Abstract: FR-PO933
Role of Fgfr2 in DNA Damage in Bladder Urothelium After Cyclophosphamide Injury
Session Information
- Development, Stem Cells, Regenerative Medicine - II
October 26, 2018 | Location: Exhibit Hall, San Diego Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Development, Stem Cells, and Regenerative Medicine
- 501 Development, Stem Cells, and Regenerative Medicine: Basic
Authors
- Narla, Sridhar Tatarao, University of Pittsburgh, Pittsburgh, Pennsylvania, United States
- Schaefer, Caitlin M., University of Pittsburgh, Pittsburgh, Pennsylvania, United States
- Bushnell, Daniel S., University of Pittsburgh, Pittsburgh, Pennsylvania, United States
- Bates, Carlton M., Children's Hospital of Pittsburgh, Pittsburgh, Pennsylvania, United States
Background
Stimulation of fibroblast growth factor 2 (Fgfr2) (IIb isoform) by keratinocyte growth factor (KGF), reduces injury in many epithelial cells. Deletion of Fgfr2IIIb prolongs cyclophosphamide (CPP)-induced bladder urothelial injury. Our purpose was to identify roles for Fgfr2 in bladder urothelial repair after CPP- injury.
Methods
We first subjected wildtype mice to a 150 mg/kg intraperitoneal (IP) dose of CPP to characterize the injury/ repair and to determine expression of Fgfr2 after injury. To determine roles of Fgfr2 in CPP-induced injury, we used a Tamoxifen inducible ShhCre line to knockout Fgfr2 in all urothelial layers (ShhcreFgfr2KO), and subjected Cre-negative and knockout mice to IP CPP. We performed general histological staining, in situ hybridization, and immunostaining.
Results
In controls, we observed major loss of urothelium, hemorrhage, and inflammation 1-day post CPP. We noted urothelial hyperplasia from 3-10 days post injury. In situ hybridization showed increased Fgfr2 expression at 1 and 3 days post injury in control urothelium. We also observed evidence of DNA damage by γH2AX immunostaining in control urothelium at 3-7 days, which was reduced by 10 days. In the knockout studies, we observed significant decreases in weight at 1 and 3 days post-CPP in ShhcreFgfr2KO vs. Cre-negative mice. We noted similar urothelial loss, hemorrhage and inflammation 1-day post CPP in Cre-negative and mutant bladders. At 3 days post injury, ShhcreFgfr2KO bladders had ongoing marked hemorrhage and inflammation with reduced uroplakin staining vs. Cre-negative mice. While mutants and Cre-negative mice had expansion of Keratin 14 (Krt14)-positive presumptive progenitor cells across basal layers 3 days-post CPP, Cre negative mice had 6-7 urothelial cell layers while ShhcreFgfr2KO had only 3-4 cell layers. Also, mutants had many (mostly Krt14+) cells that were hypertrophic with enlarged nuclei (suggesting a cell cycle block). Furthermore, mutants had 2-fold increases in γH2AX+ nuclei vs. Cre negative mice.
Conclusion
Together, Fgfr2 appears dispensable for Krt14+ progenitor cell expansion after CPP-injury, but appears required for DNA repair and proliferation/cell cycle progression/differentiation of Krt14+ progenitors. Administration of KGF to wildtype bladders injured by CPP may enhance DNA repair.
Funding
- NIDDK Support