Fig. 4

GOAC and puromycin aminonucleoside (PAN) injury. a Representation of GOAC albumin permselectivity assay following PAN injury: PAN (5-day induction, 10 mg/ml) is applied to channel C followed by albumin-FITC (40 mg/ml, yellow) and flow-through collected in channel F. b–d Bright field showing albumin leakage (yellow) after 5 min (left columns) and 60 min (right columns) in hAKPC-P + hGEC chip (b), in hiPOD + hGEC chip (c), in hpPOD + hGEC (d) after PAN injury. Marked albumin leakage occurs following podocyte injury. e–j Damage to podocytes was assessed by F-actin staining in hAKPC-P + hGEC chip before (e) and after PAN damage (f), in hiPOD + hGEC chip before (g) and after PAN damage (h), and in hpPOD + hGEC before (i) and after PAN damage (j) confirming widespread disruption of the cytoskeleton. Nuclei are stained with DAPI (blue); actin filaments stained with phalloidin (green). All pictures: scale bar = 50 µm; k box plot graph of fluorescein absorbance (expressed as log) in filtrate collected after 60 min after 5 days exposure to PAN. For all experimental groups, a marked and statistically significant increase was found in albumin permeability following injury. Number of replicates for chips used in k is as follows: hAKPC-P + hGEC chip: #12 and #4; hiPOD + hGEC chip: #6 and #4; hpPOD + hGEC chip: #7 and #3. Significant differences were determined by a one-way ANOVA and Holm–Sidak post hoc test, *p < 0.05, **p < 0.01, ***p < 0.001. [To improve clarity, the following significant differences were not drawn in the graph: hiPOD + hGEC + PAN vs. hAKPC-P + hGEC p < 0.001; hiPOD + hGEC + PAN vs. hpPOD + hGEC p < 0.001; hpPOD + hGEC + PAN vs. hAKPC-P + hGEC p < 0.001; hpPOD + hGEC + PAN vs. hiPOD + hGEC p < 0.001; hAKPC-P + hGEC + PAN vs. hpPOD + hGEC p < 0.001; hAKPC-P + hGEC + PAN vs. hiPOD + hGEC p < 0.05; hiPOD + hGEC + PAN vs. hAKPC-P + hGEC + PAN p < 0.05.] Box plots show the median, the 25th and 75th percentiles, whiskers (median ± 1.5 times interquartile range), and outliers (solid circle)