FIGURE 7:

Evolution of lymphoblast membrane surface area during transendothelial migration and cell spreading. (A) Time lapse of a lymphoblast transmigrating between human aortic endothelial cells. Scale bar, 20 μm. Images were taken every 15 s. The projected surface area S_proj is represented in yellow before (leftmost image) and after (rightmost image) transendothelial migration. The pore diameter, D_pore, is estimated by taking the image in which the lymphoblast width is identical above and below the pore (yellow arrows). (B) D_pore during transendothelial migration as a function of the lymphoblast’s projected diameter before transendothelial migration (computed using D₀ = S_proj/2π, and D₀ is an equivalent diameter for a sphere whose projected area is S_proj). (C) Histogram of the duration of transmigration. The mean duration is 3 ± 2 min (mean ± SD). (D) Scanning electron microscopy images of lymphoblasts spreading on a substrate coated with anti-CD3 plus anti-CD28 activating antibodies. Scale bars, 100 μm, 20 μm, 10 μm (left to right). Yellow arrows indicate spread cells. (E) Boxplots of the apparent membrane surface area of T-lymphocytes under both passive (white-filled box) and active (blue-filled box) deformations. The bottom and top of the box indicate the 25th and 75th percentiles, respectively. Red plus signs indicate outliers. From left to right, resting T-lymphocytes initially (column 1, A₀, n = 14) and at rupture (column 2, A*, n = 14) aspirated using a micropipette, lymphoblasts at rest (column 3, A₀, n = 14) and at rupture (column 4, A*, n = 14) aspirated using a micropipette, lymphoblasts spread on anti-CD3 plus anti-CD28 monoclonal antibodies (column 5, aCD3, n = 17), and lymphoblasts after transendothelial migration (column 6, TEM, n = 15). ***p < 0.001. n.s., p > 0.05.