Mediation of entry of human immunodeficiency virus-1 into alveolar macrophages by CD4 without facilitation by surfactant-associated protein-A.

The mechanism of uptake of human immunodeficiency virus-1 (HIV-1) into alveolar macrophages (AM), freshly isolated blood monocytes (MN), and cultured MN (CM) was investigated focusing on the role of CD4 and of surfactant-associated protein A (SP-A). By radioimmunoassay which obviated the problems of auto- and nonspecific fluorescence of more differentiated macrophages, each of the macrophage populations studied expressed CD4. Semiquantitative polymerase chain reaction was performed to assess uptake of HIV-1(JR-FL) into cells. OKT4a (directed against CD4) blocked uptake of HIV-1 into CM, AM, and MN by 67 to 100%. OKT4 (directed against another epitope of CD4) had a smaller and less consistent effect (0-90%), and control antibodies showed minimal effects and only at supersaturating concentrations. SP-A had no effect on uptake of HIV-1 into AM. SP-A also had no consistent effect on production of HIV-1(JR-FL) by AM infected in vitro (p24 antigen ELISA). Thus CD4 is the major receptor for HIV-1 in mononuclear phagocytes, including AM, and SP-A does not modulate entry.

Mycobacterium tuberculosis (MTB)-stimulated production of nitric oxide by human alveolar macrophages and relationship of nitric oxide production to growth inhibition of MTB.

SETTING: Although nitric oxide (NO) is a major proximate mediator of microbicidal activity in murine macrophages against intracellular pathogens including mycobacteria, its production by and effector role in human macrophages is not clear. OBJECTIVE: To determine the capacity of Mycobacterium tuberculosis (MTB) to stimulate NO in human monocytes (MN) and alveolar macrophages (AM) and to assess the relationship between NO production and intracellular growth of MTB. DESIGN: NO production (measured as nitrite) by MTB (H37Ra)-infected macrophages and intracellular growth of MTB were measured in cells from 17 healthy subjects. RESULTS: MTB (5:1, MTB:cells) stimulated little to no NO by MN, but induced NO in AM at days 4 and 7 after infection. There was, however, variability in the response by AM to MTB: among seven subjects MTB-induced NO was low (4 +/- 2 microM, mean +/- SE); six subjects were moderate (56 +/- 11); four subjects were high (502 +/- 167). NO synthase inhibitors inhibited the production of NO by AM but did not significantly affect the intracellular growth of MTB, although a trend towards increased intracellular growth was seen on day 4 of culture. Intracellular growth of MTB in AM from low NO producers was significantly higher than that in AM from moderate NO producers, P < or = 0.05. Inducible NO synthase (iNOS) mRNA by RT-PCR was constitutively expressed by both MN and AM, but was further stimulated by MTB in AM > MN; MTB-induced iNOS protein was present in both MN and AM by Western blot analysis. CONCLUSION: Thus, MTB-infected human AM are capable of producing NO and NO production correlates with intracellular growth inhibition of MTB in AM suggesting that NO may serve either directly or indirectly as a mycobactericidal mediator in human tissue macrophages.

Enhanced production of human immunodeficiency virus type 1 by in vitro-infected alveolar macrophages from otherwise healthy cigarette smokers.

Since cellular activation is required for replication of human immunodeficiency virus type 1 (HIV-1), the capacity of alveolar macrophages (AM) from smokers, which are relatively activated, and nonsmokers to support the production of HIV-1JR-FL was examined. Peak HIV-1 p24 antigen level in culture supernatants of infected AM from 13 smokers was significantly higher than that of 13 nonsmokers: 31,394 +/- 8295 versus 7037 +/- 2550 pg/mL (mean +/- SE; P < .002). This difference could not be explained on the basis of viral entry, extent of reverse transcription, or release of monokines, including tumor necrosis factor-alpha, interleukin-1 beta or -6, and granulocyte-macrophage colony-stimulating factor. HIV-1 production by blood monocytes from smokers and nonsmokers infected in vitro was negligible. Thus, cigarette smoking selectively increases the susceptibility of AM to productive infection with HIV-1. This finding provides a biologic plausibility to observations that smoking may enhance the progression of AIDS.

Disparate cytochemical characteristics and production of cytokines and prostaglandin E2 by human mononuclear phagocytes from the blood, lung, and peritoneal cavity.

The cytochemical characteristics and the production of tumor necrosis factor-alpha (TNF-alpha), interleukin-1 beta (IL-1 beta), and prostaglandin E2 (PGE2) by peritoneal macrophages were compared with those of blood monocytes and alveolar macrophages. The comparative percentages of mononuclear phagocytes positive for peroxidase were as follows: blood monocytes > peritoneal macrophages > alveolar macrophages. The comparative percentages of cells positive for nonspecific esterase were as follows: alveolar macrophages > peritoneal macrophages = blood monocytes. The intensity of staining for nonspecific esterase was highest in alveolar macrophages and lowest in blood monocytes. Constitutive release of TNF, IL-1 beta, and PGE2 was minimal by each cell type. Lipopolysaccharide-stimulated TNF production by alveolar macrophages was approximately five times greater than that of monocytes and 10 times greater than that of peritoneal macrophages. By contrast, lipopolysaccharide-stimulated blood monocytes produced significantly more IL-1 beta than did peritoneal or alveolar macrophages. Lipopolysaccharide-stimulated production of PGE2 by peritoneal macrophages was significantly less than that of alveolar macrophages or blood monocytes. Thus peritoneal macrophages release relatively low levels of IL-1 beta, TNF, and PGE2 in response to lipopolysaccharide. Peritoneal and alveolar macrophages differ with respect to both cytochemical characteristics and lipopolysaccharide-stimulated production of TNF and PGE2 but are similar in their limited capacity to produce IL-1 beta.

Relationship between load of virus in alveolar macrophages from human immunodeficiency virus type 1-infected persons, production of cytokines, and clinical status.

The level of human immunodeficiency virus type 1 (HIV-1) in lymphocytes and mononuclear phagocytes (MP) from the blood and pulmonary alveoli from 14 HIV-1-infected subjects during early (asymptomatic) and late (AIDS) stages of disease and the relationship between virus burden in MP and cytokine expression were assessed. Among asymptomatic subjects, HIV-1 was undetectable or low in both blood monocytes and alveolar macrophages (AM). Among subjects with AIDS, there was a significant increase of HIV-1 in AM but not monocytes. The level of HIV-1 in blood lymphocytes was higher than in either monocytes or AM. AM (but not monocytes) expressed increased levels of lipopolysaccharide-stimulated cytokine mRNA (tumor necrosis factor-alpha, interleukin-1 beta, interleukin-6) during both early and late stages of HIV-1 infection regardless of virus load. AM thus may serve as a reservoir for virus in late stages of disease yet contribute to the immunopathogenesis of lung disease in both early and late stages through increased cytokine expression.

Characterization of suppressor function of human alveolar macrophages for T lymphocyte responses to phytohemagglutinin: cellular selectivity, reversibility, and early events in T cell activation.

The predominant immunoregulatory activity of alveolar macrophages (AM) on T lymphocytes is to suppress their responses to antigenic and mitogenic stimuli. The suppressive activity of human AM for T cell responses to phytohemagglutinin (PHA) was further characterized. At ratios of AM to T lymphocytes of 0.4:1 to 1.6:1, AM inhibited the blastogenic response (3H-thymidine uptake into DNA) to PHA by 26 to 87%, respectively. Blood monocytes precultured in vitro for 5 to 7 days inhibited responses to PHA similarly. Freshly isolated blood monocytes, peritoneal macrophages, and A-549 epithelial and CCD-18Lu fibroblast cell lines failed to inhibit T lymphocyte responses. AM were capable of suppressing PHA-induced blastogenesis of purified CD4 cells without the addition of other cells. Cell contact was required for suppression of CD4 cells, as demonstrated using dual chambers. T cells precultured with AM with or without PHA retained the ability to respond to PHA compared with control T cells not precultured with AM. Kinetic experiments showed that AM needed to be added at the initiation of a 3-day culture period for suppression to occur. Analysis of the T cell DNA cycle revealed that AM decreased the percentage of cells entering the synthesis phase of DNA production. Flow cytometry also was used to assess the effect of AM on early markers of T cell activation. AM inhibited the percentage of T cells expressing the interleukin-2 receptor 46 to 83% and the transferrin receptor 58 to 78% at 24 to 48 h after stimulation with PHA. There was no effect of AM on expression of HLA-DR.(ABSTRACT TRUNCATED AT 250 WORDS)

Accessory function of human mononuclear phagocytes for lymphocyte responses to the superantigen staphylococcal enterotoxin B.

The role of the cytokines IL-1 alpha, IL-1 beta, and IL-6 and the cell adhesion molecules ICAM-1, LFA-1 (alpha and beta), and Mac-1 as accessory molecules for stimulation of T cells by the superantigen staphylococcal enterotoxin B (SEB) was examined. Both blood monocytes and alveolar macrophages were used as accessory cells because these cells differ in patterns of cytokine expression and thus potentially in accessory cell function for superantigens. The blastogenic response of highly purified T cells to SEB was reconstituted with either monocytes or alveolar macrophages. IL-1 secretion was increased comparably in monocytes and alveolar macrophages by SEB, but IL-6 was not stimulated by SEB. IL-1 alpha plus IL-1 beta reconstituted the response of T cells to SEB but required the addition of accessory cells. The cell adhesion molecules ICAM-1 and LFA-1 but not Mac-1 also functioned as accessory molecules for SEB-induced cluster formation and lymphocyte blastogenesis. Thus, not only must this superantigen bind to Class II MHC on accessory cells as is well known, but also SEB requires at least certain cytokines (IL-1 alpha and IL-1 beta) produced by accessory cells and cell adhesion molecules (ICAM-1 and LFA-1) for activation of T lymphocytes.

Serum keratan sulfate. Quantitative and qualitative comparisons in inflammatory versus noninflammatory arthritides.

The concentration of keratan sulfate (KS) epitope was measured in the serum of patients with osteoarthritis (OA) or rheumatoid arthritis (RA) by enzyme-linked immunosorbent assay and compared with that in the serum of patients with primary fibromyalgia syndrome (PFS) and of controls who had no joint disease. By Student's tau-test, the mean serum KS concentrations in OA and RA patients measured with monoclonal antibodies (MAb) 5-D-4 and 2-D-3 were significantly increased over those in the PFS and normal groups; similar findings were observed using a nonparametric test, except that levels in RA patients showed no difference from those in PFS patients and normal subjects. There was no significant correlation between joint scores or disease duration and KS levels in OA or RA patients. Gel filtration of sera revealed mainly large, polydisperse KS-bearing fragments which eluted in a broad profile. KS purified from sera by immunoaffinity chromatography consisted mainly of high-density proteoglycans. Electrophoresis of pooled high-density KS fractions in polyacrylamide-agarose gels followed by Western blotting with MAb 5-D-4 showed diffuse bands with relative mobilities corresponding to large proteoglycans. These findings are consistent with attachment of KS to protein core fragments of various sizes; KS in patient sera is comparable in size with that in normal sera. Elevations of serum KS levels occur in the presence of cartilage degradation, but do not quantitatively define the extent or duration of articular involvement.

Restricted expression of anti-type II collagen antibody isotypes in mice suppressed for collagen-induced arthritis.

The present study details the time course and isotype distribution of the immune response to type II collagen in collagen-induced arthritic mice and mice suppressed for collagen-induced arthritis. The serum of arthritic mice was observed to contain significantly higher (p less than 0.005) concentrations of antibodies to type II collagen than that of mice suppressed for arthritis at all times tested. For the arthritic mice anti-type II collagen antibodies ranged from 0.2 +/- 0.2 (SD) to 6.1 +/- 0.7 mg/ml (g/l). Serum values for mice suppressed for arthritis ranged from 0.05 +/- 0.04 to 0.6 +/- 0.04 mg/ml. Analysis of the isotypes of these responses showed an expression of anticollagen molecules restricted to the IgG1 subclass in mice suppressed for collagen arthritis throughout the time course (p less than 0.01). The data indicate that mice suppressed for collagen-induced arthritis can mount a primary and secondary immune response to the arthrogenic stimuli. This response, however, is mainly restricted to the IgG1 subclass of antibodies. This restricted subclass expression of anticollagen antibodies may represent a mechanism of suppression of arthritis in the murine model of collagen-induced arthritis.