During multiple sclerosis (MS), the main axon cystoskeleton proteins, neurofilaments (NF), are altered, and their release into the cerebrospinal fluid correlates with disease severity. The role of NF in the extraaxonal location is unknown. Therefore, we tested whether synthetic peptides corresponding to the tubulin-binding site (TBS) sequence identified on light NF chain (NFL-TBS.40–63) and keratin (KER-TBS.1–24), which could be released during MS, modulate remyelination in vitro. Biotinylated NFL-TBS.40–63, NFL-Scramble2, and KER-TBS.1–54 (1–100 μM, 24 hr) were added to rat oligodendrocyte (OL) and astrocyte (AS) cultures, grown in chemically defined medium. Proliferation and differentiation were characterized by using specific antibodies (A2B5, CNP, MBP, GFAP) and compared with untreated cultures. Lysophosphatidyl choline (LPC; 2 × 10–5 M) was used to induce OL death and to test the effects of TBS peptides under these conditions. NFL-TBS.40–63 significantly increased OL differentiation and maturation, with more CNP+ and MBP+ cells characterized by numerous ramified processes, along with myelin balls. When OL were challenged with LPC, concomitant treatment with NFL-TBS.40–63 rescued more than 50% of OL compared with cultures treated with LPC only. Proliferation of OL progenitors was not affected, nor were AS proliferation and differentiation. NFL-TBS.40–63 peptide induces specific effects in vitro, increasing OL differentiation and maturation without altering AS fate. In addition, it partially protects OL from demyelinating injury. Thus release of NFL-TBS.40–63 caused by axonal damage in vivo could improve repair through increased OL differentiation, which is a prerequisite for remyelination.