We aimed to ascertain the role of microRNAs (miRNAs) in regulating human endometrial cancer stem cells (HuECSCs). The expression level of miRNA-134 (miR-134), a member of the DLK1-DIO3 genomic imprinted miRNA cluster, differed significantly between HuECSCs and human endometrial cancer cells (HuECCs). miR-134 inhibited HuECSCs proliferation and migration by targeting protein O-glucosyltransferase 1 (POGLUT1) expression. Exogenous miR-134 overexpression downregulated POGLUT1 and Notch pathway proteins in HuECSCs in vitro. miR-134 overexpression affected the G2/M phase of HuECSCs and suppressed the growth of xenograft tumours formed. Thus, endogenous miR-134 regulation in HuECSCs may suppress tumourigenesis in human endometrial carcinoma.