Screening Of Biomarkers In Brain Tissue By Metabonomics Based On Mice Model Infected By Borrelia Garinii SZ

Lyme disease(LD),also called as Lyme Borreliosis,which is a tick-borne of natural zoonotic disease infections caused by Borrelia burgdorferi.Metabonomics is used to analyze low molecular weight metabolites based on a particular organism or cell in a specific physiological period,and then try to find metabolites and physiological and pathological changes in the relative relationship between the subjects.This study was focused on biomarkers selection by metabolomics analysis and to study the potential pathogenic mechanism of LD infection.1.To determine the mouse brain tissue successful infection with Lyme disease spirochetes and pathological changes,80 mice BALB / c female mice were randomly divided into four experimental groups and four control groups,each group has 10 mice.The experimental group was infected with 0.2 ?l Borrelia garinii SZ strains and the control group was infected with 0.2 ?l PBS.The brain tissues were harvested at the 7d,14 d,28d and 35 d of each one experimental and control group.Partial brain tissue was cultured with BSK-H,the microscopic examination showed that the spirochetes growth well after one week.At the same time,brain tissues pathology slices were also done at 4 time points.After slide fixed hematoxylin-eosin stained,the results showed that the experimental group compared with the control group had obvious pathological changes,including the neuronal volume increased,the fibrous tissue dissolution and other pathological changes.It indicated that mice have been successfully infected with Lyme disease spirochetes and have obvious pathological changes.The remaining brain tissues were then used to perform metabolic analysis.2.To test the sample quality and explore the sample gas chromatography-mass spectrometry with the best metabolite extraction method.At first,we should do preliminary experiments.The samples of the brain tissue were divided into 8 groups,10 samples in each group were divided into four time points.The experimentalmethod was well carried out by metabolites extraction,derivatization treatment,sample detection and system stability test.The results showed that the samples were good quality and the instrument platform is good and the system is stable.We can proceed with formal testing.3.In order to screen the latent biomarkers of metabolic differences in the brain tissue of mice infected with the Borrelia garinii SZ strain.A total of 32 metabolites were screened out by 8 groups of formal testing after data pretreatment,principal component analysis,orthogonal partial least squares-discriminant analysis,differential compound screening and identification.The 32 biomarkers identified by this experiment were identified as potential biomarkers.4.To further understand these 32 potential biomarkers are involved in which metabolic pathways.A total of 64 KEGG numbers were obtained from 32 potential biomarkers through a localized database and matched to the metabolic pathways of the corresponding substances through the KEGG pathway site.Because of the large number of access diagrams,we use the Cytoscape software to enrich the control channel of the top 10 and integrated the mapping of the biomarkers selected in this experiment,and then consult the relevant literature to study the interaction between substances and pathways.The results show that the main metabolomic pathway of Lyme disease spirochete is glycolytic pathway,Lyme disease spirochete only rely on glycolysis to produce ATP,the bacteria can not encode some complete access to the implementation of such as fatty acids,amino acids,nucleic acid biosynthesis.This study laid a foundation for searching pathogenesis of Lyme disease spirochetes.