Effects of 2-chlorodeoxyadenosine (Cladribine) on microglial cells and astrocytes.

PÉREZ MARÍA JULIA; JUANA M. PASQUINI; AYBAR FLORENCIA; CORREALE JORGE

Estocolmo

Congreso; 35th Congress of ECTRIMS 24th Annual Conference of RIMS; 2019

European Committee for Treatment and Research in Multiple Sclerosis

Background and goals: 2-chlorodeoxyadenosine (CdA, Cladribine) is an immune reconstitution therapy which has recently shown treatment efficacy in MS. Because CdA crosses the BBB it may also exert effects on Central Nervous System (CNS) resident cells. We therefore investigated in vitro effects of CdA on primary microglia and astrocyte cell cultures.Methods: Primary cultures of microglial cells and astrocytes were prepared from neonatal C57BL/6 mice according to the McCarthy and de Vellis protocol. After harvesting, cells were treated with different concentrations of CdA (10 pM to 2 mM) for periods lasting between 6 and 72 hours. Cells were evaluated with and without lypopolisaccharide (LPS) stimulation. Proliferation was measured by 5-bromo-2´-deoxyuridine (BrdU) incorporation, and CdA-induced apoptosis detected by TUNEL assay. TNF-α, IL-1β, IL-6, and IL-10 levels were measured in culture media supernatants using ELISA.Results: CdA treatment of microglial cells in the presence of LPS stimulation inhibited proliferation in a time- and concentration-dependent manner. After 72 hours treatment, at a concentration of 10 µM, CdA inhibited microglial cell proliferation by 53%, and higher concentrations of CdA (200 µM) induced apoptosis in 58% of microglial cells. Furthermore, CdA inhibited IL-1β (37%), IL-6 (67%), and TNF-α (77%) secretion by LPS-activated microglial cells, compared to non-activated cells. IL-10 levels were not modified by the addition of CdA. These effects were only observed in microglial cells, astrocytes were not affected by CdA under any of the conditions studied.Conclusions: The data we present suggests CdA treatment alters several biological functions in microglial cells including: proliferation, apoptosis induction and cytokine release, opening an interesting window of opportunity for use of these cells as potential MS treatment targets.