Neuropathogenecity Of Relapsing Fever Borrelia Hermsii In Mice

Background and purpose Relapsing fever(RF)is a multi-systemic disease caused by Borrelia hermsii(B.hermsii)infection.In addition to bacteremia and recurrent fever,it can also cause diseases in central nervous system(CNS).The symptoms of neuroborreliosis are similar to cold,which is easily misdiagnozed as viral influenza,leading to delay in treatment and having severe consequences.However,due to lack of disease models,mechanisms of this CNS disease caused by B.hermsii infection is not well understood.Our preious studies found that T lymphocytes play key roles in B.hermsii-caused CNS diseases in our well-established mouse model of neuroborreliosis.In addition,the disease syndrome can spontaeuously recover.Based on our previous work,our present study will continue to explore mechanisms of RF neuroborreliosis induced by B.hermsii infection,including 1)to characterize effects of RF B.hermsii infection on brain-blood barrier;2)to determine roles of IL-17 signaling in RF neuroborreliosis;3)to explore host factors involved in spontaneous recovery of RF neuroborreliosis.Methods Firstly,three kinds of common(live,inactivated and lytic)forms of B.hermsii were prepared.Peripheral blood and spleen lymphocytes were treated with these stimulators to make conditioned media.Mouse brain microvascular endothelial cells(bEnd3)were stimulated with these bacteria and media,respectively.Intracellular free radicals ROS and mitochondrial damage were analyzed by proper reagents,and Western Blot was used to detect the expression of ZO-1 protein in treated cells.Then,the microvascular permeability model of blood-brain barrier was established via human or mouse vascular endothelial cells.Sodium fluorescein was used to evaluate the permeability change before and after the stimulation of the cells.Next,wild-type C57BL/6 mice and IL-17A-deficient(IL17A-/-)mice were infected with B.hermsii.Bacteremia,cytokines in serum and CNS symptoms were analyzed and compared between these two kinds of mice in order to determine roles of IL-17 signaling in this CNS diseases.Lastly,comparisons of bacteremia,antibody and neurological symptoms among C57BL/6,Balb/c and ICR mice were performed to determine effects of host genetic backgrounds on B.hermsii infection.qRT-PCR,Western Blot and immunofluorescence were performed to analyze levels of IDO in microglia(BV-2)before and after B.hermsii treatments.Results In vitro stimulation of mouse brain microvascular endothelial cells(bEnd3)showed that B.hermsii treatments induced production of intracellular ROS,which may led to mitochondrial damage.Furthermore,these treatments also downregulated the expression of tight junctional protein ZO-1 in brain microvascular endothelial cells.B.hermsii treatments in the blood-brain barrier resulted in the increased permeability,indicated by sodium fluorecein.In order to explore roles of IL-17 signaling on neuropathogenecity of CNS disease caused by B.hermsii infection,IL-17A-deficient mice were infected with B.hermsii and compared with wild-type C57BL/6 mice.We found that within the period of 21-day post infection,IL-17A-/-mice showed no CNS disease symptoms but comparable bacteremia with wild-type mice.In contrast,wild-type C57BL/6 mice showed both severe CNS diseases and high bacteremia.B.hermsii infection in different genetic backgrounds of mice caused distinct phenotypes.C57BL/6 mice showed severe CNS diseases and recurrent bacteremia.Interestingly,Balb/c and ICR mice almost showed no CNS symptom but comparable bacteremia.Detection of B.hermsii-specific antibodies showed IgM but not IgG in infected C57BL/6 mice altered in accordance with bacterima,which was not observed in Balb/c and ICR mice.In vitro stimulation of microglia showed that mRNA levels of intracellular IDO increased after B.hermsii treatment.Western blot and immunofluorecene analysis also showed the increased protein level of IDO in cells at 24 h and 48 h post B.hermsii treatment.Conclusions RF B.hermsii stimulation of cerebral microvascular endothelial cells results in production of large amount of ROS,further damage of mitochondria and decease of tight junction protein ZO-1,which finally leads to increased permeability of blood-brain barrier.IL-17 signaling plays critical roles in the CNS diseases caused by B.hermsii infection.Upregulated mRNA and protein levels of IDO in microglia may be an important factor in allevated CNS symptom at the late stage of this disease model.Finally,genetic background of hosts may be another factor in affecting the CNS disease in this murine model of B.hermsii infection.