Numerous studies have shown that aberrant microRNA (miRNA) expression is associated with the development and progression of various types of human cancer and serum miRNAs are potential biomarkers. This study examined whether some commonly deregulated miRNAs in hepatocellular carcinoma (HCC) are presented in serum of patients with HCC and can serve as diagnostic markers. Serum miRNAs (miR-21, miR-122, and miR-223) were quantified by real-time quantitative RT-PCR in 101 patients with HCC and 89 healthy controls. In addition, 48 patients with chronic type B hepatitis were also analyzed for comparison. We found that the median levels of miR-21, miR-122, and miR-223 were significantly higher in patients with HCC than those in healthy controls (P = 7.48 × 10−13, P = 6.93 × 10−9, and P = 3.90 × 10−12, respectively). However, these elevated serum miRNAs were also detected in patients with chronic hepatitis (P = 2.05 × 10−12, P = 4.52 × 10−16, and P = 1.65 × 10−11, respectively). Moreover, serum miR-21 and miR-122 in patients with chronic hepatitis were higher than in patients with HCC (P = 3.99 × 10−4 and P = 4.97 × 10−8), although no such significant difference was found for miR-223. Receiver-operator characteristic (ROC) curve analyses suggest that these serum miRNAs may be useful markers for discriminating patients with HCC or chronic hepatitis from healthy controls, but not patients with HCC from patients with chronic hepatitis. Our results indicate that serum miR-21, miR-122 and miR-223 are elevated in patients with HCC or chronic hepatitis and these miRNAs have strong potential to serve as novel biomarkers for liver injury but not specifically for HCC. © 2010 Wiley-Liss, Inc.