Cryopreservation of microglia enables single-cell RNA sequencing with minimal effects on disease-related gene expression patterns

Brenda Morsey; Meng Niu; Shetty Ravi Dyavar; Courtney V Fletcher; Benjamin G Lamberty; Katy Emanuel; Anna Fangmeier; Howard S Fox

doi:10.1016/j.isci.2021.102357

Cryopreservation of microglia enables single-cell RNA sequencing with minimal effects on disease-related gene expression patterns

iScience. 2021 Mar 25;24(4):102357. doi: 10.1016/j.isci.2021.102357. eCollection 2021 Apr 23.

Authors

Brenda Morsey¹, Meng Niu², Shetty Ravi Dyavar³, Courtney V Fletcher³, Benjamin G Lamberty¹, Katy Emanuel¹, Anna Fangmeier¹, Howard S Fox¹

Affiliations

¹ Department of Neurological Sciences, University of Nebraska Medical Center, Omaha, NE 68198, USA.
² Department of Genetics, Cell Biology and Anatomy; and Bioinformatics and Systems Biology Core, University of Nebraska Medical Center, Omaha, NE 68198, USA.
³ Antiviral Pharmacology Laboratory, UNMC Center for Drug Discovery, University of Nebraska Medical Center, Omaha, NE 68198, USA.

Abstract

Microglia play a key role in brain development, normal homeostasis, and neurodegenerative disorders. Single-cell technologies have led to important findings about microglia, with many animal model studies using single-cell RNA sequencing (scRNA-seq), whereas most human specimen studies using archived frozen brains for single-nucleus RNA sequencing (snRNA-seq). However, microglia compose a small proportion of the total brain tissue; snRNAseq depletes expression of microglia activation genes that characterize many diseases. Here we examine the use of purified, cryopreserved microglia for scRNA-seq. Comparison of scRNA-seq on paired fresh and cryopreserved microglia from rhesus monkeys revealed a high level of correlation of gene expression between the two conditions. Disease-related genes were relatively unaffected, but an increase in immediate-early gene expression was present in cryopreserved cells. Regardless, changes in immediate-early gene expression are still detectable. Cryopreservation of microglia is a suitable procedure for prospectively archiving samples.

Keywords: Cellular Neuroscience; Molecular Neuroscience; Transcriptomics.

Abstract

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