Growing evidence has suggested that inhibitor of growth 4 (ING4), a novel member of ING family proteins, plays a critical role in the development and progression of different tumors via multiple pathways. However, the function of ING4 in human osteosarcoma remains unclear. To understand its potential roles and mechanisms in inhibiting osteosarcoma, we constructed an expression vector pEGFP-ING4 and transfected the human osteosarcoma cells using this vector. We then studied the effects of over-expressed ING4 in the transfected cells on the proliferation, apoptosis and invasion of the osteosarcoma cells. The up-regulation of ING4 in the osteosarcoma cells, arising from the stable pEGFP-ING4 gene transfection, was found to significantly inhibit the cell proliferation by the cell cycle alteration with S phase reduction and G0/G1 phase arrest, induce cell apoptosis via the activation of the mitochondria pathway, and suppress cell invasion through the down-regulation of the matrix metalloproteinase 2 (MMP-2) and MMP-9 expression. In addition, increased ING4 level evoked the blockade of NF-κB signaling pathway and down-regulation of its target proteins. Our work suggests that ING4 can suppress osteosarcoma progression through signaling pathways such as mitochondria pathway and NF-κB signaling pathway and ING4 gene therapy is a promising approach to treating osteosarcoma.