Downregulated ATF3 expression inhibited growth of adrenocortical carcinoma cells and investigation of its mechanisms

Objective To investigate the effect of downregulated activating transcription factor 3 (ATF3) expression on proliferation of adrenocortical carcinoma cells and its mechanisms. Methods Immunohistochemistry and Western blotting were used to detect the expression of ATF3 in human adrenocortical tumor tissues and cells. Adrenocortical carcinoma cells, Sw-13, and NCI-H259R cells, were transfected with siATF3 using lipidosome 2000, and expression of ATF3 mRNA was determined using RT-PCR; expression of ATF3, cleaved caspase 3, caspase 3, cleaved PARP, and PARP proteins were detected using Western blotting; cell growth inhibition rate and apoptosis rate were monitored using MTT and AnnexinV-FITC/PI, respectively. Sw-13 and NCI-H259R cells were treated with NVP-BEZ235, Perifosine, BKM120, IWP-2, PP2, KN93, Everolimus respectively followed by detected expression of ATF3 mRNA by realtime PCR. The effect of ATF3 on cell proliferation after inhibition of related signaling pathways were detected by MTT. Results The ATF3 in human adrenocortical gland tumor tissues and cells showed high expression. The levels of ATF3 mRNA and protein in Sw-13 and NCI-F259R cells transfected with siATF3 were significantly reduced. Compared with the negative control group (NC siRNA), siATF3 transfection significantly inhibited the proliferation of Sw-13 and NCI-F259R cells (P<0.05), and increased the apoptosis rate (P<0.05). Western blotting shown that the levels of cleaved caspase 3 and cleaved PARP protein in siATF3 transfected cells increased significantly; and realtime PCR results indicated that the expression of ATF3 mRNA was dramatically inhibited by PP2, KN93, and IWP-2 in NCI-F259R cells compared with control group (DMSO); but ATF3 significantly promoted the proliferation activity of NCI-F259R cells which treated by PP2, KN93, and IWP-2 signaling inhibitors. Conclusion High expression of ATF3 is existed in adrenocortical carcinoma cells. Downregulated ATF3 expression may inhibit cell proliferation and activate apoptosis pathway, resulting in apoptosis in Sw-13 and NCI-F259R cells, this mechanism of action is related to activating Wnt/β-catenin, CaMKI, and SRC pathway. (Chin J Endocrinol Metab, 2018, 34: 738-745) Key words: Adrenocortical carcinoma; Activating transcription factor 3; Signal pathway; Apoptosis; Mechanism