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To further elucidate the expression, regulation and function of Signaling Lymphocytic Activation Molecule Family (SLAMF) protein members in human monocytes and macrophages. Un-differentiated monocytic THP-1 cell (u-THP-1) and differentiated THP-1 macrophage (d-THP-1) were used as culture models in the study. Responses of cells to the differentiation agents phorbol ester (25 ng/ml) and TLR (Toll-like receptor) ligands were assessed. RT-PCR and Western blot analysis were used to determine mRNA and protein level. Pro-inflammatory cytokine mRNA expression levels and phagocytosis were used as functional markers. Data analyzed using t-test, one-way or two-way ANOVA followed by post hoc test. SLAMFs were differentially expressed in THP-1 cells. Differentiation of u-THP-1 to d-THP-1 led to significantly higher SLAMF7 mRNA and protein levels than other SLAMF. In addition, TLR stimuli increased SLAMF7 mRNA expression but not protein expression. Importantly, SLAMF7 agonist antibody and TLR ligands synergistically increased the mRNA expression levels of IL-1β, IL-6 and TNF-α, but had no effect on phagocytosis. SLAMF7 knocked-down in d-THP-1 significantly lowered TLR-induced mRNA expressions of pro-inflammatory markers. SLAM family proteins are differentially regulated by differentiation and TLRs. SLAMF7 enhanced TLR-mediated induction of pro-inflammatory cytokines in monocytes and macrophages but not phagocytosis. [ABSTRACT FROM AUTHOR]

Abstract In the present study, we evaluated, under transient transfection conditions, five different cationic lipid-based transfection reagents on the activation of NF-κB, MAP kinases signaling pathways and induction of cytokines expression. We found that the reagents studied differentially regulated the NF-κB and the MAP kinases signaling pathways in the human THP-1 macrophage. Additionally, mRNA expression levels of the cytokines, IL-1β and TNF-α in THP-1 macrophage were also induced by selected test reagents. Hence, careful selection of cationic lipid-based transfection reagent for transient transfection is warranted, especially in studies of gene expression and function mediated through NF-κB- and MAP kinases-mediated pathways. Highlights • Cationic lipids-based transfection reagents trigger activation of NF-κB and MAP kinases pathways. • Cationic lipids-based transfection reagents up-regulate IL-1β, TNF-α mRNA. • The report cautions careful testing of transient transfection reagents when studying NF-κB- and MAP kinases-related pathways. [ABSTRACT FROM AUTHOR]