C-reactive protein (CRP) is a serum protein that shows rapid increases of as much as 1000-fold in concentration in response to infection, traumatic injury, or inflammation. CRP reacts with the phosphocholine moiety of pneumococcal cell wall C-polysaccharide, and this reaction can lead to complement activation in vitro and protection against pneumococcal infection in vivo. We have previously studied the chemiluminescence response of human neutrophils to Streptococcus pneumoniae as a measure of in vitro opsonophagocytosis by CRP and complement. CRP in the presence of complement was an effective opsonin for S. pneumoniae serotype 27 (Pn27), but not for serotypes 3 or 6. Because Pn27 differs from most serotypes of S. pneumoniae in containing phosphocholine in its capsular polysaccharide, we have determined the sites of CRP and C3 fixation to Pn27 and S. pneumoniae serotype 4 (Pn4), and related these to the ability of CRP and complement to opsonize these serotypes in vitro. By using a chemiluminescence (CL) assay to measure opsonophagocytosis, CRP was shown to enhance the response of human neutrophils and monocytes to Pn27 in the presence of normal human serum. The CL response of neutrophils and monocytes to Pn4 was not affected by the addition of CRP to serum. The addition of anti-capsular antibody to Pn4 and Pn27 enhanced the CL responses of both neutrophils and monocytes to both bacteria. The localization of bound CRP and C3 on Pn4 and Pn27 was determined by immunoelectron microscopy. CRP bound to Pn4 only in the cell wall region and C3 was located in this area whether or not CRP was present. Anti-capsular antibody deposited C3 in the capsule of Pn4. In contrast, Pn27 bound CRP throughout the capsule and cell wall areas. C3 was deposited in the cell wall region of Pn27 by serum alone and in the cell wall region and capsule when CRP or anti-capsular antibody was present. Because C3 fixation to the capsule was consistently associated with enhanced responses by phagocytic cells, it appears that the site of CRP binding and subsequent complement activation may be critical in the opsonophagocytosis of S. pneumoniae. These findings extend the correlation between capsular C3 and opsonization to a nonimmune system. By using CRP and different pneumococcal serotypes we have shown that the same molecules that are effective in the stimulation of phagocytic cells when bound to the capsule are not effective when bound to the cell wall.
The effects of various cytokines were examined in an in vitro model of human immunodeficiency virus type 1 (HIV-1) infection of human peripheral blood monocyte-derived macrophages (MDM). Monocytes were obtained from blood of normal donors by Ficoll/hypaque gradient centrifugation and adherence. These cells were allowed to mature in the presence of varying concentrations of cytokines. After five days in culture, cells were harvested, counted, and inoculated with S5G7, an HTLV-IIIB subclone. The cells were replated in the presence of the same concentrations of cytokines. Culture supernatants were sampled over 28 days for p24 antigen (Ag) as measured by Ag capture assay. In repeat experiments, the following observations were made: 1. MDM from some donors could be infected only in the presence of tumor necrosis factor-alpha (TNF-α), granulocyte/macrophage colony-stimulating factor (GM-CSF) or interleukin 4 (IL-4); 2. The effect of GM-CSF was variable; TNFα also enhanced HIV replication above controls; 3. IL-4 was the most potent enhancer of HIV-1 replication in MDM of the cytokines tested, inducing p24 Ag levels 75-230 times those seen in control cultures run simultaneously. This effect was dose dependent. Ag production was not observed until Day 14 postinfection in most experiments. Multinucleated giant cell formation was observed only in the presence of IL-4.
C-reactive protein (CRP) has several properties that suggest that it may function as a bacterial opsonin. CRP shows binding reactivity with pneumococcal C-polysaccharide, the cell wall carbohydrate of Streptococcus pneumoniae. In this study we have demonstrated protection of mice against serotypes 3 and 4 of S. pneumoniae infection by a single prior injection of CRP. This effect was seen both in mice that lacked antibody to phosphocholine and in normal mice. Thus the opsonic properties of CRP previously described may be related to protection against pneumococcal infection.
Components of current vaccines for Hansen's disease include Mycobacterium bovis Bacillus Calmette-Guérin (BCG) and killed Mycobacterium leprae. BCG infections in humans are rare and most often occur in immune-compromised individuals. M. leprae on the other hand, although not causing clinical disease in most exposed individuals, is capable of infecting and replicating within mononuclear phagocytes. Lymphocytes from patients with the lepromatous form of Hansen's disease exhibit defective lymphokine production when challenged in vitro with M. leprae. This may result in inefficient mononuclear phagocyte activation for oxidative killing. To study the ability of normal phagocytes to ingest and respond oxidatively to BCG and M. leprae, we measured phagocytic cell O2- release and fluorescent oxidative product formation and visually confirmed the ingestion of the organisms. BCG stimulated a vigorous O2- generation in neutrophils and monocytes and flow cytometric oxidative product generation by neutrophils occurred in the majority of cells. M. leprae, stimulated a weak but significant O2- release requiring a high concentration of organisms and long exposure. By flow cytometric analysis, most neutrophils were able to respond to both organisms with the generation of fluorescent oxidative products. Neutrophil oxidative responses to M. leprae were substantially less than responses seen from neutrophils exposed to BCG. By microscopic examination of neutrophils phagocytizing FITC-labeled bacteria, it was shown that both M. leprae and BCG were slowly ingested but that more BCG appeared to be associated with the cell membrane of more of the cells. When phagocytic cells were incubated with BCG and M. leprae for 30 min and subsequently examined by electron microscopy, few organisms were seen in either neutrophils or monocytes. This suggests that BCG are easily recognized and slowly ingested by normal phagocytic cells, the majority of which respond with a strong oxidative burst. M. leprae appeared to only weakly stimulate phagocyte oxidative responses and were also slowly phagocytized.
Journal Article Capacity of Human Neutrophils to Kill Mycobacterium tuberculosis Get access Arthur E. Brown, Arthur E. Brown Search for other works by this author on: Oxford Academic PubMed Google Scholar Timothy J. Holzer, Timothy J. Holzer Search for other works by this author on: Oxford Academic PubMed Google Scholar Burton R. Andersen Burton R. Andersen Please address requests for reprints to Dr. Burton R. Andersen, West Side VA Medical Center (m/p 151), 820 South Damen Avenue, Chicago, Illinois 60612. Search for other works by this author on: Oxford Academic PubMed Google Scholar The Journal of Infectious Diseases, Volume 156, Issue 6, December 1987, Pages 985–989, https://doi.org/10.1093/infdis/156.6.985 Published: 01 December 1987 Article history Received: 10 July 1986 Revision received: 01 June 1987 Published: 01 December 1987
Peripheral blood monocytes were pretreated with phenolic glycolipid-I (PGL-I), dimycocerosyl phthiocerol (DIM), or mycoside A, then cultured in the presence or absence of interferon-gamma (IFN-gamma). Their oxidative responses to Mycobacterium leprae, phorbol myristate acetate (PMA), and opsonized zymosan were evaluated. In response to M. leprae, monocytes pretreated with PGL-I released less O2- than nonlipid-treated control cells. The IFN-gamma augmentation of oxidative responses was suppressed only when in PGL-I-pretreated monocytes and only when the stimulus was M. leprae. This suggests that PGL-I, by affecting the IFN-gamma enhancement of phagocytic cell oxidative responses, aids further the intracellular survival of M. leprae.
Experiments were performed to determine the effects of Mycobacterium tuberculosis-derived sulfolipid I on phagocytic cells. Sulfolipid I was taken up in significant amounts by human neutrophils and in lesser amounts by monocytes and lymphocytes. Superoxide (O2-) production by neutrophils was significantly increased by sulfolipid I, but the rate of production was slower than that reported previously for other stimuli. The optimal concentration of sulfolipid I for stimulation of O2- production was 27 micrograms/ml, while higher concentrations produced less. At substimulatory levels sulfolipid I caused enhancement of O2- release from neutrophils when it was subsequently stimulated by other agents. Nonadherent monocytes from most normal donors failed to produce O2- when treated with sulfolipid I; however, adherent monocytes pretreated with gamma interferon did produce O2- with sulfolipid I stimulation. Priming for an enhanced oxidative response of activated monocytes was also observed. These sulfolipid I-induced changes in phagocytic cell function may be important in altering the ability of phagocytes to respond effectively to M. tuberculosis and may also cause exaggerated inflammatory responses.
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