Kidney Week

Abstract: FR-PO700

Impact of MicroRNA Depletion on Maintenance of Adult Bladder Urothelium

Session Information

• Pediatric Nephrology - 1
  October 25, 2024 | Location: Exhibit Hall, Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Pediatric Nephrology

• 1900 Pediatric Nephrology

Authors

• Kercsmar, Macie M., Kidney and Urinary Tract Center, Abigail Wexner Research Institute at Nationwide Children's Hospital, Columbus, Ohio, United States

Macie M. Kercsmar

• Cortado, Hanna H., Kidney and Urinary Tract Center, Abigail Wexner Research Institute at Nationwide Children's Hospital, Columbus, Ohio, United States

Hanna H. Cortado, MS, MSc

• Grounds, Kelly, Kidney and Urinary Tract Center, Abigail Wexner Research Institute at Nationwide Children's Hospital, Columbus, Ohio, United States

Kelly Grounds

• Jackson, Ashley R., Kidney and Urinary Tract Center, Abigail Wexner Research Institute at Nationwide Children's Hospital, Columbus, Ohio, United States

Ashley R. Jackson, PhD

• Becknell, Brian, Kidney and Urinary Tract Center, Abigail Wexner Research Institute at Nationwide Children's Hospital, Columbus, Ohio, United States

Brian Becknell, MD, PhD

Background

MicroRNA (miRNA) serve essential roles in epithelial cell development, maintenance, and response to injury through the regulation of mRNA and protein expression, however their functions in urothelium remains unknown. To address this knowledge gap, we engineered mice with temporally-restricted, urothelium specific inactivation of Dicer, an exonuclease required for miRNA biogenesis.

Methods

We generated tamoxifen-inducible urothelial Dicer conditional knockout mice by intercrossing Dicer^fl/fl and Upk2CreERT₂ or ShhCreERT₂ animals. A tdTomato (tdT) fluorescent protein was expressed in a Cre/LoxP dependent manner to identify cells in which Dicer inactivation had occurred. Urothelial lineage markers were evaluated by immunofluorescence microscopy, Western blotting, and QRT-PCR.

Results

Bladders from control animals showed the expected urothelial morphology with three distinct layers: Krt5+;Krt14+ basal (B) cells, Krt5+;Upk+ intermediate (I) cells, and Upk+;FABP4+ superficial (S) cells. In contrast, adult bladders with Upk2CreERT₂-specific Dicer inactivation displayed rounded and exfoliated S cells with discontinuous Fabp4 expression. This coincided with increased Krt5 and Krt14 protein expression, whereas Krt5 and Krt14 mRNA levels were unaffected. In contrast, ShhCreERT₂-specific Dicer knockout mice displayed immature B cells and abundant I cells, whereas S cells maintained FABP expression but exhibited a round and more columnar morphology.

Conclusion

Dicer serves essential roles in urothelial structural integrity by promoting the maintenance of superficial, intermediate, and basal cells. This justifies further studies to identify specific miRNA responsible for this role.