This finding suggests that RUNX1 may play an essential role in EMT. The majority of the cell clusters from PVR membranes showed robust RUNX1 expression levels, which correlated with a shift towards mesenchymal gene expression and away from an epithelial phenotype. ScRNAseq analysis revealed the preponderance of immune cells and cells with fibroblastic phenotypes in PVR membranes. Importantly, most cells that skewed towards a mesenchymal phenotype had higher summed RUNX1 and Snail 1 expression, suggesting potential cooperation between these transcription factors in driving EMT. Only a very small percentage of cells were classified as differentiated RPE suggesting that the majority of cells have undergone epithelial to mesenchymal transition (EMT) or are of immune origin. Clusters 2 and 3 expressed gene sets characteristic of fibroblasts and mesenchymal cells, and less so epithelial cells. This dominant microglial population was found in similar size in the other samples as well. Cluster 1, one of the RUNX1-expressing cell clusters, encompassing 47.1% of all cells, was clearly distinguished as microglial cells due to their robust expression of hallmark genes.
RUNX1 expression was robust in the first two clusters of cells across all three samples. Boston Scientific Corporation, Express LD Iliac/Biliary Premounted Stent System, 316L stainless steel, 0.035, 6 (up to 8 X 37 mm), 7 (up to 10 X 57 mm). Read counts were then loaded into Partek Flow software.įour main clusters of cells defined by differential gene expression were found and, although there was some variability, these clusters accounted for ≥90% of the cells in each sample. Full-Length barcoded cDNA was amplified by polymerase chain reaction to generate sufficient mass for library construction. Lysis and barcoded reverse transcription of RNAs from single cells was performed. 10X Chromium Single Cell solutions can capture between 500 10,000 cells from samples with cell concentrations of 100 2000 cells/uL. Single cells, reagents, and a single Gel Bead containing barcoded oligonucleotides were encapsulated into nanoliter-sized GEMs (Gel Bead-in-Emulsion) using the GemCode platform. This enables novel discovery of gene expression dynamics and molecular profiling of individual cell types in an affordable and simple workflow. After tissue dissociation, PVR cells were submitted for 3’ v2 Single Cell Gene Expression for analysis using the 10x Chromium Platform. Three PVR membranes were obtained from three different patients with grade C PVR undergoing surgery. To characterize the cellular composition of proliferative vitreoretinopathy (PVR) membranes surgically removed from human patients by single-cell genome wide expression profiling and to investigate the role of RUNX1 in PVR.
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