Objective To investigate the microRNAs(miRNAs) expression profiles in the bowels of patients with Hirschsprung disease (HSCR), and to explore the role of differentially expressed miRNAs in the pathogenesis of HSCR. Methods Twenty-seven HSCR tissues, including spastic segments and distending segments, were obtained from patients with HSCR during operation.Then miRNA microarrays were used to investigate the miRNA expression profiles in 6 HSCR specimens.Bioinformatics software was used to predict target genes of miRNA.Three miRNAs (miR-145-3p, miR-4505 and miR-1260a) were chosen and quantificational real-time(qRT)-PCR was performed to verify the different expression of those three miRNAs in 27 HSCR tissues. Results The expression of 26 miRNAs in an aganglionic colon segment was found to be more than two fold greater than that in ganglionic segment tissues, including 19 up-regulated miRNAs and 7 down-regulated miRNAs in patients with HSCR (all P<0.05). Target genes of miRNAs were found, such as SOX10, RET, L1CAM.qRT-PCR showed the expression of miR-145-3p(1.42±0.42, aganglionic segment vs 0.90±0.31, ganglionic segment) and miR-4505(1.30±0.30, aganglionic segment vs 0.76±0.22, ganglionic segment)displayed a statistical difference between groups (all P<0.001). Besides, the expressions of miR-145-3p(1.53±0.46, long-segment type vs 1.16±0.12, short-segment type)and miR-4505(1.42±0.26, long-segment type vs 1.00±0.16, short-segment type) showed a statistical difference between different types (all P<0.001), but miR-1260a (1.11±0.25, aganglionic segment vs 0.99±0.21, ganglionic segment)did not show differential expression between different groups(P=0.064). Conclusions Abnormal expression of miRNAs was found in HSCR spastic segments, suggesting that miRNAs may be involved in the pathogenesis of HSCR. Key words: Hirschsprung disease; microRNA; Gene expression profiling