Tracing Cell Movement in the Bladder During Regeneration After Subtotal Cystectomy

Abstract

Introduction and ObjectivesIt has been shown that the bladder has significant regenerative capabilities. In accordance with previous studies, there are differences between these regenerated organs and their original, healthy counterparts. The objective of this study was to determine the bladder regeneration pattern in mice with bladder resections.Methods50% of the bladder was removed in a subtotal cystectomy (STC), performed on 6–10 week female mice. DiI, a red dye, was applied to the suture site to mark the incision location. EdU was injected into STC and sham surgery mice 3 days post‐op. 4 days after injecting the EdU, Thymidine was injected to chase the remaining EdU from the mice. Bladders were fixed in blocks, cryosectioned and stained for analysis at 1, 4, and 8‐week time points. Images were taken on the Zeiss 880 Confocal microscope at 10× objective and tiled together. Using the tiled images, EdU pixel density was analyzed in Adobe Photoshop CS5. The bladders were divided into four cross‐sections: the suture site, both areas adjacent to the suture site, and bladder neck. Within each cross‐section, EdU labeled cells (yellow pixels) were counted and divided by each section's total number of cells (DAPI pixels) to determine the density of EdU per region.ResultsIt was observed that the sham bladder had EdU evenly distributed throughout the bladder. STC bladders originally had most of the EdU in the suture site, however by 4 weeks post‐op, there was a decrease in EdU at the suture site and an increase in the middle and neck areas. Overall EdU decreases over time as it gets metabolized.ConclusionsWe conclude that cell division is a response to STC. Within the 8wk study, labeled cells are migrating with a net movement away from the incision towards the bladder neck.Support or Funding InformationResearch Support Source: The Hartwell Foundation to EM Gong