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. 2021 Jul 19;10:e68603. doi: 10.7554/eLife.68603

Figure 5. Damage-associated PT cells undergo high ferroptotic stress after severe IRI.

(A) UMAP rendering of glutathione metabolic process in mouse and human kidneys. (B) A scheme showing glutathione-glutathione peroxidase 4 (GPX4) anti-ferroptotic defense pathway. Slc7a11 and Slc3a2 (system xc-); Gclc and Gclm (glutamate-cysteine ligase); Gss (glutathione synthetase); Gsr (glutathione reductase): and Gpx4. MDA (malondialdehyde, a lipid peroxidation product) and ACSL4 (acyl-CoA synthetase long-chain family member 4) are markers for ferroptotic stress. (C) Dot plots show the expression of genes for glutathione-GPX4 axis, Sox9, and Acsl4. (D) Immunostaining for SOX9 and MDA (6 hr post-IRI), and (E) quantification of double-positive cells in total SOX9+ cells. N = 4. (F) Immunostaining for SOX9 and ACSL4 (1 day post-IRI), and (G) quantification of double-positive cells in total SOX9+ cells. N = 4. Insets: individual fluorescence channels of the dotted box area. Note that severe ischemia (30 min) induces more ferroptotic stress markers (MDA and ACSL4) in SOX9+ cells in damaged kidneys than mild ischemia (20 min). Wild-type C57BL/6J mice were used for (D) to (G). Scale bars, 20 μm in (D) and (F). *p < 0.05. unpaired Student’s t-test.

Figure 5.

Figure 5—figure supplement 1. Gene ontology analyses identify enrichment of anti-oxidative stress defense genes in differentiated/mature PT cells.

Figure 5—figure supplement 1.

UMAP rendering of signaling pathways. Upper panels show the pathways enriched in differentiated PT cells (PT cluster). Lower panels show the pathways enriched in damage-associated PT cells (DA-PT cluster). Arrows indicate differentiated PT cell cluster (PT). Arrowheads indicate DA-PT cell cluster.
Figure 5—figure supplement 2. Characterization of human normal kidney single-nucleus RNA-seq data.

Figure 5—figure supplement 2.

(A) UMAP plots showing human normal kidney cells from GSE131882 (7,631 cells). Two normal kidney datasets were integrated and analyzed. (B) Dot plot showing the gene expression patterns of cluster-enriched canonical markers. (C) UMAP rendering of signaling pathways. Note that the signaling pathways for anti-oxidative stress, which are enriched in differentiated PT cells in mouse kidneys, are also enriched in normal differentiated PT cells in humans. Blue arrowheads: PT cells, green arrowheads, VCAM1+ PT cells.