Fig 1: Chitinase 3-like-1 (Chi3l1) functions through its receptor CD44.(A) Immunoprecipitation with anti-CD44 antibody was performed using liver homogenates obtained from wild-type (WT) and Cd44-/- mice treated with acetaminophen (APAP) for 2 hr. Input proteins and immune-precipitated proteins were blotted with the indicated antibodies. (B) Interferometry measurement of the binding kinetics of human His-Chi3l1 with human Fc-CD44. (C) His-tagged control GFP and human Chi3l1 were incubated with recombinant human CD44. Proteins bound to Chi3l1 were immune-precipitated with an anti-His antibody. Input proteins and immune-precipitated proteins were blotted with indicated antibodies. (D–F) Male WT mice were treated with APAP and Cd44-/- mice were treated with PBS or recombinant mouse Chi3l1 (rmChi3l1) plus APAP. (D) Serum levels of ALT and (E) liver histology with necrotic areas outlined were evaluated 24 hr after APAP treatment (n = 4–9 mice/group in A, B). Scale bar, 250 µm. (F) Immunofluorescence (IF) staining was performed to detect intrahepatic platelets (CD41+) 3 hr after APAP treatment (n = 3 mice/group). Scale bar, 25 µm. One-way ANOVA were performed in D.
Fig 2: Chitinase 3-like-1 (Chi3l1)/CD44 signaling in M?s upregulates podoplanin expression and platelet adhesion.(A) Male WT, Chil1-/-, Cd44-/- mice were treated with acetaminophen (APAP) (n = 4 mice/group). After 3 hr, mice were sacrificed and M?s were isolated to measure mRNA levels of various adhesion molecules, including selectin P ligand (Selplg), Cd40, melanoma cell adhesion molecule (Mcam), Fc receptor (Fcr), intercellular adhesion molecule 1 (Icam1), lymphocyte function-associated antigen 1 (Lfa1), von Willebrand factor (Vwf), and podoplanin (Pdpn). (B, C) Wild-type (WT) mice were treated with APAP. Chil1-/- and Cd44-/- mice were treated with PBS or rmChi3l1 followed by APAP challenge simultaneously and mice were sacrificed 3 hr after APAP (n = 3 mice/group). (B) M?s were isolated and mRNA levels of Pdpn in M?s were analyzed by quantitative reverse transcription polymerase chain reaction (qRT-PCR). (C) Immunofluorescence (IF) staining of liver sections for podoplanin and F4/80 is shown and the proportions of M?s that express Pdpn were quantified, Scale bar, 25 µm. (D–F) Chil1-/- mice reconstituted with rmChi3l1 were treated with either Ctrl IgG or a-podoplanin Ab for 16 hr and subsequently challenged with APAP. (D) Serum levels of ALT and (E) liver histology were evaluated 24 hr after APAP treatment (n = 6 mice/group). Scale bar, 250 µm. (F) IF staining for intrahepatic platelets (CD41+) and M?s (F4/80+) was performed 3 hr after APAP (n = 3 mice/group). Scale bar, 25 µm. One-way ANOVA were performed in A–C. Two-tailed, unpaired Student’s t-test was performed in D.
Fig 3: Deletion of Chi3l1- or CD44 does not affect liver recovery after APAP-induced injury.Male WT, Chil1-/- and Cd44-/- mice were treated with APAP. Serum ALT levels were measured at 6hrs, 24hrs, 48hrs and 72hrs after APAP treatment (n=5-8 mice/group). One-way ANOVA was performed.
Fig 4: Schematic summary of the main findings.Acetaminophen (APAP) overdose induces chitinase 3-like-1 (Chi3l1) expression, which binds CD44 on M?s and promotes M?s-mediated platelets recruitment through podoplanin/Clec-2 (C-type lectin-like receptor 2) interaction. Recruited platelets further contribute to APAP-induced liver injury (AILI).
Supplier Page from Sino Biological, Inc. for Human CD44 Protein (Fc Tag)