Release of reactive oxygen intermediates by dengue virus-induced macrophage cytotoxin

Abstract

Dengue type 2 virus (DV) induces a subpopulation of T lymphocytes of mice to produce a cytokine, cytotoxic factor (mCF), which induces H-2A positive macrophages to produce macrophage cytotoxin (CF2). The present study was undertaken to investigate the mechanism of cytotoxicity of CF2. It was observed that CF2 induced production of superoxide anion (O^-₂) and hydrogen peroxide (H₂O₂) by the spleen cells of mice in vitro and in vivo. The maximum production of O^-₂ (260 ± 10 nM/4 × 10⁶ cells) was at 45 minutes while that of H₂O₂ was at 90 minutes after inoculation of CF2. Pretreatment of mice or spleen cells with anti-CF2-antisera inhibited O^-₂ and H₂O₂ production in a dose-dependent manner. Superoxide dismutase (SOD) inhibited O^-₂ production and cytotoxicity while H₂O₂ production was increased by increasing SOD concentration in the culture. This indicated that O^-₂ production is necessary for the cytotoxic activity of CF2. Pretreatment of the cells with Ca²⁺ channel blocking drugs, nifedipine or verapamil, inhibited CF2- induced O^-₂ and H₂O₂ production in a dose-dependent manner. We have shown earlier that the cytotoxic activity of CF2 is known to be Ca²⁺ dependent and CF2-induced production of nitrite and the cytotoxicity is inhibited by N^G-monomethyl-l-arginine. Thus, it is suggested that O^-₂ and nitrite are necessary for cell killing by CF2 in a Ca²⁺ manner and the killing may possibly be by generation of peroxynitrite.