Whole-transcriptome analysis using the livers of NASH mice treated with CDDO-Me
We comprehensively analysed the RNA expression in the liver to elucidate the mechanism underlying the hepato-protective effects of CDDO-Me in CDAHFD-induced NASH mice, because CDDO-Me is involved in the various signalling pathways such as NF-κB or Akt inhibition in addition to Nrf2 activation. Among the 41,326 transcripts detected using RNA-seq analysis, 2,625 were significantly upregulated in NASH mice by more than 1.2-fold compared to those in the control mice (Figure 5a). Pathway analysis using the 130 genes upregulated by CDDO-Me in CDAHFD-fed mice showed activation of Nrf2 (gene symbol NFE2L2 )-mediated oxidative stress and xenobiotic metabolism-related pathways (Figure S6). Of the 2,625 upregulated genes, the expression of 195 genes was decreased below 80% by treatment with CDDO-Me (Figure 5a-c), and the CDDO-Me-regulated genes abundantly included leukocyte-related genes such as C5ar1 ,Ccl3 , Ccl4, Fcgr1 , and Lilra5 as shown in Table S3. Pathway analysis using these 195 downregulated genes showed that inhibition of cytokines caused a reduction in inflammatory pathways, such as recruitment and cell movement of leukocytes through NF-κB inhibition (Figure 5d) and immune cell trafficking (Figure 5e). CDAHFD-fed mice showed 167 downregulated genes (fold change < 0.8, p < 0.05) compared to those in control mice, and only 17 of the 167 genes were significantly increased by CDDO-Me treatment (Figure 5b). These data suggest that the inhibition of inflammatory cell recruitment may play an important role in the hepatoprotective effect of CDDO-Me.