Fig. 2: Mice with CLEC-2 deficient platelets (CLEC1b^fl/flPf4Cre) exhibit reduced hepatic platelet sequestration and enhanced liver healing after a toxic injury.

Wild type (Cre negative) or CLEC-2 deficient (CLEC1b^fl/flPf4Cre) Mice were injected with IP CCl₄(1 mg/kg) or IP APAP (350 mg/kg) and sacrificed 24, 48 or 72 h (IP CCl4) or 6, 24, 48 h (IP APAP) after injection. For uninjured controls, (both wild type and CLEC-2 deficient) mice were injected with mineral oil (MO) or phosphate buffered saline (PBS) alone. a Serum alanine transaminase activity at different time points comparing wild type with CLEC1b^fl/flPf4Cre mice injected with IP APAP (n = 4–8 at point of peak injury). b Serum alanine transaminase activity at the different time points comparing wild type with CLEC1b^fl/flPf4Cre mice injected with IP CCl₄ (n = 4–8). Representative non-injured controls (PBS-APAP, MO-CCl₄) from peak point of injury in each model (APAP-24 h and CCl4-48 h) are shown. c Representative images from formalin fixed liver sections stained with hematoxylin and eosin (48 h after either CCl4 or APAP injection or non-injured controls). Confocal images captured using a Zeiss LSM880 Airyscan detector, set to SR mode and with Zeiss LSM image software; brightfield images acquired at ×10, ×20 or ×40 magnification using a Leica CTR6000 microscope (Leica, UK) with QCapture software, all images representative of at least six livers (scale bar—100 µm). d High magnification (×40) confocal microscopy images of sections from acutely injured mouse livers demonstrating both reduced platelet sequestration in CLEC-2 deficient mice (bottom row) when compared to wild type mice (top row) in pericentral areas (CV-central vein) (DAPI nuclear stain-blue, P-selectin-green, macrophages-F4/80-red, platelets-CD41-purple) (scale bar—200 µm). e Percentage area of P selectin expression measured in murine liver sections (Supplementary Fig. 3) comparing wild type mice with CLEC-2 deficient mice after toxic liver injury. Percentage area measured on eight randomly selected fields from either CLEC-2 deficient or WT mice, measured using image J version 1.8 (National Institutes of Health and the Laboratory for Optical and Computational Instrumentation, University of Wisconsin). Differences assessed using either unpaired Students t test or a Mann–Whitney test *P < 0.05, **P < 0.01, ***P < 0.001. Each data point is representative of an independent animal and data is presented as mean ± SEM.