GSE183320
Single-cell Transcriptome of Human Choroidal Endothelial Cells and Macrophages in Health and Atrophic Macular Degeneration
Purpose: The goals of this study were to identify expression patterns in a large population of human donor choroids, including donors with early atrophic AMD Methods: Independent libraries were prepared for choroidal cells from twenty human donors. Macular samples from donors 1,2,3,5, 11, and 12 were acquired, analyzed, and uploaded in previous experiments (GSE135922 - donors 1,2, 11, and 12; GSE149100- donors 3 and 5). Choroidal punches were gently dissociated in collagenase II for 1 hour before cryopreservation. Thawed cells underwent endothelial cell enrichment by incubating with anti-CD31-microbeads. Final libraries were sequenced on an Illumina NovaSeq system. Sequenced reads were mapped to the human genome build hg38 with CellRanger(v6.0.0) and filtered to remove cells likely to be doublets or cells with a high proportion of mitochondrial reads. Feature selection was performed with the variance stabilizing transformation before integration was performed with Seurat (v4). Results: A total of 30416 cells were recovered after filtering with 5,607,479,828 corresponding reads. A total of 17097 cells were recovered from AMD donor libraries while 13319 cells were recovered from control donors. Clusters were assigned to known choroidal cell types based on expression of distinguishing marker genes. Differential expression analysis identified genes enriched in each population of cells originating from AMD versus control libraries. Particular emphasis was given to choriocapillaris endothelial cells. Conclusions: This study characterizes new gene expression signatures for several choroidal cells, including tissue resident macrophages, inflammatory macrophages, arterioles, and venules. Overall design: mRNA profiles for thousands of cells from the central choroid were generated from four human donor eyes using 10X Genomics Chromium single-cell system followed by sequencing on an Illumina NovaSeq.