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Evaluation of two methods to isolate Schwann cells from murine sciatic nerve
Joy E Tomlinson1, Masoud Golshadi1, Christopher J Donahue1
1Cornell University College of Veterinary Medicine, Department of Clinical Sciences, 930 Campus Road, Ithaca, NY, 14853, United States.
Journal of Neuroscience Methods
|November 23, 2019
Summary
Accurate isolation of live Schwann cells (SC) and macrophages after peripheral nerve injury (PNI) is crucial for understanding repair. New flow cytometry methods rapidly enrich these cells without culture, enabling precise gene expression analysis for PNI research.
Area of Science:
- Neuroscience
- Cell Biology
- Regenerative Medicine
Background:
- Schwann cells (SC) and macrophages are vital for peripheral nerve injury (PNI) response.
- Accurate cell isolation is key to understanding PNI repair mechanisms.
- Current methods like culture enrichment alter gene expression, limiting transcriptomic analysis.
Purpose of the Study:
- To develop rapid, culture-free flow cytometry techniques for isolating live SC and macrophages from injured peripheral nerves.
- To enable precise gene expression analysis of SC and macrophages post-PNI.
Main Methods:
- Two flow cytometry techniques were employed for cell isolation.
- Fluorescent activated cell sorting (FACS) using eGFP transgenic SC.
- Isolation by exclusion of non-SC populations from the injury site.
Main Results:
- Successful enrichment of SC was confirmed by gene expression analysis.
- Lineage markers (Map1b, p75-NTR, S100b) were enriched in sorted SC.
- eGFP-based sorting showed enhanced enrichment of mature myelinating genes in SC.
Conclusions:
- The developed flow cytometry methods provide rapid, culture-free enrichment of SC and macrophages from injured nerves.
- These techniques facilitate accurate gene expression analysis for studying PNI.
- eGFP-based sorting offers improved enrichment, potentially capturing specific SC subsets.
