The association of transforming growth factor beta 1 gene polymorphisms with the emphysema phenotype of COPD in Japanese.

OBJECTIVE: The transforming growth factor beta-1 gene (TGFB1) is one of the most promising candidate genes for chronic obstructive pulmonary disease (COPD). Several case-control studies have been performed and generated inconsistent results. The possible reasons for these discrepancies include the diversity of ethnic populations and the heterogeneity of COPD, including emphysema and airway disease. We designed this study to investigate the association of single nucleotide polymorphisms (SNPs) of TGFB1 with the emphysema phenotype in the Japanese population. METHODS: Eight SNPs in TGFB1 (rs2241712, rs1982072, and rs1800469 in the promoter region; rs1982073 in exon 1; rs2241716 and rs4803455 in intron 2; rs6957 and rs2241718 in the 3' region) were genotyped by allelic discrimination assays in 70 COPD patients with emphysema phenotype and 99 healthy smokers. The emphysema phenotype was identified by high-resolution computed tomography imaging using Goddard's method. RESULTS: The frequency of one significant haplotype structured by the eight SNPs was significantly higher in the emphysema group (10%) than in the healthy smokers (4%, p=0.02). In the emphysema group, the predicted value of forced expiratory volume in 1 second after bronchodilator administration was significantly associated with the minor alleles of the two SNPs (rs1800469 and rs1982073, p=0.007 and 0.032, respectively), however, the low attenuation area and carbon monoxide diffusing capacity were not associated with the SNPs. In addition, the rs1800469T and rs1982073C alleles were significantly more prevalent in patients with severe and very severe airflow limitation than in those with mild and moderate airflow limitation (p=0.007 and 0.041, respectively). CONCLUSIONS: One significant haplotype of TGFB1 is associated with the emphysema phenotype in the Japanese population. Two TGFB1 SNPs (rs1800469 and rs1982073) are associated with the severity of COPD in patients with emphysema phenotype.

Effects of expiratory pressure on nitric oxide in exhaled breath. Is exhaled nitric oxide really unaffected by pressure?

The measurements of exhaled nitric oxide (ENO) concentrations in several previous reports have been quite disparate but the cause of this variability is unclear. In the present study, we have attempted to elucidate the effects of expiratory pressure upon ENO values by taking measurements at pressures ranging from 2 to 10 cmH2O in control subjects and in both smokers and asthmatics. Differences in ENO concentrations (delta pNO) were then estimated and the concentration levels were found to increase with elevated expiratory pressure levels in both the control volunteers and in the asthmatics (under 2 and 3 L/min flow rates). These results indicate that changes in expiratory pressure indeed affect ENO concentrations. The measurement of ENO concentrations in human patients must therefore be undertaken using standard procedures that must incorporate expiratory pressure levels in order to properly interpret ENO values.

Serine protease inhibitors modulate smoke-induced chemokine release from human lung fibroblasts.

Smoking is associated with lung inflammation and a protease-antiprotease imbalance. We previously reported that cigarette smoke extract (CSE) stimulates human lung fibroblasts to release chemotactic cytokines. We hypothesized that serine protease inhibitors might modulate lung fibroblast release of chemotactic cytokines in response to CSE. To test this hypothesis, serine protease inhibitors (FK706, alpha1-antitrypsin, methoxysuccinyl-Ala-Ala-Pro-Val chloromethyl ketone, or Nalpha-p-tosyl-L-lysine chloromethyl ketone) were evaluated for their capacity to attenuate the release of neutrophil chemotactic activity (NCA) and monocyte chemotactic activity (MCA) from human fetal lung fibroblasts by the blind-well chemotactic chamber. Metalloproteinases and cysteine proteinases were not examined in this study. Similarly, the release and gene expression of chemokines and nuclear factor-kappaB (NF-kappaB) activation were measured by means of enzyme-linked immunosorbent assay and reverse transcriptase-polymerase chain reaction. Release of NCA, MCA, chemotactic chemokines including interleukin-8, granulocyte colony-stimulating factor, monocyte chemoattractant protein-1, and granulocyte-macrophage colony-stimulating factor, and the expression of interleukin-8 and monocyte chemoattractant protein-1 mRNA were attenuated by FK706. Furthermore, FK706 suppressed NF-kappaB activation. These data suggest that serine protease inhibitors attenuate the CSE-induced release of NCA and MCA from human fetal lung fibroblasts and that the inhibitory action of antiproteases might depend on NF-kappaB signaling pathway.

Methotrexate stimulates lung epithelial cells to release inflammatory cell chemotactic activities.

Methotrexate-induced pneumonitis has been reported as an infrequent but potentially serious complication of therapy in a variety of malignant and benign conditions. Because inflammatory cell infiltration is concerned with the development of methotrexate-induced pneumoinitis, and because airway epithelial cells participate in the orchestration of lung inflammation, the authors determined whether methotrexate might stimulate airway epithelial cells (A549 cells) to release neutrophil, monocyte, and eosinophil chemotactic activities (NCA, MCA, and ECA). A549 cells released NCA, MCA, and ECA in a dose- and time-dependent manner in response to methotrexate. Partial characterization revealed the heterogeneity of NCA, MCA, and ECA. The release of chemotactic activity was blocked by lipoxygenase inhibitors and cycloheximide. NCA was inhibited by leukotriene (LT) B(4) receptor antagonist, and anti-interleukin (IL)-8 and granulocyte colony-stimulating factor (G-CSF) antibodies. MCA was attenuated by LTB(4) receptor antagonist, and anti-monocyte chemoattractant protein (MCP)-1 and granulocyte-macrophage CSF (GM-CSF) antibodies. ECA was attenuated by LTB(4) receptor antagonist, and anti-IL-8 and GM-CSF antibodies. The release of IL-8, G-CSF, MCP-1, GM-CSF, and LTB(4) from A549 cells significantly increased in response to methotrexate. The mRNA expression of IL-8 and MCP-1 was augmented by methotrexate stimulation. These data suggest that type II epithelial cells may modulate inflammatory cell recruitment into the lung by releasing NCA, MCA, and ECA in response to methotrexate.

Decreased level of vascular endothelial growth factor in bronchoalveolar lavage fluid of normal smokers and patients with pulmonary fibrosis.

Vascular endothelial growth factor (VEGF) plays multifunctional roles in both the development of vasculature and the maintenance of vascular function. A decrease in VEGF reduces angiogenesis and induces apoptosis of vascular endothelial cells. Inhibition of the VEGF receptor causes endothelial cell apoptosis and emphysema. We postulated that VEGF concentrations might be reduced in patients with chronic lung diseases. The level of VEGF was evaluated by enzyme-liked immunosorbent assay in bronchoalveolar lavage fluid (BALF) from normal smokers, nonsmoking volunteers, idiopathic pulmonary fibrosis, pulmonary fibrosis associated with a connective tissue disease, and sarcoidosis. The isoforms of VEGF in BALF were determined by high-performance liquid chromatography. VEGF in nonsmoking volunteers was detectable at a high concentration. In contrast, VEGF in most of the normal smokers was below the detectable limit. The VEGF found in nonsmoking volunteers BALF was VEGF165. VEGF was significantly decreased in idiopathic pulmonary fibrosis, pulmonary fibrosis associated with a connective tissue disease, and sarcoidosis compared with nonsmoking volunteers. The smoking patients showed a further decrease in VEGF. These data suggest that the decrease in VEGF in smokers and patients with chronic lung diseases may reduce angiogenesis and induce apoptosis of vascular endothelial cells.

Histamine and serotonin stimulate eotaxin production by a human lung fibroblast cell line.

Histamine and serotonin are important inflammatory mediators in the pathophysiology of asthma, and asthmatic patients have higher plasma histamine and serotonin levels than nonasthmatic control subjects. Eotaxin, a potent eosinophil-specific chemotactic factor, is increased in the lower respiratory tract of allergic patients. Recently, lung fibroblasts have been reported to produce eotaxin and are suggested to be the major cellular source of eotaxin. We postulated that lung fibroblasts might release eotaxin in response to histamine or serotonin. To test this hypothesis, we evaluated the potential of histamine or serotonin to induce the release of eotaxin by the human fetal lung fibroblast cell line, HFL-1. HFL-1 released eotaxin in response to histamine and serotonin in a dose- and time-dependent manner (p < 0.05). Histamine or serotonin treatment of HFL-1 augmented the expression of eotaxin mRNA. Eosinophil chemotactic activity by HFL-1 supernatant fluids was inhibited by anti-human eotaxin-neutralizing antibody. These findings lead to the hypothesis that lung-fibroblast-derived eotaxin may in part be responsible for the eosinophil infiltration observed in allergic disease of the airways.

Human bronchial epithelium expresses interleukin-9 receptors and releases neutrophil chemotactic factor.

Growing evidence obtained from human genomic analysis and antigen-challenged transgenic mice suggests that interleukin-9 (IL-9) is a candidate factor in immunoglobulin E (IgE) production and thus is thought to be associated with bronchial inflammation and bronchial hyperresponsiveness (BHR). To evaluate the expression of the IL-9 receptor and its effect on the IL-9 human bronchial cell line BEAS-2B cells, reverse transcriptase-polymerase chain reaction (RT-PCR), immunohistochemical investigation, and chemotaxis assay were performed. The components of the IL-9 receptor, consisting of IL-9 receptor alpha (CD129) and IL-2 receptory ((1)132), were expressed on BEAS-2B cells as determined by RT-PCR and flow cytometry. BEAS-2B cells exposed to IL-9 released neutrophil chemotactic activity (NCA) in a time- and dose-dependent manner, and the presence of granulocyte colony-stimulating factor (G-CSF) was also detected. This factor is primarily involved in NCA for the measurement of cytokines and in the inhibition assay of neutrophil chemotaxis. These findings suggest that bronchial epithelial cells may express IL-9 receptors, and that IL-9 may induce airway inflammation through the release of G-CSF from bronchial epithelial cells.

Hemodynamic changes during hemodialysis: role of nitric oxide and endothelin.

BACKGROUND: Etiology of dialysis induced hypotension and hypertension remains speculative. There is mounting evidence that nitric oxide (NO) and endothelin (ET-1) may play a vital role in these hemodynamic changes. We examined the intradialytic dynamic changes in NO and ET-1 levels and their role in the pathogenesis of hypotension and rebound hypertension during hemodialysis (HD). METHODS: The serum nitrate + nitrite (NT), fractional exhaled NO concentration (FENO), L-arginine (L-Arg), NGNG-dimethyl-L-arginine (ADMA) and endothelin (ET-1) profiles were studied in 27 end-stage renal disease (ESRD) patients on HD and 6 matched controls. The ESRD patients were grouped according to their hemodynamic profile; Group I patients had stable BP throughout HD, Group II had dialysis-induced hypotension, and Group III had intradialytic rebound hypertension. RESULTS: Pre-dialysis FENO was significantly lower in the dialysis patients compared to controls (19.3 +/- 6.3 vs. 28.6 +/- 3.4 ppb, P < 0.002). Between the experimental groups, pre-dialysis FENO was significantly higher in Group II (24.1 +/- 6.7 ppb) compared to Group I (17.8 +/- 5.6 ppb) and Group III (16.1 +/- 4.2 ppb; P < 0.05). Post-dialysis, FENO increased significantly from the pre-dialysis values (19.3 +/- 6.3 vs. 22.6 +/- 7.9 ppb; P=0.001). Pre-dialysis NT (34.4 +/- 28.2 micromol/L/L) level was not significantly different from that of controls (30.2 +/- 12.3 micromol/L/L). Serum NT decreased from 34.4 +/- 28.2 micromol/L/L at initiation of dialysis to 10.0 +/- 7.4 micormol/L/L at end of dialysis (P < 0.001). NT concentration was comparable in all the three groups at all time points. Pre-dialysis L-Arg (105.3 +/- 25.2 vs. 93.7 +/- 6.0 micromol/L/L; P < 0.05) and ADMA levels were significantly higher in ESRD patients (4.0 +/- 1.8 vs. 0.9 +/- 0.2 micromol/L/L; P < 0.001) compared to controls. Dialysis resulted in significant reduction in L-Arg (105.3 +/- 25.2 vs. 86.8 +/- 19.8 micromol/L/L; P < 0.005) and ADMA (4.0 +/- 1.8 vs. 1.6 +/- 0.7 micromol/L/L; P < 0.001) concentrations. Pre-dialysis ET-1 levels were significantly higher in ESRD patients compared to the controls (8.0 +/- 1.9 vs. 12.7 +/- 4.1 pg/mL; P < 0.002), but were comparable in the three study groups. Post-dialysis ET-1 levels did not change significantly in Group I compared to pre-dialysis values (14.3 +/- 4.3 vs.15.0 +/- 2.4 pg/mL, P=NS). However, while the ET-1 concentration decreased significantly in Group II (12.0 +/- 4.0 vs. 8.7 +/- 1.8 pg/mL, P < 0.05), it increased in Group III from pre-dialysis levels (12.8 +/- 3.8 vs. 16.7 +/- 4.5 pg/mL, P=0.06). CONCLUSION: Pre-dialysis FENO is elevated in patients with dialysis-induced hypotension and may be a more reliable than NT as a marker for endogenous NO activity in dialysis patients. Altered NO/ET-1 balance may be involved in the pathogenesis of rebound hypertension and hypotension during dialysis.