Fig. 6: Deletion of PDPN in vivo attenuates mechanical adaptation of the FRC network in response to immune challenge.
From: Lymph node homeostasis and adaptation to immune challenge resolved by fibroblast network mechanics
a, Tamoxifen and immunization strategy for PdgframGFPΔPDPN mice. Draining LNs (Dr) and non-draining LNs (nDr). b, LN paracortex maximum z-projection. GFP (PdgframGFP, green), PDPN (FRCs, magenta). Scale bars, 50 µm (zoom, 30 µm). c, Representative histograms of surface protein expression for PDPN in control and PdgframGFPΔPDPN steady-state LNs. d, LN mass after CFA/OVA immunization. Box plot indicates median, interquartile range, and minimum/maximum. Two-way ANOVA with Dunnett’s test, *P = 0.030. n indicates LNs on day 0 (n = 10), day 0ΔPDPN (n = 9), day 3 (n = 6), day 3ΔPDPN (n = 5), day 5 (n = 4), and day 5ΔPDPN (n = 4). e, Representative image of GFP+ (green), PDPN− (magenta) FRC cell body in day 5 after immunization in PdgframGFPΔPDPN LNs from two independent experiments. Scale bar, 25 µm. f, Representative images of actomyosin and ECM structures within control and PdgframGFPΔPDPN FRCs 5 days after immunization. Arrowheads and asterisks mark PDPN− and PDPN+ FRCs respectively; perlecan (magenta), PDPN (yellow), F-actin (cyan), and pMLC (red). Scale bars, 10 µm. g, Recoil curves of network displacement (µm) (mean ± SEM) for control and PdgframGFPΔPDPN mice. h, Initial recoil velocity (µm s−1) after CFA/OVA immunization in control and PdgframGFPΔPDPN mice. Box plot indicates median, interquartile range, and minimum/maximum. Two-way ANOVA with Sidak’s multiple comparisons, *P < 0.05, ****P = 1.00E^−6. Each point represents an ablation. i, Laser ablation of the FRC network throughout inflammation in control and PdgframGFPΔPDPN mice. PDGFRα+mGFP+ (FRCs) ablation ROI (white box) and cut site (red dotted line). Scale bars, 50 µm. Recoil displacement (arrowheads) with pre- (green) and postcut (magenta) overlay. Scale bars, 10 µm. g–i, n indicates ablation at day 0 (n = 48), day 0ΔPDPN (n = 44), day 3 (n = 18), day 3ΔPDPN (n = 18), day 5 (n = 28), and day 5ΔPDPN (n = 21) over three independent experiments.
