Fig. 2: Fragment-seq application to investigate gene zonation during liver metastasis.

a Schematics of liver microanatomy and fragment zonation approach. Created with BioRender.com. b Differentially expressed genes (DEGs) of LECs in spatially ordered fragments. Left, fragment-seq (1384 fragments across 9 samples). Right, Molecular Cartography (MC) (155 areas across 4 samples). Colored dots represent significantly enriched genes; red, enriched in pericentral zones; yellow, enriched in periportal zones. Gene labels indicate genes significantly enriched in both analyses. p_FDR: false discovery rate adjusted p-value. c Boxplots showing gene expression in LECs of spatially ordered fragments (n = L1–L3: 137, L4: 214, L5: 402, L6: 409, L7: 196, L8–L10: 26 fragments across 9 samples). Black dots represent individual fragments. d Representative images of MC of Plpp1 (red) and Galnt15 (yellow) are shown as an overlay with DAPI signal (white). CV central vein, PV portal vein. e Boxplots of Vcam1 gene expression in Kupffer cells (KCs) of spatially ordered fragments or spatial areas. Left, fragment-seq (like in c) (n = L1–L3: 115, L4: 206, L5: 379, L6: 387, L7: 182, L8–L10: 26 fragments across 9 samples); right, MC comparing pericentral and periportal. f Predicted ligand–receptor (L–R) interactions between KCs and T cells in grouped fragments from pericentral or periportal origin (n = 9 samples). Interaction scores were calculated from grouped fragment-seq data by CellPhoneDB, which uses a permutation test to generate p-values (unadjusted) indicating significantly enriched L–R interactions. Interactions referenced in the main text are highlighted with red squares and white numbers indicate interaction scores. g Representative MC of Vcam1 (purple), Itgb1 (yellow), and Cyp2f2 (red) shown as an overlay with DAPI signal (white). For MC in d, e, and g the complete dataset (as shown in d) was from pericentral n = 89; periportal n = 66 areas across four samples from four separate experiments. For b, c, and e, we used a negative binomial generalized log-linear model (‘glmQLFTest‘ function of edgeR), which uses a (two-sided) empirical Bayes quasi-likelihood F-test. P-values (Benjamini–Hochberg adjusted) of <0.05 were considered significant (***<0.001). For b, c, e, and f the source data are provided as a Source Data file. For all box and whiskers plots the middle line represents the median; the upper and lower lines are the first and third quartile (Q1 and Q3); the whiskers indicate the upper and lower limits of data spread by subtracting 1.5* interquartile range (IQR) from Q1 and adding 1.5* IQR to Q3. L–R: ligand–receptor.