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1.
Horm Metab Res ; 26(10): 465-9, 1994 Oct.
Article in English | MEDLINE | ID: mdl-7851869

ABSTRACT

There is evidence that iodoarachidonates are mediators of iodide in thyroid autoregulation, however, their occurrence in vivo has not yet been demonstrated. We therefore tried to identify delta-iodolactone (5-Hydroxy-6-iodo-8,11,14-eicosatrienoic delta-lactone, IL-delta) in thyroid tissue from a patient with Graves' disease treated with high doses of iodide. Lipids were extracted from thyroid tissue, purified by reversed phase chromatography and analyzed by gas chromatography--tandem mass spectrometry (GC-MSMS). The retention time in gas chromatography and fragmentation pattern in tandem mass spectrometry were determined with biochemically synthesized non-deuterated and deuterated IL-delta. According to retention time (13.44 min) and specific fragments (m/z 303, m/z 259) the occurrence of IL-delta could be demonstrated in the extract of iodide treated goiter. In vitro, potassium iodide (40 microM) as well as IL-delta (1.0 microM) significantly inhibited the proliferation of human thyroid follicular cells induced by phorbol ester TPA (12-O-tetradecanoylphorbol 13-acetate). These results demonstrate for the first time that Il-delta is present in iodide treated human thyroid. As cell proliferation is under negative control of IL-delta, a crucial role in thyroid involution following iodide treatment may be possible.


Subject(s)
Arachidonic Acids/analysis , Cell Division/drug effects , Goiter/drug therapy , Iodides/therapeutic use , Thyroid Gland/growth & development , Arachidonic Acids/isolation & purification , Gas Chromatography-Mass Spectrometry/methods , Humans , In Vitro Techniques , Thyroid Gland/cytology
2.
Arthritis Rheum ; 34(4): 481-5, 1991 Apr.
Article in English | MEDLINE | ID: mdl-2012627

ABSTRACT

Alterations in renal eicosanoid levels have been postulated as a factor in cyclosporin A (CSA) nephrotoxicity. The effects of CSA on renal eicosanoid excretion in rheumatoid arthritis were studied over a 24-week period, during which treatment with nonsteroidal antiinflammatory drugs was discontinued. The initial dosage of CSA was 4 mg/kg/day; at week 24, the mean dosage of CSA was 3.9 mg/kg/day. At week 24, the mean (+/- SD) serum creatinine level (1.04 +/- 0.24 mg/dl) was 32% above the baseline value; renal blood flow had decreased by 21% (P less than 0.03) and the glomerular filtration rate had decreased by 16%. There was a significant increase (P less than 0.03) in the 2,3-dinor thromboxane B2 level at week 2, but there was no significant change in the levels of the other eicosanoids. This study demonstrates that after CSA treatment, there is a selective increase in a thromboxane metabolite that parallels an increase in renal vascular resistance, even in the absence of nonsteroidal antiinflammatory drugs, and with unimpaired formation of other vasodilator eicosanoids.


Subject(s)
Arthritis, Rheumatoid/drug therapy , Cyclosporins/adverse effects , Eicosanoids/urine , Adult , Aged , Arthritis, Rheumatoid/metabolism , Creatinine/blood , Female , Glomerular Filtration Rate/drug effects , Humans , Hypertension/chemically induced , Kidney/drug effects , Kidney/metabolism , Male , Middle Aged , Renal Circulation/drug effects , Thromboxane B2/analogs & derivatives , Thromboxane B2/urine
3.
J Biol Chem ; 266(6): 3800-7, 1991 Feb 25.
Article in English | MEDLINE | ID: mdl-1899867

ABSTRACT

The role of endogenous arachidonic acid and its metabolites as mediators of cell growth was studied in rat mesangial cells. Inhibitors of the cytochrome P450 monooxygenase and lipoxygenase systems (nordihydroguaiaretic acid (NDGA), SK&F 525A, and ketoconazole) significantly reduced serum-stimulated cell growth as determined by cell counts and incorporation of [3H]thymidine. Inhibition of cyclooxygenase or lipoxygenases alone had no effect on cell growth. Stimulation with arginine vasopressin, epidermal growth factor, or phorbol myristate acetate increased [3H]thymidine incorporation and mRNA levels of the immediate-early response genes c-fos and Egr-1. These increases in [3H]thymidine incorporation and mRNA levels were reduced by NDGA and ketoconazole. NDGA, SK&F 525A, and ketoconazole had no effect on cellular ATP levels. Indomethacin had no effect upon cell growth. 14,15-Epoxyeicosatrienoic acid potentiated the effect of arginine vasopressin to enhance [3H]thymidine incorporation. Reverse-phase high pressure liquid chromatography analysis of lipid extracts from cells prelabeled with [3H]arachidonic acid resulted in the detection of a radioactive peak which eluted with lipoxygenase and monooxygenase products, with the same retention time as vicinal dihydroxyeicosatrienoic acids. This peak increased after stimulation with arginine vasopressin or epidermal growth factor and was reduced by preincubation with NDGA. Furthermore, analysis of unlabeled cell extracts by gas chromatography-mass spectrometry revealed the presence of a compound with epoxyeicosatrienoic acid-like characteristics. These results indicate that mesangial cells in culture likely produce products of the cytochrome P450 monooxygenase system that are important endogenous mediators of the growth response to mitogenic agents.


Subject(s)
Arachidonic Acids/metabolism , Glomerular Mesangium/metabolism , RNA, Messenger/analysis , Animals , Arachidonic Acid , Arginine Vasopressin/pharmacology , Blotting, Northern , Cells, Cultured , Chromatography, High Pressure Liquid , Cytochrome P-450 Enzyme Inhibitors , Epidermal Growth Factor/pharmacology , Gas Chromatography-Mass Spectrometry , Glomerular Mesangium/cytology , Indomethacin/pharmacology , Ketoconazole/pharmacology , Male , Masoprocol/pharmacology , Pyridines/pharmacology , Rats , Rats, Inbred WKY , Tetradecanoylphorbol Acetate/pharmacology
4.
Biochim Biophys Acta ; 1081(2): 197-201, 1991 Jan 28.
Article in English | MEDLINE | ID: mdl-1998738

ABSTRACT

Chronic smoking is a major risk factor of atherosclerosis and coronary heart disease. The measurement of three major thromboxane A2 metabolites, 11-dehydrothromboxane B2, 2,3-dinorthromboxane B2 and thromboxane B2, in the urines of 13 apparently healthy smokers (average 39 years, range 27-56 years) showed significantly elevated excretion rates for all thromboxane A2 metabolites as compared to 10 apparently healthy age-matched non-smokers (average 37 years, range 26-56 years). Importantly, characteristic alterations in the thromboxane A2 metabolite pattern were found in the urines of smokers. The contribution of 2,3-dinorthromboxane B2 to total measured excretion of thromboxane A2 metabolites was 59.2% in smokers (404.0 +/- 53.0 pg/mg creatinine) versus 19.4% in non-smokers (85.2 +/- 8.3 pg/mg creatinine), that of 11-dehydrothromboxane B2 35.7% in smokers (673.2 +/- 88.9 pg/mg creatinine) as compared to 75.5% in non-smokers (332.6 +/- 30.9 pg/mg creatinine). The contribution of thromboxane B2 (57.5 +/- 7.7 pg/mg creatinine in smokers versus 21.9 +/- 1.5 pg/mg creatinine in non-smokers) was similar at 5.1%. The excretion of cotinine, the major urinary metabolite of nicotine that correlates well with the reported daily cigarette consumption (r = 0.97, P less than 0.0001), showed a good correlation to thromboxane A2 metabolite excretion (2,3-dinorthromboxane B2: r = 0.92, P less than 0.0001; 11-dehydrothromboxane B2; r = 0.87, P less than 0.0001).


Subject(s)
Smoking/metabolism , Thromboxane A2/metabolism , Adult , Chromatography, Liquid , Cotinine/urine , Female , Gas Chromatography-Mass Spectrometry , Humans , Male , Middle Aged , Nicotine/metabolism
5.
Endocrinology ; 127(1): 337-43, 1990 Jul.
Article in English | MEDLINE | ID: mdl-2163315

ABSTRACT

Iodolactone (6-iodo-8,11,14-eicosatrienoic-delta-lactone), an iodinated derivative of arachidonic acid, was found to be synthesized in rat thyroid slices; however, the physiological role of this compound is still unknown. We tried to detect iodolactone in isolated porcine thyroid follicles and investigated the effects of in vitro synthesized iodolactone on epidermal growth factor-induced thyroid cell proliferation and TSH-induced cAMP formation. In vitro synthesis of iodolactone was performed with lactoperoxidase-catalyzed iodination of arachidonic acid in the presence of trace amounts of [125I]- and [3H]arachidonic acid. After purification by silica gel chromatography, HPLC of the reaction products revealed one main peak containing trace amounts of both [125I]- and [3H]arachidonic acid. With gas chromatography-mass spectrometry (GC-MS) a molecular mass of 391 m/z, corresponding to the derivatization product of iodolactone, was found. An ethanol-chloroform extract of isolated thyroid follicles preincubated with KI (10 microM) and arachidonic acid (1 microM) revealed peaks in HPLC and GC comparable with those of in vitro synthesized iodolactone. This indicates the ability of thyroid follicles to form iodolactone. Iodolactone (0.1-1.0 microM) dose-dependently inhibited epidermal growth factor-induced thyroid cell growth. This growth-inhibiting effect of iodolactone was 50-fold more pronounced than the inhibitory effect of KI (4 X 10(-5) microM) on thyroid cell proliferation. In contrast to the effect of iodide, the inhibitory effect of iodolactone on thyroid cell growth could not be abolished by methimazole (1 mM). Basal as well as TSH (0.5 U/liter)-induced cAMP formation were not changed by iodolactone. These experiments suggest a physiological role of iodolactone as a mediator of the known inhibitory effect of iodide on thyroid growth.


Subject(s)
Arachidonic Acids/pharmacology , Cyclic AMP/biosynthesis , Iodides/pharmacology , Thyroid Gland/cytology , Animals , Arachidonic Acid , Arachidonic Acids/analysis , Arachidonic Acids/biosynthesis , Arachidonic Acids/metabolism , Cell Division/drug effects , Chromatography, High Pressure Liquid , Epidermal Growth Factor/pharmacology , Gas Chromatography-Mass Spectrometry , Lactoperoxidase/metabolism , Methimazole/pharmacology , Swine , Thyroid Gland/drug effects , Thyroid Gland/metabolism , Thyrotropin/pharmacology
6.
Biochim Biophys Acta ; 993(2-3): 259-65, 1989 Dec 08.
Article in English | MEDLINE | ID: mdl-2597697

ABSTRACT

Urinary immunoreactive thromboxane (irTXB2) has been found helpful in acute settings with altered renal, but also extrarenal thromboxane formation. As only trace amounts of systemically formed thromboxane are excreted unmetabolized, the nature of urinary irTXB2 was explored. The two most abundant metabolites of systemic thromboxane, 2,3-dinor-TXB2 and 11-dehydro-TXB2, crossreacted about 70% and less than 1%, respectively, with a widely used thromboxane antiserum. After solid-phase extraction of urine samples and separation on reversed-phase HPLC, the bulk of immunoreactivity always eluted as one peak shown to correspond to 2,3-dinor-TXB2. Much less was found in fractions where TXB2 eluted. Therefore, urines were read against calibration curves constructed with 2,3-dinor-TXB2. This direct estimation gave good recoveries for standard 2,3-dinor-TXB2 and correlated well, both in healthy controls and in patients at increased risk or with overt vascular disease, to values obtained after solid phase extraction, purification on reversed-phase HPLC and quantitation by either gas-chromatography mass-spectrometry or radioimmunoassay. Patients with multiple cardiovascular risk factors but free from detectable vascular disease excreted significantly more irTXB2 than age-matched controls with non-vascular conditions or normals. Therefore, urinary irTXB2 measured with this antiserum represents 2,3-dinor-TXB2, reflecting the systemic formation of TXB2. This simple approach is feasible for screening thromboxane formation in large series of patients. Its acumen in detecting the early development of vascular disease and its relation to established risk factors deserves large-scale prospective testing.


Subject(s)
Thromboxane B2/urine , Vascular Diseases/urine , Chromatography, High Pressure Liquid , Gas Chromatography-Mass Spectrometry , Humans , Radioimmunoassay , Risk Factors , Thromboxane B2/analogs & derivatives
7.
J Chromatogr ; 497: 1-16, 1989 Dec 29.
Article in English | MEDLINE | ID: mdl-2625446

ABSTRACT

2,3-Dinorthromboxane B2 and 11-dehydrothromboxane B2, the two major metabolites of thromboxane B2, are considered to be indices of thromboxane A2 activity in humans. The determination of these metabolites in urine was comparatively performed by gas chromatography-mass spectrometry and gas chromatography-mass spectrometry-mass spectrometry using the corresponding chemically synthesized tetradeuterated analogues as internal standards. The urine samples of five females and two males, all healthy, were prepurified by solid-phase extraction. The corresponding pentafluorobenzyl ester derivatives were repurified by high-performance liquid chromatography. The concentrations of 2,3-dinorthromboxane B2 and 11-dehydrothromboxane B2 ranged from 21 to 266 pg/ml and 47 to 942 pg/ml, respectively. The ratio of urinary 2,3-dinorthromboxane B2 to 11-dehydrothromboxane B2 varied from 1:3 to 1:5, except for one sample with nearly equal concentrations of 2,3-dinorthromboxane B2 and 11-dehydrothromboxane B2.


Subject(s)
Thromboxane B2/analogs & derivatives , Thromboxanes/metabolism , Chemical Phenomena , Chemistry , Chromatography, High Pressure Liquid , Gas Chromatography-Mass Spectrometry/methods , Humans , Thromboxane B2/urine
8.
Am J Cardiol ; 64(18): 1185-8, 1989 Nov 15.
Article in English | MEDLINE | ID: mdl-2510490

ABSTRACT

To explore the effect of timing on the antiplatelet action of aspirin, a constant mean amount of 40 mg aspirin/day was administered either as a split regimen of 20 mg twice daily, a single dose of 40 mg or a doubled dose of 80 mg every other day for 1 week each and compared to a current standard low dose regimen of 324 mg/day. Bleeding time, serum thromboxane, collagen-stimulated platelet aggregation and associated thromboxane formation and excretion of thromboxane and prostacyclin metabolites were measured both at peak and trough action of the drug. The inhibitory effects on platelet aggregation and associated thromboxane formation were significantly less marked with the split dose regimen, intermediate with the single dose of 40 mg aspirin/day and best with the alternate day doubled dose, but still inferior to the effects of 324 mg/day. Thromboxane excretion was suppressed by greater than 80% with all regimens. Prostacyclin metabolite excretion was similar for all 40 mg/day regimens with about 40% suppression at trough and 60% at peak drug action, respectively. Suppression was more pronounced after 324 mg/day. For best platelet inactivation at comparable sparing of prostacyclin formation, low doses of aspirin should be administered in pulsed rather than split regimens.


Subject(s)
Aspirin/administration & dosage , Epoprostenol/biosynthesis , Platelet Aggregation Inhibitors , Thromboxanes/biosynthesis , Aspirin/pharmacology , Blood Coagulation/drug effects , Dose-Response Relationship, Drug , Epoprostenol/urine , Humans , Thromboxanes/blood , Thromboxanes/urine
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