Effect of the tumor suppressor gene ING4 on the proliferation of MCF-7 human breast cancer cells

Abstract

Inhibitor of growth 4 (ING4) is a member of the ING family and acts as a tumor suppressor protein. To investigate the impact of ING4 on breast cancer proliferation, the present study examined the antitumor effects caused by upregulation in the expression of ING4 and its possible mechanism of effect in MCF-7 cells. A plasmid-based expression system encoding the ING4 gene was used to construct a stable cell line and overexpress ING4 in MCF-7 cells. Real-time PCR and western blot analysis were used to detect the mRNA and protein expression levels of ING4, respectively. Cell growth was examined by methylthiazolyltetrazolium (MTT) assay. Cell cycle distribution and cell apoptosis were measured by flow cytometry. The expression of p21, p53 and bax genes were tested by real-time PCR and western blot analysis. The stably transfected cell lines pcDNA3.1(+)/ING4 (with the ING4 gene) and pcDNA3.1(+) (an empty vector) were established. The expression levels of ING4 mRNA and protein in the stable cell line expressing pcDNA3.1(+)/ING4 were significantly higher than those of the two control cell lines. The cell proliferation of stably transfected cells was inhibited, and the inhibitory rate was 62.58±2.93%. Based on the changes in cell cycle distribution in stably transfected cells compared with two control cell lines, a number of cells were blocked in the G0/G1 phase 67.82±3.78% (P<0.05). In addition, the apoptotic rate was significantly higher, at 31.51±3.02% (P<0.05). Real-time PCR revealed that p21 and bax mRNA expression were increased (P<0.01), but the expression of p53 did not change significantly (P>0.05) in the stably transfected cell lines. Western blot analysis results of p21, bax and p53 were in accordance with real-time PCR results. ING4 upregulation may inhibit breast cancer cell proliferation and accelerate the process of apoptosis. It is suggested that ING4 plays a significant role in the suppression of breast cancer progression.