40; intermediate differentiation, n = 15, AFC = 3.25; P = 0.0081). Finally, we determined a possible association of ABC expression with tumor size. Up-regulation of ABCB6 and ABCC2 was significantly higher in patients with tumors <30 mm than in patients with tumors >31 mm (<30 mm, n = 4; >31 mm, n = 15), with AFC

values of respectively 4.6 and 2.3 for ABCB6 (P = 0.0144) and 4.2 and 1.5 for ABCC2 (P = 0.0022). Rapamycin We hypothesized that ABC gene expression might be regulated by cellular miRNAs, i.e., ABC genes up-regulation in HCC would be the consequence of the down-regulation of cellular miRNAs. In order to obtain miRNA expression signatures, RNA was isolated from 10 HCC and three HL samples. To minimize variation in the miRNA profile, only 10 HCC with alcohol etiology were selected from the 19 available (FR01, FR03, FR05, FR06, FR07, FR08, FR10, FR11, FR14, and FR18). miRNA expression was determined by Taqman 384-well microfluidic array including 378 cellular miRNAs and six control wells and data Caspase inhibitor were normalized to mammalian

U6 RNA. In total, 361 out of 378 miRNAs were detectable. Changes in miRNA expression between 2-and 40-fold were considered up-regulation and changes between 0- and 0.5-fold were considered down-regulation. Average miRNA expression was compared in HCC and HL groups by two-tailed t test. miRNA expression in HCC compared with HL control was significantly higher for 11 cellular miRNAs and lower for 79 miRNAs, which accounted for respectively 3% and 22% of the detectable miRNAs (Fig. 2; Fig. S2). Analysis of the conservation of the 90 dysregulated miRNAs revealed that 87 were conserved up to the mouse, and 25 up to the chicken (Table S6). Next a subset of miRNAs quantified with the microfluidic array was cross-examined on all samples: 19 paired HCC and AHL, and three HL. Six miRNAs were selected: miR-135b, miR-145, miR-199a-3p/a/b, and miR-296 because they were consistently down-regulated in the 10 HCC patient samples (low standard deviation). Expression of these six miRNAs was quantified using single miRNA Taqman assays (Fig. 3). First, miRNA expression

in HL from pancreatic 上海皓元 cancer patients and AHL from liver cancer patients was similar (Fig. 3). This indicated that the miRNA profile is not affected in HL tissues despite the different background of these samples. Second, differences in miRNA expression observed between paired AHL and HCC samples were significant for miR-145, miR-199a-3p, miR-199a-5p, and miR-199b, hence confirming the miRNA signature in HCC from the microfluidic array. These differences were also significant for miR-135b when the two patients presenting the highest variation in each group are excluded (Fig. 3; miR-135b; FR01; FR12, FR13, and FR19; P = 0.0070). miR-296 presented a down-regulated profile but the differences were not statistically significant.