MicroRNAs (miRNAs) are a recently discovered class of post-transcriptional regulators of gene expression with critical functions in health and disease. Their role in the pathogenesis of hypersplenism, however, is completely unknown. To determine whether miRNA expression is altered in splenic macrophages associated with hypersplenism due to portal hypertension in hepatitis-B-virus (HBV)-related cirrhosis, we analyzed the entire miRNAome in macrophages from normal and portal hypertensive spleen samples by microarray and Real-Time PCR. In this study, we identified 99 miRNA differences in expression in splenic macrophages associated with hypersplenism due to portal hypertension in HBV-related cirrhosis. Among the miRNAs identified in this study, hsa-miR-615-3p was significantly up-regulated in hypersplenism. Dynamic changes in miRNA expression occurred during the pathogenesis of portal hypertension-induced hypersplenism in HBV-related cirrhosis. The miRNAs then are novel regulatory RNAs in hypersplenism in patients with HBV-related cirrhosis.