Journal: bioRxiv
Article Title: A Csf1r lineage gives rise to dermal lymphatic endothelial cells
doi: 10.64898/2026.03.17.712362
Figure Lengend Snippet: A-C Bulk RNA-seq analysis of tdTomato (TOM)+ and TOM- endothelial cells (ECs) from whole E12.5 Csf1r-iCre;Rosa tdTom mouse embryos (dataset GSE117978). A Volcano plot of significantly differentially expressed transcripts with more than 100 counts per transcript; relevant genes are named; grey and red data points represent transcripts with at least twofold over- or under-representation, respectively ( i.e. , red data points are over-represented and grey data points under-represented in TOM+ ECs compared to TOM-ECs). B,C Relative expression levels for markers typical of lymphatic endothelial cells (LECs) ( B ) or tdTomato and Csf1r ( C ); mean ± SD, n = 3 embryos; NS, not significant, *P < 0.05, **P < 0.01 (Benjamini–Hochberg’s multiple comparisons test for P value adjustment, Padj). D RT-qPCR expression analysis for the indicated genes relative to Actb , showing fold change expression in TOM+ ECs relative to TOM- ECs; mean ± SD, n = 3 embryos; *P < 0.05, **P < 0.01 (two-tailed unpaired t-test). E Immunofluorescence staining with the indicated markers of E15.5 Csf1r-iCre ; Rosa tdTom dorsal dermis (scale bar: 200 μm); the square indicates an area shown at higher magnification in the adjacent panels (scale bar: 25 μm), and shown for the different markers also in grey scale. F Immunofluorescence staining with the indicated markers for quantification of TOM+ LECs in E13.5 and E15.5 Csf1r-iCre ; Rosa tdTom dermis (scale bars: 100 μm); the squares indicate areas shown at higher magnification in the adjacent panels. The bar plot shows the fraction of TOM+ PROX1+ cells in E13.5 and E15.5 Csf1r-iCre ; Rosa tdTom dermis. Mean ± SD, n = 4 embryos; each dot represents the value for one embryo. G Immunofluorescence staining with the indicated markers of adult Csf1r-iCre ; Rosa tdTom ear dermis (scale bar: 100 μm). The square indicates an area shown at higher magnification in the adjacent panel, and shown for the different markers also in grey scale. Arrows indicate TOM+ LECs; arrowheads indicate TOM+ macrophages; curved arrows indicate TOM+ BECs; empty symbols indicate lack of expression for the indicated marker.
Article Snippet: Embryonic dorsal dermis was dissected from formaldehyde-fixed embryos and then incubated in PBS containing 2% serum-free protein block (DAKO), 2% bovine serum albumin and 0.4% Triton X-100 before staining with a combination of the following primary antibodies: goat anti-mouse NRP2 (R&D Systems #AF567, 1:100), rabbit anti-mouse PROX1 (Biolegend # 925202, 1:50), goat anti-mouse PROX1 (R&D Systems # AF2727, 1:100), rabbit anti-mouse LYVE1 (Angiobio # 11-034, 1:100), rat anti-mouse PECAM1 (BD Pharmigen # 553370, 1:50), rat anti-mouse EMCN (Santacruz # sc-65495, 1:50), goat anti-mouse FLT4 (R&D Systems # AF743, 1:100), rat anti-mouse TER119 (Biolegend # 116241, 1:100), rabbit anti-mouse GFP (MBL # 598, 1:200), goat anti-mouse TOM (Origene #AB1140-100, 1:250), or rat anti-mouse TOM (Chromotek #5F8, 1:200).
Techniques: RNA Sequencing, Expressing, Quantitative RT-PCR, Two Tailed Test, Immunofluorescence, Staining, Marker