Abstract:

Objective To discuss the effect of CD40 ligand （CD40L） on the biological behavior of the human monocytic leukemia THP-1 cells through long non-coding RNA（lncRNA） linc00239，and to clarify its potential mechanism. Methods The linc00239 over-expression vector （pcDNA-linc00239） and interference vector （sh-linc00239） were constructed and transfected into the THP-1 cells.Real-time fluorescence quantitative PCR （RT-qPCR） method was used to detect the transfection efficiency. The THP-1 cells were divided into control group， vector group， pcDNA-linc00239 group， sh-linc00239 group， vector+CD40L group， pcDNA-linc00239+CD40L group， and sh-linc00239+CD40L group. RT-qPCR method was used to detect the expression levels of linc00239 in the cells in various groups； CCK-8 assay was used to detect the proliferation activities of the cells in various groups；flow cytometry was used to detect the percentages of the cells at different cell cycles and the apoptotic rates of the cells in various groups；RT-qPCR and Western blotting methods were used to to detect the expression levels of B-cell lymphoma-2 （Bcl-2） and Bcl-2-associated X protein （Bax） mRNA and proteins in the cells in various groups； Western blotting method was used to detect the expression levels of protein kinase B （AKT） and phosphorylated AKT （p-AKT） proteins in the cells in various groups，and the ratio of p-AKT/AKT was calculated. Results Compared with vector group， the proliferation activity of the cells and the percentage of the cells at G₂ phase in pcDNA-linc00239 group were significantly increased （P<0.05 or P<0.01）， the expression levels of linc00239， Bcl-2 mRNA and protein， and the ratio of p-AKT/AKT were significantly increased （P<0.05 or P<0.01），the percentage of the cells at G₁ phase， apoptotic rate， and expression levels of Bax mRNA and protein in the cells were significantly decreased （P<0.05）； compared with vector group， the proliferation activity of the cells and percentage of the cells at G₂ phase， expression levels of linc00239， Bcl-2 mRNA and protein， and ratio of p-AKT/AKT in the cells in sh-linc00239 group and vector+CD40L group were significantly decreased （P<0.05 or P<0.01）， while the percentage of the cells at G₁ phase， apoptotic rate， and the expression levels of Bax mRNA and protein in the cells were significantly increased （P<0.05 or P<0.01）；compared with pcDNA-linc00239 group， the proliferation activity of the cells and percentage of cells at G₂ phase in pcDNA-linc00239+CD40L group were significantly decreased （P<0.05 or P<0.01）， the expression levels of linc00239， Bcl-2 mRNA and protein，and ratio of p-AKT/AKT were significantly decreased （P<0.05 or P<0.01），while the percentage of cells at G₁ phase， apoptotic rate， and the expression levels of Bax mRNA and protein were significantly increased （P<0.05 or P<0.01）；compared with sh-linc00239 group， the proliferation activity of the cells and percentage of cells at G₂ phase in sh-linc00239+CD40L group were significantly decreased （P<0.05 or P<0.01）， the expression levels of linc00239， Bcl-2 mRNA and protein， and ratio of p-AKT/AKT were significantly decreased （P<0.05 or P<0.01），and the percentage of the cells at G₁ phase， apoptotic rate， and expression levels of Bax mRNA and protein were significantly increased （P<0.05 or P<0.01）. Conclusion CD40L can inhibit the proliferation and cell cycle progression of the THP-1 cells through linc00239 and induce the apoptosis.

Key words: CD40 ligand, Long non-coding RNA, linc00239, Acute myeloid leukemia, Cell cycle, Apoptosis