[Effects of PIM1 Gene on Proliferation, Apoptosis and JAK2/STAT3 Signaling Pathway of Acute Myeloid Leukemia U937 Cells].

OBJECTIVE: To investigate the effects of the serine/threonine kinase family member 1 (PIM1) gene on the proliferation and apoptosis of acute myeloid leukemia (AML) U937 cells, and the regulation effect on Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) pathway. METHODS: Bone marrow mononuclear cells from newly diagnosed adult AML patients and patients with iron deficiency anemia were collected and PIM1 mRNA expression was detected by RT-qPCR. AML cell line U937 cells were divided into U937 group (U937 cells were cultured normally), Si-PIM1 group (U937 cells were transfected with low expression adenovirus vector containing PIM1 mRNA), Si-NC group (U937 cells were transfected with low expression adenovirus vector without PIM1 mRNA), coumermycin A1 (CoA1) group (JAK2 activator CoA1 was added to U937 cells at a concentration of 20 µmol/L), and Si-PIM1+CoA1 group (U937 cells were transfected with adenoviral vector containing low expression of PIM1 mRNA and added with CoA1 at a concentration of 20 µmol/L). After culture for 24 h, the expressions of PIM1 mRNA and protein, JAK2/STAT3 pathway, cell cycle and apoptosis-related proteins in U937 cells were detected by RT-qPCR and Western blot, the cell proliferation activity was detected by MTT assay, and flow cytometry was used to detect cell cycle changes and apoptosis rate. RESULTS: The PIM1 mRNA expression level in bone marrow mononuclear cells in AML patients was higher than that in patients with iron deficiency anemia (P < 0.05). Compared with U937 group, PIM1 mRNA and protein, phosphorylated JAK2 (p-JAK2)/JAK2, phosphorylated STAT3 (p-STAT3)/STAT3, Cyclin D1, cyclin-dependent kinase 2 (CDK2) protein, cell proliferation activity, S phase and G 2/M phase proportions were decreased in Si-PIM1 group (all P < 0.05), while p27, Caspase-3 protein, G0/G1 phase proportion and apoptosis rate were increased (all P < 0.05). However, the changes of above indicators in CoA1 group were just opposite to those in Si-PIM1 group, indicating that CoA1 could reverse the effect of Si-PIM1 on U937 cells. There were no significant differences in above indexes of U937 cells between U937 group, Si-PIM1+CoA1 group and Si-NC group (P >0.05). CONCLUSION: Knockdown of PIM1 gene expression can inhibit U937 cell proliferation and promote apoptosis, in order to alleviate ALM process, which may be related to the inhibition of JAK2/STAT3 pathway activation.

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

OBJECTIVE: 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. METHODS: Human AML THP-1 cells were cultured in vitro and divided into control (normally cultured THP-1 cells), pcDNA3.1 ï¼»THP-1 cells transfected with IGHG1 overexpression (pcDNA3.1-IGHG1) negative control plasmidï¼½, pcDNA3.1-IGHG1 (THP-1 cells transfected with plasmid containing pcDNA3.1-IGHG1), LY364947 ï¼»transforming growth factor-ß (TGF-ß)/signal transduction protein (Smad) inhibitor LY364947 20 µmol/L treated THP-1 cellsï¼½, si-NC ï¼»THP-1 cells transfected with IGHG1-small interfering RNA (siRNA) negative controlï¼½, 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-ß/Smad signaling pathway related proteins. RESULTS: 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-ß1, 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-ß/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-ß1, 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-ß/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-ß1, 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). CONCLUSION: 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-ß/Smad pathway activation.