Neural stem cells-based therapies have shown great potential for central nervous system regeneration, with three-dimensional (3D) culture systems representing a key technique for tissue engineering applications, as well as disease modeling and drug screenings. Self-assembling peptides (SAPs), providing biomimetic synthetic microenvironments regulating cellular functionality and tissue repair, constitute a suitable tool for the production of complex tissue-like structures in vitro. However, one of the most important drawbacks in 3D cultures, obtained via animal-derived substrates and serumrich media, is the reproducibility and tunability of a standardized methodology capable to coax neural differentiation of different human cell lines. In this work we cultured four distinct human neural stem cell (hNSC) lines in 3D synthetic multifunctionalized hydrogel (named HYDROSAP) for up to 6 weeks. Three-dimensional cultures of differentiating hNSCs exhibited a progressive differentiation and maturation over time. All hNSCs-derived neurons in 3D culture system exhibited randomly organized entangled networks with increasing expression of GABAergic and glutamatergic phenotypes and presence of cholinergic ones. Oligodendrocytes formed insulating myelin sheaths positive for myelin basic protein (MBP). In summary, results demonstrated a successfully standardized and reproducible 3D cell culture system for hNSC differentiation and maturation in serum-free conditions useful for future therapies.

NOTCH1 is a relevant gene for outcome in chronic lymphocytic leukemia (CLL), being recurrently targeted by coding mutations deleting the PEST (proline [P], glutamic acid [E], serine [S], threonine [T]) domain (involved in NOTCH1 degradation) and thus generating a more stable protein whose accumulation is associated with poor prognosis. 1 Recently, Puente et al identified one recurrent (chr9:136495700T>C) and two additional less frequent mutations (136495691T>G and 136495693T>C) affecting NOTCH1 3 0-untranslated region (UTR) in 5.6% of cases. 2,3 By causing aberrant splicing events, these variants led to PEST domain deletion and were associated with short time to first treatment (TTFT) and poor overall survival, similarly to coding muta