Directed Migration of Positively Selected Thymocytes Visualized in Real Time
Figure 4
Positive Selection Leads to an Increased Frequency of MRhi Thymocytes Migrating away from the Thymic Capsule
(A) A histogram showing the frequency distribution of average motility rates for positively selecting (blue, P14) and nonselecting (black, 5CC7) transgenic thymocytes compiled from over 1,200 P14 and 875 5CC7 thymocytes from, respectively, four and three independently imaged thymic lobes. Data (from Figure 2A) from wild-type (WT) thymocytes (red) were overlaid for comparison. P14 cells moving at motility rates greater than 13 μm/min were 34% of total imaged thymocytes (Videos S6 and S7) compared to approximately 1% of wild-type cortical thymocytes. Analysis of 5CC7 thymocytes showed nearly complete absence of cells moving at motility rates greater than 13 μm/min, a value that differed significantly (p = 0.002) from wild-type cortical thymocytes. (Video S8)
(B) Image showing trajectories of representative P14 MRhi cells. Note tracks for P14 thymocytes are relatively linear compared to the tracks of wild-type thymocytes (see Figure 1).
(C) Bar graph showing the average displacement per cell moved in each direction over a 3-min time interval (left). Data was computed from more than 100 P14 MRhi cortical thymocytes compiled from four independent experiments. Data from individual runs are shown in Figure S5.
(D) Results of step analysis on 412 P14 thymocytes as a function of motility rate are shown (left). Results of step analysis on 123 5CC7 thymocytes are shown for comparison (right). P14 cells moving at MR greater than 13 μm/min showed strong bias for movement in the –z direction (away from capsule) whereas 5CC7 thymocytes showed random use of both +z and –z directions.