Neuroblastoma (NB) is childhood malignancy that retains characteristics of cancer stem cells (CSCs). Targeting the CSCs is one of the therapeutic strategies proposed to achieve complete remission of NB. beta-carotene (BC), an active precursor of retinoids, is a well-known antioxidant reported to possess anti-CSCs effects. Here, we investigated the involvement of retinoic acid receptors (RARs) in the anti-CSCs effects of BC. Treatment with BC or retinoic acid (RA) upregulated RAR beta mRNA expression in two NB cell lines. Inhibition of RAR beta using siRNA up-regulated gene expression of delta-like 1 homologue (DLK1), a marker of CSCs. To understand the molecular mechanisms of RAR beta-mediated inhibition of DLK1, four retinoic acid receptor elements (RAREs) were identified in the promoter of DLK1. Chromatin immunoprecipitation assays indicated that RAR beta bound directly to a RARE in the DLK1 promoter region. Knock-down of RAR beta also increased the self-renewal capacity of NB cells, which was suppressed by BC. Taken together, this study provided evidence that the therapeutic anti-CSCs effects of BC depend on RAR beta and its ability to interact with and down-regulate the CSCs marker, DLK1. (C) 2016 Elsevier Inc. All rights reserved.