Assessment of exhaled carbon monoxide in exacerbations of chronic obstructive pulmonary disease.

Introduction Exhaled carbon monoxide (eCO) has been widely implicated as a pulmonary biomarker in respiratory diseases. The aim of this study was to investigate whether the treatment of patients with severe acute exacerbation of chronic obstructive pulmonary disease (AECOPD) could be aided by monitoring the changes in eCO. Methods The levels of eCO along with routine clinical parameters were analyzed in 29 current smoker and 33 ex-smoker COPD patients, first at the time of hospital admission, and again at discharge following the standard treatment. Patients with AECOPD were also stratified according to sputum bacteria. Results At exacerbation, the levels of eCO were increased in current smokers compared to ex-smokers (6.0 [2.0-9.5] versus 1.0 [1.0-2.0] ppm, p < 0.001). Similarly, eCO levels were higher in smokers after treatment (7.0 [2.0-12.5] versus 1.0 [1.0-2.0] ppm, p < 0.001). Treatment of AECOPD did not affect eCO concentrations. The levels of eCO were not statistically different between bacterial and non-bacterial AECOPD either. Investigating a subgroup of current smoker patients (n = 15), there was a significant correlation between the levels of eCO and blood carboxyhemoglobin concentrations both at exacerbation and discharge. No associations were found between eCO and lung function or blood gas parameters. Conclusion Our results suggest that monitoring eCO during the treatment of AECOPD is of limited clinical value.

Exercise changes volatiles in exhaled breath assessed by an electronic nose.

Exercise-caused metabolic changes can be followed by monitoring exhaled volatiles; however it has not been previously reported if a spectrum of exhaled gases is modified after physical challenge. We have hypothesized that changes in volatile molecules assessed by an electronic nose may be the reason for the alkalization of the exhaled breath condensate (EBC) fluid following physical exercise.Ten healthy young subjects performed a 6-minute running test. Exhaled breath samples pre-exercise and post-exercise (0 min, 15 min, 30 min and 60 min) were collected for volatile pattern ("smellprint") determination and pH measurements (at 5.33 kPa CO2), respectively. Exhaled breath smellprints were analyzed using principal component analysis and were related to EBC pH.Smellprints (p=0.04) and EBC pH (p=0.01) were altered during exercise challenge. Compared to pre-exercise values, smellprints and pH differed at 15 min, 30 min and 60 min following exercise (p<0.05), while no difference was found at 0 min post-exercise. In addition, a significant correlation was found between volatile pattern of exhaled breath and EBC pH (p=0.01, r=-0.34).Physical exercise changes the pattern of exhaled volatiles together with an increase in pH of breath. Changes in volatiles may be responsible for increase in EBC pH.

Adenosine triphosphate in exhaled breath condensate of healthy subjects and patients with chronic obstructive pulmonary disease.

OBJECTIVES: The effect of hypoxic relapse of chronic obstructive pulmonary disease (COPD) on lung adenosine triphosphate (ATP) concentration was studied measuring ATP in exhaled breath condensate (EBC). SUBJECTS: Thirty COPD patients with severe exacerbation, thirteen healthy non-smokers and thirteen healthy smokers. METHODS: ATP was detected using a luciferin-luciferase assay, dilution of airway droplets in EBC was assessed measuring sample conductivity. RESULTS: ATP concentrations were similar in COPD patients, non-smoking and smoking healthy individuals (141 +/- 44, 115 +/- 21 and 90 +/- 15 pM; p = 0.66). After treatment oxygenation of COPD patients improved (6.85 +/- 1.29 kPa vs. 8.20 +/- 1.28 kPa, p < 0.001), but EBC ATP concentration was similar to that of admission (p = 0.84). There was no correlation between EBC ATP concentration and airway droplet dilution. CONCLUSION: ATP detected in EBC indicates the presence of ATP in airway lining fluid. Lack of difference in ATP concentration between health and COPD suggests that airway ATP level is under complex control of multiple factors.

Angiotensin type 1 and type 2 receptor blockade in chronic allograft nephropathy.

Angiotensin-II (Ang-II) type 1 (AT(1)) receptor blockers may delay the progression of chronic allograft nephropathy (CAN). However, neither the optimal time for initiating AT(1) receptor blockade in order to delay CAN potentially nor the role of Ang-II type 2 (AT(2)) receptors under AT(1) receptor blockade is known. Both AT receptors can regulate p53 expression and apoptosis. We investigated what time of initiation with AT(1) blockers most effectively delayed CAN as well as the role of the AT(2) receptor, and how angiotensin receptor blockade affected apoptosis and its regulating factors in this context in a rat model. Kidneys of Fisher (F344) rats were transplanted into Lewis rats. Animals were treated with AT(1) (candesartan) and/or AT(2) (PD123319) receptor antagonists, a calcium channel blocker, or vehicle (treatment periods: day -7 before to week 24 after transplantation (long term), week 12 to week 24 (late), day -7 to day +5 (early)) and observed the animals for 24 weeks. Reduction of proteinuria, grade of CAN, and number of apoptotic cells was most pronounced in animals receiving long-term AT(1) receptor blockade. A combined AT(1)/AT(2) blocker treatment reduced CAN similarly to AT(1) blocker treatment alone. The number of apoptotic cells and the level of p53 mRNA were significantly lower in long-term AT(1) blocker-treated animals. In summary, AT(1) receptor blockade delayed the progression of CAN, particularly in animals treated long term. Reduction of apoptosis could be related to these beneficial effects. The AT(2) receptor does not appear to play an important role in CAN.

Contribution of androgens to chronic allograft nephropathy is mediated by dihydrotestosterone.

BACKGROUND: Donor and recipient gender influence long-term allograft outcome after kidney transplantation. Sex hormones are likely to contribute to these gender-related differences. The present study investigated the role of androgens and their inhibition on the development of chronic allograft nephropathy. METHODS: Male or female Fisher (F344) kidneys were orthotopically transplanted into intact male Lewis recipients. Animals were treated either with testosterone, the antiandrogen flutamide, the 5alpha-reductase inhibitor finasteride, or vehicle. Twenty weeks after transplantation animals were harvested for histology, immunohistology, and molecular analysis. RESULTS: Testosterone treatment resulted in an increased proteinuria as well as profound glomerulosclerosis, tubulointerstitial fibrosis, and mononuclear cell infiltration that paralleled enhanced intragraft mRNA levels of transforming growth factor-beta (TGF-beta) and platelet-derived growth factor-A and -B chain (PDGF-A and -B). In contrast, flutamide and finasteride reduced glomerulosclerosis as well as the inflammatory cell infiltration associated with decreased TGF-beta, PDGF-A, and -B chain mRNA expression. No gender-related donor differences were noted between the groups. CONCLUSIONS: Our data suggest that dihydrotestosterone mediates the adverse effects of androgens on chronic allograft nephropathy. The inhibition of androgens improves long-term allograft outcome after kidney transplantation.

Adenovirus-mediated Bcl-2 gene transfer inhibits apoptosis and promotes survival of allogeneic transplanted hepatocytes.

BACKGROUND: Donor hepatocyte apoptosis that is induced by host cytotoxic T lymphocytes (CTLs) limits the application of hepatocyte transplantation. Hepatocytes from Bcl-2 transgenic mice can resist the lethal effect of anti-Fas antibody. However, the anti-apoptotic effect of Bcl-2 expression on allogeneic transplanted hepatocytes remains elusive. This study tested the feasibility of Bcl-2 gene transfer as an approach to inhibit CTL-mediated apoptosis in allogeneic transplanted hepatocytes. METHODS: An adenovirus vector that encoded human Bcl-2 gene (AdCMVhBcl-2) was used to transfect cultured rat hepatocytes, which were then transplanted into allogeneic spleens. DNA fragmentation and caspase-3 activation were examined by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling assay and immunohistochemistry for active caspase-3, respectively. Cocultivation of hepatocytes and allogeneic CD8(+) T lymphocytes was performed, and cytotoxicity on hepatocytes was examined by alanine transaminase release. RESULTS: Bcl-2 gene transfer inhibited apoptosis and increased liver-associated enzyme activities in allogeneic transplanted hepatocytes, which were associated with inhibition of caspase-3 activation. Alanine transaminase release in hBcl-2 modified hepatocytes was lower compared with controls, which could not be further decreased by inhibition of Fas ligand and granzyme B. CONCLUSIONS: Adenovirus-mediated Bcl-2 gene transfer blocks CTL-mediated apoptosis in allogeneic hepatocytes by inhibition of caspase-3 activation. Bcl-2 gene transfer could be used to promote survival of transplanted hepatocytes.

Extracellular signal-regulated kinase and the small GTP-binding protein p21Rac1 are involved in the regulation of gene transcription by angiotensin II.

To study the role of extracellular-signal-regulated kinase (ERK) cascade and the small GTP-ase proteins in the activation of the c-fos promoter by angiotensin II (AII), transient transfection experiments were performed in CHO cells stably expressing the rat AT(1A) receptor. In this system AII activated ERK in 1 min and also increased the transcriptional activity of the c-fos promoter-luciferase reporter gene construct. The activation of the promoter proved to be dependent on the Ras-Raf-ERK cascade as cotransfection of expression vectors known to specifically inhibit this cascade blocked the effect of AII. Dominant-negative p21Rac1 mutant partially blocked the activation of the c-fos promoter by AII. However, activation of the c-fos promoter was independent of protein kinase C (PKC) as bisindolylmaleimide I, a specific PKC inhibitor did not block the effect of AII. These results suggest that AII activates the transcription of the c-fos through the Ras-Raf-ERK cascade. Furthermore, p21Rac1 is involved in the modulation of the c-fos promoter by AII.

Angiotensin II: a regulator of inflammation during renal disease?

It has been recently recognized that besides its vasoactive actions Angiotensin II (Ang II) exerts various immunomodulatory effects that may contribute to renal injury and to the progression of renal disease. Consistent with this concept, Ang II facilitates macrophage recruitment into the kidney either directly or through the-upregulation of different chemotactic molecules such as RANTES (Regulated on Activation Normal T Expressed and Secreted), monocyte chemoattractant protein-1 (MCP-1) and osteopontin. Infiltrating macrophages not only produce a number of cytokines, growth factors and proinflammatory Mediators, but also synthesize Ang II intacellularly which increases tissue levels of the hormone within the kidney. Finally, specific binding sites for Ang II have been demonstrated on macrophages and increasing evidence indicates that Ang II directly modulates many of the cellular functions of these cells during the course of renal disease. Together these data suggest that Ang II plays an important role in modulating inflammatory responses in the kidney.

Apoptosis induction and inhibition of cellular proliferation by angiotensin II: possible implication and perspectives.

The renin-angiotensin system plays a pivotal role in the regulation of fluid, electrolyte metabolism and blood pressure. Molecular cloning and pharmacological studies have defined two major classes of Angiotensin II (Ang II) receptors, designated AT1 and AT2. Recently, it has been well recognized that Ang II, beside its classical physiological actions, is a profibrogenic peptide and displays characteristics of a growth factor. The emerging picture suggests that angiotensin receptor subtypes exert opposing features in many aspects of their biological function, most importantly in cellular growth and proliferation. Accordingly, the proliferative and/or growth-promoting effects of Ang II are thought to be mediated by AT1 receptor, whereas the AT2 receptor subtype may have growth-inhibitory properties. The novel finding that Ang II is able to induce apoptosis by AT2 receptors in diverse cell types is of great scientific interest, as recent studies revealed a role for apoptosis as a deliberate form of cell death in the pathogenesis of various cardiovascular diseases such as heart failure and vascular remodeling. Furthermore apoptotic cell death might occur during the development of progressive glomerulosclerosis. It is tempting to speculate that autocrine-paracrine vasoactive substances such as Ang II might regulate these apoptotic processes during pathogenic conditions.

Influence of alternatively and classically activated macrophages on fibrogenic activities of human fibroblasts.

Activated macrophages regulate fibrogenesis by providing cytokines and growth factors that modulate the proliferation and collagen synthesis of fibroblasts. However, macrophages can be activated in a classical pathway induced by LPS or IFN-gamma and an alternative pathway induced by IL-4 or glucocorticoid. Differently activated macrophages display distinct biological features. To clarify the difference between these two subsets of macrophages in the regulatory mechanisms controlling fibrogenesis, human peripheral blood monocytes were used as the source of macrophages and cocultivation of differently activated macrophages and a fibroblast cell line, WI-38, was performed. Alternatively activated macrophages increased the proliferation index and collagen synthesis of cocultivated WI-38 cells in comparison to untreated monocytes, while classically activated macrophages markedly reduced collagen production of cocultivated WI-38 cells. Additionally, mRNA expression and protein production of TGF-beta(1), PDGF-AA, and PDGF-BB were elevated in alternatively activated macrophages in parallel to their profibrogenic effects. In contrast, expression and production of TNF-alpha, as well as MMP-7, were enhanced in classically activated macrophages. These findings suggested that alternatively activated macrophages enhance fibrogenesis of fibroblasts by providing profibrogenic factors, while classically activated macrophages inhibit fibrogenesis of fibroblasts by releasing antifibrogenic or fibrolytic factors.

Effect of angiotensin-converting enzyme inhibition on growth factor mRNA in chronic renal allograft rejection in the rat.

BACKGROUND: Despite considerable progress in immunosuppression, the incidence of chronic renal allograft rejection has not decreased. Recent studies have revealed that angiotensin-converting enzyme (ACE) inhibition ameliorates graft arteriosclerosis, glomerulosclerosis, and tubular atrophy. Moreover, it decreases systemic and glomerular capillary hydrostatic pressure in a rat kidney allograft model. We evaluated the effects of the ACE inhibitor enalapril on cytokine and growth factor expression in chronically rejecting rat kidney allografts. METHODS: Kidneys of Fisher (F344) rats were orthotopically transplanted into Lewis (Lew) rats. To prevent acute rejection, cyclosporine A (1.5 mg/kg/day) was given to all recipients during the first 10 days after transplantation. Enalapril (60 mg/L) or vehicle was added to the drinking water 10 days after transplantation. Animals were harvested 20 weeks after transplantation for histologic and immunohistologic studies, as well as for evaluation of cytokine and growth factor mRNA by semiquantitative polymerase chain reaction. RESULTS: Controls developed severe signs of chronic rejection, such as glomerular and vascular lesions, associated with a large number of infiltrating leukocytes. Enalapril-treated animals developed less proteinuria and other signs of chronic rejection. The mRNA levels of transforming growth factor-beta 1 (TGF-beta 1), platelet-derived growth factor A and B chain (PDGF A and B), insulin-like growth factor-I (IGF-I), interleukin-1 (IL-1), and monocyte chemoattractant protein-1 (MCP-1) were significantly reduced in the enalapril group and were most pronounced for IL-1 and PDGF A. In addition, we found an increased level of renal angiotensinogen mRNA after treatment with enalapril. CONCLUSIONS: Treatment with enalapril attenuated the development of proteinuria, ameliorated morphological damage, decreased leukocyte infiltration, and prevented a rise in renal mRNA levels of growth factors and cytokines in kidney grafts in a rat model of chronic renal allograft rejection.

Interleukin-2 enhances susceptibility of colon cancer cells to FasR mediated apoptosis by up-regulating Fas receptor level and down-regulating FAP-1 expression.

Colon cancer cells are resistant to FasR-mediated apoptosis, which contributes to their evasion from immune attack by CTLs and NK cells. FasR refractoriness of the malignancies was reported to result from loss of Fas receptors and overepression of Fas-associated phosphatase-1 (FAP-1). Therefore, we investigated the effects of recombinant IL-2 on FasR and FAP-1 expression of colon cancer cells, and its influence on the sensitivity of colon cancer cells to FasR-mediated apoptosis. Colon cancer cell lines SW480, HT-29 and CaCo2 were incubated with IL-2 for different periods of time. Sensitivity of the cells to FasR-mediated apoptosis was assessed by measuring their apoptosis after treated with agonistic anti-FasR MAb CH-11. Additionally, Fas receptor levels were examined by immunofluorescence and mRNA expression of FasR and FAP-1 was investigated by RT-PCR: IL-2 incubation increased CH11-induced apoptosis in all colon cancer cell lines in a time-dependent manner. In parallel, IL-2 up-regulated Fas receptor expression on HT29 and CaCo2 cells at both protein and mRNA levels, which were low before treatment, while it served to maintain high expression of FasR on SW480 cells. Additionally, IL-2 down-regulated FAP-1 mRNA expression on SW480 and CaCo2 cells, which was high before treatment. However, low expression of FAP1 on HT-29 cells remained stable after IL-2 treatment. Thus, IL-2 enhances susceptibility of colon cancer cells to FasR-mediated apoptosis by up-regulating Fas receptor levels and by down-regulating FAP-1 expression, which accounts for its therapeutic effects on abolishing immune evasion in colon cancer cells.