Toll-like receptor 4 expression of human peripheral blood mononuclear cells treated by recombinant hIFN-α-2b-BCG and its role of immune activation

Abstract

To examine the toll-like receptor 4 (TLR4) expression of human peripheral blood mononuclear cells (hPBMC) treated with recombinant bacillus Calmette-Guérin (rBCG) and its role of immune activation. hPBMC was treated with recombinant human interferon (hIFN)-α-2b-BCG (rBCG) or wild BCG (wBCG) in vitro and the TLR4 expression detected by flow cytometry. The TLR4 functional blocking antibodies were applied for intervening the TLR4 signaling pathway of hPBMC. Then rBCG, wBCG, hIFN-α-2b and phosphate-buffered solution (PBS) were used to stimulate the hPBMC of blocking and non-blocking groups. The changes of human tumor necrosis factor-alpha (hTNF-α), human interleukin-12 (hIL-12) and hIFN-γ between the blocking and non-blocking groups by ELISA. The expression of TLR4 in hPBMC treated with rBCG or wBCG were stronger than that treat with PBS (all P < 0.05). In TLR4 non-blocking groups the expressions of hTNF-α and hIFN-γ were rBCG group > wBCG group > hIFN-α-2b group > PBS group, the expressions of hIL-12 was hIFN-α-2b group > PBS group > rBCG group > wBCG group (all P < 0.05). Application of TLR4 functional blocking antibodies to intervene hPBMC 48 h, comparing the changes in rBCG group, wBCG group, hIFN-α-2b group and PBS group, the expressions of hIFN-γ in TLR4 blocking groups (27.3 ± 1.2, 20.6 ± 0.9, 20.3 ± 0.8, 18.4 ± 0.7)were significantly inhibited than those in non-blocking groups (84.6 ± 1.3, 34.0 ± 1.0, 24.9 ± 0.9, 22.9 ± 0.7) (all P < 0.05). The expressions of hTNF-α in TLR4 blocking groups (1431 ± 28, 1032 ± 21, 104 ± 6, 109 ± 4) were significantly inhibited than those in non-blocking groups (1553 ± 28, 1065 ± 31, 343 ± 6, 299 ± 4) (all P < 0.05). The expressions of hIL-12 in TLR4 blocking groups (0.646 ± 0.005, 0.592 ± 0.015, 0.638 ± 0.008, 0.595 ± 0.019) were significantly inhibited than those in non-blocking groups (1.120 ± 0.012, 0.946 ± 0.015, 1.254 ± 0.011, 1.112 ± 0.024) (all P < 0.05). rBCG regulates the secretion of Th1 cytokines through the TLR4 signaling pathway.