Fig 1: Treatment with CART upregulates the expression of Aß metabolism-associated enzymes in Aß1–42-exposed primary cortical neurons. (A) CART significantly increased cell viability in Aß-exposed cultured primary cortical neurons. Relative protein expression levels of IDE, NEP and LRP-1 in cultured primary cortical neurons exposed to Aß with or without CART were (B) determined via western blotting and (C) semi-quantified. (D) Relative mRNA expression levels of IDE, NEP, LRP-1 and RAGE in cultured primary cortical neurons exposed to Aß with or without CART were measured via reverse transcription-quantitative PCR. Data are presented as the mean ± SEM (n=6 per group). **P<0.01 and ***P<0.001 vs. control; #P<0.05, ##P<0.01 and ###P<0.001 vs. APP/PS1. CART, cocaine amphetamine regulated transcript; Aß, ß-amyloid protein; IDE, insulin-degrading enzyme; NEP, neprilysin; LRP-1, low-density lipoprotein receptor-related protein 1; RAGE, receptor for advanced glycation end products; WT, wild-type.
Fig 2: Silencing of circ-PRKDC inhibits tumor growth in vivo. SW480 cells transfected with sh-NC or sh-circ-PRKDC were subcutaneously injected into nude mice. (A and B) Tumor volume was measured every 4 days. (C) After the mice were killed, the xenograft tumors were weighed. (D and E) The levels of circ-PRKDC, miR-198 and DDR1 in xenograft tumors were tested using qRT-PCR or Western blot. (F) The levels of CyclinD1, ß-catenin and c-Myc in xenograft tumors were measured by Western blot. *P < 0.05, **P < 0.01, ****P < 0.0001.
Fig 3: DDR1 is a target of miR-198. (A) The predicted binding sites of miR-198 and DDR1 3'UTR were exhibited. (B) Luciferase activity was detected in SW480 and HCT116 cells introduced with WT-DDR1 3'UTR or MUT-DDR1 3'UTR and miR-NC or miR-198. (C and D) The mRNA and protein levels of DDR1 were measured in CRC tissues and normal tissues. (E) The correlation between miR-198 and DDR1 in CRC tissues was tested via Spearman correlation analysis. (F and G) DDR1 mRNA and protein levels were examined in FHC, SW480 and HCT116 cells. (H and I) RIP assay was used to confirm the relationship between miR-198 and DDR1. (J) Expression of miR-198 was detected in SW480 and HCT116 cells transduced with miR-NC or miR-198. (K–M) DDR1 mRNA and protein levels were measured in SW480 and HCT116 cells transfected with miR-NC, miR-198, anti-miR-NC or anti-miR-198. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.
Fig 4: HDAC1 promotes KLF4 expression by inhibiting NEP expression.A Co-expression analysis with MEM to determine the co-expression relationship between HDAC1 and NEP (p = 6.76e−05). B Co-expression analysis with MEM to determine the co-expression relationship between NEP and KLF4 (p = 6.76e−05). C A box plot from the expression data of the microarray dataset GSE3644 to determine expression of KLF4. The blue box on the left represents the expression of normal samples, and the red box on the right represents the expression of pancreatitis samples. D GeneMANIA predicted KLF4 related genes and constructed a PPI network diagram. The triangle gene in the middle is KLF4, and the outer circle gene is the predicted related gene. The redder the color of the gene, the higher the core degree, and vice versa the bluer the core, the lower the degree. E KOBAS was performed for KEGG enrichment analysis of KLF4 and its related genes. The vertical axis represents the enriched pathway. The horizontal axis and the size of the bubble represent the number of genes enriched on the pathway. Significance-logP value, the greater the significance, the redder the bubbles, and the smaller the significance, the bluer the bubbles. F Western blot analysis to detect the expression of NEP and KLF4 in AP. G Western blot analysis to detect the expression of HDAC1, NEP, and KLF4 in AP after silencing NEP. H Western blot analysis to detect the expression of NEP and KLF4 in AP. I ChIP detected the enrichment of HDAC1 and H3K27ac in the NEP promoter of each group. J ChIP detected the enrichment of HDAC1 and H3K27ac in the NEP promoter of each group after silencing NEP. K ChIP detected the enrichment of HDAC1 and H3K27ac in the NEP promoter of each group. L RT-qPCR detected HDAC1 mRNA expression. *p < 0.05 compared with the Mock, WT or sh-NC group. The comparison of the measurement data (mean standard ± deviation) between two groups were tested by independent sample t-test.
Fig 5: Downregulation of HDAC1 inhibits KLF4 expression by upregulating NEP expression thus promoting pancreatic acinar cell proliferation.A Western blot analysis was performed to detect HDAC1, NEP, and ATF4. B EdU was applied to detect pancreatic acinar cell proliferation after sh-HDAC1 or sh-NEP treatment. C TUNEL was performed to detect pancreatic acinar cell apoptosis after sh-HDAC1 or sh-NEP treatment. D Flow cytometry was performed to detect pancreatic acinar cell apoptosis after sh-HDAC1 or sh-NEP treatment. E Western blot analysis was performed to detect expression of Bcl-2, Bax, and cleaved-caspase3. F ELISA was conducted to test levels of IL-1ß, IL-6, TNF-a, and IL-10. *p < 0.05 compared with the sh-NC + oe-NC group; #p < 0.05 compared with the sh-HDAC1 + oe-NC group. The comparison of the measurement data (mean standard±deviation) among multiple groups was analyzed by one-way analysis of variance with Tukey’s post hoc test.
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