Cd62p rb40

To investigate the induction of CSF-1 in vivo, mice were injected i.v. with freshly prepared hemin (8.8–35 μmol/kg body weight), RBC lysate (17.5 μmol heme/kg body weight), or MDP (InvivoGen, 1 mg/kg body weight/d), while PBS (200 μL/20 g body weight) served as a control. In some experiments, hemin was combined with hemopexin at a 1:1 ratio. For the administration of exogenous CSF-1, mice were injected s.c. in the loose skin over the neck with recombinant human CSF-1 (0.5 mg/kg body weight/d, PeproTech) or PBS as a control for 4 consecutive days. To block endogenous CSF activity, mice were injected i.p. with a blocking antibody against CSF-1 (5A1), CSF-2 (MP1-22E9), or isotype control (1 mg/kg body weight, Bio X Cell). For blockade of CMo migration in vivo, mice were treated with Ultra-LEAF (Low Endotoxin, Azide-Free) blocking antibodies against CD62p (RB40.34, BD), CD106 (M/K-2.7, Bio X Cell), E selectin (9A9, Bio X Cell), CD11b (M1/70, Biolegend), ICAM-1 (YN1/1.7.4, Biolegend), or isotype control (Bio X Cell) (5 mg/kg body weight, i.v.). Mice were sacrificed at various time points after treatment, and blood samples and liver tissues were collected for subsequent analysis.

The Animal Care and Use Committee of the Medical University of Vienna and the Austrian Ministry of Sciences approved all performed mouse experiments (BMBWF-66.009/0402-V/3b/2018, BMBWF-66.009/0258-WF/V/3b/2017).
For all in vivo experiments, 8- to 12-week-old C57BL/6J wild-type sex-matched mice were used. Predominantly, female mice were used for IVC ligation as the surgery was optimized for females due to a lower size/fat ratio and to be consistent with the 3R’s (Replacement, Reduction, Refinement) we kept the variability marginal. For experiments with P-selectin inhibition and neutrophil depletion also male mice were included for confirmation and indicated as green points in the figures (Figures S2, S4, and S6). Littermates were randomly assigned to treatment or control groups. The treatment group received the monoclonal rat anti-mouse P-selectin antibody (CD62P, RB40.34, 2.4 µg/g, BD Biosciences), while the control group received rat IgG1, λ isotype control (2.4 µg/g, BD Biosciences) intravenously, both diluted in sterile PBS. We could confirm that this previously published concentration^{20 (link)} was sufficient to block aggregates of platelets with neutrophils and monocytes after ex vivo activation with convulxin, with no additional effect by increasing the dose to 4.0 µg/g (data not shown).