Project description:Transcriptional profiling of NHDF Cells comparing control untreated fibroblasts with fibroblasts coincubated with three different species of the Borrelia burgdorferi sensu lato group. Overall design: Four-conditions experiment (unstimulated NHDF; NHDF stimulated by Borrelia burgdorferi sensu stricto strain IBS19; NHDF stimulated by Borrelia garinii strain IBS6; NHDF stimulated by Borrelia garinii strain IBS17). There is 6 Biological replicates for each condition (i.e. 6 different batches of NHDF cells from 6 independent human donor), independently grown and harvested.
Project description:The similarity of Lyme borreliosis to other diseases and the complex pathogenesis cause diagnostic and therapeutic difficulties. Changes at the cellular and molecular level after Borrelia sp. infection remain still poorly understood. Therefore, the present study focused on the gene expression in human dermal fibroblasts in differentiation of infection with Borrelia garinii, Borrelia afzelii and Borrelia burgdorferi sensu stricto spirochetes. For microarray analysis 10 samples were used: 3 control samples - K, 2 samples of NHDF cells infected with Borrelia garinii - G, 2 samples of NHDF cells infected with Borrelia afzelii - A and 3 samples of NHDF cells infected with Borrelia burgdorferi sensu stricto - SS.
Project description:miRNA profiles of astrocytes infected with Borrelia burgdorferi for 24 hours, 48 hours, and 24 hour uninfected controls were generated by deep sequencing, in duplicate, using Illumina MiSeq. Overall design: miRNA profiles of astrocytes infected with Borrelia burgdorferi for 24 hours, 48 hours, and 24 hour uninfected controls were generated by deep sequencing, in duplicate, using Illumina MiSeq.
Project description:Background: Lyme borrelia genotypes differ in their capacity to cause disseminated disease. Gene array analysis was employed to profile the host transcriptome induced by Borrelia burgdorferi strains with different capacities for causing disseminated disease in the blood of C3H/HeJ mice during early infection. Results: Borrelia burgdorferi B515, a clinical isolate that causes disseminated infection in mice, differentially regulated 236 transcripts (P<0.05 by ANOVA, with fold change of at least 2). The 216 significantly induced transcripts included IFN-responsive genes and genes involved in immunity and inflammation. In contrast, B. burgdorferi B331, a clinical isolate that causes transient skin infection but does not disseminate in C3H/HeJ mice, stimulated changes in only a few genes (1 induced, 4 repressed). Transcriptional regulation of type I IFN and IFN-related genes was measured by quantitative RT-PCR in mouse skin biopsies collected from the site of infection 24 hours after inoculation with B. burgdorferi. The mean values for transcript of Ifnb, Cxcl10, Gbp1, Ifit1, Ifit3, Irf7, Mx1, and Stat2, were found to be significantly increased in B. burgdorferi strain B515-infected mice relative to the control group. In contrast, transcription of these genes was not significantly changed in response to B. burgdorferi strain B331 or B31-4, a mutant that is unable to disseminate. Conclusions: These results establish a positive association between the disseminating capacity of B. burgdorferi and early type I IFN induction in a murine model of Lyme disease.
Project description:Borrelia burgdorferi, the agent of Lyme disease, promotes pro-inflammatory changes in endothelium that lead to the recruitment of leukocytes. The host immune response to infection results in increased levels of IFN-gamma in the serum and lesions of Lyme disease patients that correlate with greater severity of disease. Therefore, the effect of IFN-gamma on the gene expression profile of primary human endothelial cells exposed to B. burgdorferi was determined. B. burgdorferi and IFN-gamma synergistically augmented the expression of 34 genes, seven of which encode chemokines. Six of these (CCL7, CCL8, CX3CL1, CXCL9, CXCL10, and CXCL11) attract T lymphocytes, and one (CXCL2) is specific for neutrophils. Synergistic production of the attractants for T cells was confirmed at the protein level. IL-1beta, TNF-alpha, and LPS also cooperated with IFN-gamma to induce synergistic production of CXCL10 by endothelium, indicating that IFN-gamma potentiates inflammation in concert with a variety of mediators. An in vitro model of the blood vessel wall revealed that an increased number of human T lymphocytes traversed endothelium exposed to B. burgdorferi and IFN-gamma, as compared to unstimulated endothelial monolayers. In contrast, addition of IFN-gamma diminished the migration of neutrophils across B. burgdorferi-activated endothelium. IFN-gamma thus alters gene expression by endothelium exposed to B. burgdorferi in a manner that promotes recruitment of T cells and suppresses that of neutrophils. This modulation may facilitate the development of chronic inflammatory lesions in Lyme disease. Experiment Overall Design: Human umbilical vein endothelial cells (HUVEC) were stimulated with Interferon-gamma (IFN-g), Borrelia burgdorferi or both IFN-g and Borrelia or were left unstimulated. Affymetrix HGU133 plus 2.0 slides were used in duplicate for each condition.
Project description:RNA was isolated from late-log pahse wild-type Borrelia burgdorferi B31 and the bb0647 mutant grown in BSKH media at 37 degree, 5% CO2. cDNA was synthesized, labeled and hybridized to the 70mer oligonucleotide B. burgdorferi array. Slides were scanned using axon scanner and image were analyzed using GenePix 6.0. Dta were further analysed using the professional software Acuity 4.o, based on a ratio-based normalization. Samples were labeled by either Cy3 or Cy5. 5 hybs were performed, including dye-swaps.
Project description:We performed comparative transcriptomic analysis of the outer membrane vesicles (OMVs) released from B. burgdorferi. We identified a total of ~1200 unique transcripts with at least one mapped read from the bacterial cell and its OMVs. Overall design: Sequencing of ribosomal depleted Borrelia burgdorferi strain B31 and its OMVs isolated using two methods, ultracentrifugation and size-exclusion chromatography