1F,G). As described above, it has been reported that the scirrhous find more type of cancers

undergo EMT.11 To evaluate whether or not the EMT-related genes are expressed in S-HCCs, we applied the core EMT interactome genes that have been previously identified elsewhere.19 When we performed the GSEA, significant enrichment of the EMT core gene signature (EMT_CORE_UP; n = 91) was observed in S-HCCs, compared to HCCs (Fig. 4A). This finding was further validated by examining the expression of individual EMT-related genes by the real-time PCR method. The expression of the key EMT molecules, Snail and Twist, was significantly higher in S-HCCs than in HCCs (P < 0.05) (Fig. 4B,C; Supporting Table 3). Protein expression of Snail was significantly more prevalent in S-HCCs (10 of 14; 71%) and CCs (11 of 19; 58%) than in HCCs (6 of 24; 25%) (P =

0.006) (Fig. 4D). In immunohistological evaluation, the expression of Snail protein was mainly localized to the periphery of the tumor nests, next to the fibrous stroma this website (Fig. 4E; Supporting Fig. 2). In addition, Snail and K19/EpCAM expression was colocalized in most S-HCCs (8 of 10 double-positive cases; 80%), which were mainly located on the periphery of the tumor nests, next to the fibrous stroma cells (Fig. 4F). These findings support that the expression of EMT genes are not merely the attributes of fibrotic components, but the results of the cross-talk interaction

between epithelial and fibrotic components within the tumors. We also observed that mRNA levels of Snail and Twist were well correlated with those of SPC markers, including selleckchem K19, EpCAM, cMET, CD133, and Oct3/4, suggesting the distinctive association of EMT with the stem-cell-like features of S-HCC (Supporting Fig. 3). TGF-β binds two types of receptors, TGFβRI and TGFβRII, to form active signaling complexes. The phosphorylated TGFβRI transmits signals intracellularly by phosphorylating the transcription factors, Smad2 and Smad3, which then forms a complex with Smad4. The Smad complex moves into the nucleus, where it regulates the expression of target genes.20 TGF-β is known to promote tumor-cell invasion and metastasis as a potent stimulator of EMT.21-23 With this concern, we next evaluated whether the TGF-β signaling is differentially expressed among tumor subtypes according to the expression of EMT-related molecules. We observed that the expression of TGF-β-signaling molecules, including TGF-β, TGFβRI, and Smad4, was significantly higher in S-HCCs than in HCCs or CCs (P < 0.05) (Fig. 5A-D; Supporting Table 3). Smad4 protein was expressed in most S-HCCs (12 of 14; 86%), but not significantly more than other types (Fig. 5E).