Abstract: SA-PO486
Intrahepatic Bile Duct Morphogenesis in Pkhd1 Mutant Mice
Session Information
- Cystic Kidney Diseases: Basic/Translational
November 09, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Genetic Diseases of the Kidneys
- 1001 Genetic Diseases of the Kidneys: Cystic
Authors
- Outeda, Patricia, University of Maryland, Baltimore, Maryland, United States
- Watnick, Terry J., University of Maryland School of Medicine, Baltimore, Maryland, United States
Background
ARPKD is a complex disease caused by mutations in the PKHD1 gene (encoding the Fibrocystin/Polyductin protein, or FPC). The predominant clinical manifestations are enlarged echogenic kidneys with distal tubule dilatation and cysts in the biliary tract with congenital hepatic fibrosis. The pathognomonic liver defect in ARPKD is the ductal plate malformation (DPM), which is characterized by the presence of increased numbers of tortuous, irregular bile ducts within the portal tract and fibrosis. In mice, as in humans, the biliary tract development is initiated by the induction of bi-potential hepatoblasts adjacent to the portal vein mesenchyme expressing Sox9 that eventually will form the primitive ductal structures (PDS) by recruiting Sox9− HNF4α+ cells from the surrounding parenchyma that will mature into symmetric structures with all the cells Sox9+ HNF4α−. From around E17 onwards, in a process termed ductal plate remodeling, these dilatations become surrounded by portal mesenchyme to eventually give rise to mature IHBD (intrahepatic biliary ducts). To date, few studies have characterized the DPM associated with ARPKD in either humans or mouse models and the precise cellular defect remains undefined.
Methods
To gain insights into the mechanism altered during the development of the DPM we harvested livers from homozygous Pkhd1Δ3-4 and control littermates at E18.5, P0, P7, and P15 and performed H&E staining.
Results
We observed dilated biliary ducts in homozygous mutants starting at P7. Immunofluorescence staining using Sox9 and Ki67 revealed an increased number of Sox9+ cells surrounding the portal vein in mutant homozygous mice although the number of proliferative cells Sox9+ was the same. In order to gain further insights into the role of FPC regulating the biliary tract development we isolated biliary duct cells using anti-EpCAM+ (cholangiocytes specific marker) coated magnetic beads from homozygous mutants and control littermates at P5 (N=16) and performed RNA sequencing (RNA-seq). We identified more than 50 genes that were differentially expressed (Log2 (FC)≤ ± 0.25 and a corrected p-value of <0.05).
Conclusion
Our study revealed that mutant mice develop a biliary expansion independent of the biliary precursors’ proliferation. We also detected transcriptional changes in Pkhd1 mutant cholangiocytes supporting the importance of FPC signaling in the biliary tract development.