Identification of podocytes in the primary culture of rat decapsulated glomeruli

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The origin of cobblestone-like polygonal cells in glomerular cell culture remains controversial about whether they are dedifferentiated podocytes or derived from parietal epithelial cells (PECs) of Bowman's capsule. Almost no outgrowths from glomeruli devoid of Bowman's capsule (decapsulated glomeruli) complicate the identification of cells in culture. Based on the assumption that podocytes are damaged during the isolation of glomeruli by the conventional sieving method, we tried to isolate glomeruli by the modified sieving method without the mechanical force by pressing with a spatula. As a result, the rates of cell outgrowths from decapsulated glomeruli were conspicuously improved. The morphology of the cells was distinctly different from cobblestone-like polygonal appearance. They extended long cytoplasmic processes, which often crossed over adjacent cells. Polygonal cells rarely grew out from decapsulated glomeruli, but did more frequently from encapsulated glomeruli. Cells growing out from decapsulated glomeruli showed intense staining for podocyte-specific markers (podocalyxin and synaptopodin), but no staining for markers specific to mesangial cells (Thy-1), endothelial cells (von Willebrand factor, RECA-1) or PECs (pan cadherein), indicating that these cells were of podocyte origin. On the other hand, ciliated polygonal cells growing out from encapsulated glomeruli were positive for pan cadherin along cell-cell contact, supporting the idea that polygonal cells originate from PECs. In addition, some of phenotypes specific to podocytes in vivo were expressed in cultured PECs and tubular epithelial cells. These findings indicate that all podocyte-specific markers in vivo are not necessarily specific to podocytes in culture.