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Immunomagnetic purification of mES derived neuronal precursors increases neuronal migration and differentiation in vivo

ABSTRACT: All established protocols for differentiation of mouse and human pluripotent stem cells into specific neural subpopulations generate a considerable cellular heterogeneity that hampers experimental and clinical progress. In order to obtain a homogenous population of neuronal precursor cells and to streamline the differentiation of embryonic stem cells (ESCs), we assessed PSA-NCAM, a surface glycoprotein that is specifically expressed on immature neurons. We developed an optimized strategy for magnetic isolation of PSA-NCAM positive neuronal precursors from differentiated ESC cultures and characterized their neuronal differentiation potential in vitro. PSA-NCAM enrichment at an early step of neural differentiation increased the number of ES cell derived neurons and reduced cellular diversity. Gene expression analysis revealed that mainly genes involved in neuronal activity were over-represented after purification. The in vivo potential of in vitro derived PSA-NCAM+ enriched precursors was functionally characterized by grafting into the forebrain of adult mice. Analysis for several neuronal and glia markers at 10 or 40 days post graft showed a distinct differentiation pattern. While unsorted control cells gave rise to a mixed population composed of immature precursors, early postmitotic neurons or glial cells, the majority of PSA-NCAM+ enriched cells differentiated into NeuN positive neurons. Furthermore, when in contact with the rostral migratory stream, higher numbers of cells integrated into the stream and migrated towards the olfactory bulb when the PSA-NCAM enriched population was grafted. Thus, enrichment of neuronal precursors based on PSA-NCAM expression represents a general and straightforward approach to narrow cellular heterogeneity during neuronal differentiation of pluripotent cells. Two conditions (step 4, step 5), each represented by three biological replicates of control and enriched cells (Cy5); mESC was used as common reference (Cy3)

ORGANISM(S): Mus musculus  

SUBMITTER: Angélique Desoeuvre   Josephine Ecklebe  Harold Cremer  Stefan Tomiuk  Dominik Eckardt  Andreas Bosio  Marie-Catherine Tiveron  Serena Barral 

PROVIDER: E-GEOD-35125 | ArrayExpress | 2013-12-21



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Efficient neuronal in vitro and in vivo differentiation after immunomagnetic purification of mESC derived neuronal precursors.

Barral Serena S   Ecklebe Josephine J   Tomiuk Stefan S   Tiveron Marie-Catherine MC   Desoeuvre Angélique A   Eckardt Dominik D   Cremer Harold H   Bosio Andreas A  

Stem cell research 20121102 2

The cellular heterogeneity that is generated during the differentiation of pluripotent stem cells into specific neural subpopulations represents a major obstacle for experimental and clinical progress. To address this problem we developed an optimized strategy for magnetic isolation of PSA-NCAM positive neuronal precursors from embryonic stem cells (ESCs) derived neuronal cultures. PSA-NCAM enrichment at an early step of the in vitro differentiation process increased the number of ES cell derive  ...[more]

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