Lymphatic drainage is defective and obstructed by nodules in S1pr1 ^iΔLEC mice. (A and B) FITC-conjugated dextran (2,000 kD) was injected into the muscle layer of the ileum and/or the Peyer’s patches of anesthetized 3-mo-old (A) or 10-mo-old (B) S1pr1^f/f and S1pr1^iΔLEC mice that were treated with TM from P1–7. The flow of fluorescent dye through the mesenteric lymphatic vessels was visualized by live imaging. (A) The dye rapidly drains through the lymphatic vessels in control mice. In contrast, the dye abruptly stopped (white arrow) or appeared to bypass certain locations (yellow arrows) in mutant mice. The graphs show that the distance travelled by the dye and the rate at which the dye travelled were significantly reduced in S1pr1^iΔLEC mice. (B) Time in seconds after injection is indicated on the top right corner of the panels. The dye rapidly drains through the lymphatic vessels in control mice (white arrows). In contrast, the dye accumulated in nodules that were connected to the lymphatic vessels in the mutant mice (white arrowheads). Retrograde flow was also observed between the nodules (yellow arrows). The nodules were observed both in pre-collecting vessels (red arrowheads) and in collecting lymphatic vessels (red arrow). The videos were analyzed to quantify the distance travelled by the dye and the rate of travel. The graphs show that these parameters were significantly reduced in S1pr1^iΔLEC mice. Statistics: Images are representative of (A) n = 3 S1pr1^f/f and n = 4 S1pr1^iΔLEC mice; (B) n = 4 S1pr1^f/f and n = 5 S1pr1^iΔLEC mice. Some samples were analyzed by injection at multiple sites. Each dot in the graph indicates an individual injection. Graphs were plotted as mean ± SEM. Unpaired t tests were performed for the statistical analyses. *P < 0.05; ****P < 0.0001.