Abstract: FR-PO923
SIX2+CITED1+ Human Nephron Progenitors: Novel Insights of Wilms Tumor Biology
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
- Petrosyan, Astgik, Children's Hospital Los Angeles, Los Angeles, California, United States
- Thornton, Matthew Edward, University of Southern California, Los Angeles, California, United States
- Marcheque, Julia, Children's Hospital Los Angeles, Los Angeles, California, United States
- Grubbs, Brendan, University of Southern California, Los Angeles, California, United States
- Da Sacco, Stefano, Children's Hospital Los Angeles, Los Angeles, California, United States
- Perin, Laura, Children's Hospital Los Angeles, Los Angeles, California, United States
Background
Wilms tumor (WT) accounts for 95% of renal malignancies in children and is characterized by uncontrolled proliferation of nephron progenitors (NP) without generation of functional nephrons. Due to inability of isolating these human NP, little is known about WT formation and specifically the involvement of NP in tumor progression. Using our validated Smartflares technique we isolated NP expressing SIX2 and CITED1 (the master genes regulating nephrogenesis) from WT samples and from human fetal kidneys (hFK) and compared them in terms of gene expression by RNA-seq. We also performed in vivo and in vitro experiments to study regulation of self-renewal vs differentiation of NP.
Methods
WT (#3) and hFK (#3) samples were digested to single cell suspension, incubated with Smartflare-probe and SIX2+CITED1+ cells were immediately sorted and processed for RNA-seq. Using a nephrogenic specific media, we established conditions for long-term culture of NP cells and study in vitro mechanism of self-renewal vs differentiation of NP from both WT and hFK. We also transplanted NP in vivo to study tumorogenesis.
Results
We confirmed expression of SIX2+CITED1+ cells in WT samples within the blastema and ~ 9% of the total population was expressing these markers (while ~0.2% was present in hFK). WT-NP present a similar pattern of expression of developmental genes as hFK-NP but increased expression of EYA1, SALL4 and decreased expression of LEF1 and FOXD1 suggesting their self-renewal state. WT-NP showed, as expected, upregulation of pluripotency genes (OCT-4, NANOG) and modulation of cell cycle regulators indicating their shorter G1 phase (typical of pluripotent/cancer stem cells). WT-NP presented modulation of integrin signaling, which plays an important role in exit to differentiation during nephrogenesis. WT-NP and hFK-NP were cultured in vitro for more than 28d maintaining expression of SIX2 and CITED1. Modulation of integrin outside-in signaling reduced SIX2 and CITED1 long-term in vitro and reduced tumor growth in vivo.
Conclusion
This work represents the first characterization of SIX2+CITED1+ cells from WT and suggests the importance of matrix-cell interaction in development and tumor formation. These studies can help to increase our knowledge on human nephrogenesis and the development of new strategies aimed at halting tumor progression.
Funding
- Private Foundation Support