Surfactant associated protein-A inhibits human lymphocyte proliferation and IL-2 production.

The hyporesponsive state of lung-derived mononuclear leukocytes has been, in part, attributed to the effects of the lipid rather than the protein components of pulmonary surfactant. In the present study, however, the results suggest that purified preparations of pulmonary surfactant-associated protein A (SP-A) suppress both phytohemagglutinin (PHA, 1 microgram/ml)- and anti-CD-3 (1 to 10 ng/ml) activated proliferation of human peripheral blood and tonsillar mononuclear cells in a dose-dependent manner at concentrations as low as 50 pM (6.25 micrograms/ml) when added at the initiation of cultures. Addition of SP-A to PHA-stimulated peripheral blood mononuclear cells (PBMC) as late as 24 to 36 h after PHA was also capable of suppressing [3H]thymidine incorporation measured at 72 h. In contrast, concanavalin A (Con A; 2 micrograms/ml)-stimulated PBMC proliferation was slightly augmented by the addition of SP-A. Analysis of the supernatants of PHA-stimulated cultures treated with SP-A revealed that accompanying the inhibition of proliferation was a corresponding decline in measurable interleukin-2 (IL-2) concentrations, from 154 pg/ml for the PHA-treated cells to 57.8, 28.4, 5.2, and less than 2 pg/ml of IL-2 when SP-A was added at 6.25, 12.5, 25, and 50 micrograms/ml, respectively. We suggest that the action of SP-A on PHA-stimulated human PBMC may involve the blocking of a costimulatory signal crucial for in vitro T-cell activation.

Multiparameter flow cytometric DNA analysis of effusions: a prospective study of 36 cases compared with routine cytology and immunohistochemistry.

Single-parameter flow cytometry (SFCM) is limited in its ability to detect aneuploid and diploid malignant cells or accurately estimate S-phase fractions (SPF) in effusions because of the high degree of contamination by benign mesothelial cells and inflammatory cells. We examined 36 pleural and peritoneal fluids by conventional cytology and multiparameter FCM (MFCM) to analyze the DNA content of cells expressing epithelial markers cytokeratin, epithelial membrane antigen, carcinoembryonic antigen, BRST-1, or BRST-3 (B72.3) and compared the results to those found with SCFM. The cases were also studied by immunohistochemistry using the same antibody panel. By routine cytology, 14 of the 36 cases were classified as carcinomas, 11 as reactive, 1 as mesothelioma, and 10 as suspicious. MFCM allowed reclassification of 5 of the 10 suspicious cases as carcinomas and the remaining 5 as reactive cases based on ploidy and marker expression. Whereas SFCM detected only 13 nondiploid carcinomas, MFCM detected 4 diploid and 15 nondiploid carcinomas. All reactive cases were diploid by SFCM or MFCM. The mesothelioma case showed were distinct peaks by MFCM, a diploid peak with SPF of 13.4% and a tetraploid peak with SPF of 36.1%. The SPF of the nondiploid carcinomas ranged from 5.9 to 50.4% and diploid carcinomas, from 3.5 to 14.5% when gated on epithelial cells. The reactive cases had SPF ranging from 0.4 to 4.4%.(ABSTRACT TRUNCATED AT 250 WORDS)

Calcitriol and its synthetic analogue MC 903 inhibit the interleukin-2-induced migration of human lymphocytes.

Sarcoidosis is characterized by the accumulation of activated lymphocytes in the lungs and other organs and by the spontaneous production of the active form of vitamin D, calcitriol. We hypothesized that calcitriol may modulate the responsiveness of human lymphocytes to the relevant biologic mediator, interleukin-2 (IL-2). After culture for 48 h with phytohemagglutinin, human peripheral blood lymphocytes migrated through nitrocellulose filters, secured in microchemotaxis chambers, in response to IL-2. When calcitriol at 1 nM was included in the cultures, the migratory response to IL-2 was completely abrogated. This inhibitory effect was seen despite the fact that cultured lymphocytes continued to express the IL-2 receptor and other activation markers. A similar but more rapid effect could be demonstrated by including calcitriol in the lower well during our 3-h chemokinesis assay. Calcitriol blocked IL-2-induced lymphocyte migration in a dose-dependent fashion. The synthetic noncalcemic vitamin D analogue MC 903 was equally effective in this assay. IL-2-induced migration could also be prevented by the protein kinase C inhibitor H-7, but calcitriol appeared to be at least 1,000 times more potent. Our studies suggest that calcitriol is a potent natural immunomodulator with rapid suppressive effects that may be mediated through protein kinase C. Synthetic analogues such as MC 903 may offer exciting therapeutic opportunities.

Effect of platelet activating factor (PAF) on the migration of human lymphocytes.

BACKGROUND: There is growing evidence to suggest the importance of the lymphocyte in the pathogenesis of asthma, particularly in the late phase reactions and ongoing bronchial hyperreactivity. Platelet activating factor (PAF) has also been identified as a potentially important mediator in asthma. METHODS: The migration of human peripheral blood lymphocytes obtained from normal volunteers in response to PAF and the effect of PAF antagonists was studied in a well standardised in vitro assay using nitrocellulose micropore filters in a microchemotaxis chamber. RESULTS: PAF is a potent stimulus to in vitro human lymphocyte migration; at an optimal concentration of 1 nM it augmented lymphocyte chemokinesis to 310% (SE 33%) of control values. The response to PAF appears to be specific since lyso-PAF and other related membrane phospholipids had no effect. PAF-induced migration could be abrogated by specific PAF receptor antagonists such as WEB 2086 (100 nM), and was partially blocked by the cyclooxygenase inhibitor flurbiprofen at a concentration of 1 microM. CONCLUSIONS: PAF stimulates the in vitro migration of human lymphocytes through a specific PAF receptor. Part of the response may be due to the generation of cyclooxygenase products. PAF may play a part in the recruitment of lymphocytes to asthmatic airways.

Necrotizing sarcoid granulomatosis with pleural involvement. Clinical and radiographic features.

The clinical, functional, radiologic, and pathologic characteristics of seven cases of necrotizing sarcoid granulomatosis (NSG) are presented. The population consisted of five women and two men, with an average age of 36 years. Each patient's predominant presenting complaint was pleuritic chest pain. Pulmonary function testing demonstrated a variety of abnormal patterns. Computed tomography (CT) of the chest showed solitary or multiple nodules in all patients, occasionally associated with pulmonary infiltrates in the lower lobes. Pleural involvement was seen on CT scanning in six patients and mediastinal adenopathy was present in five. Biopsy specimens of the lung lesions revealed confluent epithelioid granulomata associated with necrosis and vasculitis. Pleural involvement by confluent granulomata was a prominent feature in four patients. Follow-up has ranged from 6 months to 4 years. All patients are now asymptomatic, the majority having received prednisone. One patient received methotrexate as a steroid-sparing measure. We conclude that NSG is distinguishable from sarcoidosis as a clinicopathologic entity in which pleural involvement is a frequent finding. Treatment with steroids appears to hasten recovery.

Theophylline delays skeletal muscle fatigue during progressive exercise.

In this study, 31P nuclear magnetic resonance spectroscopy (NMRS) was used to examine the effect of theophylline on human forearm muscle metabolism during progressive exercise. Six healthy men (37 +/- 14 yr of age) were assigned to either a control (CTRL) group (n = 3), or a theophylline treatment (THEO) group (n = 3). Each subject performed two dynamic wrist flexion exercise tests to fatigue, with at least 72 h separating each trial. The THEO group repeated the protocol after receiving 300 mg of sustained-release theophylline every 12 h. 31P spectra were acquired every 36 s throughout exercise, and the relative contributions of the phosphate metabolites and pH were determined. Power output at the onset, or threshold of intracellular acidosis (IT), was identified for each subject from changes in phosphocreatine (PCr) metabolism and pH. Power at maximal exercise and at the IT was found to be reproducible in the CTRL group. After theophylline administration, the maximal power attained by the THEO group increased significantly by 19% (p < 0.05), from 2.25 +/- 0.2 to 2.68 +/- 0.15 W. A similar trend occurred in the onset of the IT, which was also prolonged by 19%, from 1.33 +/- 0.18 to 1.58 +/- 0.22 W. Therapeutic concentrations of theophylline significantly increased the endurance of the forearm musculature, apparently by delaying the onset of intracellular metabolic acidosis. These findings suggest an enhancement of oxidative capacity of the muscle.

Thoracic magnetic resonance imaging in the evaluation of HIV-1/AIDS pneumonitis.

Magnetic resonance imaging of the thorax was performed on ten occasions in eight HIV-positive patients with a clinical picture suggestive of Pneumocystis carinii pneumonia. The diagnosis of PCP was subsequently confirmed on six occasions. Patients without PCP had low MRI profusion scores, while four of six patients with PCP had MRI profusion scores greater than 6/21. Neither the chest roentgenogram appearance nor computer-generated T1 and T2 relaxation times could reliably distinguish between these two groups. Magnetic resonance imaging may be useful in the early and noninvasive diagnosis of PCP in HIV-positive patients.

Lymphocyte chemokinetic factors derived from human tonsils: modulation by 1,25-dihydroxyvitamin D3 (calcitriol).

Although interleukin (IL)-2 may in part be responsible for lymphocyte accumulation to sites of active sarcoidosis, other cytokines that control such recruitment are not well characterized. Similarly, the pathogenic rationale for the ability of sarcoid macrophages to produce 1,25-dihydroxycholecalciferol (calcitriol) is not understood. We studied the release of chemokinetic lymphokines from human nylon wool-non-adherent tonsillar lymphocytes (HNTLs) employing a standard in vitro lymphocyte migration assay. If mitogen-stimulated HNTL supernatants were fractionated by high-performance liquid chromatography, five positive and one negative chemokinetic factors could be identified. The five lymphocyte chemoattractant factors (LCFs) ranged in mol wt from 5 to 35 kD and stimulated the in vitro migration of nonsensitized human lymphocytes by 200 to 500%. The LCFs appeared distinct from IL-2, IL-1, or gamma-interferon. Co-incubation of HNTLs with mitogen and 1 nM calcitriol prevented the production or release of two of the LCFs and significantly decreased the quantity of a third LCF. Calcitriol also resulted in the appearance of a second negative chemokinetic factor, lymphocyte migration inhibitory factor (LyMIF). Combined with our previous studies demonstrating that calcitriol interferes with IL-2-induced lymphocyte migration, these results provide a rationale for an anti-inflammatory role for calcitriol in sarcoidosis and other granulomatous disorders. These experiments also demonstrate that the control of lymphocyte recruitment to inflammatory foci is multifactorial.

Inhibitors of membrane transmethylation reactions prevent the lymphocyte chemokinetic response.

The methylation of membrane phospholipids has been shown to occur following receptor-mediated activation of leukocytes. The present studies show that the human lymphocyte response to two positive chemokinetic signals (bradykinin and lymphocyte chemoattractant factor) can be interrupted by inhibitors of S-adenosyl-L-methionine-mediated transmethylation reactions. The chemokinetic response to the nonphysiologic stimulant colchicine is not affected. We speculate that phospholipid methylation accompanies receptor-mediated lymphocyte migration, and may facilitate activation of second messenger systems.

Bradykinin augments the in vitro migration of nonsensitized lymphocytes.

Bradykinin and related peptides have been considered important as mediators of acute inflammation, but their role in the cell-mediated immune response has not been extensively investigated. We have examined the effect of physiological concentrations of these autacoids on the migration of nonsensitized lymphocytes employing an in vitro micropore filter assay system. Bradykinin at a concentration of 0.01-1 nM significantly stimulated the migration of both human peripheral blood and rat splenic lymphocytes. Related naturally-occurring kinins (kallidin, MetLys-bradykinin, desArg9-bradykinin) had a similar effect but other autacoids (angiotensins I and II, histamine, serotonin) were inactive. Bradykinin was more active on freshly harvested lymphocytes than on cells incubated for up to 48 h with or without mitogen. Bradykinin appeared to act predominantly as a chemokinetic agent (augmenting random nondirectional motility), as determined by checkerboard analysis. Bradykinin appeared to exert its effect on lymphocytes predominantly through the B1 class of bradykinin receptors. The migratory response to bradykinin could not be attributed to the release of arachidonic acid metabolites, but stimulated migration could be significantly inhibited by the histamine type 2 receptor antagonist cimetidine. These studies provide a novel mechanism whereby nonsensitized lymphocytes may be recruited to sites of delayed-type hypersensitivity reactions.

Gold-naproxen pneumonitis. A toxic drug interaction?

A patient with rheumatoid arthritis developed restrictive lung disease and blood eosinophilia. Gold pneumonitis was suspected but the patient did not improve until naproxen was discontinued as well. Lymphocyte transformation studies suggested hypersensitivity to gold. We hypothesize that naproxen unmasked and perpetuated the manifestations of gold hypersensitivity in our patient.

Rat lymphokines control the migration of nonsensitized lymphocytes.

We investigated whether mediators released from rat splenic mononuclear cells could control the in vitro migration of nonsensitized resting rat lymphocytes. Rat splenocytes stimulated with concanavalin A, other mitogens, or histamine release three lymphokines that alter rat lymphocyte migration. A positive chemokinetic factor, termed lymphocyte chemoattractant factor (LCF), has a molecular weight (MW) between 50 and 70 kDa. Two negative chemokinetic lymphokines can also be identified; lymphocyte migration inhibitory factor (LyMIF, MW 25-45 kDa) and a high MW inhibitor (HWMI, MW greater than 70 kDa). Lymphokines were destroyed by heat as well as by treatment with neuraminidase and trypsin. The action of LCF and LyMIF was prevented by phenylmethylsulfonylfluoride, a specific serine esterase inhibitor, and the action of LyMIF was also blocked by alpha-L-fucose. The discovery of these mediators provides the opportunity to study the importance of such chemokinetic lymphokines in animal models of disease.

The nylon wool adherence of rat mononuclear cells: physical, chemical and biological influences.

The application of mononuclear cell populations to a nylon wool (NW) column is a common early procedure in the selection of T lymphocytes for further study. The technique as presently employed in various laboratories does not appear to be standardized, and surprisingly little is known about the effects of physicochemical alterations and biologic mediators on the interaction of lymphocytes with the NW substrate. In this study the factors controlling the adherence of rat splenocytes to NW were examined. NW adherence was shown to be independent of loaded cell concentration, column packing or pH, but was very dependent on NW column size, wash volume, incubation time and ambient temperature. The addition of protein to media did not alter lymphocyte NW adherence, and the interaction did not appear to depend on intact cellular glycolysis or protein synthesis or on microtubular or microfilament function. Because of the theoretical importance of surface adherence to cell motility, the effects of various agents that alter lymphocyte migration were tested in the lymphocyte NW assay. Of the positive chemokinetic factors tested, only casein altered (decreased) NW adherence. Of the negative chemokinetic principles tested only the human lymphokine LyMIF altered (increased) NW adherence. The studies show that the NW-nonadherent cell pool may be a heterogeneous population depending on the physical conditions of the assay, and the NW adherence of rat splenocytes is not an all-or-none phenomenon but can be altered by physical and biological factors. This makes the standardization of the assay of critical importance, particularly if one wishes to compare results of subsequent experiments between laboratories.

Hypophosphatemia and respiratory failure: prolonged abnormal energy metabolism demonstrated by nuclear magnetic resonance spectroscopy.

Hypophosphatemia has been shown to cause acute respiratory failure. The mechanism is believed to be due to decreased high-energy substrate availability at the cellular level leading to respiratory muscle dysfunction. However, direct measurement of these substrates has not been previously studied. A patient with hypophosphatemic respiratory failure is described in whom phosphocreatine and pH were continuously monitored using nuclear magnetic resonance spectroscopy. This revealed a defect in muscle metabolism that required several weeks to recover despite prompt correction of the serum phosphate level.

Proton magnetic resonance imaging to stage activity of interstitial lung disease.

Patients with active interstitial lung disease (ILD) are often treated with high-dose corticosteroids, although such therapy is not universally effective and has significant risks. Clinicians have utilized various ancillary diagnostic techniques to help with difficult treatment decisions. Since magnetic resonance imaging (MRI) has the theoretic and experimental potential of differentiating various stages of ILD, we prospectively studied 34 adult patients in a 0.15 Tesla resistive magnet body imager. The most severely affected patients, who were thought to warrant a steroid trial, had the greatest MRI image intensity, and dramatic improvement was seen following successful treatment. Qualitative MRI data were as useful as any other ancillary diagnostic technique (radiographic, physiologic, scintigraphic) in predicting clinical course. Computer-generated relaxation times were not sufficiently precise to differentiate active from inactive disease. Although limited by availability and cost, MRI appears to be a useful ancillary diagnostic technique in ILD patients facing immunomodulating therapy.

Magnetic resonance imaging to diagnose intralobar pulmonary sequestration.

Confirmation of the diagnosis of intralobar pulmonary sequestration has usually required angiographic demonstration of the systemic arterial supply. We report a young man who presented with non-resolving pneumonia where magnetic resonance imaging (MRI) suggested the correct diagnosis by demonstrating two arteries arising from the aorta supplying the sequestrum. MRI appears to offer a safe, noninvasive alternative for the diagnosis of sequestration.

Lung cancer following therapy for Hodgkin's disease.

We describe a patient in whom lung cancer developed several years after he had received combined-modality therapy for Hodgkin's disease. The literature concerning second malignant diseases, particularly thoracic tumours, that occur following combined-modality therapy for cancer is reviewed. It is important to recognize these entities, because chest symptoms or findings on x-ray films may be misinterpreted as representing late recrudescence of the first neoplastic disease.

Functional characteristics of histamine receptor-bearing mononuclear cells. II. Identification and characterization of two histamine-induced human lymphokines that inhibit lymphocyte migration.

Although functional histamine receptors have generally been restricted to those human T lymphocytes expressing suppressor cell functions, more recent evidence suggests that histamine receptor-bearing human T lymphocytes are functionally heterogeneous and capable of other immunomodulatory activities. Lymphocyte chemoattractant factor (LCF) is a cationic sialoprotein with an apparent m.w. of 56,000, whose production is limited to histamine-type 2 receptor-bearing human T cells. LCF is selectively chemokinetic for T lymphocytes, and presumably contributes to the recruitment of unsensitized effector lymphocytes at inflammatory sites. In addition to LCF, Sephadex G-100 gel filtration of histamine-induced lymphocyte supernatants revealed two regions of migration inhibitory activity for human blood T and rat splenic lymphocytes. These regions corresponded to m.w. of 70,000 to 80,000 (LyMIF75K) and 30,000 to 40,000 (LyMIF35K). LyMIF75K had a single pI of 7.5 to 8.0, and its biologic activity was sensitive to trypsin but not to neuraminidase or heat (56 degrees C). LyMIF35K had a single pI of 8.5 to 8.8, and its biologic activity was sensitive to neuraminidase and heat but not to trypsin. These LyMIFs therefore appeared to be distinct from one another and physicochemically different from other migration inhibitory lymphokines. All three lymphokine activities appeared within 4 hr of incubation. The minimum concentration of histamine required to stimulate production of the LyMIF was 10(-6) M. Lymphocytes that did not adhere to a histamine affinity matrix were unable to produce either LyMIF upon subsequent stimulation with histamine or concanavalin A (Con A). Lymphocytes incubated with histamine and diphenhydramine produced LCF but neither LyMIF, whereas cells incubated with histamine in the presence of cimetidine produced both LyMIF but not LCF. These data suggest that a subset of lymphocytes defined by the presence of histamine-type 1 receptors are capable of producing two distinct species of lymphocyte migration inhibitory activity. These cells may contribute to the immobilization of effector T lymphocytes chemokinetically attracted to certain inflammatory sites.