Characterization of the Early Proliferative Response of the Rodent Bladder to Subtotal Cystectomy: A Unique Model of Mammalian Organ Regeneration

Abstract

Subtotal cystectomy (STC; surgical removal of ∼75% of the rat urinary bladder) elicits a robust proliferative response resulting in complete structural and functional bladder regeneration within 8-weeks. The goal of these studies was to characterize the early cellular response that mediates this regenerative phenomenon, which is unique among mammalian organ systems. STC was performed on eighteen 12-week-old female Fischer F344 rats. At 1, 3, 5 and 7-days post-STC, the bladder was harvested 2-hours after intraperitoneal injection of bromodeoxyuridine (BrdU). Fluorescent BrdU labeling was quantified in cells within the urothelium, lamina propria (LP), muscularis propria (MP) and serosa. Cell location was confirmed with fluorescently co-labeled cytokeratin, vimentin or smooth muscle actin (SMA), to identify urothelial, interstitial and smooth muscle cells, respectively. Expression of sonic hedgehog (Shh), Gli-1 and bone morphogenic factor-4 (BMP-4) were evaluated with immunochemistry. Three non-operated rats injected with BrdU served as controls. Less than 1% of cells in the bladder wall were labeled with BrdU in control bladders, but this percentage significantly increased by 5-8-fold at all time points post-STC. The spatiotemporal characteristics of the proliferative response were defined by a significantly higher percentage of BrdU-labeled cells within the urothelium at 1-day than in the MP and LP. A time-dependent shift at 3 and 5-days post-STC revealed significantly fewer BrdU-labeled cells in the MP than LP or urothelium. By 7-days the percentage of BrdU-labeled cells was similar among urothelium, LP and MP. STC also caused an increase in immunostaining for Shh, Gli-1 and BMP-4. In summary, the early stages of functional bladder regeneration are characterized by time-dependent changes in the location of the proliferating cell population, and expression of several evolutionarily conserved developmental signaling proteins. This report extends previous observations and further establishes the rodent bladder as an excellent model for studying novel aspects of mammalian organ regeneration.

Dynamic reciprocity in cell–scaffold interactions

“…To date, growth factors incorporated into scaffolds have been selected based on known activities in a variety of organ systems. Ongoing studies by a number of groups are characterizing the molecular events that occur both in regenerating tissue following injury [67, 69, 96] and during development [67] to better understand how this knowledge could be exploited for tissue engineering.…”

The Pharmacology of Regenerative Medicine

“…Understanding the mechanisms and characteristics of functional recovery will be a key to designing improved therapeutics for bladder and organ regeneration in the future. (C) Colocalization in cells of incorporated BrdU (bromodeoxyuridine), indicative of proliferation, and specific markers for smooth muscle (SMA, smooth muscle actin) in the muscularis propria (MP) of the regenerating bladder of a female rat [the panel was reproduced from Peyton et al (2012); additional details can be found in the manuscript as well]. Confocal z-stack reconstruction imaging was performed at 600Â magnification, where offset pictures are digitally zoomed.…”

“…A more recent follow up study (Peyton et al, 2012) used fluorescent bromodeoxyuridine (BrdU) labeling to quantify the spatiotemporal characteristics of the proliferative response that mediates the functional regeneration observed by Burmeister et al (2010), as occurs during the critical first week post-STC. In this study, less than 1% of cells in the bladder wall were labeled with BrdU in control bladders under resting conditions (i.e., no damage), but this percentage increased significantly, by 5-to 8-fold, at all time points post-STC.…”

The potential utility of non-invasive imaging to monitor restoration of bladder structure and function following subtotal cystectomy (STC)

“…Several studies have suggested that removal of a large part of the bladder without replacement results in some degree of functional bladder regeneration in both rats [ 17 – 22 ] and humans [ 23 – 32 ]. As such, we have focused our recent efforts on developing rodent models and methods that can provide additional insight into the cellular and molecular mechanisms responsible for functional bladder regeneration in mammals.…”