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Fig. 5. Proximal Tubule Phenotype upon Inhibition of Cholesterol Synthesis. (A–B׳) Immunofluorescence analysis of untreated controls and embryos treated with 125 μM Mevinolin at stage 40 using anti-Caveolin-1 (red) antibody and ECL (green) with the gray-scale image showing Caveolin-1 staining alone and the color panels showing merged images. DAPI (blue) was used to visualize nuclei. (C) Quantification of the number of Caveolin-1-positive foci identified in (A,B). The number of embryos analyzed is indicated in the individual bars. Data were analyzed by Student׳s t-Test and asterisk represent a significance of p<0.05. (D) Bar diagram of the number of proximal tubular cells in untreated controls and Mevinolin-treated embryos at stages 40. The number of embryos analyzed is indicated in the individual bars. Data were analyzed by Student׳s t-Test and the three asterisks indicate a significance of p<0.001. (E–I׳) Untreated controls and Mevinolin-treated embryos were processed for 3G8 and 4A6 immunohistochemistry at stage 40 (E–F׳) or whole mount in situ hybridization with β1-Na/K ATPase (G,G׳), Pmp70 (H,H׳) and Catalase (I,I׳) at stage 39. (J,J׳) Model for the role of lipid rafts in proximal tubule elongation in the wild-type situation (J) or in the absence of scp2 or upon treatment with Mevinolin (J׳). See discussion for details. |
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cat.2 (catalase, gene 2) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 39, lateral view, anterior left, dorsal up. |
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Supplementary Fig. S7.Scp2 Morphant Phenotype. (A) Quantification of the whole mount immunostainings with 4A6 shown in Fig. 3K N. The embryos were categorized in three groups representing increasing severity of the phenotype. The amount of MOs injected and the number of embryos analyzed is indicated. (B-E׳) Whole mount in situ hybridization of uninjected and scp2-MO1+2-injected embryos for Nphs1 (B,B׳), Sglt1k (C,C׳), Nkcc2 (D,D׳) and Ncc (E,E׳) at stage 39. (F–H׳) Uninjected controls and scp2-MO1+2-injected embryos were stained with DAB to visualize endogenous peroxidase activity at stage 40 (F,F׳) and by whole mount in situ hybridization using the peroxisomal marker genes, Catalase (G,G׳) and Pmp70 (H,H׳) at stage 39. (I,I׳) Basigin whole mount in situ hybridization of uninjected controls and scp2 morphants at stage 39. |
