Objective Anoikis resistance is closely associated with tumor invasion and metastasis. This study aimed at determining the effects of anoikis resistance on the proliferation and survival of U87 glioma cells and revealing their mechanisms in order to contribute to the understanding of glioma metastasis.Methods The enhanced green fluorescent protein-expressing human glioma cell line U87 was established to monitor the formation of cell aggregates in suspension culture. CCK-8 proliferation assay, colony formation assay, flow cytometry analysis of cell cycle, and real-time PCR were used to determine the effects of anoikis resistance on the proliferation and survival of U87 cells.Results The anoikis-resistant U87 cells had significantly increased cell proliferation, enhanced colony formation ability, significantly reduced percentage of cells in G1 phase, and significantly increased percentage of cells in S phase. Moreover, the stem cell-related genes OCT4 and SOX1 and proto-oncogene CCND1 were up-regulated in anoikis-resistant U87 cells (P<0.05).Conclusions Anoikis-resistant glioma cells are less dependent on survival environment with more robust proliferation and self-renewal capacity.