Supplementary Figure 2: LGR4 and LGR5 lineage tracing.
From: The RSPO–LGR4/5–ZNRF3/RNF43 module controls liver zonation and size
(a) Scheme depicting lineage tracing in Lgr4ki/R26-LacZ and Lgr5ki/R26-LacZ mice. (b) LacZ staining in Lgr4ki/R26-LacZ mice showing LGR4 + hepatocytes in the liver (magnified area: parenchyma) after 10 days or 10 months of tracing. (c) LacZ staining in Lgr5ki/R26-LacZ mice showing no LGR5 + hepatocytes in the liver (magnified area: parenchyma) after 10 days or 18 months of tracing. (d) Distribution of Lgr5ki/R26-LacZ + hepatocytes after 10 days or 18 months of tracing. n = 3 mice per group. These data involved assessment of 210 cells (10 days) and 330 cells (18 months) from the indicated mice. (e) Axin2 ISH and β-Gal staining in Lgr5ki/R26-LacZ mice after 10 days of tracing showing Axin2 + /LacZ + hepatocytes. (f) LacZ staining, Axin2 and Lgr5 co-ISH in consecutive liver sections of Lgr5ki/R26-LacZ mice after 10 days of tracing. (g) Percent of hepatocytes expressing Axin2 and Lgr5 in Lgr5ki/R26-LacZ + and Lgr5ki/R26-LacZ- hepatocytes. n = 4 mice. (h) Scheme depicting lineage tracing in Lgr5ki/tdTOM mice 2 days post-PH. (i) tdTOM and Ki67 staining in consecutive liver sections of Lgr5ki/R26-tdTOM mice 2 days post-PH. Arrowheads point at tdTOM + /Ki67 + hepatocytes. (j) tdTOM + /Ki67 + hepatocytes quantified in Lgr5ki/R26-tdTOM mice 2 days post-PH. n = 4 mice. (k) Scheme depicting EdU injections in WT mice. (l) GS and EdU co-staining in control mice. Arrowheads point at EdU + hepatocytes. (m) EdU + hepatocytes quantified in liver zones of control mice. n = 5 mice. (n) Distribution of EdU + hepatocytes in the indicated liver zones. n = 5 mice. These data involved assessment of (d) 210 cells (10 days) and 330 cells (18 months); (g) 1970 LacZ- and 87 LacZ + cells; (j) 14681 tdTOM- and 49 tdTOM + cells; (m) 2650 cells; and (n) 577 cells from the indicated mice. CV, central vein; PV, portal vein. The images in (b,c) and (e,f,i,l) are representative for 3 and 4 mice each, respectively. Data represent mean ± s.d. ns, not significant; two-tailed unpaired t-test (j) and one-way ANOVA with Tukey’s test (m) were used. Scale bars, (b,c,f) 100 μm, (magnifications in b,c) 50 μm, (e,l, magnification in f) 20 μm and (i) 50 μm.
