Early Secreted Antigenic Target of 6 kDa (ESAT-6) Protein of Mycobacterium tuberculosis Induces Interleukin-8 (IL-8) Expression in Lung Epithelial Cells via Protein Kinase Signaling and Reactive Oxygen Species

Early secreted antigenic target of 6 kDa (ESAT-6) of Mycobacterium tuberculosis is critical for the virulence and pathogenicity of M. tuberculosis. IL-8, a major chemotactic cytokine for neutrophils and T lymphocytes, plays important roles in the development of lung injury. To further understand the role of ESAT-6 in lung pathology associated with tuberculosis development, we studied the effects of ESAT-6 on the regulation of IL-8 expression in lung epithelial cells. ESAT-6 induced IL-8 expression by increasing IL-8 gene transcription and mRNA stability. ESAT-6 induction of IL-8 promoter activity was dependent on nuclear factor-kappa B (NF-kappa B) and activator protein-1 (AP-1) binding and sensitive to pharmacological inhibition of PKC and ERK and p38 MAPK pathways. ESAT-6 activated ERK and p38 MAPK phosphorylation and rapidly induced reactive oxygen species (ROS) production. Dimethylthiourea but not mannitol inhibited IL-8 induction by ESAT-6, further supporting the involvement of ROS in the induction of IL-8 expression. Exposure of mice to ESAT-6 induced localized inflammatory cell aggregate formation with characteristics of early granuloma concomitant with increased keratinocyte chemoattractant CXCL1 staining in bronchiolar and alveolar type II epithelial cells and alveolar macrophages. Our studies have identified a signal transduction pathway involving ROS, PKC, ERK, and p38 MAPKs and NF-kappa B and AP-1 in the ESAT-6 induction of IL-8 expression in lung epithelial cells. This has important implications for the understanding of lung innate immune responses to tuberculosis and the pathogenesis of lung injury in tuberculosis.