Down syndrome (DS) patients with early-onset dementia share similar neurodegenerative features with Alzheimer's disease (AD). To recapitulate the AD cell model, DS induced pluripotent stem cells (DS-iPSCs), reprogrammed from mesenchymal stem cells in amniotic fluid, were directed toward a neuronal lineage. Neuroepithelial precursor cells with high purity and forebrain characteristics were robustly generated on day 10 (D10) of differentiation. Accumulated amyloid deposits, Tau protein hyperphosphorylation and Tau intracellular redistribution emerged rapidly in DS neurons within 45 days but not in normal embryonic stem cell-derived neurons. N-butylidenephthalide (Bdph), a major phthalide ingredient of Angelica sinensis, was emulsified by pluronic F127 to reduce its cellular toxicity and promote canonical Wnt signaling. Interestingly, we found that F127-Bdph showed significant therapeutic effects in reducing secreted Aβ40 deposits, the total Tau level and the hyperphosphorylated status of Tau in DS neurons. Taken together, DS-iPSC derived neural cells can serve as an ideal cellular model of DS and AD and have potential for high-throughput screening of candidate drugs. We also suggest that Bdph may benefit DS or AD treatment by scavenging Aβ aggregates and neurofibrillary tangles.
Establishing an efficient differentiation procedure is prerequisite for the cell transplantation of pluripotent stem cells. Activating fibroblast growth factor (FGF) signals and inhibiting the activin/nodal pathway are both conserved principles to direct the neural induction (NI) of developing embryos and human embryonic stem cells (hESCs). Wnt signal and OCT4 expression are critical for the hESC pluripotency; however, their roles in cell differentiation are largely unclear. We demonstrate that in the presence of FGF2 and activin inhibitor SB431542, applying a small-molecule Wnt agonist, BIO, efficiently and rapidly steers the NI of all our tested hESCs. A human induced pluripotent stem cell (iPSC), which is refractory for efficient neural conversion by FGF2, effectively differentiated to SOX1 + cells after the BIO/SB431542/FGF2 treatment. In addition, BIO promoted cell survival and transiently sustained OCT4 expression at the early NI stage with FGF2 and SB431542. Interestingly, at the late NI stage, the OCT4 level rapidly declined in the treated hESCs and consequently initiated the formation of neural rosettes with forebrain neuron characteristics. This study illustrates the distinct effects of Wnt activation on maintaining pluripotency and committing neural lineages at the early and late NI stages of hESCs and iPSCs, respectively.
Pluripotent human embryonic stem cells (hESCs) can be efficiently directed to become immature neuroepithelial precursor cells (NPCs) and functional mature neural cells, including neurotransmitter-secreting neurons and glial cells. Investigating the susceptibility of these hESCs-derived neural cells to neurotrophic viruses, such as Japanese encephalitis virus (JEV), provides insight into the viral cell tropism in the infected human brain. We demonstrate that hESC-derived NPCs are highly vulnerable to JEV infection at a low multiplicity of infection (MOI). In addition, glial fibrillary acid protein (GFAP)-expressing glial cells are also susceptible to JEV infection. In contrast, only a few mature neurons were infected at MOI 10 or higher on the third day post-infection. In addition, functional neurotransmitter-secreting neurons are also resistant to JEV infection at high MOI. Moreover, we discover that vimentin intermediate filament, reported as a putative neurovirulent JEV receptor, is highly expressed in NPCs and glial cells, but not mature neurons. These results indicate that the expression of vimentin in neural cells correlates to the cell tropism of JEV. Finally, we further demonstrate that membranous vimentin is necessary for the susceptibility of hESC-derived NPCs to JEV infection.
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