Transgenic zebrafish study.

(A) Schematic view of the Tol2 transposon-based plasmid carrying a 185 bp zebrafish nphs2 promoter. (B-D) Transient GFP expression in zebrafish podocytes at 4 dpf. Lateral view (B), confocal image (C), and dorsal view (D). Pronephros and dorsal aorta are indicated by arrows and an arrowhead, respectively. (E) GFP expression rate in 4 dpf-embryos injected with different constructs. The plasmid without inserts was used as a control for the baseline. The HCS3-A or -C sequence was subcloned upstream of the promoter. They are schematically shown in left panel. The bar graph illustrates expression rate (%) and total number of G₀ embryos for assessment are indicated in parentheses.

Glomerular expression analysis of four nearby genes.

(A) Immunofluorescence staining of mouse kidney sections. Positive signals of staining and locations of glomeruli are indicated by arrows and arrowheads, respectively. (B) Positive and negative controls of mouse kidney immunofluorescence staining for IL-20rb. LPS-treated mouse kidney was used as a positive control. Staining without primary antibody to IL-20rb was used as a negative control. (C) Western blotting analysis. Nck1 and calnexin, an internal control, are shown in the left side. Stag1, Tmem22, IL-20rb and β-action, an internal control, are shown in the right side. Glo, the glomerular lysate; ROK, the rest of kidney, indicating lysates from kidney that lacks glomeruli fractions. Molecular weight is indicated by number of kDa. (D) Double immunostaining of mouse kidney sections with a podocyte marker nephrin (green) and Nck1 (red). Yellow color pointed by arrows indicates partial colocaliztion of staining of Nck1 and nephrin staining. (E) qPCR analysis. mRNA expression of four genes from isolated glomeruli of adult C57BL/6 mouse kidney was quantified using the TaqMan method.