Role of miR‑let‑7c‑5p/c‑myc signaling axis in the committed differentiation of leukemic THP‑1 cells into monocytes/macrophages

Ruijing Sun; Chaozhe Wang; Yufang Wang; Yunhua Wu; Pengchao Du; Xiaolin Sun; Qing Li; Kehong Bi; Guosheng Jiang

doi:10.3892/ol.2023.13989

Role of miR‑let‑7c‑5p/c‑myc signaling axis in the committed differentiation of leukemic THP‑1 cells into monocytes/macrophages

Oncol Lett. 2023 Jul 31;26(3):403. doi: 10.3892/ol.2023.13989. eCollection 2023 Sep.

Authors

Ruijing Sun¹, Chaozhe Wang¹, Yufang Wang², Yunhua Wu¹, Pengchao Du¹, Xiaolin Sun³, Qing Li³, Kehong Bi⁴, Guosheng Jiang^{1

5}

Affiliations

¹ Department of Immunology, School of Basic Medical Sciences, Binzhou Medical University, Yantai, Shandong 264003, P.R. China.
² Department of Laboratory Medicine, Fushan District People's Hospital, Yantai, Shandong 265500, P.R. China.
³ Department of Laboratory Medicine, Zibo First Hospital, Zibo, Shandong 255200, P.R. China.
⁴ Department of Hematology, The First Affiliated Hospital of Shandong First Medical University (Shandong Academy of Medical Sciences), Jinan, Shandong 250062, P.R. China.
⁵ Department of Precision Molecular Laboratory Medicine, Zhangqiu District People's Hospital of Jinan Affiliated to Jining Medical University, Jinan, Shandong 250200, P.R. China.

Abstract

In a preliminary experiment, it was found that c-myc expression was decreased following the differentiation of THP-1 cells into monocytes/macrophages induced by phorbol 12-myristate 13 acetate (PMA) + lipopolysaccharide (LPS) + interferon (IFN)-γ. The expression of miR-let-7c-5p was then found to be elevated by cross-sectional analysis using TargetScan and PubMed and differential microarray analysis. The present study aimed to investigate the role of the miR-let-7c-5p/c-myc signaling axis in the committed differentiation of THP-1 leukemic cells into monocytes/macrophages induced by PMA + LPS + IFN-γ. Human THP-1 leukemic cells were induced to differentiate into monocytes/macrophages by PMA + LPS + IFN-γ. Following induction for 48 h, the growth density of the THP-1 cells was observed directly under an inverted microscope, cell proliferation was measured using Cell Counting Kit-8 assay and the cell cycle and the expression of differentiation-related antigens (CD11b and CD14) were measured using flow cytometry. The mRNA expression of miR-let-7c-5p and c-myc was detected using reverse transcription-quantitative PCR and the protein expression of c-myc was detected using western blot analysis. Dual luciferase reporter gene analysis was used to detect the targeted binding of miR-let-7c-5p on the 3'UTR of c-myc. The relative expression of miR-let-7c-5p and c-myc genes in THP-1 cells induced by PMA + LPS + IFN-γ was found to be up- and downregulated respectively, and expression of miR-let-7c-5p was negatively correlated with the expression of c-myc gene. Dual luciferase reporter gene assays confirmed that miR-let-7c-5p targeted the 3'UTR of c-myc and inhibited luciferase activity. Following transfection with miR-let-7c-5p mimics, the expression of c-myc was markedly downregulated and the proliferative ability of the THP-1 cells was decreased, while the expression rate of CD11b and CD14 was significantly increased. The rescue experiment revealed that the effects of miR-let-7c-5p mimics on the proliferation and differentiation of THP-1 cells were attenuated by transfection with c-myc overexpression vector. Together, the findings of the present study demonstrated that miR-let-7c-5p can target the 3'UTR region of c-myc and that the miR-let-7c-5p/c-myc signaling axis is one of the critical pathways involved in the directional differentiation of leukemic cells into monocytes/macrophages.

Keywords: c-myc; induced differentiation; leukemic cells; macrophage; microRNA-let-7c-5p.

Grants and funding

The present study was supported by the ‘Twelfth Five-Year’ National Science and Technology Support Program (grant no. 2013BAI07B02), Natural Science Foundation of China (grant no. 81573467), Natural Science Foundation of Shandong (grant nos. ZR2020QH160 and ZR2021MH080), The Foundation for Jinan's Clinical Science and Technology Innovation (grant no. 202134001). Cultivation Fund of the first affiliated hospital of Shandong First Medical University (Shandong Qianfoshan Hospital; grant no. QIPY2020NSFC0819).