Lon Mutant of Brucella abortus Induces Tumor Necrosis Factor-Alpha in Murine J774.A1 Macrophage.

ABSTRACT: OBJECTIVES:The objective of this study was to isolate a Brucella lon mutant and to analyze the cytokine response of B. lon mutant during macrophage infection. METHODS:A wild-type Brucella abortus strain was mutagenized by Tn5 transposition. From the mouse macrophage J774.A1 cells, total RNA was isolated at 0 hours, 6 hours, 12 hours, and 24 hours after infection with Brucella. Using mouse cytokine microarrays, we measured transcriptional levels of the cytokine response, and validated our results with a reverse transcriptase-polymerase chain reaction (RT-PCR) assay to confirm the induction of cytokine messenger RNA (mRNA). RESULTS:In host J774.A1 macrophages, mRNA levels of T helper 1 (Th1)-type cytokines, including tumor necrosis factor-alpha (TNF-?), interferon-gamma (IFN-?), interleukin-2 (IL-2), and IL-3, were significantly higher in the lon mutant compared to wild-type Brucella and the negative control. TNF-? levels in cell culture media were induced as high as 2 ?g/mL after infection with the lon mutant, a greater than sixfold change. CONCLUSION:In order to understand the role of the lon protein in virulence, we identified and characterized a novel B. lon mutant. We compared the immune response it generates to the wild-type Brucella response in a mouse macrophage cell line. We demonstrated that the B. lon mutants induce TNF-? expression from the host J774.A1 macrophage.

SUBMITTER: Park S 

PROVIDER: S-EPMC3922098 | biostudies-literature | 2013 Dec

REPOSITORIES: biostudies-literature