Fig. 5: Podocyte-specific UCH-L1 deletion attenuates ubiquitin accumulations and podocyte injury in experimental MN. | Nature Communications

Fig. 5: Podocyte-specific UCH-L1 deletion attenuates ubiquitin accumulations and podocyte injury in experimental MN.

From: Non-functional ubiquitin C-terminal hydrolase L1 drives podocyte injury through impairing proteasomes in autoimmune glomerulonephritis

Fig. 5

Experimental MN was induced by i.v. injection of AP-abs in UCH-L1-deficient (Δpod) and littermate (Ctrl) mice. Kidneys were collected on day 14. a Urinary albumin to creatinine ratio. Pooled values of 6 independent experiments, mean + /-SEM, per time n ≤ 15 (Ctrl) or n ≤ 18 (Δpod), **p = 0.0022, Two-Way ANOVA. b Stainfree SDS-PAGE from 1 experiment of creatinine-adapted urines depicts loss of high molecular weight proteins in Ctrl mice, albumin ~63 kDa. c Immunoblot quantification of glomerular nephrin abundance normalized to β-actin. Pooled values of 2 independent experiments, mean + /-SEM, n = 6 (Ctrl) or n = 7 (Δpod), *p = 0.014, two-tailed Mann Whitney U test. d STED analyses of nephrin meanders. e Electron microscopy (EM) demonstrates effaced foot processes in Ctrl and mostly preserved foot processes in Δpod mice, PC = podocyte, red arrowheads = foot processes. f Quantification of foot process effacement by PEMP analyses of filtration slit density (FSD) length per area (µm−1), pooled values of 1 experiment, mean + /-SEM, n = 15 (Ctrl) or n = 12 (Δpod), *p = 0.0321, two-tailed Mann Whitney U test. g Podocyte number per glomerular tuft area assessed by staining for p57 (podocyte marker). Pooled values of 3 independent experiments, mean + /-SEM, n = 271 (Ctrl) or n = 332 (Δpod) glomeruli, **p = 0.0014, two-tailed Unpaired t-test. h Glomerular immunoblots for polyubiquitinated proteins (pUB) normalized to β-actin. Pooled values of 3 independent experiments, mean + /-SEM, n = 8 (Ctrl) or n = 11 (Δpod), *p = 0.0328, two-tailed Mann Whitney U test. i Representative confocal images of 2 independent experiments with n = 3 showing enhanced signal for ubiquitinated proteins (green) in Ctrl podocytes with occasional ubiquitin aggregates (arrowheads). Nephrin (red) was used to depict the glomerular filtration barrier and DNA was stained with Hoechst (blue). Podocytes are marked by asterisks. j Representative confocal images of 2 independent experiments with n = 3 showing inclusion bodies (aggresomes, marked by arrowheads) in Ctrl podocytes. Wheat germ agglutinin (WGA, red) was used to highlight the podocyte plasma membrane. DNA was stained with Hoechst (blue). Asterisks demarcate podocytes. Source data are provided as a Source Data file.

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