Homo sapiens
Protocols: Human umbilical vein endothelial cells (HUVEC), purchased from Lonza (Cologne, Germany), were cultured in endothelial basal growth media (EBM) from Lonza, supplemented with hydrocortisone (1 µg/ml), bovine brain extract (12 µg/ml), gentamicin (50 µg/ml) human epidermal growth factor (10 ng/ml) and 10 % fetal calf serum. To simulate an unhealthy diet cells were seeded into cell culture dishes and incubated with the cultivation media for two days. Subsequently cells were treated with media, which was supplemented with either 0.4 mg/ml fructose (con) or 0.4 mg/ml fructose and 1 mg/ml LDL (high LDL). This media was refreshed every second to third day up for 5 to 7 days depending on the experiment. Total RNA-Isolation was performed by using the RNeasy kit from QIAGEN (Hilden, Germany). Cells were lysed using trizol and total RNA was precipitated by adding isopropanol. After centrifugation of this mixture, the liquid phase (RNA) was transferred to a QIAspin column. Subsequently RNA was washed and eluted following the manufacture guidelines. The RNA concentration was measured using a NanoDrop 2000c from Thermo Fisher (Schwerte, Germany) and RNA integrity as well as purity was analyzed by agarose gel electrophoresis. 1. Normalization: Dilution to 200 ng in 50 μL 2. Library Preparation: 50 μL of total RNA sample input. The Library preparation was performed according to the TruSeq® Stranded mRNA Sample Preparation Guide (Part # 15031047 Rev. E) on Sciclone NGS roboter (PerkinElmer).