Search

We used a denaturing gradient gel electrophoresis (DGGE) procedure with 40-nucleotide guanine- and cytosine-rich sequences in the polymerase chain reaction (PCR) and sequencing analysis to analyze the T cell antigen receptor (TCR)-Vgamma gene repertoire of infiltrating T lymphocytes in pulmonary lymphoproliferative disorders. Six of 15 low-grade mucosa-associated lymphoid tissue (MALT) lymphomas and 8 of 15 cases of lymphocytic interstitial pneumonia (LIP) showed some oligoclonal bands for TCR-Vgamma genes on DGGE. Sequencing analysis demonstrated plural oligoclonal TCR-Vgamma clones among the oligoclonal PCR products on DGGE, leading to the conclusion that conventional antigen-specific oligoclonal expansions may play some role in the pathogenesis of pulmonary lymphoproliferative disorders. The frequency of oligoclonal infiltrating T cell expansions in human immunodeficiency virus (HIV)-related LIP (100%) was significantly higher than in low-grade pulmonary MALT lymphomas (40%) or in HIV-negative LIP (30%). Because recent evidence demonstrates that the V3 loop in the proviral amino acid sequences of mononuclear cells from bronchoalveolar lavage is more homogeneous than those from peripheral blood, this homogeneity might result in oligoclonal expansions of infiltrating T lymphocytes as a consequence of ongoing reactions against lung-specific viral strains.

Pulmonary surfactant is a complex mixture of phospholipids and proteins, which is present in the alveolar lining fluid and is essential for normal lung function. Alterations in surfactant composition have been reported in several interstitial lung diseases (ILDs). Furthermore, a mutation in the surfactant protein C gene that results in complete absence of the protein has been shown to be associated with familial ILD. The role of surfactant in lung disease is therefore drawing increasing attention following the elucidation of the genetic basis underlying its surface expression and the proof of surfactant abnormalities in ILD.

To elucidate the immunogenetic background of idiopathic inflammatory myopathies (IIM) such as polymyositis (PM), dermatomyositis (DM) and any overlapping subsets, with other collagen vascular diseases, HLA class I antigens and class II alleles were determined and compared from individuals with various clinical and serological features of IIM, including pulmonary interstitial lesions (PI). Seventy-three Japanese patients with myositis (32 PM, 18 DM, 23 overlapped subsets) and 62 healthy unrelated controls were enrolled onto the study. Statistical differences between groups were determined by the Fisher's exact probability test. Serum fluorescent antinuclear antibody, rheumatoid factor (RF), anti-SS-A/Ro antibody, anti-Jo1 antibody and anti-U1 RNP antibody were examined using routine methods. PI was detected by chest X-ray and/or computed tomography. In patients with DM, the frequency of the HLA-DRB1*1302-DQA1*0102-DQB1*0604 haplotype was significantly higher than in the healthy controls (42.1% vs 17.7%), and in the patients with PM (42.1% vs 9.4%). Furthermore, the frequency of the HLA-DRB1*0405-DQA1*03-DQB1*0401 haplotype was higher in the PM patients with PI than in the controls (50.0% vs 17.7%), and PM without PI (50.0% vs 5.5%). These results suggest that in terms of HLA class II association, Japanese DM and PM, and PM with and without PI, belong to different clinical groups.

Dyskeratosis congenita (DC) is a rare inherited multisystem disorder characterised by lesions of the skin and appendages. Bone marrow failure occurs in 80% of patients. The gene for the X-linked form of DC has been identified on Xq28 and designated as DKC1. Pulmonary manifestations have rarely been reported. It is not known whether there is a respiratory disease peculiar to these patients and, if so, whether it is associated with a specific genetic mutation. A 40 year old Egyptian man with pulmonary disease and his symptom free 35 year old brother both presented with mucocutaneous lesions characteristic of DC. In the older brother chest imaging revealed generalised intralobular interstitial thickening and honeycombing. Pulmonary function tests showed a restrictive pattern. Open lung biopsy specimens of lung tissue showed various degrees of fibrosis consistent with usual interstitial pneumonia of chronic idiopathic pulmonary fibrosis. The younger brother was free of pulmonary lesions. Both had a novel missense mutation 5C-->T in exon 1 of the DKC1 gene. It is concluded that pulmonary disease in DC may be underestimated, possibly because most patients die at an early age of bone marrow failure. No relationship between genotype and phenotype could be established in the patients studied. The genetic diagnosis of DC is now available, which may enable it to be diagnosed in patients with restrictive pulmonary disease and minimal cutaneous signs.

Lymphoid interstitial pneumonitis (LIP), a frequent pulmonary complication in HIV-infected pediatric patients, is characterized histologically by marked infiltration of lymphoid cells. We sought to evaluate the nature and pathogenesis of the lymphoid infiltrates and to examine the relationship of LIP to pulmonary MALT lymphoma that has been described in pediatric HIV positive patients. To examine the potential contribution of chemokines and cytokines to the inflammatory cell recruitment in tissues involved by lymphoid interstitial pneumonitis from HIV-infected pediatric patients, RNA was extracted from paraffin-embedded tissues from five lung biopsies in four pediatric HIV-positive patients and from five control, normal lung biopsies in five HIV-negative patients and was analyzed by semiquantitative RT-PCR for the expression of cytokines (TNF-alpha, GM-CSF, IFN-gamma, IL-4, IL-6, IL-10, and IL-18) and chemokines (IP-10, Mig, regulated upon activation, normal T expressed and secreted [RANTES], and MIP1-alpha and beta) after normalization for G3PDH. Expression of IL-18 was increased, as well as expression of IFN-gamma-inducible chemokines IP-10 and Mig in LIP tissues compared with controls. RANTES and MIP1-alpha and -beta were also increased in pediatric LIP lesions compared with controls. In contrast, expression of TNF-alpha, GM-CSF, IL-10, and IL-6 was variable in LIP tissues and controls. In addition, clonality of the B-cell population was evaluated by VDJ-PCR. A polyclonal B-cell population was shown in all five biopsies from five patients with LIP; and in one patient with concurrent LIP and MALT lymphoma, a band of increased intensity was observed in the LIP biopsy that was identical in size to the monoclonal band in the concurrent MALT lymphoma biopsy. These results provide evidence of high-level expression of certain chemokines in lymphoid interstitial pneumonitis tissues and suggest that chemokines and cytokines may play an important role in the recruitment of inflammatory cell infiltrates into these tissues. In addition, LIP may represent an early stage of MALT lymphoma or an immunologic response to a chronic antigenic stimulus that may provide a milieu or microenvironment for the evolution of a monoclonal B-cell population.

The immunosuppressive activity of interleukin-10 (IL-10) makes this cytokine a potentially important clinical tool to reduce inflammatory responses in various diseases. Its efficacy as a therapeutic modality is dependent on the responsiveness of immune cells. We report that macrophages from mice chronically infected with the LP-BM5 retrovirus had a reduced capacity to respond to IL-10 in vitro. The ability of IL-10 to inhibit lipopolysaccharide-induced production of tumor necrosis factor (TNF) alpha and IL-6 was significantly reduced in both alveolar and peritoneal macrophages from infected versus uninfected mice. IL-10 hyporesponsiveness was not related to direct infection by the retrovirus, because bone marrow-derived macrophages infected in vitro with LP-BM5 were as responsive to IL-10 as were uninfected bone marrow-derived macrophages. TNF-alpha appeared to contribute to development of IL-10 hyporesponsiveness, because exposure of normal macrophages to TNF-alpha but not interferon-gamma reduced macrophage responsiveness to IL-10. Reverse transcriptase-PCR and flow cytometry demonstrated normal expression of the alpha and beta chains of the IL-10 receptor in macrophages from infected mice, suggesting that IL-10 hyporesponsiveness is not related to a change in receptor expression. The potential role of reduced IL-10 responsiveness in the chronicity of inflammation in this and other diseases is discussed.

Understanding of the cellular and cytokine interactions associated with inflammation and fibrosis in interstitial lung diseases (ILDs) has increased substantially during the past few years. Presently, many agents are known to have the ability to induce ILDs, although only a small percentage of exposed individuals will develop the disease. In addition, the majority of ILDs are of unknown origin and many are labelled "idiopathic". Therefore, host susceptibility, genetic factors and, possibly, environmental cofactors may be important for the clinical expression of ILDs. The present review reports evidence of the genetic predisposition to develop ILDs of unknown origin, more specifically sarcoidosis, idiopathic pulmonary fibrosis (IPF), lymphangioleio-myomatosis and ILDs, in systemic sclerosis. For instance, for sarcoidosis and IPF several histocompatibility antigens have been associated with the development and/or the clinical presentation of the disease. Furthermore, there are also several types of ILD that are associated with inherited disorders, of which the tuberous sclerosis complex is only one example. This clearly indicates that pulmonary fibrosis can be influenced by genetic factors. Familial occurrence of sarcoidosis and IPF is also well known, although the exact modes of inheritance are debatable. Several studies have shown that extrinsic factors, such as single or multiple fibrosing agents, probably contribute to the development of clinical ILDs of unknown origin. It is probable that some of these studies deal with patients who do not have classical IPF, as recently defined by the American Thoracic Society (ATS)/European Respiratory Society (ERS) consensus. Therefore, the true role of these extrinsic factors in the development of IPF, or even sarcoidosis, remains speculative. With the help of animal studies and, more specifically, by using knock-out mice, it may be possible in the near future to unravel at least some of the genes that are responsible for the increased susceptibility of the development of interstitial lung diseases.

Epidemiological studies have suggested that many rare diseases with respiratory involvement have a genetic component. Molecular advances have increased the understanding of the pathophysiology of these diseases which has led to better diagnostic and prognostic methods. There may be many genes responsible for diseases such as primary ciliary dyskinesia and systemic lupus erythematosus in addition to the effect of modifier genes. The genotype:phenotype correlation in these diseases remains to be elucidated. In some diseases, such as familial dysautonomia and sickle cell, the gene has been identified which allows for accurate pre-natal testing. Further, in diseases where the genetic abnormality is known, such as chronic granulomatous disease, gene therapy remains a realistic prospect and phase I studies are about to commence or currently underway. This article reviews those rare diseases in which there is or is likely to be a significant genetic contribution.

A case is described of Niemann-Pick type C2 disease presenting an infantile pneumopathic phenotype known to occur in this recently established, second, minor complementation group of Niemann-Pick type C (NPC) disease. However, the pulmonary involvement was unique, being dominated, in addition to the usual storage macrophage infiltration of the alveolar and septal compartments, by irregular emphysema attributed to storage cell migration into the bronchiolar lumen. The latter modified considerably the X-ray findings and hindered the initial clinical diagnosis. Otherwise, the storage phenotype, including the range of stored lipids, storage distribution, and cell and organ pathology, was found to be identical to that in the whole Niemann-Pick type C disease group dominated by NPC1. The biochemical findings (cholesterol esterification level) corresponded to the classical biochemical phenotype. Emphysema should thus be considered as a variant of the pulmonary NPC2 storage process, governed most probably by an epigenetic mechanism responsible for storage macrophage migration into the bronchiolar compartment.

Objective: To determine the contribution of surfactant protein abnormalities to the development of chronic lung injury in a familial form of interstitial lung disease., Study Design: An 11-year-old girl, her sister, and their mother who were diagnosed with chronic interstitial lung disease underwent laboratory investigation of surfactant protein expression in bronchoalveolar lavage fluid and lung biopsy specimens. Nineteen patients with idiopathic pulmonary fibrosis and 9 patients who were investigated for pulmonary malignancy but who did not have interstitial lung disease served as control subjects., Results: The 3 family members were found to have absent surfactant protein C (SP-C) and decreased levels of SP-A and SP-B in bronchoalveolar lavage fluid (BALF). Immunostaining for pulmonary surfactant proteins in lung biopsy specimens obtained from both children demonstrated a marked decrease of pro-SP-C in the alveolar epithelial cells but strong staining for pro-SP-B, SP-B, SP-A, and SP-D. No deviations from published surfactant protein B or C coding sequences were identified by DNA sequence analysis. All control subjects had a detectable level of SP-C in the BALF., Conclusion: The apparent absence of SP-C and a decrease in the levels of SP-A and SP-B are associated with familial interstitial lung disease.

An insertion/deletion (I/D) polymorphism of the angiotensin-converting enzyme has previously been studied extensively in relationship to cardiovascular and renal disease. The deletion/deletion (D/D) genotype is associated with a poor outcome in immunoglobulin (Ig) A nephropathy. However, the association of this genetic marker in cardiovascular and renal disease has generated controversy, with the exception of the rate of progression and therapeutic responsiveness in IgA nephropathy. Many of the same cytokines and polypeptide mediators involved in fibrosis of the cardiovascular and renal systems have been shown to be involved in pulmonary fibrosis. We examined the I/D polymorphism of the angiotensin-converting enzyme in a group of 24 patents with interstitial pneumonia and moderate to severe pulmonary fibrosis defined by radiographic studies, pulmonary function tests, and histologic findings. The incidence of the D allele in this study population was 69.0%, which is approximately 15.0% higher than the incidence in the general population of 54.0%. The incidence of the D/D genotype was 42.0%, which is approximately 11.0% greater than that in the general population (31.0%). The distribution of the D/D, I/D, and insertion/insertion genotypes of these 24 patients was not significantly different from that of historical controls (P =.1; chi(2) test); there were marginally significantly more D alleles among the 48 observed alleles than would be expected (P =.04).

The p63 gene has high homology with p53, but more complex physiologic functions, including the regulation of the maintenance of basal cells in stratified epithelia. These cells in fact express high levels of the deltaN-terminal truncated isoforms of the p63 gene that can act as dominant-negative inhibiting the activity of p53. Basal cells in human bronchi and bronchioli seem to use the same strategy, since they constitutively express high levels of p63, at variance with alveolar pneumocytes. Over-expression of these isoforms in airway basal cells can inhibit important functions of the p53-pathway, including cell cycle arrest and apoptosis. This finding underlines the key role of p63 in epithelial renewal in human lung, with important implications in the understanding of the mechanisms of tissue remodelling occurring in diffuse lung diseases.

The two-step polymerase chain reaction (PCR) and sequencing analysis was used to analyze the immunoglobulin heavy chain variable (Ig V(H)) genes of open-chest biopsy or autopsy samples from five patients with Epstein-Barr virus-negative human immunodeficiency virus (HIV)-related lymphoid interstitial pneumonia (LIP), and the results were compared with those for Ig V(H) genes from five HIV-negative LIP patients. The findings of this study are consistent with the different immunological situations of HIV-related and HIV-negative LIP. (a) The Ig V(H)3 subgroup was underexpressed in three of five cases of HIV-related LIP. In contrast, none of the HIV-negative cases showed this abnormality. Because the Ig V(H)3 subgroup encodes the largest portion of Ig V(H) genes, a depletion of B cells expressing Ig V(H)3 genes reflects a major alteration in the B-cell compartment. (b) All HIV-related LIP cases demonstrated two or three oligoclonal populations. HIV-negative cases showed minor monoclonal or polyclonal populations, but not oligoclonal ones. These oligoclonal populations suggest the coexistence of several occult clonal B-cell populations in HIV-related LIP. (c) Some oligoclonal clones in HIV-related LIP showed mutated framework regions not demonstrated in HIV-negative clones. This degree of variation exceeds the usual mutation rate for frameworks, suggesting a role for framework residues in antigen binding. (d) The frequency of D-D fusions of minor oligoclonal clones (HIV-related LIP) is higher than that of minor monoclonal clones (HIV-negative LIP). Such D-D fusions may enhance the probability of expression of antibodies capable of binding HIV glycoproteins.

To elucidate which cells are synthesizing tenascin in usual interstitial pneumonia (UIP) we have analyzed thoracoscopic or open lung biopsies from 30 patients with UIP by mRNA in situ hybridization, using (35)S-labeled tenascin RNA probes. The phenotype of the cells expressing tenascin mRNA was confirmed by immunohistochemical stainings of serial sections with antibodies against alpha-smooth muscle actin and human cytokeratin. The results demonstrate that tenascin is expressed at the foci of recent lesions consisting of intralumenal or incorporating loose fibrotic buds. The cells expressing tenascin mRNA were located in and underneath the newly formed epithelium. Immunohistochemical stainings showed that the cells in the newly formed epithelium were strongly cytokeratin positive, and thus evidently regenerating type 2 pneumocytes, while the cells underneath the newly formed epithelium were alpha-smooth muscle actin positive and apparently myofibroblasts. Tenascin mRNA expression was clearly stronger and more frequent in myofibroblasts than in type 2 pneumocytes, however. Weak tenascin mRNA expression was also found in metaplastic bronchiolar-type epithelium and alveolar macrophages. Our results are thus in good agreement with the previous studies showing that tenascin is actively synthesized at the early fibrotic lesions in UIP. Furthermore, results demonstrate that the interaction between the epithelium and the underlying connective tissue plays a significant role in tenascin synthesis and that myofibroblasts are mainly responsible for its synthesis in fibroblastic foci of UIP.

There is increasing evidence implicating activated macrophages in the pathogenesis of interstitial and other lung diseases. We investigated whether there was a unique pattern of cell surface expression that constituted a disease-specific phenotype on alveolar macrophages from patients with interstitial lung disease (ILD). Macrophage cell surface receptor expression of 19 selected markers was assessed by indirect immunofluorescence and flow cytometry in bronchoalveolar lavage (BAL) fluids from patients with idiopathic pulmonary fibrosis (IPF, n = 4), scleroderma (SCL-ILD, n = 14), mild asthma (n = 7), allergy without asthma (n = 2), and normal subjects (n = 9). There was increased expression of adhesion receptors (CD11c, CD29, CD36, CD44, CD49e, CD54), receptors involved in signal transduction and/or inflammation (CD13, CD45, CD53), and other markers (CD9, CD52, CD71, CD98, HLA Class I) on macrophages from ILD patients compared to the non-ILD group. Most markers upregulated on macrophages in ILD were significantly inversely correlated with clinical parameters of disease activity such as FEV(1), FVC, and DL(CO) and positively correlated with numbers of BAL neutrophils and eosinophils. Increased expression of several cell surface markers suggests that activated alveolar macrophages may contribute to the pathophysiology of IPF and SCL-ILD., (Copyright 2000 Academic Press.)

We report a case of de novo myelodysplastic syndrome with clonal eosinophilia (MDS-Eo) and eosinophilic pulmonary interstitial infiltration, confirmed by autopsy. Cytogenetic study using Giemsa banding identified 47,XY,+1,der(1;7)(q10;p10),+8 in the marrow cells. Simple Giemsa staining revealed the same chromosomal aberration in metaphase spreads with eosinophilic granules, indicating the clonal proliferation of eosinophils. To our knowledge, our case is the 6th reported case of MDS-Eo with cytogenetically confirmed clonal eosinophilia, and the first autopsy of MDS-Eo. A review of the literature combined with our findings suggests that this type of chromosomal aberration might be involved in the as yet unknown pathogenesis of MDS-Eo.

In the present study, K-ras mutation was investigated in 156 neogenetic epithelia that appeared in the lesion of subpleural fibrosis in order to elucidate the close relationship of lung cancer development with pulmonary interstitial pneumonia. The neogenetic epithelia were histologically subclassified into six types: (i) ciliated bronchial epithelium (CBE); (ii) squamous metaplastic epithelium (SME); (iii) cuboidal immature epithelium (CIE); (iv) stratified immature epithelium (SIE); (v) mucus cell epithelium (MCE); and (vi) intestinal metaplastic epithelium (IME). K-ras mutation was detected in 9.6% of neogenetic epithelia overall; 21% of CIE, 12% of SIE, 16% of SME, but not in other types of neogenetic epithelia. Immunohistochemically, CIE and SIE frequently expressed surfactant apoprotein and SME was characteristic to carcinoembryonic antigen expression. According to Ki-67 immunostain, CIE, SIE and SME are likely to grow faster than other histological types of epithelia. K-ras mutation was seen exclusively in codon 12 with predominant G to A and G to C substitutions without any G to T transversions, results which are somewhat different to previous studies in lung cancers. The present study clearly demonstrated that K-ras mutation appeared in certain histological types of neogenetic epithelia, but raised the question of whether K-ras mutation in neogenetic epithelia during the inflammatory reparative process might be an early genetic event in lung carcinogenesis.

Fas is expressed in various cells and transduces the cell death signal. p21 is a mediator of p53-dependent G1 arrest associated with deoxyribonucleic acid (DNA) damage. The upregulation of p53 and p21 associated with DNA damage in idiopathic pulmonary fibrosis has been described previously. In this study, p53, p21, and Fas expression and DNA damage were examined in interstitial pneumonia associated with collagen vascular diseases (CVD-IP). DNA damage was assessed by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate biotin nick end-labelling (TUNEL) and p53, p21 and Fas proteins were detected by immunohistochemistry in 13 cases of CVD-IP, 13 of sarcoidosis, seven of hypersensitivity pneumonitis (HP) and eight control patients with normal lung parenchyma. TUNEL-positive signals were found in bronchiolar or alveolar epithelial cells in 11 of 13 (85%) specimens of CVD-IP, but not in sarcoidosis, HP or controls, except for a case of chronic HP with pulmonary fibrosis. p53, p21 and Fas were detected in bronchiolar or alveolar epithelial cells in nine (69%), 10 (77%) and 12 (92%) of 13 specimens of CVD-IP, respectively, but not in sarcoidosis, HP or controls, except for a case of chronic HP. These results suggest that the upregulation of p53, p21 and Fas in bronchiolar and alveolar epithelial cells associated with deoxyribonucleic acid damage may participate in the process of pulmonary fibrosis in interstitial pneumonia associated with collagen vascular diseases and chronic hypersensitivity pneumonitis.

Interstitial lung diseases, with or without pulmonary hypertension and epidermolysis bullosa are rare in infancy. Pathogenetic correlations between these disease are not known and their coincidence has not been reported, yet. We report on a seven weeks old boy of consanguine parents with typical skin efflorescences of epidermolysis bullosa, tachydyspnoea and cyanosis. Echocardiography and cardiac catheterisation revealed pulmonary hypertension, which persisted under therapy with oxygen and nifedipin. Lung biopsy showed interstitial and peribronchiolar increased lymphocytes and lymphfollicels, a mild intraalveolar desquamation and a media hypertrophy of the arteries. A combined therapy of prednisone and nifedipine normalised the pulmonary hypertension and the oxygen saturation. The activity of the epidermolysis bullosa showed no correlation with the interstitial lung disease or with the therapy. A connection between both diseases is discussed.

The p53 gene is well known as a tumour suppressor gene. In addition, the mutated p53 gene is detected in a variety of human cancers including lung cancer, and is considered as an oncogene. Lung cancer is also frequently associated with interstitial lung diseases. Therefore, it may be possible to hypothesize that there might be some abnormality of p53 gene in interstitial lung diseases. This work examined the relationship between the p53 protein and gene in lung tissues of 28 patients with interstitial lung diseases. Among 28 patients, 13 cases were pathologically diagnosed to have usual interstitial pneumonia (UIP), 12 cases were diagnosed as having collagen vascular lung diseases, and three cases were diagnosed to have a non-specific interstitial pneumonia. Twenty-three tissue samples were obtained by open lung biopsy and five samples were taken by autopsy. Paraffin-embedded tissues were treated by microwave, and stained with an anti-p53 antibody (DO7) by the Avidin-Biotin-Peroxidase (ABC) method. In selected patients, mutations in exons 5-8 of the p53 gene were also examined by single-strand conformation polymorphism (SSCP) analysis and DNA sequence. In addition, the presence of anti-p53 antibodies in patients' sera was screened for by ELISA. Fifteen samples (53.6%) revealed overexpression of the p53 protein in the nuclei of alveolar epithelial cells. However, SSCP or sequence analysis, which was performed in 13 tissues, showed no mutations in exons 5-8 of the p53 gene. In conclusion, p53 proteins were overexpressed in interstitial lung diseases, and the expressed p53 protein was considered to be wild-type. This wild-type p53 protein may play a role in blocking the transformation of proliferative epithelial cells.

Unlabelled: An infant with an unusual clinical presentation and course of Niemann-Pick disease type C (NPC) is described. The baby presented with severe pulmonary involvement and hepatosplenomegaly at the age of 3 months and died of respiratory failure at the age of 7 months. Cell hybridization studies revealed that the infant belonged to the rare genetic complementation group 2. To our knowledge, this is the third reported case with early-lethal pulmonary involvement in NPC. All three reported patients belong to the minor complementation group 2. An elder brother of the present case had also died at 6 weeks of age from NPC with severe pulmonary involvement. The two complementation groups cannot be distinguished from each other by clinical, cellular and biochemical criteria except for severe pulmonary involvement which may be characteristic of the second group., Conclusion: NPC may present in early infancy with severe pulmonary involvement mimicking interstitial pneumonia. Such manifestation may be characteristic of the rare genetic complementation group 2.

We analyzed the third complementarity-determining region (CDR3) of the immunoglobulin heavy chain (IgH) gene in five patients with lymphoid interstitial pneumonia (LIP) through a two-step polymerase chain reaction (PCR) and sequencing analysis. By sequencing analysis of the PCR products, morphologic LIP could be divided into two groups: a polyclonal type and a minor monoclonal type. Because of their high frequencies, minor monoclonal clones seemed to be neoplastic clones hidden in normally reactive lymphocyte clones. Consequently, only the polyclonal type might have represented true LIP. Sequencing of the PCR products from open-chest biopsy and transbronchial lung biopsy (TBLB) specimens obtained 8 yr later in one patient with minor monoclonal type LIP confirmed this possibility. In true LIP, six of 20 lymphocyte clones showed 67 to 86% homology with lymphocyte clones derived from fetal tissue. In three of these six clones, the D-region (N-D-N) lengths were very short, whereas four clones showed a high homology with autoreactive lymphocytes (rheumatoid factor, anti-DNA antibody, and G6-positive lymphocytes). Since rheumatoid factors, anti-DNA antibodies, and G6 are autoreactive antibodies, immature B cells stimulated by autoantigens might play some role in the pathogenesis of true LIP.

We have studied a 38-year-old man with a prior diagnosis of Holt-Oram syndrome, who presented with diabetes mellitus. He had recently taken prednisone for idiopathic interstitial lung disease and trimethoprim-sulfamethoxazole for sinusitis. Thrombocytopenia progressed to pancytopenia. The patient had skeletal, cardiac, renal, cutaneous, endocrine, hepatic, neurologic, and hematologic manifestations of Fanconi anemia (FA). Chest radiographs showed increased interstitial markings at age 25, dyspnea began in his late 20s, and he stopped smoking at age 32. At age 38, computerized tomography showed bilateral upper lobe fibrosis, lower lobe honeycombing, and bronchiectasis. Pulmonary function tests, compromised at age 29, showed a moderately severe obstructive and restrictive pattern by age 38. Serum alpha-1 antitrypsin level was 224 (normal 85-213) mg/dL and PI phenotype was M1. Karyotype was 46,XY with a marked increase in chromosome aberrations induced in vitro by diepoxybutane. The early onset and degree of pulmonary disease in this patient cannot be fully explained by environmental or known genetic causes. The International Fanconi Anemia Registry (IFAR) contains no example of a similar pulmonary presentation. Gene-environment (ecogenetic) interactions in FA seem evident in the final phenotype. The pathogenic mechanism of lung involvement in FA may relate to oxidative injury and cytokine anomalies.

Diffuse (interstitial) lung disease comprises a wide variety of relatively uncommon conditions, which present with characteristic clusters of clinical features and often with aberrant lung function. These diseases cause major morbidity and mortality due to lung injury and fibrosis. Some of these diseases are of known aetiology, whereas others are not. It has been suggested that the environment is a major contributing factor in this group of diseases. However, since not all individuals exposed to a common environment develop interstitial diseases, we can hypothesize that there is a genetic predisposition to their development. Therefore, if we can identify individuals who are genetically predisposed to develop diseases characterized by lung injury and fibrosis, then management strategies can be designed which will attempt to identify early disease and, in the longer term, to develop targeted genetic interventional approaches to treatment.

Diffuse (interstitial) lung disease comprises a wide variety of conditions, individually relatively uncommon but collectively being found in approximately 50 per 100,000 population. Some of these diseases are of known aetiology but others are not. It has been suggested that the environment is a major contributory factor in this group of diseases. However, since not all individuals exposed to a common environment develop interstitial diseases, it can be hypothesised that there is a genetic predisposition to their development. These diseases cause major morbidity and mortality due to lung injury and fibrosis. It follows that, if individuals who are genetically predisposed to develop diseases characterised by lung injury and fibrosis can be identified, then management strategies can be designed which will attempt to identify and treat early disease and, in the longer term, to develop targeted genetic interventional approaches to treatment.

We describe a male infant with biopsy-confirmed interstitial lung disease (ILD) who responded to chloroquine, after he failed to improve on oral corticosteroids or cyclophosphamide. The infant presented at 8 days of age with respiratory distress and cyanosis. Lung biopsy at 8 weeks of age was consistent with desquamative interstitial pneumonitis (DIP). He was treated with corticosteroids at 2 weeks of age because of a family history of two siblings who died during infancy and who had DIP on postmortem examination. At 8.5 months, our patient was treated with cyclophosphamide because of lack of response to corticosteroids therapy. At 14 months of age, he began treatment with chloroquine in addition to corticosteroids and had a dramatic response within 3 weeks. The patient has been maintained successfully on continuous treatment with chloroquine alone for more than 9 years since this treatment was started.

Etiology of idiopathic interstitial pneumonia (IIP) remains unknown. While many studies focused on the analysis of individual patients, available information is limited for the further study. Having experienced a case, 3 of 5 brothers suffered from IIP and 2 of them complicated with lung cancer, intensive familial history taking with chest photos and functional information for fibrotic lung was performed. Among 37 families in which at least 2 members were examined, 14 families (37.8%) had minimum 2 members with fibrotic lungs. Unexpectedly high familial accumulation of fibrotic lung indicates the possibility of multi-gene involvement for the pathogenesis. We need to start group studies to register sib-pairs with fibrotic lungs and systematize the automated genetic analysis for the responsible genes.

Patient 1: A 64-year-old woman was admitted for further examination after reticulonodular shadows were found on a chest X-ray film. Idiopathic interstitial pneumonia (IIP) was diagnosed. Patient 2: The 60-year-old sister of patient 1 was admitted for further examination after reticulonodular shadows were found on a chest X-ray film. IIP was diagnosed. About half a year later, her proximal interphalangeal joints had become swollen and the result of a rheumatoid hemagglutination test was positive. Therefore, the pneumonia was suspected to have been caused by a collagen-vascular disease (CVD), rheumatoid arthritis. Patient 3: The 64-year-old brother of patient 1 was examined. A chest X-ray film revealed reticulonodular shadows that were strongly suggestive of IIP. The remaining three siblings were examined. In a 62-year-old sister, the chest X-ray film was normal, but the level of anti-nuclear antigen was elevated. The fact that the level of this antigen was high in these four siblings and that the 60-year-old sister later suffered from rheumatoid arthritis suggested the presence of a factor predisposing to CVD in these siblings. The interstitial pneumonia in these siblings may have been related to CVD.

Tumor necrosis factor alpha (TNF alpha) and lymphotoxin (TNF beta) are cytokines with numerous similar immunoregulatory effects. In pulmonary fibrosis, TNF alpha/beta appears to play a critical role in the regulation of the fibroproliferative response. The human TNF alpha/beta genes are located between the HLA-B and the HLA-DR region. In man 5.5 kb and 10.5 kb of TNF alpha/beta restriction fragment length polymorphic (RFLP) bands have recently been identified by means of enzyme NcoI. TNF alpha/beta production is reported to be correlated to the NcoI RFLP. In some collagen diseases, a decreasing frequency of the 10.5 kb band is reported. The two allele NcoI RFLP of the TNF alpha region, yielding bands of 5.5 kb and 10.5 kb, was investigated in normal Japanese subjects and patients with IIP. The frequency of these bands was not different between normal subjects and patients with IIP. However, in female patients, the frequency of the 5.5 kb band was significantly increased compared with those in the male patients and normal subjects. The increase of this band may have some relation to the pathogenesis of idiopathic interstitial pneumonia.

We report on 2 sibs with desquamative interstitial pneumonia. The female died at age 1 7/12 years despite use of prednisolone and methylprednisolone, while the male, now age 3 years. is alive with oxygen support. The occurrence of desquamative interstitial pneumonia in sibs born to normal parents suggests that in some cases the disease is an autosomal-recessive trait.

We describe an immunocompetent 12-year-old boy with chronic EBV infection and lymphoid interstitial pneumonitis. Lymph node biopsies showed effacement of the architecture with polymorphic cellular infiltrates, consisting predominantly of T cells and natural killer cells. No clonal rearrangement of TCR or immunoglobulin genes was seen. DNA was extracted from hilar lymph nodes; sequencing of the carboxy terminal region of the latent membrane protein 1 (LMP-1) oncogene revealed a 69 base-pair deletion and four point mutations. Immunosuppressive treatment with prednisone and cyclosporine reversed the lymphadenopathy.

Although the pathological patterns of interstitial pneumonia associated with collagen vascular disease (CVD-IP) resemble those of usual interstitial pneumonia in idiopathic interstitial pneumonia (IIP), the clinical features of CVD-IP and IIIP are quite different. We evaluated the differences between these conditions, with regard to the expression of genes in cells obtained by bronchoalveolar lavage. The reverse transcription-polymerase chain reaction was used to measure the levels of mRNA for IL-1 beta, TNF-alpha, IL-8, TGF-beta, PDGF-B, and IGF-1, and no significant differences were found between patients with CVD-IP and those with IIP. However, differential display analysis revealed a fragment that can be considered to have been derived from an unknown gene mRNA, and this was found only in patients with pulmonary fibrosis associated with progressive systemic sclerosis. Expression of specific genes may differentiate CVD-IP from IIP.

The two main reactive pulmonary lymphoid disorders are lymphoid interstitial pneumonia and follicular bronchitis/bronchiolitis, both pathological entities with a variety of aetiologies. We reviewed the morphological and immunohistochemical features of 26 cases with one or other of these two diagnoses, to explore the possibility that they represented overlapping patterns of hyperplasia of the bronchopulmonary immune system. The polymerase chain reaction was used to determine the clonality of the infiltrates. Histologically, there was a spectrum of changes with two main components. An interstitial infiltrate of mainly T lymphocytes, plasma cells and histiocytes predominated in lymphoid interstitial pneumonia, whilst lymphoid follicles predominated around airways in follicular bronchitis/bronchiolitis. Classification of the disorder rested on which component the pathologists believed to be dominant. In two cases, histology and immunohistochemistry suggested lymphoma, and in one of these cases this diagnosis was confirmed by the polymerase chain reaction. One case of lymphoid interstitial pneumonia produced three bands. The remainder produced polyclonal patterns when samples were adequate. Clinically, there was no clear difference between patients with the two disorders, or patients with pathological features of both.

The agouti locus on mouse chromosome 2 encodes a secreted cysteine-rich protein of 131 amino acids that acts as a molecular switch to instruct the melanocyte to make either yellow pigment (phaeomelanin) or black pigment (eumelanin). Mutations that up-regulate agouti expression are dominant to those causing decreased expression and result in yellow coat color. Other associated effects are obesity, diabetes, and increased susceptibility to tumors. To try to define important functional domains of the agouti protein, we have analyzed the molecular defects present in a series of recessive viable agouti mutations. In total, six alleles (amJ, au, ada, a16H, a18H, ae) were examined at both the RNA and DNA level. Two of the alleles, a16H and ae, result from mutations in the agouti coding region. Four alleles (amJ, au, a18H, and ada) appear to represent regulatory mutations that down-regulate agouti expression. Interestingly, one of these mutations, a18H, also appears to cause an immunological defect in the homozygous condition. This immunological defect is somewhat analogous to that observed in motheaten (me) mutant mice. Short and long-range restriction enzyme analyses of homozygous a18H DNA are consistent with the hypothesis that a18H results from a paracentric inversion where one end of the inversion maps in the 5' regulatory region of agouti and the other end in or near a gene that is required for normal immunological function. Cloning the breakpoints of this putative inversion should allow us to identify the gene that confers this interesting immunological disorder.

We investigated the effects of alveolar macrophage (Am) conditioned media from interstitial lung disease (ILD) patients on lung fibroblast proliferation and procollagen mRNA expression in cells. 13 patients with ILD and 8 patients with bronchial carcinoma (BC) received bronchoalveolar lavage (BAL). The BAL fluid was collected and centrifuged to obtain cells of two groups. The cells were washed and incubated at 37 degrees C and 5% CO2 for 2 hours in DMEM with 10% FCS. Then the adherent Am was added with DMEM with 0.1% FCS. (2ml/10(6) cells), and cultured at 37 degrees C and 5% CO2 for 24 hours. The supernatant was obtained by centrifugation and stored in -20 degrees C for experiment. Lung fibroblasts were planted in 96 well microtitle plate and cultured to confluence. Each well was added 100 microliters Am conditioned medium and incubated for 16 hours, then added 5 uci 3H-TdR for measuring the fibroblast proliferation and incubated till 24 hours. The cells were collected and counted. The DMEM with 0.1% FCS was used as control. When stimulated with Am conditioned media from ILD patients, fibroblast proliferation increased 71%, but for media from BC patients, it just increased 14% (P < 0.05). After stimulated with conditioned media for 24 hours, the cells were trypzied, collected, ashed and isolated total RNA with guanine method. Several samples' total RNA was subjected to electrophoresis in 1% agaroseformaldehyde gel to test specificity of procollagen al (I) and al (III) cDNA probes. 10 micrograms total RNA of each sample was bloted to nitrit membrane and baked at 80 degrees C for 2 hours.

The presence of the HTLV-I gene in peripheral blood mononuclear cells was studied by polymerase chain reaction in 42 patients including 16 with lung cancer, 12 with diffuse panbronchiolitis (DPB), 11 with idiopathic interstitial pneumonia (IIP), and 3 with pneumoconiosis and hematological malignancy. Sequences equal to a part of the pX gene were found in 44% of the lung cancer cases, 50% of the DPB cases, 55% of the IIP cases, and 100% of the cases of pneumoconiosis and leukemia. In the lung cancer cases, detection of the pX gene was frequently associated with the existence of diffuse interstitial pulmonary shadows. The pX gene was detected in 100% of patients with anti-HTLV-I antibody, 50% of patients with HTLV-I-related reaction and 14% of patients who tested seronegative. It may be inferred from the results that respiratory diseases that produce diffuse interstitial pulmonary shadows are closely associated with HTLV-I infection and that the HTLV-I-related reaction to the immunofluorescent test might reflect the latent infection state of HTLV-I.

Idiopathic interstitial pneumonia (IIP), which is referred to as "idiopathic pulmonary fibrosis (IPF)" in US and European countries or "cryptogenic fibrosing alveolitis (CFA)" in UK, is considered to result from an uncontrolled, chronic inflammatory process in the lower respiratory tract and alveoli of the lung. It is conceivable that some hereditary background may contribute to the disease in susceptible individuals. In IPF, alveolitis involving activation of alveolar macrophages (AM), neutrophils, T- and B-lymphocytes and/or other inflammatory cell types precedes the injury to the lung parenchyma and interstitial fibrosis. Activation and accumulation of cellular components in the disease seem to be modulated by a variety of biological substances such as cytokines, chemotactic factors and adhesion molecules. Exposure to inorganic dusts or viral infection also may trigger activation of AM. The injury to the alveolar epithelium, capillary endothelial cells and extracellular matrix mediated by oxidants and/or proteases produced by AM and neutrophils results in the derangement of the lung parenchyma. Subsequent fibrotic change of the lung interstitium is characterized by the accumulation of mesenchymal cells such as fibroblasts and deposition of their connective tissue produces within the alveolar walls. In this process, AM is considered to play a major role by secreting growth factors for mesenchymal cells, although alveolar epithelial and endothelial cells also seem to participate in activating mesenchymal cell proliferation in IPF. Possibly, connective tissue synthesis and/or degradation may be altered in the process of IPF, although there is no direct evidence for this so far. Thus, IPF is a chronic inflammatory disease in which inflammatory processes of the alveoli injure the lung parenchyma and modulate the proliferation of mesenchymal cells which result in the fibrotic change. The pathogenesis of the disease involves multiple cellular components of the lower respiratory tract through modulation of gene expression and protein synthesis of a variety of biological factors.

To clarify the pathogenesis of idiopathic interstitial pneumonia (IIP), we compared the data of interstitial pneumonia (IP) associated with collagen vascular disease (CVD) with those of idiopathic pulmonary fibrosis (IPF). These disorders are found to have similar clinical, radiographic, and morphological findings, and lung function. However, the in vitro behavior of fibroblasts from CVD-IP and IPF, and the detection of soluble ICAM-1 were different. Further, in the T cell receptor repertoire, the predominance of different genes was found in bronchoalveolar lavage cells from CVD and IPF. These differences could partly explain the variability of IP pattern seen in CVD. In summary, our results indicate the possibility that CVD-IF and IPF result from different mechanisms although they share the same spectrum of pulmonary changes.

We have studied the gene expression of ten growth factors in bronchoalveolar lavage (BAL) cells from patients with lung fibrosis by using a reverse-transcription-DNA polymerase chain reaction (RT-PCR) technique. IL-1 beta mRNA was detected in almost all of the samples, and TGF beta and IGF-I mRNA were detected in some. The BAL supernatant from the patients was found to have mitogenic activity for lung fibroblasts. Moreover, human recombinant IL-1 beta, TGF beta and IGF-I were found to promote the proliferation of lung fibroblasts. These data suggest that IL-1 beta, TGF beta and IGF-I gene expression in BAL cells might be involved in the development of lung fibrosis.