Fig 2: Pretreatment with uPAR monoclonal antibody alleviates acute kidney injury in septic mice. A, B Representative images of PAS staining, TUNEL staining in kidney tissues of mice injected with Saline(100 μl), LPS (10 mg/kg) for 48 h (with or without pretreatment with 500 μg/kg uPAR monoclonal antibody), along with corresponding quantitative analyses (n = 3 mice/group; 3–10 fields/mouse). C Serum levels of SCr and BUN of mice in different groups (n = 4 mice/group). D-F ROS staining of mouse kidney frozen sections, cleaved Caspase-3 (cl-Caspase-3) immunofluorescence staining across different groups, and corresponding quantitative analyses were performed (n = 3 mice/group; 3 fields/mouse). Data were presented as mean ± SEM. Statistical significance was determined by one-way ANOVA, with P < 0.05 considered significant (*P < 0.05, **P < 0.01)

Fig 3: suPAR mediates signal transduction through RAGE binding. A Protein levels of P-PERK, PERK, P-eIF2α, ATF4, CHOP, Nrf2, BAX and BCL-2 of HK-2 cells treated with suPAR(10 ng/ml) and/or FPS—ZM1(100 nM) or TLR4—IN—C34(10 μM). B The schematic diagram of the Protein docking between uPAR and RAGE. The ARG-116, SER-117, GLU-37, GLU-39, GLY-10, and ASP-11 of uPAR form hydrogen bonds with the GLN-24, ASN-81, GLY-10, THR-109, and LYS-107 of RAGE respectively. The lengths of the hydrogen bonds are 2.5 Å, 2.6 Å, 2.4 Å, 3.2 Å, 3.3 Å, and 3.5 Å respectively. C Direct suPAR-RAGE interaction confirmed by pull-down assay coupled with western blotting. D Representative confocal images showed the colocalization of suPAR (green) and RAGE (red) in HK-2 cells. Scale bar: 25 μm. Data were presented as mean ± SEM from three independent experiments (n = 3). Statistical significance was determined by Kruskal–Wallis test with Dunn's post hoc analysis, with P < 0.05 considered significant (*P < 0.05, **P < 0.01)

Fig 4: Increased suPAR/uPAR protein and mRNA levels in the kidneys of SA-AKI mice. A Serum levels of SCr, BUN, and suPAR in mice injected with saline (100 μL), LPS (10 mg/kg, 24 h), or subjected to CLP (24 h) (n = 6 mice/group). B-C Representative images of PAS staining, TUNEL staining, and uPAR immunohistochemical staining in kidney tissues across experimental groups, along with quantitative analyses (n = 3 mice/group; 3–10 fields/mouse). D-E uPAR immunofluorescence staining and quantitative assessment were performed in different groups of mice, with LTL staining used to visualize the brush border of renal tubular epithelial cells (White arrows: brush border discontinuity) (n = 3 mice/group; 3 fields/mouse). F Protein expression levels of suPAR/uPAR in kidney tissues from different groups (n = 3 mice/group; Mann–Whitney U test). G mRNA expression levels of uPAR in kidney tissues from different groups (n = 6 mice/group). Data were presented as mean ± SEM. Statistical significance was determined by one-way ANOVA or unpaired two-tailed Student's t-test unless noted, with P < 0.05 considered significant (*P < 0.05, **P < 0.01)