Study On The Mechanism Of Inflammasome Activation By MAPK2-knocked Out Toxoplasma Gondii

Toxoplasma gondii is a kind of specific intracellular opportunistic pathogenic parasite,which can infect all warm-blooded animals and lead to serious zoonosis.Toxoplasmosis is a worldwide important disease.For most healthy people,the clinical symptoms of T.gondii infections is invisibility,but it can cause devastating diseases in the developing fetus and immunocompromised individuals.Innate immunity plays an important role in protecting the host against Toxoplasma gondii infection,by regulating IL-1? and IL-18 maturation and activation to mediate inflammatory response.Mitogen activated protein kinase(MAPK)pathway,which is the major signal transduction system of Toxoplasma gondii,can regulate the growth of parasite.Research on the effect of mitogen-activated protein kinase 2(MAPK2)of T.gondii on inflammasome and the relative mechanism are of great significance.Research objective:To determine the effect of MAPK2 delection on T.gondii and the mechanism of virulence,and to lay the foundation for the prevention and treatment of Toxoplasmosis.Research method:1.Three groups of mice were infected intraperitoneally with ?mapk2,GT1 and PBS,to investigate the effect of MAPK2 deletion on parasite virulence in mice through observing the survial time of mice,assaying the serum levels of IL-12,IFN-?,TNF-a,and IL-6.2.RT-PCR and Western blotting to compare the difference of NF-?B and STAT activation between parental GT-1 or ?mapk2 parasites.Gene expression levels of IL-10 and IFN-? were detected by quantitative reverse transcription(RT)-PCR.Research results:1.The survival time was significant prolonged in the ?mapk2-infected mice in comparison with the mice infected with GT-1 parasites.We determined the levels of serum cytokines in mice infected with the different parasites,showing the parental GT-1 and ?mapk2 parasites could cause high and low levels of up-regulations of the proinflammatory cytokines IL-12,IFN-?,TNF-?,and IL-6.These results showed that ?mapk2 parasites tended to balance the pro-inflammatory and anti-inflammatory responses,which may be associated with the decreased virulence in mice.2.We determined the inflammatory cytokines in infected mice,showedthe ?mapk2 parasites induced low level of mature IL-18,NLRP1,NLRP3,ASC,and caspase-1 as compared to the parental GT-1 parasites.We also evaluated mRNA level of IL-10 in liver and spleen tissues of infected mice by qRT-PCR,revealing that IL-10 was significantly increased in mice infected with ?mapk2,compared with GT-1 parasites.Western blotting demonstrated that ?mapk2 parasites caused an increase in STAT1 phosphorylation,and a concomitant reduction in STAT3 phosphorylation in comparison with the parental GT-1 parasites,showed no significant difference of the activation of MAPK and NF-?B signaling in infected mice.Conclusion:In this study,?mapk2 parasites negatively regulates virulence of T.gondii by secreting high levels of IFN-? and IL-10 transcripts in infected mice.Suppression of inflammasome activation by IFN-? is through the enhancement of IL-10 production,which requires the STAT1-mediated synergistic action.IL-10,in turn,controls pro-IL-18 amounts in a STAT3 dependent manner.