Fig 2: Myofibroblast-restricted OASIS deletion resulted in decreased kidney fibrosis after UUO. A, Generation of myofibroblasts-restricted OASIS KO (cKO) mice using Cre-loxP system. B, cKO and control mice were subjected to UUO. At Day 7 after UUO, quantitative PCR was performed for transcript expression of OASIS/Creb3l1. The expression of the transcripts was normalized to that of Gapdh. ** P < .01 by Student t test. C, Myofibroblasts were isolated from kidney of cKO and control. The transcript expression of OASIS/Creb3l1 was examined by PCR. P: positive control, N: negative control. D, Sirius Red (upper and middle panel) or Masson's trichrome (lower panel) staining was performed. Representative images are shown. Bar: 50 µm. E, Sirius Red positive area was measured. F, Hydroxyproline assay was performed. G, Immunofluorescence analysis was performed with anti-Ki-67 antibody, anti-a-SMA antibody and DAPI. Representative images are shown. Bar: 50 µm. H and I, The number of a-SMA+ cells (H) and the number of Ki-67 + a-SMA+ cells (I) was quantitatively evaluated. J-O, Quantitative PCR was performed for transcript expression of Tgfb1, Col 1, Col 3, Mmp2, Mmp9, and Bst2. The expression of the transcripts was normalized to that of Gapdh. * P < .05, ** P < .01 by one-way ANOVA followed by Dunnett test. Data are shown as mean ± SD (n = 9 for control, n = 11 for cKO)

Fig 3: The graphical abstract of this study. OASIS is induced by TGF-ß1 and promotes collagen synthesis in myofibroblasts. Furthermore, OASIS contributes to progressive kidney fibrosis by facilitating migration and proliferation of myofibroblasts, accompanied by increased Bst2 expression

Fig 4: Blocking Bst2, a candidate downstream pro-fibrotic effector of OASIS, attenuated kidney fibrosis. A, Myofibroblasts were isolated from kidneys of WT and OASIS KO mice at Day 7 after UUO. Twenty-four hours after TGF-ß1 treatment, the total mRNA was extracted from the cells and microarray analysis was performed (n = 2 in each group). Heat map shows the genes with >1.5-fold change between WT and OASIS KO cells with a P-value of <0.2. B, The expression of Bst2 transcript was measured at indicated time points after UUO by quantitative RT-PCR. The expression of the transcript was normalized to that of Gapdh. Data are shown as mean ± SD (n = 5). C, At Day 7 after UUO, the transcript expression of Bst2 in kidney of WT and OASIS KO mice was analyzed by quantitative PCR. Data are shown as mean ± SD (n = 7 for WT, n = 6 for KO). D, Upper panel, cAMP responsive element-like sequence was cited from JASPAR and was picked up in the upstream region of Bst2 gene. The line indicates same sequence. Lower panel, isolated myofibroblasts were treated with TGF-ß1 and chromatin immunoprecipitation assay was conducted. E-G, C57BL/6J mice were intraperitoneally treated with anti-Bst2 antibody or control IgG? at Day 1 after UUO. E, Seven days after UUO, Sirius Red (upper and middle panel) or Masson's trichrome (lower panel) staining was performed. Representative images are shown. Bar: 50 µm. F, Sirius Red positive area was measured. G, Hydroxyproline assay was performed. Data are shown as mean ± SD (n = 8). H, OASIS KO or WT mice were intraperitoneally treated with anti-Bst2 antibody or control IgG? at Day 1 after UUO. Hydroxyproline assay was performed at Day 7 after UUO. Data are shown as mean ± SD (n = 6 for WT, n = 5 for KO). * P < .05, ** P < .01 by one-way ANOVA followed by Dunnett test