Fibroblast expression of transmembrane protein smoothened governs microenvironment characteristics after acute kidney injury
Yuan Gui, … , Youhua Liu, Dong Zhou
Yuan Gui, … , Youhua Liu, Dong Zhou
Published May 7, 2024
Citation Information: J Clin Invest. 2024;134(13):e165836. https://doi.org/10.1172/JCI165836.
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Research Article Nephrology

Fibroblast expression of transmembrane protein smoothened governs microenvironment characteristics after acute kidney injury

Abstract

The smoothened (Smo) receptor facilitates hedgehog signaling between kidney fibroblasts and tubules during acute kidney injury (AKI). Tubule-derived hedgehog is protective in AKI, but the role of fibroblast-selective Smo is unclear. Here, we report that Smo-specific ablation in fibroblasts reduced tubular cell apoptosis and inflammation, enhanced perivascular mesenchymal cell activities, and preserved kidney function after AKI. Global proteomics of these kidneys identified extracellular matrix proteins, and nidogen-1 glycoprotein in particular, as key response markers to AKI. Intriguingly, Smo was bound to nidogen-1 in cells, suggesting that loss of Smo could affect nidogen-1 accessibility. Phosphoproteomics revealed that the ‘AKI protector’ Wnt signaling pathway was activated in these kidneys. Mechanistically, nidogen-1 interacted with integrin β1 to induce Wnt in tubules to mitigate AKI. Altogether, our results support that fibroblast-selective Smo dictates AKI fate through cell-matrix interactions, including nidogen-1, and offers a robust resource and path to further dissect AKI pathogenesis.

Authors

Yuan Gui, Haiyan Fu, Zachary Palanza, Jianling Tao, Yi-Han Lin, Wenjian Min, Yi Qiao, Christopher Bonin, Geneva Hargis, Yuanyuan Wang, Peng Yang, Donald L. Kreutzer, Yanlin Wang, Yansheng Liu, Yanbao Yu, Youhua Liu, Dong Zhou

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Figure 1

Fibroblast-specific ablation of Smo mitigates ischemic AKI.

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Fibroblast-specific ablation of Smo mitigates ischemic AKI. (A) Generati...
(A) Generation of Smo fibroblast-specific deletion mice. (B) Quantitative real-time PCR (qPCR) analysis (n = 4) and Western blot assay showing Smo levels in primary fibroblasts isolated from Gli1-Smo+/+ and Gli1-Smo–/– or Pdgfr-β-Smo+/+ and Pdgfr-β-Smo–/– kidneys. (C) The mouse baseline body weight (BW), kidney-to-body weight (KW/BW) ratios, serum creatinine (Scr), and (D) blood pressure levels (n = 6). At 1 day after IRI, (E) Scr levels in Gli1-Smo+/+ and Gli1-Smo–/– mice (n = 16-17). (F) Periodic Acid–Schiff (PAS) staining showing kidney morphological changes in Gli1-Smo+/+ and Gli1-Smo–/– mice. Blue asterisks indicate injured tubules. Representative micrographs of TUNEL or IHC staining against CD45 and CD3 in Gli1-Smo+/+ and Gli1-Smo–/– kidneys. Scale bar: 25 μm. (G) Western blots assay of NGAL, FasL, and Bad in Gli1-Smo+/+ and Gli1-Smo–/– kidneys (Sham, n = 3; IRI, n = 5). (H) qPCR analysis showing Tnf-α, Mcp1, and Rantes mRNA levels in Gli1-Smo+/+ and Gli1-Smo–/– kidneys (Sham, n = 3; IRI, n = 6). (I) Scr levels in Pdgfr-β-Smo+/+ and Pdgfr-β-Smo–/– mice (n = 15-16). (J) Western blots showing NGAL, FasL, and Bad levels in Pdgfr-β-Smo+/+ and Pdgfr-β-Smo–/– kidneys. (K) qPCR analysis showing Tnf-α, Mcp1, and Rantes mRNA levels in Pdgfr-β-Smo+/+ and Pdgfr-β-Smo–/– kidneys (Sham, n = 3; IRI, n = 6). (L) Representative micrographs of PAS, TUNEL, and IHC staining against CD45 in Pdgfr-β-Smo+/+ and Pdgfr-β-Smo–/– kidneys. Scale bar: 25 μm. Arrows indicate positive cells. DAPI is a nuclear counterstain. For all Western blot panels, numbers indicate individual animals in a given group. †P < 0.05 versus sham control, *P < 0.05 versus Gli1-Smo+/+ or Pdgfr-β-Smo+/+ IRI mice. Dots indicate individual animals in a given group. Graphs are presented as means ± SEM. Differences among groups were analyzed using unpaired t tests or 1-way ANOVA followed by the Student-Newman-Keuls test.