Sonic hedgehog connects podocyte injury to mesangial activation and glomerulosclerosis
Dong Zhou, Haiyan Fu, Yang Han, Lu Zhang, Shijia Liu, Lin Lin, Donna B. Stolz, Youhua Liu
Dong Zhou, Haiyan Fu, Yang Han, Lu Zhang, Shijia Liu, Lin Lin, Donna B. Stolz, Youhua Liu
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Research Article Nephrology

Sonic hedgehog connects podocyte injury to mesangial activation and glomerulosclerosis

Abstract

Glomerular disease is characterized by proteinuria and glomerulosclerosis, two pathologic features caused by podocyte injury and mesangial cell activation, respectively. However, whether these two events are linked remains elusive. Here, we report that sonic hedgehog (Shh) is the mediator that connects podocyte damage to mesangial activation and glomerulosclerosis. Shh was induced in glomerular podocytes in various models of proteinuric chronic kidney diseases (CKD). However, mesangial cells in the glomeruli, but not podocytes, responded to hedgehog ligand. In vitro, Shh was induced in podocytes after injury and selectively promoted mesangial cell activation and proliferation. In a miniorgan culture of isolated glomeruli, Shh promoted mesangial activation but did not affect the integrity of podocytes. Podocyte-specific ablation of Shh in vivo exhibited no effect on proteinuria after adriamycin injection but hampered mesangial activation and glomerulosclerosis. Consistently, pharmacologic blockade of Shh signaling decoupled proteinuria from glomerulosclerosis. In humans, Shh was upregulated in glomerular podocytes in patients with CKD and its circulating level was associated with glomerulosclerosis but not proteinuria. These studies demonstrate that Shh mechanistically links podocyte injury to mesangial activation in the pathogenesis of glomerular diseases. Our findings also illustrate a crucial role for podocyte-mesangial communication in connecting proteinuria to glomerulosclerosis.

Authors

Dong Zhou, Haiyan Fu, Yang Han, Lu Zhang, Shijia Liu, Lin Lin, Donna B. Stolz, Youhua Liu

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Figure 4

Genetic ablation of podocyte Shh does not affect proteinuria but inhibits mesangial cell activation after injury.

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Genetic ablation of podocyte Shh does not affect proteinuria but inhibit...
(A) Generation of conditional knockout mice with podocyte-specific ablation of Shh. (B) qPCR reveals that loss of podocyte Shh repressed Gli1 expression at 1 week after ADR injection. *P < 0.05 versus controls (n = 5–7, t test). (C) Podocyte-specific ablation of Shh did not affect albuminuria at day 4 and day 7 after ADR injection (n = 12–17). (D) SDS-PAGE analysis shows the abundance and composition of urinary proteins in different groups of mice at 1 week after ADR. Urine samples after normalization to creatinine were analyzed on SDS-PAGE. The numbers (1 and 2) indicate each individual animal in a given group. The asterisk indicates the position of albumin. (E) qPCR reveals that loss of Shh in podocytes did not affect nephrin, podocin, CD2AP, podocalyxin, and WT1 expression at 1 week after ADR, compared with the controls (n = 5–7, t test). (F) Representative micrographs show that loss of Shh in podocytes affected neither nephrin and α-actinin-4 expression nor podocyte ultrastructure at 1 week after ADR. Scale bar: 20 μm. Scale bar in transmission electron microscopy (TEM) images: 500 nm. (G and H) Western blot analyses demonstrate that loss of Shh in podocyte had little effect on α-actinin-4 and PDGFR-β expression but inhibited α-SMA induction at 1 week after ADR. Representative Western blots (G) and quantitative data (H) are presented. Numbers (1 through 4) indicate each individual animal in a given group. *P < 0.05 (n = 7, t test). (I) Representative micrographs show that ablation of Shh in podocytes did not affect mesangial abundance but inhibited mesangial activation at 1 week after ADR. Kidney sections were subjected to PAS and PASM staining or double immunofluorescence staining for podocalyxin (green) and PDGFR-β (red) or podocalyxin (green) and α-SMA (red). Arrows indicate positive staining. Scale bar: 20 μm. (J) Quantitation of IF intensity (arbitrary unit) of PDGFR-β in the glomeruli of control and podo-Shh–/– mice at 7 days after ADR. (K) Quantitation of α-SMA+ cells in the glomeruli of control and podo-Shh–/– mice. *P < 0.05 versus controls, t test.