Interference of gene transfection of A20 zinc finger protein on expression of TNF-α、IL-12 and IL-10 in human monocytes

Objectives To observe effect of A20 expression on regulating the expressions of proinflammatory cytokine tumor necrosis factor-α(TNF-α) and interleukin(IL)-12,as well as its effect on the ratios of TNF-α / IL-10 and IL12 / IL-10 via transfecting A20 gene into human monocytes by cationic liposome,and to discuss the protective function of A20 zinc finger protein on monocyte inflammatory and potential regulation mechanism.Methods Ficoll separation medium was used to separate monocytes from peripheral blood.Monocytes were divided randomly into four groups : A group(control group);B group [lipopolysaccharide(LPS) intervention group];C group(A20 group) and D group(LPS and A20 intervention group).GFP gene was detected by fluorescence microscopy.Immunofluorescence and immunohistochemistry were employed to analyze the expression of A20 zinc finger protein.Reverse transcription polymerase chain reaction(RT-PCR) was used to detect A20 mRNA expression.Expression levels of supernatant TNF-α,IL-12 and IL-10 were detected by double antibody sandwich enzyme linked immunosorbent assay(ELISA).Results The mRNA and protein expressions of endogenous A20,TNF-α,IL-12 and IL-10 increased obviously after monocytes being stimulated by LPS(1 mg / L);ratios of TNF-α / IL-10 and IL-12 / IL-10 were significantly higher than those in control group(P 0.05).Comparing with control group,transfection of A20 gene into monocytes,mRNA and protein expressions of endogenous A20,TNF-α,IL-12 and IL-10 had no significant change in the absence of LPS stimulus conditions;ratios of TNF-α / IL-10 and IL-12 / IL-10 had no significant change either(P0.05).However,expressions of TNF-α and IL-12 were significantly lower in LPS and A20 intervention group than those in LPS group,whereas expression of IL-10 increased significantly,higher than those in control group and LPS group after monocytes of transfection A20 gene were stimulated by LPS(1 mg / L);ratios of TNF-α / IL-10 and IL-12 / IL-10 significantly decreased,which was lower than those in LPS group(P 0.05).Conclusions Expression of A20 has inflammatory activation dependency.Over-expression of A20 can inhibit expression of TNF-α and IL-12 and raise expression of inflammatory factor IL-10,thus inhibits the effect of inflammatory response by improving the balance between proinflammatory cytokine and anti-inflammatory cytokine.