Scalable Differentiation and Dedifferentiation Assays Using Neuron-Free Schwann Cell Cultures
This chapter describes protocols to establish simplified in vitro assays of Schwann cell (SC) differentiation in the absence of neurons. The assays are based on the capacity of isolated primary SCs to increase or decrease the expression of myelination-associated genes in response to the presence or absence of cell permeable analogs of cyclic adenosine monophosphate (cAMP). No special conditions of media or substrates beyond the administration or removal of cAMP analogs are required to obtain a synchronous response on differentiation and dedifferentiation. The assays are cost-effective and far easier to implement than traditional myelinating SC-neuron cultures. They are scalable to a variety of plate formats suited for downstream experimentation and analysis. These cell-based assays can be used as drug discovery platforms for the evaluation of novel agents controlling the onset, maintenance, and reversal of the differentiated state using any typical adherent SC population.
Key wordsPrimary cultures cAMP In vitro assays Myelin markers Cell morphology Krox-20 c-Jun O1 MAG
The author appreciates the guidance provided by Dr. Patrick Wood and the assistance provided by Drs. Jennifer Soto and Ketty Bacallao in the development of these protocols. James Guest, Kristine Ravelo, and Gonzalo Piñero are acknowledged for performing critical review of the manuscript. The work presented in this chapter was generously supported by the NIH-NINDS (NS084326), The Craig Neilsen Foundation (339576), The Miami Project to Cure Paralysis, and The Buoniconti Fund. The author declares no conflicts of interest with the contents of this article.
- 10.Pinero G, Berg R, Andersen ND, Setton-Avruj P, Monje PV (2016) Lithium reversibly inhibits Schwann cell proliferation and differentiation without inducing myelin loss. Mol Neurobiol. https://doi.org/10.1007/s12035-016-0262-z
- 11.Soto J, Monje PV (2017) Axon contact-driven Schwann cell dedifferentiation. Glia. https://doi.org/10.1002/glia.23131