The leaf essential oil hydrodistilled from Schinus molle grown in Costa Rica was characterised in terms of its chemical composition, antioxidant activity, ability to induce cytotoxicity and the mechanism of cell death involved in the process. As a result, 42 constituents, accounting for 97.2% of the total oil, were identified. The major constituents of the oil were beta-pinene and alpha-pinene. The antioxidant activity showed an IC(50) of 36.3 microg mL(-1). The essential oil was cytotoxic in several cell lines, showing that it is more effective on breast carcinoma and leukemic cell lines. The LD(50) for cytotoxicity at 48 h in K562 corresponded to 78.7 microg mL(-1), which was very similar to the LD(50) obtained when apoptosis was measured. The essential oil did not induce significant necrosis up to 200 microg mL(-1), which together with the former results indicate that apoptosis is the main mechanism of toxicity induced by S. molle essential oil in this cell line. In conclusion, the essential oil tested was weak antioxidant and induced cytotoxicity in different cell types by a mechanism related to apoptosis. It would be interesting to elucidate the role that different components of the oil play in the effect observed here, since some of them could have potential anti-tumoural effects, either alone or in combination.

Human endothelial EA.hy926 cells were incubated with BaP1, a hemorrhagic metalloproteinase purified from Bothrops asper snake venom. Since the first hour of incubation with the proteinase, cells started showing DNA fragmentation, detected by a terminal deoxynucleotidyl transferase-mediated dUDP nick-end labeling (TUNEL)-based photometric enzyme-linked immunosorbent assay (ELISA). At later times, DNA fragments were predominantly located outside the cells, evidencing plasma membrane rupture. DNA fragmentation was completely abolished by Batimastat, a potent inhibitor of metalloproteinase enzymatic activity. Apoptosis induced by BaP1 on endothelial cells was independent of two Bcl-2 family members (anti-apototic Bcl-xL and pro-apoptotic Bax), that did not show any changes in their expression during a 24 h-treatment period. Interestingly, IkappaBalpha, an inhibitor of NFkappaB, decreased after 24 h of treatment, suggesting further activation of the transcription factor. When some elements of the apoptotic extrinsic pathway were assessed, it was observed that procaspase-8 completely disappeared after 24 h of treatment with BaP1, probably indicating its activation by a death receptor, whereas caspase-8 inhibitor, cellular FLICE-inhibitory protein (cFLIP(L)), increased its expression since the first hours of BaP1 incubation. In conclusion, treatment of human endothelial cells with BaP1 induces apoptosis/anoikis, independently of Bcl-2 family members Bax and Bcl-xL and associated with caspase-8 activation and cFLIP(L) up-regulation. Apoptosis was completely dependent on BaP1 enzymatic activity. Similarities between this and other endothelial cell anoikis-related systems suggest that BaP1 and other snake venom metalloproteinases may be useful experimental tools in the study of death-related events that occur when adherent cells loose contact with extracellular matrix.

TitleSynapsin knock-down is associated with decreased neurite outgrowth, functional synaptogenesis impairment and fast high frequency neurotransmitter release. Authors

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