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Intracrine cysteinyl leukotriene receptor-mediated signaling of eosinophil vesicular transport-mediated interleukin-4 secretion.

Bandeira-Melo C, Woods LJ, Phoofolo M, Weller PF - J. Exp. Med. (2002)

Bottom Line: LTC(4)-elicited IL-4 release was pertussis toxin inhibitable, but inhibitors of the two known G protein-coupled cysLT receptors (cysLTRs) (CysLT1 and CysLT2) did not block LTC(4)-elicited IL-4 release.Thus, LTC(4) acts, via an intracellular cysLTR distinct from CysLT1 or CysLT2, as a signal transducer to selectively regulate IL-4 release.These results demonstrate that LTC(4), well recognized as a paracrine mediator, may also dynamically govern inflammatory and immune responses as an intracrine mediator of eosinophil cytokine secretion.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Harvard Thorndike Laboratories, Charles A. Dana Research Institute, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215, USA.

ABSTRACT
We investigated whether cysteinyl leukotrienes (cysLT) are intracrine signal transducers that regulate human eosinophil degranulation mechanisms. Interleukin (IL)-16, eotaxin, and RANTES stimulate vesicular transport-mediated release of preformed, granule-derived IL-4 and RANTES from eosinophils and the synthesis at intracellular lipid bodies of LTC(4), the dominant 5-lipoxygenase-derived eicosanoid in eosinophils. 5-Lipoxygenase inhibitors blocked IL-16-, eotaxin-, and RANTES-induced IL-4 release; but neither exogenous LTC(4), LTD(4), nor LTE(4) elicited IL-4 release. Only after membrane permeabilization enabled cysLTs to enter eosinophils did LTC(4) and LTD(4) stimulate IL-4, but not RANTES, release. LTC(4)-elicited IL-4 release was pertussis toxin inhibitable, but inhibitors of the two known G protein-coupled cysLT receptors (cysLTRs) (CysLT1 and CysLT2) did not block LTC(4)-elicited IL-4 release. LTC(4) was 10-fold more potent than LTD(4) and at low concentrations (0.3-3 nM) elicited, and at higher concentrations (>3 nM) inhibited, IL-4 release from permeabilized eosinophils. Likewise with intact eosinophils, LTC(4) export inhibitors, which increased intracellular LTC(4), inhibited eotaxin-elicited IL-4 release. Thus, LTC(4) acts, via an intracellular cysLTR distinct from CysLT1 or CysLT2, as a signal transducer to selectively regulate IL-4 release. These results demonstrate that LTC(4), well recognized as a paracrine mediator, may also dynamically govern inflammatory and immune responses as an intracrine mediator of eosinophil cytokine secretion.

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Effect of kinase inhibitors on LTC4- and eotaxin-induced IL-4 secretion from eosinophils. (A) Plasma membrane–permeabilized eosinophils were pretreated for 30 min with wortmannin (1 μM), LY294002 (10 μM), PD98059 (10 μM), U0126 (10 μM), SB203580 (10 μM), genistein (10 μM), herbimycin (10 μM), staurosporin (1 μM), chelerythrine (10 μM), or calphostin C (1 μM), and then stimulated for 3 h with LTC4 (3 nM). (B) Intact eosinophils were pretreated as described in A and then stimulated for 1 h with eotaxin (6 nM). Results were expressed as the percentages of eosinophils exhibiting extracellular IL-4. Values are means ± SD from three independent assays. + and * represent P < 0.01 compared with nonstimulated (A, 6 ± 3% baseline positive; B, 5 ± 2% positive) and stimulated eosinophils, respectively.
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fig7: Effect of kinase inhibitors on LTC4- and eotaxin-induced IL-4 secretion from eosinophils. (A) Plasma membrane–permeabilized eosinophils were pretreated for 30 min with wortmannin (1 μM), LY294002 (10 μM), PD98059 (10 μM), U0126 (10 μM), SB203580 (10 μM), genistein (10 μM), herbimycin (10 μM), staurosporin (1 μM), chelerythrine (10 μM), or calphostin C (1 μM), and then stimulated for 3 h with LTC4 (3 nM). (B) Intact eosinophils were pretreated as described in A and then stimulated for 1 h with eotaxin (6 nM). Results were expressed as the percentages of eosinophils exhibiting extracellular IL-4. Values are means ± SD from three independent assays. + and * represent P < 0.01 compared with nonstimulated (A, 6 ± 3% baseline positive; B, 5 ± 2% positive) and stimulated eosinophils, respectively.

Mentions: We have shown recently that LTC4 production at cytoplasmic lipid bodies within CCR3 chemokine-stimulated eosinophils depended on activation of PI3K and MAP kinases (20). With specific kinase inhibitors, we investigated the downstream signaling of LTC4- and eotaxin-induced IL-4 release from eosinophils. Both LTC4 and eotaxin-induced IL-4 release were blocked by inhibitors of PI3K (wortmannin, LY294002) and tyrosine kinases (genistein, herbimycin A), but not by PKC inhibitors (staurosporin, chelerythrine, and calphostin C; Fig. 7) . In contrast, the MAP kinase inhibitors, PD98059 and U0126 (inhibitors of the extracellular signal-regulated kinases (ERK)1/2 activating kinase MEK [MAP ERK kinase]), and SB203586 (a p38 MAP kinase inhibitor), significantly inhibited eotaxin's actions (Fig. 7 B) without affecting LTC4-induced IL-4 secretion (Fig. 7 A).


Intracrine cysteinyl leukotriene receptor-mediated signaling of eosinophil vesicular transport-mediated interleukin-4 secretion.

Bandeira-Melo C, Woods LJ, Phoofolo M, Weller PF - J. Exp. Med. (2002)

Effect of kinase inhibitors on LTC4- and eotaxin-induced IL-4 secretion from eosinophils. (A) Plasma membrane–permeabilized eosinophils were pretreated for 30 min with wortmannin (1 μM), LY294002 (10 μM), PD98059 (10 μM), U0126 (10 μM), SB203580 (10 μM), genistein (10 μM), herbimycin (10 μM), staurosporin (1 μM), chelerythrine (10 μM), or calphostin C (1 μM), and then stimulated for 3 h with LTC4 (3 nM). (B) Intact eosinophils were pretreated as described in A and then stimulated for 1 h with eotaxin (6 nM). Results were expressed as the percentages of eosinophils exhibiting extracellular IL-4. Values are means ± SD from three independent assays. + and * represent P < 0.01 compared with nonstimulated (A, 6 ± 3% baseline positive; B, 5 ± 2% positive) and stimulated eosinophils, respectively.
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fig7: Effect of kinase inhibitors on LTC4- and eotaxin-induced IL-4 secretion from eosinophils. (A) Plasma membrane–permeabilized eosinophils were pretreated for 30 min with wortmannin (1 μM), LY294002 (10 μM), PD98059 (10 μM), U0126 (10 μM), SB203580 (10 μM), genistein (10 μM), herbimycin (10 μM), staurosporin (1 μM), chelerythrine (10 μM), or calphostin C (1 μM), and then stimulated for 3 h with LTC4 (3 nM). (B) Intact eosinophils were pretreated as described in A and then stimulated for 1 h with eotaxin (6 nM). Results were expressed as the percentages of eosinophils exhibiting extracellular IL-4. Values are means ± SD from three independent assays. + and * represent P < 0.01 compared with nonstimulated (A, 6 ± 3% baseline positive; B, 5 ± 2% positive) and stimulated eosinophils, respectively.
Mentions: We have shown recently that LTC4 production at cytoplasmic lipid bodies within CCR3 chemokine-stimulated eosinophils depended on activation of PI3K and MAP kinases (20). With specific kinase inhibitors, we investigated the downstream signaling of LTC4- and eotaxin-induced IL-4 release from eosinophils. Both LTC4 and eotaxin-induced IL-4 release were blocked by inhibitors of PI3K (wortmannin, LY294002) and tyrosine kinases (genistein, herbimycin A), but not by PKC inhibitors (staurosporin, chelerythrine, and calphostin C; Fig. 7) . In contrast, the MAP kinase inhibitors, PD98059 and U0126 (inhibitors of the extracellular signal-regulated kinases (ERK)1/2 activating kinase MEK [MAP ERK kinase]), and SB203586 (a p38 MAP kinase inhibitor), significantly inhibited eotaxin's actions (Fig. 7 B) without affecting LTC4-induced IL-4 secretion (Fig. 7 A).

Bottom Line: LTC(4)-elicited IL-4 release was pertussis toxin inhibitable, but inhibitors of the two known G protein-coupled cysLT receptors (cysLTRs) (CysLT1 and CysLT2) did not block LTC(4)-elicited IL-4 release.Thus, LTC(4) acts, via an intracellular cysLTR distinct from CysLT1 or CysLT2, as a signal transducer to selectively regulate IL-4 release.These results demonstrate that LTC(4), well recognized as a paracrine mediator, may also dynamically govern inflammatory and immune responses as an intracrine mediator of eosinophil cytokine secretion.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Harvard Thorndike Laboratories, Charles A. Dana Research Institute, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02215, USA.

ABSTRACT
We investigated whether cysteinyl leukotrienes (cysLT) are intracrine signal transducers that regulate human eosinophil degranulation mechanisms. Interleukin (IL)-16, eotaxin, and RANTES stimulate vesicular transport-mediated release of preformed, granule-derived IL-4 and RANTES from eosinophils and the synthesis at intracellular lipid bodies of LTC(4), the dominant 5-lipoxygenase-derived eicosanoid in eosinophils. 5-Lipoxygenase inhibitors blocked IL-16-, eotaxin-, and RANTES-induced IL-4 release; but neither exogenous LTC(4), LTD(4), nor LTE(4) elicited IL-4 release. Only after membrane permeabilization enabled cysLTs to enter eosinophils did LTC(4) and LTD(4) stimulate IL-4, but not RANTES, release. LTC(4)-elicited IL-4 release was pertussis toxin inhibitable, but inhibitors of the two known G protein-coupled cysLT receptors (cysLTRs) (CysLT1 and CysLT2) did not block LTC(4)-elicited IL-4 release. LTC(4) was 10-fold more potent than LTD(4) and at low concentrations (0.3-3 nM) elicited, and at higher concentrations (>3 nM) inhibited, IL-4 release from permeabilized eosinophils. Likewise with intact eosinophils, LTC(4) export inhibitors, which increased intracellular LTC(4), inhibited eotaxin-elicited IL-4 release. Thus, LTC(4) acts, via an intracellular cysLTR distinct from CysLT1 or CysLT2, as a signal transducer to selectively regulate IL-4 release. These results demonstrate that LTC(4), well recognized as a paracrine mediator, may also dynamically govern inflammatory and immune responses as an intracrine mediator of eosinophil cytokine secretion.

Show MeSH
Related in: MedlinePlus