CTL motility within pancreatic islets imaged at cellular resolution.
(A–C) Pancreatic islet (green, GFP) and infiltrating CTL (magenta, CFP) populations from 3 individual mice were imaged at the highest digital magnification that would fully encompass β cell mass and associated CTLs. Dimensions: A, w/h = 0.79/d = 5/z = 22; B, w/h = 0.88/d = 5/z = 20; C, w/h = 0.91/d = 5/z = 25. Acquisition times are 10 minutes, 29 minutes, and 14 minutes, respectively, with t = 60 s. A corresponds to Supplemental Video 11. Five representative, high-resolution examples are included for each series that document CTL–β cell interactions (first frame [T1 in min] and last frame of contact [T2 in min]). Supplemental Videos 12 and 13 correspond to magnified zones from A and B, respectively. (D) Enlarged region from C showing a β cell that is almost entirely engulfed by CTLs. (E) Control experiment using mice with fluorescently labeled but antigen-deficient β cells. A small exocrine population of CTLs can be observed. One frame is shown from a 14-minute sequence during which none of the CTLs shown here infiltrated the islet (representative of duplicate control experiments). Dimensions: w/h = 0.75/d = 5/z = 24. (F–I) Kinetic CTL population parameters obtained via 3D cellular-tracking analysis in Imaris. Color coding correlates with the image sequences in A–C. Curves in I were generated via nonlinear regression analysis in GraphPad and, in combination with R2 values, reveal deviation from linearity and thus constrained motility of the intra-islet CTL population. Horizontal bars are means and associated error bars represent the SEM. Scale bars: 80 μm (A–C and E); 10 μm (D).