Browsing by Author "Johnson, Raymond M."

Now showing 1 - 6 of 6
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    Elucidating the interaction of Borrelia burgdorferi OspC with phagocytes in the establishment of lyme borreliosis
    (2015-03-20) Carrasco, Sebastian Eduardo; Yang, X. Frank; Serezani, C. Henrique; Blum, Janice Sherry, 1957-; Johnson, Raymond M.; Bauer, Margaret E.
    Lyme disease, the most prevalent vector-borne illness in the United States, is a multisystem inflammatory disorder caused by infection with the spirochete Borrelia burgdorferi (Bb). This spirochete is maintained in nature through an enzootic cycle involving ticks and small mammals. The Bb genome encodes a large number of surface lipoproteins, many of which are expressed during mammalian infection. One of these lipoproteins is the major outer surface protein C (OspC) whose production is induced during transmission as spirochetes transition from ticks to mammals. OspC is required for Bb to establish infection in mice and has been proposed to facilitate evasion of innate immunity. However, the exact biological function of OspC remains elusive. Our studies show the ospC-deficient spirochete could not establish infection in NOD-scid IL2rγnull mice that lack B cells, T cells, NK cells, and lytic complement, whereas the wild-type spirochete was fully infectious in these mice. The ospC mutant also could not establish infection in SCID and C3H mice that were transiently neutropenic during the first 48 h post-challenge. However, depletion of F4/80+ phagocytes at the skin-site of inoculation in SCID mice allowed the ospC mutant to establish infection in vivo. In phagocyte-depleted SCID mice, the ospC mutant was capable to colonize the joints and triggered neutrophilia during dissemination in a similar pattern as wild-type bacteria. We then constructed GFP-expressing Bb strains to evaluate the interaction of the ospC mutant with phagocytes. Using flow cytometry and fluorometric assay for phagocytosis, we found that phagocytosis of GFP-expressing ospC mutant spirochetes by murine peritoneal macrophages and human THP-1 cells was significantly higher than parental wild-type Bb strains, suggesting that OspC has an anti-phagocytic property. This enhancement in phagocytosis was not mediated by MARCO and CD36 scavenger receptors and was not associated with changes in mRNA levels of TNFα, IL-1β, and IL-10. Phagocytosis assays with HL60 neutrophil-like cells showed that uptake of Bb strains was independent to OspC. Together, our findings reveal that F4/80+ phagocytes are important for clearance of the ospC mutant, and suggest that OspC promotes spirochetes' evasion of macrophages in the skin of mice during early Lyme borreliosis.
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    Modeling the transcriptome of genital tract epithelial cells and macrophages in healthy mucosa versus mucosa inflamed by Chlamydia muridarum infection
    (Oxford University Press, 2015-12) Johnson, Raymond M.; Kerr, Micah S.; Department of Medicine, IU School of Medicine
    Chlamydia trachomatis urogenital serovars are intracellular bacteria that parasitize human reproductive tract epithelium. As the principal cell type supporting bacterial replication, epithelial cells are central to Chlamydia immunobiology initially as sentries and innate defenders, and subsequently as collaborators in adaptive immunity-mediated bacterial clearance. In asymptomatic individuals who do not seek medical care a decisive struggle between C. trachomatis and host defenses occurs at the epithelial interface. For this study, we modeled the immunobiology of epithelial cells and macrophages lining healthy genital mucosa and inflamed/infected mucosa during the transition from innate to adaptive immunity. Upper reproductive tract epithelial cell line responses were compared to bone marrow-derived macrophages utilizing gene expression microarray technology. Those comparisons showed minor differences in the intrinsic innate defenses of macrophages and epithelial cells. Major lineage-specific differences in immunobiology relate to epithelial collaboration with adaptive immunity including an epithelial requirement for inflammatory cytokines to express MHC class II molecules, and a paucity and imbalance between costimulatory and coinhibitory ligands on epithelial cells that potentially limits sterilizing immunity (replication termination) to Chlamydia-specific T cells activated with limited or unconventional second signals.
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    Nearly Fatal Case of Whipple's Disease in a Patient Mistakenly on Anti-TNF Therapy
    (American College of Gastroenterology, 2013-10) Klochan, Christen; Anderson, Teresa A.; Rose, Dusten; Dimitrov, Rosen K.; Johnson, Raymond M.; Department of Pathology and Laboratory Medicine, IU School of Medicine
    Whipple's disease is a rare cause of chronic diarrhea and abdominal pain that may be confused with inflammatory bowel disease. We report a Whipple's case misdiagnosed as Crohn's disease in which treatment with anti-tumor necrosis factor (anti-TNF) therapy led to nearly fatal progression. Lymph node tissue obtained during laparotomy for suspected bowel necrosis stained dramatically with periodic acid-Schiff (PAS), and electron microscopy showed a bacterium consistent with Trophyrema whipplei. The patient made a remarkable recovery complicated only by cholestatic hepatitis, which was likely a treatment-associated inflammatory response. This case serves as a reminder that all granulomatous infections should be considered prior to initiation of anti-TNF therapies.
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    Pediatric Kawasaki Disease and Adult Human Immunodeficiency Virus Kawasaki-Like Syndrome Are Likely the Same Malady.
    (Oxford UP, 2016-09) Johnson, Raymond M.; Bergmann, Kelly R.; Manaloor, John J.; Yu, Xiaoqing; Slaven, James E.; Kharbanda, Anupam B.; Department of Biostatistics, Richard M. Fairbanks School of Public Health
    Background. Pediatric Kawasaki disease (KD) and human immunodeficiency virus (HIV)+ adult Kawasaki-like syndrome (KLS) are dramatic vasculitides with similar physical findings. Both syndromes include unusual arterial histopathology with immunoglobulin (Ig)A+ plasma cells, and both impressively respond to pooled Ig therapy. Their distinctive presentations, histopathology, and therapeutic response suggest a common etiology. Because blood is in immediate contact with inflamed arteries, we investigated whether KD and KLS share an inflammatory signature in serum.Methods. A custom multiplex enzyme-linked immunosorbent assay (ELISA) defined the serum cytokine milieu in 2 adults with KLS during acute and convalescent phases, with asymptomatic HIV+ subjects not taking antiretroviral therapy serving as controls. We then prospectively collected serum and plasma samples from children hospitalized with KD, unrelated febrile illnesses, and noninfectious conditions, analyzing them with a custom multiplex ELISA based on the KLS data.Results. Patients with KLS and KD subjects shared an inflammatory signature including acute-phase reactants reflecting tumor necrosis factor (TNF)-α biologic activity (soluble TNF receptor I/II) and endothelial/smooth muscle chemokines Ccl1 (Th2), Ccl2 (vascular inflammation), and Cxcl11 (plasma cell recruitment). Ccl1 was specifically elevated in KD versus febrile controls, suggesting a unique relationship between Ccl1 and KD/KLS pathogenesis.Conclusions. This study defines a KD/KLS inflammatory signature mirroring a dysfunctional response likely to a common etiologic agent. The KD/KLS inflammatory signature based on elevated acute-phase reactants and specific endothelial/smooth muscle chemokines was able to identify KD subjects versus febrile controls, and it may serve as a practicable diagnostic test for KD.
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    The role of STAT1 in Chlamydia-induced type I interferon responses in oviduct epithelium
    (2013-12-10) Hosey, Kristen L.; Derbigny, Wilbert A; Blum, Janice Sherry, 1957-; Goebl, Mark, 1958-; Johnson, Raymond M.; Kaplan, Mark H.
    Progression of Chlamydia into upper reproductive tract epithelium and the induction of subsequent immune responses to infection are major contributors to Chlamydia-induced pathogenesis of the genital tract. We reported that C. muridarum infection of the oviduct epithelial cells (OEs) secrete IFN-β in a TLR3 dependent manner. However, we showed that the C. muridarum infected TLR3-deficient OEs were still able to secrete minimal amounts of IFN-β into the supernatants, which is suggestive that there are other signaling pathways that contribute to Chlamydia-induced IFN-β synthesis in these cells. Previous studies describing the activation of the JAK/STAT signaling pathway during Chlamydia infection of cervical epithelial cells proposes a putative role for STAT1 in the synthesis of type I IFNs during Chlamydia infection. The present study investigated the role of STAT1 in Chlamydia-induced IFN-β production in OEs. OEs were infected with Chlamydia muridarum and analyzed at 24 hours by RT-PCR and western blot to determine STAT1 expression. STAT (-/-) OEs were infected and IFN-β production measured by ELISA. Quantitative real-time PCR analyses were performed at 6 and 16 hour post-infection to elucidate the mechanisms involved in IFN-β production during infection. Fluorescent microscopy was used to observe changes in Chlamydia replication. STAT1 activation and expression were significantly increased in wild-type (WT) OEs upon infection. TLR3 (-/-) OEs showed diminished STAT1 protein activation and expression. Augmented STAT1 protein expression corresponded to STAT1 mRNA levels. ELISA analyses revealed significantly less IFN-β production in infected STAT1 (-/-) OEs compared to WT OEs. Quantitative real-time PCR data showed that gene expression of IFN-β and of type I IFN signaling components were significantly increased during late stage Chlamydia infection, dependent on STAT1. Temporal regulation and increases in expression of IFN-α subtypes during infection were STAT1-dependent. Our results implicate STAT1-mediated signaling as a contributor to the C. muridarum-induced synthesis of IFN-β and other type I IFNs in OEs. We previously described a major role for TLR3 in the early-stage Chlamydia-induced synthesis of IFN-β in OEs; the results from this study suggest a role for STAT1 in the synthesis of type I IFNs that occurs during early and late stages of infection.
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    The role of the Borrelia oxidative stress regulator protein in virulence gene expression of the Lyme disease spirochete
    (2014-02-25) Khoo, Joleyn Yean Chern; Yang, X. Frank; Spinola, Stanley M.; Bauer, Margaret E.; Johnson, Raymond M.
    The Lyme disease agent, Borrelia burgdorferi, has a complex system that allows it to thrive in the harsh and distinct environments of its tick vector and mammalian host. Although it has been known for some time that the Borrelia oxidative stress regulator protein (BosR) plays a necessary role in mammalian infectivity and functions as a transcriptional regulator of alternative sigma factor RpoS, very little is known about its mechanism of action, other than the suggestion that BosR activates rpoS transcription by binding to certain upstream regions of the gene. In our studies, we performed protein degradation assays and luciferase reporter assays for further understanding of BosR function. Our preliminary findings suggest that BosR is post-transcriptionally regulated by an unknown protease and may not need to bind to any rpoS upstream regions in order to activate transcription. We also describe the construction of luciferase reporter systems that will shed light on BosR’s mechanism of action. We postulate the provocative possibility that unlike its homologs Fur and PerR in other bacterial systems, BosR may not utilize a DNA-binding mechanism in order to fulfill its role as a transcriptional regulator to modulate virulence gene expression.