Perfil agregante em cardíacos em uso do ácido acetil salicílico / Aggregant profile in cardiac patients using acetyl-salicylic acid

As plaquetas atuam na hemostasia, processo esse responsável pelo controle da perda sangüínea em um vaso lesado. O ácido acetil salicílico, um antiagregante plaquetário, tem sido avaliado em cardíacos, quanto a essa atuação, há mais de trinta anos. Os objetivos do presente projeto foram verificar a agregação plaquetária em pacientes cardíacos e hipertensos em uso do ácido acetil salicílico (AAS), nas concentrações de 100 mg/dia e 200 mg/dia, frente a agentes agonistas adenosina difosfato, ácido aracdônico e adrenalina em diferentes concentrações; realizar os mesmos estudos em um grupo considerado normal de doadores de sangue. O grupo analisado foi constituído por 41 pacientes cardíacos e por 40 doadores de sangue, considerado grupo controle. Os resultados evidenciaram que a realização de uma padronização com pool de plasmas frescos de doadores de sangue mostrou ser válida e aplicável para a agregação plaquetária. Ao estabelecermos nossos valores de referência para a agregação plaquetária frente a diferentes agentes agregantes, diferentes concentrações e frente à ausência de agente agregante, foi possível evidenciar que a maioria dos pacientes em estudo em ambas as concentrações de AAS, demonstraram perfil hipoagregante e os pacientes que excepcionalmente não responderam do mesmo modo, muito provavelmente deve-se à variáveis pessoais.

Cytosolic phospholipase A2 is required for macrophage arachidonic acid release by agonists that Do and Do not mobilize calcium. Novel role of mitogen-activated protein kinase pathways in cytosolic phospholipase A2 regulation.

The 85-kDa cytosolic phospholipase A(2) (cPLA(2)) mediates agonist-induced arachidonic acid release and eicosanoid production. Calcium and phosphorylation on Ser-505 by mitogen-activated protein kinases (MAPKs) regulate cPLA(2). Arachidonic acid release and eicosanoid production induced by stimuli that do (A23187, zymosan) or do not (phorbol myristate acetate (PMA), okadaic acid) mobilize calcium were quantitatively suppressed in cPLA(2)-deficient mouse peritoneal macrophages. The contribution of MAPKs to cPLA(2)-mediated arachidonic acid release was investigated. Both extracellular signal-regulated kinases (ERKs) and p38 contributed to cPLA(2) phosphorylation on Ser-505. However, although ERK inhibition did not affect A23187-induced arachidonic acid release, it suppressed zymosan-, PMA-, and okadaic acid-induced arachidonic acid release under conditions where phosphorylation of cPLA(2) on Ser-505 was unaffected. This indicates an additional regulatory mechanism for the ERK pathway. A role for transcriptional regulation is suggested by data showing that cycloheximide and actinomycin D inhibited arachidonic acid release induced by zymosan, PMA and, okadaic acid but not by A23187. Our results show that MAPK pathways contribute to arachidonic acid release in macrophages through alternative mechanisms in addition to their ability to phosphorylate cPLA(2) on Ser-505 and suggest a role for new protein synthesis.

Signal transduction pathways involved in the synergistic stimulation of prostaglandin production by interleukin-1beta and tumor necrosis factor alpha in human gingival fibroblasts.

Accumulating evidence indicates that prostaglandins play an important role in the pathogenesis of periodontal disease. In this study, the effects and interactions between IL-1beta and TNFalpha on prostaglandin production and its regulation were investigated. The cytokines IL-1beta and TNFalpha stimulated prostaglandin E2 (PGE2) and prostacyclin (PGI2) production in gingival fibroblasts. Simultaneous treatment of the cells with IL-1beta and TNFalpha resulted in a synergistic stimulation of PGE2 and PGI2 formation. IL-1beta and, to a lesser extent, TNFalpha stimulated the release of 3H-arachidonic acid (3H-AA), and simultaneous addition of IL-1beta and TNFalpha further increased the release of 3H-AA from pre-labeled gingival fibroblasts. Furthermore, IL-1beta and, to a lesser extent, TNFalpha induced the expression of cyclooxygenase-2 (COX-2) mRNA. Simultaneous addition of IL-1beta and TNFalpha synergistically enhanced COX-2 mRNA levels, accompanied by a corresponding stimulation of PGE2 synthesis. Neither IL-1beta, TNFalpha, nor the combination of these two cytokines affected COX-1 mRNA levels. PMA, known to activate protein kinase C (PKC), enhanced the stimulatory effect of IL-1beta, TNFalpha, and the combination on COX-2 mRNA levels accompanied by a corresponding increase in PGE2 production. The phospholipase A2 (PLA2) inhibitor, BPB, and the PKC inhibitor, BIS, reduced PGE2 production, whereas dexamethasone, indomethacin, and NS-398 completely abolished PGE2 production induced by IL-1beta, TNFalpha, and the combination. The study indicates that the synergistic stimulation of prostaglandin production by IL-1beta, and TNFalpha is mediated partly at the level of COX-2 and partly at the level of PLA2 and that PKC is involved in the signal transduction of the synergy between the two cytokines. The synergy between IL-1beta and TNFalpha may play an important role in the inflammatory processes in gingival tissue in vivo.

Phosphorylation of cytosolic phospholipase A2 and the release of arachidonic acid in human neutrophils.

Kinases mediating phosphorylation and activation of cytosolic phospholipase A2 (cPLA2) in intact cells remain to be fully characterized. Platelet-activating factor stimulation of human neutrophils increases cPLA2 phosphorylation. This increase is inhibited by PD 98059, a mitogen-activated protein (MAP)/extracellular signal-regulating kinase (erk) 1 inhibitor, but not by SB 203580, a p38 MAP kinase inhibitor, indicating that this action is mediated through activation of the p42 MAP kinase (erk2). However, platelet-activating factor-induced arachidonic acid release is inhibited by both PD 98059 and SB 203580. Stimulation by TNF-alpha increases cPLA2 phosphorylation, which is inhibited by SB 203580, but not PD 98059, suggesting a role for p38 MAP kinase. LPS increases cPLA2 phosphorylation and arachidonic acid release. However, neither of these actions is inhibited by either PD 98059 or SB 203580. PMA increases cPLA2 phosphorylation. This action is inhibited by PD 98059 but not SB 203580. Finally, FMLP increases cPLA2 phosphorylation and arachidonic acid release. Interestingly, while the FMLP-induced phosphorylation of cPLA2 is not affected by the inhibitors of the p38 MAP kinase or erk cascades, both inhibitors significantly decrease arachidonic acid release stimulated by FMLP. SB 203580 or PD 98059 has no inhibitory effects on the activity of coenzyme A-independent transacylase.

Hemoglobin A0 and alpha-crosslinked hemoglobin (alpha-DBBF) potentiate agonist-induced platelet aggregation through the platelet thromboxane receptor.

Chemically modified hemoglobins are potential oxygen-carrying blood substitutes, but their in vivo administration has been associated with a variety of unexpected side events, including increased platelet reactivity. We studied the effects of hemoglobin A0 (HbA0) and alpha-crosslinked hemoglobin (alpha-DBBF) on platelets in vitro. Neither hemoglobin A0 nor alpha-DBBF activated platelets when added alone, but both proteins potentiated submaximal agonist-induced platelet aggregation without increasing other markers of platelet activation such as serotonin secretion. Only agonists that are known to cause release of platelet arachidonic acid (AA) were potentiated while aggregation induced by ADP, which does not release AA, was not potentiated. Blockade of the thromboxane receptor with SQ-29,548 prevented the HbA0-induced and the alpha-DBBF-induced potentiation suggesting that the AA/thromboxane signaling pathway mediates the interaction of platelets with hemoglobin.