Human natural killer cells enhance a mixed leukocyte reaction.

Natural killer cells (NK) have been reported to down-regulate the initiation of T cell responses in animal models. In the current study, highly purified CD16+ human NK cells were obtained by cell sorting and their effect on the stimulation of allogeneic T cells (MLR) determined. NK cells did not directly stimulate T cell proliferation. However, when added to a population of loosely adherent mononuclear cells (LAM), NK enhanced the ability of these accessory cells to stimulate T proliferation. This effect was not reproduced by the addition of sorted CD5 + T cells, sorted CD16- cells, or control lymphocytes to the MLR. The effect of NK on the MLR was not restricted by class II antigens and was similar to the effect of adding IL-1 to MLR cultures. These results demonstrate that human NK cells are capable of enhancing a T cell response.

Separation of potent and poorly functional human lung accessory cells based on autofluorescence.

Human alveolar macrophages obtained by bronchoalveolar lavage are usually poor accessory cells in in vitro lymphoproliferation assays. However, we recently described a subpopulation of pulmonary mononuclear cells, obtained from minced and enzyme-digested lung, which were potent stimulators of allogeneic T-lymphocyte proliferation. These cells were enriched in loosely adherent mononuclear cell (LAM) fractions, but further study of these accessory cells was hampered by the heterogeneous nature of LAM. It was observed that in the majority of lung tissue sections, most alveolar macrophages were autofluorescent, whereas most interstitial HLA-DR positive cells were not. Therefore autofluorescence was utilized to fractionate LAM in an attempt to remove alveolar macrophages and selectively purify interstitial accessory cells. LAM were separated by flow cytometry using forward and side scatter to exclude lymphocytes, and red autofluorescence to obtain brightly autofluorescent (A pos) and relatively nonautofluorescent (A neg) mononuclear cells. Although both populations contained over 80% HLA-DR positive cells, A pos cells were poor accessory cells, whereas A neg cells were extremely potent stimulators of a mixed leukocyte reaction at all stimulator ratios tested. When A pos cells were added to A neg cells, T-cell proliferation was markedly suppressed in the majority of experiments. Morphologically, A pos cells appeared similar to classical alveolar macrophages with 95% of the cells being large and intensely nonspecific esterase positive. In contrast, the majority of A neg were smaller, B-cell antigen-negative, nonspecific esterase negative, and had a distinctive morphology on Wright-stained smears. We conclude that fractionation of LAM based on autofluorescence is a powerful tool to isolate and characterize lung mononuclear cells that act either as stimulators or as suppressors of immune responses in the lung.

Mononuclear cells from human lung parenchyma support antigen-induced T lymphocyte proliferation.

We have previously demonstrated that there is a subpopulation of loosely adherent pulmonary mononuclear cells that can be isolated from minced and enzyme-digested lung tissue with a potent capacity to stimulate allogeneic T lymphocyte proliferation. We now demonstrate that these cells are also capable of stimulating an autologous mixed leukocyte reaction (AMLR) and presenting antigen to autologous T lymphocytes. These loosely adherent mononuclear cells (LAM) were more effective than either alveolar macrophages or monocytes as antigen-presenting cells. Depletion of phagocytic or Fc receptor-positive cells from the LAM population enhanced the stimulation of an reaction AMLR while preserving antigen-induced T lymphocyte proliferation. These results indicate that there are nonphagocytic, Fc receptor-negative accessory cells in human lung parenchyma capable of activating resting T cells in an AMLR and supporting antigen-specific T lymphocyte proliferation. The identity of these cells is uncertain, but the data strongly suggest that the cell is not a classical monocyte-derived macrophage. These antigen-presenting cells may be critical in the initiation of immune responses within the lung.

Autoantigen Ro52 directly interacts with human IgG heavy chain in vivo in mammalian cells.

Previously, when we used in vivo yeast two-hybrid and in vitro protein-protein interaction analyses, we demonstrated a direct interaction between autoantigen Ro52 and the human IgG heavy chain. This interaction occurred in the absence of antibody-antigen specific interaction. Here, by employing a novel strategy, we further demonstrated that Ro52 co-localized with IgG in transfected mammalian cells. The co-localization was specific to IgG1 but not IgG3. Co-immunoprecipitating IgG with Ro52 from transfected cell lysates suggested that protein complex containing Ro52 and IgG contributed to the in vivo co-localization. In addition, IgG from normal human serum was shown to bind to the surface of apoptotic keratinocytes and the binding could be competitively blocked by 50-fold excesses of IgG1, not IgG3. With a direct binding study, we also demonstrated that IgG1 could bind to the surface of apoptotic cells while IgG3 bound barely. This binding was not competed by Fcgamma fragments indicating a non-Fcgamma receptor mediated interaction. Finally, in a competition analysis the addition of GST-RFP could reduce the IgG binding to the cell surface. Thus, we suggested that the binding of IgG to the apoptotic keratinocytes might be mediated through the interactions with the surface exposed Ro52. The potential role of forming this protein complex on the apoptotic cells will be discussed.

Treatment of pneumonia in patients at risk of infection with gram-negative bacilli.

Patients with a history of chronic debilitating disease due to a variety of causes are known to be at risk for infection with coliform gram-negative bacilli when they present with community-acquired pneumonia. Empiric treatment with broad-spectrum parenteral antibiotics is often begun in such patients pending the result of blood and other cultures. The optimal duration of broad-spectrum empiric therapy in such patients when cultures fail to reveal a specific pathogen is unknown. Review was made of the charts of 131 patients with community-acquired pneumonia admitted to the hospital and treated with broad-spectrum parenteral antibiotics in whom a specific pathogen was never isolated. Overall, 89 percent of these patients were cured without complication or relapse. Patients receiving broad-spectrum therapy for four days or less due to rapid clinical improvement had a successful outcome in 95 percent of cases. It is concluded that broad-spectrum parenteral antibiotic therapy can safely be abbreviated provided response to therapy is prompt.

Increased bronchoalveolar IgG2/IgG1 ratio is a marker for human lung allograft rejection.

BACKGROUND: Lung allograft rejection (AR) is thought to involve T-helper-1 (Th-1) lymphocytes mediating both cellular immunity and alloantibody production. Th-1 lymphocytes produce gamma interferon (gamma IFN) and induce IgG2 production, suggesting that increased IgG2 production might occur during AR. The purpose of this study was to determine if locally altered bronchoalveolar IgG2/IgG1 ratios might correlate with AR. METHODS: Eighteen recipients of lung allografts underwent a total of 25 bronchoscopies for surveillance or at times of suspected infection or AR. Bronchoalveolar lavage (BAL), serum collection, and transbronchial biopsy (TB) were performed on all patients. gamma IFN, IgG1, IgG2 levels, and the ratio of IgG2/IgG1 were determined in serum and BAL and matched with TB histology. Five nonsmoking normal volunteers undergoing bronchoscopy, BAL, and serum collection served as controls. RESULTS: IgG2 was upregulated in allograft BAL during AR as determined by the ratio IgG2/IgG1 (2.91 +/- 0.79 SEM vs 0.62 +/- SEM, p < 0.019, IgG2/IgG1, AR BAL vs non-AR BAL, respectively). An IgG2/IgG1 > or = 1 in allograft BAL (95% confidence intervals 1.26 to 4.56) was 80% specific and 91% sensitive for the diagnosis of AR with a positive predictive value of 92%. A BAL IgG2/IgG1 < 1 (95% confidence interval 0.27 to 0.97) had a negative predictive value of 77%. After therapy in two patients the elevated IgG2/IgG1 ratio reversed to normal (ie, < 1) with histologic resolution of AR. CONCLUSIONS: Human lung AR is associated with a locally increased IgG2/IgG1 ratio suggesting locally upregulated Th-1 lymphocyte activity during lung AR.

Southwestern internal medicine conference: sarcoidosis: immunology and clinical management.

Sarcoidosis is a chronic inflammatory disease characterized by the presence of noncaseating granulomas in the lung and other organs. Current evidence suggests that this response is driven by a foreign antigen whose identity remains unclear. In this article, the nature of the cellular immune response is explored and the value of local markers of inflammation in predicting clinical course is examined.

Pulmonary emphysema: current concepts of pathogenesis.

Pulmonary emphysema is a major public health problem and is primarily a disease of smokers. The pathogenesis of emphysema in smokers is likely to be multifactorial and may involve protease-antiprotease imbalance, abnormal host response to injury, the inactivation of antiproteases by oxidants, and direct damage of lung tissue by pulmonary phagocytes. The data regarding current concepts of pathogenesis of emphysema in smokers are reviewed in this article.

Idiopathic pulmonary fibrosis: cellular and molecular pathogenesis.

Diffuse involvement of the pulmonary interstitium with abnormal fibrous tissue is a process that occurs in many settings. There are many possible etiologies for pulmonary fibrosis, but in the majority of individuals, a clear cause cannot be determined and a diagnosis of idiopathic pulmonary fibrosis (IPF) results. Despite limited knowledge concerning the etiology, recent advances in biomedical technology offer great promise for increasing our understanding of IPF. This review will focus on current concepts of the pathogenesis and therapy of IPF.

Syndromes of severe asthma.

Asthma responds to conventional therapy in the majority of patients. However, attention has recently focused on the 1 to 15% of asthmatics who are thought to manifest severe asthma, which responds poorly to commonly used regimens. In this review, current knowledge about the pathogenesis of severe asthma is reviewed and several distinct clinical syndromes of severe asthma are discussed.

Tuberculosis--immunopathogenesis and therapy.

Infection with Mycobacterium tuberculosis (TB) has returned to the forefront of public and medical concern because of the recent sharp increase in the number of cases. Major strides have been made in understanding the pathogenesis of TB, and some of these basic advances are being applied clinically. This review focuses on current concepts of the host response to TB, the changing epidemiology of TB, and optimal treatment strategies.

Pulmonary disease in AIDS patients.

Pulmonary disease remains a major complication of the human immunodeficiency virus (HIV). Over the past decade several changes in the pattern of disease have occurred. Pneumocystis carinii pneumonia (PCP) remains the most common opportunistic pathogen in AIDS patients, though its incidence on bronchoscopy has declined and empiric therapy often occurs without a specific diagnosis. Changes in the management of patients with PCP have included different dosages and routes of administration for chemotherapy, improved overall survival, and a recent increase in the number of patients surviving episodes of respiratory failure. In addition, infection with mycobacteria tuberculosis (M.Tb.) has emerged as a major public health problem. The pattern of M.Tb. is distinct from non-immunocompromised patients though response to therapy usually occurs.

Human pulmonary natural killer (NK) cells exhibit limited lymphokine-activated killer (LAK) activity.

The spontaneous activity of natural killer (NK) cells against most solid tumor targets is low but can be increased by incubation with interleukin 2 (IL-2). This phenomenon, termed lymphokine-activated killer (LAK) activity, has been used in recent clinical trials against some pulmonary malignancies. We compared the LAK activity of blood and lung lymphocytes after activation with IL-2. Lung lymphocytes did not develop LAK activity despite demonstrating a significant increase in NK activity against K562 targets after incubation with IL-2. This functional difference correlated with a reduced expression of Leu-19, a marker present on virtually all LAK cells derived from peripheral blood, on lung NK cells. Because pulmonary macrophages (PM) are important regulators of NK function, we next investigated whether PM could be responsible for the functional and phenotypic differences noted. Measuring NK and LAK activity in parallel, we found that the addition of PM to IL-2-activated lymphocytes resulted in a preferential suppression of LAK activity and a loss of Leu-19 expression from IL-2-activated blood lymphocytes as well as a Leu-19+ T cell clone. We conclude that pulmonary NK cells are phenotypically and functionally different from peripheral blood NK cells and that this likely reflects local regulation, perhaps by PM.

Alloantigen-induced immunoglobulin production in human lung: differential effects of accessory cell populations on IgG synthesis.

Local immunoglobulin production has been implicated in the pathogenesis of lung allograft rejection. The role of varying classes of lung accessory cells in stimulating an immunoglobulin (Ig) response in this setting as well as cytokines necessary for Ig synthesis is unknown. The purpose of the current study was to compare the accessory cell capabilities of lung dendritic cells (DC), parenchymal macrophages (PM), and alveolar macrophages (AM) in the generation of a humoral response to alloantigen. Allogeneic AM induced a dose-dependent production of IgG from peripheral blood mononuclear cells. In contrast, allogeneic DC and PM were unable to induce IgG synthesis. The inability of DC to stimulate IgG synthesis was observed despite a potent induction of T-cell proliferation and interferon-gamma (IFN-gamma) production. Additionally, supernatants from DC cultures suppressed AM-induced IgG production, suggesting that a soluble inhibitor of IgG synthesis was produced by DC-stimulated lymphocytes. AM-induced IgG synthesis was predominantly the result of IgG1 and IgG2 production. Experiments with blocking antibodies to either IFN-gamma or interleukin-4 (IL-4) revealed that both IFN-gamma and IL-4 participated in IgG synthesis, while only IFN-gamma was required for IgG2 production. These data demonstrate a discordance between the ability of lung accessory cells to induce T-cell proliferation and IgG synthesis. Furthermore, these findings suggest that local induction of either IL-4 or IFN-gamma is involved in stimulation of an IgG response to lung alloantigen.

Human alveolar macrophages inhibit receptor-mediated increases in intracellular calcium concentration in lymphocytes.

Prior studies have demonstrated that human alveolar macrophages (AM) are suppressive of lymphocyte function, through the mechanism of inhibition is unclear. In the current study, human AM inhibited receptor-mediated increases in intracellular calcium concentration ([Ca2+]i) in T cells, natural killer cells, and B cells. This effect was produced by either live or fixed AM, while peripheral blood monocytes caused a minimal reduction in [Ca2+]i. The inhibitory effect of AM was seen following 1 to 2 h of incubation with lymphocytes, was complete at 16 h, and did not affect ionomycin-mediated [Ca2+]i. Inhibition of [Ca2+]i by AM correlated with suppression of T-lymphocyte proliferation and cytotoxic T-lymphocyte activity in response to alloantigen and Staphylococcus A-induced immunoglobulin production. Our findings suggest that a membrane signal on AM is capable of inhibiting receptor-mediated signal transduction in lymphocytes and that this is likely a major mechanism by which immune responses are downregulated in the alveolus.

Human alveolar macrophages inhibit immunoglobulin production in response to direct B cell mitogen.

B lymphocytes are crucial participants in pulmonary immune defense. However, excess local antibody production is associated with accelerated lung destruction in several types of lung disease. The purpose of the current study was to study the potential role of alveolar macrophages (AM) in the local regulation of immunoglobulin (Ig) production in the lung in response to a direct B cell mitogen, Staphylococcus aureus Cowan strain (SAC). AM, when added to peripheral blood mononuclear cells, caused a dose-dependent inhibition of IgG and IgM, while not affecting IgA production in response to SAC. The mechanism of the AM-induced inhibition included both membrane-bound and soluble signals. The inhibition was not abrogated by indocin and catalase, or reversed by blocking antibodies to transforming growth factor-beta or interferon-gamma. Mononuclear cells isolated from human lung parenchyma displayed a reduced response to SAC compared with blood cells. However, depletion of macrophages from the parenchymal cells was associated with a restoration of IgG production in response to SAC. The results demonstrate that AM inhibit B cell responses to direct B cell mitogen and suggest that the effect of AM is selective for IgM and IgG.

Inhibition of lymphokine-activated killer cells by human pulmonary macrophages: discordance between up-regulation of the beta chain (p75) of the interleukin-2 receptor on CD56+ cells and limited response to interleukin-2.

Previous studies have demonstrated that interaction of interleukin-2 (IL-2) with the beta chain (p75) of the IL-2 receptor on CD56+ cells is necessary for the development of lymphokine-activated killer (LAK) activity and proliferation of CD56+ LAK cells in response to IL-2. Human pulmonary macrophages (PM) are potent inhibitors of LAK cells in vitro, and purified resident human lung lymphocytes show limited LAK activity in response to IL-2, suggesting that IL-2-p75 interactions may be altered locally in vivo. In the current study, human PM or anti-p75 inhibited LAK activity and proliferation of CD56+ cells in response to IL-2. This effect was produced by either live or paraformaldehyde-fixed PM, but not peripheral blood monocytes, suggesting that a membrane signal on PM was responsible for inhibition. Suppression of LAK function and proliferation in response to IL-2 occurred despite a rapid up-regulation of p75 on CD56+ cells after 24 h of incubation with PM. Greater than 70% of CD56+ cells expressed p75 after culture with either live or fixed PM, compared with 10 to 15% at 0 h or after 24 h of incubation in IL-2 alone. p75 dim and p75 bright cells increased equally, suggesting that p75 was being up-regulated on previously p75- cells rather than an overexpansion of one subset of p75+ cells. The increase in p75 expression in the presence of PM paralleled with an increase in IL-2 binding to these lymphocytes. These results suggest that PM inhibit the activation of LAK cells at a point distal to IL-2-p75 binding.

Pulmonary natural killer cell activity is reduced in patients with bronchogenic carcinoma.

Natural killer (NK) cells are a subpopulation of lymphocytes capable of killing a variety of tumor targets. They can limit pulmonary metastases in vivo and thus might be important effectors of tumor defense in human lung. Lymphocytes were purified from whole lung specimens obtained from patients with lung cancer undergoing curative resection, and their NK activity was compared with that of lymphocytes purified from normal lung specimens obtained from cadavers undergoing medicolegal autopsy. The NK activity of pulmonary lymphocytes obtained from the patients with lung cancer was significantly lower (p less than 0.05) than the NK activity of normal lungs. This reduction occurred despite high levels of blood NK activity in the patients with cancer, suggesting that NK cells might be locally suppressed in the lungs of patients with bronchogenic carcinoma. Because human pulmonary macrophages (PM) are known to be potent inhibitors of NK function, we investigated the role that PM might play in the reduction of NK activity in these patients. The PM obtained from the patients with lung cancer released soluble inhibitors of NK activity when stimulated with lipopolysaccharide. Release of these inhibitors was blocked by indomethacin, strongly suggesting a role for arachidonic acid metabolites as an inhibitor of pulmonary NK function. Inhibition of NK function by PM may occur in vivo, as a significant inverse correlation (r = -0.71, p less than 0.001) existed between the NK activity of lymphocytes obtained from a lung and the number of PM present in the lung.(ABSTRACT TRUNCATED AT 250 WORDS)