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Ageing of the immune system, immunosenescence, is characterized by impaired lymphopoiesis, especially B-lymphocyte maturation, and is a hallmark of chronic heart failure (CHF). MicroRNAs (miRNAs) are non-coding, small RNAs, which post-transcriptionally control gene expression of multiple target genes. The miR-181 family is known to control haematopoietic lineage differentiation. Here, we study the role of the miR-181 family in immunosenescence and CHF.We conducted a clinical study analysing peripheral blood (PB) for miRNA expression and leucocyte distribution of young healthy controls (25 ± 4 years; n = 30), aged healthy controls (64 ± 5 years; n = 13), and age-matched CHF patients (64 ± 11years; n = 18). The expression of miR-181 family members was reduced, whereas miR-34a was increased in PB of aged individuals. In particular, miR-181c was further reduced in age-matched CHF patients. In PB, we observed reduced numbers of lymphocytes, in particular cytotoxic T cells and B cells, with rising age, and the expression of miR-181 correlated with the number of B cells. Notably, in CHF patients, ischaemic heart failure was associated with a further reduction of total B cells as well as their subpopulations, such as memory B cells, compared with age-matched healthy volunteers.Ageing- and CHF-associated changes in PB leucocyte subsets are paralleled by alterations in the expression of miRNAs involved in lymphopoiesis, which might play an important role in the age-related and CHF-mediated dysregulation of immune functions resulting in immunosenescence. Furthermore, miR-181c may serve as a marker for reduced immune functions in CHF patients.