Renal proximal tubular NEMO plays a critical role in ischemic acute kidney injury
Sang Jun Han, … , Marc Schmidt-Supprian, H. Thomas Lee
Sang Jun Han, … , Marc Schmidt-Supprian, H. Thomas Lee
Published September 17, 2020
Citation Information: JCI Insight. 2020;5(19):e139246. https://doi.org/10.1172/jci.insight.139246.
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Research Article Inflammation

Renal proximal tubular NEMO plays a critical role in ischemic acute kidney injury

Abstract

We determined that renal proximal tubular (PT) NF-κB essential modulator (NEMO) plays a direct and critical role in ischemic acute kidney injury (AKI) using mice lacking renal PT NEMO and by targeted renal PT NEMO inhibition with mesoscale nanoparticle–encapsulated NEMO binding peptide (NBP MNP). We subjected renal PT NEMO–deficient mice, WT mice, and C57BL/6 mice to sham surgery or 30 minutes of renal ischemia and reperfusion (IR). C57BL/6 mice received NBP MNP or empty MNP before renal IR injury. Mice treated with NBP MNP and mice deficient in renal PT NEMO were protected against ischemic AKI, having decreased renal tubular necrosis, inflammation, and apoptosis compared with control MNP-treated or WT mice, respectively. Recombinant peptidylarginine deiminase type 4 (rPAD4) targeted kidney PT NEMO to exacerbate ischemic AKI in that exogenous rPAD4 exacerbated renal IR injury in WT mice but not in renal PT NEMO–deficient mice. Furthermore, rPAD4 upregulated proinflammatory cytokine mRNA and NF-κB activation in freshly isolated renal proximal tubules from WT mice but not from PT NEMO–deficient mice. Taken together, our studies suggest that renal PT NEMO plays a critical role in ischemic AKI by promoting renal tubular inflammation, apoptosis, and necrosis.

Authors

Sang Jun Han, Ryan M. Williams, Mihwa Kim, Daniel A. Heller, Vivette D’Agati, Marc Schmidt-Supprian, H. Thomas Lee

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Figure 12

NEMO-mediated induction of renal proximal tubular NF-κB proinflammatory signaling in mouse renal proximal tubule cells.

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NEMO-mediated induction of renal proximal tubular NF-κB proinflammatory ...
Primary cultures of renal proximal tubule cells from NEMOfl/fl WT or renal PT NEMO–deficient (NEMOfl/fl PEPCK-Cre) mice were treated with recombinant PAD4 (rPAD4, 10 μg/mL) for 6 hours (A) or 4 hours (B) or with lipopolysaccharides (LPS, 10 μg/mL) for 6 hours (C). (A) With RT-PCR, we measured the expressions of TNF-α, ICAM-1, MCP-1, IL-6, KC, and MIP-2 mRNAs. Fold increases in mRNAs normalized to GAPDH from quantitative RT-PCR reactions for each indicated mRNA (N = 3) are shown. (B) A representative immunoblotting experiment for nuclear p65 NF-κB subunit (top) and band intensity quantifications normalized to histone H3 (N = 3, bottom). (C) With RT-PCR, we measured the expression of TNF-α, MCP-1, and MIP-2 mRNAs. Fold increases in mRNAs normalized to GAPDH from quantitative RT-PCR reactions for each indicated mRNA (N = 3) are shown. For statistical analysis, 1-way ANOVA plus Tukey’s post hoc multiple-comparisons test was used to detect significant changes. *P < 0.05 versus vehicle-treated NEMOfl/fl mice. #P < 0.05 versus rPAD4-treated NEMOfl/fl mice. Error bars represent 1 SEM.