Abstract Details

Title Neuroblastoma Cell Line and Lymphocytes Talk for Cladribine Influenced Apoptosis and Inflammation Pathways in Multiple Sclerosis (MS): An “In Vitro” Study

Topic Multiple Sclerosis

Presentation(s) P2 - Poster Session 2 (5:30 PM-6:30 PM)

Poster/Presentation
Number 15-095

Objective To study neurons response to therapy and to autoreactive cells by evaluating apoptotic and inflammation pathway in Healthy Donors (HDs) and multiple sclerosis (MS) patients (pts)’ lymphocytes and neuroblastoma cell line (SHSY5Y) co-cultures with and without CdA stimulation “in vitro”.

Background MS is characterized by autoreactive lymphocytes and inflammation in the Central Nervous System (CNS). In this context, many therapies act to defeat the mechanism but few of them are able to cross the Blood Brain Barrier (BBB). Cladribine (CdA) is a purine analogue molecule able to pass BBB, but no data available about its effects in CNS and on neurons.

Design/Methods 20 HDs (43,6±20,1; 10F) and 20 naive MS pts (45,5±21,4; 11F) age and sex matched were enrolled. Co-cultures were assessed and stimulated with 20 nM of CdA for many time points (4 and 24 hours); co-cultures control (no CdA) were considered too. Flow cytometric analysis was used to assess apoptosis on both cellular types. Cellular lysates were analyzed for pro and anti-apoptotic molecules (Bax; Smac/DIABLO; Bcl-2; Cyt-c) in Western Blot. Co-culture supernatants (with/without CdA) were examinated for pro and anti- inflammatory cytokines (OPN; IL-6; TNF-α; IFN-γ; IL-4; IL-23) with ELISA test.

Results In vitro experiments showed an increase of MS lymphocytes apoptosis during CdA treatment, if compared to basal cultures (p≤0,0025) and HDs (p≤0,001). Expression of anti-apoptotic proteins was higher in MS compared to HDs (p≤0,0001) and reduced during treatment (p≤0,0001). The same was showed in HDs cells. Anti-inflammatory proteins were increased during CdA treatment in MS lymphocytes co-cultures supernatants if compared to basal (p≤0,001) while pro-inflammatory molecules were reduced (p≤0,0005). Neurons (alone/co-cultures), were not affected by therapy for apoptotic pattern, since pro-apoptotic proteins were not involved.

Conclusions These data suggest that CdA reduces the viability of peripheral and central lymphocytes by apoptosis. No apoptotic signal in neurons was detected.

Authors/Disclosures
PRESENTER	No disclosure on file
	No disclosure on file
	No disclosure on file
	No disclosure on file
	No disclosure on file
	No disclosure on file
Damiano Paolicelli	Damiano Paolicelli has received personal compensation in the range of $500-$4,999 for serving on a Speakers Bureau for Merck Serono. Damiano Paolicelli has received personal compensation in the range of $500-$4,999 for serving on a Speakers Bureau for Sanofi.
Maria Trojano (Policlinico - Bari)	Maria Trojano has received personal compensation in the range of $500-$4,999 for serving on a Scientific Advisory or Data Safety Monitoring board for Biogen. Maria Trojano has received personal compensation in the range of $500-$4,999 for serving on a Scientific Advisory or Data Safety Monitoring board for Merck. Maria Trojano has received personal compensation in the range of $500-$4,999 for serving on a Scientific Advisory or Data Safety Monitoring board for Roche. Maria Trojano has received personal compensation in the range of $500-$4,999 for serving on a Scientific Advisory or Data Safety Monitoring board for Novartis. Maria Trojano has received personal compensation in the range of $500-$4,999 for serving on a Scientific Advisory or Data Safety Monitoring board for Sanofi_Genzyme. Maria Trojano has received personal compensation in the range of $500-$4,999 for serving on a Scientific Advisory or Data Safety Monitoring board for Janssen. Maria Trojano has received personal compensation in the range of $500-$4,999 for serving on a Scientific Advisory or Data Safety Monitoring board for Sandoz. Maria Trojano has received personal compensation in the range of $500-$4,999 for serving on a Speakers Bureau for Biogen. Maria Trojano has received personal compensation in the range of $500-$4,999 for serving on a Speakers Bureau for Merck. Maria Trojano has received personal compensation in the range of $500-$4,999 for serving on a Speakers Bureau for Roche. Maria Trojano has received personal compensation in the range of $500-$4,999 for serving on a Speakers Bureau for Sanofi-Genzyme. Maria Trojano has received personal compensation in the range of $500-$4,999 for serving on a Speakers Bureau for Novartis. Maria Trojano has received personal compensation in the range of $500-$4,999 for serving on a Speakers Bureau for Janssen.

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