Generally speaking, neurons don't immortalize well because they are postmitotic. Is there a stage of your protocol at which you could isolate the dopaminergic neuronal progenitors? You may even be able to expand these without immortalization.
Tell us more about your differentiation protocol. If there is a step in which you have proliferating cells in SHH and FGF8, I would expect to find neuronal progenitors there.
I agree with Richard, it is impossible to immortalize neurons, these cells are post-mitotic. You can expand the neural progenitor cells in the presence of mitogens (bFGF and/or EGF), but only for limited passages. Late passage neural progenitors have lower potential for neurogenesis.
The "neurons" from adipose-derived stem cells, obviously are not really neurons. The term will need to change for dopaminergic-neural-progenitor . The Neurons are very complexes cells, those cells could be committed to dopaminergic cell as they have expression of ß-tubulin, nestin and hydroxylases and synthesize dopamine; and have maintained the proliferation capacity why I believe that we it will be possible to immortalize.
Hi Katherine-- Long ago, we had good luck immortalizing nestin-positive neural progenitors with the SV40 large T antigen. I would suggest finding an inducible system for T antigen that will allow you to silence the expression when it is time to differentiate to neurons.
Dear Richard Josephson, thank you very much for your message and your interesting information.
Katherine.
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