Early Secreted Antigenic Target of 6 kDa of Mycobacterium tuberculosis Stimulates Macrophage Chemoattractant Protein-1 Production by Macrophages and Its Regulation by p38 Mitogen-Activated Protein Kinases and Interleukin-4

Early secreted antigenic target of 6 kDa (ESAT-6), the major virulence factor of Mycobacterium tuberculosis, affects host immunity and the formation of granulomas likely through inflammatory cytokines. To understand its role in this regard further, we investigated the effect of ESAT-6 on macrophages by determining the production of macrophage chemoattractant protein (MCP)-1, a major chemokine associated with tuberculosis pathogenesis, by murine bone marrow-derived macrophages (BMDMs) and its regulation by protein kinases and cytokines. The results revealed that ESAT-6, but not Ag85A and culture filtrate protein 10 kDa (CFP10), induced MCP-1 production by BMDMs dose and time dependently. Inhibition of p38 but not other mitogen-activated protein kinases (MAPK) and PI3K further enhanced ESAT-6-induced MCP-1 production by BMDMs. Inhibition of p38 MAPK enhanced ESAT-6-induced MCP-1 mRNA accumulation without affecting mRNA stability. ESAT-6 also induced TNF-alpha from BMDMs and MCP-1 from mouse lung epithelial cells, and these were suppressed by p38 MAPK inhibition, implying cytokine- and cell-specific effect of p38 MAPK inhibition on ESAT-6-induced MCP-1 by macrophages. Pretreatment of BMDMs with IL-4, but not other cytokines (IL-2, IL-10, TNF-alpha, IFN-gamma and IL-1 alpha) further elevated ESAT-6-stimulated MCP-1 production although IL-4 did not induce MCP-1 without ESAT-6. Both p38 MAPK inhibitor and IL-4 did not show additive effect on ESAT-6-induced MCP-1 protein level despite such effect on MCP-1 mRNA level was evident. In conclusion, these results indicate a specific role for both p38 MAPK and IL-4 in ESAT-6-induced MCP-1 production by macrophages and suggest a pathway with significance in tuberculosis pathogenesis.