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| Author(s): |
Rafael Prada Morganti
Total Authors: 1
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| Document type: | Doctoral Thesis |
| Press: | Campinas, SP. |
| Institution: | Universidade Estadual de Campinas (UNICAMP). Faculdade de Ciências Médicas |
| Defense date: | 2008-07-24 |
| Examining board members: |
Edson Antunes;
Ivani Aparecida de Souza;
Lucia Rossetti Lopes;
Jose Luiz Donato;
Marcelo de Carvalho Ramos
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| Advisor: | Edson Antunes |
| Abstract | |
Sepsis is the second major cause of death in intensive care unit. Septic patients show abnormalities in platelet like thrombocytopenia, prolongation in coagulation time, disseminated intravascular coagulation, deep venous thrombosis and bleedings. However, the mechanisms involved in these platelet alterations are not totally clear. The objective of this work was investigate the actions of lipopolysaccharide (LPS) from E. coli and enterotoxins from Staphylococcus aureus (SEA and SEB), in human platelet adhesion and aggregation, likewise the mechanism involved in these phenomenon. The platelets were exposed to LPS, SEA or SEB in different concentrations and times of incubation in vitro evaluations of adhesion and aggregation. Moreover, experiments to elucidate the participation of nitric oxide (NO) and reactive oxygen species in the actions of LPS, SEA and SEB, likewise the intracellular sinalization (calcium mobilization) and activation of fibrinogen receptor glycoprotein IIb/IIIa (GPIIb/IIIa) were carry out. The platelets incubation with LPS, SEA and SEB for 5 to 120 min, inhibited the spontaneous adhesion to fibrinogen and this inhibition was dependent of concentration and time of incubation. The thrombin activated adhesion of platelets was concentration and time dependently also inhibited after incubation with LPS, SEA and SEB for 5 to 120 min. However, the activated platelets inhibition was lower when compared to spontaneous adhesion inhibition. This inhibition was not dependent of intracellular levels increase of cGMP and cAMP. To understand the mechanism of action involved in the platelet adhesion inhibition by LPS, SEA and SEB, the platelets were previously treated with pharmacological agents like: superoxide dismutase (superoxide anion scavenger), superoxide dismutase-polyethylene glycol (superoxide anion intracellular scavenger), catalase-polyethylene glycol (Hydrogen peroxide intracellular scavenger), puromycin and cyclohexamide (protein synthesis inhibitors). Our data suggests that the inhibitory effect of LPS neither involve superoxide anion (O2 -) and hydrogen peroxide (H2O2) increase, nor the formation of neo-proteins, and alterations on activation of glycoprotein IIb/IIIa. In contrast, LPS significantly reduce the external calcium influx, without effecting calcium internal mobilization. With regard to staphylococcal enterotoxin, the protein synthesis and rise in O2 - do not play a role in the platelet inhibition; however the increase of H2O2 intracellular concentration was involved in this response. In conclusion, platelet aggregation and adhesion was inhibited after addition of LPS, concentration and time-dependently, inhibiting calcium influx; similary, incubation with SEA or SEB inhibited platelets adhesion to fibrinogen, by a mechanism involving H2O2 formation (AU) | |