RNA-seq of acLDL high vs low 4T1-T cells
To determine whether acLDL uptake was associated with specfifc patterns of gene expression we treated vasculogenic mimicry-competent 4T1-T mammary carcinoma cells with Alexa-488 labelled acetylated low density lipoproteins (acLDL) and sorted the cells based on high or low uptake and subjected them to RNA-Seq Overall design: 4T1-T VM cells were incubated with Alexa-488 acLDL followed by FACS. Briefly, cells were trypsinized and resuspended in MACS buffer (1x PBS, 0.5% BSA, 2 mM EDTA) and put through a 40 mm cell strainer. Cells were then sorted using a Sony SH800 cell sorter using the EGFP gate taking the top 25% as high and bottom 25% as low uptake. RNA was extracted from both populations and subjected to RNA-Seq. The experiment was repeated on three seperate occasions.