Journal: Cell proliferation

Article Title: Deletion of Tgm2 suppresses BMP-mediated hepatocyte-to-cholangiocyte metaplasia in ductular reaction.

doi: 10.1111/cpr.13646

Figure Lengend Snippet: FIGURE 4 Activation of BMP signalling promotes the development and maturation of BECs to alleviate cholestasis in Tgm2/ mice. (A) Schematic diagram of the use of rhBMP7 in Tgm2/ mouse after 4-week DDC injury. (B) Western blot assay of pSMAD1/5/8, SMAD1, BMP7, and β-actin in liver extracts from WT and Tgm2/ mice (n = 3/group). (C) Quantification of pSMAD1/5/8 and BMP7 WB signals measured as integrated density using ImageJ (n = 3/group). (D) Co-staining of CK19 and OPN or SSTR2 and co-staining of CK7 and acTUB in mouse liver samples (n = 4/group). Scale bar, 20 μM. (E) Immunohistochemical staining of TGR5 was observed in WT, Tgm2/, and Tgm2/ + rhBMP7 groups after 4-week DDC injury (n = 4/group). Scale bar, 50 μM. (F) Plasma ALP, γ-GT, TBIL, and ALT levels were measured in WT and Tgm2/ mice (n ≥4/group). Comparisons between multiple groups were performed using ordinary one-way ANOVA with Dunnett's multiple comparison test. Significant difference was presented at the levels of *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001.

Article Snippet: Thin sections were stained with primary antibodies GFP (1:100 dilution, Santa Cruz, sc-9996, or Proteintech, 50,430), CK19 (1:100 dilution, Servicebio, GB12197), OPN (1:50 dilution, Servicebio, GB11500), acTUB (1:200 dilution, Sigma, T6793), SSTR2 (1:300 dilution, ABclonal, A15101), TGR5 (1:100 dilution, ELK Biotechnology; ES4906), HNF4α (1:100 dilution, Abcam, ab41898), αSMA (1:100 dilution, Servicebio, GB111364), desmin (1:100 dilution, Servicebio, GB12081), Col1a1 (1:100 dilution, Servicebio, GB11022-3), EpCAM (1:150 dilution, Servicebio, GB1127), CK7 (1:200 dilution, Abcam, ab181598), and Sox9 (1:100 dilution, Millipore, AB5535) and incubated at 4 C for 12–18 h. For double immunohistochemical staining, Alexa Fluor 555 (1:500 dilution, Invitrogen, A-21428) and Alexa Fluor 488 (1:500 dilution, Invitrogen, A11001) were used as secondary antibodies and incubated at room temperature for 1 h. DAPI (Abcam, ab104139) counterstaining was employed to demonstrate nuclei.

Techniques: Activation Assay, Western Blot, Staining, Immunohistochemical staining, Clinical Proteomics, Comparison

Journal: Cell proliferation

Article Title: Deletion of Tgm2 suppresses BMP-mediated hepatocyte-to-cholangiocyte metaplasia in ductular reaction.

doi: 10.1111/cpr.13646

Figure Lengend Snippet: FIGURE 5 Inhibition of BMP signalling suppresses the metaplasia of hepatocytes, thereby affecting the development and maturation of BECs in DR. (A) Schematic diagram of hepatocyte fate tracing after 4-week DDC diet and DMH1 injection in Tgm2CreERT2-R26T/Gf/f mice. (B) Hepatic expression levels of ID1, OPN, and SSTR2 in Tgm2CreERT2-R26T/Gf/f mouse liver (n ≥4/group). (C) ALT and TBIL levels were measured in Tgm2CreERT2-R26T/Gf/f mice (n ≥3/group). (D) Immunofluorescence co-staining of CK19, acTUB, and SSTR2 with the GFP lineage label was observed after 4-week DDC treatment, but lesser co-stained cells were observed after DMH1 injection (n = 4/group). White arrows indicated co-stained cells. Scale bar, 20 μM. (E) The percentage of marker+ cells' (i.e., cells that stained positive for CK19 or SSTR2 or acTUB) fluorescence intensity was determined in GFP lineage label by fluorescence colocalization analysis (n = 4/group). Comparisons between multiple groups were performed using ordinary one-way ANOVA with Dunnett's multiple comparison test. Significant difference was presented at the levels of *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001.

Article Snippet: Thin sections were stained with primary antibodies GFP (1:100 dilution, Santa Cruz, sc-9996, or Proteintech, 50,430), CK19 (1:100 dilution, Servicebio, GB12197), OPN (1:50 dilution, Servicebio, GB11500), acTUB (1:200 dilution, Sigma, T6793), SSTR2 (1:300 dilution, ABclonal, A15101), TGR5 (1:100 dilution, ELK Biotechnology; ES4906), HNF4α (1:100 dilution, Abcam, ab41898), αSMA (1:100 dilution, Servicebio, GB111364), desmin (1:100 dilution, Servicebio, GB12081), Col1a1 (1:100 dilution, Servicebio, GB11022-3), EpCAM (1:150 dilution, Servicebio, GB1127), CK7 (1:200 dilution, Abcam, ab181598), and Sox9 (1:100 dilution, Millipore, AB5535) and incubated at 4 C for 12–18 h. For double immunohistochemical staining, Alexa Fluor 555 (1:500 dilution, Invitrogen, A-21428) and Alexa Fluor 488 (1:500 dilution, Invitrogen, A11001) were used as secondary antibodies and incubated at room temperature for 1 h. DAPI (Abcam, ab104139) counterstaining was employed to demonstrate nuclei.

Techniques: Inhibition, Injection, Expressing, Immunofluorescence, Staining, Marker, Fluorescence, Comparison