The Directed Differentiation of Human iPS Cells into Kidney Podocytes (original) (raw)
Figure 2
iPS Podocyte Characterisation and Gene Expression.
A) Immunofluorescence microscopy showed localisation of podocin (green) in human podocytes, counterstained with DAPI. B) Human podocytes localised with synaptopodin protein (red) throughout the cytoplasm surrounding the DAPI-stained nucleus (blue). C) At 10 days of directed differentiation iPS podocytes show localisation of podocin protein, counterstained with DAPI. D) Synaptopodin (red) was expressed in the cytoplasmic matrix of iPS podocytes. E) Unlike human podocytes, iPS podocytes proliferated in culture as observed by podocin expression and DAPI (blue) and are also WT1-positive (F). Differentiated iPS cells express Pax-2 protein (G) and show nuclear co-localisation of Pax-2 and WT1 proteins (H). Mag A–D, H ×400; E–G ×200. I) qPCR showed the upregulated mRNA expression of kidney metanephric mesenchymal and podocyte genes. From day 3 (D3) to day 10 (D10) of iPS podocyte directed differentiation there was an upregulated expression of Pax2 and WT1 and podocyte-specific markers synaptopodin and nephrin at comparable levels to primary human podocytes, but different from the starting normal human mesangial cells (NHMC).