Journal: eLife

Article Title: Chitinase 3-like-1 contributes to acetaminophen-induced liver injury by promoting hepatic platelet recruitment

doi: 10.7554/eLife.68571

Figure Lengend Snippet: ( A ) Immunohistochemical (IHC) staining to detect platelets (CD41 + ) in healthy liver biopsies (Normal) and those from patients with AILI (Patient). Scale bar, 250 μm (n = 10/group). ( B ) Male C57B/6 mice treated with PBS or acetaminophen (APAP). Intravital microscopy analyses were performed around 3 hr post-APAP. Mɸs (cyan) and platelets (white) in liver sinusoids (red) are indicated. Representative images were chosen from intravital microscopy videos: https://bcm.box.com/s/15hmtryyrdl302mihrsm034ure87x4ea (Supplementary video 1, PBS treatment) and https://bcm.box.com/s/tuljfmstvv4lvoksx16fkxkpirkekynz (Supplementary video 2; n = 6–7 mice/group, 4–15 videos/mouse). ( C–E ) Male C57B/6 (wild-type [WT]) mice were treated with control IgG (Ctrl IgG) or an anti-CD41 antibody (α-CD41 Ab) either 3 hr before or 3 hr after APAP administration. ( C ) Serum levels of ALT and ( D ) liver histology with necrotic areas outlined were evaluated 24 hr after APAP treatment (n = 5 mice/group in C , D ). Scale bar, 250 μm. ( E ) Male C57B/6 (WT) and Chil1 -/- mice were treated with APAP. Additionally, Chil1 -/- mice were divided into two groups treated with either PBS or recombinant mouse Chi3l1 (rmChi3l1) simultaneously with APAP. Immunofluorescence (IF) staining was performed to detect intrahepatic platelets (CD41 + ) 3 hr after APAP treatment (n = 3 mice/group). Scale bar, 25 μm. Two-tailed, unpaired Student’s t-test was performed in A–C . One-way ANOVA were performed in E .

Article Snippet: Mice were i.p. injected with Ctrl IgG (BD Pharmingen, 559478, 50 μg/mouse) or anti-CD44 antibody (BD Pharmingen, 553131, 50 μg/mouse) in Chil1 -/- reconstituted with rmChi3l1 at 30 min prior to APAP treatment.

Techniques: Immunohistochemical staining, Immunohistochemistry, Intravital Microscopy, Recombinant, Immunofluorescence, Staining, Two Tailed Test

Journal: eLife

Article Title: Chitinase 3-like-1 contributes to acetaminophen-induced liver injury by promoting hepatic platelet recruitment

doi: 10.7554/eLife.68571

Figure Lengend Snippet: Male C57B/6 mice were treated with control IgG (Ctrl IgG) or an anti-CD41 antibody (α-CD41 Ab) either 3 hr before (pre-) or 3 hr after (post-) APAP administration. Immunofluorescence (IF) staining was performed to identify intrahepatic platelets (CD41 + ) (n = 5 mice/group). Scale bar, 25 μm.

Article Snippet: Mice were i.p. injected with Ctrl IgG (BD Pharmingen, 559478, 50 μg/mouse) or anti-CD44 antibody (BD Pharmingen, 553131, 50 μg/mouse) in Chil1 -/- reconstituted with rmChi3l1 at 30 min prior to APAP treatment.

Techniques: Immunofluorescence, Staining

Journal: eLife

Article Title: Chitinase 3-like-1 contributes to acetaminophen-induced liver injury by promoting hepatic platelet recruitment

doi: 10.7554/eLife.68571

Figure Lengend Snippet: ( A–C ) Chil1 -/- mice reconstituted with recombinant mouse Chi3l1 (rmChi3l1) were treated with either Ctrl IgG or α-CD44 Ab 30 min prior to acetaminophen (APAP) challenge. ( A ) Serum levels of ALT and ( B ) liver histology with necrotic areas outlined were evaluated 24 hr after APAP treatment (n = 4–5 mice/group). Scale bar, 250 μm. ( C ) Immunofluorescence (IF) staining was performed to detect intrahepatic platelets (CD41 + ) 3 hr after APAP treatment (n = 3 mice/group). Scale bar, 25 μm. ( D ) Flow cytometry analysis was performed to identify Chi3l1-binding cells among liver non-parenchymal cells (NPCs) isolated from wild-type (WT) mice treated with APAP for 2 hr. CD44 + cells were gated from single live cells. CD44 + cells that bind to rmChi3l1 were further gated. The Chi3l1 + CD44 + cells were then identified by markers for various cell types, including CD45 + CD146 - F4/80 + (Mɸs), CD45 - CD146 + (liver sinusoidal endothelial cells [LSECs]), and Ly6G + (neutrophils). Two-tailed, unpaired Student’s t-test was performed in A .

Article Snippet: Mice were i.p. injected with Ctrl IgG (BD Pharmingen, 559478, 50 μg/mouse) or anti-CD44 antibody (BD Pharmingen, 553131, 50 μg/mouse) in Chil1 -/- reconstituted with rmChi3l1 at 30 min prior to APAP treatment.

Techniques: Recombinant, Immunofluorescence, Staining, Flow Cytometry, Binding Assay, Isolation, Two Tailed Test

Journal: eLife

Article Title: Chitinase 3-like-1 contributes to acetaminophen-induced liver injury by promoting hepatic platelet recruitment

doi: 10.7554/eLife.68571

Figure Lengend Snippet: ( A ) Male WT, Chil1 -/- , Cd44 -/- mice were treated with acetaminophen (APAP) (n = 4 mice/group). After 3 hr, mice were sacrificed and Mɸs 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 ) Mɸs were isolated and mRNA levels of Pdpn in Mɸs 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 Mɸs that express Pdpn were quantified, Scale bar, 25 μm. ( D–F ) Chil1 -/- mice reconstituted with rmChi3l1 were treated with either Ctrl IgG or α-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 Mɸs (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 .

Article Snippet: Mice were i.p. injected with Ctrl IgG (BD Pharmingen, 559478, 50 μg/mouse) or anti-CD44 antibody (BD Pharmingen, 553131, 50 μg/mouse) in Chil1 -/- reconstituted with rmChi3l1 at 30 min prior to APAP treatment.

Techniques: Isolation, Reverse Transcription Polymerase Chain Reaction, Quantitative RT-PCR, Immunofluorescence, Staining, Two Tailed Test

Journal: eLife

Article Title: Chitinase 3-like-1 contributes to acetaminophen-induced liver injury by promoting hepatic platelet recruitment

doi: 10.7554/eLife.68571

Figure Lengend Snippet: Mɸs were isolated from wild-type (WT) mice treated with acetaminophen (APAP) for 3 hr. The cells were treated in vitro with either control IgG (Ctrl IgG) or an anti-podoplanin antibody (α-podoplanin Ab) before incubation with platelets. Immunofluorescence (IF) staining was performed to detect podoplanin on Mɸs and C-type lectin-like receptor 2 (Clec-2) on platelets. Scale bar, 25 μm.

Article Snippet: Mice were i.p. injected with Ctrl IgG (BD Pharmingen, 559478, 50 μg/mouse) or anti-CD44 antibody (BD Pharmingen, 553131, 50 μg/mouse) in Chil1 -/- reconstituted with rmChi3l1 at 30 min prior to APAP treatment.

Techniques: Isolation, In Vitro, Incubation, Immunofluorescence, Staining

Journal: eLife

Article Title: Chitinase 3-like-1 contributes to acetaminophen-induced liver injury by promoting hepatic platelet recruitment

doi: 10.7554/eLife.68571

Figure Lengend Snippet: ( A–C ) Male C57B/6 mice were treated with acetaminophen (APAP) for 3 hr, followed by intraperitoneally ( i.p. ) injection of either a control IgG (Ctrl IgG) or an anti-mouse Chi3l1 Ab (α-mChi3l1 Ab, C59). ( A ) Immunofluorescence (IF) staining for intrahepatic platelets (CD41 + ) was performed 6 hr after APAP treatment (n = 3 mice/group). Scale bar, 25 μm. ( B ) Serum levels of ALT and ( C ) liver histology were evaluated 24 hr after APAP treatment (n = 4–6 mice/group). Scale bar, 250 μm. ( D–F ) Chil1 -/- mice were treated with APAP plus PBS or recombinant human Chi3l1 (rhChi3l1) for 3 hr as indicated and APAP plus rhChi3l1 treatment group were either without treatment or treated with a control IgG (Ctrl IgG) or an anti-human Chi3l1 Ab (α-hChi3l1 Ab, C7). ( D ) IF staining was performed to identify intrahepatic platelets (CD41 + ) 6 hr after APAP treatment. Scale bar, 25 μm. ( E ) Serum levels of ALT and ( F ) liver histology were evaluated 24 hr after APAP treatment. Scale bar, 250 μm (n = 5–10 mice/group in D–F ). Two-tailed, unpaired Student’s t-test was performed in B . One-way ANOVA were performed in E .

Article Snippet: Mice were i.p. injected with Ctrl IgG (BD Pharmingen, 559478, 50 μg/mouse) or anti-CD44 antibody (BD Pharmingen, 553131, 50 μg/mouse) in Chil1 -/- reconstituted with rmChi3l1 at 30 min prior to APAP treatment.

Techniques: Injection, Immunofluorescence, Staining, Recombinant, Two Tailed Test

Journal: eLife

Article Title: Chitinase 3-like-1 contributes to acetaminophen-induced liver injury by promoting hepatic platelet recruitment

doi: 10.7554/eLife.68571

Figure Lengend Snippet:

Article Snippet: Mice were i.p. injected with Ctrl IgG (BD Pharmingen, 559478, 50 μg/mouse) or anti-CD44 antibody (BD Pharmingen, 553131, 50 μg/mouse) in Chil1 -/- reconstituted with rmChi3l1 at 30 min prior to APAP treatment.

Techniques: Recombinant, Diagnostic Assay, Intravital Microscopy

Journal: Oncotarget

Article Title: MiR-155-5p positively regulates CCL17-induced colon cancer cell migration by targeting RhoA

doi: 10.18632/oncotarget.14841

Figure Lengend Snippet: The amount of miR-155-5p and RhoA mRNA was measured in input RNA used for the RIP assays by qRT-PCR. AntagomiR-155-5p decreased relative enrichment of ( A ) miR-155-5p and ( B ) RhoA mRNA in Ago2 immunoprecipitates. Data are expressed as mean ± SEM and n = 4. Data are expressed as fold change compared to anti-IgG ctrl. U1 snRNA and beta actin were used as housekeeping genes for miR-155-5p and RhoA mRNA, respectively. # P < 0.05 versus ctrl-Ab and * P < 0.05 versus anti-Ago2-AntagomiR ctrl treated cells.

Article Snippet: Anti-Ago2 clone 1B1-E2H5 RIP-certified and ctrl-IgG were purchased from (MBL international, Woburn, MA, USA).

Techniques: Quantitative RT-PCR