Mechanism of action of novel NO-releasing furoxan derivatives of aspirin in human platelets

1. Incorporation of a nitric oxide (NO)-releasing moiety in aspirin can overcome its gastric side effects. 2. We investigated the NO-release patterns and antiplatelet effects of novel furoxan derivatives of aspirin (B8 and B7) in comparison to existing antiplatelet agents. 3. Cyclooxygenase (COX) activity was investigated in purified enzyme using an electron paramagnetic resonance-based technique. Concentration–response curves for antiplatelet agents±the soluble guanylate cyclase inhibitor, ODQ (50 μM) were generated in platelet-rich plasma (PRP) and washed platelets (WP) activated with collagen using turbidometric aggregometry. NO was detected using an isolated NO electrode. 4. The furoxan derivatives of aspirin (B8, B7) and their NO-free furazan equivalents (B16, B15; all 100 μM) significantly inhibited COX activity (P<0.01; n=6) in vitro and caused aspirin-independent, cGMP-dependent inhibition of collagen-induced platelet aggregation in WP. B8 was more potent than B7 (PRP IC(50)=0.62±0.1 μM for B8; 400±89 μM for B7; P<0.0001. WP IC(50)s=0.6±0.1 and 62±10 μM, respectively). The NO-free furazan counterparts were less potent antiplatelet agents (WP IC(50)s=54±3 μM and 62±10 μM, respectively; P<0.0001, B8 vs B16). Of the hybrids investigated, only B8 retained antiplatelet activity in PRP. 5. NO release from furoxan–aspirin hybrids was undetectable in buffer alone, but was accelerated in the presence of either plasma or plasma components, albumin (4%), glutathione (GSH; 3 μM) and ascorbate (50 μM), the effects of which were additive for B7 but not B8. NO generation from furoxans was greatly enhanced by platelet extract, an effect that could largely be explained by the synergistic effect of intracellular concentrations of GSH (3 mM) and ascorbate (1 mM). 6. We conclude that the decomposition of furoxan–aspirin hybrids to generate biologically active NO is catalysed by endogenous agents which may instil a potential for primarily intracellular delivery of NO. The blunting of the aspirin effects of furoxan hybrids is likely to be due to loss of the acetyl moiety in plasma; the observed antiplatelet effects are thereby primarily mediated via NO release. Compounds of this class might represent a novel means of inhibiting platelet aggregation by a combination of NO generation and COX inhibition

A Calsequestrin-1 mutation associated with a skeletal muscle disease alters sarcoplasmic Ca2+ release

An autosomal dominant protein aggregate myopathy, characterized by high plasma creatine kinase and calsequestrin-1 (CASQ1) accumulation in skeletal muscle, has been recently associated with a missense mutation in CASQ1 gene. The mutation replaces an evolutionarily-conserved aspartic acid with glycine at position 244 (p.D244G) of CASQ1, the main sarcoplasmic reticulum (SR) Ca2+ binding and storage protein localized at the terminal cisternae of skeletal muscle cells. Here, immunocytochemical analysis of myotubes, differentiated from muscle-derived primary myoblasts, shows that sarcoplasmic vacuolar aggregations positive for CASQ1 are significantly larger in CASQ1-mutated cells than control cells. A strong co-immuno staining of both RyR1 and CASQ1 was also noted in the vacuoles of myotubes and muscle biopsies derived from patients. Electrophysiological recordings and sarcoplasmic Ca2+ measurements provide evidence for less Ca2+ release from the SR of mutated myotubes when compared to that of controls. These findings further clarify the pathogenic nature of the p.D244G variant and point out defects in sarcoplasmic Ca2+ homeostasis as a mechanism underlying this human disease, which could be distinctly classified as "CASQ1-couplonopathy".peer-reviewe

Rassegna di tecniche relative alla depurazione delle acque. Utilizzo di macrofite e microfite nei sistemi di depurazione

Consiglio Nazionale delle Ricerche - Biblioteca Centrale - P.le Aldo Moro, 7, Rome / CNR - Consiglio Nazionale delle RichercheSIGLEITItal

Un manifesto per il rinnovamento della GD

Il contributo, a più voci, articola una proposta per il rinnovamento della Geometria Descrittiva nell'era digitale