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

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 Representative immunofluorescence staining with the indicated markers and quantification of TOM+ LECs of E15.5 Csf1r-iCre ; Rosa tdTom dermis (scale bar: 100 μm) from mice with ( Spi1 +/+ ) or without ( Spi1 -/- ) differentiated myeloid cells. The square indicates an area shown at higher magnification in the adjacent panel. The bar plot shows the fraction of TOM+ PROX1+ cells; mean ± SD, n = 5 Spi1 -/- , n = 3 Spi1 +/+ embryos; each dot represents the value from one embryo. B Immunofluorescence staining with the indicated markers of E17.5 Spi1 -/- ; Csf1r-iCre ; Rosa Yfp 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: Immunofluorescence, Staining, Expressing, Marker

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,B Strategy for the combined Csf1r-iCre -mediated lineage tracing and targeting of Prox1 ( A ) and representative immunofluorescence staining with the indicated markers of E15.5 dermis from a heterozygously targeted Csf1r-iCre ; Prox1 fl(Egfp)/+ mouse ( B ) (scale bars: 25 μm). C,D Representative images of E15.5 Prox1 fl/fl embryos with or without Csf1r-iCre (scale bars: 1 mm) ( C ) and table showing the frequency of embryos displaying the indicated phenotype ( D ); n = 20 Prox1 fl/+ , n = 22 Prox1 fl/fl , n = 22 Csf1r-iCre ; Prox1 fl/+ , n = 19 Csf1r-iCre ; Prox1 fl/fl from 11 litters. E Immunofluorescence staining with the indicated markers of E15.5 dermis of Csf1r-iCre ; Rosa tdTom ; Prox1 fl(Egfp)/+ and Csf1r-iCre ; Rosa tdTom ; Prox1 fl(Egfp)/fl(Egfp) littermate mice identifies erythrocytes in mutant dermal lymphatic vessels that are co-labelled for TOM and GFP (scale bars: 100 μm). F Representative immunofluorescence staining with the indicated markers of E15.5 Prox1 fl(Egfp)/fl(Egfp) (no Cre, normal PROX1 function), Csf1r-iCre ; Prox1 fl(Egfp)/+ (heterozygous PROX1 deficiency) and E15.5 Csf1r-iCre ; Prox1 fl(Egfp)/fl(Egfp) (homozygous PROX1 deficiency) dermis illustrates that TER119+ erythrocytes are located in NRP2+ lymphatic vessels of PROX1-deficient embryos. The square indicates an area shown at higher magnification in the adjacent panels and shown for the different markers also in grey scale (Scale bars: 100 μm). Arrows indicate TER119+ erythrocytes in lymphatic vessels.

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: Immunofluorescence, Staining, Mutagenesis

Journal: bioRxiv

Article Title: Mitotic bookmarking by Prox1 preserves mammalian neuronal lineage identity memory via promoting timely H3K27me3 restoration

doi: 10.64898/2026.02.25.707603

Figure Lengend Snippet: ( a ) Representative images of control ( Prox1 fl/fl ) and Prox1 Nes-cKO ( Nestin-CreER T2 ; Prox1 fl/fl ) hippocampus at P120 stained for Prox1 and NeuN (Neuronal nuclei), a pan-neuronal marker. Scale bar, 200 μm. ( b ) Representative swimming paths of the control and Prox1 Nes-cKO mice before and after training in Morris Water Maze test. ( c ) A schematic diagram showing CA and DG neuroepithelium near the ventricular surface, with the neurogenesis process (arrows) producing CA and DG neurons respectively. ( d–f ) Representative images of developing hippocampus at E13 stained for Prox1 and pH3 ( d , f ) and quantifications of the relative Prox1 expression levels from CA to DG ( e ; n =27 brain sections). High magnification of the boxed areas in ( d ) showing the distribution of Prox1 in dividing CA or DG NSCs ( f ). *p < 0.05, ****p < 0.0001; Student’s t-test. Scale bar, 50 μm ( d ); 25 μm ( f ). The cell body of CA and DG NSCs is encircled by dotted lines. ( g , h ) Sample images of CA and DG NSCs in interphase stained with anti-Prox1 antibody and DAPI ( g ) and quantifications of Prox1 foci per NSC ( h ). n = 47, 60 cells; ****p < 0.0001; Student’s t-test. Scale bar, 5 μm. In this and subsequent micrographs, the cell body of CA and DG NSCs is encircled by dashed lines. ( i–k ) Representative images of metaphase (top) and anaphase (bottom) CA (left) and DG (right) NSCs at E13 stained with anti-Prox1, anti-pH3 and DAPI ( i ) and quantifications of Prox1 fluorescent intensity (F.I.) on the chromosomes (chro.) or in the cytosol (cyto.) ( j , k ). Arrowheads indicate Prox1 associating with chromosomes. n = 7, 9, 11, 14 cells, respectively; NS, not significant, ****p < 0.0001, ***p < 0.001; Student’s t-test. Scale bar, 5 μm. ( l ) A schematic diagram showing Prox1 (orange) mitotic retention in DG but not CA NSCs and progenitors.

Article Snippet: Antibodies used in this study include goat anti-Prox1 (1:50, R&D Systems, AF2727), goat anti-Sox2 (1:200, R&D Systems, AF2018), rabbit anti-Prox1 (1:200, Abcam, Ab101851), rat anti-Tbr2 (1:200, Thermo Fisher Scientific, 14-4875-82), rabbit anti-NeuroD1 (1:200, Abcam, Ab213725), rabbit anti-NeuN (1:200, Abcam, Ab177487), rabbit anti-NeuroD2 (1:200, Abcam, Ab104430), rabbit anti-NeuroD6 (1:200, Abcam, Ab85824), rabbit anti-FOXG1 (1:200, Abcam, Ab18259), rat anti-Ctip2 (1:200, Abcam, Ab18465), rabbit anti-Calb2 (Calretinin) (1:200, Abcam, Ab244299) and rabbit anti-Calb1 (Calbindin) (1:200, Abcam, Ab108404).

Techniques: Control, Staining, Marker, Expressing