Alveolar Wall Area Research Articles

Rapid diagnosis of alveolar echinococcosis from lung puncture sample using metagenomic next-generation sequencing: a case report

IntroductionAlveolar echinococcosis (AE), caused by the larval forms of Echinococcus multilocularis, is a zoonotic disease affecting the liver, lungs, lymph nodes, kidneys, brain, bones, thyroid, and other organs. Diagnosing AE in a non-endemic area is usually challenging. With the rapid development and increasing application of sequencing techniques in recent years, metagenomic next-generation sequencing (mNGS) has become a powerful tool for diagnosing rare infectious diseases.Case PresentationA 45-year-old woman was admitted to the hospital for the presence of pulmonary shadows for more than 3 months. The lung computed tomography (CT) at a local hospital revealed scattered solid and quasi-circular nodules in the left upper lobe, left lower lobe, right middle lobe, and right lower lobe. The largest nodule was located in the dorsal part of the right lung, measuring 2.0 × 1.7 × 1.5 cm. Moreover, abdominal CT revealed one space-occupying lesion each in the left and right lobes. The pathological analysis of the lung biopsy specimen revealed infiltration of lymphocytes, plasma cells, and eosinophils in the alveolar wall and interstitial area. No pathogenic bacteria were observed in the sputum smear and culture tests. There were no parasite eggs in the stool. The mNGS of the lung puncture tissue revealed 6156 sequence reads matching E. multilocularis; thus, the condition was diagnosed as AE. Albendazole 400 mg was administered twice daily, and the patient was stable during follow-up.ConclusionThis case emphasizes the role of mNGS in diagnosing AE. As a novel, sensitive, and accurate diagnostic method, mNGS could be an attractive approach for facilitating early diagnosis and prompt treatment of infectious diseases, especially when the infection was caused by rare pathogens.

Curcumin effect on Bleomycin-induced pulmonary fibrosis in Mus musculus

Curcumin, a curcuminoid compound of turmeric has been demonstrated to have anti-oxidant and anti-inflammatory properties. Bleomycin (BLM) is an anti-cancer drug induced pulmonary fibrosis in human and animals. This study was aimed to investigate biological effects of curcumin on bleomycin-induced pulmonary fibrosis in mice (Mus musculus) through pathomorphological assessment. In this study, 16 mice ddy strain were divided into four groups, namely (i) control, mice were subcutaneously (SC) injected with 100 µl sterilized aquadest in dorsal skin, (ii) BLM group, injected SC with 100 µl of 1 mg/ml BLM in dorsal skin, (iii) Curcumin (CMN) group, mice were intraperitoneally (IP) injected with 100 mg/kg body weight (BW) curcumin dissolved in 0,5% carboxy methyl cellulose (CMC) and injected with 100 µl sterilized aquadest SC, (iv) BLM+CMN group, injected SC with 100 µl of BLM 1 mg/ml and injected IP with 100 mg/kg BW CMN in 0,5% CMC. All treatments were performed daily for four weeks period. The lung samples were collected and fixed in buffered neutral formalin (BNF) 10%. Histopathological evaluation was performed with hematoxylin-eosin (HE) and Masson’s trichrome (MT) stains. The results showed that BLM treatment significantly increased fibrosis area and alveolar wall area fraction as compared to control. OIn the other hand, CMN treatment significantly reduced fibrosis area and alveolar wall area fraction in mice treated with BLM. In conclusion, our study showed that CMN treatment may inhibit lung fibrogenesis in BLM-induced pulmonary fibrosis.

Bleomycin-induced lung fibrosis in IL-4-overexpressing and knockout mice.

The role of IL-4 in the development of lung fibrosis is as yet unclear. Bleomycin (Bleo) or saline (Sal) was injected intratracheally into three groups of C57BL/6J mice: transgenic animals that overexpressed IL-4 (IL-4 TG, n = 14), mice with a targeted knockout mutation of the IL-4 gene (IL-4 KO, n = 11), and wild-type (WT, n = 13) mice. At 14 days, lung fibrosis was evaluated by hydroxyproline measurement and by quantitative image analysis of fibrosis fraction and alveolar wall area fraction. Bronchoalveolar lavage cell counts in all Bleo-treated groups demonstrated an increased percentage of lymphocytes with a corresponding decrease in the percentage of macrophages. Comparing Bleo- to Sal-treated controls within each group of mice showed increases in all lung fibrosis parameters in IL-4 KO and WT, but not in any of the parameters in IL-4 TG mice. The severity of Bleo-induced fibrotic response was decreased in overexpressed IL-4 TG compared with IL-4 KO mice. These data negate a critical profibrotic role for IL-4 in Bleo-induced lung fibrosis.

Time course of bleomycin-induced lung fibrosis.

Intratracheal instillation (IT) of bleomycin is a widely used experimental model for lung fibrosis. In this study we describe the time-course of bleomycin-induced lung fibrosis in mice using computer-assisted morphometry. C57Bl/6J mice were treated with a single IT dose of bleomycin or control saline. Animals were killed 3, 6, 14 and 21 days post-IT. Lung injury was evaluated by analysis of bronchoalveolar lavage (BAL) fluid, hydroxyproline concentration in the lung, routine light microscopic examination resulting in a semiquantitative morphological index (SMI) of lung injury, and quantitative morphological measurements (fibrosis fraction and alveolar wall area fraction) aided by optimas image analysis software. Changes in BAL fluid attributed to bleomycin treatment include increased total cell count (days 14 and 21), and increased percentage of neutrophils (days 3 and 6) followed by a sustained increase in lymphocytes (days 6, 14 and 21). Hydroxyproline levels increased in bleomycin-treated mice on days 14 and 21. Median SMI grades were significantly elevated on days 3, 14 and 21. Computer-assisted morphometry demonstrated a 3-fold increase in fibrosis fraction and a 1.3-fold increase in wall area fraction in bleomycin-treated mice on day 14, with no further increase on day 21. These data also demonstrate that the most suitable time point for assessing lung fibrosis in this model is 14 days after IT instillation of bleomycin, based on the observation that at 14 days the animals developed extensive fibrosis, but had less variability in the fibrotic response and lower mortality than later at 21 days. Computer-assisted morphometry provides objective and quantitative measurements that are a useful tool for the evaluation of bleomycin-induced lung injury.

Effect of Immunomodulators on Bleomycin-Induced Lung Injury

Background: The role of lymphocytes and their subpopulations in lung fibrosis is as yet unclear. Objective: To define the role of immunomodulation in bleomycin-induced inflammatory fibrotic lung injury, by testing the effect of two known Th1 inhibitors: linomide and pentoxifylline. Methods: C57BL/6 mice were treated by a single intratracheal instillation of 0.06 mg bleomycin in 0.01 ml saline or saline alone. Treatment groups included: (1) intratracheal bleomycin and daily treatment with linomide or pentoxifylline; (2) intratracheal bleomycin and daily water; (3) intratracheal saline and daily linomide or pentoxifylline; (4) intratracheal saline and daily water. Linomide and pentoxifylline were available per os in the drinking water from 1 day prior to intratracheal instillation. Animals were studied 14 days after intratracheal instillation. Lung injury was evaluated by total and differential cell count in bronchoalveolar lavage fluid, by a semiquantitative morphological index of lung injury and a quantitative image analysis of cellularity, fibrosis fraction and alveolar wall area fraction, and by biochemical analysis of lung hydroxyproline content. Results: Linomide or pentoxifylline did not cause any lung injury in saline-treated control mice. Overt signs of lung injury were apparent in bleomycin-treated mice. These changes were not affected by daily treatment with linomide or pentoxifylline, which were given in the highest tolerable dose. Conclusion: This study does not support the use of linomide or pentoxifylline to prevent or ameliorate lung fibrosis and may suggest that drug-induced differentiation of T lymphocytes into Th1/th2 subpopulations does not affect the evolution of bleomycin-induced lung injury.

THE EFFECT OF ENOXAPARIN ON BLEOMYCIN-INDUCED LUNG INJURY IN MICE

We have evaluated the effect of enoxaparin, a potent antithrombotic drug, on bleomycin (Bleo) induced pulmonary inflammation in mice. Pulmonary injury was induced by a single intratracheal (IT) instillation of Bleo. Four groups of female C57BL/6 mice, each received one of four treatments: (1) IT Bleo and daily intraperitoneal (IP) injections of enoxaparin (EN) starting one day before IT instillation of Bleo (Bleo-EN); (2) IT Bleo and IP injections of saline (Bleo-Sal); (3) IT saline and IP enoxaparin (Sal-EN); (4) IT saline and IP saline (Sal-Sal). Animals were sacrificed 14 days after IT treatment. Lung injury was evaluated by analysis of bronchoalveolar lavage fluid and histologically by an overall semiquantitative index of lung injury and a quantitative image analysis assessing alveolar wall area fraction and fibrosis fraction. Treatment of mice with enoxaparin did not ameliorate Bleo induced lung injury. Our study does not establish a critical role of procoagulant activity in the evolution of Bleo-induced lung injury and does not support the use of antithrombotic therapy for the prevention of pulmonary fibrosis.

Effects of 0.25 PPM Nitrogen Dioxide on the Developing Mouse Lung. Part 1: Quantitation of Type 2 Cells and Measurements of Alveolar Walls

Weanling Swiss-Webster male mice were exposed intermittently to 0.25 ppm NO2. Lactate dehydrogenase stained frozen sections of gelatin-inflated lungs were used for image analysis measurements of Type 2 cells and alveolar walls at 3 test periods, immediately after 6 wk of exposure, 10 wk postexposure, and 32 wk postexposure, for a total of 196 control and 196 exposed animals. NO2 exposure resulted in a trend toward Type 2 cell hyperplasia and hypertrophy at all 3 test periods, but significant increases in number and size (mean area) of Type 2 cells did not occur until the 32-wk postexposure test (p = .002 and p <.04, respectively). Alveolar wall area, perimeters, and linear intercepts were also consistently greater for the exposed animals at all three test periods, with perimeters and intercepts at borderline levels of significance at the first test period (p = .09 and p = .08, respectively). An increase in the ratio of Type 2 cell number to alveolar wall area (excluding Type 2 cells within the walls) for the NO2-exposed animals approached significance at the 32-wk postexposure period (p = .08). The delay in the detection of significant Type 2 cell alterations until the 32-wk postexposure test seems best explained by an adverse effect of NO2 exposure on lung development and maturation during the 3–9 wk of age growth period when exposure took place. Since significant Type 2 cell alterations were present 32 wk after exposure (41 wk of age), it seems likely that injury to Type 1 cells of the alveolar epithelial lining (implicated by the Type 2 cell hyperplasia) has occurred and was not completely reversible. The concomitant occurrence of an increase in Type 2 cell size is in accord with reports of NO2 -induced edematous swelling but may also reflect a functional hypertrophy. The data add to cumulative evidence that an ambient level of NO2 has an adverse effect on the mammalian lung.

Influence of Community Air Pollution on the Lungs of Mice. Part I: Hyperplasia and Hypertrophy of Type 2 Cells, and Increase in Alveolar Wall Area

AbstractColonies of 115 mice each were exposed for 43 days to the ambient outdoor atmosphere of Los Angeles (LA) and Santa Barbara (SB), cities that frequently and infrequently exceed air quality standards, respectively The air monitoring stations closest to the LA and SB vivarial sites provided data on nitrogen dioxide (NO2), ozone (O3, sulfur dioxide, carbon monoxide, hydrocarbons, and particulates. All Los Angeles air pollutants were higher than those in Santa Barbara. In particular, the 1-h average of O3 exceeded the 1985 California state O3 standard of 0.10 ppm on 21 of the 43 days, versus just 2 for Santa Barbara. The NO2 mean for LA was 4 times higher than the SB mean, 0.10 versus 0.03 ppm, respectively Image analysis measurements of Type 2 cells and alveolar walls showed the following greater measurements for the LA animals: (a) numbers of Type 2 cells (p - .05); (b) mean area of Type 2 cells (p -.06); (c) alveolar wall area (p -.001); and (d) alveolar wall perimeter (p - .001). In addition, the r...

The effect of ozone and simulated high altitude on murine lung elastin: quantitation by image analysis.

Four subgroups of a colony of 50 mice were housed in environmental chambers supplied by particle- and pollutant-filtered air. The animals were exposed to 0.35 ppm ozone and/or simulated high altitude (380 mm Hg) for four weeks, with exposure 4 1/2 hours/day and 5 days/week. Lung elastin area and alveolar wall area were quantitated by computer assisted image analysis of paraffin embedded sections stained with aldehyde fuchsin and metanil yellow. Compared to the controls, the combination of ozone and simulated high altitude resulted in a 54.5% increase in lung elastin area (p less than 0.005), and simulated high altitude by itself increased elastin area by 19.6% (p less than 0.05). Simulated high altitude with and without ozone exposure also increased alveolar wall area (24.8%, p less than 0.01; and 9.7%, NS, respectively). Ozone exposure alone had a reverse effect: a 16.1% decrease in elastin area (p less than 0.1), and a 6.5% decrease in alveolar wall area (NS). Since an intact lung scaffolding is required for full restoration of injured alveolar epithelium and since intact lung elastin is critical for proper lung compliance, the results suggest that ozone exposure at high altitude is most likely to have an adverse effect on lung structure and function.

Image Analysis Quantitation of Type 2 Cells and Alveolar Walls Part II: Influence of 0.3 ppm Nitrogen Dioxide Exposure on the Developing Mouse Lung

Newborn male mice (45 control and 45 experimental) were tested after 6 weeks of 0.3 ppm nitrogen dioxide (NO2) exposure and at postexposure periods of 4 and 10 weeks. After 6 weeks of exposure, there was a 12.9% increase in type 2 cell number that was statistically significant (P &lt; 0.025) and an 11% increase in mean type 2 cell area that fell short of statistical significance. The ratio of cell area to alveolar wall area was also statistically significant (P = 0.05). The latter ratio, dividing a combined measurement of numbers and area of type 2 cells (type 2 cell field area) by the alveolar wall area, serves to control for lung volume. Although increases in type 2 cell number were not statistically significant for the 2 postexposure test periods, a significant increase (P &lt; 0.05) was found by analysis of variance for all 3 test periods. NO2 exposure also resulted in a significant interaction (group x time; P &lt; 0.05) of the type 2 cell field area:alveolar wall area ratio, i.e., a progressive fall for the exposed animals vs. a progressive rise for the control group. The interaction is believed to be the result of NO2-induced type 2 cell swelling followed by impairment of normal type 2 cell growth. The increase in type 2 cell numbers most likely reflects damage to the type 1 cell population since type 2 cell hyperplasia is a common denominator for diverse kinds of human lung disease where damaged type 1 cells are replaced by type 2 cells. The finding of a statistically significant interaction of the type 2 cell field area:wall area ratio and the persistence of increased alveolar wall area imply that the effects of NO2 on type 2 cells and alveolar walls were not completely reversed 10 weeks after exposure. Should the effects be irreversible, there would be the further implication of a commensurate depletion of structural and functional reserves of the lung.

Image Analysis Quantitation of Type 2 Cells and Alveolar Walls Part I: Influence of Time on the Developing Mouse Lung

Quantitative measurements of the lungs of Swiss-Webster male mice at 6, 10, and 16 weeks of age were obtained using image analysis. The measurements included numbers and area of type 2 cells, and the area, perimeters, and linear intercepts of alveolar walls. In addition, type 2 cell:alveolar wall ratios were used to compare cell and alveolar wall interrelationships with time. The most outstanding of the findings was a 21.8% increase in mean type 2 cell area with time (6 weeks vs. 16 weeks of age; P &lt; 0.05), with only a relatively slight increase (6.3%) in the number of type 2 cells. Type 2 cell (≥ 12 μm) number also increased at 16 weeks of age (P &lt; 0.06). Accompanying the increases in cell size and number was an increase in alveolar wall area (25.6%, or 21.4% with the type 2 cell field area excluded). Since the wall area increase was less than that for type 2 cell field area or type 2 cell mean area but greater than the type 2 cell number increase, there were disparate trends in the ratios comparing cell area and cell number to alveolar wall area. Significantly, type 2 cell numbers, mean type 2 cell area, and alveolar wall area had not plateaued at 16 weeks of age. Thus, there is the implication that the type 2 cell population and the alveolar walls are still undergoing changes at least up to the age of 10 weeks and possibly beyond 16 weeks of age. This is a longer period of lung development than expected from prior literature reports and raises the question of a correspondingly greater period of increased susceptibility to noxious air pollutants. An influence of an ambient level of NO2 exposure on the developing mouse lung is the subject of Part II of this investigation.

An experimental model and automated methodology for the analysis of the effects of ambient levels of air pollutants on the lung

This article discusses a series of studies designed to develop and automate a method for quantitation of type II pneumocyte populations of the lung and their changes in response to 2 ppm NO 2 exposure. The validity and sensitivity of the automated approach were strongly supported by high correlations between manual and automated cell counts ( r>0.8, P<10 -6) and between their respective baselines —numbers of alveoli and alveolar wall area ( r=0.56, P<.002)—as well as by the automated detection of significant hypertrophy of type II cells following the NO 2 exposure ( P<.025). The speed and versitility of the automated image analyzer create the capability for a comprehensive quantitation of the lungs at the cellular level, and the methodology as used could be applied immediately to the study of tissue loss and emphysema, and to the establishment of air quality standards through the measurement of cell population alterations.