Endothelial Cells and Fibroblasts Amplify the Arthritogenic Type I IFN Response in Murine Lyme Disease and Are Major Sources of Chemokines inBorrelia burgdorferi-Infected Joint Tissue

Abstract

Localized elevation in Type I IFN has been uniquely linked to the severe Lyme arthritis that develops in C3H mice infected with the spirochete Borrelia burgdorferi. In this study, the dynamic interactions that result in generation of these responses were further examined in C3H mice carrying the Type I IFN receptor gene ablation, which effectively blocks all autocrine/paracrine signaling crucial to induction of downstream effectors. Reciprocal radiation chimeras between C3H and IFNAR1−/− mice implicated both radiation-sensitive and radiation-resistant cells of the joint tissue in the pro-arthritic induction of Type I IFN. Ex vivo analysis of cells from the naïve joint revealed CD45+ cells residing in the tissue to be uniquely capable of initiating the Type I IFN response to B. burgdorferi. Type I IFN responses were analyzed in real time by lineage sorting of cells from infected joint tissue. This demonstrated that myeloid cells, endothelial cells, and fibroblasts were responsible for propagating the robust IFN response, which peaked at Day 7 post infection and rapidly resolved. Endothelial cells and fibroblasts were the dominant sources of IFN signature transcripts in the joint tissue. Fibroblasts were also the major early source of chemokines associated with PMN and monocyte/macrophage infiltration, thus providing a focal point for arthritis development. These findings suggest joint-localized interactions among related and unrelated stromal, endothelial, and myeloid cell lineages that may be broadly applicable to understanding the pathogeneses of diseases associated with type I IFN signature, including systemic lupus erythematosus and some rheumatoid arthritides.

Borrelia burgdorferi Arthritis–Associated Locus Bbaa1 Regulates Lyme Arthritis and K/B×N Serum Transfer Arthritis through Intrinsic Control of Type I IFN Production

“…Reciprocal interval specific congenic lines (ISCL) for Bbaa1 on Chr4 were generated on both B6 and C3H backgrounds as described (32) and are indicated as B6.C3- Bbaa1 (9.32–94.97 Mbp) and C3.B6- Bbaa1 (3.58–150.8 Mbp) with introgressed region of Chr4 indicated in parentheses. B6-IFNAR1 −/− mice were provided by Dr. Murali-Krisna Kaja University of Washington, Seattle, WA, and backcrossed to C3H to generate C3H-IFNAR1 −/− mice as described (19). All mice were housed in the University of Utah Animal Research Center (Salt Lake City, UT) and followed protocols approved by the institutional review committee for the care and use of mice in biomedical research.…”

“…Ankle measurements were obtained using a metric caliper before and at 4 weeks of infection. Rear ankle joints were prepared for assessment of histopathology by removal of skin and fixation of the tissue in 10% neutral buffered formalin as described (19). Decalcified joints were embedded in paraffin, sectioned at 3μm, and stained with hematoxylin and eosin.…”

“…Ankle joints were excised, immersed in RNA Later (Qiagen) and stored at −80°C. Total RNA was recovered from homogenized tissue using Trizol reagent (Invitrogen) (19). RNA recovered from tissue and cells was reverse-transcribed and transcripts were quantified using a Roche LC-480 according to our previously described protocols (32).…”

“…Previously, using global gene expression analysis, we identified an IFN signature response in the joint tissue of B. burgdorferi -infected C3H mice that preceded the development of severe arthritis and was absent from mildly arthritic B6 mice (18). Blocking the Type I IFN receptor (INFAR1), either with a neutralizing monoclonal antibody (mAb) or by gene ablation, reduced the severity of arthritis in C3H mice, formally linking Type I IFN to Lyme arthritis (19, 20). Numerous investigators have studied the Type I IFN response to B. burgdorferi in murine and human myeloid cells, and identified pathogen receptor signaling pathways involved in this response (21–25).…”

Identification of the long, edited dsRNAome of LPS-stimulated immune cells

“…BMDMs were isolated and differentiated from femurs and tibias of 7-to 8-wk-old male C57BL/6 mice (Jackson Laboratory), as described (Lochhead et al 2012). Macrophages were differentiated for 7 d in differentiation media (RPMI 1640, Life Technologies; 20% horse serum, Life Technologies; 2 mM L-glutamine, Life Technologies; 20 µg/mL gentamicin; β-mercaptoethanol, Invitrogen; 30% L929 fibroblast supernatant containing macrophage-colony stimulating factor) and stored in a 37°C + 5% CO 2 humidified incubator.…”