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Engineering of Synthetic Peptide Substrates for the Expansion and Differentiation of Human Pluripotent Stem Cell-Derived Neural Progenitor Cells


Neurodegenerative diseases, characterized by traumatic or progressive loss of neurons in the brain and spinal, present significant medical and economic challenges to society with fast growing patient populations and yet unavailability of permanent cure-giving therapies. Current drug-based treatments are highly limited in a sense that they merely alleviate symptoms, delay the onset or slow down the progression of the disease at best--even so, with low efficacy and wide variability in therapeutic outcome at the cost of many side effects. With prospect to induce permanent recovery in affected neural functions at molecular level, stem cell therapies based on neural progenitor cell (NPC) transplantation have been extensively performed in animal models of neurodegeneration. However, laminin (LN) substrates commonly used to culture these NPCs may contain animal- derived pathogens and are relatively expensive to produce. In this project, we developed optimal combinations of synthetic peptides that mimick functional sequences of extracellular matrix proteins, on which human embryonic stem cell (hESC) derived NPCs were expanded for 10 passages and differentiated into neurons after 10th passage followed by molecular profiling. Cells cultured on the optimized peptide substrates proliferated at a rate similar to those on the LN substrate, also expressing multipotent NPC markers such as NESTIN, SOX1, and SOX2 at a comparable degree to LN-based culture as verified by gene expression analysis, immunofluorescence staining and flow cytometry. Cells differentiated on the synthetic peptides expressed neuronal markers, MAP2 and B3T, at a similar level to those on LN. Therefore, our synthetic peptide combinations support the long-term expansion and neuronal differentiation of NPCs, providing cheaper and xenogenic contaminant-free alternatives for the LN substrate, and holding promise as a feasible substrate for the large-scale production of NPCs and neurons for cell- based therapies

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