Analysis of CCL21 in murine ear skin and of ICAM-1, VCAM-1, laminin, and fibrillar collagen expression in ear skin explants. (A) Representative whole-mount picture of uninflamed murine ear skin showing intracellular and extracellular CCL21 protein levels (white) in CD31⁺ (red) LYVE-1⁺ (green) capillaries and CD31⁺LYVE-1⁻ collectors. Scale bar, 100 µm. (B) Quantitative PCR analysis of Ccl21 was performed on RNA isolated from FACS-sorted endothelial cells derived from CTR and INF murine ear skin. The same gating scheme as in Fig. S2 A was used for cell isolation (n = 3 experiments; two mice per condition were pooled). (C–K) FACS analysis of ICAM-1 and VCAM-1 expression in single-cell suspensions of freshly isolated CTR ear skin or upon ear skin incubation for 4 h in medium. (C) Depiction of the gating scheme used to identify singlet, live (ZombieAqua⁻) capLECs (CD45⁻ CD31⁺ podo⁺ LYVE-1⁺), colLECs (CD45⁻ CD31⁺ podo⁺ LYVE-1⁻), and BECs (CD45⁻ CD31⁺ podo⁻ LYVE-1⁻). (D and E) Representative FACS plots of ICAM-1 expression (shaded histogram) at 0 h (CTR; D) or after 4 h (E) of incubation. (F and G) Representative FACS plots of VCAM-1 expression at 0 h (CTR; F) and after 4 h (G) of incubation. Shaded histogram, VCAM-1; empty histogram, isotype control. (H and I) Quantification of the delta median fluorescent intensities (ΔMFI: specific-isotype staining) of the experiments performed in F and G. Measurements deriving from the same explant are connected by a line. (J) Summary of MFIs measured for colLECs at time 0 (CTR) and after 4 h of incubation. (K) Summary of the ΔMFIs measured per experiment between colLECs and capLECs. Each dot represents the value from one experiment. (H and I) Paired one-way ANOVA with the Geisser–Greenhouse correction. (J and K) unpaired Student’s t test. *, P < 0.05. cap, capillaries; col, collectors; FSC-A, forward scatter area; FSC-H, forward scatter height; Max, maximum; podo, podoplanin; SSC-A, side scatter area.