A high-throughput single-cell RNA expression profiling method identifies human pericyte markers

AimsWe sought to identify and optimise a universally available histological marker for pericytes in the human brain. Such a marker could be a useful tool for researchers. Further, identifying a gene expressed relatively specifically in human pericytes could provide new insights into the biological functions of this fascinating cell type.MethodsWe analysed single-cell RNA expression profiles derived from different human and mouse brain regions using a high-throughput and low-cost single-cell transcriptome sequencing method called EasySci. Through this analysis, we were able to identify specific gene markers for pericytes, some of which had not been previously characterised. We then used commercially (and therefore universally) available antibodies to immunolabel the pericyte-specific gene products in formalin-fixed paraffin-embedded (FFPE) human brains and also performed immunoblots to determine whether appropriately sized proteins were recognised.ResultsIn the EasySci data sets, highly pericyte-enriched expression was notable for SLC6A12 and SLC19A1. Antibodies against these proteins recognised bands of approximately the correct size in immunoblots of human brain extracts. Following optimisation of the immunohistochemical technique, staining for both antibodies was strongly positive in small blood vessels and was far more effective than a PDGFRB antibody at staining pericyte-like cells in FFPE human brain sections. In an exploratory sample of other human organs (kidney, lung, liver, muscle), immunohistochemistry did not show the same pericyte-like pattern of staining.ConclusionsThe SLC6A12 antibody was well suited for labelling pericytes in human FFPE brain sections, based on the combined results of single-cell RNA-seq analyses, immunoblots and immunohistochemical studies. Pericytes are important brain cells, but existing pericyte markers (e.g. PDGFR beta immunohistochemical staining) are suboptimal for use in human brain sections. We analysed single-cell RNA expression profiles derived from human and mouse brains, and pericyte-enriched expression was notable for SLC6A12 and SLC19A1. In human brain sections, staining with SLC6A12 and SLC19A1 antibodies was far more effective than a PDGFRB antibody at staining pericyte-like cells. The SLC6A12 antibody was particularly well suited for the purpose of labelling pericytes in human brain sections. image

Automated summary

By analyzing single-cell RNA expression profiles derived from different regions of the human and mouse brain, we have identified specific gene markers for pericytes. These gene markers showed better immunolabeling for pericytes in human brain sections compared to existing markers.