Three-dimensional capillary geometry in gut tissue

Accurate characterization of capillary geometry is of the utmost importance for physiological tissue studies such as oxygen transport. We show that 3D microscopy can be used to measure tissue capillary geometry both in normal and disease states. We imaged fluorescently labeled gut mucosa capillary beds of three control rats and three rats 4 hours after i.p. injection of 9 mg/kg endotoxin. We used serial optical sectioning microscopy coupled with deconvolution to reconstruct 3D capillary geometry. Theoretical point spread functions accounting for depth into the specimen resulted in better reconstructions than experimentally measured point spread functions. We next derived the distribution of the shortest distances to the nearest capillary from all extravascular tissue voxels. In normal rats the shortest-distance distributions were remarkably constant despite widely varying capillary geometry. Furthermore, the mean of the shortest-distance distributions increased significantly for endotoxemic rats (4.8+/-0.4 microm) compared to controls (4.3+/-0.3 microm, P<0.05). Hence, serial sectioning microscopy provides an accurate venue for measuring physiologically relevant 3D capillary structure.