HSPGs glypican‐1 and glypican‐4 are human neuronal proteins characteristic of different neural phenotypes

Generating neurons from human stem cells has potential for brain damage therapy and neurogenesis modeling, but current efficacy is limited by culture heterogeneity and the lack of markers. We have previously reported the heparan sulfate proteoglycans (HSPGs) glypican‐1 (GPC1) and ‐4 (GPC4) as the markers of lineage‐specific human neural stem cells (hNSCs) and mediators of hNSC lineage potential. Here, we further examined phenotypical characteristics and GPC1 and GPC4 during neural differentiation of hNSCs in the presence of two neurogenic growth factors reported to bind to heparan sulfate: brain‐derived neurotrophic factor (BDNF) and platelet‐derived growth factor‐B (PDGF‐B). In hNSC neural cultures, GPC1 and GPC4 were expressed along neurites and cell bodies in long‐term (40–60 days) neural differentiation cultures demonstrating the areas of differential localization—suggesting potentially different functions. Neural differentiation cultures in the presence of BDNF or PDGF‐B generated phenotypically different neural cells with BDNF treatment associated with higher GPC4 versus GPC1 expression, increased heterogeneity, and differential neuron subtype marker expression to PDGF‐B cultures. PDGF‐B cultures exhibited higher levels of spontaneous activity and reduced heterogeneity over long‐term culture associated with decreased GPC4. Untreated neural cultures were highly variable, supporting the use of neuroregulatory growth factors for guided differentiation. Targeted siRNA downregulation of GPC1/4 reduced neural differentiation markers and altered response to exogenous BDNF and PDGF‐B. This work confirms GPC1 and GPC4 as regulators of human neural differentiation and supports their use as novel markers of neural cell characterization. Glypican‐1 (GPC1) and ‐4 (GPC4) have previously been reported as markers of lineage‐specific neural stem cells. Here, their pheonotypical characteristics were further examined under neuronal differentiation conditions in the presence of brain‐derived neurotrophic factor (BDNF) and platelet‐derived growth factor‐B (PDGF‐B). GPC1 and GPC4 demonstrated differential localized expression during long‐term (40–60 days) neuronal differentiation, suggesting different functions within neural cells. Under BDNF conditions, higher GPC4 to GPC1 gene expression ratio correlated with increased heterogeneity, increased proliferation, and increased GABAergic marker expression. In contrast, PDGF conditions resulted in higher GPC1 to GPC4 gene expr