Notch ligand-selective activation in CNS demyelination-remyelination

MATHIEU, P.; GOMEZ PINTO, L. ; ALMEIRA GUBIANI, M.F.; ADAMO A.M.

Chicago

Congreso; Neuroscience 2015; 2015

In the CNS, myelination is a physiological process driven by oligodendroglial cells, while demyelination is a pathological process characterized by myelin loss around axons. Demyelination is followed by remyelination, which solves functional deficits.This work focuses on Notch1 ligands and their role in the demyelination-remyelination process in a cuprizone (CPZ)-induced demyelination model. Twenty-one-day-old Wistar rats were fed a diet containing CPZ during 2 weeks. Demyelinated and control animals were sacrificed 7d before CPZ withdrawal (-7d), the day of CPZ withdrawal (0d), 7, 14 and 21d (+7d, +14d and +21d) after CPZ withdrawal and experiments were conducted for each survival time. The levels of F3/contactin and Jagged1 were determined by Western blot in the subventricular zone (SVZ) and corpus callosum (CC) of control and CPZ animals. To evaluate the participation of SVZ in the remyelination process in the CC, primary neurosphere cultures from SVZ dissected from control and CPZ animals were carried out. The cellular populations in these cultures were characterized with specific markers, i.e. Nestin for neural precursor cells, GFAP for astrocytes and NG2 for oligodendroglial progenitor cells (OPCs). Notch signaling activation and ligands F3/contactin and Jagged1 expression were determined.Results show an increase in F3/contactin expression at -7d, +7d and +21d in CC of CPZ animals as compared to controls, whereas in the SVZ the increase in F3/contactin was observed at +14d. In addition, a greater proportion of Nestin+ cells was detected in neurosphere cultures from SVZ of demyelinated and spontaneously remyelinating animals (0d, +7d) as compared to controls, which indicates an increase in OPC proliferation in response to injury. Notch signaling activation (NICD levels) was observed in NG2+ OPCs, together with an increase in the percentage of Jagged1-expressing cells, which suggests that the pool of OPCs remains preserved in these cultures. In turn, the increase observed in the percentage of F3/contactin+ cells and the concomitant decrease in NG2+ OPCs in neurosphere cultures carried out at 0d and +7d might prove F3/contactin participation in inducing OPC maturation. Worth pointing out, F3/contactin was found to colocalize with OPC marker NG2 but went undetected in GFAP+ cells. Finally, co-immunoprecipitation experiments prove F3/contactin binding to Notch, thus confirming its role as a non-canonical ligand.