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Cytosolic phospholipase A₂-? participates in lipid body formation and PGE₂ release in human neutrophils stimulated with an L-amino acid oxidase from Calloselasma rhodostoma venom.

ABSTRACT: Cr-LAAO, an L-amino acid oxidase isolated from Calloselasma rhodosthoma snake venom, has been demonstrated as a potent stimulus for neutrophil activation and inflammatory mediator production. However, the mechanisms involved in Cr-LAAO induced neutrophil activation has not been well characterized. Here we investigated the mechanisms involved in Cr-LAAO-induced lipid body (also known as lipid droplet) biogenesis and eicosanoid formation in human neutrophils. Using microarray analysis, we show for the first time that Cr-LAAO plays a role in the up-regulation of the expression of genes involved in lipid signalling and metabolism. Those include different members of phospholipase A_2, mostly cytosolic phospholipase A₂-? (cPLA₂-?); and enzymes involved in prostaglandin synthesis including cyclooxygenases 2 (COX-2), and prostaglandin E synthase (PTGES). In addition, genes involved in lipid droplet formation, including perilipin 2 and 3 (PLIN 2 and 3) and diacylglycerol acyltransferase 1 (DGAT1), were also upregulated. Furthermore, increased phosphorylation of cPLA₂-?, lipid droplet biogenesis and PGE₂ synthesis were observed in human neutrophils stimulated with Cr-LAAO. Treatment with cPLA₂-? inhibitor (CAY10650) or DGAT-1 inhibitor (A922500) suppressed lipid droplets formation and PGE₂ secretion. In conclusion, we demonstrate for the first time the effects of Cr-LAAO to regulate neutrophil lipid metabolism and signalling.

SUBMITTER: Paloschi MV

PROVIDER: S-EPMC7334223 | biostudies-literature | 2020 Jul

REPOSITORIES: biostudies-literature

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Cytosolic phospholipase A<sub>2</sub>-α participates in lipid body formation and PGE<sub>2</sub> release in human neutrophils stimulated with an L-amino acid oxidase from Calloselasma rhodostoma venom.

Paloschi Mauro Valentino MV Lopes Jéssica Amaral JA Boeno Charles Nunes CN Silva Milena Daniela Souza MDS Evangelista Jaína Rodrigues JR Pontes Adriana Silva AS da Silva Setúbal Sulamita S Rego Cristina Matiele Alves CMA Néry Neriane Monteiro NM Ferreira E Ferreira Alex Augusto AA Pires Weverson Luciano WL Felipin Kátia Paula KP Ferreira Gabriel Eduardo Melim GEM Bozza Patrícia Torres PT Zuliani Juliana Pavan JP

Scientific reports 20200703 1

Cr-LAAO, an L-amino acid oxidase isolated from Calloselasma rhodosthoma snake venom, has been demonstrated as a potent stimulus for neutrophil activation and inflammatory mediator production. However, the mechanisms involved in Cr-LAAO induced neutrophil activation has not been well characterized. Here we investigated the mechanisms involved in Cr-LAAO-induced lipid body (also known as lipid droplet) biogenesis and eicosanoid formation in human neutrophils. Using microarray analysis, we show for ...[more]

PMID: 32620771

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Project description:Pulmonary fibrosis is characterized by fibroblasts persisting in an activated form, producing excessive fibrous material that destroys alveolar structure. The second messenger molecule cyclic 3',5'-adenosine monophosphate (cAMP) has antifibrotic properties, and prostaglandin E2 (PGE2) can stimulate cAMP production through prostaglandin E (EP)2 and EP4 receptors. Although EP receptors are attractive therapeutic targets, the effects of long-term exposure to PGE2 have not been characterized. To determine the effects of long-term exposure of lung fibroblasts to PGE2, human fetal lung (HFL)-1 cells were treated for 24 h with 100 nM PGE2 or other cAMP-elevating agents. cAMP levels stimulated by acute exposure to PGE2 were measured using a fluorescent biosensor. Pretreatment for 24 h with PGE2 shifted the concentration-response curve to PGE2 rightward by approximately 22-fold but did not affect responses to the beta-adrenoceptor agonist isoproterenol. Neither isoproterenol nor forskolin pretreatment altered PGE2 responses, implying that other cAMP-elevating agents do not induce desensitization. Use of EP2- and EP4-selective agonists and antagonists suggested that PGE2-stimulated cAMP responses in HFL-1 cells are mediated by EP2 receptors. EP2 receptors are resistant to classical mechanisms of agonist-specific receptor desensitization, so we hypothesized that increased PDE activity mediates the loss of signaling after PGE2 pretreatment. PGE2 treatment upregulated messenger RNA for PDE3A, PDE3B, PDE4B, and PDE4D and increased overall PDE activity. The PDE4 inhibitor rolipram partially reversed PGE2-mediated desensitization and PDE4 activity was increased, but rolipram did not alter responses to isoproterenol. The PDE3 inhibitor cilostazol had minimal effect. These results show that long-term exposure to PGE2 causes agonist-specific desensitization of EP2 receptor-stimulated cAMP signaling through the increased expression of PDE isozymes, most likely of the PDE4 family.

Dataset Information

Cytosolic phospholipase A2-? participates in lipid body formation and PGE2 release in human neutrophils stimulated with an L-amino acid oxidase from Calloselasma rhodostoma venom.

Publications

Cytosolic phospholipase A<sub>2</sub>-α participates in lipid body formation and PGE<sub>2</sub> release in human neutrophils stimulated with an L-amino acid oxidase from Calloselasma rhodostoma venom.

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Cytosolic phospholipase A₂-? participates in lipid body formation and PGE₂ release in human neutrophils stimulated with an L-amino acid oxidase from Calloselasma rhodostoma venom.