Function Of Leptospires Lipopolysaccharide And The Role Of TLR4 Receptor On The Pathogenicity Of Leptospira

Leptospirosis is one of the most widespread zoonosis caused by infection with pathogenic Leptospira species.At present,the pathogenic mechanism of leptospirosis has not been elucidated.LPS was extracted from two leptospiral strains,i.e.the virulent strain 56606v and the attenuated strain 56606a by hot phenol method.Molecular weight of LPS of the two strains was compared using SDS-PAGE and Pro-Q?Emerald300 staining.The results showed that strain 56606a has an LPS of relatively smaller molecular weight than v-LPS.ELISA and Real-time PCR were performed to detect cytokine expression in bone marrow-derived macrophages of mice stimulated by LPS of the two Leptospira strains.Results showed higher activity of a-LPS than v-LPS.Laser scanning confocal microscopy was then used to observe the phagocytosis of 1?m beads coated with LPS by macrophages.Results showed that phagocytosis of a-LPS and v-LPS presented no significant difference.In addition,v-LPS was found to be different from the previously reported leptospiral LPS,primarily acting on TLR4,rather than TLR2,which has been regarded as the major receptor recognizing leptospiral LPS.In order to further study the host immune response in leptospirosis,we established a murine model of acute infection by infecting C57BL/6 mice with L.interrogans strain 56606v for the first time,with symptoms of jaundice and hemorrhage typical of severe human leptospirosis.WT and TLR4^-/-mice were infected with L.interrogans strain 56606v at the same time.Comparison of pathological changes and Leptospira burden in tissues were performed by HE staining,immunohistochemistry and RT-PCR.Results showed that Leptospira burden in TLR4^-/-mice was dramatically higher than WT mice,with more severe jaundice and liver injury.However,WT mice displayed higher expression of inflammatory cytokines and more severe hemorrhage.In conclusion,pathogenicity of Leptospires is possibly associated with differences in the structure and biological activity of L-LPS.For the first time,we successfully established a mouse model of acute infection of leptospirosis.We confirmed that TLR4 effectively modulate host immune response and control in vivo proliferation of leptospires through mediating cytokine production,thus influencing the severity of jaundice and liver injury caused by leptospiral burden,and hemorrhage caused by host immunopathological injury.Our study provides the experimental basis for further studies into the role of LPS-mediated signaling pathways in the pathogenesis of Leptospira infection...