Glycoengineering human neural stem cells (hNSCs) for adhesion improvement using a novel thiol-modified N-acetylmannosamine (ManNAc) analog.

This study sets the stage for the therapeutic use of AcManNTProp, an N-acetylmannosamine (ManNAc) analog that installs thiol-modified sialoglycans onto the surfaces of human neural stem cells (hNSC). First, we compared hNSC adhesion to the extracellular matrix (ECM) proteins laminin, fibronectin, and collagen and found preferential adhesion and concomitant changes to cell morphology and cell spreading for AcManNTProp-treated cells to laminin, compared to fibronectin where there was a modest response, and collagen where there was no observable increase. PCR array transcript analysis identified several classes of cell adhesion molecules that responded to combined AcManNTProp treatment and hNSC adhesion to laminin. Of these, we focused on integrin α6β1 expression, which was most strongly upregulated in analog-treated cells incubated on laminin. We also characterized downstream responses including vinculin display as well as the phosphorylation of focal adhesion kinase (FAK) and extracellular signal-related kinase (ERK). In these experiments, AcManNTProp more strongly induced all tested biological endpoints compared to AcManNTGc, showing that the single methylene unit that structurally separates the two analogs finely tunes biological responses. Together, the concerted modulation of multiple pro-regenerative activities through AcManNTProp treatment, in concert with crosstalk with ECM components, lays a foundation for using our metabolic glycoengineering approach to treat neurological disorders by favorably modulating endpoints that contribute to the viability of transplanted NSCs.