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- Patcha Brodin, Veronika, et al.
(författare)
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Differential inside-out activation of β2 integrins by leukotriene B4 and fMLP in human neutrophils
- 2004
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Ingår i: Experimental Cell Research. - : Elsevier BV. - 0014-4827 .- 1090-2422. ; 300:2, s. 308-319
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Tidskriftsartikel (refereegranskat)abstract
- We have investigated how LTB4, an endogenous chemoattractant encountered early in the inflammatory process, and fMLP, a bacteria-derived chemotactic peptide emanating from the site of infection, mediate inside-out regulation of the β2-integrin. The role of the two chemoattractants on β2-integrin avidity was investigated by measuring their effect on β2-integrin clustering and surface mobility, whereas their effect on β2-integrin affinity was measured by the expression of a high affinity epitope, a ligand-binding domain on β2-integrins, and by integrin binding to s-ICAM. We find that the two chemoattractants modulate the β2-integrin differently. LTB4 induces an increase in integrin clustering and surface mobility, but only a modest increase in integrin affinity. fMLP evokes a large increase in β2-integrin affinity as well as in clustering and mobility. Lipoxin, which acts as a stop signal for the functions mediated by pro-inflammatory agents, was used as a tool for further examining the inside-out mechanisms. While LTB4-induced integrin clustering and mobility were inhibited by lipoxin, only a minor inhibition of fMLP-induced β2-integrin avidity and no inhibition of integrin affinity were detected. The different modes of the inside-out regulation of β2-integrins suggest that distinct mechanisms are involved in the β2-integrin modulation induced by various chemoattractants.
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- Söderström, Mats, et al.
(författare)
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Novel prostaglandin D2-derived activators of peroxisome proliferator-activated receptor-γ are formed in macrophage cell cultures
- 2003
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Ingår i: Biochimica et Biophysica Acta. - : Elsevier. - 0006-3002 .- 1878-2434. ; 1631:1, s. 35-41
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Tidskriftsartikel (refereegranskat)abstract
- Incubation of RAW 264.7 murine macrophages with 9,15-dihydroxy-11-oxo-, (5Z,9alpha,13E,15(S))-Prosta-5,13-dien-1-oic acid [prostaglandin D(2) (PGD(2))] induced formation of considerable peroxisome proliferator-activated receptor-gamma (PPARgamma) activity [Nature 391 (1998) 79]. Because PGD(2) itself is a poor PPARgamma ligand, we incubated RAW 264.7 macrophage cultures with prostaglandin D(2) for 24 h and studied the ability of the metabolites formed to activate PPARgamma. PGD(2) products were extracted and fractionated by reverse phase high-performance liquid chromatography. Chemical identification was achieved by UV spectroscopy, gas-liquid chromatography/mass spectrometry and chemical syntheses of reference compounds. PGD(2) was converted to eight products, six of which were identified. Ligand-induced interaction of PPARgamma with steroid receptor coactivator-1 was determined by glutathione-S-transferase pull-down assays and PPARgamma activation was investigated by transient transfection of RAW 264.7 macrophages. In addition to the previously known ligand 11-oxo-(5Z,9,12E,14Z)-Prosta-5,9,12,14-tetraen-1-oic acid (15-deoxy-delta(12,14)-PGJ(2)), a novel PPARgamma ligand and activator viz. 9-hydroxy-11-oxo-, (5Z,9alpha,12E,14Z)-Prosta-5,12,14-trien-1-oic acid (15-deoxy-delta(12,14)-PGD(2)) was identified. The biological significance of these results is currently under investigation.
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- Wigren, Jane, 1967-, et al.
(författare)
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Differential recruitment of the coactivator proteins CREB-binding protein and steroid receptor coactivator-1 to peroxisome proliferator-activated receptor gamma/9-cis-retinoic acid receptor heterodimers by ligands present in oxidized low-density lipoprotein
- 2003
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Ingår i: Journal of Endocrinology. - : Bioscientifica. - 0022-0795 .- 1479-6805. ; 177:2, s. 207-214
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Tidskriftsartikel (refereegranskat)abstract
- Peroxisome proliferator-activated receptor gamma (PPAR?) colocalizes with oxidized low-density lipoprotein (LDL) in foam cells in atherosclerotic lesions. We have explored a potential role of oxidized fatty acids in LDL as PPAR? activators. LDL from patients suffering from intermittent claudication due to atherosclerosis was analyzed using HPLC and gas chromatography/mass spectrophotometry and found to contain 9-hydroxy-and 13-hydroxyoctadecadienoic acid (9- and 13-HODE), as well as 5-hydroxy-, 12-hydroxy- and 15-hydroxyeicosatetraenoic acid (5-, 12- and 15-HETE respectively). PPAR? was potently activated by 13(S)-HODE and 15(S)-HETE, as judged by transient transfection assays in macrophages or CV-1 cells. 5(S)- and 12(S)-HETE as well as 15-deoxy-?12,14 -prostaglandin J2 also activated PPAR? but were less potent. Interestingly, the effect of the lipoxygenase products 13(S -HODE and 15(S)-HETE as well as of the drug rosiglitazone were preferentially enhanced by the coactivator CREB-binding protein, whereas the effect of the cyclooxygenase product 15-deoxy-?12,14-prostaglandin J2 was preferentially enhanced by steroid receptor coactivator-1. We interpret these results, which may have relevance to the pathogenesis of atherosclerosis, to indicate that the lipoxygenase products on the one hand and the cyclooxygenase product on the other exert specific effects on the transcription of target genes through differential coactivator recruitment by PPAR?/9-cis retinoic acid receptor heterodimer complexes.
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- Wigren, Jane, 1967-
(författare)
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Identification of natural activators of the nuclear receptor peroxisome proliferator-activated receptor : relevance to the pathogenesis of atherosclerosis
- 1999
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Polyunsaturated fatty acids induce peroxisome proliferation. This phenomenon is mediated by the ligand-dependent transcription factor peroxisome proliferator-activated receptor (PPAR). This thesis is an investigation on the role of eicosanoids and oxidized products of linoleic acid for the activation of PPARs. Special emphasis was given to the subtype PPAR/gamma/ in the context of atherosclerosis.It had earlier been shown that arachidonic acid induces peroxisome proliferation in Morris Hepatoma 7800C1 cells. We investigated whether this effect could be attributed to a cytochrome P-450IVA product of arachidonic acid, 20-hydroxy-arachidonic acid. Arachidonic acid, but not 20-hydroxy-arachidonic acid induced lauryl-CoA oxidase activity. The effect of arachidonic acid was potentiated by all-trans retinoic acid, consistent with the notion that PPAR/RXR heterodimers mediate the effect.Several reports in the litterature were suggestive of an important role of peroxisomes in eicosanoid metabolism. However, nobody had isolated pure peroxisomes and investigated their eicosanoid metabolizing ability. We therefore investigated the ability of peroxisomes to metabolize the eicosanoid 12(S)-hydroxy-5,8,10,14-eicosatetraenoic acid (12(S)-HETE). Incubation of tritium-labeled 12(S)-HETE with isolated peroxisomes from rat liver or kidney peroxisomes demonstrated that more than 90 % of the diethyl ether extractable radioactivity was due to a single metabolite, identified as 8-hydroxy-6, 8, 12-octadecatrienoic acid (8-OH-16:3). This compound was apparently formed by two rounds of ß-oxidation. The data for the first time provided conclusive evidence for a role of peroxisomes in HETE metabolism.The second half of the thesis deals with the identification of natural PPAR/gamma/ ligands in LDL from atherosclerotic patients and in activated macrophages. Analyses of the endogenous content of selected monohydroxy fatty acids in LDL isolated from a group of patients diagnosed with intermittent claudication, showed the presence of 9- and 13-HODE, 5-, 12-, and 15-HETE. These compounds activated PPAR/gamma/ in macrophages and preferentially recruited the coactivator protein CBP to PPAR/gamma/RXR/alpha/ heterodimers. 15-deoxy-/DELTA/12,14-Prostaglandin J2 (15-deoxy-/DELTA/12,14-PGJ2) was identified as a PGD2 metabolite in macrophage cultures (see below). It induced the interaction of PPAR/gamma/RXR/alpha/ heterodimers with both CBP and SRC-1. This observation suggests that different PPAR/gamma/ ligands may induce different effects through a single kind of receptor by differential recruitment of coactivators.Although PGD2, is not a PPAR/gamma/ ligand, it induces PPAR/gamma/-mediated effects in IFN-/gamma/-stimulated RAW 264.7 macrophages, suggesting that the effects required metabolism. We therefore investigated PGD2 metabolism in macrophage cultures, and determined the capacity of these metabolites to activate PPAR/gamma/. Two novel (/DELTA/12-PGD2, 15-deoxy-/DELTA/12,14-PGD2) and two previously known PPAR/gamma/ activators (/DELTA/12-PGJ2 and 15-deoxy-/DELTA/12,14-PGJ2) were identified by mass spectrometry. The structural difference between the novel products and the previously recognized PPAR/gamma/ agonists , /DELTA/12-PGJ2 and 15-deoxy-/DELTA/12,14-PGJ2, is that they contain a 9/alpha/-hydroxy group and lack a /DELTA/9,10 double bond. Two novel PPAR/gamma/ activators were formed in equal or greater amounts and were more potent activators of PPAR/gamma/ in macrophages.
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