Vascular dysfunction by myofibroblast activation in patients with idiopathic pulmonary fibrosis and prognostic significance

In this study, we demonstrated the importance of telomerase protein expression and determined the relationships among telomerase, endothelin-1 (ET-1) and myofibroblasts during early and late remodeling of parenchymal and vascular areas in usual interstitial pneumonia (UIP) using 27 surgical lung biopsies from patients with idiopathic pulmonary fibrosis (IPF). Telomerase+, myofibroblasts α-SMA+, smooth muscle cells caldesmon+, endothelium ET-1+ cellularity, and fibrosis severity were evaluated in 30 fields covering normal lung parenchyma, minimal fibrosis (fibroblastic foci), severe (mural) fibrosis, and vascular areas of UIP by the point-counting technique and a semiquantitative score. The impact of these markers was determined in pulmonary functional tests and follow-up until death from IPF. Telomerase and ET-1 expression was significantly increased in normal and vascular areas compared to areas of fibroblast foci. Telomerase and ET-1 expression was inversely correlated with minimal fibrosis in areas of fibroblast foci and directly associated with severe fibrosis in vascular areas. Telomerase activity in minimal fibrosis areas was directly associated with diffusing capacity of the lung for oxygen/alveolar volume and ET-1 expression and indirectly associated with diffusing capacity of the lungs for carbon monoxide and severe fibrosis in vascular areas. Cox proportional hazards regression revealed a low risk of death for females with minimal fibrosis displaying high telomerase and ET-1 expression in normal areas. Vascular dysfunction by telomerase/ET-1 expression was found earlier than vascular remodeling by myofibroblast activation in UIP with impact on IPF evolution, suggesting that strategies aimed at preventing the effect of these mediators may have a greater impact on patient outcome.

Modeling pulmonary fibrosis by abnormal expression of telomerase/apoptosis/collagen V in experimental usual interstitial pneumonia

Limitations on tissue proliferation capacity determined by telomerase/apoptosis balance have been implicated in pathogenesis of idiopathic pulmonary fibrosis. In addition, collagen V shows promise as an inductor of apoptosis. We evaluated the quantitative relationship between the telomerase/apoptosis index, collagen V synthesis, and epithelial/fibroblast replication in mice exposed to butylated hydroxytoluene (BHT) at high oxygen concentration. Two groups of mice were analyzed: 20 mice received BHT, and 10 control mice received corn oil. Telomerase expression, apoptosis, collagen I, III, and V fibers, and hydroxyproline were evaluated by immunohistochemistry, in situ detection of apoptosis, electron microscopy, immunofluorescence, and histomorphometry. Electron microscopy confirmed the presence of increased alveolar epithelial cells type 1 (AEC1) in apoptosis. Immunostaining showed increased nuclear expression of telomerase in AEC type 2 (AEC2) between normal and chronic scarring areas of usual interstitial pneumonia (UIP). Control lungs and normal areas from UIP lungs showed weak green birefringence of type I and III collagens in the alveolar wall and type V collagen in the basement membrane of alveolar capillaries. The increase in collagen V was greater than collagens I and III in scarring areas of UIP. A significant direct association was found between collagen V and AEC2 apoptosis. We concluded that telomerase, collagen V fiber density, and apoptosis evaluation in experimental UIP offers the potential to control reepithelization of alveolar septa and fibroblast proliferation. Strategies aimed at preventing high rates of collagen V synthesis, or local responses to high rates of cell apoptosis, may have a significant impact in pulmonary fibrosis.

THE PREVALENCE AND PREDICTION OF PULMONARY FIBROSIS IN AN

This thesis describes an ancillary project to the Early Diagnosis of Mesothelioma and Lung Cancer in Prior Asbestos Workers study and was conducted to determine the effects of asbestos exposure, pulmonary function and cigarette smoking in the prediction of pulmonary fibrosis. 613 workers who were occupationally exposed to asbestos for an average of 25.9 (SD=14.69) years were sampled from Sarnia, Ontario. A structured questionnaire was administered during a face-to-face interview along with a low-dose computed tomography (LDCT) of the thorax. Of them, 65 workers (10.7%, 95%CI 8.12—12.24) had LDCT-detected pulmonary fibrosis. The model predicting fibrosis included the variables age, smoking (dichotomized), post FVC % splines and post- FEV1% splines. This model had a receiver operator characteristic area under the curve of 0.738. The calibration of the model was evaluated with R statistical program and the bootstrap optimism-corrected calibration slope was 0.692. Thus, our model demonstrated moderate predictive performance.

Use of algorithms for semi-automatic quantification of pulmonary fibrosis and emphysema

Obtaining a semi-automatic quantification of pathologies found in the lung, through images of high resolution computed tomography (HRCT), is of great importance to aid in medical diagnosis. Paraccocidioidomycosis (PCM) is a systemic disease that affects the lung and even after effective treatment leaves sequels such as pulmonary fibrosis and emphysema. It is very important to the area of tropical diseases that the lung injury be quantified more accurately. In this stud, we propose the development of algorithms in computational environment Matlab® able to objectively quantify lung diseases such as fibrosis and emphysema. The program consists in selecting the region of interest (ROI), and through the use of density masks and filters, obtaining the lesion area quantification in relation to the healthy area of the lung. The proposed method was tested on 15 exams of HRCT of patients with confirmed PCM. To prove the validity and effectiveness of the method, we used a virtual phantom, also developed in this research. © 2013 Springer-Verlag.

The Th17 pathway in the peripheral lung microenvironment interacts with expression of collagen V in the late state of experimental pulmonary fibrosis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Lymphatic fluctuation in the parenchymal remodeling stage of acute interstitial pneumonia, organizing pneumonia, nonspecific interstitial pneumonia and idiopathic pulmonary fibrosis

Because the superficial lymphatics in the lungs are distributed in the subpleural, interlobular and peribroncovascular interstitium, lymphatic impairment may occur in the lungs of patients with idiopathic interstitial pneumonias (IIPs) and increase their severity. We investigated the distribution of lymphatics in different remodeling stages of IIPs by immunohistochemistry using the D2-40 antibody. Pulmonary tissue was obtained from 69 patients with acute interstitial pneumonia/diffuse alveolar damage (AIP/DAD, N = 24), cryptogenic organizing pneumonia/organizing pneumonia (COP/OP, N = 6), nonspecific interstitial pneumonia (NSIP/NSIP, N = 20), and idiopathic pulmonary fibrosis/usual interstitial pneumonia (IPF/UIP, N = 19). D2-40+ lymphatic in the lesions was quantitatively determined and associated with remodeling stage score. We observed an increase in the D2-40+ percent from DAD (6.66 +/- 1.11) to UIP (23.45 +/- 5.24, P = 0.008) with the advanced process of remodeling stage of the lesions. Kaplan-Meier survival curves showed a better survival for patients with higher lymphatic D2-40+ expression than 9.3%. Lymphatic impairment occurs in the lungs of IIPs and its severity increases according to remodeling stage. The results suggest that disruption of the superficial lymphatics may impair alveolar clearance, delay organ repair and cause severe disease progress mainly in patients with AIP/DAD. Therefore, lymphatic distribution may serve as a surrogate marker for the identification of patients at greatest risk for death due to IIPs.

Vascular dysfunction by myofibroblast activation in patients with idiopathic pulmonary fibrosis and prognostic significance

In this study, we demonstrated the importance of telomerase protein expression and determined the relationships among telomerase, endothelin-1 (ET-1) and myofibroblasts during early and late remodeling of parenchymal and vascular areas in usual interstitial pneumonia (UIP) using 27 surgical lung biopsies from patients with idiopathic pulmonary fibrosis (IPF). Telomerase+, myofibroblasts alpha-SMA+, smooth muscle cells caldesmon+, endothelium ET-1+ cellularity, and fibrosis severity were evaluated in 30 fields covering normal lung parenchyma, minimal fibrosis (fibroblastic foci), severe ( mural) fibrosis, and vascular areas of UIP by the point-counting technique and a semiquantitative score. The impact of these markers was determined in pulmonary functional tests and follow-up until death from IPF. Telomerase and ET-1 expression was significantly increased in normal and vascular areas compared to areas of fibroblast foci. Telomerase and ET-1 expression was inversely correlated with minimal fibrosis in areas of fibroblast foci and directly associated with severe fibrosis in vascular areas. Telomerase activity in minimal fibrosis areas was directly associated with diffusing capacity of the lung for oxygen/alveolar volume and ET-1 expression and indirectly associated with diffusing capacity of the lungs for carbon monoxide and severe fibrosis in vascular areas. Cox proportional hazards regression revealed a low risk of death for females with minimal fibrosis displaying high telomerase and ET-1 expression in normal areas. Vascular dysfunction by telomerase/ET-1 expression was found earlier than vascular remodeling by myofibroblast activation in UIP with impact on IPF evolution, suggesting that strategies aimed at preventing the effect of these mediators may have a greater impact on patient outcome.

Increased fibroblast telomerase expression precedes myofibroblast alpha-smooth muscle actin expression in idiopathic pulmonary fibrosis

OBJECTIVE: This study sought to identify the relationship between fibroblast telomerase expression, myofibroblasts, and telomerase-mediated regulatory signals in idiopathic pulmonary fibrosis. METHODS: Thirty-four surgical lung biopsies, which had been obtained from patients with idiopathic pulmonary fibrosis and histologically classified as usual interstitial pneumonia, were examined. Immunohistochemistry was used to evaluate fibroblast telomerase expression, myofibroblast alpha-smooth muscle actin expression and the tissue expression of interleukin-4, transforming growth factor-beta, and basic fibroblast growth factor. The point-counting technique was used to quantify the expression of these markers in unaffected, collapsed, mural fibrosis, and honeycombing areas. The results were correlated to patient survival. RESULTS: Fibroblast telomerase expression and basic fibroblast growth factor tissue expression were higher in collapsed areas, whereas myofibroblast expression and interleukine-4 tissue expression were higher in areas of mural fibrosis. Transforming growth factor-beta expression was higher in collapsed, mural fibrosis and honeycombing areas in comparison to unaffected areas. Positive correlations were found between basic fibroblast growth factor tissue expression and fibroblast telomerase expression and between interleukin-4 tissue expression and myofibroblast alpha-smooth muscle actin expression. Negative correlations were observed between interleukin-4 expression and basic fibroblast growth factor tissue expression in areas of mural fibrosis. Myofibroblast alpha-smooth muscle actin expression and interleukin-4 tissue expression in areas of mural fibrosis were negatively associated with patient survival. CONCLUSION: Fibroblast telomerase expression is higher in areas of early remodeling in lung tissues demonstrating typical interstitial pneumonia, whereas myofibroblast alpha-smooth muscle actin expression predominates in areas of late remodeling. These events seem to be regulated by basic fibroblast growth factor and interleukin-4 tissue expression, respectively.

Increased fibroblast telomerase expression precedes myofibroblast α-smooth muscle actin expression in idiopathic pulmonary fibrosis

OBJECTIVE: This study sought to identify the relationship between fibroblast telomerase expression, myofibroblasts, and telomerase-mediated regulatory signals in idiopathic pulmonary fibrosis. METHODS: Thirty-four surgical lung biopsies, which had been obtained from patients with idiopathic pulmonary fibrosis and histologically classified as usual interstitial pneumonia, were examined. Immunohistochemistry was used to evaluate fibroblast telomerase expression, myofibroblast α-smooth muscle actin expression and the tissue expression of inter leu kin-4, transforming growth factor-β, and basic fibroblast growth factor. The point-counting technique was used to quantify the expression of these markers in unaffected, collapsed, mural fibrosis, and honeycombing areas. The results were correlated to patient survival. RESULTS: Fibroblast telomerase expression and basic fibroblast growth factor tissue expression were higher in collapsed areas, whereas myofibroblast expression and interleukine-4 tissue expression were higher in areas of mural fibrosis. Transforming growth factor-β expression was higher in collapsed, mural fibrosis and honeycombing areas in comparison to unaffected areas. Positive correlations were found between basic fibroblast growth factor tissue expression and fibroblast telomerase expression and between interleukin-4 tissue expression and myofibroblast α-smooth muscle actin expression. Negative correlations were observed between interleukin-4 expression and basic fibroblast growth factor tissue expression in areas of mural fibrosis. Myofibroblast α-smooth muscle actin expression and interleukin-4 tissue expression in areas of mural fibrosis were negatively associated with patient survival. CONCLUSION: Fibroblast telomerase expression is higher in areas of early remodeling in lung tissues demonstrating typical interstitial pneumonia, whereas myofibroblast α-smooth muscle actin expression predominates in areas of late remodeling. These events seem to be regulated by basic fibroblast growth factor and interleukin-4 tissue expression, respectively.