Fig. 4: TRC network tension increases upon lymph node swelling.

a, In vivo ultraviolet (UV) laser cut measurement of the TRC network at subcapsular IF regions where a high UV laser cuts the TRC network along 10 µm at three z planes after which the local recoil of the TRC network is imaged. The scissor and line indicate the cutting location and arrows indicate the recoiling FRC network. Scale bars, 20 µm. b, Representative example of TRC network recoil. Images depict stills from before (t = −1s), directly after (t = 0 s) and late after (t = 6.2 s) cutting (scale bars, 5 µm), with corresponding kymograph along the recoil axis (scale bar, x axis = 1 s and y axis = 2 µm). Scissor and line indicate cutting location and arrows show the recoiling TRC network. Dashed lines in the kymograph indicate slopes used to calculate the recoil velocity, and the vertical white line indicates the cut. c, Quantification of recoil velocity from kymographs as in b in homeostasis (day 0) and inflammation (days 2, 4, 8 and 14; n = 43, 33, 35, 51 and 36). d, 3D view of the TRC network stained for YAP/TAZ. Stack size, 20 µm. Scale bar, 20 µm. e, Representative examples of YAP/TAZ nuclear and cytoplasmic localization from TRCs of the deep T-zone. NC, nuclear to cytoplasmic. Scale bars, 2 µm. f, Quantification of YAP/TAZ NC fluorescence intensity ratio (n = 46, 19, 46, 48 and 50). Dashed line indicates an equal ratio. Data from c and f are shown as the mean ± s.e.m. where means are connected by a line. Datapoints in c represent single TRC network cuts and in f represent single measured TRCs. Statistical analysis was performed using the Kruskal-Wallis test. All experiments were repeated independently (≥5 lymph nodes from ≥3 mice and ≥2 experiments) and data were pooled for each time point. For statistical details, see Supplementary Table 1. NS, not significant. *P < 0.05, **P < 0.01, ****P < 0.0001.

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