15 deoxy δ12 14 prostaglandin j2

Human aortic endothelial cells were obtained from Cascade Biologics – Invitrogen cell culture (Carlsbad, CA, USA) and were grown in EC culture media MV plus growth supplements from Promo Cell (Heidelberg, DE, USA) supplemented with 100 units/ml penicillin and 100 μg/ml streptomycin. For experiments, 80% confluent ECs were incubated in EC basal media from Promo Cell (2% fetal calf serum, 0.4% EC growth supplements, 90 μg/ml heparin and with 100 U/ml penicillin, and 100 μg/ml streptomycin) for 24 h before treatments. THP-1 monocytes were purchased from the LGC Promochem (Middlesex, UK) and were maintained in RPMI 1640 media supplemented with 10% fetal bovine serum, 1% l-glut, and 1% P/S. The cells were passaged twice weekly in complete media at a ratio of 1:5. All cell lines were maintained in an incubator at 37°C and 5% CO₂. For cell counts and viability determination, the cells were diluted 1:1 with trypan blue and counted using a hemocytometer. 15-Deoxy-Δ^12,14-prostaglandin J₂, biotin-15-deoxy-Δ^12,14-prostaglandin J₂, prostaglandin D₂, and biotin-prostaglandin D₂ were purchased from Cayman Chemical (Ann Arbor, MI, USA). To change the solvent for experiments, prostaglandins were dried down and re-suspended in ethanol at a concentration of 1 mg/ml. For experiments, ethanol was used as vehicle control.

Methanol, acetonitrile (ACN), ethyl acetate, water, formic acid (FA) were acquired from Merck Millipore (Warsaw, Poland). Standards of Thromboxane B2, Leukotriene B4, Prostaglandin D2, Prostaglandin E2, 6-keto Prostaglandin F1α, Prostaglandin F2α, 15-deoxy-Δ12,14-Prostaglandin J2, 13,14-dihydro Prostaglandin E1, and their isotope-labeled standards were procured from Cayman Chemical Company (Ann Arbor, MI, USA).