The effect of electrospun scaffolds on the glycosaminoglycan profile of differentiating neural stem cells

Abstract

The use of electrospun scaffolds for neural tissue engineering applications allows a closer mimicry of the native tissue extracellular matrix (ECM), important for the transplantation of cells in vivo. Moreover, the role of the electrospun fiber mat topography on neural stem cell (NSC) differentiation remains to be completely understood. In this work REN-VM cells (NSC model) were differentiated on polycaprolactone (PCL) nanofibers, obtained by wet/wet electrospinning, and on flat glass lamellas. The obtained differentiation profile of NSCs was evaluated using immunofluorescence and qPCR analysis. Glycosaminoglycan (GAG) analysis was successfully emplyed to evaluate changes in the GAG profile of differentiating cells through the use of the highly sensitive liquid chromatography-tandem mass/mass spectrometry (LC-MS/MS) method. Our results show that both culture platforms allow the differentiation of REN-VM cells into neural cells (neurons and astrocytes) similarly. Moreover, LC-MS/MS analysis shows changes in the production of GAGs present both in cell cultures and conditioned media samples. In the media, hyaluronic acid (HA) was detected and correlated with cellular activity and the production of a more plastic extracellular matrix. The cell samples evidence changes in chondroitin sulfate (CS4S, CS6S, CS4S6S) and heparan sulfate (HS6S, HS0S), similar to those previously described in vivo studies and possibly associated with the creation of complex structures, such as perineural networks. The GAG profile of differentiating REN-VM cells on electrospun scaffolds was analyzed for the first time. Our results highlight the advantage of using platforms obtain more reliable and robust neural tissue-engineered transplants.

Keywords: Artificial extracellular matrix; Hyaluronic acid; LC-MS/MS; Neural differentiation; Neural tissue engineering; Polycaprolactone.