Effects of immune inhibitory receptor LILRB2 on proliferation and apoptosis of leukemia cell lines

Abstract

Objective To explore the expression, function, and relevant mechanism of the immune inhibitory receptor of leukocyte immunoglobulin-like receptors subfamily B member 2( LILRB2) in human leukemia cell lines and to provide new clues for the targeting treatment of leukemia. Methods T he expressions of LILRB2 in leukemia cell lines, such as T HP1, U937, and K562 were detected by the flow cytometry and W estern blotting. T he shRNA plasmids specific for the knockdown of LILRB2( with GFP tag) was constructed and by which the virus was packaged. T HP1 cells with high expression of LILRB2 were then infected by the virus. C ell lines with steady expression of GFP+were obtained by the flow cell sorting. T he knockdown efficiency of shRNA at both mRNA level and protein level was detected. T he changes of cell proliferation and apoptosis at different time points were observed and the relevant molecular mechanisms, such as changes of downstream regulatory signals, were analyzed after the knockdown of LILRB2. Results All three cell lines expressed LILRB2 and the expression level of LILRB2 of T HP1 cells was the highest. T he Real-T ime PC R and W estern blotting findings showed that among four constructed shRNAs plasmids, shLILRB2-717 and shLILRB2-1312 significantly downregulated the expression of LILRB2. T he inhibition of LILRB2 expression in T HP1 cells dramatically decreased the proliferation and significantly increased the apoptosis. T he results of the flow cytometry indicated that the apoptotic rates of cells of the shLILRB2-717 group and shLILRB2-1312 group at the early stage were( 7. 90 ±1. 61) % and( 24. 80 ± 1. 32) %, much higher than( 3. 96 ± 0. 48) % of the control group( Scramble group)( P 0. 05). T he apoptotic rates of cells of the two groups at the late stage were( 13. 80 ± 0. 98) % and( 23. 60 ±1. 03) %, much higher than( 10. 80 ± 0. 48) % of the Scramble group( P 0. 05). T he knockdown of LILRB2 led to significant down-regulation of expressions of SHP1 and p-SHP1, which indicated that LILRB2 might regulate the proliferation and apoptosis of leukemia cell lines through SHP1. Conclusion Immune inhibitory receptor LILRB2 can be expressed in a variety of leukemia cell lines. T he inhibition of LILRB2 expression can lead to significant down-regulation of SHP1 and p-SHP1 in T HP1 cells and mass apoptosis, which suggests that SHP1 may positively regulate the development of leukemia.