Quantification of Prostanoid Levels in Tissue Samples

Prostanoid levels were measured as previously described [41 (link)] with slight adaptations for the analysis of tissue samples. Briefly, tissues were homogenized by wet bead milling using zirconium oxide beads in 2 mL reinforced tubes (Bertin Instruments, Frankfurt, Germany) and a Precellys 24-Dual with a Cryolys cooling module (Bertin Instruments) as previously described [42 (link)]. After weighing the tissue samples, we added weight-dependent volumes of pre-cooled 25/75% (v/v) ethanol and water supplemented with 10 µM indomethacin to a final tissue concentration of 0.1 mg/µL. We then extracted 200 µL of this homogenate using a combination of protein precipitation and solid-phase extraction for analysis by liquid chromatography-tandem mass spectrometry (LC-MS/MS) (Supplementary Table S7).