Fig. 1

Zone-specific polyploidization in Per-null mouse liver. a–c A representative HE-stained image of liver in WT and Per-null mice (male, 12 weeks old; n = 6 mice for each group). High magnification photos of indicated areas of a are shown in b. Wide view of HE-stained Per-null mouse liver (c) shows clear zone-specific difference of hepatocytes. Hepatocytes encircling the central vein (CV) have larger nuclei than those encircling the portal vessels (PV). Dotted line borders between the two groups. d Zone-specific nuclear sizes stained with Hoechst-33342 of WT and Per-null mice. The bar graph shows the distribution of variant hepatic nuclear size (nuclear square: μm²) along with the CV-PV axis. The CV-PV axis was subdivided into 15 parts (left schema), and the less number indicates the region closed to central vein region. The traditional 3-zone classification of hepatic lobule divides CV-PV axis into three equal parts: namely, centrilobular zone corresponds to 1–5, midlobular zone to 6–10, and periportal zone to 11–15. Note significant increase of nuclear diameter in Per-null hepatocytes than those of WT was observed from part 3 to 9, which distributes both centrilobular and midlobular zones, and thus, we describe these cells to centro-midlobular hepatocytes (CMH) separating from periportal hepatocytes (PH). The results were generated from five mouse livers for each group, and three different lobules were analyzed in each liver. e Frequency distributions of nuclear sizes of WT and Per-null CMH (e) and PH f (n > 2500 cells from 5 mice for each group). g Developmental changes of nuclear sizes of CMH in WT and Per-null liver. The size of Hoechst-33342 stained nuclei was plotted against frequency distributions (n > 2000 cells from 3 mice for each group). Flow sorting of single hepatocytes on the basis of DNA content against cellular area shows dot plot graphs h for gating criteria and histograms i of Hoechst-33342 stained nuclei, showing the DNA content of WT and Per-null hepatocytes at 3 weeks (n = 3) and 12 weeks (n = 4) of age. Data are representative of at least three independent experiments. Two-tailed unpaired Student’s t-test with Welch correction was applied to d; Two-way ANOVA with Bonferroni’s post-test was used to (i). Values represent the mean ± SEM, *P < 0.05, **P < 0.01, ***P < 0.001. Scale bars, 100 μm in a, c, g, 20 μm in b