Fig 2: CUGBP1 in HSCs promotes murine liver fibrosis induced by BDL.Mice were administrated intravenously with AAV for 8 weeks before BDL operation. Fraxinellone 40 mg kg-1 was administrated by gavage once a day for 4 weeks after BDL operation. VA-Lip-siRNA-CUGBP1 (0.75 mg kg-1) was administrated intravenously three times a week for 4 weeks after BDL operation. (a,d) Representative microphotograph of H&E-stained, Masson-stained and Sirius Red-stained paraffin-embedded sections of liver tissues (Scale bars, 100 µm). (b,e) Ishak fibrosis staging scores of the Sirius red-stained sections. (c,f) Expression levels of liver Hyp. (b,c,e,f, mean±s.e.m.; n=6). *P<0.05, and **P<0.01 as indicated by Student's t test; NS, not significant.

Fig 3: Fraxinellone regulates TGF-ß and IFN-? signalling in BDL model.(a) Western blot analyses of a-SMA, CUGBP1, pSmad2, pSTAT1, Smad7 and a-tubulin expression in the mouse liver. (b) IHF analyses of a-SMA and CUGBP1 expression in liver cryosections (Scale bars, 100 µm). (c) IHF analysis of IFN-? expression in liver cryosections (Scale bars, 50 µm). Arrows indicate the IFN-? (green) in HSCs that were positive for both a-SMA (red) and Reelin (cyan).

Fig 4: The failure of livers of Gank LKO mice to proliferate after CCl4injections is caused by high levels of tumor-suppressor proteins, which are eliminated by Gank in control LoxP mice. (A) Expression of TSPs in LoxP and Gank LKO mice after CCl4 injury. The experiment was performed with 3 biological replicates. (B) Calculations of levels of TSPs as ratios to ß-actin using biological replicates. P values for HNF4a were as follows: 0 hours (P > .99), 24 hours (P > .99), 48 hours (P < .0002), 72 hours (P > .05), and 96 hours (P > .99). P values for Rb were as follows: 0 hours (P > .99), 24 hours (P > .18), 48 hours (P < .009), 72 hours (P > .33), and 96 hours (P > .99). P values for C/EBPa were as follows: 0 hours (P > .99), 24 hours (P > .19), 48 hours (P < .03), 72 hours (P < .009), and 96 hours (P > .86). P values for p53 were as follows: 0 hours (P < .002), 24 hours (P > .23), 48 hours (P < .02), 72 hours (P > .99), and 96 hours (P > .99). (C) Gank interacts with TSPs in livers of LoxP mice at 48 hours after CCl4 injury. Co-IP assay was applied for these studies. Nuclear extracts of LoxP and Gank LKO mice (200 µg for LoxP 0-hour time point and Gank LKO at 0 and 48 hours and 1 mg for LoxP at 48 hours) were incubated with antibodies to Gank and rabbit IgG beads (TrueBlot). After washing, IPs were loaded on the gel and probed with antibodies to Rb, p53, CUGBP1, C/EBPa, and HNF4a. The IPs were performed 3 times. Every IP was examined by Western blot with the earlier-mentioned antibodies. Right: Hypothetical mechanism for the failure of livers of Gank LKO mice to proliferate after CCl4-mediated injury. Two way analysis of variance. *P < .05, **P < .01, ***P < .001 LoxP vs Gank LKO.

Fig 5: miR-322/-503 targeted Celf1 leading to partial rescue of myoblast differentiation defects but not ribonuclear foci in DM1.A, B Celf1 protein level was repressed by miR-322/-503 overexpression (OE) in C2C12-CUG200 cells. C, D Celf1 protein level was repressed by miR-322/-503 OE in HEK293T cells. E A mutual binding site of miR-322/-503 on Celf1-3'UTR was predicted by RNA22. F miR-322/-503 significantly repressed the relative luciferase activity of luciferase reporter that contained wild-type Celf1-3'UTR. However, the repression was abolished if the predicted binding sites of miR-322/-503 was mutated (Celf1-3'UTR-mut). G, H The knockdown of Celf1 in C2C12-CUG200 cells was verified by western blot. I Celf1 knockdown significantly upregulated the expressions of myogenic markers (MyoD, MyoG, and Mef2C) and downregulated Celf1 expression during C2C12-CUG200 differentiation. All expression levels were normalized to the control group at day 0. J Celf1 knockdown promoted myotube formation during C2C12-CUG200 differentiation. Myotube formation was displayed by immunostaining of MF20 on day 6. K Celf1 knockdown increased myotube areas (left) and fusion index (right) of C2C12-CUG200 differentiation. L, M Celf1 knockdown did not repress ribonuclear foci formation in C212-CUG200 cells. The ribonuclear foci were detected with CAG probe RNA FISH. Control, pLL4.0 vector stable transfection; miR-322/-503, pLL4.0-miR-322/-503 stable transfection; pLL4.0, pLL4.0 vector transient transfection; pLL4.0-miR-322/-503, pLL4.0-miR-322/-503 transient transfection; scramble, C2C12-CUG200/scramble cells; shCelf1, C2C12-CUG200/shCelf1 cells; * statistically significant (p < 0.05); ns not statistically significant.