Abstract
Purpose :
Proliferative vitreoretinopathy (PVR) is the most common cause of failure of retinal reattachment surgery. The development of PVR occurs following a retinal tear and rhegmatogenous retinal detachment resulting in epiretinal and/or subretinal membrane formation that can redetach the retina and lead to blindness. Determining the specific cell types present in the membranes will allow us to have a better understanding of the disease process and may lead to the development of novel target therapies for the prevention of PVR. In this study, we sought to characterize the cell types and identify genes that are critical to the development of PVR.
Methods :
Four idiopathic epiretinal membranes, 1 subretinal membrane and 1 ciliary body membrane were obtained from patients during surgery and were immediately processed for single cell isolation according to our previously published protocol. Single cell suspension of PVR membranes was prepared according to the protocol as specified by the 10x Genomics sample preparation guide (CG00053) and single cell RNA-seq (scRNA-seq) was performed using 10x Genomics Chromium Single Cell 3' Reagent Kits version 2. The single cell sequencing data were then analyzed using the open-source Seurat kit. The counts matrix was filtered to only include the top 5,000 variable features before clustering.
Results :
The full dataset contains 33,694 features and 4,627 cells. Using the associated marker genes, five cell types were identified as retinal pigment epithelium (RPE), microglia, T cells, B cells and fibroblasts. Remarkably, about 56% of the total cells were categorized as immune cells. Heatmap analysis identified several genes that are involved in PVR development including genes involved in inflammation and epithelial-mesenchymal transition (EMT). Enrichment analysis based on gene ontology pathway analysis suggested that the common differential expressed genes were significantly enriched in oxidative phosphorylation and EMT.
Conclusions :
This study illustrates how scRNA-seq may be used to understand the pathophysiology of PVR. Identification of specific cell types and genes that are critical to PVR development will provide the basis for discovery and development of novel treatments and PVR prevention. Our findings also highlight the role of immune cells in PVR pathobiology.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.