Identification of microRNA-18a as a novel regulator of the insulin-like growth factor-1 in the proliferation and regeneration of deer antler

To investigate the effect of miR-18a on the regulation of the insulin-like growth factor (IGF-1) during growth of antlers in sika deer, miRNA Chip analysis, Target Scan and real-time PCR analysis were used to identify miRNAs that bind to the 3′-UTR of IGF-1. An miR-18a mimic was transfected into antler cartilage cells and the expression levels were quantified by real-time PCR. Dual luciferase assays revealed that miR-18a binds to the 3′-UTR of the IGF-1 gene thus indicating this to be a target gene regulated by miR-18a. MTT assays and cell-cycle analyses confirmed that miR-18a significantly inhibited proliferation of cartilage cells. In contrast, transfection of miR-18a inhibitors increased proliferation. Furthermore, Western blot analysis showed that over-expression of miR-18a down-regulated IGF-1 protein levels while IGF-1 expression was increased after transfection of miR-18a inhibitors. Thus, miR-1 down-regulated IGF-1 expression thus implicating miR-18a as an important regulator of antler proliferation.