Met 5a atcc crl 9444

Human endothelial umbilical vein cells (HUVEC), human cardiac microvascular endothelial cells (HCMEC) and the immortalized mesothelial cell line (MeT-5A) were purchased from established vendors (HUVEC and HCMEC from Promocell, Heidelberg, Germany; MeT-5A (ATCC^® CRL-9444™) from LGC Standards, Wesel, Germany). Endothelial cells were grown in endothelial cell growth medium (Promocell, Heidelberg, Germany) with supplements and antibiotics according to the manufacturer’s instructions. MeT-5A were cultured in Medium 199 (M199, 31150022, Gibco, Thermo Fisher Scientific, Waltham, MA, USA) supplemented with 10% (v/v) foetal bovine serum (FBS, Gibco, Thermo Fisher Scientific, Waltham, MA, USA), 1% (v/v) penicillin/streptomycin (P/S, Gibco, Thermo Fisher Scientific, Waltham, MA, USA). Human peritoneal mesothelial cells (HPMC) were isolated from four non-uremic patients and cultured as previously described [11 (link)], as approved by the Ethics Committee of the Medical Faculty, Heidelberg University (S-501/2018). Informed written consent was signed by the patients. The cells were grown in M199 medium supplemented with 10% FBS, 1% penicillin/streptomycin, 0.5 μg/mL insulin, 0.5 μg/mL transferrin, 0.4 μg/mL hydrocortisone and 2 mM L-glutamine (all from Merck, Darmstadt, Germany).

RNA from cell lines was isolated using the miRVana miRNA Isolation Kit (Thermo Fisher Scientific, Darmstadt, Germany) according to the manufacturer’s instructions. Concentration of the isolated RNA was determined using the NanoDrop ND-100 spectrophotometer (Thermo Fisher Scientific). Isolated RNA of the immortalized human mesothelial cell line MeT-5A (ATCC® CRL-9444™) was purchased from tgcBIOMICS GmbH (Bingen, Germany).
RNA from 0.5 ml plasma samples was isolated using the miRVana PARIS kit (Thermo Fisher Scientific) according to the manufacturer’s instructions, modified by adding 5 μl carrier RNA MS2 (Roche, Mannheim, Germany). Amount of free hemoglobin (Hb) in plasma was measured by spectral analysis using a NanoDrop ND-100 spectrophotometer (Thermo Fisher Scientific). Absorbance was measured at 415 nm (total Hb), 450 nm (bilirubin), and 700 nm (sample turbidity). Hemoglobin concentrations were quantified using the formula Hb (mg/dl) = 154.7 x A₄₁₅–130.7 x A₄₅₀–123.9 x A₇₀₀ [12 (link), 13 (link)]. Hemoglobin values (g/l) of the plasma samples used in the subsequent performance analyses are presented in Additional File 3.