Margit V. Szabari

Lung volumes and respiratory mechanics in elastase-induced emphysema in mice

Absolute lung volumes such as functional residual capacity, residual volume (RV), and total lung capacity (TLC) are used to characterize emphysema in patients, whereas in animal models of emphysema, the mechanical parameters are invariably obtained as a function of transrespiratory pressure (Prs). The aim of the present study was to establish a link between the mechanical parameters including tissue elastance (H) and airway resistance (Raw), and thoracic gas volume (TGV) in addition to Prs in a mouse model of emphysema. Using low-frequency forced oscillations during slow deep inflation, we tracked H and Raw as functions of TGV and Prs in normal mice and mice treated with porcine pancreatic elastase. The presence of emphysema was confirmed by morphometric analysis of histological slices. The treatment resulted in an increase in TGV by 51 and 44% and a decrease in H by 57 and 27%, respectively, at 0 and 20 cmH(2)O of Prs. The Raw did not differ between the groups at any value of Prs, but it was significantly higher in the treated mice at comparable TGV values. In further groups of mice, tracheal sounds were recorded during inflations from RV to TLC. All lung volumes but RV were significantly elevated in the treated mice, whereas the numbers and size distributions of inspiratory crackles were not different, suggesting that the airways were not affected by the elastase treatment. These findings emphasize the importance of absolute lung volumes and indicate that tissue destruction was not associated with airway dysfunction in this mouse model of emphysema.

Birefringence microscopy platform for assessing airway smooth muscle structure and function in vivo

A birefringence microscopy technique provides structural and functional information about airway smooth muscle in healthy and mild asthmatic subjects. Advanced analysis of asthma Not all individuals who have respiratory reactions to allergens progress to asthma. In this issue, Cho et al. found that although allergic asthmatics and allergic nonasthmatics both experienced inflammation after allergen challenge, asthmatics had more mucin and type 2 cytokines, and allergen-specific T cells sampled from the airspace had increased innate type 2 receptors. Using orientation-resolved optical coherence tomography, described by Adams et al., they demonstrated that allergic asthmatics also had increased airway smooth muscle mass. This technique allows for in vivo imaging of airway smooth muscle structure and function, which could shed light on the pathogenesis of many respiratory diseases. The inability to visualize airway smooth muscle (ASM) cells in vivo is a major obstacle in understanding their role in normal physiology and diseases. At present, there is no imaging modality available to assess ASM in vivo. Confocal endomicroscopy lacks the penetration depth and field of view, and conventional optical coherence tomography (OCT) does not have sufficient contrast to differentiate ASM from surrounding tissues. We have developed a birefringence microscopy platform that leverages the micro-organization of tissue to add further dimension to traditional OCT. We have used this technology to validate ASM measurements in ex vivo swine and canine studies, visualize and characterize volumetric representations of ASM in vivo, and quantify and predict ASM contractile force as a function of optical retardation. We provide in vivo images and volumetric assessments of ASM in living humans and document structural disease variations in subjects with mild asthma. The opportunity to link inflammatory responses to ASM responses and to link ASM responses to clinical responses and outcomes could lead to an increased understanding of diseases of the airway and, ultimately, to improved patient outcomes.

Emphysema and Mechanical Stress-Induced Lung Remodeling

Transpulmonary pressure and the mechanical stresses of breathing modulate many essential cell functions in the lung via mechanotransduction. We review how mechanical factors could influence the pathogenesis of emphysema. Although the progression of emphysema has been linked to mechanical rupture, little is known about how these stresses alter lung remodeling. We present possible new directions and an integrated multiscale view that may prove useful in finding solutions for this disease.

Functional and morphological assessment of early impairment of airway function in a rat model of emphysema

The aim of this study was to evaluate airway structure-function relations in elastase-induced emphysema in rats. Sprague-Dawley rats were treated intratracheally with 50 IU porcine pancreatic elastase (PPE, n = 8) or saline (controls, n = 6). Six weeks later, lung volumes [functional residual capacity (FRC), residual volume (RV), and total lung capacity (TLC)] and low-frequency impedance parameters (Newtonian resistance, R(N); tissue damping; tissue elastance, H) were measured, and tracheal sounds were recorded during slow inflation to TLC following in vivo degassing. The lungs were fixed and stained for standard morphometry, elastin, and collagen. In the PPE group, FRC and RV were higher [4.53 ± 0.7 (SD) vs. 3.28 ± 0.45 ml; P = 0.003 and 1.06 ± 0.35 vs. 0.69 ± 0.18 ml; P = 0.036, respectively], and H was smaller in the PPE-treated rats than in the controls (1,344 ± 216 vs. 2,178 ± 305 cmH(2)O/l; P < 0.001), whereas there was no difference in R(N). The average number of crackles per inflation was similar in the two groups; however, the crackle size distributions were different and the lower knee of the pressure-volume curves was higher in the PPE group. Microscopic images revealed different alveolar size distributions but similar bronchial diameters in the two groups. The treatment caused a slight but significant decrease in the numbers of alveolar attachments, no difference in elastin and slightly increased mean level and heterogeneity of collagen in the bronchial walls. These results suggest that tissue destruction did not affect the conventionally assessed airway resistance in this emphysema model, whereas the alterations in the recruitment dynamics can be an early manifestation of impaired airway function.

Acute mechanical forces cause deterioration in lung structure and function in elastase-induced emphysema.

The relation between the progression of chronic obstructive pulmonary disease (COPD) and exacerbations is unclear. Currently, no animal model of acute exacerbation of COPD (AECOPD) exists. The objectives of this study were to evaluate the effects of mechanical forces induced by deep inspirations (DIs) on short-term deterioration of lung structure and function to mimic AECOPD. At 2, 7, or 21 days after treatment with elastase, mice were ventilated with or without DIs (35 cmH(2)O airway pressure for 3 s, 2 times/min) for 1 h. Functional residual capacity (FRC) was measured with body plethysmography, and respiratory compliance, resistance, and hysteresivity were obtained via forced oscillations. From hematoxylin and eosin-stained sections, equivalent airspace diameters (D), alveolar wall thickness (W(t)), number of septal ruptures (N(sr)), and attachment density (A(d)) around airways were determined. FRC, compliance, and hysteresivity statistically significantly increased with time, and both increased due to DIs. Interestingly, DIs also had an effect on FRC, compliance, resistance, and hysteresivity in control mice. The development of emphysema statistically significantly increased D and W(t) in time, and the DIs caused subtle differences in D. At 21 days, the application of DIs changed the distribution of D, increased W(t) and N(sr), and decreased A(d). These results suggest that once a critical remodeling of the parenchyma has been reached, acute mechanical forces lead to irreversible changes in structure and function, mimicking COPD exacerbations. Thus, the acute application of DIs in mice with emphysema may serve as a useful model of AECOPD.

Lung structure and function in elastase-treated rats: A follow-up study

Structural and functional longitudinal alterations of the lungs were followed in an emphysema model. Rats were treated with porcine pancreatic elastase (PPE, n=21) or saline (controls, C, n=19). Before the treatment and 3, 10, 21 and 105 days thereafter, absolute lung volumes (FRC, TLC and RV) and tissue mechanical parameters (elastance: H; damping: G) were determined. At 3, 21 and 105 days the lungs were fixed in subgroups of rats. From histological samples the equivalent diameter of airspaces (Dalv), elastin (Mec) and collagen densities were assessed. In the PPE group, FRC and RV were higher from 3 days after treatment compared to controls (p<0.001), while TLC exhibited a delayed increase. H and G decreased in the PPE group throughout the study (p<0.001). Higher Mec (p<0.001) and late-phase inflammation were observed at 105 days. We conclude that during the progression of emphysema, septal failures increase Dalv which decreases H; this reveals a strong structure-function relationship.

Optical coherence tomography (OCT) imaging of dynamic airway behavior in an asthma model (Conference Presentation)

To better understand bronchoconstriction in asthma, it is critical to dynamically visualize airway behavior in vivo. However, currently available imaging techniques do not have sufficient temporal and spatial resolution to investigate airway dynamics. We propose to use endobronchial Optical Coherence Tomography (OCT) to provide real-time cross-sectional images of airway dynamics with a high spatial resolution. Our aim was to study the structure and function of spatially distinct airways during tidal breathing (TB), breath-holds (BH) at end inspiration, and in a response to single deep inspiration (DI) and multiple DI (MDI) in a preclinical sheep asthma model. Anesthetized and mechanically ventilated sheep (n=3) were imaged with OCT in 4 dependent and 4 non-dependent airways at baseline and in methacholine constricted airways. We assessed airway morphology during TB, BH, DI and MDI maneuvers. The change in airway lumen area was found to be greater in the dependent airways compared to the non-dependent airways during TB (dependent: +14.9%, non-dependent: +6%) at baseline. Similarly, the dependent airways dilated more than the non-dependent airways in response to BH (dependent: +7.9%, non-dependent: +5.7%) in relaxed condition. Conversely, in the constricted lung, the DI and MDI maneuvers dilated the non-dependent airways (+13.6% DI, +44% MDI) more than the dependent airways (+6% DI, +15.5% MDI). Overall, dependent airways were more distensible than non-dependent airways during TB and BH, while this behavior was reversed following DI and MDI maneuvers in constricted airways possibly due to a greater local methacholine delivery due to gravitational dependencies on perfusion.

Automated segmentation and quantification of airway mucus with endobronchial optical coherence tomography

We propose a novel suite of algorithms for automatically segmenting the airway lumen and mucus in endobronchial optical coherence tomography (OCT) data sets, as well as a novel approach for quantifying the contents of the mucus. Mucus and lumen were segmented using a robust, multi-stage algorithm that requires only minimal input regarding sheath geometry. The algorithm performance was highly accurate in a wide range of airway and noise conditions. Mucus was classified using mean backscattering intensity and grey level co-occurrence matrix (GLCM) statistics. We evaluated our techniques in vivo in asthmatic and non-asthmatic volunteers.

Exploiting the relationship between birefringence and force to measure airway smooth muscle contraction with PS-OCT (Conference Presentation)

The ability to observe airway dynamics is fundamental to forming a complete understanding of pulmonary diseases such as asthma. We have previously demonstrated that Optical Coherence Tomography (OCT) can be used to observe structural changes in the airway during bronchoconstriction, but standard OCT lacks the contrast to discriminate airway smooth muscle (ASM) bands- ASM being responsible for generating the force that drives airway constriction- from the surrounding tissue. Since ASM in general exhibits a greater degree of birefringence than the surrounding tissue, a potential solution to this problem lies in the implementation of polarization sensitivity (PS) to the OCT system. By modifying the OCT system so that it is sensitive to the birefringence of tissue under inspection, we can visualize the ASM with much greater clarity and definition. In this presentation we show that the force of contraction can be indirectly measured by an associated increase in the birefringence signal of the ASM. We validate this approach by attaching segments of swine trachea to an isometric force transducer and stimulating contraction, while simultaneously measuring the exerted force and imaging the segment with PS-OCT. We then show how our results may be used to extrapolate the force of contraction of closed airways in absence of additional measurement devices. We apply this technique to assess ASM contractility volumetrically and in vivo, in both asthmatic and non-asthmatic human volunteers.

A study of airway smooth muscle in asthmatic and non-asthmatic airways using PS-OCT (Conference Presentation)

Present understanding of the pathophysiological mechanisms of asthma has been severely limited by the lack of an imaging modality capable of assessing airway conditions of asthma patients in vivo. Of particular interest is the role that airway smooth muscle (ASM) plays in the development of asthma and asthma related symptoms. With standard Optical Coherence Tomography (OCT), imaging ASM is often not possible due to poor structural contrast between the muscle and surrounding tissues. A potential solution to this problem is to utilize additional optical contrast factors intrinsic to the tissue, such as birefringence. Due to its highly ordered structure, ASM is strongly birefringent. Previously, we demonstrated that Polarization Sensitive OCT(PS-OCT) has the potential to be used to visualize ASM as well as easily segment it from the surrounding (weakly) birefringent tissue by exploiting a property which allows it to discriminate the orientation of birefringent fibers. We have already validated our technology with a substantial set of histological comparisons made against data obtained ex vivo. In this work we present a comprehensive comparison of ASM distributions in asthmatic and non-asthmatic human volunteers. By isolating the ASM we parameterize its distribution in terms of both thickness and band width, calculated volumetrically over centimeters of airway. Using this data we perform analyses of the asthmatic and non-asthmatic airways using a broad number and variety and subjects.