共查询到20条相似文献,搜索用时 15 毫秒
1.
Wolfgang Hagmann Claudio Denzlinger Stephan Rapp Gisbert Weckbecker Dietrich Keppler 《Prostaglandins & other lipid mediators》1986,31(2)
Mercapturic acid formation, an established pathway in the detoxication of xenobiotics, is demonstrated for cysteinyl leukotrienes generated in rats
after endotoxin treatment. The mercapturate N-acetyl-leukotriene E4 (N-acetyl-LTE4) represented a major metabolite eliminated into bile after injection of [3H]LTC4 as shown by cochromatography with synthetic N-acetyl-LTE4 in four different HPLC solvent systems. The identity of endogenoud N-acetyl-LTE4 elicited by endotoxin
was additionally verified by enzymatic deacetylation followed by chemical N-acetylation. The deacetylation was catalyzed by penicillin amidase. Endogenous cysteinyl leukotrienes were quantified by radioimmunoassay after HPLC separation. A N-acetyl-LTE4 concentration of 80 nmol/l was determined in bile collected between 30 and 60 min after endotoxin injection. Under this condition, other cysteinyl leukotrienes detected in bile by radioimmunoassay amounted to less than 5% of N-acetyl-LTE4. The mercapturic acid pathway, leading from the glutathione conjugate LTC4 to N-acetyl-LTE4, thus plays an important role in the deactivation and elimination of these potent endogenous mediators. 相似文献
2.
F. Kohi D.K. Agrawal J.B. Cheng A.K. Bewtra R.G. Townley 《Prostaglandins & other lipid mediators》1987,33(1)
A simple and sensitive radioreceptor assay (RRA) for leukotrienes (LTs) was developed using a highly specific [3H]leukotriene D4 (LTD4) binding to guinea pig lung membrane homogenates. The assay can detect down to 0.15 pmol of LTD4. The values for fifty percent inhibition of bound [3H]LTD4 was 1.5 nM for LTD4, 45 nM for LTC4 and 24 nm for LTE4. LTB4 at 3.0 × 10−5 M had no effect on [3H]LTD4 binding. The RRA for LTs in the absence of serine-borate complex was bi-specific for both LTC4 and LTD4. However, in the presence of 20 nM serine-borate this method was highly specific for LTD4. Recovery rate averaged 87.2% after ethanol extraction and evaporation of known amounts of LTD4. When the radioreceptor assay and radioimmunoassay data for leukotriene levels in the samples were compared to each other, an excellent correlation was observed with a correlation coefficient ‘r’ of 0.992. The assay was also validated by quantitation of LTs released from human granulocytes stimulated with calcium ionophore, A23187. The method is simpler, less expensive, and more specific for LTD4 than the other methods such as high pressure liquid chromatography and radioimmunoassay and is suitable for routine measurement of either LTD4 specifically or LTC4 plus LTD4 simultaneously in one cell system. 相似文献
3.
Jose Sraer Marcelle Bens Jean-Paul Oudinet Raymond Ardaillou 《Prostaglandins & other lipid mediators》1986,31(5)
Since leukotriene C4 (LTC4) may be locally synthesized by bone marrow-derived cells infiltrating the kidney in inflammatory renal diseases we examined the in vitro metabolism of exogenously added |3H| LTC4 by rat glomeruli and papilla using radiometric HPLC. Homogenized as well as intact glomeruli converted |3H| LTC4 mainly into |3h| LTE4 (83%) and, at a smaller extent, into |3H| LTD4 (4%). Intact |3H| LTC4 represented 13% of the sum of radioactive leukotrienes. Addition of L-cysteine resulted in accumulation of LTD4. In contrast, there was nearly no conversion of |3H| LTC4 (87% ntact) in the presence of homogenized papilla. The metabolism of |3H| LTC4 by the glomeruli was time- and temperature- dependent. The 10,000 g supernatant and pellet of homogenized glomeruli both retained the ability to metabolize |3H| LTC4. The papillary 10,000 g supernatant was inactive, as found for the total homogenate, whereas the papillary 10,000 g pellet separated from its supernatant could transform |3H| LTC4 into its metabolites, LTD4 and LTE4. Addition of increasing amounts of papillary 10,000 g supernatant to homogenized glomeruli progressively protected |3H| LTC4 from its bioconversion. These results demonstrate that both glomeruli and papilla possess the γ-glutamyl transpeptidase and dipeptidase necessary to process LTC4. However, the enzyme activity of the papilla is unmasked only when the inhibitor present in the 10,000 g supernatant is separated from the enzyme present in the pellet. 相似文献
4.
《Prostaglandins, Leukotrienes and Medicine》1983,10(1):11-17
Leukotriene C4 (LTC4) and prostaglandin E2 (PGE2) have been studied for their effects on vascular permeability in rats. LTC4 and/or PGE2 were dissolved in 0.3% ethanol and were administered subcutaneously (0.1 ml) in the plantar surface of one of the hind paws of different series of rats. The changes in vascular permeability were measured by the radioactive marker (HSA. I125) method. LTC4 administered in dose of 2 × 10−8M produced marked increase (77 and 133%) in the vascular permeability (local edemogenic effect). PGE2 administered in a dose of 10−6M also produced significant increase (38 and 40%) in the vascular permeability. However, PGE2 in the same dose either administered along with LTC4 or administered at 30 minutes after the injection of LTC4 (2 × 10−8M) did not have any potentiating effect on the edemogenic response of LTC4. 相似文献
5.
Wei Zhang Xuexin Zhang José C. González-Cobos Judith A. Stolwijk Khalid Matrougui Mohamed Trebak 《The Journal of biological chemistry》2015,290(8):5015-5027
Leukotriene-C4 synthase (LTC4S) generates LTC4 from arachidonic acid metabolism. LTC4 is a proinflammatory factor that acts on plasma membrane cysteinyl leukotriene receptors. Recently, however, we showed that LTC4 was also a cytosolic second messenger that activated store-independent LTC4-regulated Ca2+ (LRC) channels encoded by Orai1/Orai3 heteromultimers in vascular smooth muscle cells (VSMCs). We showed that Orai3 and LRC currents were up-regulated in medial and neointimal VSMCs after vascular injury and that Orai3 knockdown inhibited LRC currents and neointimal hyperplasia. However, the role of LTC4S in neointima formation remains unknown. Here we show that LTC4S knockdown inhibited LRC currents in VSMCs. We performed in vivo experiments where rat left carotid arteries were injured using balloon angioplasty to cause neointimal hyperplasia. Neointima formation was associated with up-regulation of LTC4S protein expression in VSMCs. Inhibition of LTC4S expression in injured carotids by lentiviral particles encoding shRNA inhibited neointima formation and inward and outward vessel remodeling. LRC current activation did not cause nuclear factor for activated T cells (NFAT) nuclear translocation in VSMCs. Surprisingly, knockdown of either LTC4S or Orai3 yielded more robust and sustained Akt1 and Akt2 phosphorylation on Ser-473/Ser-474 upon serum stimulation. LTC4S and Orai3 knockdown inhibited VSMC migration in vitro with no effect on proliferation. Akt activity was suppressed in neointimal and medial VSMCs from injured vessels at 2 weeks postinjury but was restored when the up-regulation of either LTC4S or Orai3 was prevented by shRNA. We conclude that LTC4S and Orai3 altered Akt signaling to promote VSMC migration and neointima formation. 相似文献
6.
Jin T. Hong Mary W. Wilson Howard P. Glauert 《Journal of biochemical and molecular toxicology》1995,10(5):239-243
Peroxisome proliferators induce hepatocellular carcinomas in rodents by an unknown mechanism. γ-Glutamyltranspeptidase (GGT), a biochemical marker for identifying putative preneoplastic lesions in the liver, is highly expressed in phenobarbital (PB)-promoted altered hepatic foci but not in those promoted by peroxisome proliferators. One of the substrates of GGT is the eicosanoid LTC4. Because peroxisome proliferators and PB have differing effects on eicosanoid metabolism in vivo, we hypothesized that PB would similarly increase LTC4 concentrations, whereas the peroxisome proliferator ciprofibrate (CIP) would not. Cultured hepatocytes were treated with the peroxisome proliferator ciprofibrate (CIP: 100 and 400 μM) or PB (PB: 0.5 and 2 mM). Competitive radioimmunoassay (RIA) was used to determine the concentration of LTC4 in extracts of cultured hepatocytes. CIP decreased the concentration of LTC4 throughout the culture period, but PB increased the LTC4 concentration. Both doses of CIP significantly inhibited the induction of GGT activity at 48 and 72 hours, whereas PB enhanced GGT activity. We therefore hypothesized that LTC4, a substrate of GGT, may induce GGT activity. LTC4, however, did not enhance GGT activity and inhibited it at very high concentrations. The results of this experiment show that CIP and PB have different effects on GGT activity and LTC4 concentration. LTC4, however, does not induce GGT in cultured hepatocytes. © 1996 John Wiley & Sons, Inc. 相似文献
7.
Summary The leukotriene, LTC4, exerts a stimulatory effect on chloride transport in the frog cornea. In the work described here, the mechanism of action of LTC4 to stimulate chloride transport was studied.In corneas pretreated with indomethacin, the effect of LTC4 was abolished, suggesting the involvement of cyclo-oxygenase products in the response. Incubation of corneas with LTC4 resulted in a significant stimulation in PGE2 synthesis, as determined by TLC-autoradiography and radioimmunoassay. In addition, LTC4 was found to stimulate cAMP synthesis in the cornea, and this stimulation was blocked with indomethacin. PGE2 was previously shown by us to be the dominant cyclo-oxygenase product formed in the frog cornea, and is capable of stimulating cAMP and chloride transport. We suggest that LTC4 stimulation of chloride transport is mediated via activation of the cyclooxygenase pathway, resulting in enhanced PGE2 synthesis. Elevated PGE2 levels induce cAMP synthesis, and ultimately, the stimulation of chloride transport. Further, the activation of cyclo-oxygenase was found to be dependent on phospholipase A2 activity. This was shown by the inhibition of the LTC4 effect in the presence of quinacrine. Similarly, inhibition of the LTC4 effect in the presence of trifluoperazine suggests that cyclo-oxygenase activation by LTC4 may be mediated via calmodulin. We have previously demonstrated that the frog cornea has the biosynthetic capacity to produce LTC4. Therefore LTC4 may function as an endogenous regulator of chloride transport in this tissue. 相似文献
8.
Christopher W. Benjamin Nancy K. Hopkins Thomas D. Oglesby Robert R. Gorman 《Biochemical and biophysical research communications》1983,117(3):780-787
Leukotriene C4 (LTC4) and, to a lesser extent, leukotriene D4 (LTD4) concentration dependently stimulate prostacyclin (PGI2) biosynthesis in cultured human umbilical vein endothelial cells. PGI2 biosynthesis was quantitated by radioimmunoassay and its structure confirmed by gas chromatography/mass spectrometry. Preincubation of endothelial cells with LTC4 resulted in desensitization to subsequent LTC4 stimulation. However, PGI2 biosynthesis in response to thrombin, PGH2 and arachidonic acid was not inhibited by preincubation with LTC4. The C-6-sulfidopeptide leukotriene receptor level antagonist FPL-55712 attenuates LTC4, but not thrombin-stimulated PGI2 biosynthesis. These data suggest that human umbilical vein endothelial cells have a C-6-sulfidopeptide leukotriene receptor, and that stimulation of this receptor results in PGI2 biosynthesis. 相似文献
9.
Akinori Ueno Kunio Tanaka Makoto Katori Masashi Hayashi Yoshinobu Arai 《Prostaglandins & other lipid mediators》1981,21(4):637-648
The activity of synthetic LTC4 was tested in guinea-pig ileum and was 200 times more potent than histamine in contraction of the ileum (3 × 10?11 M- 3 × 10?9 M). The activities of LTC4 and LTD4 in increased vascular permeability in guinea pigs, rats and rabbits were compared with those histamine, bradykinin and prostaglandin (PG) E2. LTC4 was approximately equipotent to bradykinin on a molar basis in guinea pigs and rats and 5–100 times more potent than histamin. LTD4 was about 10 times more potent than LTC4 in guinea pigs and as equipotent to LTC4 in rats. On the contrary, in rabbits, neither LTC4 (upto 30 nmole/site) nor LTD4 (1 nmole/site) induced the dye exduation. These results show that species difference is present in activity of LTC4 and LTD4 in vascular permeability. Furthermore, in guinea pigs, the vascular permeability increased by LTC4 was not affected after pretreatment with pyrilamine (2.5 mg/kg, i.v.), and LTC4 and LTD4 did not potenciate the activity of bradykinin in vascular permeability. 相似文献
10.
C. Omini L. Daffonchio G. Brunelli F. Berti 《Prostaglandins & other lipid mediators》1985,29(6):1009-1020
The interaction of leukotriene C4 (LTC4) with the contractile activity of histamine (H), serotonin (5HT) and norepinephrine (NE) has been investigated in isolated vascular preparations. Threshold concentration of LTC4 (5 × 10−9 M) significantly potentiated the vasoconstricting effect of these compounds on guinea-pig pulmonary artery (GPPA). This phenomenon was long-lasting for H since it was still present 40 min after LTC4 had been washed. FPL-55712 (10−5M) counteracted the increased H response on GPPA induced by LTC4. Potentiation of H activity due to LTC4 was also observed on guinea-pig thoracic aorta (GPTA) indicating that LTC4-induced hyperreactivity is not a phenomenon restricted to the pulmonary vascular bed. In the experiments carried out in presence of indomethacin (3 × 10−6M), LTC4 still potentiated H-induced vasoconstriction on GPPA, however the time course of the phenomenon was significantly shorter than that observed in absence of the cyclooxygenase inhibitor. The contractile activity of H and NE on guinea-pig portal vein (GPPV) was not potentiated by LTC4 These results demonstrate that LTC4 induces hyperreactivity of the arterial vascular tissue to vasoactive compounds and suggest that cysteinyl-leukotrienes may have pathological significance in the hemodynamic changes occurring during anaphylactic reactions. Preliminary experiments carried out on human intralobar pulmonary artery strongly support this hypothesis. 相似文献
11.
Xuexin Zhang Wei Zhang José C. González-Cobos Isaac Jardin Christoph Romanin Khalid Matrougui Mohamed Trebak 《The Journal of general physiology》2014,143(3):345-359
Orai proteins contribute to Ca2+ entry into cells through both store-dependent, Ca2+ release–activated Ca2+ (CRAC) channels (Orai1) and store-independent, arachidonic acid (AA)-regulated Ca2+ (ARC) and leukotriene C4 (LTC4)-regulated Ca2+ (LRC) channels (Orai1/3 heteromultimers). Although activated by fundamentally different mechanisms, CRAC channels, like ARC and LRC channels, require stromal interacting molecule 1 (STIM1). The role of endoplasmic reticulum–resident STIM1 (ER-STIM1) in CRAC channel activation is widely accepted. Although ER-STIM1 is necessary and sufficient for LRC channel activation in vascular smooth muscle cells (VSMCs), the minor pool of STIM1 located at the plasma membrane (PM-STIM1) is necessary for ARC channel activation in HEK293 cells. To determine whether ARC and LRC conductances are mediated by the same or different populations of STIM1, Orai1, and Orai3 proteins, we used whole-cell and perforated patch-clamp recording to compare AA- and LTC4-activated currents in VSMCs and HEK293 cells. We found that both cell types show indistinguishable nonadditive LTC4- and AA-activated currents that require both Orai1 and Orai3, suggesting that both conductances are mediated by the same channel. Experiments using a nonmetabolizable form of AA or an inhibitor of 5-lipooxygenase suggested that ARC and LRC currents in both cell types could be activated by either LTC4 or AA, with LTC4 being more potent. Although PM-STIM1 was required for current activation by LTC4 and AA under whole-cell patch-clamp recordings in both cell types, ER-STIM1 was sufficient with perforated patch recordings. These results demonstrate that ARC and LRC currents are mediated by the same cellular populations of STIM1, Orai1, and Orai3, and suggest a complex role for both ER-STIM1 and PM-STIM1 in regulating these store-independent Orai1/3 channels. 相似文献
12.
S. Nicosia H.J. Crowley D. Oliva A.F. Welton 《Prostaglandins & other lipid mediators》1984,27(3):483-494
Leukotriene (LTC4) is one of the components of Slow Reacting Substance of Anaphylaxis (SRS-A) and is a potent constrictor of guinea pig ilea. The contraction is likely to be a receptor-mediated process. Here we report the existence of specific binding sites for 3H-LTC4 in a crude membrane preparation from guinea pig ileal longitudinal muscle.At 4°C in the presence of 20 mM Serine-borate, binding increases linearly with protein concentration, reaches equilibrium in 10 minutes, and is reversible upon addition of 3 × 10?5M unlabelled LTC4. The dissociation curve is consistent with the existence of more than one class of binding site. Ca++ and Mg++ greatly enhance the binding of 3H-LTC4 at equilibrium. In the presence of 5mM CaCl2 and MgCl2 not only LTC4 (IC50 10?7M), but also LTD4 (albeit with much lower affinity, IC50 = 6 × 10?5M) and the SRS-A antagonist FPL 55712 (IC50 = 10?5M) can compete with 3H-LTC4 for its binding sites. FPL 55712 only displaces 60–70% of the total amount bound, while LTC4 displaces 90–95%.These studies indicate that multiple classes of binding sites exist for 3H-LTC4 in guinea pig ileal longitudinal muscle, and that at least part of these binding sites might be related to the ability of LTC4 to contract guinea pig ilea. 相似文献
13.
Mara L. Leimanis Joel Karwatsky Elias Georges 《International Journal of Biochemistry and Molecular Biology》2011,2(1):39-46
Several studies have shown that the multidrug resistant protein MRP2 mediates the transport of chemotherapeutic drugs and normal cell metabolites, including Leukotriene C (LTC4); however direct binding of the LTC4 to MRP2 has not been demonstrated. In this study, a photoreactive analog of LTC4 (IAALTC4) was used to demonstrate its direct binding to MRP2. Our results show specific photoaffinity labeling of MRP2 with IAALTC4 in plasma membranes from MDCKIIMRP2 cells. The photoaffinity labeling signal of MRP2 with IAALTC4 was much lower than that of MRP1, consistent with previous studies whereby the measured Km values of MRP1 and MRP2 for LTC4 were 1 μM and 0.1 μM LTC4, respectively. Competition of IAALTC4 photoaffinity labeling to MRP2 with MK571, a well characterized inhibitor of MRP2 function, showed ~75% reduction in binding in the presence of 50 μM excess MK571. Interestingly, unmodified LTC4 enhanced the photoaffinity labeling of IAALTC4 to MRP2, whereas excess GSH and Quercetin had no significant effect. Mild tryptic digestion of photoaffinity labeled MRP2 revealed several photoaffinity labeled peptides that localized the IAALTC4 binding to a 15 kDa amino acid sequence that contains transmembrane 16 and 17. Together these results provide the first demonstration of direct LTC4 binding to MRP2. 相似文献
14.
John F. Burka 《Prostaglandins & other lipid mediators》1985,29(4):529-535
Ovalbumin (OA) and arachidonic acid (AA) were used to induce contractions of sensitized guinea-pig tracheal and lung preparations in the presence and absence of indomethacin. Leukotriene (LT)C4-like material released from these tissues was extracted from the bathing fluid and measured by radioimmunoassay. Challenge with either OA or AA induced release of LTC4-like material from both parenchyma and trachea, AA inducing a greater release than OA although OA induced greater contractions. This suggested that OA-induced the synthesis of other bronchoconstrictor compounds than LTC4. Although indomethacin enhanced OA- and AA-induced contractions of trachea, there was no enhancement of the release of LTC4-like material, suggesting enhancement by indomethacin was a result of the inhibition of the synthesis of prostaglandin E2 and not diversion of AA into the lipoxygenase pathway. Indomethacin had no effect on OA-induced contractions of parenchyma, but attenuated those induced by AA. Indomethacin had no modulatory effect on the release of LTC4-like material in the parenchyma. The results demonstrate that indomethacin does not result in increased synthesis of LTs in the airways. 相似文献
15.
Cornelius F. H. Mueller Marc Ulrich Becher Sebastian Zimmer Sven Wassmann Barbara Keuler Georg Nickenig 《Molecular and cellular biochemistry》2010,333(1-2):261-267
The multidrug resistance-related protein-1 (MRP1) is important for the management of oxidative stress in vascular cells in vivo. Substrates of MRP1 are, among others, glutathione and the leukotriene C4 (LTC4), an eicosanoid and mediator of inflammation. Angiotensin (Ang) II infusion results in MRP1?/? mice compared to wild-type mice in improved endothelial function and reduced reactive oxygen species (ROS) formation. However, the interaction between Ang II, LTC4 and MRP1 is not completely understood and has never been investigated in vitro. Ang II induced in vascular smooth muscle cells (VSMC) the release of LTC4 and the generation of ROS. Pharmacologic inhibition of MRP1 via MK 571 significantly reduced Ang II-induced ROS release (L012-luminescence) in VSMC. The release of ROS after Ang II stimulation is inhibited, to a comparable degree, by blockade of the Cys-LT1 receptor with montelukast. Incubation of VSMC with recombined LTC4 and Ang II caused enhanced rates of proliferation in VSMC. This effect can be rescued by either MRP1 or Cys-LT1 receptor inhibition. Accordingly, stimulation of VSMC with LTC4 reduces intracellular levels of glutathione, but does not affect apoptosis. LTC4 stimulation results in a significant activation of MRP1, but does not alter MRP1 expression. These findings indicate a connection between Ang II, MRP1 and LTC4. Both, MRP1 and LTC4, are potentially promising targets for atheroprotective therapy. 相似文献
16.
Xuexin Zhang José C. González-Cobos Rainer Schindl Martin Muik Brian Ruhle Rajender K. Motiani Jonathan M. Bisaillon Wei Zhang Marc Fahrner Margarida Barroso Khalid Matrougui Christoph Romanin Mohamed Trebak 《Molecular and cellular biology》2013,33(18):3715-3723
We recently showed, in primary vascular smooth muscle cells (VSMCs), that the platelet-derived growth factor activates canonical store-operated Ca2+ entry and Ca2+ release-activated Ca2+ currents encoded by Orai1 and STIM1 genes. However, thrombin activates store-independent Ca2+ selective channels contributed by both Orai3 and Orai1. These store-independent Orai3/Orai1 channels are gated by cytosolic leukotriene C4 (LTC4) and require STIM1 downstream LTC4 action. However, the source of LTC4 and the signaling mechanisms of STIM1 in the activation of this LTC4-regulated Ca2+ (LRC) channel are unknown. Here, we show that upon thrombin stimulation, LTC4 is produced through the sequential activities of phospholipase C, diacylglycerol lipase, 5-lipo-oxygenease, and leukotriene C4 synthase. We show that the endoplasmic reticulum-resident STIM1 is necessary and sufficient for LRC channel activation by thrombin. STIM1 does not form sustained puncta and does not colocalize with Orai1 either under basal conditions or in response to thrombin. However, STIM1 is precoupled to Orai3 and Orai3/Orai1 channels under basal conditions as shown using Forster resonance energy transfer (FRET) imaging. The second coiled-coil domain of STIM1 is required for coupling to either Orai3 or Orai3/Orai1 channels and for LRC channel activation. We conclude that STIM1 employs distinct mechanisms in the activation of store-dependent and store-independent Ca2+ entry pathways. 相似文献
17.
The effects of chemically-synthesised leukotrienes C4 and D4 (5(S) hydroxy-6(R)-δ-glutamylcysteinylglycinyl-7,9,11,14-eicosa-4tetraenoic acid, LTC4; 5(S) hydroxy-6(R)-cysteinylglycinyl-7,9,11,14-eicosatetraenoic acid, LTD4) on the microvasculature have been measured in guinea-pig skin using [125I]-albumin accumulation to measure plasma exudation and 133Xe clearance to measure blood flow changes. As previously shown using biosynthetic material, LTD4 caused vasoconstriction resulting in reduced blood flow. Similarly, LTC4 was found to have vasoconstrictor activity but was more potent and had a steeper dose-response curve than LTD4. There was no evidence of conversion of exogenous arachidonic acid to vaso-constrictor activity in the skin (in the absence of another stimulus): intradermally injected arachidonic acid produced vasodilatation, but induced little change in blood flow in animals pretreated with indomethacin. The vasodilator effect of arachidonic acid is presumed to be due to conversion to either PGE2 or PGI2. These results suggest that cyclo-oxygenase is normally active in the skin, whilst lipoxygenase requires activation in some way. As reported in a previous study, LTD4 induced plasma exudation when injected into the skin, but pronounced responses could only be induced by LTD4 mixed with a vasodilator prostaglandin such as PGE2. In contrast, LTC4 induced no exudation when tested alone and little when PGE2 was added. However, evidence was obtained that LTC4 has some permeability-increasing activity which is marked by its potent vasoconstrictor activity. 相似文献
18.
Dennis J. Heavey Roy J. Soberman Robert A. Lewis Bern Spur K. Frank Austen 《Prostaglandins & other lipid mediators》1987,33(5)
A sensitive and specific assay has been developed for measurement of total sulfidopeptide leukotriense (LT) in plasma. LTC4 and LTD4 in plasma are converted to LTE4 which is then extracted by C18 Sep-Pak binding and elution. Total LTE4 in resolved by reverse phase high performance liquid chromatography (RP-HPLC) and quantitated by radioimmunoassay (RIA). A [3H]LTE4 internal standard is added to the starting plasma sample to allow RP-HPLC to be assayed for LTE4-like immunoreactivity. The correlation between the measured increase in LTE4 concentration after addition of incremental amounts of LTC4 and LTE4 to plasma was 0.989 and 0.978, respectively, with slopes of 1.05 and 1.11. Addition of 51 pg/ml LTE4 to 5 ml plasma was detectable; the measured increase was 48 ± 12 pg/ml (mean ± SE, n = 7). The intra-assay coefficient of variation for 341 pg/ml of added LTC4 was 3.2% (n = 6). Sulfidopeptide leukotrienes could not be detected in blood samples taken from 12 normal volunteers in whom the theoretical detection limit, calculated from the sensitivity of the RIA, the overall recovery of LTE4, and the volume of plasma extracted, was 83 ± 4 pg LTE4/ml plasma (0.19 ± 0.01 pmol sulfidopeptide leukotriene/ml plasma; mean ± SE). 相似文献
19.
In order to examine the modulation of leukotriene (LT) release, the PAF-acether-mediated stimulation of these compounds in rat lung was studied. Release of LTC4, LTD4 and LTE4 in both perfused and chopped lung preparations was measured using HPLC and radioimmunoassay. Pre-incubation or pre-infusion of the tissue with indomethacin and PGE2 was conducted to investigate the effect of cyclooxygenase inhibitors and products on the lipoxygenase pathway. In addition, the effects of LT levels of pre-incubation with vasoactive intenstinal polypeptide (VIP) in chopped lung were observed.In perfused rat lung, indomethacin reduced the levels of LTC4 relative to LTD4 as measured in the first 2 min after stimulation of the lung by PAF-acether. Chopped lung preparations, incubated for 15 min. exhibited higher levels of LTC4 and LTD4 in indomethacin-treated samples, this increases being effectively reversed by PGE2.In the VIP pre-incubation experiments clear inhibition of peptido -leukotriene synthesis was observed, with no LTC4 and only low levels of LTD4 and LTE4 observed in VIP-incubated samples. In preliminary experiments using rabbit C5a des arg and PAF-acether on rabbit lung parenchyma strips to stimulaet LT release, disodium cromoglycate pre-incubation was observed to inhibit this release.Inhibition of the 5-lipoxygenase pathway of PGE2 is supported by these experiments. VIP appears to act as an inhibitor of LTC4 and LTD4 biosynthesis or release in this model. Too little is known that peptidergic actions to postulate a mechanism by which a neuroendocrine peptide exerts control of release of arachidonate metabolites; however, VIP is associated with muscarinic stimulation (1) and has been found in mast cells (2). 相似文献
20.
Leukotriene (LTC4) is one of the components of Slow Reacting Substance of Anaphylaxis (SRS-A) and is a potent constrictor of guinea pig ilea. The contraction is likely to be a receptor-mediated process. Here we report the existence of specific binding sites for 3H-LTC4 in a crude membrane preparation from guinea pig ileal longitudinal muscle.At 4°C in the presence of 20 mM Serine-borate, binding increases linearly with protein concentration, reaches equilibrium in 10 minutes, and is reversible upon addition of 3 × 10−5M unlabelled LTC4. The dissociation curve is consistent with the existence of more than one class of binding site. Ca++ and Mg++ greatly enhance the binding of 3H-LTC4 at equilibrium. In the presence of 5mM CaCl2 and MgCl2 not only LTC4 (IC50 10−7M), but also LTD4 (albeit with much lower affinity, IC50 = 6 × 10−5M) and the SRS-A antagonist FPL 55712 (IC50 = 10−5M) can compete with 3H-LTC4 for its binding sites. FPL 55712 only displaces 60–70% of the total amount bound, while LTC4 displaces 90–95%.These studies indicate that multiple classes of binding sites exist for 3H-LTC4 in guinea pig ileal longitudinal muscle, and that at least part of these binding sites might be related to the ability of LTC4 to contract guinea pig ilea. 相似文献