Extended Data Fig. 5: Membrane access of fibroblastic reticular cells is increased through the CLEC-2/PDPN signaling axis.
From: Lymph node homeostasis and adaptation to immune challenge resolved by fibroblast network mechanics
a) Schematic of osmotic shock experiment, inducing swelling by altering osmolarity. b) Representative stills of FRC in isotonic control media. Scale bar, 25 μm. c-d) Time course of c) PDPN T34A FRCs with or without CLEC-2 treatment in hypotonic conditions, and d) time course of PDPN CTRL vs PDPN shRNA KD FRCs in extreme hypotonic conditions (H20). Stills (left) show swelling of cells in hypotonic media. White dotted circle marks initial size before swelling and is compared to swelling at t = 20 (black dotted circle). Scale bar, 25 μm. Change in diameter ratio over time (middle, mean ± SEM). Swelling ratio comparisons between control and c) CLEC-2 treatment or d) PDPN shRNA KD FRCs at 15 min post swelling (orange dotted line, right). Box plot indicates median, interquartile range and minimum/maximum. One-way ANOVA with Tukey’s multiple comparisons. c) n = individual cells where CLEC-2- ISO (n = 42), CLEC-2- HYPO (n = 47), CLEC-2+ ISO (n = 34), CLEC-2+ HYPO (n = 29) analyzed over 5 independent experiments. d) n = individual cells where Control ISO (n = 19), Control H20 (n = 20), PDPN KD ISO (n = 19), PDPN KD H20 (n = 5) analyzed over 5 independent experiments. This figure supports Fig. 4.
