Poster Presentations P3 S557

Cerebrolysin as reported. This beneficial effect continued on after the termina-

tion of Cerebrolysin treatment as evidenced by improved water maze where

behavioral performance was still improved after 3 month of suspension of

the Cerebrolysin therapy (age 6 months) in comparison to vehicle-treated con-

trols. However after 6 months of suspension of Cerebrolysin the behavioral

deficits reversed (age 9 months). The amelioration of the behavioral deficits

at in the 0 and 3 months after suspension of CBL therapy correlated with im-

proved synaptic complexity, while after 6 month of suspending the CBL ther-

apy synaptic pathology was detected. Conclusions: The results indicate that

the beneficial effects of Cerebrolysin persist for some time after the cessation

of Cerebrolysin treatment.

P3-358 CD200 NEGATIVELY REGULATES b-AMYLOID-

INDUCED MICROGLIAL ACTIVATION AND

NEUROINFLAMMATION IN VIVO AND IN VITRO

Anthony Lyons, Marina A. Lynch, Trinity College Institute for Neurosci-

ence, Dublin, Ireland. Contact e-mail: lyonsan@tcd.ie

Background: It is becoming increasingly apparent that Alzheimer’s Disease

(AD) is associated with neuroinflammatory changes, typified by activation of

microglial cells and an alteration in the balance between pro- and anti-inflam-

matory cytokines in the brain. It is now known that microglial activation is reg-

ulated by interaction with other cell types but the mechanisms involved are

poorly understood. We have previously demonstrated that CD200-CD200R

interaction is involved in immune homeostasis in the brain. Therefore we

have investigated the anti-inflammatory role of CD200 both in vitro and in

vivo and predicted that knocking out CD200 would result in microglial activa-

tion, while activation of CD200 receptor by CD200Fc would ameliorate in-

flammatory changes induced by b-Amyloid (Ab). Methods: Mixed glial

cultures were prepared from wildtype and CD200-/- mice and expression of

CD40 and CD68 on microglia was assessed by flow cytometry and real time

PCR. Tissue from wildtype and CD200-/- mice was also assessed for differ-

ences in pro-inflammatory markers. Having identified deficits in CD200 as

a hallmark of neuroinflammation, CD200Fc was then investigated as a possible

therapeutic against Ab-induced toxicity. Results: We found significant in-

creases in CD40 and CD68 expression in cultures prepared from CD200-/-

mice relative to wild type mice. Similarly, CD40 and CD68 mRNA was in-

creased in cells prepared from CD200-/- mice. In parallel with these changes,

we observed that CD40 and CD68 mRNA were both increased in hippocampal

tissue prepared from CD200-/- mice and these changes were associated with

increased mRNA expression of the pro-inflammatory cytokines, IL-1b and

TNFa. These data indicate that the absence of CD200 and CD200R activation

resulted in inflammatory changes and this leads to the hypothesis that

CD200Fc, by interacting with CD200R, should ameliorate Ab-induced in-

flammatory changes. We found that Ab significantly increased CD40 and

CD68 and production of proinflammatory cytokines in mixed glial cultures

and that co-incubation with CD200Fc attenuated these Ab-induced changes.

Conclusions: These findings, together with the observation that CD200 is de-

creased in post-mortem brains of Alzheimer’s disease patients, suggest that

methods of modulating CD200 expression may represent a novel therapeutic

strategy for the treatment of Alzheimer’s disease.

P3-359 MITOCHONDRIAL REACTIVE OXYGEN SPECIES

PRECONDITIONING PROTECTS BRAIN CELLS

AGAINST HYPERGLYCEMIA

Sonia C. Correia, Renato X. Santos, Maria S Santos, Paula I. Moreira,

Center for Neuroscience and Cell Biology, Coimbra, Portugal.

Contact e-mail: sonia.s.correia84@gmail.com

Background: Hyperglycemia, a key feature of diabetes mellitus, is associated

withneuronalandvasculardeterioration and ithasbeensuggestedas apredispos-

ing factor for several pathologies including vascular dementia and Alzheimer’s

disease. On the other hand, mitochondria and hypoxia inducible factor-1 alpha

(HIF-1a) have been intimately involved in the protective signaling processes of

preconditioning-induced brain tolerance. Methods: The current study was

aimed to evaluate the role of mitochondrial reactive oxygen species (ROS)

and HIF-1a signaling pathway in cyanide preconditioning-induced protection

against high glucose levels in brain endothelial and neuronal cells. Rat brain

endothelial and human teratocarcinoma (NT2) cells were treated with cyanide

to stimulate ROS production. Results: Cyanide significantly enhanced ROS

levels, particularly hydrogen peroxide (H2O2), without affecting cell viability.

Furthermore, HIF-1a, vascular endothelial growth factor (VEGF) and endothe-

lial nitric oxide synthase (eNOS) protein levels increased following cyanide

treatment. This cyanide- induced immediate preconditioning protected cells

against high glucose-mediated damage by preventing BAX translocation to mi-

tochondria, caspase-3 activation and cell death. High glucose-dependent de-

crease in HIF-1, VEGF, eNOS and glucose transporter 1 (GLUT-1) protein

levels was also prevented by cyanide preconditioning. The free radical scaven-

ger N-acetyl-L-cysteine and the HIF-1a inhibitor 2-methoxyestradiol com-

pletely abolished the protective effects of induced by cyanide preconditioning.

However, cyanide was unable to protect mitochondrial DNA-depleted NT2

(NT2 rho 0) cells against the toxicity induced by high glucose levels. Conclu-

sions: Altogether our results suggest that the protection afforded by cyanide pre-

conditioning involves a crosstalk between mitochondrial ROS and HIF-1a

signaling pathways. Therefore, mitochondria and HIF-1a signaling pathways

could represent feasible therapeutic targets to counteract neuronal and vascular

dysfunction promoted by diabetes and associated complications.

P3-360 MULTI-NUTRIENT SUPPLEMENTATION INDUCES

CHANGES IN SYNAPTIC PROTEIN EXPRESSION IN

PC12 CELLS, RELEVANT TO ALZHEIMER’S

DISEASE

Paul J. M. Savelkoul1, Mandy M. Merkes1, Almar A. Kuipers1,

Amanda J. Kiliaan2, Robert J. J. Hageman1, Laus M. Broersen1,

Patrick J. Kamphuis1, 1Nutricia Advanced Medical Nutrition, Danone

Research, Center of Specialized Nutrition, Wageningen, Netherlands;2Department Anatomy, Donders Centre for Neuroscience, RadboudUniversity Nijmegen Medical Centre, Nijmegen, Netherlands.

Contact e-mail: paul.savelkoul@danone.com

Background: Alzheimer’s disease (AD) is a progressive neurodegenerative

disease leading to a gradual decline in cognitive and behavioural function.

Although the etiology of AD is not yet completely known, it is clear that

in addition to Abeta plaque and tangle formation, loss of dendritic spines

and synaptic connections are a hallmark of AD. Previous work has demon-

strated that several nutrients such as DHA, EPA, uridine monophosphate

(UMP), choline, vitamins B6 and B12, folate, phospholipids, vitamin C

and E, and selenium (combined in Fortasyn�Connect) act synergistically

in improving synapse formation. Recently, this specific combination of nu-

trients was tested in a proof-of-concept study that resulted in an improvement

in memory in patients with mild AD (Scheltens et al, 2010). Methods: To

explore the molecular mechanisms in which these nutrient combinations

stimulate synaptogenesis, we tested their effects on synaptic protein expres-

sion in rat pheochromocytoma cells (PC12). When exposed to nerve growth

factor (NGF), PC12 cells undergo neuronal differentiation characterized by

neurite outgrowth and expression of neuronal proteins. We used this cell

model to test the effects of multi-nutrient supplementation on gene expres-

sion of several proteins involved in synapse formation. Cells were differen-

tiated with NGF and supplemented with or without combinations of PUFAs,

UMP, choline, B-vits, PLs and vitamin C and E and selenium. Results: After

supplementation for 6 days, mRNA was isolated and differences in gene ex-

pression of synaptic proteins were determined using quantitative Real-Time

PCR. Analysis of the RT-PCR data showed an increase in several synaptic pro-

tein mRNAs related to neurite outgrowth and synapse formation. Conclusions:

In conclusion, these data show that supplementation with specific nutrient com-

binations known to increase neurite outgrowth, changes synaptic gene expres-

sion that subsequently could induce synapse formation. In addition, these data

suggest that a multi-nutrient approach might offer an effective method in the

management of AD. Fortasyn is a trademark of N.V. Nutricia

P3-361 ANTI-INFLAMMATORY PROPERTIES OF

LADOSTIGIL AND ITS METABOLITES IN

PRIMARY MICROGLIA.

Rony Panarsky, Marta Weinstock, Hebrew University, Jerusalem, Israel.Contact e-mail: apropo73@hotmail.com