Vascular endothelial growth factor mRNA and protein expression in airway epithelial cell lines in vitro.

Vascular endothelial growth factor (VEGF) plays multifunctional roles in vascular permeability, repair and remodelling processes, in addition to the maintenance of vascular structure and function. In the present study, the potential of airway epithelial cell lines, BEAS-2B cells and A549 cells, to release and express VEGF in unstimulated and stimulated conditions was evaluated. The secretion and expression of VEGF were evaluated by enzyme-linked immunosorbant assay and by reverse transcriptase-polymerase chain reaction. The isoforms of released VEGF were determined by high-performance liquid chromatography. BEAS-2B cells and A549 cells released VEGF constitutively. Interleukin (IL)-1beta and tumour necrosis factor (TNF)-alpha augmented the release of VEGF in a time- and dose-dependent manner. The released VEGF was 165 amino acid residues in either condition. Pseudomonas aeruginosa lipopolysaccharide (LPS), interferon (IFN)-gamma, smoke extract (SE), neutrophil elastase (NE), and bradykinin stimulated the release of VEGF. Keracinocyte growth factor (KGF), which reduces vascular permeability, also stimulated both cells to release VEGF. VEGF messenger ribonucleic acid (mRNA) was expressed both time- and dose-dependently at 2 h, and declined after 2 h in response to IL-1beta and TNF-alpha. The expression of VEGF mRNA in airway epithelial cells was also augmented by LPS, IFN-gamma, SE, NE, and KGF stimulation. These data suggest that airway epithelial cells may regulate the maintenance of vascular structure and function, as well as vascular permeability, repair and remodelling processes, in a variety of lung conditions by expressing vascular endothelial growth factor.

Theophylline inhibits TNF-alpha-induced CD4 expression on human eosinophils and CD4+ eosinophil migration.

BACKGROUND: Increasing evidence regarding asthma suggests that CD4+ cells are preferentially recruited to sites of bronchial inflammation. Interleukin (IL)-16 has been reported as playing an important role in the accumulation of CD4+ cells. We have shown that the CD4 molecule is expressed on normal human eosinophils by tumor necrosis factor (TNF)-alpha stimulation. METHODS: We evaluated the effects of theophylline, KF19514 [a selective phosphodiesterase (PDE) IV inhibitor] and dexamethasone on CD4 expression on eosinophils and eosinophil migration in response to IL-16, a natural soluble ligand of the CD4 molecule. RESULTS: The maximum eosinophil migration was observed when eosinophils were cultured with TNF-alpha at 10 ng/ml for 18 h and the concentration of IL-16 was 10 pg/ml. CD4+ eosinophil migration in response to IL-16 was mostly, if not fully, chemokinetic and this migration was significantly inhibited by Fab of anti-CD4 monoclonal antibody. Theophylline (10(-4)-10(-3) M), KF19514 (10(-7)-10(-6) M) and dexamethasone (10(-8)- 10(-6) M) significantly inhibited CD4 expression on eosinophils induced by TNF-alpha. Theophylline (10(-3) M) and KF19514 (10(-6) M) inhibited CD4+ eosinophil migratory responses induced by IL-16, but 10(-6) M dexamethasone did not. Theophylline and KF19514 augmented the intracellular adenosine-3',5'-cyclic monophosphate (cAMP) concentration in eosinophils, suggesting modulation by cAMP of CD4 expression and eosinophil migration. CONCLUSIONS: These data suggest that TNF-alpha-induced CD4+ eosinophils may contribute to eosinophil migratory responses induced by IL-16. Theophylline and selective PDE IV inhibitor may prevent airway inflammation by downregulating CD4 expression on eosinophils and inhibiting eosinophil migration through CD4 and IL-16 interaction.

[Two cases of pulmonary disease with perinuclear anti-neutrophil cytoplasmic antibody].

We encountered two cases of perinuclear anti-neutrophil cytoplasmic antibody (p-ANCA). The first was a case of idiopathic interstitial pneumonia diagnosed in a 73-year-old man since 1998. He was admitted to our hospital because of renal failure and anemia. The serum level of p-ANCA on admission was 264 EU, and specimens obtained by percutaneous renal biopsy showed crescentic glomerulonephritis and vasculitis due to p-ANCA. He was treated with prednisolone pulse therapy and prednisolone (PSL), however interstitial pneumonia occurred during PSL tapering. We treated him for pulmonary fibrosis with plasmapheresis, methylprednisolone (mPSL) and cyclophosphamide (CPA), which suppressed the progress of the interstitial pneumonia. The second case was one of massive pulmonary hemorrhage in a 68-year-old man who was admitted to our hospital. Physical examination revealed anemia: the laboratory data, renal failure; and the serum level of p-ANCA was elevated to 611 EU. The specimens obtained by percutaneous renal biopsy showed crescentic glomerulonephritis and vasculitis. The renal failure was not improved by PSL, but, together with the inflammation, responded to the combination of PSL and CPA. However, both patients died of serious infection. They were regarded as compromised patients because of the therapy mentioned above. No standard therapy has been established against p-ANCA positive pulmonary disease with renal failure. The treatment should control the progression of interstitial pneumonitis and pulmonary hemorrhage. It is important to consider the possibility of serious infection.

Chemotaxis of human CD4+ eosinophils.

We reviewed the biological functions of CD4+ eosinophils, which are observed in peripheral blood, sputum and bronchoalveolar lavage fluid of various diseases. We have shown that CD4 molecules on human eosinophils are induced by tumor necrosis factor-alpha (TNF-alpha) stimulation. Interleukin-16 (IL-16) has been reported to bind a natural soluble ligand for the CD4 molecule. We reported that TNF-alpha-stimulated eosinophils migrate in a time- and dose-dependent manner against IL-16. Theophylline and dexamethasone significantly inhibited CD4 expression. Theophylline inhibited CD4+ eosinophil chemotaxis, but dexamethasone did not. Theophylline may prevent airway inflammation by downregulating the expression of CD4 molecule and CD4+ eosinophil migration. However, dexamethasone may inhibit airway inflammation through the downregulation of CD4 expression.

Concurrent left-sided spontaneous pneumothorax in Japanese monogerminal twins.

Monogerminal twin brothers aged 17 were admitted because of concurrent left-sided spontaneous pneumothorax. A familial background of this common disease in association with the human leukocyte antigen (HLA) has been suggested; however, the actual mode of inheritance in association with HLA typing is still uncertain. HLA analysis in this family revealed the HLA-A24, B61 (40), Cw3, DR4, DR53, and DQ3 haplotype in the twins and their father, who also had a medical history of spontaneous pneumothorax. We report these cases as an extremely rare example of familial occurrence of spontaneous pneumothorax.

[A case of renal cell carcinoma complicated with interstitial pneumonitis, complete A-V block and pleural effusion during interferon-alpha therapy].

A 76-year-old man with postoperative renal cell carcinoma accompanied by multiple lung metastasis was being treated with recombinant interferon-alpha. After administration of 3 MU/day on 3 days/week for 1 month, he complained of headache and tinnitus. During continuous treatment for 3 months, he complained of appetite loss, low-grade fever and dyspnea. He was then referred to our Department of Internal Medicine. Electrocardiography indicated a complete A-V block, and chest radiography (CXR) showed a reticular shadow in both lower lung fields and bilateral pleural effusion. Chest computed tomography (CT) indicated subpleural emphysematous changes, multiple nodules, consolidation shadow with ground glass opacity in both lower lobes, and bilateral pleural effusion. The findings in the bronchoalveolar lavage (BAL) fluid included increases in the numbers of lymphocytes and eosinophils. We reached a diagnosis of interferon-alpha-induced pneumonitis on the basis of the patient's clinical course, and the CXR, chest CT and BAL fluid findings. Treatment with methylprednisolone pulse therapy for 3 days and then administration of prednisolone for 1 month resulted in marked improvement in the complete A-V block and interstitial pneumonitis. At day 7 after discontinuation of prednisolone, the serum level of C-reactive protein increased, and CXR showed bilateral pleural effusion. We therefore believe that the pleural effusion was probably also induced by interferon-alpha. Interferon is an effective drug for chronic hepatitis C and malignant diseases. Many complications have been reported during interferon therapy. However, although these complications, such as interstitial pneumonitis, complete A-V block and pleural effusion, have rarely been reported, careful attention is required during interferon therapy in case any appear.

[A case of intrathoracic schwannoma derived from the left vagus nerve].

We report a case of schwannoma asymptomatic for 9 years, derived from the left vagus nerve in the middle mediastinum. This spindle-shaped tumor caused paralysis of the left recurrent nerve as an initial clinical manifestation with cough. T2-weighted magnetic resonance imaging (MRI) showed a neurogenic tumor with a characteristic target appearance and with constituents of different intensities: mucinous material in the peripheral zone and solitary tissue in the central zone. But, this different intensity is not directly reflected by the histopathologic features of Antoni types A and B. These findings suggest that MRI is useful for determining the parent nerve of a neurogenic tumor and is helpful for the diagnosis.

Spontaneous B-cell IgE production in a patient with remarkable eosinophilia and hyper IgE.

BACKGROUND: The pathophysiology of eosinophilia and hyper-IgE is not fully elucidated yet. OBJECTIVE: To clarify the pathophysiology of a patient with remarkable eosinophilia and hyper IgE, we examined cytokine levels in serum, surface antigens of peripheral blood eosinophils and IgE production in vitro. RESULTS: Concentrations of tumor necrosis factor-alpha (TNF-alpha), interleukin-3 (IL-3), interleukin-4 (IL-4), interleukin-5 (IL-5), and granulocyte/macrophage-colony stimulating factor (GM-CSF) in the serum were 21 pg/mL, <15 pg/mL, <15 pg/mL, 8 pg/mL, and <5 pg/mL pg/mL, respectively. Newly expressed surface antigens CD4, CD25, CD69, and HLA-DR, but not CD54, were observed on peripheral blood eosinophils. Extremely high levels of IgE secretion was found in the patient's mononuclear cells without stimuli; this was not enhanced by IL-4 or IL-4 plus anti-CD40 monoclonal antibody stimulation. Furthermore, highly purified B cells spontaneously produced large amounts of IgE and the production was not enhanced in addition of his T cells. CONCLUSION: The eosinophils were activated, and the B cells spontaneously produced IgE independently of T cells or cytokines, suggesting that intrinsic abnormality of B cells leading to dysregulated production of IgE in this disease.

[A case of limited Wegener granulomatosis with hypereosinophilia].

A 51-year-old female was admitted to Nagano Matsushiro General Hospital because of fever, cough and dyspnea on exertion. Her laboratory data revealed leukocytosis with hypereosinophilia, a high erythrocyte sedimentation rate and c-reactive protein. Chest radiography revealed an infiltration shadow with a cavity in the right upper lobe. A lung abscess was diagnosed and antibiotics were administered. Laboratory results showed improvement, but chest radiography continued to show cavities. She was admitted to our hospital because of fever, left pleural effusion and progression of cavities on chest radiographs. She showed no abnormalities of the upper airway or kidney, and was negative for c-antineutrophil cytoplasmic antibody (c-ANCA). Because a positive c-ANCA was seen on day 8 of hospitalization, L-type limited Wegener granulomatosis (WG) was diagnosed according to Gross et al. Prednisolone (PSL) was administered, which improved the anemia, eosinophilia and the cavities. On day 7 of PSL administration, of the left pneumothorax occurred as a complication caused by perforation of the left chest cavity, but her clinical course was good after a cavernectomy was performed. Some studies have reported that limited WG shows a negative c-ANCA, and that antibiotic therapy improves inflammation. The L-type of limited WG revealed a low-grade positive ratio and titer of c-ANCA. Moreover, L-type limited WG responds well to therapy. We therefore selected PSL administration only against L-type limited WG. We have reported L-type limited WG with eosinophilia and the negative effects of c-ANCA at an early clinical stage.

Interferon-gamma increases CD62L expression on human eosinophils.

BACKGROUND: L-selectin (CD62L) and Mac-1 (CD11b/CD18) play a crucial role in the infiltration of eosinophils. However, changes in CD62L and CD11b expression on eosinophils after stimulation with cytokines were little studied. METHODS: Eosinophils in peripheral blood of healthy volunteers were purified and cultured with interleukin-5 (IL-5), granulocyte/macrophage colony-stimulating factor (GM-CSF) or interferon-gamma (IFN-gamma). After up to 24 h incubation, CD62L and CD11b expression were examined using flow cytometry. The effects of dexamethasone (Dex), cycloheximide (CHX) or theophylline on CD62L expression on IFN-gamma-stimulated eosinophils were also studied. RESULTS: IL-5 or GM-CSF downregulated CD62L and upregulated CD11b expression on eosinophils after 30 min stimulation. Conversely, IFN-gamma upregulated CD62L after 12 h stimulation in a time- and dose-dependent manner, and had no effect on CD11b expression. Dex, CHX or theophylline dose-dependently decreased CD62L expression on IFN-gamma-stimulated eosinophils. CONCLUSIONS: IFN-gamma is a particular cytokine which can increase CD62L expression on circulating or infiltrated human eosinophils. It is suggested that protein synthesis and intracellular cAMP participate in the increase in L-selectin expression on IFN-gamma-stimulated eosinophils.

Eosinophil active cytokines and surface analysis of eosinophils in Churg-Strauss syndrome.

There are few reports regarding the measurement of cytokines and surface analysis of eosinophils in Churg-Strauss syndrome (CSS). To examine the pathophysiology of CSS, concentrations of cytokines in serum and bronchoalveolar lavage fluid (BALF), and surface antigens on peripheral blood eosinophils were analyzed in five patients with CSS. Concentrations of cytokines (interleukin-1 beta (IL-1 beta), tumor necrosis factor-alpha (TNF-alpha), interleukin-3 (IL-3), interleukin-5 (IL-5) and granulocyte/macrophage colony stimulating factor (GM-CSF) were measured using ELISA. Surface antigens on eosinophils in peripheral blood were analyzed using flow cytometry. A concentration of interleukin-5 (IL-5) and TNF-alpha in serum was detected in five cases; however IL-1 beta, GM-CSF, and IL-3 were detected in 3 of 5, 2 of 5, and 1 of 5 patients, respectively. In BALF, TNF-alpha and IL-5 were detected in 2 of 3 and 1 of 3 patients, respectively; however, neither IL-1 beta, GM-CSF, nor IL-3 was detected in any. Newly expressed surface antigens such as CD25, CD4, and CD69 were observed on peripheral blood eosinophils in five cases. CD54 and HLA-DR were expressed in 4 of 5 and 3 of 5 patients, respectively. Eosinophils in peripheral blood are activated to various degrees, possibly depending on cytokine stimulation. This eosinophil activation may be related to the clinical stage of CSS.

[ADOC regimen for unresectable advanced thymic cancer].

Between 1996 and 1998, we treated 6 patients with unresectable and advanced thymic cancer (stages IVa and IVb). All received 50 mg/m2 of cisplatin and 40 mg/m2 of doxorubicin intravenously (i.v.) on day 1,0.6 mg/m2 of vincristine i.v. on day 3, and 700 mg/m2 of cyclophosphamide i.v. on day 4; ADOC regimen, respectively at 3-4 week intervals. Four patients obtained a partial response (PR) after ADOC chemotherapy and the overall clinical response rate was 67%. No life-threatening side effects were noted. In 2 patients, cisplatin plus VP-16 chemotherapy failed to demonstrate any benefits prior to the ADOC regimen. Radiotherapy was initiated after the achievement of PR in the other 2 patients. ADOC chemotherapy appears to be an effective treatment for thymic cancer.

A case of pathophysiologic study in Kimura's disease: measurement of cytokines and surface analysis of eosinophils.

BACKGROUND: Kimura's disease is a rare but distinctive eosinophilic inflammatory disorder of unknown etiology; few reported case studies have focused on the immunopathologic background of this unique disease. OBJECTIVE: To define better the immunopathogenetic features of Kimura's disease, we attempted to quantitatively analyze values of cytokines and soluble interleukin-2 receptor (sIL-2R) in peripheral blood (PB), as well as perform surface immunophenotypic analysis of eosinophils from a Japanese patient with chronic relapsing Kimura's disease. RESULTS: Granulocyte macrophage-colony stimulating factor (GM-CSF), tumor necrosis factor-alpha (TNF-alpha) and sIL-2R were elevated, and newly expressed antigens on eosinophils CD4, CD25, and HLA-DR were found to be involved in the pathophysiology of this disorder. CONCLUSIONS: Kimura's disease may be a disease in which activated lymphocytes release cytokines, and these released cytokines, such as GM-CSF and TNF-alpha cause eosinophil activation. These processes may be related to the pathogenesis of this disorder.

Predominant implication of IL-5 in acute eosinophilic pneumonia: comparison with chronic eosinophilic pneumonia.

BACKGROUND: Acute eosinophilic pneumonia (AEP) is a rare disease with unknown etiology. To examine pathophysiology of AEP we measured the cell number of eosinophils and eosinophil active cytokines in the peripheral blood and bronchoalveolar lavage fluid (BALF) of AEP patients and compared the levels with those measured in chronic eosinophilic pneumonia (CEP) patients. METHODS: Cell number of eosinophils in peripheral blood and BALF from patients with AEP (n = 3) and CEP (n = 3) were measured. Eosinophil active cytokines in serum and BALF from the patients were measured using ELISA. RESULTS: Eosinophil cell number in peripheral blood was 274-1,377/mm3 in AEP and 526-2,500/mm3 in CEP. The percentages of BALF eosinophils were high in AEP and CEP. Eosinophilia disappeared after methylprednisolone pulse therapy (1 g for 3 days) in AEP, however the cell number of eosinophils gradually increased after methylprednisolone pulse therapy and then spontaneously decreased to within normal range without any further medication. The concentrations of IL-5 in AEP were very high in serum and in BALF, however the concentrations in CEP were low in serum and BALF. CONCLUSION: AEP is a disease in which eosinophil active cytokine IL-5 is predominantly involved; CEP is not. The factors involving eosinophil infiltration to inflammatory loci differ between AEP and CEP.