Figure 4

Expression analysis of Klotho protein in kidney and heart tissue. (A) Columns 1–3 (Human Tissue): Immunoblot of Klotho and GAPDH in tissue samples derived from kidney (K), non-failing control hearts (C), and CMP hearts (D). Gel was loaded with 50 µg protein. Note that in the kidney two bands at 130 kDa and 65 kDa are visible, as opposed to only one band at 65 kDa in the heart. (B) Columns 4–7 (Mouse Tissue): Immunoblot of Klotho in KO K (−/−) and WT K (+/+) kidney as controls and KO H (−/−) and WT H (+/+) hearts. Two different antibodies, KM2076 (first row, recognition site: AA 55–261), and ab181373 (second row, recognition site: AA 400–500) with recognition sites in the KL1 region of human Klotho were used. The gels were cropped for conciseness. Full length gels are presented in Supplementary Figure S5. (B) Relative amount of ADAM10, ADAM17, and BACE1 mRNA levels related to the reference gene RPL32 in human kidney (n = 4), controls hearts (n = 10) and CMP hearts (n = 10). Shown are all individual data points, lines show mean ± SD. ^***P ≤ 0.001. (C) Human Tissue A-D: Immunohistochemistry staining of Klotho protein in kidney (A), control hearts (B), and CMP hearts from patients with low (C) and high levels (D) of serum FGF23, respectively. Mouse Tissue E-F: Immunohistochemistry staining of Klotho protein in kidney (E), and heart tissue from mice. Scale bar represents 50 µM.