Surfactant protein A mediates mycoplasmacidal activity of alveolar macrophages by production of peroxynitrite

We have previously shown that surfactant protein A (SP-A) mediates in vitro killing of mycoplasmas by alveolar macrophages (AMs) from resistant C57BL/ 6 mice through a nitric oxide ( NO)-dependent mechanism. Herein, SP-A-defic ient [SP-A(-/-)] and inducible NO synthase; deficient [iNOS(-/-)] mice were infected intranasally with 10(5) or 10(7) colony-forming units of Mycoplas ma pulmonis, SP-A(-/-) mice were as susceptible to mycoplasmal infection as highly susceptible C3H/He mice, and far more susceptible than resistant C5 7BL/6 mice. iNOS(-/-) mice had significantly greater numbers of mycoplasmas and severity of lung lesions than iNOS(+/+) controls, In vitro, AMs isolat ed from C57BL/6 mice, activated with IFN-gamma, incubated with SP-A (25 mu g/ml), and infected with 10(10) colony-forming units of M. pulmonis, killed mycoplasmas within 6 h, Mycoplasmal killing was abrogated by 1,000 units/m l of copper-zinc superoxide dismutase, In the absence of AMs, incubation of M. pulmonis with the peroxynitrite generator 3-morpholinosynodiomine . HCl (SIN-1) effected complete killing of mycoplasmas by 90 min in a dose-depen dent manner. Addition of copper-zinc superoxide dismutase (3,000 units/ml), which converts SIN-1 to a NO donor, prevented this killing. Neither of the reactive oxygen species generated by xanthine oxidase (10 milliunits/ mi, plus 500 mu M xanthine and 100 mu M FeCl3), nor . NO generated by 1-propana mine-3-(2-hydroxy-2-nitroso-1-propylhydrazine (PAPA NONOate) (100 mu M) kil led mycoplasmas. These data establish that peroxynitrite generation by AMs is necessary for the killing of a pathogen in vitro and in vivo.