Bronchoalveolar lavage fluid cytology in patients with Pneumocystis carinii pneumonia

OBJECTIVE: To evaluate bronchoalveolar lavage (BAL) cytology and organism burden in patients with Pneumocystis carinii pneumonia (PCP) who were infected with the human immunodeficiency virus (HIV) and in those with other immunodeficiencies. STUDY DESIGN: BAL fluid samples from patients with PCP were selected (HIV-infected patients, n = 15; patients with other immunodeficiencies, n = 11). May-Grünwald-Giemsa-stained cytocentrifuge preparations were evaluated. Foamy alveolar casts (FACs) and P carinii clusters were counted. RESULTS: The numbers of FACs and P carinii clusters in BAL fluid samples of HIV-infected patients were significantly higher as compared to those in samples from patients with other immunodeficiencies. Striking cytologic findings observed in half the samples from both patient groups included the presence of foamy alveolar macrophages, activated lymphocytes, plasma cells and reactive type II pneumocytes. Furthermore, a peculiar cell type, "nonidentified cell" (NIC), was observed almost exclusively in BAL fluid samples from HIV-infected patients. CONCLUSION: BAL fluid samples from HIV-infected patients with PCP displayed higher organism burdens as compared to those from patients with other immunodeficiencies. Moreover, cytologic findings suggestive of noninfectious lung conditions were common in BAL fluid samples obtained from patients with PCP. Further study is required to elucidate the identity of the NIC cell type.     

Differential cell analysis of cytocentrifuged bronchoalveolar fluid samples affected by the area counted

OBJECTIVE: To investigate variations in the differential cell counts between the quadrants of cytocentrifuged bronchoalveolar lavage (BAL) fluid preparations and to evaluate the diagnostic impact of these differences in interstitial lung diseases (ILD). STUDY DESIGN: BAL fluid samples obtained from 30 patients suspected of having ILD or pneumonia were cytocentrifuged and additionally stained with May-Grünwald-Giemsa stain. Two observers differentiated 200 cells in each quadrant as well as in a circular pattern around the center of the cytocentrifuge spot. RESULTS: Lymphocytes and alveolar macrophages were not randomly distributed on the cytocentrifuge spot. Ten samples of patients with histologically confirmed ILD were selected to test the diagnostic impact using a validated computer program. The predicted diagnosis did not correspond to the histologic diagnosis for one quadrant from 1 of these 10 samples (sarcoidosis instead of idiopathic pulmonary fibrosis), whereas the differential cell counts performed around the center of the cytocentrifuge spot provided the correct diagnosis in all cases. CONCLUSION: BAL fluid differential cell counts varied between the quadrants of the cytocentrifuge spot. The center of the cytocentrifuge spot appeared to be the most reliable area. Therefore, cell counting is recommended in a circular pattern around the center of the cytocentrifuge spot.     

Leukocyte immunophenotypes in bronchoalveolar lavage fluid and peripheral blood of paracoccidioidomycosis, sarcoidosis and silicosis.

Leukocyte subsets in bronchoalveolar lavage (BAL) fluid and peripheral blood of patients with paraccoccidioidomycosis, sarcoidosis and silicosis were characterized using monoclonal antibodies and an immunoperoxidase technique. In paraccocidioidomycosis, the number of T-helper/inducer CD4-positive lymphocytes was lower in peripheral blood than in BAL fluid. Additional analysis showed that the expression of HLA-DR was very similar in alveolar macrophages, lung and blood T-cells. In sarcoidosis and silicosis there were higher proportions of T-helper/inducer cells in peripheral blood than in BAL fluid. The alterations in the T-helper/inducer/T-suppressor/cytoxic CD4/CD8 ratio in sarcoidosis and silicosis were more appreciable in peripheral blood than in BAL fluid, contrasting with the results in paracoccidioidomycosis. The expression of HLA-DR by alveolar macrophages in sarcoidosis was the highest of all the disease studied. No statistically significant differences were observed between chronic multifocal and chronic unifocal paracoccidioidomycosis disease, stage II and stage III sarcoidosis, and chronic and accelerated silicosis. The three granulomatous diseases analyzed had a few alveolar macrophages expressing the CD4 molecule on their surface. These findings and the technique of analyzing both peripheral blood and BAL leukocyte subsets may help to understand the pathogenesis of interstitial lung diseases.     

IL-12 and IL-18 are increased and stimulate IFN-gamma production in sarcoid lungs

Sarcoidosis is a systemic chronic granulomatous disease of unknown cause. Recent investigations revealed that the cytokine profile in inflamed lesions of sarcoidosis is Th1 dominant. To obtain better immunopathologic understanding of sarcoidosis, we examined the expression of IL-12 and IL-18 and their roles in IFN-gamma production in pulmonary sarcoidosis. Sarcoid cases had significantly elevated levels of IL-12 (p40 and p70) and IL-18 in bronchoalveolar lavage (BAL) fluids compared with healthy subjects. IL-12 p70 and IL-18 were immunohistochemically expressed in the epithelioid cells and giant cells of sarcoid granulomas. Significant induction of IFN-gamma, IL-12 p70, and IL-18 was observed from sarcoid BAL fluid cells with LPS stimulation, whereas LPS tended to induce only IL-12 p70 in BAL fluid cells from healthy subjects. Sarcoid cases had significantly greater IFN-gamma induction with LPS stimulation than healthy subjects did. IL-18 mRNA expression was observed in freshly isolated sarcoid BAL fluid cells as well as in LPS-stimulated sarcoid BAL fluid cells, but IFN-gamma and IL-12 mRNA expression was observed only in LPS-stimulated BAL fluid cells. Treatment with anti-IL-12- and anti-IL-18-neutralizing Abs significantly inhibited IFN-gamma production from LPS-stimulated BAL fluid cells of sarcoid cases. Coadministration of rIL-12 or rIL-18 induced greater IFN-gamma production in sarcoid BAL fluid cells than in normal BAL fluid cells. We concluded that bioactive IL-12 and IL-18 were produced in sarcoid BAL fluid cells and synergistically induced IFN-gamma production, indicating important cytokines in the Th1 response of sarcoidosis.     

Bronchoalveolar lavage fluid differential cell count. How many cells should be counted?

OBJECTIVE: To investigate the number of cells to be counted in cytocentrifuged bronchoalveolar lavage (BAL) fluid preparations in order to reach a reliable enumeration of each cell type. STUDY DESIGN: A total of 136 BAL fluid samples for patients with suspected pneumonia or interstitial lung disease were investigated. Differential cell counts were performed on May-Grünwald-Giemsa-stained cytocentrifuged preparations by 2 observers, each differentiating 500 cells. Reliability for the enumeration of each cell type was expressed as phi value, as calculated in generalizability theory. RESULTS: For polymorphonuclear neutrophils (PMNs), alveolar macrophages, lymphocytes and eosinophils, an acceptable phi value of > or = .95 was reached at a count of 300 cells by 1 observer. Mast cells reached a phi value of only .674 at a count of 500 cells by 1 observer, precluding a reliable count. At a count of 500 cells by 1 observer, squamous epithelial cells, bronchial epithelial cells and plasma cells displayed phi values of .868, .903 and .816, respectively. CONCLUSION: At a count of 300 cells, PMNs, alveolar macrophages, lymphocytes and eosinophils are reliably enumerated in cytocentrifuged BAL fluid samples.     

Cytokine profile and proteome analysis in bronchoalveolar lavage of patients with sarcoidosis, pulmonary fibrosis associated with systemic sclerosis and idiopathic pulmonary fibrosis

The aim of this study was to analyze the type of immune response (Th1, Th2) and protein composition of bronchoalveolar lavage (BAL) of patients with sarcoidosis, pulmonary fibrosis associated with systemic sclerosis (SSc) and idiopathic pulmonary fibrosis (IPF). Flow cytometry analysis of intracellular cytokines revealed different patterns: in IPF and SSc Th2 profiles were predominant, whereas in sarcoidosis Th1 prevailed. The proteomic analysis of BAL fluid (BALF) showed that there were quantitative differences between the three diseases. These were more evident between sarcoidosis and IPF, confirming our previous observations, whereas SSc had an intermediate profile between the two, however with some peculiarities. Comparison of BALF protein maps, constructed with the same quantity of total proteins, enabled us to identify the main profiles of the three diseases: an increase in plasma protein prevalent in sarcoidosis and also present in SSc, though for fewer proteins with respect to IPF and a greater abundance of low molecular weight proteins, mainly locally produced, in IPF. These findings are in line with the different pathogenesis of these diseases: IPF is considered a prevalently fibrotic disorder limited to the lung, with intense local production of functionally different proteins, whereas sarcoidosis and SSc are systemic immunoinflammatory diseases.     

Depressed urokinase activity in bronchoalveolar lavage fluid from patients with sarcoidosis,silicosis or idiopathic pulmonary Rbrosis: relationship to disease severity

Intraalveolar fibrinolysis, is regulated by the concerted actions of plasmin, plasminogen activators (PAs), and their specific inhibitors (PAIs). This event is considered as a critical step in the pathogenesis of pulmonary fibrosis. The aim of this study was to evaluate whether local PA activity can be held as a marker of fibrosis in chronic interstitial lung disorders (ILD). Changes in both PA activity and PA-related proteins (urokinase-type PA (uPA), tissue-type PA (tPA), PAI-1 and PAI-2) were assessed in bronchoalveolar fluid (BALF) of 60 subjects: 18 healthy controls, 18 non-fibrotic sarcoidosis patients, 16 patients with idiopathic pulmonary fibrosis (IPF) and eight silicotic patients with established fibrosis. We observed a significant decrease of BALF PA activity in the three groups of patients as compared with controls. Reduction in BALF PA activity was compatible with lower uPA protein levels associated, especially in IPF patients, with an increased occurrence of PAI-1 and PAI-2 antigens. Soluble tPA antigen was never detected either in control subjects or in patients. Most importantly, the reduction in BALF PA activity and uPA protein levels was found to be most severe in patients with advanced fibrotic disease, namely IPF, while moderate and only weak alterations were found in silicosis and non-fibrotic sarcoidosis, respectively. In addition, significant positive correlations were found between BALF PA activity and functional impairment as assessed by TLC % and DL_CO%. Finally, the reduction in uPA and PA activity levels observed in BALF from sarcoidosis patients was found to be proportional to the degree of BAL lymphocytosis. These findings indicate that an intense reduction in BALF PA activity is associated with severe stages of the parenchymal disease, possibly reflecting the degree of the fibrotic process.     

Macrophage derived chemokine (CCL22), thymus and activation-regulated chemokine (CCL17), and CCR4 in idiopathic pulmonary fibrosis

Background

Idiopathic pulmonary fibrosis (IPF) is a chronically progressive interstitial lung disease of unknown etiology. Previously, we have demonstrated the selective upregulation of the macrophage-derived chemokine CCL22 and the thymus activation-regulated chemokine CCL17 among chemokines, in a rat model of radiation pneumonitis/pulmonary fibrosis and preliminarily observed an increase in bronchoalveolar (BAL) fluid CCL22 levels of IPF patients.

Methods

We examined the expression of CCR4, a specific receptor for CCL22 and CCL17, in bronchoalveolar lavage (BAL) fluid cells, as well as the levels of CCL22 and CCL17, to elucidate their pathophysiological roles in pulmonary fibrosis. We also studied their immunohistochemical localization.

Results

BAL fluid CCL22 and CCL17 levels were significantly higher in patients with IPF than those with collagen vascular diseases and healthy volunteers, and there was a significant correlation between the levels of CCL22 and CCL17 in patients with IPF. CCL22 levels in the BAL fluid did not correlate with the total cell numbers, alveolar lymphocytes, or macrophages in BAL fluid. However, the CCL22 levels significantly correlated with the numbers of CCR4-expressing alveolar macrophages. By immunohistochemical and immunofluorescence analysis, localization of CCL22 and CCR4 to CD68-positive alveolar macrophages as well as that of CCL17 to hyperplastic epithelial cells were shown. Clinically, CCL22 BAL fluid levels inversely correlated with DLco/VA values in IPF patients.

Conclusion

We speculated that locally overexpressed CCL22 may induce lung dysfunction through recruitment and activation of CCR4-positive alveolar macrophages.     

Diffuse alveolar damage. Morphologic features in bronchoalveolar lavage fluid

OBJECTIVE: To evaluate the overall cytologic characteristics of diffuse alveolar damage (DAD) in bronchoalveolar lavage (BAL) specimens in search of features that could be useful in cytologic diagnosis. STUDY DESIGN: We evaluated BAL samples from patients with DAD obtained simultaneously with transbronchial biopsies (n = 8) or open lung biopsies (n = 2) or within 24 hours of autopsy (n = 2). The material was processed routinely for cytologic and histologic evaluation. RESULTS: The smears were moderately to highly cellular. All cases had large numbers of alveolar macrophages and/or desquamated alveolar cells. The epithelial component displayed various degrees of nuclear atypia. Some epithelial clusters were three-dimensional, with peripheral cells showing clear cytoplasm, protruding outwards and resembling hobnails. Other aggregates appeared two-dimensional, as sheets of cells with flattened and dense cytoplasm (squamotized). Both types of cell clusters were often associated with dense, basophilic or amphophilic, amorphous extracellular material. Counterparts of all the cytologic features were observed in the histologic material, including atypia of the alveolar lining with hobnailing, squamotization, amorphous extracellular material and hyaline membranes. CONCLUSION: The cytologic features of BAL represent a constellation of alveolar cell injury. Based on these features, DAD can be correctly diagnosed or suggested in BAL samples in the appropriate clinical setting.     

Multinucleated giant cells in bronchoalveolar lavage

OBJECTIVE: To determine the frequency, morphology and possible diagnostic significance of multinucleated giant cells (MGC) in bronchoalveolar lavage (BAL). STUDY DESIGN: Retrospectively we examined 671 BAL specimens. Enlarged cells having > or = 10 nuclei were defined as MGC. Cytomorphologic features were described. BAL specimens containing MGC were grouped according to clinicohistologic diagnosis into sarcoidosis, asbestosis, other interstitial lung diseases and different chronic, noninterstitial lung diseases. RESULTS: MGC were present in 10.7% of BAL specimens and occurred in low numbers. MGC were classified into Langhans' or foreign-body-type MGC (LF-MGC), alveolar macrophage-like MGC (AM-MGC) and nonspecific MGC (NS-MGC). LF-MGC were found most often in patients with sarcoidosis. AM-MGC were found in all groups of patients. NS-MGC were found most often in patients with asbestosis and other interstital lung diseases. CONCLUSION: MGC in BAL are not encountered frequently and are not numerous. Based on cytomorphologic features, three types of MGC can be distinguished.     

Enhanced expression of Fas Ligand (FasL) in the lower airways of patients with fibrotic interstitial lung diseases (ILDs)

The exact role of FasL, and particularly its soluble and membrane-bound forms, in the development of chronic ILDs and lung fibrosis has not been extensively explored. We aimed at analyzing membrane-bound FasL expression on alveolar macrophages (AM) and lymphocytes (AL) as well as soluble FasL (sFasL) levels in bronchoalveolar lavage (BAL) from ILDs patients, incl. pulmonary sarcoidosis (PS), hypersensitivity pneumonitis (HP), silicosis, asbestosis, idiopathic pulmonary fibrosis (IPF), nonspecific interstitial pneumonia (NSIP), and healthy subjects (n = 89, 12, 7, 8, 23, 6, 17, respectively). In IPF, significantly increased percentage of AM FasL(+) and CD8(+)FasL(+) cells as well as sFasL levels in BAL were found. Increased sFasL levels were also observed in HP. NSIP and asbestosis were characterized by higher AM FasL(+) relative number; CD8(+)FasL(+) population was expanded in asbestosis only. There was a significant decline in AL FasL(+) percentage in PS and HP. Vital capacity was negatively correlated with sFasL levels, AM FasL(+) and CD8(+)FasL(+) cell relative count. CD4(+)FasL(+) and CD8(+)FasL(+) percentage strongly correlated with BAL neutrophilia, an unfavorable prognostic factor in lung fibrosis. The concurrent comparative BAL analysis of FasL expression indicates that FasL(+) AM and AL (mainly Tc cells) comprise an important element of the fibrotic process, mostly in IPF. FasL might play a crucial role in other fibrosis-complicated ILDs, like NSIP and asbestosis.     

Detection of Alveolar Fibrocytes in Idiopathic Pulmonary Fibrosis and Systemic Sclerosis

Background

Fibrocytes are circulating precursors for fibroblasts. Blood fibrocytes are increased in patients with idiopathic pulmonary fibrosis (IPF). The aim of this study was to determine whether alveolar fibrocytes are detected in broncho-alveolar lavage (BAL), to identify their prognostic value, and their potential association with culture of fibroblasts from BAL.

Methods

We quantified fibrocytes in BAL from 26 patients with IPF, 9 patients with Systemic Sclerosis(SSc)-interstitial lung disease (ILD), and 11 controls. BAL cells were cultured to isolate alveolar fibroblasts.

Results

Fibrocytes were detected in BAL in 14/26 IPF (54%) and 5/9 SSc patients (55%), and never in controls. Fibrocytes were in median 2.5% [0.4–19.7] and 3.0% [2.7–3.7] of BAL cells in IPF and SSc-ILD patients respectively. In IPF patients, the number of alveolar fibrocytes was correlated with the number of alveolar macrophages and was associated with a less severe disease but not with a better outcome. Fibroblasts were cultured from BAL in 12/26 IPF (46%), 5/9 SSc-ILD (65%) and never in controls. The detection of BAL fibrocytes did not predict a positive culture of fibroblasts.

Conclusion

Fibrocytes were detected in BAL fluid in about half of the patients with IPF and SSc-ILD. Their number was associated with less severe disease in IPF patients and did not associate with the capacity to grow fibroblasts from BAL fluid.     

Cytology of Bronchoalveolar Lavage In Some Rare Pulmonary Disorders: Pulmonary Alveolar Proteinosis and Amiodarone Pulmonary Toxicity

Cytological patterns of bronchoalveolar lavage (BAL) in pulmonary alveolar proteinosis (PAP) and amiodarone pulmonary toxicity (APT) are presented together with light and electron microscopy (EM). the differential cell count of BAL in both diseases is similar in that alveolar macrophages predominate. However, the cytology of PAP is characterized by scanty macrophages and alveolar epithelial cells in abundant periodic acid-Schiff (PAS)-positive extracellular material. the gross appearance of the BAL fluid is therefore opaque. In contrast, the cytology of APT is characterized by foamy alveolar macrophages with numerous lamellar bodies in their cytoplasm, and the BAL fluid is clear.     

Analysis of Trace Elements in Bronchoalveolar Lavage of Patients with Diffuse Lung Diseases

Airborne trace elements are implicated in the etio-pathogenesis of a large number of pulmonary diseases. The aim of this study was to evaluate the reliability and effectiveness of direct determination of Cd, Cr, Cu, Fe, Mn, Ni, Pb, V, and Zn concentrations in bronchoalveolar lavage (BAL) samples from patients with sarcoidosis, idiopathic pulmonary fibrosis, and Langerhans cell histiocytosis and healthy (smoking and non-smoking) controls. A total of 44 individuals were recruited among sarcoidosis, idiopathic pulmonary fibrosis, and Langerhans cell histiocytosis patients and healthy (smoking and non-smoking) controls. Average Mn concentrations in BAL from patients were 45% lower than in controls (p < 0.01) and remarkable decreases in average concentrations of Cr, Ni and Zn were also found in BAL from patients with idiopathic pulmonary fibrosis and Langerhans cell histiocytosis. As these diseases are characterized by the enhanced activation of certain immunomodulatory cells and by generation of free radicals, the depressed Mn, Zn, Cr and Ni concentrations in BAL from patients may be due to oxidative stress. These preliminary results indicate that assessment of the elemental composition of BAL is a promising approach to study the pathogenesis of diffuse lung diseases and Langerhans cell histiocytosis.     

Determinants of bronchoalveolar lavage cellularity in idiopathic pulmonary fibrosis.

To investigate factors that determine bronchoalveolar lavage (BAL) cellularity in patients with idiopathic pulmonary fibrosis (IPF), we compared BAL cells in patients with IPF (n = 83) to both nonsmoking (n = 111) and smoking (n = 19) normal volunteers. Patients with IPF had higher concentrations of BAL total cells and alveolar macrophages than nonsmoking volunteers and more BAL neutrophils and eosinophils than normal volunteers regardless of smoking status. Among patients with IPF, the numbers of alveolar macrophages, neutrophils, or eosinophils were strongly associated with either smoking status or pack-years of cigarette smoking. In fact, after accounting for cigarette smoking, using multivariate analysis, the only additional factors that were found to be associated with BAL cellularity were age (macrophages and eosinophils) and the percent predicted forced expired volume in 1 s (neutrophils). Additional multivariate models failed to identify a significant relationship between BAL cellularity and either the type of immunosuppressive therapy or other physiological measures of lung function. We conclude that cigarette smoking strongly influences BAL cellularity in patients with IPF. These findings suggest that cigarette smoking may have a role in the pathogenesis of IPF or may adversely affect the prognosis in patients with IPF.     

Evidence for local dendritic cell activation in pulmonary sarcoidosis

Background

Sarcoidosis is a granulomatous disease characterized by a seemingly exaggerated immune response against a difficult to discern antigen. Dendritic cells (DCs) are pivotal antigen presenting cells thought to play an important role in the pathogenesis. Paradoxically, decreased DC immune reactivity was reported in blood samples from pulmonary sarcoidosis patients. However, functional data on lung DCs in sarcoidosis are lacking. We hypothesized that at the site of disease DCs are mature, immunocompetent and involved in granuloma formation.

Methods

We analyzed myeloid DCs (mDCs) and plasmacytoid DCs (pDCs) in broncho-alveolar lavage (BAL) and blood from newly diagnosed, untreated pulmonary sarcoidosis patients and healthy controls using 9-color flowcytometry. DCs, isolated from BAL using flowcytometric sorting (mDCs) or cultured from monocytes (mo-DCs), were functionally assessed in a mixed leukocyte reaction with naïve allogeneic CD4+ T cells. Using Immunohistochemistry, location and activation status of CD11c⁺DCs was assessed in mucosal airway biopsies.

Results

mDCs in BAL, but not in blood, from sarcoidosis patients were increased in number when compared with mDCs from healthy controls. mDCs purified from BAL of sarcoidosis patients induced T cell proliferation and differentiation and did not show diminished immune reactivity. Mo-DCs from patients induced increased TNFα release in co-cultures with naïve allogeneic CD4⁺ T cells. Finally, immunohistochemical analyses revealed increased numbers of mature CD86⁺ DCs in granuloma-containing airway mucosal biopsies from sarcoidosis patients.

Conclusion

Taken together, these finding implicate increased local DC activation in granuloma formation or maintenance in pulmonary sarcoidosis.     

Local expansion of allergen-specific CD30+Th2-type gamma delta T cells in bronchial asthma.

BACKGROUND: T lymphocytes infiltrating airways during the allergic immune response play a fundamental role in recruiting other specialized cells, such as eosinophils, by secreting interleukin 5 (IL-5), and promoting local and systemic IgE synthesis by producing IL-4. Whether these presumed allergen-specific T cells are of mucosal or systemic origin is still a matter of conjecture. MATERIALS AND METHODS: Immunophenotype, IL-4 production, and in vitro proliferative response to specific or unrelated allergens were analyzed in the bronchoalveolar lavage (BAL) fluid lymphocyte suspensions obtained from untreated patients with allergic asthma. Healthy subjects and patients affected by pulmonary sarcoidosis, a granulomatous lung disease characterized by infiltrating Th1 CD4+ lymphocytes, served as controls. RESULTS: The proportions of gamma delta T lymphocytes, mostly CD4+ or CD4- (-)CD8-, was higher in asthmatic subjects than in controls (p < 0.05). Most BAL gamma delta CD4+ lymphocytes of asthmatic patients displayed the T cell receptor (TCR)-gamma delta V delta 1 chain. While CD30 antigen coexpression on the surface of BAL alpha beta(+) T lymphocytes was low (ranging from 5 to 12%), about half of pulmonary gamma delta T cells coexpressed it. These cells produced IL-4 and negligible amounts of interferon-gamma (IFN gamma), and proliferated in vitro in response to purified specific but not unrelated allergens. In contrast, control or sarcoidosis gamma delta T cells never displayed the CD30 surface molecule or produced significant quantities of IL-4. CONCLUSIONS: These findings not only confirm our previous hypothesis that the allergen-specific Th2-type lymphocytes found in the lungs of asthmatic patients are gamma delta T cells belonging to airway mucosal immunocytes, but also strongly support the notion that asthma is a local rather than a systemic disease.     

Stage related morphometry of sarcoid granulomas and inflammatory cell types in broncho-alveolar lavage.

The epithelioid granulomas, interstitial and intra-alveolar mononuclear inflammatory infiltrates and the cellular compartments obtained by broncho-alveolar lavage (BAL) were measured in 40 patients with pulmonary involvement of sarcoidosis. The granulomas were divided into a central (epithelioid-cellular) zone and into a peripheral (lymphocytic-fibrotic) zone, and the density of various inflammatory cells was measured in both compartments. The cases were grouped and analyzed according to the radiological and clinical stages. The results are as follows: neutrophilic granulocytes seen in the BAL probably originate from the sarcoid granulomas in patients with stage 1 of the disease and may derive from other compartments of the lung parenchyma in patients with stage 2 or stage 3. Lymphocytes seen in the BAL of patients with stage 1 or stage 2 of the disease probably derive from intra-alveolar lymphocytic agglutinations. They originate from the sarcoid granulomas only in patients with stage 3. Macrophages seen in the BAL probably derive from the sarcoid granulomas independent from the stage of sarcoidosis. No relationship was found between the morphometric parameters and the clinical outcome of the disease.     

RNA interference as a gene silencing therapy for mutant MYOC protein in primary open angle glaucoma

Bronchoalveolar lavage (BAL) is a useful diagnostic tool in interstitial lunge diseases (ILD). However, differential cell counts are often non specific and immunocytochemistry is time consuming. Staining of glyoproteins by periodic acid Schiff (PAS) reaction may help in discriminating different forms of ILD. In addition, PAS staining is easy to perform. BAL cells from patients with idiopathic pulmonary fibrosis (IPF) (n = 8), sarcoidosis (n = 9), and extrinsic allergic alveolitis (EAA) (n = 2) were investigated. Cytospins from BAL cells were made and cells were stained using Hemacolor quick stain and PAS staining. Lymphocytic alveolitis was found in sarcoidosis and EAA whereas in IPF both lymphocytes and neutrophils were increased. PAS positive cells were significantly decreased in EAA compared to IPF and sarcoidosis (25.5% ± 0.7% vs 59.8% ± 25.1% and 64.0% ± 19.7%, respectively) (P < 0.05). No significant correlation between PAS positive cells and inflammatory cells was observed. These results suggest that PAS staining of BAL cells may provide additional information in the differential diagnosis of ILD. Further studies ware warranted to evaluate PAS staining in larger numbers of BAL from patients with ILD.     

Shedding of soluble ICAM-1 into the alveolar space in murine models of acute lung injury

Intercellular adhesion molecule-1 (ICAM-1; CD54) is an adhesion molecule constitutively expressed in abundance on the cell surface of type I alveolar epithelial cells (AEC) in the normal lung and is a critical participant in pulmonary innate immunity. At many sites, ICAM-1 is shed from the cell surface as a soluble molecule (sICAM-1). Limited information is available regarding the presence, source, or significance of sICAM-1 in the alveolar lining fluid of normal or injured lungs. We found sICAM-1 in the bronchoalveolar lavage (BAL) fluid of normal mice (386 +/- 50 ng/ml). Additionally, sICAM-1 was spontaneously released by murine AEC in primary culture as type II cells spread and assumed characteristics of type I cells. Shedding of sICAM-1 increased significantly at later points in culture (5-7 days) compared with earlier time points (3-5 days). In contrast, treatment of AEC with inflammatory cytokines had limited effect on sICAM-1 shedding. BAL sICAM-1 was evaluated in in vivo models of acute lung injury. In hyperoxic lung injury, a reversible process with a major component of leak across the alveolar wall, BAL fluid sICAM-1 only increased in parallel with increased alveolar protein. However, in lung injury due to FITC, there were increased levels of sICAM-1 in BAL that were independent of changes in BAL total protein concentration. We speculate that after lung injury, changes in sICAM-1 in BAL fluid are associated with progressive injury and may be a reflection of type I cell differentiation during reepithelialization of the injured lung.