Abstract
This manuscript describes step-by-step procedures to establish and manage fresh and cryopreserved cultures of nerve-derived human Schwann cells (hSCs) at the desired scale. Adaptable protocols are provided to propagate hSC cultures through serial passaging and perform routine manipulations such as enzymatic dissociation, purification, cryogenic preservation, live-cell labeling, and gene delivery. Expanded hSCs cultures are metabolically active, proliferative, and phenotypically stable for at least three consecutive passages. Cell yields are expected to be variable as determined by the rate of growth of individual batches and the rounds of subculture. The purity, however, can be maintained high at >95% hSC regardless of passage. The cells obtained in this manner are suitable for various applications, including small drug screens, in vitro modeling of neurodevelopmental processes, and cell transplantation. One caveat of this protocol is that continued expansion of same-batch hSC populations is eventually restricted due to senescence-linked growth arrest.
| Original language | English |
|---|---|
| Pages (from-to) | e4882 |
| Journal | Bio-protocol |
| Volume | 13 |
| Issue number | 22 |
| DOIs | |
| State | Published - Nov 20 2023 |
| Externally published | Yes |
Bibliographical note
©Copyright : © 2023 The Authors; This is an open access article under the CC BY-NC license.Funding
Natalia Andersen, Ketty Bacallao, and Kaiwen Peng provided expert technical assistance. Gabriela Aparicio assisted with data analysis and figure preparation, Lingxiao Deng with lab support, Valeria Nogueira with illustrations, Thomas Dolan with video editing, and Louise Pay with English editing. Patrick M. Wood and Mary Bunge contributed guidance and critical materials, including hybridoma cell lines and human Schwann cells, during our early investigations. The protocols described here were developed during a >15-year period supported by funding (to P.V.M.) from the National Institutes of Health NIH-NINDS (NS084326), the Craig H Neilsen Foundation, The Miami Project to Cure Paralysis and the Buoniconti Foundation from University of Miami, and the Indiana State Department of Health (grants 33997 and 43547). Grant NIH/NS009923 (to M.B., P.M.W., and P.V.M.) provided seminal funding for cell labeling and banking studies. P.V.M. is currently affiliated to and receives support from the Department of Neurosurgery from UK. The contents of this article are the responsibility of the author and do not necessarily represent the official views of the funding agencies. We are greatly indebted to the generosity of the anonymous individuals who donated their tissues for research.