Fig 2: Organ-specific mRNA and protein expression levels of OCLN, JAM-A and ZO-1 in canine duodenum, lung, liver and kidney. mRNA expression levels of (A) OCLN, (B) JAM-A and (C) ZO-1 were greatest in the duodenum, moderate in the lung and kidney, and low in the liver. Protein expression levels of (D) OCLN, (E) JAM-A and (F) ZO-1 followed a similar pattern to mRNA expression levels. mRNA and protein expression were normalized to ß-actin. Data are presented as the mean ± standard deviation. OCLN, occludin; JAM-A, junction adhesion molecule A; ZO-1, zona occludens 1; ACTB, ß-actin; D, duodenum; Lu, lung; Li, liver; K, kidney.

Fig 3: Lovastatin selectively induces ?Np63 protein and mRNA in HaCaT cells. Top panels; Representative western blots of ?Np63 (72 kDa) and ß-actin (42 kDa) as loading control for HaCaT, FaDu and SCC-25 cells treated for 24 h with the indicated concentrations of lovastatin (Lov). Relative ?Np63 densitometry measurements normalized to ß-actin are indicated, with 0 used as control. Lower panels; RT-qPCR for ?Np63 mRNA shown as fold change after normalization to ACTB mRNA, with 0 used as control. *p < 0.05; **p < 0.01 compared to untreated cells

Fig 4: p63-TID interferes with p63/HDAC1 interactions and enhances human cardiac reprogramming. (A) FLAG co-immunoprecipitation assay in 293T cells transfected with HDAC1, p63-FLAG and/or p63-TID vectors, showing TID interference in p63-HDAC1 binding. Beta-actin was used as loading control. IB immunoblot, IP immunoprecipitation. (B) Cardiomyocyte and fibroblast marker gene expression assessed by qRT-PCR after indicated treatments (n = 3, ****p < 0.0001). (C) Representative flow cytometry plots for cardiac troponin T positive (cTnT +) human cardiac fibroblasts 2 weeks after their treatment with shp63 or p63-TID with or without Hand2/Myocardin (H/M). (D) Quantification of the percentage of cTnT+ cells treated as indicated, as assessed by flow cytometry (n = 3; ****p < 0.0001). (E,F) Representative immunofluorescence staining for DAPI (blue), GFP (green), and cardiomyocyte markers (red) cTnT (E) or a-actinin (F) after 2 weeks. Scale bar = 100 µm. (G,H) Quantification of cells expressing cardiomyocyte markers cTnT+ and a-actinin+ 2 weeks after indicated treatments, as assessed by immunofluorescence labelling (n = 3, ***p < 0.001, **p < 0.01). (I) Representative high-magnification images of cTnT and a-actinin staining in cells treated with p63-TID and H/M demonstrating sarcomeric structures, most clearly visible in a-actinin labeled cells. Scale bar = 25 µm. (J) Quantification of cells with well-developed sarcomeres as a percentage of total a-actinin+ cells after 4 weeks of shp63+ GMT, shp63+ H/M, or p63-TID+ H/M transduction (n = 3; **p < 0.01, ***p < 0.001).

Fig 5: Effect of NaB on high-cholesterol-induced ROS generation, BACE1 expression, and Aß accumulation through SOD1 expression.a SK-N-MC cells were transfected with NRF2 siRNA or NT siRNA for 24 h, and pretreated with NaB for 30 min prior to treatment of high cholesterol for 24 h. The expression levels of SOD1, SOD2, catalase, and GPX4 were analyzed by western blot. ß-actin was used as a loading control. n = 3. *p < 0.05 versus control with NT siRNA transfection, #p < 0.05 versus high cholesterol treatment with NT siRNA transfection. b Cells were pretreated with NaB for 30 min prior to treatment of high cholesterol for 24 h. mRNA expression levels of SOD1, SOD2, catalase, and GPX4 were analyzed by quantitative real-time PCR. Data were normalized by the ACTB mRNA expression levels. n = 4. *p < 0.05 versus control, #p < 0.05 versus high cholesterol treatment. c Cells were pretreated with NaB for 30 min prior to treatment of high cholesterol for 24 h. The expression levels of SOD1, SOD2, catalase, and GPX4 were analyzed by western blot. ß-actin was used as a loading control. n = 4. *p < 0.05 versus control, #p < 0.05 versus high cholesterol treatment. d Brains of 20 weeks mice fed with HFD for 14 weeks were extracted. The expression levels of SOD1 were analyzed by western blot. ß-actin was used as a loading control. n = 3. *p < 0.05 versus control. e Cells were pretreated with NaB and ATN-224 (4 µM) for 30 min prior to treatment of high cholesterol for 72 h where ROS with DCF-DA was measured by flowcytometer. Total cell counts = 1.0 × 104 cells. n = 4. f Cells were pretreated with NaB and ATN-224 for 30 min prior to treatment of high cholesterol for 24 h. The expression levels of BACE1 were analyzed by western blot. ß-actin was used as a loading control. n = 4. g Cells were pretreated with NaB and ATN-224 for 30 min prior to treatment of high cholesterol for 24 h and immunostained with BACE1 antibody. Scale bars are 8 µm (magnification, × 1,000). n = 3. H Cells were pretreated with NaB and ATN-224 for 30 min prior to treatment of high cholesterol for 72 h. Aß concentration of medium samples was detected by using ELISA kit. Data are presented as a mean ± S.E.M. n = 4. *p < 0.05 versus control, #p < 0.05 versus high cholesterol treatment, $p < 0.05 versus treatment of high cholesterol and NaB. All blot and immunofluorescence images shown are representative.