MicroRNAs (miRNAs), important regulators of cellular processes, show specific expression signatures in different blood cell lineages and stages of hematopoietic stem cell (HSC) differentiation, indicating their role in the control of hematopoiesis. Because neonatal blood displays various features of immaturity, we might expect differential miRNA regulation. Herein, we determined miRNA expression profiles of umbilical cord blood (UCB) cell lineages and compared them to those of bone marrow (BM) and peripheral blood (PB) cell counterparts. Further, we determined mRNA expression profiles using whole-genome microarrays. An approach combining bioinformatic prediction of miRNA targets with mRNA expression profiling was used to search for putative targets of miRNAs with potential functions in UCB. We pointed out several differentially expressed miRNAs and associated their expression with the target transcript levels. miR-148a expression was suppressed in HSCs and its level inversely correlated with the previously verified target, DNA methyltransferase 3B, suggesting dependence of de novo DNA methylation in HSCs on miR-148a. Prolonged cell survival of UCB HSCs may be associated with low expression of miR-143 and miR-145 and up-regulation of their downstream targets (high expression of c-MYC and miR-17-92 and following repression of TGFBR2). In HSCs, we monitored significant up-regulation of eight miRNAs, which were previously verified as regulators of HOX genes. Further, miR-146b may be associated with immaturity of neonatal immune system because it is strongly up-regulated in UCB granulocytes and T lymphocytes compared to PB cell counterparts. Comparative analysis revealed 13 miRNAs significantly altered between UCB and BM CD34(+) cells. In UCB CD34(+) cells, we monitored up-regulation of miR-520h, promoting differentiation of HSCs into progenitor cells, and reduction of miR-214, whose expression might support HSC survival. In conclusion, UCB cells show specific miRNA expression patterns, indicating different regulation in these cells.