase. In contrast, platelet sensitivity to ADP in vitro was reduced. The urinary 2,3-dinor-6-keto-PGF1 alpha levels were increased during the interventions. Despite the cardiovascular and biochemical signs of beta-adrenoceptor blockade at rest and during the interventions, metoprolol failed to influence platelet function in vivo, as measured by ex vivo filtragometry, or urinary HMW beta-TG or 11-dehydro-TxB2 levels. It tended rather to enhance the stress response measured by ex vivo filtragometry. Platelet aggregability in vitro and urinary 2,3-dinor-6-keto-PGF1 alpha levels were not altered by metoprolol.(ABSTRACT TRUNCATED AT 250 WORDS)

6-Ketoprostaglandin F1 alpha↗
PubMed1992

Urinary excretion of leukotriene E4 and 11-dehydro-thromboxane B2 in response to bronchial provocations with allergen, aspirin, leukotriene D4, and histamine in asthmatics.

In vivo production of thromboxane (TX) A2 and the cysteinyl-containing leukotrienes (LT) C4, D4, and E4 in correlation to airway responses was studied. Bronchial provocation with specific allergen in atopic asthmatics was followed by a significant increase in urinary concentration of immunoreactive LTE4 (34 +/- 6 before versus 56 +/- 7 ng/mmol creatinine after allergen challenge; n = 5) and 11-dehydro-TXB2 (164 +/- 29 versus 238 +/- 25 ng/mmol creatinine). In the presence of the leukotriene-antagonist ICI-204,219, which significantly increased the PD20 for allergen, the increment in urinary excretion of LTE4 was even higher (60 +/- 8 versus 288 +/- 128 ng/mmol creatinine; n = 5). In contrast, provocation with histamine (n = 5) did not provoke release of leukotrienes or thromboxane, nor was inhalation of LTD4 (n = 7) associated with increased urinary concentration of 11-dehydro-TXB2. Furthermore, bronchoconstriction induced by inhalation of lysine-aspirin in aspirin-sensitive asthmatics (n = 4) was followed by increased levels of LTE4 in the urine, whereas the levels of 11-dehydro-TXB2 remained the same. Finally, the basal levels of LTE4 in the urine of nine aspirin-sensitive asthmatics were elevated as compared with 15 other asthmatics (112 +/- 54 versus 38 +/- 20 ng/mmol creatinine; p less than 0.001). The findings support that the cysteinyl-leukotrienes are potential mediators of allergen-induced asthma and that the release of LTE4 and 11-dehydro-TXB2 into the urine appeared to be a direct and dose-dependent effect of the antigen-antibody reaction.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1990

15(S)-hydroxyeicosatetraenoic acid is the major arachidonic acid metabolite in human bronchi: association with airway epithelium.

15(S)-Hydroxy-5,8,11,13-eicosatetraenoic acid (15-HETE) was by far the most abundant metabolite of arachidonic acid in chopped human bronchi, as identified by reverse phase HPLC, uv spectrometry, and GC/MS. The quantitation of monohydroxyeicosatetraenoic acids (mono-HETEs) was performed by the use of 16(S)-hydroxy-9(Z),12(Z),14(E)-heneicosatrienoic acid as internal standard. Thus, significant amounts of 15-HETE were obtained in incubations of bronchi in buffer alone, but the addition of exogenous arachidonic acid (3-100 microM), dose-dependently increased the formation, with maximal levels reached at around 10 min. In contrast, challenge with ionophore A23187 or anti-human IgE did not stimulate the production of 15-HETE in the bronchi. Nordihydroguaiaretic acid inhibited the production of 15-HETE, whereas indomethacin did not. Small amounts of 8,15-diHETEs were detected in incubations with exogenous 15H(P)ETE. Lipoxins were however not detected under any of the incubation conditions used. Furthermore, removal of the airway epithelium substantially diminished the production of 15-HETE in the bronchi. Finally, bronchi were obtained from three patients with asthma, and the amounts of 15-HETE in these specimens were significantly higher than those found in tissues from nonasthmatics. Also, in peripheral lung parenchyma and pulmonary blood vessels 15-HETE was the major mono-HETE after stimulation with arachidonic acid but the levels were about 10 times lower than in the bronchi. As another difference, challenge of the parenchyma with the ionophore A23187 made 5-HETE the predominant mono-HETE. Taken together, airway epithelium appears to be the major source of 15-HETE in the human lung and the findings in specimens of asthmatics raise the possibility that 15-HETE somehow is involved in airway inflammation.

Arachidonic Acid↗
PubMed1990

Identification and biological activity of dihydroleukotriene B4: a prominent metabolite of leukotriene B4 in the human lung.

Exogenous [3H]leukotriene B4 (LTB4) was converted into several polar and non-polar metabolites in the chopped human lung. One of the major metabolites was identified as 5(S),12-dihydroxy-6,8,14-eicosatrienoic acid (10,11-dihydro-LTB4) by means of co-chromatography with authentic standards, ultraviolet spectrometry and gas chromatography-mass spectrometry. Analysis of chiral straight phase HPLC revealed the presence of both the 12(S) and 12(R) epimers of dihydro-LTB4. Dihydro-LTB4 was also formed from endogenously generated LTB4 in ionophore A23187 stimulated incubations. The dihydro metabolites were approximately 100 times less potent than LTB4 in causing guinea pig lung strip contraction and leukocyte-dependent inflammation in the hamster cheek pouch in vivo.

Animals↗
PubMed1987

Quantitative determination of prostaglandins E1, E2 and E3 in frog tissue.

A method was developed for quantitative determination of endogenous production of prostaglandin (PG)E1, PGE2 and PGE3 by Rana temporaria lung, heart and urinary bladder homogenates, since these tissues contain the precursors, 8,11,14-eicosatrienoic, arachidonic and 5,8,11,14,17-eicosapentaenoic acids. Following homogenization and shaking at 22 degrees C for 30 min, media were extracted by XAD-2, treated with sodium hydroxide in order to convert PGE compounds into PGB compounds, purified by thin-layer chromatography, and analyzed by high-performance liquid chromatography with homo-PGE1 as an internal standard. The ratio of prostaglandins E1, E2 and E3 compared to the ratio of fatty acid precursors in tissue suggested that the tissue content of precursor is not the only factor determining the type of prostaglandin synthesized.

Alprostadil↗
PubMed1987

Quality control of antibodies with special reference to prostaglandins.

Sources of error in the immunoassay of prostaglandins are reviewed. First, the specificity of the antibody, in terms of cross-reactions with structurally related substances, is often tested against irrelevant compounds, i.e. available compounds that do not occur in the biological material under study; major metabolises occurring in much larger amounts are overlooked. Hence, the reported high specificity of some antibodies may be apparent only. Second, many eicosanoids occur in two or more chemical forms in equilibrium in aqueous media. Antibodies may recognize one form preferentially; thus comparison of data from different laboratories is difficult. In addition numerous factors may interfere with the antigen-antibody binding in a non-immunological way. The most common effect is inhibition of antigen-antibody binding, but enhancement of binding sometimes occurs.

Journal Article↗
PubMed1986

Identification of 11-dehydro-TXB2 as a suitable parameter for monitoring thromboxane production in the human.

In order to identify suitable parameters for measurement of thromboxane production in vivo, the metabolism of TXB2 was studied in the human. [3H8]-TXB2 was given intravenously to a healthy human volunteer. Blood samples were collected for 50 min after the injection, and urine was collected for 24 hours. The urinary and blood metabolic profiles were visualized by the use of two-dimensional TLC and autoradiography. Identification of metabolites was achieved with GC/MS and in some cases by cochromatography with reference compounds in TLC and GC. In blood, unmetabolized TXB2 was the dominating compound during the first 30 min. Three less polar metabolites appeared, two of which were identified as 11-dehydro-TXB2 and 11,15-didehydro-13,14-dihydro-TXB2, respectively. The third compound was tentatively identified as 15-dehydro-13,14-dihydro-TXB2. Since 11-dehydro-TXB2 was one of the major metabolites in blood as well as urine, it was deemed suitable as target for measurement of thromboxane production in vivo. The advantages of 11-dehydro-TXB2 over its parent compound, TXB2, were demonstrated in experiments where unlabeled TXB2 was injected i.v. to a human volunteer, and the blood and urinary levels of both compounds were then followed by radioimmunoassay. Measured levels of 11-dehydro-TXB2 were found to give a more reliable picture of metabolic events than TXB2, the latter compound to a large extent reflecting technical difficulties during blood sample collection.

Autoradiography↗