Accumulation of surfactant protein D in human pulmonary alveolar proteinosis

Surfactant protein D (SP-D) is a collagenous calcium-dependent carbohydrate-binding protein that is structurally related to the serum mannose-binding proteins and pulmonary surfactant protein A. SP-D was initially characterized as a biosynthetic product of freshly isolated rat type II cells and first purified in chemical amounts from bronchoalveolar lavage of rats with silica-induced alveolar lipoproteinosis. The present studies describe the characterization of human SP-D isolated from therapeutic bronchoalveolar lavage of patients with pulmonary alveolar proteinosis. Human proteinosis SP-D was extracted from the 10,000 x g pellet of bronchoalveolar lavage with 100 mmol/L glucose or ethylenediamine tetraacetic acid, and specifically bound to and eluted from maltosyl-agarose. The protein cross-reacted with monospecific antibodies to rat SP-D by enzyme-linked immunosorbent assay and immunoblot and eluted near the position of rat SP-D on reverse-phase high performance liquid chromatography. When chromatographed on 4% agarose (A-15M) in the presence of ethylenediamine tetraacetic acid, the solubilized human proteinosis SP-D eluted near the void volume and earlier than rat SP-D dodecamers or human SP-D multimers in the lavage supernatant. Two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting of proteins in the lavage pellet with antibodies to the carbohydrate-binding domain of proteinosis human SP-D demonstrated covalently cross-linked multimers of SP-D monomers (43 kd, reduced) and multimers of trimeric components stabilized by disulfide and non-disulfide bonds. These studies describe the isolation and biochemical characterization of human SP-D and demonstrate the abnormal accumulation of this protein in the air spaces of patients with alveolar proteinosis.     

Continuous intra-arterial blood gas monitoring during bronchopulmonary lavage for pulmonary alveolar proteinosis

We report a clinical experience of continuous intra-arterial blood gas monitoring (PARATREND 7:P 7) during bronchopulmonary lavage. A 47-year-old, 86-kg male was suffering from myelodysplastic syndrome associated with alveolar proteinosis. Bronchopulmonary lavage was scheduled to alleviate the symptom. P 7 and a continuous cardiac output monitor were used with an EKG monitor, a pulse oximeter, and a esophageal stethoscope. Anesthesia was maintained using sevoflurane and fentanyl. The patient's trachea was intubated with a double-lumen endobronchial tube (39-F Broncho-cath; Mallinckrodt, Ireland). After denitrogenation and degassing, warmed isotonic saline was infused into his left lung. The volume chosen was 2,200 ml which approximated the sum of the patient's functional residual capacity and tidal volume. The lowest point of PO2 appeared at the point 'degassed'. P 7 showed a consecutive rise of PO2 associated with infusion of large volume of saline. Although the response to change in PO2 of P 7 is slower than pulse oximeter, frequent blood sampling can be avoided. We think that P 7 with pulse oximeter are useful for the safe management of pulmonary lung lavage.     

Radiological evidence of involvement of dust exposure in pathogenesis of pulmonary alveolar proteinosis

A 52-year-old man was admitted because of increasing dyspnea on exersion and presence of pulmonary infiltrates. The patient had pulmonary tuberculosis at the age of 31, which resulted in volume loss and calcified foci in the upper lobe of his left lung. As a construction worker for more than 20 years, he had been exposed to inorganic dusts. Chest radiographs showed a symmetrical consolidation of infiltrates in both lungs with the exception of the left upper lobe, where no apparent infiltrates were shown. A computed tomographic scan of the chest revealed widely panlobular consolidation with the exception of the left upper lobe. A diagnosis of pulmonary alveolar proteinosis (PAP) was established by analysis of bronchoalveolar lavage fluid. Although the patient underwent segmental bronchoalveolar lavage four times under general anesthesia, he suffered frequent pulmonary infection and died two years after the onset his symptoms. Interestingly, the patient had a markedly narrowed orifice in the left upper lobe, as demonstrated by fiberoptic bronchoscopy. Chest radiographs of this lung field revealed no infiltrative shadows. These results suggest that some inhalative agent was involved in the pathogenesis of PAP in this case. In addition, significantly increased levels of KL-6 detected in both serum and bronchoalveolar lavage fluid were attributable to overproduction of KL-6 by Type II pneumocytes that had been stimulated or damaged by PAP.     

Attenuated hematopoietic response to granulocyte-macrophage colony-stimulating factor in patients with acquired pulmonary alveolar proteinosis

The pathogenesis of acquired pulmonary alveolar proteinosis (PAP), a rare lung disease characterized by excessive surfactant accumulation within the alveolar space, remains obscure. Gene-targeted mice lacking the hematopoietic growth factor granulocyte-macrophage colony-stimulating factor (GM-CSF) or the signal-transducing beta-common chain of the GM-CSF receptor have impaired surfactant clearance and pulmonary pathology resembling human PAP. We therefore investigated the hematopoietic effects of GM-CSF in patients with PAP. The hematologic response of 5 infants with congenital PAP to 5 microgram/kg/d was of normal magnitude. By contrast, despite normal expression of GM-CSF receptor alpha- and beta-common chains on peripheral blood myelomonocytic cells (n = 6) and normal binding affinity of bone marrow mononuclear cells for GM-CSF (n = 3), each of the 12 patients with acquired PAP treated displayed impaired responses to GM-CSF; 5 microgram/kg/d produced only minor eosinophilia, and doses of 7.5 to 20 microgram/kg were required to induce >/=1.5-fold neutrophil increments in the 3 patients who underwent dose-escalation. However, neutrophilic responses to 5 microgram/kg granulocyte colony-stimulating factor (G-CSF) were normal (n = 4). In vitro, the proportion of hematopoietic progenitors responsive to GM-CSF (16.1% +/- 8.9%; P = .042) or interleukin-3 (IL-3; 19.3% +/- 7.7%; P = .063), both of which utilize the beta-common chain of the GM-CSF receptor complex, were reduced among patients with acquired PAP (n = 4) compared with normal bone marrow donor controls (47.2% +/- 25.9% and 40.9% +/- 18.6%, respectively). In the one individual who had complete resolution of lung disease during the period of study, this was temporally associated with correction of this defective in vitro response to GM-CSF and IL-3 on serial assessment. These data establish that patients with acquired PAP have an associated impaired responsiveness to GM-CSF that is potentially pathogenic in the development of their lung disease. Based on these observations, we propose a model of the pathogenesis of acquired PAP that suggests the disease arises as a consequence of an acquired clonal disorder within the hematopoietic progenitor cell compartment.     

Elevated levels of lung surfactant protein A in sera from patients with idiopathic pulmonary fibrosis and pulmonary alveolar proteinosis

An enzyme-linked immunosorbent assay using monoclonal antibodies to human lung surfactant protein A (SP-A) was applied to sera from patients with lung diseases. We examined whether SP-A appears in the sera of patients with diseases that are known to cause alterations in surfactant composition in bronchoalveolar lavage fluids, and we characterized the SP-A that was found. The level of SP-A in sera from 57 healthy volunteers was 45 +/- 3 ng/ml (mean +/- SEM). The levels in patients with idiopathic pulmonary fibrosis (IPF) (205 +/- 23 ng/ml, n = 32) and pulmonary alveolar proteinosis (PAP) (285 +/- 23 ng/ml, n = 6) were significantly higher than those in healthy control subjects (p < 0.01), whereas those of sarcoidosis (n = 16), pneumonia (n = 14), and tuberculosis (n = 14) were 52 +/- 27 ng/ml, 65 +/- 11 ng/ml, and 49 +/- 23 ng/ml, respectively. Electrophoresis and immunoblotting analysis demonstrated that the fraction isolated from serum of a patient with PAP or IPF by anti-SP-A immunoaffinity column chromatography consisted chiefly of human IgG and IgM, and that it also contained SP-A. Furthermore, IgG was found in preparation of purified human SP-A. SP-A was demonstrated to bind to nonimmune IgG coated onto microtiter wells. Gel filtration analysis revealed that serum SP-A was eluted at fractions of larger molecular size than was the purified SP-A. These findings suggest that SP-A appears in the bloodstream as a complex with immunoglobulin in IPF and in PAP.     

Severe aplastic anaemia relapsing during a pregnancy; spontaneous remission following termination

IFN-gamma and TNF-alpha plasma levels were measured before and after local treatment in 27 patients. Twenty healthy subjects served as controls. Plasma concentrations of IFN-gamma and TNF-alpha were significantly higher before treatment (178.7 +/- 11.9 pg/ml and 31.9 +/- 11.6 pg/ml, respectively) compared to the control group (139.6 +/- 7.86 pg/ml and 17.1 +/- 7.7 pg/ml, respectively). After treatment IFN-gamma levels were significantly decreased (151.3 +/- 8.3 pg/ml) toward the control group values and TNF-alpha levels were observed even lower than in the controls (11.48 +/- 6.8 pg/ml). No correlations were found between age, duration of psoriasis and plasma levels of cytokines. However, IFN-gamma levels were related, although not significantly, to disease severity (evidenced by the PASI score). The data support the important proinflammatory role of IFN-gamma and TNF-alpha in the clinical manifestation of psoriasis.     

The clinical and physiological effect of whole-lung lavage in pulmonary alveolar proteinosis: a ten-year experience

We have utilized whole-lung lavage in the successful treatment of 18 patients with pulmonary alveolar proteinosis. Our ten-year experience includes serial evaluations of patients with disabling lung dysfunction who had a total of 49 whole-lung lavages under general anesthesia. Clinical and physiological responses were documented both before and after each lavage. There were no complications or deaths. All patients were radiographically, physiologically, and symptomatically improved within hours after the procedures. Five patients required from two to four repeat lavages one to three years later. The treatment of this disorder has included a wide variety of techniques. We attribute our results to the use of a lung lavage technique that includes: (1) unilateral whole-lung lavages at two to three day intervals; (2) isotonic saline as the lavage solution; (3) use of a mechanical chest percussor during lavage; and (4) measuring the total thoracic compliance of each side in the immediate postlavage period as a guide for extubation. We conclude that whole-lung lavage is a safe, highly effective, repetitively applicable treatment for pulmonary alveolar proteinosis.     

Adenovirus-mediated granulocyte-macrophage colony-stimulating factor improves lung pathology of pulmonary alveolar proteinosis in granulocyte-macrophage colony-stimulating factor-deficient mice

Mutation of the granulocyte-macrophage colony-stimulating factor (GM-CSF) gene by homologous recombination causes progressive pulmonary alveolar proteinosis (PAP) in GM-CSF-deficient mice (GM-/-). The present study tested whether adenovirus-mediated expression of GM-CSF alters the progression of PAP in GM-/- mice. Adult mice were pretreated with an anti-T cell receptor (TCR) antibody to block T cell-mediated immune response, followed by intratracheal instillation of deltaE1-E3 replication-deficient adenovirus expressing mouse GM-CSF (Av1mGM). Mice were killed 1, 3, and 5 weeks after treatment to assess lungs for GM-CSF, surfactant protein B (SP-B), alveolar macrophage maturation, and type II cell proliferation. GM-CSF was detected in BAL fluid from GM-/- mice 1 week after Av1mGM treatment, and GM-CSF mRNA was detected by RT-PCR through 5 weeks. Five weeks after Av1mGM treatment, PAP was improved and SP-B decreased as assessed by ELISA and immunostaining. Increased numbers of alveolar macrophages stained with alpha-naphthyl acetate esterase (alpha-NAE) following treatment with Av1mGM. Local expression of GM-CSF with a recombinant adenovirus ameliorated PAP in the GM-/- mice in association with enhanced maturation of alveolar macrophages.     

Pulmonary alveolar proteinosis in a child: HRCT findings before and after bronchoalveolar lavage

We demonstrate the high-resolution computed tomography (HRCT) findings of pulmonary alveolar proteinosis in a child before and after bronchoalveolar lavage. The CT pattern in our case differs from the pattern described in previous reports. We found a more homogeneous distribution of the pulmonary changes and a "crazy paving" pattern. High-resolution CT may be helpful in the differential diagnosis of this rare disease and in the follow-up of the pulmonary changes after bronchoalveolar lavage.     

Lungs of patients with idiopathic pulmonary alveolar proteinosis express a factor which neutralizes granulocyte-macrophage colony stimulating factor

Mice deficient in granulocyte-macrophage colony stimulating factor (GM-CSF) develop pulmonary alveolar proteinosis (PAP). We found that bronchoalveolar lavage fluid (BALF) from 11 patients with idiopathic pulmonary alveolar proteinosis (IPAP) suppressed the growth of peripheral blood monocytes and TF-1 cells, a cell line dependent on either GM-CSF or interleukin-3 (IL-3). The inhibitory effect of PAP-BALF occurred only when TF-1 cells were cultured with GM-CSF but not when cultured with IL-3, suggesting that PAP-BALF contains a factor that specifically interferes with GM-CSF function. 125I-GM-CSF binding to TF-1 cells was prevented in the presence of BALF from IPAP patients. Furthermore, cross-linking of 125I-GM-CSF to IPAP-BALF produced two major bands on SDS-PAGE; these bands were not observed in normal BALF. These data suggest that IPAP is caused by expression of binding factor(s) which inhibit GM-CSF function in the lung.     

Unusual relapse of adult T-cell leukemia/lymphoma after spontaneous remission

Signalling via the receptor Notch, delivered by the ligands Delta and Serrate, plays a key role in many cell fate decisions in both Drosophila and vertebrate development (for review seeArtavanis-Tsakonas, S., Matsuno, K. and Fortini, M.E., 1995. Notch signalling. Science 268, 225-232; Lewis, J., 1996. Neurogenic genes and vertebrate neurogenesis. Curr. Opin. Neurobiol. 6, 3-10; Blair, S.S., 1997. Limb development: marginal fringe benefits. Curr Biol. 7, 686-690; Irvine, K.D. and Vogt, T.F., 1997. Dorsal-ventral signaling in limb development. Curr. Opin. Cell Biol. 9, 867-876). Recently vertebrate homologues of Notch (Notch1; Myat, A., Henrique, D., Ish-Horowicz, D. and Lewis, J., 1996. A chick homologue of Serrate and its relationship with Notch and Delta homologues during central neurogeneis. Dev. Biol. 174, 233-247) and Serrate (Serrate1 and 2; Myat, A., Henrique, D., Ish-Horowicz, D. and Lewis, J., 1996. A chick homologue of Serrate and its relationship with Notch and Delta homologues during central neurogeneis. Dev. Biol. 174, 233-247; Hayashi, H., Mochii, M., Kodama, R., Hamada, Y., Mizuno, N., Eguchi, G. and Tachi, C., 1996. Isolation of a novel chick homolog of Serrate and its coexpression with Notch-1 in chick development. Int. J. Dev. Biol. 40, 1089-96; Laufer, E., Dahn, R., Orozco, O.E., Yeo, C.Y., Pisenti, J., Henrique, D., Abbott, U., Fallon, J.F. and Tabin, C., 1996. Expression of Radical fringe in limb-bud ectoderm regulates apical ectodermal ridge formation. Nature 386, 366-373; Rodriguez-Esteban, C., Schwabe, J.W., De La Pena, J., Foys, B., Eshelman, B. and Izpisua-Belmonte, J.C., 1997. Radical fringe positions the apical ectodermal ridge at the dorsoventral boundary of the vertebrate limb. Nature 386, 360-366) were shown to be expressed in early chick limb mesenchyme and apical ridge. However, later expression patterns of these genes and of Delta 1 (Henrique, D. , Adam, J., Myat, A., Chitnis, A., Lewis, J. and Ish-Horowicz, D., 1995. Expression of a Delta homologue in prospective neurons in the chick. Nature 375, 787-790) in vertebrate limbs have not been documented. We have used whole mount in-situ hybridization to document expression patterns of Notch1, Serrate1, Serrate2 and Delta1 within the mesenchyme of the developing chick limb up to stage 31 of development. We show these genes are expressed, in different combinations, in the vasculature, the musculature and the tissues of the handplate.     

Persistence of pulmonary pathology and abnormal lung function in IL-3/GM-CSF/IL-5 beta c receptor-deficient mice despite correction of alveolar proteinosis after BMT

Mice deficient for the IL-3/GM-CSF/IL-5 beta c receptor (beta cR KO) develop lung disease similar to that seen in human pulmonary alveolar proteinosis (PAP) which includes lymphocytic infiltration around airways and vessels and the progressive accumulation of surfactant and macrophages within the alveolar space. We investigated bone marrow transplantation (BMT) as a curative treatment of PAP in beta cR KO mice by semiquantitative histologic analysis and evaluation of pulmonary function. BMT from wild-type (WT) donors into lethally irradiated beta cR KO recipients (WT --> KO) led to the complete resolution of alveolar protein accumulation and to normalization of BAL fluid cellularity and macrophage morphology. However, detailed microscopic analysis of lung tissue revealed the persistence of significant cellular infiltrates in WT --> KO recipients which were equivalent to those seen in KO --> KO animals. Evaluation of pulmonary function demonstrated that only dynamic compliance (Cdyn) and not airway conductance (G[L]) was significantly improved in the WT --> KO group compared to KO --> KO animals and that both of these measurements remained significantly abnormal when compared to WT --> WT controls. We conclude, that although BMT for PAP reverses alveolar macrophage and protein accumulation, it does not decrease the interstitial inflammatory component of this disease. The importance of this residual pathology is demonstrated by the incomplete correction of alveolar function (Cdyn) and lack of improvement in increased airway resistance (G[L]). These findings may have important implications with regard to the extent that BMT can be considered a potential curative procedure for this clinical disorder.     

Quantity and structure of surfactant proteins vary among patients with alveolar proteinosis

Alveolar proteinosis (AP) is an idiopathic condition characterized by excess alveolar surfactant. Although the surfactant proteins (SP) are known to be aberrant, little is known of their variation between patients or their abundance relative to the lipids. We have examined surfactant composition in lavage fluid from 16 normal subjects and 13 patients with AP, one of whom was lavaged on 11 occasions over approximately 13 mo. In this patient we have examined composition on each occasion and in each sequential lavage aliquot. Composition was constant between right and left lung, but it differed markedly between patients. The cholesterol/disaturated phospholipid ratios (CHOL/DSP) were invariably elevated, on average by approximately 7-fold, whereas the SP-A/DSP and SP-B/DSP ratios were generally elevated, in some cases by as much as approximately 40- and approximately 100-fold, respectively. Although AP lavage generally contained more non-thiol-dependent SP-A aggregates and low Mr isoforms, the two-dimensional immunochemical staining patterns varied between patients and right and left lung. In the patient lavaged on multiple occasions, the SP-A/DSP and SP-B/DSP ratios progressively decreased as the patient's condition resolved. Because the SP-B/SP-A ratio was normal in all cases, we suggest that structural changes to the proteins occurred secondarily and that caution must be used in comparing functional data derived using SP-A obtained from patients with AP.     

A case of spontaneous remission of paragonimiasis miyazakii

A 52-year-old man was admitted with fever and chest pain. Chest X-ray showed a soft infiltration in the right lung and bilateral pleural effusions. A strong tuberculin reaction was elicited. Significant laboratory findings included eosinophilia (37% in peripheral blood and 78% in pleural fluid) and elevated IgE levels (577 IU/ml in sera and 6700 IU/ml in pleural fluid). Adenosine deaminase activity in the pleural fluid was high. No helminth eggs were detected after repeated examination of the pleural fluid and sputum. No definitive diagnosis was made. Three months of chemotherapy with INH and rifampicin resulted in little improvement. Corticosteroid was then administered orally under a tentative diagnosis of idiopathic eosinophilic pleurisy, which proved to be a successful treatment and resulted in a marked reduction of pleural fluid volume. Two years after discharge, the patient's chest X-ray was normal and laboratory findings were normal including the eosinophil count and IgE level. The pleural fluid obtained at the first admission and kept frozen was subjected to immunological analysis for anti-parasite antibody activity. The pleural fluid showed an unexpectedly high titer of antibody activity (x6400 dilution) against Paragonimus miyazakii antigen assayed by double diffusion Ouchterlony method. Examination of the sera obtained from the patient two years after discharge, however, revealed no detectable antibody activity against the parasite antigens assayed either by Ouchterlony or ELISA method. We concluded from the clinical as well as laboratory findings that the patient had recovered from Paragonimiasis miyazakii without specific intervention for the disease.     

Successful thrombolysis after prosthetic pulmonary valve obstruction under aspirin monotherapy

In a 22-year-old woman with recent onset of left-sided chest pain and exertional dyspnea, echocardiography revealed obstruction of a St. Jude Medical bileaflet prosthetic valve (size 23 mm) in the pulmonary position. Oral anticoagulation had been replaced for the previous 7 years by aspirin as the sole antithrombotic treatment. The valve had been inserted 16 years ago for pulmonary atresia. Valve function was restored by systemic application of 9 million units of urokinase.     

Successful pregnancy after conditioning with cyclophosphamide and fractionated total body irradiation

We report the case of a 24-year-old man who received high-dose cyclophosphamide (CY) 120 mg/kg over 2 days and twice daily fractionated total body irradiation (TBI) over 3 days(1,320 cGy) prior to allogeneic bone marrow transplantation. Seven and one-half years later he fathered a normal child who has developed normally so far.