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Explore the effects of beta-amyloid on erythrocyte shape and deformability, impacting oxygen delivery to the brain. Investigate eNOS signaling pathway involvement and associated alterations in erythrocyte morphology.
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MECHANISMS RESPONSIBLE FOR BETA-AMYLOID-DEPENDENT REDUCTION OF ERYTHROCYTE DEFORMABILITYFrancesco MisitiUniversity of Cassino and Southern LazioCassino (Italy)
Conflict of Interest • Francesco Misiti • Has no real or apparent conflicts of interest to report
Erythrocytes and Alzheimer • Ab are associatedwithcerebralbloodvessels • Ab are transferredtoblood • Ab are producedbyplatelets • Bloodcells are exposedtoAb(1-42) and (1-40) • ErythrocytesinteractwithAb(1-42) • Abalterserythrocyteshape and deformability • Abimpairsoxygen delivery tobrain
Effects of Aβ on erythrocyte morphology Amyloid Control
Membrane AChE activity as a marker of membrane integrity * P < 0.05 vs. control
Objective Clarify the roleplayedbyNitricOxidesynthase (eNOS) signalingpathway in the Ab-dependentalterationoferythrocytemorphology • NitricOxide: a regulatory factor of erythrocyte mechanical properties • NitricOxidesynthase (eNOS) isexpressed in erythrocytes • PKC a andCaspase 3:responsible for regulation on eNOS function • PentosePhosphatePathway (PPP):responsibleforerythrocyteantioxidantdefense
Western blot analysis showing the conformational species of Ab Piacentini et al. J Neurochem (2008)
Western blot analysis showing the conformational species of Ab Piacentini et al. J Neurochem (2008)
Immuno-histochemical staining of erythrocyte activated nitric oxide synthase (eNOS) Control Amyloid * P< 0.05 vs. control;
Time course of the effects of Aβ on nitrite and nitrate levels in erythrocytes suspensions * P < 0.05 vs. control; # P < 0.05 vs.Aβ at 24 hours.
Time course of the effects of Aβ on nitrite and nitrate levels in erythrocytes suspensions * P < 0.05 vs. control; # P < 0.05 vs.Aβ at 24 hours.
AChE inhibitors abrogate the effects of Aβ on nitrite and nitrate levels in erythrocytes suspensions * P < 0.05 vs. control; # P < 0.05 vs.Aβ at 24 hours.
PKCaactivationbyAb cytosol (80 kDa) PKCa (80 kDa)PKCa membrane Ctr Ab + PMA + PMA amyloid control
PKCaactivationbyAb cytosol (80 kDa) PKCa (80 kDa)PKCa membrane Ctr Ab + PMA + PMA amyloid control
Caspase 3 activationbyAb : activity Clementi et al. Int J Biochem Cell Biol. 39: 727-735 (2007)
Caspase 3 activation by Ab: WB --32kDa --20 kDa Ctr Ab Misiti et al. Biochem Cell Biol.86: 1-8 (2008)
Band 3 degradation by Caspase 3 --32kDa --20 kDa Ctr Ab CtrAbAb Misiti et al. Biochem Cell Biol.86: 1-8 (2008) Mandal et al. JBC 278: 52551-52558 (2003)
Reduction in antioxidant defense (Pentose Phosphate Pathway) CtrAb Clementi et al. Int. J. Biochem Cell Biol., 36(10):2066-76 (2004) ** P < 0.05 vs.Ab at 24 hours
Ab dependent reduced stimulus for endogenous NO synthesis in the vasculature CtrAb Misiti et al. Biochem Cell Biol.86: 1-8 (2008)
Conclusions Ab/erythrocyteinteractioninduces : • animbalance in the antioxidantdefense (PPP fluxreduction) • Caspase 3 activation • PKCaactivation • eNOSdown-regulation • NO levelsreduction
Conclusions Ab/erythrocyteinteractioninduces : • animbalance in the antioxidantdefense (PPP fluxreduction) • Caspase 3 activation • PKCaactivation • eNOSdown-regulation • NO levelsreduction
Acknowledgements University of Cassino and Southern Lazio- Cassino Cristiana Carelli Alinovi Catholic University-School of Medicine- Rome Bruno Giardina Research National Council (CNR)-ICRM & ISM Beatrice Sampaolese Marco Girasole :
