Figure 4.

Regeneration of nephrons derived from transplanted rat NPCs in an adult Lewis rat with TAC and MP treatment. (a) Schematic of the experimental protocol. (b) E13.5 Six2-iDTR mouse MNB-containing rat RPCs and DT was transplanted into the vicinity of the aorta of an adult Lewis rat with TAC (2 mg/kg/day s.c.) and MP (5 mg/kg/day i.p.) treatment. Twenty-one days after transplantation, GFP expression was evaluated and identified (scale bars, upper column: 2 mm, lower column: 250 µm). The arrows represent the infiltration of host blood vessels. (c) Frozen sections were prepared from the collected MNB. We observed GFP-positive glomerular and tubular structures (left and middle column: scale bars, 300 µm). HE staining demonstrated that the glomeruli and tubules had regenerated from the transplanted rat NPCs (right column; scale bar, 100 µm). (d) Frozen sections were analysed by immunostaining. We identified neo-glomeruli (left upper column; scale bars,100 µm) and tubules (left lower column; sale bars, 50 µm) that originated from transplanted rat GFP-NPCs. Glomeruli had nephrin-positive epithelial cells lining CD31-positive vascular endothelial cells (right upper column; scale bar, 10 µm). (e) Aligned podocytes and a glomerular basement membrane in the neo-glomeruli were observed by transmission electron microscopy (left column, second from left). Slit diaphragm structures were observed (second from left arrowheads), and red blood cells could be confirmed in the neo-glomeruli (middle column, arrow). Neo-tubules displayed brush borders (second from right, right column arrows). Scale bars: left column, 2 µm; second from left, 500 nm; middle column, 500 nm; second from right, 2 μm; right column, 500 nm. Representative images of six specimens from three independent transplantation experiments are indicated.