Fig. 4

Effect of hinokiflavone on the transcriptome of N/P(14) cells. The cells were treated with hinokiflavone (5 µM) for 24 h. Total RNA was extracted using TRIzol reagent and subjected to next-generation sequencing (NGS) and bioinformatics analyses (n = 2). The gene set enrichment analysis was performed by using the GSEA software (version 4.0.3) (https://www.gsea-msigdb.org/gsea/index.jsp). The top 10 KEGG pathways displaying the most significant enrichment in differentially expressed genes were shown (a), and the base excision repair (BER) pathway is significantly enriched (b–c). (d) Effects of hinokiflavone on BER-related genes in N/P(14) cells. Cells were treated with hinokiflavone (5 µM) for 24 h and mRNA levels determined via RT-qPCR. Data are expressed as mean±S.D (n = 3). Statistical analyses were performed by unpaired two-tailed Student’s t-test. ***p < 0.001 compared with control group. (e–g) The effects of hinokiflavone (HNK) on the levels of BER-related proteins in N/P(14) cells were analyzed by western blotting. The band intensities of each protein were quantified using ImageJ software and normalized to that of GAPDH. Fold changes compared to the control (0) group are expressed as the mean±S.D. (n = 3). One-way ANOVA with Dunnett’s multiple comparison was used to evaluate statistical significance.