Heat shock protein 70 is a potential virulence factor in murine toxoplasma infection via immunomodulation of host NF-kappa B and nitric oxide

Abstract

We propose that the 70-kDa heat shock protein (HSP70) protects virulent Toxoplasma gondii from the effects of the host by immunomodulation. This hypothesis was tested using quercetin and antisense oligonucleotides targeting the start codon of the virulent T. gondii HSP70 gene. Oligonucleotides were transiently transfected into two virulent (RH, ENT) and two avirulent (ME49, C) strains of T. gondii, significantly reducing HSP70 expression in treated parasites. Virulent parasites with reduced HSP70 expression displayed reduced proliferation in vivo, as measured by the number of tachyzoites present in spleens of infected mice. They also exhibited an enhanced rate of conversion from tachyzoites to bradyzoites in vitro. Our results implicate HSP70 as a means by which virulent strains of T. gondii evade host proinflammatory responses: when RAW 264.7 cells were exposed to parasites with reduced HSP70 expression, differential expression of inducible NO synthase (iNOS) and cell NO production were observed between infections with normal and HSP70-deficient T. gondii. iNOS message levels were significantly increased when host cells were infected with HSP70 reduced virulent tachyzoites and HSP70-related inhibition of iNOS transcription resulted in altered host NO production by virulent T. gondii infection. Virulent parasites expressing reduced levels of HSP70 initiated significantly more NF-kappa B activation in host splenocytes than infections with untreated parasites. Neither proliferative ability nor conversion from tachyzoites to bradyzoites was affected by lack of HSP70 in avirulent strains of T. gondii. Furthermore, avirulent T. gondii strains induced high levels of host iNOS expression and NO production, regardless of HSP70 expression in these parasites, and inhibition of HSP70 had no significant effects on translocation of NF-kappa B to the nucleus. Therefore, the 70-kDa parasite stress protein may be part of an important survival strategy by which virulent strains down-regulate host parasiticidal mechanisms.