Abstract
Amniotic fluid is in continuity with multiple developing organ systems, including the kidney. Committed, but still stem-like cells from these organs may thus appear in amniotic fluid. We report having established for the first time a stem-like cell population derived from human amniotic fluid and possessing characteristics of podocyte precursors. Using a method of triple positive selection we obtained a population of cells (hAKPC-P) that can be propagated in vitro for many passages without immortalization or genetic manipulation. Under specific culture conditions, these cells can be differentiated to mature podocytes. In this work we compared these cells with conditionally immortalized podocytes, the current gold standard for in vitro studies. After in vitro differentiation, both cell lines have similar expression of the major podocyte proteins, such as nephrin and type IV collagen, that are characteristic of mature functional podocytes. In addition, differentiated hAKPC-P respond to angiotensin II and the podocyte toxin, puromycin aminonucleoside, in a way typical of podocytes. In contrast to immortalized cells, hAKPC-P have a more nearly normal cell cycle regulation and a pronounced developmental pattern of specific protein expression, suggesting their suitability for studies of podocyte development for the first time in vitro. These novel progenitor cells appear to have several distinct advantages for studies of podocyte cell biology and potentially for translational therapies.
Highlights
The visceral epithelial cell is the pivotal cell maintaining normal structure and function of the kidney glomerulus [1]
In order to characterize the specificity of our selection method, we compared hAKPC-P to human immortalized podocytes [7], human lung fibroblasts, mouse kidney cortex cells and human bone marrow mesenchymal stem cells (Fig. 1, Fig. S1–S2). hIPod had 0.22% triple positive cells before re-differentiation (Fig. 1E–G) and mKC had 0.02% triple positive cells (Fig. S1A–C)
The number of studies reporting the differentiation of stem cells into podocytes, both in vivo and in vitro, is quite limited
Summary
The visceral epithelial cell (podocyte) is the pivotal cell maintaining normal structure and function of the kidney glomerulus [1]. Loss of podocytes is associated with progression of kidney disease in humans and experimental animals [2,3] since there is at most a limited possibility to replace these post-mitotic cells [4]. When a sufficient number of nephrons are lost for any reason, progressive glomerular sclerosis ensues leading to total kidney failure. Despite many characteristics typical of epithelial cells, its location, architecture and function are singular [4]. Attachment of podocyte foot processes to the glomerular basement membrane (GBM) makes direct isolation of podocytes difficult, so in vitro studies of these cells depend largely on cell culture systems
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