Abstract
When modeling disease in the laboratory, it is important to use clinically relevant models. Patient-derived human brain cells grown in vitro to study and test potential treatments provide such a model. Here, we present simple, highly reproducible coordinated procedures that can be used to routinely culture most cell types found in the human brain from single neurosurgically excised brain specimens. The cell types that can be cultured include dissociated cultures of neurons, astrocytes, microglia, pericytes and brain endothelial and neural precursor cells, as well as explant cultures of the leptomeninges, cortical slice cultures and brain tumor cells. The initial setup of cultures takes ~2 h, and the cells are ready for further experiments within days to weeks. The resulting cells can be studied as purified or mixed population cultures, slice cultures and explant-derived cultures. This protocol therefore enables the investigation of human brain cells to facilitate translation of neuroscience research to the clinic.
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Data availability
The data in our protocol figures are composed entirely of photomicrographs and electrophysiological recordings displaying the results anticipated with the use of the protocols. Previous uses of the protocols are available in the literature and clearly cited, providing further examples of the types of data that can be obtained from using this protocol.
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Acknowledgements
We are tremendously grateful to the generous neurosurgical brain tissue donors and their families, without whom this research could not be performed; thank you so much. They are the real drivers of this work, and it is because of their special gifts of brain tissue that we can undertake this work for the benefit of future patients. We also thank our collaborators and research technicians: R. Faull, M. Curtis, J. Montgomery, H. Gibbons, M. Cheung, Y. Jung, S.-L. Kim, I. Semenyajenko and M. Eszes from Auckland University; and C. Turner, P. Bergin, E. Walker, L. Roberts and S. Tharakan from Auckland Hospital and LabPlus. This work was supported by the Hugh Green Foundation (funders of the Hugh Green Biobank), a Programme Grant from the Health Research Council of New Zealand, the Sir Thomas and Lady Duncan Trust, the Coker Charitable Trust, and Brain Research New Zealand to M.D. T.I.-H.P., the Centre for Brain Research Douglas Neurosurgical Postdoctoral Research Fellow, is funded by the Douglas Charitable Trust through a School of Medicine Foundation Grant to M.D. (director of the Hugh Green Biobank).
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T.I.-H.P. conceived, optimized and established the neuronal, neural progenitor, microglia, pericyte and endothelial cultures; optimized the tissue-collection process and wrote most of the manuscript. L.C.D.S. conceived, optimized and established the pericyte and endothelial cultures and wrote parts of the manuscript. M.A. conceived, optimized and established the pericyte and mixed glial cultures and wrote parts of the manuscript. Z.R.W. optimized the microglial cultures and wrote parts of the manuscript. J.R. conceived, optimized and established the pericyte and microglial cultures and wrote parts of the manuscript. K.L. conceived, optimized and established the neuronal and brain slice cultures and wrote parts of the manuscript. D.J. conceived, optimized and established the mixed glial and explant cultures. A.S. optimized the mixed glial cultures and inflammatory experiments. S.F. conceived, optimized and established the mixed glial and explant cultures. J.C., P.H. and P.S. are neurosurgeons and were part of the clinical research team; they aided in design and coordination of experiments, consented and provided tissue and optimized the biopsy tissue-collection process. E.M.is a neurosurgeon and was part of the clinical research team. He conceived these projects with M.D. and consented and provided biopsy tissue. M.D.was the study leader and Principal Investigator. He conceived, led and funded the entire program and supervised T.I.-H.P., L.C.D.S., M.A., Z.R.W., J.R., K.L., D.J., A.S. and S.F. in establishing the Hugh Green Biobank.
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M.D. has developed a platform called ‘Neurovalida’ for CNS drug discovery and target validation.
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Nature Protocols thanks Henner Koch, Debomoy K. Lahiri, Jonas Ort and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.
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Key references using this protocol
Park, T. I.-H. et al. Brain Commun. 2, fcaa171 (2020): https://doi.org/10.1093/braincomms/fcaa171
Rustenhoven, J. et al. Sci. Rep. 6, 19371 (2016): https://doi.org/10.1038/srep19371
Jansson, D. et al. Commun. Biol. 4, 260 (2021): https://doi.org/10.1038/s42003-021-01787-x
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Park, T.IH., Smyth, L.C.D., Aalderink, M. et al. Routine culture and study of adult human brain cells from neurosurgical specimens. Nat Protoc 17, 190–221 (2022). https://doi.org/10.1038/s41596-021-00637-8
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DOI: https://doi.org/10.1038/s41596-021-00637-8
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