Fig 2: The renal expression of DKK1 positively correlates with accumulation of ß-catenin in 5/6 Nx rats. (A) Representative images of Western blot analysis for the expression of DKK1 and ß-catenin in renal tissues of the sham group and the 5/6 Nx groups that were untreated or treated with DKK1-S or DKK1-AS. (B) Quantitative analysis of Western blots for ß-catenin in renal tissues of different groups (n = 6 for each group). * p < 0.05 versus the sham group, # p < 0.05 versus the untreated 5/6 Nx group. (C) Immunofluorescence analysis of ß-catenin in kidney sections from the sham-operated rats and the 5/6 Nx rats left untreated or treated with DKK1-S or DKK-AS. Scale bars: 100 µm. (D) Immunofluorescence images of renal glomeruli stained for ß-catenin in different treated groups. Scale bars: 50 µm. (E) Quantification of relative fluorescence intensities for ß-catenin in renal glomeruli of different groups (n = 6 for each group). * p < 0.05 versus the sham group, # p < 0.05 versus the untreated 5/6 Nx group.

Fig 3: Elevated levels of serum DKK1 are associated with fast progression to ESRD in patients with CKD stages 4–5. (A) Serum DKK1 levels in normal controls (n = 15) and in patients with CKD stages 4–5 (n = 50). Data are presented as mean ± SD. (B) Western blot analysis for the expression of DKK1, fibronectin (FBN) and TGF-ß1 in renal tissues from non-CKD and CKD subjects. Quantitative data are expressed as mean ± SD. * p < 0.05 versus control subjects (n = 6). (C) Chromogenic IHC staining for DKK1, FBN and TGF-ß1 in renal glomeruli of representative non-CKD subjects (#1 and #2) and CKD subjects (#1 and #2). Scale bars: 20 µm. (D) Chromogenic IHC staining for DKK1, FBN and TGF-ß1 in renal tubules of representative non-CKD subjects (#1 and #2) and CKD subjects (#1 and #2). Scale bars: 20 µm. (E) One minus dialysis-free survival analysis of CKD 4–5 patients with high serum levels of DKK1 (=1526.4 pg/mL; n = 22) or low serum levels of DKK1 (<1526.4 pg/mL; n = 28) in an 8-year follow-up. (F) Stratification by 3-year follow-up in CKD 4–5 patients with low or high serum levels of DKK1.

Fig 4: Reduced Notch signaling, but increased Wnt signaling in Ccn1-deficient mice.Ccn1?Lgr5 mice were analyzed at day 5-post corn oil or TM. a Immunofluorescence staining of Notch1 intracellular domain (N1-ICD, red) in the crypt of the jejunum. DAPI is counterstaining. Representative images are shown from three independent replicates. Bars: 20 µm. b Immunoblot analysis of indicated protein expression (N1-ICD, N2-ICD, Jag1, DLL1, and DLL2) in the jejunum. ß-actin as a loading control. Representative images are shown from at least three replicates. c qPCR analysis of Notch target genes (Hes1, Hey1, and HeyL) and Atoh1. Data represent the mean ± SEM of four biological replicates. One-sided t-test. ***p < 0.001. d Immunofluorescence staining of active ß-catenin in the crypt of Ccn1?Lgr5 mice. Images are representatives of three independent replicates. Bars: 20 µm. e Immunoblot analysis of active and total ß-catenin. Images are representative of three replicates. f, g qPCR analysis of Wnt target genes (Axin2, Cd44, Ephb2, and Myc; f), Wnt ligands (Wnt3a, Wnt5a, and Wnt3b; g), and a secreted Wnt antagonist Dkk1 (g). Data represent the mean ± SEM of four biological replicates. One-sided t-test. *p < 0.05; **p < 0.01; ***p < 0.001. Source data and the exact p-values are provided in a Source Data file.

Fig 5: DKK1 acts as a profibrotic mediator in mouse mesangial cells. (A) Effects of increasing amounts of recombinant human DKK1 (100, 200 and 400 ng/mL) on the expression of FBN, COL-IV, TGF-ß1, a-SMA, DKK1 and ß-catenin in mouse mesangial cells. The quantitative results from Western blots are plotted as mean ± SD (n = 3). * p < 0.05 versus the untreated control. (B) Effects of increasing amounts of recombinant mouse DKK1 (100, 200 and 400 ng/mL) on the expression of the indicated proteins in mouse mesangial cells. The quantitative data from Western blots are plotted as mean ± SD (n = 3). * p < 0.05 versus the untreated control. (C) Quantitative RT-PCR analysis of FBN, TGF-ß1 and DKK1 mRNAs expressed in mouse mesangial cells that were left untreated or treated with increasing amounts of recombinant human DKK1 (100, 200 and 400 ng/mL). * p < 0.05 versus the untreated control (n = 3). (D) Quantitative RT-PCR analysis of FBN, TGF-ß1 and DKK1 mRNAs expressed in mouse mesangial cells that were left untreated or treated with increasing amounts of recombinant mouse DKK1 (100, 200 and 400 ng/mL). * p < 0.05 versus the untreated control (n = 3).