Xylazine rompun

Endothelial cell basal medium and pericyte growth medium were purchased from PromoCell (Heidelberg, Germany). Dulbecco’s modified Eagle’s medium (DMEM) low glucose was from Lonza (Cologne, Germany). Human TNF-α was from Provitro (Berlin, Germany). MA was purchased from Santa Cruz Inc. (Heidelberg, Germany). Fluorescein isothiocyanate-labeled dextran 150,000, rhodamine 6 G and peroxidase-labeled-streptavidin were purchased from Sigma-Aldrich (Munich, Germany). Mayer’s hemalaun solution was from Merck (Darmstadt, Germany). 3-Amino-9-ethylcarbazole was obtained from Abcam (Cambridge, UK). Ketamine (Ursotamin) was from Pharmacia GmbH (Erlangen, Germany) and xylazine (Rompun) was from Bayer (Leverkusen, Germany).

LPS (Escherichia coli, serotype 055: B5) was from Sigma-Aldrich. NF449 (4,4′,4″,4″′-(carbonylbis(imino-5,1,3-benzenetriylbis(carbonylimino))) tetrakis-benzene-1,3-disulphonic acid, octasodium salt) and SAR7334 were provided by Tocris Bioscience. Xylazine (Rompun) and ketamine (Imalgene 1,000) were from Bayer and Merial, respectively. Clodronate liposomes (clodronate 5 mg/mL) were purchased from LIPOSOMA. The allogeneic anti-mouse MHC class I (H-2K^d/H-2D^d) mAb (clone 34-1-2S, IgG2a) was from e-Biosciences or Bio X Cell and FITC-labeled anti-Ly6C (clone AL-21) from BD Biosciences. Rat anti-mouse Ly6G (clone 1A8), A700-labeled anti-CD45 (clone 30-F11) and A647-labeled anti-CD11b (clone M1/70) were from Biolegend. Anti-CCR2 (clone MC-21) was kindly provided by Dr. Matthias Mack (University of Regensburg, Germany)^{20 (link)}.

7–9 weeks old mice were transferred individually to metabolic cages (12:12 h light–dark cycle, 28 ± 1 °C) for three days with free access to water and regular rodent diet (LabDiet® 5001, Forth Worth, TX, USA). After 24 h acclimation, urine samples were collected on days 2 and 3. Subsequently, mice were used for perfusion fixation or direct organ harvest, as described below. Urine was stored at − 80 °C and a protease inhibitor (1:1000, P8340, Protease Inhibitor Cocktail, Sigma Aldrich, Denmark) were added when thawed. Mice used for immunohistochemistry and fluorescent microscopy were anesthetized by i.p. injection with 10 mg/kg Xylazine (Rompun, Bayer Healthcare, Shawnee Mission, KS) and 50 mg/kg Ketamine (Ketalar, Pfizer, Sandwich, Kent, UK). Blood samples were taken by cardiac puncture through the apex before mice were flushed with 1 × PBS and fixed with 4% paraformaldehyde by retrograde perfusion via the left ventricle. Subsequently, mice were fixed for an additional 6 h in paraformaldehyde and transferred to 1 × PBS. Mice used for western blotting were likewise anesthetized, blood samples were taken, and relevant organs were removed and snap-frozen in liquid nitrogen.