The Effects of IGHG1 on the Proliferation and Apoptosis of Human Acute Myeloid Leukemia THP-1 Cells

Abstract

To investigate the effects of the immunoglobulin G1 heavy chain constant region (IGHG1) on the proliferation and apoptosis of acute myeloid leukemia (AML) THP-1 cells and its possible mechanism of action. Human AML THP-1 cells were cultured in vitro and divided into control (normally cultured THP-1 cells), pcDNA3.1 ［THP-1 cells transfected with IGHG1 overexpression (pcDNA3.1-IGHG1) negative control plasmid］, pcDNA3.1-IGHG1 (THP-1 cells transfected with plasmid containing pcDNA3.1-IGHG1), LY364947 ［transforming growth factor-β (TGF-β)/signal transduction protein (Smad) inhibitor LY364947 20 μmol/L treated THP-1 cells］, si-NC ［THP-1 cells transfected with IGHG1-small interfering RNA (siRNA) negative control］, si-IGHG1 (THP-1 cells transfected with IGHG1-siRNA), and si-IGHG1+LY364947 (IGHG1-siRNA and LY364947 co-treated THP-1 cells) a total of 7 groups. Fluorescence quantitative PCR was used to detect the expression of IGHG1 and immunoglobulin G (IgG) mRNA of THP-1 cells in each group; CCK-8 was used to detect THP-1 cells proliferation activity; flow cytometry was used to detect THP-1 cells apoptosis and cell cycle in each group; Western blot was used to detect the THP-1 cells proliferation, apoptosis and the expression of TGF-β/Smad signaling pathway related proteins. Compared with the control group, after overexpression of IGHG1, the expression of IGHG1 and IgG mRNA, cell proliferation viability, S phase cell ratio, expressions of Cyclin D1, B cell lymphoma-2 (Bcl-2), IgG, TGF-β1, phosphorylated Smad3 (p-Smad3)/Smad3 protein in THP-1 cells were significantly increased (P<0.05), the apoptosis rate, G0/G1 phase cell ratio, expression of p21, Bcl-2 related X protein (Bax), Caspase-3 protein were significantly reduced (P<0.05); after inhibiting TGF-β/Smad signaling pathway or silencing IGHG1, the expression of IGHG1 and IgG mRNA, cell proliferation viability, S phase cell ratio, expression of Cyclin D1, Bcl-2, IgG, TGF-β1, p-Smad3/Smad3 protein in THP-1 cells were significantly reduced (P<0.05), the apoptosis rate, G0/G1 phase cell ratio, expressions of p21, Bax, and Caspase-3 protein were significantly increased (P<0.05); and compared with silencing IGHG1, after co-treatment of IGHG1 gene silencing and TGF-β/Smad pathway inhibition, the expression of IGHG1 and IgG mRNA, cell proliferation viability, S phase cell ratio, expressions of Cyclin D1, Bcl-2, IgG, TGF-β1, p-Smad3/Smad3 protein in THP-1 cells were significantly reduced (P<0.05), the apoptosis rate, G0/G1 phase cell ratio, expression of p21, Bax, and Caspase-3 protein were significantly increased (P<0.05). Silencing IGHG1 gene can down-regulate the expression of IgG, inhibit the proliferation of human AML THP-1 cells, block cell cycle progression, and induce cell apoptosis; its mechanism may be related to the inhibition of the TGF-β/Smad pathway activation.