Intracellular Infection by the Human Granulocytic Ehrlichiosis Agent Inhibits Human Neutrophil Apoptosis (original) (raw)
Related papers
Biochemical and Biophysical Research Communications, 1997
Recently, Goodman et al. (1) reported the culture of Human granulocytic ehrlichiosis (HGE) is an emergan HGE pathogen in human promyelocytic HL-60 leuing and occasionally fatal human infectious disease kemic cells. It was noted, however, that there was an whose pathogenesis is largely unknown. Goodman et apparent cytopathic effect associated with HGE infecal. (1) recently described the successful cultivation of tion that ultimately resulted in complete lysis of the the HGE infectious agent in human promyelocytic HL-HL-60 cells (1). The mechanism(s) responsible for 60 leukemic cells. It was reported in the same study HGE-mediated cell death has not been determined. that infectivity invariably led to host cell death, al-Herein, we show by flow cytometry that infection by though the mechanism by which HGE infection trigan HGE pathogen results in disruption of the cell cycle gers cellular self-destruction is as yet undetermined. phase distribution in HL-60 cells, primarily by blocking In this communication, we show that in vitro passage the G 1-to-S transition. In addition, using a variety of of HGE pathogen-infected blood elicits a significantly biochemical, immunohistochemical and flow cytometric dysfunctional G 1-to-S transition. Moreover, we provide techniques, we demonstrate that the HGE pathogen evidence that the cytopathic properties of the HGE triggers a cytopathic response in target cells by its abilpathogen are attributed to its ability to induce ity to induce apoptosis (programmed cell death). Inapoptosis in host HL-60 cells. Determination of specific protein expression changes by Western blot analysis duced cellular self-destruction correlates with reduced showed that HGE infection resulted in reduced expres-expression of PCNA, pRB and bcl-2. sion of PCNA and pRB, both of which play a role in cell cycling. Moreover, the steady state level of bcl-2, MATERIALS AND METHODS which protects eukaryotic cells against apoptosis, is suppressed by exposure to the HGE agent. These Blood samples were obtained from patients who had clinical sympresults suggest that this pathogen HGE induces toms suggestive of HGE. Six samples (8%) were able to infect HLapoptosis in HL-60 cells by a mechanism involving the 60 cells and showed characteristic intragranulocytic inclusions (morshut-off of multiple cell cycle and apoptosis regulatory ulae) in peripheral leukocytes. events.
2000
Human granulocytic ehrlichiosis (HGE) is an emerging febrile systemic disease caused by the HGE agent, an obligatory intracellular bacterium of granulocytes. The pathogenicity-and immunity-related mechanisms of HGE are unknown. In this study, several cytokines generated in human peripheral blood leukocytes (PBLs) incubated with the HGE agent or a recombinant 44-kDa major surface protein (rP44) of the HGE agent were examined by reverse transcription-PCR and a capture enzyme-linked immunosorbent assay. The HGE agent induced expression of interleukin-1 (IL-1), tumor necrosis factor alpha (TNF-␣), and IL-6 mRNAs and proteins in PBLs in a dose-dependent manner to levels as high as those resulting from Escherichia coli lipopolysaccharide stimulation. The kinetics of induction of these three cytokines in PBLs by rP44 and by the HGE agent were similar. Proteinase K treatment of the HGE agent or rP44 eliminated the ability to induce these three cytokines. Induction of these cytokine mRNAs was not dependent on superoxide generation. These results suggest that P44 proteins have a major role in inducing the production of proinflammatory cytokines by PBLs. Expression of IL-8, IL-10, gamma interferon, transforming growth factor , and IL-2 mRNAs in response to the HGE agent was not remarkable. Among PBLs, neutrophils and lymphocytes expressed IL-1 mRNA but not TNF-␣ or IL-6 mRNA in response to the HGE agent, whereas monocytes expressed all three of these cytokine mRNAs. These observations suggest that induction of proinflammatory-cytokine gene expression by the major outer membrane protein of the HGE agent in monocytes, which are not the primary host cells of the HGE agent, contributes to HGE pathogenesis and immunomodulation.
Infection and Immunity, 2000
Human granulocytic ehrlichiosis (HGE) is an emerging febrile systemic disease caused by the HGE agent, an obligatory intracellular bacterium of granulocytes. The pathogenicity-and immunity-related mechanisms of HGE are unknown. In this study, several cytokines generated in human peripheral blood leukocytes (PBLs) incubated with the HGE agent or a recombinant 44-kDa major surface protein (rP44) of the HGE agent were examined by reverse transcription-PCR and a capture enzyme-linked immunosorbent assay. The HGE agent induced expression of interleukin-1 (IL-1), tumor necrosis factor alpha (TNF-␣), and IL-6 mRNAs and proteins in PBLs in a dose-dependent manner to levels as high as those resulting from Escherichia coli lipopolysaccharide stimulation. The kinetics of induction of these three cytokines in PBLs by rP44 and by the HGE agent were similar. Proteinase K treatment of the HGE agent or rP44 eliminated the ability to induce these three cytokines. Induction of these cytokine mRNAs was not dependent on superoxide generation. These results suggest that P44 proteins have a major role in inducing the production of proinflammatory cytokines by PBLs. Expression of IL-8, IL-10, gamma interferon, transforming growth factor , and IL-2 mRNAs in response to the HGE agent was not remarkable. Among PBLs, neutrophils and lymphocytes expressed IL-1 mRNA but not TNF-␣ or IL-6 mRNA in response to the HGE agent, whereas monocytes expressed all three of these cytokine mRNAs. These observations suggest that induction of proinflammatory-cytokine gene expression by the major outer membrane protein of the HGE agent in monocytes, which are not the primary host cells of the HGE agent, contributes to HGE pathogenesis and immunomodulation.
Human Granulocytic Ehrlichiosis Agent Inhibits Superoxide Anion Generation by Human Neutrophils
Infection and Immunity, 2000
The human granulocytic ehrlichiosis (HGE) agent, which replicates in neutrophils, was found not to induce superoxide anion (O2 −) generation or extracellular release by human peripheral blood neutrophils, as measured by a luminol-dependent chemiluminescence assay or a cytochrome c reduction assay, respectively. Furthermore, the HGE agent completely prevented O2 − release by neutrophils upon stimulation with phorbol myristate acetate (PMA), formylmethionyl-leucyl-phenylalanine, or Escherichia coli. The inhibition was HGE agent dose dependent, required ehrlichial contact with the host cells, and was reversible upon removal of the extracellular HGE agent bound to the host cells prior to PMA stimulation. Structural integrity of or new protein synthesis by the HGE agent was not required for the inhibition; carbohydrate but not surface protein of the HGE agent was required. The HGE agent did not prevent O2 − generation in human peripheral blood monocytes derived from the same individual. ...
The Journal of Infectious Diseases, 2000
Human granulocytic ehrlichiosis (HGE) results in fever, pancytopenia, and mild liver injury. We used a mouse model to examine immunity in the pathogenesis of HGE. HGE agent-infected C3H/HeJ mice were necropsied over 21 days. Histologic, immunohistologic, and serologic analyses, blood culture, tissue and blood polymerase chain reaction (PCR), cell counts, serum chemistries, and plasma cytokine ELISAs were performed. No clinical signs were detected. Ehrlichiae were identified in neutrophils in hematopoietic tissues maximally on day 7. Interleukin (IL)-10 levels were high throughout, whereas interferon (IFN)-g levels peaked on days 7 and 10 and dropped thereafter. Hepatic lymphohistiocytic aggregates with apoptoses were maximal at day 14. HGE-agent infection of mice induces pathologic changes similar to those in infected humans, despite differences in cytokine profile. The IFN-g peak prior to maximal pathologic change, when ehrlichiae are absent in tissues, suggests a role for host immunity in the pathogenesis of HGE. Human granulocytic ehrlichiosis (HGE) often presents with fever, headache, myalgias, leukopenia, thrombocytopenia, anemia, and elevations in levels of hepatic transaminases [1-4]. In severe cases, gastroenteritis with nausea and vomiting, cough, confusion or peripheral neuropathy, possible adult respiratory distress syndrome, and opportunistic infections have been documented [3, 5-7]. The spectrum of clinical illness is disproportionate to the number and anatomic distribution of organisms detected in blood or tissues in humans and in animal models [8, 9]. This suggests that another mechanism may be responsible in part for pathologic and clinical manifestations of HGE. A major pitfall in the study of HGE has been the lack of a suitable small animal model that reproduces aspects of human infection. Although horses develop reproducible and easily recognized signs similar to those in humans [9], multiple attempts to elicit clinical signs in murine models of HGE have failed,
Involvement of Tumor Necrosis Factor in Human Granulocyte-Mediated Killing of WEHI 164 Cells
International Archives of Allergy and Immunology, 1989
Human polymorphonuclear leukocytes (PMNLs) kill WEHI 164 clone 13 cells in an 18-hour 5lCr release assay. Antibody to human tumor necrosis factor (TNF) blocks the lysis of targets mediated by human granulocytes. PMNLs triggered by sensitive targets not only displayed cytotoxic activity, but also released a soluble factor capable of selectively lysing WEHI 164 cells. The killing of these cells by supernatants of trig gered granulocytes was totally inhibited by anti-TNF antibody. These experiments suggest that the killing of WEHI 164 sarcoma cells by human PMNLs involves TNF or TNF-like molecules.
Journal of Inflammation Research, 2019
We postulate that the extensive cell and tissue damage inflicted by many infectious, inflammatory and post-inflammatory episodes is an enled result of a synergism among the invading microbial agents, host neutrophils and dead and dying cells in the nidus. Microbial toxins and other metabolites along with the plethora of pro-inflammatory agents released from activated neutrophils massively recruited to the infectious sites and high levels of cationic histones, other cationic peptides, proteinases and Th1 cytokines released from activated polymorphonuclear neutrophils (PMNs) and from necrotized tissues may act in concert (synergism) to bring about cell killing and tissue destruction. Multiple, diverse interactions among the many potential pro-inflammatory moieties have been described in these complex lesions. Such infections are often seen in the skin and aerodigestive tract where the tissue is exposed to the environment, but can occur in any tissue. Commonly, the tissue-destructive infections are caused by group A streptococci, pneumococci, Staphylococcus aureus, meningococci, Escherichia coli and Shigella, although many other microbial species are seen on occasion. All these microbial agents are characterized by their ability to recruit large numbers of PMNs. Given the complex nature of the disease process, it is proposed that, to treat these multifactorial disorders, a "cocktail" of anti-inflammatory agents combined with non-bacteriolytic antibiotics and measures to counteract the critical toxic role of cationic moieties might prove more effective than a strategy based on attacking the bacteria alone.
Journal of Inflammation Research
We postulate that the extensive cell and tissue damage inflicted by many infectious, inflammatory and post-inflammatory episodes is an enled result of a synergism among the invading microbial agents, host neutrophils and dead and dying cells in the nidus. Microbial toxins and other metabolites along with the plethora of pro-inflammatory agents released from activated neutrophils massively recruited to the infectious sites and high levels of cationic histones, other cationic peptides, proteinases and Th1 cytokines released from activated polymorphonuclear neutrophils (PMNs) and from necrotized tissues may act in concert (synergism) to bring about cell killing and tissue destruction. Multiple, diverse interactions among the many potential pro-inflammatory moieties have been described in these complex lesions. Such infections are often seen in the skin and aerodigestive tract where the tissue is exposed to the environment, but can occur in any tissue. Commonly, the tissue-destructive infections are caused by group A streptococci, pneumococci, Staphylococcus aureus, meningococci, Escherichia coli and Shigella, although many other microbial species are seen on occasion. All these microbial agents are characterized by their ability to recruit large numbers of PMNs. Given the complex nature of the disease process, it is proposed that, to treat these multifactorial disorders, a "cocktail" of anti-inflammatory agents combined with non-bacteriolytic antibiotics and measures to counteract the critical toxic role of cationic moieties might prove more effective than a strategy based on attacking the bacteria alone.