Abstract: FR-PO253
Effect of Integrin Signaling Blockade on Self-Renewal and Differentiation of Human Nephrogenic Progenitors In Vitro
Session Information
- Stem Cells
November 03, 2017 | Location: Hall H, Morial Convention Center
Abstract Time: 10:00 AM - 10:00 AM
Category: Developmental Biology and Inherited Kidney Diseases
- 402 Stem Cells
Authors
- Petrosyan, Astgik, Children's Hospital Los Angeles, Los Angeles, California, United States
- Karg?n, Sinem, Children's Hospital Los Angeles, Los Angeles, California, United States
- Thornton, Matthew Edward, University of Southern California, Los Angeles, California, United States
- Grubbs, Brendan, University of Southern California, Los Angeles, California, United States
- De Filippo, Roger E., Children's Hospital Los Angeles, Los Angeles, California, United States
- Perin, Laura, Children's Hospital Los Angeles, Los Angeles, California, United States
- Da Sacco, Stefano, Children's Hospital Los Angeles, Los Angeles, California, United States
Background
Mammalian kidney development is controlled through the proliferation and differentiation of a specific population of nephron progenitors (NP) characterized by the expression of CITED1 and SIX2. The mechanisms regulating the balance between self-renewal and renal differentiation in NP are still elusive, impairing our ability to effectively expand NP in vitro for long term. In particular, the effects of extracellular matrix (ECM) composition and ECM-NP interaction are poorly understood.
Methods
We have investigated the relationship between ECM and self-renewal traits in NP isolated from human fetal kidneys (hFK). NP were isolated using our established RNA Smartflare protocol. Nephrogenic characteristics were confirmed by RNA-seq and nephrogenic potential by in vitro differentiation and dissociation/reaggregation assays. By immunofluorescence we have characterized the ECM present within the nephrogenic niche of hFK. Subsequently we have tested NP expansion on these ECM substrates and assessed effects on signaling cascade by PCR array.
Results
Among others, laminin alpha 5, collagen 16 and collagen 18 where found to be highly expressed within the cap mesenchyme of developing hFK. In vitro, laminin was confirmed to better preserve self-renewal properties in NP, as confirmed by maintenance of higher co-expression of SIX2 and CITED1 in cultured NP, both in short and long term experiments. Interestingly, blocking integrin mediated ECM-NP interaction with specific antibodies lead to an increase in SIX2 and CITED1 co-expression, suggesting an important role of integrin mediated signaling pathway on balance between renal specification vs self-renewal. Effect of integrin blocking in NP on downstream WNT signaling was further confirmed by PCR array, suggesting a direct role of ECM-NP interaction on self-renewal.
Conclusion
Our data indicate a strong link between ECM-NP during human renal development. NP-laminin interaction appears to play an essential role on nephron endowment by directly controlling self-renewal/differentiation balance. These results could provide not only a tool for the optimization of in vitro NP expansion but also a platform to advance our understanding of human renal development and nephron cell commitment.
Funding
- Private Foundation Support